發(fā)布時間：2018-02-07 15:21

  本文關(guān)鍵詞： 糖尿病1型2型 血糖 尿糖 體重 大鼠模型 聽性腦干反應(yīng)(ABR) Cx26 Cx30 Western blot 耳聾 毛細(xì)胞 HE染色 免疫熒光染色 共聚焦　出處：《河北醫(yī)科大學(xué)》2013年博士論文　論文類型：學(xué)位論文

【摘要】：第一部分：復(fù)制糖尿病大鼠模型 背景：隨著我國國民經(jīng)濟(jì)的發(fā)展，人民生活水平的不斷提高，生活方式的改變以及社會的老齡化，糖尿病的患病率逐漸增高，嚴(yán)重威脅我國人民的健康。糖尿病是一種胰島素相對或絕對缺乏而使血糖水平異常增高、脂肪和蛋白質(zhì)代謝紊亂的一種代謝性疾病，糖尿病的并發(fā)癥之一是聽力損害。糖尿病性聽力下降嚴(yán)重影響著人們的生活質(zhì)量，因此越來越受到各方面的高度重視。為研究糖尿病性聽力下降的發(fā)病相關(guān)機(jī)制，必需建立適時的糖尿病動物模型，我們依據(jù)國內(nèi)外公認(rèn)的糖尿病動物模型建模標(biāo)準(zhǔn)，復(fù)制動物模型并監(jiān)測其血糖、體重、尿糖等相關(guān)指標(biāo)，為進(jìn)一步研究提供實驗和理論依據(jù)。 方法：120只Wistar大鼠隨機(jī)分為4組：1型糖尿病實驗組（n=40）、2型糖尿病實驗組（n=40）、1型糖尿病對照組（n=20），2型糖尿病對照組（n=20）。實驗組的Wistar大鼠用兩種不同飼養(yǎng)方法進(jìn)行喂養(yǎng)。不同時間段取尾靜脈全血用血糖儀測定血糖，并檢測體重、尿糖等指標(biāo)。每月對造模后糖尿病大鼠進(jìn)行觀察，包括精神狀態(tài)、活動度、毛發(fā)光澤度、攝食、尿量、耳廓反射等情況。1型糖尿病大鼠隨機(jī)血糖＞16.65mmol/L為成模標(biāo)準(zhǔn)；2型糖尿病大鼠隨機(jī)空腹血糖≥7.8mmol/L為成模標(biāo)準(zhǔn)。 結(jié)果：1實驗動物一般情況 兩型實驗組大鼠均呈現(xiàn)明顯糖尿病癥狀，出現(xiàn)飲水、飲食、尿量增多及消瘦等癥狀，毛發(fā)發(fā)黃無光澤，精神萎靡，活動遲緩，雙側(cè)耳廓反應(yīng)欠靈敏，耳鏡檢查外耳道清潔,正常。正常對照組大鼠無飲水、飲食、尿量增多及消瘦等糖尿病表現(xiàn)，生長及營養(yǎng)狀況良好，雙耳情況正常。 2實驗動物血糖變化 1型糖尿病實驗組大鼠在實驗開始時（0月）隨機(jī)血糖與對照組相比無統(tǒng)計學(xué)差異（P＞0.05），造模后1、2、3、4、5月血糖實驗組與對照組相比升高顯著(P＜0.01)。 2型糖尿病實驗組大鼠在特殊飼料喂養(yǎng)8周后，空腹血糖與對照組相比明顯升高(P＜0.05)，注射STZ后1個月至5個月，血糖升高顯著（與對照組相比P0.01）。 3實驗動物尿糖變化 兩型模型對照組實驗期尿糖均陰性，兩組實驗組造模后尿糖均為(+++或++++)，符合糖尿病動物成模模型標(biāo)準(zhǔn)。 4實驗動物體重變化 1型糖尿病實驗組大鼠體重變化表現(xiàn)為0月時實驗組與對照組無顯著差異（P＞0.05），造模后1、2、3、4、5月實驗組大鼠體重呈持續(xù)降低趨勢，與對照組相比差異顯著(P＜0.01)。 2型糖尿病實驗組大鼠體重變化表現(xiàn)為特殊飼料喂養(yǎng)4周、8周時，實驗組大鼠體重與對照相比顯著增加（P0.01）。注射STZ后1個月，實驗組大鼠體重比注射前有所降低（與特殊飼料喂養(yǎng)8周相比，（P0.05），與對照相比無顯著差異（P＞0.05）；從成模后2月開始，實驗組大鼠的體重呈下降趨勢，與對照組相比差異顯著(P0.01)。 