Supplement to Serotonin Signaling in Mitral Valve Homeostasis, Maintenance and Restoration

Children'S Hosp Of Philadelphia, Philadelphia PA

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Abstract

Abstract/Summary This one-year Supplement project will investigate the novel relationship of Piezo1, a mechanical sensing channel never studied previously concerning mitral valve prolapse (MVP) or mitral regurgitation (MR) to serotonin (5HT) mechanisms that contribute to the progression of MR. We previously demonstrated that mitral valve interstitial cell (MVIC) serotonin transporter (SERT) activity was inhibited by either the diet drug, Fenfluoramine or Fluoxetine, a selective 5HT reuptake inhibitor (SSRI), thus resulting in diminished 5HT clearance and enhanced 5HT receptor (HTR) activity. In addition, we showed in clinical studies of MR surgical patients that SSRI use was significantly associated with MR surgery at a younger age, and qRTPCR studies of human MR leaflets, compared to normal human mitral valve (MV) leaflets showed diminished SERT activity in MR. Piezo1 and increased 5HT secretion: Prior research has reported that intestinal Piezo1 activation results in increased 5HT, stimulating our research interest. Feasibility studies presented in this proposal demonstrate that Piezo1 agonists increase HTR signaling in MVIC. Hypothesis: MR progression results from increased HTR2B signaling due to diminished SERT expression and increased 5HT secretion, caused by activation of the mechanically sensitive Ca++ channel, Piezo1. Specific Aim 1: To study the effects of Piezo1 activation and inhibition on 5HT mechanisms in human MVIC, and in a mouse model of Fluoxetine induced MR. We will use the specific Piezo1 agonist, Yoda1, to model mechanical stimulation of Piezo1 in MVIC, and we will also study the effects of inhibition of Piezo1, HTR2B, and tryptophan hydroxylase-1 (TPH1) to reduce HTR signaling. MVIC endpoints include differences in extracellular matrix (ECM) production, expression of 5HT related genes, collagens 1&3, and Piezo1 and 2. Studies of our Fluoxetine mouse mitral valvulopathy model will investigate mitigation of the valvulopathy with both a HTR2B inhibitor and the Piezo1 inhibitor, Dooku1, with endpoints including: Ventricular remodeling per echocardiography, ventricular and valve morphology, and qRTPCR studies of 5HT related genes, and Piezo1&2. Specific Aim 2: To study human and sheep mitral valve (MV) leaflets, from both MR (human only) and normal MV (sheep and human) in cycle stretch studies with our mechanical bioreactor system. We will compare static and cyclic-stretch conditions, with exposure to inhibitors of Piezo1, HTR2B, and TPH1 compared to untreated, assessing differences in expression of ECM proteins, 5HT related genes and Piezo1 and 2. RNA sequencing studies will be carried out on bioreactor samples before and after mechanical stimulation with informatics analyses to identify gene expression patterns and pathways, compared to our human MV data. The proposed studies for this Supplement are responsive to NOT-HL-23-078 because they address the underlying basic mechanisms of fibrosis responsible for progression of MVP to MR, and utilize a relevant animal model.