Chronic venous disease (CVD) is one of the most prevalent yet underrated disorders worldwide. High heritability estimates of CVD indicate prominent genetic components in its etiology and pathology. Mutations in human forkhead box C2 (FoxC2) gene are strongly associated with valve failure in saphenous and deep veins of lower extremities. We explored the association of genetic variants of FoxC2 as well as FoxC2 mRNA and protein expression levels with CVD of lower limbs. We systematically sequenced the single coding exon, 5′ and 3′ flanking regions of FoxC2 gene in 754 study subjects which includes 382 patients with CVD and 372 healthy subjects. Four novel and three reported polymorphisms were identified in our cohort. Three variants in 5′ flanking region and one in 3′ flanking region of FoxC2 gene were significantly associated with CVD risk. FoxC2 mRNA in vein tissues from 22 patients was 4±1.42 fold increased compared to saphenous veins from 20 normal subjects (p<0.01). FoxC2 protein was also significantly upregulated in varicose veins compared to control samples. The c.-512C>T (rs34221221: C>T) variant which is located in the FoxC2 putative promoter region was further analyzed. Functional analysis of c.-512C>T revealed increased mRNA and protein expression in patients with homozygous TT genotype compared to heterozygous CT and wild CC genotypes. Luciferase assay indicated higher transcriptional activity of mutant compared to wild genotype of this variant. These findings suggested that c.-512C>T variant of FoxC2 was strongly associated with susceptibility to CVD and also that this variant resulted in FoxC2 overexpression. To obtain a mechanistic insight into the role of upregulated FoxC2 in varicosities, we overexpressed FoxC2 in venous endothelial cells and observed elevated expression of arterial markers Dll4 and Hey2 and downregulation of venous marker COUP-TFII. Our study indicates altered FoxC2-Notch signaling in saphenous vein wall remodeling in patients with varicose veins.

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Phone surveys are a rapid and cost-effective way to collect primary data for research, monitoring and evaluation purposes. But for these data to be precise, reliable and unbiased, women’s perspectives must be accurately represented. Throughout 2020, we conducted seven household surveys in rural India to understand households’ experiences of the COVID-19 pandemic and contemporaneous relief programmes. Given social distancing protocols, we conducted these surveys over the phone, using household phone numbers collected during earlier, face-to-face research. Analysing metadata from these surveys (along with women’s responses to questions about phone use), we determine how gaps in phone access inhibit women’s representation in phone surveys. We find that the prevalence of male management of household phones significantly reduces access to female respondents. This is a problem for two reasons. Firstly, men are usually the first to pick up the phone: in two surveys in which we tracked the gender of the person who picked up, men picked up 63.2% and 71.1% of the time, respectively. Moreover, only a small minority of those we reached by phone were able and willing to pass the phone to a household member of the opposite gender, when prompted (with no statistically significant difference between pass rates for women and men). This low immediate pass rate, in combination with low female pickup, led to fewer women respondents. As such, we recommend that researchers dedicate time and resources to taking appointments and making call-backs to reach more women. We also show that the use of female enumerators improves households’ willingness to participate in women-centred surveys, and call for more investment into female enumerator teams.

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