Regulatory control of the Na-Cl co-transporter NCC and its therapeutic potential for hypertension

This is the author accepted manuscript. The final version is available from Elsevier via the DOI in this record.Hypertension is the largest risk factor for cardiovascular disease, the leading cause of mortality worldwide. As blood pressure regulation is influenced by multiple physiological systems, hypertension cannot be attributed to a single identifiable etiology. Three decades of research into Mendelian forms of hypertension implicated alterations in the renal tubular sodium handling, particularly the distal convoluted tubule (DCT)-native, thiazide-sensitive Na–Cl cotransporter (NCC). Altered functions of the NCC have shown to have profound effects on blood pressure regulation as illustrated by the over activation and inactivation of the NCC in Gordon’s and Gitelman syndromes respectively. Substantial progress has uncovered multiple factors that affect the expression and activity of the NCC. In particular, NCC activity is controlled by phosphorylation/dephosphorylation, and NCC expression is facilitated by glycosylation and negatively regulated by ubiquitination. Studies have even found parvalbumin to be an unexpected regulator of the NCC. In recent years, there have been considerable advances in our understanding of NCC control mechanisms, particularly via the pathway containing the with-no-lysine [K] (WNK) and its downstream target kinases, SPS/Ste20-related proline-alanine-rich kinase (SPAK) and oxidative stress responsive 1 (OSR1), which has led to the discovery of novel inhibitory molecules. This review summarizes the currently reported regulatory mechanisms of the NCC and discusses their potential as therapeutic targets for treating hypertension.National Institutes of HealthUniversity of Exeter Medical Schoo

Targeting the WNK-SPAK/OSR1 Pathway and Cation-Chloride Cotransporters for the Therapy of Stroke

This is the final version. Available on open access from MDPI via the DOI in this recordData Availability Statement: Not applicable.Stroke is one of the major culprits responsible for morbidity and mortality worldwide, and the currently available pharmacological strategies to combat this global disease are scanty. Cation-chloride cotransporters (CCCs) are expressed in several tissues (including neurons) and extensively contribute to the maintenance of numerous physiological functions including chloride homeostasis. Previous studies have implicated two CCCs, the Na+-K+-Cl− and K+-Cl− cotransporters (NKCCs and KCCs) in stroke episodes along with their upstream regulators, the with-no-lysine kinase (WNKs) family and STE20/SPS1-related proline/alanine rich kinase (SPAK) or oxidative stress response kinase (OSR1) via a signaling pathway. As the WNK-SPAK/OSR1 pathway reciprocally regulates NKCC and KCC, a growing body of evidence implicates over-activation and altered expression of NKCC1 in stroke pathology whilst stimulation of KCC3 during and even after a stroke event is neuroprotective. Both inhibition of NKCC1 and activation of KCC3 exert neuroprotection through reduction in intracellular chloride levels and thus could be a novel therapeutic strategy. Hence, this review summarizes the current understanding of functional regulations of the CCCs implicated in stroke with particular focus on NKCC1, KCC3, and WNK-SPAK/OSR1 signaling and discusses the current and potential pharmacological treatments for strokeCommonwealth PhD ScholarshipUniversity of ExeterNational Institutes of Health (NIH

Angiotensin II Receptor Blockers in the Management of Hypertension in Preventing Cognitive Impairment and Dementia - A Systematic Review

This is the final version. Available on open access from MDPI via the DOI in this recordHypertension is a known risk factor for cognition-related pathologies including dementia. The National Institute of Health and Care Excellence (NICE) guidelines recommend angiotensin (Ang) II receptor blockers (ARBs) or angiotensin-converting enzyme inhibitors (ACEIs) as a first-line treatment for hypertension. Although both ARBs and ACEIs show neuroprotective effects, ACEIs show contradictory side effects; therefore, ARBs may be a more viable option. However, trials assessing the effects of ARBs on cognition are scarce and conflicting. Therefore, the aim of this review is to conduct a systematic review and synthesise data on the influence of ARBs on cognition and dementia prevention. Five databases were searched from 1992-2022 to produce 13 randomised controlled trials (RCTs) involving 26,907 patients that compared associations of ARBs against placebos or other antihypertensives on cognition or probable dementia with a minimum duration of 3 months. ARBs showed greater cognitive benefits when compared to hydrochlorothiazide (HCTZ), beta blockers (BB), and ACEIs. Our findings showed that although ARBs are superior to some antihypertensives such as ACEIs, thiazide and beta blockers, they made no difference in comparison to the placebo in all but one sample of patients. The positive effects on cognitive performances are equal to calcium channel blockers (CCBs) and lower than statin. The neuroprotective effects of ARBs are also more beneficial when ARBs are taken at the same time as a statin. Due to these inconsistencies, robust conclusions cannot be made. Future trials are warranted and, if successful, could have positive economic implications and consequently improve quality of life.National Natural Science Foundation of ChinaAlzheimer’s Research UKRoyal Societ

WNK-SPAK/OSR1-NCC kinase signaling pathway as a novel target for the treatment of salt-sensitive hypertension

