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  本文選題：慢性前列腺炎 + 慢性非細菌前列腺炎�。� 參考：《安徽醫(yī)科大學》2014年博士論文

【摘要】：目的采用17p-雌二醇聯(lián)合去勢建立SD大鼠慢性非細菌性前列腺炎模型(Chronic Nonbacterial Prostatitis,CNBP),評價造模結(jié)果。 方法取2月齡SPF級雄性Sprague Dawley(SD)大鼠80只,隨機分為8組,即A組：空白對照組(blank control)(n=10);B組：生理鹽水組(saline)(n=10);C組：假手術(shù)組(sham operation)(n=10);D組：單純?nèi)萁M(Castration)(n=10);E組：單純17β-雌二醇(17β-Estradiol)(0.25mg/2ml/kg)(n=10);F組：17p-雌二醇(0.15mg/2ml/kg)(低劑量組)聯(lián)合去勢(n=10)、G組：17β-雌二醇(0.20mg/2ml/kg)(中劑量組)聯(lián)合去勢(n=10)；H組：17β-雌二醇(0.25mg/2ml/kg)(高劑量組)聯(lián)合去勢(n=10)。按照國際前列腺炎組織學診斷與分度標準評估造模結(jié)果,并分析不同濃度17β-雌二醇聯(lián)合去勢建模效果。 結(jié)果大體形態(tài)觀察可見H組大鼠前列腺與周圍組織粘連較緊密,充血水腫,前列腺包膜不完整。光鏡下觀察見前列腺組織結(jié)構(gòu)破壞,伴有不均勻淋巴樣組織增生,前列腺導(dǎo)管破壞或者部分基底膜破壞,炎性細胞多成群狀分布,可見淋巴小結(jié)或濾泡形成。D組、E組、F組及G組光鏡下可見散在淋巴細胞及單核細胞浸潤,表現(xiàn)為輕度或中度炎癥。A組、B組、C組大鼠前列腺大體形態(tài)觀察可見包膜完整腺體表面較光滑、平整。光鏡下觀察未見明顯組織炎癥表現(xiàn)。在評價炎癥的核心指標方面,H組大鼠慢性前列腺炎組織學炎癥分級與D組、E組、F組及G組比有統(tǒng)計學差異(P0.05),表現(xiàn)出中、重度的炎癥反應(yīng)。而D組、E組、F組及G組兩兩相比無統(tǒng)計學差異(P0.05)。炎癥范圍而言,H組大鼠慢性前列腺炎炎癥分級同D組、E組、F組及G組比有統(tǒng)計學差異(P0.05),表現(xiàn)出多灶性或彌漫性的炎癥反應(yīng),而其他組表現(xiàn)為局灶性或多灶性表現(xiàn)。 結(jié)論17β-雌二醇聯(lián)合去勢可以誘導(dǎo)建立SD大鼠慢性非細菌性前列腺炎(CNBP)模型,建模成功后組織病理學表現(xiàn)與臨床非細菌慢性前列腺炎病理表現(xiàn)相似,其組織病理炎癥與17β-雌二醇濃度有一定的正相關(guān)性。單一采用17p-雌二醇或者去勢可以誘導(dǎo)大鼠前列腺產(chǎn)生輕度的、局灶性炎癥,臨床代表性較差。從評價炎癥的核心指標來說,單一采用17β-雌二醇或者去勢手術(shù)誘導(dǎo)的SD大鼠的慢性非細菌性前列腺炎效果不夠穩(wěn)定,不適合作為動物模型。 背景及目的前列腺炎,特別是慢性前列腺炎(CP)一直是泌尿外科和男科的主要問題。該病發(fā)病機制尚不明確,而且治愈率低、復(fù)發(fā)率高,可以影響男性性功能及生殖功能,給患者身心健康及家庭帶來不利影響,因此了解其發(fā)病機制對于慢性前列腺炎的治療十分必要。目前有關(guān)慢性前列腺病因?qū)W研究多集中于病原學、尿液返流、上皮功能異常、神經(jīng)內(nèi)分泌異常等方面。近年相關(guān)研究表明,慢性前列腺炎的發(fā)生與神經(jīng)內(nèi)分泌之間有一定的聯(lián)系。前列腺神經(jīng)內(nèi)分泌細胞(Neuroendocrine cell, NEC)又稱胺前體攝取及脫羧(Amine Precursor Uptake And Decarboxylation,APUD)細胞,研究發(fā)現(xiàn)前列腺神經(jīng)內(nèi)分泌細胞存在分泌上皮細胞之間,可以通過分泌激素多肽及5-羥色胺發(fā)揮作用,調(diào)節(jié)前列腺細胞生長及分泌活動。本文主要通過采用17β-雌二醇聯(lián)合去勢手術(shù)(雙側(cè)睪丸切除)建立SD大鼠非細菌性慢性前列腺炎(CNBP)模型,研究神經(jīng)內(nèi)分泌細胞重要標志性蛋白：嗜鉻粒蛋白A(Chromogranin A, CgA)、神經(jīng)醇化酶(Neuronal specific enolase, NSE)、神經(jīng)生長因子(Nerve growth factor, NGF)及P物質(zhì)(Substance P, SP)在慢性前列腺炎組織中的改變及其相關(guān)性,以了解神經(jīng)分泌分化在慢性前列腺炎發(fā)病中的作用。 方法利用建立的SD大鼠的慢性非細菌性前列腺炎動物模型,分別利用免疫組化組織化學(Immunohistochemical,IHC)、免疫印跡(Western blot, WB)、酶聯(lián)免疫吸附法(Enzyme-linked immunosorbent assay, ELLS A)及實時定量PCR(Real-time quantitative PCR, qPCR)從蛋白及基因表達水平研究A組、B組、C組、D組、E組、H組不同組別大鼠神經(jīng)內(nèi)分泌細胞重要標志蛋白：嗜鉻粒蛋白A(CgA)、神經(jīng)醇化酶(NSE)、神經(jīng)生長因子(NGF)及炎癥相關(guān)P物質(zhì)(SP)的變化,并分析嗜鉻粒蛋白A(CgA)、神經(jīng)醇化酶(NSE)、神經(jīng)生長因子(NGF)分別與P物質(zhì)(SP)的相關(guān)性。 結(jié)果 1.免疫組織化學結(jié)果顯示CgA、CgA、NSE、SP在模型組H組表達較A組、B組、C組、D組、E組增強。 2.免疫印跡半定量研究發(fā)現(xiàn)神經(jīng)內(nèi)分泌細胞代表性蛋白CgA、NSE、NGF在慢性前列腺炎模型組(H組)表達上調(diào),與其A組、B組、C組、D組、E組比較有統(tǒng)計學差異(P0.05)。 3.酶聯(lián)免疫吸附法定量檢測大鼠血清模型組(H組)CgA、NSE、NGF、SP物質(zhì)含量在慢性前列腺炎模型組(H組)表達上調(diào),與其它各組兩者相比有統(tǒng)計學意義(P0.05)。相關(guān)性分析得出Cg A與SP的表達成正相關(guān)性(γ,=0.35,P0.01)；NSE與SP的表達成正相關(guān)(γ=0.29, P0.01); NGF與SP的表達正相關(guān)(γ,=0.50,P0.01)。 4.實時定量PCR定量研究發(fā)現(xiàn)CgA (F=109.4)、NSE (F=68.4)、NGF (F=65.4). SP (F=61.1)在慢性非細菌性前列腺炎模型組中表達上調(diào),與其他各組相比有統(tǒng)計學意義(P0.05)。相關(guān)性分析得出CgA與SP的表達成正相關(guān)性(γ=0.45, P0.01); NSE與SP的表達成正相關(guān)(γ=0.53,P0.01)；NGF與SP的表達正相關(guān)(γ=0.37,P0.01) 結(jié)論 1.17β-雌二醇皮下注射聯(lián)合去勢手術(shù)建立慢性非細菌性前列腺炎模型組中神經(jīng)內(nèi)分泌細胞標志性蛋白CgA、NSE、NGF表達上調(diào),提示慢性前列腺炎中存在神經(jīng)內(nèi)分泌分化現(xiàn)象,推測由17p-雌二醇和(或)去勢誘導(dǎo)產(chǎn)生。 2. CgA、NSE、NGF分別與SP存在相關(guān)性,提示神經(jīng)內(nèi)分泌分化可能與炎癥相關(guān),SP是重要的致痛因子,推測神經(jīng)內(nèi)分泌細胞可能參與引起慢性非細菌性前列腺炎的神經(jīng)原性疼痛。
[Abstract]:Objective to establish 17p- SD (Chronic) Nonbacterial Prostatitis (CNBP) model by using estradiol combined with castration, and to evaluate the results of modeling.
