本文選題：腎臟　切入點(diǎn)：晝夜節(jié)律　出處：《北京協(xié)和醫(yī)學(xué)院》2015年博士論文

【摘要】：[研究背景]腎臟是具有節(jié)律的器官,參與調(diào)控血壓及水鹽排泄等生理晝夜節(jié)律的形成。臨床上慢性腎臟病患者表現(xiàn)出多種節(jié)律紊亂如血壓節(jié)律紊亂、睡眠節(jié)律紊亂、不寧腿綜合征等。雖其發(fā)生機(jī)制并不明確,但這些節(jié)律紊亂部分在腎移植術(shù)后得到改善。越來(lái)越多的證據(jù)表明,外周組織臟器本身的生物鐘系統(tǒng)參與生理節(jié)律的形成、穩(wěn)定與調(diào)節(jié)。我們前期研究已經(jīng)證實(shí)阿霉素大鼠表現(xiàn)腎病綜合征的同時(shí)呈現(xiàn)出鹽敏高血壓和血壓晝夜節(jié)律紊亂,且協(xié)同表現(xiàn)為尿鈉排泄晝夜節(jié)律的紊亂。本研究進(jìn)一步嘗試探究慢性腎臟病大鼠出現(xiàn)的血壓和尿鈉晝夜節(jié)律紊亂的時(shí)間生物學(xué)機(jī)制,探討腎臟本身固有的生物鐘基因和腎臟特有鐘控基因如鈉離子轉(zhuǎn)運(yùn)相關(guān)基因在慢性腎臟病中的節(jié)律變化及其意義。[研究方法]經(jīng)SD大鼠尾靜脈注射阿霉素建立腎病綜合征大鼠模型為腎病組,對(duì)照組為注射同體積生理鹽水的SD大鼠。適應(yīng)性飼養(yǎng)2周,嚴(yán)格執(zhí)行開(kāi)關(guān)燈時(shí)間,早8：00開(kāi)照明燈(記為ZT00：00或CT08：00),晚20：00關(guān)燈(記為ZT12：00或CT20：00)。ZT00:00-ZT12:00定義為日間(Light Period), ZT12:00-24:00定義為夜間(Dark Period)。分別收集日夜尿,記錄日夜尿量,分測(cè)日夜尿鈉、尿鉀、尿氯的濃度,計(jì)算日夜間尿量、尿鈉、尿鉀和尿氯的排泄率。分別于CT10am,2pm,6pm,10pm, 2am,6am處死對(duì)照組和腎病組大鼠各3只,分別取大鼠腎組織、血漿標(biāo)本,應(yīng)用實(shí)時(shí)定量PCR方法檢測(cè)腎組織鐘基因CLOCK、BMAL1、Per1、Per2、Cryl、 Cry2及鐘控基因NHE3、aENaC、NCC、Ptges、VlaR、V2R mRNA表達(dá)；測(cè)定6個(gè)時(shí)間點(diǎn)大鼠血漿腎素活性、醛固酮和血漿血管緊張素II水平。應(yīng)用部分傅里葉分析合并逐步回歸的分析方法對(duì)以上數(shù)據(jù)進(jìn)行節(jié)律性分析。[研究結(jié)果]1、對(duì)照組正常大鼠尿量、尿鈉及尿鉀排泄率呈夜高日低的趨勢(shì),腎臟組織內(nèi)鐘基因CLOCK、BMAL1、Per1、 Per2、Cry1、Cry2mRNA均表現(xiàn)晝夜節(jié)律(p0.05)及鐘基因下游與水鹽重吸收和排泄相關(guān)的鐘控基因NHE3、aENaC、NCC、 Ptges、VlaR、V2R mRNA也表現(xiàn)明顯晝夜節(jié)律(p0.05),各基因峰值均處于大鼠夜間活動(dòng)期,與大鼠腎臟水鹽排泄的夜高日低的節(jié)律一致。2、腎病組大鼠尿鈉排泄呈日高夜低的節(jié)律(p0.05)。腎臟組織內(nèi)鐘基因BMAL1表達(dá)呈24小時(shí)和12小時(shí)節(jié)律,鐘基因Cry1表達(dá)呈4.8小時(shí)節(jié)律,鐘控基因NCC、V2R表達(dá)均呈12小時(shí)節(jié)律,上述基因晝夜節(jié)律峰值均移至日間大鼠休息期(p0.05)。而鐘基因CLOCK、Per1、Per2、Cry2及鐘控基因NHE3、aENaC Ptges、 VlaR表達(dá)晝夜節(jié)律均消失(p0.05)。3、對(duì)照組SD大鼠外周血漿腎素活性水平呈12小時(shí)節(jié)律(p0.05),血漿醛固酮濃度水平呈24小時(shí)節(jié)律(p0.05),血漿血管緊張素II濃度水平無(wú)節(jié)律(p0.05)；腎病組大鼠血漿腎素活性及醛固酮濃度節(jié)律消失(p0.05),而血漿血管緊張素II濃度仍無(wú)節(jié)律(p0.05)。[研究結(jié)論]正常大鼠腎臟固有生物鐘基因以及腎臟鐘控基因表達(dá)的晝夜節(jié)律穩(wěn)定而保守,與正常大鼠尿鈉排泄節(jié)律具有時(shí)相協(xié)同性。阿霉素腎病大鼠表現(xiàn)為尿鈉排泄的晝夜節(jié)律倒置,其腎臟局部的鐘基因表達(dá)節(jié)律紊亂,且鐘基因下游與水鹽重吸收和排泄相關(guān)的鐘控基因NHE3, aENaC, NCC, Ptges, VlaR, V2R mRNA的表達(dá)時(shí)相和節(jié)律特點(diǎn)均發(fā)生改變。這是首次證實(shí)在慢性腎病狀態(tài)下,大鼠腎臟鐘基因系統(tǒng)紊亂,與其血壓和尿鈉晝夜節(jié)律紊亂偶聯(lián)出現(xiàn),表明腎臟生物鐘分子系統(tǒng)在腎臟正常生理功能的晝夜節(jié)律中具有重要作用。
[Abstract]:[background] with the rhythm of the kidney is the organ formation, involved in the regulation of blood pressure and the excretion of salt water and other physiological circadian rhythm in patients with chronic kidney disease. The clinical manifestations of a variety of disorders such as blood pressure circadian rhythm disorder, sleep rhythm disorder, restless legs syndrome. Although its pathogenesis is not clear, but the rhythm section improvement in renal transplantation. More and more evidence that the formation of biological clock peripheral organs themselves participate in physiological rhythm, stability and regulation. Our previous studies have demonstrated that rats with adriamycin and nephrotic syndrome showed salt sensitive hypertension and blood pressure circadian rhythm disorder, and collaborative performance for urinary sodium the excretion of circadian rhythm disorders. This study further attempts to explore the biological mechanism of time chronic kidney disease rats blood pressure and urinary sodium circadian rhythm disorder, the body of the kidney The intrinsic circadian clock genes and kidney specific clock controlled genes such as sodium transport related genes rhythm changes in chronic kidney disease and its significance. Methods] SD rat tail vein injection of adriamycin nephrotic syndrome rat model for nephropathy group, the control group was injected with the same