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Amirsasan R, Akbarzadeh M, Akbarzadeh S. Exercise and colorectal cancer: prevention and molecular mechanisms. Cancer Cell Int 2022; 22:247. [PMID: 35945569 PMCID: PMC9361674 DOI: 10.1186/s12935-022-02670-3] [Citation(s) in RCA: 7] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/03/2022] [Accepted: 08/02/2022] [Indexed: 12/16/2022] Open
Abstract
Exercise and physical activity have been shown to be strongly associated with a decreased incidence rate of various chronic diseases especially numerous human malignancies. A huge number of clinical trials and meta-analysis have demonstrated that exercise is significantly effective in lowering the risk of colorectal cancer. In addition, it is suggested as an effective therapeutic modality against this cancer type. Therefore, in this review, we will review comprehensibly the effects of exercise in preventing, treating, and alleviating the adverse effects of conventional therapeutic options in colorectal cancer. Moreover, the possible mechanisms underlying the positive effects of exercise and physical activity in colorectal cancer, including regulation of inflammation, apoptosis, growth factor axis, immunity, epigenetic, etc. will be also discussed.
Exercise is an effective post-treatment management program in colorectal cancer survivals Exercise improves muscle strength, cardiorespiratory fitness, emotional distress, physical activity, fatigue, and sleep quality in colorectal patients undergoing chemotherapy Targeting and modulating insulin-like growth factor (IGF) system, inflammation, apoptosis, immunity, epigenetic, Leptin and Ghrelin, and signaling pathways are major underlying mechanisms for preventive effects of exercise in colorectal cancer
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Affiliation(s)
- Ramin Amirsasan
- Department of Exercise Physiology, Faculty of Physical Education and Sport Sciences, University of Tabriz, Tabriz, Iran
| | - Maryam Akbarzadeh
- Department of Biology, Faculty of Natural Sciences, University of Tabriz, Tabriz, Iran
| | - Shabnam Akbarzadeh
- Department of Exercise Physiology, Faculty of Physical Education and Sport Sciences, University of Tabriz, Tabriz, Iran.
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Insulin-Like Growth Factor 1 (IGF-1) Signaling in Glucose Metabolism in Colorectal Cancer. Int J Mol Sci 2021; 22:ijms22126434. [PMID: 34208601 PMCID: PMC8234711 DOI: 10.3390/ijms22126434] [Citation(s) in RCA: 84] [Impact Index Per Article: 21.0] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/29/2021] [Revised: 06/04/2021] [Accepted: 06/14/2021] [Indexed: 02/07/2023] Open
Abstract
Colorectal cancer (CRC) is one of the most common aggressive carcinoma types worldwide, characterized by unfavorable curative effect and poor prognosis. Epidemiological data re-vealed that CRC risk is increased in patients with metabolic syndrome (MetS) and its serum components (e.g., hyperglycemia). High glycemic index diets, which chronically raise post-prandial blood glucose, may at least in part increase colon cancer risk via the insulin/insulin-like growth factor 1 (IGF-1) signaling pathway. However, the underlying mechanisms linking IGF-1 and MetS are still poorly understood. Hyperactivated glucose uptake and aerobic glycolysis (the Warburg effect) are considered as a one of six hallmarks of cancer, including CRC. However, the role of insulin/IGF-1 signaling during the acquisition of the Warburg metabolic phenotypes by CRC cells is still poorly understood. It most likely results from the interaction of multiple processes, directly or indirectly regulated by IGF-1, such as activation of PI3K/Akt/mTORC, and Raf/MAPK signaling pathways, activation of glucose transporters (e.g., GLUT1), activation of key glycolytic enzymes (e.g., LDHA, LDH5, HK II, and PFKFB3), aberrant expression of the oncogenes (e.g., MYC, and KRAS) and/or overexpression of signaling proteins (e.g., HIF-1, TGF-β1, PI3K, ERK, Akt, and mTOR). This review describes the role of IGF-1 in glucose metabolism in physiology and colorectal carcinogenesis, including the role of the insulin/IGF system in the Warburg effect. Furthermore, current therapeutic strategies aimed at repairing impaired glucose metabolism in CRC are indicated.
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Oruç Z, Kaplan MA. Effect of exercise on colorectal cancer prevention and treatment. World J Gastrointest Oncol 2019; 11:348-366. [PMID: 31139306 PMCID: PMC6522766 DOI: 10.4251/wjgo.v11.i5.348] [Citation(s) in RCA: 62] [Impact Index Per Article: 10.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/24/2019] [Revised: 04/17/2019] [Accepted: 05/06/2019] [Indexed: 02/05/2023] Open
Abstract
In recent years, because of improved cancer screening, detection and treatment modalities, a rapid increase in the population of colorectal and other cancer survivors has been observed. The increasing population has justified the requirement of preventive strategies such as lifestyle modifications with regard to obesity, physical activity, diet and smoking. Physical activity may prevent approximately 15% of the colon cancers. Furthermore, several observational studies have demonstrated the efficacy and dose-dependent and anti-cancer effects of exercise on decreasing the mortality and risk of recurrence before and after the colorectal cancer (CRC) diagnosis. However, the required exercise dose, type and intensity are yet unclear. The results of randomised prospective studies are expected to determine the optimal amount, type and intensity of exercise and formulate the most appropriate exercise plan and guidelines, according to the requirements and comorbidities of the patients. In addition, recent studies have focused on the molecular and genetic mechanisms underlying the effect of physical activity on disease outcomes and recurrence rates. This review aimed to investigate the effects of physical activity and the biological basis of these effects in preventing the risk and recurrence of CRC and decreasing the hazards of cancer and cancer treatment.
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Affiliation(s)
- Zeynep Oruç
- Department of Medical Oncology, Mersin City Hospital, Mersin 33000, Turkey
| | - Muhammed Ali Kaplan
- Department of Medical Oncology, Faculty of Medicine, Dicle University, Diyarbakır 21280, Turkey
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Calcium homeostasis and ER stress in control of autophagy in cancer cells. BIOMED RESEARCH INTERNATIONAL 2015; 2015:352794. [PMID: 25821797 PMCID: PMC4363509 DOI: 10.1155/2015/352794] [Citation(s) in RCA: 166] [Impact Index Per Article: 16.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 08/20/2014] [Revised: 11/21/2014] [Accepted: 11/24/2014] [Indexed: 01/29/2023]
Abstract
Autophagy is a basic catabolic process, serving as an internal engine during responses to various cellular stresses. As regards cancer, autophagy may play a tumor suppressive role by preserving cellular integrity during tumor development and by possible contribution to cell death. However, autophagy may also exert oncogenic effects by promoting tumor cell survival and preventing cell death, for example, upon anticancer treatment. The major factors influencing autophagy are Ca2+ homeostasis perturbation and starvation. Several Ca2+ channels like voltage-gated T- and L-type channels, IP3 receptors, or CRAC are involved in autophagy regulation. Glucose transporters, mainly from GLUT family, which are often upregulated in cancer, are also prominent targets for autophagy induction. Signals from both Ca2+ perturbations and glucose transport blockage might be integrated at UPR and ER stress activation. Molecular pathways such as IRE 1-JNK-Bcl-2, PERK-eIF2α-ATF4, or ATF6-XBP 1-ATG are related to autophagy induced through ER stress. Moreover ER molecular chaperones such as GRP78/BiP and transcription factors like CHOP participate in regulation of ER stress-mediated autophagy. Autophagy modulation might be promising in anticancer therapies; however, it is a context-dependent matter whether inhibition or activation of autophagy leads to tumor cell death.
