青海地区藏汉民族HLA-G5-URR基因多态性及血浆sHLA-G与子痫前期的关系
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摘要
目的:检测青海高原地区不同海拔藏汉民族子痫前期(PE)患者人类白细胞抗原-G(HLA-G)5'上游调控区(5-URR)的基因多态性及血浆可溶性HLA-G(s HLA-G)与PE的关系。方法:选择汉族PE患者(汉族PE组)87例、汉族正常妊娠妇女(汉族正常组)101例、藏族PE患者(藏族PE组)52例、藏族正常妊娠妇女(藏族正常组)108例为研究对象,采用全血基因DNA提取、聚合酶联反应技术(PCR)、琼脂糖电泳及测序方法检测4组HLA-G5-URR基因SNP位点,对检测到的位点进行等位基因及基因型分型的比较。酶联免疫吸附法(Elisa)检测各组30例血浆中s HLA-G蛋白浓度,并分析HLA-G基因5-URR区域差异性分布的SNP位点多态性对s HLA-G表达的影响。结果:(1)HLA-G基因5-URR区域检测到6个SNP,分别为+124位点、+250位点、+274位点、+289位点、+308位点及+465位点,其中+274位点基因型和等位基因频率在汉族正常组与汉族PE组的差异有统计学意义(基因型P=0.007;等位基因频率P=0.006);+289位点基因型的分布在汉族正常组与藏族正常组、汉族PE组与藏族PE组间差异有统计学意义(P=0.010;P=0.002);+289位点等位基因频率在汉族PE组与藏族PE组间差异有统计学意义(P=0.003)。(2)汉族正常组、汉族PE组、藏族正常组、藏族PE组的s HLA-G蛋白浓度分别为2.63±0.88 ng/L、1.77±0.44 ng/L、2.27±0.57 ng/L、1.44±0.51 ng/L,4组间两两比较,差异均有统计学意义(P<0.05)。(3)HLA-G基因5-URR区域+274位点和+289位点的3种基因型AA、AG、GG在4组内的s HLA-G蛋白表达水平比较,差异均无统计学意义(P>0.05)。结论:HLA-G基因5-URR+274位点和+289位点的多态性可能与PE的易感性有关,尤其是+289位点的基因多态性可能是导致青海地区藏族人群易患PE的因素之一。高原慢性低氧环境可能影响s HLA-G蛋白的表达,可能参与了青海高原地区PE的发生;不能完全排除HLA-G基因5-URR多态性与HLA-G蛋白表达的关系。
Objective: To investigate the pathogenesis of preeclampsia( PE) in Tibetan and Han nationalities through testing the gene polymorphism in 5'-upstream regulatory region( 5'URR) of Human Leukocyte Antigen-G( HLA-G) and the plasma soluble human leucocyte antigen G( s HLA-G).Methods: Extraction of whole blood DNA gene,polymerase chain reaction( PCR),agarose gel electrophoresis and sequencing were used to detect the single nucleotide polymorphism( SNP) of 5-URR of HLA-G gene in 87 Han and 52 Tibetan with PE and 101 Han and108 Tibetan controls.Then the allele and genotype frequencies of SNPs were analysed.The s HLA-G protein in 30 plasma cases of each group was detected by enzyme-linked immunosorbent assay( Elisa).Results:(1)6 SNPs were detected: +124 locus、+250 locus、+274 locus、+289 locus、+308 locus and +465 locus.Genotype distribution and allele frequency of +274 locus had significant difference between control group and PE group of Han nationality( Genotype P=0. 007; Allele frequency P=0. 006); The genotype distribution of +289 locus had significant difference between normal groups of Tibetan and Han nationalities( P = 0. 010),and PE groups of this two nationalities,too( P = 0. 002); The allele frequency of +289 locus had significant difference between PE groups of Han nationality and Tibetan( P = 0. 003).(2)The concentration of s HLA-G protein in control and PE group of Han nationality,and control and PE groups of Tibetan were 2. 63±0. 88 ng/L、1. 77±0. 44 ng/L、2. 27±0. 57 ng/L、1. 44±0. 51 ng/L,respectively. The paired comparison in the 4 groups had significant differences( P < 0. 05).(3) In the four groups,there were no significant differences in the expression of s HLA-G protein in AA,AG and GG genotypes at +274 locus and +289 locus of the HLA-G gene 5-URR( P>0. 05).Conclusions: The polymorphism of +274 locus and +289 locus in 5-URR of HLA-G gene may be related to the susceptibility to PE,especially the polymorphism of +289 locus in Qinghai Tibetan population; The chronic hypoxic environment in the plateau area may affect the expression of s HLA-G protein,participating in the pathogenesis of PE in Qinghai plateau; It cannot be completely ruled out that the relationship between polymorphism of HLA-G 5-URR and the expression of HLA-G protein.
