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Multiple Inherited Thrombophilic Gene Polymorphisms in Spontaneous Abortions in Turkish Population.

Yalcintepe S, Ozdemir O, Hacivelioglu SO, Akurut C, Koc E, Uludag A, Cosar E, Silan F - Int J Mol Cell Med (2015)

Bottom Line: For this purpose, the Factor V Leiden (FVL) (rs6025), Prothrombin G20210A (rs1799963), MTHFR C677T (rs1801133), PAI-1 4G/5G (rs1799889), ACE I/D (rs1799752), eNOS E298D (rs1799983), and Apo E E2/E3/E4 (rs429358) polymorphisms were genotyped and correlated in spontaneously aborted fetal materials, their mothers and fertile women.Target SNPs for each gene were analyzed by real time-PCR technique after genomic DNA isolation from maternal blood-EDTA, control group blood-EDTA and spontaneously aborted fetal tissues.Some cases had a single thrombophilic polymorphism, but the rest of the patients and fetal materials had combined thrombophilic polymorphisms.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Genetics, Faculty of Medicine, Canakkale Onsekiz Mart University, Canakkale, Turkey. ; Genetics Diagnosis Center, Numune Training and Education Hospital, Adana, Turkey.

ABSTRACT
The aim of this study was to investigate the possible role of multiple inherited thrombophilic gene variations in women with unexplained spontaneous abortions. For this purpose, the Factor V Leiden (FVL) (rs6025), Prothrombin G20210A (rs1799963), MTHFR C677T (rs1801133), PAI-1 4G/5G (rs1799889), ACE I/D (rs1799752), eNOS E298D (rs1799983), and Apo E E2/E3/E4 (rs429358) polymorphisms were genotyped and correlated in spontaneously aborted fetal materials, their mothers and fertile women. Twenty three abortion materials, 22 women with ≥1 unexplained fetal loss, and 22 control subjects with at least two healthy term infants as a control group were studied. Target SNPs for each gene were analyzed by real time-PCR technique after genomic DNA isolation from maternal blood-EDTA, control group blood-EDTA and spontaneously aborted fetal tissues. Some cases had a single thrombophilic polymorphism, but the rest of the patients and fetal materials had combined thrombophilic polymorphisms. The PAI-1 4G/5G+4G/4G (P= 0.0017), 4G/4G (P= 0.0253), eNOS 894GT+894TT (P=0.0011) genotypes and T allele (P=0.0185), Apo E E3/E4+E3/E2+E2/E4 (P<0.0001) genotypes, E2 (P<0.0001) and E4 (P<0.0001) alleles were higher in spontaneously aborted fetal materials when compared to their mothers and control group. The Factor V Leiden rs6025, Prothrombin G20210A, MTHFR C677T, ACE I/D genotypes were different for each group but not statistically significant due to relatively small size of the samples (P>0.05). Our results indicated that combined thrombophilic gene variations may be associated with increased risk for spontaneous abortions and results need to be confirmed by larger sample size.

No MeSH data available.


Related in: MedlinePlus

Shows the Real-time-PCR profiles of MTHFR C677T genotypes that evaluated for the studied groups. a: Homozygous CC genotype (Tm: 63 0C). b: Heterozygous CT genotype (Tm: 54.5 0C and 63 0C). c: Homozygous TT genotype (Tm: 54.5 0C
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Figure 1: Shows the Real-time-PCR profiles of MTHFR C677T genotypes that evaluated for the studied groups. a: Homozygous CC genotype (Tm: 63 0C). b: Heterozygous CT genotype (Tm: 54.5 0C and 63 0C). c: Homozygous TT genotype (Tm: 54.5 0C

Mentions: Total genomic DNA was isolated from either 10-15 mg of abortion material tissue by the QIAGEN QIAamp DNA Mini Kit or peripheral blood samples containing EDTA from the mothers and control cases by the QIAGEN QIAamp DNA Blood Mini Kit. The polymorphic alleles were amplified by real- time polymerase chain reaction (PCR) technique in Light Cycler 2.0 (Roche, Switzerland) using Light Mix kits (TIB MOBIOL, Germany). Briefly, Light Cycler, Roche Fast Start Master mix, master mix (water, PCR- grade, Mg+2, stock solution, Primer mix and HybProbe mix) and DNA template were used for real-time amplification. The multiple PCR consisted of a denaturation step of 10 min at 95 oC, followed by 45 cycles of 5 s at 95 oC,10 s at 60 oC, and 15 s at 72 oC, and a melting step of 20 s at 95 oC, 20 s at 40 oC, a continuous mode at 85 oC, a cooling step of 30 s at 40 oC for all four regions. A software program (Light Cycler 2.0, Roche, Switzerland) was used for the detection of the mutated and normal genotype profiles of the target gene in the groups included in the study. The wild, heterozygous and homozygous profiles were analysed in channel 640 with melting curve analysis (Figure 1).


