子癇前期外周血及胎盤組織中HNP3、PAPPA2的研究
發(fā)布時(shí)間：2019-06-03 02:07
【摘要】：目的:篩選子癇前期患者與正常妊娠孕婦血清中的差異蛋白,對(duì)相關(guān)差異蛋白進(jìn)行檢測(cè)并初步探討其在子癇前期發(fā)病過程中可能的作用機(jī)制,以期篩選出能夠早期預(yù)測(cè)子癇前期的潛在血清生物標(biāo)志物。方法:收集2015年6月至2016年6月在青島大學(xué)附屬醫(yī)院產(chǎn)科住院治療的重度子癇前期患者30例,設(shè)為病例組;同期收集30例正常妊娠,因社會(huì)因素、產(chǎn)道異常、胎位不正等原因行擇期剖宮產(chǎn)的孕婦,設(shè)為對(duì)照組。懫血分離血清提取總蛋白,經(jīng)iTRAQ標(biāo)記后,用質(zhì)譜儀進(jìn)行鑒定,獲得差異表達(dá)的蛋白質(zhì)。差異蛋白的研究采用定量蛋白質(zhì)組學(xué)技術(shù),差異蛋白功能分析采用生物信息學(xué)的方法。同期收集另外30例重度子癇前期患者,作為重度組;30例輕度子癇前期患者,作為輕度組;30例正常妊娠孕婦,作為正常組。采血分離血清、收集胎盤組織。選取有統(tǒng)計(jì)學(xué)意義的差異蛋白:中性粒細(xì)胞防御素3(HNP3)、冠毛素2(PAPPA2),應(yīng)用酶聯(lián)免疫吸附試驗(yàn)以及實(shí)時(shí)熒光定量PCR技術(shù)分別檢測(cè)其在三組研究對(duì)象外周血及胎盤組織中的表達(dá)。結(jié)果:(1)共篩選出血清差異蛋白234個(gè),病例組與對(duì)照組相比表達(dá)豐度相差1.6倍以上或0.625倍以下且經(jīng)t檢驗(yàn)確認(rèn),差異有統(tǒng)計(jì)學(xué)意義的差異蛋白點(diǎn)共25個(gè),其中表達(dá)上調(diào)的蛋白點(diǎn)有16個(gè),另9個(gè)蛋白點(diǎn)表達(dá)下調(diào);(2)重度組及輕度組子癇前期患者血清中HNP3水平分別為(144.0±22.35)ng/m L、(98.9±11.70)ng/m L均高于正常組(83.8±11.71)ng/m L(P0.0001、P=0.045,P0.05),差異有統(tǒng)計(jì)學(xué)意義;且重度組高于輕度組(P=0.0074,P0.01),差異具有統(tǒng)計(jì)學(xué)意義;(3)重度組及輕度組子癇前期患者胎盤組織中HNP3m RNA的相對(duì)表達(dá)量分別為(4.856±0.077)、(1.978±0.156)均高于正常組(1.001±0.219)(P=0.001、P=0.032,P0.05),差異有統(tǒng)計(jì)學(xué)意義;重度組與輕度組相比(P=0.015,P0.05),差異亦具有統(tǒng)計(jì)學(xué)意義;(4)子癇前期患者血清中HNP3水平與胎盤組織中HNP3m RNA相對(duì)表達(dá)量無相關(guān)性,兩者相關(guān)系數(shù)r=0.554,P=0.626,P0.05,無統(tǒng)計(jì)學(xué)意義;(5)重度組及輕度組子癇前期患者血清中PAPPA2水平分別為(549.1±36.68)pg/m L、(166.1±16.41)pg/m L均高于正常組(138.1±9.05)pg/m L(P0.0001、P=0.014,P0.05),差異有統(tǒng)計(jì)學(xué)意義;且重度組高于輕度組(P=0.0015,P0.01),差異有統(tǒng)計(jì)學(xué)意義;(6)重度組及輕度組子癇前期患者胎盤組織中PAPPA2m RNA的相對(duì)表達(dá)量分別為(13.136±0.230)、(4.523±0.282)均高于正常組(1.000±0.169)(P0.0001、P=0.015,P0.05),差異有統(tǒng)計(jì)學(xué)意義;比較重度組與輕度組(P=0.001,P0.01),差異具有統(tǒng)計(jì)學(xué)意義;(7)子癇前期患者血清中PAPPA2水平與胎盤組織中PAPPA2m RNA相對(duì)表達(dá)量呈正相關(guān),兩者相關(guān)系數(shù)r=0.944,P=0.005,P0.05,相關(guān)有統(tǒng)計(jì)學(xué)意義;(8)子癇前期患者血清中HNP3與PAPPA2水平呈正相關(guān),兩者相關(guān)系數(shù)r=0.852,P0.0001,相關(guān)有統(tǒng)計(jì)學(xué)意義。結(jié)論:(1)iTRAQ聯(lián)合LC-MS/MS技術(shù)為子癇前期早期血清標(biāo)志物的篩選與鑒定提供了有效的技術(shù)支持;(2)中性粒細(xì)胞防御素3(HNP3)和冠毛素2(PAPPA2)可能參與了子癇前期的發(fā)生發(fā)展;(3)中性粒細(xì)胞防御素3(HNP3)和冠毛素2(PAPPA2)在子癇前期的發(fā)病過程中可能存在相互作用;(4)冠毛素2(PAPPA2)有可能成為早期預(yù)測(cè)子癇前期的潛在血清生物標(biāo)志物。
[Abstract]:Objective: To screen the differential protein in the serum of pre-eclampsia and the normal pregnant women, to test the related differential protein and to explore the possible mechanism of the early preeclampsia in the early stage of preeclampsia, with a view to screening the potential serum biomarkers in the early preeclampsia. Methods:30 cases of severe preeclampsia were collected from June,2015 to June,2016 in the hospital of the Affiliated Hospital of the University of Wisconsin, and 30 cases of normal pregnancy were collected in the same period. Set as the control group. The total protein of the serum was isolated from the serum. After the iTRAQ marker, the protein with differential expression