【摘要】：目的：初步闡明耐力運動在大鼠運動性支氣管痙攣發(fā)病過程中對腎上腺髓質(zhì)嗜鉻細(xì)胞結(jié)構(gòu)和功能的影響及其作用機制。 方法：對大鼠進(jìn)行中強度或高強度跑臺耐力運動,并分別予NGF抗體、力竭運動或卵清蛋白(ovalbumin,OVA)致敏、激發(fā)干預(yù),120只雄性Sprague-Dawley大鼠隨機分為15組,每組8只,分別為：正常對照組,力竭運動組,中速訓(xùn)練組,中速訓(xùn)練+NGF抗體組,中速訓(xùn)練+力竭運動組,中速訓(xùn)練+NGF抗體+力竭運動組,高速訓(xùn)練組,高速訓(xùn)練+NGF抗體組,高速訓(xùn)練+力竭運動組,高速訓(xùn)練+NGF抗體+力竭運動組,OVA組,OVA+中速訓(xùn)練組,OVA+中速訓(xùn)練+NGF抗體組,OVA+高速訓(xùn)練組,OVA+高速訓(xùn)練+NGF抗體組。檢測大鼠氣道阻力以及支氣管肺泡灌洗液中白細(xì)胞比例；ELISA檢測大鼠血清中腎上腺素(Epinephrine, EPI)、神經(jīng)生長因子(Nerve growth factor, NGF)、皮質(zhì)酮(Corticosterone, CORT)水平；光鏡觀察大鼠支氣管肺組織以及腎上腺髓質(zhì)的形態(tài)學(xué)改變；電鏡觀察大鼠腎上腺髓質(zhì)嗜鉻細(xì)胞的超微結(jié)構(gòu)；免疫組化法檢測腎上腺髓質(zhì)中苯乙醇胺-N-甲基轉(zhuǎn)移酶(Phenylethanolamine N-methyl transferase, PNMT)、外周蛋白(Peripherin)的表達(dá)；Western blot法檢測p-ERK/ERK、p-p38/p38, p-JNK/JNK以及peripherin蛋白表達(dá)量；Real-time PCR以及western blot法分別檢測JMJD3以及Mash1的mRNA和蛋白表達(dá)量。 結(jié)果：1.與正常組以及中速訓(xùn)練組比,高速訓(xùn)練組大鼠氣道內(nèi)淋巴細(xì)胞浸潤明顯增加,氣道阻力升高；大鼠腎上腺髓質(zhì)嗜鉻細(xì)胞出現(xiàn)空泡樣變性,嗜鉻顆粒減少,出現(xiàn)細(xì)胞突起。高速訓(xùn)練組大鼠血清中NGF水平升高,EPI水平下降,腎上腺中PNMT表達(dá)下降,peripherin表達(dá)上升,同時p-ERK/ERK、JMJD3、Mash1表達(dá)均升高；NGF抗體干預(yù)能抑制高強度耐力運動所致的上述病理生理改變。 2.與正常組對比,力竭運動組大鼠氣道阻力增高,腎上腺髓質(zhì)中血竇擴張,其血清中EPI、CORT水平升高,腎上腺中PNMT、p-ERK表達(dá)增高,而血清中NGF、腎上腺中JMJD3及Mash1表達(dá)無明顯差異。與高速訓(xùn)練組比較,高速訓(xùn)練+力竭運動組大鼠氣道淋巴細(xì)胞浸潤增加,氣道阻力升高,腎上腺中peripherin、p-ERK、JMJD3、Mash1的表達(dá)均升高。但血清中EPI、CORT、NGF水平均無明顯改變。經(jīng)NGF抗體干預(yù)后,高速訓(xùn)練+NGF抗體+力竭運動組大鼠上述病理生理改變均明顯改善。 3.與正常組以及高速訓(xùn)練組比較,OVA組大鼠氣道中淋巴細(xì)胞、巨噬細(xì)胞、嗜酸性粒細(xì)胞、中性粒細(xì)胞增多,氣道阻力明顯升高,血清中NGF水平升高,CORT水平下降。與正常對照組比較,OVA組大鼠腎上腺髓質(zhì)嗜鉻顆粒減少,有細(xì)胞突起形成,血清EPI水平下降,大鼠腎上腺中PNMT表達(dá)下降,peripherin表達(dá)上升,同時p-ERK/ERK、JMJD3、Mash1表達(dá)均升高。與OVA組對比,OVA+高速訓(xùn)練組大鼠上述病理生理改變進(jìn)一步加重,嗜鉻細(xì)胞出現(xiàn)脫顆粒改變,纖維組織增多。而經(jīng)過NGF抗體以及中強度耐力運動干預(yù),OVA+高速訓(xùn)練+NGF抗體組以及OVA+中速訓(xùn)練組大鼠的病理生理改變可部分逆轉(zhuǎn)。 結(jié)論：1.高強度耐力運動能夠引起大鼠腎上腺嗜鉻細(xì)胞向神經(jīng)細(xì)胞表型轉(zhuǎn)化,其發(fā)生機制與NGF介導(dǎo)的p-ERK-JMJD3-Mashl通路活化密切關(guān)聯(lián)； 2.劇烈運動激發(fā)能夠引起高強度耐力運動大鼠支氣管痙攣,其機制與腎上腺髓質(zhì)嗜鉻細(xì)胞發(fā)生神經(jīng)細(xì)胞表型轉(zhuǎn)化導(dǎo)致腎上腺素分泌水平降低密切關(guān)聯(lián)； 3.高強度耐力運動干預(yù)與OVA干預(yù)相互作用,使大鼠腎上腺髓質(zhì)嗜鉻細(xì)胞發(fā)生神經(jīng)細(xì)胞表型轉(zhuǎn)化過程加劇,而中強度耐力運動有逆轉(zhuǎn)作用。圖38幅,表14個,參考文獻(xiàn)67篇
[Abstract]:Objective: to elucidate the effect and mechanism of endurance exercise on the structure and function of adrenal medullary chromaffin cells during the pathogenesis of exercise-induced bronchospasm in rats.
