• Characterization of Variants in the Hypertension-Associated Gene STK39

      Dorff, Sarah E.; Chang, Y. P. Christy (2012)
      Persistent elevation of blood pressure (BP), termed hypertension (HTN), affects one in three adults in the US and more than one billion individuals worldwide. A genome-wide association study (GWAS) by our group identified intronic sequence variants that show association to systolic BP (SBP) in the gene STK39. The protein encoded by STK39, SPAK, plays a critical role in the maintenance of vascular volume, and physiologic and genetic evidence solidifies the significance of SPAK in BP regulation. Therefore, it is important to identify and characterize all functional variants in the STK39 gene. We sequenced exons and noncoding conserved elements in STK39 in a subset of Amish Family Diabetes Study (AFDS) participants and identified four coding variants. In exon 1, there are three insertion/deletion (in/del) polymorphisms that alter the length of a relatively-uncharacterized proline-alanine rich domain. In exon 11, the nonsynonymous SNP rs56031549 creates a putative novel phosphorylation site by replacing an alanine with a threonine in the regulatory domain of SPAK. These four variants have minor allele frequencies ranging from 0.02 to 0.28 in European Americans (EA). In the AFDS, the 21- and 3-nucleotide in/dels show association to SBP under an additive model (p < 0.05). We also detected suggestive evidence of association between SBP and the 3-nucleotide deletion in another group of Amish subjects (additive model, p = 0.07). Analysis of these variants in the outbred EA population of the GenNet study showed a trend towards significance for rs56031549 to SBP (p = 0.07). Functional characterization based on an in vitro kinase assay revealed that the STK39 haplotype of the 21-nucleotide deletion and 6-nucleotide deletion (9 amino acid deletion) resulted in greater kinase activity that is not attributable to the phosphorylation status of SPAK's active site. Thus, the association between the 21-nucleotide deletion and BP might lead to a variant SPAK protein with greater kinase activity that increases renal sodium reabsorption by cotransporters such as NCC and NKCC2. This finding suggests that pharmaceutical manipulation of SPAK activity may be a novel and effective approach for the treatment of HTN.
    • The effectiveness of patient medication information leaflets for African-American hypertensive patients

      Bradley, Lynette Renee; Beardsley, Robert S. (1999)
      The utilization of patient information leaflets (PILs) for medication has increased tremendously over the past decade. While previous research has demonstrated positive health outcomes for patients using theses educational resources, there has been no such research conducted with a group of chronically ill, minority patients. This study assessed two methods of presenting a PIL to African American hypertensive patients attending an ambulatory care clinic in Baltimore, Maryland. At baseline, patient knowledge regarding a newly prescribed antihypertensive medication, satisfaction with medical care, and adherence to the medication regimen were assessed. Patients were randomized into one of four PIL educational groups: (1) easy-to-read (ETR) PIL; (2) ETR PIL with verbal consultation by a pharmacist; (3) standard PIL; or (4) standard PIL with verbal consultation by a pharmacist. The ETR PILs were written at the 5-7th grade level and the standard PILs were written at the 11th grade level. Patient were contacted 7 days and 3 months after receiving an intervention. Level of knowledge about the medication was assessed at the 7 day and 3 month interviews. Patients were asked about their medical care satisfaction and adherence at the three month interview. Medical appointment adherence and blood pressure control were assessed from the medical record measurements for the six month period pre-intervention and post-intervention. Ninety-seven patients were approached to enter the study. Ninety-two patients were recruited and six were lost to follow-up. Based on a maximum score of 6, the ETR PIL group knowledge scores were 5.00 (baseline), 5.25 (7-day), and 5.45 (3 months); the ETR + verbal group scores were 4.91, 5.26, and 5.64; the standard PIL group scores were 4.76, 5.28, and 5.40; and the standard + verbal group scores were 5.04, 5.59, and 5.41, respectively. Although not statistically significant, patient satisfaction scores (maximum of 70) improved for all groups increasing from an average of 53.7 to 55.1, with the exception of the standard + verbal group whose average decreased from 54.5 to 53.8. The medication adherence scores (maximum of 5) improved for the ETR groups by 0.49 and decreased for the standard PIL groups by 0.13, % = 1.93, p = 0.05. Medical appointment adherence decreased for all groups from 92.7% to 89.6%. Patient blood pressure measurements were reduced for the entire sample. There was a significant effect of an ETR PIL in the change of diastolic blood pressure (T = -1.80, p = 0.05) and systolic blood pressure (T = -1.70, p = 0.05) when gender, type of antihypertensive prescribed, adherence with taking the antihypertensive medication, and time diagnosed with hypertensive were controlled for in the statistical model. This research attempted to examine the effect of presenting a PIL, with and without verbal consultation, to a group of patients with a chronic illness. The study suggests that the ETR groups and the PIL + verbal groups tended to have better improvements than the standard PIL and PIL only groups in the patient outcomes studied. This research illustrates that ETR PILs and/or PILs with verbal counseling may contribute to enhanced patient outcomes for a group of inner-city African American hypertensive patients.
    • A physiologic and molecular characterization of an animal model of salt-sensitive arterial hypertension

