Dongchao Lu

BACKGROUND: Exercise training has been demonstrated to provide direct endogenous cardioprotection and to reduce cardiovascular disease risk. Skeletal muscle functions not only as an organ for movement but also as a vital secretory organ. However, how muscle-heart crosstalk contributes to exercise-induced cardioprotection remains largely unclear. Here, we investigated the cardioprotective effects of exercise-induced small extracellular vesicles (sEVs) derived from skeletal muscle in myocardial ischemia/reperfusion (MI/R) injury in mice. METHODS: An adeno-associated virus 8-dMCK (double-muscle creatine kinase)-CD63-EGFP (enhanced green fluorescent protein) reporter system was used to track skeletal muscle-derived sEVs in the heart of mice after 4 weeks of exercise. To determine whether sEVs from exercised skeletal muscle mediate cardioprotection, cardiac sEV abundance was increased by intramyocardial injection and decreased by generating skeletal muscle-specific Rab27a knockout mice. The molecular mechanism was elucidated by profiling sEV-enriched microRNAs and validating their functions in cardiomyocytes and mouse MI/R injury models. The translational potential was then assessed in human exercise cohorts. Finally, the therapeutic efficacy was evaluated using miRNA agomirs and adeno-associated virus 8-mediated, skeletal muscle-specific overexpression of miRNAs in MI/R injury models. RESULTS: Exercise promoted the entry of skeletal muscle-derived sEVs into the heart. Skeletal muscle-derived sEVs from exercised mice significantly attenuated MI/R injury by reducing cardiomyocyte apoptosis and improving mitochondrial respiration. Exercise-mediated protection against MI/R injury was attenuated after inhibiting the secretion of sEVs from skeletal muscle. Mechanistically, exercise-enriched sEVs delivered miR-451a and miR-143-3p to cardiomyocytes, thereby modulating the PERK (protein kinase R-like endoplasmic reticulum kinase)-ATF4-CHOP stress-response pathway and the USP46/P53 axis, thus reducing cardiomyocyte death. Skeletal muscle-specific overexpression of miR-451a/143-3p recapitulated the cardioprotective effects of exercise in vivo. Furthermore, exercising humans also showed elevated levels of miR-451a/143-3p in their skeletal muscle and circulating sEVs. CONCLUSIONS: Our study reveals a novel exercise-induced cardioprotective mechanism via muscle-heart crosstalk, highlighting a promising therapeutic approach for treating cardiovascular disease.

BACKGROUND: Extracellular vesicles (EVs) are involved in exercise-induced cardiac protection. However, the effects and underlying mechanisms of tissue-specific molecular cargo packaged within these EVs, including PIWI (P-element induced wimpy testis)-interacting RNA (piRNA), remain poorly understood. In particular, the mechanistic contribution of exercised intracardiac EV-associated piRNAs to doxorubicin-induced cardiotoxicity (DCT) has not been defined. METHODS: Transgenic reporter mice and a cardiomyocyte-specific Rab27a (RAB27A, member RAS oncogene family) knockout strategy were used to investigate the contribution of cardiomyocyte-derived EVs to exercise-induced cardioprotection against DCT. To screen functionally relevant cardioprotective piRNA cargo, heart tissues from patients with dilated cardiomyopathy and experimental DCT models were analyzed, including cardiomyocyte-specific knockout mice, human embryonic stem cell–derived cardiomyocytes, and primary murine cardiomyocytes. RESULTS: We found that cardiomyocyte-derived EVs post-exercise were enriched for a cardiac-specific protective piRNA (piR-mmu-57256903), designated as an exercise-induced protective piRNA (EPPIR). EPPIR levels were significantly reduced in heart tissue from patients with dilated cardiomyopathy and DCT models. Functionally, EPPIR protects the heart against DCT by regulating KDM6B (lysine [K]-specific demethylase 6B)-H3K27me3 (trimethylated histone H3 at lysine 27)-Dtna epigenetic axis. In addition, EPPIR acts as a cardiomyocyte-specific suppressor of Tp53 (tumor protein p53). CONCLUSIONS: We identify a previously unrecognized role for cardiomyocyte-derived EV-associated piRNA EPPIR in mediating exercise-induced cardioprotection. EPPIR exerts its protective effects through coordinated regulation of the KDM6B-Dtna axis and cardiomyocyte-specific suppressor of Tp53, providing mechanistic insight and highlighting a potential therapeutic strategy for DCT.

