【摘要】：胰島素抵抗與高血壓、糖尿病、血脂異常、心腦血管疾病、肥胖等多種疾病密切相關(guān),是導(dǎo)致這些疾病的“共同土壤”。近年來(lái)的研究表明：胰島素抵抗是心腦血管疾病中眾多代謝性異常的始動(dòng)原因和致病基礎(chǔ),而血管內(nèi)皮功能障礙是動(dòng)脈粥樣硬化、高血壓、心腦血管疾病發(fā)生的病理基礎(chǔ)和早期階段。胰島素抵抗和血管內(nèi)皮功能障礙二者之間存在相互影響、相互促進(jìn)的關(guān)系,形成惡性循環(huán),在動(dòng)脈粥樣硬化的病理過(guò)程中扮演著重要的角色。大量的臨床研究報(bào)道證實(shí),針刺在這些疾病的治療中具有不可替代的優(yōu)勢(shì),其作用機(jī)理與改善這些疾病中存在的胰島素抵抗現(xiàn)象密切相關(guān)。通過(guò)本課題組的前期研究,我們發(fā)現(xiàn)針刺能有效改善胰島素的敏感性,能從受體后水平調(diào)整和改善胰島素抵抗?fàn)顟B(tài)。本研究是以胰島素抵抗與血管內(nèi)皮功能障礙相互作用為基礎(chǔ),設(shè)計(jì)實(shí)驗(yàn)觀察針刺干預(yù)對(duì)胰島素抵抗模型大鼠的內(nèi)皮功能影響；以自發(fā)性胰島素抵抗模型-OLETF大鼠及同系正常大鼠-LETO大鼠為研究對(duì)象,用ELISA法、硝酸還原酶法、硫代巴比妥酸等等多種檢測(cè)方法,檢測(cè)空腹血糖(FPG)、血清胰島素(FINS)、C肽、胰島素敏感指數(shù)(ISI)、一氧化氮(NO)、丙二醛(MDA)等血液指標(biāo),觀察針刺干預(yù)對(duì)OLETF大鼠胰島素抵抗及內(nèi)皮功能障礙共同基礎(chǔ)的氧化應(yīng)激的影響,探討針刺對(duì)胰島素抵抗及血管內(nèi)皮功能障礙的作用機(jī)制。全文分為文獻(xiàn)研究、實(shí)驗(yàn)研究及小結(jié)三大部分。文獻(xiàn)研究：從胰島素抵抗的概念、胰島素抵抗的發(fā)病機(jī)制、血管內(nèi)皮的生理功能、胰島素抵抗于內(nèi)皮功能障礙之間的關(guān)系及共同病理基礎(chǔ)、中醫(yī)藥防治胰島素抵抗的研究現(xiàn)狀、胰島素的動(dòng)物模型及評(píng)價(jià)等多方面闡述了胰島素抵抗的最新研究進(jìn)展。通過(guò)對(duì)大量與胰島素抵抗相關(guān)的文獻(xiàn)進(jìn)行綜合分析,本文提出了今后的研究方向,認(rèn)為針刺是改善胰島素抵抗的有效方法之一,對(duì)胰島素抵抗具有一定的逆轉(zhuǎn)效應(yīng),應(yīng)加強(qiáng)對(duì)其作用機(jī)理的深入研究及探討。實(shí)驗(yàn)研究：目的：觀察針刺對(duì)自發(fā)性胰島素抵抗模型OLETF大鼠的氧化應(yīng)激的影響,探討針刺干預(yù)對(duì)改善胰島素抵抗及血管內(nèi)皮功能障礙的作用機(jī)理。方法：實(shí)驗(yàn)選用雄性O(shè)LETF大鼠16只,正常雄性LETO大鼠8只。將8只雄性LETO大鼠作為空白組,16只雄性O(shè)LETF大鼠按體重根據(jù)隨機(jī)數(shù)字表分為：模型組和針刺組,每組各8只。空白組和模型組不予針刺干預(yù),針刺組則給予針刺干預(yù)。針刺穴位：腎俞、內(nèi)關(guān)、足三里和三陰交穴(均取雙側(cè))。針刺方式選用疏密波的電針(疏波2Hz,密波50Hz,調(diào)制頻率10次／分),加電穴位選取雙側(cè)三陰交和足三里穴,電針刺激強(qiáng)度以大鼠肢體輕微抖動(dòng)為標(biāo)準(zhǔn),針刺時(shí)間每日1次,每次持續(xù)20min,針刺干預(yù)持續(xù)4周。針刺前4周及針刺4周療程中,每周稱(chēng)重一次。在治療結(jié)束后,各組大鼠禁食不禁水12h。次日早晨取材。先用10%水合氯醛按體重將大鼠麻醉,采用眼眶靜脈竇采血,離心取上清,檢測(cè)針刺干預(yù)后FPG、FINS、C-P、NO、ET、SOD及MDA等指標(biāo),用葡萄糖氧化酶法測(cè)定FPG；用ELISA法測(cè)定FINS、C-P、ET;用硝酸還原酶法測(cè)定NO；用WST-1微孔板法測(cè)定SOD；硫代巴比妥酸(TBA法)測(cè)定丙二醛(MDA)；用1/(CFPG×JFINS)公式計(jì)算ISI。取完血液樣品后,將大鼠四肢固定于鼠板上,沿腹中線開(kāi)腹,從下往上分離出主動(dòng)脈,截取胸主動(dòng)脈,沿血管腔縱向剪開(kāi),置于4%多聚甲醛固定,HE染色。觀察針刺作用前后,OLETF大鼠主動(dòng)脈形態(tài)學(xué)改變情況。結(jié)果：1針刺對(duì)胰島素抵抗模型一般狀況及體重的影響在實(shí)驗(yàn)過(guò)程中,正常對(duì)照組大鼠體重增加,精神狀態(tài)良好,反應(yīng)靈敏,動(dòng)作自如,毛發(fā)有光澤。針刺前4周開(kāi)始可見(jiàn)模型組及針刺組大鼠性情變溫順,嗜睡少動(dòng),大鼠皮毛蓬亂、無(wú)光澤,大便量較正常組偏少,色灰,質(zhì)較軟,腹部肥胖。針刺4周后,可見(jiàn)針刺組大鼠體重仍有增長(zhǎng),表現(xiàn)出精神萎靡,反應(yīng)遲鈍,動(dòng)作遲緩等較前有所好轉(zhuǎn)。2針刺對(duì)大鼠血清FPG、FINS、ISI、C-P的影響模型組的FPG、FINS、C-P相比空白組顯著升高(P0.01),ISI較空白組顯著降低(P0.01)。針刺組的C-P相比空白組顯著降低(P0.05),FPG、FINS和ISI與空白組相比無(wú)統(tǒng)計(jì)學(xué)差異。針刺組的FPG、FINS、C-P較模型組顯著降低(P0.01),ISI相比模型組顯著升高(P0.01)3針刺對(duì)大鼠血清NO與ET-1的影響模型組的血清NO的水平相比空白組顯著降低(P0.01),模型組ET-1水平相比空白組顯著升高(P0.01)。針刺組的NO、ET-1水平相較空白組無(wú)統(tǒng)計(jì)學(xué)差異(P0.05)。針刺組的血清NO水平相比模型組顯著升高(P0.05),ET-1水平與模型組相比明顯降低(P0.01)。4針刺對(duì)大鼠血清SOD及MDA的影響模型組的血清SOD活力水平相比空白組明顯降低(P0.05),模型組MDA水平相比空白組顯著升高(P0.01)。針刺組的SOD活力水平、MDA相較空白組無(wú)統(tǒng)計(jì)學(xué)差異(P0.05)。針刺組的血清SOD活力相比模型組顯著升高(P0.05),MDA水平與模型組相比明顯降低(P0.01)。5針刺對(duì)大鼠主動(dòng)脈的形態(tài)學(xué)改變各組大鼠主動(dòng)脈HE染色后于數(shù)碼成像顯微鏡下以200倍觀察,可見(jiàn)空白對(duì)照組大鼠的血管內(nèi)膜光滑完整、連續(xù),無(wú)局部缺損或增厚,彈力纖維、血管平滑肌細(xì)胞排列整齊,未見(jiàn)浸潤(rùn)的淋巴細(xì)胞等病理改變(見(jiàn)圖9)。而模型組大鼠的主動(dòng)脈內(nèi)皮層不連續(xù),明顯失去內(nèi)膜的完整性,內(nèi)皮下層呈疏松或斷裂狀態(tài),血管平滑肌細(xì)胞增生、肥大且排列紊亂(見(jiàn)圖10)。當(dāng)給予一定療程針刺干預(yù)治療后,針刺組大鼠的主動(dòng)脈內(nèi)膜層基本連續(xù)完整,未見(jiàn)明顯增厚,內(nèi)皮下間隙稍寬,但程度相比模型組減輕,彈力纖維及平滑肌細(xì)胞排列尚整齊,病理?yè)p傷均得到不同程度的改善(見(jiàn)圖11),提示針刺治療具有保護(hù)主動(dòng)脈內(nèi)皮損傷的作用。結(jié)論：研究結(jié)果表明,針刺可以通過(guò)改善大鼠的胰島素抵抗及氧化應(yīng)激狀態(tài),調(diào)整血管內(nèi)皮功能障礙,逆轉(zhuǎn)血管內(nèi)皮病理改變。
