cytochrome-c-t and 2-5-hexanedione

It has been shown that the conditioned medium of bone mesenchymal stem cells (BMSC-CM) can inhibit apoptosis of neural cells exposed to 2,5-hexanedione (HD), but its protective mechanism remains unclear. To investigate the underlying mechanism, VSC4.1 cells were given HD and 5, 10 and 15% BMSC-CM (v/v) in the current experiment. Our data showed that BMSC-CM concentration-dependently attenuated HD-induced cell apoptosis. Moreover, BMSC-CM remarkably decreased the mitochondrial cytochrome c (Cyt C) release and the caspase-3 activity in HD-given VSC4.1 cells. Given a relatively high expression of NGF in BMSCs and BMSC-CM, we hypothesized that NGF might be an important mediator of the protection of BMSC-CM against apoptosis induced by HD. To verify our hypothesis, the VSC4.1 cells were administrated with NGF and anti-NGF antibody in addition to HD. As expected, NGF could perfectly mimic BMSC-CM's protective role and these beneficial effects were abolished by anti-NGF antibody intervention. To further explore its mechanism, inhibitors of TrkA and Akt were given to the VSC4.1 cells and NGF/Akt/Bad pathway turned out to be involved in anti-apoptotic role of BMSC-CM. Based on these findings, it was revealed that BMSC-CM beneficial role was mediated by NGF and relied on the Akt/Bad pathway.

Topics: Animals; Apoptosis; bcl-Associated Death Protein; Caspase 3; Culture Media, Conditioned; Cytochromes c; Hexanones; Mesenchymal Stem Cells; Microscopy, Electron, Transmission; Mitochondria; Nerve Growth Factor; Neurons; Proto-Oncogene Proteins c-akt; Rats; Receptor, trkA; Signal Transduction

2,5-hexanedione (HD) induces apoptosis of nerve cells. However,the mechanism of HD-induced apoptosis remains unknown. Mesenchymal stem cells (MSCs) are multipotential stem cells with the ability to differentiate into various cell types. This study is designed to investigate the apoptosis induced by HD in rat bone marrow MSCs (BMSCs) and the related underlying mechanisms. The fifth generation of MSCs was treated with 0, 10, 20 and 40 mM HD respectively. The viability of BMSCs was observed by MTT. Apoptosis were estimated by Hoechst 33342 staining and TUNEL assay. The disruption of mitochondrial transmembrane potential (MMP) was examined by JC-1 staining. Moreover, the expression of Bax and Bcl-2, cytochrome c release, and caspase-3 activity were determined by real-time RT-PCR, Western blot and Spectrophotometry. Our results showed that HD induced apoptosis in MSCs in a dose dependent manner. Moreover, HD downregulated the Bcl-2 expression,upregulated the Bax expression and the Bax/Bcl-2 ratio, promoted the disruption of MMP, induced the release of cytochrome c from mitochondria to cytosol, and increased the activity of caspase-3 in MSCs. These results indicate that HD induces apoptosis in MSCs and the activated mitochondria-dependent caspase-3 pathway may be involved in the HD-induced apoptosis.

Topics: Animals; Apoptosis; bcl-2-Associated X Protein; Bone Marrow Cells; Caspase 3; Cytochromes c; Dose-Response Relationship, Drug; Hexanones; Male; Membrane Potential, Mitochondrial; Mesenchymal Stem Cells; Mitochondria; Neurotoxins; Proto-Oncogene Proteins c-bcl-2; Rats; Rats, Sprague-Dawley; Signal Transduction