本文關鍵詞： 縮泉丸聯用腎氣丸 補腎縮尿法 膀胱過度活動癥 豚鼠 膀胱 神經-ICC-平滑肌網絡　出處：《廣州中醫(yī)藥大學》2016年碩士論文　論文類型：學位論文

【摘要】：目的：膀胱過度活動癥(Overactive Bladder,OAB)作為一種以下尿路病理和尿急癥狀為特征的病癥,其明顯對患者的日常生活和社會活動造成極大的干擾,已經成為困擾人們的一大疾病。神經-ICC-平滑肌網絡作為新近認為是OAB的其中一個致病原因,日益受到研究人員關注的研究熱點�？s泉丸的功效為補腎縮尿,固脫止遺。腎氣丸功效為溫補腎陽,振奮陽氣,縮泉丸與腎氣丸合用在治療OAB上有良好的療效。本課題以神經-ICC-平滑肌網絡為切入點,研究補腎縮尿法對OAB的作用機制。方法：本研究以膀胱出口梗阻模型作為OAB的研究模型。將豚鼠分為正常組、模型組、陽性組、縮泉丸組、腎氣丸組和縮泉丸聯用腎氣丸組六個組。分成三部分1、補腎縮尿法對豚鼠OAB的治療作用：測各組的排尿次數、排尿量、尿動力參數和離體肌條功能參數。2、研究補腎縮尿法對OAB豚鼠膀胱神經-ICC-平滑肌網絡結構的影響：通過組織免疫熒光法觀察膀胱ICC、乙酰膽堿能神經(VAChT)、P物質肽能神經的相對含量變化,以及透射電鏡觀察整體膀胱神經-ICC-平滑肌網絡結構；3、研究補腎縮尿法對OAB豚鼠膀胱神經-ICC-平滑肌網絡信號傳導的影響：通過組織免疫熒光雙標觀察ICC與M3、P2X5受體亞型的共表達情況,觀察補腎縮尿法是否作用于ICC上的M3和P2X5受體,以及采用熒光定量PCR法、westernblotting法檢測ICC-平滑肌網絡縫隙連接蛋白Cx43mRNA及蛋白的表達。結果：1、模型組能顯著增加豚鼠排尿次數,增加膀胱漏尿點壓力(BLPP),最大膀胱壓(MVP)及殘余尿量(PVR)的作用(P0.05)；模型組顯著提高了膀胱肌條對激動劑的刺激敏感性,對由于KCL激動劑引起的收縮和Carbachol引起的收縮有顯著性提高(P0.05)；模型組ICC相對表達量顯著升高,而肽能神經的相對表達量顯著降低(P0.05),模型組膀胱神經-ICC-平滑肌網絡之間的連接更為緊密；ICC上神經受體M3和P2X5表達量在造模后顯著升高(P0.05),縫隙連接蛋白Cx43的mRNA和蛋白表達量顯著升高(P0.01)。2、補腎縮尿法對豚鼠OAB的治療作用：用托特羅定、縮泉丸或縮泉丸聯用腎氣丸組治療能減少OAB模型豚鼠的排尿次數(P0.05),托特羅定、縮泉丸、腎氣丸以及縮泉丸聯用腎氣丸組有不同程度降低BLPP、MVP及PVR的作用(P0.05)。托特羅定、縮泉丸以及縮泉丸聯用腎氣丸組均有不同程度顯著性降低由于KCL激動劑所引起的肌條收縮作用(P0.05),并在Carbachol不同濃度影響下的收縮作用均有顯著性降低(P0.05)。3、補腎縮尿法對OAB豚鼠膀胱神經-ICC-平滑肌網絡結構的影響：免疫熒光結果顯示,給予縮泉丸、腎氣丸和縮泉丸聯用腎氣丸有不同程度降低ICC細胞和提高神經纖維P物質的相對表達量,其中縮泉丸聯用腎氣丸效果最為顯著(P0.05,P0.01)。透射電鏡圖顯示,分別給予托特羅定,縮泉丸,腎氣丸或補腎縮尿法治療后,均有不同程度改善神經-ICC-平滑肌結構間的聯系,聯系減少。4、補腎縮尿法對OAB豚鼠膀胱神經-ICC-平滑肌網絡信號傳導的影響：對神經受體與ICC的免疫熒光雙標結果顯示,補腎縮尿法治療后,ICC細胞上的M3神經受體表達顯著下降(P0.05)；縮泉丸聯用腎氣丸干預后,膀胱縫隙連接蛋白Cx43mRNA和蛋白表達量呈極顯著下降(P0.01),證明縮泉丸聯用腎氣丸能夠通過減少沖動信號的傳遞而達到調節(jié)膀胱功能。結論：1、模型組能增加排尿次數,伴有平滑肌不穩(wěn)定收縮,并對激動劑的敏感性伴有顯著性升高,代表造模成功。其中膀胱ICC數量升高,代表OAB的發(fā)生與ICC數量變化有關。2、縮泉丸聯用腎氣丸組明顯改善OAB豚鼠排尿次數,能夠減少膀胱漏尿點壓力(BLPP)、最大膀胱壓(MVP)及殘余尿量(PVR),改善膀胱肌條張力活動,初步證明補腎縮尿法具有改善OAB膀胱病理狀態(tài)的作用。3、模型組ICC數量明顯增加,神經纖維數量明顯增加,透射電鏡結果顯示神經-ICC-平滑肌網絡聯系更加緊密。經縮泉丸聯用腎氣丸組治療能夠改善膀胱神經-ICC-平滑肌網絡的聯系,改善去神經化的病理狀態(tài),預示給藥后藥物能通過降低ICC細胞和改變神經纖維數量從而起到補腎縮尿的作用。4、縮泉丸聯用腎氣丸組能減少神經受體的數量,減低縫隙連接蛋白的表達量,起到減少OAB豚鼠膀胱神經-ICC-平滑肌網絡信號傳導。說明補腎縮尿法通過改善神經-ICC-平滑肌網絡之間的聯系起到補腎縮尿的作用。
[Abstract]:Objective: overactive bladder (Overactive Bladder, OAB) as a kind of urinary tract and bladder symptoms following pathological condition characterized by its obvious, on the patient's daily life and social activities caused great disturbance, has become a major disease of people. -ICC- neural network as a newly smooth muscle is considered a cause OAB which has been the research focus of attention of researchers. The effect of suoquanwan for nourishing kidney and reducing urine gutuo bushenzhiyi. Efficacy of Shenqi Pill for warming Yang, activating Yang Qi, suoquanwan and Shenqi Pill combination has good curative effect in the treatment of OAB. The -ICC- neural network for smooth muscle the starting point, study the mechanism of nourishing kidney and reducing urine of OAB. Methods: in this study, bladder outlet obstruction model as the research model of OAB. The guinea pigs were divided into normal group, model