Vascular endothelial growth factor (VEGF) is a pro-angiogenic factor. Variability in VEGF expression, induced by specific VEGFA variants, are involved in angiogenesis-related disorders. This study examined the genotype distribution and functional role (VEGF expression) of rs699947, rs833061, rs1570360, rs2010963, rs833068, rs833070, rs3025020, and rs3025039 VEGFA variants and their haplotypes in 519 healthy Bahraini individuals of both genders. The distribution of the eight VEGFA polymorphisms screened was in Hardy-Weinberg equilibrium. The minor allele frequencies of rs699947 (0.42), rs833061 (0.32), rs1570360 (0.31), rs2010963 (0.33), rs833068 (0.37), rs833070 (0.42), rs3025020 (0.33), and rs3025039 (0.13) were generally compared to those established for Caucasians. Of the variants tested, rs3025020 was associated with increased VEGF serum levels (p=0.019), while rs3025039 was associated with decreased levels (p=0.038). Linkage analysis identified two VEGFA blocks, the first, spanning 16 kb, was not associated with altered VEGF levels, while the second, spanning 3 kb containing rs3025020 and rs3025039, was linked with higher VEGF expression, of which the (-583)T/(+936)T haplotype (p=0.008) was linked with higher VEGF levels compared to the (-583)C/(+936)C (all wild-type) haplotype. These results support the association of rs30250202 and rs3025039, and specific VEGF haplotypes, with altered VEGF serum levels, although the exact functional mechanisms remain to be elucidated.

STUDY QUESTION Is recurrent spontaneous miscarriage (RSM) associated with changes in vascular endothelial growth factor (VEGF) serum levels, and with polymorphisms in the VEGFA gene? SUMMARY ANSWER Reduced serum VEGF levels, and VEGFA −460T/C (rs833061), 398G/A (rs833068), −583T/C (rs3025020) variants, were associated with RSM. WHAT IS KNOWN ALREADY Reduced expression of VEGF has been linked with spontaneous miscarriage, likely due to defective fetal and placental angiogenesis. Since VEGF production is in part inherited, VEGFA polymorphisms associated with altered VEGF secretion have been investigated for their association with RSM, often with variable conclusions. STUDY DESIGN, SIZE, DURATION A retrospective case–control study, which was conducted between January 2011 and April 15, 2012. PARTICIPANTS/MATERIALS, SETTING, METHODS Subjects comprised 296 women with RSM (mean age: 31.6 ± 5.4 year), and 305 age-matched (mean age: 31.6 ± 4.9 year) control Arab women, who had attended outpatient obstetrics and gynecology clinics in two teaching hospitals in Bahrain. VEGFA −2578C/A (rs699947), −460T/C (rs833061), −1154G/A (rs15703060), −634G/C (rs2010963), 398G/A (rs833068), 497G/A (rs833070), −583T/C (rs3025020) and 936C/T (rs3025039) genotyping was done by real-time PCR, with defined clusters; VEGF serum levels were measured by ELISA. MAIN RESULTS AND THE ROLE OF CHANCE Higher minor allele frequency (MAF) and genotype distribution of −460T/C [corrected P (P_c) = 0.003], 398G/A (P_c = 0.016) and −583T/C (P_c < 0.001) single nucleotide polymorphisms (SNPs) were seen in RSM cases than control women. Increased RSM risk was seen with homozygous −460T/C and 398G/A SNPs and with heterozygous −583T/C, which had a stronger effect when homozygous. Serum VEGF levels were significantly reduced in RSM cases compared with control women (P = 0.016), and correlated with −460T/C, 398G/A and −583T/C genotypes. Haploview analysis revealed heterogeneity in linkage disequilibrium between VEGFA variants, and two blocks were identified: Block 1 comprising −2578C/A, −460T/C and −1154G/A, while Block 2 contained −634G/C, 398G/A, 497G/A, −583T/C and 936C/T. Both negatively and positively RSM-associated 3-locus (Block 1) and 5-locus (Block 2) VEGFA haplotypes were identified, after controlling for a number of covariates. LIMITATIONS, REASONS FOR CAUTION The study was retrospective and can only demonstrate association and not a cause–effect relationship. Furthermore, it was limited to Bahraini Arabs,thereby necessitating parallel studies on other ethnic groups. WIDER IMPLICATIONS OF THE FINDINGS Reduced VEGF secretion, and specific VEGFA variants may contribute to the pathogenesis of RSM. However, the association of VEGFA SNPs with RSM appears to be independent of their association with altered VEGF serum levels. The differential association of VEGFA variants with RSM is in line with previous findings on the contribution of ethnicity/racial background to genetic association studies.