Smooth muscle Acid-sensing ion channel 1a as a therapeutic target to reverse hypoxic pulmonary hypertension.

Acid-sensing ion channel 1a (ASIC1a) is a voltage-independent, non-selective cation channel that conducts both Na+ and Ca2+. Activation of ASIC1a elicits plasma membrane depolarization and stimulates intracellular Ca2+-dependent signaling pathways in multiple cell types, including vascular smooth muscle (SM) and endothelial cells (ECs). Previous studies have shown that increases in pulmonary vascular resistance accompanying chronic hypoxia (CH)-induced pulmonary hypertension requires ASIC1a to elicit enhanced pulmonary vasoconstriction and vascular remodeling. Both SM and EC dysfunction drive these processes; however, the involvement of ASIC1a within these different cell types is unknown. Using the Cre-LoxP system to generate cell-type-specific Asic1a knockout mice, we tested the hypothesis that SM-Asic1a contributes to CH-induced pulmonary hypertension and vascular remodeling, whereas EC-Asic1a opposes the development of CH-induced pulmonary hypertension. The severity of pulmonary hypertension was not altered in mice with specific deletion of EC-Asic1a (TekCre-Asic1a fl/fl). However, similar to global Asic1a knockout (Asic1a -/-) mice, mice with specific deletion of SM-Asic1a (MHCCreER-Asic1a fl/fl) were protected from the development of CH-induced pulmonary hypertension and right heart hypertrophy. Furthermore, pulmonary hypertension was reversed when deletion of SM-Asic1a was initiated in conditional MHCCreER-Asic1a fl/fl mice with established pulmonary hypertension. CH-induced vascular remodeling was also significantly attenuated in pulmonary arteries from MHCCreER-Asic1a fl/fl mice. These findings were additionally supported by decreased CH-induced proliferation and migration of pulmonary arterial smooth muscle cells (PASMCs) from Asic1a -/- mice. Together these data demonstrate that SM-, but not EC-Asic1a contributes to CH-induced pulmonary hypertension and vascular remodeling. Furthermore, these studies provide evidence for the therapeutic potential of ASIC1a inhibition to reverse pulmonary hypertension.

Infection prevention strategies are highly protective in COVID-19 units while main risks to healthcare professionals come from coworkers and the community.

BACKGROUND: Early evaluations of healthcare professional (HCP) COVID-19 risk occurred during insufficient personal protective equipment and disproportionate testing, contributing to perceptions of high patient-care related HCP risk. We evaluated HCP COVID-19 seropositivity after accounting for community factors and coworker outbreaks. METHODS: Prior to universal masking, we conducted a single-center retrospective cohort plus cross-sectional study. All HCP (1) seen by Occupational Health for COVID-like symptoms (regardless of test result) or assigned to (2) dedicated COVID-19 units, (3) units with a COVID-19 HCP outbreak, or (4) control units from 01/01/2020 to 04/15/2020 were offered serologic testing by an FDA-authorized assay plus a research assay against 67 respiratory viruses, including 11 SARS-CoV-2 antigens. Multivariable models assessed the association of demographics, job role, comorbidities, care of a COVID-19 patient, and geocoded socioeconomic status with positive serology. RESULTS: Of 654 participants, 87 (13.3%) were seropositive; among these 60.8% (N = 52) had never cared for a COVID-19 patient. Being male (OR 1.79, CI 1.05-3.04, p = 0.03), working in a unit with a HCP-outbreak unit (OR 2.21, CI 1.28-3.81, p < 0.01), living in a community with low owner-occupied housing (OR = 1.63, CI = 1.00-2.64, p = 0.05), and ethnically Latino (OR 2.10, CI 1.12-3.96, p = 0.02) were positively-associated with COVID-19 seropositivity, while working in dedicated COVID-19 units was negatively-associated (OR 0.53, CI = 0.30-0.94, p = 0.03). The research assay identified 25 additional seropositive individuals (78 [12%] vs. 53 [8%], p < 0.01). CONCLUSIONS: Prior to universal masking, HCP COVID-19 risk was dominated by workplace and community exposures while working in a dedicated COVID-19 unit was protective, suggesting that infection prevention protocols prevent patient-to-HCP transmission. Prior to universal masking, HCP COVID-19 risk was dominated by workplace and community exposures while working in a dedicated COVID-19 unit was protective, suggesting that infection prevention protocols prevent patient-to-HCP transmission.

