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[大鼠严重烫伤早期胰岛素分泌及其信号转导机制的变化]

[Changes of insulin secretion and its signal transduction mechanism at early stage of severe scald in rats].

作者信息

Zhang B H, Shen Z A, Sun P C, Zheng B, Li D W, Liu Z X, Xu S B, Sun J C, Zhang W

机构信息

Department of Burns and Plastic Surgery, the Fourth Medical Center of PLA General Hospital, Beijing 100048, China.

出版信息

Zhonghua Shao Shang Za Zhi. 2020 Apr 20;36(4):280-287. doi: 10.3760/cma.j.cn501120-20190702-00289.

Abstract

To observe the changes of insulin secretion in the early stage of severe scald in rats, and to explore its signal transduction mechanism. 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. (1) At PIH 72, the rat FBG levels in SIA and SIB groups were normal and similar (>0.05). Compared with the levels of those two groups, the rat FBG level in SA group was increased significantly (<0.01), while the level in SB group showed no obvious change (>0.05). Compared with that in SA group, the rat FBG level in SB group was decreased significantly (<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 (>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 (<0.01), while the proportion of insulin vesicles was increased significantly (<0.01); the number of docked granules per 10 μm membrane of rat islet beta cells in SB group was obviously decreased (<0.05), while the proportion of insulin vesicles did not change obviously (>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 (<0.01), while the proportion of insulin vesicles was significantly decreased (<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 (<0.05) but was decreased significantly in SA group (<0.01), while the level in SB group did not change obviously (>0.05). Compared with the level in SIB group, the phosphorylation levels of Akt in SA and SB groups were decreased significantly (<0.01). Compared with that in SA group, the phosphorylation level of Akt in SB group was increased significantly (<0.05). 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.

摘要

观察大鼠严重烫伤早期胰岛素分泌的变化,并探讨其信号转导机制。将24只7周龄雄性Wistar大鼠采用随机数字表法分为单纯假伤(SIA)组、假伤+BPV(HOpic)(SIB)组、单纯烫伤(SA)组和烫伤+BPV(HOpic)(SB)组,每组6只。SA组和SB组大鼠通过将腹部浸入94℃热水6秒、背部浸入12秒造成50%体表面积的全层烫伤。SIA组和SIB组大鼠分别将背部和腹部浸入37℃温水中6秒和12秒进行假伤。伤后0(即刻)至2天,SB组和SIB组大鼠每天腹腔注射磷脂酰肌醇3激酶/蛋白激酶B(PI3K/Akt)信号通路增强剂BPV(HOpic)溶液(0.5mg/mL),剂量为0.6mg/kg,SA组和SIA组大鼠每天腹腔注射相同体积的二甲基亚砜。伤后72小时(PIH 72),采集大鼠尾血用血糖仪测定空腹血糖(FBG),取大鼠胰腺组织样本,苏木精-伊红染色观察胰岛病理表现,透射电镜观察胰岛β细胞超微结构,计数每10μm胰岛β细胞膜对接颗粒数并计算胰岛素囊泡比例,Western blotting检测胰腺PI3K/Akt信号通路中Akt的磷酸化水平。数据采用单因素方差分析和最小显著差法检验。(1)PIH 72时,SIA组和SIB组大鼠FBG水平正常且相近(>0.05)。与这两组相比,SA组大鼠FBG水平显著升高(<0.01),而SB组水平无明显变化(>0.05)。与SA组相比,SB组大鼠FBG水平显著降低(<0.01)。(2)PIH 72时,SIA组和SIB组大鼠胰岛形态完整,胰岛细胞分布规则。与SIA组和SIB组相比,SA组大鼠胰岛形态不完整,胰岛内胰岛素囊泡常见,胰岛细胞分布不规则,部分胰岛β细胞胞质淡染或呈半透明状;SB组胰岛形态无明显变化。与SA组相比,SB组胰岛形态相对完整,胰岛内胰岛素囊泡较少见,胰岛细胞分布相对规则,胰岛β细胞淡染或半透明胞质较少。(3)PIH 72时,SIA组和SIB组大鼠每10μm胰岛β细胞膜对接颗粒数及胰岛素囊泡比例相近(>0.05)。与SIA组和SIB组相比,SA组大鼠每10μm胰岛β细胞膜对接颗粒数显著减少(<0.01),而胰岛素囊泡比例显著增加(<0.01);SB组大鼠每10μm胰岛β细胞膜对接颗粒数明显减少(<0.05),而胰岛素囊泡比例无明显变化(>0.05)。与SA组相比,SB组大鼠每10μm胰岛β细胞膜对接颗粒数显著增加(<0.01),而胰岛素囊泡比例显著降低(<0.01)。(4)PIH 72时,SIA、SIB、SA和SB组Akt的磷酸化水平分别为0.91±0.03、0.98±0.03、0.78±0.08和0.87±0.08。与SIA组相比,SIB组Akt磷酸化水平明显升高(<0.05),而SA组显著降低(<0.01),SB组水平无明显变化(>0.05)。与SIB组相比,SA组和SB组Akt磷酸化水平显著降低(<0.01)。与SA组相比,SB组Akt磷酸化水平显著升高(<0.05)。大鼠严重烫伤早期,胰腺PI3K/Akt信号通路活性及胰岛素分泌功能降低。提高大鼠胰腺PI3K/Akt信号通路活性可改善胰岛素分泌功能,恢复血糖生理水平。

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