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Qi L, Duan R, Zhou J, Guo Y, Zhang C. Novel osteogenic peptide from bovine bone collagen hydrolysate: Targeted screening, molecular mechanism, and stability analysis. Food Chem 2024; 459:140359. [PMID: 38996641 DOI: 10.1016/j.foodchem.2024.140359] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/05/2024] [Revised: 05/07/2024] [Accepted: 07/03/2024] [Indexed: 07/14/2024]
Abstract
This study aimed to screen for a novel osteogenic peptide based on the calcium-sensing receptor (CaSR) and explore its molecular mechanism and gastrointestinal stability. In this study, a novel osteogenic peptide (Phe-Ser-Gly-Leu, FSGL) derived from bovine bone collagen hydrolysate was successfully screened by molecular docking and synthesised by solid phase peptide synthesis for further analysis. Cell experiments showed that FSGL significantly enhanced the osteogenic activity of MC3T3-E1 cells by acting on CaSR, including proliferation (152.53%), differentiation, and mineralization. Molecular docking and molecular dynamics further demonstrated that FSGL was a potential allosteric activator of CaSR, that turned on the activation switch of CaSR by closing the Venus flytrap (VFT) domain and driving the two protein chains in the VFT domain to easily form dimers. In addition, 96.03% of the novel osteogenic peptide FSGL was stable during gastrointestinal digestion. Therefore, FSGL showed substantial potential for enhancing the osteogenic activity of osteoblasts. This study provided new insights for the application of CaSR in the targeted screening of osteogenic peptides to improve bone health.
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Affiliation(s)
- Liwei Qi
- Key Laboratory of Agro-Products Processing, Ministry of Agriculture and Rural Affairs, Institute of Food Science and Technology, Chinese Academy of Agricultural Sciences, Beijing 100193, China
| | - Ruipei Duan
- Key Laboratory of Agro-Products Processing, Ministry of Agriculture and Rural Affairs, Institute of Food Science and Technology, Chinese Academy of Agricultural Sciences, Beijing 100193, China
| | - Jiaojiao Zhou
- Key Laboratory of Agro-Products Processing, Ministry of Agriculture and Rural Affairs, Institute of Food Science and Technology, Chinese Academy of Agricultural Sciences, Beijing 100193, China
| | - Yujie Guo
- Key Laboratory of Agro-Products Processing, Ministry of Agriculture and Rural Affairs, Institute of Food Science and Technology, Chinese Academy of Agricultural Sciences, Beijing 100193, China.
| | - Chunhui Zhang
- Key Laboratory of Agro-Products Processing, Ministry of Agriculture and Rural Affairs, Institute of Food Science and Technology, Chinese Academy of Agricultural Sciences, Beijing 100193, China.
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Qi L, Zhang H, Guo Y, Zhang C, Xu Y. Novel Calcium-Binding Peptide from Bovine Bone Collagen Hydrolysates and Its Potential Pro-Osteogenic Activity via Calcium-Sensing Receptor (CaSR). Mol Nutr Food Res 2024; 68:e2200726. [PMID: 38161238 DOI: 10.1002/mnfr.202200726] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/24/2022] [Revised: 07/25/2023] [Indexed: 01/03/2024]
Abstract
SCOPE This paper aims to explore the osteogenic activity and potential mechanism of the peptide-calcium chelate, and provides a theoretical basis for peptide-calcium chelates as functional foods to prevent or improve osteoporosis. METHODS AND RESULTS In this research, a novel peptide (Phe-Gly-Leu, FGL) with a high calcium-binding capacity is screened from bovine bone collagen hydrolysates (CPs), calcium binding sites of which mainly included carbonyl, amino and carboxyl groups. The FGL-Ca significantly enhances the osteogenic activity of MC3T3-E1 cells (survival rate, differentiation, and mineralization). The results of calcium fluorescence labeling and molecular docking show that FGL-Ca may activate calcium-sensing receptor (CaSR), leading to an increase in intracellular calcium concentration, then enhancing osteogenic activity of MC3T3-E1 cells. Further research found that FGL-Ca significantly promotes the mRNA and protein expression levels of CaSR, transforming growth factor β (TGF-β1), TGF-β-type II receptor (TβRII), Smad2, Smad3, osteocalcin (OCN), alkaline phosphatase (ALP), osteoprotegrin (OPG), and collagen type I (COLI). Subsequently, in the signal pathway intervention experiment, the expression levels of genes and proteins related to the TGF-β1/Smad2/3 signaling pathway that are promoted by FGL-Ca are found to decrease. CONCLUSIONS These results suggest that FGL-Ca may activate CaSR, increase intracellular calcium concentration, and activate TGF-β1/Smad2/3 signaling pathway, which may be one of the potential mechanisms for enhancing osteogenic activity.
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Affiliation(s)
- Liwei Qi
- Key Laboratory of Agro-Products Processing, Ministry of Agriculture and Rural Affairs, Institute of Food Science and technology, Chinese Academy of Agricultural Sciences, Beijing, 100193, China
| | - Hongru Zhang
- Key Laboratory of Agro-Products Processing, Ministry of Agriculture and Rural Affairs, Institute of Food Science and technology, Chinese Academy of Agricultural Sciences, Beijing, 100193, China
- Laboratory of Biomass and Green Technologies, University of Liege-Gembloux Agro-Bio Tech, Passage des déportés 2, B-5030, Gembloux, Belgium
| | - Yujie Guo
- Key Laboratory of Agro-Products Processing, Ministry of Agriculture and Rural Affairs, Institute of Food Science and technology, Chinese Academy of Agricultural Sciences, Beijing, 100193, China
| | - Chunhui Zhang
- Key Laboratory of Agro-Products Processing, Ministry of Agriculture and Rural Affairs, Institute of Food Science and technology, Chinese Academy of Agricultural Sciences, Beijing, 100193, China
| | - Yang Xu
- Inner Mongolia Mengtai Biological Engineering Co., Ltd., Shengle Economic Park, Helinger County, Hohhot, Inner Mongolia, 010000, China
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Tang LQ, Fraebel J, Jin S, Winesett SP, Harrell J, Chang WH, Cheng SX. Calcium/calcimimetic via calcium-sensing receptor ameliorates cholera toxin-induced secretory diarrhea in mice. World J Gastroenterol 2024; 30:268-279. [PMID: 38314127 PMCID: PMC10835527 DOI: 10.3748/wjg.v30.i3.268] [Citation(s) in RCA: 1] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/02/2023] [Revised: 12/01/2023] [Accepted: 01/02/2024] [Indexed: 01/18/2024] Open
Abstract
BACKGROUND Enterotoxins produce diarrhea through direct epithelial action and indirectly by activating the enteric nervous system. Calcium-sensing receptor (CaSR) inhibits both actions. The latter has been well documented in vitro but not in vivo. The hypothesis to be tested was that activating CaSR inhibits diarrhea in vivo. AIM To determine whether CaSR agonists ameliorate secretory diarrhea evoked by cholera toxin (CTX) in mice. METHODS CTX was given orally to C57BL/6 mice to induce diarrhea. Calcium and calcimimetic R568 were used to activate CaSR. To maximize their local intestinal actions, calcium was administered luminally via oral rehydration solution (ORS), whereas R568 was applied serosally using an intraperitoneal route. To verify that their actions resulted from the intestine, effects were also examined on Cre-lox intestine-specific CaSR knockouts. Diarrhea outcome was measured biochemically by monitoring changes in fecal Cl- or clinically by assessing stool consistency and weight loss. RESULTS CTX induced secretory diarrhea, as evidenced by increases in fecal Cl-, stool consistency, and weight loss following CTX exposure, but did not alter CaSR, neither in content nor in function. Accordingly, calcium and R568 were each able to ameliorate diarrhea when applied to diseased intestines. Intestinal CaSR involvement is suggested by gene knockout experiments where the anti-diarrheal actions of R568 were lost in intestinal epithelial CaSR knockouts (villinCre/Casrflox/flox) and neuronal CaSR knockouts (nestinCre/Casrflox/flox). CONCLUSION Treatment of acute secretory diarrheas remains a global challenge. Despite advances in diarrhea research, few have been made in the realm of diarrhea therapeutics. ORS therapy has remained the standard of care, although it does not halt the losses of intestinal fluid and ions caused by pathogens. There is no cost-effective therapeutic for diarrhea. This and other studies suggest that adding calcium to ORS or using calcimimetics to activate intestinal CaSR might represent a novel approach for treating secretory diarrheal diseases.
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Affiliation(s)
- Lie-Qi Tang
- Department of Pediatrics, University of Florida, Gainesville, FL 32610, United States
| | - Johnathan Fraebel
- Department of Pediatrics, University of Florida, Gainesville, FL 32610, United States
- College of Medicine, University of Florida, Gainesville, FL 32610, United States
| | - Shi Jin
- Department of Pediatrics, University of Florida, Gainesville, FL 32610, United States
| | - Steven P Winesett
- Applied Physiology and Kinesiology, University of Florida, Gainesville, FL 32610, United States
- Brain Rehabilitation Research Center, Malcom Randall VA Medical Center, Gainesville, FL 32610, United States
| | - Jane Harrell
- Department of Pediatrics, University of Florida, Gainesville, FL 32610, United States
| | - Wen-Han Chang
- Department of Medicine, Endocrine Research Unit, Veterans Affairs Medical Center, University of California, San Francisco, San Francisco, CA 94121, United States
| | - Sam Xianjun Cheng
- Department of Pediatric Gastroenterology, Hepatology, and Nutrition, University of Florida Shands Children’s Hospital, Gainesville, FL 32608, United States
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Qi L, Zhang H, Guo Y, Zhang C, Xu Y. A novel calcium-binding peptide from bovine bone collagen hydrolysate and chelation mechanism and calcium absorption activity of peptide-calcium chelate. Food Chem 2023; 410:135387. [PMID: 36621334 DOI: 10.1016/j.foodchem.2023.135387] [Citation(s) in RCA: 9] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/24/2022] [Revised: 12/11/2022] [Accepted: 01/02/2023] [Indexed: 01/05/2023]
Abstract
A novel calcium-binding peptide from bovine bone collagen hydrolysate was screened based on a new target-the calcium-sensing receptor (CaSR), and its chelation mechanism and calcium absorption activity were investigated. Glu-Tyr-Gly exhibited superior binding affinities to CaSR because of its interaction with the active sites of the CaSR Venus Flytrap (VFT) domain. Glu-Tyr-Gly-Ca may exist in five potential chelation modes and its potential chelation mechanism was that calcium ions were located in the center and surrounded by ionic bonds (carboxyl group) and coordination bonds (carbonyl, amino, and carboxyl group). Glu-Tyr-Gly-Ca was slightly damaged in the intestinal fluid and at different temperatures, whereas it was severely damaged in the gastric fluid and acidic conditions. The results of the calcium dialysis percentage and Caco-2 cells experiments showed that Glu-Tyr-Gly-Ca possessed good calcium transport activity and bioavailability. The findings provided theoretical basis for Glu-Tyr-Gly-Ca as potential calcium supplement to improve intestinal calcium absorption.
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Affiliation(s)
- Liwei Qi
- Key Laboratory of Agro-Products Processing, Ministry of Agriculture and Rural Affairs, Institute of Food Science and Technology, Chinese Academy of Agricultural Sciences, Beijing 100193, China
| | - Hongru Zhang
- Key Laboratory of Agro-Products Processing, Ministry of Agriculture and Rural Affairs, Institute of Food Science and Technology, Chinese Academy of Agricultural Sciences, Beijing 100193, China; Laboratory of Biomass and Green Technologies, University of Liege-Gembloux Agro-Bio Tech, Passage des déportés 2, B-5030 Gembloux, Belgium
| | - Yujie Guo
- Key Laboratory of Agro-Products Processing, Ministry of Agriculture and Rural Affairs, Institute of Food Science and Technology, Chinese Academy of Agricultural Sciences, Beijing 100193, China.
| | - Chunhui Zhang
- Key Laboratory of Agro-Products Processing, Ministry of Agriculture and Rural Affairs, Institute of Food Science and Technology, Chinese Academy of Agricultural Sciences, Beijing 100193, China.
| | - Yang Xu
- Inner Mongolia Mengtai Biological Engineering Co., Ltd, Shengle Economic Park, Helinger County, Hohhot, Inner Mongolia 010000, China
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Salinity-dependent expression of calcium-sensing receptors in Atlantic salmon (Salmo salar) tissues. J Comp Physiol A Neuroethol Sens Neural Behav Physiol 2021; 207:505-522. [PMID: 34114081 DOI: 10.1007/s00359-021-01493-3] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/31/2020] [Revised: 05/23/2021] [Accepted: 05/26/2021] [Indexed: 10/21/2022]
Abstract
Multiple reports suggest that calcium-sensing receptors (CaSRs) are involved in calcium homeostasis, osmoregulation, and/or salinity sensing in fish (Loretz 2008, Herberger and Loretz 2013). We have isolated three unique full-length CaSR cDNAs from Atlantic salmon (Salmo salar) kidney that share many features with other reported CaSRs. Using anti-CaSR antibodies and PCR primers specific for individual salmon CaSR transcripts we show expression in osmoregulatory, neuroendocrine and sensory tissues. Furthermore, CaSRs are expressed in different patterns in salmon tissues where mRNA and protein expression are modified by freshwater or seawater acclimation. For example, in seawater, CaSR mRNA and protein expression is increased significantly in kidney as compared to freshwater. Electrophysiological recordings of olfactory responses produced upon exposure of salmon olfactory epithelium to CaSR agonists suggest a role for CaSRs in chemoreception in this species consistent with other freshwater, anadromous, and marine species where similar olfactory responses to divalent and polyvalent cations have been reported. These data provide further support for a role of CaSR proteins in osmoregulatory and sensory functions in Atlantic salmon, an anadromous species that experiences a broad range of environmental salinities in its life history.
