ABSTRACT
Lamin A/C gene (LMNA) mutations contribute to severe striated muscle laminopathies, affecting cardiac and skeletal muscles, with limited treatment options. In this study, we delve into the investigations of five distinct LMNA mutations, including three novel variants and two pathogenic variants identified in patients with muscular laminopathy. Our approach employs zebrafish models to comprehensively study these variants. Transgenic zebrafish expressing wild-type LMNA and each mutation undergo extensive morphological profiling, swimming behavior assessments, muscle endurance evaluations, heartbeat measurement, and histopathological analysis of skeletal muscles. Additionally, these models serve as platform for focused drug screening. We explore the transcriptomic landscape through qPCR and RNAseq to unveil altered gene expression profiles in muscle tissues. Larvae of LMNA(L35P), LMNA(E358K), and LMNA(R453W) transgenic fish exhibit reduced swim speed compared to LMNA(WT) measured by DanioVision. All LMNA transgenic adult fish exhibit reduced swim speed compared to LMNA(WT) in T-maze. Moreover, all LMNA transgenic adult fish, except LMNA(E358K), display weaker muscle endurance than LMNA(WT) measured by swimming tunnel. Histochemical staining reveals decreased fiber size in all LMNA mutations transgenic fish, excluding LMNA(WT) fish. Interestingly, LMNA(A539V) and LMNA(E358K) exhibited elevated heartbeats. We recognize potential limitations with transgene overexpression and conducted association calculations to explore its effects on zebrafish phenotypes. Our results suggest lamin A/C overexpression may not directly impact mutant phenotypes, such as impaired swim speed, increased heart rates, or decreased muscle fiber diameter. Utilizing LMNA zebrafish models for drug screening, we identify L-carnitine treatment rescuing muscle endurance in LMNA(L35P) and creatine treatment reversing muscle endurance in LMNA(R453W) zebrafish models. Creatine activates AMPK and mTOR pathways, improving muscle endurance and swim speed in LMNA(R453W) fish. Transcriptomic profiling reveals upstream regulators and affected genes contributing to motor dysfunction, cardiac anomalies, and ion flux dysregulation in LMNA mutant transgenic fish. These findings faithfully mimic clinical manifestations of muscular laminopathies, including dysmorphism, early mortality, decreased fiber size, and muscle dysfunction in zebrafish. Furthermore, our drug screening results suggest L-carnitine and creatine treatments as potential rescuers of muscle endurance in LMNA(L35P) and LMNA(R453W) zebrafish models. Our study offers valuable insights into the future development of potential treatments for LMNA-related muscular laminopathy.
Subject(s)
Animals, Genetically Modified , Carnitine , Creatine , Lamin Type A , Muscle, Skeletal , Mutation , Zebrafish , Animals , Lamin Type A/genetics , Lamin Type A/metabolism , Muscle, Skeletal/metabolism , Muscle, Skeletal/pathology , Muscle, Skeletal/drug effects , Creatine/metabolism , Carnitine/metabolism , Disease Models, Animal , Laminopathies/genetics , Laminopathies/metabolism , Swimming , Transcriptome , HumansABSTRACT
Carassius auratus complex formula (CACF) is a traditional Chinese medicine known for its antidiabetic effects. Hepatocellular carcinoma (HCC) is a major cause of cancer-related deaths worldwide, and there are currently no effective therapies for advanced HCC. This study explores the comprehensive effects and possible mechanisms of CACF on HCC. The results show that CACF reduces the viability of hepatoma cells in vitro, while benefiting normal hepatocytes. In addition, CACF inhibits hepatoma cell growth in a zebrafish xenotransplantation model and decreases lipid accumulation, represses inflammation and cell proliferation markers in fatty acid translocase (CD36) transgenic zebrafish, and inhibits the expression of cell proliferation and ß-catenin downstream targets in telomerase (tert) transgenic zebrafish models. Ingenuity Pathway Analysis reveals that CACF exerts multiple functions, including reduction of inflammation and inhibition of lipid transporter and PPAR signaling pathway. Surprisingly, CACF also regulates the expression of genes and reduces coronavirus infection and pathogenesis in a zebrafish model. CACF treatment is validated to regulate the expression of genes for anti-coronavirus activity. Mechanistically, CACF stabilizes G-quadruplex and reduces cell senescence associated ß-galactosidase activity. In summary, CACF may be a promising therapeutic agent with multiple functions including anticancer, anti-inflammation, and anti-microorganisms in a zebrafish model.
