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Adnan M, Siddiqui AJ, Hamadou WS, Snoussi M, Badraoui R, Ashraf SA, Jamal A, Awadelkareem AM, Sachidanandan M, Hadi S, Khan MA, Patel M. Deciphering the Molecular Mechanism Responsible for Efficiently Inhibiting Metastasis of Human Non-Small Cell Lung and Colorectal Cancer Cells Targeting the Matrix Metalloproteinases by Selaginella repanda. PLANTS (BASEL, SWITZERLAND) 2021; 10:979. [PMID: 34068885 PMCID: PMC8156211 DOI: 10.3390/plants10050979] [Citation(s) in RCA: 13] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 03/28/2021] [Revised: 05/07/2021] [Accepted: 05/08/2021] [Indexed: 12/23/2022]
Abstract
Selaginella species are known to have antimicrobial, antioxidant, anti-inflammatory, anti-diabetic as well as anticancer effects. However, no study has examined the cytotoxic and anti-metastatic efficacy of Selaginella repanda (S. repanda) to date. Therefore, this study aimed to evaluate the potential anti-metastatic properties of ethanol crude extract of S. repanda in human non-small-cell lung (A-549) and colorectal cancer (HCT-116) cells with possible mechanisms. Effect of S. repanda crude extract on the growth, adhesion, migration and invasion of the A-549 and HCT-116 were investigated. We demonstrated that S. repanda crude extract inhibited cell growth of metastatic cells in a dose and time dependent manner. Incubation of A-549 and HCT-116 cells with 100-500 µg/mL of S. repanda crude extract significantly inhibited cell adhesion to gelatin coated surface. In the migration and invasion assay, S. repanda crude extract also significantly inhibited cellular migration and invasion in both A-549 and HCT-116 cells. Moreover, reverse transcription-polymerase chain reaction, and real-time PCR (RT-PCR) analysis revealed that the activity and mRNA level of matrix metalloproteinase-9 (MMP-9), matrix metalloproteinase-2 (MMP-2) and membrane type 1-matrix metalloproteinase (MT1-MMP) were inhibited. While the activity of tissue inhibitor matrix metalloproteinase 1 (TIMP-1); an inhibitor of MMPs was stimulated by S. repanda crude extract in a concentration-dependent manner. Therefore, the present study not only indicated the inhibition of motility and invasion of malignant cells by S. repanda, but also revealed that such effects were likely associated with the decrease in MMP-2/-9 expression of both A-549 and HCT-116 cells. This further suggests that S. repanda could be used as a potential source of anti-metastasis agent in pharmaceutical development for cancer therapy.
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Affiliation(s)
- Mohd Adnan
- Department of Biology, College of Science, University of Hail, Hail P.O. Box 2440, Saudi Arabia; (M.A.); (A.J.S.); (W.S.H.); (M.S.); (R.B.); (A.J.)
| | - Arif Jamal Siddiqui
- Department of Biology, College of Science, University of Hail, Hail P.O. Box 2440, Saudi Arabia; (M.A.); (A.J.S.); (W.S.H.); (M.S.); (R.B.); (A.J.)
| | - Walid Sabri Hamadou
- Department of Biology, College of Science, University of Hail, Hail P.O. Box 2440, Saudi Arabia; (M.A.); (A.J.S.); (W.S.H.); (M.S.); (R.B.); (A.J.)
| | - Mejdi Snoussi
- Department of Biology, College of Science, University of Hail, Hail P.O. Box 2440, Saudi Arabia; (M.A.); (A.J.S.); (W.S.H.); (M.S.); (R.B.); (A.J.)
| | - Riadh Badraoui
- Department of Biology, College of Science, University of Hail, Hail P.O. Box 2440, Saudi Arabia; (M.A.); (A.J.S.); (W.S.H.); (M.S.); (R.B.); (A.J.)
- Section of Histology-Cytology, Medicine Faculty of Tunis, University of Tunis El Manar, La Rabta-Tunis 1007, Tunisia
| | - Syed Amir Ashraf
- Department of Clinical Nutrition, College of Applied Medial Sciences, University of Hail, Hail P.O. Box 2440, Saudi Arabia; (S.A.A.); (A.M.A.)
| | - Arshad Jamal
- Department of Biology, College of Science, University of Hail, Hail P.O. Box 2440, Saudi Arabia; (M.A.); (A.J.S.); (W.S.H.); (M.S.); (R.B.); (A.J.)
| | - Amir Mahgoub Awadelkareem
- Department of Clinical Nutrition, College of Applied Medial Sciences, University of Hail, Hail P.O. Box 2440, Saudi Arabia; (S.A.A.); (A.M.A.)
| | - Manojkumar Sachidanandan
- Department of Oral Radiology, College of Dentistry, University of Hail, Hail P.O. Box 2440, Saudi Arabia;
| | - Sibte Hadi
- School of Forensic and Applied Sciences, University of Central Lancashire, Preston PR1 2HE, UK;
| | - Mushtaq Ahmad Khan
- Department of Microbiology and Immunology, College of Medicine and Health Sciences, UAE University, Al Ain 17666, United Arab Emirates
| | - Mitesh Patel
- Bapalal Vaidya Botanical Research Centre, Department of Biosciences, Veer Narmad South Gujarat University, Surat 394230, India
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Pattanayak S, Acharya R, Mishra N, Kumar A, Bose P, Pattnaik A, Mukhopadhyay K, Sunita P. Naringin, a natural flavonone glycoside attenuates N-nitrosodiethylamine- induced hepatocellular carcinoma in sprague-dawley rats. Pharmacogn Mag 2021. [DOI: 10.4103/pm.pm_94_21] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/04/2022] Open
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Okazaki I, Noro T, Tsutsui N, Yamanouchi E, Kuroda H, Nakano M, Yokomori H, Inagaki Y. Fibrogenesis and Carcinogenesis in Nonalcoholic Steatohepatitis (NASH): Involvement of Matrix Metalloproteinases (MMPs) and Tissue Inhibitors of Metalloproteinase (TIMPs). Cancers (Basel) 2014; 6:1220-55. [PMID: 24978432 PMCID: PMC4190539 DOI: 10.3390/cancers6031220] [Citation(s) in RCA: 41] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/07/2014] [Revised: 04/24/2014] [Accepted: 05/15/2014] [Indexed: 01/18/2023] Open
Abstract
Nonalcoholic steatohepatitis (NASH) is emerging worldwide because life-styles have changed to include much over-eating and less physical activity. The clinical and pathophysiological features of NASH are very different from those of HBV- and HCV-chronic liver diseases. The prognosis of NASH is worse among those with nonalcoholic fatty liver diseases (NAFLD), and some NASH patients show HCC with or without cirrhosis. In the present review we discuss fibrogenesis and the relationship between fibrosis and HCC occurrence in NASH to clarify the role of MMPs and TIMPs in both mechanisms. Previously we proposed MMP and TIMP expression in the multi-step occurrence of HCC from the literature based on viral-derived HCC. We introduce again these expressions during hepatocarcinogenesis and compare them to those in NASH-derived HCC, although the relationship with hepatic stem/progenitor cells (HPCs) invasion remains unknown. Signal transduction of MMPs and TIMPs is also discussed because it is valuable for the prevention and treatment of NASH and NASH-derived HCC.
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Affiliation(s)
- Isao Okazaki
- Department of Internal Medicine, Sanno Hospital, International University of Health and Welfare, Tokyo 107-0052, Japan.
| | - Takuji Noro
- Department of Surgery, International University of Health and Welfare Hospital, Tochigi 329-2763, Japan.
| | - Nobuhiro Tsutsui
- Department of Surgery, International University of Health and Welfare Hospital, Tochigi 329-2763, Japan.
| | - Eigoro Yamanouchi
- Department of Radiology, International University of Health and Welfare Hospital, Tochigi 329-2763, Japan.
| | - Hajime Kuroda
- Department of Pathology, International University of Health and Welfare Hospital, Tochigi 329-2763, Japan.
| | - Masayuki Nakano
- Department of Pathology, Ofuna Chuo Hospital, Kanagawa 247-0056, Japan.
| | - Hiroaki Yokomori
- Department of Internal Medicine, Kitasato University Medical Center, Saitama 364-8501, Japan.
| | - Yutaka Inagaki
- Department of Regenerative Medicine, Tokai University School of Medicine and Institute of Medical Sciences, Isehara 259-1193, Japan.