group, positive group, suoquanwan group, Shenqi Pill group and Suoquan Shenqi Pill group six group combination pills. Divided into three parts 1, the treatment effect of tonifying kidney and reducing urine of guinea pig OAB: measured the frequency of urination, urine volume, urine dynamic parameters and function parameters from.2 body muscle, effects of nourishing kidney and reducing urine of guinea pig bladder smooth muscle -ICC- OAB neural network structure. By immunofluorescence observation of bladder ICC, cholinergic nerve (VAChT), changes in the relative content of P substance peptidergic nerve, and transmission electron microscopy. The whole bladder smooth muscle -ICC- neural network structure; 3, effect of nourishing kidney and reducing urine of guinea pig bladder smooth muscle -ICC- OAB neural network signal transduction through tissue immunofluorescence observation ICC and M3 co expression of P2X5 receptor subtypes, M3 receptor and P2X5 observation of nourishing kidney and reducing urine method is applied on the ICC, and the fluorescent quantitative PCR method was used to detect ICC- smooth muscle westernblotting joint network The expression of gap junction protein Cx43mRNA and protein. Results: 1, the model group significantly increased the frequency of urination in guinea pigs, increase bladder leak point pressure (BLPP), maximum bladder pressure (MVP) and residual urine volume (PVR) effect (P0.05); model group significantly increased the bladder muscle strips of excited stimulation sensitivity agent, can significantly improve the induced by KCL agonist induced contraction and contraction of Carbachol (P0.05); model group ICC expression was significantly increased, while the relative expression of peptidergic nerve decreased significantly (P0.05), the connection between the model group of bladder smooth muscle -ICC- neural network more closely; ICC nerve receptors M3 and P2X5 expression increased significantly in the model (P0.05), mRNA and protein expression of gap junction protein Cx43 was significantly higher (P0.01).2, the treatment effect of tonifying kidney and reducing urine of guinea pig OAB: Todd Luoding, suoquanwan or Suoquan pill combined with Shenqi Pill group Treatment can reduce the frequency of urination of guinea pig OAB model (P0.05), Todd Luoding, suoquanwan, Shenqiwan and combined with Shenqi Pill group to different degrees. BLPP suoquanwan, MVP and the role of PVR (P0.05). Luoding Todd, suoquanwan and Suoquan pill combined with Shenqi Pill group had different degree due to significantly reduced muscle KCL agonist induced contraction (P0.05), and the contraction had significant effects on Carbachol under different concentrations decreased (P0.05).3, effects of nourishing kidney and reducing urine of guinea pig bladder smooth muscle -ICC- OAB neural network structure: the immunofluorescence results showed that given Suoquan pill, Qi pill and