【摘要】：目的探討鈣超載在Aβ25-35引起HT22細(xì)胞Per2表達(dá)變化中的作用。方法體內(nèi)研究將C57BL/6小鼠雙側(cè)海馬注射Aβ25-35(300μmol/kg)和無菌水后,利用跑輪行為學(xué)實驗評估小鼠的晝夜節(jié)律。體外研究將HT22細(xì)胞分為對照組、Aβ25-35處理組、Nim+Aβ25-35處理組。以MTT法檢測細(xì)胞存活率,實時熒光定量PCR檢測per2 mRNA表達(dá)水平,Western Blotting檢測PER2蛋白相對表達(dá)量,免疫熒光染色觀察PER2蛋白原位表達(dá)情況,流式細(xì)胞術(shù)檢測胞內(nèi)鈣離子濃度。結(jié)果體內(nèi)研究表明Aβ25-35導(dǎo)致C57BL/6小鼠在跑輪行為學(xué)實驗中表現(xiàn)出明顯的晝夜節(jié)律紊亂。離體研究發(fā)現(xiàn),5、10、15、20、25、30μmol/L Aβ25-35劑量組細(xì)胞存活率較對照組顯著降低(P0.05)。經(jīng)20μmol/L Aβ25-35處理后,per2 mRNA及PER2蛋白水平在CT16較對照組顯著降低(P0.05)。經(jīng)鈣離子拮抗劑尼莫地平預(yù)處理后,由Aβ25-35在CT16所致per2 mRNA及PER2蛋白水平的降低均顯著升高(P0.05)。結(jié)論鈣超載在Aβ25-35引起HT22細(xì)胞Per2異常表達(dá)中發(fā)揮重要作用。
[Abstract]:Objective to investigate the role of calcium overload in the changes of Per2 expression in HT22 cells induced by A 尾 25-35. Methods A 尾 25-35 (300 渭 mol/kg) and aseptic water were injected into the hippocampus of C57BL/6 mice in vivo. HT22 cells were divided into two groups in vitro: control group, A 尾 25-35 group, Nim A 尾 25-35 group. The cell survival rate was detected by MTT method, the per2 mRNA expression level was detected by real-time fluorescence quantitative PCR, the relative expression of PER2 protein was detected by, Western Blotting, the expression of PER2 protein in situ was observed by immunofluorescence staining, and the intracellular calcium concentration was detected by flow cytometry. Results in vivo studies showed that A 尾 25-35 resulted in obvious circadian rhythm disorder in C57BL/6 mice. In vitro study, the cell survival rate of the group at the dose of 10 ~ 10 ~ 15A 尾 25 ~ (35) 渭 mol/L A 尾 25 ~ 35 was significantly lower than that of the control group (P0.05). After treated with 20 渭 mol/L A 尾 25-35, the levels of per2 mRNA and PER2 protein in CT16 were significantly lower than those in control group (P0.05). Pretreatment with Nimodipine, a calcium antagonist, significantly increased the levels of per2 mRNA and PER2 protein in CT16 induced by A 尾 25-35 (P0.05). Conclusion calcium overload may play an important role in the abnormal expression of Per2 in HT22 cells induced by A 尾 25-35.
【作者單位】： 山西醫(yī)科大學(xué)基礎(chǔ)醫(yī)學(xué)院病理教研室;山西醫(yī)科大學(xué)基礎(chǔ)醫(yī)學(xué)院形態(tài)學(xué)實驗室;
【基金】：國家自然科學(xué)基金資助項目(81471343) 山西醫(yī)科大學(xué)科技創(chuàng)新基金資助項目(01201307)
【分類號】：R749.16
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本文編號：2290482