In chick skeletal muscle cell primary cultures, we previously demonstrated that 1alpha,25(OH)2-vitamin D3 [1alpha,25(OH)2D3], the hormonally active form of vitamin D, increases the phosphorylation and activity of the extracellular signal-regulated mitogen-activated protein (MAP) kinase isoforms ERK1 and ERK2, their subsequent translocation to the nucleus and involvement in DNA synthesis stimulation. In this study, we show that other members of the MAP kinase superfamily are also activated by the hormone. Using the muscle cell line C2C12 we found that 1alpha,25(OH)2D3 within 1 min phosphorylates and increases the activity of p38 MAPK. The immediately upstream mitogen-activated protein kinase kinases 3/6 (MKK3/MKK6) were also phosphorylated by the hormone suggesting their participation in p38 activation. 1Alpha,25(OH)2D3 was able to dephosphorylate/activate the ubiquitous cytosolic tyrosine kinase c-Src in C2C12 cells and studies with specific inhibitors imply that Src participates in hormone induced-p38 activation. Of relevance, 1alpha,25(OH)2D3 induced in the C2C12 line the stimulation of mitogen-activated protein kinase activating protein kinase 2 (MAPKAP-kinase 2) and subsequent phosphorylation of heat shock protein 27 (HSP27) in a p38 kinase activation-dependent manner. Treatment with the p38 inhibitor, SB203580, blocked p38 phosphorylation caused by the hormone and inhibited the phosphorylation of its downstrean substrates. 1Alpha,25(OH)2D3 also promotes the phosphorylation of c-jun N-terminal protein kinases (JNK 1/2), the response is fast (0.5-1 min) and maximal phosphorylation of the enzyme is observed at physiological doses of 1alpha,25(OH)2D3 (1 nM). The relative contribution of ERK-1/2, p38, and JNK-1/2 and their interrelationships in hormonal regulation of muscle cell proliferation and differentiation remain to be established.
17b-Estradiol (E 2 ) stimulates the mitogen-activated protein kinases (MAPKs) in various cellular types. We have shown that the hormone activates extracellular-regulated kinase (ERK) and p38 MAPK in skeletal muscle cells. However, the functions of MAPK modulation by the estrogen in muscle cells have not been studied yet. We have recently reported antiapoptotic actions of E 2 in C2C12 cells. Here, the role of MAPKs mediating the hormone effect in muscle cells was investigated. The results showed that cells exposed to 0 . 5 mM hydrogen peroxide (H 2 O 2 ) presented cytoskeleton disorganization, mitochondrial redistribution, and picnotic/ fragmented nuclei. Pretreatment with 10K8 M E 2 prevented these morphological apoptotic characteristics, except in the presence of ERK or p38 MAPK inhibitors, U0126 and SB203580 respectively. Mitochondrial membrane integrity was also studied. Preincubation of cultures with 10 K8 M E 2 abrogated H 2 O 2 effects such as Janus Green oxidation, presence of cytochrome c oxidase activity in the cytoplasm, and SMAC/DIABLO release from mitochondria. When MAPKs were inhibited, the hormone could not prevent mitochondrial membrane damage exerted by oxidative stress. Blocking experiments with small interfering RNAs confirmed that both ERK and p38 MAPKs mediate the antiapoptotic effects of the hormone at the mitochondrial level. Further, some of the molecular mechanisms involved were also investigated. Thus, E 2 was able to induce AKT (Ser473) and BAD (Ser112) phosphorylation in C2C12 cells in the absence or in the presence of H 2 O 2 but not when the cultures were incubated with H 2 O 2 and MAPK inhibitors. Altogether, these results show that E 2 exerts a survival action in skeletal muscle cells involving ERK and p38 MAPK activation.
Background: Contribution of resorption to bone anabolism by PTH receptor signaling in osteocytes is unknown. Results: Pharmacologic/genetic approaches demonstrated that remodeling-or modeling-based bone formation differentially operate in specific surfaces. Conclusion: Resorption is critical for anabolism in periosteal/endocortical bone surfaces, but tempers bone gain in cancellous bone. Significance: Targeting bone compartment-specific actions of PTH receptor signaling could enhance the therapeutic potential of the pathway.
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