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World J Gastroenterol 2003;9(7):1607-1610 World Journal of Gastroenterology Copyright © 2003 by The WJG Press ISSN 1007-9327
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Effect of P-selectin monoclonal antibody on metastasis of gastric cancer and immune function Jin-Lian Chen, Wei-Xiong Chen, Jin-Shui Zhu, Ni-Wei Chen, Tong Zhou, Ming Yao, Dong-Qing Zhang, Yun-Lin Wu Jin-Lian Chen, Wei-Xiong Chen, Jin-Shui Zhu, Ni-Wei Chen, Department of Gastroenterology, Shanghai Jiao-Tong University affiliated Sixth People’s Hospital, Shanghai 200233, China Tong Zhou, Yun-Lin Wu, Ruijin Hospital, Shanghai Second Medical University, Shanghai 200025, China Ming Yao, Shanghai Cancer Institute, Shanghai 200233, China Dong-Qing Zhang, Shanghai Institute of Immunology, Shanghai 200025, China Supported by the Youth Science Foundation of Shanghai Public Administration No.131984Y15 Correspondence to: Jin-Lian Chen, Department of Gastroenterology, Shanghai Sixth People’s Hospital, Shanghai Jiao-Tong University affiliated Sixth People’s Hospital, Shanghai 200233, China.
[email protected] Telephone: +86-21-64369181 Received: 2003-03-02 Accepted: 2003-03-29
Abstract AIM: To investigate the effect of cell adhesion molecule Pselectin monoclonal antibody (Mab) on metastasis and immune function of mice orthototopically implanted with human gastric cancer tissue. METHODS: SCID mice were implanted orthotopically with SGC-7901 human gastric carcinoma tissue. Starting from day 3 after operation, animals were given intravenously PBS or P-selectin Mab (100 µg/injection) (for both normal mice and tumor-implanted mice with tumors), twice weekly for 3 weeks. Two animals in each group were sacrificed randomly at the 1st, 2nd, 4th week and 6th week. While T cell and B cell transformation indices were determined with the 3H TdR infiltration method, the NK cell activity was detected by the LDH release method. RESULTS: The metastatic rate in the P-selectin Mab treated group was lower than that in the PBS treated group (with tumors). The NK activity of normal mice increased over time. The immune functions (T, B cell function, NK activity) of the tumor group in the 6th week were significantly lower than those in the 4th week, but the change was attenuated by Pselectin Mab. CONCLUSION: P-selectin Mab could suppress the metastasis of gastric cancer with no adverse effect on host immune function. Chen JL, Chen WX, Zhu JS, Chen NW, Zhou T, Yao M, Zhang DQ, Wu YL. Effect of P-selectin monoclonal antibody on metastasis of gastric cancer and immune function. World J Gastroenterol 2003; 9(7): 1607-1610
http://www.wjgnet.com/1007-9327/9/1607.asp
INTRODUCTION Gastric carcinoma is one of the most frequent tumors in China. Tumor metastasis is very common clinically. Cell adhesion molecules have been implicated to be crucial elements in the
process of metastasis[1-28]. P-selectin is an adhesion molecules that mediates the cell to cell interaction of platelets and endothelial cells with neutrophils and monocytes as well as tumor cells[29]. Our previous study indicates that P-selectin expression is related to aggressive behavior, dissemination and poor prognosis of human gastric carcinomas, and Pselectin monoclonal artibody can inhibit gastric carcinoma metastasis[30-32]. The present study was performed to investigate effects of P-selectin monoclonal antibody on metastasis and immune function in SCID mouse metastaic models of human gastric cancer constructed by orthotopic implantation of histologically intact tumor tissue.
MATERIALS AND METHODS Animal model Forty-eight male SCID mice obtained from Shanghai Cancer Institute were 7-8 weeks old with weight of 20-25 g. Human gastric cancer SGC-7901, a poorly-differentiated adenocarcinoma line, was originally derived from a primary tumor and maintained by passage in nude mice subcutaneously. SCID mice were randomly divided into experimental group (n=24) and normal group (n=24). Animal models in experimental group were made using orthotopic implantation of histologically intact tissue of human gastric carcinoma[33]. Tumors were resected aseptically. Necrotic tumor tissues were removed and the remaining non-necrotic tumor tissues were minced into pieces about 5-7 mm in diameter in Hank’s balanced salt solution. Each of the tumor pieces was weighed and adjusted to be 150 mg with scissors. Mice were anesthetized with 4.3 % trichloraldehyde hydrate and an incision was made through the left upper abdominal pararectal line and peritoneal cavity was carefully exposed and a part of the serosal membrane in the middle of the greater curvature of the glandular stomach was mechanically injured by using scissors. A tumor piece of 150 mg was fixed on each injured site of the serosal surface. The stomach was then returned to the peritoneal cavity, and the abdominal wall and skin were closed. 3 d later, all animals implanted with intact tumor tissues received i.v. injection of PBS (group3, n=12) or P-selectin antibody (Suzhou Medical College; 100 µg/injectiom; group4, n=12) twice weekly for 3 weeks. Animals in normal group received i.v. injections of PBS (group1, n=12) or P-selectin antibody (100 µg/injection; group2, n=12). Sample collection and pathological examination Two mice in each group were sacrificed randomly on weeks 1, 2,and 4. The remaining animals were sacrificed at 6th week and the tumors growing on the stomach wall were removed and examined histologically. Tissues from all organs were examined for metastasis after careful macroscopic examination. The spleen of mice was harvested for detection of immune function. Lymphocyte transformation test Using 3H TdR infiltration method, T cell transformation function was detected with Con A (5 µg/ml, Sigma), B cell function with LPS (50 µg/ml, Sigma). The spleen was made
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into suspension and mononuclear cells were acquired with lymphocyte-separating fluid. The cell suspension was adjusted to 2×106/ml with RPMI 1640 liquid containing 10 % calf serum (GIBCO). Then 200 µl of the cell suspension was put into each well in 96-well plates. One plate was for Con A group, and another plate for LPS group. There were negative control group and experimental group for Con A or LPS group respectively, and each test had 5 repetitive wells and was cultivated under 5 % CO2 at 37 . The cell suspension in Con A group was cultivated for 3 d, while it was done for 5 d in Con A group. All wells were added up 3H TdR 16 h before the end of cultivation. They were collected in filtration paper, and given 0.5 ml of scintillating liquid to detect cpm value with βscintillator, which was presented with SI. SI amounts to cpm of experimental group/cpm of empty group.
NK activity NK activity was detected by the 4 h LDH release method. NK cell activity (%)= Experimental group’s OD-Natural release group’s OD Max release group’s OD-Natural release group’s OD Statistical methods Comparisons among groups were performed by the student’s test and χ2 test. A value of P
