casein-kinase-ii and Hypertrophy

Bone morphogenetic protein 2 regulates chondrogenesis and cartilage formation. However, it also induces chondrocyte hypertrophy and cartilage matrix degradation. We recently designed three peptides CK2.1, CK2.2, and CK2.3 that activate the BMP signaling pathways by releasing casein kinase II (CK2) from distinct sites at the bone morphogenetic protein receptor type Ia (BMPRIa). Since BMP2 is a major regulator of chondrogenesis and the peptides activated BMP signaling in a similar way, we evaluated the effect of these peptides on chondrogenesis and cartilage formation. C3H10T1/2 cells were stimulated with CK2.1, CK2.2, and CK2.3 and evaluated for the chondrogenic and osteogenic potential. For chondrogenesis, Alcian blue staining was performed. Additionally, collagen types II and X expression was measured. For osteogenesis, osteocalcin and von Kossa staining were performed. From the three peptides, CK2.1 was the most promising peptide to induce chondrogenesis but not osteogenesis. To investigate the effect of CK2.1 on articular cartilage formation in vivo, we injected CK2.1 into the tail vein of mice. Injection of CK2.1 into the tail vein of mice led to increased articular cartilage formation but not BMD. In sharp contrast, injection of BMP2 led to increased BMD and expression of collagen type X, a marker of chondrocyte hypertrophy. MMP13 expression was unchanged. Our study demonstrates that CK2.1 drives chondrogenesis and cartilage formation without induction of chondrocyte hypertrophy. Peptide CK2.1 may, therefore, be a valuable therapeutic for cartilage degenerative diseases. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:876-885, 2017.

Topics: Animals; Bone Density; Bone Morphogenetic Protein 2; Bone Morphogenetic Protein Receptors, Type I; Cartilage, Articular; Casein Kinase II; Cell Differentiation; Chondrogenesis; Female; Hypertrophy; Matrix Metalloproteinase 13; Mice; Mice, Inbred C3H; Mice, Inbred C57BL; Microscopy, Fluorescence; Osteogenesis; Peptides; Smad Proteins

Cardiac hypertrophy is regulated by a complex interplay of pro- and anti-hypertrophic factors. Here, we report a novel anti-hypertrophic pathway composed of catalase, protein kinase CK2 (CK2), and apoptosis repressor with caspase recruitment domain (ARC). Our results showed that ARC phosphorylation levels, CK2 activity, and catalase expression levels were decreased in the hearts of the angiotensinogen transgenic mice and in cardiomyocytes treated with the hypertrophic stimuli, including phenylephrine, tumor necrosis factor-alpha, and angiotensin II. To understand the role of ARC in hypertrophy, we observed that enforced expression of ARC could inhibit hypertrophy. Knockdown of endogenous ARC or inhibition of its phosphorylation could sensitize cardiomyocytes to undergoing hypertrophy. The phosphorylatable, but not the nonphosphorylatable, ARC could inhibit hypertrophy. Thus, ARC is able to inhibit hypertrophy in a phosphorylation-dependent manner. In exploring the molecular mechanism by which CK2 activity is reduced, we found that CK2 was carbonylated in angiotensinogen transgenic mice and in cardiomyocytes treated with the hypertrophic stimuli. The decrease in catalase expression led to an elevated level of reactive oxygen species. The latter oxidatively modified CK2, resulting in its carbonylation. CK2 lost its catalytic activity upon carbonylation. ARC is phosphorylated by CK2, and ARC phosphorylation levels were reduced as a consequence of the decrease of CK2 activity. To understand the molecular mechanism by which ARC inhibits hypertrophy, we observed that ARC could inhibit the activation of mitochondrial permeability transition. These results suggest that catalase, CK2, and ARC constitute an anti-hypertrophic pathway in the heart.

