Genetic deletion of p66shc and/or cyclophilin D results in decreased pulmonary vascular tone

Abstract

Aims The pulmonary vascular tone and hypoxia-induced alterations of the pulmonary vasculature may be regulated by the mitochondrial membrane permeability transition pore (mPTP) that controls mitochondrial calcium load and apoptosis. We thus investigated, if the mitochondrial proteins p66shc and cyclophilin D (CypD) that regulate mPTP opening affect the pulmonary vascular tone. Methods and Results Mice deficient for p66shc (p66shc-/-), CypD (CypD-/-), or both proteins (p66shc/CypD-/-) exhibited decreased pulmonary vascular resistance (PVR) compared to wild-type mice determined in isolated lungs and in vivo. In contrast, systemic arterial pressure was only lower in CypD-/- mice. As cardiac function and pulmonary vascular remodelling did not differ between genotypes, we determined alterations of vascular contractility in isolated lungs and calcium handling in pulmonary arterial smooth muscle cells (PASMC) as underlying reason for decreased PVR. Potassium chloride (KCl)-induced pulmonary vasoconstriction and KCl-induced cytosolic calcium increase determined by Fura-2 were attenuated in all gene-deficient mice. In contrast, KCl-induced mitochondrial calcium increase determined by the genetically encoded Mito-Car-GECO and calcium retention capacity were increased only in CypD-/- and p66shc/CypD-/- mitochondria indicating that decreased mPTP opening affected KCl-induced intracellular calcium peaks in these cells. All mouse strains showed a similar pulmonary vascular response to chronic hypoxia, while acute hypoxic pulmonary vasoconstriction was decreased in gene-deficient mice indicating that CypD and p66shc regulates vascular contractility but not remodelling. Conclusions We conclude that p66shc specifically regulates the pulmonary vascular tone, while CypD also affects systemic pressure. However, only CypD acts via regulation of mPTP opening and mitochondrial calcium regulation. Translational Perspective Pulmonary hypertension is a progressive disease of the pulmonary vasculature ultimately resulting in right heart failure. Thus, therapeutic options targeting specifically the pulmonary vasculature are urgently needed. Our study describes for the first time the role of the proteins p66shc and CypD in the regulation of the pulmonary vascular tone. As the effect of p66shc-/- was specific for the pulmonary vasculature, it is an interesting target for future research on therapies for pulmonary vascular diseases like pulmonary hypertension.

Lack of Contribution of p66shc to Pressure Overload-Induced Right Heart Hypertrophy

“…However, our data did not indicate differences between WT and p66KO hearts regarding hemodynamic, functional, and morphological parameters suggesting that p66shc deletion per se had no impact on the development of RV hypertrophy and function upon pressure overload. Similarly, our recent work demonstrated that p66shc deletion does not influence cardiac function in hypoxia-induced pulmonary hypertension [54]. Our previous data also exclude a contribution for p66shc in myocardial ischemia/reperfusion injury and the protection from it by ischemic preconditioning, both conditions in which ROS play important roles [55].…”

Deletion of classical transient receptor potential 1, 3 and 6 alters pulmonary vasoconstriction in chronic hypoxia-induced pulmonary hypertension in mice

“…Afterwards, the arteries were flushed with PSS until the baseline was reached, and then equilibrated for 30 min in Ca 2+ -free PSS (in mM, NaCl 119, KCl 4.7, MgSO 4 1.17, KH 2 PO 4 1.18, MnCl 2 2.5, EGTA 10, EDTA 0.03, Glucose 5.5, NaHCO 3 25, pH 7.4). The arterial stiffness was assessed as previously described ( Gierhardt et al, 2022 ): The distance between the wires was increased stepwise while recording the corresponding force in mN. Using the DMT normalization feature in Labchart (ADInstruments, Dunedin, New Zealand), the resulting stress-strain curve displaying the relationship between the logarithmically transformed resting wall tension and the corresponding internal diameter was calculated via regression analysis.…”

Stress/cell death pathways, neuroinflammation, and neuropathic pain

“…p66shc, a shc adaptor protein family member, participates in modulating mitochondrial homeostasis, apoptosis, and production of autophagosomes. 124 siRNA-packaged nanoparticles relieve neuropathic pain through decreasing autophagic, mitophagic, and neuroinflammatory processes both in the L5 spinal cord of SNL-evoked neuropathic pain rats and H 2 O 2 -exposed neuronal cells. 125…”

“…Carvacrol ameliorates these alterations, demonstrating that NLRP3 suppression by carvacrol potentially puts down to autophagic activation through Keap1/Nrf2/p62 and augmented mitochondrial quality control. p66shc, a shc adaptor protein family member, participates in modulating mitochondrial homeostasis, apoptosis, and production of autophagosomes 124 . p66shc siRNA‐packaged nanoparticles relieve neuropathic pain through decreasing autophagic, mitophagic, and neuroinflammatory processes both in the L5 spinal cord of SNL‐evoked neuropathic pain rats and H 2 O 2 ‐exposed neuronal cells 125 …”