558 research outputs found

    The Insulin receptor catalyzes the tyrosine phosphorylation o Caveolin 1

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    Our previous studies revealed that insulin stimulates the tyrosine phosphorylation of caveolin in 3T3L1 adipocytes. To explore the mechanisms involved in this event, we evaluated the association of the insulin receptor with caveolin. The receptor was detected in a Triton-insoluble low density fraction, co-sedimenting with caveolin and flotillin on sucrose density gradients. We also detected the receptor in caveolin-enriched rosette structures by immunohistochemical analysis of plasma membrane sheets from 3T3L1 adipocytes. Insulin stimulated the phosphorylation of caveolin-1 on Tyr14. This effect of the hormone was not blocked by overexpression of mutant forms of the Cbl-associated protein that block the translocation of phospho-Cbl to the caveolin-enriched, lipid raft microdomains. Moreover, this phosphorylation event was also unaffected by inhibitors of the MAPK and phosphatidylinositol 3-kinase pathways. Although previous studies demonstrated that the Src family kinase Fyn was highly enriched in caveolae, an inhibitor of this kinase had no effect on insulin-stimulated caveolin phosphorylation. Interestingly, overexpression of a mutant form of caveolin that failed to interact with the insulin receptor did not undergo phosphorylation. Taken together, these data indicate that the insulin receptor directly catalyzes the tyrosine phosphorylation of caveolin. Previous article in issu

    Increased SK3 expression in DM1 lens cells leads to impaired growth through a greater calcium-induced fragility

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    Although cataract is a characteristic feature of myotonic dystrophy type 1 (DM1), little is known of the underlying mechanisms. We generated four lens epithelial cell lines derived from DM1 cataracts and two from age-matched, non-DM cataracts. Small-pool PCR revealed typical large triplet repeat expansions in the DM1 cells. Furthermore, real-time PCR analysis showed reduced SIX5 expression and increased expression of the Ca2+-activated K+ channel SK3 in the DM1 cells. These cells also exhibited longer population doubling times which did not arise through reduced proliferation, but rather increased cell death as shown by increased release of lactate dehydrogenase (LDH). Using 86Rb+ as a tracer for K+, we found no difference in the resting K+ influx or efflux kinetics. In all cases, the ouabain sensitive component of the influx contributed ~50% of the total. However, stimulating internal Ca2+ by exposure to ionomycin not only caused greater stimulation of K+ (86Rb) efflux in the DM1 cells but also induced a higher rate of cell death (LDH assay). Since both the hyper-stimulation of K+ efflux and cell death were reduced by the highly specific SK inhibitor apamin, we suggest that increased expression of SK3 has a critical role in the increased Ca2+-induced fragility in DM1 cells. The present data, therefore, both help explain the lower epithelial cell density previously observed in DM1 cataracts and provide general insights into mechanisms underlying the fragility of other DM1-affected tissues

    Zinc Finger Protein, Hzf, Is Required for Megakaryocyte Development and Hemostasis

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    Using an expression gene trapping strategy, we recently identified a novel gene, hematopoietic zinc finger (Hzf), which encodes a protein containing three C2H2-type zinc fingers that is predominantly expressed in megakaryocytes. Here, we have examined the in vivo function of Hzf by gene targeting and demonstrated that Hzf is essential for megakaryopoiesis and hemostasis in vivo. Hzf-deficient mice exhibited a pronounced tendency to rebleed and had reduced α-granule substances in both megakaryocytes and platelets. These mice also had large, faintly stained platelets, whereas the numbers of both megakaryocytes and platelets were normal. These results indicate that Hzf plays important roles in regulating the synthesis of α-granule substances and/or their packing into α-granules during the process of megakaryopoiesis

    Rate of telomere shortening and cardiovascular damage: a longitudinal study in the 1946 British Birth Cohort.

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    AIM: Cross-sectional studies reported associations between short leucocyte telomere length (LTL) and measures of vascular and cardiac damage. However, the contribution of LTL dynamics to the age-related process of cardiovascular (CV) remodelling remains unknown. In this study, we explored whether the rate of LTL shortening can predict CV phenotypes over 10-year follow-up and the influence of established CV risk factors on this relationship. METHODS AND RESULTS: All the participants from the MRC National Survey of Health and Development (NSHD) with measures of LTL and traditional CV risk factors at 53 and 60-64 years and common carotid intima-media thickness (cIMT), cardiac mass and left ventricular function at 60-64 years were included. LTL was measured by real-time polymerase chain reaction and available at both time points in 1033 individuals. While LTL at 53 years was not linked with any CV phenotype at 60-64 years, a negative association was found between LTL and cIMT at 60-64 years (β = -0.017, P = 0.015). However, the strongest association was found between rate of telomere shortening between 53 and 60-64 years and values of cIMT at 60-64 years (β = -0.020, P = 0.006). This association was not affected by adjustment for traditional CV risk factors. Cardiac measurements were not associated with cross-sectional or longitudinal measures of LTL. CONCLUSION: These findings suggest that the rate of progression of cellular ageing in late midlife (reflected by the rate of LTL attrition) relates to vascular damage, independently from contribution of CV risk factor exposure

    Killing Tensors and Conformal Killing Tensors from Conformal Killing Vectors

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    Koutras has proposed some methods to construct reducible proper conformal Killing tensors and Killing tensors (which are, in general, irreducible) when a pair of orthogonal conformal Killing vectors exist in a given space. We give the completely general result demonstrating that this severe restriction of orthogonality is unnecessary. In addition we correct and extend some results concerning Killing tensors constructed from a single conformal Killing vector. A number of examples demonstrate how it is possible to construct a much larger class of reducible proper conformal Killing tensors and Killing tensors than permitted by the Koutras algorithms. In particular, by showing that all conformal Killing tensors are reducible in conformally flat spaces, we have a method of constructing all conformal Killing tensors (including all the Killing tensors which will in general be irreducible) of conformally flat spaces using their conformal Killing vectors.Comment: 18 pages References added. Comments and reference to 2-dim case. Typos correcte

    Rapid Prototyping of Flexible Structures for Tissue Engineered Ear Reconstruction

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    The tissue engineered ear has been an iconic symbol of the field since 1991, when the report of an engineered ear in a mouse model was first published A tissue engineered ear has an inherent advantage over conventional approaches because the structure is derived from the patient's own cartilage. In this approach, autologous auricular chondrocytes are harvested from the patient and grown within an ear-shaped scaffold. However, as the scaffold degrades or remodels, the ear-shaped structure undergoes significant distortion, resulting in a skewed ear shape that is smaller and often unrecognizable In order to maintain the desired ear geometry, a composite scaffold concept was developed Methods Several functional requirements for the manufacturing process were identified. First, the wire framework must be created with arbitrary three dimensional (3D) control, and with a diameter significantly smaller than the thickness of normal ear cartilage, which is about 2 mm. The bending stiffness must be sufficiently high so that shape is maintained during neocartilage maturation and sufficiently low such that flexibility of the overall structure is not impaired. The material must be approved for clinical use, and must not cause an inflammatory reaction. Finally, the manufacturing process must be capable of producing single, custom parts without significant cost burden. Plastic surgeons identified titanium and stainless steel as preferred materials due to their long history of success in medical implants Three manufacturing processes were identified that are capable of producing arbitrary shapes with the listed metals: wire bending, direct metal laser sintering (DMLS) Results Ear frameworks produced using DMLS and EBM technology are shown in Interpretation Ear frameworks produced using DMLS and EBM technology are shown i
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