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Brown JC, Winters-Stone K, Lee A, Schmitz KH. Cancer, physical activity, and exercise. Compr Physiol 2013; 2:2775-809. [PMID: 23720265 DOI: 10.1002/cphy.c120005] [Citation(s) in RCA: 214] [Impact Index Per Article: 17.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022]
Abstract
This review examines the relationship between physical activity and cancer along the cancer continuum, and serves as a synthesis of systematic and meta-analytic reviews conducted to date. There exists a large body of epidemiologic evidence that conclude those who participate in higher levels of physical activity have a reduced likelihood of developing a variety of cancers compared to those who engage in lower levels of physical activity. Despite this observational evidence, the causal pathway underlying the association between participation in physical activity and cancer risk reduction remains unclear. Physical activity is also a useful adjunct to improve the deleterious sequelae experienced during cancer treatment. These deleterious sequelae may include fatigue, muscular weakness, deteriorated functional capacity, and many others. The benefits of physical activity during cancer treatment are similar to those experienced after treatment. Despite the growing volume of literature examining physical activity and cancer across the cancer continuum, a number of research gaps exist. There is little evidence on the safety of physical activity among all cancer survivors, as most trials have selectively recruited participants. The specific dose of exercise needed to optimize primary cancer prevention or symptom control during and after cancer treatment remains to be elucidated.
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Affiliation(s)
- Justin C Brown
- University of Pennsylvania, Perelman School of Medicine, Philadelphia, Pennsylvania, USA
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The insulin receptor substrate 1 (IRS1) in intestinal epithelial differentiation and in colorectal cancer. PLoS One 2012; 7:e36190. [PMID: 22558377 PMCID: PMC3338610 DOI: 10.1371/journal.pone.0036190] [Citation(s) in RCA: 52] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/15/2010] [Accepted: 04/01/2012] [Indexed: 12/13/2022] Open
Abstract
Colorectal cancer (CRC) is associated with lifestyle factors that affect insulin/IGF signaling, of which the insulin receptor substrate 1 (IRS1) is a key transducer. We investigated expression, localization and pathologic correlations of IRS1 in cancer-uninvolved colonic epithelium, primary CRCs with paired liver metastases and in vitro polarizing Caco2 and HT29 cells. IRS1 mRNA and protein resulted higher, relative to paired mucosa, in adenomas of familial adenomatous polyposis patients and in CRCs that overexpressed c-MYC, ß-catenin, InsRß, and IGF1R. Analysis of IRS1 immunostaining in 24 cases of primary CRC with paired colonic epithelium and hepatic metastasis showed that staining intensity was significantly higher in metastases relative to both primary CRC (P<0.01) and colonic epithelium (P<0.01). Primary and metastatic CRCs, compared to colonic epithelium, contained significantly higher numbers of IRS1-positive cells (P = 0.013 and P = 0.014, respectively). Pathologic correlations in 163 primary CRCs revealed that diffuse IRS1 staining was associated with tumors combining differentiated phenotype and aggressive markers (high Ki67, p53, and ß-catenin). In Caco 2 IRS1 and InsR were maximally expressed after polarization, while IGF1R was highest in pre-polarized cells. No nuclear IRS1 was detected, while, with polarization, phosphorylated IRS1 (pIRS1) shifted from the lateral to the apical plasma membrane and was expressed in surface cells only. In HT29, that carry mutations constitutively activating survival signaling, IRS1 and IGF1R decreased with polarization, while pIRS1 localized in nuclear spots throughout the course. Overall, these data provide evidence that IRS1 is modulated according to CRC differentiation, and support a role of IRS1 in CRC progression and liver metastatization.
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Perakslis E, Tuesca A, Lowman A. Complexation hydrogels for oral protein delivery: an in vitro assessment of the insulin transport-enhancing effects following dissolution in simulated digestive fluids. JOURNAL OF BIOMATERIALS SCIENCE-POLYMER EDITION 2012. [DOI: 10.1163/156856207794761989] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/19/2022]
Affiliation(s)
- Eric Perakslis
- a Biomaterials and Drug Delivery Laboratory, Department of Chemical and Biological Engineering, Drexel University, Philadelphia, PA 19104, USA; Centocor Research & Development, Radnor, PA 19087, USA
| | - Anthony Tuesca
- b Biomaterials and Drug Delivery Laboratory, Department of Chemical and Biological Engineering, Drexel University, Philadelphia, PA 19104, USA
| | - Anthony Lowman
- c Biomaterials and Drug Delivery Laboratory, Department of Chemical and Biological Engineering, Drexel University, Philadelphia, PA 19104, USA
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Abstract
AbstractAcromegaly is associated with an increased prevalence of colonic polyps. The aim of this study was to evaluate the prevalence and recurrence rate of colonic polyps in acromegalic patients. Ninety-six acromegalic patients and 100 irritable bowel syndrome patients (IBS) were enrolled in the study. Twenty patients who were cured exclusively by surgery, and 20 patients that could not be hormonally controlled were re-examined colonoscopically after 36 months. Twenty-nine of 96 acromegalic patients (30.2%) had colonic polyps. In the IBS group, 10 (10.0%) had colonic polyps. The prevalence of colonic polyps was significantly higher in acromegaly. The group of acromegalic patients with and without polyps did not differ significantly with regard to plasma GH, IGF-I, fasting insulin levels and glycemic status. The presence of colonic polyps was correlated with increased patient age and male gender. We did not observe a difference in terms of polyp recurrence frequencies in the patients cured by surgery compared to uncontrolled patients. Acromegalic patients have a higher prevalence of colonic polyps than that of control subjects. We could not identify any factors that could predict polyps within the acromegalic patients - but age and male sex.
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Abstract
Colorectal cancer is the third most common cancer globally and is a leading cause of cancer death. Gastric cancer contributes significantly to the global cancer burden, particularly in low- and middle-income countries. We reviewed the literature for studies on physical activity or exercise and colon, rectal, and gastric cancers as well as colonic adenomatous polyps. We identified 52 studies of colon cancer, 31 studies of rectal cancer, 23 studies of colon polyps, and 16 studies of gastric cancer. Of the 52 studies of physical activity and colon cancer, 37 found a statistically significant association between increased levels of physical activity and decreased colon cancer risk in at least one comparison. Accumulated evidence suggests that physical activity is associated with a 25% reduction in colon cancer risk. In line with previous reports, we found no indication that the association was more pronounced for occupational versus recreational physical activity, with both resulting in a risk reduction of about 22%. Evidence for other domains of physical activity (i.e., transportation or household physical activity) is limited. Evidence is emerging that individuals who are consistently active across the lifetime may obtain greater risk reductions than those who are only active in recent years. Despite consistent associations with colon cancer, evidence is more limited though suggestive that physical activity reduces risk of colon adenomas or adenoma recurrence. There is clear evidence that physical activity is not associated with rectal or gastric cancers.
