Changes of insulin secretion and its signal transduction mechanism at early stage of severe scald in rats
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摘要: 目的 观察严重烫伤大鼠早期胰岛素分泌功能变化情况并探讨其信号转导机制。 方法 将24只7周龄雄性Wistar大鼠按随机数字表法分为单纯假伤组、假伤+BPV(HOpic)组、单纯烫伤组和烫伤+BPV(HOpic)组,每组6只。单纯烫伤组、烫伤+BPV(HOpic)组大鼠在94 ℃热水中浸浴腹部6 s、背部12 s,造成50%体表总面积Ⅲ度烫伤;单纯假伤组、假伤+BPV(HOpic)组大鼠在37 ℃温水中浸浴腹部6 s、背部12 s,模拟致伤。伤后0(即刻)~2 d,烫伤+BPV(HOpic)组、假伤+BPV(HOpic)组大鼠每天一次性腹腔注射0.5 mg/mL磷脂酰肌醇3-激酶/蛋白激酶B(PI3K/Akt)信号通路增强剂BPV(HOpic)溶液0.6 mg/kg,单纯烫伤组、单纯假伤组大鼠每天一次性腹腔注射等体积二甲基亚砜。伤后72 h,剪尾法取大鼠尾血,用血糖仪测定空腹血糖;取大鼠胰腺组织,苏木精-伊红染色观察胰岛组织病理学表现,透射电镜观察胰岛β细胞超微结构以计数每10微米细胞膜上锚接胰岛素颗粒并计算胰岛素空泡占比,蛋白质印迹法检测胰腺PI3K/Akt信号通路中Akt磷酸化水平。对数据行单因素方差分析和LSD检验。 结果 (1)伤后72 h,单纯假伤组与假伤+BPV(HOpic)组大鼠空腹血糖水平均正常且相近(
P >0.05),与该2组比较,单纯烫伤组大鼠空腹血糖水平显著升高(P <0.01),烫伤+BPV(HOpic)组大鼠空腹血糖水平没有明显变化(P >0.05)。与单纯烫伤组比较,烫伤+BPV(HOpic)组大鼠空腹血糖水平显著降低(P <0.01)。(2)伤后72 h,单纯假伤组与假伤+BPV(HOpic)组大鼠胰岛形态完整且胰岛细胞排列整齐,与该2组比较,单纯烫伤组大鼠胰岛形态不完整,胰岛内空泡样变多见,细胞排列不规则,部分胰岛β细胞胞质淡染或透亮;烫伤+BPV(HOpic)组大鼠胰岛形态变化不明显。与单纯烫伤组比较,烫伤+BPV(HOpic)组大鼠胰岛形状较为完整,胰岛内空泡样变较少,细胞排列较规则,胰岛β细胞胞质淡染或透亮较少。(3)伤后72 h,单纯假伤组与假伤+BPV(HOpic)组大鼠胰岛β细胞中每10微米细胞膜上锚接胰岛素颗粒数和胰岛素空泡占比均相近(P >0.05),与该2组比较,单纯烫伤组大鼠胰岛β细胞中每10微米细胞膜上锚接胰岛素颗粒数显著减少(P <0.01),胰岛素空泡占比显著增加(P <0.01);烫伤+BPV(HOpic)组大鼠胰岛β细胞中每10微米细胞膜上锚接胰岛素颗粒数明显减少(P <0.05),胰岛素空泡占比没有明显变化(P >0.05)。与单纯烫伤组比较,烫伤+BPV(HOpic)组大鼠胰岛β细胞中每10微米细胞膜上锚接胰岛素颗粒数显著增多(P <0.01),胰岛素空泡占比显著减少(P <0.01)。(4)伤后72 h,单纯假伤组、假伤+BPV(HOpic)组、单纯烫伤组、烫伤+BPV(HOpic)组大鼠胰腺Akt磷酸化水平分别为0.91±0.03、0.98±0.03、0.78±0.08、0.87±0.08。与单纯假伤组比较,假伤+BPV(HOpic)组大鼠胰腺Akt磷酸化水平明显升高(P <0.05),单纯烫伤组大鼠胰腺Akt磷酸化水平显著降低(P <0.01),烫伤+BPV(HOpic)组大鼠胰腺Akt磷酸化水平无明显变化(P >0.05);与假伤+BPV(HOpic)组比较,单纯烫伤组与烫伤+BPV(HOpic)组大鼠胰腺Akt磷酸化水平均显著降低(P <0.01);与单纯烫伤组比较,烫伤+BPV(HOpic)组大鼠胰腺Akt磷酸化水平明显升高(P <0.05)。 结论 严重烫伤大鼠早期胰腺PI3K/Akt信号通路活性降低,胰岛素分泌功能下降;提高胰腺PI3K/Akt信号通路活性可以改善早期胰岛素分泌功能,恢复血糖生理水平。-
关键词:
- 烧伤 /
- 胰岛素 /
- 胰腺 /
- 血糖 /
- 磷脂酰肌醇3-激酶/蛋白激酶B信号通路 /
- BPV(HOpic)
Abstract: Objective To observe the changes of insulin secretion in the early stage of severe scald in rats, and to explore its signal transduction mechanism. Methods Twenty-four male Wistar rats aged 7 weeks were divided into sham injury alone (SIA) group, sham injury+ BPV (HOpic) (SIB) group, scald alone (SA) group, and scald+ BPV (HOpic) (SB) group using the random number table, with 6 rats in each group. Full-thickness scald of 50% total body surface area was inflicted in rats of SA and SB groups by a 6-s immersion of the abdomen and a 12-s immersion of the back in 94 ℃ hot water. Rats in SIA and SIB groups received sham injuries through immersion of the back and abdomen in 37 ℃ warm water for 6 and 12 seconds respectively. From 0 (immediately) to 2 day (s) after injury, the rats in groups SB and SIB were intraperitoneally injected with the phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) signaling pathway enhancer BPV (HOpic) solution (0.5 mg/mL) at the dosage of 0.6 mg/kg once a day, and the rats in groups SA and SIA were intraperitoneally injected with the same volume of dimethyl sulfoxide once a day. At post injury hour (PIH) 72, the tail blood of rats was sampled for measuring fasting blood glucose (FBG) with a glucometer, and the pancreatic tissue samples of rats was harvested for observing the pathological manifestations of islets by hematoxylin-eosin staining, counting the docked granules per 10 μm membrane of islet beta cells and calculating