本文選題：HMGB1 + 氧化狀態(tài)��； 參考：《第二軍醫(yī)大學(xué)》2017年碩士論文

【摘要】：抑郁癥是一種常見的心理障礙,但發(fā)病機(jī)制尚未完全闡明�；诩�(xì)胞因子學(xué)說,我們對(duì)高遷移率蛋白族1(High-mobility group box 1,HMGB1)在抑郁癥發(fā)病中的作用及可能機(jī)制進(jìn)行研究。HMGB1作為晚期炎性介質(zhì),廣泛參與了自身免疫疾病的發(fā)病過程。最近的研究發(fā)現(xiàn)HMGB1在一些神經(jīng)炎性疾病中也有關(guān)鍵作用。我們的前期研究證實(shí),在腹腔注射低劑量細(xì)菌脂多糖(lipopolysaccharide,LPS)誘導(dǎo)的抑郁模型中,中樞HMGB1呈主動(dòng)釋放。HMGB1是一種非組蛋白染色體結(jié)合蛋白,細(xì)胞處于穩(wěn)態(tài)時(shí)主要位于細(xì)胞核;當(dāng)遭受到適當(dāng)?shù)男盘?hào)刺激時(shí),則經(jīng)過一定的化學(xué)修飾分泌到細(xì)胞外,這是HMGB1的主動(dòng)釋放。在此過程中,由于其所處的環(huán)境不同,其第23位、45位及106位半胱氨酸所連接的巰基會(huì)發(fā)生氧化形成三種不同形式的HMGB1:完全還原型HMGB1(fully reduced HMGB,fr-HMGB1),二硫鍵型HMGB1(disulfide HMGB1,ds-HMGB1),完全氧化型HMGB1(fully oxidized HMGB1,ox-HMGB1)。Fr-HMGB1主要與細(xì)胞膜表面的糖基化終產(chǎn)物受體(receptor for advanced glycation end-products,RAGE)及CXC趨化因子受體4(CXC-chemokine receptor 4,CXCR4)受體結(jié)合發(fā)揮趨化作用;ds-HMGB1主要與Toll樣受體家族(toll-like family of receptors,TLRs)部分成員結(jié)合促進(jìn)炎癥反應(yīng);ox-HMGB1無免疫活性。最近的研究表明,細(xì)胞外HMGB1的生物活性是由其氧化狀態(tài)決定的。本研究的目的主要在于明確HMGB1在抑郁癥發(fā)病中的作用,并在此基礎(chǔ)上探討導(dǎo)致抑郁的HMGB1的氧化狀態(tài)及受體機(jī)制。通過慢性不可預(yù)知應(yīng)激(chronic unpredictable mild stress,CUMS)構(gòu)建動(dòng)物模型,4周后進(jìn)行抑郁行為學(xué)檢測,包括懸尾實(shí)驗(yàn)(tail suspension test,TST)、糖水偏愛度檢測(sucrose preference test,SPT)及開場實(shí)驗(yàn)(open field test,OPT)。采用酶聯(lián)免疫吸附試驗(yàn)(enzyme-linked immunosorbent assay,ELISA)和蛋白質(zhì)免疫印跡(western blot,WB)的方法對(duì)外周和中樞HMGB1的蛋白含量進(jìn)行檢測。對(duì)導(dǎo)致抑郁的HMGB1的氧化狀態(tài)及機(jī)制進(jìn)行研究的部分,采用側(cè)腦室注射的方法,將三種商品化的重組HMGB1:ds-HMGB1,fr-HMGB1及非氧化型HMGB1(non-oxidizable chemokine-HMGB1,nonoxid-HMGB1)分別給予小鼠進(jìn)行研究。與fr-HMGB1相比,nonoxid-HMGB1也具有趨化作用,但在體內(nèi)不會(huì)發(fā)生氧化狀態(tài)的改變。通過中樞將這三種HMGB1以200ng/5μL/只的劑量給予小鼠,20h后對(duì)小鼠進(jìn)行抑郁行為學(xué)檢測;為了進(jìn)一步研究其致抑郁行為的受體機(jī)制,在給予HMGB1之前半小時(shí)分別用相應(yīng)的受體拮抗劑TLR4/RAGE/CXCR4(TAK-242/FPS-ZM1/AMD3100)(3mg/kg)進(jìn)行拮抗,檢測其對(duì)小鼠抑郁行為的影響。最后,通過測定中樞海馬炎性因子腫瘤壞死因子-α(Tumor necrosis factor-α,TNF-α)和髓鞘形成相關(guān)蛋白-髓鞘堿性蛋白(myelin basic protein,MBP)的表達(dá)初步探究HMGB1與受體結(jié)合后致抑郁的下游機(jī)制。我們的研究表明:在CUMS構(gòu)建的抑郁模型中,與對(duì)照組相比,CUMS組外周及中樞HMGB1的表達(dá)均顯著增高。中樞分別給予ds-HMGB1與fr-HMGB1均可誘導(dǎo)小鼠出現(xiàn)抑郁樣行為;相應(yīng)的受體拮抗劑能改善抑郁樣行為。TAK-242也可以改善fr-HMGB1誘導(dǎo)的抑郁樣行為。而non-oxidHMGB1不能誘導(dǎo)小鼠出現(xiàn)抑郁樣行為。經(jīng)檢測兩種氧化狀態(tài)的HMGB1均能引起TNF-α表達(dá)的增加,且與抑郁行為學(xué)指標(biāo)相一致。在進(jìn)一步檢測中,我們發(fā)現(xiàn)中樞給予ds-HMGB1可以引起小鼠海馬組織MBP表達(dá)量的降低;而TLR4拮抗劑對(duì)ds-HMGB1引起的效應(yīng)具有MBP保護(hù)作用。綜上所述:兩種氧化狀態(tài)HMGB1在抑郁癥的發(fā)病中起重要作用,這種作用可能是通過激活炎癥通路誘導(dǎo)中樞炎癥,進(jìn)而導(dǎo)致髓鞘損傷引起的。
[Abstract]:Depression is a common mental disorder, but the pathogenesis has not been fully elucidated. Based on the cytokine theory, we study the role and possible mechanism of high mobility protein family 1 (High-mobility group box 1, HMGB1) in the pathogenesis of depression..HMGB1 is a late inflammatory medium and is widely involved in the pathogenesis of autoimmune diseases. Recent studies have found that HMGB1 also plays a key role in some neuroinflammatory diseases. Our previous study confirmed that in the depressive model induced by low dose lipopolysaccharide (LPS) intraperitoneal injection of bacterial lipopolysaccharide (LPS), central HMGB1 actively releases.HMGB1 as a non histone chromosome binding protein, the main position of the cell at the time of homeostasis. In the nucleus; when a proper signal is stimulated, a certain chemical modification is secreted to the extracellular, which is the active release of HMGB1. In this process, the sulfhydryl groups connected by its twenty-third, 45 and 106 - bit cysteine will be oxygenation to form three different forms of HMGB1: completely prototype HMGB1 (f Ully reduced HMGB, fr-HMGB1), two sulfur bond HMGB1 (disulfide HMGB1, ds-HMGB1), completely oxidized HMGB1 (fully oxidized