PKCε is a regulator of hypertrophic differentiation of chondrocytes in osteoarthritis.

OBJECTIVE: Osteoarthritis (OA) is a common and highly debilitating degenerative disease whose complex pathogenesis and the multiplicity of the molecular processes involved, hinder its complete understanding. Protein Kinase C (PKC) novel isozyme PKCε recently proved to be an interesting molecule for further investigations as it can represent an intriguing, new actor in the acquisition of a OA phenotype by the chondrocyte. DESIGN: PKCε was modulated in primary chondrocytes from human OA patient knee cartilage samples by means of short hairpin RNA (ShRNA) and the expression of cartilage specific markers observed at mRNA and protein level. The involvement of Histone deacetylases (HDACs) signaling pathway was also investigated through the use of specific inhibitors MS-275 and Inhibitor VIII. RESULTS: PKCε loss induces up-regulation of Runt-domain transcription factor (RUNX2), Metalloproteinase 13 (MMP13) and Collagen X (COL10) as well as an enhanced calcium deposition in OA chondrocyte cultures. In parallel, PKCε knock-down also leads to SOX9 and Collagen II (COL2) down-modulation and to a lower deposition of glycosaminoglycans (GAGs) in the extracellular matrix (ECM). This novel regulatory role of PKCε over cartilage hypertrophic phenotype is exerted via an HDAC-mediated pathway, as HDAC2 and HDAC4 expression is modulated by PKCε. HDAC2 and HDAC4, in turn, are at least in part responsible for the modulation of the master transcription factors RUNX2 and SOX9, key regulators of chondrocyte phenotype. CONCLUSIONS: PKCε prevents the phenotypic progression of the OA chondrocyte, acting on cartilage specific markers through the modulation of the transcription factors SOX9 and RUNX2. The loss of PKCε enhances, in fact, the OA hypertrophic phenotype, with clear implications in the pathophysiology of the disease.

PKCε as a novel promoter of skeletal muscle differentiation and regeneration.

INTRODUCTION: Satellite cells are muscle resident stem cells and are responsible for muscle regeneration. In this study we investigate the involvement of PKCε during muscle stem cell differentiation in vitro and in vivo. Here, we describe the identification of a previously unrecognized role for the PKCε-HMGA1 signaling axis in myoblast differentiation and regeneration processes. METHODS: PKCε expression was modulated in the C2C12 cell line and primary murine satellite cells in vitro, as well as in an in vivo model of muscle regeneration. Immunohistochemistry and immunofluorescence, RT-PCR and shRNA silencing techniques were used to determine the role of PKCε and HMGA1 in myogenic differentiation. RESULTS: PKCε expression increases and subsequently re-localizes to the nucleus during skeletal muscle cell differentiation. In the nucleus, PKCε blocks Hmga1 expression to promote Myogenin and Mrf4 accumulation and myoblast formation. Following in vivo muscle injury, PKCε accumulates in regenerating, centrally-nucleated myofibers. Pharmacological inhibition of PKCε impairs the expression of two crucial markers of muscle differentiation, namely MyoD and Myogenin, during injury induced muscle regeneration. CONCLUSION: This work identifies the PKCε-HMGA1 signaling axis as a positive regulator of skeletal muscle differentiation.

Protein kinase Cɛ inhibition restores megakaryocytic differentiation of hematopoietic progenitors from primary myelofibrosis patients.