Multiple Inherited Thrombophilic Gene Polymorphisms in Spontaneous Abortions in Turkish Population.

Yalcintepe S, Ozdemir O, Hacivelioglu SO, Akurut C, Koc E, Uludag A, Cosar E, Silan F - Int J Mol Cell Med (2015)

Shows the Real-time-PCR profiles of MTHFR C677T genotypes that evaluated for the studied groups. a: Homozygous CC genotype (Tm: 63 0C). b: Heterozygous CT genotype (Tm: 54.5 0C and 63 0C). c: Homozygous TT genotype (Tm: 54.5 0C
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4499574&req=5

Figure 1: Shows the Real-time-PCR profiles of MTHFR C677T genotypes that evaluated for the studied groups. a: Homozygous CC genotype (Tm: 63 0C). b: Heterozygous CT genotype (Tm: 54.5 0C and 63 0C). c: Homozygous TT genotype (Tm: 54.5 0C
Mentions: Total genomic DNA was isolated from either 10-15 mg of abortion material tissue by the QIAGEN QIAamp DNA Mini Kit or peripheral blood samples containing EDTA from the mothers and control cases by the QIAGEN QIAamp DNA Blood Mini Kit. The polymorphic alleles were amplified by real- time polymerase chain reaction (PCR) technique in Light Cycler 2.0 (Roche, Switzerland) using Light Mix kits (TIB MOBIOL, Germany). Briefly, Light Cycler, Roche Fast Start Master mix, master mix (water, PCR- grade, Mg+2, stock solution, Primer mix and HybProbe mix) and DNA template were used for real-time amplification. The multiple PCR consisted of a denaturation step of 10 min at 95 oC, followed by 45 cycles of 5 s at 95 oC,10 s at 60 oC, and 15 s at 72 oC, and a melting step of 20 s at 95 oC, 20 s at 40 oC, a continuous mode at 85 oC, a cooling step of 30 s at 40 oC for all four regions. A software program (Light Cycler 2.0, Roche, Switzerland) was used for the detection of the mutated and normal genotype profiles of the target gene in the groups included in the study. The wild, heterozygous and homozygous profiles were analysed in channel 640 with melting curve analysis (Figure 1).

Bottom Line: For this purpose, the Factor V Leiden (FVL) (rs6025), Prothrombin G20210A (rs1799963), MTHFR C677T (rs1801133), PAI-1 4G/5G (rs1799889), ACE I/D (rs1799752), eNOS E298D (rs1799983), and Apo E E2/E3/E4 (rs429358) polymorphisms were genotyped and correlated in spontaneously aborted fetal materials, their mothers and fertile women.Target SNPs for each gene were analyzed by real time-PCR technique after genomic DNA isolation from maternal blood-EDTA, control group blood-EDTA and spontaneously aborted fetal tissues.Some cases had a single thrombophilic polymorphism, but the rest of the patients and fetal materials had combined thrombophilic polymorphisms.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Genetics, Faculty of Medicine, Canakkale Onsekiz Mart University, Canakkale, Turkey. ; Genetics Diagnosis Center, Numune Training and Education Hospital, Adana, Turkey.

ABSTRACT
The aim of this study was to investigate the possible role of multiple inherited thrombophilic gene variations in women with unexplained spontaneous abortions. For this purpose, the Factor V Leiden (FVL) (rs6025), Prothrombin G20210A (rs1799963), MTHFR C677T (rs1801133), PAI-1 4G/5G (rs1799889), ACE I/D (rs1799752), eNOS E298D (rs1799983), and Apo E E2/E3/E4 (rs429358) polymorphisms were genotyped and correlated in spontaneously aborted fetal materials, their mothers and fertile women. Twenty three abortion materials, 22 women with ≥1 unexplained fetal loss, and 22 control subjects with at least two healthy term infants as a control group were studied. Target SNPs for each gene were analyzed by real time-PCR technique after genomic DNA isolation from maternal blood-EDTA, control group blood-EDTA and spontaneously aborted fetal tissues. Some cases had a single thrombophilic polymorphism, but the rest of the patients and fetal materials had combined thrombophilic polymorphisms. The PAI-1 4G/5G+4G/4G (P= 0.0017), 4G/4G (P= 0.0253), eNOS 894GT+894TT (P=0.0011) genotypes and T allele (P=0.0185), Apo E E3/E4+E3/E2+E2/E4 (P<0.0001) genotypes, E2 (P<0.0001) and E4 (P<0.0001) alleles were higher in spontaneously aborted fetal materials when compared to their mothers and control group. The Factor V Leiden rs6025, Prothrombin G20210A, MTHFR C677T, ACE I/D genotypes were different for each group but not statistically significant due to relatively small size of the samples (P>0.05). Our results indicated that combined thrombophilic gene variations may be associated with increased risk for spontaneous abortions and results need to be confirmed by larger sample size.

No MeSH data available.


Related in: MedlinePlus