本文選題：卡那霉素 + 速尿�。� 參考：《福建醫(yī)科大學(xué)》2011年碩士論文

【摘要】：感音神經(jīng)性聾嚴(yán)重影響著人類的健康和生存質(zhì)量,但目前尚無有效治療方法,毛細(xì)胞再生是治療感音神經(jīng)性聾的關(guān)鍵。隨著分子生物學(xué)、分子遺傳學(xué)、基因工程技術(shù)等的不斷發(fā)展,尤其是近年來基因調(diào)控內(nèi)耳毛細(xì)胞再生方面的研究逐步深入,為治療感音神經(jīng)性聾開辟了一條嶄新的道路。本研究通過建立藥物性聾動物模型,并進(jìn)行Hath1基因和DAPT聯(lián)合導(dǎo)入內(nèi)耳誘導(dǎo)毛細(xì)胞再生,探索感音神經(jīng)性聾藥物治療的方法。 第一部分速尿和卡那霉素聯(lián)合用藥后大鼠聽功能和內(nèi)耳毛細(xì)胞損害觀察目的:探討速尿和硫酸卡那霉素聯(lián)合用藥快速制備大鼠藥物性聾動物模型的方法。方法:實(shí)驗(yàn)組大鼠靜脈注射速尿和/或肌肉注射硫酸卡那霉素,正常對照組不予任何處理。給藥3天、7天、2個(gè)月后行聽覺腦干誘發(fā)電位(auditory brainstem response ABR)閾值檢測,給藥7天后行耳蝸掃描電鏡、全耳蝸基底膜鋪片及內(nèi)耳冰凍切片免疫熒光染色和內(nèi)耳冰凍切片HE染色觀察。結(jié)果:聯(lián)合應(yīng)用不同劑量速尿和硫酸卡那霉素后,大鼠ABR閾值出現(xiàn)不同程度的提高,給藥3天、7天、2個(gè)月后閾值兩兩比較均無統(tǒng)計(jì)學(xué)意義;掃描電鏡下耳蝸毛細(xì)胞出現(xiàn)由底回到頂回、由外毛細(xì)胞到內(nèi)毛細(xì)胞的不同程度損傷。而單用速尿或硫酸卡那霉素的大鼠并不出現(xiàn)閾值提高或毛細(xì)胞損傷。耳蝸內(nèi)、外毛細(xì)胞完全損傷的大鼠,支持細(xì)胞大部分完好,前庭毛細(xì)胞未見損傷。結(jié)論:速尿和硫酸卡那霉素具有明顯的耳毒性協(xié)同作用,聯(lián)合用藥可導(dǎo)致大鼠聽功能嚴(yán)重受損并且用藥2個(gè)月致聾大鼠聽力無恢復(fù),但對耳蝸支持細(xì)胞和前庭毛細(xì)胞損傷較輕;速尿和卡那霉素聯(lián)合用藥是建立大鼠藥物性聾動物模型的一種簡單、快速且較可靠的方法。 第二部分Hath1基因和DAPT聯(lián)合導(dǎo)入內(nèi)耳誘導(dǎo)毛細(xì)胞再生的初步研究目的:探討Hath1基因和DAPT聯(lián)合導(dǎo)入內(nèi)耳后,Hath1在內(nèi)耳的表達(dá)及誘導(dǎo)毛細(xì)胞再生的情況。方法:正常大鼠與致聾大鼠(ABR閾值 110dB SPL)各15只,經(jīng)背側(cè)入路暴露聽泡,去除部分聽泡骨暴露鼓階,于鼓階近圓窗處打孔微量進(jìn)液器導(dǎo)入Hath1基因,隨后用膠囊滲透壓泵經(jīng)鼓階持續(xù)導(dǎo)入DAPT 4周。行ABR閾值檢測,耳蝸掃描電鏡觀察及全耳蝸基底膜鋪片熒光染色觀察。 結(jié)果:所有大鼠非導(dǎo)入耳ABR閾值導(dǎo)入前后無改變,正常大鼠導(dǎo)入耳ABR閾值較導(dǎo)入前提高約10dB SPL,致聾大鼠導(dǎo)入耳ABR閾值導(dǎo)入前后無改變;與導(dǎo)入前相比較,所有正常大鼠與致聾大鼠的導(dǎo)入耳沒有發(fā)現(xiàn)毛細(xì)胞增多或再生。結(jié)論:Hath1基因和DAPT聯(lián)合內(nèi)耳導(dǎo)入未能誘導(dǎo)藥物性聾大鼠耳蝸毛細(xì)胞再生,致聾大鼠毛細(xì)胞再生和聽力改善可能需要多基因聯(lián)合導(dǎo)入。
[Abstract]:Sensorineural hearing loss seriously affects human health and quality of life, but there is no effective treatment. Hair cell regeneration is the key to the treatment of sensorineural hearing loss.With the development of molecular biology, molecular genetics and genetic engineering technology, especially the research on gene regulation of hair cell regeneration in inner ear, it opens a new way for the treatment of sensorineural hearing loss.In this study, the animal model of drug-induced deafness was established, and the hair cell regeneration was induced by the combination of Hath1 gene and DAPT to induce hair cell regeneration, and the method of drug therapy for sensorineural hearing loss was explored.Part I observation of auditory function and hair cell damage in inner ear of rats after combined therapy of furosemide and kanamycin objective: to study the method of rapid establishment of drug induced deafness model in rats by combination of furosemide and kanamycin sulfate.Methods: rats in the experimental group were given furosemide intravenously and / or kanamycin sulfate intramuscularly.The threshold value of auditory brainstem evoked potential (ABAEP) brainstem response ABR was detected after 3 days and 7 days, and cochlea scanning electron microscope was used after 7 days of administration. Immunofluorescence