ROS-mediated lysosomal membrane permeabilization is involved in bupivacaine-induced death of rabbit intervertebral disc cells

Cai, Xianyi; Liu, Yunlu; Hu, Yiqiang; Liu, Xianzhe; Jiang, Hongyan; Yang, Shuai; Shao, Zengwu; Xia, Yun; Xiong, Liming

doi:10.1016/j.redox.2018.06.010

Cited by 62 publications

(56 citation statements)

References 44 publications

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“…The expression of ROS has been confirmed to be increased during the progress of IVDD, suggesting that ROS may play an essential role in the pathological process of IVDD (15). In the present study, the intervertebral disc with severe degenerated degree expressed a higher level of MDA compared with the mild, suggesting the degree of degeneration is positively related to the content of ROS.…”

Section: Discussionsupporting

confidence: 67%

Hydrogen peroxide induces nucleus pulposus cell apoptosis by ATF4/CHOP signaling pathway

Liu

2020

Exp Ther Med

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Oxidative stress induces excessive apoptosis resulting in the reduction of intervertebral disc cells, the consequent reduction of extracellular matrix (ECM) synthesis, and compositional changes, which is the pathological basis for intervertebral disc degeneration (IVDD). The present study explored the activating transcription factor 4 (ATF4)/C/EBP homologous protein (CHOP) signaling pathway in the H 2 O 2-induced nucleus pulposus (NP) cell apoptosis. Human degenerated intervertebral discs were collected from Lumbar disc surgery. NP cells isolated from the tissues were cultured with H 2 O 2 to induce apoptosis in vitro. Malondialdehyde (MDA) analysis was performed to determine the reactive oxygen species (ROS) of the tissue. Western blot analysis and reverse transcription-polymerase chain reaction (RT-PCR) were performed to analyze collagen II, ATF4, CHOP, and caspase-9 gene expression. Flow cytometry was used to determine the apoptotic ratio of NP cells. siRNA was also used to silence ATF4 and CHOP gene expression. NP tissues in higher degenerated degree underwent much more MDA, expressed less collagen II, more ATF4, CHOP, and caspase-9 compared with the mildly degenerated tissues. H 2 O 2 induced NP cell apoptosis by upregulating expression of ATF4, CHOP and caspase-9. The silencing of ATF4 or CHOP alleviated NP cell apoptosis by suppressing caspase-9 expression. Inhibiting caspase-9 did not affect ATF4 and CHOP expression but protected NP cells from apoptosis. In this study, we found H 2 O 2 could promote NP cell apoptosis by activating the ATF4/CHOP signaling pathway resulting in the upregulation of caspase-9. Interdict of ATF4, CHOP, or caspase-9 contributed to the reduction of apoptosis caused by H 2 O 2 .

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Section: Discussionsupporting

confidence: 67%

Hydrogen peroxide induces nucleus pulposus cell apoptosis by ATF4/CHOP signaling pathway

Liu

2020

Exp Ther Med

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“…The RIPK1 inhibitor Necrostatin-1 (Nec-1) (Selleck, Houston, TX), MLKL inhibitor Necrosulfonamide (NSA) (Selleck, Houston, TX), and RIPK3 inhibitor GSK872 (Selleck, Houston, TX) were dissolved in DMSO solution. Before exposure to FAC, the MC3T3-E1 osteoblastic cells were incubated with or without NAC (1 mM), Nec-1 (20 μ M), GSK872 (4 μ M), or NSA (4 μ M) [ 9 , 23 – 25 ]. Then, after FAC (200 μ M) treatment for 120 h, all samples were collected and analyzed by a microplate reader, flow cytometry, western blots, and confocal microscopy.…”

Section: Methodsmentioning

confidence: 99%

“…Briefly, after being treated as described above, the culture medium was discarded and the osteoblastic cells were harvested and washed thrice by PBS. Then, cells were resuspended and incubated with 20 μ M H2DCF-DA at 37°C for 20 min in the dark [ 25 ]. Subsequently, serum-free medium was used to rinse the cells in order to remove the residual dyes.…”

Section: Methodsmentioning

confidence: 99%

ROS-Mediated Necroptosis Is Involved in Iron Overload-Induced Osteoblastic Cell Death

