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Objective To investigate the different expression of various subtypes of human leukocyte antigen-G(HLA-G)in placenta of patients complicated with severe pre-eclampsia.Methods Ten placental samples from early-onset severe pre-eclamptic pregnancies and ten from late-onset severe preeclamptic pregnancies were collected as study group; ten placental samples from preterm pregnancies and ten from normal pregnancies were collected as control group.The levels of HLA-G protein in the four groups were measured by western blot and immunohistochemistry.Results(1)HLA-G1 protein decreased significantly in both the early-onset(2.4 ± 0.6 versus 2.9 ± 1.1,P ＜ 0.05)and the late-onset pre-eclampsia groups (3.5 ± 2.1 versus 4.2 ± 2.4,P ＜ 0.05).(2)HLA-G5 protein increased in the late-onset pre-eclampsia groups(1.8 ± 1.1 versus 1.1 ± 0.9,P ＜ 0.05); the increase in the early-onset pre-eclampsia group is not obvious(1.6 ± 0.9 versus 1.4 ± 0.7,P ＞ 0.05).(3)The level of HLA-G1 protein in placenta from patients complicated with premature labor is lower(2.9 ± 1.1 versus 4.2 ± 2.4,P ＜ 0.05); HLA-G5 protein does not change significantly(1.4 ± 0.7 versus 1.1 ± 0.9,P ＞ 0.05).(4)HLA-G1 and G5 proteins mainly express in the placenta extravillous cytotrophoblast cells.There is also a high level of expression around the blood vessels and in the extraembryonic mesoderm.Conclusions(1)HLA-G1 decreased significantly in both the early-onset and late-onset pre-eclamptic patients.(2)HLA-G5 increased in both the early-onset and late-onset pre-eclamptic patients,and the increase in the late-onset pre-eclamptic patients is obvious.(3)In late pregnancy,the level of HLA-G1 is lower in patients complicated with premature labor,this may be the result of its earlier pregnancy week; HLA-G5 does not change significantly.(4)HLA-G1 and G5 mainly express in the placenta extravillous cytotrophoblast cells.
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Objectives To detect the expression of human leukocyte antigen-G (HLA-G) in tissues from pregnant women with preeclampsia and discuss the relationship between HLA-G and preeclampsia.Methods Pregnant women with preeclampsia in Maternal and Child Health Hospital of Shaanxi Province from March 2009 to December 2009 were included.Eight were included into mild preeclampsia groups and 22 were included into severe preeclampsia group.And 30 age-matched normal pregnancies were referred as the control group.All women in the three groups received cesarean section.The soluble HLA-G (sHLA-G)levels in peripheral blood,umbilical blood and amniotic fluid were examined by ELISA ; the expressions of HLA-G protein in placenta,fetal membrane and umbilical cord were examined by western blot.Results ( 1 ) The sHLA-G levels in peripheral blood,umbilical blood and amniotic fluid in each group.The sHLA-G levels in peripheral blood in mild and severe preeclampsia group were (50 + 14) and (30+6) μg/L respectively,and the sHLA-G levels in umbilical blood were (34 ± 10) and (26 ±8)μg/L respectively.All were significantly lower than those in the control group ( P ＜ 0.01 ),which were (100 ± 16) and (70±9) μg/L respectively.There was also statistical difference between mild and severe preeclampsia group (P ＜0.01 ).Although the sHLA-G level in umbilical blood of severe preeclampsia group was lower than that in mild preeclampsia group,there was no statistical difference ( P＞0.05 ).The sHLA-G levels in amniotic fluid in mild and severe preeclampsia groups were (26±7 ) and (25 ± 5 ) μg/L respectively,which were lower than that in the control group (27±6) μg/L,but the differences were not significant ( P＞0.05 ).There was no statistical difference between mild and severe preeclampsia groups ( P＞0.05 ).(2) The expression levels of HLA-G protein in placenta,fetal membrane and umbilical cord in each group.The expression levels of HLA-G in placenta and fetal membrane in the control group were 1.59 ± 0.36 and 0.42 ± 0.09 respectively.The expression of HLA-G in placenta was significantly higher than that in fetal membrane ( P＜0.05 ).The expression level of HLA-G in umbilical cord in the control group was 0.24±0.17,statistically different from those in placenta and fetal membrane,respectively (P＜0.01 ).The expression levels of HLA-G in placenta in mild and severe preeclampsia groups were 0.78 + 0.21 and 0.29 ± 0.17 respectively,significantly different from the control group ( P ＜ 0.01 ).There was no expression of HLA-G in fetal membrane and umbilical cord in mild and severe preeclampsia groups.Conclusions The expressions of HLA-G in the peripheral blood,umbilical blood and placenta in women with preeclampsia are significantly lower than those in normal pregnant women.The abnormal expression of HLA-G might be associated with the pathogenesis of preeclampsia.
