Biology subjects

Yutzey, K. E.

Publications and source records attributed to Yutzey, K. E..

5 recordsLinked to original sources

bioRxiv2023

PROX1 inhibits PDGF-B expression to prevent myxomatous degeneration of heart valves

BackgroundCardiac valve disease (CVD) is observed in 2.5% of the general population and 10% of the elderly people. Effective pharmacological treatments are currently not available, and patients with severe CVD require surgery. PROX1 and FOXC2 are transcription factors that are required for the development of lymphatic and venous valves. We found that PROX1 and FOXC2 are expressed in a subset of valvular endothelial cells (VECs) that are located on the downstream (fibrosa) side of cardiac valves. Whether PROX1 and FOXC2 regulate cardiac valve development and disease is not known. MethodsWe used histology, electron microscopy and echocardiography to investigate the structure and functioning of heart valves from Prox1{Delta}VEC mice in which Prox1 was conditionally deleted from VECs. Isolated valve endothelial cells and valve interstitial cells were used to identify the molecular mechanisms in vitro, which were tested in vivo by RNAScope, additional mouse models and pharmacological approaches. The significance of our findings was tested by evaluation of human samples of mitral valve prolapse (MVP) and aortic valve insufficiency. ResultsHistological analysis revealed that the aortic and mitral valves of Prox1{Delta}VEC mice become progressively thick and myxomatous. Echocardiography revealed that the aortic valves of Prox1{Delta}VEC mice are stenotic. FOXC2 was downregulated and platelet-derived growth factor-B (PDGF-B) was upregulated in the VECs of Prox1{Delta}VEC mice. Conditional knockdown of FOXC2 and conditional overexpression of PDGF-B in VECs recapitulated the phenotype of Prox1{Delta}VEC mice. PDGF-B was also increased in mice lacking FOXC2 and in human MVP and insufficient aortic valve samples. Pharmacological inhibition of PDGF-B signaling with imatinib partially ameliorated the valve defects of Prox1{Delta}VEC mice. ConclusionPROX1 antagonizes PDGF-B signaling partially via FOXC2 to maintain the extracellular matrix composition and prevent myxomatous degeneration of cardiac valves. Novelty and SignificanceWhat Is Known? O_LIThe transcription factors PROX1 and FOXC2 are critical regulators of lymphatic and venous valve development. C_LIO_LIPROX1 and FOXC2 are expressed in the downstream valvular endothelial cells of heart valves. C_LI What Is New? O_LIDeletion of Prox1 from the valvular endothelial cells of mice results in enlarged and myxomatous aortic and mitral valves. Aortic valves of the mutant (Prox1{Delta}VEC) mice were stenotic. C_LIO_LIFOXC2 is partially responsible for the phenotype of Prox1{Delta}VEC mice. C_LIO_LIPROX1 and FOXC2 inhibit the expression of the cytokine PDGF-B in heart valves. C_LIO_LIHyperactivation of PDGF-B signaling results in aortic and mitral valve thickening. C_LIO_LIInhibition of PDGF-B signaling ameliorates aortic valve stenosis in Prox1{Delta}VEC mice. C_LIO_LIPDGFB is overexpressed and PROX1 is downregulated in human mitral valve prolapse (MVP) samples. C_LI Our findings suggest that PROX1 is an inhibitor of myxomatous valve disease that afflicts ~10% of the elderly population. We have also identified PDGF-B as a potential target for treating myxomatous valve disease.