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Hui Q, Zhao X, Lu P, Liu S, Nyachoti M, O K, Yang C. Molecular distribution and localization of extracellular calcium-sensing receptor (CaSR) and vitamin D receptor (VDR) at three different laying stages in laying hens (Gallus gallus domesticus). Poult Sci 2021; 100:101060. [PMID: 33752067 PMCID: PMC8010884 DOI: 10.1016/j.psj.2021.101060] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/18/2020] [Revised: 11/02/2020] [Accepted: 02/11/2021] [Indexed: 01/06/2023] Open
Abstract
The extracellular calcium-sensing receptor (CaSR) and vitamin D receptor (VDR) play important roles in regulating calcium mobilization, calcium absorption, and calcium homeostasis, and they could be potential therapeutic targets to osteoporosis in laying hens. The present study investigated the molecular distribution of CaSR and VDR and the localization of CaSR in the kidney, proventriculus (true stomach), duodenum, jejunum, ileum, colon, cecum, shell gland, and tibia of laying hens at 3 different laying stages (19, 40, and 55 wk). The results showed that the relative mRNA abundance of CaSR in the kidney, ileum, proventriculus, duodenum, and colon was higher (P < 0.05) than the other tissues at 40 and 55 wk. The relative mRNA abundance of CaSR in the tibia was higher (P < 0.05) at 55 wk than at 40 wk. However, there were no significant differences in the relative protein abundance of CaSR among all tested tissues at peak production or in each tissue at the 3 different laying stages (P > 0.05). The relative mRNA abundance of VDR was higher (P < 0.05) in the small intestine (duodenum, jejunum, and ileum) when compared with other tissues at the 3 different laying stages. The relative protein abundance of VDR in the duodenum was higher (P < 0.05) than that in the proventriculus, colon, and cecum. There were no significant differences in the VDR expression among the tested tissues at the 3 different laying stages (P > 0.05). The immunohistochemical results showed that the positive staining was found widely in each tissue. Moreover, different laying stages did not affect the localization of CaSR except for the tibia tissue. In conclusion, similar to VDR, CaSR was widely expressed not only in the gut but also in the tibia and shell gland in laying hens. The expression level of CaSR and VDR in all tested tissues was unchanged at the different laying stages.
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Affiliation(s)
- Qianru Hui
- Department of Animal Science, University of Manitoba, Winnipeg, Manitoba R3T 2N2, Canada
| | - Xiaoya Zhao
- Department of Animal Science, University of Manitoba, Winnipeg, Manitoba R3T 2N2, Canada
| | - Peng Lu
- Department of Animal Science, University of Manitoba, Winnipeg, Manitoba R3T 2N2, Canada
| | - Shangxi Liu
- Department of Animal Science, University of Manitoba, Winnipeg, Manitoba R3T 2N2, Canada
| | - Martin Nyachoti
- Department of Animal Science, University of Manitoba, Winnipeg, Manitoba R3T 2N2, Canada
| | - Karmin O
- Department of Animal Science, University of Manitoba, Winnipeg, Manitoba R3T 2N2, Canada; CCARM, St. Boniface Hospital Research Centre, Winnipeg, Manitoba R2H 2A6, Canada
| | - Chengbo Yang
- Department of Animal Science, University of Manitoba, Winnipeg, Manitoba R3T 2N2, Canada.
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Wu X, Ju L, Song Y, Bai L, Yuan M, Xu W, Li J, Xu T, Pei L, Sun J. Mechanism of Colonic Slow Wave Rhythm Regulated by Electro-acupuncture Determined using Calcium-Sensitive Receptor. ACUPUNCTURE ELECTRO 2021. [DOI: 10.3727/036012921x16112663844842] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/24/2022]
Abstract
The calcium-sensitive receptor (CaSR) plays a role in several biological processes. However, its role in intestinal motility remains unclear. In this study, we aimed to determine the effect of electro-acupuncture (EA) at Shangjuxu (ST37) on CaSR in colonic dysplasia mice, and to explore
the possible mechanism of EA regulating colonic movement. The mice were injected with nicardipine or hexamethonium bromide to induce colonic dysplasia. Intestinal transit function was assessed by twelve hours fecal granules and fecal water content percentage, while colonic slow wave was assessed
by multi-channel physiological signal acquisition system, immunofluorescence and laser confocal microscopy were used to examine CaSR expression in the gastrointestinal (GI) tract of the mice. We found that the number of fecal particles, the frequency and amplitude of colonic slow wave were
disrupted after nicardipine or hexamethonium bromide injection. In addition, CaSR expression in control group was mainly distributed in intestinal epithelial cells, and the morphological structure of mucosal layer was regular. Compared with control group, the structure of mucosal layer in
nicardipine group and hexamethonium bromide group were all disorderly, the expression and fluorescence intensity of CaSR in nicardipine group were visible, but in hexamethonium bromide group were weakened. After EA intervention, these disorders were ameliorated, which suggested that EA at
ST37 could therefore regulate colonic motility disorders via the involvement of CaSR.
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Wang Y, Alkhalidy H, Liu D. The Emerging Role of Polyphenols in the Management of Type 2 Diabetes. Molecules 2021; 26:molecules26030703. [PMID: 33572808 PMCID: PMC7866283 DOI: 10.3390/molecules26030703] [Citation(s) in RCA: 48] [Impact Index Per Article: 12.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/13/2020] [Revised: 01/25/2021] [Accepted: 01/26/2021] [Indexed: 12/12/2022] Open
Abstract
Type 2 diabetes (T2D) is a fast-increasing health problem globally, and it results from insulin resistance and pancreatic β-cell dysfunction. The gastrointestinal (GI) tract is recognized as one of the major regulatory organs of glucose homeostasis that involves multiple gut hormones and microbiota. Notably, the incretin hormone glucagon-like peptide-1 (GLP-1) secreted from enteroendocrine L-cells plays a pivotal role in maintaining glucose homeostasis via eliciting pleiotropic effects, which are largely mediated via its receptor. Thus, targeting the GLP-1 signaling system is a highly attractive therapeutic strategy to treatment T2D. Polyphenols, the secondary metabolites from plants, have drawn considerable attention because of their numerous health benefits, including potential anti-diabetic effects. Although the major targets and locations for the polyphenolic compounds to exert the anti-diabetic action are still unclear, the first organ that is exposed to these compounds is the GI tract in which polyphenols could modulate enzymes and hormones. Indeed, emerging evidence has shown that polyphenols can stimulate GLP-1 secretion, indicating that these natural compounds might exert metabolic action at least partially mediated by GLP-1. This review provides an overview of nutritional regulation of GLP-1 secretion and summarizes recent studies on the roles of polyphenols in GLP-1 secretion and degradation as it relates to metabolic homeostasis. In addition, the effects of polyphenols on microbiota and microbial metabolites that could indirectly modulate GLP-1 secretion are also discussed.
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Affiliation(s)
- Yao Wang
- Department of Human Nutrition, Foods and Exercise, College of Agricultural and Life Sciences, Virginia Tech, Blacksburg, VA 24060, USA;
| | - Hana Alkhalidy
- Department of Nutrition and Food Technology, Jordan University of Science and Technology, Irbid 22110, Jordan;
| | - Dongmin Liu
- Department of Human Nutrition, Foods and Exercise, College of Agricultural and Life Sciences, Virginia Tech, Blacksburg, VA 24060, USA;
- Correspondence: ; Tel.: +1-540-231-3402; Fax: +1-540-231-3916
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Abstract
Sensing and responding to changes in nutrient levels, including those of glucose, lipids, and amino acids, by the body is necessary for survival. Accordingly, perturbations in nutrient sensing are tightly linked with human pathologies, particularly metabolic diseases such as obesity, type 2 diabetes mellitus, and other complications of metabolic syndromes. The conventional view is that amino acids are fundamental elements for protein and peptide synthesis, while recent studies have revealed that amino acids are also important bioactive molecules that play key roles in signaling pathways and metabolic regulation. Different pathways that sense intracellular and extracellular levels of amino acids are integrated and coordinated at the organismal level, and, together, these pathways maintain whole metabolic homeostasis. In this review, we discuss the studies describing how important sensing signals respond to amino acid availability and how these sensing mechanisms modulate metabolic processes, including energy, glucose, and lipid metabolism. We further discuss whether dysregulation of amino acid sensing signals can be targeted to promote metabolic disorders, and discuss how to translate these mechanisms to treat human diseases. This review will help to enhance our overall understanding of the correlation between amino acid sensing and metabolic homeostasis, which have important implications for human health.
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Affiliation(s)
- Xiaoming Hu
- CAS Key Laboratory of Nutrition, Metabolism and Food Safety, Shanghai Institute of Nutrition and Health, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China
| | - Feifan Guo
- CAS Key Laboratory of Nutrition, Metabolism and Food Safety, Shanghai Institute of Nutrition and Health, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China
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Xu J, Zeug A, Riederer B, Yeruva S, Griesbeck O, Daniel H, Tuo B, Ponimaskin E, Dong H, Seidler U. Calcium-sensing receptor regulates intestinal dipeptide absorption via Ca 2+ signaling and IK Ca activation. Physiol Rep 2020; 8:e14337. [PMID: 31960592 PMCID: PMC6971415 DOI: 10.14814/phy2.14337] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022] Open
Abstract
Although absorption of di- and tripeptides into intestinal epithelial cells occurs via the peptide transporter 1 (PEPT1, also called solute carrier family 15 member 1 (SLC15A1)), the detailed regulatory mechanisms are not fully understood. We examined: (a) whether dipeptide absorption in villous enterocytes is associated with a rise in cytosolic Ca2+ ([Ca2+ ]cyt ), (b) whether the calcium sensing receptor (CaSR) is involved in dipeptide-elicited [Ca2+ ]cyt signaling, and (c) what potential consequences of [Ca2+ ]cyt signaling may enhance enterocyte dipeptide absorption. Dipeptide Gly-Sar and CaSR agonist spermine markedly raised [Ca2+ ]cyt in villous enterocytes, which was abolished by NPS-2143, a selective CaSR antagonist and U73122, an phospholipase C (PLC) inhibitor. Apical application of Gly-Sar induced a jejunal short-circuit current (Isc), which was reduced by NPS-2143. CaSR expression was identified in the lamina propria and on the basal enterocyte membrane of mouse jejunal mucosa in both WT and Slc15a1-/- animals, but Gly-Sar-induced [Ca2+ ]cyt signaling was significantly decreased in Slc15a1-/- villi. Clotrimazole and TRM-34, two selective blockers of the intermediate conductance Ca2+ -activated K+ channel (IKCa ), but not iberiotoxin, a selective blocker of the large-conductance K+ channel (BKCa ) and apamin, a selective blocker of the small-conductance K+ channel (SKCa ), significantly inhibited Gly-Sar-induced Isc in native tissues. We reveal a novel CaSR-PLC-Ca2+ -IKCa pathway in the regulation of small intestinal dipeptide absorption, which may be exploited as a target for future drug development in human nutritional disorders.
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Affiliation(s)
- Jingyu Xu
- Department of Gastroenterology, Hepatology and EndocrinologyHannover Medical SchoolHannoverGermany
- Research GastroenterologyAffiliated Hospital of Zunyi Medical UniversityZunyiChina
| | - Andre Zeug
- Cellular NeurophysiologyHannover Medical SchoolHannoverGermany
| | - Brigitte Riederer
- Department of Gastroenterology, Hepatology and EndocrinologyHannover Medical SchoolHannoverGermany
| | - Sunil Yeruva
- Department of Gastroenterology, Hepatology and EndocrinologyHannover Medical SchoolHannoverGermany
| | | | - Hannelore Daniel
- Nutritional PhysiologyTechnical University of MunichFreisingGermany
| | - Biguang Tuo
- Research GastroenterologyAffiliated Hospital of Zunyi Medical UniversityZunyiChina
| | | | - Hui Dong
- Department of MedicineUniversity of California, San DiegoLa JollaCAUSA
| | - Ursula Seidler
- Department of Gastroenterology, Hepatology and EndocrinologyHannover Medical SchoolHannoverGermany
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Wongdee K, Rodrat M, Teerapornpuntakit J, Krishnamra N, Charoenphandhu N. Factors inhibiting intestinal calcium absorption: hormones and luminal factors that prevent excessive calcium uptake. J Physiol Sci 2019; 69:683-696. [PMID: 31222614 PMCID: PMC10717634 DOI: 10.1007/s12576-019-00688-3] [Citation(s) in RCA: 28] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/04/2019] [Accepted: 06/09/2019] [Indexed: 12/11/2022]
Abstract
Besides the two canonical calciotropic hormones, namely parathyroid hormone and 1,25-dihydroxyvitamin D [1,25(OH)2D3], there are several other endocrine and paracrine factors, such as prolactin, estrogen, and insulin-like growth factor that have been known to directly stimulate intestinal calcium absorption. Generally, to maintain an optimal plasma calcium level, these positive regulators enhance calcium absorption, which is indirectly counterbalanced by a long-loop negative feedback mechanism, i.e., through calcium-sensing receptor in the parathyroid chief cells. However, several lines of recent evidence have revealed the presence of calcium absorption inhibitors present in the intestinal lumen and extracellular fluid in close vicinity to enterocytes, which could also directly compromise calcium absorption. For example, luminal iron, circulating fibroblast growth factor (FGF)-23, and stanniocalcin can decrease calcium absorption, thereby preventing excessive calcium uptake under certain conditions. Interestingly, the intestinal epithelial cells themselves could lower their rate of calcium uptake after exposure to high luminal calcium concentration, suggesting a presence of an ultra-short negative feedback loop independent of systemic hormones. The existence of neural regulation is also plausible but this requires more supporting evidence. In the present review, we elaborate on the physiological significance of these negative feedback regulators of calcium absorption, and provide evidence to show how our body can efficiently restrict a flood of calcium influx in order to maintain calcium homeostasis.
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Affiliation(s)
- Kannikar Wongdee
- Faculty of Allied Health Sciences, Burapha University, Chonburi, Thailand
- Center of Calcium and Bone Research (COCAB), Faculty of Science, Mahidol University, Bangkok, Thailand
| | - Mayuree Rodrat
- Center of Calcium and Bone Research (COCAB), Faculty of Science, Mahidol University, Bangkok, Thailand
- Department of Physiology, Faculty of Science, Mahidol University, Rama VI Road, Bangkok, 10400, Thailand
| | - Jarinthorn Teerapornpuntakit
- Center of Calcium and Bone Research (COCAB), Faculty of Science, Mahidol University, Bangkok, Thailand
- Department of Physiology, Faculty of Medical Science, Naresuan University, Phitsanulok, Thailand
| | - Nateetip Krishnamra
- Center of Calcium and Bone Research (COCAB), Faculty of Science, Mahidol University, Bangkok, Thailand
- Department of Physiology, Faculty of Science, Mahidol University, Rama VI Road, Bangkok, 10400, Thailand
| | - Narattaphol Charoenphandhu
- Center of Calcium and Bone Research (COCAB), Faculty of Science, Mahidol University, Bangkok, Thailand.
- Department of Physiology, Faculty of Science, Mahidol University, Rama VI Road, Bangkok, 10400, Thailand.