      Lighthall, Geoffrey Kenton (1993)
      Over 60 million Americans are afflicted with essential hypertension--a potentially dangerous elevation of arterial blood pressure. Although the etiology of this disorder is largely unknown, two factors are of undisputed importance: a permissive genetic background and the prolonged ingestion of salt in amounts greater than 50 meq/day. The salt-sensitive (S) and salt resistant (R) rats developed by Lewis Dahl provide a convenient genetic model to address hypertension experimentally and have been used extensively to study various physiologic systems mediating salt-sensitive arterial hypertension. The studies to be presented were based on the proposition that the comparative physiology of the Dahl rat model of hypertension might suggest new pathways toward identifying genes responsible for the hypertensive phenotype, and that such are worthy of exploration. As the renal handling of sodium and water plays a primary role in the pathogenesis of Dahl hypertension, subtractive and differential hybridization methods were used to identify kidney messages expressed at a higher rate in either the R normotensive or S hypertensive rat. Messenger RNA expression was addressed quantitatively by dot blot, northern blot, and RNase protection assays in the two strains of rat for recombinant phage appearing to bear differentially expressed messages. Likewise, expression of messages for a number of known genes whose physiologic role suggests an involvement in hypertension were also evaluated. After screening {dollar}{lcub}>{rcub}10\sp4{dollar} plaques (from both S and R kidney cDNA libraries) by different experimental paradigms, over fifty messages appeared to be differentially expressed. A second round of quantitative dot blot analysis narrowed the field to six clones showing significant differences in expression between S and R rats. These latter clones have been sequenced resulting in the identification of several previously undescribed cDNAs. The temporal nature of their expression in the kidneys of S and R rats following salt loading has been partially evaluated. Future work will characterize the genes encoding these differentially expressed cDNAs and examine their linkage to hypertension in segregating populations.
    • Primary Prevention of Atrial Fibrillation Using Renin-Angiotensin-Aldosterone System Inhibitors among Medicare Beneficiaries with Hypertension

