Osteoarthritis (OA) is characterized by a slowly progressing degradation of the matrix and destruction of articular cartilage. Apoptosis of chondrocyte is accounted for the mechanism of OA. Nitric oxide (NO), as a stimulus, has been shown to induce chondrocyte apoptosis by activating the matrix metalloproteinases (MMPs), increasing the expression of cyclooxygenase 2 (COX-2) and the level of prostaglandin E2 (PGE2), inhibiting the proteoglycan synthesis and type II collagen expression. In this study, sodium nitroprusside (SNP) was administered to be the NO donor to explore the mechanism of NO-induced apoptosis of rabbit chondrocytes obtained from six weeks old New Zealand rabbits. CCK-8 assay revealed the inhibitory effect of SNP on cell viability. We used flow cytometry (FCM) to assess the form of cell death by Annexin-V/propidium iodide (PI) double staining, and evaluate the change of mitochondrial membrane potential (ΔΨm). We found that the SNP induced chondrocyte apoptosis in a dose- and time-dependent manner and an observable reduction of ΔΨm. In conclusion, our findings indicate that SNP induces apoptosis of rabbit chondrocytes via a mitochondria-mediated pathway.
Resveratrol (RV), a naturally occurring phytoalexin, is known to possess a wide spectrum of chemopreventive and
chemotherapeutic effects in various stages of human tumors. p38, a member of the mitogen-activated protein kinase
(MAPK) superfamily, is always activated by some extracellular stimulus to regulate many cellular signal transduction
pathways, such as apoptosis, proliferation, and inflammation and so on. In this report, we assessed the effect of
SB203580, a specific inhibitor of p38 MAPK signaling pathway, on the RV-induced apoptosis in human lung
adenocarcinoma (A549) cells. CCK-8 assay showed that pretreatment with SB203580 significantly enhanced the
cytotoxicity of RV, which was further verified by analyzing the phosphatidylserine externalization using flow cytometry.
In order to further confirm whether SB203580 accelerated apoptosis via the intrinsic apoptosis pathway, we analyzed the
dysfunction of mitochondrial membrane potential (Δψm) of cells stained with rhodamine 123 by using flow cytometry
after treatment with RV in the absence and presence of SB203580. Our data for the first time reported that p38 inhibitor
SB203580 enhanced the RV-induced apoptosis via a mitochondrial pathway.
The degenerative joint disease such as osteoarthritis (OA) is closely associated with the death of chondrocytes in
apoptosis fashion. Hydrogen peroxide (H2O2), higher expression following acute damage in OA patients, has been shown
to be up-regulated during apoptosis in a bulk of experimental models. This study was aimed to explore the mechanism of
H2O2-induced rabbit chondrocytes apoptosis. Articular cartilage was biopsied from the joints of 6 weeks old New
Zealand rabbits. Cell Counting Kit (CCK-8) assay was used to assess the inhibitory effect of H2O2 on cell viability. H2O2
treatment induced a remarkable reduction of cell viability. We used flow cytometry to assess the form of cell death with
Annexin-V/PI double staining, and found that H2O2 treatment induced apoptosis in a dose-and time-dependent manner.
Exposure of chondrocytes to 1.5 mM of H2O2 for 2 h induced a burst apoptosis that can be alleviated by N-acetyl
cysteine (NAC) pretreatment, an anti-oxidant amino-acid derivative. Loss of mitochondria membrane potential (▵Ψm)
was evaluated using confocal microscopy imaging and flow cytometry (FCM). H2O2 treatment induced a marked
reduction of ▵Ψm, and the abrupt disappearance of ▵Ψm occurred within 5 minutes. These results indicate that H2O2
induces a rapid apoptosis via a mitochondrial pathway in rabbit chondrocytes.
Dihydroartemisinin (DHA), a front-line antimalarial herbal compound, has been shown to possess promising anticancer activity with low toxicity. We have previously reported that DHA induced caspase-3-dependent apoptosis in human lung adenocarcinoma cells. However, the cellular target and molecular mechanism of DHA-induced apoptosis is still poorly defined. We use confocal fluorescence microscopy imaging, fluorescence resonance energy transfer, and fluorescence recovery after photobleaching techniques to explore the roles of DHA-elicited reactive oxygen species (ROS) in the DHA-induced Bcl-2 family proteins activation, mitochondrial dysfunction, caspase cascade, and cell death. Cell Counting Kit-8 assay and flow cytometry analysis showed that DHA induced ROS-mediated apoptosis. Confocal imaging analysis in a single living cell and Western blot assay showed that DHA triggered ROS-dependent Bax translocation, mitochondrial membrane depolarization, alteration of mitochondrial morphology, cytochrome c release, caspase-9, caspase-8, and caspase-3 activation, indicating the coexistence of ROS-mediated mitochondrial and death receptor pathway. Collectively, our findings demonstrate for the first time that DHA induces cell apoptosis by triggering ROS-mediated caspase-8/Bid activation and the mitochondrial pathway, which provides some novel insights into the application of DHA as a potential anticancer drug and a new therapeutic strategy by targeting ROS signaling in lung adenocarcinoma therapy in the future.
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