Gap Junctions Formed By Connexin43 Between BMMSCs And RPMI8226 Mediates MM Cells Drug Sensitive

Objective The aim of study is to analysis the expression of connexin-43(Cx43) on bone marrow mesenchymal stem cells(BMMSCs) and myeloma cell line RPMI8226 in vitro, explore whether they can form gap junction intercellular communication(GJIC) between the cells under the condition of coculture and what will be happen to RPMI8226 cells when the GJIC was blocken, and to investigate the role of Cx43 and GJIC formed between BMMSCs and RPMI8226 on drug sensitive of MM cells.Methods RT-PCR and immunohistochemical assays were employed to detect Cx43 expression on BMMSCs and RPMI8226, the cell proliftion, cell cycle, apoptosis, secretion of cytokines and Bcl-2、Bcl-xl、Survivin、Cyclin D1 of RPMI8226 cells were analysis by means of growth curve,flow cytometry, Annexin V/PI double staining, cytometric beads array(CBA) and western blotting pre- and post- coincubation with BMMSCs. The changes of biological behavior of RPMI8226 cells and Bcl-2、Bcl-xl、 Survivin、Cyclin D1 expression was explored after the GJIC between BMMSCs and RPMI8226 was blocken with 18 alpha glycyrrhetinic acid(α-GA).Results Both BMMSCs and RPMI8226 cells express Cx43. Our results demonstrated that BMMSCs can promote RPMI8226 cells proliferation and protect RPMI8226 cells from antineoplastic drugs. The cells apoptosis caused by Bortezomib(BTZ) decreased significantly(p<0.05) in comparison with the controls. Cell cycle analysis showed that the proportion of RPMI8226 cells in S phase increased significantly than those in controls after 24 h co-cultured with BMMSCs(p<0.05). The protective effect of BMMSCs on RPMI8226 was partially abroated, when the cocultured cells were incubated in medium with 18 alpha glycyrrhetinic acid(α-GA) and the sensitivity of RPMI8226 to BTZ can be partly recoverd;The cell cycle analysis showed that the proportion of cells in S phase also decreased(p<0.05). The level of IL- 6, IL- 10 and TGF-βin supernatant increased significantly(p<0.05), especially the levels of IL- 6 and IL- 10 increased significantly as compared with the controls(p<0.01) after coincubation for 24 hours, but the level of b FGF and IL-17 didn’t change before and after co-culture(p>0.05). The level of IL-6, IL-10 and TGF-βdecreased significantly(p<0.05) when the cells were cultured in medium containing GJIC blocker, α-GA. Our data also demonstrated that the expression of apoptosis related protein Bcl-xl、Survivin、Cyclin D1 was upregulated(p<0.05) when the RPMI 8226 was cultured with BMMSCS and the expression Bcl-xl、Survivin、Cyclin D1 decreased again(p<0.05) when the GJIC was blocken with α-GA, but there was no change of Bcl-2 expression(p>0.05).Conclusion There is functional GJIC between BMMSCs and RPMI8226 cells, which might have great impact on the MM cell survival and chemoresistance.