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Objective To investigate the role of human leukocyte antigen-G ( HLA-G ) on the invasion and the molecular mechanism involved in this cellular progress in HTR-8/SVneo cell line. Methods There were three groups: groups of transfection, negative control and blank control, which corresponding to treatment by HLA-G specific siRNA, negative siRNA and only lipofectamine 2000 using lipofection technology in HTR-8/SVneo cell line. The efficiency of down-regulated of HLA-G was detected by reverse transcription-polymerase chain reaction and western blot analysis in mRNA and protein level,respectively. Changes of p38 mitogen-activated protein kinases (p-p38MAPK)/p38MAPK protein levels and the cell invasion were respectively detected by western blot analysis and transwell test. Results ( 1 ) The mRNA levels of HLA-G transfection group, negative control group and blank control group were 0. 26 ±0. 08, 0. 71 ±0. 11, 0. 79 ±0. 07, respectively. There was significant difference between transfection group and negative control group ( P ＜ 0. 01 ), while there was no significant difference between negative control group and blank control group ( P ＞ 0. 05 ). The efficiencies of down-regulated of HLA-G were ( 69. 8 ±6. 3)%, ( 14. 9 ± 2. 2 )%, 0 in transfection group, negative control group and blank control group respectively in mRNA level. (2)In protein levels, HLA-G were 0. 20 ±0. 15, 0. 75 ±0. 12, 0. 76 ±0. 21 in transfection group, negative control group and blank control group, respectively. There was significant difference between transfection group and negative control group ( P ＜ 0. 01 ), whereas there was no significant difference between negative control group and blank control group ( P ＞ 0. 05 ). The efficiencies of down-regulated of HLA-G were (81. 1 ± 14.4)%, ( 18.0 ± 7.7)%, 0 in transfection group, negative control group and blank control group respectively. ( 3 ) The invasive number of transfection group, negative control group and blank control group were 57 ± 38,364 ± 79 and 260 ± 84, respectively, with a significant difference between transfection group and negative control group (P ＜ 0. 01 ). There was no significant difference between negative control group and blank control group ( P ＞ 0. 05 ). ( 4 ) The p-p38MAPK/p38MAPK values of the HLA-G transfection group, negative control group and blank control group were 0. 74 ±0.04, 0. 47 ± 0. 09 and 0. 36 ± 0. 21, respectively. HLA-G transfection group was significantly different compared with the other two groups( P ＜0. 01 ). (5)Without or with SB203580, the p-p38MAPK/ p38MAPK values of the HLA-G transfection group were 0. 89 ± 0. 09 and 0. 16 ± 0. 04, the values of negative control group and blank control group were 0.76 ±0.08, 0. 14 ±0.03 and 0.51 ±0.05, 0.03 ±0.01, respectively. There was significant difference between without SB203580 and with SB203580 ( P ＜ 0. 01 ). (6)Without or with SB203580, the invasive number of transfection group were 51 ± 13 and 90 ± 21 ,respectively,which was significantly different ( P ＜ 0. 01 ). The invasive number of negative control group and blank control group were 290 ± 52, 298 ± 33 and 290 ± 73, 264 ± 64, respeczively, which was no significant difference between without SB203580 and with SB203580 (P ＞ 0. 05 ). Conclusions HLA-G gene may regulate invasion of trophoblast-derived cell line HTR-8/SVneo via p38MAPK signaling pathway. The lower expression of HLA-G in trophoblast cells may lead to the occurrence of pathologic pregnancy.
