PHD3 mediates denervation skeletal muscle atrophy through Nf‐κB signal pathway

Abstract

Skeletal muscle is the largest organ of the body, the development of skeletal muscle is very important for the health of the animal body. Prolyl hydroxylases (PHDs) are the classical regulator of the hypoxia inducible factor (HIF) signal pathway, many researchers found that PHDs are involved in the muscle fiber type transformation, muscle regeneration, and myocyte differentiation. However, whether PHDs can impact the protein turnover of skeletal muscle is poorly understood. In this study, we constructed denervated muscle atrophy mouse model and found PHD3 was highly expressed in the atrophic muscles and there was a significant correlation between the expression level of PHD3 and skeletal muscle weight which was distinct from PHD1 and PHD2. Then, the similar results were getting from the different weight muscles of normal mice. To further verify the relationship between PHD3 and skeletal muscle protein turnover, we established a PHD3 interference model by injecting PHD3 sgRNA virus into tibialis anterior muscle (TA) muscle of MCK‐Cre‐cas9 mice and transfecting PHD3 shRNA lentivirus into primary satellite cells. It was found that the Knock‐out of PHD3 in vivo led to a significant increase in muscle weight and muscle fiber area (P < .05). Besides, the activity of protein synthesis signal pathway increased significantly, while the protein degradation pathway was inhibited evidently (P < .05). In vitro, the results of 5‐ethynyl‐2′‐deoxyuridine (EdU) and tetramethylrhodamine ethyl ester (TMRE) fluorescence detection showed that PHD3 interference could lead to a decrease in cell proliferation and an increase of cell apoptosis. After the differentiation of satellite cells, the production of puromycin in the interference group was higher than that in the control group, and the content of 3‐methylhistidine in the interference group was lower than that in the control group (P < .05) which is consistent with the change of protein turnover signal pathway in the cell. Mechanistically, there is an interaction between PHD3, NF‐κB, and IKBα which was detected by immunoprecipitation. With the interfering of PHD3, the expression of the inflammatory signal pathway also significantly decreased (P < .05). These results suggest that PHD3 may affect protein turnover in muscle tissue by mediating inflammatory signal pathway. Finally, we knocked out PHD3 in denervated muscle atrophy mice and LPS‐induced myotubes atrophy model. Then, we found that the decrease of PHD3 protein level could alleviate the muscle weight and muscle fiber reduction induced by denervation in mice. Meanwhile, the protein level of the inflammatory signal pathway and the content of 3‐methylhistidine in denervated atrophic muscle were also significantly reduced (P < .05). In vitro, PHD3 knock‐out could alleviate the decrease of myotube diameter induced by LPS, and the expression of protein synthesis pathway was also significantly increased (P < .05). On the contrary, the expression level of protein degradation and inflammatory signal pathway was significantly...

Morroniside ameliorates inflammatory skeletal muscle atrophy via inhibiting canonical and non-canonical NF-κB and regulating protein synthesis/degradation

“…Our and other studies have shown that NF-κB signaling is involved in RA-, aging-, and denervation-related muscle atrophy in animal models (Li et al, 2020;Li et al, 2021b). To investigate if this signaling pathway in response to TNFα is associated with muscle atrophy and grip strength in human, we conducted a pathway-based genetic association study in population, and found that the weighted genetic risk score of NF-κB pathway in response to TNFα (including 198 genes) was significantly associated with grip strength, suggesting this TNFα-induced NF-κB pathway is related to muscle atrophy in human (Supplementary Table S3; Table 1).…”

Combined effects of functional overload and denervation on skeletal muscle mass and its regulatory proteins in mice

“…Jun N-terminus kinase (JNK) and STE20-like protein kinase activation induced by oxidative and genotoxic stress causes the nuclear translocation of FoxO independent of Akt regulation (Hay, 2011). It has been reported that DEN increased the JNK phosphorylation in mouse muscle (Li et al, 2021). These findings suggest that FoxOmediated ubiquitin-proteasome system-dependent proteolysis in concurrent loaded muscle is independent of Akt regulation.…”

Muscle cell‐derived cytokines in skeletal muscle regeneration

“…To that end, the advent of CRISPR/Cas9 gene editing technology should enable more rapid generation of mice with muscle‐specific ablation of cytokines. For instance, the recently reported transgenic mice with Cas9 expression driven by human skeletal actin (HSA)‐Cre [177] and muscle creatine kinase (MCK)‐Cre [178] would offer a convenient way to knock out or knock down cytokine genes of interest in muscle by delivering specific sgRNAs. Muscle‐specific activation of cytokine gene expression would also be informative and potentially applicable in therapeutics, and the CRISPR activation technology (CRISPRa) provides a powerful tool [179].…”