結(jié)論：本實驗成功復(fù)制了近似于人類糖尿病癥征的兩種糖尿病大鼠模型，此兩種模型建立方法簡便易行，是糖尿病慢性并發(fā)癥發(fā)病機(jī)制研究的理想動物模型。 第二部分：糖尿病大鼠聽功能變化與Cx26和Cx30在耳蝸中表達(dá)關(guān)系動態(tài)觀察 背景：耳聾是糖尿病慢性并發(fā)癥之一。糖尿病性耳聾的主要特點是雙側(cè)的對稱性神經(jīng)性耳聾，表現(xiàn)為蝸性和或蝸后性耳聾，以高頻聽力下降為主，常伴有眩暈和前庭功能減退。國內(nèi)外研究糖尿病性耳聾的發(fā)病機(jī)制主要從神經(jīng)病變、微血管病變、血液流變學(xué)異常、免疫反應(yīng)、遺傳學(xué)等方面切入，但其發(fā)病機(jī)制至今尚不清楚,給干預(yù)或治療糖尿病性耳聾帶來了極大困難，這也是醫(yī)學(xué)一直關(guān)注的問題。目前關(guān)于糖尿病內(nèi)耳病變的報道多以外毛細(xì)胞散在缺失為主，其原因有待探討。糖尿病患者的聽力損害可能是耳蝸的微觀結(jié)構(gòu)和中樞聽覺功能發(fā)生病變，因此，本研究將從宏觀和微觀兩方面對此機(jī)理進(jìn)行研究。連接蛋白是細(xì)胞間信號傳遞通道，屬于縫隙連接，目前研究最多的連接蛋白是Cx26和Cx30。縫隙連接是細(xì)胞間通訊的結(jié)構(gòu)基礎(chǔ)，是細(xì)胞間電解質(zhì)、第二信使、和代謝產(chǎn)物等信息傳遞的重要通道。內(nèi)耳的縫隙連接除了輸送營養(yǎng)物質(zhì)、參與K+循環(huán)、交換代謝產(chǎn)物以外，還可能對內(nèi)耳感覺上皮細(xì)胞的功能進(jìn)行調(diào)控，維持內(nèi)耳環(huán)境的穩(wěn)定。K+是內(nèi)耳淋巴液中產(chǎn)生聽力的必不可少的離子，Cx26、Cx30又是K+轉(zhuǎn)運的離子通道之一，因此Cx26，Cx30與K+是聽力產(chǎn)生的必要條件。 正常的哺乳動物耳蝸組織中存在Cx26及Cx30，維持耳蝸的正常功能。糖尿病性耳聾模型大鼠的耳蝸組織中Cx26及Cx30的表達(dá)情況如何？其表達(dá)量與聽力下降是否有關(guān)？國內(nèi)外尚無相關(guān)報道。因此，本研究采用糖尿病大鼠動物模型進(jìn)行聽性腦干反應(yīng)（ABR）監(jiān)測，探尋其聽力功能動態(tài)變化特點。利用Western blot方法，檢測Cx26和Cx30在糖尿病實驗組大鼠中的表達(dá)量，，通過定量分析明確Cx26和Cx30表達(dá)量與動物聽覺功能變化的相關(guān)關(guān)系，為探討糖尿病性耳聾的發(fā)病機(jī)制提供分子生物學(xué)理論實驗依據(jù)。 方法：對照組和實驗組大鼠均采用美國ICS-CHARTR型聽性腦干監(jiān)測儀分別進(jìn)行聽性腦干反應(yīng)(ABR)測試，ABR閾值顯著提高(≥40dBnHL)或波潛伏期及波間期延長作為入選糖尿病性耳聾的標(biāo)準(zhǔn)，進(jìn)入下一步的實驗觀察。在預(yù)定的時間點將動物麻醉后斷頭，快速取出耳蝸，提取耳蝸蛋白，使用Western blot方法，檢測Cx26和Cx30在耳蝸組織中的表達(dá)趨勢，驗證其表達(dá)是否與實驗組動物聽功能變化相一致。 結(jié)果：11型糖尿病大鼠ABR指標(biāo)變化主要是Ⅱ波、Ⅴ波潛伏期、Ⅰ-Ⅴ、Ⅲ-Ⅴ波間期的變化，造模后1個月對照組與實驗組聽力無明顯差異（P＞0.05）；造模后2、3、4、5個月對照組與實驗組Ⅱ波、Ⅴ波潛伏期有顯著差異（P＜0.01）、Ⅰ-Ⅴ、Ⅲ-Ⅴ波間期（P＜0.05）有差異。 22型糖尿病大鼠ABR指標(biāo)變化主要是Ⅲ波、Ⅴ波潛伏期、Ⅰ-Ⅲ、Ⅰ-Ⅴ波間期的變化，造模后1個月對照組與實驗組聽力無顯著差異（P＞0.05）；造模后2、3、4、5個月對照組與實驗組Ⅲ波、Ⅴ波潛伏期、Ⅰ-Ⅲ、Ⅰ-Ⅴ波間期（P＜0.01）具有顯著差異。 