Enrichment Of Gastric Cancer Stem Cells By Chemotherapy And Screening Of Surface Markers In Nude Mice

ObjectivesTo enrich gastric cancer stem cells by chemotherapy with cisplatin and to screensurface markers of gastric cancer stem cells in nude mice.MethodsHuman poorly differentiated gastric cancer cells BGC-823were injectedsubcutaneously to the right scapular region of nude mice. Nude mice with tumor weredivided into2groups randomly, treat group and control group. Cisplatin wasdelivered through tail vein in treat group and0.9%NaCl was delivered through tailvein in control group. A part of tumor tissue was transplanted to next generation nudemice after the first period of chemotherapy, and continued the second period oftreatment after the new mice generated tumor, we finally got the enriched tissue ofgastric cancer stem cells and the control gastric cancer tissue. Total protein of twogroups was extracted, then to examine high-throughput protein chip (ASB600) thatinclude656antibodies, and screened the differentially expressed proteins through theresult of protein chip. Then, verifying chip result by western blot in two group tissues.BGC-823cells formed spheroid with serum-free medium containing growth factor.Finally, flow cytometry (FCM) detected the proportion of cell mass between adherentcells and spheroid.Results1. Obtained the tissue of enriched gastric cancer stem cells (treat group) and thecontrol gastric cancer tissue (control group). After chemotherapy, volume and weightof tumor have significant difference between treat group and control group in the two generations nude mice, and tumors notably reduced in chemotherapy group (All ofthem are P<0.05).2. There are different proteins between treat group and control group throughASB600protein chip, the up-regulation proteins in treat group include HLA-DQ,PMP22, Claudin7and so on; the down-regulation proteins in treat group includeHLA-DR, CD14, CD16, CD56and so on. They were verified by Western blot intissue samples, the result is consistent with protein microarray.3. BGC-823cells form spheroid with serum-free medium containing epidermalgrowth factor (EGF), basic fibroblast growth factor (bFGF), B27and insulin.4. Flow cytometry (FCM) shows that there are more PMP22+cells andHLA-DQ+cells in spheroid than adherent cells (P<0.05).Conclusions1. Gastric cancer stem cells can be enriched in nude mice by chemotherapy.2. There is different expression between gastric cancer tissue by chemotherapyand cancer tissue.3. BGC-823cells can form spheroid with serum-free medium supplementedgrowth factor.4. PMP22and HLA-DQ probably become potential surface markers of gastriccancer stem cell.