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Byambaragchaa M, de la Cruz J, Yang SH, Hwang SG. Anti-metastatic potential of ethanol extract of Saussurea involucrata against hepatic cancer in vitro. Asian Pac J Cancer Prev 2014; 14:5397-402. [PMID: 24175833 DOI: 10.7314/apjcp.2013.14.9.5397] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/10/2022] Open
Abstract
The rates of morbidity and mortality of hepatocellular carcinoma (HCC) have not lessened because of difficulty in treating tumor metastasis. Mongolian Saussurea involucrata (SIE) possesses various anticancer activities, including apoptosis and cell cycle arrest. However, detailed effects and molecular mechanisms of SIE on metastasis are unclear. Thus, the present study was undertaken to investigate antimetastatic effects on HCC cells as well as possible mechanisms. Effects of SIE on the growth, adhesion, migration, aggregation and invasion of the SK-Hep1 human HCC cell line were investigated. SIE inhibited cell growth of metastatic cells in dose- and time-dependent manners. Incubation of SK-Hep1 cells with 200-400 μg/mL of SIE significantly inhibited cell adhesion to gelatin-coated substrate. In the migration (wound healing) and aggregation assays, SIE treated cells showed lower levels than untreated cells. Invasion assays revealed that SIE treatment inhibited cell invasion capacity of HCC cells substantially. Quantitative real time PCR showed inhibitory effects of SIE on MMP-2/-9 and MT1-MMP mRNA levels, and stimulatory effects on TIMP-1, an inhibitor of MMPs. The present study not only demonstrated that invasion and motility of cancer cells were inhibited by SIE, but also indicated that such effects were likely associated with the decrease in MMP-2/-9 expression of SK-Hep1 cells. From these results, it was suggested that SIE could be used as potential anti-tumor agent.
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Affiliation(s)
- Munkhzaya Byambaragchaa
- Division of Animal Life and Environmental Science, Hankyong National University, Anseong, Philippines E-mail :
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Bayo J, Fiore E, Aquino JB, Malvicini M, Rizzo M, Peixoto E, Andriani O, Alaniz L, Piccioni F, Bolontrade M, Podhajcer O, Garcia MG, Mazzolini G. Increased migration of human mesenchymal stromal cells by autocrine motility factor (AMF) resulted in enhanced recruitment towards hepatocellular carcinoma. PLoS One 2014; 9:e95171. [PMID: 24736611 PMCID: PMC3988162 DOI: 10.1371/journal.pone.0095171] [Citation(s) in RCA: 39] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/04/2013] [Accepted: 03/24/2014] [Indexed: 12/26/2022] Open
Abstract
Background and Aims Several reports described the migration of human mesenchymal stromal cells (MSCs) towards tumor-released factors. Autocrine motility factor (AMF) is produced by several tumors including hepatocellular carcinoma (HCC). The aim of this study was to analyze AMF involvement on MSC migration towards human HCC. Methods Production of AMF by HCC tumors was evaluated by western analysis. The effects of AMF on MSCs from different sources (bone marrow, adipose tissue and perivascular cells from umbilical cord) were analyzed using in vitro migration assay; metalloproteinase 2 (MMP2) activity and expression of critical genes were studied by zymography and qRT-PCR, respectively. To assess AMF involvement on the in vivo MSC migration, noninvasive fluorescence imaging was performed. To test the effect of AMF-primed MSCs on tumor development, in vitro proliferation and spheroids growth and in vivo tumor volume were evaluated. Results AMF produced by HCC was found to induce migration of different MSCs in vitro and to enhance their MMP2 activity. Stimulation of MSCs with recombinant AMF (rAMF) also induced the in vitro adhesion to endothelial cells in coincidence with changes in the expression levels of MMP3, AMF receptor, caveolin-1, and -2 and GDI-2. Importantly, stimulation of MSCs with rAMF increased the in vivo migration of MSCs towards experimental HCC tumors. AMF-priming of MSCs did not induce a pro-tumorigenic effect on HCC cells neither in vivo nor in vitro. Conclusion AMF plays a role in MSC recruitment towards HCC. However, its ability to increase MSC migration to HCC for therapeutic purposes merits further evaluation.
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Affiliation(s)
- Juan Bayo
- Gene Therapy Laboratory, Facultad de Ciencias Biomédicas, Universidad Austral, Derqui-Pilar, Buenos Aires, Argentina
| | - Esteban Fiore
- Gene Therapy Laboratory, Facultad de Ciencias Biomédicas, Universidad Austral, Derqui-Pilar, Buenos Aires, Argentina
| | - Jorge B. Aquino
- Gene Therapy Laboratory, Facultad de Ciencias Biomédicas, Universidad Austral, Derqui-Pilar, Buenos Aires, Argentina
- CONICET (Consejo Nacional de Investigaciones Científicas y Técnicas), Buenos Aires, Argentina
| | - Mariana Malvicini
- Gene Therapy Laboratory, Facultad de Ciencias Biomédicas, Universidad Austral, Derqui-Pilar, Buenos Aires, Argentina
| | - Manglio Rizzo
- Gene Therapy Laboratory, Facultad de Ciencias Biomédicas, Universidad Austral, Derqui-Pilar, Buenos Aires, Argentina
| | - Estanislao Peixoto
- Gene Therapy Laboratory, Facultad de Ciencias Biomédicas, Universidad Austral, Derqui-Pilar, Buenos Aires, Argentina
| | - Oscar Andriani
- Liver Unit, Hospital Universitario Austral, Universidad Austral, Derqui-Pilar, Argentina
| | - Laura Alaniz
- Gene Therapy Laboratory, Facultad de Ciencias Biomédicas, Universidad Austral, Derqui-Pilar, Buenos Aires, Argentina
- CONICET (Consejo Nacional de Investigaciones Científicas y Técnicas), Buenos Aires, Argentina
| | - Flavia Piccioni
- Gene Therapy Laboratory, Facultad de Ciencias Biomédicas, Universidad Austral, Derqui-Pilar, Buenos Aires, Argentina
| | - Marcela Bolontrade
- CONICET (Consejo Nacional de Investigaciones Científicas y Técnicas), Buenos Aires, Argentina
- Molecular and Cellular Therapy Laboratory, Fundación Instituto Leloir, Buenos Aires, Argentina
| | - Osvaldo Podhajcer
- CONICET (Consejo Nacional de Investigaciones Científicas y Técnicas), Buenos Aires, Argentina
- Molecular and Cellular Therapy Laboratory, Fundación Instituto Leloir, Buenos Aires, Argentina
| | - Mariana G. Garcia
- Gene Therapy Laboratory, Facultad de Ciencias Biomédicas, Universidad Austral, Derqui-Pilar, Buenos Aires, Argentina
- CONICET (Consejo Nacional de Investigaciones Científicas y Técnicas), Buenos Aires, Argentina
| | - Guillermo Mazzolini
- Gene Therapy Laboratory, Facultad de Ciencias Biomédicas, Universidad Austral, Derqui-Pilar, Buenos Aires, Argentina
- CONICET (Consejo Nacional de Investigaciones Científicas y Técnicas), Buenos Aires, Argentina
- Liver Unit, Hospital Universitario Austral, Universidad Austral, Derqui-Pilar, Argentina
- * E-mail:
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Abstract
Matrix metalloproteinases (MMPs) were originally identified as matrixin proteases that act in the extracellular matrix. Recent works have uncovered nontraditional roles for MMPs in the extracellular space as well as in the cytosol and nucleus. There is strong evidence that subspecialized and compartmentalized matrixins participate in many physiological and pathological cellular processes, in which they can act as both degradative and regulatory proteases. In this review, we discuss the transcriptional and translational control of matrixin expression, their regulation of intracellular sorting, and the structural basis of activation and inhibition. In particular, we highlight the emerging roles of various matrixin forms in diseases. The activity of matrix metalloproteinases is regulated at several levels, including enzyme activation, inhibition, complex formation and compartmentalization. Most MMPs are secreted and have their function in the extracellular environment. MMPs are also found inside cells, both in the nucleus, cytosol and organelles. The role of intracellular located MMPs is still poorly understood, although recent studies have unraveled some of their functions. The localization, activation and activity of MMPs are regulated by their interactions with other proteins, proteoglycan core proteins and / or their glycosaminoglycan chains, as well as other molecules. Complexes formed between MMPs and various molecules may also include interactions with noncatalytic sites. Such exosites are regions involved in substrate processing, localized outside the active site, and are potential binding sites of specific MMP inhibitors. Knowledge about regulation of MMP activity is essential for understanding various physiological processes and pathogenesis of diseases, as well as for the development of new MMP targeting drugs.
