血红素氧合酶1对大鼠心肌细胞急性损伤的保护作用

目的 了解血红素氧合酶1(HO-1)对脂多糖(LPS)诱导的心肌细胞损伤的保护作用及其机制. 方法分离培养SD乳鼠心肌细胞,根据细胞悬液中所加刺激物的不同分为对照组(A组):常规培养;LPS组(B组):培养液中加入终浓度为30 μmol/L的LPS,作用1 h;LPS+氯化血红素(heroin)组(C组):加入终浓度为5 μmol/L的heroin,作用1 h后再加入30 μmol/L的LPS作用1 h;LPS+ZnPP组(D组):加入终浓度为3 μmol/L的ZnPPⅨ,作用1 h后再加入30 μmol/L的LPS作用1 h.硫代巴比妥酸比色法及黄嘌呤氧化酶法测定各组心肌细胞乳酸脱氢酶(LDH)、丙二醛(MDA)及超氧化物歧化酶(SOD)含量;检测心肌细胞节律、存活率及凋亡率;用反转录-PCR法检测HO-1 mRNA表达;蛋白质印迹法检测心肌细胞HO-1、肿瘤坏死因子α(TNF-α)、核因子кB(NF-кB)的表达. 结果 B、C、D组LDH和MDA值分别为(113±15)、(79±13)、(154±22)U/L和(1.88±0.36)、(1.16±0.32)、(2.84±0.44)mmol/L,高于A组(69±10)U/L、(0.87±0.25)mmol/L,P<0.05],而以上3组的SOD值(17.8±1.8)、(22.5±2.4)、(13.4±1.5)U/mL]却低于A组(24.3±3.6)U/mL,P<0.05].B、C、D组心肌细胞节律,凋亡率均高于A组(P<0.05),存活率低于A组(P<0.05).B、C、D组心肌细胞HO-1 mRNA表达均高于A组(P<0.05),其中C组最高.B、C、D组心肌细胞HO-1、TNF-α、NF-кB值均高于A组(P<0.05),其中C组的HO-1值最高,D组TNF-α、NF-кB值最高. 结论 LPS对心肌细胞有明显的损伤作用.HO-1可能通过抗炎性反应、减轻氧化应激以及减少细胞凋亡途径,对心肌细胞产生保护作用.

The protection of Heme oxygenase-1 from acute cardiocyte injury in rats

Objective To observe the protection of Heme oxygenase-1 ( HO-1 ) from lipopolysaccha- ride (LPS) -induced eardiocyte injury and its mechanism. Methods Cardiocyte was isolated from SD neo- nate rat and cultured in vitro, and was divided into control group ( normal culture) , LPS group (with stim- ulation of 30 μmoL/L LPS for 1 hour), LPS + Hemin group( with same treatment to LPS group after stimula- tion of 5 μmoL/L Heroin for 1 hour ) ,and LPS + ZnPP group ( with same treatment to LPS group after stimu- lation of 3 μmoL/L ZnPP for 1 hour). The level of lactic-dehydrogenase( LDH), malondialdehyde (MDA), superoxide dismutase (SOD) were measured by thio-barbituric acid and xanthine oxidase techniques. The cell heart rhythm, survival rate and apoptosis rate were examined. The expressions of nuclear factor KB ( NF- кB) ,HO-1 and tumor necrosis factor-alpha (TNF-α) were measured with Western blotting. The HO-1 mR- NA was examined by RT-PCR. Results The level of LDH and MDA in LPS, LPS + Heroin, and LPS + ZnPP groups were(113±15), (79±13), (154±22)U/L, and(1.88±0.36),(1.16±0.32),(2.84± 0.44) mmoL/L respectively, which were all obviously higher than those in control group (69±10) U/L, (0.87±0.25) mmol/L, P <0.05]. The level of SOD in LPS, PS + Heroin, and LPS + ZnPP groups ( 17.8 ± 1.8, 22.5 ± 2.4, 13.4 ± 1.5 U/mL, respectively) was all obviously lower than that in control group (24.3 ± 3.6 U/mL, P < 0. 05 ). The apoptosis rate and heart rhythm were obviously higher and surviv- al rate significantly lower in LPS, LPS + Heroin, and LPS + ZnPP groups than those in control group ( P < 0. 05). The level of HO-1 mRNA in LPS, LPS + Heroin, and LPS + ZnPP groups was higher than that in con- trol group( P <0.01), among which LPS + Heroin group was the highest. The level of HO-1, TNF-α and NF-кB in LPS, LPS + Heroin, and LPS +ZnPP groups was higher than those in control group( P <0.05), among which the level of HO-1 protein in LPS + Heroin group was the highest, the level of TNF-α and NF- кB in LPS + ZnPP group was highest. Conclusion LPS can induce cardiocyte injury,which can be inhibi- ted through the anti-inflammatory, anti-oxidant, and anti-apoptosis functions by HO-1.