- Institute of Molecular Biosciences, Mahidol University, Nakhon Pathom, Thailand.
- The Academy of Science, The Royal Society of Thailand, Dusit, Bangkok, Thailand.
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Wang W, Yu S, Huang S, Deng R, Ding Y, Wu Y, Li X, Wang A, Wang S, Chen W, Lu Y. A Complex Role for Calcium Signaling in Colorectal Cancer Development and Progression. Mol Cancer Res 2019; 17:2145-2153. [PMID: 31366605 DOI: 10.1158/1541-7786.mcr-19-0429] [Citation(s) in RCA: 37] [Impact Index Per Article: 6.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/26/2019] [Revised: 06/27/2019] [Accepted: 07/29/2019] [Indexed: 11/16/2022]
Abstract
Clinical data suggest that many malignant cancers are associated with hypercalcemia. Hypercalcemia can facilitate the proliferation and metastasis of gastric and colon tumors, and has been considered a hallmark of end-stage disease. However, it has also been reported that dietary calcium or vitamin D supplementation could reduce the risk of many types of cancers. In particular, the intestines can absorb considerable amounts of calcium via Ca2+-permeable ion channels, and hypercalcemia is common in patients with colorectal cancer. Thus, this review considers the role of calcium signaling in the context of colorectal cancer and summarizes the functions of specific regulators of cellular calcium levels in the proliferation, invasion, metastasis, cell death, and drug resistance of colorectal cancer cells. The data reveal that even a slight upregulation of intracellular Ca2+ signaling can facilitate the onset and progression of colorectal cancer, while continuous Ca2+ influx and Ca2+ overload may cause tumor cell death. This dual function of Ca2+ signaling adds nuance to the debate over the hallmarks of colorectal cancer, and may even provide new directions and strategies for clinical interventions.
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Affiliation(s)
- Wei Wang
- Jiangsu Key Laboratory for Pharmacology and Safety Evaluation of Chinese Materia Medica, School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, P.R. China
| | - Suyun Yu
- Jiangsu Key Laboratory for Pharmacology and Safety Evaluation of Chinese Materia Medica, School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, P.R. China
| | - Shuai Huang
- Jiangsu Key Laboratory for Pharmacology and Safety Evaluation of Chinese Materia Medica, School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, P.R. China
| | - Rui Deng
- Jiangsu Key Laboratory for Pharmacology and Safety Evaluation of Chinese Materia Medica, School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, P.R. China
| | - Yushi Ding
- Jiangsu Key Laboratory for Pharmacology and Safety Evaluation of Chinese Materia Medica, School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, P.R. China
| | - Yuanyuan Wu
- Jiangsu Key Laboratory for Pharmacology and Safety Evaluation of Chinese Materia Medica, School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, P.R. China.,Jiangsu Collaborative Innovation Center of Traditional Chinese Medicine Prevention and Treatment of Tumor, Nanjing University of Chinese Medicine, Nanjing, P.R. China
| | - Xiaoman Li
- Jiangsu Key Laboratory for Pharmacology and Safety Evaluation of Chinese Materia Medica, School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, P.R. China.,Jiangsu Collaborative Innovation Center of Traditional Chinese Medicine Prevention and Treatment of Tumor, Nanjing University of Chinese Medicine, Nanjing, P.R. China
| | - Aiyun Wang
- Jiangsu Key Laboratory for Pharmacology and Safety Evaluation of Chinese Materia Medica, School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, P.R. China.,Jiangsu Collaborative Innovation Center of Traditional Chinese Medicine Prevention and Treatment of Tumor, Nanjing University of Chinese Medicine, Nanjing, P.R. China
| | - Shijun Wang
- Shandong Co-Innovation Center of TCM Formula, College of Traditional Chinese Medicine, Shandong University of Traditional Chinese Medicine, Shandong, P.R. China
| | - Wenxing Chen
- Jiangsu Key Laboratory for Pharmacology and Safety Evaluation of Chinese Materia Medica, School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, P.R. China. .,Jiangsu Collaborative Innovation Center of Traditional Chinese Medicine Prevention and Treatment of Tumor, Nanjing University of Chinese Medicine, Nanjing, P.R. China
| | - Yin Lu
- Jiangsu Key Laboratory for Pharmacology and Safety Evaluation of Chinese Materia Medica, School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, P.R. China. .,Jiangsu Collaborative Innovation Center of Traditional Chinese Medicine Prevention and Treatment of Tumor, Nanjing University of Chinese Medicine, Nanjing, P.R. China
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13
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Xian Y, Zhao X, Wang C, Kang C, Ding L, Zhu W, Hang S. Phenylalanine and tryptophan stimulate gastrin and somatostatin secretion and H +-K +-ATPase activity in pigs through calcium-sensing receptor. Gen Comp Endocrinol 2018; 267:1-8. [PMID: 29782837 DOI: 10.1016/j.ygcen.2018.05.022] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/16/2018] [Revised: 04/28/2018] [Accepted: 05/17/2018] [Indexed: 11/20/2022]
Abstract
In rodents and humans, aromatic amino acids increase gut hormone secretion and H+-K+-ATPase activity by modulating calcium-sensing receptor (CaSR). However, the role of CaSR and its related signaling molecules in amino acid-induced gut hormone secretion in swine has not been investigated. Here, we examined whether a CaSR-dependent pathway modulated gastrin and somatostatin (SS) secretion and H+-K+-ATPase activity in pigs. Perfusion of pig stomach tissues in the presence of extracellular 80 mM l-phenylalanine (Phe) or 20 mM l-tryptophan (Trp) and a CaSR agonist cinacalcet triggered gastrin and SS secretion and H+-K+-ATPase activity (P < 0.05) and increased CaSR expression (P < 0.05). This effect of Phe and Trp was dependent on Ca2+ (P < 0.05) and was abolished after treatment with NPS 2143, an inhibitor of CaSR, and 2-aminoethyl diphenyl borinate, an inhibitor of CaSR downstream signaling molecule inositol 1,4,5-triphosphate receptor (IP3R). These findings indicate that Phe and Trp induce Ca2+-dependent gastrin and SS secretion and H+-K+-ATPase activity through CaSR and its downstream signaling molecule IP3R.
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Affiliation(s)
- Yihan Xian
- Laboratory of Gastrointestinal Microbiology, Nanjing Agriculture University, Nanjing, Jiangsu 210095, China
| | - Xiuying Zhao
- Laboratory of Gastrointestinal Microbiology, Nanjing Agriculture University, Nanjing, Jiangsu 210095, China
| | - Chao Wang
- Laboratory of Gastrointestinal Microbiology, Nanjing Agriculture University, Nanjing, Jiangsu 210095, China
| | - Cuicui Kang
- Laboratory of Gastrointestinal Microbiology, Nanjing Agriculture University, Nanjing, Jiangsu 210095, China
| | - Liren Ding
- Laboratory of Gastrointestinal Microbiology, Nanjing Agriculture University, Nanjing, Jiangsu 210095, China
| | - Weiyun Zhu
- Laboratory of Gastrointestinal Microbiology, Nanjing Agriculture University, Nanjing, Jiangsu 210095, China
| | - Suqin Hang
- Laboratory of Gastrointestinal Microbiology, Nanjing Agriculture University, Nanjing, Jiangsu 210095, China.
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14
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Zhao X, Xian Y, Wang C, Ding L, Meng X, Zhu W, Hang S. Calcium-sensing receptor-mediated L-tryptophan-induced secretion of cholecystokinin and glucose-dependent insulinotropic peptide in swine duodenum. J Vet Sci 2018; 19:179-187. [PMID: 29284209 PMCID: PMC5879066 DOI: 10.4142/jvs.2018.19.2.179] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/07/2017] [Revised: 12/17/2017] [Accepted: 12/26/2017] [Indexed: 12/31/2022] Open
Abstract
This study aimed to elucidate the effect of tryptophan (Trp) on gut hormone secretion as well as the roles of the calcium-sensing receptor (CaSR) and its downstream signaling pathway in gut hormone secretion by assessing swine duodenal perfusion in vitro. Swine duodenum was perfused with Krebs-Henseleit buffer as a basal solution. Various concentrations (0, 10, and 20 mM) of Trp were applied to investigate its effect on gut hormone secretion. A CaSR antagonist was used to detect the involvement of CaSR and its signal molecules. The 20 mM Trp concentration promoted the secretion of cholecystokinin (CCK) and glucose-dependent insulinotropic peptide (GIP), elevated the mRNA level of CaSR, and upregulated the protein levels of CaSR, protein kinase C (PKC), and inositol trisphosphate receptor (IP3R). However, NPS 2143, an inhibitor of CaSR, attenuated the CCK and GIP release, reduced the mRNA level of CaSR, and decreased the protein levels of CaSR, PKC, and IP3R with 20 mM Trp perfusion. The results indicate that CCK and GIP secretion can be induced by Trp in swine duodenum in vitro, and the effect is mediated by CaSR and its downstream signal molecules PKC and IP3R.
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Affiliation(s)
- Xiuying Zhao
- Laboratory of Gastrointestinal Microbiology, Nanjing Agricultural University, Nanjing 210095, China
| | - Yihan Xian
- Laboratory of Gastrointestinal Microbiology, Nanjing Agricultural University, Nanjing 210095, China
| | - Chao Wang
- Laboratory of Gastrointestinal Microbiology, Nanjing Agricultural University, Nanjing 210095, China
| | - Liren Ding
- Laboratory of Gastrointestinal Microbiology, Nanjing Agricultural University, Nanjing 210095, China
| | - Xianglong Meng
- Laboratory of Gastrointestinal Microbiology, Nanjing Agricultural University, Nanjing 210095, China
| | - Weiyun Zhu
- Laboratory of Gastrointestinal Microbiology, Nanjing Agricultural University, Nanjing 210095, China
| | - Suqin Hang
- Laboratory of Gastrointestinal Microbiology, Nanjing Agricultural University, Nanjing 210095, China
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15
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Calcium-sensing receptor in nutrient sensing: an insight into the modulation of intestinal homoeostasis. Br J Nutr 2018; 120:881-890. [DOI: 10.1017/s0007114518002088] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022]
Abstract
AbstractThe animal gut effectively prevents the entry of hazardous substances and microbes while permitting the transfer of nutrients, such as water, electrolytes, vitamins, proteins, lipids, carbohydrates, minerals and microbial metabolites, which are intimately associated with intestinal homoeostasis. The gut maintains biological functions through its nutrient-sensing receptors, including the Ca-sensing receptor (CaSR), which activates a variety of signalling pathways, depending on cellular context. CaSR coordinates food digestion and nutrient absorption, promotes cell proliferation and differentiation, regulates energy metabolism and immune response, stimulates hormone secretion, mitigates secretory diarrhoea and enhances intestinal barrier function. Thus, CaSR is crucial to the maintenance of gut homoeostasis and protection of intestinal health. In this review, we focused on the emerging roles of CaSR in the modulation of intestinal homoeostasis including related underlying mechanisms. By elucidating the relationship between CaSR and animal gut homoeostasis, effective and inexpensive methods for treating intestinal health imbalance through nutritional manipulation can be developed. This article is expected to provide experimental data of the effects of CaSR on animal or human health.
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16
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Tang L, Jiang L, McIntyre ME, Petrova E, Cheng SX. Calcimimetic acts on enteric neuronal CaSR to reverse cholera toxin-induced intestinal electrolyte secretion. Sci Rep 2018; 8:7851. [PMID: 29777154 PMCID: PMC5959902 DOI: 10.1038/s41598-018-26171-4] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/20/2017] [Accepted: 05/08/2018] [Indexed: 01/19/2023] Open
Abstract
Treatment of acute secretory diarrheal illnesses remains a global challenge. Enterotoxins produce secretion through direct epithelial action and indirectly by activating enteric nervous system (ENS). Using a microperfused colonic crypt technique, we have previously shown that R568, a calcimimetic that activates the calcium-sensing receptor (CaSR), can act on intestinal epithelium and reverse cholera toxin-induced fluid secretion. In the present study, using the Ussing chamber technique in conjunction with a tissue-specific knockout approach, we show that the effects of cholera toxin and CaSR agonists on electrolyte secretion by the intestine can also be attributed to opposing actions of the toxin and CaSR on the activity of the ENS. Our results suggest that targeting intestinal CaSR might represent a previously undescribed new approach for treating secretory diarrheal diseases and other conditions with ENS over-activation.
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Affiliation(s)
- Lieqi Tang
- Department of Pediatrics, University of Florida, Gainesville, FL, 32610, USA
| | - Lingli Jiang
- Department of Pediatrics, University of Florida, Gainesville, FL, 32610, USA
| | - Megan E McIntyre
- Department of Pediatrics, University of Florida, Gainesville, FL, 32610, USA
| | - Ekaterina Petrova
- Department of Pediatrics, University of Florida, Gainesville, FL, 32610, USA
| | - Sam X Cheng
- Department of Pediatrics, University of Florida, Gainesville, FL, 32610, USA. .,Department of Pediatrics, Division of Gastroenterology, Hepatology and Nutrition, University of Florida, Gainesville, FL, 32610, USA.
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17
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Gregório SF, Fuentes J. Regulation of Bicarbonate Secretion in Marine Fish Intestine by the Calcium-Sensing Receptor. Int J Mol Sci 2018; 19:E1072. [PMID: 29617283 PMCID: PMC5979614 DOI: 10.3390/ijms19041072] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/21/2017] [Revised: 03/26/2018] [Accepted: 04/01/2018] [Indexed: 12/29/2022] Open
Abstract
In marine fish, high epithelial intestinal HCO₃− secretion generates luminal carbonate precipitates of divalent cations that play a key role in water and ion homeostasis. The present study was designed to expose the putative role for calcium and the calcium-sensing receptor (CaSR) in the regulation of HCO₃− secretion in the intestine of the sea bream (Sparus aurata L.). Effects on the expression of the CaSR in the intestine were evaluated by qPCR and an increase was observed in the anterior intestine in fed fish compared with unfed fish and with different regions of intestine. CaSR expression reflected intestinal fluid calcium concentration. In addition, anterior intestine tissue was mounted in Ussing chambers to test the putative regulation of HCO₃− secretion in vitro using the anterior intestine. HCO₃− secretion was sensitive to varying calcium levels in luminal saline and to calcimimetic compounds known to activate/block the CaSR i.e., R 568 and NPS-2143. Subsequent experiments were performed in intestinal sacs to measure water absorption and the sensitivity of water absorption to varying luminal levels of calcium and calcimimetics were exposed as well. It appears, that CaSR mediates HCO₃− secretion and water absorption in marine fish as shown by responsiveness to calcium levels and calcimimetic compounds.
