【摘要】：目的研究探討褪黑素對(duì)脂多糖(LPS)誘導(dǎo)心肌肥厚的保護(hù)作用及機(jī)制。方法 (1)培養(yǎng)的心肌細(xì)胞分為五組:對(duì)照組、LPS組、LPS+褪黑素(1.5mg/L)組、LPS+褪黑素(3mg/L)組、LPS+褪黑素(6mg/L)組,在倒置相差顯微鏡下觀察心肌細(xì)胞形態(tài)學(xué)改變;(2)通過(guò)RC DCTM Protein Assay Kit檢測(cè)各組心肌細(xì)胞蛋白水平。(3)通過(guò)酶聯(lián)免疫吸附試驗(yàn)(ELISA)檢測(cè)各組心肌細(xì)胞腫瘤壞死因子(TNF-α)的分泌水平;(4)利用Realtime PCR和Western blot檢測(cè)各組心肌細(xì)胞腫瘤壞死因子受體1(TNFR1)基因和蛋白表達(dá)水平變化;(5)采用Ca2+探針Fura-2/AM在影像系統(tǒng)下記錄各組心肌細(xì)胞Ca2+內(nèi)流情況。(6)Western blot檢測(cè)各組心肌細(xì)胞內(nèi)CaN和CaMKⅡδB蛋白水平變化。結(jié)果 LPS能夠誘導(dǎo)心肌肥厚、促進(jìn)TNF-a的分泌和TNFR1的表達(dá)及增加Ca2+內(nèi)流。在LPS誘導(dǎo)損傷之前預(yù)先給予不同濃度的褪黑素,能夠減弱LPS誘導(dǎo)的心肌肥厚,降低心肌細(xì)胞TNF-a的分泌和TNFR1,Ca N和Ca MKⅡδB的表達(dá)和Ca2+內(nèi)流,并且在濃度(1.5mg/L、3mg/L、6mg/L)范圍內(nèi)呈劑量依賴(lài)性。結(jié)論 在體外試驗(yàn)中褪黑素通過(guò)下調(diào)TNF-α和TNFR1的表達(dá)和維持Ca2+內(nèi)穩(wěn)態(tài)拮抗LPS誘導(dǎo)的心肌肥厚,發(fā)揮保護(hù)作用。
[Abstract]:Objective to study the protective effect and mechanism of melatonin on myocardial hypertrophy induced by lipopolysaccharide (LPS). Methods (1) the cultured cardiomyocytes were divided into five groups: control group, LPS melatonin (1.5mg/L) group, LPS melatonin (3mg/L) group, LPS melatonin (6mg/L) group. The morphological changes of cardiomyocytes were observed under inverted phase contrast microscope. (2) the protein level of cardiomyocytes in each group was detected by RC DCTM Protein Assay Kit. (3) the secretion of tumor necrosis factor 偽 (TNF- 偽) in cardiomyocytes was detected by enzyme-linked immunosorbent assay (ELISA). (4) the expression of tumor necrosis factor receptor 1 (TNFR1) gene and protein in cardiomyocytes of each group were detected by Realtime PCR and Western blot. (5) the Ca2 influx of cardiomyocytes in each group was recorded by Ca2 probe Fura-2/AM under imaging system. (6) the levels of CaN and CaMK 鈪，

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