免疫相关性血细胞减少症患者IgG亚型的研究

发布时间：2018-06-04 08:16

  本文选题：免疫相关 + 血细胞减少　； 参考：《天津医科大学》2015年博士论文

【摘要】：目的检测免疫相关性血细胞减少(immuno-related Immuno-related hemtocytopenia,IRH)患者骨髓造血细胞膜上抗体免疫球蛋白G(immunoglobulin G,IgG)亚型,检测骨髓上清IgG各亚型含量,探索IRH患者IgG的优势亚型,为进一步揭示IRH患者的发病机制提供理论上的依据。方法1.研究对象为初治IRH患者(实验组),以意义未名的血细胞减少症患者(ICUS)及正常人分别为病例对照组及正常对照组,进行以下研究分析:(1)采用流式细胞术(FCM)检测IRH患者骨髓细胞(CDl5+、CD34+及GlycoA+细胞)膜抗体(IgM、IgG);(2)FCM检测IgG(+)的IRH患者及正常人(CDl5+及GlycoA+细胞)膜抗体(IgG1、IgG3);(3)采用免疫组化方法(IHC)对Ig G(+)的IRH患者,ICUS组及正常对照组骨髓有核细胞进行IgG亚型(IgG1、IgG2、IgG3、IgG4)染色。2.选取骨髓造血细胞膜抗体IgG阳性初治IRH患者为实验组,以IRH缓解病人、ICUS初治病人及正常人分别为病例及正常对照组,进行下述研究:(1)采用流式微球捕获技术(CBA)对骨髓上清IgG亚型(IgG1,IgG2,IgG3,IgG4)含量进行检测;(2)采用FCM对外周血CD5+CD19+/CD19+比例进行检测;(3)RT-PCR的方法检测外周血淋巴细胞IgG1的mRNA表达。结果1.IRH患者骨髓造血细胞膜抗体IgG阳性率58.69%(27/46)明显高于病例对照组0%及正常对照组0%,骨髓造血细胞膜抗体IgM阳性率80.43%(37/46)。2.实验组CD15+细胞IgG1的阳性率明显高于正常人((3.13±1.80)%vs(0.87±0.38)%),P〈0.05);实验组CD15+细胞IgG3的阳性率明显高于正常人((2.73±2.09)%vs(0.72±0.42)%,P〈0.05);实验组GlyCoA+细胞IgG1的阳性率明显高于正常人((3.97±2.21)%vs(0.87±0.41)%,P〈0.05);实验组GlyCoA+细胞IgG3的阳性率明显高于正常人((2.24±1.82)%vs(0.71±0.50)%,P〈0.05)。3.应用免疫组化方法分别对实验组,ICUS患者及正常人骨髓有核细胞进行IgG亚型(IgG1,IgG2,IgG3,IgG4)免疫组化染色,11/15名IRH患者计数100个有核细胞IgG1阳性细胞≥3个,在ICUS患者及正常人骨髓有核细胞涂片染色中IgG1阳性细胞未见,对上述三组患者骨髓有核细胞涂片进行IgG2,IgG3,IgG4免疫组化染色,阳性细胞未见。4.应用CBA方法对实验组、IRH恢复病人、正常人及ICUS患者骨髓上清IgG亚型(IgG1,IgG2,IgG3,IgG4)进行检测。IRH组IgG1(6998.51±2498.38)mg/L明显高于IRH恢复组((4702.27±1235.25)mg/L,P〈0.01)正常对照组((5613.20±1640.93)mg/L,P〈0.05)和ICUS组((5166.20±1761.64)mg/L,P〈0.01)。IRH组IgG3(637.63±252.93)mg/L明显高于IRH恢复组((415.47±196.60)mg/L,P〈0.01)正常对照组((422.85±212.69)mg/L,P〈0.05)和ICUS组((485.70±197.29)mg/L,P〈0.05)。IgG2四组比较无统计学意义,IgG4四组比较无统计学意义。IRH组IgG((10903.29±2576.10)mg/L)明显高于IRH恢复组((8468.27±1957.58)mg/L,P〈0.05)。5.实验组CD5+CD19+/CD19+明显高于正常对照组及ICUS组((23.34±9.36)%)vs(13.67±5.10)%vs(12.40±3.63)%(P0.01),实验组CD5+CD19+/CD19+与骨髓上清IgG1水平呈明显正相关关系(r=0.392,P=0.043)。6.实验组,骨髓上清IgG1水平与中性粒细胞绝对值呈明显负相关关系(r=-0.459,P=0.016);骨髓上清IgG3水平与血红蛋白成明显负相关关系(r=-0.389,P=0.045)。7.外周血淋巴细胞IgG1基因相对表达量在实验组,恢复组,正常对照组分别为((5.27±4.83)vs(2.32±2.03)vs(1.58±1.50)),实验组明显高于正常对照组(P〈0.05〉,恢复组与正常对照组比较无统计学意义。结论1.IRH患者的骨髓造血细胞膜上可检测到IgG抗体。通过流式细胞学方法检测到骨髓造血膜抗体IgG阳性组CD15+及GlyCoA+细胞膜抗体IgG1及IgG3升高明显,免疫组化法证实IRH患者骨髓细胞膜抗体IgG亚型以IgG1为主,提示在IRH中破坏造血的IgG亚型可能为IgG1及IgG3。2.BMMNC-Ab IgG阳性患者骨髓上清IgG1与IgG3升高明显,IgG1与CD5+CD19+/CD19+比例呈明显正相关关系,IgG1与患者白细胞呈负相关关系,IgG3与患者血红蛋白呈明显负相关关系。通过RT-PCR方法证实IRH患者外周血淋巴细胞IgG1 mRNA相对表达量较正常对照组增高。以上提示IRH患者IgG1生成增多,且为血细胞破坏的主要因素。
[Abstract]:Objective to detect the subtype of antibody immunoglobulin G (immunoglobulin G, IgG) on the hematopoietic cell membrane of Immuno-related Immuno-related hemtocytopenia (IRH) and to detect the content of IgG subtypes in the bone marrow supernatant, and to explore the dominant subtype of IgG in the IRH patients, providing a theory to further reveal the pathogenesis of the IRH patients. Methods 1. subjects were the primary treatment of IRH