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Objective To explore the effect of hydrogen peroxide (H2O2) on the expression of human leukocyte antigen-G ( HLA-G) in placental trophoblasts in pregnant women with pre-eclampsia.Methods Forty pregnant women,delivered through cesarean section in the Department of Obstetrics of and Gynecology,the First Affiliated Hospital of Nanjing Medical University from October 2008 to October 2009,were enrolled,including 20 women with pre-eclampsia and 20 healthy gravidas (control group).Colorimetry and western blot were applied,respectively,to determine the level of H2O2 and the expression of HLA-G protein in placental tissues and the correlation between them were analyzed.After 24 hours of seeding,JEG-3 cells (the HLA-G positive cell line of choriocarcinoma) were divided into two groups:intervention group (exposure to 175 μmol/L H2O2) and control group (without H2O2).Immunofluorescence and western blot were used to investigate the expression of HLA-G protein in JEG-3 cells at 24 hours and 48 hours after incubation.Results (1 )The level of H2O2 in placenta in the pre-eclampsia group was significantly higher than that in control group[(105 ±13) nmol·mg-1·prot-1 vs (62 ± 18) nmol·mg-1 ·prot-1,P < 0.05].(2) The expression of HLA-G protein in placenta of the pre-eclampsia group was reduced by88%compared with that of the control (0.20 ± 0.08 vs 1.67 ± 0.65,P < 0.05).( 3) Negative correlation was found between HLA-G level and H2O2 expression in the placenta in both groups(r =-0.895,P =0.000).(4) Compared with the control group,the expression of HLA-G protein in JEG-3 cells,after 24 hours and 48 hours exposure to H2O2,reduced by 39% and 80%,respectively,(3.21 ±0.33 vs 1.95 ±0.25 and 0.65 ±0.08,P <0.05,respectively) and 67% reduction was detected from 24 hours to 48 hours of H2O2 exposure (P <0.05).Fluorescence microscope observed reduced expression of HLA-G in JEG-E cells in the intervention group at 48 hours compared to the control group (P<0.05).Conclusion High level of H2O2 could down-regulate HLA-G expression in the placental trophoblasts in pre-eclampsia which may be involved in the pathogenesis of pre-eclampsia.
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Objective To investigate the relationship between human leukocyte antigen-G( HLA-G) gene Exon 8 14 bp deletion polymorphism and the pathogenesis of severe pre-eclampsia.Methods Forty-two pregnant women with severe pre-eclampsia,who admitted to the Third Affiliated Hospital of Zhengzhou University from October 2008 to February 2009,and their newborns were chosen as the severe pre-eclampsia group.Another 45 healthy gravidas at the third trimester and their newborns were chosen as the control.All gravidas in both groups were Han Nationality.HLA-G Exon 8 genotyping was detected by PCR in both groups and the allele frequencies and genotype frequencies were compared between the two groups.The genotype frequencies of maternal-neonatal pairs were also analyzed.Results ( 1 ) In the severe pre-eclampsia group,14% of the maternal-neonatal pairs were homozygote of 14 bp deletion,and significantly higher frequency 33% (15/45) was found in the control group (P =0.038).(2) No significant difference was found in the allele frequencies and genotype frequencies of HLA-G 14 bp deletion polymorphism among all the mothers between the two groups (P >0.05).(3) The + 14 bp and-14 bp allele frequencies of HLA-G 14 bp deletion polymorphism in newborns in the severe pre-eclampsia group were 44% (37/84) and56% (47/84),respectively,and 30% (27/90) and 70% (63/90) in the control group.Although there was no significant difference between the two groups,but differences in trends was identified (χ2= 3.678 P = 0.055) ; The genotype (-14 bp/-14 bp) frequency of neonates in the severe pre-eclampsia group showed no difference compared with that in the control group[29% (12/42) vs 49% (22/45)],but differences in trends was also found (P =0.052).Conclusions HLA-G 14 bp deletion polymorphism is associated with the susceptibility of severe pre-eclampsia in Chinese Han nationality.Maternal-fetal genotype pairs of-14 bp/-14 bp may have reduced risk of severe pre-eclampsia.
