Apoptosis signal‐regulating kinase 1 inhibition in in vivo and in vitro models of pulmonary hypertension

Abstract

Pulmonary arterial hypertension, group 1 of the pulmonary hypertension disease family, involves pulmonary vascular remodelling, right ventricular dysfunction and cardiac failure. Oxidative stress, through activation of mitogen-activated protein kinases is implicated in these changes. Inhibition of apoptosis signal-regulating kinase 1, an apical mitogen-activated protein kinase, prevented pulmonary arterial hypertension developing in rodent models. Here, we investigate apoptosis signal-regulating kinase 1 in pulmonary arterial hypertension by examining the impact that its inhibition has on the molecular and cellular signalling in established disease. Apoptosis signal-regulating kinase 1 inhibition was investigated in in vivo pulmonary arterial hypertension and in vitro pulmonary hypertension models. In the in vivo model, male Sprague Dawley rats received a single subcutaneous injection of Sugen SU5416 (20 mg/kg) prior to two weeks of hypobaric hypoxia (380 mmHg) followed by three weeks normoxia (Sugen/hypoxic), then animals were either maintained for three weeks on control chow or one containing apoptosis signal-regulating kinase 1 inhibitor (100 mg/kg/day). Cardiovascular measurements were carried out. In the in vitro model, primary cultures of rat pulmonary artery fibroblasts and rat pulmonary artery smooth muscle cells were maintained in hypoxia (5% O2) and investigated for proliferation, migration and molecular signalling in the presence or absence of apoptosis signal-regulating kinase 1 inhibitor. Sugen/hypoxic animals displayed significant pulmonary arterial hypertension compared to normoxic controls at eight weeks. Apoptosis signal-regulating kinase 1 inhibitor decreased right ventricular systolic pressure to control levels and reduced muscularised vessels in lung tissue. Apoptosis signal-regulating kinase 1 inhibition was found to prevent hypoxia-induced proliferation, migration and cytokine release in rat pulmonary artery fibroblasts and also prevented rat pulmonary artery fibroblast-induced rat pulmonary artery smooth muscle cell migration and proliferation. Apoptosis signal-regulating kinase 1 inhibition reversed pulmonary arterial hypertension in the Sugen/hypoxic rat model. These effects may be a result of intrinsic changes in the signalling of adventitial fibroblast.

Connexin 43 plays a role in proliferation and migration of pulmonary arterial fibroblasts in response to hypoxia

“… 37 In the current study, rPASMCs proliferated in response to 1% FBS to a similar degree under both normoxic and hypoxic conditions. While our finding that rPASMCs do not proliferate in response to hypoxia is in agreement with some previous studies, 17 , 38 others have shown hypoxic-induced proliferation of rPASMCs. You et al.…”

“…17 In vitro rPAFs derived from sugen5416/ hypoxic rats displayed increased proliferation, migration and p38 MAPK expression; these effects were not present in rPAFs derived from sugen5416/hypoxic rats dosed with the ASK1 inhibitor, GS-444217. 17 Using lung tissue from these sugen5416/hypoxic rats, we investigated if the changes seen in vivo and in vitro also coincided with an altered connexin expression. We observed no changes in GJA1 (Cx43) gene expression in lung tissue from sugen5416/hypoxic rats.…”

“…17 In the sugen5416/hypoxic rat model of PH, inhibiting ASK-1 decreased right ventricular pressure and vascular remodelling in vivo . 17 In vitro rPAFs derived from sugen5416/hypoxic rats displayed increased proliferation, migration and p38 MAPK expression; these effects were not present in rPAFs derived from sugen5416/hypoxic rats dosed with the ASK1 inhibitor, GS-444217. 17 Using lung tissue from these sugen5416/hypoxic rats, we investigated if the changes seen in vivo and in vitro also coincided with an altered connexin expression.…”

“…2 It has also been shown that inhibition of the MAPK pathway leads to decreased proliferation, migration and p38 MAPK expression in rat PAFs (rPAFs) and also attenuates the development of PH in sugen5416/hypoxic rats. 17 Apoptosis signal-regulating kinase-1 (ASK-1) is an upstream regulator of the MAPK pathway. In response to oxidative stress, ASK-1 undergoes auto-phosphorylation and activation of downstream p38 MAPK.…”

Oxidative Stress, Kinase Activation, and Inflammatory Pathways Involved in Effects on Smooth Muscle Cells During Pulmonary Artery Hypertension Under Hypobaric Hypoxia Exposure

“…Reactive oxygen species are also involved in regulation of cell signaling through the mitogen-activated protein kinase (MAPK) pathway, which is associated with a variety of cellular effects, including proliferation, survival, death, and transformation ( Son et al, 2013 ). One study on rats exposed to hypobaric hypoxia showed that the constitutive MAPK termed apoptosis signal-regulating kinase 1 (ASK1), which is known to be critical in the remodeling processes of pulmonary artery hypertension, participates in PASMC proliferation through activation of p38 MAPK and endothelin-1 (ET-1) in response to oxidative stress ( Wilson et al, 2020 ).…”