patients (experimental group), a case control group and a normal control group with unnamed blood cytopenia (ICUS) and normal controls. The following studies were performed: (1) a flow cytometry (FCM) was used to detect the membrane antibodies (IgM, IgG) of IRH patients (CDl5+, CD34+ and GlycoA+ cells); (2) F. CM detection of IgG (+) of IRH patients and normal people (CDl5+ and GlycoA+ cells) membrane antibody (IgG1, IgG3); (3) using immunohistochemical method (IHC) for Ig G (+) IRH patients, ICUS and normal control group marrow nucleated cells to select the bone marrow hematopoietic cell membrane antibody positive initial treatment group as the experimental group. RH remission patients, ICUS initial treatment patients and normal people were respectively cases and normal controls, and the following study was carried out: (1) the content of IgG subtype (IgG1, IgG2, IgG3, IgG4) in bone marrow supernatant (CBA) was detected by flow microsphere capture (CBA); (2) CD5 +CD19+/CD19+ ratio of FCM peripheral blood was detected; (3) RT-PCR method detected peripheral blood lymphatic fine. Results the mRNA expression of cell IgG1 was 58.69% (27/46) in patients with bone marrow hematopoietic cells (27/46) significantly higher than that in case control group 0% and 0% in normal control group. The positive rate of IgM positive rate of hematopoietic cell membrane antibody in bone marrow 80.43% (37/46).2. experimental group was significantly higher than that of normal people (3.13 + 1.80)%vs (0.87 + 0.38)%), P < 0.05); The positive rate of IgG3 in CD15+ cells was significantly higher than that of normal people (2.73 + 2.09)%vs (0.72 + 0.42)%, P < 0.05). The positive rate of IgG1 in GlyCoA+ cells in experimental group was significantly higher than that of normal people (3.97 + 2.21)%vs (0.87 + 0.41)%, P < 0.05), and the positive rate of GlyCoA+ cell IgG3 in experimental group was significantly higher than that of normal people (2.24 + 1.82)%vs (0.71 +%)%, P Immunohistochemistry method was used to immunohistochemical staining of IgG subtypes (IgG1, IgG2, IgG3, IgG4) in the experimental group, ICUS patients and normal human marrow nucleated cells. The 11/15 name IRH patients counted 100 nucleated cell IgG1 positive cells more than 3, and the IgG1 positive cells were not seen in the ICUS patients and normal human bone marrow smear smear staining, and the three groups of patients were observed. Bone marrow smears were smeared for IgG2, IgG3, and IgG4 immunohistochemical staining. The positive cells did not see the.4. application CBA method to the experimental group, IRH recovery patients, normal and ICUS patients with IgG subtype (IgG1, IgG2, IgG3, IgG4) were significantly higher than those of the recovery group (4702.27 + 1235.25) (4702.27 + 1235.25). The normal control group ((5613.20 + 1640.93) mg/L, P < 0.05) and group ICUS (5166.20 + 1761.64) mg/L, P < 0.01).IRH IgG3 (637.63 + 252.93) mg/L were significantly higher than the IRH recovery group (415.47 + 196.60) mg/L, P < 0.01) normal control group (422.85 + 212.69) mg/L. G4 four groups had no statistically significant.IRH group IgG ((10903.29 + 2576.10) mg/L) significantly higher than the IRH recovery group (8468.27 + 1957.58) mg/L, P < 0.05).5. experimental group CD5+CD19+/CD19+ significantly higher than the normal control group and ICUS group (23.34 + 9.36)%) vs (13.67 + 5.10)%vs (12.40 + 3.63)%. The positive correlation (r=0.392, P=0.043).6. experimental group, the IgG1 level of the bone marrow supernatant was negatively correlated with the absolute neutrophils (r=-0.459, P=0.016); the IgG3 level of the bone marrow supernatant was negatively correlated with the hemoglobin (r=-0.389, P=0.045).7. peripheral blood lymphocyte IgG1 gene relative expression in the experimental group, the recovery group, normal. The control group was (5.27 + 4.83) vs (2.32 + 2.03) vs (1.58 + 1.50)), and the experimental group was significantly higher than that of the normal control group (P < 0.05 >). There was no statistical significance between the recovery group and the normal control group. Conclusion the IgG antibody could be detected on the bone marrow hematopoietic cell membrane of the 1.IRH patients. Through the flow cytometry, the IgG positive group of bone marrow hematopoietic membrane antibody CD was detected in CD. The increase of 15+ and GlyCoA+ cell membrane antibodies IgG1 and IgG3 was obvious. The immunohistochemical method confirmed that the IgG subtype of the bone marrow cell membrane antibody of IRH patients was dominated by IgG1, suggesting that the IgG subtype that destroyed the hematopoiesis in IRH was likely to be higher in IgG1 and IgG3.2.BMMNC-Ab IgG positive bone marrow supernatants. There was a negative correlation between IgG1 and leukocyte in patients. IgG3 was negatively correlated with hemoglobin in patients. The relative expression of IgG1 mRNA in peripheral blood lymphocytes of IRH patients was higher than that in normal control group by RT-PCR method. The above suggested that IgG1 production in IRH patients increased and was the main factor of blood cell destruction.
【学位授予单位】：天津医科大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：R55

【共引文献】

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2 刘惠;免疫相关性全血细胞减少症患者骨髓造血细胞膜靶抗原的初步研究[D];天津医科大学;2010年

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4 任悦;免疫相关性全血细胞减少症患者遗传易感性的初步研究[D];天津医科大学;2014年

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2 邴丽娟;骨髓间接Coombs实验方法的应用及其对免疫相关性血细胞减少症患者的临床诊断意义[D];天津医科大学;2012年

3 张江勃;自身免疫性骨髓衰竭症患者淋巴细胞和CD34~+细胞端粒长度研究[D];天津医科大学;2014年

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