摘要
目的探讨内质网应激（ERS）相关蛋白及其介导的细胞凋亡在大鼠难免性压疮形成中的作用。方法将40只雄性SD大鼠按照随机数字表法分为正常对照组和1个受压周期组、3个受压周期组、6个受压周期组、9个受压周期组，每组8只。本实验模拟临床压疮的处理，向大鼠大腿股薄肌部位施加22．47kPa压力，持续施压2．0h，放松0。5h，即1个受压周期。1个受压周期组大鼠模拟压疮处理1次；3个受压周期组大鼠1d内模拟压疮处理3次；6个受压周期组大鼠每天模拟压疮处理3次，持续2d；9个受压周期组大鼠每天模拟压疮处理3次，持续3d；正常对照组大鼠未进行施压处理。模拟完成后，处死各受压组大鼠，取受压中心部位肌肉组织行HE染色观察组织形态学变化；Hoeehst33258染色检测每平方毫米受压肌肉组织中凋亡细胞核数量；蛋白质印迹法检测ERS相关蛋白即结合蛋白（BIP）、蛋白质二硫键异构酶（PDI）、转录因子CCAAT增强子结合蛋白同源蛋白（CHOP）、半胱氨酸天冬氨酸蛋白酶12（caspase-12）的水平，数据以目的蛋白与内参照GAPDH灰度比值表示。正常对照组大鼠取相同部位肌肉组织进行上述检测。对数据进行单因素方差分析，两两比较行LSD-t检验。结果形态学观察显示，与正常对照组比较，各受压组大鼠局部受压肌肉组织呈现炎性细胞浸润、肌纤维溶解、空泡变性等病理改变。Hoechst33258荧光染色显示，1个受压周期组、3个受乐周期组、6个受压周期组、9个受压周期组大鼠每平方毫米受压肌肉组织中凋亡细胞核数量分别为（14．5±4．4）、（11．0±2．9）、（13．8±5．1）、（21．3±6．0）个，均明显多于正常对照组[（2．7±1．4）个，t值为4．223—6．000，P值均小于0．01]。蛋白质印迹法检测显示，正常对照组和1个受压周期组、3个受压周期组、6个受压周期组、9个受压周期组大鼠BIP蛋白表达分别为0．64±0．12、1．20±0．34、1．59±0．24、1．17±0．28、1．44±0．33，PDI蛋白表达分别为0．48±0．15、0．61±0．19、1．23±0．38、0．37±0．19、0．29±0．15。组间2项指标比较差异均有统计学意义（F值分别为5．32、7．95，P〈0．05或P〈0．01）。蛋白质印迹法检测显示，正常对照组和1个受压周期组、3个受压周期组、6个受压周期组、9个受压周期组大鼠CHOP蛋白表达分别为0．58±0．18、1．48±0．27、1．03±0．21、0．95±0．30、1．69±0．34，easpase—12蛋白表达分别为0．55±0．12、1．08±0．31、0．69±0．24、1．79±0．20、2．06±0．47。组间2项指标比较差异均有统计学意义（F值分别为8．17、15．48，P值均小于0．01）。结论ERS相关蛋白及其介导的细胞凋亡可能参与难免性压疮的形成。
Objective To explore the effects of endoplasmic reticulum stress （ERS） related pro- teins and their mediated apoptosis in the formation of deep tissue injury of pressure ulcer in rats. Methods Forty male Sprague-Dawley rats were divided into normal control group and groups A, B, C, D according to the random number table, with 8 rats in each group. Rats in group A were loaded with 22.47 kPa pressure with a special pressure apparatus for 2.0 h in the region over graciles, and then unloaded for 0.5 h. Rats ingroup B were treated with the same manoeuvre as that in group A for 3 times in one day. Rats in groups C and D were treated with the same manoeuvre as that in group B for 2 and 3 days. Rats in normal control group were free from pressure loading. Rats in groups A, B, C, and D were sacrificed after pressure load- ing, and then the central part of pressure loaded muscular tissues were harvested for observation of histomor- phological change with HE staining; apoptotic nucleoli per millimeter pressure loaded muscular tissue were counted with Hoechst 33258 staining; the levels of binding protein （BIP） , protein disulfide isomerase （PDI） , C/EBP homologous protein （CHOP） , and caspase-12 were assessed with Western blotting （ denoted as gray level ratio of target protein to GAPDH）. The were harvested for determination of all the indexes as same parts of graciles of rats in normal control group above. Data were processed with one-way analysis of variance, LSD- t test was applied for paired comparison. Results （ 1 ） Histomorpbological observation. Some pathological changes, including inflammatory cell infiltration, myofibers lysis, and vacuolar degenera- tion, etc. were observed in pressure loaded muscular tissue of rats in groups A, B, C, and D, but not in the same parts of graciles muscle of rats in normal control group. Compared with those in normal control group [ （2.7 ± 1.4） per millimeter muscular tissue ], the number of apoptotic nuclei was significantly increased in pressure loaded muscular tissue of rats in groups A, B, C, and D [（14.5±4.4）, （11.0±2.9）, （13.8± 5.1 ）, （21.3 ± 6.0） per millimeter pressure loaded muscular tissue, with t values from 4. 223 to 6. 000, P values all below 0.01 ）. （2） Western blotting. The protein expressions of BIP and PDI in rats of normal control group and groups A, B, C, D were respectively 0.64 ±0.12, 1.20 ±0.34, 1.59 ±0.24, 1.17 ± 0.28, 1.44±0.33; 0.48±0.15, 0.61±0.19, 1.23±0.38, 0.37±0.19, 0.29±0.15, and they showed significant statistical difference （ with F values respectively 5.32, 7.95, P 〈 0.05 or P 〈 0.01 ）. The protein expressions of CHOP and caspase-12 in rats of normal control group and groups A, B, C, D were respectively0.58±0.18, 1.48 ±0.27, 1.03 ±0.21, 0.95 ±0.30, 1.69 ±0.34; 0.55 ±0. 12, 1.08 ± 0.31, 0.69 ± 0.24, 1.79 ± 0.20, 2.06 ± 0.47, with significant statistical difference （ with F values respectively 8.17, 15.48, P values all below 0.01 ）. Conclusions ERS related proteins and their apop- totic pathway may play an important role in the formation of deep tissue injury of pressure ulcer in rats.
出处
《中华烧伤杂志》
CAS
CSCD
北大核心
2013年第5期448-453,共6页
Chinese Journal of Burns
基金
国家自然科学基金（81372064）
关键词
细胞凋亡
内质网
应激
压疮
深部组织损伤
Apoptosis
Endoplasmic reticulum
Stress
Pressure ulcer
Deep tissue injury