Objective: To investigate the pathogenesis of preeclampsia( PE) in Tibetan and Han nationalities through testing the gene polymorphism in 5'-upstream regulatory region( 5'URR) of Human Leukocyte Antigen-G( HLA-G) and the plasma soluble human leucocyte antigen G( s HLA-G).Methods: Extraction of whole blood DNA gene,polymerase chain reaction( PCR),agarose gel electrophoresis and sequencing were used to detect the single nucleotide polymorphism( SNP) of 5-URR of HLA-G gene in 87 Han and 52 Tibetan with PE and 101 Han and108 Tibetan controls.Then the allele and genotype frequencies of SNPs were analysed.The s HLA-G protein in 30 plasma cases of each group was detected by enzyme-linked immunosorbent assay( Elisa).Results:(1)6 SNPs were detected: +124 locus、+250 locus、+274 locus、+289 locus、+308 locus and +465 locus.Genotype distribution and allele frequency of +274 locus had significant difference between control group and PE group of Han nationality( Genotype P=0. 007; Allele frequency P=0. 006); The genotype distribution of +289 locus had significant difference between normal groups of Tibetan and Han nationalities( P = 0. 010),and PE groups of this two nationalities,too( P = 0. 002); The allele frequency of +289 locus had significant difference between PE groups of Han nationality and Tibetan( P = 0. 003).(2)The concentration of s HLA-G protein in control and PE group of Han nationality,and control and PE groups of Tibetan were 2. 63±0. 88 ng/L、1. 77±0. 44 ng/L、2. 27±0. 57 ng/L、1. 44±0. 51 ng/L,respectively. The paired comparison in the 4 groups had significant differences( P < 0. 05).(3) In the four groups,there were no significant differences in the expression of s HLA-G protein in AA,AG and GG genotypes at +274 locus and +289 locus of the HLA-G gene 5-URR( P>0. 05).Conclusions: The polymorphism of +274 locus and +289 locus in 5-URR of HLA-G gene may be related to the susceptibility to PE,especially the polymorphism of +289 locus in Qinghai Tibetan population; The chronic hypoxic environment in the plateau area may affect the expression of s HLA-G protein,participating in the pathogenesis of PE in Qinghai plateau; It cannot be completely ruled out that the relationship between polymorphism of HLA-G 5-URR and the expression of HLA-G protein.
引文
[1]Hviid TV.Non Classical human leukocyte antigen(HLA)tissue types from implantation to transplantation[J].Ugeskr Laeger,2006,168(5):461-466.
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[4]Brenol CV,Veit TD,Chies JA,et al.The role of the HLA-G gene and molecule on the clinical expression of rheumatologic diseases[J].Rev Bras Reumatol,2012,52(1):82-91.
[5]Robinson J,Halliwell JA,Hayhurst JD,et al.The IPD and IMGT/HLAdatabase:Allele variant databases[J].Nucleic Acids Res,2015,28(43):D423-D431.
[6]Nilsson LL,Djurisic S,Andersen AM,et al.Distribution of HLA-G extended haplotypes and one HLA-E polymorphism in a large-scale study of mother-child dyads with and without severe preeclampsia and eclampsia[J].HLA,2016,88(4):172-186.
[7]张展,张楠,张琳琳,等.HLA-G基因非编码区多态性与重度子痫前期的相关性[J].中国妇幼保健,2011,26(14):2169-2172.
[8]Noci I,Fuzzi B,Rizzo R,et al.Embryonic soluble HLA-Gas a marker of developmental potential in embryos[J].Hum Rep rod,2005,20(1):138-146.
[9]Darmochwal-Kolarz D,Kolarz B,Rolinski J,et al.The concentrations of soluble HLA-G protein are elevated during mid-gestation and decreased in pre-eclampsia[J].Folia Histochem Cytobiol,2012,50(2):286-291.
[10]He Y,Chen S,Huang H,et al.Association between decreased plasma levels of soluble human leukocyte antigen-G and severepre-eclampsia[J].J Perinat Med,2016,44(3):283-290.
[11]赵三存,李志斌,贺同强,等.人类白细胞抗原G的表达与子痫前期发病的关系[J].中华妇产科杂志,2011,46(10):758-762.
[12]谢莹莺,李凤莲,赵生珠,等.青海地区子痫前期患者血清可溶性人类白细胞抗原G检测及预诊指标的建立[J].青海医学院学报,2013,34(2):133-136.