developmental biology↗

bioRxiv2023

APOE-NOTCH Axis Governs Elastogenesis During Human Cardiac Valve Remodeling

BackgroundValve remodeling is a complex process involving extracellular matrix organization, development of trilaminar structures, and physical elongation of valve leaflets. However, the cellular and molecular mechanisms regulating valve remodeling and their roles in congenital valve disorders remain poorly understood. MethodsSemilunar valves and atrioventricular valves from healthy and age-matched human fetal hearts with pulmonary stenosis (PS) were collected. Single-Cell RNA-sequencing (scRNA-seq) was performed to determine the transcriptomic landscape of multiple valvular cell subtypes in valve remodeling and disease. Spatial localization of newly-identified cell subtypes was determined via immunofluorescence and RNA in situ hybridization. The molecular mechanisms mediating valve development was investigated utilizing primary human fetal heart valve interstitial cells (VICs) and endothelial cells (VECs). ResultsscRNA-seq analysis of healthy human fetal valves identified a novel APOE+ elastin-producing VIC subtype (Elastin-VICs) spatially located underneath VECs sensing the unidirectional flow. Knockdown of APOE in fetal VICs resulted in significant elastogenesis defects. In pulmonary valve with PS, we observed decreased expression of APOE and other genes regulating elastogenesis such as EMILIN1 and LOXL1, as well as elastin fragmentation. These findings suggested the crucial role of APOE in regulating elastogenesis during valve remodeling. Furthermore, cell-cell interaction analysis revealed that JAG1 from unidirectional VECs activates NOTCH signaling in Elastin-VICs through NOTCH3. In vitro Jag1 treatment in VICs increased elastogenesis, while similar observations were found in VICs co-cultured with VECs in the presence of unidirectional flow. Notably, we found that the JAG1-NOTCH3 signaling pair was drastically reduced in the PS valves. Lastly, we demonstrated that APOE is indispensable for JAG1-induced NOTCH activation in VICs, reinforcing the presence of a synergistic intrinsic and external regulatory network involving APOE and NOTCH signaling that is responsible for regulating elastogenesis during human valve remodeling. ConclusionscRNA-seq analysis of human fetal valves identified a novel Elastin-VIC subpopulation, and revealed mechanism of intrinsic APOE and external NOTCH signaling in regulating elastogenesis during cardiac valve remodeling. These mechanisms may contribute to deciphering the pathogenesis of elastin malformation in congenital valve diseases. Clinical Perspective What Is New?O_LIHigh-resolution single-cell transcriptome atlas generated from healthy human fetal heart valves and valves affected by pulmonary stenosis during the early phase of valve remodeling prior to birth. C_LIO_LIA unique subset of valve interstitial cells (VICs) that produce elastin (Elastin-VICs) was identified. C_LIO_LIElastin-VICs specifically located underneath the valve endothelial cells (VECs) sensing unidirectional flow, and played a crucial role in elastin maturation via the expression of APOE. C_LIO_LIElastin-VICs communicated with adjacent VECs via the JAG1-NOTCH signaling, facilitating elastin formation and valve remodeling. C_LI What Are the Clinical Implications?O_LIElastin-VICs from patient valvular tissues with Pulmonary Stenosis exhibit decreased APOE-NOTCH signaling and elastin fragmentation. C_LIO_LIDirect targeting of APOE and NOTCH signaling could be a novel approach to promote elastin fiber formation and valve remodeling in patients with valvular defects. C_LI

bioinformatics↗

bioRxiv2022

Cardiac fibroblasts regulate cardiomyocyte hypertrophy through dynamic regulation of type I collagen

RationaleCardiomyocytes and fibroblasts in the heart communicate through both secreted growth factors as well as through sensing the structural properties of the extracellular matrix that each helps generate. Previous studies have shown that defects in fibroblast activity during disease stimulation result in altered cardiomyocyte hypertrophy, although the role that collagen might play in this communication is unknown. ObjectiveHere we investigated how type I collagen maturation and disease-responsive matrix expansion in the heart by cardiac fibroblasts impacts cardiac fibrosis and cardiomyocyte hypertrophy. Methods and ResultsWe generated and characterized Col1a2-/- mice using standard gene-targeting. Col1a2-/- mice were viable, although by young adulthood their hearts showed alterations in extracellular matrix mechanical properties, as well as an unanticipated activation of cardiac fibroblasts and induction of a progressive fibrotic response. This included increases in fibroblast number and a progressive cardiac hypertrophy, with reduced functional performance by 9 months. Col1a2-loxP targeted mice were also generated and crossed with the tamoxifen-inducible Postn-MerCreMer knock-in mice to delete the Col1a2 gene in myofibroblasts post-pressure overload injury, to more specifically implicate fibroblasts as effectors of cardiomyocyte hypertrophy in vivo. Opposite to the gradual induction of cardiac hypertrophy observed in germline Col1a2-/- mice as they matured developmentally, adult fibroblast-specific deletion of Col1a2 during pressure overload protected these mice from cardiac hypertrophy in the first week with a delayed fibrotic response. However, this reduction in hypertrophy due to myofibroblast-specific Col1a2 deletion was gradually lost over 2 and 6 weeks of pressure overload as augmented fibrosis returned. ConclusionsDefective type I collagen in the developing heart alters the structural integrity of the extracellular matrix that leads to fibroblast expansion, activation, fibrosis and hypertrophy with progressive cardiomyopathy in adulthood. However, acute deletion of type I collagen production for the first time in the adult heart during pressure overload prevents ECM expansion and inhibits cardiomyocyte hypertrophy, while gradual restoration of fibrosis again permitted hypertrophy comparable to controls.