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Affiliation(s)
- Sílvia F Gregório
- Centre of Marine Sciences (CCMar), Universidade do Algarve, Campus de Gambelas, 8005-139 Faro, Portugal.
| | - Juan Fuentes
- Centre of Marine Sciences (CCMar), Universidade do Algarve, Campus de Gambelas, 8005-139 Faro, Portugal.
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18
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Rasmussen AQ, Jørgensen NR, Schwarz P. Identification and Functional Characterization of a Novel Mutation in the Human Calcium-Sensing Receptor That Co-Segregates With Autosomal-Dominant Hypocalcemia. Front Endocrinol (Lausanne) 2018; 9:200. [PMID: 29743878 PMCID: PMC5930847 DOI: 10.3389/fendo.2018.00200] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/14/2017] [Accepted: 04/10/2018] [Indexed: 12/31/2022] Open
Abstract
The human calcium-sensing receptor (CASR) is the key controller of extracellular Cao2+ homeostasis, and different mutations in the CASR gene have been linked to different calcium diseases, such as familial hypocalciuric hypercalcemia, severe hyperparathyroidism, autosomal-dominant hypocalcemia (ADH), and Bartter's syndrome type V. In this study, two generations of a family with biochemically and clinically confirmed ADH who suffered severe muscle pain, arthralgia, tetany, abdominal pain, and fatigue were evaluated for mutations in the CASR gene. The study comprises genotyping of all family members, functional characterization of a potential mutant receptor by in vitro analysis related to the wild-type receptor to reveal an association between the genotype and phenotype in the affected family members. The in vitro analysis of functional characteristics includes measurements of inositol trisphosphate accumulation, Ca2+ mobilization in response to [Ca2+]o-stimulation and receptor expression. The results reveal a significant leftward shift of inositol trisphosphate accumulation as a result of the "gain-of-function" mutant receptor and surprisingly a normalization of the response in (Ca2+)i release in the downstream pathway and additionally the maximal response of (Ca2+)i release was significantly decreased compared to the wild type. However, no gross differences were seen in D126V and the D126V/WT CASR dimeric >250 kDa band expression compared to the WT receptor, however, the D126V and D126V/WT CASR immature ~140 kDa species appear to have reduced expression compared to the WT receptor. In conclusion, in this study, a family with a clinical diagnosis of ADH in two generations was evaluated to identify a mutation in the CASR gene and reveal an association between genotype and phenotype in the affected family members. The clinical condition was caused by a novel, activating, missense mutation (D126V) in the CASR gene and the in vitro functional characteristics of the mutation co-segregated with their individual phenotype.
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Affiliation(s)
- Anne Qvist Rasmussen
- Research Centre of Ageing and Osteoporosis, Department of Endocrinology, Rigshospitalet, Copenhagen, Denmark
- *Correspondence: Anne Qvist Rasmussen,
| | - Niklas Rye Jørgensen
- Department of Clinical Biochemistry, Rigshospitalet, Copenhagen, Denmark
- Institute of Clinical Research, University of Southern, Odense, Denmark
| | - Peter Schwarz
- Research Centre of Ageing and Osteoporosis, Department of Endocrinology, Rigshospitalet, Copenhagen, Denmark
- Faculty of Health Sciences, Copenhagen University, Copenhagen, Denmark
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19
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Sánchez M, Suárez L, Andrés MT, Flórez BH, Bordallo J, Riestra S, Cantabrana B. Modulatory effect of intestinal polyamines and trace amines on the spontaneous phasic contractions of the isolated ileum and colon rings of mice. Food Nutr Res 2017; 61:1321948. [PMID: 28659731 PMCID: PMC5475348 DOI: 10.1080/16546628.2017.1321948] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/22/2016] [Accepted: 03/20/2017] [Indexed: 11/08/2022] Open
Abstract
Background: Gastrointestinal motility modulatory factors include substances of the intestinal content, such as polyamines and trace amines (TAs), the focus of this study. Methods: The amines of food, intestinal content and from faecal bacteria of Swiss mice were determined by HPLC and functionally characterised in isolated distal ileum and medial colon rings. Results: Mouse food and intestinal content contain polyamines (spermidine>putrescine>spermine) and TAs (isoamylamine>cadaverine). Intestinal bacteria mainly produce putrescine and cadaverine. The amines inhibited the spontaneous motility of the ileum (0.1-3 mM) and colon rings (0.01-3 mM, with lower IC50), with: spermine~isoamylamine~spermidine. Spermine inhibition was tetrodotoxin (TTX)-insensitive, while isoamylamine was TTX-sensitive, suggesting neural control. Mainly in the ileum, isoamylamine (3 mM) elicited acute effects modified by TTX, atropine and propranolol, and suppressed by spermine (3 mM), not being localized at the smooth muscle level. The amines assayed (3 mM), except putrescine and cadaverine in the ileum and isoamylamine in the colon, antagonised acetylcholine (ACh, 0.1 mM)-elicited phasic contractions. Isoamylamine and spermine in colon relaxed KCl (100 mM)-elicited tonic contractions, suggesting an effect on smooth muscle, but did not justify the suppression of motility caused by spermine and isoamylamine. Conclusions: Polyamines and TAs of the intestinal content might act on chemosensors and modulate intestinal peristalsis.
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Affiliation(s)
- Manuel Sánchez
- Farmacología, Departamento de Medicina, Universidad de Oviedo, Oviedo, Spain.,Instituto Universitario de Oncología del Principado de Asturias (IUOPA), Oviedo, Spain
| | - Lorena Suárez
- Instituto Universitario de Oncología del Principado de Asturias (IUOPA), Oviedo, Spain
| | - María Teresa Andrés
- Laboratorio de Microbiología Oral, Escuela de Estomatología, Universidad de Oviedo, Oviedo, Spain
| | - Blanca Henar Flórez
- Farmacología, Departamento de Medicina, Universidad de Oviedo, Oviedo, Spain
| | - Javier Bordallo
- Farmacología, Departamento de Medicina, Universidad de Oviedo, Oviedo, Spain.,Instituto Universitario de Oncología del Principado de Asturias (IUOPA), Oviedo, Spain
| | - Sabino Riestra
- Servicio de Aparato Digestivo, Unidad de Enfermedad Inflamatoria Intestinal, Hospital Universitario Central de Asturias (HUCA), Oviedo, Spain
| | - Begoña Cantabrana
- Farmacología, Departamento de Medicina, Universidad de Oviedo, Oviedo, Spain.,Instituto Universitario de Oncología del Principado de Asturias (IUOPA), Oviedo, Spain
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20
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Bumetanide increases Cl--dependent short-circuit current in late distal colon: Evidence for the presence of active electrogenic Cl- absorption. PLoS One 2017; 12:e0171045. [PMID: 28152000 PMCID: PMC5289505 DOI: 10.1371/journal.pone.0171045] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/28/2016] [Accepted: 01/13/2017] [Indexed: 12/17/2022] Open
Abstract
Mammalian colonic epithelia consist of cells that are capable of both absorbing and secreting Cl-. The present studies employing Ussing chamber technique identified two opposing short-circuit current (Isc) responses to basolateral bumetanide in rat distal colon. Apart from the transepithelial Cl--secretory Isc in early distal colon that was inhibited by bumetanide, bumetanide also stimulated Isc in late distal colon that had not previously been identified. Since bumetanide inhibits basolateral Na+-K+-2Cl- cotransporter (NKCC) in crypt cells and basolateral K+-Cl- cotransporter (KCC) in surface epithelium, we proposed this stimulatory Isc could represent a KCC-mediated Cl- absorptive current. In support of this hypothesis, ion substitution experiments established Cl- dependency of this absorptive Isc and transport inhibitor studies demonstrated the involvement of an apical Cl- conductance. Current distribution and RNA sequencing analyses revealed that this Cl- absorptive Isc is closely associated with epithelial Na+ channel (ENaC) but is not dependent on ENaC activity. Thus, inhibition of ENaC by 10 μM amiloride or benzamil neither altered the direction nor its activity. Physiological studies suggested that this Cl- absorptive Isc senses dietary Cl- content; thus when dietary Cl- was low, Cl- absorptive Isc was up-regulated. In contrast, when dietary Cl- was increased, Cl- absorptive Isc was down-regulated. We conclude that an active Cl- extrusion mechanism exists in ENaC-expressing late distal colon and likely operates in parallel with ENaC to facilitate NaCl absorption.
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21
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Mahmoudi T, Karimi K, Arkani M, Farahani H, Nobakht H, Dabiri R, Asadi A, Zali MR. Parathyroid hormone gene rs6256 and calcium sensing receptor gene rs1801725 variants are not associated with susceptibility to colorectal cancer in Iran. Asian Pac J Cancer Prev 2017; 15:6035-9. [PMID: 25124570 DOI: 10.7314/apjcp.2014.15.15.6035] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/10/2022] Open
Abstract
BACKGROUND Substantial evidence from epidemiological studies has suggested that increased levels of calcium may play a protective role against colorectal cancer (CRC). Given the vital role of calcium sensing receptor (CaSR) and parathyroid hormone (PTH) in the maintenance of calcium homeostasis, we explored whether the rs1801725 (A986S) variant located in exon 7 of the CaSR gene and the rs6256 variant located in exon 3 of PTH gene might be associated with CRC risk. MATERIALS AND METHODS In this study 860 subjects including 350 cases with CRC and 510 controls were enrolled and genotyped using PCR-RFLP methods. RESULTS We observed no significant difference in genotype or allele frequencies between the cases with CRC and controls for both CaSR and PTH genes either before or after adjustment for confounding factors including age, BMI, sex, smoking status, and family history of CRC. Furthermore, no evidence for effect modification of any association of rs1801725 and rs6256 variants and CRC by BMI, sex, or tumor site was observed. In addition, there was no significant difference in genotype and allele frequencies between the normal weight (BMI<25 kg/m2) cases and overweight/ obese (BMI≥25 kg/m2) cases for the two SNPs. CONCLUSIONS These data indicated that the CaSR gene A986S variant is not a genetic contributor to CRC risk in the Iranian population. Furthermore, our results suggest for the first time that PTH gene variant does not affect CRC risk. Nonetheless, further studies with larger sample size are needed to validate these findings.
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Affiliation(s)
- Touraj Mahmoudi
- Gastroenterology and Liver Diseases Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran E-mail :
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22
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Dabiri R, Mahmoudi T, Farahani H, Nobakht H, Zali MR. Alanine to Serine Variant at Position 986 of Calcium Sensing Receptor and Colorectal Cancer Risk. IRANIAN JOURNAL OF CANCER PREVENTION 2016; 9:e8098. [PMID: 27761213 PMCID: PMC5056015 DOI: 10.17795/ijcp-8098] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Subscribe] [Scholar Register] [Received: 01/13/2016] [Revised: 04/04/2016] [Accepted: 08/06/2016] [Indexed: 11/25/2022]
Abstract
BACKGROUND With regard to the effect of calcium against colorectal cancer (CRC) and considering the key role of calcium sensing receptor (CaSR) in calcium homeostasis, this study investigated whether CaSR gene rs1801725 or A986S variant was associated with susceptibility to CRC risk. METHODS This study was conducted as a case-control study and 303 cases with CRC and 354 controls were enrolled. All 657 subjects were genotyped for CaSR gene A986S variant using PCR-RFLP method. RESULTS No significant difference was observed for the A986S variant of CaSR gene in either genotype or allele frequencies between the cases and the controls and this lack of difference remained non-significant even after adjustment for age, BMI, sex, smoking status, and family history of CRC. No evidence for the effect modification of the association A986S variant and CRC by BMI, sex, or tumor site was also observed. Furthermore, the risk of obesity in relation to the A986S variant in the controls and the cases was separately analyzed and we observed no significant difference between normal weight (BMI < 25kg/m2) and overweight/obese (BMI ≥ 25kg/m2) subjects. CONCLUSIONS Our findings do not support a role for effect of the CaSR gene A986S variant on CRC risk; nevertheless, this finding requires confirmation and the role of the gene variant in carcinogenesis needs to be further investigated.
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Affiliation(s)
- Reza Dabiri
- Internal Medicine Department, Semnan University of Medical Sciences, Semnan, IR Iran
| | - Touraj Mahmoudi
- Gastroenterology and Liver Diseases Research Center, Research Institute for Gastroenterology and Liver Diseases, Shahid Beheshti University of Medical Sciences, Tehran, IR Iran
| | - Hamid Farahani
- Department of Physiology, School of Medicine, Qom University of Medical Sciences, Qom, IR Iran
| | - Hossein Nobakht
- Internal Medicine Department, Semnan University of Medical Sciences, Semnan, IR Iran
| | - Mohammad Reza Zali
- Gastroenterology and Liver Diseases Research Center, Research Institute for Gastroenterology and Liver Diseases, Shahid Beheshti University of Medical Sciences, Tehran, IR Iran
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23
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Uranga JA, López-Miranda V, Lombó F, Abalo R. Food, nutrients and nutraceuticals affecting the course of inflammatory bowel disease. Pharmacol Rep 2016; 68:816-26. [PMID: 27267792 DOI: 10.1016/j.pharep.2016.05.002] [Citation(s) in RCA: 94] [Impact Index Per Article: 10.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/10/2016] [Revised: 05/07/2016] [Accepted: 05/09/2016] [Indexed: 12/20/2022]
Abstract
Inflammatory bowel diseases (ulcerative colitis; Crohn's disease) are debilitating relapsing inflammatory disorders affecting the gastrointestinal tract, with deleterious effect on quality of life, and increasing incidence and prevalence. Mucosal inflammation, due to altered microbiota, increased intestinal permeability and immune system dysfunction underlies the symptoms and may be caused in susceptible individuals by different factors (or a combination of them), including dietary habits and components. In this review we describe the influence of the Western diet, obesity, and different nutraceuticals/functional foods (bioactive peptides, phytochemicals, omega 3-polyunsaturated fatty acids, vitamin D, probiotics and prebiotics) on the course of IBD, and provide some hints that could be useful for nutritional guidance. Hopefully, research will soon offer enough reliable data to slow down the spread of the disease and to make diet a cornerstone in IBD therapy.