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Affiliation(s)
- Ferdinando Mannello
- Department of Biomolecular Sciences, Section of Clinical Biochemistry, Unit of Cell Biology, University Carlo Bo of Urbino, Via O. Ubaldini 7, 61029 Urbino (PU), Italy.
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Ho CC, Lai KC, Hsu SC, Kuo CL, Ma CY, Lin ML, Yang JS, Chung JG. Benzyl isothiocyanate (BITC) inhibits migration and invasion of human gastric cancer AGS cells via suppressing ERK signal pathways. Hum Exp Toxicol 2010; 30:296-306. [PMID: 20498032 DOI: 10.1177/0960327110371991] [Citation(s) in RCA: 43] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022]
Abstract
Metastasis suppressors and associated other regulators of cell motility play a critical initial role in tumor invasion and metastases. Benzyl isothiocyanate (BITC) is a hydrolysis compound of glucotropaeolin in dietary cruciferous vegetables. BITC has been found to exhibit prevention of cancers in laboratory animals and might also be chemoprotective in humans. Here, the purpose of this study was to investigate the effects of BITC on cell proliferation, migration, invasion and mitogen-activated protein kinase (MAPK) pathways of AGS human gastric cancer cells. Wound healing and Boyden chamber (migration and invasion) assays demonstrated that BITC exhibited an inhibitory effect on the abilities of migration and invasion in AGS cancer cells. BITC suppressed cell migration and invasion of AGS cells in a dose-dependent manner. Results from Western blotting indicated that BITC exerted an inhibitory effect on the ERK1/2, Ras, GRB2, Rho A, iNOS, COX-2 for causing the inhibitions of MMP-2, -7 and -9 then followed by the inhibitions of invasion and migration of AGS cells in vitro. BITC also promoted MKK7, MEKK3, c-jun, JNK1/2, VEGF, Sos1, phosphoinositide 3-kinase (PI3K), PKC, nuclear factor-kappaB (NF-κB) p65 in AGS cells. Results from real-time polymerized chain reaction (PCR) showed that BITC inhibited the gene expressions of MMP-2,-7 -9, FAK, ROCK1 and RhoA after BITC treatment for 24 and 48 hours in AGS cells. Taken together, the finding may provide new mechanisms and functions of BITC, which inhibit migration and invasion of human gastric cancer AGS cells.
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Affiliation(s)
- Chin-Chin Ho
- Department of Nursing, Central Taiwan University of Science and Technology, Taichung, Taiwan
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Hwang ES, Lee HJ. Benzyl isothiocyanate inhibits metalloproteinase-2/-9 expression by suppressing the mitogen-activated protein kinase in SK-Hep1 human hepatoma cells. Food Chem Toxicol 2008; 46:2358-64. [PMID: 18502015 DOI: 10.1016/j.fct.2008.03.016] [Citation(s) in RCA: 32] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/03/2006] [Revised: 03/17/2008] [Accepted: 03/17/2008] [Indexed: 11/20/2022]
Abstract
Benzyl isothiocyanate (BITC) is a hydrolysis compound of glucotropaeolin in cruciferous vegetables. Many studies have reported that BITC prevents cancers in laboratory animals and might also be chemoprotective in humans. The purpose of this study was to investigate the effects of BITC on cell proliferation, metastasis, and MAPK pathways of SK-Hep1 human hepatocellular carcinoma cells. BITC suppressed SK-Hep1 cell proliferation in a dose-dependent manner, and exposure to 1 and 5 microM BITC reduced cell proliferation by 25% and 30%, respectively. The expression of matrix metalloproteinase (MMP)-2, MMP-9, and membrane type-1/MMP (MT-1/MMP) is a known risk factor for metastatic disease. Gelatin zymography analysis revealed a significant downregulation of MMP-2/-9 protein expression in SK-Hep1 cells treated with 0.1-5 microM BITC. BITC treatment caused dose-dependent decreases in MMP-2/-9 and MT1-MMP mRNA levels as determined by RT-PCR. BITC also increased the mRNA levels of tissue inhibitors of matrix metalloproteinases-2 (TIMP-2) 1.3- and 1.5-fold after a 24 h exposure to 1 and 5 microM BITC, respectively. Increased TIMP-2 expression is mediated by the downregulation of MMP-2 and MT1-MMP. BITC inhibited the phosphorylation activities of all three major mitogen-activated protein kinases (MAPKs) in a dose-dependent manner. BITC at 5 microM reduced the ERK1/2 phosphorylation activity by 50% and p38 activity by 70%. BITC also reduced the p-JNK1/2 level by 30% and 70% at 1 and 5 microM treatments, respectively. These data may represent anti-metastatic activities of BITC through the suppression of MAPKs in SK-Hep1 cells.
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Affiliation(s)
- Eun-Sun Hwang
- Center for Agricultural Biomaterials, College of Agriculture and Life Sciences, Seoul National University, 599 Gwanangno, Gwanak-gu, Seoul 151-921, Republic of Korea
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Marongiu F, Doratiotto S, Montisci S, Pani P, Laconi E. Liver repopulation and carcinogenesis: two sides of the same coin? THE AMERICAN JOURNAL OF PATHOLOGY 2008; 172:857-64. [PMID: 18321999 DOI: 10.2353/ajpath.2008.070910] [Citation(s) in RCA: 41] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/17/2022]
Abstract
Liver repopulation by transplanted normal hepatocytes has been described in a number of experimental settings. Extensive repopulation can also occur from the selective proliferation of endogenous normal hepatocytes, both in experimental animals and in the human liver. This review highlights the intriguing association between clinical and experimental conditions related to liver repopulation and an increased risk for development of hepatocellular carcinoma. It is suggested that any microenvironment that is able to sustain the clonal growth of normal transplanted (or endogenous) hepatocytes is also geared to select for the emergence of rare resistant cells with an altered phenotype. Whereas the first pathway leads to liver repopulation with normal histology, the latter results in the growth of focal proliferative lesions and carries an increased risk of neoplastic disease. The implications of this association are discussed, both in terms of pathogenetic significance and possible therapeutic exploitation.
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Affiliation(s)
- Fabio Marongiu
- Dipartimento di Scienze e Tecnologie Biomediche, Sezione di Patologia Sperimentale, Università di Cagliari, Cagliari, Italy
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Csepregi A, Röcken C, Hoffmann J, Gu P, Saliger S, Müller O, Schneider-Stock R, Kutzner N, Roessner A, Malfertheiner P, Ebert MPA. APC promoter methylation and protein expression in hepatocellular carcinoma. J Cancer Res Clin Oncol 2007; 15:1415-28. [PMID: 20629990 PMCID: PMC4373337 DOI: 10.1111/j.1582-4934.2010.01124.x] [Citation(s) in RCA: 52] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022]
Abstract
PURPOSE We investigated the impact of promoter methylation on APC protein expression in patients with hepatocellular carcinoma (HCC). MATERIALS AND METHODS 50 patients [HCC (n=19), liver metastasis (n=19), cholangiocellular cancer (n=7), and benign liver tumors (n=5)] were studied for methylation using Methylight analysis. APC mutation was investigated by protein truncation test and direct sequencing of genomic DNA. The protein expression was evaluated by immunohistochemistry and Western blot analysis. RESULTS The APC promoter was hypermethylated in 81.8% of non-cancerous liver tissue samples. All HCC samples and ten patients with liver metastasis (52.6%) exhibited APC promoter methylation. The degree of methylation was significantly higher in samples from HCC compared to the non-cancerous liver tissue samples (63.1% vs. 24.98%; p=0.001). The level of APC protein expression was significantly reduced in HCC samples compared to that of the corresponding non-tumor liver tissue (p<0.05). CONCLUSIONS Promoter methylation of the APC gene seems to be of significance in hepatocarcinogenesis and results in reduced protein expression in HCC. Interestingly, APC promoter methylation is also present in the vast majority of non-cancerous liver tissue whose (patho)physiological function remains unresolved.
