ABSTRACT
Objective: Gastric cancer [GC] is widely associated with chronic inflammation. The pro inflammatory microenvironment provides conditions that disrupt stem/progenitor cell proliferation and differentiation. The signal transducer and activator of transcrip-tion-3 [STAT3] signaling pathway is involved in inflammation and also contributes to the maintenance of embryonic stem cell [ESCs] pluripotency. Here, we have investi-gated the activation status of STAT3 in GC stem-like cells [GCSLCs]
Materials and Methods: In this experimental research, CSLCs derived from the human GC cell line MKN-45 and patient specimens, through spheroid body formation, characterized and then assayed for the STAT3 transcription factor expression in mRNA and protein level further to its activation
Results: Spheroid cells showed higher potential for spheroid formation than the parental cells. Furthemore, stemness genes NANOG, c-MYC and SOX-2 were over expressed in spheroids of MKN-45 and in patient samples. In MKN-45 spheroid cells, epithelial mesenchymal transition [EMT] related markers CDH2, SNAIL2, TWIST and VIMENTIN were upregulated [P<0.05], but we observed no change in expression of the E-cadherin epithelial marker. These cells exhibited more resistance to docetaxel [DTX] when compared with parental cells [P<0.05] according to the MTS assay. Although immunostaining and Western blotting showed expression of the STAT3 protein in both spheroids and parents, the mRNA level of STAT3 in spheroids was higher than the parents. Nuclear translocation of STAT3 was accompanied by more intensive phospho-STAT3 [p-STAT3] in spheroid structures relative to the parent cells according to flow cytometry analysis [P<0.05]
Conclusion: The present findings point to STAT3 over activation in GCSLCs. Complementary experiments are required to extend the role of STAT3 in stemness features and invasion properties of GCSCs and to consider the STAT3 pathway for CSC targeted therapy
ABSTRACT
Objective: Cutaneous melanoma is the most hazardous malignancy of skin cancer with a high mortality rate. It has been reported that cancer stem cells [CSCs] are responsible for malignancy in most of cancers including melanoma. The aim of this study is to compare two common methods for melanoma stem cell enriching; isolating based on the CD133 cell surface marker and spheroid cell culture
Materials and Methods: In this experimental study, melanoma stem cells were enriched by fluorescence activated cell sorting [FACS] based on the CD133 protein expression and spheroid culture of D10 melanoma cell line,. To determine stemness features, the mRNA expression analysis of ABCG2, c-MYC, NESTIN, OCT4-A and -B genes as well as colony and spheroid formation assays were utilized in unsorted CD133+, CD133- and spheroid cells. Significant differences of the two experimental groups were compared using student's t tests and a two-tailed value of P<0.05 was statistically considered as a significant threshold
Results: Our results demonstrated that spheroid cells had more colony and spheroid forming ability, rather than CD133+ cells and the other groups. Moreover, melanospheres expressed higher mRNA expression level of ABCG2, c-MYC, NESTIN and OCT4-A compared to other groups [P<0.05]
Conclusion: Although CD133+ derived melanoma cells represented stemness features, our findings demonstrated that spheroid culture could be more effective method to enrich melanoma stem cells