Crucial role for pathology residents in laboratory self-inspection, a single Institute's experience.

BACKGROUND: Training in patient safety, quality, and management is a key component of Graduate Medical Education (GME) training in all specialties. However, residency programs, especially Pathology programs, often find it challenging to create strong learning opportunities in these areas. OBJECTIVES: Focused quality assurance (QA) projects are one approach to teach and engage trainees in these key areas. Residents have been historically involved in different QA projects in our department but mainly in small secondary roles. Leading a large QA project that can enhance residents' management skills and improve clinical operations in our laboratory was the main objective of our project. DESCRIPTION: A new process for laboratory self-inspection led by residents was implemented that simulates the exact process of a formal outside College of American Pathologists (CAP) inspection. We aim to prove that resident-led QA activities not only have profound educational benefit but can also result in significant performance and operational improvement. RESULTS: For this paper, we focus on the Histology laboratory since the ramifications from the self-inspection process during a three year period were profound leading to change in management, workflow changes, and notable improvement in staff morale. CONCLUSION: The self-inspection process exposed the residents to operational issues and corrective actions that provided them the opportunity to take a more active role in laboratory management and helped prepare them for post-graduation challenges. It also helped the department identify and rectify many operational issues, confirmed by the enumeration of CAP deficiencies and significant improvement of staff morale.

Loss of acid-sensing ion channel 2 enhances pulmonary vascular resistance and hypoxic pulmonary hypertension.

Acid-sensing ion channels (ASICs) are voltage-insensitive cation channels that contribute to cellular excitability. We previously reported that ASIC1 in pulmonary artery smooth muscle cells (PASMC) contribute to pulmonary vasoreactivity and vascular remodeling during the development of chronic hypoxia (CH)-induced pulmonary hypertension. However, the roles of ASIC2 and ASIC3 in regulation of pulmonary vasoreactivity and the development of CH-induced pulmonary hypertension are unknown. We tested the hypothesis that ASIC2 and ASIC3 contribute to increased pulmonary vasoreactivity and development of CH-induced pulmonary hypertension using ASIC2- and ASIC3-knockout (-/-) mice. In contrast to this hypothesis, we found that ASIC2-/- mice exhibit enhanced CH-induced pulmonary hypertension compared with WT and ASIC3-/- mice. This response was not associated with a change in ventilatory sensitivity or systemic cardiovascular function but was instead associated with direct changes in pulmonary vascular reactivity and pulmonary arterial morphology in ASIC2-/- mice. This increase in reactivity correlated with enhanced pulmonary arterial basal tone, elevated basal PASMC [Ca2+] and store-operated calcium entry (SOCE) in PASMC from ASIC2-/- mice. This increase in PASMC [Ca2+] and vasoreactivity was dependent on ASIC1-mediated Ca2+ influx but was not contingent upon an increase in ASIC1 mRNA or protein expression in PASMC from ASIC2-/- mice. Together, the results from this study demonstrate an important role for ASIC2 to regulate pulmonary vascular reactivity and for ASIC2 to modulate the development of CH-induced pulmonary hypertension. These data further suggest that loss of ASIC2 enhances the contribution of ASIC1 to overall pulmonary vascular reactivity.NEW & NOTEWORTHY This study demonstrates that loss of ASIC2 leads to increased baseline pulmonary vascular resistance, enhanced responses to a variety of vasoconstrictor stimuli, and greater development of hypoxic pulmonary hypertension. Furthermore, these results suggest that loss of ASIC2 enhances the contribution of ASIC1 to pulmonary vascular reactivity.