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Affiliation(s)
- Antal Csepregi
- Department of Gastroenterology, Hepatology, and Infectious Diseases, Otto-von-Guericke University, Leipziger Strasse 44, 39120 Magdeburg, Germany.
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Mamori S, Nagatsuma K, Matsuura T, Ohkawa K, Hano H, Fukunaga M, Matsushima M, Masui Y, Fushiya N, Onoda H, Searashi Y, Takagi I, Tagiri H. Useful detection of CD147 (EMMPRIN) for pathological diagnosis of early hepatocellular carcinoma in needle biopsy samples. World J Gastroenterol 2007; 13:2913-7. [PMID: 17589939 PMCID: PMC4171141 DOI: 10.3748/wjg.v13.i21.2913] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To make clear whether CD147 (EMMPRIN) expression in pathological tumor samples with a fine-needle aspiration biopsy is useful for pathological diagnosis of early hepatocellular carcinoma (HCC).
METHODS: Twenty-two patients (15 men and 7 women; median age 68 years, range 56-81 years) underwent a liver tissue biopsy in order to make a diagnosis of HCC. Paraffin-embedded liver biopsy tissue samples from 22 patients were stained with anti-CD147 antibody, murine monoclonal antibody 12C3 (MAb12C3) for immunohistochemical analysis. An immunohistochemical analysis of CD147 was performed and the degree of staining compared between tumor and non-tumor tissue. In addition, the degree of staining within tumor tissue was compared according to a number of clinicopathological variables.
RESULTS: The degree of staining of CD147 was significantly higher in tumor tissues than non-tumor tissues, even in tumors less than 15 mm in diameter. The expression of this protein was significantly elevated in HCC tissue specimens from patients with a low value of serum AST and γ-GTP.
CONCLUSION: CD147 serves potentially as a pathological target for cancer detection of early HCC.
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Affiliation(s)
- Satoshi Mamori
- Division of Gastroenterology and Hepatology, Department of Internal Medicine, The Jikei University School of Medicine, 3-25-8 Nishi-shinbashi, Minato-ku, Tokyo 105-8461, Japan.
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Ku XM, Liao CG, Li Y, Yang XM, Yang B, Yao XY, Wang L, Kong LM, Zhao P, Chen ZN. Epitope mapping of series of monoclonal antibodies against the hepatocellular carcinoma-associated antigen HAb18G/CD147. Scand J Immunol 2007; 65:435-43. [PMID: 17444954 DOI: 10.1111/j.1365-3083.2007.01930.x] [Citation(s) in RCA: 30] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/23/2022]
Abstract
The hepatocellular carcinoma-associated antigen HAb18G/CD147, a member of CD147 family, could promote tumour invasion and metastasis via inducing the secretion of matrix metalloproteinases (MMP). Anti-CD147 monoclonal antibodies (MoAb) have exhibited obvious inhibitory effect on MMP induction. However, none of the epitopes of these MoAb has been reported. We previously prepared five MoAb against HAb18G/CD147, named HAb18, 3B3, 1B3, 5A5 and 4D2. To map the epitopes of these MoAb, a series of truncated fragments of extracellular region of HAb18G/CD147 was expressed in Escherichia coli and the MoAb-binding affinity to these fragments was examined with an enzyme-linked immunosorbent assay and Western blot. The residues (39)LTCSLNDSATEV(50), (36)KILLTCS(42) and (22)AAGTVFTTVEDL(33) were determined to be the epitopes of HAb18, 3B3 and 1B3, respectively, which were further proved by a dot-blot analysis with synthesized peptides and bioinformatics epitope prediction. The binding regions of MoAb 5A5 and 4D2 were located at residues E(120)-R(203). Then we constructed and expressed full-length HAb18G/CD147 and truncated HAb18G/CD147 without residues A(22)-V(50) in COS-7 cells. Gelatin zymography and Boyden chamber assay showed that the COS-7 cells expressing truncated HAb18G/CD147 failed to induce MMP production and enhance the cells' invasive potential, compared with the cells expressing full-length HAb18G/CD147. Taken together with the obviously inhibitory effects of HAb18 on the function of full-length HAb18G/CD147, these findings suggest that residues (22)AAGTVFTTVEDLGSKILLTCSLNDSATEV(50) may play a critical role in the functions of HAb18G/CD147 on MMP secretion and tumour invasion. These key residues can be used as potential drug target in cancer therapy.
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Affiliation(s)
- X-M Ku
- Cell Engineering Research Center & Department of Cell Biology, State Key Laboratory of Cancer Biology, The Fourth Military Medical University, Xi'an, China
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Ip YC, Cheung ST, Fan ST. Atypical localization of membrane type 1-matrix metalloproteinase in the nucleus is associated with aggressive features of hepatocellular carcinoma. Mol Carcinog 2007; 46:225-30. [PMID: 17219425 DOI: 10.1002/mc.20270] [Citation(s) in RCA: 54] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/10/2022]
Abstract
Membrane type 1-matrix metalloproteinase (MT1-MMP) is a versatile proteinase and recent studies indicated it could be internalized. Our earlier study found that it is overexpressed in hepatocellular carcinoma (HCC) and could promote intrahepatic metastasis. The present study was conducted to examine its subcellular localization and its clinicopathological significance in HCC after curative partial hepatectomy. Localization of MT1-MMP in 101 pairs of HCCs and their adjacent liver tissues, and 8 normal liver tissues was examined by the immunohistochemical method. MT1-MMP protein was localized at membrane and cytoplasm of hepatocytes in the normal and tumor adjacent liver tissues. In contrast, the HCCs were highly heterogeneous with variable degrees of membrane, cytoplasmic, and even nuclear staining. Interestingly, patients with presence of nuclear MT1-MMP were associated with poor overall survival (log-rank test, P=0.043) and large tumor size (>5 cm) (Fisher's exact test, P=0.031). Subcellular distribution was further demonstrated by Western blotting and immunofluorescence with Hep3B stable transfectant overexpressing MT1-MMP. Western blot analyses of subcellular fractions confirmed a differential partitioning of various post-translationally modified MT1-MMP in these fractions. Different antibodies corroborated the presence of MT1-MMP in the nuclear fraction. Concomitant nuclear presence of MMP2 with MT1-MMP further indicated its potential involvement in the nuclear functions. MT1-MMP co-localized with caveolin-1 at the perinuclear region, suggesting nuclear translocation of MT1-MMP via caveolae-mediated endocytosis. In summary, the association of nuclear MT1-MMP with aggressive tumor features including poor prognosis and large tumor expands its functional repertoire and further indicates a new functional role of MMPs within nuclei of tumor cells.
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Affiliation(s)
- Ying Chi Ip
- Department of Surgery, Centre for the Study of Liver Disease, The University of Hong Kong, Pokfulam, Hong Kong, China
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14
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Tsai WC, Chao YC, Lee WH, Chen A, Sheu LF, Jin JS. Increasing EMMPRIN and matriptase expression in hepatocellular carcinoma: tissue microarray analysis of immunohistochemical scores with clinicopathological parameters. Histopathology 2006; 49:388-95. [PMID: 16978202 DOI: 10.1111/j.1365-2559.2006.02516.x] [Citation(s) in RCA: 42] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/26/2022]
Abstract
AIMS To examine the expression of extracellular matrix metalloprotease inducer (EMMPRIN) and matriptase in hepatocellular carcinoma (HCC) and to correlate this with tumour progression. METHODS AND RESULTS Immunohistochemical analysis of EMMPRIN and matriptase was performed on tissue microarrays of 122 cases of HCC with various histological grades and/or clinical parameters. The expression of EMMPRIN and matriptase was undetectable in normal liver parenchyma of all eight control cases. However, among the 122 HCC cases, EMMPRIN and matriptase immunoreactivity was seen on the cell membrane and in the cytoplasm. The average immunostaining scores of EMMPRIN were 88 for grade I HCC, 195 for grade II HCC and 293 for grade III HCC. Of 85 HCC cases in 122 with detailed clinical TNM stages, the average immunostaining scores of EMMPRIN were 75 for stage T1, 177 for stage T2, 260 for stage T3 and 313 for stage T4 cases of HCC. In addition, the average immunostaining scores of matriptase were 84 for grade I HCC, 187 for grade II HCC, 302 for grade III HCC, and 72 for stage T1, 181 for stage T2, 224 for stage T3 and 284 for stage T4 cases of HCC. More advanced M and N stages of HCC were associated with higher intensity, greater percentages of tumour staining and immunostaining scores of EMMPRIN and matriptase. Higher EMMPRIN and matriptase immunostaining scores in HCCs also correlated significantly with tumour grading and TNM stages. CONCLUSIONS Our findings demonstrate for the first time that EMMPRIN and matriptase are overexpressed in HCC. These may be novel biomarkers for the diagnosis and treatment of HCC.
