Study on the mechanism of Wuzi-Yanzong-Wan-medicated serum interfering with the mitochondrial permeability transition pore in the GC-2 cell induced by atractyloside / 中国天然药物

Wuzi-Yanzong-Wan (WZYZW) is a classic prescription for male infertility. Our previous investigation has demonstrated that it can inhibit sperm apoptosis via affecting mitochondria, but the underlying mechanisms are unclear. The purpose of the present study was to explore the actions of WZYZW on mitochondrial permeability transition pore (mPTP) in mouse spermatocyte cell line (GC-2 cells) opened by atractyloside (ATR). At first, WZYZW-medicated serum was prepared from rats following oral administration of WZYZW for 7 days. GC-2 cells were divided into control group, model group, positive group, as well as 5%, 10%, 15% WZYZW-medicated serum group. Cyclosporine A (CsA) was used as a positive control. 50 μmol·L-1 ATR was added after drugs incubation. Cell viability was assessed using CCK-8. Apoptosis was detected using flow cytometry and TUNEL method. The opening of mPTP and mitochondrial membrane potential (MMP) were detected by Calcein AM and JC-1 fluorescent probe respectively. The mRNA and protein levels of voltage-dependent anion channel 1 (VDAC1), cyclophilin D (CypD), adenine nucleotide translocator (ANT), cytochrome C (Cyt C), caspase 3, 9 were detected by RT-PCR (real time quantity PCR) and Western blotting respectively. The results demonstrated that mPTP of GC-2 cells was opened after 24 hours of ATR treatment, resulting in decreased MMP and increased apoptosis. Pre-protection with WZYZ-medicated serum and CsA inhibited the opening of mPTP of GC-2 cells induced by ATR associated with increased MMP and decreased apoptosis. Moreover, the results of RT-qPCR and WB suggested that WZYZW-medicated serum could significantly reduce the mRNA and protein levels of VDAC1 and CypD, Caspase-3, 9 and CytC, as well as a increased ratio of Bcl/Bax. However, ANT was not significantly affected. Therefore, these findings indicated that WZYZW inhibited mitochondrial mediated apoptosis by attenuating the opening of mPTP in GC-2 cells. WZYZW-medicated serum inhibited the expressions of VDAC1 and CypD and increased the expression of Bcl-2, which affected the opening of mPTP and exerted protective and anti-apoptotic effects on GC-2 cell induced by ATR.

Role of cyclophilinD in induction of the regulated necrosis / 基础医学与临床

CyclophilinD ( CypD) , a member of cyclophilins family displays a variety of biological function .CypD lo-cates in the mitochondrial matrix .As an component of mitochondrial permeability transition pore ( mPTP ) , CypD plays an important role in the regulating of mPTP opening , which leads to mitochondrial dysfunction and CypD-de-pendent regulated necrosis .Elucidation of the mechanisms of CypD in regulating mPTP and the regulated necrosis will provide novel insights in injury-relevant diseases , such as cardiac or cerebral ischemia .

Regulatory necrosis:a novel way to recognize and prevent injury-relevant diseases / 中国药理学通报

Necrosis is also tightly controlled by signaling path-ways,thus it is called as regulated necrosis,which includes ne-croptosis,ferroptosis,parthanatos and CypD-mediated necrosis. It has been shown that regulated necrosis is closely related to the occurrence,development and prognosis of injury-relevant disea-ses such as myocardial infarction,stroke,neurodegenerative dis-eases.It will be significant for prevention and therapy of injury-relevant diseases to clarify the signal transductions and regulatory mechanisms for the regulated necrosis.