Tian

Qin

et al. 2020

Oxidative Medicine and Cellular Longevity

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Excess iron has been reported to lead to osteoblastic cell damage, which is a crucial pathogenesis of iron overload-related osteoporosis. However, the cytotoxic mechanisms have not been fully documented. In the present study, we focused on whether necroptosis contributes to iron overload-induced osteoblastic cell death and related underlying mechanisms. Here, we showed that the cytotoxicity of iron overload in osteoblastic cells was mainly due to necrosis, as evidenced by the Hoechst 33258/PI staining, Annexin-V/PI staining, and transmission electronic microscopy. Furthermore, we revealed that iron overload-induced osteoblastic necrosis might be mediated via the RIPK1/RIPK3/MLKL necroptotic pathway. In addition, we also found that iron overload was able to trigger mitochondrial permeability transition pore (mPTP) opening, which is a critical downstream event in the execution of necroptosis. The key finding of our experiment was that iron overload-induced necroptotic cell death might depend on reactive oxygen species (ROS) generation, as N-acetylcysteine effectively rescued mPTP opening and necroptotic cell death. ROS induced by iron overload promote necroptosis via a positive feedback mechanism, as on the one hand N-acetylcysteine attenuates the upregulation of RIPK1 and RIPK3 and phosphorylation of RIPK1, RIPK3, and MLKL and on the other hand Nec-1, siRIPK1, or siRIPK3 reduced ROS generation. In summary, iron overload induced necroptosis of osteoblastic cells in vitro, which is mediated, at least in part, through the RIPK1/RIPK3/MLKL pathway. We also highlight the critical role of ROS in the regulation of iron overload-induced necroptosis in osteoblastic cells.

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“…When the lysosome ruptures, the AO leaks out and redistributes in the cytoplasm. Hence, AO redistribution can be used to determine the lysosomal membrane stability [25,26]. Microglia cells were stained with acridine orange (5 μM) and the excess fluorescent dye were washed off by performing centrifugation twice at 1500 rpm for 3 min in the 2ml incubation medium.…”

Section: Plos Onementioning

confidence: 99%

Betanin purification from red beetroots and evaluation of its anti-oxidant and anti-inflammatory activity on LPS-activated microglial cells

et al. 2020

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Microglial activation can release free radicals and various pro-inflammatory cytokines, which implicates the progress of a neurodegenerative disease. Therefore suppression of microglial activation can be an appropriate strategy for combating neurodegenerative diseases. Betanin is a red food dye that acts as free radical scavenger and can be a promising candidate for this purpose. In this study, purification of betanin from red beetroots was carried out by normal phase colum chromatography, yielding 500 mg of betanin from 100 g of red beetroot. The purified betanin was evaluated by TLC, UV-visible, HPLC, ESI-MASS, FT-IR spectroscopy. Investigation on the inhibitory effect of betanin on activated microglia was performed using primary microglial culture. The results showed that betanin significantly inhibited lipopolysaccharide induced microglial function including the production of nitric oxide free radicals, reactive oxygen species, tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6) and interleukin-1 beta (IL-1β). Moreover, betanin modulated mitochondrial membrane potential, lysosomal membrane permeabilization and adenosine triphosphate. We further investigated the interaction of betanin with TNF-α, IL-6 and Nitric oxide synthase (iNOS or NOS2) using in silico molecular docking analysis. The docking results demonstrated that betanin have significant negative binding energy against active sites of TNF-α, IL-6 and iNOS.

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ROS-mediated lysosomal membrane permeabilization is involved in bupivacaine-induced death of rabbit intervertebral disc cells

Cited by 62 publications

References 44 publications

Hydrogen peroxide induces nucleus pulposus cell apoptosis by ATF4/CHOP signaling pathway

Hydrogen peroxide induces nucleus pulposus cell apoptosis by ATF4/CHOP signaling pathway

ROS-Mediated Necroptosis Is Involved in Iron Overload-Induced Osteoblastic Cell Death

Betanin purification from red beetroots and evaluation of its anti-oxidant and anti-inflammatory activity on LPS-activated microglial cells

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