ABSTRACT
To investigate the changes in activation and proliferation of Tregs after targeted RNA inter-ference of Kvl. 3 channel genes and in vitro administration of eplerenone (EPL). Methods After lenti-virus vector was transfected to regulatory T cells (Tregs) of rats, qPCR and whole-cell patch-clamp methods were used to detect gene knockout efficiency, and EOSA method was used to detect cytokine secretion IL-10 and TGF-p levels of Tregs group,Tregs + EPL group,RNAi-Tregs group,and RNAi-Tregs + EPL group. Results Lentivirus vector was successfully transfected into Tregs cells, and the mRNA level and current density of Kvl. 3 channel was 78% and 71.3% respectively; compared with Tregs group, extracellular and intracellular TGF-p levels in RNAi-Tregs group were significantly reduced (P < 0. 01), and extracellular and intracellular TGF-(3 levels in Tregs + EPL group were also reduced (P < 0. 05 ) ; compared with RNAi-Tregs group, extracellular and in-tracellular TGF-fJ levels in RNAi-Tregs + EPL group showed no change. However, IL-10 levels in Tregs group,Tregs + EPL group, RNAi-Tregs group, RNAi-Tregs + EPL group showed no significant change. Conclusions Kvl.3 channel mediates the activation and proliferation of Tregs cells, while EPL can reduce the activation and proliferation of Tregs cells by directly inhibiting Kvl.3 channel and reducing the secretion of TGF-fi levels, further indicating that EPL is a specific blocker of Kvl. 3 channel.
ABSTRACT
Aim To investigate the effect of TRPV4 on hepatic fibrosis of rats . Methods Liver fibrosis model of rats was induced by 50% CCl4 twice a week for 12 weeks. HE and Masson staining were used to evaluate the degree of hepatic fibrosis, and the levels of α-smooth muscle actin(α-SMA) and TRPV4 were detec-ted in fibrotic liver tissue by Western blot. HSC-T6 cells were activated by transforming growth factor β1 (TGF-β1),and the protein levels of α-SMA, TRPV4 were detected by Western blot. After using Ru and transfected with TRPV4-siRNA, HSC-T6 was stimula-ted with TGF-β1, the levels of α-SMA, TRPV4 and phosphorylation level of Akt were determined by West-ern blot. Results TRPV4 was highly expressed in model liver tissues and in activated HSC-T6 induced by TGF-β1 . The levels of α-SMA and phosphorylation of Akt decreased in TGF-β1-induced HSC, used with Ru or transfected with TRPV4-siRNA. Conclusions The expression of TRPV4 increases in fibrotic livers and ac-tivated hepatic stellate cells. Knockdown of TRPV4 can suppress the activation of hepatic stellate cells in-duced by TGF-β1 , and decrease the phosphorylation levels of Akt.
ABSTRACT
We have previously reported that a CD_(20) monoclonal antibody (HI_(47)) had obvious inhibition to ~3H-TdR incorporation of tonsil B cells in SAC drived system. This paper describes that McAb HI_(47) also markedly inhibited ~3H-UdR uptake and enlargement of B lymphocytes induced by SAC. However, it had no effect on B cell activation in PMA and anti-immunoglobul in antibody drived systems and the proliferation of established T, B and myeloid cell lines. The non-specific inhibition of HI_(47) Fc fragment had been ruled out by the experiments with irrelative MoAbs, which Ig subclass was same as HI_(47) (IgG_3),and with HI_(47) F (ab')2 fragment. Exposure of tonsil B cell to either SAC or PMA resulted in an increase in expression of HI_(47) antigen. McAb HI_(47), neverthless, induced reduction of HI_(47) antigen expressed on B cells stimulated by SAC. but not by PMA. In conclusion, antigen HI_(47) (CD_(20)) only involved in the SAC-activation pathway. The modulation of HI47 antigen by McAb HI47 may be one of the mechanisms of the depressed SAC activation.