Alteration of Piezo1 signaling in type 2 diabetic mice: focus on endothelium and BKCa channel

Pflügers Archiv - European Journal of Physiology https://doi.org/10.1007/s00424-024-02983-4 ION CHANNELS, RECEPTORS AND TRANSPORTERS Alteration of Piezo1 signaling in type 2 diabetic mice: focus on endothelium and BKCa channel Chae Eun Haam1 · Sooyeon Choi1 · Seonhee Byeon1 · Eun Yi Oh1 · Soo‑Kyoung Choi1 · Young‑Ho Lee1 Received: 13 May 2024 / Revised: 18 June 2024 / Accepted: 21 June 2024 © The Author(s) 2024 Abstract Piezo1 mechanosensitive ion channel plays a important role in vascular physiology and disease. This study aimed to elucidate the altered signaling elicited by Piezo1 activation in the arteries of type 2 diabetes. Ten- to 12-week-old male C57BL/6 (control) and type 2 diabetic mice (db−/db−) were used. The second-order mesenteric arteries (~ 150 μm) were used for isometric tension experiments. Western blot analysis and immunofluorescence staining were performed to observe protein expression. Piezo1 was significantly decreased in mesenteric arteries of type 2 diabetic mice compared to control mice, as analyzed by western blot and immunofluorescence staining. Piezo1 agonist, Yoda1, concentration-dependently induced relaxation of mesenteric arteries in both groups. Interestingly, the relaxation response was significantly greater in control mice than in d b−/db− mice. The removal of endothelium reduced relaxation responses induced by Yoda1, which was greater in control mice than d b−/db− mice. Furthermore, the relaxation response was reduced by pre-treatment with various types of K+ channel blockers in endothelium-intact arteries in control mice. In endothelium-denuded arteries, pre-incubation with charybdotoxin, an Ca2+-activated K+ channel (BKCa channel) blocker, significantly attenuated Yoda1-induced relaxation in db−/db− mice, while there was no effect in control mice. Co-immunofluorescence staining showed co-localization of Piezo1 and BKCa channel was more pronounced in d b−/db− mice than in control mice. These results indicate that the vascular responses induced by Piezo1 activation are different in the mesenteric resistance arteries in type 2 diabetic mice. Keywords Piezo1 · Yoda1 · Type 2 diabetes · BKCa channel · Vasorelaxation Abbreviations ACh Acetylcholine SNP Sodium nitroprusside NO Nitric oxide sGC Soluble guanylyl cyclase COX Cyclooxygenase L-NNA Nω-nitro-L-arginine ODQ 1H-[1,2,4]oxadiazolo [4,3,-a] quinoxalin-1-one INDO Indomethacin ChTX Charybdotoxin BKCa Large-conductance Ca2+-activated K+ channels * Soo‑Kyoung Choi * Young‑Ho Lee 1 Department of Physiology, Yonsei University College of Medicine, 50 Yonseiro, Seodaemun‑gu, Seoul 03722, Korea TRAM-34 Rriarylmethane-34 IKCa Intermediate-conductance Ca2+-activated K+ channels SKCa Small-conductance Ca2+-activated K+ channels DMSO Dimethyl sulfoxide cGMP Guanosine 3′,5′-cyclic monophosphate cAMP Adenosine 3′,5′-cyclic monophosphate W/O Wash out SEM Standard error of the mean ANOVA Analysis of variance PCC Pearson’s correlation coefficient Introduction Diabetes mellitus (DM) is a chronic metabolic disease characterized by high blood sugar resulting from impaired insulin secretion, defective insulin action, or a combination of the two [14]. Vascular dysfunction, specifically the compromised endothelium-dependent relaxation, is Vol.:(0123456789) Pflügers Archiv - European Journal of Physiology linked to diabetes and is recognized as a crucial factor in the onset of vascular complications associated with diabetes [28]. Endothelial dysfunction is a central event in the pathogenesis of diabetes, and it greatly affects the development of future vascular complications [27]. However, the mechanisms underlying this damage remain incompletely understood. Therefore, it is crucial to comprehend the mechanisms responsible for endothelial dysfunction caused by diabetes mellitus and to identify treatments that can enhance or restore endothelial function to prevent diabetic vascular complications. Mechanotransduction plays a pivotal role in vascular development, physiology, and disease [17]. Piezo1, a mechanically sensitive non-selective cation channel, was identified as an essential protein expressed in endothelial and vascular smooth muscle cells [6, 9, 16, 26]. The Piezo1 is considered a sensor for shear stress in vascular structures and is crucial for embryonic development [25]. Furthermore, Piezo1 plays important roles for vasculogenesis, valve morphogenesis, and the regulation of vascular tone [2, 24]. Interestingly, Piezo1 exerts atheroprotective effects by regulating nitric oxide (NO) release by the endothelium. Conversely, the activation of Piezo1 through high hydrostatic pressure not only disturbs the barrier function of lung endothelial cells but also leads to arterial remodeling under hypertensive conditions. Additionally, it induces a pro-atherogenic response when exposed to turbulent flow [8]. A previous study demonstrated that dysregulation of Piezo1 occurs in multiple blood lineages in patients with type 2 diabetes mellitus (T2DM). They also reported that elevated Piezo1 activity induces prothrombotic cellular responses in red blood cells, neutrophils, and platelets. Inhibition of Piezo1 protected against thrombosis in zebrafish genetic models and human blood samples, particularly in hyperglycemic conditions [36]. Although the significance of Piezo1 in vascular function has been studied, no reports have investigated the involvement of Piezo1 in diabetic vascular dysfunction. We hypothesized that activation of Piezo1 in mesenteric resistance arteries induces distinct vascular responses in control and diabetic mice, with differences in the underlying aspects and mechanisms governing Piezo1-induced responses between the two groups. was purchased from Alomone Labs (Jerusalem, Israel). All drugs and reagents, including Yoda1, were procured from Sigma-Aldrich (St. Louis, MO, USA). Experimental animals Male C57BL/6 and db−/db− (10 weeks) supplied by the Central Lab Animal Inc. (Seoul, Republic of Korea). The db−/db− mice are characterized by a mutation in the leptin receptor gene, werve as a well-established animal model for type 2 diabetes. The mice were accommodated in a climatecontrolled chamber with conditions set at a temperature of 22.0 ± 2°C, humidity maintained at 55 ± 5%, and a 12-h light/dark cycle, and had free access to food and tap water. All experiments were approved by the Animal Care and Use Committees at the Yonsei University College of Medicine (protocol number 2023-0016), and experimental procedures were performed according to the Guide for the Care and Use of Laboratory Animals published by the US National Institutes of Health (NIH publication no. 85-23, 2011). Tissue preparation In all experiments, mice were euthanized using isoflurane inhalation. To confirm death, the mice were carefully checked for several signs, such as no response to toe pinch, no palpable heartbeat, and color change opacity in the eyes. We used mesenteric resistance arteries t (...truncated)