硫化氢对压疮缺血/再灌注损伤大鼠的保护作用及机制

Protective effect of hydrogen sulfide on pressure ulcer rats with ischemia/reperfusion injury and its mecha-nisms

目的：探讨外源性硫化氢（ H2 S）对压疮缺血/再灌注（ I/R）损伤大鼠的保护作用及其相关机制。方法将40只SD大鼠按随机数字表法分为4组：正常对照组、I/R模型组及低、高剂量H2 S干预组,每组10只。低、高剂量H2 S干预组分别给予10、30μmol/kg的硫氢化钠（ NaHS）腹腔注射1周,正常对照组与模型组接受等量的9 g/L盐水,然后正常对照组不作受压处理,其余3组均于压疮装置上受压,3个I/R循环后处死大鼠,通过 HE 染色判断压疮病理改变程度,利用亚甲基分光光度法分析血清 H2 S 水平,行酶联免疫吸附法（ ELISA）检测血清肿瘤坏死因子-α（ TNF-α）、白细胞介素-6（ IL-6）、细胞间黏附分子-1（ ICAM-1）、髓过氧化物酶（ MPO）水平,测定肌肉匀浆液中丙二醛（ MDA）、超氧化物歧化酶（ SOD）、谷胱甘肽过氧化物酶（ GSH-Px）、过氧化氢酶（ CAT）活性,采用原位切口末端标记法（ TUNEL）计算肌细胞凋亡指数（ AI）,Western blot分析肌肉组织Bcl-2、Bax表达情况。结果与正常对照组比较,I/R模型组血清TNF-α、IL-6、ICAM-1、MPO水平、肌肉匀浆液中MDA活性、AI及肌肉组织 Bax 表达水平增加（P均〈0.01）,血清H2S水平、肌肉匀浆液SOD、GSH-Px、CAT活性及肌肉组织 Bcl-2表达水平降低（P均〈0.01）。与I/R模型组比较,经低、高剂量H2S干预治疗后,病理积分、血清TNF-α、IL-6、ICAM-1、MPO水平、肌肉匀浆液中MDA活性、AI及肌肉组织 Bax 表达水平逐渐减少（ P均〈0．05、0.01）,血清H2S水平、肌肉匀浆液SOD、GSH-Px、CAT活性及肌肉组织 Bcl-2表达水平逐渐增高（P均〈0．05、0.01）。结论外源性H2 S对压疮I/R损伤大鼠产生良好的保护作用,其机制可能与抑制炎性反应、增强抗氧化能力及减少细胞凋亡有关。

Objective To explore the protective effect and mechanism of exogenous hydrogen sulfide （ H2 S） on pressure ulcer rats with ischemia/reperfusion （ I/R） injury. Methods Forty SD rarts were randomly divided into 4 groups,namely normal control group,I/R model group,low-dose H2 S intervention group and high-dose H2 S interven-tion group. Each group had 10 animals. The low-and the high-dose H2 S intervention groups were injected intraperitone-ally 10 and 30μmol/kg of sodium hydrosulfide （ NaHS） for 1 week,respectively;however,the normal control group and I/R model group were treated with same volume 9 g/L saline. Then,the normal control group did not sustain any pres-sure,but other 3 groups were pressurized. After 3 circulations of ischemia/reperfusion,all rats were sacrificed,and path-ological changes in pressure ulcer rats were evaluated by HE staining. Serum H2 S contents were analyzed through meth-ylene spectrophotometric method. Serum levels of tumor necrosis factor-α（TNF-α）,interleukin-6 （IL-6）,intercellu-lar adhesion molecule -1 （ICAM-1）,and myeloperoxidase （MPO） were detected by enzyme linked immunosorbent assay （ ELISA ） . The activities of malondialdehyde （ MDA ） , superoxide dismutase （ SOD ） , glutathione-peroxidase （ GSH-Px） and catalase （ CAT） in the homogenate of muscle were examined. Apoptotic index （ AI） was calculated by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling （ TUNEL） . The expressions of Bcl-2 and Bax in muscle tissue were evaluated by Western blot. Results In comparison with the normal control group,serum TNF-α, IL-6,ICAM-1 and MPO levels,MDA activity in the homogenate of muscles,AI and Bax expression level were in-creased （all P〈0. 01）,while serum H2S levels,the activities of SOD,GSH-Px and CAT as well as Bcl-2 expression levels were reduced in the I/R model group （all P〈0. 01）. Upon intervention therapy of low-or high-dose H2S,path-ological scores,serum TNF-α,IL-6,ICAM-1 and MPO levels,MDA activity in the homogenate of muscle,AI and Bax expression levels were gradually reduced while serum H2 S levels,the activities of SOD,GSH-Px and CAT and Bcl-2 expression levels were gradually enhanced when compared to those of the I/R model group （all P〈0. 05,0. 01）. Con-clusions Exogenous H2 S has better protection against pressure ulcer rats with I/R injury. The mechanism may be as-sociated with inhibition of inflammatory reaction and enhancement of antioxidative ability as well as reduction of apopto-sis.