This is the author accepted manuscript. The final version is available from Wiley via the DOI in this record.Hypertension is the most prevalent health condition worldwide, affecting ~1 billion people. Gordon’s syndrome is a form of secondary hypertension that can arise due to a number of possible mutations in key genes that encode proteins in a pathway containing the With No Lysine [K] (WNK) and its downstream target kinases, SPS/Ste20-related proline-alanine-rich kinase (SPAK) and oxidative stress responsive kinase 1 (OSR1). This pathway regulates the activity of the thiazide-sensitive sodium chloride cotransporter (NCC), which is responsible for NaCl reabsorption in the distal nephron. Therefore, mutations in genes encoding proteins that regulate the NCC proteins disrupt ion homeostasis and cause hypertension by increasing NaCl reabsorption. Thiazide diuretics are currently the main treatment option for Gordon’s syndrome. However, they have a number of side effects, and chronic usage can lead to compensatory adaptations in the nephron that counteract their action. Therefore, recent research has focused on developing novel inhibitory molecules that inhibit components of the WNK-SPAK/OSR1-NCC pathway, thereby reducing NaCl reabsorption and restoring normal blood pressure. In this review we provide an overview of the currently reported molecular inhibitors of the WNK-SPAK/OSR1-NCC pathway and discuss their potential as treatment options for Gordon’s syndrome.University of ExeterNational Natural Science Foundation of Chin

Study of the functions and activities of neuronal K-Cl co-transporter KCC2 using western blotting

This is the final version. Available from MyJove Corporation via the DOI in this recordPotassium chloride cotransporters 2 (KCC2) is a member of the solute carrier family 12 (SLC12) of cation-chloride-cotransporters (CCCs), found exclusively in the neuron and is essential for the proper functioning of Cl- homeostasis and consequently functional GABAergic inhibition. Failure in proper regulation of KCC2 is deleterious and has been associated with the prevalence of several neurological diseases, including epilepsy. There has been considerable progress with regard to understanding the mechanisms involved in the regulation of KCC2, accredited to the development of techniques that enable researchers to study its functions and activities; either via direct (assessing kinase regulatory sites phosphorylation) or indirect (observing and monitoring GABA activity) investigations. Here, the protocol highlights how to investigate KCC2 phosphorylation at kinase regulatory sites - Thr906 and Thr1007- using western blotting technique. There are other classic methods used to directly measure KCC2 activity, such as rubidium ion and thallium ion uptake assay. Further techniques such as patch-clamp-electrophysiology are used to measure GABA activity; hence, indirectly reflecting activated and/or inactivated KCC2 as informed by the assessment of intracellular chloride ion homeostasis. A few of these additional techniques will be briefly discussed in this manuscript.Royal Society (Charity)National Institutes of Healt

NF-κB Signaling-Mediated Activation of WNK-SPAK-NKCC1 Cascade in Worsened Stroke Outcomes of Ang II-Hypertensive Mice

This is the author accepted manuscript. The final version is available from Lippincott, Williams & Wilkins via the DOI in this recordData availability: The data that support the findings of this study are available within the article and its Data Supplement.BACKGROUND: Worsened stroke outcomes with hypertension comorbidity are insensitive to blood pressure-lowering therapies. In an experimental stroke model with comorbid hypertension, we investigated causal roles of ang II (angiotensin II)-mediated stimulation of the brain WNK (with no lysine [K] kinases)-SPAK (STE20/SPS1-related proline/alanine-rich kinase)-NKCC1 (Na-K-Cl cotransporter) complex in worsened outcomes. METHODS: Saline- or ang II-infused C57BL/6J male mice underwent stroke induced by permanent occlusion of the distal branches of the middle cerebral artery. Mice were randomly assigned to receive either vehicle dimethyl sulfoxide/PBS (2 mL/kg body weight/day, IP), a novel SPAK inhibitor, 5-chloro-N-(5-chloro-4-((4-chlorophenyl)(cyano)methyl)-2-methylphenyl)-2-hydroxybenzamide (ZT-1a' 5 mg/kg per day, IP) or a NF-κB (nuclear factor-κB) inhibitor TAT-NBD (transactivator of transcription-NEMO-binding domain' 20 mg/kg per day, IP). Activation of brain NF-κB and WNK-SPAK-NKCC1 cascade as well as ischemic stroke outcomes were examined. RESULTS: Stroke triggered a 2- to 5-fold increase of WNK (isoforms 1, 2, 4), SPAK/OSR1 (oxidative stress-responsive kinase 1), and NKCC1 protein in the ang II-infused hypertensive mouse brains at 24 hours after stroke, which was associated with increased nuclear translocation of phospho-NF-κB protein in the cortical neurons (a Pearson correlation r of 0.77, P<0.005). The upregulation of WNK-SPAK-NKCC1 cascade proteins resulted from increased NF-κB recruitment on Wnk1, Wnk2, Wnk4, Spak, and Nkcc1 gene promoters and was attenuated by NF-κB inhibitor TAT-NBD. Poststroke administration of SPAK inhibitor ZT-1a significantly reduced WNK-SPAK-NKCC1 complex activation, brain lesion size, and neurological function deficits in the ang II-hypertensive mice without affecting blood pressure and cerebral blood flow. CONCLUSIONS: The ang II-induced stimulation of NF-κB transcriptional activity upregulates brain WNK-SPAK-NKCC1 cascade and contributes to worsened ischemic stroke outcomes, illustrating the brain WNK-SPAK-NKCC1 complex as a therapeutic target for stroke with comorbid hypertension.Veteran AffairsNational Institutes of Health (NIH)AH