Methods: the rats were treated with moderate or high strength endurance exercise, and were treated with NGF antibody, exhaustive exercise or ovalbumin (OVA) sensitization and excitation intervention. 120 male Sprague-Dawley rats were randomly divided into 15 groups, 8 rats in each group, which were the normal control group, the exhaustive exercise group, the medium speed training group and the medium speed training +NGF antibody group. Speed training + exhausted exercise group, medium speed training +NGF antibody + exhausting exercise group, high speed training group, high speed training +NGF antibody group, high speed training + exhaustive exercise group, high speed training +NGF antibody + exhausted exercise group, OVA group, OVA+ medium speed training group, OVA+ medium speed training +NGF antibody group, OVA+ high-speed training group, OVA+ high-speed training +NGF antibody group. Test rats gas. ELISA, Epinephrine, EPI, Nerve growth factor, NGF, and corticosterone (Corticosterone, CORT) in rat serum, and the morphological changes of bronchopulmonary tissues and adrenal medulla in rats were observed by light microscopy. The rats were observed by electron microscopy. The ultrastructure of the chromaffin cells in the adrenal medulla; the expression of Phenylethanolamine N-methyl transferase (PNMT) (PNMT) and the peripheral protein (Peripherin) in the adrenal medulla by immunohistochemistry; the Western blot method was used to detect p-ERK/ERK, p-p38/p38, p-JNK/JNK, and the expression of peripherin protein. The mRNA and protein expression levels of JMJD3 and Mash1 were detected by Western blot.
Results: 1. compared with the normal group and the medium speed training group, the infiltration of lymphocytes in the airway of the rats in the high speed training group increased obviously, the airway resistance increased, and the chromaffin cells in the adrenal medulla were vacuolated, the chromaffin granules decreased and the cell protruded. The level of NGF in the high speed training group was higher, the level of EPI decreased, and the PN in the adrenal gland was decreased. The expression of MT decreased, the expression of peripherin increased, while the expression of p-ERK/ERK, JMJD3 and Mash1 all increased, and the intervention of NGF antibody could inhibit the above pathophysiological changes caused by high intensity endurance exercise.
2. compared with the normal group, the airway resistance of the rats in the exhaustive exercise group increased and the blood sinus dilated in the adrenal medulla, the level of EPI and CORT in the serum increased, the expression of PNMT and p-ERK in the adrenal gland increased, but there was no significant difference in the expression of JMJD3 and Mash1 in the adrenal gland. Compared with the high speed training group, the high speed training + exhaustive exercise group had lymphatic airway lymphatic. The expression of peripherin, p-ERK, JMJD3, and Mash1 in the adrenal gland increased, but the levels of EPI, CORT and NGF in the serum were not significantly changed. After the prognosis of NGF antibody, the above pathophysiological changes in the rats with high speed training +NGF antibody + exhausting exercise group were obviously improved.
3. compared with the normal group and the high speed training group, the lymphocyte, macrophage, eosinophil, neutrophilic granulocyte, neutrophils increased in the OVA group, increased the airway resistance, the level of NGF in the serum and the level of CORT decreased. Compared with the normal control group, the chromaffin granules in the renal gland medulla of the OVA group were reduced, the formation of cell protruded and the serum EPI was formed. The expression of PNMT in the adrenal gland decreased and the expression of peripherin increased, while the expression of p-ERK/ERK, JMJD3 and Mash1 increased. Compared with the OVA group, the above pathophysiological changes were further aggravated in the OVA+ high speed training group, and the chromaffin cells were degranulation, and the fibrous fabric increased. After NGF antibody and moderate strength endurance exercise intervention, the expression of chromaffin cells was increased. The pathophysiological changes of OVA+ high speed training +NGF antibody group and OVA+ medium speed training group can be partially reversed.
Conclusion: 1. high intensity endurance exercise can induce the phenotype of adrenal chromaffin cells to be transformed into neural cell phenotype, and the mechanism is closely related to the activation of NGF mediated p-ERK-JMJD3-Mashl pathway.
2. intense exercise induced bronchial spasm in rats with high intensity endurance exercise, which is closely related to the reduction of adrenaline secretion by the phenotypic transformation of the adrenal medullary chromaffin cells.
3. the interaction between high strength endurance exercise intervention and OVA intervention makes the phenotypic transformation process of the adrenal medullary chromaffin cells aggravated, while the middle intensity endurance exercise has a reversal effect. Figure 38, table 14, 67 references
【學(xué)位授予單位】：中南大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2013
【分類號】：R562.25

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