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Tripkovic I, Tripkovic A, Strnad M, Capkun V, Zekan L. Role of insulin-like growth factor-1 in colon cancerogenesis: a case-control study. Arch Med Res 2007; 38:519-25. [PMID: 17560457 DOI: 10.1016/j.arcmed.2007.01.012] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/30/2006] [Accepted: 01/22/2007] [Indexed: 10/23/2022]
Abstract
BACKGROUND Our aim was to establish whether individuals who developed colon cancer have elevated blood levels of insulin-like growth factor-1 (IGF-1). METHODS This was a case/control study in which 52 patients with colon cancer and a corresponding control group were investigated. Data on age, weight, height, and sex of subjects were recorded and levels of IGF-1 and growth hormone, as well as insulin and C-peptide levels, were measured in the morning before eating, 90 min after breakfast and again 90 min after lunch. RESULTS We found significantly higher levels of IGF-1 in blood of colon cancer patients compared to the control group. No differences in the levels of growth hormone, insulin and C-peptide in blood were found between colon cancer patients and the control group. It was found that the increase of IGF-1 level was followed by a 3.15-fold increased risk for developing colon cancer. There were no differences in the levels of IGF-1 in blood in all three measurements in the group of colon cancer patients, whereas differences were found in the control group. We found differences in the levels of insulin and C-peptide in blood in all three measurements in both groups of patients. No differences were found in the levels of growth hormone in blood in all three measurements in both groups of patients. CONCLUSIONS The results of this study suggest a positive correlation between the increased levels of IGF-1 and colon cancer and are thus consistent with the hypothesis that the level of IGF-1 plays an important role in the development of colon cancer.
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Affiliation(s)
- Ingrid Tripkovic
- Department of Epidemiology, Teaching Institute of Public Health of Split and Dalmatia Country, Split, Croatia.
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Samad AKA, Taylor RS, Marshall T, Chapman MAS. A meta-analysis of the association of physical activity with reduced risk of colorectal cancer. Colorectal Dis 2005; 7:204-13. [PMID: 15859955 DOI: 10.1111/j.1463-1318.2005.00747.x] [Citation(s) in RCA: 227] [Impact Index Per Article: 11.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/11/2022]
Abstract
BACKGROUND Physical activity may be associated with reduced risk of colorectal cancer. The main aim of this paper is to review the available evidence for a link between exercise and large bowel cancer. METHODS A Cochrane-type methodology was performed. Data extracted included, type of study, type of physical activity measured and the numerical results. The risk ratios (RR) of the studies have been pooled according to the type of study, type of exercise, type of cancer and sex. Pooling was undertaken using fixed effect meta-analysis. A random effect meta-analysis was used where substantial heterogeneity existed. RESULT Data from 19 cohort studies showed a statistically significant reduction in the risk of colon cancer in physically active males, RR being 0.79 (95% CI 0.72-0.87) and 0.78 (95% CI 0.68-0.91) for occupational and recreational activities, respectively. In women only recreational activities are protective against colon cancer (RR = 0.71, 95%CI 0.57-0.88). Case-control studies showed significantly reduced risks of colon cancer in both sexes irrespective of the type of activity. No protection against rectal cancer is seen in either sex. CONCLUSION There is considerable evidence that physical activity is associated with reduced risk of colon cancer in both males and females.
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Affiliation(s)
- A K A Samad
- Department of Surgery, Good Hope Hospital, Sutton Coldfield, UK
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Watanabe K, Terada K, Jinriki T, Sato J. Effect of insulin on cephalexin uptake and transepithelial transport in the human intestinal cell line Caco-2. Eur J Pharm Sci 2004; 21:87-95. [PMID: 14706815 DOI: 10.1016/j.ejps.2003.10.002] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/26/2022]
Abstract
We investigated whether cephalexin transport in Caco-2 cells is regulated by insulin. After the insulin pretreatment, cephalexin uptake, and transport as well as PEPT1 mRNA and protein expression in the cells were measured. Cephalexin uptake was significantly increased by the insulin pretreatment. Insulin significantly increased cephalexin saturable uptake, but had no significant effect on the non-saturable one. PEPT1 protein expression on the apical membrane, but not PEPT1 mRNA expression, was increased by the insulin pretreatment. The enhancement of cephalexin uptake by the insulin pretreatment was inhibited by genistein, a tyrosine kinase inhibitor, and colchicine, an agent that disrupts protein translocation. Apical-to-basolateral transport of cephalexin has increased by the insulin pretreatment at the apical side and long-term insulin pretreatment at the basolateral side. It is considered that insulin mainly binds to its receptor on the apical and basolateral membranes, thereby promoting PEPT1 translocation from the intracellular pool to the apical membrane surface; consequently, PEPT1 protein expression on the apical membrane is increased.
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Affiliation(s)
- Kazuhiro Watanabe
- Hokkaido College of Pharmacy, 7-1 Katsuraoka-cho, Otaru, Hokkaido 047-0264, Japan.
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Ichikawa H, Peppas NA. Novel complexation hydrogels for oral peptide delivery: in vitro evaluation of their cytocompatibility and insulin-transport enhancing effects using Caco-2 cell monolayers. J Biomed Mater Res A 2003; 67:609-17. [PMID: 14566804 PMCID: PMC4467685 DOI: 10.1002/jbm.a.10128] [Citation(s) in RCA: 69] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022]
Abstract
Poly[methacrylic acid-grafted-poly(ethylene glycol)] [P(MAA-g-EG)] is a complexation hydrogel molecularly designed for oral peptide delivery. In this work, the cytotoxicity and insulin-transport enhancing effect of P(MAA-g-EG) microparticles on intestinal epithelial cells were evaluated using Caco-2 cell monolayers. A series of P(MAA-g-EG) microparticles with different polymer compositions were prepared by a photo-initiated free radical solution polymerization and subsequent pulverization. The hydrogel microparticles were preswollen in either Ca2+-containing (CM+) or Ca2+-free medium (CM-; pH 7.4) and applied to the apical side of the Caco-2 monolayers. No significant cytotoxic effects, as determined by a calorimetric assay with P(MAA-g-EG) microparticles preswollen in the CM+, were observed at doses ranging from 3 to 31 mg/cm2 of cell monolayer. Transepithelial electrical resistance (TEER) measurements showed that the P(MAA-g-EG) microparticles induced a Ca2+ concentration-dependent lowering in TEER values. The reduction effect in CM- media was greater than that in CM+ media (17 +/- 2% reduction in CM+ and 45 +/- 3% reduction in CM-, respectively). Insulin transport in the presence of the preswollen P(MAA-g-EG) microparticles was also strongly depended on the Ca2+ concentration in the medium. The respective estimated permeability for insulin alone and the insulin with hydrogels in CM+ were 0.77 and 1.16 x 10(-8) cm/s, whereas those in CM- were 1.18 and 24.78 x 10(-8) cm/s. The results demonstrate that the P(MAA-g-EG) hydrogel microparticles could be used as a cytocompatible carrier possessing the transport-enhancing effect of insulin on the intestinal epithelial cells.
