1. Inhibition of Caspase-1-dependent pyroptosis attenuates copper-induced apoptosis in chicken hepatocytes
Zhaoxin Tang, Jiaqiang Pan, Yuyin Lin, Jianying Guo, Lianmei Hu, Jianzhao Liao, Ying Li, Fan Yang, Qingyue Han, Feiyang Ma, Wenlan Yu, Xinyan Ma Ecotoxicol Environ Saf . 2019 Jun 15;174:110-119. doi: 10.1016/j.ecoenv.2019.02.069.
The purpose of this study was to investigate the effects of copper (Cu) on hepatocyte pyroptosis and the relationship between pyroptosis and apoptosis in the mechanisms of Cu toxicity. Primary chicken hepatocytes were cultured in different concentrations of Cu sulfate (CuSO4) (0, 10, 50, and 100 μM), N-acetylcysteine (NAC) (1 mM), and Z-YVAD-fluoromethylketone (Z-YVAD-FMK) (10 μM) for 24 h, and the combination of Cu and NAC or Z-YVAD-FMK for 24 h. Cellular morphology and function, cell viability, mitochondria membrane potential (MMP), apoptosis rate, mRNA expression of pyroptosis-related and apoptosis-related genes, and Caspase-1, Caspase-3 proteins expression were determined. These results indicated that Cu markedly induced the mRNA expression of pyroptosis-related genes (Caspase-1, IL-1β, IL-18, and NLRP3) and Caspase-1 protein expression. Furthermore, contents of Caspase-1, IL-1β, and IL-18 in the supernatant fluid of culture hepatocytes were significantly increased in hepatocytes. NAC relieved excess Cu-caused the changes of above genes and proteins. Additionally, Z-YVAD-FMK, caspase-1 inhibitor, which attenuated Cu-induced the increased lactic dehydrogenase (LDH), aspartate amino transferase (AST), alanine aminotransferase (ALT) activities. Furthermore, treatment with Cu and Z-YVAD-FMK could down-regulate the mRNA levels of Caspase-3, Bak1, Bax, and CytC and Caspase-3 protein expression, up-regulate the mRNA expression of Bcl2, increase the MMP and reduce cell apoptosis compared to treatment with Cu in hepatocytes. Collectively, these finding evidenced that excess Cu induced pyroptosis by generating ROS in hepatocytes, and the inhibition of Caspase-1-dependent pyroptosis might attenuate Cu-induced apoptosis.
2. Caspase-dependent apoptosis in light-induced retinal degeneration
Isabelle Ranchon-Cole, Olivier Perche, Michel Doly Invest Ophthalmol Vis Sci . 2007 Jun;48(6):2753-9. doi: 10.1167/iovs.06-1258.
Purpose:To study the apoptotic mechanism involved in our model of light-induced retinal degeneration.Methods:Rats were injected intravitreally with PBS, 2% dimethyl sulfoxide (DMSO), caspase inhibitor Z-VAD-FMK (1.06 mM), Z-YVAD-FMK (0.16 mM), or Z-DEVD-FMK (2 mM) before they were placed in constant light (3400 lux) for 24 hours. Additional controls included rats that were uninjected or were punctured with a dry needle. Electroretinograms were recorded before injection and 1 day after the cessation of exposure to constant light. A group of rats was killed for apoptotic cell detection in the outer nuclear layer. Fifteen days later, the remaining rats were killed for histology, and the outer nuclear layer (ONL) thickness was measured. Caspase-1, caspase-3, and calpain activities were measured before and 1 day after exposure to the damaging light.Results:ZVAD, YVAD, and DEVD inhibited caspase-1 and -3 activities, but not calpain activity, from the beginning and up to 1 day after light exposure. In untreated, dry needle-punctured, PBS, DMSO, and YVAD groups, light exposure significantly reduced retinal function and ONL thickness and increased by 51-fold the number of apoptotic cells. ZVAD and DEVD preserved retinal function to 86% and 78%, respectively, and reduced by three times the number of apoptotic photoreceptors. ONL thickness was more preserved in ZVAD (to 72%) than in DEVD (to 56%).Conclusions:In the authors' model of retinal degeneration, photoreceptor cells die through a caspase-dependent mechanism. However, the molecular events involved during and after light exposure seemed to implicate different proteases.
3. Fine particulate matter-induced lung inflammation is mediated by pyroptosis in mice
Jing Jiang, Zhen An, Jinge Du, Jingfang Zhang, Jie Song, Chunzhi Wang, Lin Zhang, Yanmei Ma, Weidong Wu, Juan Li Ecotoxicol Environ Saf . 2021 Aug;219:112351. doi: 10.1016/j.ecoenv.2021.112351.
Background:Exposure to ambient air-borne fine particulate matter (PM2.5) increases respiratory morbidity and mortality. The mechanisms underlying PM2.5-induced adverse effects remain unclear. This study aimed to uncover the molecular mechanisms of PM2.5-induced lung toxicity using a mouse model.Methods:Scanning electron microscopy and inductively coupled plasma mass spectrometry were used to examine and analyze PM2.5morphology and element compositions, respectively. Twenty four male mice were randomly divided into three groups: control (PBS), PM2.5(4.0 mg/kg b.w.), and PM2.5+ Z-YVAD-FMK. In the latter group, the pan-caspase inhibitor (Z-YVAD-FMK) was intraperitoneally injected into mice at a dose of 12.5 mg/kg body weight prior to intratracheal instillation of PM2.5(4.0 mg/kg b.w.) every other day for a total of 3 times (n = 8 in each group). Bronchoalveolar lavage fluids (BALFs) were collected 24 h after the last instillation of PM2.5. Levels of total proteins (TP), lactate dehydrogenase (LDH), IL-1β and IL-18 were analyzed for biomarkers of cell injury and inflammation. Additionally, histological alterations of lung tissues were assessed by hematoxylin-eosin staining. mRNA and protein expression of Caspase1, NLRP3 and GSDMD were examined by real-time fluorescent quantitative PCR and immunohistochemical staining.Results:Exposure to PM2.5increased levels of TP, LDH, IL-1β, IL-18 and inflammatory cell counts in lung. The mRNA and protein expression of Caspase1, NLRP3 and GSDMD were increased. Inhibition of the NALRP3/Caspase-1 signaling pathway ameliorated PM2.5-induced lung injury and inflammation, partially through suppressing pyroptosis in lung.Conclusion:PM2.5exposure induces lung injury and inflammation, which is mediated by the NALRP3/Caspase-1 signaling pathway.
