本文關(guān)鍵詞：NogoA-PirB通路對(duì)腦出血大鼠軸突再生和神經(jīng)功能的影響　出處：《西南醫(yī)科大學(xué)》2017年碩士論文　論文類型：學(xué)位論文

  更多相關(guān)文章： 腦出血 PirB NogoA 軸突再生 神經(jīng)元細(xì)胞 星形膠質(zhì)細(xì)胞

【摘要】：目的:通過構(gòu)建大鼠腦出血模型,觀察NogoA和Pir B兩種蛋白在腦出血大鼠腦組織中不同時(shí)間點(diǎn)的定量表達(dá)、PirB表達(dá)的時(shí)空特點(diǎn)、軸突再生情況、大鼠神經(jīng)功能缺損情況,探討PirB的表達(dá)對(duì)腦出血損傷大鼠軸突再生和神經(jīng)功能的影響、NogoA和PirB蛋白表達(dá)的相關(guān)性,推測(cè)Nogo A-PirB通路與神經(jīng)功能缺損的相關(guān)性,探討腦出血損傷后腦組織的修復(fù)機(jī)制,進(jìn)一步探索腦出血后治療靶點(diǎn),為臨床治療研究提供理論依據(jù)。方法:雄性SD大鼠75只,隨機(jī)分為假手術(shù)(Sham)組(n=25)和腦出血(ICH)組(n=50),兩組分別按斷頭取腦時(shí)間點(diǎn)不同分為1d、3d、7d、14d、28d共5個(gè)亞組,各時(shí)間點(diǎn)腦出血大鼠隨機(jī)分為Western Blot組和組織免疫熒光組。Sham組大鼠顱骨鉆孔后不注射血液,腦出血組以大鼠自體鼠尾動(dòng)脈不凝血50ul,經(jīng)改良二次注入法建立大鼠腦出血模型,術(shù)后3小時(shí)通過神經(jīng)功能評(píng)分判定造模成功與否。造模成功后,各時(shí)間點(diǎn)大鼠采用Garcia神經(jīng)功能障礙評(píng)分方法進(jìn)行評(píng)分后斷頭取腦。Western Blot組腦組織取出后立即液氮冷凍,隨后轉(zhuǎn)入-80℃冰箱凍存?zhèn)溆?研磨組織提取蛋白后,采用蛋白免疫印跡法(Western-blot)測(cè)定PirB和NogoA蛋白的表達(dá);組織免疫熒光組大鼠深度麻醉后予以PBS和4%多聚甲醛灌注后取腦,取出腦組織經(jīng)后續(xù)處理后行石蠟切片和冰凍切片,分別行HE染色和免疫熒光。結(jié)果:(1)、腦出血組神經(jīng)功能缺失評(píng)分明顯低于對(duì)應(yīng)時(shí)間點(diǎn)Sham組,以3d組神經(jīng)功能缺失評(píng)分最低,以上結(jié)果統(tǒng)計(jì)學(xué)差異具有顯著性(P0.05)。(2)腦組織形態(tài)結(jié)構(gòu)改變:Sham組腦組織中可見神經(jīng)細(xì)胞完整。腦出血組1d亞組病灶內(nèi)充滿紅細(xì)胞,病灶周圍水腫明顯伴少量炎細(xì)胞浸潤(rùn);3d亞組血腫紅細(xì)胞減少,病灶周圍神經(jīng)元腫脹壞死明顯,大量炎細(xì)胞浸潤(rùn);7d亞組血腫進(jìn)一步吸收,周圍水腫有所減輕,炎細(xì)胞減少,膠質(zhì)細(xì)胞大量增生;14d、28d亞組血腫吸收,見明顯膠原纖維增生及修復(fù)瘢痕。(3)NogoA和PirB定量分析:腦出血組各時(shí)間點(diǎn)NogoA、PirB表達(dá)均高于對(duì)應(yīng)時(shí)間點(diǎn)的Sham組;二者在腦出血后表達(dá)明顯增加,并在腦出血后7天達(dá)到表達(dá)高峰,持續(xù)高水平表達(dá)至28d,差異具有統(tǒng)計(jì)學(xué)意義(P0.05)。(4)、雙標(biāo)免疫熒光:PirB在腦出血組各時(shí)間點(diǎn)均有表達(dá),主要表達(dá)在神經(jīng)元細(xì)胞、軸突及星形膠質(zhì)細(xì)胞;腦出血組出現(xiàn)明顯軸突斷裂、紊亂等損傷表現(xiàn),軸突長(zhǎng)度較正常對(duì)照組明顯縮短,持續(xù)至腦出血后28天無明顯改善,與Sham組差異具有統(tǒng)計(jì)學(xué)意義(P0.05)。(5)、腦出血組各時(shí)間點(diǎn)NogoA、PirB表達(dá)與神經(jīng)功能評(píng)分和軸突長(zhǎng)度呈負(fù)相關(guān)。結(jié)論:(1)正常成年大鼠腦組織中可見少量NogoA、PirB表達(dá),主要表達(dá)于神經(jīng)元及星形膠質(zhì)細(xì)胞,維持神經(jīng)功能的穩(wěn)定。(2)腦出血可誘導(dǎo)NogoA、PirB表達(dá)增加,NogoA-PirB通路可抑制軸突再生,阻礙腦出血后的神經(jīng)功能恢復(fù)。
[Abstract]:Objective: to construct a rat model of intracerebral hemorrhage, the expression of NogoA and Pir were two B in the quantitative protein at different time points in rats with intracerebral hemorrhage, spatial and temporal characteristics of PirB expression, axonal regeneration, the neurological deficit in rats, to explore the influence of the injured axon regeneration and neural function of rats with cerebral hemorrhage. The expression of PirB, NogoA and PirB protein expression, speculated that the correlation between Nogo A-PirB pathway and neural function defect, explore the repair mechanism of the injured brain tissue in cerebral hemorrhage, cerebral hemorrhage and further explore the therapeutic targets, provide a theoretical basis for clinical treatment. Methods: 75 male SD rats were randomly divided into sham operation (Sham) group (n=25) and cerebral hemorrhage (ICH) group (n=50), two groups were decapitated at different time points of 1D, 3D, 7d, 14d, 28d a total of 5 sub groups, each time point in cerebral hemorrhage rats were randomly divided into Western group and Blot group of immune The fluorescent group.Sham rats after injection of blood drilling skull, cerebral hemorrhage group with rat tail artery blood autologous 50ul modified two injection method to establish