Pathological hypertrophy amelioration by PRAS40-mediated inhibition of mTORC1.

Mechanistic target of rapamycin complex 1 (mTORC1), necessary for cellular growth, is regulated by intracellular signaling mediating inhibition of mTORC1 activation. Among mTORC1 regulatory binding partners, the role of Proline Rich AKT Substrate of 40 kDa (PRAS40) in controlling mTORC1 activity and cellular growth in response to pathological and physiological stress in the heart has never been addressed. This report shows PRAS40 is regulated by AKT in cardiomyocytes and that AKT-driven phosphorylation relieves the inhibitory function of PRAS40. PRAS40 overexpression in vitro blocks mTORC1 in cardiomyocytes and decreases pathological growth. Cardiomyocyte-specific overexpression in vivo blunts pathological remodeling after pressure overload and preserves cardiac function. Inhibition of mTORC1 by PRAS40 preferentially promotes protective mTORC2 signaling in chronic diseased myocardium. In contrast, strong PRAS40 phosphorylation by AKT allows for physiological hypertrophy both in vitro and in vivo, whereas cardiomyocyte-specific overexpression of a PRAS40 mutant lacking capacity for AKT-phosphorylation inhibits physiological growth in vivo, demonstrating that AKT-mediated PRAS40 phosphorylation is necessary for induction of physiological hypertrophy. Therefore, PRAS40 phosphorylation acts as a molecular switch allowing mTORC1 activation during physiological growth, opening up unique possibilities for therapeutic regulation of the mTORC1 complex to mitigate pathologic myocardial hypertrophy by PRAS40.

Differential regulation of cellular senescence and differentiation by prolyl isomerase Pin1 in cardiac progenitor cells.

Autologous c-kit(+) cardiac progenitor cells (CPCs) are currently used in the clinic to treat heart disease. CPC-based regeneration may be further augmented by better understanding molecular mechanisms of endogenous cardiac repair and enhancement of pro-survival signaling pathways that antagonize senescence while also increasing differentiation. The prolyl isomerase Pin1 regulates multiple signaling cascades by modulating protein folding and thereby activity and stability of phosphoproteins. In this study, we examine the heretofore unexplored role of Pin1 in CPCs. Pin1 is expressed in CPCs in vitro and in vivo and is associated with increased proliferation. Pin1 is required for cell cycle progression and loss of Pin1 causes cell cycle arrest in the G1 phase in CPCs, concomitantly associated with decreased expression of Cyclins D and B and increased expression of cell cycle inhibitors p53 and retinoblastoma (Rb). Pin1 deletion increases cellular senescence but not differentiation or cell death of CPCs. Pin1 is required for endogenous CPC response as Pin1 knock-out mice have a reduced number of proliferating CPCs after ischemic challenge. Pin1 overexpression also impairs proliferation and causes G2/M phase cell cycle arrest with concurrent down-regulation of Cyclin B, p53, and Rb. Additionally, Pin1 overexpression inhibits replicative senescence, increases differentiation, and inhibits cell death of CPCs, indicating that cell cycle arrest caused by Pin1 overexpression is a consequence of differentiation and not senescence or cell death. In conclusion, Pin1 has pleiotropic roles in CPCs and may be a molecular target to promote survival, enhance repair, improve differentiation, and antagonize senescence.

Notch activation enhances lineage commitment and protective signaling in cardiac progenitor cells.

Phase I clinical trials applying autologous progenitor cells to treat heart failure have yielded promising results; however, improvement in function is modest, indicating a need to enhance cardiac stem cell reparative capacity. Notch signaling plays a crucial role in cardiac development, guiding cell fate decisions that underlie myocyte and vessel differentiation. The Notch pathway is retained in the adult cardiac stem cell niche, where level and duration of Notch signal influence proliferation and differentiation of cardiac progenitors. In this study, Notch signaling promotes growth, survival and differentiation of cardiac progenitor cells into smooth muscle lineages in vitro. Cardiac progenitor cells expressing tamoxifen-regulated intracellular Notch1 (CPCeK) are significantly larger and proliferate more slowly than control cells, exhibit elevated mTORC1 and Akt signaling, and are resistant to oxidative stress. Vascular smooth muscle and cardiomyocyte markers increase in CPCeK and are augmented further upon ligand-mediated induction of Notch signal. Paracrine signals indicative of growth, survival and differentiation increase with Notch activity, while markers of senescence are decreased. Adoptive transfer of CPCeK into infarcted mouse myocardium enhances preservation of cardiac function and reduces infarct size relative to hearts receiving control cells. Greater capillary density and proportion of vascular smooth muscle tissue in CPCeK-treated hearts indicate improved vascularization. Finally, we report a previously undescribed signaling mechanism whereby Notch activation stimulates CPC growth, survival and differentiation via mTORC1 and paracrine factor expression. Taken together, these findings suggest that regulated Notch activation potentiates the reparative capacity of CPCs in the treatment of cardiac disease.