Methods 80 rats of 2 month old SPF grade Sprague Dawley (SD) were randomly divided into 8 groups, namely group A, blank control (n=10), B group: normal saline group (saline), C group: sham operation group; simple 17 beta estradiol (17 beta) ) (n=10); group F: 17p- estradiol (0.15mg/2ml/kg) (low dose group) combined castration (n=10), group G: 17 beta estradiol (0.20mg/2ml/kg) (medium dose group) combined castration (n=10); H group: 17 beta estradiol (0.25mg/2ml/kg) (0.25mg/2ml/kg) (high dose group) combined castration (n =10). Evaluation of model results according to the histological diagnosis and graduation criteria of international prostatitis and analysis Different concentrations of 17 beta estradiol combined with castration were used to model the effect.
Results the general morphological observation showed that the prostate and surrounding tissue were closely adhered to the prostate in the H group, and the hyperemia and edema were not complete. The damage of the prostate tissue structure was observed under the light microscope, accompanied by the uneven lymphoid tissue hyperplasia, the destruction of the prostatic catheter or the broken part of the basement membrane, and the distribution of inflammatory cells in groups, and the lymph nodes were visible. Or follicular formation in group.D, group E, group F and group G showed infiltration of lymphocytes and mononuclear cells under light microscope, which showed mild or moderate inflammation in group.A, B group, group B and group C, the whole glandular surface of the prostate gland was smooth and smooth. No obvious tissue inflammation was observed under the light microscope. The core index of inflammation was evaluated. The histologic grade of chronic prostatitis in group H was significantly different from group D, group E, group F and G group (P0.05), showing a moderate and severe inflammatory response. There was no statistical difference between group D, E, F and G group 22 (P0.05). Differences (P0.05) showed multifocal or diffuse inflammatory response, while the others showed focal or multifocal manifestations.
Conclusion the combined castration of 17 beta estradiol can induce the establishment of a chronic non bacterial prostatitis (CNBP) model in SD rats. The histopathological performance of the model is similar to that of the clinical non bacterial chronic prostatitis after successful modeling. The histopathological inflammation is positively correlated with the concentration of 17 beta estradiol. A single 17p- estradiol or castration is used alone. It can induce mild, focal inflammation and poor clinical representation in the prostate of rats. From the core indicators of inflammation, the effect of a single 17 beta estradiol or castrated SD rat chronic non bacterial prostatitis is not stable and is not suitable as an animal model.
Background and objective prostatitis, especially chronic prostatitis (CP), has always been the main problem in the Department of Urology and the male family. The pathogenesis of the disease is not clear, the cure rate is low, the recurrence rate is high, the male sexual function and reproductive function can be affected, and the patients' physical and mental health and family are adversely affected. Therefore, the pathogenesis of the disease is understood to be chronic. The treatment of prostatitis is very necessary. At present, the study of chronic prostatic etiology focuses on etiology, urine reflux, abnormal epithelial function, and neuroendocrine abnormalities. In recent years, related studies have shown that there is a certain