Effect Of Umbilical Cord And Amniotic Mesenchymal Stem Cells On Proliferation And Metastasis Of Gastric Cancer

Objective(s):Gastric cancer is one of the malignant tumors that threaten human health seriously,with a high incidence in recent years.It is the third leading cause of cancer related death in China and even the second globally.Most of the patients were diagnosed at the advanced stage,thus missed the best time for surgical treatment.Gastric cancer was resistant to various treatment methods,resulting in a low 5-year survival rate.Due to its high morbility and mortality threaten human health seriously and the traditional treatment methods have limitations,it is an urgent need to develop new treatments.Mesenchymal stem cells(MSCs)is a type of pluripotent stem cells that derived from diverse sources and has the potential of multi-directional differentiation,therefore it has been a hot spot in the field of clinical and scientific research recently.Because of its biological functions involved in anti-apoptosis,immunomodulation,anti-inflammation and pro-angiogenesis,MSCs have been proved as a potential method to treat a variety of diseases such as cardiovascular disease,diabetes,Alzheimer's disease,multiple sclerosis,etc.Meanwhile,since MSC has a good potentiality to migratate to tumor sites and can interact with tumor microenvironment(TME)to participate in the regulation of tumor development process,more and more studies have been conducted on MSC and tumor in recent years,especially the use of human mesenchymal stem cslls(hMSCs)to intervene in the development of tumor cells.Previous studies had shown that MSC could promote the proliferation and metastasis of tumor cells,but contrary studies had also been reported.Therefore,this study would further explored the effect of human umbilical cord mesenchymal stem cells(hUCMSCs)and human amniotic mesenchymal stem cells(hAMSCs)on the proliferation and metastasis of gastric adenocarcinoma cancer cell lines,namely BGC-823 and HGC27 in vivo and in vitro,and the possible underlying mechanisms.The present study would evaluate the possibility of MSC in the treatment of gastric cancer.Methods:1.UCMSC,AMSC,BGC-823 and HGC27 were resuscitated from liquid nitrogen refrigeration and cultured them.2.Phenotypic analysis of hAMSCs and hUCMSCs induced differentiation,the collection of mesenchymal stem cells conditioned medium.2.1 Phenotyping of hAMSCs and hUCMSCs by flow.2.2 Multi-differentiation capabilities of hAMSCs and hUCMSCs.2.3 After the cells grew approximately 80-90%confluent,the cells were sub-cultured,the third-generation cells in good growth state were taken for the follow-up experiment.The conditioned medium-umbilical cord mesenchymal stem cell(cm-UCMSC)and the conditioned medium-amniotic mesenchymal stem cell(cm-AMSC)was collected.3.Cell proliferation experiments(Cell Counting Kit-8,CCK8).3.1 CCK8 assay was performed to evaluate the proliferation of gastric canlcer cells BGC-823 and HGC27,after the cells were exposed to cm-UCMSC and cm-AMSC.OD values at 450nm was measured by a microplate spectrophotometer.4.Cell scratch experiment.4.1 Cell scratch was used to detect the effect of cm-UCMSC and cm-AMSC on BGC-823 and HGC27 migration in vitro.5.Matrigel invasion assay.5.1 Matrigel invasion assay was performed to determine the invasion of BGC-823 and HGC27 after incubation with cm-UCSC and cm-AMSC in vitro.6.Effect of mesenchymal stem cells on metastasis of gastric cancer cells in BGC-823 xenograft nude mice model.7.Western Blot showed that MSC promoted gastric cancer BGC-823 and HGC27 proliferation through the activation of ERK signaling pathway.Results:1.The results of CCK8 assay showed that the proliferation of BGC-823 and HGC27 cells were promoted after 60%and 100%cm-UCSC and cm-AMSC exposure.2.The results of cell scratch experiment showed that cm-UCMSC had no significant effect on the horizontal migration of BGC-823,while cm-AMSC inhibited the horizontal migration of BGC-823.However,both cm-UCMSC and cm-AMSC can promote the horizontal migration of HGC27.3.The results of Matrigel invasion assay showed that cm-UCMSC had no significant effect on the invasion of BGC-823,while cm-AMSC had an inhibitory effect on the invasion of BGC-823.Both cm-UCMSC and cm-AMSC can promote the invasion of HGC27.These results were consistant with those of scratch assay.4.The in vivo study demonstrated that UCMSC and AMSC had no significant effect on the metastasis of BGC-823 cancer xenografts.5.Western blot showed that the pERK/ERK ratio was higher than that of the control group after BGC-823 and HGC27 were cultured with 100%cm-UCMSC and cm-AMSC.Conclusions:1.UCMSC and AMSC can promote the proliferation of BGC-823 and HGC27 with their high concentrations of mediums.2.UCMSC had no significant effect on the migration and invasion of gastric cancer cell BGC-823 in vitro,while AMSC inhibited the migration and invasion of gastric cancer cell BGC-823 in vitro.However,UAMCS and AMSC could significantly promote the migration and invasion of HGC27 cells in vitro.3.UCMSC and AMSC had no significant promoted or inhibitory effects on BGC-823 cell metastasis in nude mice xenograft model.4.The mechanism study found that the activation of ERK pathway might be involved in UCMSC and AMSC's promoted effects of proliferation of BGC-823 and HGC27.5.The safety of UCMSC and AMSC on gastric cancer treatment need be further considered cautiously according to the results of our present study.