研究组蛋白去乙酰化酶抑制剂丙戊酸钠对50%总体表面积(TBSA)Ⅲ度烫伤大鼠脑损伤的保护作用。

采用随机数字表法将80只清洁级雄性SD大鼠随机分为4组：假伤对照组，单纯烫伤组，2-甲基-2-戊烯酸(2M2P)组和丙戊酸钠(VPA)组，每组20只。假伤对照组大鼠浸入37 ℃水浴中制备假伤模型，单纯烫伤组、2M2P组和VPA组采用96 ℃水浴浸泡大鼠背部15 s、双下肢15 s、腹部8 s，制作50%TBSA Ⅲ度烫伤模型。烫伤后即刻，2M2P组和VPA组分别腹腔注射300 mg/kg 2M2P和VPA，假伤对照组和单纯烫伤组分别腹腔注射3 mL/kg 0.9%氯化钠溶液。各组大鼠分别于烫伤后4、8 h 2个时间点以腹主动脉采血法被处死，断脑取脑组织，采用酶联免疫吸附测定(ELISA)检测脑组织S-100β蛋白表达水平，干/湿重法测定大鼠脑组织含水率，采用蛋白质印迹法检测脑组织中热休克蛋白(HSP)70、组蛋白H3赖氨酸9位乙酰化(H3K9ac)及缺氧诱导因子-1α(HIF-1α)的蛋白表达水平。另取80只大鼠用于观察各组烫伤后4、24 h生存率(大鼠分组、烫伤模型制备以及治疗方案同上)。数据比较采用单因素方差分析、t检验和χ²检验。

烫伤后4、8 h，4组大鼠脑组织S-100β蛋白水平和含水率比较，差异均有统计学意义(P<0.05)。烫伤后4、8 h，单纯烫伤组、2M2P组和VPA组大鼠脑组织S-100β蛋白表达水平和含水率与假伤对照组比较，差异均有统计学意义(P<0.05)；烫伤后4、8 h，2M2P组和VPA组大鼠S-100β蛋白表达水平和含水率与单纯烫伤组比较，差异均有统计学意义(P<0.05)；烫伤后4、8 h，VPA组脑组织S-100β蛋白表达水平分别为(0.54±0.05)、(0.79±0.05)μg/L，与2M2P组[(1.02±0.07)、(1.41±0.08) μg/L]比较均显著降低，差异均有统计学意义(t＝2.369、2.251，P<0.05)。烫伤后4、8 h，VPA组脑组织含水率分别为(78.62±0.58)%、(81.25±0.57)%，与2M2P组[(80.15±0.64)%、(83.15±0.61)%]比较均显著降低，差异均有统计学意义(t＝2.054、2.191，P<0.05)。烫伤后4、8 h，4组大鼠脑组织HSP70、H3K9ac、HIF-1α蛋白表达水平比较，差异均有统计学意义(P<0.05)。烫伤后4、8 h，单纯烫伤组、2M2P组和VPA组大鼠脑组织HSP70、H3K9ac、HIF-1α蛋白表达水平与假伤对照组比较，差异均有统计学意义(P<0.05)。烫伤后4、8 h，2M2P组和VPA组大鼠脑组织HSP70、H3K9ac、HIF-1α蛋白水平与单纯烫伤组比较，差异均有统计学意义(P<0.05)；烫伤后4、8 h，VPA组大鼠脑组织HSP70蛋白水平分别为0.63±0.05、0.81±0.04，与2M2P组(0.41±0.03、0.59±0.04)比较，VPA组大鼠脑组织HSP70蛋白水平升高更显著，差异均有统计学意义(t＝2.413、2.152，P<0.05)；烫伤后4、8 h，VPA组大鼠脑组织H3K9ac蛋白水平分别为1.26±0.09、1.58±0.07，与2M2P组(1.06±0.06、1.24±0.07)比较升高更显著，差异均有统计学意义(t＝2.031、2.167，P<0.05)；烫伤后4、8 h，VPA组HIF-1α蛋白表达水平分别为2.23±0.21、1.86±0.23，与2M2P组(3.01±0.24、2.74±0.21)比较均显著降低，差异均有统计学意义(t＝2.157、2.425，P<0.05)。烫伤后4、24 h，单纯烫伤组大鼠的存活率为80.0%(16/20)和0(0/20)；2M2P组为90%(18/20)和20%(4/20)；VPA组为100%(20/20)和40%(8/20)。烫伤后4、24 h，VPA组和2M2P组大鼠存活率均显著高于单纯烫伤组，差异均有统计学意义(χ²＝11.31、11.96，P<0.05)。烫伤后4、24 h，VPA组大鼠存活率均高于2M2P组，差异均有统计学意义(χ²＝11.32、12.05，P<0.05)。

VPA能减轻50%TBSAⅢ度烫伤大鼠的脑损伤并提高生存率，其机制可能与VPA降低脑组织S-100β的蛋白表达水平和脑组织含水率，促进脑组织HSP70和H3K9ac蛋白表达和抑制HIF-1α的活性有关。

To study the effect of histone deacetylase inhibitor valproate on cerebral injury of the rats with 50% total body surface area (TBSA) full thickness burn.