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Affiliation(s)
- Hideki Ichikawa
- Biomaterials, Drug Delivery and Molecular Recognition Laboratories, Department of Chemical Engineering, University of Texas at Austin, Austin, Texas 78712-0231, USA.
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Adibi SA. Regulation of expression of the intestinal oligopeptide transporter (Pept-1) in health and disease. Am J Physiol Gastrointest Liver Physiol 2003; 285:G779-88. [PMID: 14561585 DOI: 10.1152/ajpgi.00056.2003] [Citation(s) in RCA: 127] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
The abundance of the oligopeptide transporter (Pept-1) in the brush-border membrane of the intestinal epithelium is the central mechanism for regulation of transport of products of protein digestion (dipeptides and tripeptides) and peptidomimetic drugs (for example, beta-lactam antibiotics). Within the past few years, there has been substantial progress in identifying the factors controlling this regulation and the mechanisms of their actions. The purpose of this report is to review this progress. The studies of individual substrates and hormones in a human intestinal cell line (Caco-2) have shown that dipeptides, certain amino acids, insulin, and leptin increase and epidermal growth factor and triiodothyronine decrease the membrane population of Pept-1. In the case of dipeptides, epidermal growth factor, and thyroid hormone, there are parallel changes in the gene expression brought about by alteration of transcription and/or stability of Pept-1 mRNA. In contrast, the treatment with insulin and leptin does not induce any alteration in the Pept-1 gene expression, and the mechanism of increased protein expression appears to be increased trafficking from a preformed cytoplasmic pool to the apical membrane. In vivo studies in rats have shown modulation of protein and gene expressions of the intestinal oligopeptide transporter during the day and during development and in nutritional and metabolic alterations, such as high-protein diet, fasting, and diabetes. Patients with intestinal diseases, such as ulcerative colitis, Crohn's disease, and short-bowel syndrome, may have induction of the Pept-1 expression in their colon. Finally, pharmacological studies have shown that the expression of Pept-1 can be upregulated by agents such as 5 fluorouracil and downregulated by agents such as cyclosporine. In conclusion, the above studies have produced a wealth of new information on regulation of a key transporter in the intestine. This information may have useful applications in nutritional and pharmacological treatments, for example, in diabetic patients needing enteral nutrition or in ulcerative colitis patients needing the suppression of the intestinal inflammation.
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Affiliation(s)
- Siamak A Adibi
- Emeritus Professor of Medicine, Univ. of Pittsburgh, 601 Kaufmann Bldg., 3471 Fifth Ave., Pittsburgh, PA 15213, USA.
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Nielsen CU, Amstrup J, Nielsen R, Steffansen B, Frokjaer S, Brodin B. Epidermal growth factor and insulin short-term increase hPepT1-mediated glycylsarcosine uptake in Caco-2 cells. ACTA PHYSIOLOGICA SCANDINAVICA 2003; 178:139-48. [PMID: 12780388 DOI: 10.1046/j.1365-201x.2003.01113.x] [Citation(s) in RCA: 27] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/20/2022]
Abstract
AIMS Little is known about the physiological regulation of the human intestinal di/tri-peptide transporter, hPepT1. In the present study we evaluated the effects of epidermal growth factor (EGF) and insulin on hPepT1-mediated dipeptide uptake in the intestinal cell line Caco-2. METHODS Caco-2 cells were grown on filters for 23-27 days. Apical dipeptide uptake was measured using [14C]glycylsarcosine([14C]Gly-Sar). HPepT1 mRNA levels were investigated using RT-PCR, cytosolic pH was determined using the pH-sensitive fluorescent probe BCECF. RESULTS Basolateral application of EGF increased [14C]Gly-Sar uptake with an ED50 value of 0.77 +/- 0.25 ng mL-1 (n = 3-6) and a maximal stimulation of 33 +/- 2% (n = 3-6). Insulin stimulated [14C]Gly-Sar uptake with an ED50 value of 3.5 +/- 2.0 ng mL-1 (n = 3-6) and a maximal stimulation of approximately 18% (n = 3-6). Gly-Sar uptake followed simple Michaelis-Menten kinetics. Km in control cells was 0.98 +/- 0.11 mM (n = 8) and Vmax was 1.86 +/- 0.07 nmol cm-2 min-1 (n = 8). In monolayers treated with 200 ng mL-1 of EGF, Km was 1.11 +/- 0.05 mM (n = 5) and Vmax was 2.79 +/- 0.05 nmol cm-2 min-1 (n = 5). In monolayers treated with 50 ng mL-1 insulin, Km was 1.03 +/- 0.08 mM and Vmax was 2.19 +/- 0.06 nmol cm-2 min-1 (n = 5). Kinetic data thus indicates an increase in the number of active transporters, following stimulation. The incrased Gly-Sar uptake was not accompanied by changes in hPepT1 mRNA, nor by measurable changes in cytosolic pH. CONCLUSIONS Short-term stimulation with EGF and insulin caused an increase in hPepT1-mediated uptake of Gly-Sar in Caco-2 cell monolayers, which could not be accounted for by changes in hPepT1 mRNA or proton-motive driving force.
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Affiliation(s)
- C U Nielsen
- Department of Pharmaceutics, Danish University of Pharmaceutical Sciences, Universitetsparken 2, DK-2100 Copenhagen, Denmark
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Chang CK, Ulrich CM. Hyperinsulinaemia and hyperglycaemia: possible risk factors of colorectal cancer among diabetic patients. Diabetologia 2003; 46:595-607. [PMID: 12764580 DOI: 10.1007/s00125-003-1109-5] [Citation(s) in RCA: 83] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/10/2003] [Revised: 03/19/2003] [Indexed: 12/15/2022]
Abstract
Hyperinsulinaemia and hyperglycaemia are two possible risk factors for colorectal cancer, which constitutes the third leading cause of cancer death in Western countries. Molecular evidence as well as animal models provide support for these associations: Insulin has been shown to be an important growth factor for colonic carcinoma cells, and both insulin and insulin-like growth factor-1 receptors have been detected in colon cancer tissue. The insulin-signal transduction pathway is involved in the regulation of gene expression and apoptosis. The role of hyperglycaemia in carcinogenesis could include pathways via luminal factors (related to fecal bile acid concentrations, stool bulk, and prolonged transit time) or circulatory factors (via glucose as the only energy source for neoplastic cells). This review summarizes the epidemiologic literature with respect to hyperinsulinaemia and hyperglycaemia as risk factors for colorectal cancer, and aims to integrate the biological and epidemiological evidence. Epidemiologic findings to date indicate a slightly increased risk of colorectal cancer for diabetic patients; however, there are some inconsistencies. Possible explanations for these inconsistencies include inadequate information about patients' diabetic disease and treatment states. We suggest that future studies should take medical history, staging and treatment for hyperinsulinaemia and hyperglycaemia into account to further our understanding of the role of hyperglycaemia and hyperinsulinaemia in colorectal carcinogenesis.