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Affiliation(s)
- José Antonio Uranga
- Área de Histología y Anatomía Patológica, Depto. de Ciencias Básicas de la Salud, Facultad de Ciencias de la Salud, Universidad Rey Juan Carlos (URJC), Madrid, Spain; Unidad Asociada I+D+i al Instituto de Investigación en Ciencias de la Alimentación (CIAL) del Consejo Superior de Investigaciones Científicas (CSIC), Madrid, Spain; Grupo de Excelencia Investigadora URJC-Banco de Santander-Grupo Multidisciplinar de Investigación y Tratamiento del Dolor (i+DOL). Facultad de Ciencias de la Salud, Universidad Rey Juan Carlos (URJC), Madrid, Spain
| | - Visitación López-Miranda
- Unidad Asociada I+D+i al Instituto de Investigación en Ciencias de la Alimentación (CIAL) del Consejo Superior de Investigaciones Científicas (CSIC), Madrid, Spain; Grupo de Excelencia Investigadora URJC-Banco de Santander-Grupo Multidisciplinar de Investigación y Tratamiento del Dolor (i+DOL). Facultad de Ciencias de la Salud, Universidad Rey Juan Carlos (URJC), Madrid, Spain; Área de Farmacología y Nutrición, Depto. de Ciencias Básicas de la Salud, Facultad de Ciencias de la Salud, URJC, Madrid, Spain; Unidad Asociada I+D+i al Instituto de Química Médica (IQM) del CSIC, Madrid, Spain
| | - Felipe Lombó
- Grupo de Investigación "Biotecnología de Nutracéuticos y Compuestos Bioactivos-BIONUC", Instituto Universitario de Oncología del Principado de Asturias, Universidad de Oviedo, Oviedo, Spain
| | - Raquel Abalo
- Unidad Asociada I+D+i al Instituto de Investigación en Ciencias de la Alimentación (CIAL) del Consejo Superior de Investigaciones Científicas (CSIC), Madrid, Spain; Grupo de Excelencia Investigadora URJC-Banco de Santander-Grupo Multidisciplinar de Investigación y Tratamiento del Dolor (i+DOL). Facultad de Ciencias de la Salud, Universidad Rey Juan Carlos (URJC), Madrid, Spain; Área de Farmacología y Nutrición, Depto. de Ciencias Básicas de la Salud, Facultad de Ciencias de la Salud, URJC, Madrid, Spain; Unidad Asociada I+D+i al Instituto de Química Médica (IQM) del CSIC, Madrid, Spain.
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Romero P, Schmitteckert S, Wouters MM, Houghton LA, Czogalla B, Sayuk GS, Boeckxstaens GE, Guenther P, Holland-Cunz S, Niesler B. No association between the common calcium-sensing receptor polymorphism rs1801725 and irritable bowel syndrome. BMC MEDICAL GENETICS 2015; 16:110. [PMID: 26654249 PMCID: PMC4676826 DOI: 10.1186/s12881-015-0256-0] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Subscribe] [Scholar Register] [Received: 05/08/2015] [Accepted: 11/27/2015] [Indexed: 12/14/2022]
Abstract
BACKGROUND The calcium-sensing receptor (CaSR) is a calcium (Ca(2+)) sensitive G protein-coupled receptor implicated in various biological processes. In particular, it regulates Ca(2+)/Mg(2+)- homeostasis and senses interstitial Ca(2+) levels and thereby controls downstream signalling cascades. Due to its expression in the gut epithelium, the enteric nervous system and smooth muscles and its key function in regulation and coordination of muscular contraction and secretion, it represents an excellent candidate gene to be investigated in the pathophysiology of irritable bowel syndrome (IBS). Disturbed CaSR structure and function may impact gastrointestinal regulation of muscular contraction, neuronal excitation and secretion and consequently contribute to symptoms seen in IBS, such as disordered defecation as well as disturbed gut motility and visceral sensitivity. METHODS We have therefore genotyped the functional CASR SNP rs1801725 in three case control samples from the UK, Belgium and the USA. RESULTS Genotype frequencies showed no association in the three genotyped case-control samples, neither with IBS nor with IBS subtypes. CONCLUSIONS Although we could not associate the SNP to any of the established bowel symptom based IBS subtypes we cannot rule out association to altered Ca(2+) levels and disturbed secretion and gut motility which were unfortunately not assessed in the patients genotyped. This underlines the necessity of a more detailed phenotyping of IBS patients and control individuals in future studies.
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Affiliation(s)
- Philipp Romero
- Department of Surgery, Division of Paediatric Surgery, University of Heidelberg, Heidelberg, Germany.
| | - Stefanie Schmitteckert
- Department of Human Molecular Genetics, Institute of Human Genetics, University of Heidelberg, Im Neuenheimer Feld 366, Heidelberg, 69120, Germany.
| | | | - Lesley A Houghton
- University of Manchester, Manchester, UK & Mayo Clinic, Jacksonville, USA.
| | - Bastian Czogalla
- Department of Human Molecular Genetics, Institute of Human Genetics, University of Heidelberg, Im Neuenheimer Feld 366, Heidelberg, 69120, Germany.
| | | | | | - Patrick Guenther
- Department of Surgery, Division of Paediatric Surgery, University of Heidelberg, Heidelberg, Germany.
| | | | - Beate Niesler
- Department of Human Molecular Genetics, Institute of Human Genetics, University of Heidelberg, Im Neuenheimer Feld 366, Heidelberg, 69120, Germany.
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Mine Y, Zhang H. Calcium-sensing receptor (CaSR)-mediated anti-inflammatory effects of L-amino acids in intestinal epithelial cells. JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY 2015; 63:9987-9995. [PMID: 26551350 DOI: 10.1021/acs.jafc.5b03749] [Citation(s) in RCA: 40] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/05/2023]
Abstract
Calcium-sensing receptor (CaSR) plays an essential role in sensing nutrients and monitoring ion balance in the human gut. However, no discovery of CaSR-mediated anti-inflammatory effect of l-amino acids (l-AAs) on the gut system has been reported. The aim of this study is to screen and identify the anti-inflammatory activity of various l-AAs in intestinal epithelial cells (IECs) and stepwise illustrate a possible molecular mechanism for anti-inflammation. We used Caco-2 and HT-29 cell lines to evaluate the anti-inflammatory activity of l-AAs and revealed that l-tryptophan (l-Trp) and l-valine (l-Val) have strong anti-inflammatory activity consistent in both cell lines. l-Trp treatment (5 mM) reduced TNF-α-induced IL-8 secretion from HT-29 or Caco-2 cells to about 50 or 40%, respectively. l-Trp also significantly inhibited the expression of phosphorylation of JNK or IκBα to around 50% in HT-29 cells. However, the above inhibitory effects of l-Trp on inflammatory responses in TNF-α-induced HT-29 cells were abrogated by NPS-2143. The result of CaSR antagonist NPS-2143 pretreatment study suggests l-Trp exerts anti-inflammatory effects on IECs through CaSR activation. The involvement of β-arrestin2 was then found to block tumor necrosis factor (TNF)-α-induced signaling pathways after CaSR activated by l-Trp. These results validate a novel mechanism underlying CaSR agonistic l-AAs exerting anti-inflammatory effects on human intestinal epithelia.
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Affiliation(s)
- Yoshinori Mine
- Department of Food Science, University of Guelph , Guelph, Ontario N1G 2W1, Canada
| | - Hua Zhang
- Department of Food Science, University of Guelph , Guelph, Ontario N1G 2W1, Canada
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26
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Pipino C, Di Tomo P, Mandatori D, Cianci E, Lanuti P, Cutrona MB, Penolazzi L, Pierdomenico L, Lambertini E, Antonucci I, Sirolli V, Bonomini M, Romano M, Piva R, Marchisio M, Pandolfi A. Calcium sensing receptor activation by calcimimetic R-568 in human amniotic fluid mesenchymal stem cells: correlation with osteogenic differentiation. Stem Cells Dev 2015; 23:2959-71. [PMID: 25036254 DOI: 10.1089/scd.2013.0627] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
Human amniotic fluid mesenchymal stem cells (hAFMSCs) are promising for therapeutic applications in bone damage. Calcium sensing receptor (CaSR), a G protein-coupled receptor, plays a physiological role in the regulation of bone metabolism. Thus, the bone CaSR could be targeted by calcimimetic agonists, which may be potentially helpful in treating bone diseases. The aim of our study was to characterize CaSR expression in hAFMSCs and to assess the activity of calcimimetic R-568 during in vitro osteogenesis. Using western blotting, immunofluorescence, and flow cytometry, we consistently observed constitutive CaSR in osteo-differentiating hAFMSCs. Notably, both R-568 and calcium significantly enhanced hAFMSC osteogenic differentiation after exposure to osteogenic medium. To provide further evidence of the involvement of CaSR in osteogenesis, we correlated its expression with that of established osteogenic markers, that is, alkaline phosphatase (ALP), runt-related transcription factor 2 (Runx2), and osteopontin (OPN), and novel, not yet completely defined regulators of osteogenesis. Among these are β-catenin and Slug, which are mediators of Wnt signaling, and nuclear factor of activated T cells c1 (NFATc1), which plays a critical role in calcium/calcineurin signaling. Taken together, our results demonstrate that CaSR is expressed in hAFMSCs, positively correlates with osteogenic markers, and is activated by R-568. Notably, downregulation of CaSR by RNA interference supports the conclusion that CaSR activation plays a central role in hAFMSC osteogenesis. Thus, this study provides significant information on the mechanisms of hAFMSC osteogenesis, which could provide additional molecular basis for the use of calcimimetics in bone regenerative medicine.
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Affiliation(s)
- Caterina Pipino
- 1 Department of Experimental and Clinical Sciences, School of Medicine and Health Sciences, "G. d'Annunzio" University Chieti-Pescara , Chieti, Italy
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Zhang H, Kovacs-Nolan J, Kodera T, Eto Y, Mine Y. γ-Glutamyl cysteine and γ-glutamyl valine inhibit TNF-α signaling in intestinal epithelial cells and reduce inflammation in a mouse model of colitis via allosteric activation of the calcium-sensing receptor. Biochim Biophys Acta Mol Basis Dis 2015; 1852:792-804. [PMID: 25558818 DOI: 10.1016/j.bbadis.2014.12.023] [Citation(s) in RCA: 116] [Impact Index Per Article: 11.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/14/2014] [Revised: 12/16/2014] [Accepted: 12/27/2014] [Indexed: 10/24/2022]
Abstract
BACKGROUND The extracellular calcium-sensing receptor (CaSR) is distributed throughout the gastrointestinal tract, and its activation has been shown to promote intestinal homeostasis, suggesting that CaSR may be a promising target for novel therapies to prevent chronic intestinal inflammation such as inflammatory bowel disease (IBD). The γ-glutamyl dipeptides γ-glutamyl cysteine (γ-EC) and γ-glutamyl valine (γ-EV) are dietary flavor enhancing compounds, and have been shown to activate CaSR via allosteric ligand binding. The aim of this study was to examine the anti-inflammatory effects of γ-EC and γ-EV in vitro in intestinal epithelial cells and in a mouse model of intestinal inflammation. RESULTS In vitro, treatment of Caco-2 cells with γ-EC and γ-EV resulted in the CaSR-mediated reduction of TNF-α-stimulated pro-inflammatory cytokines and chemokines including IL-8, IL-6, and IL-1β, and inhibited phosphorylation of JNK and IκBα, while increasing expression of IL-10. In vivo, using a mouse model of dextran sodium sulfate (DSS)-induced colitis, γ-EC and γ-EV treatment ameliorated DSS-induced clinical signs, weight loss, colon shortening and histological damage. Moreover, γ-EC and γ-EV reduced the expression of TNF-α, IL-6, INF-γ, IL-1β, and IL-17, and increased the expression of IL-10 in the colon, in a CaSR-dependent manner. The CaSR-mediated anti-inflammatory effects of γ-EC were abrogated in β-arrestin2 knock-down Caco-2 cells, and involvement of β-arrestin2 was found to inhibit TNF-α-dependent signaling via cross-talk with the TNF-α receptor (TNFR). CONCLUSIONS Thus CaSR activation by γ-EC and γ-EV can aid in maintaining intestinal homeostasis and reducing inflammation in chronic inflammatory conditions such as IBD.
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Affiliation(s)
- Hua Zhang
- Department of Food Science, University of Guelph, Guelph, Ontario N1G 2W1, Canada
| | | | - Tomohiro Kodera
- Ajinomoto Co. Ltd., 1-1 Suzuki-cho, Kawasaki-ku, Kawasaki, Kanagawa 210-8681, Japan
| | - Yuzuru Eto
- Ajinomoto Co. Ltd., 1-1 Suzuki-cho, Kawasaki-ku, Kawasaki, Kanagawa 210-8681, Japan
| | - Yoshinori Mine
- Department of Food Science, University of Guelph, Guelph, Ontario N1G 2W1, Canada.
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28
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Zhang H, Hu CAA, Kovacs-Nolan J, Mine Y. Bioactive dietary peptides and amino acids in inflammatory bowel disease. Amino Acids 2014; 47:2127-41. [DOI: 10.1007/s00726-014-1886-9] [Citation(s) in RCA: 53] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/20/2014] [Accepted: 11/27/2014] [Indexed: 12/21/2022]
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Cheng SX, Lightfoot YL, Yang T, Zadeh M, Tang L, Sahay B, Wang GP, Owen JL, Mohamadzadeh M. Epithelial CaSR deficiency alters intestinal integrity and promotes proinflammatory immune responses. FEBS Lett 2014; 588:4158-66. [PMID: 24842610 PMCID: PMC4234694 DOI: 10.1016/j.febslet.2014.05.007] [Citation(s) in RCA: 63] [Impact Index Per Article: 5.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/31/2014] [Revised: 04/30/2014] [Accepted: 05/02/2014] [Indexed: 12/22/2022]
Abstract
The intestinal epithelium is equipped with sensing receptor mechanisms that interact with luminal microorganisms and nutrients to regulate barrier function and gut immune responses, thereby maintaining intestinal homeostasis. Herein, we clarify the role of the extracellular calcium-sensing receptor (CaSR) using intestinal epithelium-specific Casr(-/-) mice. Epithelial CaSR deficiency diminished intestinal barrier function, altered microbiota composition, and skewed immune responses towards proinflammatory. Consequently, Casr(-/-) mice were significantly more prone to chemically induced intestinal inflammation resulting in colitis. Accordingly, CaSR represents a potential therapeutic target for autoinflammatory disorders, including inflammatory bowel diseases.