Regulation of cardiac hypertrophic signaling by prolyl isomerase Pin1.

RATIONALE: Cardiac hypertrophy results from the complex interplay of differentially regulated cascades based on the phosphorylation status of involved signaling molecules. Although numerous critical regulatory kinases and phosphatases have been identified in the myocardium, the intracellular mechanism for temporal regulation of signaling duration and intensity remains obscure. In the nonmyocyte context, control of folding, activity, and stability of proteins is mediated by the prolyl isomerase Pin1, but the role of Pin1 in the heart is unknown. OBJECTIVE: To establish the role of Pin1 in the heart. METHODS AND RESULTS: Here, we show that either genetic deletion or cardiac overexpression of Pin1 blunts hypertrophic responses induced by transaortic constriction and consequent cardiac failure in vivo. Mechanistically, we find that Pin1 directly binds to Akt, mitogen activated protein kinase (MEK), and Raf-1 in cultured cardiomyocytes after hypertrophic stimulation. Furthermore, loss of Pin1 leads to diminished hypertrophic signaling of Akt and MEK, whereas overexpression of Pin1 increases Raf-1 phosphorylation on the autoinhibitory site Ser259, leading to reduced MEK activation. CONCLUSIONS: Collectively, these data support a role for Pin1 as a central modulator of the intensity and duration of 2 major hypertrophic signaling pathways, thereby providing a novel target for regulation and control of cardiac hypertrophy.

Malignancy Risk of Thyroid Nodules That Are Not Classifiable by the American Thyroid Association Ultrasound Risk Stratification System: A Systematic Review and Meta-Analysis.

Background: Sonographic evaluation is fundamental to thyroid nodule assessment. The American Thyroid Association (ATA) ultrasound risk stratification system (USRSS) is widely used, but the appearance of some nodules has been considered nonclassifiable (NC-ATA). The risk of malignancy (RoM) of NC-ATA nodules varies widely between studies, leading to uncertainty in clinical management. The aim of this study was to comprehensively evaluate the prevalence and malignancy risk of NC-ATA nodules. Methods: A systematic review was performed searching PubMed/MEDLINE and EMBASE to identify original studies of thyroid nodules classified using the ATA USRSS from 2016 to 2022 and reporting the outcome of NC-ATA nodules. Meta-analysis was conducted to obtain pooled RoM estimates and meta-regression sensitivity analyses were used to explore sources of between-study heterogeneity. Results: Of 6377 screened studies, 135 underwent full-text review, and 16 studies reporting 21,271 nodules were included. Within these, the pooled prevalence of NC-ATA nodules was 7.8% (1872 nodules; [confidence interval; CI 5.1-11.1]). The pooled RoM estimate for NC-ATA nodules was 20.3% [CI 13.0-28.7] and there was significant heterogeneity between studies (I2 = 92.8%, p < 0.001). NC-ATA nodule RoM estimates were significantly different by study type: single-center versus multicenter studies (24.8% vs. 12.3%, respectively, p = 0.031) and study design: retrospective versus prospective studies (25.1% vs. 8.5%, respectively, p = 0.003). No significant difference was observed in RoM based on inclusion of <1 cm nodules or geographic region. Meta-regression analysis showed study design and use of surgical histology for diagnostic criteria contributed significantly to differences in the reported RoM estimates. Conclusion: In this first meta-analysis comprehensively assessing the RoM of NC-ATA nodules, the malignancy risk was found to be comparable with the current ATA USRSS intermediate suspicion category. Significant heterogeneity was observed between studies and limits the interpretation of these results. In future iterations of the ATA USRSS that seek into incorporate categorization of NC-ATA nodules, these meta-analysis data may help to inform proper malignancy risk stratification. The study protocol was registered on PROSPERO, the international prospective register of systematic reviews (CRD42020182498), on July 14, 2020.