本文關(guān)鍵詞：補益論對形覺剝奪性弱視大鼠視皮質(zhì)神經(jīng)元中c-fos蛋白表達的研究　出處：《長春中醫(yī)藥大學》2015年碩士論文　論文類型：學位論文

  更多相關(guān)文章： 單眼形覺剝奪性弱視大鼠 針刺 c-fos蛋白

【摘要】：目的:通過P-VEP檢測驗證針刺治療弱視的有效性,免疫熒光染色技術(shù)觀察單眼形覺剝奪性弱視大鼠和正常發(fā)育大鼠視皮質(zhì)神經(jīng)元中c-fos蛋白的表達情況,探討在視覺發(fā)育可塑期內(nèi),正常大鼠及單眼剝奪性弱視大鼠視皮質(zhì)17區(qū)功能形態(tài)的變化規(guī)律。從分子水平來探索補益論對弱視治療的效果。為針刺治療弱視提供理論基礎(chǔ),在臨床預防與治療弱視提供新的思路。方法:實驗選取剛出生14天,健康wistar大鼠18只,雌雄不限(由吉林大學動物中心提供,清潔動物)。將此18只大鼠隨機分為正常組(NC)、單眼形覺剝奪模型組(MD1)、單眼形覺剝奪針刺組(MD2),各6只。根據(jù)大鼠視覺發(fā)育可塑期為出生后14d~45d為依據(jù)[22],在大鼠14d時,NC組不給予任何處理,MD1與MD2組行單眼眼瞼縫合術(shù),建立單眼形覺剝奪弱視模型。術(shù)后每天早晚檢查眼瞼創(chuàng)口情況,檢查縫線是否有脫落,每天滴抗生素眼液、涂紅霉素眼膏以預防感染。如縫線抓脫或傷口感染導致漏光或眼瞼閉合不嚴密的動物棄用。形覺剝奪性弱視動物模型的標準為術(shù)眼上下瞼粘連。各組幼鼠均在同等環(huán)境下散放喂養(yǎng)。單眼形覺剝奪針刺治療組在術(shù)后第7天開始給予針刺治療,單眼形覺剝奪模型組只進行單眼剝奪縫合術(shù),正常組不給予任何處置。針刺治療組選取“睛明穴”、“攢竹穴”、“腎俞穴”、“脾俞穴”等4個穴位。睛明穴為眼內(nèi)角直刺0.3cm,攢竹穴斜刺0.5cm,脾俞穴、腎俞穴直刺0.6cm。采用平補平瀉手法,1次/1d進行治療,1次/15min,治療周期為7天。檢測治療后各組幼鼠的P-VEP的變化。MD組大鼠心臟灌注成功后分別切取MD1、MD2單眼形覺剝奪弱視眼對側(cè)視皮層(VC)的視皮質(zhì)17區(qū),NC組切取任意一側(cè)視皮質(zhì)17區(qū)。所得視皮質(zhì)17區(qū)標本均浸入4%多聚甲醛磷酸鹽緩沖液中,放入4℃冰箱里后固定24h以上。經(jīng)石蠟切片后行免疫熒光染色技術(shù)觀察各組視皮質(zhì)17區(qū)組織的c-fos蛋白表達的情況。結(jié)果:1.正常組P-VEP檢測結(jié)果波峰明顯、穩(wěn)定性好、正常變異小。MD1與MD2相比P100潛伏期明顯延長,振幅降低。MD1與NC、MD2比較,其P-VEP的振幅明顯降低,潛伏期延長,波形不穩(wěn)定,并且經(jīng)統(tǒng)計學分析兩組差異有統(tǒng)計學意義(P0.05);MD2的P-VEP檢測結(jié)果與MD1結(jié)果比較,其波形較穩(wěn)定,潛伏期縮短、振幅提高,經(jīng)統(tǒng)計學分析,兩組之間P100潛伏期與P100振幅比較差異有統(tǒng)計學意義(P0.05)。2.免疫熒光染色結(jié)果:針刺治療結(jié)束后,在光鏡下觀察MD2大鼠視皮質(zhì)17區(qū)內(nèi)可見到c-fos陽性表達神經(jīng)元分布散在、均勻,著色顯強熒光;MD1視皮質(zhì)17區(qū)內(nèi)c-fos免疫陽性神經(jīng)元密度及陽性細胞熒光數(shù)量都明顯低于NC與MD2,經(jīng)統(tǒng)計學分析,此兩組差異有統(tǒng)計學意義(P0.05)。MD1與MD2大鼠視皮質(zhì)17區(qū)內(nèi)c-fos免疫陽性神經(jīng)元密度及陽性細胞熒光數(shù)量比較,兩組差異有統(tǒng)計學意義(P0.05)。結(jié)論:1.視覺發(fā)育敏感期內(nèi)補益論對單眼形覺剝奪大鼠的P-VEP波形改變具有明顯的改善作用。2.視覺發(fā)育關(guān)鍵期內(nèi)補益論對單眼剝奪弱視大鼠治療有明顯的效果。3.補益論對單眼剝奪大鼠視皮質(zhì)17區(qū)內(nèi)組織中c-fos蛋白表達變化,提示針刺對弱視視功能改善的機制可能是通過c-fos蛋白表達的改變而實現(xiàn)的,c-fos參與了視覺發(fā)育及視覺發(fā)育可塑性調(diào)節(jié),影響了大鼠視覺系統(tǒng)的可塑性變化。
[Abstract]:Objective: to verify the validity of P-VEP detection by acupuncture treatment of amblyopia, immunofluorescence staining to observe the expression of monocular visual deprivation amblyopia rats and the normal development of rat visual cortical neurons in c-fos protein, investigate in rat visual development period, changes in normal rats and single eye amblyopia rat visual cortex the 17 district function form. From the molecular level to explore the benefit on the effect of treatment of amblyopia. Provide a theoretical basis for acupuncture treatment of amblyopia, and provide a new way of thinking in the clinical prevention and treatment of amblyopia. Methods: this experiment was born just 14 days, 18 healthy Wistar rats, male and female (provided by the animal center of the Jilin University clean. This animal) 18 rats were randomly divided into normal group (NC), monocular deprivation model group (MD1), monocular deprivation acupuncture group (MD2), 6 rats in each rat. According to the visual development period after birth can be plastic 14d~45d According to [22], in 14d rats, NC group were not given any treatment, MD1 and MD2 group underwent monocular lid suture, establish the model of monocular deprivation amblyopia. Postoperative eyelid wound every day sooner or later check, check whether there is a suture off, every drop of antibiotic eye drops, with Erythromycin Eye Ointment such as suture grasping to prevent infection. Removal or wound