Eighty male clean-grade SD rats were divided into four groups according to the random number table method: sham injury control group, simple scald group, 2-methyl-2-pentenoic acid (2M2P) group and sodium valproate (VPA) group, with 20 rats in each group. Rats in the sham injury group were immersed in a 37 ℃ water bath to prepare a sham injury model. In the simple scald group, 2M2P group and VPA group, the rats were immersed in a 96 ℃ water bath for 15 s on the back, 15 s on both lower limbs, and 8 s on the abdomen to create 50% TBSA full thickness burn rats models. Immediately after scalding, the 2M2P group and the VPA group were intraperitoneally injected with 300 mg/kg 2M2P and VPA, respectively, and the sham injury control group and the simple scald group were intraperitoneally injected with 3 mL/kg 0.9% sodium chloride solution, respectively. Rats in each group were sacrificed by abdominal aorta blood sampling at 4 and 8 h after scalding, respectively. The expression level of S-100β protein in brain tissue was detected by enzyme-linked immunosorbent assay (ELISA), the water content of rat brain tissue was determined by dry/wet weight method, and the protein expression levels of heat shock protein (HSP) 70, histone H3 lysine 9 acetylation (H3K9ac) and hypoxia-inducible factor-1α (HIF-1α) in brain tissue were detected by Western blotting. Another 80 rats were used to observe the survival rate of 4 and 24 h after scald in each group (rat grouping, scald model prepare and treatment plan were the same as above). Data were compared with one-way ANOVA, t test and chi-square test.

At 4 and 8 h after scalding, there were statistically significant differences in the levels of S-100β protein and water content in the brain tissue of the four groups (P< 0.05). At 4 and 8 h after scalding, the expression levels of S-100β protein and water content in the brain tissue of the rats in the simple scald group, 2M2P group and VPA group were statistically significantly different from those in the sham-injured control group(P<0.05). At 4 and 8 h after scalding, the expression levels of S-100β protein and water content in the 2M2P group and the VPA group were statistically significantly different from those in the simple scalding group (P<0.05). At 4 and 8 h after scalding, the expression levels of S-100β protein in the brain tissue of the VPA group were (0.54±0.05), (0.79±0.05) μg/L, respectively, compared with [(1.02±0.07), (1.41±0.08) μg/L] in the 2M2P group were significantly decreased, and the differences were statistically significant (t=2.369, 2.251; P<0.05). At 4 and 8 h after scalding, the water content of brain tissue in the VPA group were (78.62±0.58)% and (81.25±0.57)%, respectively, which were significantly lower than those in the 2M2P group [(80.15±0.64)% and (83.15±0.61)%], the differences were statistically significant (t=2.054, 2.191; P<0.05). At 4 and 8 h after scalding, the protein expression levels of HSP70, H3K9ac and HIF-1α in the brain tissue of the four groups of rats were compared, and the differences were statistically significant (P<0.05). At 4 and 8 h after scalding, the protein expression levels of HSP70, H3K9ac and HIF-1α in the brain tissue of the rats in the simple scald group, the 2M2P group and the VPA group were statistically significantly different from those in the sham-injured control group (P<0.05). At 4 and 8 h after scalding, the protein levels of HSP70, H3K9ac and HIF-1α in the brain tissue of the 2M2P group and VPA group were statistically significantly different from those in the simple scalding group (P<0.05). At 4 and 8 h after scalding, the levels of HSP70 protein in the brain tissue of the VPA group were 0.63±0.05, 0.81±0.04, respectively, compared with the 2M2P group (0.41±0.03, 0.59±0.04), the level of HSP70 protein in the brain tissue of the rats in the VPA group increased significantly, and the differences were statistically significant (t=2.413, 2.152; P<0.05). At 4 and 8 h after scalding, the levels of H3K9ac protein in the brain tissue of the VPA group were 1.26±0.09, 1.58±0.07, which were higher than those in the 2M2P group (1.06±0.06, 1.24±0.07), the differences were statistically significant (t=2.031, 2.167; P<0.05); the expression levels of HIF-1α protein in the VPA group were 2.23±0.21 and 1.86±0.23, respectively, which were significantly lower than those in the 2M2P group (3.01±0.24, 2.74±0.21), the differences were statistically significant (t=2.157, 2.425; P<0.05). At 4 and 24 h after scalding, the survival rates of the rats in the simple scald group were 80.0% (16/20) and 0 (0/20); those in the 2M2P group were 90% (18/20) and 20% (4/20); the VPA group was 100% (20/20) and 40% (8/20). At 4 and 24 h after scalding, the survival rates of the rats in the VPA group and 2M2P group were significantly higher than those in the simple scald group, and the differences were statistically significant (χ²=11.31, 11.96; P<0.05). At 4 and 24 h after scalding, the survival rates of rats in the VPA group were higher than those in the 2M2P group, and the differences were statistically significant (χ²=11.32, 12.05; P<0.05).

VPA can reduce the brain injury of 50% TBSA full thickness burn rats and improve the survival rate. The mechanism may be related to VPA reduces the protein expression level of S-100β and the water content of brain tissue, promots the expression of HSP70 and H3K9ac proteins in brain tissue and inhibits the activity of HIF-1α.