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Affiliation(s)
- Sam X Cheng
- Division of Gastroenterology, Department of Pediatrics, University of Florida, Gainesville, FL 32607, USA
| | - Yaíma L Lightfoot
- Department of Infectious Diseases and Pathology, University of Florida, Gainesville, FL 32608, USA; Division of Gastroenterology, Hepatology & Nutrition, Department of Medicine, University of Florida, Gainesville, FL 32610, USA
| | - Tao Yang
- Department of Infectious Diseases and Pathology, University of Florida, Gainesville, FL 32608, USA; Division of Gastroenterology, Hepatology & Nutrition, Department of Medicine, University of Florida, Gainesville, FL 32610, USA
| | - Mojgan Zadeh
- Department of Infectious Diseases and Pathology, University of Florida, Gainesville, FL 32608, USA; Division of Gastroenterology, Hepatology & Nutrition, Department of Medicine, University of Florida, Gainesville, FL 32610, USA
| | - Lieqi Tang
- Division of Gastroenterology, Department of Pediatrics, University of Florida, Gainesville, FL 32607, USA
| | - Bikash Sahay
- Department of Infectious Diseases and Pathology, University of Florida, Gainesville, FL 32608, USA; Division of Gastroenterology, Hepatology & Nutrition, Department of Medicine, University of Florida, Gainesville, FL 32610, USA
| | - Gary P Wang
- Division of Infectious Diseases and Global Medicine, Department of Medicine, University of Florida, Gainesville, FL 32610, USA
| | - Jennifer L Owen
- Department of Physiological Sciences, College of Veterinary Medicine, University of Florida, Gainesville, FL 32610, USA
| | - Mansour Mohamadzadeh
- Department of Infectious Diseases and Pathology, University of Florida, Gainesville, FL 32608, USA; Division of Gastroenterology, Hepatology & Nutrition, Department of Medicine, University of Florida, Gainesville, FL 32610, USA.
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30
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Dal Prà I, Chiarini A, Pacchiana R, Gardenal E, Chakravarthy B, Whitfield JF, Armato U. Calcium-Sensing Receptors of Human Astrocyte-Neuron Teams: Amyloid-β-Driven Mediators and Therapeutic Targets of Alzheimer's Disease. Curr Neuropharmacol 2014; 12:353-64. [PMID: 25342943 PMCID: PMC4207075 DOI: 10.2174/1570159x12666140828214701] [Citation(s) in RCA: 30] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/20/2014] [Revised: 06/19/2014] [Accepted: 06/26/2014] [Indexed: 12/24/2022] Open
Abstract
It is generally assumed that the neuropathology of sporadic (late-onset or nonfamilial) Alzheimer’s disease (AD) is driven by the overproduction and spreading of first Amyloid-βx-42 (Aβ42) and later hyperphosphorylated (hp)-Tau oligomeric “infectious seeds”. Hitherto, only neurons were held to make and spread both oligomer types; astrocytes would just remove debris. However, we have recently shown that exogenous fibrillar or soluble Aβ peptides specifically bind and activate the Ca2+-sensing receptors (CaSRs) of untransformed human cortical adult astrocytes and postnatal neurons cultured in vitro driving them to produce, accrue, and secrete surplus endogenous Aβ42. While the Aβ-exposed neurons start dying, astrocytes survive and keep oversecreting Aβ42, nitric oxide (NO), and vascular endothelial growth factor (VEGF)-A. Thus astrocytes help neurons’ demise. Moreover, we have found that a highly selective allosteric CaSR agonist (“calcimimetic”), NPS R-568, mimics the just mentioned neurotoxic actions triggered by Aβ●CaSR signaling. Contrariwise, and most important, NPS 2143, a highly selective allosteric CaSR antagonist (“calcilytic”), fully suppresses all the Aβ●CaSR signaling-driven noxious actions. Altogether our findings suggest that the progression of AD neuropathology is promoted by unceasingly repeating cycles of accruing exogenous Aβ42 oligomers interacting with the CaSRs of swelling numbers of astrocyte-neuron teams thereby recruiting them to overrelease additional Aβ42 oligomers, VEGF-A, and NO. Calcilytics would beneficially break such Aβ/CaSR-driven vicious cycles and hence halt or at least slow the otherwise unstoppable spreading of AD neuropathology
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Affiliation(s)
- I Dal Prà
- Histology & Embryology Section, Department of Life & Reproduction Sciences, University of Verona Medical School, Verona, Venetia, Italy
| | - A Chiarini
- Histology & Embryology Section, Department of Life & Reproduction Sciences, University of Verona Medical School, Verona, Venetia, Italy
| | - R Pacchiana
- Histology & Embryology Section, Department of Life & Reproduction Sciences, University of Verona Medical School, Verona, Venetia, Italy
| | - E Gardenal
- Histology & Embryology Section, Department of Life & Reproduction Sciences, University of Verona Medical School, Verona, Venetia, Italy
| | - B Chakravarthy
- National Research Council of Canada, Ottawa, Ontario, Canada
| | - J F Whitfield
- National Research Council of Canada, Ottawa, Ontario, Canada
| | - U Armato
- Histology & Embryology Section, Department of Life & Reproduction Sciences, University of Verona Medical School, Verona, Venetia, Italy
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Emerging roles of the extracellular calcium-sensing receptor in nutrient sensing: control of taste modulation and intestinal hormone secretion. Br J Nutr 2014; 111 Suppl 1:S16-22. [PMID: 24382107 DOI: 10.1017/s0007114513002250] [Citation(s) in RCA: 41] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/16/2022]
Abstract
The extracellular Ca-sensing receptor (CaSR) is a sensor for a number of key nutrients within the body, including Ca ions (Ca²⁺) and L-amino acids. The CaSR is expressed in a number of specialised cells within the gastrointestinal (GI) tract, and much work has been done to examine CaSR's role as a nutrient sensor in this system. This review article examines two emerging roles for the CaSR within the GI tract--as a mediator of kokumi taste modulation in taste cells and as a regulator of dietary hormone release in response to L-amino acids in the intestine.
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32
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Márquez L, Fuentes J. In vitro characterization of acid secretion in the gilthead sea bream (Sparus aurata) stomach. Comp Biochem Physiol A Mol Integr Physiol 2013; 167:52-8. [PMID: 24126049 DOI: 10.1016/j.cbpa.2013.10.016] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/14/2013] [Revised: 10/06/2013] [Accepted: 10/07/2013] [Indexed: 01/19/2023]
Abstract
The gastric acid secretion of juvenile Sparus aurata was characterized in Ussing chambers; secretion rates were determined by a pH-stat method at pH5.50 and bioelectrical parameters were measured in current-clamped tissues. The basal secretion equaled to 535±87nmol·cm(-2)·h(-1). Serosal carbachol 100μM produced an increase (ΔJH(+)) of 725±133nmol·cm(-2)·h(-1) from basal secretion, this effect being inhibited by mucosal omeprazole 100μM. Basal secretion was also sensitive to the combination of serosal forskolin (FK) 10μM+serosal isobutylmethylxanthine (IBMX) 100μM (ΔJH(+)=793±239nmol·cm(-2)·h(-1)); this effect was insensitive to mucosal omeprazole 100mM but inhibited by mucosal bafilomycin A1 100nM. The effect of carbachol proceeded within a few minutes (<10min), whereas the effect of FK+IBMX was gradual, taking 40min to reach the maximum. The addition of mucosal gadolinium (Gd(3+)) 100μM, a potent calcium-sensing receptor (CaR) agonist, stimulated the basal secretion (ΔJH(+)=340±81nmol·cm(-2)·h(-1)). The present results indicate that the acid secretion mechanism in the sea bream stomach is regulated by muscarinic and CaR-like receptors, cAMP is implicated in the signal transduction, and at least two proton pumps, a HK-ATPase and a V-ATPase contribute to acid secretion.
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Affiliation(s)
- Lorenzo Márquez
- Núcleo de Investigación en Producción Alimentaria/Escuela de Acuicultura, Facultad de Recursos Naturales, Universidad Católica de Temuco, Avda. Rudecindo Ortega 02950, PO Box 15-D, Temuco, Chile.
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Di Tomo P, Pipino C, Lanuti P, Morabito C, Pierdomenico L, Sirolli V, Bonomini M, Miscia S, Mariggiò MA, Marchisio M, Barboni B, Pandolfi A. Calcium sensing receptor expression in ovine amniotic fluid mesenchymal stem cells and the potential role of R-568 during osteogenic differentiation. PLoS One 2013; 8:e73816. [PMID: 24040082 PMCID: PMC3767786 DOI: 10.1371/journal.pone.0073816] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/07/2013] [Accepted: 07/24/2013] [Indexed: 02/07/2023] Open
Abstract
Amniotic fluid-derived stem (AFS) cells have been identified as a promising source for cell therapy applications in bone traumatic and degenerative damage. Calcium Sensing Receptor (CaSR), a G protein-coupled receptor able to bind calcium ions, plays a physiological role in regulating bone metabolism. It is expressed in different kinds of cells, as well as in some stem cells. The bone CaSR could potentially be targeted by allosteric modulators, in particular by agonists such as calcimimetic R-568, which may potentially be helpful for the treatment of bone disease. The aim of our study was first to investigate the presence of CaSR in ovine Amniotic Fluid Mesenchymal Stem Cells (oAFMSCs) and then the potential role of calcimimetics in in vitro osteogenesis. oAFMSCs were isolated, characterized and analyzed to examine the possible presence of CaSR by western blotting and flow cytometry analysis. Once we had demonstrated CaSR expression, we worked out that 1 µM R-568 was the optimal and effective concentration by cell viability test (MTT), cell number, Alkaline Phosphatase (ALP) and Alizarin Red S (ARS) assays. Interestingly, we observed that basal diffuse CaSR expression in oAFMSCs increased at the membrane when cells were treated with R-568 (1 µM), potentially resulting in activation of the receptor. This was associated with significantly increased cell mineralization (ALP and ARS staining) and augmented intracellular calcium and Inositol trisphosphate (IP3) levels, thus demonstrating a potential role for calcimimetics during osteogenic differentiation. Calhex-231, a CaSR allosteric inhibitor, totally reversed R-568 induced mineralization. Taken together, our results demonstrate for the first time that CaSR is expressed in oAFMSCs and that calcimimetic R-568, possibly through CaSR activation, can significantly improve the osteogenic process. Hence, our study may provide useful information on the mechanisms regulating osteogenesis in oAFMSCs, perhaps prompting the use of calcimimetics in bone regenerative medicine.
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Affiliation(s)
- Pamela Di Tomo
- Department of Experimental and Clinical Sciences, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- Aging Research Center, Ce.S.I., “University G. d’Annunzio” Foundation Chieti, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- StemTeCh Group Chieti, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
| | - Caterina Pipino
- Department of Experimental and Clinical Sciences, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- Aging Research Center, Ce.S.I., “University G. d’Annunzio” Foundation Chieti, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- StemTeCh Group Chieti, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
| | - Paola Lanuti
- Aging Research Center, Ce.S.I., “University G. d’Annunzio” Foundation Chieti, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- StemTeCh Group Chieti, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- Department of Medicine and Aging Science, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
| | - Caterina Morabito
- Aging Research Center, Ce.S.I., “University G. d’Annunzio” Foundation Chieti, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- StemTeCh Group Chieti, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- Department of Neuroscience and Imaging, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
| | - Laura Pierdomenico
- Aging Research Center, Ce.S.I., “University G. d’Annunzio” Foundation Chieti, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- StemTeCh Group Chieti, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- Department of Medicine and Aging Science, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
| | - Vittorio Sirolli
- Department of Medicine and Aging Science, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
| | - Mario Bonomini
- Department of Medicine and Aging Science, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
| | - Sebastiano Miscia
- Aging Research Center, Ce.S.I., “University G. d’Annunzio” Foundation Chieti, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- StemTeCh Group Chieti, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- Department of Medicine and Aging Science, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
| | - Maria Addolorata Mariggiò
- Aging Research Center, Ce.S.I., “University G. d’Annunzio” Foundation Chieti, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- StemTeCh Group Chieti, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- Department of Neuroscience and Imaging, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
| | - Marco Marchisio
- Aging Research Center, Ce.S.I., “University G. d’Annunzio” Foundation Chieti, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- StemTeCh Group Chieti, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- Department of Medicine and Aging Science, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
| | - Barbara Barboni
- StemTeCh Group Chieti, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- Department of Comparative Biomedical Science, University of Teramo, Teramo, Italy
| | - Assunta Pandolfi
- Department of Experimental and Clinical Sciences, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- Aging Research Center, Ce.S.I., “University G. d’Annunzio” Foundation Chieti, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- StemTeCh Group Chieti, University “G. d’Annunzio” Chieti-Pescara, Chieti, Italy
- * E-mail:
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Lembrechts R, Brouns I, Schnorbusch K, Pintelon I, Kemp PJ, Timmermans JP, Riccardi D, Adriaensen D. Functional expression of the multimodal extracellular calcium-sensing receptor in pulmonary neuroendocrine cells. J Cell Sci 2013; 126:4490-501. [PMID: 23886943 DOI: 10.1242/jcs.131656] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022] Open
Abstract
The Ca(2+)-sensing receptor (CaSR) is the master regulator of whole-body extracellular free ionized [Ca(2+)]o. In addition to sensing [Ca(2+)]o, CaSR integrates inputs from a variety of different physiological stimuli. The CaSR is also expressed in many regions outside the [Ca(2+)]o homeostatic system, including the fetal lung where it plays a crucial role in lung development. Here, we show that neuroepithelial bodies (NEBs) of the postnatal mouse lung express a functional CaSR. NEBs are densely innervated groups of neuroendocrine epithelial cells in the lung representing complex sensory receptors in the airways and exhibiting stem cell characteristics. qRT-PCR performed on laser microdissected samples from GAD67-GFP mouse lung cryosections revealed exclusive expression of the CaSR in the NEB microenvironment. CaSR immunoreactivity was present at NEB cells from postnatal day 14 onwards. Confocal imaging of lung slices revealed that NEB cells responded to an increase of [Ca(2+)]o with a rise in intracellular Ca(2+) ([Ca(2+)]i); an effect mimicked by several membrane-impermeant CaSR agonists (e.g. the calcimimetic R-568) and that was blocked by the calcilytic Calhex-231. Block of TRPC channels attenuated the CaSR-dependent increases in [Ca(2+)]i, suggesting that Ca(2+) influx through TRPC channels contributes to the total [Ca(2+)]i signal evoked by the CaSR in NEBs. CaSR also regulated baseline [Ca(2+)]i in NEBs and, through paracrine signaling from Clara-like cells, coordinated intercellular communication in the NEB microenvironment. These data suggest that the NEB CaSR integrates multiple signals converging on this complex chemosensory unit, and is a key regulator of this intrapulmonary airway stem cell niche.