was obtained by mass spectrometer. The research of differential protein is based on the quantitative proteomics technology, and the functional analysis of the differential protein is a bioinformatics method. In the same period,30 cases of severe preeclampsia were collected as the severe group,30 of the 30 patients with mild preeclampsia were treated as mild group, and 30 normal pregnant women were used as the normal group. Blood was collected to separate the serum and the placental tissue was collected. The differentially expressed proteins were selected: Neutrophil Defensin 3 (HNP3), PAPP-2 (PAPPA2), Enzyme-linked Immunosorbent Assay (ELISA) and real-time fluorescence quantitative PCR (RT-PCR). Results: (1) A total of 234 serum differential proteins were selected, and the expression abundance was more than 1.6 times or 0.625 times or 0.625 times as compared with the control group. The levels of HNP3 in serum of patients with severe and mild preeclampsia were (144.0-22.35) ng/ mL, (98.9-11.70) ng/ mL higher than that of normal group (83.8-11.71) ng/ mL (P0.01, P = 0.045, P0.05). The relative expression of HNP3m in the placenta was higher than that in the normal group (1.001 and 0.219) (P = 0.001, P = 0.032, P0.05). (4) There was no correlation between the level of HNP3 in serum of preeclampsia and the relative expression of HNP3m in placenta, and the correlation coefficient was r = 0.554, P = 0.626, P <0.05, and there was no statistical significance. (5) The levels of PAPPA2 in serum of severe and mild group of preeclampsia were (549.1, 36.68) pg/ mL, (166.1-16.41) pg/ m L were higher than that of normal group (138.1-9.05) pg/ m L (P = 0.014, P0.05), and the difference was statistically significant; and the severe group was higher than that of mild group (P = 0.0015, P0.01). (6) The relative expression of PAPPA2 mRNA in the placenta of the patients with severe and mild preeclampsia were (13.136, 0.230) and (4.523, 0.282) higher than that in the normal group (1.000-0.169) (P = 0.015, P <0.05). (7) The level of PAPPA2 in the serum of preeclampsia was positively correlated with the relative expression of PAPPA2 mRNA in the placenta tissue, the correlation coefficient r = 0.944, P = 0.005, P 0.05, and the correlation was statistical significance; (8) The level of HNP3 in the serum of preeclampsia was positively correlated with the level of PAPPA2, and the correlation coefficient was r = 0.852, P <0001, and the correlation was of statistical significance. Conclusion: (1) The combination of iTRAQ and LC-MS/ MS provides effective technical support for screening and identification of early-stage serum markers in pre-eclampsia. (2) Neutrophil-Defensin 3 (HNP3) and Seaponin 2 (PAPPA2) may be involved in the development of pre-eclampsia. (3) Neutrophil-3 (HNP3) and hypertrichostatin 2 (PAPPA2) may interact in the pathogenesis of pre-eclampsia; (4) the preeclampsia-2 (PAPPA2) is likely to be a potential serum biomarker for early preeclampsia.