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Affiliation(s)
- W-C Tsai
- Department of Pathology and Division of Gastroenterology, Tri-Service General Hospital, National Defence Medical Centre, Taipei, Taiwan, R.O.C
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15
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Terada T, Iwai M, Kawakami S, Yamashita F, Hashida M. Novel PEG-matrix metalloproteinase-2 cleavable peptide-lipid containing galactosylated liposomes for hepatocellular carcinoma-selective targeting. J Control Release 2006; 111:333-42. [PMID: 16488046 DOI: 10.1016/j.jconrel.2005.12.023] [Citation(s) in RCA: 134] [Impact Index Per Article: 7.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/07/2005] [Revised: 12/26/2005] [Accepted: 12/26/2005] [Indexed: 01/18/2023]
Abstract
In order to obtain an HCC-selective drug delivery system, a novel functional lipid, which is cleaved by the protease activity of matrix metalloproteinase-2 (MMP-2), was developed. The amino group of dioleoylphosphatidylethanolamine (DOPE) was conjugated with PEGylated MMP-2 substrate peptide (Gly-Pro-Leu-Gly-Ile-Ala-Gly-Gln), and MMP-2-cleavable PEG-Peptide-DOPE (PEG-PD) was synthesized. When PEG-PD was incorporated in galactosylated liposomes (Gal-PEG-PD-liposomes), we expected that Gal-PEG-PD-liposomes would not be taken up by normal hepatocytes due to the steric hindrance effect, but would be activated around HCC cells by secreted MMPs. In the pretreatment by hMMP2 (1, 5, and 10mug/ml), an hMMP2 concentration-dependent higher uptake of Gal-PEG-PD-liposomes was observed in HepG2 cells, suggesting PEG-PD cleavage. In the presence of an excess of galactose, the uptake of Gal-PEG-PD-liposomes with hMMP2 was significantly inhibited, suggesting asialoglycoprotein receptor-mediated uptake of Gal-PEG-PD-liposomes following the PEG-PD cleavage. Pretreatment of Gal-PEG-PD-liposomes with the conditioned medium of B16BL6, which contained secreted MMPs, enhanced the binding to HepG2 cells, as in the case of hMMP-2 treatment. Moreover, the cytotoxicity of N(4)-octadecyl-1-beta-d-arabinofuranosylcytosine (NOAC) incorporated Gal-PEG-PD-liposomes was enhanced by hMMPs (5mug/ml) and its cytotoxicity was significantly reduced by the presence of an excess of galactose in HepG2 cells. In conclusion, Gal-PEG-PD-liposomes were successfully developed for novel HCC-selective targeting.
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Affiliation(s)
- Takeshi Terada
- Department of Drug Delivery Research, Graduate School of Pharmaceutical Sciences, Kyoto University, Kyoto 606-8501, Japan
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16
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Huang Y, Jiang J, Dou K, Chen Z. HAb18G/CD147 enhances the secretion of matrix metalloproteinases (MMP) via cGMP/NO-sensitive capacitative calcium entry (CCE) and accordingly attenuates adhesion ability of fibroblasts. Eur J Cell Biol 2005; 84:59-73. [PMID: 15724816 DOI: 10.1016/j.ejcb.2004.09.003] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/25/2022] Open
Abstract
The present study examined the effect of hepatoma-associated antigen HAb18G (homologous to CD147) expression on the NO/cGMP-regulated Ca2+ mobilization to induce matrix metalloproteinases (MMP) production and attenuate adhesion ability of mouse fibroblast NIH/3T3 cells. HAb18G/CD147 cDNA was transfected into fibroblast 3T3 cells to obtain a cell line stably expressing HAb18G/CD147, t3T3, as demonstrated by immunofluorescence staining and flow cytometry assays. 8-Bromo-cGMP inhibited the thapsigargin-induced Ca2+ entry in 3T3 cells, whereas an inhibitor of protein kinase G, KT5823 (1 microM), led to an increase in Ca2+ entry. Expression of HAb18G/CD147 in t3T3 cells decreased the inhibitory response to cGMP. A similar effect on the Ca2+ entry was observed in 3T3 cells in response to an NO donor, (+/-)-S-nitroso-N-acetylpenicillamine (SNAP). The inhibitory effect of SNAP on the thapsigargin-induced Ca2+ entry was also reduced in HAb18G/CD147-expressing t3T3 cells, indicating a role for HAb18G/CD 147 in NO/cGMP-regulated Ca2+ entry. Results of gelatin zymography assays showed that addition of extracellular Ca2+ induced MMP (MMP-2, MMP-9) release and activation in a dose-dependent manner, and expression of HAb18G/CD147 enhanced the secretion of MMP-2 and MMP-9 in 3T3 cells. 8-Bromo-cGMP and SNAP reduced the production of MMP in 3T3 cells but not in t3T3 with HAb18G/CD147 expression. RT-PCR experiments substantiated that the expression of MMP-2 and MMP-9 mRNA in HAb18G/CD 147-expressing t3T3 cell was significantly greater than that in 3T3 cells. Experiments investigating adhesion potentials demonstrated that HAb18G/CD147-expressing t3T3 cells pretreated with Ca2+ attached to Matrigel-coated culture plates significantly less efficiently than 3T3 cells. The proportion of attached cells could be increased by treatment with 8-bromo-cGMP and SNAP in 3T3 cells, but not in t3T3. These results suggest that HAb18G/CD147 attenuates adhesion potentials in fibroblasts by enhancing the secretion of MMP through NO/cGMP-sensitive capacitative Ca2+ entry.
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Affiliation(s)
- Yong Huang
- Cell Engineering Research Centre & Department of Cell Biology, The Fourth Military Medical University, Xi'an 710032, PR China
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Affiliation(s)
- David Semela
- Institute of Clinical Pharmacology, University of Bern, 35 Murtenstrasse, Bern CH-3010, Switzerland
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Iijima T, Minami Y, Nakamura N, Onizuka M, Morishita Y, Inadome Y, Noguchi M. MMP-2 activation and stepwise progression of pulmonary adenocarcinoma: Analysis of MMP-2 and MMP-9 with gelatin zymography. Pathol Int 2004; 54:295-301. [PMID: 15086833 DOI: 10.1111/j.1440-1827.2004.01623.x] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Abstract
Small pulmonary adenocarcinomas can be classified on the basis of their histological characteristics and prognosis, and when classified as such, the prognosis of replacing-type adenocarcinoma with active fibroblast proliferation is significantly worse than adenocarcinoma without fibroblast proliferation. In order to clarify the biological mechanisms of the key to the morphological changes associated with active fibroblast proliferation, we examined the activities of matrix metalloproteinase (MMP)-2 and MMP-9, which are important enzymes in the stromal invasion by cancers. The active MMP-2 and MMP-9 content of 40 pulmonary adenocarcinomas that were less than 20 mm in diameter was measured by the gelatin zymography method. The quantity of active MMP-2 in the pulmonary adenocarcinomas with active fibroblast proliferation was higher than in the pulmonary adenocarcinomas without proliferation (P < 0.001), but there were no correlations between the histological features and the activation of MMP-9. The presence of active fibroblast proliferation in small pulmonary adenocarcinomas suggests that the cancer cells have acquired the ability to invade through the action of active MMP-2, and this is thought to be one of the reasons for the worse prognosis of pulmonary adenocarcinoma with active fibroblast proliferation.
