Biology subjects

Shattock, M. J.

Publications and source records attributed to Shattock, M. J..

3 recordsLinked to original sources

bioRxiv2025

Targeting C42 in PKGIα alleviates diastolic dysfunction in HFpEF

BackgroundIn our exploration of novel therapeutic strategies for treating heart failure with preserved ejection fraction (HFpEF), we focused on cysteine 42 (C42) in cGMP-dependent protein kinase I (PKGI), which plays a crucial role in regulating blood pressure and diastolic relaxation. Our findings demonstrate that targeting C42 on PKGI with urolithin A effectively limits diastolic dysfunction and cardiac remodeling in a multi-hit mouse model of HFpEF, while also improving the kinetics of relaxation and contraction in human-engineered heart tissue. MethodsWe evaluated various polyphenols, including urolithin A, for their capacity to oxidize and activate PKGI. This assessment involved recombinant protein, isolated rat aortic smooth muscle cells, and myography experiments using isolated mesenteric vessels. Additionally, we investigated the ability of urolithin A to stimulate PKGI-dependent phosphorylation of phospholamban (PLN) in neonatal rat ventricular myocytes (NRVMs). A multi-hit mouse model was employed to induce HFpEF in wild-type and C42S PKGI knock-in (KI) mice. Following this, the mice were given vehicle or urolithin A by gavage, and heart function was evaluated using echocardiography. The relevance to human physiology was assessed using human-engineered heart tissue. ResultsThe polyphenols quercetin, fisetin, and urolithin A were shown to induce C42-dependent activation of PKGI. This mechanism of kinase activation led to vasorelaxation, which was attenuated in mesenteric vessels isolated from KI mice. Additionally, urolithin A treatment promoted PKGI-dependent phosphorylation of phospholamban in isolated cardiomyocytes. In a multi-hit model of HFpEF, urolithin A reversed diastolic dysfunction and cardiac remodeling in wild-type mice, but not in KI mice. These findings were shown to be likely relevant to humans, as urolithin A also enhanced the kinetics of relaxation and contraction in human-engineered heart tissue. ConclusionsTargeting C42 in PKGI with urolithin A presents a promising new strategy for the treatment of HFpEF.

biochemistry↗

bioRxiv2024

Sarcolemmal and mitochondrial membrane potentials measured ex vivo and in vivo in the heart by pharmacokinetic modelling of sestamibi

We present a compartmental modelling approach to analyse radioactive time activity curves for first pass kinetics of [99mTc]sestamibi in the heart. Reparametrizing the kinetic equations using the Nernst membrane-potential equation provides a novel means of non-invasively estimating the sarcolemmal (Em) and mitochondrial ({Delta}{Psi}m) membrane potentials in the heart. A Markov Chain Monte Carlo (MCMC) fitting approach was applied to data derived from established interventions in Langendorff perfused rat hearts where the sarcolemmal membrane was depolarised using hyperkalaemic Krebs Henseleit buffers; the mitochondrial membrane was depolarised using carbonylcyanide-3-chlorophenylhydrazone (CCCP); or both membranes were depolarised using their combination. Translating this approach to single photon emission planar scintigraphy kinetics from healthy rats allowed an estimate of these membrane potentials (voltages) in vivo for the first time; the values were Em = -62 {+/-} 5 mV and {Delta}{Psi}m = -151 {+/-} 5 mV (n = 4, mean {+/-} SD).

biochemistry↗

bioRxiv2022

zDHHC5 expression is increased in cardiac hypertrophy and reduced in heart failure but this does not correlate with changes in substrate palmitoylation

S-palmitoylation is an essential lipid modification catalysed by zDHHC-palmitoyl acyltransferases that regulates the localisation and activity of substrates in every class of protein and tissue investigated to date. In the heart, S-palmitoylation regulates sodiumcalcium exchanger (NCX1) inactivation, phospholemman (PLM) inhibition of the Na+/K+ ATPase, Nav1.5 influence on membrane excitability and membrane localisation of heterotrimeric G-proteins. The cell surface localised enzyme zDHHC5 palmitoylates NCX1 and PLM and is implicated in injury during anoxia/reperfusion. Information is lacking about how palmitoylation remodels in cardiac diseases. We investigated expression of zDHHC5 in animal models of left ventricular hypertrophy (LVH) and heart failure (HF), along with HF tissue from humans. zDHHC5 expression was rapidly elevated during onset of LVH, whilst HF was associated with decreased zDHHC5 expression. Paradoxically, palmitoylation of the zDHHC5 substrate NCX1 was significantly reduced in LVH but increased in human HF. Overexpression of zDHHC5 in rabbit ventricular cardiomyocytes was not sufficient to drive changes in palmitoylation of zDHHC5 substrates or overall cardiomyocyte contractility, suggesting changes in zDHHC5 expression in disease may not be a primary driver of pathology. zDHHC5 itself is regulated by post-translational modifications, including palmitoylation in its Cterminal tail, and we found the palmitoylation of zDHHC5 may be increased in heart failure in the same manner as NCX1, suggesting additional regulatory mechanisms such as acyl-CoA availability may be involved. Importantly, this study provides the first evidence that palmitoylation of cardiac substrates is altered in the setting of HF, and that expression of zDHHC5 is dysregulated in both hypertrophy and HF.

biochemistry↗