32型糖尿病大鼠模型中，耳蝸Cx26、Cx30蛋白表達(dá)水平與對照組大鼠相比有顯著下降，隨著造模時間的延長，Cx26和Cx30蛋白表達(dá)逐漸降低，Cx26蛋白較Cx30蛋白變化更為明顯。 結(jié)論： 1兩型糖尿病性聾大鼠從造模2個月開始，大鼠的聽力出現(xiàn)下降。兩型糖尿病大鼠都有不同程度的聽力損害，腦干聽誘發(fā)電位可作為早期診斷 其聽力下降的指標(biāo)之一。 2從造模2個月開始，實驗組耳蝸組織中的Cx26、Cx30蛋白的表達(dá)水平有改變，Cx26和Cx30蛋白表達(dá)均有下降趨勢，Cx26和Cx30變化與聽功能改變相一致。 第三部分：糖尿病性耳聾大鼠Cx26、Cx30表達(dá)變化與耳蝸毛細(xì)胞損傷關(guān)系初步研究 背景：隨著糖尿病患者人數(shù)的增加，由糖尿病引起的聽力損害的報道也逐漸增多。糖尿病聽力損害起病隱匿，易與老年性耳聾混淆，待發(fā)現(xiàn)時常發(fā)展為不可逆病變，嚴(yán)重影響患者的生活質(zhì)量。對糖尿病性耳聾發(fā)病機(jī)制的深入研究，是預(yù)防糖尿病性耳聾不可缺少的環(huán)節(jié)。 在糖尿病性耳聾大鼠模型中，尤其是2型糖尿病性耳聾大鼠模型，隨著時間的延長，聽力損失越明顯，影響聽覺功能。維持耳蝸正常機(jī)械-電轉(zhuǎn)換功能以及保證聽力的正常傳導(dǎo)作用的分子是縫隙連接蛋白，糖尿病性耳聾大鼠耳蝸中的連接蛋白，在成模后兩個月發(fā)生變化，病變時間越長，其表達(dá)降低越顯著。為進(jìn)一步證實聽力損失與耳蝸組織中的Cx26和Cx30相關(guān)性，本研究采用熒光免疫染色方法、HE染色，檢測Cx26和Cx30在實驗組大鼠中的分布是否與耳蝸組織的蛋白表達(dá)水平及聽力損失相一致，并進(jìn)行了全耳蝸鋪片，觀察毛細(xì)胞損失情況與聽力損失的相關(guān)性。進(jìn)一步探討了Cx26、Cx30表達(dá)與糖尿病性耳聾的發(fā)病機(jī)制關(guān)系，從分子水平為糖尿病性聾的發(fā)病機(jī)制提供新的研究方向和理論實驗依據(jù)。 方法：大鼠麻醉后斷頭，取出耳蝸，固定、脫鈣、石蠟包埋、切片。挑選結(jié)構(gòu)完整耳蝸石蠟切片，分別進(jìn)行HE染色及免疫組織化學(xué)染色，檢測Cx26和Cx30在糖尿病性耳聾大鼠耳蝸的表達(dá)分布，激光共聚焦顯微鏡觀察耳蝸的結(jié)構(gòu)形態(tài)及Cx26和Cx30的表達(dá)情況是否與耳蝸組織的病理變化相一致，最后通過全耳蝸鋪片來觀察毛細(xì)胞損失情況與聽力損失的相關(guān)性。 結(jié)果：1兩型糖尿病大鼠HE染色耳蝸側(cè)壁組織、基底膜均出現(xiàn)實驗組較正常組變薄，實驗組大鼠耳蝸組織血管紋中管腔樣結(jié)構(gòu)與對照組耳蝸組織血管紋相比較出現(xiàn)減少，耳蝸側(cè)壁及基底膜嗜伊紅染色實驗組較正常組深染，實驗組與對照組大鼠耳蝸螺旋韌帶中和螺旋神經(jīng)節(jié)細(xì)胞數(shù)量比較有統(tǒng)計學(xué)差異（P㩳0.05），實驗組大鼠耳蝸螺旋韌帶中和螺旋神經(jīng)節(jié)細(xì)胞數(shù)量少于對照組。 2兩型糖尿病大鼠造模后1月實驗組和對照組耳蝸內(nèi)基底膜、血管紋和螺旋韌帶中Cx26、Cx30表達(dá)無統(tǒng)計學(xué)差異（P㧐0.05），造模后2、3、4、5月實驗組和對照組耳蝸內(nèi)基底膜、血管紋和螺旋韌帶中Cx26、Cx30表達(dá)水平顯著低于對照組（P㩳0.05）。 3兩型糖尿病大鼠模型造模后1月實驗組和對照組大鼠耳蝸鋪片比較，內(nèi)外毛細(xì)胞均無損傷。 