infection caused leaked or lagophthalmus strict animal abandoned. Form deprivation amblyopia animal model standard for eye surgery on lower eyelid adhesions. The pups were in the same environment scattered feeding. Monocular deprivation acupuncture treatment group in seventh days after surgery was given acupuncture treatment, monocular deprivation the model group only monocular deprivation suture, the normal group was not given any treatment. Acupuncture group selected "Jingming", "zanzhu acupoint", "Shenshu", "4 points Pishu points. Jingming eye angle into 0.3cm, Acupuncture cuanzhu acupoint 0.5cm, spleen Shu point, Shenshu Point into the 0.6cm. the use of reinforcing reducing techniques, 1 /1d for 1 /15min, the treatment period is 7 days. After the treatment in different group P-VEP changes group.MD perfused rat heart were harvested after successful MD1 deprivation amblyopia on the side MD2 monocular cortex (VC) of the visual cortex area 17, NC group cut from any side of the visual cortex area 17. The visual cortex area 17 specimens were immersed in 4% paraformaldehyde phosphate buffer, into the refrigerator after 4 degrees Celsius above 24h. The fixed paraffin sections after immunofluorescence staining to observe the expression of each group the visual cortex area 17 tissue c-fos protein. Results: 1. normal results of P-VEP wave obviously, good stability, small variation in.MD1 compared with MD2 normal P100 latency was prolonged and the amplitude decreased.MD1 and NC, MD2, P-VEP significantly decreased the amplitude, latency prolonged, waveform Stable, and statistically significant differences between the two groups (P0.05); the results of P-VEP MD2 compared with MD1 results, the waveform is stable, the latency and amplitude increase, by statistical analysis, the comparison between the two groups P100 latency and amplitude of P100 were statistically significant (P0.05).2. immunofluorescence staining results: Acupuncture Treatment after MD2 rats was observed under light microscope in the visual cortex area 17 can be seen in the positive expression of c-fos neurons in the distribution, uniform coloring was strong fluorescence MD1; visual cortex area 17 the number of fluorescent c-fos immunoreactive neurons and positive cell density were significantly lower than those of NC and MD2, statistical analysis, statistical the significance of the difference between the two groups (P0.05) in visual cortex area 17 fluorescence quantitative comparison of c-fos immunoreactive neurons and density of.MD1 positive cells and MD2 rats, there was significant difference between two groups (P0.05). Conclusion: 1. visual development sensitive During the period of tonic on monocular visual deprivation P-VEP waveform changes of rats can significantly improve the effect of.2. on the critical period of visual development of tonic monocular deprivation amblyopia rats treatment has obvious effect on.3. and on the expression of c-fos protein in monocularly deprived rats visual cortex area 17 in the organization change, as the mechanism of acupuncture to improve the function of amblyopia may be through changes in the expression of c-fos was achieved, c-fos is involved in the visual development and visual plasticity regulation effect of plasticity of the visual system of rats.

【學位授予單位】：長春中醫(yī)藥大學
【學位級別】：碩士
【學位授予年份】：2015
【分類號】：R777.44

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