Topics: Angiotensin II; Angiotensinogen; Animals; Apoptosis Regulatory Proteins; Cardiotonic Agents; Casein Kinase II; Catalase; Cell Membrane Permeability; Down-Regulation; Gene Expression Regulation, Enzymologic; Hypertrophy; Mice; Mice, Transgenic; Mitochondria, Heart; Muscle Proteins; Myocytes, Cardiac; Oxidation-Reduction; Phenylephrine; Phosphorylation; Protein Carbonylation; Protein Processing, Post-Translational; Rats; Rats, Wistar; Reactive Oxygen Species; Tumor Necrosis Factor-alpha

Emerging data suggest that urokinase-type plasminogen activator (UPA), beyond its role in pericellular proteolysis, may also act as a mitogen. We investigated the function of endogenous UPA in mediating the mitogenic effects of platelet-derived growth factor (PDGF) and basic fibroblast growth factor (bFGF) on human vascular smooth muscle cells (SMC). Growth-arrested SMC constitutively expressed UPA, but UPA expression and secretion increased several times upon stimulation with either PDGF or bFGF. Inhibition of endogenous UPA with a polyclonal antibody significantly reduced DNA synthesis and proliferation of PDGF or bFGF stimulated SMC, this effect already being evident when the cells entered S-phase. The proliferative activity of endogenous UPA was dependent on a functional catalytic domain as demonstrated by inhibition experiments with a specific monoclonal antibody (394OA) and p-aminobenzamidine, respectively. In contrast, neither plasmin generation nor binding of UPA to its receptor (CD87) were required for UPA-mediated mitogenic effects. The results demonstrate that endogenous UPA is not only overexpressed in SMC upon stimulation with PDGF/bFGF, but also mediates the mitogenic activity of the growth factors in a catalytic-domain-dependent manner. Specific inhibition of this UPA domain may represent an attractive target for pharmacological interventions in atherogenesis and restenosis after angioplasty.

Topics: Arteriosclerosis; Casein Kinase II; Catalytic Domain; Cell Division; Cells, Cultured; DNA; Dose-Response Relationship, Drug; Drug Administration Schedule; Enzyme Inhibitors; Fibrinolysin; Fibroblast Growth Factor 2; Graft Occlusion, Vascular; Humans; Hypertrophy; Muscle, Smooth, Vascular; Platelet-Derived Growth Factor; Protein Serine-Threonine Kinases; Receptors, Cell Surface; Receptors, Urokinase Plasminogen Activator; Up-Regulation; Urokinase-Type Plasminogen Activator

Hypertrophy of vascular smooth muscle cells (VSMC) is an important adaptive response of hypertension. Drug intervention studies have implicated a role for angiotensin II (A-II) in the mediation of VSMC hypertrophy in vivo, and A-II is a potent hypertrophic agent for VSMC in culture. Our laboratory has previously shown that A-II-induced hypertrophy of cultured VSMC is due in part to generalized increases in protein synthesis and increased content of rRNA. The aim of the present study was to determine if A-II stimulates rRNA gene synthesis and whether the rRNA transcription factor, upstream binding factor (UBF), is involved. Nuclear run-on analysis demonstrated that A-II induced a greater than 5-fold increase in rRNA gene synthesis within 6 h of stimulation. A-II also stimulated a rapid increase in UBF phosphorylation as well as nucleolar localization, but no changes in the content of UBF. Phosphoamino acid analysis showed that phosphorylation occurred only on serine residue(s). Results demonstrate that increased transcription of ribosomal DNA contributes to the A-II-induced increase in protein synthesis and VSMC hypertrophy, and suggest that an important regulatory event in this pathway may be the phosphorylation and/or nucleolar localization of UBF.

Topics: Angiotensin II; Animals; Casein Kinase II; Cells, Cultured; DNA-Binding Proteins; Hypertrophy; Muscle, Smooth, Vascular; Phosphorylation; Pol1 Transcription Initiation Complex Proteins; Protein Serine-Threonine Kinases; Rats; RNA, Ribosomal, 18S; Serine; Transcription Factors; Transcription, Genetic