4兩型糖尿病大鼠模型造模后2月實驗組和對照組大鼠耳蝸鋪片比較，實驗組較對照組內(nèi)、外毛細(xì)胞排列散亂，但無缺失；造模后3月實驗組較正常組內(nèi)、外毛細(xì)胞排列散亂，其程度較2月時嚴(yán)重，但內(nèi)外毛細(xì)胞無缺失；造模后4、5月實驗組較正常組內(nèi)、外毛細(xì)胞排列散亂，并出現(xiàn)內(nèi)外毛細(xì)胞的缺失。這些均與實驗組耳蝸組織中的Cx26和Cx30蛋白表達(dá)水平較正常組在明顯下降一致，與ABR聽力測試結(jié)果相符合。 結(jié)論： 1Cx26及Cx30是聽力產(chǎn)生的必不可少的連接蛋白，推測糖尿病大鼠聽力損害可能與Cx26、Cx30相關(guān)。 2兩型型糖尿病大鼠造模后1月實驗組和對照組耳蝸內(nèi)基底膜、血管紋和螺旋韌帶中Cx26、Cx30表達(dá)水平無化，造模后2、3、4、5月實驗組和對照組耳蝸內(nèi)基底膜、血管紋和螺旋韌帶中Cx26、Cx30表達(dá)顯著低于對照組。與聽力變化較為一致。 3耳蝸內(nèi)外毛細(xì)胞實驗組較對照組排列散亂，隨時間延長毛細(xì)胞損傷程度加重，與蛋白表達(dá)及聽力測試結(jié)果均一致。兩型糖尿病大鼠模型在造模后2月Cx26和Cx30在耳蝸組織中的表達(dá)較對照組均有降低，隨時間延長其降低程度越顯著。提示糖尿病性耳聾的聽力減退可能與Cx26和Cx30表達(dá)降低有關(guān)。
[Abstract]:Part one: replicating diabetic rat model
Background: with the development of our national economy, improve the people's living standards, changes in lifestyle and social aging, the prevalence of diabetes increased gradually, a serious threat to people's health in China. Diabetes is an absolute or relative lack of insulin to abnormally high blood sugar levels, a fat metabolic disease and protein metabolism disorders, one of the complications of diabetes is diabetic hearing impairment. The hearing loss seriously affects people's quality of life, so it attracts more and more attention from various aspects. For the onset mechanism of diabetes hearing loss, must establish the animal model of diabetes in time, based on our modeling standard animal model of diabetes at home and abroad accepted, to establish the animal model and monitor their blood glucose, body weight, urine and other related indicators, to provide experimental and theoretical basis for the further study.