Skeletal muscle diseases pose a serious threat to human health, yet effective pharmacological treatments remain scarce. RNA-based therapeutics are emerging as a promising avenue: an RNA drug that restores muscle function has already been approved and is in clinical use, highlighting the potential of RNA therapy in this approach. This review surveys the current landscape of RNA therapies for muscle disorders, focusing on the correction of aberrant messenger RNA (mRNA) metabolism, RNA modifications, and the therapeutic manipulation of noncoding RNAs in skeletal muscle diseases. We outline RNA-directed strategies in skeletal muscle, identify the advantages and challenges of RNA therapy in muscle diseases, and discuss future directions for the field.

Pulmonary Fibrosis (PF) is a life‐threatening illness that is characterized by progressive scarring in the lung interstitium. There is an urgent need for new PF therapies because current treatments only slow down the progression of fibrosis, and the median life expectancy post‐diagnosis is only 4–6 years. Since PF patients frequently exhibit telomere attrition, overexpressing telomerase, the enzyme responsible for synthesizing telomeres, represents a compelling therapeutic option. In this study, we in vitro transcribed human telomerase reverse transcriptase (hTERT) mRNA using modified nucleosides (modRNA). ModRNA hTERT treatment led to transient activation of telomerase activity in a dose‐dependent manner in MRC‐5 cells and, importantly, in primary human alveolar type II pneumocytes. Consequently, the proliferative capacity was increased, concomitant with reduced DNA damage and elongated telomere length. Notably, the induction of cellular immune response was only detectable at the highest modRNA concentration and returned to normal levels within 48 h. Next, we demonstrated that circularized, exonuclease‐resistant modRNA hTERT extended the transient expression profile, which may be clinically advantageous. Finally, we provided therapeutic proof of concept in organotypic 3D ex vivo human precision‐cut lung slices derived from end‐stage PF patients. Intriguingly, a single modRNA hTERT treatment inhibited senescence, as indicated by significantly lower levels of senescence‐associated β‐galactosidase. Pro‐inflammatory markers (IL6 and IL8) and, concurrently, the key fibrosis mediators TGFβ and COL1A1 were markedly reduced after modRNA and circular RNA hTERT treatment. In conclusion, the data presented herein provide initial evidence for the potential of RNA‐based hTERT therapy for treating human lung fibrosis.

Circular RNAs (circRNAs) have emerged as pivotal regulators in the posttranscriptional regulatory landscape, exhibiting unique properties such as increased stability and a range of biological functions due to their covalently closed-loop structure. Despite advancements in circRNA research, the field meets a significant challenge in the absence of a standardized nomenclature system, leading to confusion and hindering effective scientific communication. This chapter outlines the historical development, current state, and the necessity for a unified naming system in circRNA research. We highlight the role of databases in shaping circRNA nomenclature and discuss the implications of nomenclature ambiguity, especially with the identification of multiple circRNAs from single gene loci. We propose a systematic approach to naming circRNAs that incorporates the “circ” prefix, references to GENCODE/Ensembl transcripts, and annotations for unique structural features such as retained introns or alternative splicing. The adoption of such a nomenclature system is essential for enhancing clarity, ensuring reproducibility, and advancing our understanding of circRNA investigation.

Pulmonary Fibrosis (PF) is a life-threatening illness that is characterized by progressive scarring in the lung interstitium. There is an urgent need for new PF therapies because current treatments only slow down the progression of fibrosis and the median life expectancy post-diagnosis is only 4 to 6 years. Since PF patients frequently exhibit telomere attrition, overexpressing telomerase, the enzyme responsible for synthesizing telomeres represents a compelling therapeutic option. In this study, we in vitro transcribed human telomerase reverse transcriptase (hTERT) mRNA using modified nucleosides (modRNA). ModRNA hTERT treatment led to transient activation of telomerase activity in a dose-dependent manner in MRC-5 cells and, importantly, in primary human alveolar type II pneumocytes (ATII cells). Consequently, the proliferative capacity was increased, concomitant with reduced DNA damage and elongated telomere length. Notably, induction of cellular immune response was only detectable at the highest modRNA concentration, and returned to normal levels within 48 hours. Next, we demonstrated that circularized, exonuclease-resistant modRNA hTERT extended the transient expression profile which may be clinically advantageous. Finally, we provided therapeutic proof of concept in organotypic 3D ex vivo human precision-cut lung slices derived from end-stage PF patients. Intriguingly, a single modRNA hTERT treatment inhibited senescence as indicated by significantly lower levels of senescence-associated β-galactosidase, and pro-inflammatory IL6 and IL8. Concurrently, the key fibrosis mediators TGFβ and COL1A1 were markedly reduced. In conclusion, the data presented herein provide initial evidence for the potential of RNA-based hTERT therapy for treating human lung fibrosis.

Alessia Costa, Hannah Jill Hunkler, Shambhabi Chatterjee, Sarah Cushman, Erika Hilbold, Ke Xiao, Dongchao Lu, Julia Leonardy, Malte Juchem, Marida Sansonet