Department of Anesthesiology, Zhejiang Cancer Hospital. Hangzhou, Zhejiang Province, 310022, China.
Laboratory of Anesthesia and Perioperative Medicine, Hangzhou Institute of Medicine (HIM), Chinese Academy of Sciences, Hangzhou, Zhejiang 310018, China.
Int J Med Sci. 2023 Sep 18;20(12):1570-1583. doi: 10.7150/ijms.83207. eCollection 2023.
The aim of this study was to investigate whether calcium-sensing receptor (CaSR) was involved in HRF-mediated exacerbation of MI/R injury through NLRP3 inflammasome activation and pyroptosis. , a rat MI/R model was established by ligating the left coronary artery, and short-term HRF exposure was induced during reoxygenation. Then, TUNEL, H&E, Masson staining, immunohistochemical (IHC) and serum levels of lactate dehydrogenase (LDH) and creatine kinase isoenzyme (CK), as well as the expression levels of CaSR and pyroptosis-related proteins in heart tissues, were measured. H9c2 cells were cultured to create a hypoxia/reoxygenation (H/R) model and exposed to different concentrations of RF. After pretreatment with the CaSR activator gadolinium chloride (GdCl) and inhibitor NPS2143 in the H/R model and treatment with HRF, we compared cellular viability, TUNEL, cytosolic [Ca], the levels of LDH and CK, pyroptosis-related proteins and CaSR in H9c2 cells. We further researched the mechanisms of CaSR-mediated pyroptosis in the H/R+HRF model by CaSR-shRNA, Ac-YVAD-CMK, MCC950 and NAC. We found that HRF significantly increased CaSR expression, rate of cell death, levels of CK and LDH, and exacerbated pyroptosis in MI/R model. , HRF increased CaSR expression, decreased viability, enhanced cytosolic [Ca] and exacerbated pyroptosis in H/R cells. Pretreated with GdCl worsen these changes, and NPS2143, MCC950, Ac-YVAD-CMK, NAC and sh-CaSR can reversed these effects. Exposure to HRF for a short time exacerbates MI/R-induced injury by targeting CaSR to increase cytosolic [Ca] and ROS levels, which mediate the NLRP3 inflammasome and pyroptosis.
本研究旨在探讨钙敏感受体（CaSR）是否通过 NLRP3 炎性小体激活和细胞焦亡参与 HRF 介导的 MI/R 损伤加重。通过结扎左冠状动脉建立大鼠 MI/R 模型，并在再氧合期间诱导短暂的 HRF 暴露。然后，通过 TUNEL、H&E、Masson 染色、免疫组织化学（IHC）和血清乳酸脱氢酶（LDH）和肌酸激酶同工酶（CK）水平，以及心脏组织中 CaSR 和细胞焦亡相关蛋白的表达水平来测量。培养 H9c2 细胞以建立缺氧/复氧（H/R）模型，并暴露于不同浓度的 RF。在 H/R 模型和 HRF 处理中用 CaSR 激活剂钆氯化物（GdCl）和抑制剂 NPS2143 预处理后，我们比较了 H9c2 细胞的细胞活力、TUNEL、胞浆[Ca]、LDH 和 CK 水平、细胞焦亡相关蛋白和 CaSR。我们进一步通过 CaSR-shRNA、Ac-YVAD-CMK、MCC950 和 NAC 在 H/R+HRF 模型中研究了 CaSR 介导的细胞焦亡的机制。结果发现，HRF 显著增加了 MI/R 模型中 CaSR 的表达、细胞死亡率、CK 和 LDH 水平，并加重了细胞焦亡。结果发现，HRF 增加了 H/R 细胞中 CaSR 的表达，降低了细胞活力，增强了胞浆[Ca]，并加重了细胞焦亡。用 GdCl 预处理会加重这些变化，而 NPS2143、MCC950、Ac-YVAD-CMK、NAC 和 sh-CaSR 可以逆转这些效应。短暂暴露于 HRF 会通过靶向 CaSR 增加胞浆[Ca]和 ROS 水平来加重 MI/R 诱导的损伤，从而介导 NLRP3 炎性小体和细胞焦亡。