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Affiliation(s)
- Robrecht Lembrechts
- Laboratory of Cell Biology and Histology, Department of Veterinary Sciences, University of Antwerp, BE-2020 Antwerp, Belgium
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Peterlik M, Kállay E, Cross HS. Calcium nutrition and extracellular calcium sensing: relevance for the pathogenesis of osteoporosis, cancer and cardiovascular diseases. Nutrients 2013; 5:302-27. [PMID: 23340319 PMCID: PMC3571650 DOI: 10.3390/nu5010302] [Citation(s) in RCA: 53] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/30/2012] [Revised: 01/10/2013] [Accepted: 01/11/2013] [Indexed: 02/07/2023] Open
Abstract
Through a systematic search in Pubmed for literature, on links between calcium malnutrition and risk of chronic diseases, we found the highest degree of evidence for osteoporosis, colorectal and breast cancer, as well as for hypertension, as the only major cardiovascular risk factor. Low calcium intake apparently has some impact also on cardiovascular events and disease outcome. Calcium malnutrition can causally be related to low activity of the extracellular calcium-sensing receptor (CaSR). This member of the family of 7-TM G-protein coupled receptors allows extracellular Ca2+ to function as a "first messenger" for various intracellular signaling cascades. Evidence demonstrates that Ca2+/CaSR signaling in functional linkage with vitamin D receptor (VDR)-activated pathways (i) promotes osteoblast differentiation and formation of mineralized bone; (ii) targets downstream effectors of the canonical and non-canonical Wnt pathway to inhibit proliferation and induce differentiation of colorectal cancer cells; (iii) evokes Ca2+ influx into breast cancer cells, thereby activating pro-apoptotic intracellular signaling. Furthermore, Ca2+/CaSR signaling opens Ca2+-sensitive K+ conductance channels in vascular endothelial cells, and also participates in IP(3)-dependent regulation of cytoplasmic Ca2+, the key intermediate of cardiomyocyte functions. Consequently, impairment of Ca2+/CaSR signaling may contribute to inadequate bone formation, tumor progression, hypertension, vascular calcification and, probably, cardiovascular disease.
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Affiliation(s)
- Meinrad Peterlik
- Department of Pathophysiology, Medical University of Vienna, Waehringer Guertel 18-20, A-1090 Vienna, Austria.
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Cheng SX. Calcium-sensing receptor inhibits secretagogue-induced electrolyte secretion by intestine via the enteric nervous system. Am J Physiol Gastrointest Liver Physiol 2012; 303:G60-70. [PMID: 22517767 PMCID: PMC3404579 DOI: 10.1152/ajpgi.00425.2011] [Citation(s) in RCA: 42] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
Bacterial toxins such as cholera toxin induce diarrhea by both direct epithelial cell generation of cyclic nucleotides as well as stimulation of the enteric nervous system (ENS). Agonists of the extracellular calcium-sensing receptor (CaSR) can reduce toxin-stimulated fluid secretion in ENS-absent colonic epithelial crypts by increasing phosphodiesterase-dependent cyclic-nucleotide degradation. Here we show that the CaSR is also highly expressed in tetrodotoxin (TTX)-sensitive neurons comprising the ENS, suggesting that CaSR agonists might also function through neuronal pathways. To test this hypothesis, rat colon segments containing intact ENS were isolated and mounted on Ussing chambers. Basal and cyclic nucleotide-stimulated electrolyte secretions were monitored by measuring changes in short-circuit current (I(sc)). CaSR was activated by R-568 and its effects were compared in the presence and absence of TTX. Consistent with active regulation of anion secretion by the ENS, a significant proportion of I(sc) in the proximal and distal colon was inhibited by serosal TTX, both at basal and under cyclic AMP-stimulated conditions. In the absence of TTX, activation of CaSR with R-568 significantly reduced basal I(sc) and cyclic AMP-stimulated I(sc); it also completely reversed the cAMP-stimulated secretory responses if the drug was applied after the forskolin stimulation. Such inhibitory effects of R-568 were either absent or significantly reduced when serosal TTX was present, suggesting that this agonist exerts its antisecretory effect on the intestine by inhibiting ENS. The present results suggest a new model for regulating intestinal fluid transport in which neuronal and nonneuronal secretagogue actions are modulated by the inhibitory effects of CaSR on the ENS. The ability of a CaSR agonist to reduce secretagogue-stimulated Cl(-) secretion might provide a new therapeutic approach for secretory and other ENS-mediated diarrheal conditions.
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Affiliation(s)
- Sam X. Cheng
- 1Department of Pediatrics, School of Medicine, Yale University, New Haven, Connecticut; and ,2Department of Pediatrics, School of Medicine, University of Florida, Gainesville, Florida
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Abstract
Recent advances highlight that nutrient receptors (such as T1R1/T1R3 heterodimer, Ca sensing receptor and GPR93 for amino acids and protein, GPR40, GPR41, GPR43 and GPR120 for fatty acids, T1R2/T1R3 heterodimer for monosaccharides) are expressed in the apical face of the gut and sense nutrients in the lumen. They transduce signals for the regulation of nutrient transporter expressions in the apical face. Interestingly, they are also localised in enteroendocrine cells (EEC) and mainly exert a direct control on the secretion in the lamina propria of gastro-intestinal peptides such as cholecystokinin, glucagon-like peptide-1 and peptide YY in response to energy nutrient transit and absorption in the gut. This informs central nuclei involved in the control of feeding such as the hypothalamus and nucleus of the solitary tract of the availability of these nutrients and thus triggers adaptive responses to maintain energy homoeostasis. These nutrient receptors then have a prominent position since they manage nutrient absorption and are principally the generator of the first signal of satiation mechanisms mainly transmitted to the brain by vagal afferents. Moreover, tastants are also able to elicit gut peptides secretion via chemosensory receptors expressed in EEC. Targeting these nutrient and tastant receptors in EEC may thus be helpful to promote satiation and so to fight overfeeding and its consequences.
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Riccardi D, Kemp PJ. The Calcium-Sensing Receptor Beyond Extracellular Calcium Homeostasis: Conception, Development, Adult Physiology, and Disease. Annu Rev Physiol 2012; 74:271-97. [DOI: 10.1146/annurev-physiol-020911-153318] [Citation(s) in RCA: 111] [Impact Index Per Article: 8.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Affiliation(s)
- Daniela Riccardi
- Division of Pathophysiology and Repair, School of Biosciences, Cardiff University, Cardiff, CF10 3AX, United Kingdom; ,
| | - Paul J. Kemp
- Division of Pathophysiology and Repair, School of Biosciences, Cardiff University, Cardiff, CF10 3AX, United Kingdom; ,
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Romero P, Niesler B, Schmitz-Winnenthal H, Fitze G, Holland-Cunz S. Is there a link between the calcium sensing receptor and Hirschsprung's disease? A mutational analysis. J Pediatr Surg 2012; 47:551-5. [PMID: 22424352 DOI: 10.1016/j.jpedsurg.2011.10.010] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/12/2011] [Revised: 08/23/2011] [Accepted: 10/03/2011] [Indexed: 10/28/2022]
Abstract
INTRODUCTION Hirschsprung's disease (HD) is a congenital malformation of the hindgut characterized by the absence of enteric ganglion cells in the submucosal and myenteric plexuses. Hirschsprung's disease is routinely treated by resection of the aganglionic bowel and pull-through procedure of the proximal ganglionated bowel to the anorectal region. Occasionally, after resection of the aganglionic bowel, HD patients still experience persistent disturbances in gut motility. The etiology of HD as well as the underlying pathomechanism for postoperative disturbances in gut motility is unclear. Molecular analysis of putative candidate genes may help to clarify the pathophysiology of these conditions. In the present study, we investigated the correlation between mutations and polymorphisms in the calcium sensing receptor (CaSR) and HD patients. METHODS Mutational screening of the CaSR coding sequence was performed via polymerase chain reaction and direct sequencing of the amplified samples from 63 HD patients and 100 controls. RESULTS We identified 3 common polymorphisms (p.A986S, p.G990R, and p.Q1011E) residing in exon 7 and 1 polymorphism in intron 5 (IVS 5-88 T>C) of the CaSR gene. Overall, 16 patients (25%) and 25 controls (25%) were carriers of the p.A986S polymorphism (P = 1). The incidence of p.R990G polymorphism of patients was twice as high as in the control group (P = .17). Furthermore, we verified a 4 times higher incidence of p.Q1011E polymorphism carriers in patients compared to the control group (P = .1). CONCLUSION We found a higher incidence of R990G and Q1011E polymorphisms in HD patients compared to controls. However, there was no statistically significant association between HD and the 3 polymorphic variants in the intracellular signaling region of CaSR. It will be important to further investigate genetic variations in CaSR in more patient cohorts to better characterize the function of this gene and to establish a correlation between CaSR variants and HD.
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Affiliation(s)
- Philipp Romero
- Department of Surgery, Division of Pediatric Surgery, University of Heidelberg, Im Neuenheimer Feld 110, 69120 Heidelberg, Germany.
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40
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Steinert RE, Beglinger C. Nutrient sensing in the gut: interactions between chemosensory cells, visceral afferents and the secretion of satiation peptides. Physiol Behav 2011; 105:62-70. [DOI: 10.1016/j.physbeh.2011.02.039] [Citation(s) in RCA: 66] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/20/2010] [Revised: 02/24/2011] [Accepted: 02/25/2011] [Indexed: 01/01/2023]
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Milk Alkali and Hydrochlorothiazide: A Case Report. Case Rep Med 2011; 2011:729862. [PMID: 21738535 PMCID: PMC3114559 DOI: 10.1155/2011/729862] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/09/2010] [Revised: 02/12/2011] [Accepted: 03/28/2011] [Indexed: 11/26/2022] Open
Abstract
Hypercalcemia is a relatively common clinical problem in both outpatient and inpatient settings. Primary pathophysiology is the entry of calcium that exceeds its excretion into urine or deposition in bone into circulation. Among a wide array of causes of hypercalcemia, hyperparathyroidism and malignancy are the most common, accounting for greater than 90 percent of cases. Concordantly, there has been a resurgence of milk-alkali syndrome associated with the ingestion of large amounts of calcium and absorbable alkali, making it the third leading cause of hypercalcemia (Beall and Scofield, 1995 and Picolos et al., 2005). This paper centers on a case of over-the-counter calcium and alkali ingestion for acid reflux leading to milk alkali with concordant use of thiazide diuretic for hypertension.
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Rey O, Young SH, Jacamo R, Moyer MP, Rozengurt E. Extracellular calcium sensing receptor stimulation in human colonic epithelial cells induces intracellular calcium oscillations and proliferation inhibition. J Cell Physiol 2010; 225:73-83. [PMID: 20648625 DOI: 10.1002/jcp.22198] [Citation(s) in RCA: 57] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/08/2022]
Abstract
The extracellular Ca(2+)-sensing receptor (CaR) is increasingly implicated in the regulation of multiple cellular functions in the gastrointestinal tract, including secretion, proliferation and differentiation of intestinal epithelial cells. However, the signaling mechanisms involved remain poorly defined. Here we examined signaling pathways activated by the CaR, including Ca(2+) oscillations, in individual human colon epithelial cells. Single cell imaging of colon-derived cells expressing the CaR, including SW-480, HT-29, and NCM-460 cells, shows that stimulation of this receptor by addition of aromatic amino acids or by an elevation of the extracellular Ca(2+) concentration promoted striking intracellular Ca(2+) oscillations. The intracellular calcium oscillations in response to extracellular Ca(2+) were of sinusoidal pattern and mediated by the phospholipase C/diacylglycerol/inositol 1,4,5-trisphosphate pathway as revealed by a biosensor that detects the accumulation of diacylglycerol in the plasma membrane. The intracellular calcium oscillations in response to aromatic amino acids were of transient type, that is, Ca(2+) spikes that returned to baseline levels, and required an intact actin cytoskeleton, a functional Rho, Filamin A and the ion channel TRPC1. Further analysis showed that re-expression and stimulation of the CaR in human epithelial cells derived from normal colon and from colorectal adenocarcinoma inhibits their proliferation. This inhibition was associated with the activation of the signaling pathway that mediates the generation of sinusoidal, but not transient, intracellular Ca(2+) oscillations. Thus, these results indicate that the CaR can function in two signaling modes in human colonic epithelial cells offering a potential link between gastrointestinal responses and food/nutrients uptake and metabolism.
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Affiliation(s)
- Osvaldo Rey
- Unit of Signal Transduction and Gastrointestinal Cancer, Division of Digestive Diseases, Department of Medicine, CURE, Digestive Diseases Research Center, Molecular Biology Institute and Jonsson Comprehensive Cancer Center, David Geffen School of Medicine, University of California at Los Angeles, California 90095-1786, USA.
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Peleg S, Sellin JH, Wang Y, Freeman MR, Umar S. Suppression of aberrant transient receptor potential cation channel, subfamily V, member 6 expression in hyperproliferative colonic crypts by dietary calcium. Am J Physiol Gastrointest Liver Physiol 2010. [PMID: 20508153 DOI: 10.1152/ajp-gi.00193.2010] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/16/2023]
Abstract
Dietary calcium is believed to reduce colon cancer risk, but the mechanism by which this occurs is poorly understood. Employing the Citrobacter rodentium-induced transmissible murine colonic hyperplasia (TMCH) model, we previously showed that a high-calcium diet (hCa) significantly abrogated hyperplasia in the distal colons of NIH-Swiss mice. Here, we explored the mechanism of dietary protection by hCa by analyzing the expression of genes involved in the regulation of Ca uptake/flux in the intestinal epithelium, including the Ca-sensing receptor, vitamin D receptor, Ca binding protein, and transient receptor potential cation channels, subfamily V, members 5 and 6 (TRPV5/6). Interestingly, while TRPV6 expression increased significantly during TMCH, the expression of the other gene products was unchanged. This elevated TRPV6 expression was significantly abrogated by a hCa diet. Immunofluorescence revealed apical membrane localization of TRPV6 in the normal colon, whereas during TMCH we observed intense apical pole and cytoplasmic staining along the entire longitudinal crypt axis, including the expanded proliferating zone. The hCa diet reversed this effect. In humans, overexpression of TRPV6 was associated with early-stage colon cancer, and in colon carcinoma cells, inhibition of TRPV6 expression by small interfering RNA inhibited their proliferation and induced apoptosis. TRPV6 small interfering RNA also diminished the transcriptional activity of the calcium-dependent nuclear factors in activated T cells. Thus the aberrant overexpression of TRPV6 contributes to colonic crypt hyperplasia in mice and to colon cancer cell proliferation in humans. Therefore, it is likely that suppression of TRPV6 by a hCa diet is required for its protective effects in the colon.
