高温高湿环境对大鼠汗腺影响的实验研究

发布时间：2018-03-08 14:17

  本文选题：高温高湿　切入点：抗氧化　出处：《广州中医药大学》2017年硕士论文　论文类型：学位论文

【摘要】：背景:汗腺排泄汗液的功能在机体稳态中具有不可替代的作用,是人体体温调节主要方法。汗液的主要成份98～99%是水,1~2%为无机盐和有机物,汗液的有效蒸发是汗液持续分泌的必要前提。当空气相对湿度增大,其中的水分越接近饱和,人体皮肤表面汗液的蒸发就会困难,若同时周围环境温度升高,则机体热量不易消散,汗液的分泌是降低机体核心温度的重要途径,一旦该途径受阻,核心温度升高,机体的内部器官也将受到热损伤。湿热环境通常是指环境温度在36℃以上,相对湿度超过60%[1]。临床资料显示,高温高湿环境常常可引发中暑,中暑患者可出现高热、皮肤干燥、情绪错乱,甚至全身惊厥发作的症状[2]。其中皮肤干燥的症状则意味着汗液分泌减少,减少的原因以及高温高湿环境对汗腺功能的影响机制尚不清楚。本实验观察高温高湿环境对Sprague-Dawley(SD)大鼠足垫皮肤汗腺细胞功能的影响,并探讨汗腺细胞功能发生变化的可能机制。第一部分高温高湿环境对大鼠汗腺细胞影响的初步探讨目的:初步探讨高温高湿环境对汗腺功能的影响并对其可能的机制进行研究。方法:14只Sprague-Dawley(SD)大鼠随机分为常温常湿(NTH)组和高温高湿(HTH)组,每组7只,HTH组大鼠进行高温高湿干预2h,NTH组室温放置2h,72h后1%戊巴比妥钠麻醉,收集大鼠心脏血液,检测血液中乙酰胆碱(ACH)的水平;剥离左后足底皮肤用10%福尔马林固定,应用HE染色观察组织形态和免疫组织化学法检测乙酰胆碱转移酶(ChAT)的表达;剥离右后足底皮肤,标本-80℃冻存,应用生物化学方法检测组织总超氧化物歧化酶(SOD)活力以及蛋白免疫印迹(Western Blot)法检测M3受体的表达。结果:光镜下HTH组汗液分泌部的上皮细胞胞质中空泡减少,且伴有汗腺细胞凋亡增多,HTH组的凋亡指数为(70.7±14.0)%,显著高于NTH组[(58.6±11.7)%],差异有统计学意义(t=2.478,P=0.021);Western Blot的结果显示,HTH组应激后M3受体的相对表达量为(1.21±0.16)%,显著高于NTH组[(1.81 士0.41)%,t=2.711,P=0.035];进一步检测到HTH组SD大鼠血清中乙酰胆碱的表达明显弱于NTH组;并且HTH组超氧化物歧化酶(SOD)活力为(13.65 ±5.14)U/mg,较NTH组[(25.89± 11.05)U/mg]显著降低(t=2.633,P=0.023)。结论:高温高湿环境可引起汗腺结构形态变化,其对汗腺细胞的损伤可能是由于组织抗氧化能力减退导致。第二部分维生素E保护高温高湿应激大鼠汗腺细胞的研究目的:探讨高温高湿应激后维生素E(VE)是否能改善皮肤组织的抗氧化能力,从而减少高温高湿应激对汗腺细胞的损害。方法:21只SD大鼠随机分为常温常湿(NTH)组,高温高湿(HTH)组和VE干预高温高湿(VHTH)组,每组7只。HTH组和VHTH组大鼠进行高温高湿干预2h,干预后HTH组给予生理盐水1ml灌胃,VHTH组给予VE灌胃(100IU/kgVE配置生理盐水1ml);NTH组室温放置2h后给予生理盐水1ml灌胃。72h后1%戊巴比妥钠麻醉,剥离左后足底皮肤用10%福尔马林固定,应用HE染色观察组织形态、免疫组织化学法检测天冬氨酸特异性半胱氨酸蛋白酶-3(Caspase-3)和热休克蛋白70(HSP70)的表达;剥离右后足底皮肤,标本-80℃冻存,应用生物化学方法检测组织总超氧化物歧化酶(SOD)活力以及蛋白免疫印迹(Western Blot)法检测HSP70的表达。结果:光镜下可观察到经高温高湿干预的HTH组和VHTH组皮肤汗腺分泌部腺腔扩张,NTH组分泌部腔隙较小;但是HTH组分泌部腺上皮细胞排列疏松,上皮细胞核下空泡减少,核仁淡染,NTH组和VHTH组分泌部腺上皮细胞排列整齐,分泌部腔隙较小,上皮细胞核下空泡分泌旺盛(核下空泡较多),腺上皮胞浆丰富,核仁深染;VHTH组组织 SOD 的活性为(35.18 士 8.19)U/mg,与 HTH 组[(13.65 ±5.14)U/mg]相比显著升高,与NTH组[(25.90±11.05)U/mg]相比无显著性差异;免疫组织化学法检测组织Caspase-3和HSP70的表达结果显示VHTH组的表达水平均显著高于HTH组,且与NTH组无显著差异;蛋白免疫印迹(Western Blot)法检测组织HSP70的表达,分析结果与免疫组织化学法的检测结果相一致。结论:VE能够改善高温高湿环境引起的氧化还原失衡,减少湿热应激对大鼠汗腺的损害,对汗腺有保护作用。
[Abstract]:Background: the excretion of sweat gland function plays an irreplaceable role in the homeostasis of body, is the main method of regulating body temperature. The main ingredients of sweat 98 ~ 99% water, 1~2% as inorganic salt and organic matter, effective evaporation sweat is necessary to sweat secreted continuously. When the relative humidity increases, the water the more close to saturation, human skin surface evaporation of sweat will be difficult, if the ambient temperature rises, the body is not easy to dissipate heat, sweat secretion is an important way to reduce the body's core temperature, once the way is blocked, the core temperature rises, the body's internal organs will also be subject to thermal damage. Usually refers to the hot and humid environment the ambient temperature is over 36 DEG C, relative humidity is more than 60%[1]. clinical data showed that the environment can often lead to heat stroke in high temperature and high humidity, heat stroke can occur in patients with high fever, dry skin, emotional disorders, and even systemic Seizure symptoms including [2]. dry skin symptoms were meant to reduce the secretion of sweat, and the reason for the reduction of high temperature and high humidity on the sweat gland function influence mechanism is not clear. The effects of high temperature and high humidity on the Sprague-Dawley (SD) effect of sweat gland cell function foot pad of rats, and to investigate the possible mechanism of sweat gland cell function change. Preliminary study on the effect of the first part of high temperature and high humidity on rat sweat gland cells Objective: To investigate the high temperature and high humidity on the sweat gland function effect and to study its possible mechanism. Methods: 14 Sprague-Dawley (SD) rats were randomly divided into normal group (NTH) and high temperature wet high humidity (HTH) group, 7 rats in each group, the rats in group HTH of high temperature and high humidity intervention 2h, NTH group for 2h at room temperature, 1% pentobarbital sodium 72h after anesthesia, the blood collecting heart of rats, detection of acetyl choline in blood (ACH) level After stripping; left plantar