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Affiliation(s)
- C K Chang
- Department of Epidemiology, School of Public Health and Community Medicine, University of Washington, WA 98195, Seattle, USA
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Buyse M, Berlioz F, Guilmeau S, Tsocas A, Voisin T, Péranzi G, Merlin D, Laburthe M, Lewin MJ, Rozé C, Bado A. PepT1-mediated epithelial transport of dipeptides and cephalexin is enhanced by luminal leptin in the small intestine. J Clin Invest 2001; 108:1483-94. [PMID: 11714740 PMCID: PMC209419 DOI: 10.1172/jci13219] [Citation(s) in RCA: 152] [Impact Index Per Article: 6.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/25/2022] Open
Abstract
Dietary proteins are mostly absorbed as di- and tripeptides by the intestinal proton-dependent transporter PepT1. We have examined the effects of leptin on PepT1 function in rat jejunum and in monolayers of the human enterocyte-like 2 cell Caco-2. Leptin is produced by the stomach and secreted in the gut lumen. We show here that PepT1 and leptin receptors are expressed in Caco-2 and rat intestinal mucosal cells. Apical (but not basolateral) leptin increased Caco-2 cell transport of cephalexin (CFX) and glycylsarcosine (Gly-Sar), an effect that was associated with increased Gly-Sar uptake, increased membrane PepT1 protein, decreased intracellular PepT1 content, and no change in PepT1 mRNA levels. The maximal velocity (Vmax) for Gly-Sar transport was significantly increased by leptin, whereas the apparent Michaelis-Menten constant (Km) did not change. Furthermore, leptin-stimulated Gly-Sar transport was completely suppressed by colchicine, which disrupts cellular translocation of proteins to plasma membranes. Intrajejunal leptin also induced a rapid twofold increase in plasma CFX after jejunal perfusion with CFX in the rat, indicating enhanced intestinal absorption of CFX. These data revealed an unexpected action of gastric leptin in controlling ingestion of dietary proteins.
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Affiliation(s)
- M Buyse
- Institut National de la Santé et de la Recherche Médicale Unité 410, Faculté de Médecine Xavier Bichat, Paris, France
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18
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Giovannucci E. Insulin, insulin-like growth factors and colon cancer: a review of the evidence. J Nutr 2001; 131:3109S-20S. [PMID: 11694656 DOI: 10.1093/jn/131.11.3109s] [Citation(s) in RCA: 658] [Impact Index Per Article: 27.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/02/2023] Open
Abstract
Insulin and insulin-like growth factor (IGF) axes are major determinants of proliferation and apoptosis and thus may influence carcinogenesis. In various animal models, modulation of insulin and IGF-1 levels through various means, including direct infusion, energy excess or restriction, genetically induced obesity, dietary quality including fatty acid and sucrose content, inhibition of normal insulin secretion and pharmacologic inhibition of IGF-1, influences colonic carcinogenesis. Human evidence also associates high levels of insulin and IGF-1 with increased risk of colon cancer. Clinical conditions associated with high levels of insulin (noninsulin-dependent diabetes mellitus and hypertriglyceridemia) and IGF-1 (acromegaly) are related to increased risk of colon cancer, and increased circulating concentrations of insulin and IGF-1 are related to a higher risk of colonic neoplasia. Determinants and markers of hyperinsulinemia (physical inactivity, high body mass index, central adiposity) and high IGF-1 levels (tall stature) are also related to higher risk. Many studies indicate that dietary patterns that stimulate insulin resistance or secretion, including high consumption of sucrose, various sources of starch, a high glycemic index and high saturated fatty acid intake, are associated with a higher risk of colon cancer. Although additional environmental and genetic factors affect colon cancer, the incidence of this malignancy was invariably low before the technological advances that rendered sedentary lifestyles and obesity common, and increased availability of highly processed carbohydrates and saturated fatty acids. Efforts to counter these patterns are likely to have the most potential to reduce colon cancer incidence, as well as cardiovascular disease and diabetes mellitus.
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Affiliation(s)
- E Giovannucci
- Channing Laboratory, Department of Medicine, Harvard Medical School and Brigham and Women's Hospital, Boston, MA 02115, USA.
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19
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Nishii T, Kono S, Abe H, Eguchi H, Shimazaki K, Hatano B, Hamada H. Glucose intolerance, plasma insulin levels, and colon adenomas in Japanese men. Jpn J Cancer Res 2001; 92:836-40. [PMID: 11509114 PMCID: PMC5926827 DOI: 10.1111/j.1349-7006.2001.tb01169.x] [Citation(s) in RCA: 35] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022] Open
Abstract
Hyperinsulinemia may be related to colon carcinogenesis. Several studies have suggested that diabetes mellitus is related to increased risk of colon cancer. We examined cross-sectionally the relation of fasting plasma insulin levels and glucose tolerance status to colon adenomas. In a consecutive series of 951 men undergoing total colonoscopy for a health examination at the Japan Self Defense Forces Fukuoka Hospital from April 1998 to August 1999, we identified 233 cases of colon adenomas and 497 controls with normal colonoscopy. Glucose tolerance status was determined by a 75-g oral glucose tolerance test, and subjects were classified as normal, impaired glucose tolerance (IGT) or non-insulin dependent diabetes mellitus (NIDDM). Plasma insulin levels were measured after subjects had fasted overnight. Logistic regression analysis and analysis of covariance was used to control for age and obesity. While plasma insulin levels were unrelated to colon adenomas, NIDDM was associated with a significantly increased risk of colon adenomas. There was no association between IGT and colon adenomas. NIDDM was more strongly associated with proximal colon adenomas. The findings suggest that long-term hyperinsulinemic status associated with NIDDM may increase the risk of colon adenomas, and subsequently of colon cancer.
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Affiliation(s)
- T Nishii
- Self Defense Forces Fukuoka Hospital, Kasuga-shi, Fukuoka 816-0826, Japan.