Methods: 120 Wistar rats were randomly divided into 4 groups: type 1 diabetes mellitus group (n=40), type 2 diabetes mellitus group (n=40), type 1 diabetes control group (n=20), type 2 diabetes control group (n=20). Wistar rats in the experimental group with two different feeding methods of feeding. Different time from tail vein whole blood glucose was determined by blood glucose meter, and the body weight, glucose and other indicators. The observation of monthly DM rats, including mental state, activity, hair gloss, food intake, urine volume, and auricle reflex type.1 diabetic rats were randomized into model 16.65mmol/L blood sugar. Standard; type 2 diabetes mellitus rats were fasting blood glucose than 7.8mmol/L to become the standard model.
Results: 1 the general situation of experimental animals
Two types of experimental rats showed obvious symptoms of diabetes, drinking water, diet, urine volume increased and weight loss and other symptoms of yellow hair, no gloss, listlessness, slow reaction, bilateral ear less sensitive, ear canal clean and microscopic examination, normal. Rats in normal control group without drinking water, diet, urine volume increased and weight loss and other diabetic symptoms, growth and nutritional status, ears were normal.
2 blood glucose changes in experimental animals
In the experimental group of type 1 diabetes, there was no significant difference in the blood glucose at the beginning of the experiment (0 months) between the experimental group and the control group (P > 0.05). The blood glucose in the experimental group was significantly higher than that in the control group after 1,2,3,4,5 months (P < 0.01).
In type 2 diabetes mellitus group, fasting blood glucose increased significantly after 8 weeks of special feed (P < 0.05), and blood sugar increased significantly from 1 months to 5 months after STZ injection (P0.01 compared with control group).
3 experimental animal urine changes
Type two model control group the urine were negative, two groups of experimental group after modeling the urine were (+ + + + + + +, or) with diabetes animal model standard model.
4 body weight changes in experimental animals
The body weight changes in the experimental group of type 1 diabetes showed no significant difference between the experimental group and the control group at 0 months (P > 0.05). After modeling for 1,2,3,4,5 months, the body weight of the experimental group showed a decreasing trend, which was significantly different from that of the control group (P < 0.01).
Weight change in type 2 diabetic rats in the experimental group showed a special diet for 4 weeks, 8 weeks, body weight of rats in the experimental group compared with the control group increased significantly (P0.01). 1 months after STZ injection, body weight of rats in the experimental group was lower than that before injection (compared with the special diet for 8 weeks (P0.05). Compared with the control, no significant difference (P > 0.05); from the model after the beginning of February, the experimental group rats weight decreased, compared with the control group had significant difference (P0.01).
Conclusion: the two diabetic rat models similar to human diabetes symptoms were successfully replicated in the experiment. The two models are simple and convenient, and are ideal animal models for studying the pathogenesis of diabetic chronic complications.
