Dataset Information

Impairment of Flow-Sensitive Inwardly Rectifying K⁺ Channels via Disruption of Glycocalyx Mediates Obesity-Induced Endothelial Dysfunction.

ABSTRACT:

Objective

To determine if endothelial dysfunction in a mouse model of diet-induced obesity and in obese humans is mediated by the suppression of endothelial Kir (inwardly rectifying K⁺) channels. Approach and Results: Endothelial dysfunction, observed as reduced dilations to flow, occurred after feeding mice a high-fat, Western diet for 8 weeks. The functional downregulation of endothelial Kir2.1 using dominant-negative Kir2.1 construct resulted in substantial reductions in the response to flow in mesenteric arteries of lean mice, whereas no effect was observed in arteries of obese mice. Overexpressing wild-type-Kir2.1 in endothelium of arteries from obese mice resulted in full recovery of the flow response. Exposing freshly isolated endothelial cells to fluid shear during patch-clamp electrophysiology revealed that the flow-sensitivity of Kir was virtually abolished in cells from obese mice. Atomic force microscopy revealed that the endothelial glycocalyx was stiffer and the thickness of the glycocalyx layer reduced in arteries from obese mice. We also identified that the length of the glycocalyx is critical to the flow-activation of Kir. Overexpressing Kir2.1 in endothelium of arteries from obese mice restored flow- and heparanase-sensitivity, indicating an important role for heparan sulfates in the flow-activation of Kir. Furthermore, the Kir2.1-dependent component of flow-induced vasodilation was lost in the endothelium of resistance arteries of obese humans obtained from biopsies collected during bariatric surgery.

Conclusions

We conclude that obesity-induced impairment of flow-induced vasodilation is attributed to the loss of flow-sensitivity of endothelial Kir channels and propose that the latter is mediated by the biophysical alterations of the glycocalyx.

SUBMITTER: Fancher IS

PROVIDER: S-EPMC7503211 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Background/aimsShear stress plays major roles in developmental angiogenesis, particularly in blood vessel remodeling and maturation but little is known about the shear stress sensors involved in this process. Our recent study identified endothelial Kir2.1 channels as major contributors to flow-induced vasodilation, a hallmark of the endothelial flow response. The goal of this study is to establish the role of Kir2.1 in the regulation of retinal angiogenesis.MethodsThe retina of newly born Kir2.1+/- mice were used to investigate the sprouting angiogenesis and remodeling of newly formed branched vessels. The structure, blood density and mural cell coverage have been evaluated by immunohistochemistry of the whole-mount retina. Endothelial cell alignment was assessed using CD31 staining. The experiments with flow-induced vasodilation were used to study the cerebrovascular response to flow.ResultsUsing Kir2.1-deficient mice, we show that the retinas of Kir2.1+/- mice have higher vessel density, increased lengths and increased number of the branching points, as compared to WT littermates. In contrast, the coverage by αSMA is decreased in Kir2.1+/- mice while pericyte coverage does not change. Furthermore, to determine whether deficiency of Kir2.1 affects vessel pruning, we discriminated between intact and degraded vessels or "empty matrix sleeves" and found a significant reduction in the number of empty sleeves on the peripheral part of the retina or "angiogenic front" in Kir2.1+/- mice. We also show that Kir2.1 deficiency results in decreased endothelial alignment in retinal endothelium and impaired flow-induced vasodilation of cerebral arteries, verifying the involvement of Kir2.1 in shear-stress sensing in retina and cerebral circulation.ConclusionThis study shows that shear-stress sensitive Kir2.1 channels play an important role in pruning of excess vessels and vascular remodeling during retinal angiogenesis. We propose that Kir2.1 mediates the effect of shear stress on vessel maturation.

Project description:Background: Endurance athletes are prone to bradyarrhythmias, which in the long-term may underscore the increased incidence of pacemaker implantation reported in this population. Our previous work in rodent models has shown training-induced sinus bradycardia to be due to microRNA (miR)-mediated transcriptional remodeling of the HCN4 channel, leading to a reduction of the "funny" (I f) current in the sinoatrial node (SAN). Objective: To test if genetic ablation of G-protein-gated inwardly rectifying potassium channel, also known as I KACh channels prevents sinus bradycardia induced by intensive exercise training in mice. Methods: Control wild-type (WT) and mice lacking GIRK4 (Girk4 -/-), an integral subunit of I KACh were assigned to trained or sedentary groups. Mice in the trained group underwent 1-h exercise swimming twice a day for 28 days, 7 days per week. We performed electrocardiogram recordings and echocardiography in both groups at baseline, during and after the training period. At training cessation, mice were euthanized and SAN tissues were isolated for patch clamp recordings in isolated SAN cells and molecular profiling by quantitative PCR (qPCR) and western blotting. Results: At swimming cessation trained WT mice presented with a significantly lower resting HR that was reversible by acute I KACh block whereas Girk4 -/- mice failed to develop a training-induced sinus bradycardia. In line with HR reduction, action potential rate, density of I f, as well as of T- and L-type Ca2+ currents (I CaT and I CaL ) were significantly reduced only in SAN cells obtained from WT-trained mice. I f reduction in WT mice was concomitant with downregulation of HCN4 transcript and protein, attributable to increased expression of corresponding repressor microRNAs (miRs) whereas reduced I CaL in WT mice was associated with reduced Cav1.3 protein levels. Strikingly, I KACh ablation suppressed all training-induced molecular remodeling observed in WT mice. Conclusion: Genetic ablation of cardiac I KACh in mice prevents exercise-induced sinus bradycardia by suppressing training induced remodeling of inward currents I f, I CaT and I CaL due in part to the prevention of miR-mediated transcriptional remodeling of HCN4 and likely post transcriptional remodeling of Cav1.3. Strategies targeting cardiac I KACh may therefore represent an alternative to pacemaker implantation for bradyarrhythmias seen in some veteran athletes.

Dataset Information

Impairment of Flow-Sensitive Inwardly Rectifying K+ Channels via Disruption of Glycocalyx Mediates Obesity-Induced Endothelial Dysfunction.

Objective

Conclusions

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Impairment of Flow-Sensitive Inwardly Rectifying K⁺ Channels via Disruption of Glycocalyx Mediates Obesity-Induced Endothelial Dysfunction.