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Affiliation(s)
- Sara Peleg
- The University of Texas, M. D. Anderson Cancer Center, Houston, 77030, USA.
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Peleg S, Sellin JH, Wang Y, Freeman MR, Umar S. Suppression of aberrant transient receptor potential cation channel, subfamily V, member 6 expression in hyperproliferative colonic crypts by dietary calcium. Am J Physiol Gastrointest Liver Physiol 2010; 299:G593-601. [PMID: 20508153 PMCID: PMC2950683 DOI: 10.1152/ajpgi.00193.2010] [Citation(s) in RCA: 34] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/22/2010] [Accepted: 05/26/2010] [Indexed: 01/31/2023]
Abstract
Dietary calcium is believed to reduce colon cancer risk, but the mechanism by which this occurs is poorly understood. Employing the Citrobacter rodentium-induced transmissible murine colonic hyperplasia (TMCH) model, we previously showed that a high-calcium diet (hCa) significantly abrogated hyperplasia in the distal colons of NIH-Swiss mice. Here, we explored the mechanism of dietary protection by hCa by analyzing the expression of genes involved in the regulation of Ca uptake/flux in the intestinal epithelium, including the Ca-sensing receptor, vitamin D receptor, Ca binding protein, and transient receptor potential cation channels, subfamily V, members 5 and 6 (TRPV5/6). Interestingly, while TRPV6 expression increased significantly during TMCH, the expression of the other gene products was unchanged. This elevated TRPV6 expression was significantly abrogated by a hCa diet. Immunofluorescence revealed apical membrane localization of TRPV6 in the normal colon, whereas during TMCH we observed intense apical pole and cytoplasmic staining along the entire longitudinal crypt axis, including the expanded proliferating zone. The hCa diet reversed this effect. In humans, overexpression of TRPV6 was associated with early-stage colon cancer, and in colon carcinoma cells, inhibition of TRPV6 expression by small interfering RNA inhibited their proliferation and induced apoptosis. TRPV6 small interfering RNA also diminished the transcriptional activity of the calcium-dependent nuclear factors in activated T cells. Thus the aberrant overexpression of TRPV6 contributes to colonic crypt hyperplasia in mice and to colon cancer cell proliferation in humans. Therefore, it is likely that suppression of TRPV6 by a hCa diet is required for its protective effects in the colon.
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Affiliation(s)
- Sara Peleg
- The University of Texas, M. D. Anderson Cancer Center, Houston, 77030, USA.
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Nguyen-Yamamoto L, Bolivar I, Strugnell SA, Goltzman D. Comparison of active vitamin D compounds and a calcimimetic in mineral homeostasis. J Am Soc Nephrol 2010; 21:1713-23. [PMID: 20651168 DOI: 10.1681/asn.2009050531] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022] Open
Abstract
The differential effects between cinacalcet and active vitamin D compounds on parathyroid function, mineral metabolism, and skeletal function are incompletely understood. Here, we studied cinacalcet and active vitamin D compounds in mice expressing the null mutation for Cyp27b1, which encodes 25-hydroxyvitamin D-1α-hydroxylase, thereby lacking endogenous 1,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)]. Vehicle-treated mice given high dietary calcium had hypocalcemia, hypophosphatemia, and marked secondary hyperparathyroidism. Doxercalciferol and 1,25(OH)(2)D(3) each normalized these parameters and corrected both the abnormal growth plate architecture and the diminished longitudinal bone growth observed in these mice. In contrast, cinacalcet suppressed serum parathyroid hormone (PTH) cyclically and did not correct the skeletal abnormalities and hypocalcemia persisted. Vehicle-treated mice given a "rescue diet" (high calcium and phosphorus, 20% lactose) had normal serum calcium and PTH levels; cinacalcet induced transient hypocalcemia and mild hypercalciuria. The active vitamin D compounds and cinacalcet normalized the increased osteoblast activity observed in mice with secondary hyperparathyroidism; cinacalcet, however, increased the number and activity of osteoclasts. In conclusion, cinacalcet reduces PTH in a cyclical manner, does not eliminate hypocalcemia, and does not correct abnormalities of the growth plate. Doxercalciferol and 1,25(OH)(2)D(3) reduce PTH in a sustained manner, normalize serum calcium, and improve skeletal abnormalities.
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Affiliation(s)
- Loan Nguyen-Yamamoto
- Department of Medicine, McGill University and McGill University Health Centre, Montreal, Quebec, Canada
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Sun YH, Li YQ, Feng SL, Li BX, Pan ZW, Xu CQ, Li TT, Yang BF. Calcium-sensing receptor activation contributed to apoptosis stimulates TRPC6 channel in rat neonatal ventricular myocytes. Biochem Biophys Res Commun 2010; 394:955-61. [PMID: 20307499 DOI: 10.1016/j.bbrc.2010.03.096] [Citation(s) in RCA: 36] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/02/2010] [Accepted: 03/16/2010] [Indexed: 02/07/2023]
Abstract
Capacitative calcium entry (CCE) refers to the influx of calcium through plasma membrane channels activated on depletion of endoplasmic sarcoplasmic/reticulum (ER/SR) Ca(2+) stores, which is performed mainly by the transient receptor potential (TRP) channels. TRP channels are expressed in cardiomyocytes. Calcium-sensing receptor (CaR) is also expressed in rat cardiac tissue and plays an important role in mediating cardiomyocyte apoptosis. However, there are no data regarding the link between CaR and TRP channels in rat heart. In this study, in rat neonatal myocytes, by Ca(2+) imaging, we found that the depletion of ER/SR Ca(2+) stores by thapsigargin (TG) elicited a transient rise in cytoplasmic Ca(2+) ([Ca(2+)](i)), followed by sustained increase depending on extracellular Ca(2+). But, TRP channels inhibitor (SKF96365), not L-type channels or the Na(+)/Ca(2+) exchanger inhibitors, inhibited [Ca(2+)](i) relatively high. Then, we found that the stimulation of CaR with its activator gadolinium chloride (GdCl(3)) or by an increased extracellular Ca(2+)([Ca(2+)](o)) increased the concentration of intracelluar Ca(2+), whereas, the sustained elevation of [Ca(2+)](i) was reduced in the presence of SKF96365. Similarly, the duration of [Ca(2+)](i) increase was also shortened in the absence of extracellular Ca(2+). Western blot analysis showed that GdCl(3) increased the expression of TRPC6, which was reversed by SKF96365. Additionally, SKF96365 reduced cardiomyocyte apoptosis induced by GdCl(3). Our results suggested that CCE exhibited in rat neonatal myocytes and CaR activation induced Ca(2+)-permeable cationic channels TRPCs to gate the CCE, for which TRPC6 was one of the most likely candidates. TRPC6 channel was functionally coupled with CaR to enhance the cardiomyocyte apoptosis.
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Affiliation(s)
- Yi-hua Sun
- Department of Clinical Laboratory, The Second Affiliated Hospital of Harbin Medical University, Harbin 150086, China
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Bacchetta J, Ranchin B, Brunet AS, Bouvier R, Duquesne A, Edery P, Fabien N, Peretti N. Autoimmune hypoparathyroidism in a 12-year-old girl with McKusick cartilage hair hypoplasia. Pediatr Nephrol 2009; 24:2449-53. [PMID: 19626344 DOI: 10.1007/s00467-009-1256-0] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/15/2009] [Revised: 04/28/2009] [Accepted: 05/26/2009] [Indexed: 11/27/2022]
Abstract
McKusick type metaphyseal chondrodysplasia, or cartilage hair hypoplasia (CHH), is a rare autosomal recessive osteochondrodysplasia secondary to a mutation in the RMRP gene. In addition to the metaphyseal chondrodysplasia and the short-limb dwarfism, patients may present with a multisystemic disease, associating immune deficiency with recurrent infantile or childhood infections, hematological abnormalities, and gastrointestinal dysfunction. The probability of malignancy is increased in these patients, as are disimmune manifestations. We report on a 12-year-old girl with a new mutation of the RMRP gene and a severe multisystemic CHH (hematological and pulmonary lesions, severe immune deficiency, arthritis, pancreatic insufficiency, malabsorption, chronic diarrhea) receiving parenteral nutrition who presented with acute symptomatic hypocalcemia and hypercalciuria associated with the presence of autoantibodies directed against the calcium-sensor receptor. At the same time, there was an important escalation of diarrhea. Corticosteroids led to a progressive improvement of biological signs (hypocalcemia, hypoparathyroidism). By contrast, gastrointestinal symptoms and malabsorption did not improve. To our knowledge, this is the first report of autoimmune hypoparathyroidism in CHH.
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Affiliation(s)
- Justine Bacchetta
- Service de Néphrologie et Rhumatologie Pédiatriques, Centre de Référence des Maladies Rénales Rares, Hôpital Femme Mère Enfant, Hospices Civils de Lyon, 59 Bd Pinel, 69677 Bron Cedex, France.
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Ohsu T, Amino Y, Nagasaki H, Yamanaka T, Takeshita S, Hatanaka T, Maruyama Y, Miyamura N, Eto Y. Involvement of the calcium-sensing receptor in human taste perception. J Biol Chem 2009; 285:1016-22. [PMID: 19892707 DOI: 10.1074/jbc.m109.029165] [Citation(s) in RCA: 212] [Impact Index Per Article: 13.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
By human sensory analyses, we found that various extracellular calcium-sensing receptor (CaSR) agonists enhance sweet, salty, and umami tastes, although they have no taste themselves. These characteristics are known as "kokumi taste" and often appear in traditional Japanese cuisine. Although GSH is a typical kokumi taste substance (taste enhancer), its mode of action is poorly understood. Here, we demonstrate how the kokumi taste is enhanced by the CaSR, a close relative of the class C G-protein-coupled receptors T1R1, T1R2, and T1R3 (sweet and umami receptors). We identified a large number of CaSR agonist gamma-glutamyl peptides, including GSH (gamma-Glu-Cys-Gly) and gamma-Glu-Val-Gly, and showed that these peptides elicit the kokumi taste. Further analyses revealed that some known CaSR agonists such as Ca(2+), protamine, polylysine, L-histidine, and cinacalcet (a calcium-mimetic drug) also elicit the kokumi taste and that the CaSR-specific antagonist, NPS-2143, significantly suppresses the kokumi taste. This is the first report indicating a distinct function of the CaSR in human taste perception.
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Affiliation(s)
- Takeaki Ohsu
- Pharmaceutical Research Laboratories, Ajinomoto Company, Incorporated, Kawasaki 210-8681, Japan
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Riccardi D, Finney BA, Wilkinson WJ, Kemp PJ. Novel regulatory aspects of the extracellular Ca2+-sensing receptor, CaR. Pflugers Arch 2009; 458:1007-22. [PMID: 19484257 DOI: 10.1007/s00424-009-0681-z] [Citation(s) in RCA: 28] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/23/2009] [Revised: 04/30/2009] [Accepted: 05/05/2009] [Indexed: 01/15/2023]
Abstract
The capacity to sense and adapt to changes in environmental cues is of paramount importance for every living organism. From yeast to man, cells must be able to match cellular activities to growth environment and nutrient availability. Key to this process is the development of membrane-bound systems that can detect modifications in the extracellular environment and to translate these into biological responses. Evidence gathered over the last 15 years has demonstrated that many of these cell surface "sensors" belong to the G protein-coupled receptor superfamily. Crucial to our understanding of nutrient sensing in mammalian species has been the identification of the extracellular Ca(2+)/cation-sensing receptor, CaR. CaR was the first ion-sensing molecule identified in man and genetic studies in humans have revealed the importance of the CaR in mineral ion metabolism. Latter, it has become apparent that the CaR also plays an important role outside the Ca(2+) homeostatic system, as an integrator of multiple environmental signals for the regulation of many vital cellular processes, from cell-to-cell communication to secretion and cell survival/cell death. Recently, novel aspects of receptor function reveal an unexpected role for the CaR in the regulation of growth and development in utero.
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Hao S, Zhao H, Darzynkiewicz Z, Battula S, Ferreri NR. Expression and function of NFAT5 in medullary thick ascending limb (mTAL) cells. Am J Physiol Renal Physiol 2009; 296:F1494-503. [PMID: 19369291 PMCID: PMC2692445 DOI: 10.1152/ajprenal.90436.2008] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/25/2008] [Accepted: 04/08/2009] [Indexed: 01/31/2023] Open
Abstract
The contribution of nuclear factor of activated T cells 5 (NFAT5) to the regulation of tumor necrosis factor-alpha (TNF) production in medullary thick ascending limb (mTAL) cells is unclear. RT-PCR analysis was performed on primary cultures of mouse mTAL cells and freshly isolated mTAL tubules to determine which NFAT isoforms are present in this nephron segment. Primer pairs were designed, based on published sequences for mouse NFAT1-5, to produce fragments of approximately 200 bp. Analysis of PCR products by gel electrophoresis and subsequent DNA sequencing indicated that cells and tubules contained mRNA for all five NFAT isoforms. The relative expression of NFAT isoforms was then determined using quantitative real-time RT-PCR. The data indicate that NFAT isoforms 5 >/= 1 are the predominant isoforms present in mTAL cells and tubules. Western blot analysis demonstrated constitutive expression of NFAT5 in nuclear extracts from mTAL tubules and primary culture cells; expression in mTAL cells also was detected by immunofluorescence. Expression of NFAT5 was increased in mTAL cells transiently transfected with an NFAT5 overexpression vector (pcDNA3.1-NFAT5), resulting in increased basal and calcium-sensing receptor (CaR)-mediated TNF production. Transient transfection of mTAL cells with a small hairpin RNA vector that targeted exon 8 of NFAT5 (U6-N5 ex8) significantly inhibited TNF promoter activity. Transient transfection with U6-N5 ex8 also reduced nuclear expression of NFAT5, TNF mRNA accumulation, and attenuated CaR-mediated activation of Cl(-) entry into polarized mTAL cells. Collectively, these data suggest that activation of NFAT5 is part of a TNF-dependent pathway that inhibits apical Cl(-) influx in the mTAL after activation of CaR.
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Affiliation(s)
- Shoujin Hao
- Department of Pharmacology, New York Medical College, Valhalla, NY 10595, USA
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