the proportion of insulin vesicles through the observation of the ultrastructure of islet beta cells by transmission electron microscope, and detecting the phosphorylation level of Akt in the pancreatic PI3K/Akt signaling pathway by Western blotting. Data were statistically analyzed with one-way analysis of variance and least significant difference test. Results (1) At PIH 72, the rat FBG levels in SIA and SIB groups were normal and similar (P >0.05). Compared with the levels of those two groups, the rat FBG level in SA group was increased significantly (P <0.01), while the level in SB group showed no obvious change (P >0.05). Compared with that in SA group, the rat FBG level in SB group was decreased significantly (P <0.01). (2) At PIH 72, the morphology of rat islets was complete and the islet cells distributed regularly in SIA and SIB groups. Compared with those in SIA and SIB groups, the morphology of rat islets was incomplete, the insulin vesicles in islets were common, the islet cells distributed irregularly, and the cytoplasm of some islet beta cells was lightly stained or translucent in SA group; the morphology of islets in SB group did not change obviously. Compared with those in SA group, the morphology of islets was comparatively complete, the insulin vesicles in islets were less common, the islet cells distributed comparatively regularly, and the lightly stained or translucent cytoplasm of islet beta cells was less in SB group. (3) At PIH 72, the number of docked granules per 10 μm membrane of rat islet beta cells and the proportion of insulin vesicles in SIA and SIB groups were similar (P >0.05). Compared with those in SIA and SIB groups, the number of docked granules per 10 μm membrane of rat islet beta cells in SA group was decreased significantly (P <0.01), while the proportion of insulin vesicles was increased significantly (P <0.01); the number of docked granules per 10 μm membrane of rat islet beta cells in SB group was obviously decreased (P <0.05), while the proportion of insulin vesicles did not change obviously (P >0.05). Compared with those in SA group, the number of docked granules per 10 μm membrane of rat islet beta cells in SB group was significantly increased (P <0.01), while the proportion of insulin vesicles was significantly decreased (P <0.01). (4) At PIH 72, the phosphorylation levels of Akt in SIA, SIB, SA, and SB groups were 0.91±0.03, 0.98±0.03, 0.78±0.08, and 0.87±0.08, respectively. Compared with that in SIA group, the phosphorylation level of Akt was increased obviously in SIB group (P <0.05) but was decreased significantly in SA group (P <0.01), while the level in SB group did not change obviously (P >0.05). Compared with the level in SIB group, the phosphorylation levels of Akt in SA and SB groups were decreased significantly (P <0.01). Compared with that in SA group, the phosphorylation level of Akt in SB group was increased significantly (P <0.05). Conclusions At the early stage post severe scald in rats, the activity of the pancreatic PI3K/Akt signaling pathway and the function of insulin secretion are reduced. Improving the activity of the pancreatic PI3K/Akt signaling pathway in rats can ameliorate the function of insulin secretion and recover the physiological level of blood glucose.
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