skin with 10% formalin fixed, HE staining was used to observe morphology and immunohistochemical detection of choline acetyltransferase (ChAT) expression after stripping; right plantar skin specimens, -80 C frozen by biochemical method to determine the total superoxide dismutase (SOD) activity and immune protein Western blot (Western Blot) was used to detect the expression of M3 receptor. Results: in group HTH, the secretion of sweat vacuoles in the cytoplasm of epithelial cells and is associated with reduced light, sweat gland cells apoptosis, the apoptosis index in HTH group was (70.7 + 14)%, significantly higher than the NTH Group [(58.6 + 11.7) is detected statistically significant difference (t=2.478, P=0.021); Western Blot showed that HTH group after stress, the relative expression of M3 receptor was (1.21 + 0.16)%, significantly higher than the NTH Group [(1.81 + 0.41)%, t=2.711, P=0.035]; further to detect serum HTH acetylcholine SD rats in group table Damien was weaker than that of NTH group and HTH group; superoxide dismutase (SOD) activity was (13.65 + 5.14) U/mg, compared with the NTH Group [(25.89 + 11.05) U/mg] decreased significantly (t=2.633, P=0.023). Conclusion: high temperature and high humidity can cause the morphological changes of the structure of the sweat glands, sweat gland cells. Injury may be due to the antioxidant capacity of tissues leads to decline. In the second part, vitamin E protects the wet high temperature stress of sweat gland cells of rats Objective: To investigate the effect of high temperature and high humidity stress after vitamin E (VE) can improve skin antioxidant capacity, thereby reducing the high temperature and high humidity stress damage to the sweat gland cells. Methods: 21 SD rats were randomly divided into the normal temperature and humidity (NTH) group, high temperature and high humidity (HTH) group and VE treated with high temperature and high humidity (VHTH) group, 7 rats in each group.HTH group and VHTH group rats were subjected to high temperature and high humidity intervention 2h, HTH intervention group 1ml saline, VHTH group received VE Ig (100IU/kgVE distribution The physiological saline 1ml); NTH group for 2h at room temperature after 1% pentobarbital sodium 1ml saline gavage.72h after anesthesia, the left foot after peeling skin with 10% formalin fixed, observe tissue morphology by HE staining, immunohistochemical method to detect caspase -3 (Caspase-3) and heat shock protein 70 (HSP70) expression; stripping right plantar skin specimens, -80 C frozen by biochemical method to determine the total superoxide dismutase (SOD) activity and protein immunoblotting (Western Blot) method was used to detect the expression of HSP70. Results: under the light microscope can be observed in the HTH group and VHTH group of skin the secretion of sweat glands in high temperature and high humidity intervention glandular cavity expansion, NTH group of secretory lacunar smaller; but the group HTH secreting part of glandular epithelial cells arranged loosely, epithelial cell vacuoles decreased, nucleolus stained, NTH group and VHTH group of secretory gland epithelial cells arranged Neat, lacunar epithelial cells secreting part of small cavity secretion (nuclear vacuoles more), glandular epithelium with abundant cytoplasm and nucleolus stained; VHTH group activity of SOD is (35.18 + 8.19) U/mg, and HTH Group [(13.65 + 5.14) U/mg] significantly increased compared with group NTH, [(25.90 11.05) there was no significant difference between U/mg] results; detect the expression by immunohistochemistry and Caspase-3 HSP70 showed the expression level of VHTH group were significantly higher than HTH group, and no significant difference with NTH group; Western blot (Western Blot) HSP70 expression detection method, detection and analysis results of immunohistochemical method the results are consistent. Conclusion: VE can improve the oxidation caused by high temperature and high humidity environment reducing imbalances, reduce the damage of damp heat stress on rats has a protective effect on the sweat glands, sweat glands.

【学位授予单位】：广州中医药大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R594.12
，

本文编号：1584241