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Affiliation(s)
- Tatsuo Iijima
- Department of Pathology, Institute of Basic Medical Science, University of Tsukuka, Japan
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19
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Yu FL, Liao MH, Lee JW, Shih WL. Induction of hepatoma cells migration by phosphoglucose isomerase/autocrine motility factor through the upregulation of matrix metalloproteinase-3. Biochem Biophys Res Commun 2004; 314:76-82. [PMID: 14715248 DOI: 10.1016/j.bbrc.2003.12.056] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/26/2022]
Abstract
Phosphoglucose isomerase/autocrine motility factor (PGI/AMF) catalyzes the isomerization between glucose-6-phosphate and fructose-6-phosphate, and is involved in cytokine activity, mitogenesis, differentiation, oncogenesis, and tumor metastasis. Presently, we demonstrate that exogenous PGI/AMF stimulates the migration of Huh7 and HepG2 hepatoma cells, but not Hep3B cells. Inhibition of PGI/AMF by PGI/AMF specific inhibitor 5-phospho-D-arabinonate markedly repressed the cellular migration. RT-PCR was used to examine the expression profile of matrix metalloproteinases (MMPs). MMP-3 transcripts, protein level, and secreted form were significantly upregulated in PGI/AMF-treated Huh7 and HepG2 cells, but not in Hep3B cells. MMP-3 inhibition abolished the PGI/AMF-induced cell motility. The observations are consistent with a downstream mediation role of MMP-3 in PGI/AMF-stimulated tumor cell metastasis.
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Affiliation(s)
- Feng-Ling Yu
- Department of Veterinary Medicine, National Pingtung University of Science and Technology, Pingtung, Taiwan, ROC
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20
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21
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Hsu C, Chen CN, Chen LT, Wu CY, Yang PM, Lai MY, Lee PH, Cheng AL. Low-Dose Thalidomide Treatment for Advanced Hepatocellular Carcinoma. Oncology 2003; 65:242-9. [PMID: 14657598 DOI: 10.1159/000074477] [Citation(s) in RCA: 76] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022]
Abstract
OBJECTIVE To analyze the efficacy of oral thalidomide in the treatment of advanced hepatocellular carcinoma (HCC). METHODS Sixty-eight patients with unresectable and nonembolizable HCC were consecutively enrolled in a compassionate treatment program of oral thalidomide. Tumor response and treatment-related toxicity were prospectively followed. Thalidomide was given at a starting dose of 200 mg per day. The dose was gradually escalated in 100-mg steps up to 600 mg per day if no limiting toxicities developed. RESULTS Sixty-three patients were evaluable for response. One complete and 3 partial responses, defined by World Health Organization criteria, were seen, with a response rate of 6.3% (95% CI 0-12.5). The duration of response was 50+, 24.6, 11.6+ and 8.7+ weeks, respectively. All 4 responders had a dramatic decrease in alpha-fetoprotein (alpha-FP) levels. Another 6 of the 42 patients with elevated alpha-FP levels before treatment had a more than 50% decrease in their alpha-FP levels after thalidomide treatment. Totally 10 patients had an objective response to thalidomide. The median overall survival for all of the 68 patients was 18.7 weeks (95% CI 11.8- 25.6) with a 1-year survival rate of 27.6%. The median overall survival of the 10 patients with an objective response to thalidomide was 62.4 weeks (95% CI 31.2-93.6 weeks). All responders responded at a dose equal to or less than 300 mg per day. Toxicities of thalidomide were generally manageable, and only 16, 6, and 0 patients developed grade 2, 3, and 4 toxicities, respectively. CONCLUSION Low-dose thalidomide is safe and induces unequivocal tumor response in a minority of patients with advanced HCC.
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Affiliation(s)
- Chiun Hsu
- Department of Oncology, National Taiwan University Hospital, Taipei, Taiwan, ROC
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22
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Zhang Z, Fang SG, Gao Y, Jiang ZS, Sun EW. Effect of batimastat on Matrix metallopro-teinase-2 mRNA in rat hepatocellular carcinoma. Shijie Huaren Xiaohua Zazhi 2003; 11:716-718. [DOI: 10.11569/wcjd.v11.i6.716] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM To study the dynamic changes of matrix metalloproteinase-2 mRNA in liver tissue during the experimental hepatocarcin-ogenesis and the effect of Batimastat on it.
METHODS Hepatocellular carcinoma was induced with the administration of diethy Initrosoamine(DENA) in rats. BB-94 was intraperitoneally injected to treat the experimental models. Reverse transcription polymerase chain reaction was used for quantitative analysis of MMP-2 mRNA expression level during the induction and therapy.
RESULTS The expression of MMP-2mRNA was increased throughout the hepatocarcinogenesis, and reach had its maximum plateau in the early hepatocarcinogenesis stage. BB-94 no effect on MMP-2mRNA expression.
CONCLUSION MMP-2mRNA expression is increasing during hepatocarcinogenesis, and BB-94 has no effect on MMP-2mRNA expression.
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Affiliation(s)
- Zhi Zhang
- Department of Organ Transplantation, Zhujiang Hospital, First Military Medical University, Guangzhou 510282, Guangdong Province China
| | - Shi-Gang Fang
- Department of general surgery, Zhujiang Hospital, First Military Medical University, Guangzhou 510282, Guangdong Province China
| | - Yi Gao
- Department of Organ Transplantation, Zhujiang Hospital, First Military Medical University, Guangzhou 510282, Guangdong Province China
| | - Ze-Sheng Jiang
- Department of Organ Transplantation, Zhujiang Hospital, First Military Medical University, Guangzhou 510282, Guangdong Province China
| | - Er-Wei Sun
- Department of Organ Transplantation, Zhujiang Hospital, First Military Medical University, Guangzhou 510282, Guangdong Province China
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Schoedel KE, Tyner VZ, Kim TH, Michalopoulos GK, Mars WM. HGF, MET, and matrix-related proteases in hepatocellular carcinoma, fibrolamellar variant, cirrhotic and normal liver. Mod Pathol 2003; 16:14-21. [PMID: 12527708 DOI: 10.1097/01.mp.0000043521.96995.db] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022]
Abstract
Fibrolamellar variant is an uncommon subcategory of hepatocellular carcinoma with a better prognostic outcome. Proteinases and growth factors that are involved in the remodeling of extracellular matrix may influence the behavior of cancers. To determine whether these factors contribute to the distinct etiologies of fibrolamellar hepatocellular carcinoma and traditional hepatocellular carcinoma, we assayed hepatocyte growth factor, the hepatocyte growth factor receptor, and two hepatocyte growth factor activators, hepatocyte growth factor activator and urokinase-type plasminogen activator, in hepatocellular carcinoma, fibrolamellar hepatocellular carcinoma, cirrhotic liver and normal liver. In addition, we examined the urokinase-type plasminogen activator receptor, the type 1 plasminogen activator inhibitor, plasmin, fibrinogen, and the type IV matrix metalloproteinases. Eighteen hepatocellular carcinomas and 11 fibrolamellar hepatocellular carcinomas were obtained as paraffin embedded sections from the University of Pittsburgh Department of Pathology. Frozen tissues from a subset of cases (9 hepatocellular carcinomas, 4 fibrolamellar hepatocellular carcinomas, 12 cirrhotic livers and 2 normal livers) were also available for analysis. Antibodies against urokinase-type plasminogen activator, urokinase-type plasminogen activator receptor, hepatocyte growth factor and hepatocyte growth factor receptor were used to analyze immunoperoxidase stained slides from the paraffin blocks. Western blot analyses using antibodies against hepatocyte growth factor, hepatocyte growth factor receptor, phosphotyrosine, hepatocyte growth factor activator, urokinase-type plasminogen activator receptor, urokinase-type plasminogen activator, plasminogen activator inhibitor-1, fibrinogen and plasmin were performed on membrane-enriched fractions from the frozen tissue, as was collagen zymography for matrix metalloproteinase-2 and matrix metalloproteinase-9. The most notable findings are as follows: hepatocyte growth factor activator was only detected in malignant tissue but not cirrhotic liver or normal liver. Although hepatocyte growth factor was detected in most samples, it was significantly elevated in 5/9 hepatocellular carcinomas. Furthermore, 8/9 fibrolamellar hepatocellular carcinomas demonstrated hepatocyte growth factor receptor levels similar to normal, whereas 8/9 hepatocellular carcinomas and 11/12 cirrhotic livers exhibited either an increase or decrease. In contrast, active matrix metalloproteinase-2, which was absent in normal liver, was elevated in fibrolamellar hepatocellular carcinoma as compared to cirrhotic liver and conventional hepatocellular carcinoma. Surprisingly, 10/12 cirrhotic livers and 2/4 fibrolamellar hepatocellular carcinomas but only 1/9 hepatocellular carcinomas were enriched for plasmin. The combined data suggest that the hepatocyte growth factor and plasmin systems tend to be operative in hepatocellular carcinoma and cirrhotic liver, more than fibrolamellar hepatocellular carcinoma. Furthermore, matrix turnover appears to be a more prominent feature of fibrolamellar hepatocellular carcinoma. These findings provide insight into the behavioral differences between hepatocellular carcinoma and the fibrolamellar variant.
