Skeletal muscle wasting represents both a common phenotype of aging and a feature of pathological conditions such as chronic kidney disease (CKD). Although both clinical data and genetic experiments ...in mice suggest that hyperphosphatemia accelerates muscle wasting, the underlying mechanism remains unclear. Here, we showed that inorganic phosphate (Pi) dose-dependently decreases myotube size, fusion index, and myogenin expression in mouse C2C12 skeletal muscle cells. These changes were accompanied by increases in reactive oxygen species (ROS) production and Nrf2 and p62 expression, and reductions in mitochondrial membrane potential (MMP) and Keap1 expression. Inhibition of Pi entry, cytosolic ROS production, or Nrf2 activation reversed the effects of high Pi on Nrf2, p62, and myogenin expression. Overexpression of Nrf2 respectively increased and decreased the promoter activity of
-Luc and
-Luc reporters. Analysis of nuclear extracts from gastrocnemius muscles from mice fed a high-Pi (2% Pi) diet showed increased Nrf2 phosphorylation in sham-operated and 5/6 nephrectomized (CKD) mice, and both increased p62 phosphorylation and decreased myogenin expression in CKD mice. These data suggest that high Pi suppresses myogenic differentiation
and promotes muscle atrophy
through oxidative stress-mediated protein degradation and both canonical (ROS-mediated) and non-canonical (p62-mediated) activation of Nrf2 signaling.
The aim of the study was to investigate the use of a hyaluronic acid/polycaprolactone material for meniscal tissue engineering and to evaluate the tissue regeneration after the augmentation of the ...implant with expanded autologous chondrocytes. Eighteen skeletally mature sheep were treated. The animals were divided into three groups: cell-free scaffold, scaffold seeded with autologous chondrocytes, and meniscectomy alone. The implant was sutured to the capsule and to the meniscal ligament. At a 12-month gross assessment, histology and histomorphometry were used to assess the meniscus implant, knee joint, and osteoarthritis development. All implants showed excellent capsular ingrowth at the periphery. The implant gross assessment showed significant differences between cell-seeded and cell-free groups (p=0.011). The histological analysis indicated a cellular colonization throughout the implanted constructs. Avascular cartilaginous tissue formation was significantly more frequent in the cell-seeded constructs. Joint gross assessment showed that sheep treated with scaffold implantation achieved a significant higher score than those underwent meniscectomy (p<0.0005), and the Osteoarthritis Research Society International score showed that osteoarthritic changes were significantly less in the cell-seeded group than in the meniscectomy group (p=0.047), even though results were not significantly superior to those of the cell-free scaffold. Seeding of the scaffold with autologous chondrocytes increases its tissue regeneration capacity, providing a better fibrocartilaginous tissue formation. The study suggests the potential of the novel hyaluronic acid/polycaprolactone scaffold for total meniscal substitution, although this approach has to be further improved before being applied into clinical practice.
Mammalian Hedgehog (HH) signalling pathway plays an essential role in tissue homeostasis and its deregulation is linked to rheumatological disorders. UBR5 is the mammalian homologue of the E3 ...ubiquitin-protein ligase Hyd, a negative regulator of the Hh-pathway in Drosophila. To investigate a possible role of UBR5 in regulation of the musculoskeletal system through modulation of mammalian HH signaling, we created a mouse model for specific loss of Ubr5 function in limb bud mesenchyme. Our findings revealed a role for UBR5 in maintaining cartilage homeostasis and suppressing metaplasia. Ubr5 loss of function resulted in progressive and dramatic articular cartilage degradation, enlarged, abnormally shaped sesamoid bones and extensive heterotopic tissue metaplasia linked to calcification of tendons and ossification of synovium. Genetic suppression of smoothened (Smo), a key mediator of HH signalling, dramatically enhanced the Ubr5 mutant phenotype. Analysis of HH signalling in both mouse and cell model systems revealed that loss of Ubr5 stimulated canonical HH-signalling while also increasing PKA activity. In addition, human osteoarthritic samples revealed similar correlations between UBR5 expression, canonical HH signalling and PKA activity markers. Our studies identified a crucial function for the Ubr5 gene in the maintenance of skeletal tissue homeostasis and an unexpected mode of regulation of the HH signalling pathway.
Inflammation induced by infectious and noninfectious triggers in the kidney may lead to end stage renal disease (ESRD). Toll-like receptor 9 (TLR-9) a receptor for CpG DNA is involved in activation ...of immune cells in renal disease and may contribute to chronic inflammatory disease progression through an interleukin-6 (IL-6) dependent pathway. Previous studies indicate that -1237T/C confers regulatory effects on TLR-9 transcription. To date the effect of TLR-9 polymorphisms on ESRD remains unknown. We performed a case-control study and genotyped 630 ESRD patients and 415 controls for -1237T/C, -1486T/C and 1635G/A by real-time PCR assays and assessed plasma concentration of IL-6 by ELISA. Haplotype association analysis was performed using the Haploview package. A luciferase reporter assay and real-time PCR were used to test the function of the -1237T/C promoter polymorphism. A significant association between -1237T/C in TLR-9 and ESRD was identified. The TCA, TTA and CCA haplotype of TLR-9 were associated with ESRD. ESRD patients carrying -1237TC had a higher mean plasma IL-6 level when compared with -1237TT. The TLR-9 transcriptional activity of the variant -1237CC allele is higher than the -1237TT allele. The results indicate that in a Han Chinese population the presence of the C allele of -1237T/C in the TLR-9 gene increases susceptibility towards development of ESRD. In vitro studies demonstrate that -1237T/C may be involved in the development of ESRD through transcriptional modulation of TLR-9.
