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Theranostics ; 11(16): 7879-7895, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34335970


Rationale: Previous studies have shown that human embryonic stem cell-derived cardiomyocytes improved myocardial recovery when administered to infarcted pig and non-human primate hearts. However, the engraftment of intramyocardially delivered cells is poor and the effectiveness of clinically relevant doses of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) in large animal models of myocardial injury remains unknown. Here, we determined whether thymosin ß4 (Tb4) could improve the engraftment and reparative potency of transplanted hiPSC-CMs in a porcine model of myocardial infarction (MI). Methods: Tb4 was delivered from injected gelatin microspheres, which extended the duration of Tb4 administration for up to two weeks in vitro. After MI induction, pigs were randomly distributed into 4 treatment groups: the MI Group was injected with basal medium; the Tb4 Group received gelatin microspheres carrying Tb4; the CM Group was treated with 1.2 × 108 hiPSC-CMs; and the Tb4+CM Group received both the Tb4 microspheres and hiPSC-CMs. Myocardial recovery was assessed by cardiac magnetic resonance imaging (MRI), arrhythmogenesis was monitored with implanted loop recorders, and tumorigenesis was evaluated via whole-body MRI. Results: In vitro, 600 ng/mL of Tb4 protected cultured hiPSC-CMs from hypoxic damage by upregulating AKT activity and BcL-XL and promoted hiPSC-CM and hiPSC-EC proliferation. In infarcted pig hearts, hiPSC-CM transplantation alone had a minimal effect on myocardial recovery, but co-treatment with Tb4 significantly enhanced hiPSC-CM engraftment, induced vasculogenesis and the proliferation of cardiomyocytes and endothelial cells, improved left ventricular systolic function, and reduced infarct size. hiPSC-CM implantation did not increase incidence of ventricular arrhythmia and did not induce tumorigenesis in the immunosuppressed pigs. Conclusions: Co-treatment with Tb4-microspheres and hiPSC-CMs was safe and enhanced the reparative potency of hiPSC-CMs for myocardial repair in a large-animal model of MI.

Infarto do Miocárdio/terapia , Miócitos Cardíacos/metabolismo , Timosina/farmacologia , Animais , Diferenciação Celular , Proliferação de Células , Células Cultivadas , China , Modelos Animais de Doenças , Células Endoteliais/patologia , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Infarto do Miocárdio/metabolismo , Miocárdio/patologia , Regeneração , Transplante de Células-Tronco/métodos , Suínos , Timosina/metabolismo , Timosina/fisiologia
Artigo em Inglês | MEDLINE | ID: mdl-33446047


Three-dimensional (3D) printed scaffolds have recently emerged as an innovative treatment option for patients with critical-sized skin wounds. Current approaches to managing life-threatening wounds include skin grafting and application of commercially sourced skin substitutes. However, these approaches are not without several challenges. Limited donor tissue and donor site morbidity remain a concern for tissue grafting, while engineered skin substitutes fail to fully recapitulate the complex native environment required for wound healing. The implementation of 3D printed dermal scaffolds offers a potential solution for these shortcomings. Spatial control over scaffold structure, the ability to incorporate multiple materials and bioactive ingredients, enables the creation of conditions specifically optimized for wound healing. Three-dimensional bioprinting, a subset of 3D printing, allows for the replacement of lost cell populations and secreted active compounds that contribute to tissue repair and recovery. The replacement of damaged and lost cells delivers beneficial effects directly, or synergistically, supporting injured tissue to recover its native state. Despite encouraging results, the promise of 3D printed scaffolds has yet to be realized. Further improvements to current material formulations and scaffold designs are required to achieve the goal of clinical adoption. Herein, we provide an overview of 3D printing techniques and discuss several strategies for healing of full-thickness wounds by using 3D printed acellular scaffolds or bioprinted cellular scaffolds, aimed at translating this technology to the clinical management of skin lesions. We identify the challenges associated with designing and optimizing printed tissue replacements, and discuss the future perspectives of this emerging option for managing patients who present with critical-sized life-threatening cutaneous wounds.

PLoS One ; 13(10): e0204853, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30273379


The transcription factor NF-E2 Related Factor-2 (NRF2) is an important drug target. Activation of NRF2 has chemopreventive effects in cancer and exerts beneficial effects in a number of diseases, including neurodegenerative diseases, inflammatory diseases, hepatosteatosis, obesity and insulin resistance. Hence, there have been great efforts to discover and characterize novel NRF2 activators. One reported NRF2 activator is the labdane diterpenoid andrographolide. In this study, we identified the mechanism through which andrographolide activates NRF2. We showed that andrographolide inhibits the function of KEAP1, a protein that together with CUL3 and RBX1 forms an E3 ubiquitin ligase that polyubiquitinates NRF2. Andrographolide partially inhibits the interaction of KEAP1 with CUL3 in a manner dependent on Cys151 in KEAP1. This suggests that andrographolide forms Michael acceptor dependent adducts with Cys151 in KEAP1 in vivo, leading to inhibition of NRF2 ubiquitination and consequently accumulation of the transcription factor. Interestingly, we also showed that at higher concentrations andrographolide increases NRF2 protein expression in a Cys151 independent, but likely KEAP1 dependent manner, possibly through modification of other Cys residues in KEAP1. In this study we also screened secondary metabolites produced by endophytes isolated from non-flowering plants for NRF2-inducing properties. One of the extracts, ORX 41, increased both NRF2 protein expression and transcriptional activity markedly. These results suggest that endophytes isolated from non-flowering or other plants may be a good source of novel NRF2 inducing compounds.

