Members of the transforming growth factor beta (TGF-β) family are secreted proteins that regulate skeletal development. TGF-β signaling is critical in embryonic development of the annulus fibrosus (AF) of the intervertebral disc (IVD). To address the question of the role of TGF-β signaling in postnatal development and maintenance of the skeleton, we generated mice in which Tgfbr2 was deleted at 2-weeks of age in Aggrecan (Acan)-expressing cells using inducible Cre/LoxP recombination.Localization of Cre recombination was visualized by crossing Acantm1(cre/ERT2)Crm mice to fluorescent mTmG reporter mice. Acantm1(cre/ERT2)Crm mice were mated to Tgfbr2LoxP/LoxP mice and Cre recombinase was activated by tamoxifen injection at 2-weeks postnatally. Following tamoxifen injection, mice were aged to 3, 6, and 12-months and control mice were compared to the experimental (cKO) group. Mice were initially analyzed using X-ray and skeletal preparations. Sternocostal joints and IVD tissues were further analyzed histologically by hematoxylin and eosin (H&E), Safranin O, and Picrosirius Red staining as well as Col10 immunostaining.Cre recombination was observed in the IVD and sternocostal joints. X-ray analysis revealed osteophyte formation within the disc space of 12-month-old cKO mice. Skeletal preparations confirmed calcification within the IVD and the sternocostal joints in cKO mice. H&E staining of cKO IVD revealed disorganized growth plates, delay in the formation of the bony endplate, and Col10 staining in the AF indicative of ectopic endochondral bone formation. Furthermore, proteoglycan loss was observed and collagen bundles within the inner AF were thinner and less organized. Alterations in the IVD were apparent beginning at 3 months and were progressively more visible at 6 and 12 months. Similarly, histological analysis of cKO sternocostal joints revealed joint calcification, proteoglycan loss, and disorganization of the collagen architecture at 12 months of age.TGF-β signaling is important for postnatal development and maintenance of fibrocartilaginous IVD and sternocostal joints.
Abstract Intraflagellar transport proteins (IFT) are required for hedgehog (Hh) signalling transduction that is essential for bone development, however, how IFT proteins regulate Hh signalling in osteoblasts (OBs) remains unclear. Here we show that deletion of ciliary IFT80 in OB precursor cells (OPC) in mice results in growth retardation and markedly decreased bone mass with impaired OB differentiation. Loss of IFT80 blocks canonical Hh–Gli signalling via disrupting Smo ciliary localization, but elevates non-canonical Hh–Gαi–RhoA–stress fibre signalling by increasing Smo and Gαi binding. Inhibition of RhoA and ROCK activity partially restores osteogenic differentiation of IFT80 -deficient OPCs by inhibiting non-canonical Hh–RhoA–Cofilin/MLC2 signalling. Cytochalasin D, an actin destabilizer, dramatically restores OB differentiation of IFT80 -deficient OPCs by disrupting actin stress fibres and promoting cilia formation and Hh–Gli signalling. These findings reveal that IFT80 is required for OB differentiation by balancing between canonical Hh–Gli and non-canonical Hh–Gαi–RhoA pathways and highlight IFT80 as a therapeutic target for craniofacial and skeletal abnormalities.
TGF-beta is now recognized as an important factor regulating normal mammary gland development as well as breast cancer. In development, TGF-beta regulates branching morphogenesis and differentiation by acting on both epithelial and stromal cells. TGF-beta also regulates apoptosis and matrix remodeling during involution at the end of the pregnancy cycle. TGF-beta has biphasic effects on tumor progression, acting as a tumor suppressor in early stages of cancer and promoting invasion and metastasis at later stages. Furthermore, TGF-beta may play a role in tumor progression through effects on the microenvironment. The tumor promoting effects of TGF-beta may provide a therapeutic target for late stage breast cancer via TGF-beta antagonists like the soluble receptors recently described. Future experiments will uncover the precise mechanisms of TGF-beta action in development and neoplastic disease providing more opportunities for prevention and treatment of breast disease.
Several functional parameters can be used to define respiratory dysfunctions. Many of these variables provide specific information about particular physiopathologic phenomena, but they seldom provide a general overview of the ventilatory function. Nevertheless, both pharmacologic and epidemiologic research need functional parameters that precisely specify the respiratory defect. The new parameter we discuss takes into account the variations in volumes as related to flow variations: it is the product of two traditional spirometric parameters (FEV1 X MMEF). This value gives information on the patient's capacity to exhale air from the lungs for a fixed variation of intrathoracic pressure as well as data on the acceleration pattern of the expired volume in 1 s, hence the name, acceleration index (AI). AI was used to predict bronchodilation in two groups of patients with severe airway flow limitations. On the basis of our findings three conclusions can be drawn. This new spirometric parameter provides an inexpensive and rapid means of obtaining a highly sensitive definition of the patient's respiratory dysfunction which is useful in screening patients. Applied to patients with severe airways obstruction, AI can be used to immediately predict the reversibility of air flow limitation and then to define the limits of the activity of bronchodilators. Finally, a basal value of AI below 1.000 l/s-1 indicates the probable irreversibility of the airways obstruction, regardless of drug or route of administration. Such cases suggest a prevailing picture of emphysema rather than reversible bronchospasm.
Ribosome biogenesis is essential for cell growth and proliferation and is commonly elevated in cancer. Accordingly, numerous oncogene and tumor suppressor signaling pathways target rRNA synthesis. In breast cancer, non-canonical Wnt signaling by Wnt5a has been reported to antagonize tumor growth. Here, we show that Wnt5a rapidly represses rDNA gene transcription in breast cancer cells and generates a chromatin state with reduced transcription of rDNA by RNA polymerase I (Pol I). These effects were specifically dependent on Dishevelled1 (DVL1), which accumulates in nucleolar organizer regions (NORs) and binds to rDNA regions of the chromosome. Upon DVL1 binding, the Pol I transcription activator and deacetylase Sirtuin 7 (SIRT7) releases from rDNA loci, concomitant with disassembly of Pol I transcription machinery at the rDNA promoter. These findings reveal that Wnt5a signals through DVL1 to suppress rRNA transcription. This provides a novel mechanism for how Wnt5a exerts tumor suppressive effects and why disruption of Wnt5a signaling enhances mammary tumor growth in vivo.
Primary cilia are slender, microtubule based structures found in the majority of cell types with one cilium per cell. In articular cartilage, primary cilia are required for chondrocyte mechanotransduction and the development of healthy tissue. Loss of primary cilia in Col2aCre;ift88(fl/fl) transgenic mice results in up-regulation of osteoarthritic (OA) markers and development of OA like cartilage with greater thickness and reduced mechanical stiffness. However no previous studies have examined whether loss of primary cilia influences the intrinsic mechanical properties of articular cartilage matrix in the form of the modulus or just the structural properties of the tissue. The present study describes a modified analytical model to derive the viscoelastic moduli based on previous experimental indentation data. Results show that the increased thickness of the articular cartilage in the Col2aCre;ift88(fl/fl) transgenic mice is associated with a reduction in both the instantaneous and equilibrium moduli at indentation strains of greater than 20%. This reveals that the loss of primary cilia causes a significant reduction in the mechanical properties of cartilage particularly in the deeper zones and possibly the underlying bone. This is consistent with histological analysis and confirms the importance of primary cilia in the development of a mechanically functional articular cartilage.
