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PD4 is a monoclonal antibody prepared from mouse immunized with human gastric cancer cells and could specifically reacted with different cancer tissues. Recently the antigen was identified and it is a protein P37 from Mycoplasma hyorhinis (M.Hyo.). To confirm the mycoplasma existence in gastric cancer tissues and explore its clinical significance as well as its activity in malignant transformation, sixty-one specimens from gastric cancer patients were examined by nested PCR, immunohistochemistry, and direct mycoplasmal culture. 32D cells were infected with the mycoplasmas and the colony formation assay was performed and the potential mechanisms were investigated. We found that M. hyorhinis was positive in 39 specimens (63.9%) detected by PCR and in 28 specimens (45.9%) by immunohistochemistry. 19 specimens, which were M. hyorhinis positive in PCR, were also M. fermentans positive. No other mycoplasmas were detected. Fifty-two specimens were successfully cultured and eight (15.4%) were positive for mycoplasmas (5 for M. hyorhinis, 2 for M. fermentans, and 1 for both species). Analysis of PCR results suggested that M. hyorhinis infection in gastric cancer is correlated with lymph node metastasis, the Lauren's classification and the TNM stage (I-IV) (P ≤ 0.05). 32D cells, infected with M. hyorhinis or M. fermentans for 6 weeks, were able to form colonies on soft agar, with deregulation of a number of genes involved in carcinogenesis, proliferation, and metabolism. Because p37 is the main antigen of M hyo. and to investigate its function in mycoplasma promoting cell malignance, p37 gene was cloned and well-expressed in eukaryotic cells. After stably integrated in AGS and HEK 293/TRE/TO cell lines, p37 gene apparently promoted cells' colony forming abilities in soft agar. Compared to control cells, the stable p37 AGS cell became smaller, more spherical and adhesion ability among cells became weaker. Furthermore, in stable p37 AGS cell, the phosphorylation of β-catenin was downregulated and β-catenin was stabilized and transferred into nucleus, as well as the expression of ICAM-1 was downregulated. Taken together, M. hyo. exist in gastric carcinoma and possibly relate with metastasis, and p37 maybe involve in cell proliferation and colony formation enhanced by m hyo. infection. Key words: mycoplasma hyorhinis, infection; gastric cancer; promotion

[Proc Amer Assoc Cancer Res, Volume 47, 2006]