[Abstract]:Insulin resistance is closely related to many diseases such as hypertension, diabetes, dyslipidemia, cardiovascular and cerebrovascular diseases, obesity and so on. It is a "common soil" that causes these diseases. Recent studies have shown that insulin resistance is the origin and pathogenesis of many metabolic abnormalities in cardiovascular and cerebrovascular diseases, and vascular endothelial dysfunction is a movement. The pathological basis and early stage of atherosclerotic, hypertensive, cardio cerebral vascular diseases. There is a mutual influence between the two cases of insulin resistance and vascular endothelial dysfunction, the relationship of mutual promotion and the formation of a vicious circle, which plays an important role in the pathological process of atherosclerosis. A large number of clinical studies have confirmed that acupuncture has been used. It has an irreplaceable advantage in the treatment of these diseases, and its mechanism is closely related to the improvement of insulin resistance in these diseases. Through the earlier study of this group, we found that acupuncture can effectively improve the sensitivity of insulin and can adjust and improve the state of insulin resistance from the post receptor level. This study is based on this study. On the basis of the interaction of insulin resistance and vascular endothelial dysfunction, the effect of acupuncture intervention on the endothelial function of rats with insulin resistance model was designed by the design experiment. The -OLETF rats and -LETO rats of the same normal rats were used as the research object. The ELISA, nitrate reductase, thiobarbituric acid and so on were used. The method of detecting fasting blood glucose (FPG), serum insulin (FINS), C peptide, insulin sensitivity index (ISI), nitric oxide (NO), malondialdehyde (MDA) and other blood indexes, to observe the effects of acupuncture intervention on the oxidative stress of insulin resistance and endothelial dysfunction in OLETF rats, and to explore the effect of acupuncture on insulin resistance and vascular endothelial function. The full text is divided into literature research, experimental research and summary three parts. Literature study: the concept of insulin resistance, the pathogenesis of insulin resistance, the physiological function of vascular endothelium, the relationship between insulin resistance and endothelial dysfunction and the common pathophysiological basis, the study of the prevention and treatment of insulin resistance in traditional Chinese Medicine The latest research progress in insulin resistance is described in many aspects, such as animal model and evaluation of insulin. Through a comprehensive analysis of a large number of literature related to insulin resistance, this paper puts forward the future research direction. It is considered that acupuncture is one of the effective methods to improve insulin resistance and has a certain reversal effect on insulin resistance. In order to investigate the effect of acupuncture on oxidative stress in spontaneous insulin resistance model OLETF rats and to explore the mechanism of acupuncture intervention on improving insulin resistance and vascular endothelial dysfunction. Methods: 16 male OLETF rats, normal male rats were selected. 