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20
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Affiliation(s)
- Y I Kim
- Division of Gastroenterology Department of Medicine University of Toronto and St. Michael's Hospital Department of Nutritional Sciences University of Toronto Toronto, Canada
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21
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Morishita M, Kajita M, Suzuki A, Takayama K, Chiba Y, Tokiwa S, Nagai T. The dose-related hypoglycemic effects of insulin emulsions incorporating highly purified EPA and DHA. Int J Pharm 2000; 201:175-85. [PMID: 10878324 DOI: 10.1016/s0378-5173(00)00411-7] [Citation(s) in RCA: 17] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/18/2022]
Abstract
The dose-related pharmacological effects of insulin emulsion incorporating highly purified eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) were investigated. Water-in-oil-in-water multiple emulsions (insulin dose, 0, 10, 25 and 50 IU/kg) incorporating 2% DHA or EPA were administered directly into the colonic and rectal loops in situ. Serum insulin levels rose and serum glucose levels decreased in an insulin dose-related fashion. The relationship of insulin dose and C(max) or AUC(insulin) was linear at the rectum, but a non-linear relationship was observed at the colon. The trend was more predominant in DHA. In the in vivo rectal absorption experiment using emulsions incorporating 2% DHA, 5 IU/kg of insulin emulsion produced a rapid, transitory increase in serum insulin levels and strong reduction of serum glucose levels. The pharmacological availability determined from the dose-response curve by s.c. administration of insulin reached 43.2+/-26.3% (mean+/-S.D.). Mucosal irritation caused by administration of emulsions incorporating 2% EPA or DHA was evaluated by a lactate dehydrogenase (LDH) release study, and compared with those of the emulsion incorporating 2% oleic or linolenic acid. Only when emulsion incorporating 2% oleic acid was applied in the intestine did significant LDH release into the mesenteric veins occur. Our results indicate that emulsion incorporating highly purified long-chain polyunsaturated fatty acid, especially DHA, has the potential of becoming the formulation for enteral delivery of insulin.
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Affiliation(s)
- M Morishita
- Department of Pharmaceutics, Hoshi University, Ebara 2-4-41, Shinagawa-ku, 142 8501, Tokyo, Japan.
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22
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Hakam A, Yeatman TJ, Lu L, Mora L, Marcet G, Nicosia SV, Karl RC, Coppola D. Expression of insulin-like growth factor-1 receptor in human colorectal cancer. Hum Pathol 1999; 30:1128-33. [PMID: 10534157 DOI: 10.1016/s0046-8177(99)90027-8] [Citation(s) in RCA: 176] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
The activation of the insulinlike growth factor 1/IGF-1 receptor system (IGF1/IGF1-R) has recently emerged as critical event in transformation and tumorigenicity of several murine and human tumors. Expression of IGF1 and of IGF1-R has been demonstrated in normal and neoplastic intestinal cell lines of rats and humans. However, the modulation of IGF1-R expression during the progression from normal colonic mucosa to adenoma, to carcinoma, and to metastasis, has not been evaluated. In this retrospective study, we investigated the expression of IGF1-R in 12 colonic adenomas (AD), 36 primary colorectal adenocarcinomas (CA), and in 27 corresponding metastases (MT). Normal colonic mucosa (N) was adjacent to the CA in 34 cases. Formalin-fixed, paraffin-embedded tissues of each case were immunostained using the avidin-biotin-peroxidase method. We used an anti-IGF1-R rabbit polyclonal antibody (Santa Cruz Biotechnology, CA; dilution 1:100). Positive staining was quantitated by CAS-200. Moderate to strong cytoplasmic immunostaining was observed in 34 of 36 CA (96%), and in 25 of 27 MT (93%). In all of the positive MTs, the intensity of the staining was always strong. In 10 of 12 ADs (83%), only a faint cytoplasmic stain was identified. Normal mucosa when present was negative. Strong IGF1-R positivity correlated with higher grade and higher-stage tumors (P < .01). These data suggest a role of IGF1-R expression during the progression of colorectal adenoma to carcinoma. An increased number of IGF1-R receptors may favor the metastasis of colorectal cancer.
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Affiliation(s)
- A Hakam
- Department of Pathology, Moffitt Cancer Center and Research Institute, University of South Florida, College of Medicine, Tampa 33612-9497, USA
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23
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Thamotharan M, Bawani SZ, Zhou X, Adibi SA. Hormonal regulation of oligopeptide transporter pept-1 in a human intestinal cell line. THE AMERICAN JOURNAL OF PHYSIOLOGY 1999; 276:C821-6. [PMID: 10199812 DOI: 10.1152/ajpcell.1999.276.4.c821] [Citation(s) in RCA: 130] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
The intestinal oligopeptide transporter (cloned as Pept-1) has major roles in protein nutrition and drug therapy. A key unstudied question is whether expression of Pept-1 is hormonally regulated. In this experiment, we investigated whether insulin has such a role. We used a human intestinal cell monolayer (Caco-2) as the in vitro model of human small intestine and glycylglutamine (Gly-Gln) as the model substrate for Pept-1. Results showed that addition of insulin at a physiological concentration (5 nM) to incubation medium greatly stimulates Gly-Gln uptake by Caco-2 cells. This stimulation was blocked when genistein, an inhibitor of tyrosine kinase, was added to incubation medium. Studies of the mechanism of insulin stimulation showed the following. 1) Stimulation occurred promptly (30-60 min) after exposure to insulin. 2) There was no significant change in the Michaelis-Menten constant of Gly-Gln transport, but there was a nearly twofold increase in its maximal velocity. 3) Insulin effect persisted even when Golgi apparatus, which is involved in trafficking of newly synthesized Pept-1, was dismantled. 4) However, there was complete elimination of insulin effect by disruption of microtubules involved in trafficking of preformed Pept-1. 5) Finally, with insulin treatment, there was no change in Pept-1 gene expression, but the amount of Pept-1 protein in the apical membrane was increased. In conclusion, the results show that insulin, when it binds to its receptor, stimulates Gly-Gln uptake by Caco-2 cells by increasing the membrane population of Pept-1. The mechanism appears to be increased translocation of this transporter from a preformed cytoplasmic pool.
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Affiliation(s)
- M Thamotharan
- Clinical Nutrition Research Unit, Department of Medicine, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15260, USA
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24
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Abstract
Several dietary and other lifestyle factors have been implicated in the development of colorectal cancer. However, the precise nature and actual magnitude of the relationship between individual nutrient intakes and other lifestyle factors and colorectal cancer risk are not clear. A unifying hypothesis has recently been proposed that explains why obesity, physical inactivity, alcohol, and consumption of a typical Western diet increase colorectal cancer risk. This hypothesis suggests that these dietary and other lifestyle factors are associated with insulin resistance and hyperinsulinemia and that hyperinsulinemia, in turn, may stimulate growth of colorectal tumors. Two recently published large prospective epidemiologic studies indicate a significant increase in colorectal cancer risk in subjects with diabetes mellitus, thereby supporting this hypothesis.