the rat model of intracerebral hemorrhage, 3 hours after the operation to determine the model is successful or not by the neurological score. After the success of the model, each time point rats with Garcia nerve dysfunction score method were decapitated after.Western Blot in brain tissue were removed immediately after freezing in liquid nitrogen, then transferred to the -80 C refrigerator and cryopreserved, grinding tissue protein extraction, by Western blotting (Western-blot) to detect the expression of PirB and NogoA protein; immunohistochemical rats after deep anesthesia to be PBS and 4% paraformaldehyde perfusion after brain, brain tissue removed by subsequent treatment after paraffin and frozen sections were stained by HE staining and immunofluorescence. Results: (1), cerebral hemorrhage group nerve function lack of clear evaluation Significantly lower than that of the corresponding time points in Sham group, 3D group with neurological deficit score was the lowest, the above results have significant difference (P0.05). (2) morphological changes of brain tissue, brain tissue Sham was found in nerve cells. The complete group of 1D subgroup of lesions filled with red blood cells of cerebral hemorrhage, edema around the lesion with obvious a small amount of inflammatory cell infiltration; 3D subgroup hematoma decreased red blood cells around the lesion, swelling of neuron necrosis, inflammatory cell infiltration; 7d subgroup hematoma further absorption, peripheral edema reduced, inflammatory cells decreased, glial cell proliferation; 14d, absorption of 28d subgroup of hematoma, obvious hyperplasia of collagen fiber and scar repair (. 3) PirB: quantitative analysis of NogoA and NogoA group at different time points of cerebral hemorrhage, the expression of PirB was higher than that of Sham group at corresponding time points; the two was significantly increased in cerebral hemorrhage after expression, and the expression peak in up to 7 days after cerebral hemorrhage, the sustained high level table Up to 28d, the difference was statistically significant (P0.05). (4), double immunofluorescence: the expression of PirB in the group at each time point were cerebral hemorrhage, mainly expressed in neurons, axons and astrocytes; cerebral hemorrhage group showed axonal fracture, disorder of injury, axonal length was significantly shortened and until 28 days after cerebral hemorrhage had no obvious improvement, with statistically significant difference with Sham group (P0.05). (5), NogoA group at different time points of cerebral hemorrhage, the expression of PirB was negatively correlated with the score of nerve function and axon length. Conclusion: (1) in brain tissue of rats with a small amount of NogoA, normal adult the expression of PirB was mainly expressed in neurons and astrocytes, nerve function to maintain stability. (2) NogoA can induce cerebral hemorrhage, increase the expression of PirB, NogoA-PirB pathway can inhibit axon regeneration, hindrance to God after intracerebral hemorrhage by function recovery.

【學(xué)位授予單位】：西南醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R743.34

【參考文獻(xiàn)】

相關(guān)期刊論文 前1條

1 周迎春;遷榮軍;饒競(jìng);翁密霞;易序霞;;Expression of PirB in Normal and Injured Spinal Cord of Rats[J];Journal of Huazhong University of Science and Technology(Medical Sciences);2010年04期

，

本文編號(hào)：1421065