Proteínas Culina/metabolismo , Diterpenos/farmacologia , Endófitos/química , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Extratos Vegetais/farmacologia , Sítios de Ligação/efeitos dos fármacos , Briófitas/química , Proteínas de Transporte/metabolismo , Proteínas Culina/química , Diterpenos/química , Gleiquênias/química , Regulação da Expressão Gênica/efeitos dos fármacos , Células HEK293 , Humanos , Proteína 1 Associada a ECH Semelhante a Kelch/química , Fator 2 Relacionado a NF-E2 , Extratos Vegetais/química , Ligação Proteica/efeitos dos fármacos , Metabolismo Secundário , Ubiquitinação
Circulation ; 138(24): 2798-2808, 2018 12 11.
Artigo em Inglês | MEDLINE | ID: mdl-30030417


BACKGROUND: The adult mammalian heart has limited ability to repair itself after injury. Zebrafish, newts, and neonatal mice can regenerate cardiac tissue, largely by cardiac myocyte (CM) proliferation. It is unknown whether hearts of young large mammals can regenerate. METHODS: We examined the regenerative capacity of the pig heart in neonatal animals (ages 2, 3, or 14 days postnatal) after myocardial infarction or sham procedure. Myocardial scar and left ventricular function were determined by cardiac magnetic resonance imaging and echocardiography. Bromodeoxyuridine pulse-chase labeling, histology, immunohistochemistry, and Western blotting were performed to study cell proliferation, sarcomere dynamics, and cytokinesis and to quantify myocardial fibrosis. RNA-sequencing was also performed. RESULTS: After myocardial infarction, there was early and sustained recovery of cardiac function and wall thickness in the absence of fibrosis in 2-day-old pigs. In contrast, older animals developed full-thickness myocardial scarring, thinned walls, and did not recover function. Genome-wide analyses of the infarct zone revealed a strong transcriptional signature of fibrosis in 14-day-old animals that was absent in 2-day-old pigs, which instead had enrichment for cytokinesis genes. In regenerating hearts of the younger animals, up to 10% of CMs in the border zone of the myocardial infarction showed evidence of DNA replication that was associated with markers of myocyte division and sarcomere disassembly. CONCLUSIONS: Hearts of large mammals have regenerative capacity, likely driven by cardiac myocyte division, but this potential is lost immediately after birth.

Coração/fisiologia , Infarto do Miocárdio/patologia , Animais , Animais Recém-Nascidos , Citocinese/genética , Ecocardiografia , Fibrose , Imagem Cinética por Ressonância Magnética , Infarto do Miocárdio/diagnóstico por imagem , Miocárdio/patologia , Miócitos Cardíacos/fisiologia , Regeneração , Suínos , Troponina I/análise , Função Ventricular Esquerda
J Cardiovasc Transl Res ; 11(5): 375-392, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-29557052


Cardiomyocytes derived from human pluripotent stem cells (hPSCs) are emerging as an invaluable alternative to primarily sourced cardiomyocytes. The potentially unlimited number of hPSC-derived cardiomyocytes (hPSC-CMs) that may be obtained in vitro facilitates high-throughput applications like cell transplantation for myocardial repair, cardiotoxicity testing during drug development, and patient-specific disease modeling. Despite promising progress in these areas, a major disadvantage that limits the use of hPSC-CMs is their immaturity. Improvements to the maturity of hPSC-CMs are necessary to capture physiologically relevant responses. Herein, we review and discuss the different maturation strategies undertaken by others to improve the morphology, contractility, electrophysiology, and metabolism of these derived cardiomyocytes.

Diferenciação Celular , Descoberta de Drogas/métodos , Miócitos Cardíacos/fisiologia , Células-Tronco Pluripotentes/fisiologia , Transplante de Células-Tronco/métodos , Animais , Cardiotoxicidade , Fármacos Cardiovasculares/farmacologia , Linhagem da Célula , Células Cultivadas , Humanos , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/transplante , Fenótipo , Células-Tronco Pluripotentes/efeitos dos fármacos , Células-Tronco Pluripotentes/metabolismo , Células-Tronco Pluripotentes/transplante , Testes de Toxicidade/métodos