      Yin, Xianghua; Zhan, Min (2016)
      Statement of the Problem: Renin-angiotensin-aldosterone-system inhibitors (RAASIs) have long been associated with reduced risk of new-onset AF in patients with hypertension. However, previous studies have not properly accounted for the presence of competing risks in the usual care setting. Hypothesis: The study was designed to determine the effects of RAASIs on the hazard and cumulative incidence (sub-distribution hazard) of newly documented AF and to test whether the effects were significantly different from those of beta-blockers (BBs) or calcium-channel blockers (CCBs). Methods: A propensity score (PS)-matched retrospective cohort study was conducted in a random 5% sample of the Medicare beneficiary population from 2007 to 2011 with hypertension treated with antihypertensive drug monotherapy, consisting of 50,307 beneficiaries. Beneficiaries on RAASI-based monotherapies were matched 1:1 with beneficiaries on BB-based monotherapies (n=13,242) and CCB-based monotherapies (n=10,843) based on PS. All beneficiaries were free of baseline AF and compelling indications for RAASIs. Competing risk analyses were performed. Cox proportional cause-specific hazard regression was used to estimate the effects of RAASIs on newly documented AF and all-cause mortality without AF (i.e., competing risks). Fine-Gray Models were used to examine whether there was a significant difference in the cumulative incidence of newly documented AF between beneficiaries treated with RAASIs and BBs/CCBs, accounting for all-cause mortality without AF as competing risks. Results: The adjusted cause-specific hazard ratio (95% confidence interval [CI]) in the RAASI vs. BB groups was 0.69 (95% CI: 0.58 to 0.81) for newly documented AF. The adjusted sub-distribution hazard ratio (95% CI) in the RAASI vs. BB groups was 0.69 for newly documented AF (95% CI: 0.59 to 0.81). The adjusted cause-specific hazard ratio (95% CI) in the RAASI vs. CCB groups was 0.55 (95% CI: 0.46 to 0.66) for newly documented AF. The adjusted sub-distribution hazard ratio (95% CI) in the RAASI vs. CCB groups was 0.54 for newly documented AF (95% CI: 0.45 to 0.65). Conclusions: RAASI-based monotherapies were associated with not only a reduced hazard of newly documented AF but also with a reduced sub-distribution hazard of newly documented AF for Medicare beneficiaries with hypertension enrolled in the Part D program.
    • Transcriptional and Post-Transcriptional Regulation of Genes Critical for Sodium Reabsorption and Blood Pressure Control in the Kidney

      Mercado, Carlo Jose Santos; Chang, Y. P. Christy (2017)
      Serine/threonine kinases (WNK1, WNK4, SPAK, OSR1) and cation co-transporters (NKCC2, NCC) are members of a multi-kinase network that determines renal Na+ reabsorption and blood pressure (BP) regulation. The importance of these proteins is highlighted by their roles in monogenic forms of hyper- and hypotension, animal models, and by the efficacy of BP-lowering medications that target this pathway. While post-translational regulation of these proteins has been well established, regulation of these genes at the transcript level is not completely understood. In this study, we examined both human and mouse kidney transcriptomes to uncover novel transcriptional and post-transcriptional regulation of two genes in this pathway. First, STK39 encodes for Ste20-related proline alanine rich kinase (SPAK), which phosphorylates and activates cation co-transporters. Variants within STK39 are associated with susceptibility to essential hypertension, and SPAK null mice are hypotensive and mimic Gitelman syndrome, a rare monogenic salt-wasting human disorder. Mice exhibit nephron segment-specific expression of full length SPAK and N-terminally truncated SPAK isoforms with impaired kinase function. We established that while humans also express transcript isoforms similar to those found in mice, they differ in abundance and are transcribed from human-specific promoters. Second, SLC12A3 encodes for the thiazide-sensitive Na+-Cl- co-transporter (NCC), and rare mutations in this gene cause Gitelman syndrome. In humans and mice, alternative polyadenylation of NCC pre-mRNA results in a longer 3'UTR isoform, while alternative splicing within the final exon leads to an exon-exon junction downstream of the termination codon. Both of these events generate potential substrates for nonsense-mediated mRNA decay (NMD). By suppressing NMD, we demonstrated that NCC transcript abundance is partially determined by post-transcriptional processing of its final exon. Finally, dietary K+ manipulation differentially alters SPAK and NCC transcript and protein isoform abundance, demonstrating dynamic physiological regulation of gene expression in response to salt reabsorptive needs. In summary, genes in this pathway undergo complex transcriptional and post-transcriptional regulation, resulting in the differential expression of novel alternative transcripts that contribute to the fine-tuning of BP control.