The second part: the dynamic observation of the relationship between the changes of the auditory function of diabetic rats and the expression of Cx26 and Cx30 in the cochlea
Background: deafness is one of the chronic complications of diabetes. The main characteristics of diabetic deafness is the symmetry of bilateral nerve deafness, cochlear and performance or retrocochlear deafness, with high-frequency hearing loss, often accompanied by dizziness and vestibular dysfunction. The pathogenesis of diabetic deafness at home and abroad mainly from neuropathy, micro vascular disease, abnormal blood rheology, immune response, genetics and other aspects, but its pathogenesis is still unclear, brought great difficulties to the intervention or treatment of diabetic deafness, which is the medicine has been a concern. At present about diabetic inner ear pathological reports outside hair cells scattered in the absence of the main, to causes of hearing impairment. Patients with diabetes may be the micro structure and the central auditory function of cochlear lesions, therefore, this study will be two from the macro and micro aspects of this machine Make a research on the connection. Protein is the signal transmission path between cells belonging to gap junctions, one of the most connected protein is Cx26 and Cx30. is the structural basis of gap junction intercellular communication, one cell electrolyte, second messenger and metabolites information important channel. The inner gap junctions in addition to transporting nutrients in K+, circulation, exchange of metabolites, may also be the inner ear sensory epithelial cell function regulation, maintain the stability of the.K+ environment is essential for inner ear hearing ear ion, Cx26, and Cx30 is one of the K+ ion channel transport in the Cx26, Cx30 and K+ is a necessary condition for the hearing.
The presence of Cx26 and Cx30 in normal mammalian cochlea, maintain the normal function of the cochlea. How the expression of Cx26 and Cx30 in the cochlear tissue of diabetic deafness rats in? And the expression level of hearing loss is about? There is no relevant reports at home and abroad. Therefore, this study used the animal model of diabetic rat auditory brainstem reaction monitoring (ABR), to explore the dynamic changes of hearing function. By using Western blot method, the expression detection of Cx26 and Cx30 in diabetic rats in the experimental group, through quantitative analysis of the expression of Cx30 and Cx26 clear correlation and dynamic changes of auditory function, provide experimental basis for the theory of molecular biology to explore the pathogenesis of diabetes deafness.
Methods: the control group and the experimental groups of rats were ICS-CHARTR respectively monitor auditory brainstem auditory brainstem response (ABR) test, the ABR threshold was significantly increased (more than 40dBnHL) or latencies and wave interval prolongation was selected as diabetic deafness, enter into the next experiment. The breakage in the predetermined anesthesia the time will come out quickly after the animal cochlea and cochlear protein extraction, using Western blot method, the expression of Cx26 and Cx30 in the detection of trends in cochlear tissues, verify whether the expression and experimental group animal auditory function changes consistent.
Results: the changes of ABR index in type 11 diabetic rats is mainly II wave, V wave latency, I-V, changes of III-V wave interval, made 1 months after the experimental group and the control group hearing had no significant difference (P > 0.05); model 2,3,4,5 months after the experimental group and the control group II V wave wave, the incubation period had significant difference (P < 0.01), I-V, III-V wave interval (P < 0.05) difference.
The change of ABR index in type 22 diabetic rats is mainly III wave, V wave latency of I-III, changes of I-V wave interval model, 1 months after the experimental group and the control group hearing no significant difference (P > 0.05); after modeling 2,3,4,5 months the control group and the experimental group III wave. V wave latency of I-III, I-V wave interval (P < 0.01) has significant differences.
In type 32 diabetic rats, the expression level of Cx26 and Cx30 protein in cochlea decreased significantly compared with control rats. With the prolongation of modeling time, the expression of Cx26 and Cx30 decreased gradually, and Cx26 protein changed more significantly than Cx30 protein.
Conclusion:
1, type two diabetic deafness rats started hearing loss for 2 months from model building. All types two diabetic rats had different degrees of hearing loss. Brainstem auditory evoked potentials could be used as early diagnosis.
One of the indicators of its hearing loss.
2, from the 2 months after modeling, the expression level of Cx26 and Cx30 protein in the cochlea tissue of experimental group changed, and the expression of Cx26 and Cx30 decreased. The change of Cx26 and Cx30 was consistent with the change of auditory function.
