[Establishment of mouse stably knockout of MYSM1 in MSC cell line C3H10T1/2 and its effect on the immune regulation in vitro].

Objective: To aimed at the establishment of mouse stably knockout of MYSM1 mesenchymal stem cell(MSC) line C3H10T1/2, and to investigate its immunological capacity of MSC in vitro.

Methods: To establish the stably transfected MSC cell line by using CRISPR-Cas9 technology. Then the Flow cytometry, quantitative PCR and Western blot were employed to detect whether the MYSM1 have been knockout yet. Furthermore, the immune modulatory effect of MYSM1 MSC was tested by addition of MYSM1 MSC supernatant into spleen lymphocyte and Foxp3 culture. The mRNA expression of inflammatory cytokines such as interleukin-4, interferon-γ and interleukin-17 were detected by quatitatine PCR.

Results: The expression of MYSM1 was steadily knock out in MSC. In addition, MYSM1 MSC showed a stronger inhibitory effect on the expression of inflammatory cytokines. Therefore, the MYSM1 has been stably knocked out in C3H10T1/2.

Conclusion: The mouse stably knockout of MYSM1 mesenchymal stem cells has been successfully established, the knock-out of MYSM1 in MSC can induce more potent immunosuppressive effects on cellular immune reaction in vitro. Our data laid a foundation for the further MSC-based applications in immune related diseases.