■由于血液动力学超负荷导致的心肌细胞肥大最终导致心力衰竭。水蛭素已被广泛用于心血管疾病的治疗,NLRP3炎性体被证明可诱导心肌细胞焦亡。然而,其抑制心肌细胞肥大的机制尚不清楚.
■基于NLRP3炎性体激活和线粒体自噬,探讨水蛭素抑制心肌细胞肥大的机制。
■1μMAngII用于H9C2细胞的心脏肥大建模,和细胞活力通过CCK-8测定定量,以筛选合适的水蛭素作用浓度。之后,我们用0、0.3、0.6和1.2mM水蛭素培养AngII诱导的H9C2细胞24小时,分别。接下来,我们用差别定标记H9C2细胞,并用荧光显微镜观察。IL-1β,IL-18,IL-6,TNF-α,ANP,BNP,β-MHC,和mtDNA通过qRT-PCR分析;ROS通过流式细胞术定量;SOD,MDA,用ELISA法检测GSH-Px;NLRP3、ASC、caspase-1,pro-caspase-1,IL-1β,IL-18,PINK-1,Parkin,Beclin-1,LC3-Ⅰ,LC3-Ⅱ,p62,通过蛋白质印迹定量。
■发现水蛭素减少了AngII诱导的H9C2细胞的浅表面积,抑制了AngII诱导的ANP上调,BNP,和β-MHC。此外,水蛭素下调NLRP3炎性体相关细胞因子的表达,含有IL-1β,IL-18、IL-6、TNF-α。它还下调了mtDNA和ROS的表达,NLRP3炎性体活化相关蛋白的表达水平降低,包括NLRP3,ASC,caspase-1,pro-caspase-1,IL-1β,IL-18;并增加PINK-1,Parkin,Beclin-1,LC3-Ⅱ/LC3-Ⅰ,在AngII诱导的H9C2细胞中p62。
■水蛭素促进了线粒体自噬的过程,抑制炎症和氧化应激的发展,并抑制NLRP3炎性体和PINK-1/Parkin通路的激活。
■水蛭素具有抑制心脏肥大的活性,可能受益于NLRP3炎性体的抑制和PINK-1/Parkin相关线粒体自噬的激活。
UNASSIGNED: Cardiomyocyte hypertrophy due to hemodynamic overload eventually leads to heart failure.
Hirudin has been widely used in the treatment of cardiovascular diseases and NLRP3 inflammasome was proven to induce cardiomyocyte pyroptosis. However, the mechanism by which it inhibits cardiomyocyte hypertrophy remains unclear.
UNASSIGNED: To explore the mechanism of
hirudin inhibiting cardiomyocyte hypertrophy based on NLRP3 inflammasome activation and mitophagy.
UNASSIGNED: 1 μM AngII was used for cardiac hypertrophy modeling in H9C2 cells, and cell viability was quantified by CCK-8 assay to screen the appropriate action concentrations of hirudin. After that, we cultured AngII induced-H9C2 cells for 24 h with 0, 0.3, 0.6, and 1.2 mM hirudin, respectively. Next, we marked H9C2 cells with phalloidine and observed them using fluorescence microscope. IL-1β, IL-18, IL-6, TNF-α, ANP, BNP, β-MHC, and mtDNA were analyzed by qRT-PCR; ROS were quantified by Flow cytometry; SOD, MDA, and GSH-Px were detected by ELISA; and proteins including NLRP3, ASC, caspase-1, pro-caspase-1, IL-1β, IL-18, PINK-1, Parkin, beclin-1, LC3-Ⅰ, LC3-Ⅱ, p62, were quantified by western blotting.
UNASSIGNED: It was discovered that
hirudin reduced the superficial area of AngII-induced H9C2 cells and inhibited the AngII-induced up-regulation of ANP, BNP, and β-MHC. Besides,
hirudin down-regulated the expressions of NLRP3 inflammasome-related cytokines, containing IL-1β, IL-18, IL-6, TNF-α. It also down-regulated the expression of mtDNA and ROS, decreased the expression levels of NLRP3 inflammasome activation related proteins, including NLRP3, ASC, caspase-1, pro-caspase-1, IL-1β, IL-18; and increased the expressions of PINK-1, Parkin, beclin-1, LC3-Ⅱ/LC3-Ⅰ, p62 in AngII-induced H9C2 cells.
UNASSIGNED: Hirudin promoted the process of mitophagy, inhibited the development of inflammation and oxidative stress, and inhibited the activation of the NLRP3 inflammasome and the PINK-1/Parkin pathway.
UNASSIGNED: Hirudin has the activity to suppress cardiac hypertrophy may benefit from the inhibition of NLRP3 inflammasome and activating of PINK-1/Parkin related-mitophagy.