Signaling networks and transcription factors regulating mechanotransduction in bone.

Mechanical stimulation has a critical role in the development and maintenance of the skeleton. This function requires the perception of extracellular stimuli as well as their conversion into intracellular biochemical responses. This process is called mechanotransduction and is mediated by a plethora of molecular events that regulate bone metabolism. Indeed, mechanoreceptors, such as integrins, G protein-coupled receptors, receptor protein tyrosine kinases, and stretch-activated Ca(2+) channels, together with their downstream effectors coordinate the transmission of load-induced signals to the nucleus and the expression of bone-related genes. During the past decade, scientists have gained increasing insight into the molecular networks implicated in bone mechanotransduction. In the present paper, we consider the major signaling cascades and transcription factors that control bone and cartilage mechanobiology and discuss the influence of the mechanical microenvironment on the determination of skeletal morphology.

Lysyl oxidase interacts with AGE signalling to modulate collagen synthesis in polycystic ovarian tissue.

Connective tissue components--collagen types I, III and IV--surrounding the ovarian follicles undergo drastic changes during ovulation. Abnormal collagen synthesis and increased volume and density of ovarian stroma characterize the polycystic ovary syndrome (PCOS). During the ovulatory process, collagen synthesis is regulated by prolyl hydroxylase and lysyl oxidase (LOX) activity in ovarian follicles. LOX catalyzes collagen and elastin cross-linking and plays essential role in coordinating the control of ovarian extracellular matrix (ECM) during follicular development. We have recently shown accumulation of advanced glycation end products (AGEs), molecules that stimulate ECM production and abnormal collagen cross-linking, in ovarian tissue. However, the possible link between LOX and AGEs-induced signalling in collagen production and stroma formation in ovarian tissue from PCOS remains elusive. The present study investigates the hypothesis of AGE signalling pathway interaction with LOX gene activity in polycystic ovarian (PCO) tissue. We show an increased distribution and co-localization of LOX, collagen type IV and AGE molecules in the PCO tissue compared to control, as well as augmented expression of AGE signalling mediators/effectors, phospho(p)-ERK, phospho(p)-c-Jun and nuclear factor κB (NF-κB) in pathological tissue. Moreover, we demonstrate binding of AGE-induced transcription factors, NF-κB and activator protein-1 (AP-1) on LOX promoter, indicating a possible involvement of AGEs in LOX gene regulation, which may account for the documented increase in LOX mRNA and protein levels compared to control. These findings suggest that deposition of excess collagen in PCO tissue that induces cystogenesis may, in part, be due to AGE-mediated stimulation of LOX activity.

Mechanotransduction in osteoblast regulation and bone disease.

Osteoblasts are key components of the bone multicellular unit and have a seminal role in bone remodeling, which is an essential function for the maintenance of the structural integrity and metabolic capacity of the skeleton. The coordinated function of skeletal cells is regulated by several hormones, growth factors and mechanical cues that act via interconnected signaling networks, resulting in the activation of specific transcription factors and, in turn, their target genes. Bone cells are responsive to mechanical stimuli and this is of pivotal importance in developing biomechanical strategies for the treatment of osteodegenerative diseases. Here, we review the molecular pathways and players activated by mechanical stimulation during osteoblastic growth, differentiation and activity in health, and consider the role of mechanostimulatory approaches in treating various bone pathophysiologies.

Neurons expressing the highest levels of gamma-synuclein are unaffected by targeted inactivation of the gene.

Homologous recombination in ES cells was employed to generate mice with targeted deletion of the first three exons of the gamma-synuclein gene. Complete inactivation of gene expression in null mutant mice was confirmed on the mRNA and protein levels. Null mutant mice are viable, are fertile, and do not display evident phenotypical abnormalities. The effects of gamma-synuclein deficiency on motor and peripheral sensory neurons were studied by various methods in vivo and in vitro. These two types of neurons were selected because they both express high levels of gamma-synuclein from the early stages of mouse embryonic development but later in the development they display different patterns of intracellular compartmentalization of the protein. We found no difference in the number of neurons between wild-type and null mutant animals in several brain stem motor nuclei, in lumbar dorsal root ganglia, and in the trigeminal ganglion. The survival of gamma-synuclein-deficient trigeminal neurons in various culture conditions was not different from that of wild-type neurons. There was no difference in the numbers of myelinated and nonmyelinated fibers in the saphenous nerves of these animals, and sensory reflex thresholds were also intact in gamma-synuclein null mutant mice. Nerve injury led to similar changes in sensory function in wild-type and mutant mice. Taken together, our data suggest that like alpha-synuclein, gamma-synuclein is dispensable for the development and function of the nervous system.

Peripheral sensory neurons survive in the absence of alpha- and gamma-synucleins.

