The Action of 2-Aminoethyldiphenyl Borinate on the Pulmonary Arterial Hypertension and Remodeling of High-Altitude Hypoxemic Lambs

Abstract

Calcium signaling is key for the contraction, differentiation, and proliferation of pulmonary arterial smooth muscle cells. Furthermore, calcium influx through store-operated channels (SOCs) is particularly important in the vasoconstrictor response to hypoxia. Previously, we found a decrease in pulmonary hypertension and remodeling in normoxic newborn lambs partially gestated under chronic hypoxia, when treated with 2-aminoethyldiphenyl borinate (2-APB), a non-specific SOC blocker. However, the effects of 2-APB are unknown in neonates completely gestated, born, and raised under environmental hypoxia. Accordingly, we studied the effects of 2-APB-treatment on the cardiopulmonary variables in lambs under chronic hypobaric hypoxia. Experiments were done in nine newborn lambs gestated, born, and raised in high altitude (3,600 m): five animals were treated with 2-APB [intravenous (i.v.) 10 mg kg–1] for 10 days, while other four animals received vehicle. During the treatment, cardiopulmonary variables were measured daily, and these were also evaluated during an acute episode of superimposed hypoxia, 1 day after the end of the treatment. Furthermore, pulmonary vascular remodeling was assessed by histological analysis 2 days after the end of the treatment. Basal cardiac output and mean systemic arterial pressure (SAP) and resistance from 2-APB- and vehicle-treated lambs did not differ along with the treatment. Mean pulmonary arterial pressure (mPAP) decreased after the first day of 2-APB treatment and remained lower than the vehicle-treated group until the third day, and during the fifth, sixth, and ninth day of treatment. The net mPAP increase in response to acute hypoxia did not change, but the pressure area under the curve (AUC) during hypoxia was slightly lower in 2-APB-treated lambs than in vehicle-treated lambs. Moreover, the 2-APB treatment decreased the pulmonary arterial wall thickness and the α-actin immunoreactivity and increased the luminal area with no changes in the vascular density. Our findings show that 2-APB treatment partially reduced the contractile hypoxic response and reverted the pulmonary vascular remodeling, but this is not enough to normalize the pulmonary hemodynamics in chronically hypoxic newborn lambs.

Crucial Role of Stromal Interaction Molecule-Activated TRPC-ORAI Channels in Vascular Remodeling and Pulmonary Hypertension Induced by Intermittent Hypoxia

“…We studied the effects of 2-APB, a drug with inhibitory actions on STOC ( Bootman et al, 2002 , Castillo-Galán et al, 2016 , 2022 ), in a rat model of CIH-induced pulmonary hypertension ( Del Rio et al, 2010 , 2012 , 2014 ; Iturriaga et al, 2014 ; Castillo-Galán et al, 2020 ). Consistently with previous studies, rats exposed to CIH for 28 days, develop vascular remodeling characterized by an increased thickness of the medial layer in pulmonary arteries and an elevated right ventricular systolic pressure ( Nisbet et al, 2009 ; Jin et al, 2014 , 2016 ; Iturriaga and Castillo-Galán, 2019 ; Castillo-Galán et al, 2020 ).…”

“…In this study, 2-APB was administered with osmotic pumps at a delivery rate of 10 mg/kg per day, which is similar to the daily dose range used by Smith et al (2015) , Castillo-Galán et al (2016 , 2022) and Jain et al (2021) to block STOC in vivo . Indeed, Smith et al (2015) treated chronic hypoxic mice with 2-APB (1 mg/kg/day i.p.)…”

“…to reduce pulmonary hypertension and right ventricular enlargement. Castillo-Galán et al (2016 , 2022) , administered 2-APB (10 mg/kg/day i.v.) to neonatal lambs with a partial or full gestation in high altitude to ameliorate the pulmonary hypertension and vascular remodeling, and Jain et al (2021) administrated 2-APB (1 mg/kg per day i.p.)…”

“…On the other hand, an auxiliary subunit of the family of stromal interaction molecules [stromal interaction molecule (STIM) 1 and 2] acts as a calcium sensor activating pore-forming subunits in response to calcium depletion from sarcoplasmic reticulum deposits ( Fuchs et al, 2010 ; Johnstone et al, 2010 ; Wray and Burdyga, 2010 ; Yang et al, 2010 ). Indeed, a growing body of evidence suggests that STIM-activated TRPC-ORAI Ca 2+ channels (STOC) play a key role in PASMC remodeling and hypercontractility, which led to pulmonary hypertension in animals exposed to sustained hypoxia ( Weigand et al, 2005 ; Ng et al, 2008 ; Castillo-Galán et al, 2016 , 2022 )…”

“…Recently, we found that CIH produced the upregulation of mRNA and protein levels of TRPC1, TRPC4, TRPC6, ORAI1, and STIM1 subunits in the rat lung tissue, which paralleled the vascular remodeling and pulmonary hypertension ( Castillo-Galán et al, 2020 ), suggesting that STOC participate in the vascular alterations induced by CIH. Accordingly, to assess the contribution of STOC on the CIH-induced pulmonary vascular alterations, we study the effects of the STOC non-selective inhibitor 2-aminoethyldiphenylborinate (2-APB) ( Bootman et al, 2002 ; Smith et al, 2015 ; Castillo-Galán et al, 2016 , 2022 ; Jain et al, 2021 ) on the development of pulmonary hypertension and vascular remodeling induced by CIH in a well stablished preclinical model of OSA ( Del Rio et al, 2010 , 2012 , 2014 ; Iturriaga et al, 2014 , Iturriaga and Castillo-Galán, 2019 ). We also determined the effects of 2-APB on systemic and lung oxidative stress induced by CIH, because 2-APB ameliorates the oxidative stress in a heart ischemia-reperfusion model in mice ( Morihara et al, 2017 ) and rat ( Shen et al, 2021 ).…”