The Effect Of Proliferation Inhibition And Mechanism Of N,N’-dicyclohexyl-N-linolenic Acid Ureide On Lymphocytes

Objective:Organ transplantation is the only effective method for clinical treatment of organ failure,and the major obstacle in succeeding the organ transplantation is to overcome the immune-rejection.At present,anti-rejection drugs after organ transplantation are not effective enough for immunosuppressive effects or reduce side effects.Thus,there is a great urge to develop a more advanced,targeted and safer immunosuppressant.In our laboratory,several monomers were extracted from Radix Isatidis,and the monomer components with immunosuppressive feature were screened out and validated by pharmacodynamic experiments.On this basis,to overcome the poor solubility of these compound,we have been structurally remodified them to obtain a new compound MSR405.In this study,we researched on the immunosuppressive effects and possible mechanisms of MSR405 through a series of experiments in vitro.Methods:Firstly,The inhibitory effect and mechanism of MSR405 on lymphocyte were investigated in vitro by using lymphocyte transformation assay,anergy assay,and apoptosis assay.Secondly,we also used RNA-seq technology to analyse the inhibition effect of MSR405 on lymphocyte cell cycle.Finally,based on the sequening results and futhuer investigation,we varified our hypothesis,the possible mechanism pathways of MSR405,with Western Blot and qRT-PCR.Results:Firstly,we found that MSR405 was able to suppress lymphocytes proliferation in a dose-dependent manner in vitro.Secondly,RNA-seq data displayed that the inhibitied lymphocytes proliferation is closely associated with cell cycle arrest.FACS revealed that MSR405 increased the ratio of G0/G1 phase of lymphocytes,decreased the proportion of S phase,and the ratio of G2/M phase did not change.Lastly,the results indicated that the negative changes of the cell cycle-related effectors(such as,CDK2,Cyclin E,CDK4,Cyclin D1 and Rb)was achived by suppressing the activity of TGF-P and Akt.Therefore,we proposed that MSR405 inhibite the lymphocyte proliferation through cell cycle arrest.Conclusion:For the first time,we demonstrated that MSR405 can suppress lymphocyte proliferation in vitro via inhibited TGF-β/Smad and Pdkl/Akt signaling pathway and thereby affecting cell cycle regulation.This study nominated a highly potential candidate as an immunosuppressive compound for or gan transplantation.