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Affiliation(s)
- Karen E Schoedel
- Department of Pathology, University of Pittsburgh Medical Center, Pittsburgh, Pennsylvania 15261, USA
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24
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Giannelli G, Bergamini C, Marinosci F, Fransvea E, Quaranta M, Lupo L, Schiraldi O, Antonaci S. Clinical role of MMP-2/TIMP-2 imbalance in hepatocellular carcinoma. Int J Cancer 2002; 97:425-31. [PMID: 11802202 DOI: 10.1002/ijc.1635] [Citation(s) in RCA: 144] [Impact Index Per Article: 6.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022]
Abstract
An imbalance between the proteolytic activity of matrix metalloproteinase-2 (MMP-2) and the tissue inhibitor of MMP-2 (TIMP-2) is responsible for degradation of extracellular matrix (ECM) components and plays a critical role in tumor invasion and in metastasis formation. The occurrence of intra-hepatic metastasis, which severely affects prognosis and long-term survival, is commonly observed in the course of hepatocellular carcinoma (HCC). We investigated the expression of MT1-MMP in tissues, whereas both MMP-2 and TIMP-2 were evaluated in the sera and tissues (primary and metastatic nodules) of HCC patients with and without metastasis, whose clinical outcome was followed over a 2-year period. MT1-MMP expression was similar among primary nodule tissues of patients with and without metastasis. Serum and tissue levels of MMP-2 were not statistically different between patients with and without metastasis, but MMP-2 was concentrated at the invasive edge of the metastatic tissue. On the contrary, serum and tissue levels of TIMP-2 were significantly higher in HCC patients without metastasis than in those with. This situation was not only observed in the primary HCC tissues, but also in the metastatic nodules. These results correlate with the clinical outcome, because more than 90% of the patients with high levels of TIMP-2 were still alive after 2 years, whereas less than 30% with low levels of TIMP-2 had survived. Furthermore, we found a strict correlation between tissue and serum levels of TIMP-2, this suggesting that a MMP-2/TIMP-2 imbalance and in particular TIMP-2 levels, could represent an important prognostic factor in patients with HCC.
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Affiliation(s)
- Gianluigi Giannelli
- Department of Internal Medicine, Immunology and Infectious Diseases, Section of Internal Medicine, University of Bari Medical School, Bari, Italy.
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Yano H, Hara A, Murase S, Hayashi K, Ando H, Shinoda J, Shimokawa K, Sakai N. Expression of hepatocyte growth factor and matrix metalloproteinase-2 in human glioma. Brain Tumor Pathol 2002; 18:7-12. [PMID: 11517976 DOI: 10.1007/bf02478919] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/06/2023]
Abstract
Hepatocyte growth factor (HGF) has a stimulatory effect on the synthesis of matrix metalloproteinase-2 (MMP-2), which is involved in glioma invasion. In this study, to clarify the correlation between the expression of HGF and MMP-2 in glioma tissues, immunohistochemical analysis of HGF and MMP-2 was performed in 11 cases of astrocytoma, 10 cases of anaplastic astrocytoma, and 9 cases of glioblastoma. As a result, expression of HGF and MMP-2 was correlated with the grade of malignancy (P = 0.0181 and 0.0001, respectively), and a significant correlation between the immunoreactivity of HGF and that of MMP-2 was observed (P < 0.05). Immunofluorescence study revealed the concomitant expression of HGF and MMP-2 in glioma tissue. In cultured glioma cell lines (SNB-19, U87MG, and U373MG), exogenous recombinant HGF effectively acted on the production of the active and latent forms of MMP-2 protein in a dose-dependent manner. Active MMP-2 increased more effectively than the latent form. Taken together, these results suggest that HGF may promote glioma invasion in vivo by production of MMP-2.
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Affiliation(s)
- H Yano
- Department of Neurosurgery, Gifu University School of Medicine, Japan.
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Jiang JL, Zhou Q, Yu MK, Ho LS, Chen ZN, Chan HC. The involvement of HAb18G/CD147 in regulation of store-operated calcium entry and metastasis of human hepatoma cells. J Biol Chem 2001; 276:46870-7. [PMID: 11591720 DOI: 10.1074/jbc.m108291200] [Citation(s) in RCA: 106] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/29/2023] Open
Abstract
The present study examined the effect of hepatoma-associated antigen HAb18G (homologous to CD147) expression on the NO/cGMP-regulated Ca(2+) mobilization and metastatic process of human hepatoma cells. HAb18G/CD147 cDNA was transfected into human 7721 hepatoma cells to obtain a cell line stably expressing HAb18G/CD147, T7721, as demonstrated by Northern blot and immunocytochemical studies. 8-Bromo-cGMP (cGMP) inhibited the thapsigargin-induced Ca(2+) entry in a concentration-dependent manner in 7721 cells. The cGMP-induced inhibition was abolished by an inhibitor of protein kinase G, KT5823 (1 microm). However, expression of HAb18G/CD147 in T7721 cells decreased the inhibitory response to cGMP. A similar concentration-dependent inhibitory effect on the Ca(2+) entry was observed in 7721 cells in response to a NO donor, (+/-)-S-nitroso-N-acetylpenicillamine (SNAP). The inhibitory effect of SNAP on the thapsigargin-induced Ca(2+) entry was significantly reduced in HAb18G/CD147-expressing T7721 cells, indicating a role for HAb18G/CD147 in NO/cGMP-regulated Ca(2+) entry. Experiments investigating metastatic potentials demonstrated that HAb18G/CD147-expressing T7721 cells attached to the Matrigel-coated culture plates and invaded through Matrigel-coated permeable filters at the rate significantly greater than that observed in 7721 cells. Both the attachment and invasion rates could be suppressed by SNAP, and the inhibitory effect of SNAP could be reversed by NO inhibitor, N(G)-nitro-l-arginine methyl ester. The sensitivity of the attachment and invasion rates to cGMP was significantly reduced in T7721 cells as compared with 7721 cells when cells were pretreated with thapsigargin. The difference in the sensitivity between the two cells could be abolished by a Ca(2+) channel blocker, Ni(2+) (3 mm). These results suggest that HAb18G/CD147 enhances metastatic potentials in human hepatoma cells by disrupting the regulation of store-operated Ca(2+) entry by NO/cGMP.
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Affiliation(s)
- J L Jiang
- Epithelial Cell Biology Research Center, Department of Physiology, Faculty of Medicine, The Chinese University of Hong Kong, Shatin, NT, Hong Kong, China
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Kumaki F, Matsui K, Kawai T, Ozeki Y, Yu ZX, Ferrans VJ, Travis WD. Expression of matrix metalloproteinases in invasive pulmonary adenocarcinoma with bronchioloalveolar component and atypical adenomatous hyperplasia. THE AMERICAN JOURNAL OF PATHOLOGY 2001; 159:2125-35. [PMID: 11733363 PMCID: PMC1850594 DOI: 10.1016/s0002-9440(10)63064-7] [Citation(s) in RCA: 46] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 10/18/2022]
Abstract
To investigate the association between the expression of matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) and the clinicopathological features in lepidic and invasive components of adenocarcinoma of the lung, we performed immunostaining for type IV collagen, various MMPs, and TIMPs in 27 cases of invasive adenocarcinomas and 5 cases of atypical adenomatous hyperplasia of alveolar epithelial cells (AAH). Mean extent of lepidic growth was 61% and the survival was significantly better in cases with 50% or more lepidic component. The preservation of type IV collagen in lepidic areas correlated inversely with lymphatic or vascular invasion (P = 0.02 and 0.002, respectively). Five-year survival was reduced in cases showing destruction of type IV collagen (P = 0.004) or expression of MMP-2 (P = 0.008) in lepidic areas. MMP-2 co-localized with MT-1-MMP (its activating enzyme) and TIMP-2 in neoplastic cells. Reactivity for other MMPs and TIMPs did not correlate with destruction of type IV collagen or prognosis. Type IV collagen was preserved in all cases of AAH. MMP-2, but not MT-1-MMP, was expressed in two of the five cases of AAH. Immunostaining for type IV collagen MMP-2 is useful in evaluating the prognosis of the lung.