relationship between the occurrence of chronic prostatitis and neuroendocrine cells. Neuroendocri NE cell, NEC), also known as amine precursor uptake and decarboxylation (Amine Precursor Uptake And Decarboxylation, APUD) cells, found that the prostate neuroendocrine cells exist between secretory epithelial cells, and can play a role by secreting hormone peptides and 5- serotonin to regulate the growth and secretion of anterior gland cells. This article mainly adopts 17 The non bacterial chronic prostatitis (CNBP) model of SD rats was established with beta estradiol combined with castration (bilateral testicle excision) to study the important markers of neuroendocrine cells: chromogranin A (Chromogranin A, CgA), neuroglycolase (Neuronal specific enolase, NSE), nerve growth factor (Nerve growth) and substances ( Substance P (SP) changes in chronic prostatitis tissues and their correlation, in order to understand the role of neurosecretory differentiation in the pathogenesis of chronic prostatitis.
Methods using the established model of chronic non bacterial prostatitis in SD rats, Immunohistochemical, IHC, Western blot, WB, enzyme linked immunosorbent assay (Enzyme-linked immunosorbent assay, ELLS A) and real-time quantitative PCR were used. The expression level of A group, group B, group C, group D, group E, group H, and group H were important markers of neuroendocrine cells: chromaffin A (CgA), neuroalkinase (NSE), nerve growth factor (NGF) and P substance in inflammation (SP), and the analysis of chromaffin protein, neurotrophic factor and nerve growth factor respectively The correlation of substance (SP).
Result
1. immunohistochemical results showed that CgA, CgA, NSE and SP in the model group H group were higher than those in A group, B group, C group, D group and E group.
2. the semi quantitative study of immunoblotting found that the expression of representative protein CgA, NSE, NGF in the chronic prostatitis model group (group H) was up regulated, and there was a statistically significant difference between the group A, the B group, the C group, the D group and the E group (P0.05).
The expression of CgA, NSE, NGF, SP in the model group (group H) of chronic prostatitis (group H) was up regulated by 3. enzyme linked immunosorbent assay (group H), compared with other groups (P0.05). The correlation analysis showed that the expression of Cg A and SP was positive correlation (gamma, =0.35, P0.01), and positive correlation (gamma, =0.35, P0.01). 9, P0.01); NGF is positively correlated with the expression of SP (gamma, =0.50, P0.01).
4. quantitative study of real-time quantitative PCR found that CgA (F=109.4), NSE (F=68.4), NGF (F=65.4). SP (F=61.1) was up regulated in the chronic non bacterial prostatitis model group (P0.05). The correlation analysis showed that CgA was positively correlated with the SP table. 3, P0.01); NGF is positively correlated with the expression of SP (gamma =0.37, P0.01).
conclusion
1.17 beta estradiol subcutaneous injection combined with castration to establish a chronic nonbacterial prostatitis model group, the expression of neuroendocrine cell marker protein CgA, NSE, NGF expression was up-regulated, suggesting the existence of neuroendocrine differentiation in chronic prostatitis, which is presumed to be induced by 17p- estradiol and (or) castration.
2. CgA, NSE, and NGF are associated with SP, suggesting that neuroendocrine differentiation may be associated with inflammation. SP is an important pain causing factor. It is suggested that neuroendocrine cells may be involved in neurogenic pain causing chronic abacterial prostatitis.