Role of acid-sensing ion channels in hypoxia- and hypercapnia-induced ventilatory responses.

Previous reports indicate roles for acid-sensing ion channels (ASICs) in both peripheral and central chemoreception, but the contributions of ASICs to ventilatory drive in conscious, unrestrained animals remain largely unknown. We tested the hypotheses that ASICs contribute to hypoxic- and hypercapnic-ventilatory responses. Blood samples taken from conscious, unrestrained mice chronically instrumented with femoral artery catheters were used to assess arterial O2, CO2, and pH levels during exposure to inspired gas mixtures designed to cause isocapnic hypoxemia or hypercapnia. Whole-body plethysmography was used to monitor ventilatory parameters in conscious, unrestrained ASIC1, ASIC2, or ASIC3 knockout (-/-) and wild-type (WT) mice at baseline, during isocapnic hypoxemia and during hypercapnia. Hypercapnia increased respiratory frequency, tidal volume, and minute ventilation in all groups of mice, but there were no differences between ASIC1-/-, ASIC2-/-, or ASIC3-/- and WT. Isocapnic hypoxemia also increased respiratory frequency, tidal volume, and minute ventilation in all groups of mice. Minute ventilation in ASIC2-/- mice during isocapnic hypoxemia was significantly lower compared to WT, but there were no differences in the responses to isocapnic hypoxemia between ASIC1-/- or ASIC3-/- compared to WT. Surprisingly, these findings show that loss of individual ASIC subunits does not substantially alter hypercapnic or hypoxic ventilatory responses.

Contribution of reactive oxygen species to the pathogenesis of pulmonary arterial hypertension.

Pulmonary arterial hypertension is associated with a decreased antioxidant capacity. However, neither the contribution of reactive oxygen species to pulmonary vasoconstrictor sensitivity, nor the therapeutic efficacy of antioxidant strategies in this setting are known. We hypothesized that reactive oxygen species play a central role in mediating both vasoconstrictor and arterial remodeling components of severe pulmonary arterial hypertension. We examined the effect of the chemical antioxidant, TEMPOL, on right ventricular systolic pressure, vascular remodeling, and enhanced vasoconstrictor reactivity in both chronic hypoxia and hypoxia/SU5416 rat models of pulmonary hypertension. SU5416 is a vascular endothelial growth factor receptor antagonist and the combination of chronic hypoxia/SU5416 produces a model of severe pulmonary arterial hypertension with vascular plexiform lesions/fibrosis that is not present with chronic hypoxia alone. The major findings from this study are: 1) compared to hypoxia alone, hypoxia/SU5416 exposure caused more severe pulmonary hypertension, right ventricular hypertrophy, adventitial lesion formation, and greater vasoconstrictor sensitivity through a superoxide and Rho kinase-dependent Ca2+ sensitization mechanism. 2) Chronic hypoxia increased medial muscularization and superoxide levels, however there was no effect of SU5416 to augment these responses. 3) Treatment with TEMPOL decreased right ventricular systolic pressure in both hypoxia and hypoxia/SU5416 groups. 4) This effect of TEMPOL was associated with normalization of vasoconstrictor responses, but not arterial remodeling. Rather, medial hypertrophy and adventitial fibrotic lesion formation were more pronounced following chronic TEMPOL treatment in hypoxia/SU5416 rats. Our findings support a major role for reactive oxygen species in mediating enhanced vasoconstrictor reactivity and pulmonary hypertension in both chronic hypoxia and hypoxia/SU5416 rat models, despite a paradoxical effect of antioxidant therapy to exacerbate arterial remodeling in animals with severe pulmonary arterial hypertension in the hypoxia/SU5416 model.