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Affiliation(s)
- Y I Kim
- Department of Medicine, St. Michael's Hospital, University of Toronto, Ontario, Canada
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25
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Lindmark T, Schipper N, Lazorová L, de Boer AG, Artursson P. Absorption enhancement in intestinal epithelial Caco-2 monolayers by sodium caprate: assessment of molecular weight dependence and demonstration of transport routes. J Drug Target 1998; 5:215-23. [PMID: 9606011 DOI: 10.3109/10611869808995876] [Citation(s) in RCA: 59] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/13/2022]
Abstract
Sodium caprate (C10), a medium chain fatty acid, is used clinically to enhance rectal absorption of the low molecular weight (MW) drug ampicillin. The main aim of this study was to investigate whether C10 also enhances the permeability of high MW model drugs in a model of the intestinal epithelium. The second aim was to present visual evidence of the route of enhanced transport across the epithelial cell layer. The studies were performed in Caco-2 monolayers cultured on permeable supports. The effects of non-toxic concentrations (< or = 13 mM) of C10 on drug transport across the monolayers was studied using monodisperse 14C-polyethylene glycols (MW 238-502; 14C-PEGs), 125I-Arg5-vasopressin (MW 1,208), 125I-insulin (MW 6,000) and FITC-labelled dextrans (MW 4,400 and 19,600; FD4 and FD20 respectively) as model drugs. Electron and confocal laser scanning microscopy were used to demonstrate transport routes across the epithelium. 10 mM C10 increased the permeability of all 14C-PEGs to approximately the same extent. 13 mM C10 increased the permeability of 125I-Arg8-vasopressin 10-fold. Only small increases in FD4 and FD20 permeabilities were observed. After C10 exposure, both tight junctions with normal morphology and those with dilatations showed an increased permeability to ruthenium red, indicating that C10 enhanced the paracellular transport of molecules with a MW < 1,000. Confocal microscopy showed that C10 increased the transport of FD4 and FD20 by the paracellular route. In conclusion, non-toxic concentrations of C10 can be used to enhance the permeability of drugs of MW up to approximately 1,200. Enhancement of the absorption of molecules larger than 4,000 is quantitatively insignificant. The enhanced permeability occurred via the paracellular pathway.
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Affiliation(s)
- T Lindmark
- Department of Pharmacy, Uppsala University, Sweden
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26
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Chang LL, Stout LE, Wong WD, Buls JG, Rothenberger DA, Shier WT, Sorenson RL, Bai JP. Immunohistochemical localization of insulin-degrading enzyme along the rat intestine, in the human colon adenocarcinoma cell line (Caco-2), and in human ileum. J Pharm Sci 1997; 86:116-9. [PMID: 9002470 DOI: 10.1021/js960035q] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/03/2023]
Abstract
Insulin-degrading enzyme (IDE) has been implicated in the intracellular degradation of insulin in insulin target cells. Knowledge of the existence of this enzyme in the intestine will be beneficial to the achievement of clinical oral efficacy of insulin. A comparative study was conducted with rat intestine, human colon adenocarcinoma (Caco-2) cells, and human ileum. Confocal microscopy analysis using the anti-IDE antibody showed that IDE was localized in the mucosal cells of rat and human intestines, as well as in Caco-2 cells. Immunostaining of this enzyme was homogeneous throughout the cell excluding nucleus, indicating a typical cytosolic distribution in rat and human enterocytes and in Caco-2 cells.
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Affiliation(s)
- L L Chang
- Department of Pharmaceutics, College of Pharmacy, University of Minnesota, Minneapolis 55455, USA
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27
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Shah D, Shen WC. Transcellular delivery of an insulin-transferrin conjugate in enterocyte-like Caco-2 cells. J Pharm Sci 1996; 85:1306-11. [PMID: 8961144 DOI: 10.1021/js9601400] [Citation(s) in RCA: 59] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/03/2023]
Abstract
Insulin, acylated with dimethylmaleic anhydride, was conjugated to transferrin (Tf) via a disulfide linkage. The molar insulin: Tf ratio in the conjugate was 3:1. The insulin-Tf conjugate (insulin-Tf) was tested for the transport of insulin across enterocyte-like Caco-2 cell monolayers by the process of transferrin receptor (TfR)-mediated transcytosis. The uptake of insulin-Tf in Caco-2 cells was TfR-mediated but no insulin receptor-mediated. Transport studies showed that insulin-Tf transport was 5- to 15-fold higher than free insulin transport across Caco-2 cells in both apical-to-basal and basal-to-apical directions. Brefeldin A (BFA), an agent that we have previously shown to cause an increase in TfR transcytosis, further enhanced the transport of the conjugated insulin three-fold in both directions; thus, a combination of the conjugate and BFA can cause a net 45-fold increase in the apical-to-basolateral transport of insulin across Caco-2 cell monolayers. The transported conjugate was intact as indicated by elution on a Sephadex G-50 column. Insulin in the transcytosed conjugate, unlike the original dimethylmaleyl insulin, was capable of binding to anti-insulin antibodies, indicating that free amino groups of insulin were regenerated either during or after the transcytotic process. Because Caco-2 cell monolayers provide a good model for intestinal epithelium, the insulin-Tf conjugate in combination with BFA can be a rational approach to increase the oral absorption of insulin in vivo.
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Affiliation(s)
- D Shah
- Department of Pharmaceutical Sciences, School of Pharmacy, University of Southern California, Los Angeles 90033, USA
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28
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Turner JR, Lencer WI, Carlson S, Madara JL. Carboxy-terminal vesicular stomatitis virus G protein-tagged intestinal Na+-dependent glucose cotransporter (SGLT1): maintenance of surface expression and global transport function with selective perturbation of transport kinetics and polarized expression. J Biol Chem 1996; 271:7738-44. [PMID: 8631815 DOI: 10.1074/jbc.271.13.7738] [Citation(s) in RCA: 53] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/01/2023] Open
Abstract
The Na+-dependent glucose transporter (SGLT1) mediates absorption of luminal glucose by the intestine. However, available intestinal cell lines that recapitulate a monolayer phenotype only express SGLT1 at low levels. Thus, to facilitate studies of the biology of SGLT1 function in epithelial monolayers, we engineered an epitope-tagged construct containing the YTDIEMNRLGK sequence (from the vesicular stomatitis virus G protein). The tag was placed at the carboxyl terminus since this is the least conserved portion of SGLT1. Transiently transfected COS-1 cells demonstrated surface expression of the immunoreactive protein and enhanced Na+-dependent glucose uptake that was phloridzin-sensitive (a specific competitive inhibitor of SGLT1). However, subsequent detailed analyses of epitope-tagged SGLT1 using stably transfected clones derived from the Caco-2 human intestinal epithelial cell line revealed substantial effects of the epitope on critical functions of SGLT1. When compared with native SGLT1 transfectants, the apparent Km for sugar transport was increased 23-fold (313 microM to 7.37 mM for native versus epitope-tagged SGLT1). In contrast, the apparent KNa for epitope-tagged SGLT1 was similar to that for native SGLT1. Permeabilization studies indicated that the C-terminal epitope tag was intracellular and thus could not directly disrupt extracellular ligand-binding sites. Immunolocalization and functional assays designed to detect polarized surface expression indicated that epitope tagging resulted in loss of apical targeting and enrichment of basolateral expression. Functional isolation of the small apical pool of epitope-tagged SGLT1 (by selective inhibition of basolateral epitope-tagged SGLT1) revealed that, despite the documented kinetic alterations in sugar transport, epitope-tagged SGLT1 could promote absorptive Na+ currents. These data show that 1) the C terminus of SGLT1 is intracellular; 2) disruption of protein structure by addition of a C-terminal tag leads to selective modifications of SGLT1 function; 3) the kinetics of sugar transport can be altered independently of influences on the Na+-binding site of SGLT1; and 4) the weak basolateral targeting sequence present within the epitope tag is dominant over endogenous SGLT1 apical targeting information and can direct polytopic membrane protein localization. The data also caution that subtle effects of foreign sequences must be considered when epitope tagging polytopic membrane proteins.