Summary
Chondrosarcomas are malignant cartilage tumours. They are poorly responsive to chemotherapy and radiotherapy. Treatment is usually limited to surgical resection; however, survival of patients ...with high‐grade chondrosarcoma is poor, even with wide surgical resection. Induction of apoptosis in chondrosarcoma cells, either directly or by enhancement of the response to chemotherapeutic drugs and radiation, may be a route by which outcome can be improved. In this article, we review potential molecular targets that regulate chondrocyte apoptosis and discuss the experimental evidence for their utility.
The use of stem cells to support tissue repair is facilitated by loading of the therapeutic cells with magnetic nanoparticles (MNPs) enabling magnetic tracking and targeting. Current methods for ...magnetizing cells use artificial MNPs and have disadvantages of variable uptake, cellular cytotoxicity and loss of nanoparticles on cell division. Here we demonstrate a transgenic approach to magnetize human mesenchymal stem cells (MSCs). MSCs are genetically modified by transfection with the mms6 gene derived from Magnetospirillum magneticum AMB-1, a magnetotactic bacterium that synthesises single-magnetic domain crystals which are incorporated into magnetosomes. Following transfection of MSCs with the mms6 gene there is bio-assimilated synthesis of intracytoplasmic magnetic nanoparticles which can be imaged by MR and which have no deleterious effects on cell proliferation, migration or differentiation. The assimilation of magnetic nanoparticle synthesis into mammalian cells creates a real and compelling, cytocompatible, alternative to exogenous administration of MNPs.
In breast cancer, increased expression of the cytoskeletal adaptor protein Kindlin-1 has been linked to increased risks of lung metastasis, but the functional basis is unknown. Here, we show that in ...a mouse model of polyomavirus middle T antigen-induced mammary tumorigenesis, loss of Kindlin-1 reduced early pulmonary arrest and later development of lung metastasis. This phenotype relied on the ability of Kindlin-1 to bind and activate β integrin heterodimers. Kindlin-1 loss reduced α4 integrin-mediated adhesion of mammary tumor cells to the adhesion molecule VCAM-1 on endothelial cells. Treating mice with an anti-VCAM-1 blocking antibody prevented early pulmonary arrest. Kindlin-1 loss also resulted in reduced secretion of several factors linked to metastatic spread, including the lung metastasis regulator tenascin-C, showing that Kindlin-1 regulated metastatic dissemination by an additional mechanism in the tumor microenvironment. Overall, our results show that Kindlin-1 contributes functionally to early pulmonary metastasis of breast cancer.
These findings provide a mechanistic proof in mice that Kindin-1, an integrin-binding adaptor protein, is a critical mediator of early lung metastasis of breast cancer.
.
The fascinating prospect to direct tissue regeneration by magnetic activation has been recently explored. In this study we investigate the possibility to boost bone regeneration in an experimental ...defect in rabbit femoral condyle by combining static magnetic fields and magnetic biomaterials. NdFeB permanent magnets are implanted close to biomimetic collagen/hydroxyapatite resorbable scaffolds magnetized according to two different protocols . Permanent magnet only or non-magnetic scaffolds are used as controls. Bone tissue regeneration is evaluated at 12 weeks from surgery from a histological, histomorphometric and biomechanical point of view. The reorganization of the magnetized collagen fibers under the effect of the static magnetic field generated by the permanent magnet produces a highly-peculiar bone pattern, with highly-interconnected trabeculae orthogonally oriented with respect to the magnetic field lines. In contrast, only partial defect healing is achieved within the control groups. We ascribe the peculiar bone regeneration to the transfer of micro-environmental information, mediated by collagen fibrils magnetized by magnetic nanoparticles, under the effect of the static magnetic field. These results open new perspectives on the possibility to improve implant fixation and control the morphology and maturity of regenerated bone providing “in site” forces by synergically combining static magnetic fields and biomaterials.
Angiosarcomas are a rare group of tumours which have poor prognosis and limited treatment options. The development of new therapies has been hampered by a lack of good preclinical models. Here, we ...describe the development of an autochthonous mouse model of angiosarcoma driven by loss of p53 in VE-cadherin-expressing endothelial cells. Using
to drive recombination in adult endothelial cells, mice developed angiosarcomas with 100% penetrance upon homozygous deletion of
with a median lifespan of 325 days. In contrast, expression of the R172H mutant p53 resulted in formation of thymic lymphomas with a more rapid onset (median lifespan 151 days). We also used
-expressing mice, allowing us to target predominantly pericytes, as these have been reported as the cell of origin for a number of soft tissue sarcomas.
also results in low levels of recombination in venous blood endothelial cells in multiple tissues during development. Upon deletion of
in
-expressing mice (
mice), 65% developed lymphomas and 21% developed pleomorphic undifferentiated soft tissue sarcomas. None developed angiosarcomas. In contrast, 75% of
mice developed angiosarcomas, with 60% of these mice also developing lymphomas. The median lifespan of the
mice was 151 days. Re-implantation of angiosarcoma tumour fragments from
mice provided a more consistent and rapid model of angiosarcoma than the two spontaneous models. The ability to passage tumour fragments through the mouse provides a novel model which is amenable to preclinical studies and will help the development of potential new therapies for angiosarcoma.
PDF