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Affiliation(s)
- Fumiyuki Kumaki
- National Defense Medical College, Tokorozawa, Saitama, Japan; the Pathology Section,†
| | - Kazuhiro Matsui
- National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland; and the Department of Pulmonary and Mediastinal Pathology,‡
| | - Toshiaki Kawai
- National Defense Medical College, Tokorozawa, Saitama, Japan; the Pathology Section,†
| | - Yuichi Ozeki
- National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland; and the Department of Pulmonary and Mediastinal Pathology,‡
| | - Zu-Xi Yu
- National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland; and the Department of Pulmonary and Mediastinal Pathology,‡
| | - Victor J. Ferrans
- National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland; and the Department of Pulmonary and Mediastinal Pathology,‡
| | - William D. Travis
- Armed Forces Institute of Pathology, Washington, District of Columbia
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Torimura T, Ueno T, Kin M, Harad R, Nakamura T, Sakamoto M, Kumashiro R, Yano H, Kojiro M, Sata M. Laminin deposition to type IV collagen enhances haptotaxis, chemokinesis, and adhesion of hepatoma cells through beta1-integrins. J Hepatol 2001; 35:245-53. [PMID: 11580147 DOI: 10.1016/s0168-8278(01)00127-1] [Citation(s) in RCA: 20] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/04/2022]
Abstract
BACKGROUND/AIMS In hepatocellular carcinoma, laminin deposition to type IV collagen along the sinusoids is observed with the development of arterial network, coinciding with intrahepatic metastasis. We investigated the influence of laminin deposition to type IV collagen on hepatoma cell adhesion, motility and secretion of matrix metalloproteinases (MMPs), which are indispensable behaviors for tumor metastasis. METHODS Hepatoma cell lines (KYN-1, -2 and -3) were used. The expression of integrin subunit mRNAs in hepatoma cells was confirmed by RT-PCR. The influence of laminin addition to type IV collagen on the adhesion, chemokinesis, and migration of KYN-1, -2 and -3 was evaluated by the haptotactic migration, phagokinetic track motility, and cell adhesion assays. The effects of integrin subunits on these activities were evaluated using the function-blocking antibodies for integrins. Phosphorylation of MEK1/2 and secretion of MMPs were investigated by Western blotting and gelatin zymography. RESULTS Integrin alpha1, alpha2, alpha3, alpha6 and beta1 subunit mRNAs were detected. The combination of type IV collagen and laminin enhanced the migration, chemokinesis, and adhesion of hepatoma cells compared to that of type IV collagen when used alone. The enhanced activity was significantly suppressed by function-blocking antibodies for integrin alpha1, alpha2, alpha3, alpha6 and beta1 subunits. Hepatoma cells cultured on the combination of type IV collagen and laminin showed phosphorylation of MEK1/2 and increased secretion of MMPs. CONCLUSIONS The addition of laminin to type IV collagen enhances hepatoma cell adhesion and motility through beta1-integrins.
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Affiliation(s)
- T Torimura
- The Second Department of Medicine, Kurume University School of Medicine, Japan.
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Tang ZY. Hepatocellular carcinoma--cause, treatment and metastasis. World J Gastroenterol 2001; 7:445-54. [PMID: 11819809 PMCID: PMC4688653 DOI: 10.3748/wjg.v7.i4.445] [Citation(s) in RCA: 303] [Impact Index Per Article: 12.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/13/2001] [Revised: 07/20/2001] [Accepted: 07/27/2001] [Indexed: 02/06/2023] Open
Abstract
In the recent decades, the incidence of hepatocellular carcinoma (HCC) has been found to be increasing in males in some countries. In China, HCC ranked second of cancer mortality since 1990s. Hepatitis B and C viruses (HBV and HCV) and dietary aflatoxin intake remain the major causative factors of HCC. Surgery plays a major role in the treatment of HCC, particularly for small HCC. Down-staging unresectable huge HCC to smaller HCC and followed by resection will probably be a new approach for further study. Liver transplantation is indicated for small HCC, however, some issues remain to be solved. Different modes of regional cancer therapy for HCC have been tried. Systemic chemotherapy has been disappointing in the past but the future can be promising. Biotherapy, such as cytokines, differentiation inducers, anti-angiogenic agents, gene therapy and tumor vaccine will probably play a role, particularly in the prevention of tumor recurrence. HCC invasiveness is currently the major target of study. Tremendous works have been done at the molecular level, which will provide clues for biomarker of HCC progression as well as targets for intervention.
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Affiliation(s)
- Z Y Tang
- Liver Cancer Institute of Fudan University, 136 Yixueyuan Road, Zhongshan Hospital, Shanghai 200032, China.
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Akalin I, Güllü IH, Kurdoglu M, Marangoz S. Why hepatocellular carcinoma cells are unlikely to metastasize: is there a role for tissue inhibitor of metalloproteinase-1? Med Hypotheses 2001; 57:221-3. [PMID: 11461177 DOI: 10.1054/mehy.2001.1271] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022]
Abstract
The question 'Why hepatocellular carcinoma cells are unlikely to metastasize although they have a high proliferative activity?' is a major point of interest from a cancer physiopathological viewpoint. Recent articles about the roles and relationships of some cytokines with matrix degrading enzymes and their inhibitors in various types of normal tissues and malignancies give rise to another question: 'Does tissue inhibitor of metalloproteinase-1 prevent the extrahepatic metastasis of hepatocellular carcinoma cells?' On the basis of many evidences, it is highly probable that under the effect of a possible inducing mechanism of the cytokines interleukin-6, -1 beta and transforming growth factor beta, the increase in concentration of tissue inhibitor of metalloproteinase-1 in hepatocellular carcinoma cause increased type I collagen accumulation and consequent prevention of cellular detachment, which explains why highly proliferative malignant hepatocytes have less metastatic ability.
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Affiliation(s)
- I Akalin
- Institute of Oncology, Hacettepe University, Ankara, Turkey.
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Duivenvoorden WC, Hirte HW, Singh G. Quantification of matrix metalloproteinase activity in plasma of patients enrolled in a BAY 12-9566 phase I study. Int J Cancer 2001; 91:857-62. [PMID: 11275992 DOI: 10.1002/1097-0215(200002)9999:9999<::aid-ijc1135>3.0.co;2-m] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/24/2022]
Abstract
The expression of matrix metalloproteinases (MMPs) is often associated with invasiveness or grade of tumours. Increased blood levels of MMP proteins, including MMP-1, MMP-2, MMP-3 and MMP-9 have been detected in various types of cancers. With the exception of one study, MMPs in serum and plasma have been determined using ELISA. In the present study we measured the activity of the MMPs found in human plasma samples using gelatin enzymography and fluorimetric degradation assays. We used plasma samples from healthy control subjects and cancer patients enrolled in a dose-finding study for the MMP inhibitor, BAY 12-9566, to assess the activity of MMPs found in plasma and screen for efficacy of the MMP inhibitor. BAY 12-9566 has inhibitory activity toward MMP-2, MMP-3 and MMP-9. Patients with advanced solid tumours were enrolled in our study and plasma was collected on day 1 before dosing and at steady-state of the drug on day 15. Our results show that BAY 12-9566 was effective in lowering the plasma gelatinolytic activity in the group of 29 patients when considering the data obtained from a fluorimetric gelatinase assay. The data obtained from gelatin enzymography, however, did not reach significance. The fluorimetric degradation assay could be a useful tool to screen plasma from cancer patients in other clinical trials assessing MMP inhibitors.
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Affiliation(s)
- W C Duivenvoorden
- Hamilton Regional Cancer Centre, McMaster University, 699 Concession Street, Hamilton, Ontario, Canada L8V 5C2
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