staining of the whole cochlear basement membrane and frozen sections of the inner ear and HE staining of the frozen sections of the inner ear were observed.Results: after different doses of furosemide and kanamycin sulfate, the threshold value of ABR in rats increased in different degree, after 3 days and 7 days, there was no significant difference between the two groups after 2 months.Under scanning electron microscope, the hair cells of cochlea were damaged from the bottom to the apical gyrus, and from the outer hair cells to the inner hair cells.However, rats treated with furosemide or kanamycin sulfate had no threshold elevation or hair cell injury.In the rat cochlea, most of the Sertoli cells were intact and the vestibular hair cells were not damaged.Conclusion: furosemide and kanamycin sulfate have obvious ototoxic synergistic effects. Combined administration of kanamycin can cause severe impairment of auditory function in rats and no recovery of hearing in deaf rats after 2 months of administration, but the damage to cochlear Sertoli cells and vestibular hair cells is mild.The combination of furosemide and kanamycin is a simple, rapid and reliable method to establish a rat model of drug-induced deafness.Part two: preliminary study on the induction of hair cell regeneration by Hath1 gene and DAPT in inner ear objective: to investigate the expression of Hath1 in inner ear and the induction of hair cell regeneration in inner ear by Hath1 gene and DAPT.Methods: 15 normal rats and 15 deafness rats were exposed to auditory bubble via dorsal approach, and some of the auditory alveolar bone were removed. The Hath1 gene was introduced into the microinjector at the near round window.Then the capsule osmotic pump was continuously introduced into DAPT for 4 weeks.The threshold value of ABR, the scanning electron microscope of cochlea and the fluorescence staining of the whole cochlea basement membrane were observed.Results: there was no change in the threshold of ABR before and after the introduction of ABR in all rats. The threshold of ABR in normal rats was higher than that in the premise of introducing ABR. The threshold of ABR in deaf rats was not changed before and after the introduction of ABR, and compared with that before introduction.Hair cells were not found to increase or regenerate in all normal and deaf rats.Conclusion the combination of DAPT and 1% Hath1 gene can not induce cochlear hair cell regeneration in drug-induced deafness rats. The regeneration of hair cells and hearing improvement in deafness rats may require the combination of multiple gene transduction.
【學(xué)位授予單位】：福建醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2011
【分類號】：R764.35

【引證文獻(xiàn)】

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1 宋麗華;齊力;王敏;遲玉濤;許小敏;;Hath1基因真核表達(dá)載體的構(gòu)建及其在SH-SY5Y細(xì)胞中的表達(dá)[J];吉林大學(xué)學(xué)報(bào)(醫(yī)學(xué)版);2013年04期

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本文編號：1760307