Essential Hypertension (EH) is a medical condition characterized by persistent high blood pressure, and it is one of the most significant public health problems globally, causing approximately 9.4 million deaths annually. The prevalence of EH varies according to genetic ancestry and affects differently specific populations, with 17% in the Americas, 19.2% in the Western Pacific, 23.2% in Europe, 25.1% in Southeast Asia, 26.3% in the Eastern Mediterranean, and 27.2% in Africa. EH is a multifactorial disease, that is, it is determined by genetic factors and influenced by environmental factors. Although genetic factors are estimated to contribute to around 30-60% of the variation in blood pressure, the genetic complexity of hypertension is not yet fully understood due to the limited knowledge of candidate genes, susceptibility loci, and population-specific differences. Various approaches such as candidate gene-based, genome-wide linkage analysis (GWLA), and family- and population-based genome-wide association studies (GWAS) have been used to identify genetic factors contributing to EH, but part of the heritability of blood pressure-related phenotypes is not explained by the genetic factors known so far, mainly due to methodological limitations. The main objective of this study was to investigate the genetic basis of EH by mapping regions of interest (ROI) and investigate candidate genes and variants influencing the Essential Hypertension in African-derived individuals from partially isolated populations of quilombo remnants in Vale do Ribeira (Sao Paulo - Brazil), which were previously well characterized from the clinical, genealogical and population genetics point of view. Samples from 431 individuals (167 affected, 261 unaffected and 3 with an unknown phenotype) from eight quilombo remnants populations were genotyped using SNP array with approximately 650.000 SNPs. The global ancestry proportions of these populations were estimated to be 47%, 36%, and 16% for African, European, and Native American ancestries, respectively. In addition, genealogical information from 673 individuals was used to construct six pedigrees comprising a total of 1104 individuals. The mapping strategy consisted of a multi-level computational approach. Pedigrees were constructed (GenoPro v.3) based on interviews and kinship coefficient (King v.2.2, MORGAN v.3.4 and PBAP v.1). The dataset was pruned (King v.2.2 and PBAP v.1) to obtain three non-overlapping markers subpanels (PBAP v.2). Haplotype phasing and local ancestry (SNPFlip v.0.0.6, SHAPEIT2 and RFMIX v.2) were performed to obtain SNP allele frequency (ADMIXFRQ v.1) to account for admixture. Genome-wide and dense linkage analyses were performed using the three subpanels of markers (MORGAN v.3.4). Finally, we performed fine-mapping using family-based association studies (GENESIS v.2.24) based on population (MINIMAC v.4) and pedigree (GIGI2) imputed data, and EH-related genes and variant investigation (relying on publicly available databases). The linkage analysis strategy resulted in the mapping of 22 ROIs with LOD score 1.45, containing markers co-segregating with the phenotype. The LOD score range was 1.45-3.03 considering all the linked segments and these 22 ROIs encompassed 2363 genes. As our first fine-mapping strategy, we identified 60 EH-related genes as potential candidates to contribute to high blood pressure in quilombo remnants pedigrees. In addition, as our second fine-mapping strategy, we identified 118 suggestive or significant variants through family-based association studies. Considering only the common results between the two strategies, we found that 14 genes - PHGDH and S100A10 (ROI1), MFN2 (ROI2), RYR2, EDARADD and MTR (ROI3), SERTAD2 (ROI4), LPP (ROI5), KCNT1 (ROI11), TENM4 (ROI13), P2RX1, ZZEF1 and RPA1 (ROI18), and ALPK2 (ROI20) - were identified within the mapped regions with strong and sufficient evidence in the literature attesting relatedness with the phenotype. These genes harbor 29 SNPs that were either within them or very close, and they were pointed out by family-based association studies as showing suggestive or significant association with hypertension. We also identified 46 other genes within the mapped regions, but with less evidence of relatedness to the phenotype since they were not replicated through family-based association studies. Overall, the results obtained in this study revealed, through a complementary approach - combining admixture-adjusted genome-wide linkage analysis based on Markov chain Monte Carlo (MCMC) methods, association studies on imputed data, and in silico investigations - genetic regions, variants and candidate genes that shed light on the genetic basis of essential hypertension. These genes are responsible for encoding proteins that play crucial roles in regulating blood pressure, including the regulation of sodium and potassium levels, and the aldosterone pathway, among others. Our findings reveal genes and variants with distinct potential to explain the genetic etiology of essential hypertension observed in quilombo remnant populations.

The authors have declared no competing interest.

We acknowledge funding from CEPID-FAPESP (Research Center on the Human Genome and Stem Cells, grants 1998/14254-2 and 2013/08028-1, and FAPESP/INCT-CNPq 2014/50931-3 led by Dr. Mayana Zatz). This research also received support from grant FAPESP 2012/18010-0, titled Balancing selection in the human genome: detection, causes, and consequences, led by Dr. Diogo Meyer (IB/USP-Brazil). Additionally, we are grateful to CAPES for the sandwich Ph.D. fellowship #88887.371219/2019-00 and CNPq for the Ph.D. fellowship #142193/2017-8. We also acknowledge funding from the Marshall University Joan C. Edwards School of Medicine, WV-INBRE grant (NIH P20GM103434), Bench-to-Bedside Pilot grant (Nato) under the West Virginia Clinical and Translational Science Institute (WV-CTSI) grant (NIH 5U54GM104942), and Nato startup fund.

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Ethics committee/IRB of University of Sao Paulo (Brazil)/Institute of Biomedical Sciences gave ethical approval for this work under the protocol number 111/2001. Ethics committee/IRB of University of Sao Paulo (Brazil)/Institute of Biosciences gave ethical approval for this work under the protocol number 012/2004 and 034/2005.

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