The third part: a preliminary study on the relationship between the changes of Cx26, Cx30 expression and cochlear hair cell damage in diabetic deafness rats
Background: with the increase in the number of patients with diabetes, hearing impairment caused by diabetes are increasingly reported. Diabetic hearing impairment onset occult, easily confused with senile deafness, often to be found for the development of irreversible lesions, seriously affecting the quality of life of patients. Further research on diabetes deafness pathogenesis, prevention diabetes deafness is an indispensable link.
In the model of diabetic deafness rats, especially the model of type 2 diabetic deafness rats, with the extension of time, the hearing loss is more obvious, effect of auditory function. To maintain the molecular normal cochlea mechano electrical conversion function and ensure the normal hearing conduction is the gap junction protein, connexin in diabetic rat cochlear deafness the change in the two months after modeling, the longer the disease, its expression decreased more significantly. In order to further confirm the Cx26 and Cx30 correlation between hearing loss and cochlear tissues, this study used immunofluorescence staining, HE staining, the distribution of detection of Cx26 and Cx30 in experimental rats with cochlear tissue protein expression level and hearing loss are consistent, and the whole cochlea preparation and observation of the relationship between hearing loss of hair cell loss. To further explore the expression of Cx30 and Cx26, diabetes mellitus The pathogenesis of deafness provides new research direction and theoretical experimental basis from molecular level to the pathogenesis of diabetic deafness.
Methods: the rats were anesthetized and decapitated, remove the cochlea, fixed, decalcified, paraffin embedded, sliced. Select the structural integrity of cochlear paraffin sections, HE staining and immunohistochemical staining respectively, to detect Cx26 and Cx30 expression in diabetic rat cochlear deafness distribution and expression were observed by confocal cochlear morphology and structure Cx26 and Cx30 microscopy and pathological changes of cochlea is consistent, correlation finally through the whole cochlear sections to observe the situation and hearing loss of hair cell loss.
Results: 1 type two diabetic rats HE staining of cochlear lateral wall tissue, basement membrane appeared in the experimental group compared with the normal group, thinning, lumen like structure stria cochlea tissue in the experimental group rats compared with control group, the cochlea stria appears to reduce the lateral wall of the cochlea and basement membrane eosinophilic staining in experimental group compared with normal group anachromasis, experimental group and control group, the number of rat cochlea spiral ligament and spiral ganglion cells were statistically significant difference (P? 0.05), the number of spiral ligament rats and spiral ganglion cells than the control group.
2 rats of type two diabetes model in January after the experimental group and the control group in the cochlear basilar membrane Cx26, stria vascularis and spiral ligament, the expression of Cx30 had no significant difference (P? 0.05), modeling 2,3,4,5 months after the experimental group and the control group in the cochlear basilar membrane Cx26, stria vascularis and spiral ligament, the expression level of Cx30 was significantly lower than the control group (P? 0.05).
3 the model of type two diabetic rat model was made in the experimental group and the control group was compared with the cochlear paving in the control group. There was no damage to the inner and outer hair cells.
4 type two diabetic rat model after the model February experimental group and control group rats cochlea preparation, experimental group than in the control group, the outer hair cells arranged in disorder, but no lack of modeling after March; the experimental group compared with the normal group, the outer hair cells were irregular and scattered, the degree is serious in February, but the outer hair cell missing; modeling 4,5 months after the experimental group compared with the normal group, the outer hair cells were irregular and scattered, and lack of inner and outer hair cells. These were the experimental group and the cochlear tissue Cx26 and the expression level of Cx30 protein decreased significantly compared with normal group in uniform, consistent with the ABR listening test results.
Conclusion:
1Cx26 and Cx30 are essential connexin in hearing generation. It is presumed that hearing impairment in diabetic rats may be associated with Cx26, Cx30

【學(xué)位授予單位】：河北醫(yī)科大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2013
【分類號】：R587.2;R764.43

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