样本资源库中肝病患者和HIV感染者外周血单个核细胞分离效果比较
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摘要
目的样本资源库中不同患者分离的外周血单个核细胞(peripheral blood mononuclear cell,PBMC)保存效果不同,从而影响后续科学研究。对肝病患者和人类免疫缺陷病毒(human immunodeficiency virus,HIV)感染者PBMC分离效果进行分析比较,有助于更好的对患者PBMC细胞质量进行控制。方法 121例标本根据疾病诊断分为肝病患者35例,HIV感染者51例,对照组35例。淋巴细胞分离液密度梯度离心法分离PBMC,吖啶橙/碘化丙啶双染色法检测PBMC的活细胞、总细胞和细胞活性。结果 3组人群PBMC总细胞数结果显示对照组分离细胞最多(1.54×106,9.03×105~3.53×106),其次为HIV感染者人群(1.08×106,3.72×105~2.82×106),最后为肝病患者(8.49×105,3.61×105~2.30×106),3组之间差异具有统计学意义(P<0.05)。活细胞结果显示对照组分离后活细胞浓度最高(1.53×106,8.79×105~3.51×106),其次为HIV感染者人群(1.06×106,3.46×105~2.77×106),最后为肝病患者(8.44×105,3.54×105~2.25×106),3组之间差异具有统计学意义(P<0.05)。细胞活性结果显示对照组细胞活性(98.40±0.39)高于肝病患者(97.28±0.33)和HIV感染者(96.86±0.37),3组之间差异具有统计学意义(P<0.05)。结论肝病患者和HIV感染者在分离PBMC活细胞、总细胞和细胞活性方面均要低于正常对照组。
Objective The quality of isolated peripheral blood mononuclear cells( PBMC)can influence the preservation of PBMC,thus affect the subsequent scientific research. Liver disease individuals and HIV infected individuals are the main study population in hepatitis/AIDS Biobank.Analysis the PBMC of this two groups could help to better control the quality of PBMC. Methods One hundred and twenty-one samples including 35 liver disease individuals,51 HIV infected individuals and 35 control were analysed. Ficoll density gradient centrifugation to isolate PBMC,AO/PI double staining method to detect the living cells,total cells and cell viability. Results The total number of cells in the control group( 1. 54×106,9. 03×105—3. 53×106) were the highest,followed by HIV infected individuals( 1. 08×106,3. 72×105—2. 82×106),and then the liver disease individuals( 8. 49×105,3. 61×105—2. 30×106),P<0. 05. Living cells in the control group were the highest( 1. 53 × 106,8. 79×105—3. 51×106),followed by HIV infected individuals( 1. 06×106,3. 46×105—2. 77×106),and the liver disease individuals( 8. 44×105,3. 54×105—2. 25×106),P<0. 05. The cell viability of the control( 98. 40±0. 39) was higher than that of liver disease individuals( 97. 28±0. 33) and HIV infected individuals( 96. 86±0. 37),P<0. 05. Conclusion The PBMC isolated from liver disease individuals and HIV infected individuals were different with control,both the number and the cell viability were lower than those of control.
Objective The quality of isolated peripheral blood mononuclear cells( PBMC)can influence the preservation of PBMC,thus affect the subsequent scientific research. Liver disease individuals and HIV infected individuals are the main study population in hepatitis/AIDS Biobank.Analysis the PBMC of this two groups could help to better control the quality of PBMC. Methods One hundred and twenty-one samples including 35 liver disease individuals,51 HIV infected individuals and 35 control were analysed. Ficoll density gradient centrifugation to isolate PBMC,AO/PI double staining method to detect the living cells,total cells and cell viability. Results The total number of cells in the control group( 1. 54×106,9. 03×105—3. 53×106) were the highest,followed by HIV infected individuals( 1. 08×106,3. 72×105—2. 82×106),and then the liver disease individuals( 8. 49×105,3. 61×105—2. 30×106),P<0. 05. Living cells in the control group were the highest( 1. 53 × 106,8. 79×105—3. 51×106),followed by HIV infected individuals( 1. 06×106,3. 46×105—2. 77×106),and the liver disease individuals( 8. 44×105,3. 54×105—2. 25×106),P<0. 05. The cell viability of the control( 98. 40±0. 39) was higher than that of liver disease individuals( 97. 28±0. 33) and HIV infected individuals( 96. 86±0. 37),P<0. 05. Conclusion The PBMC isolated from liver disease individuals and HIV infected individuals were different with control,both the number and the cell viability were lower than those of control.
引文
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[4]李海燕,张雪娇,邵雪梅,等.疾病资源库建设的重要性及现状分析[J].中华医院管理杂志,2010,(11):801-804.
[5]孙焕芹,刘梦佳,戴北宸,等.用转化医学理念指导样本资源库建设[J].北京医学,2012,(6):495-496.
[6]孙坚萍,孙焕芹,刘宁,等.传染病样本资源库质量控制体系建设经验[J].中华医院管理杂志,2016,(9):698-699.
[7]Bolen CR,Uduman M,Kleinstein SH.Cell subset prediction for blood genomic studies[J].BMC Bioinformatics,2011,12:258.
[8]Wieczorek L,Krebs SJ,Kalyanaraman V,et al.Comparable antigenicity and immunogenicity of oligomeric forms of a novel,acute HIV-1 subtype c gp145 envelope for use in preclinical and clinical vaccine research[J].J Virol,2015,89(15):7478-7493.
[9]Wetzel KS,Yi Y,Elliott ST,et al.CXCR6-Mediated simian immunodeficiency virus sivagmsab entry into sabaeus African green monkey lymphocytes implicates widespread use of non-ccr5 pathways in natural host infections[J].J Virol,2017,91(4).pii:e01626-16
[10]Donahue RN,Lepone LM,Grenga I,et al.Analyses of the peripheral immunome following multiple administrations of avelumab,a human IgG 1 anti-PD-L1 monoclonal antibody[J].J Immunother Cancer,2017,5:20.
[11]Chéret A,Nembot G,Mélard A,et al.Intensive five-drug antiretroviral therapy regimen versus standard triple-drug therapy during primary HIV-1 infection(OPTIPRIM-ANRS147):a randomised,open-label,phase 3 trial[J].Lancet Infect Dis,2015,15(4):387-396.
[12]郜恒骏.中国生物样本库向标准化迈进[J].中国医药生物技术,2015,10(6):481-483.
[13]杜莉利.生物样本库的标准化建设[J].转化医学杂志,2016,5(6):324-326.
[14]Malm L,Tybring G,Moritz T,et al.Metabolomic quality assessment of EDTA plasma and serum samples[J].Biopreserv Biobank,2016,14(5):416-423.
[15]Grievink HW,Luisman T,Kluft C,et al.Comparison of three isolation techniques for human peripheral blood mononuclear cells:cell recovery and viability,population composition,and cell functionality[J].Biopreserv Biobank,2016,14(5):410-415.
[16]Klatzmann D,Champagne E,Chamaret S,et al.T-lymphocyte T4 molecule behaves as the receptor for human retrovirus LAV[J].Nature,1984,312(5996):767-768.
[17]田瑛,邱志峰,李太生.慢性乙型肝炎患者和乙型肝炎病毒携带者外周血T细胞亚群的变化和意义[J].中华医学杂志,2005(47):3354-3358.
[18]强占荣,易彩霞,刘颖,等.慢性HBV感染者外周血淋巴细胞亚群及血清IgG和补体检测的临床意义[J].内科,2016,11(1):1-3.