Among the three classic Philadelphia chromosome-negative myeloproliferative neoplasms, primary myelofibrosis (PMF) is the most severe in terms of disease biology, survival and quality of life. Abnormalities in the process of differentiation of PMF megakaryocytes (MKs) are a hallmark of the disease. Nevertheless, the molecular events that lead to aberrant megakaryocytopoiesis have yet to be clarified. Protein kinase Cɛ (PKCɛ) is a novel serine/threonine kinase that is overexpressed in a variety of cancers, promoting aggressive phenotype, invasiveness and drug resistance. Our previous findings on the role of PKCɛ in normal (erythroid and megakaryocytic commitment) and malignant (acute myeloid leukemia) hematopoiesis prompted us to investigate whether it could be involved in the pathogenesis of PMF MK-impaired differentiation. We demonstrate that PMF megakaryocytic cultures express higher levels of PKCɛ than healthy donors, which correlate with higher disease burden but not with JAK2V617F mutation. Inhibition of PKCɛ function (by a negative regulator of PKCɛ translocation) or translation (by target small hairpin RNA) leads to reduction in PMF cell growth, restoration of PMF MK differentiation and inhibition of PKCɛ-related anti-apoptotic signaling (Bcl-xL). Our data suggest that targeting PKCɛ directly affects the PMF neoplastic clone and represent a proof-of-concept for PKCɛ inhibition as a novel therapeutic strategy in PMF.

PKCε is a negative regulator of PVAT-derived vessel formation.

RATIONALE: Vessel formation is a crucial event in tissue repair after injury. Thus, one assumption of innovative therapeutic approaches is the understanding of its molecular mechanisms. Notwithstanding our knowledge of the role of Protein Kinase C epsilon (PKCε) in cardio-protection and vascular restenosis, its role in vessel progenitor differentiation remains elusive. OBJECTIVE: Given the availability of PKCε pharmacological modulators already tested in clinical trials, the specific aim of this study is to unravel the role of PKCε in vessel progenitor differentiation, with implications in vascular pathology and vasculogenesis. METHODS AND RESULTS: Mouse Peri-Vascular Adipose Tissue (PVAT) was used as source of mesenchymal vessel progenitors. VEGF-induced differentiation of PVAT cells down-regulates both PKCε and p-PAK1 protein expression levels. PKCε overexpression and activation: i) reduced the expression levels of SMA and PECAM in endothelial differentiation of PVAT cells; ii) completely abrogated tubules formation in collagen gel assays; iii) increased the expression of p-PAK1. CONCLUSION: PKCε negatively interferes with vessel progenitor differentiation via interaction with PAK-1.

Chronic expanding hematoma might be a potential insidious challenge for orthopedic surgeon.

Chronic expanding hematomas can cause alarm both to the physician and to the patient because they simulate the growth of a malignant tumor. It is not always possible to bring back the cause to a specific traumatic event that the patient can remember. At this purpose, it is important to have a precise diagnosis when dealing with a growing mass and to exclude any malignancy before processing any treatment. In this article, we report the case of a young patient admitted to our department with a suspected soft tissue sarcoma, but imaging study and histological examination revealed to be hematoma. The authors want to emphasize the necessity of performing a complete clinical and instrumental study when surgeon has to deal with a growing mass. From accurate analysis of imaging, it is often possible to discriminate between malignancy and other benign forms.

Gap balancing versus measured resection technique using a mobile-bearing prosthesis in computer-assisted surgery.

Navigation has been developed to help surgeons install implants more accurately and reproducibly; at the same time, this tool is able to record quantitative information such as joint range of motion, laxity and kinematics intra-operatively. As for standard surgery, two strategies are possible to achieve either femoral component rotation or overall prosthetic alignment: a measured gap resection approach, in which bone landmarks are used to guide resections equal to the distal and posterior thickness of the femoral component, or a gap-balancing technique, in which equal collateral ligament tension in flexion and extension is tried to find before as a guide to final bone cuts. The purpose of this paper is to compare the two different methods in a 67 patients group submitted to the same procedure using mobile-bearing (MB) prosthesis in order to analyse the effect of both techniques on joint line maintenance, axial limb restoration and components position. The gap group (GG) consists of 31 patients in whom the arthroplasty was performed using a navigated gap-balancing technique. The measured group (MG) consists of 36 patients in whom a computer-assisted measured resection technique was used. The results of imaging and the number of outliers were not statistically different (P = 0.56) for the mechanical axis and prosthetic positioning between the two groups. The gap technique showed a statistically significant alteration of the post-operative value when compared with the measured resection technique, (P = 0.036). The mean elevation of the joint line was 4.09 mm for the GG and 3.50 mm in the MG.