【學位授予單位】：安徽醫(yī)科大學
【學位級別】：博士
【學位授予年份】：2014
【分類號】：R697.33;R-332

【參考文獻】

相關(guān)期刊論文 前6條

1 周曉輝,韓蕾,周智恒,劉忠德,楊吉相,呂延偉,尤春來;免疫性慢性非細菌性前列腺炎大鼠模型的形態(tài)學與分子生物學特性[J];中華男科學雜志;2005年04期

2 樊松;梁朝朝;郝宗耀;張賢生;張翼飛;周駿;楊勇飛;唐智國;;新型鈣通道TRPV5在慢性前列腺炎患者前列腺上皮細胞中的表達及其意義[J];中國男科學雜志;2007年10期

3 李冬梅;胡文娟;劉向云;張曉芳;閆晗;謝淑武;吳建輝;孫祖越;;大鼠自體免疫性前列腺炎模型的實驗研究[J];實驗動物與比較醫(yī)學;2008年03期

4 樊松;梁朝朝;張賢生;郝宗耀;周駿;張翼飛;江長琴;;慢性非細菌性前列腺炎小鼠前列腺組織TRPV5的表達[J];江蘇醫(yī)藥;2011年07期

5 張亞強;王炎;李敏;盧建新;高筱松;劉兵;龐然;;大鼠前列腺組織抗原蛋白誘導(dǎo)自身免疫性前列腺炎模型的建立[J];中國中西醫(yī)結(jié)合外科雜志;2008年06期

6 梁朝朝,張學軍,王克孝;前列腺炎病因?qū)W研究進展[J];中華泌尿外科雜志;2003年06期

，

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