1.Effects of miR-30 a on cell cycle and dendritic cell-suppressive function of MSCs
Erling HU ; Dan LIU ; Huishuang MIAO ; Yahong HUANG ; Yayi HOU
Chinese Journal of Immunology 2015;(4):462-466,471
Objective:Mesenchymal stem cells( MSCs) have self-renewal capacity and potential to differentiate into the cells.It was reported that the expression of miR-30a changed in some immune diseases.But it remains unclear the effect of miR-30a on the im-munoregulatory functions of MSCs.Here we studied the impact of miR-30a on the phenotype,cell viability,apoptosis,cell cycle and im-munoregulatory functions of MSCs.Methods: The mixed enzyme methods were used for the isolation of human umbilical cord MSCs.Flow cytometry(FCM)was used to investigate the effect of overexpressed miR-30a on the phenotype of MSCs.CCK-8 was used to examine the cell viability of miR-30a-overexpressed MSCs.Annexin V/PI was used for the detection of apoptosis of MSCs.Q-PCR and Western blot were used to investigate the effect of miR-30a on the expression of Cyclin E2( CCNE2).CCNE2 was one putative target of miR-30a predicted by Targetscan database.The effects of miR-30a-overexpressed MSCs on the maturation of dendritic cells(DCs)were determined.Results:Overexpression of miR-30a blocked the cell cycle of MSCs in the G0/G1 phase by inhibiting the expression of CCNE2,but did not affect the phenotype, cell viability and apoptosis of MSCs.When co-cultured with DCs, although MSCs down-regulated the expression of CD40 and CD86 on DCs,overexpression of miR-30a more significantly enhanced the suppressive impact of MSCs on the maturation of DCs.Conclusion: miR-30a affects the cell cycle of MSCs and enhances its immunosuppressive effect on DCs.
2.Functions of gene phoN1 in Shigella flexneri 2a:preliminary study
Xiaoyu NIU ; Wei HU ; Erling FENG ; Xiankai LIU ; Mei ZHANG ; Li ZHU
Military Medical Sciences 2014;(1):35-38
Objective To explore the function of gene phoN1 in Shigella flexneri.Method Using the λ-Red recombi-nant system, phoN1was knocked out from S.flexneri 2a strain 301.Comparative proteomics was performed to analyze the differences between mutant and wild-type strains in protein expression profiles .Sereny tests and competitive infection assays were carried out to compare the virulence of mutant and wild-type strains .Results The deletion mutant of phoN1 was suc-cessfully constructed .No significant difference between the two strains was found in the comparative proteomics analyses . The function of gene phoN1 might be unrelated to the invasion ability of S.flexneri according to the results of Sereny tests and competitive infection assays .Conclusion Gene phoN1 might be of no use for the in vitro survival and host cell invasion of S.flexneri.