EXPRESSION OF MATRIX METALLOPROTEINASE-7 AND FAS LIGAND: THEIR APOPTOSIS-INDUCING EFFECT ON GASTRIC CANCER CELLS

Objective: To investigate the expression of matrix metalloproteinase-7 (MMP-7) and Fas ligand (FasL) in gastric cancer and explore their role in progression of gastric cancer. Methods: Formalin-fixed paraffin and embedded tissues of primary gastric cancer and adjacent non-tumor mucosa from 113 cases were evaluated for MMP-7, FasL and Capase-3 expression by streptavidin-peroxidase (S-P) immunohistochemistry. The expression of the first two proteins in cancer cells of primary foci was compared with clinicopathological parameters of tumors. We also observed the correlation of MMP-7 and FasL expression with Caspase-3 expression in cancer cells of primary foci. Results: MMP-7 positive immunostaining was less frequently detected in adjacent epithelial cells than in cancer cells of primary foci of gastric cancer (P<0.05, 29.2% vs 69.0%), and so was FasL (P<0.05, 34.5% vs 54.0%). MMP-7 expression was associated with tumor size, Borrmann抯 classification, invasive depth, metastasis and TNM staging (P<0.05), but not with growth pattern, Lauren抯 classification, or histological classification (P>0.05). FasL expression was correlated with tumor size, invasive depth, metastasis, Lauren抯 classification, histological classification (P<0.05), while not with Borrmann抯 classification, TNM staging or growth pattern (P>0.05). Cancer cells of primary foci expressed less Caspase-3 than their adjacent epithelial cells (P<0.05, 32.7% vs 50.4%). There was an obvious correlation between FasL, MMP-7 and Caspase-3 expression in cancer cells of primary foci (P<0.05). Co-expression of MMP-7 and FasL paralleled with Caspase-3 expression in cancer cells of primary foci (P<0.05). Conclusion: MMP-7 and FasL expression was up-regulated in gastric carcinogenesis and was principally involved in progression of gastric cancer. FasL expression could reflect the differentiation of gastric cancer cells and underlie the molecular mechanisms of different pathways of gastric tumorigenesis. Co-expression of MMP-7 and FasL could have apoptosis-inducing effect on gastric cancer cells.

EXPRESSION OF MATRIX METALLOPROTEINASE-7 INVOLVING IN GROWTH, INVASION, METASTASIS AND ANGIOGENESIS OF GASTRIC CANCER

Objective. To investigate the role of matrix metalloproteinase-7 (MMP-7) expression in caricinogenesisand progression of gastric cancer.Methods. We studied MMP-7 expression and microvessel density (MVD) in adjacent mucosa and pri-mary foci of 113 cases of gastric cancer by streptavidin-biotin-immunoperoxidase method using anti-MMP-7 and anti-CD34 antibodies. MMP-7 expression and mean MVD were compared with clinicopatholog-ical features of gastric cancer, with the relationship between MMP-7 expression and MVD concerned in gastric cancer.Results. MMP-7 showed positive expression in adjacent mucosa of gastric cancer (29.20%, 33/113),less than that in gastric cancer (69.03%, 78/113). MMP-7 expression in primary foci of gastric cancerwas positively correlated with tumor size, invasive depth, metastasis and TNM staging (P＜0.05), but notwith differentiation or growth pattern of gastric cancer (P＞0.05). Positive correlation of mean MVD withtumor size, invasive depth, metastasis and TNM staging was found (P＜0.05), despite no relationshipbetween mean MVD and differentiation of gastric cancer (P＞0.05). Mean MVD was dependent on MMP-7expression in gastric cancer (P＜0.05).Conclusion. Up-regulated expression of MMP-7 played an important role in carcinogenesis and pro-gression by participating in growth, invasion, metastasis and angiogenesis of gastric cancer. MMP-7 ex-pression could be regarded as an effective and objective marker to reflect the biological behaviors of gas-tric cancer.

Expression of Matrix Metalloproteinase-7 in Serous Ovarian Tumors

Objective:To explore the expression of matrix metalloproteinase-7 in serous ovarian tumors. Methods: Expression of MMP- 1 in 6 normal ovaries, 12 serous cystadenomas of ovary, 6 borderline cystadenomas of ovary, and 22 serous cystadenocarcinomas of ovary were studied by immunohistoc/temical SP staining. Results: No expression of MMP- 1 was detected in normal ovaries. In most serous ovarian tumors, expression of MMP-7 was detected in both the cytoplasm of tumor cells and stroma, although it was reported in other tumors that MMP- 7 was mainly expressed in the cytoplasm of tumor cells. The expression level of MMP-7 in the cytoplasm of tumor cells was statistically insignificant between serous cystadenomas of ovary, borderline cystadenomas of ovary, and serous cystadenocarcinomas of ovary. But in the stroma, the expression level of MMP-7 in borderline cystadenomas of ovary and serous cystadenocarcinomas of ovary was significantly higher than that in serous cystadenomas of ovary (P<0. 05). In borderline cystadenomas and serous cystadenocarcinomas of ovary, expression of MMP-7 could also be detected in the nuclei of some tumor cells. Conclusion: MMP-7 may play an important role in the progression of serous ovarian tumors.

