Aging‐related hyperphosphatemia impairs myogenic differentiation and enhances fibrosis in skeletal muscle

Abstract

Background Hyperphosphatemia has been related to the development of sarcopenia in aging mice. We describe the intracellular mechanisms involved in the impairment of the myogenic differentiation promoted by hyperphosphatemia and analyse these mechanisms in the muscle from older mice. Methods C 2 C 12 cells were grown in 2% horse serum in order to promote myogenic differentiation, in the presence or absence of 10 mM beta-glycerophosphate (BGP) for 7 days. Troponin T, paired box 7 (Pax-7), myogenic factor 5 (Myf5), myogenic differentiation 1 (MyoD), myogenin (MyoG), myocyte enhancer factor 2 (MEF2C), P300/CBP-associated factor (PCAF), histone deacetylase 1 (HDAC1), fibronectin, vimentin, and collagen I were analysed at 48, 72, and 168 h, by western blotting or by immunofluorescence staining visualized by confocal microscopy. Studies in mice were performed in 5-and 24-month-old C57BL6 mice. Three months before sacrifice, 21-month-old mice were fed with a standard diet or a low phosphate diet, containing 0.6% or 0.2% phosphate, respectively. Serum phosphate concentration was assessed by a colorimetric method and forelimb strength by a grip test. Fibrosis was observed in the tibialis anterior muscle by Sirius Red staining. In gastrocnemius muscle, MyoG, MEF2C, and fibronectin expressions were analysed by western blotting. Results Cells differentiated in the presence of BGP showed near five times less expression of troponin T and kept higher levels of Pax-7 than control cells indicating a reduced myogenic differentiation. BGP reduced Myf5 about 50% and diminished MyoD transcriptional activity by increasing the expression of HDAC1 and reducing the expression of PCAF. Consequently, BGP reduced to 50% the expression of MyoG and MEF2C. A significant increase in the expression of fibrosis markers as collagen I, vimentin, and fibronectin was found in cells treated with BGP. In mice, serum phosphate (17.24 ± 0.77 mg/dL young; 23.23 ± 0.81 mg/dL old; 19.09 ± 0.75 mg/dL old with low phosphate diet) correlates negatively (r = À0.515, P = 0.001) with the muscular strength (3.13 ± 0.07 gf/g young; 1.70 ± 0.12 gf/ g old; 2.10 ± 0.09 gf/g old with low phosphate diet) and with the expression of MyoG (r = À0.535, P = 0.007) and positively with the expression of fibronectin (r = 0.503, P = 0.001) in gastrocnemius muscle. The tibialis anterior muscle from old mice showed muscular fibrosis. Older mice fed with a low phosphate diet showed improved muscular parameters relative to control mice of similar age. Conclusions Hyperphosphatemia impairs myogenic differentiation, by inhibiting the transcriptional activity of MyoD, and enhances the expression of fibrotic genes in cultured myoblasts. Experiments carried out in older mice demonstrate a close relationship between age-related hyperphosphatemia and the decrease in the expression of myogenic factors and the increase in factors related to muscle fibrosis.

Age-Related Changes in the Matrisome of the Mouse Skeletal Muscle

“…Fibronectin, for instance, is one of the most widespread glycoproteins of the ECM, playing a role in various processes such as cellular adhesion, spreading and migration, as well as cellular development and differentiation. The increased abundance of fibronectin in old gastrocnemius is in agreement with very recent data demonstrating an association between high fibronectin levels in the aged gastrocnemius muscle, the reduced muscular strength, and myogenic regeneration and differentiation [62].…”

Histone Deacetylase 1 Expression and Regulatory Network in Lung Adenocarcinoma Based on Data Mining and Implications for Targeted Treatment

“…The protein encoded by HDAC1 is part of the histone deacetylase complex. Current drug progresses related to HDAC1 show that HDACi can increase the acetylation degree of intracellular histones and nonhistones, improve the expression level of P21, P53, and other genes, and then achieve the effect of inhibiting tumor cells [ 36 , 37 ]. This kind of drug can also show a good synergistic therapeutic effect when combined with a variety of chemotherapy drugs and has low toxicity in the effective inhibitory dose range, which has made HDACi a new targeted antitumor drug with wide application.…”

Spatial transcriptomics reveal markers of histopathological changes in Duchenne muscular dystrophy mouse models

“…Vim was expressed in non-fibrotic spots, but at significantly lower levels compared to fibrotic spots (log2(FC) = 1.11, adjusted p-value = 1.71e-10). Vim encodes for the myofibroblastic protein vimentin and is expected to be upregulated in areas of skeletal muscle fibrosis [42][43][44] . Given the role of Vim in focal adhesion, where regulatory signals and mechanical force is transmitted between the extracellular matrix (ECM) and an interacting cell 45 , expression was found in healthy tissue areas as well (Figure 4E-DBA/2J).…”