本文關(guān)鍵詞：一氧化氮對(duì)懸浮紅細(xì)胞保存質(zhì)量影響及糖酵解代謝關(guān)系的研究　出處：《北京協(xié)和醫(yī)學(xué)院》2015年碩士論文　論文類型：學(xué)位論文

  更多相關(guān)文章： 硝普鈉 L-精氨酸 一氧化氮 密度梯度離心 D.L甘油醛

【摘要】：紅細(xì)胞(red blood cell, RBC)輸注效果與紅細(xì)胞保存質(zhì)量有著密切的關(guān)系。針對(duì)紅細(xì)胞保存質(zhì)量難以再提高的問題,有研究結(jié)果顯示,給保存期長(zhǎng)的紅細(xì)胞補(bǔ)充一氧化氮(nitric oxide, NO)可以明顯改善紅細(xì)胞的變形性及輸注后效果。本實(shí)驗(yàn)室在前期研究的基礎(chǔ)上,推測(cè)NO可能通過某些特定細(xì)胞群的作用來提高整體紅細(xì)胞保存質(zhì)量。目的：研究外源性NO藥物供體硝普鈉(sodium nitroprusside, SNP)和前體L-精氨酸(L-arginie, L-arg)對(duì)紅細(xì)胞不同保存時(shí)期質(zhì)量的影響,且探究NO與紅細(xì)胞糖酵解代謝的關(guān)系以此來闡明NO作用于紅細(xì)胞的部分機(jī)理。方法：密度梯度離心法分離已孵育1h不同濃度的SNP和L-arg的紅細(xì)胞,通過檢測(cè)紅細(xì)胞變形性和滲透脆性來分別確立SNP和L-arg的最佳添加濃度。在不同保存時(shí)期添加最佳濃度的NO的紅細(xì)胞經(jīng)密度梯度分離后,檢測(cè)分層后各層紅細(xì)胞數(shù)目比例、丙酮酸激酶、變形性、滲透脆性等指標(biāo)。D.L甘油醛(D.L glyceraldehyde, DL-GA)是目前公認(rèn)的最理想的糖酵解抑制劑,它能特異性抑制丙酮酸激酶(pyruvate kinase,PK)和己糖激酶(hexokinase, HK)活性從而達(dá)到抗糖酵解的目的。將保存末期的紅細(xì)胞內(nèi)分為6組,1組為對(duì)照組,2組為D.L甘油醛,3組為SNP,4組為D.L甘油醛加SNP,5組為L(zhǎng)-arg,6組為D.L甘油醛加L-arg,孵育2h后密度梯度分離后洗滌2次所得的各層紅細(xì)胞及未分層紅細(xì)胞,分別檢測(cè)變形性、PK、HK活性。結(jié)果：保存初期和中期的紅細(xì)胞在添加10l,M的SNP或L-arg后,其變形性和滲透脆性改善效果明顯,末期最佳濃度則為100μM。保存末期的紅細(xì)胞數(shù)目比例在加入NO后2,3層紅細(xì)胞增加,各層紅細(xì)胞的天齡也在加入NO后有明顯降低,其他指標(biāo)均發(fā)現(xiàn)第4層細(xì)胞和保存末期紅細(xì)胞在加入NO后有明顯改善,其他紅細(xì)胞沒有明顯的改變或規(guī)律性。保存末期的SNP或L-arg能明顯提高各層紅細(xì)胞的變形性、PK和HK活性,在此基礎(chǔ)上添加D.L甘油醛能夠降低上述指標(biāo)。結(jié)論：保存初期和中期的紅細(xì)胞SNP和L-arg的最佳濃度均為10μM,末期最佳濃度為NO僅對(duì)老化或保存損傷紅細(xì)胞的保存質(zhì)量有明顯改善,對(duì)年輕紅細(xì)胞沒有規(guī)律性作用。NO能通過促進(jìn)紅細(xì)胞的糖酵解代謝從而提高紅細(xì)胞的保存質(zhì)量。
[Abstract]:Red blood cells (red blood cell, RBC) and the effect of infusion of red blood cell preservation quality are closely related. The red blood cell preservation quality difficult to increase the problem, some researches show that for long period of preservation of the red blood cells of nitric oxide (nitric oxide, NO) could significantly improve the erythrocyte deformability and transport after the injection effect. The laboratory on the basis of previous studies, suggesting that NO may through some specific cell populations to improve the overall quality of preservation of red blood cells. Objective: To study the effect of exogenous NO donor drugs (sodium nitroprusside, SNP) and precursor L- arginine (L-arginie, L-arg) on the quality of red cells to save time, and the relationship of NO and explore the mechanism of erythrocyte glycolysis metabolism in order to clarify the role of NO in the red blood cells. Methods: density gradient centrifugation had been incubated with different concentrations of 1H SNP and L-arg red blood cells The cell, and osmotic fragility to establish SNP and L-arg respectively, the optimal concentration of deformation through the detection of red blood cells. The best adding concentration of NO in different preservation period of red blood cells by density gradient separation, detection of stratified layers of RBC ratio, pyruvate kinase, deformability, osmotic fragility index of glyceraldehyde (.D.L D.L glyceraldehyde, DL-GA) is currently recognized as the most ideal inhibitor of glycolysis, which can specifically inhibit pyruvate kinase (pyruvate kinase, PK) and hexokinase (hexokinase, HK) activity so as to achieve the Antiglycolytic. Will the end of storage in the red blood cells were divided into 6 groups, 1 group the 2 group was D.L group, L-glyceraldehyde, 3 groups of SNP, the 4 group was D.L glyceraldehyde plus SNP, 5 were L-arg, 6 were D.L glyceraldehyde L-arg incubated 2H density gradient separation after washing 2 times the red blood cells and red blood cells were detected without stratification, deformation PK, the activity of HK. Results: the preservation of the early and mid red blood cells in the addition of 10L, M SNP or L-arg, the deformation and permeability improvement effect is obvious, the best concentration was increased at the end of the 2,3 layer 100 M. red blood cell preservation at the end of the red blood cell number ratio after joining NO, each layer of red the day old cells also decreased significantly after joining NO, the other indexes were found in fourth cell layers and preservation of red blood cells was significantly improved in the end after joining the NO, the other red cells did not change or obvious regularity. The end of storage of SNP or L-arg can significantly improve the deformation of each layer of red blood cells, and PK the activity of HK, based on the addition of D.L can reduce the index of glyceraldehyde. Conclusion: the optimum concentration of preservation of the early and mid red blood cells SNP and L-arg were 10 M, at the end of the best concentration of NO on aging or preservation injury of red blood cell preservation quality has improved significantly, for years Light erythrocytes have no regular effect on.NO by promoting the glycolysis and metabolism of red blood cells to improve the quality of red blood cells.

【學(xué)位授予單位】：北京協(xié)和醫(yī)學(xué)院
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：R457.1

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