Transforming growth factor-beta (TGF-β)/bone morphogenic protein (BMP) signaling is involved in a vast majority of cellular processes and is fundamentally important throughout life. TGF-β/BMPs have widely recognized roles in bone formation during mammalian development and exhibit versatile regulatory functions in the body. Signaling transduction by TGF-β/BMPs is specifically through both canonical Smad-dependent pathways (TGF-β/BMP ligands, receptors and Smads) and non-canonical Smad-independent signaling pathway (e.g. p38 mitogen-activated protein kinase pathway, MAPK). Following TGF-β/BMP induction, both the Smad and p38 MAPK pathways converge at the Runx2 gene to control mesenchymal precursor cell differentiation. The coordinated activity of Runx2 and TGF-β/BMP-activated Smads is critical for formation of the skeleton. Recent advances in molecular and genetic studies using gene targeting in mice enable a better understanding of TGF-β/BMP signaling in bone and in the signaling networks underlying osteoblast differentiation and bone formation. This review summarizes the recent advances in our understanding of TGF-β/BMP signaling in bone from studies of genetic mouse models and human diseases caused by the disruption of TGF-β/BMP signaling. This review also highlights the different modes of cross-talk between TGF-β/BMP signaling and the signaling pathways of MAPK, Wnt, Hedgehog, Notch, and FGF in osteoblast differentiation and bone formation.

Soluble factors such as polypeptide growth factors, mitogenic lipids, inflammatory cytokines, and hormones are known regulators of cell proliferation. However, the effect of mechanical stimuli on cell proliferation is less well understood. Here we examined the effect of low intensity pulsed ultrasound (US), which is used to promote wound healing, on the proliferation of primary human foreskin fibroblasts and the underlying signaling mechanisms. We show that a single 6 -11-min US stimulation increases bromodeoxyuridine incorporation. In addition, an increase in the total cell number is observed after sequential US stimulation. US induced stress fiber and focal adhesion formation via activation of Rho. We further observed that US selectively induced activation of extracellular signal-regulated kinase (ERK) 1/2. Inhibition of Rho-associated coiled-coil-containing protein kinase (ROCK) prevented US-induced ERK1/2 activation, demonstrating that the Rho/ROCK pathway is an upstream regulator of ERK activation in response to US. Consequently, activation of ROCK and MEK-1 was required for US-induced DNA synthesis. Finally, an integrin ␤ 1 blocking antibody as well as a RGD peptide prevented US-induced DNA synthesis. In addition, US slightly increased phosphorylation of Src at Tyr 416 , and Src activity was found to be required for ERK1/2 activation in response to US. In conclusion, our data demonstrate for the first time that US promotes cell proliferation via activation of integrin receptors and a Rho/ROCK/Src/ERK signaling pathway.Signal transduction mechanisms of receptor tyrosine kinase or heptahelical receptors have been studied extensively over the last years. However, the effect of acoustic pulsed energy on cell growth and the signal transduction mechanisms induced by this type of mechanical stimulation are not well understood. Previous studies have shown that mechanical stress such as stretch and shear stress can induce DNA synthesis in certain cell types and that extracellular signal-regulated kinase (ERK) 1 1/2 can be activated by mechanical stress (1-4).Low intensity pulsed ultrasound (US) is a special type of acoustic pulsed energy that is increasingly used as a supplementary therapy to promote bone and wound healing (5). US, transmitting as an acoustic pressure wave and applying mechanical stress indirectly to the tissues, has been reported to promote osteogenesis and protein synthesis, calcium uptake, and DNA synthesis in different cells (6 -9). US-induced DNA synthesis seems cell type-dependent; US promotes DNA synthesis in human osteoblasts, gingival fibroblasts, and periosteal cells (6, 8, 10), but not in chondrocytes (11,12). However, the molecular mechanisms by which US induces DNA synthesis or even cell proliferation are largely unknown.There is growing evidence that integrins are promising candidates for sensing extracellular matrix-derived mechanical stimuli and converting them into biochemical signals (13,14). Integrin-associated signaling pathways include an increase in tyrosine phosphorylation of...

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Previously, RNA transcripts of cDNA clones of hepatitis C virus (HCV) genotypes 1a (strains H77, HCV-1, and HC-TN), 1b (HC-J4, Con1, and HCV-N), and 2a (HC-J6 and JFH1) were found to be infectious in chimpanzees. However, only JFH1 was infectious in human hepatoma Huh7 cells. We performed genetic analysis of HCV genotype 3a (strain S52) and 4a (strain ED43) prototype strains and generated full-length consensus cDNA clones (pS52 and pED43). Transfection of Huh7.5 cells with RNA transcripts of these clones did not yield cells expressing HCV Core. However, intrahepatic transfection of chimpanzees resulted in robust infection with peak HCV RNA titers of ϳ5.5 log 10 international units (IU)/ml. Genomic consensus sequences recovered from serum at the times of peak viral titers were identical to the sequences of the parental plasmids. Both chimpanzees developed acute hepatitis with elevated liver enzymes and significant necroinflammatory liver changes coinciding with detection of gamma interferon-secreting, intrahepatic T cells. However, the onset and broadness of intrahepatic T-cell responses varied greatly in the two animals, with an early (week 4) multispecific response in the ED43-infected animal (3 weeks before the first evidence of viral control) and a late (week 11) response with limited breadth in the S52-infected animal (without evidence of viral control). Autologous serum neutralizing antibodies were not detected during the acute infection in either animal. Both animals became persistently infected. In conclusion, we generated fully functional infectious cDNA clones of HCV genotypes 3a and 4a. Proof of functionality of all genes might further the development of recombinant cell culture systems for these important genotypes.Hepatitis C virus (HCV) is a small, enveloped virus with a single-stranded RNA genome, approximately 9.6 kb in length. The genome consists of 5Ј and 3Ј untranslated regions (UTRs) and a single open reading frame (ORF), encoding structural proteins (Core, E1, and E2), p7, and nonstructural proteins (NS2, NS3, NS4A, NS4B, NS5A, and NS5B) (22). Due to significant genetic heterogeneity, HCV was classified into 7 major genotypes and numerous subtypes, differing Ͼ30% and Ͼ20%, respectively, at the nucleotide level and at the amino acid level. Strains/isolates differ in 2 to 10% at the nucleotide/ amino acid level, and quasispecies typically differ in up to 2% at the nucleotide/amino acid level (70). As a main cause of liver cirrhosis and hepatocellular carcinoma, chronic HCV infection poses a major public health burden. There is no vaccine available, and combination therapy with alpha interferon and ribavirin is characterized by many side effects and contraindications, as well as low efficacy (22). Research on the HCV life cycle and new therapeutics requires well-characterized experimental models and reagents representing the different virus variants.Chimpanzees, the only animal model of HCV infection mirroring immunopathogenesis and viral persistence observed in human infections (4, 80), can be ...