【學(xué)位授予單位】：青島大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R714.244
本文編號(hào)：2491589
[Abstract]:Objective: To screen the differential protein in the serum of pre-eclampsia and the normal pregnant women, to test the related differential protein and to explore the possible mechanism of the early preeclampsia in the early stage of preeclampsia, with a view to screening the potential serum biomarkers in the early preeclampsia. Methods:30 cases of severe preeclampsia were collected from June,2015 to June,2016 in the hospital of the Affiliated Hospital of the University of Wisconsin, and 30 cases of normal pregnancy were collected in the same period. Set as the control group. The total protein of the serum was isolated from the serum. After the iTRAQ marker, the protein with differential expression was obtained by mass spectrometer. The research of differential protein is based on the quantitative proteomics technology, and the functional analysis of the differential protein is a bioinformatics method. In the same period,30 cases of severe preeclampsia were collected as the severe group,30 of the 30 patients with mild preeclampsia were treated as mild group, and 30 normal pregnant women were used as the normal group. Blood was collected to separate the serum and the placental tissue was collected. The differentially expressed proteins were selected: Neutrophil Defensin 3 (HNP3), PAPP-2 (PAPPA2), Enzyme-linked Immunosorbent Assay (ELISA) and real-time fluorescence quantitative PCR (RT-PCR). Results: (1) A total of 234 serum differential proteins were selected, and the expression abundance was more than 1.6 times or 0.625 times or 0.625 times as compared with the control group. The levels of HNP3 in serum of patients with severe and mild preeclampsia were (144.0-22.35) ng/ mL, (98.9-11.70) ng/ mL higher than that of normal group (83.8-11.71) ng/ mL (P0.01, P = 0.045, P0.05). The relative expression of HNP3m in the placenta was higher than that in the normal group (1.001 and 0.219) (P = 0.001, P = 0.032, P0.05). (4) There was no correlation between the level of HNP3 in serum of preeclampsia and the relative expression of HNP3m in placenta, and the correlation coefficient was r = 0.554, P = 0.626, P <0.05, and there was no statistical significance. (5) The levels of PAPPA2 in serum of severe and mild group of preeclampsia were (549.1, 36.68) pg/ mL, (166.1-16.41) pg/ m L were higher than that of normal group (138.1-9.05) pg/ m L (P = 0.014, P0.05), and the difference was statistically significant; and the severe group was higher than that of mild group (P = 0.0015, P0.01). (6) The relative expression of PAPPA2 mRNA in the placenta of the patients with severe and mild preeclampsia were (13.136, 0.230) and (4.523, 0.282) higher than that in the normal group (1.000-0.169) (P = 0.015, P <0.05). (7) The level of PAPPA2 in the serum of preeclampsia was positively correlated with the relative expression of PAPPA2 mRNA in the placenta tissue, the correlation coefficient r = 0.944, P = 0.005, P 0.05, and the correlation was statistical significance; (8) The level of HNP3 in the serum of preeclampsia was positively correlated with the level of PAPPA2, and the correlation coefficient was r = 0.852, P <0001, and the correlation was of statistical significance. Conclusion: (1) The combination of iTRAQ and LC-MS/ MS provides effective technical support for screening and identification of early-stage serum markers in pre-eclampsia. (2) Neutrophil-Defensin 3 (HNP3) and Seaponin 2 (PAPPA2) may be involved in the development of pre-eclampsia. (3) Neutrophil-3 (HNP3) and hypertrichostatin 2 (PAPPA2) may interact in the pathogenesis of pre-eclampsia; (4) the preeclampsia-2 (PAPPA2) is likely to be a potential serum biomarker for early preeclampsia.
【學(xué)位授予單位】：青島大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R714.244
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