Autoantibodies against MMP-7 as a novel diagnostic biomarker in esophageal squamous cell carcinoma

AIM： To evaluate the diagnostic values of serum autoan tibodies against matrix metalloproteinase-7 （MMP-7） in patients with esophageal squamous cell carcinoma （ESCC）. METHODS： The MMP-7 cDNA was cloned from ESCC tissues, and MMP-7 was expressed and purified from a prokaryotic system. MMP-7 autoanUbodies were then measured in sera from 50 patients with primary ESCC and 58 risk-matched controls, using a reverse capture enzyme-linked immunosorbent assay （ELISA） in which autoantibodies to MMP-7 bound to the purified MMP-7 proteins. In addition, MMP-7 autoantibody levels in sera from 38 gastric cancer patients and from control serum samples were also tested. RESULTS： The optimum conditions for recombinant MMP-7 protein expression were determined as 0.04 mmol/L Isopropyl-13-D-Thiogalactopyranoside （IPTG）induction at 37℃ for four hours. The levels of serum autoantibodies against MMP-7 were significantly higher in patients with ESCC than in the matched-control samples （OD450 = 1.69 ±0.08 vs OD450 = 1.55 ± 0.10, P 〈 0.001）. The area under the receiver operating character- istic （ROC） curve was 0.87. The sensitivity and specificity for detection of ESCC were 78.0% and 81.0%, respectively, when the OD450 value was greater than 1.65. Although the levels of autoantibodies against MMP-7 were also significantly higher in patients with gastric cancer compared to control samples （OD450 = 1.62± 0.06 vs OD450 = 1.55±0.10, P 〈 0.001）, the diagnostic accuracy was less significant than in ESCC patients. The area of ROC curve was 0.75, whereas the sensitivity and specificity were 60.5% and 71.7%, respectively, when the cut-off value of OD450 was set at 1.60. CONCLUSION： Serum autoantibody levels of MMP-7 may be a good diagnostic biomarker for esophageal squamous cell carcinoma,

Prospective study of MMP7 serum levels in the diagnosis of cholangiocarcinoma

AIM: To determine whether the serum level of matrix metalloproteinase-7 (MMP7) has the potential to diagnosis cholangiocarcinoma from benign biliary tract diseases. METHODS: This study was performed according to the PRoBE (a prospective-specimen-collection, retrospectiveblinded-evaluation) design. A total of 187 patients with obstructive jaundice were consecutively enrolled. After the diagnostic status of these patients was ascertained, their levels of serum MMP7 were assayed and compared with serum carbohydrate antigen 19-9 (CA19-9). This was conducted in a blinded case (cholangiocarcinoma)control (benign biliary tract disease) setup. RESULTS: MMP7 and CA19-9 serum levels were significantly elevated in cholangiocarcinoma patients (P < 0.001). The area under the curve (AUC) from a receiver operating characteristic (ROC) curve analysis for thediagnosis of cholangiocarcinoma, using MMP7 was more accurate than CA19-9 (AUC = 0.84, 95% CI: 0.778-0.903 for MMP7 and AUC = 0.79, 95% CI: 0.708-0.868 for CA19-9). The sensitivity and specificity of serum MMP7 (cut-off value of 5.5 ng/mL) was 75% and 78%, respectively, while the sensitivity and specificity of serum CA19-9 (cut-off value of 100 U/mL) was 68% and 87%, respectively. CONCLUSION: Serum values of MMP7 and CA19-9 appear to be useful biomarkers for differentiating cholangiocarcinoma from benign biliary tract obstructive diseases.