HMGB1), mainly with the receptor of the cell membrane surface, and chemokine receptor 4 receptor. In combination with chemotaxis, ds-HMGB1 is mainly associated with some members of the Toll like receptor family (Toll-like family of receptors, TLRs) to promote inflammatory response; ox-HMGB1 has no immune activity. Recent studies have shown that the biological activity of extracellular HMGB1 is determined by its oxidation state. The purpose of this study is to identify HMGB1 in the pathogenesis of depression. On this basis, we explored the oxidative status and receptor mechanism of HMGB1 causing depression. The animal model was constructed by chronic unpredictable stress (chronic unpredictable mild stress, CUMS), and the depressive behavior was detected after 4 weeks, including the tail suspension test (tail suspension test, TST), sugar water preference test (sucrose preference) EST, SPT) and the opening experiment (open field test, OPT). Enzyme linked immunosorbent assay (enzyme-linked immunosorbent assay, ELISA) and protein immunoblotting (Western blot) were used to detect the protein content of the peripheral and central nervous systems. The part of the study of the oxidative state and mechanism of depression, using the lateral brain Three commercialized recombinant HMGB1:ds-HMGB1, fr-HMGB1 and non oxidative HMGB1 (non-oxidizable chemokine-HMGB1, nonoxid-HMGB1) were given to mice respectively. Compared with fr-HMGB1, nonoxid-HMGB1 also had chemotaxis, but there was no change in the oxidative state in the body. The three HMGB1 were 20 in the center. The dose of 0ng/5 mu L/ was given only to mice, and the mice were tested for depressive behavior after 20h. In order to further study the receptor mechanism of their depressive behavior, the corresponding receptor antagonist TLR4/RAGE/CXCR4 (TAK-242/FPS-ZM1/AMD3100) (3mg/kg) was used to detect the effect of the receptor antagonist on the depressive behavior of the mice at half an hour before the HMGB1. Later, through the determination of the expression of Tumor necrosis factor- alpha (TNF- alpha) and myelin formation related protein myelin basic protein (myelin basic protein, MBP) in the central hippocampal inflammatory factors, the downstream mechanism of depression after the combination of HMGB1 and receptor is preliminarily explored. Our study showed that in the depressive model constructed by CUMS, and the control The expression of peripheral and central HMGB1 in the group CUMS was significantly higher than that in the group. Both ds-HMGB1 and fr-HMGB1 could induce depressive behavior in mice, and the corresponding receptor antagonists could improve the depressive behavior.TAK-242 and improve the depressive behavior induced by fr-HMGB1, while non-oxidHMGB1 could not induce depressive behavior in mice. HMGB1, which was detected in two oxidation states, could cause an increase in the expression of TNF- alpha and was in accordance with the depressive behavioral index. In further detection, we found that the central administration of ds-HMGB1 could cause a decrease in the expression of MBP in the hippocampus of the mice; and the TLR4 antagonist had the effect of MBP protection on the effect of ds-HMGB1. To sum up, two kinds of oxygen are described. The chemical state HMGB1 plays an important role in the pathogenesis of depression, which may be caused by the activation of the inflammatory pathway to induce central inflammation, which may lead to myelin injury.

【學(xué)位授予單位】：第二軍醫(yī)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R749.4

【參考文獻(xiàn)】

相關(guān)期刊論文 前1條

1 趙勇;張璐;蔡麗萍;徐晨;;小鼠側(cè)腦室埋管給藥方法的優(yōu)化[J];實(shí)驗(yàn)動(dòng)物與比較醫(yī)學(xué);2014年02期

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本文編號(hào)：1840982