BK channels in rat and human pulmonary smooth muscle cells are BKα-ß1 functional complexes lacking the oxygen-sensitive stress axis regulated exon insert.

A loss of K+ efflux in pulmonary arterial smooth muscle cells (PASMCs) contributes to abnormal vasoconstriction and PASMC proliferation during pulmonary hypertension (PH). Activation of high-conductance Ca2+-activated (BK) channels represents a therapeutic strategy to restore K+ efflux to the affected PASMCs. However, the properties of BK channels in PASMCs-including sensitivity to BK channel openers (BKCOs)-are poorly defined. The goal of this study was to compare the properties of BK channels between PASMCs of normoxic (N) and chronic hypoxic (CH) rats and then explore key findings in human PASMCs. Polymerase chain reaction results revealed that 94.3% of transcripts encoding BKα pore proteins in PASMCs from N rats represent splice variants lacking the stress axis regulated exon insert, which confers oxygen sensitivity. Subsequent patch-clamp recordings from inside-out (I-O) patches confirmed a dense population of BK channels insensitive to hypoxia. The BK channels were highly activated by intracellular Ca2+ and the BKCO lithocholate; these responses require BKα-ß1 subunit coupling. PASMCs of CH rats with established PH exhibited a profound overabundance of the dominant oxygen-insensitive BKα variant. Importantly, human BK (hBK) channels in PASMCs from human donor lungs also represented the oxygen-insensitive BKα variant activated by BKCOs. The hBK channels showed significantly enhanced Ca2+ sensitivity compared with rat BK channels. We conclude that rat BK and hBK channels in PASMCs are oxygen-insensitive BKα-ß1 complexes highly sensitive to Ca2+ and the BKCO lithocholate. BK channels are overexpressed in PASMCs of a rat model of PH and may provide an abundant target for BKCOs designed to restore K+ efflux.

Furegrelate, a thromboxane synthase inhibitor, blunts the development of pulmonary arterial hypertension in neonatal piglets.

The development of pulmonary arterial hypertension (PAH) in pediatric patients has been linked to the production of the arachidonic acid metabolite, thromboxane A(2) (TxA(2)). The present study evaluated the therapeutic effect of furegrelate sodium, a thromboxane synthase inhibitor, on the development of PAH in a neonatal piglet model. Three-day-old piglets were exposed to 21 days of normoxia (N; 21% F(I)O(2)) or chronic hypoxia (CH; 10% F(I)O(2)). A third group of piglets received the oral TxA(2) synthase inhibitor, furegrelate (3 mg/kg, 2 or 3 times daily) at the induction of CH. In vivo hemodynamics confirmed a 2.55-fold increase of the pulmonary vascular resistance index (PVRI) in CH piglets (104±7 WU) compared to N piglets (40±2 WU). The CH piglets treated twice daily with furegrelate failed to show improved PVRI, but furegrelate three times daily lowered the elevated PVRI in CH piglets by 34% to 69±5 WU and ameliorated the development of right ventricular hypertrophy. Microfocal X-ray computed tomography (CT) scanning was used to estimate the diameter-independent distensibility term, α (% change in diameter per Torr). Pulmonary arterial distensibility in isolated lungs of CH piglets (α=1.0±0.1% per Torr) was lower than that of N piglets (α=1.5±0.1% per Torr) indicative of vascular remodeling. Arterial distensibility was partially restored in furegrelate-treated CH piglets (α =1.2±0.1% per Torr) and microscopic evidence showing muscularization of small pulmonary arteries also was less prominent in these animals. Finally, isolated lungs of furegrelate-treated piglets showed lower basal and vasodilator-induced transpulmonary pressures compared to CH animals. These findings suggest that pharmacological inhibition of TxA(2) synthase activity by furegrelate blunts the development of hypoxia-induced PAH in an established neonatal piglet model primarily by preserving the structural integrity of the pulmonary vasculature.