Suoquan pill combined with Shenqi Pill with different degrees of reduction of ICC cells and increase the relative expression of nerve fiber P amount of substance, which Suoquan pill combined with Shenqi Pill was the most significant (P0.05, P0.01). Transmission electron microscope showed that were given Luoding Todd, suoquanwan, kidney Gas pill or method of nourishing kidney and reducing urination after treatment were improved -ICC- nerve smooth muscle structure between the contact to reduce.4, effects of nourishing kidney and reducing urine of guinea pig bladder smooth muscle -ICC- OAB neural network signal transduction: immunofluorescence staining of nerve receptor and ICC show that the method of nourishing kidney and reducing urine after the treatment, the expression of M3 nerve receptors in ICC cells decreased significantly (P0.05); Suoquan pill combined with Shenqi Pill intervention, bladder gap junction protein Cx43mRNA and protein expression was significantly decreased (P0.01), proof of Suoquan pill combined with Shenqi Pill can transfer by reducing the impulse signal and to adjust the bladder function. Conclusion: 1 the model group, can increase the frequency of urination, accompanied by unstable contraction of smooth muscle, and sensitivity to agonist was associated with a significant increase, on behalf of the successful establishment of the model. The number of ICC increased the number of bladder, occurrence and change of ICC on behalf of OAB are .2, Suoquan pill combined with Shenqi Pill group significantly improved OAB guinea pig urination, can reduce bladder leak point pressure (BLPP), maximum bladder pressure (MVP) and residual urine volume (PVR), improve the tension activity of bladder muscle strips, preliminary proof method has the effect of nourishing kidney and reducing urine.3 improve the pathological state of bladder OAB the number of ICC in the model group increased significantly, the number of nerve fibers increased significantly. The TEM results showed that the -ICC- neural network is more closely linked. The smooth muscle of Suoquan pill combined with Shenqi Pill group therapy can improve bladder smooth muscle -ICC- neural network, improve the pathological state of denervation, that after administration of drugs by reducing effect the number of.4 ICC cells and the change of nerve fibers to nourishing kidney and reducing urine, Suoquan pill combined with Shenqi Pill group can reduce the number of neural receptors, the expression of gap junction protein decreased, to reduce OAB in guinea pig bladder nerve -ICC- The smooth muscle network signal transduction. It shows that the tonifying kidney and urination method plays the role of tonifying kidney and reducing urine by improving the connection between the neural -ICC- smooth muscle network.

【學位授予單位】：廣州中醫(yī)藥大學
【學位級別】：碩士
【學位授予年份】：2016
【分類號】：R277.5

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