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The immune tolerance or immune escape plays important roles in the genesis and progression of malignant gynecological neoplasms.Human leukocyte antigen G(HLA-G)is an important member of major histocompatiblity complex class Ⅰ.HLA-G seems to affect almost every aspect of human immunity.It can inhibit immunological function,and play an important role in immune escape of malignant gynecological neoplasms.
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Objective To investigate the alterations in killer cell immunoglobulin-like receptors (KIRs)2D and their specific HLA-Cw ligands in patients with ankylosing spondylitis(AS)and determine whether the changes were correlate to the pathogenesis of AS.Methods Polymerase chain reaction of sequence specific primerB(PCR-SSP)was employed for genotyping the presence or absence of five KIR2D genes(KIR2DL1,2DS1,KIR2DL2,2DL3,2DS2)as well as HLA-Cw01-08 alleles from genomic DNA in 105 individuals with AS,together with 51 individuals with osteoarthritis(OA)and 120 healthy controls.Then HLA-C10-08 was divided into two groups.HLA-Cwasn and HLA-Cwlys to calculate the frequency of KIRID genotype.HLA-Gu alleles and KIR/HLA-Cw genotypes.Results The frequencies of HLA-Cwlys genes were significandy higher in patients with AS(0.269 7)compared with those in OA controls(0.148 2)and healthy controls(0.138 8,P=0.024,P=0.001,respectively).The frequency of KIR2DS1/HLA-Cwlys combination Was also markedly higher in AS group(26.67%)than that in OA controls(11.76%)and healthy controls(13.33%,P=0.039,P=0.018,respectively).Condusion The data suggest that the HLA-Cwlys allele may be associated with genetic susceptibility to AS and moreover.in the existence of HLA-Cwlys.the individuals with KIR2DS1 gene are likely to be at increased risk of AS.
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Objective To investigate the different expressions of various subtypes of human leukocyte antigen-G(HLA-G)mRNA in placentas among early and late-onset severe preeclamptic and normal pregnant women. Methods Eleven placental samples from early-onset severe preeclamptic pregnancies and fourteen from late-onset severe preeclamptic pregnancies were collected as study group;eight placental samples from normal pregnancies were collected as control group.The levels of HLA-G mRNA in the three groups were measured by nested RT-PCR. Results Compared with the counterparts in the control group,the leveI of HLA-G1 subtype was significantly decreased (median:Early-onset group:1.37,P<0.05;Late-onset group:24.90,P<0.05;Control group:46.67,P<0.05)and the level of HLA-G5 subtype increased(median:Early-onset group 19.23;Late-onset group:3.65;Control group:1.33)in both early and late-onset severe preeclamptic group.The level of HLA-G2 subtype increased(median:Early-onset group:0;Late-onset group:21.59,P<0.05;Control group:5.39)in the late-onset severe preeclamptic group. Conclusion The reduced expression of HLA-G membrane-bound subtype mRNA in the placenta and the increased expression of HLA-G soluble subtype mRNA in the placenta may be related tO the pathogenesis of preeclampsia.
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Objective To study the role of human leukocyte antigen-G (HLA-G) in the carcinogenesis and development of choriocarcinoma. Methods We designed and synthesized a double strand small interference RNA (siRNA) of HLA-G, then transfected it into a HLA-G overexpressed choriocarcinoma cell line, JEG-3 by lipofectine; HLA-G mRNA level was detected by real time RT-PCR; HLA-G protein level was detected by western blot. The living cells of JEG-3 were counted under microscope after transfection by siRNA. Results Double strand siRNA of HLA-G effectively downregulated the mRNA level and the protein level of HLA-G. The mRNA levels by Ct value at different concentration (1.0, 2.5, 5.0 ?g/L) were 20.67?0.02, 21.37?0.03,21.43?0.02, respectively. They were significantly different compared with the control groups which were 20.33?0.01, 20.37?0.02,20.40?0.03, respectively ( P