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Affiliation(s)
- J R Turner
- Division of Gastrointestinal Pathology, Department of Pathology, Brigham and Women's Hospital, Boston, Massachusetts 02115, USA
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29
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Bading JR, Mircheff AK, Kan-Mitchell J. Evidence of sodium-dependent glucose transport in human erythroleukemia cells. Life Sci 1996; 58:1445-52. [PMID: 8622570 DOI: 10.1016/0024-3205(96)00114-2] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
Sodium-dependent transport of D-glucose has been reported only in epithelial cells of small intestine and kidney, and well-differentiated tumors thereof. We observed a two-fold decrease (p < 0.05) in the intracellular distribution volume (Vic, defined as steady-state intracellular uptake divided by extracellular concentration) of the non-metabolized D-glucose analog 3-O-methylglucose (3-O-MG) when logarithmically growing K562 cells (an anaplastic human erythroleukemia) were incubated 3 h in choline-substituted, phosphate buffered saline (PBS) rather than Na+ PBS, each containing a glucose concentration ([Glu]) of 5.6 mM. Electromechanically measured cellular volume Vc differed < 10% between the different media. In Na+ PBS, Vic (3-O-MG) was approximately twice Vc and declined progressively when [Glu] was reduced to 2.8 and 0.1 mM. We conclude that, in a balanced salt medium containing glucose as the only energy source, K562 cells express a concentrative mechanism having characteristics consistent with Na(+)-dependent transport of glucose.
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Affiliation(s)
- J R Bading
- Department of Radiology, University of Southern California, Los Angeles 90033, USA
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30
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Park JH, Corkins MR, Vanderhoof JA, Caruso NM, Hrbek MJ, Schaffer BS, Slentz DH, McCusker RH, MacDonald RG. Expression of insulin-like growth factor-II and insulin-like growth factor binding proteins during Caco-2 cell proliferation and differentiation. J Cell Physiol 1996; 166:396-406. [PMID: 8592000 DOI: 10.1002/(sici)1097-4652(199602)166:2<396::aid-jcp18>3.0.co;2-9] [Citation(s) in RCA: 24] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
The components of the insulin-like growth factor (IGF) axis and their roles in regulating proliferation and differentiation of the human colon adenocarcinoma cell line, Caco-2, have been investigated. Caco-2 cells proliferated in serum-free medium at 75% the rate observed in medium containing 10% fetal bovine serum. IGF-I (10 nM) increased Caco-2 cell growth in serum-free medium, but not to the rate seen with serum. Multiple IGF-II mRNA species were produced by Caco-2 cells, but IGF-I mRNA was undetectable. Secretion of radioimmunoassayable IGF-II corresponded with steady-state levels of IGF-II mRNA, neither of which was observed to change markedly over the course of 16 days of Caco-2 cell differentiation. Levels of sucrase-isomaltase mRNA, a marker for enterocytic differentiation, increased 12-fold between days 5 and 16 of culture. Northern blotting of total RNA and ligand blot and immunoblot analyses of serum-free conditioned medium revealed that Caco-2 cells produce several IGF binding proteins (IGFBPs), including IGFBP-2, -3, and -4, as well as a 31,000 M(r) species that was not identified. The pattern of IGFBP secretion changed dramatically during Caco-2 cell differentiation: IGFBP-3 and IGFBP-2 increased 8.5-fold and 5-fold, respectively, whereas IGFBP-4 and the 31,000 M(r) species decreased 43% and 90%. Caco-2 cell clones stably transfected with a human IGFBP-4 cDNA construct exhibited a 60% increase in steady-state level of IGFBP-4 mRNA, and secreted twice as much IGFBP-4 protein as controls. Moreover, IGFBP-4-overexpressing cells proliferated at only 25% the rate of control cells in serum-free medium, in conjunction with a 70% increase in expression of sucrase-isomaltase. In summary, these studies indicate that a complex IGF axis is involved in autocrine regulation of Caco-2 cell proliferation and differentiation.
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Affiliation(s)
- J H Park
- Department of Pediatrics, Creighton University, Omaha, Nebraska, USA
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31
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Bai JP, Hsu MJ, Shier WT. Insulin-degrading enzyme in a human colon adenocarcinoma cell line (Caco-2). Pharm Res 1995; 12:513-7. [PMID: 7596985 DOI: 10.1023/a:1016241610649] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/26/2023]
Abstract
The activity of insulin-degrading enzyme (IDE), a thiol metalloprotease degrading insulin in many insulin target cells, was determined in human colon adenocarcinoma (Caco-2) cells. Insulin-degrading activity was localized in the cytosol of Caco-2 cells, accounting for 88% of total activity. Western blots and immunoprecipitation showed that IDE was present in the cytosol of Caco-2 cells and contributed to more than 93% cytosolic insulin-degrading activity. Cytosolic insulin degradation was strongly inhibited by IDE inhibitors, including N-ethylmaleimide, 1,10-phenanthroline, p-chloromericuribenzoate, and EDTA, but was not significantly or not as extensively inhibited by strong inhibitors of proteasome, i.e., chymostatin, soybean trypsin inhibitor, leupeptin, and Dip-F. These results suggest that IDE is present in Caco-2 cells, that Caco-2 IDE has properties similar to those of its counterparts in insulin-target tissues, and that it significantly contributes to intracellular insulin degradation.
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Affiliation(s)
- J P Bai
- Department of Pharmaceutics, College of Pharmacy, University of Minnesota, Minneapolis 55455, USA
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32
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Abstract
Some factors related to Westernization or industrialization increase risk of colon cancer. It is believed widely that this increase in risk is related to the direct effects of dietary fat and fiber in the colonic lumen. However, the fat and fiber hypotheses, at least as originally formulated, do not explain adequately many emerging findings from recent epidemiologic studies. An alternative hypothesis, that hyperinsulinemia promotes colon carcinogenesis, is presented here. Insulin is an important growth factor of colonic epithelial cells and is a mitogen of tumor cell growth in vitro. Epidemiologic evidence supporting the insulin/colon-cancer hypothesis is largely indirect and based on the similarity of factors which produce elevated insulin levels with those related to colon cancer risk. Specifically, obesity--particularly central obesity, physical inactivity, and possibly a low dietary polyunsaturated fat to saturated fat ratio--are major determinants of insulin resistance and hyperinsulinemia, and appear related to colon cancer risk. Moreover, a diet high in refined carbohydrates and low in water-soluble fiber, which is associated with an increased risk of colon cancer, causes rapid intestinal absorption of glucose into the blood leading to postprandial hyperinsulinemia. The combination of insulin resistance and high glycemic load produces particularly high insulin levels. Thus, hyperinsulinemia may explain why obesity, physical inactivity, and a diet low in fruits and vegetables and high in red meat and extensively processed foods, all common in the West, increase colon cancer risk.
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Affiliation(s)
- E Giovannucci
- Channing Laboratory, Department of Medicine, Harvard Medical School, Boston, MA 02115, USA
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