Immunohistochemical molecular markers as predictors of curability of endoscopically resected submucosal colorectal cancer

AIM: To clarify the usefulness of immunohistochemical molecular markers in predicting lymph node metastasis of submucosal colorectal cancer. METHODS: We examined microvessel density, lymphatic vessel density, the Ki-67 labeling index, expression of MUC1 and Matrix metalloproteinase-7 (MMP-7) in tumor cells, and expression of cathepsin D in stromal cells at the invasive front by immunostaining of samples resected from 214 patients with submucosal colorectal cancer. Pathologic features were assessed on hematoxylin-eosin- stained samples. We evaluated the relations between clinicopathologic/immunohistochemical features and lymph node metastasis. RESULTS: Lesions of the superficial type, with an unfavorable histologic grade, budding, lymphatic involvement, high microvessel density (≥ 40), high lymphatic vessel density (≥ 9), high Ki-67 labeling index (≥ 42), and positivity of MUC1, cathepsin D, and MMP-7 showed a significantly high incidence of lymph node metastasis. Multivariate analysis revealed that high microvessel density, unfavorable histologic grade, cathepsin D positivity, high lymphatic vessel density, superficial type, budding, and MUC1 positivity were independent risk factors for lymph node metastasis.A combined examination with four independent immunohistochemical markers (microvessel density, cathepsin D, lymphatic vessel density, and MUC1) revealed that all lesions that were negative for all markers or positive for only one marker were negative for lymph node metastasis. CONCLUSION: Analysis of a combination of immuno- histochemical molecular markers in endoscopically resected specimens of submucosal colorectal cancer allows prediction of curability regardless of the pathologic features visible of hematoxylin-eosin-stained sections.

Pygopus 2 promotes kidney cancer OS-RC-2 cells proliferation and invasion in vitro and in vivo

Objective:Human Pygopus 2(Pygo2)was recently discovered to be a component of the Wnt signaling pathway required for b-catenin/Tcf-mediated transcription.But the role of Pygo2 in malignant cell proliferation and invasion has not yet been determined.Methods:Lentivirus-mediated small interfering RNA(siRNA)and vector-based overexpression were used to study the function of Pygo2 in OS-RC-2 cells.The resulted cells were subject to Western blotting assay,MTT assay,colony formation and cell invasion assays.Furthermore,renal cell carcinoma(RCC)models were established in BALB/c nude mice inoculated with OS-RC-2 cells.Immunohistochemistry(IHC)staining of matrix metalloproteinase-7(MMP-7),matrix metalloproteinase-9(MMP-9)and vascular endothelial growth factor(VEGF)was performed in tumor tissue.Results:Pygo2 gene was successful knocked down and overexpressed in RCC OS-RC-2 cells by using an shRNA and overexpressing vector,respectively.Overexpression of Pygo2 effectively promoted cell proliferation,colony formation and invasion in vitro.Knockdown of Pygo2 obviously inhibited xenograft tumor growth in nude mice.In addition,overexpression of Pygo2 increased the levels of MMP-7,MMP-9 and VEGF in the xenograft tumors.Conclusion:Pygo2 has a role in promoting cell proliferation,invasion and metastasis,and may regulate angiogenesis via the Wnt/b-catenin signaling pathway.

Effects of Weipixiao(胃痞消)on Wnt Pathway-Associated Proteins in Gastric Mucosal Epithelial Cells from Rats with Gastric Precancerous Lesions*

Objective： To study the effects of Weipixiao （胃痞消, WPX） on Wnt pathway-associated proteins in gastric mucosal epithelial cells from rats with gastric precancerous lesions （GPL）. Methods： Sprague Dawley rats were randomly divided into control, model, vitacoenzyme （0.2 g·kg-l·day-1）, WPX high-dose （H-WPX, 15 g·kg-l·day-1）, WPX medium-dose （M-WPX, 7.5 g·kg-1·day-1） and WPX low-dose （L-WPX, 3.75 g·kg-l·day-1） groups. After successfully establishing the GPL model, the rats were consecutively administered WPX or vitacoenzyme by gastrogavage for 10 weeks. Differential expression of Leucine-rich repeat-containing G-protein- coupled receptor 5 （Lgr5）, matrix metalloproteinase-7 （MMP-7）, Wntl, Wnt3a, and 13 -catenin in gastric mucosal epithelial cells in all groups were immunohistochemically detected, and the images were taken and analyzed semiquantitatively by image pro plus 6.0 software. Results： Gastric epithelium in the model group showed significantly higher expression levels of Lgr5, MMP-7, Wntl, Wnt3a and 13 -catenin than those of the control group （P〈0.01）. Interestingly, we also observed Lgr5＋ cells, which generally located at the base of the gastric glandular unit, migrated to the luminal side of gastric epithelium with GPL. The expression levels of Lgr5, MMP-7, Wntl, and 13-catenin were all down-regulated in the L-WPX group as compared with those of both model and vitacoenzyme groups （P〈0.05）. A similar, but nonsignificant down-regulation in expression level of Wnt3a was noted in all WPX groups （P〉0.05）. Conclusion： Our findings suggested that the therapeutic mechanisms of WPX in treating GPL might be related with its inhibitory effects on the expressions of Lgr5, MMP-7, Wntl, β -catenin and the aberrant activation of Wnt/β -catenin pathway.