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Modifications in the interactions of the muscle cytoskeleton with the cell membrane occur during cell growth and adaptation, although the mechanisms regulating these interactions are unknown. We have observed that myotendinous junctions (MTJs), which are the primary sites of turnover of the thin filament-membrane associations in skeletal muscle, are greatly enriched in receptors for PDGF. The high concentration of PDGF receptors at MTJs suggested to us that receptor binding may initiate cytoskeletal remodeling in skeletal muscle. We tested this possibility by examining the organization and phosphorylation of cytoskeletal components of L6 myocytes after PDGF stimulation. We have found that 10 min after PDGF stimulation, L6 myoblasts exhibit no stress fibers discernible by phalloidin binding, and that vinculin relocates from focal contacts into a diffuse cytoplasmic distribution. After 60 min of incubation, these changes are largely reversed. Indirect immunofluorescence shows that at 10-min PDGF stimulation, there are no changes in the distribution of talin, the beta 1 subunit of integrin, pp125FAK or desmin. Phosphotyrosine distribution changes upon stimulation from focal contacts to being located both in focal contacts and granules concentrated in perinuclear regions. These granules also immunolabel with anti-PDGF receptor Immunoprecipitations with anti-phosphotyrosine show that polypeptides at 180 and 230 kD show the greatest increase in tyrosine phosphorylation after PDGF stimulation. Immunoblots of anti-phosphotyrosine precipitates show that these polypeptides are the PDGF receptor and talin. We also examined the possibility that the cytoskeletal reorganization observed may result from calpain activation caused by elevated intracellular calcium induced by PDGF stimulation. However, immunoblots of control and stimulated cells show no decrease in the inactive calpain proenzyme or increase in the proteolytic, autolyzed forms of calpain pursuant to stimulation. Furthermore, stimulation produces no increase in the proportion of the 190-kD talin fragment characteristic of calpain-mediated cleavage. The retention of talin and integrin at focal contacts after talin phosphorylation, while vinculin is redistributed, indicate that phosphorylation of talin in PDGF-stimulated cells leads to separation of talin-vinculin associations but not talin-integrin associations. We propose that PDGF binding to PDGF receptors at MTJs may provide one means of regulating myofibril associations with the muscle cell membrane.

The vascular endothelium is critical for induction of appropriate lineage differentiation in organogenesis. In this study, we report that dysfunctional pulmonary endothelium, resulting from the loss of matrix Gla protein (MGP), causes ectopic hepatic differentiation in the pulmonary epithelium. We demonstrate uncontrolled induction of the hepatic growth factor (HGF) caused by dysregulated cross talk between pulmonary endothelium and epithelium in Mgp -null lungs. Elevated HGF induced hepatocyte nuclear factor 4 α (Hnf4a), which competed with NK2 homeobox 1 (Nkx2.1) for binding to forkhead box A2 (Foxa2) to drive hepatic differentiation in Mgp -null airway progenitor cells. Limiting endothelial HGF reduced Hnf4a, abolished interference of Hnf4a with Foxa2, and reduced hepatic differentiation in Mgp -null lungs. Together, our results suggest that endothelial-epithelial interactions, maintained by MGP, are essential in pulmonary cell differentiation., (© 2017 Yao et al.)

In the degenerative disease Duchenne muscular dystrophy, inflammatory cells enter muscles in response to repetitive muscle damage. Immune factors are required for muscle regeneration, but chronic inflammation creates a profibrotic milieu that exacerbates disease progression. Osteopontin (OPN) is an immunomodulator highly expressed in dystrophic muscles. Ablation of OPN correlates with reduced fibrosis and improved muscle strength as well as reduced natural killer T (NKT) cell counts. Here, we demonstrate that the improved dystrophic phenotype observed with OPN ablation does not result from reductions in NKT cells. OPN ablation skews macrophage polarization toward a pro-regenerative phenotype by reducing M1 and M2a and increasing M2c subsets. These changes are associated with increased expression of pro-regenerative factors insulin-like growth factor 1, leukemia inhibitory factor, and urokinase-type plasminogen activator. Furthermore, altered macrophage polarization correlated with increases in muscle weight and muscle fiber diameter, resulting in long-term improvements in muscle strength and function in mdx mice. These findings suggest that OPN ablation promotes muscle repair via macrophage secretion of pro-myogenic growth factors., (© 2016 Capote et al.)

Dystrophin-deficient muscles experience large reductions in expression of nitric oxide synthase (NOS), which suggests that NO deficiency may influence the dystrophic pathology. Because NO can function as an antiinflammatory and cytoprotective molecule, we propose that the loss of NOS from dystrophic muscle exacerbates muscle inflammation and fiber damage by inflammatory cells. Analysis of transgenic mdx mice that were null mutants for dystrophin, but expressed normal levels of NO in muscle, showed that the normalization of NO production caused large reductions in macrophage concentrations in the mdx muscle. Expression of the NOS transgene in mdx muscle also prevented the majority of muscle membrane injury that is detectable in vivo, and resulted in large decreases in serum creatine kinase concentrations. Furthermore, our data show that mdx muscle macrophages are cytolytic at concentrations that occur in dystrophic, NOS-deficient muscle, but are not cytolytic at concentrations that occur in dystrophic mice that express the NOS transgene in muscle. Finally, our data show that antibody depletions of macrophages from mdx mice cause significant reductions in muscle membrane injury. Together, these findings indicate that macrophages promote injury of dystrophin-deficient muscle, and the loss of normal levels of NO production by dystrophic muscle exacerbates inflammation and membrane injury in muscular dystrophy.