cell biology↗

bioRxiv2019

Cardiomyocyte cell cycling, maturation, and growth by multinucleation in postnatal swine

AimsCardiomyocyte (CM) cell cycle arrest, decline of mononucleated-diploid CMs, sarcomeric maturation, and extracellular matrix remodeling are implicated in loss of cardiac regenerative potential in mice after birth. Recent studies show a 3-day neonatal regenerative capacity in pig hearts similar to mice, but postnatal pig CM growth dynamics are unknown. We examined cardiac maturation in postnatal pigs and mice, to determine the relative timing of developmental events underlying heart growth and regenerative potential in large and small mammals.\n\nMethods and ResultsLeft ventricular tissue from White Yorkshire-Landrace pigs at postnatal day (P)0 to 6 months (6mo) was analyzed to span birth, weaning, and adolescence in pigs, compared to similar physiological timepoints in mice. Collagen remodeling increases by P7 in postnatal pigs, but sarcomeric and gap junctional maturation only occur at 2mo. Also, there is no postnatal transition to beta-oxidation metabolism in pig hearts. Mononucleated CMs, predominant at birth, persist to 2mo in swine, with over 50% incidence of mononucleated-diploid CMs at P7-P15. Extensive multinucleation with 4-16 nuclei per CM occurs beyond P30. Pigs also exhibit increased CM length relative to multinucleation, preceding increase in CM width at 2mo-6mo. Further, robust CM mitotic nuclear pHH3 activity and cardiac cell cycle gene expression is apparent in pig left ventricles up to 2mo. By contrast, in mice, these maturational events occur concurrently in the first two postnatal weeks alongside loss of cardiac regenerative capacity.\n\nConclusionsCardiac maturation occurs over a 6mo postnatal period in pigs, despite a similar early-neonatal heart regenerative window as mice. Postnatal pig CM growth includes increase in CM length alongside multinucleation, with CM cell cycle arrest and loss of mononucleated-diploid CMs occurring at 2mo-6mo. These CM characteristics are important to consider for pig preclinical studies and may offer opportunities to study aspects of heart regeneration unavailable in other models.

developmental biology↗

bioRxiv2019

SCAR FORMATION AND DECREASED CARDIAC FUNCTION FOLLOWING ISCHEMIA/REPERFUSION INJURY IN 1-MONTH-OLD SWINE

Studies in mice show a brief neonatal period of cardiac regeneration with minimal scar formation. Less is known about reparative mechanisms in large mammals. A transient cardiac injury approach (ischemia/reperfusion, IR) was used in weaned postnatal day (P)30 pigs, to assess regenerative repair in young large mammals. Female and male P30 pigs were subjected to cardiac ischemia (1 hour) by occlusion of the left anterior descending artery followed by reperfusion, or to sham operation. Following IR, myocardial damage occurred, with cardiac ejection fraction significantly decreased 2 hours post-ischemia. No improvement or worsening of cardiac function to the 4 week study end-point was observed. Histology demonstrated cardiomyocyte (CM) cell cycling at 2-months-of-age in multinucleated CMs in both sham-operated and IR pigs. Regional scar formation and inflammation in the epicardial region proximal to injury were observed 4 weeks post-IR. Sex differences were found, suggestive of females creating a greater fibrotic response with worse cardiac function, highlighting the importance of representing both sexes in cardiac injury studies. Together, our results describe an effective novel cardiac injury model in P30 swine, at a time when CMs are still cycling. Pigs subjected to IR show myocardial damage with a prolonged decrease in cardiac function, formation of a small, regional scar with increased inflammation. These data demonstrate that P30 pigs do not regenerate myocardium, even in the presence of CM mitotic activity, but form a scar after transient IR injury.\n\nNEW & NOTEWORTHYHere, we report for the first time ischemia/reperfusion (IR) cardiac injury in 1-month-old (P30) pigs. This model of IR injury highlights lack of cardiac regeneration, even in the presence of cardiomyocyte (CM) cell cycling in young swine. An effective injury approach is described for use in large mammals to investigate cardiac function, CM cell cycling, extracellular matrix (ECM) remodeling, and gene expression changes, while highlighting the importance of studying both sexes.

developmental biology↗