8 male LETO rats. 8 male LETO rats were taken as blank group, and 16 male OLETF rats were divided into two groups according to the weight of random numbers: model group and acupuncture group, 8 rats in each group. The blank group and model group were not needled, and acupuncture group was given acupuncture intervention. Acupuncture points: Shenshu, Neiguan, Zusanli and three yin acupoints (both sides). The electroacupuncture (sparse wave 2Hz, dense wave 50Hz, modulation frequency 10 times) and the acupoint of electric acupuncture were selected to select the bilateral and Zusanli points. The stimulation intensity of the electroacupuncture was taken as the standard of the mild limb jitter of the rats. The acupuncture time was 1 times a day, each time 20min was sustained, and the acupuncture intervention lasted for 4 weeks. In the course of 4 weeks before acupuncture and 4 weeks of acupuncture, weight was weighed once a week. After the end of the treatment, the rats in each group were forbidden to feed the water 12h. the following morning. First, the rats were anaesthetized with 10% chloral chloral hydrate and the rats were anaesthetized by the orbital venous sinus and centrifuged to pick up the supernatant. The indexes of FPG, FINS, C-P, NO, ET, SOD and MDA were measured, and FPG was measured by the method of glucosinase. The FINS, C-P, and nitrate were measured by ELISA. NO was determined by enzyme method, SOD was measured with WST-1 microplate method, and malondialdehyde (MDA) was measured by thiobarbituric acid (TBA method). The rat limbs were fixed on the rat plate with 1/ (CFPG x JFINS) formula, and the aorta was separated from the middle line, the aorta was isolated from the lower abdomen, the thoracic aorta was intercepted, and the 4% polymethylene was cut along the blood tube and placed in the polymethyl methylene. The morphological changes in the aorta of OLETF rats before and after acupuncture were observed. Results: 1 the effect of acupuncture on the general condition of insulin resistance model and the influence of weight on the body weight of the insulin resistance model in the normal control group was increased, the mental state was good, the reaction was sensitive, the movement was free and the hair was luster. The visible model of the 1 weeks before the acupuncture was started. The sexual emotion of the rats in the group and the acupuncture group was gentle, sleepy, shaggy, shiny, shiny, less stool than the normal group, gray, soft and abdominal fat. After 4 weeks of acupuncture, the body weight of the rats in the acupuncture group was still growing, showing the mental retardation, the slow reaction, the slow motion and so on,.2 acupuncture on the rat serum FPG, FINS, ISI, C The effect of -P on FPG, FINS, C-P in the blank group was significantly higher than that in the blank group (P0.01), and ISI was significantly lower than that in the blank group (P0.01). The C-P in the acupuncture group was significantly lower than that in the blank group (P0.05). FPG, FINS and ISI were not significantly different from those in the blank group. 