[13]Yie SM,Li LH,Xiao R,et al.A single base-pair mutation in the3'-untranslated region of HLA-G mRNA is associated with pre-eclampsia[J].Mol Hum Reprod,2008,14(11):649-653.
[14]Hviid TV.HLA-G in human reproduction:aspects of genetics,function and pregnancy complications[J].Hum Reprod Update,2006,12(3):209-232.
[15]Hviid TV,Rizzo R,Christiansen OB,et al.HLA-G and IL-10 in serum in relation to HLA-G genotype and polymorphisms[J].Immunogenetics,2004,56(3):135-141.
[16]Loisel DA,Billstrand C,Murray K,et al.The maternal HLA-G 1597ΔCnull mutation is associated with increased risk of pre-eclampsia and reduced HLA-G expression during pregnancy in African-American women[J].Hum Immunol,2015,76(4):260-271.
[17]谢莹莺,李凤莲,刘春燕,等.青海地区子痫前期患者HLA-G mR-NA及蛋白表达研究[J].实用妇产科杂志,2012,28(8):671-674.
[2]Darmochwal-Kolarz D,Kolarz B,Rolinski J,et al.The concentrations of soluble HLA-G protein are elevated during mid-gestation and decreased in pre-eclampsia[J].Folia Histochem Cytobiol,2012,50(2):286-291.
[3]Hviid TV,Rizzo R,Melchiorri L,et al.Polymorphism in the 5'upstream regulatory and 3'untranslated regions of the HLA-G gene in relation to soluble HLA-G and IL-10 expression[J].Human Immunology,2006,67(1):53.
[4]Brenol CV,Veit TD,Chies JA,et al.The role of the HLA-G gene and molecule on the clinical expression of rheumatologic diseases[J].Rev Bras Reumatol,2012,52(1):82-91.
[5]Robinson J,Halliwell JA,Hayhurst JD,et al.The IPD and IMGT/HLAdatabase:Allele variant databases[J].Nucleic Acids Res,2015,28(43):D423-D431.
[6]Nilsson LL,Djurisic S,Andersen AM,et al.Distribution of HLA-G extended haplotypes and one HLA-E polymorphism in a large-scale study of mother-child dyads with and without severe preeclampsia and eclampsia[J].HLA,2016,88(4):172-186.
[7]张展,张楠,张琳琳,等.HLA-G基因非编码区多态性与重度子痫前期的相关性[J].中国妇幼保健,2011,26(14):2169-2172.
[8]Noci I,Fuzzi B,Rizzo R,et al.Embryonic soluble HLA-Gas a marker of developmental potential in embryos[J].Hum Rep rod,2005,20(1):138-146.
[9]Darmochwal-Kolarz D,Kolarz B,Rolinski J,et al.The concentrations of soluble HLA-G protein are elevated during mid-gestation and decreased in pre-eclampsia[J].Folia Histochem Cytobiol,2012,50(2):286-291.
[10]He Y,Chen S,Huang H,et al.Association between decreased plasma levels of soluble human leukocyte antigen-G and severepre-eclampsia[J].J Perinat Med,2016,44(3):283-290.
[11]赵三存,李志斌,贺同强,等.人类白细胞抗原G的表达与子痫前期发病的关系[J].中华妇产科杂志,2011,46(10):758-762.
[12]谢莹莺,李凤莲,赵生珠,等.青海地区子痫前期患者血清可溶性人类白细胞抗原G检测及预诊指标的建立[J].青海医学院学报,2013,34(2):133-136.
[13]Yie SM,Li LH,Xiao R,et al.A single base-pair mutation in the3'-untranslated region of HLA-G mRNA is associated with pre-eclampsia[J].Mol Hum Reprod,2008,14(11):649-653.
[14]Hviid TV.HLA-G in human reproduction:aspects of genetics,function and pregnancy complications[J].Hum Reprod Update,2006,12(3):209-232.
[15]Hviid TV,Rizzo R,Christiansen OB,et al.HLA-G and IL-10 in serum in relation to HLA-G genotype and polymorphisms[J].Immunogenetics,2004,56(3):135-141.
[16]Loisel DA,Billstrand C,Murray K,et al.The maternal HLA-G 1597ΔCnull mutation is associated with increased risk of pre-eclampsia and reduced HLA-G expression during pregnancy in African-American women[J].Hum Immunol,2015,76(4):260-271.
[17]谢莹莺,李凤莲,刘春燕,等.青海地区子痫前期患者HLA-G mR-NA及蛋白表达研究[J].实用妇产科杂志,2012,28(8):671-674.