volume of saline. The adaptability of SD rats 2 weeks of feeding, the strict implementation of switch time, 8:00 in the morning lights (ZT00:00 or CT08:00), 20:00 at night lights (ZT12:00 or CT20:00).ZT00:00-ZT12:00 (Light Period) is defined as the day, ZT12:00-24:00 is defined as the night (Dark Period) were collected and recorded. Urine, day and night urine volume, urine test is divided into day and night the sodium chloride concentration, urine potassium, urine, urine sodium, urine volume calculation at night, urine and urine excretion rate of potassium chloride. In CT10am, 2pm, 6pm, 10pm, 2am, 6am were control group and nephritic rats 3 rats in each group were taken from rat kidney Fabric, plasma specimens, detection of clock gene CLOCK, renal tissue using real-time quantitative PCR method for BMAL1, Per1, Per2, Cryl, Cry2 and aENaC, clock controlled genes NHE3, NCC, Ptges, VlaR, V2R mRNA expression; Determination of 6 time points in rat plasma renin activity, and plasma aldosterone angiotensin Zhang Su II level. The application of Fourier analysis analysis combined with stepwise regression analysis. The results of the rhythm of]1 on the above data, the control group of normal rats urine volume, urinary sodium and urinary potassium excretion rate was on the night high low trend, kidney tissue clock gene CLOCK, BMAL1, Per1, Per2, Cry1, Cry2mRNA all day and night rhythm (P0.05) and clock genes downstream and water salt reabsorption and excretion of clock controlled genes NHE3, aENaC, NCC, Ptges, VlaR, V2R and mRNA also showed obvious circadian rhythm (P0.05), the gene in rat nocturnal peak period, and the kidney of rats with salt and water excretion, night high low the Same rhythm.2 nephritic rats urinary sodium excretion was high and low night rhythm (P0.05). Renal tissue in clock gene expression of BMAL1 was 24 hours and 12 hours of rhythm, clock gene Cry1 expression was 4.8 hour rhythm, clock controlled genes NCC, V2R expression showed a 12 hour rhythm, the peak circadian gene all rats to day rest period (P0.05). And the clock genes CLOCK, Per1, Per2, Cry2 and aENaC Ptges clock controlled genes NHE3, VlaR, the expression of circadian rhythm disappeared (P0.05.3), the control group of SD rats peripheral levels of plasma renin activity was 12 hour rhythm (P0.05), plasma aldosterone concentration level a 24 hour rhythm (P0.05), plasma angiotensin II concentration level rhythm (P0.05); nephropathy rats plasma renin activity and aldosterone (P0.05), and the rhythm of plasma angiotensin II concentration was no rhythm (P0.05). The research conclusion of normal rat kidney inherent biological Zhong Ji Because the clock controlled gene expression in the kidney and the circadian rhythm of stable and conservative, and the excretion of urine sodium in rats with normal circadian phase. Sometimes cooperativity in adriamycin induced nephropathy rats showed circadian inversion of urinary sodium excretion, renal local clock gene expression rhythm disorder, and clock genes downstream and water salt reabsorption and excretion the clock controlled genes NHE3, aENaC, NCC, Ptges, VlaR, mRNA and V2R expression rhythm are changed. This is confirmed for the first time in the state of chronic kidney disease, rat kidney system clock gene disorders, their blood pressure and urine sodium circadian rhythm disorder coupling, indicate the important role of circadian rhythm in the kidney the normal physiological function of the kidney biological clock in molecular systems.

【學(xué)位授予單位】：北京協(xié)和醫(yī)學(xué)院
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2015
【分類(lèi)號(hào)】：R692

【共引文獻(xiàn)】

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