3 the effect of acupuncture on serum NO and ET-1 in rats was significantly lower than that in the blank group (P0.01), and the level of ET-1 in the model group was significantly higher than that in the blank group (P0.01). The level of ET-1 in the acupuncture group was not significantly different than that in the blank group (P0.05). The serum NO level of the acupuncture group was significantly higher than that in the model group (P0.05), and the ET-1 level was significantly higher than that in the model group (P0.05). The model group significantly reduced (P0.01).4 acupuncture on the serum SOD and MDA in rats, the serum SOD activity level of the model group was significantly lower than that in the blank group (P0.05). The MDA level in the model group was significantly higher than that in the blank group (P0.01). The SOD activity level of the acupuncture group was not statistically significant (P0.05) compared with the blank group (P0.05). The serum SOD activity phase of the acupuncture group was not significant (P0.05). Compared with the model group (P0.05), the level of MDA was significantly lower than that of the model group (P0.01). The morphological changes of the aorta in the rat aorta were compared with the model group. After HE staining, the aorta of rats was observed 200 times under the digital imaging microscope. It could be seen that the vascular intima of the blank control group was intact, continuous, no local defect or thickening, elastic fiber, The vascular smooth muscle cells were arranged neatly with no infiltrating lymphocytes and other pathological changes (see Figure 9). But the aortic endothelium in the model group was discontinuous, obviously lost the integrity of the intima, the underlayer of the endothelium was loose or fractured, the vascular smooth muscle cells proliferated and hypertrophy and disorder (see Figure 10). When given a certain course of acupuncture intervention treatment After the treatment, the aortic intima of the rats in the acupuncture group was basically complete, no obvious thickening and a slightly wider space under the endothelium, but the extent of the endothelium was lighter than the model group. The arrangement of elastic fibers and smooth muscle cells was neatly arranged, and the pathological damage was improved in different degrees (see Figure 11). The effect of acupuncture treatment on the protection of aortic endothelial injury was suggested. The results show that acupuncture can adjust the dysfunction of vascular endothelium and reverse the pathological changes of vascular endothelium by improving the state of insulin resistance and oxidative stress in rats.
【學(xué)位授予單位】：廣州中醫(yī)藥大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2015
【分類(lèi)號(hào)】：R245

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