發(fā)布時(shí)間：2018-03-08 14:17

  本文選題：高溫高濕　切入點(diǎn)：抗氧化　出處：《廣州中醫(yī)藥大學(xué)》2017年碩士論文　論文類型：學(xué)位論文

【摘要】：背景:汗腺排泄汗液的功能在機(jī)體穩(wěn)態(tài)中具有不可替代的作用,是人體體溫調(diào)節(jié)主要方法。汗液的主要成份98～99%是水,1~2%為無機(jī)鹽和有機(jī)物,汗液的有效蒸發(fā)是汗液持續(xù)分泌的必要前提。當(dāng)空氣相對(duì)濕度增大,其中的水分越接近飽和,人體皮膚表面汗液的蒸發(fā)就會(huì)困難,若同時(shí)周圍環(huán)境溫度升高,則機(jī)體熱量不易消散,汗液的分泌是降低機(jī)體核心溫度的重要途徑,一旦該途徑受阻,核心溫度升高,機(jī)體的內(nèi)部器官也將受到熱損傷。濕熱環(huán)境通常是指環(huán)境溫度在36℃以上,相對(duì)濕度超過60%[1]。臨床資料顯示,高溫高濕環(huán)境常常可引發(fā)中暑,中暑患者可出現(xiàn)高熱、皮膚干燥、情緒錯(cuò)亂,甚至全身驚厥發(fā)作的癥狀[2]。其中皮膚干燥的癥狀則意味著汗液分泌減少,減少的原因以及高溫高濕環(huán)境對(duì)汗腺功能的影響機(jī)制尚不清楚。本實(shí)驗(yàn)觀察高溫高濕環(huán)境對(duì)Sprague-Dawley(SD)大鼠足墊皮膚汗腺細(xì)胞功能的影響,并探討汗腺細(xì)胞功能發(fā)生變化的可能機(jī)制。第一部分高溫高濕環(huán)境對(duì)大鼠汗腺細(xì)胞影響的初步探討目的:初步探討高溫高濕環(huán)境對(duì)汗腺功能的影響并對(duì)其可能的機(jī)制進(jìn)行研究。方法:14只Sprague-Dawley(SD)大鼠隨機(jī)分為常溫常濕(NTH)組和高溫高濕(HTH)組,每組7只,HTH組大鼠進(jìn)行高溫高濕干預(yù)2h,NTH組室溫放置2h,72h后1%戊巴比妥鈉麻醉,收集大鼠心臟血液,檢測血液中乙酰膽堿(ACH)的水平;剝離左后足底皮膚用10%福爾馬林固定,應(yīng)用HE染色觀察組織形態(tài)和免疫組織化學(xué)法檢測乙酰膽堿轉(zhuǎn)移酶(ChAT)的表達(dá);剝離右后足底皮膚,標(biāo)本-80℃凍存,應(yīng)用生物化學(xué)方法檢測組織總超氧化物歧化酶(SOD)活力以及蛋白免疫印跡(Western Blot)法檢測M3受體的表達(dá)。結(jié)果:光鏡下HTH組汗液分泌部的上皮細(xì)胞胞質(zhì)中空泡減少,且伴有汗腺細(xì)胞凋亡增多,HTH組的凋亡指數(shù)為(70.7±14.0)%,顯著高于NTH組[(58.6±11.7)%],差異有統(tǒng)計(jì)學(xué)意義(t=2.478,P=0.021);Western Blot的結(jié)果顯示,HTH組應(yīng)激后M3受體的相對(duì)表達(dá)量為(1.21±0.16)%,顯著高于NTH組[(1.81 士0.41)%,t=2.711,P=0.035];進(jìn)一步檢測到HTH組SD大鼠血清中乙酰膽堿的表達(dá)明顯弱于NTH組;并且HTH組超氧化物歧化酶(SOD)活力為(13.65 ±5.14)U/mg,較NTH組[(25.89± 11.05)U/mg]顯著降低(t=2.633,P=0.023)。結(jié)論:高溫高濕環(huán)境可引起汗腺結(jié)構(gòu)形態(tài)變化,其對(duì)汗腺細(xì)胞的損傷可能是由于組織抗氧化能力減退導(dǎo)致。第二部分維生素E保護(hù)高溫高濕應(yīng)激大鼠汗腺細(xì)胞的研究目的:探討高溫高濕應(yīng)激后維生素E(VE)是否能改善皮膚組織的抗氧化能力,從而減少高溫高濕應(yīng)激對(duì)汗腺細(xì)胞的損害。方法:21只SD大鼠隨機(jī)分為常溫常濕(NTH)組,高溫高濕(HTH)組和VE干預(yù)高溫高濕(VHTH)組,每組7只。HTH組和VHTH組大鼠進(jìn)行高溫高濕干預(yù)2h,干預(yù)后HTH組給予生理鹽水1ml灌胃,VHTH組給予VE灌胃(100IU/kgVE配置生理鹽水1ml);NTH組室溫放置2h后給予生理鹽水1ml灌胃。72h后1%戊巴比妥鈉麻醉,剝離左后足底皮膚用10%福爾馬林固定,應(yīng)用HE染色觀察組織形態(tài)、免疫組織化學(xué)法檢測天冬氨酸特異性半胱氨酸蛋白酶-3(Caspase-3)和熱休克蛋白70(HSP70)的表達(dá);剝離右后足底皮膚,標(biāo)本-80℃凍存,應(yīng)用生物化學(xué)方法檢測組織總超氧化物歧化酶(SOD)活力以及蛋白免疫印跡(Western Blot)法檢測HSP70的表達(dá)。結(jié)果:光鏡下可觀察到經(jīng)高溫高濕干預(yù)的HTH組和VHTH組皮膚汗腺分泌部腺腔擴(kuò)張,NTH組分泌部腔隙較小;但是HTH組分泌部腺上皮細(xì)胞排列疏松,上皮細(xì)胞核下空泡減少,核仁淡染,NTH組和VHTH組分泌部腺上皮細(xì)胞排列整齊,分泌部腔隙較小,上皮細(xì)胞核下空泡分泌旺盛(核下空泡較多),腺上皮胞漿豐富,核仁深染;VHTH組組織 SOD 的活性為(35.18 士 8.19)U/mg,與 HTH 組[(13.65 ±5.14)U/mg]相比顯著升高,與NTH組[(25.90±11.05)U/mg]相比無顯著性差異;免疫組織化學(xué)法檢測組織Caspase-3和HSP70的表達(dá)結(jié)果顯示VHTH組的表達(dá)水平均顯著高于HTH組,且與NTH組無顯著差異;蛋白免疫印跡(Western Blot)法檢測組織HSP70的表達(dá),分析結(jié)果與免疫組織化學(xué)法的檢測結(jié)果相一致。結(jié)論:VE能夠改善高溫高濕環(huán)境引起的氧化還原失衡,減少濕熱應(yīng)激對(duì)大鼠汗腺的損害,對(duì)汗腺有保護(hù)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.

【學(xué)位授予單位】：廣州中醫(yī)藥大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R594.12
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本文編號(hào)：1584241