Physiological functions of alpha-synuclein, a protein implicated in certain types of neurodegeneration, and two other members of the same family, beta-synuclein and gamma-synuclein, are not clearly understood. It has been suggested that synucleins are involved in intracellular processes associated with survival of neurons and their response to stress, and that changes of synuclein ratio might have deteriorating effects on neurons. In wild-type mice, sensory neurons of the peripheral nervous system express alpha-synuclein and notably high levels of gamma-synuclein, but targeted inactivation of either of these genes has no effect on these neurons. Here we produced double, alpha-synuclein/gamma-synuclein null mutant mice, which develop normally, are fertile, and show no obvious signs of pathology in adulthood. Survival of alpha/gamma-synuclein-deficient peripheral sensory neurons in vivo and in primary tissue culture is indistinguishable from survival of wild-type neurons. The absence of two synucleins does not lead to expression in sensory neurons of the third member of the family, beta-synuclein. Therefore, our results demonstrate that neurons with normally high levels of synuclein(s) can develop and survive normally in the absence of any of these proteins. This suggests that other intraneuronal mechanisms and pathways effectively compensate the loss of synuclein function in null mutant animals.

Colon carcinogenesis: Learning from NF-kappaB and AP-1.

Colorectal cancer (CRC) is among the most common types of cancer attributed to genetic alterations. Its manifestation implicates NF-kappaB and AP-1 signaling pathways by virtue of their regulative role on the genetic control of cell cycle and apoptosis as well as by their capacity to be constitutively activated or exogenously induced by growth factors, cytokines, stress signals and oncoproteins. In CRC, the positive impact of NF-kappaB and AP-1 on the transcription of angiogenic and invasive factors strongly implicates these transcription factors in the transition of benign carcinomas towards a metastatic phenotype. Furthermore, the deregulated function of NF-kappaB and AP-1 in CRC cells affects inflammatory cascades, manifested by the ample production of inflammatory mediators. In this perspective, inhibition of NF-kappaB and AP-1 signaling mechanisms has become a rational target in the development of novel therapeutic approaches against CRC.

Functional alterations in mechanical loading of condylar cartilage induces changes in the bony subcondylar region.

Bone remodeling is orchestrated by cells of the osteoblast lineage and involves an intricate network of cell-cell and cell-matrix interactions. This dynamic process engages systemic hormones, locally produced cytokines and growth factors, as well as the mechanical environment of the cells. In growing subjects, the mandibular condyle consists of both articular and growth components and the presence of progenitor cells is verified by their anabolic responses to growth hormones. The pathways of chondrocyte and osteoblast differentiation during endochondral bone formation are interconnected and controlled by key transcription factors. The present study was undertaken to explore the possibility and the extent by which the mechano-transduction events in chondrocytes are 'sensed' in the subchondral bony area under altered functional loading. To this end, the involvement of the JNK/ERK-AP-1/Runx2 signaling axe was investigated by immunohistochemistry in temporomandibular joints of young rats subjected to different functional mastication loads. Our results showed that mechanical load triggers differentiation phenomena through the induction of master tissue regulators, namely the expression and/or activation of the JNK-c-Jun signaling pathway components and c-Fos in subchondral osteoblasts, as well as the activation of ERK/MAPK and the cellular expression of the transcription factor Runx2 in subchondral osteoblasts.

MG-63 osteoblast-like cells enhance the osteoprotegerin expression of PC-3 prostate cancer cells.

BACKGROUND: Osteoprotegerin (OPG) expression participates in the pathophysiology of osteoblastic metastasis in prostate cancer. MATERIALS AND METHODS: We investigated whether the expression of OPG of PC-3 prostate cancer cells grown in 3-D collagen gels is stimulated by co-culture with MG-63 osteoblast-like cells. The expression of Runx2 (Cbfa1) and OPG were assessed by reverse transcription-polymerase chain reaction and Western blot analysis. RESULTS: OPG and Runx2 were expressed in both PC-3 and MG-63 cells. OPG expression was remarkably enhanced in PC-3 cells grown in co-culture with MG-63 cells in a time-dependent manner. Runx2 expression of PC-3 cells was not altered by their co-culture with MG-63 cells. OPG expression of PC-3 cells was altered neither by insulin-like growth factor I (IGF-1), transforming growth factor beta1 (TGFbeta1), interleukin 6 (IL-6) nor by dexamethasone and zoledronic acid exogenously added to PC-3 cells. CONCLUSION: The enhancement of the OPG expression in PC-3 cells by MG-63 cells is not mediated by IGF-1, IL-6 and TGFbeta1.

Autoantibodies to alpha-synuclein in inherited Parkinson's disease.

Neurodegeneration in Parkinson's disease (PD) is accompanied by a local immune reaction in the affected brain regions. It is well established that alpha-synuclein is directly implicated in the pathogenesis of PD. Development of the disease is often associated with changes of expression and cellular compartmentalisation of this protein; moreover, its oligomers or protofibrils are often released to the CSF and plasma of patients. Aggregated alpha-synuclein can trigger the activation of microglia; however, its capacity to induce production of specific autoantibodies (AAb) has not been assessed. In this study, we examined the presence of AAb against synuclein family members in the peripheral blood serum of PD patients and control individuals. Presence of AAb against beta-synuclein or gamma-synuclein showed no association with PD. Multi-epitopic AAb against alpha-synuclein were detected in 65% of all patients tested and their presence strongly correlated with an inherited mode of the disease but not with other disease-related factors. The frequency of the presence of AAb in the studied group of patients with sporadic form of PD was not significantly different from the frequency in the control group but very high proportion (90%) of patients with familial form of the disease were positive for AAb against alpha-synuclein. We hypothesise that these AAb could be involved in pathogenesis of the inherited form of PD.