Deficiency of annexins A5 and A6 induces complex changes in the transcriptome of growth plate cartilage but does not inhibit the induction of mineralization

Initiation of mineralization during endochondral ossification is a multistep process and has been assumed to correlate with specific interactions of annexins A5 and A6 and collagens. However, skeletal development appears to be normal in mice deficient for either A5 or A6, and the highly conserved structures led to the assumption that A5 and A6 may fulfill redundant functions. We have now generated mice deficient of both proteins. These mice were viable and fertile and showed no obvious abnormalities. Assessment of skeletal elements using histologic, ultrastructural, and peripheral quantitative computed tomographic methods revealed that mineralization and development of the skeleton were not significantly affected in mutant mice. Otherwise, global gene expression analysis showed subtle changes at the transcriptome level of genes involved in cell growth and intermediate metabolism. These results indicate that annexins A5 and A6 may not represent the essential annexins that promote mineralization in vivo

Global comparative transcriptome analysis of cartilage formation in vivo

<p>Abstract</p> <p>Background</p> <p>During vertebrate embryogenesis the initial stages of bone formation by endochondral ossification involve the aggregation and proliferation of mesenchymal cells into condensations. Continued growth of the condensations and differentiation of the mesenchymal cells into chondrocytes results in the formation of cartilage templates, or anlagen, which prefigure the shape of the future bones. The chondrocytes in the anlagen further differentiate by undergoing a complex sequence of maturation and hypertrophy, and are eventually replaced by mineralized bone. Regulation of the onset of chondrogenesis is incompletely understood, and would be informed by comprehensive analyses of <it>in vivo </it>gene expression.</p> <p>Results</p> <p>Tibial and fibular pre-condensed mesenchyme was microdissected from mouse hind limbs at 11.5 dpc, and the corresponding condensations at 12.5 dpc and cartilage anlagen at 13.5 dpc. Total RNA was isolated, and cRNA generated by linear amplification was interrogated using mouse whole genome microarrays. Differential expression was validated by quantitative PCR for <it>Agc1</it>, <it>Bmp8a</it>, <it>Col2a1</it>, <it>Fgfr4</it>, <it>Foxa3</it>, <it>Gdf5</it>, <it>Klf2</it>, <it>Klf4</it>, <it>Lepre1</it>, <it>Ncad</it>, <it>Sox11</it>, and <it>Trpv4</it>. Further, independent validation of the microarray data was achieved by <it>in situ </it>hybridization to analyse the expression of <it>Lepre1</it>, <it>Pcdh8</it>, <it>Sox11</it>, and <it>Trpv4 </it>from 11.5 dpc to 13.5 dpc during mouse hind limb development. We found significant differential expression of 931 genes during these early stages of chondrogenesis. Of these, 380 genes were down-regulated and 551 up-regulated. Our studies characterized the expression pattern of gene families previously associated with chondrogenesis, such as adhesion molecules, secreted signalling molecules, transcription factors, and extracellular matrix components. Gene ontology approaches identified 892 differentially expressed genes not previously identified during the initiation of chondrogenesis. These included several <it>Bmp, Gdf, Wnt, Sox and Fox </it>family members.</p> <p>Conclusion</p> <p>These data represent the first global gene expression profiling analysis of chondrogenic tissues during <it>in vivo </it>development. They identify genes for further study on their functional roles in chondrogenesis, and provide a comprehensive and important resource for future studies on cartilage development and disease.</p

S100A8 and S100A9 in experimental osteoarthritis

INTRODUCTION: The objective was to evaluate the changes in S100A8 S100A9, and their complex (S100A8/S100A9) in cartilage during the onset of osteoarthritis (OA) as opposed to inflammatory arthritis. METHODS: S100A8 and S100A9 protein localization were determined in antigen-induced inflammatory arthritis in mice, mouse femoral head cartilage explants stimulated with interleukin-1 (IL-1), and in surgically-induced OA in mice. Microarray expression profiling of all S100 proteins in cartilage was evaluated at different times after initiation of degradation in femoral head explant cultures stimulated with IL-1 and surgically-induced OA. The effect of S100A8, S100A9 or the complex on the expression of aggrecan (Acan), collagen II (Col2a1), disintegrin and metalloproteases with thrombospondin motifs (Adamts1, Adamts 4 &Adamts 5), matrix metalloproteases (Mmp1, Mmp3, Mmp13 &Mmp14) and tissue inhibitors of metalloproteinases (Timp1, Timp2 &Timp3), by primary adult ovine articular chondrocytes was determined using real time quantitative reverse transcription polymerase chain reaction (qRT-PCR). RESULTS: Stimulation with IL-1 increased chondrocyte S100a8 and S100a9 mRNA and protein levels. There was increased chondrocyte mRNA expression of S100a8 and S100a9 in early but not late mouse OA. However, loss of the S100A8 staining in chondrocytes occurred as mouse OA progressed, in contrast to the positive reactivity for both S100A8 and S100A9 in chondrocytes in inflammatory arthritis in mice. Homodimeric S100A8 and S100A9, but not the heterodimeric complex, significantly upregulated chondrocyte Adamts1, Adamts4 and Adamts 5, Mmp1, Mmp3 and Mmp13 gene expression, while collagen II and aggrecan mRNAs were significantly decreased. CONCLUSIONS: Chondrocyte derived S100A8 and S100A9 may have a sustained role in cartilage degradation in inflammatory arthritis. In contrast, while these proteins may have a role in initiating early cartilage degradation in OA by upregulating MMPs and aggrecanases, their reduced expression in late stages of OA suggests they do not have an ongoing role in cartilage degradation in this non-inflammatory arthropathy

Влияние моделей обратного тока насыщения диода на выходные характеристики двух-диодной модели солнечной батареи в среде Matlab Simulink

В работе проводилось моделирование солнечной батареи в программном обеспечении MATLAB Simulink. Изучались выходные параметры солнечной батареи в зависимости от модели обратного тока насыщения диода. Полученными результатами являются вольт-амперные и вольт-ваттные характеристики, показывающие влияние модели обратного тока насыщения диода на выходные параметры солнечной батареи.The work was carried out modeling of the solar battery in the software MATLAB Simulink. The output parameters of the solar battery were studied depending on the model of the reverse current saturation of the diode. The results are current-voltage and voltage-watt characteristics, showing the influence of the model of the inverse diode saturation current on the output parameters of the solar battery

Cross-Sample Validation Provides Enhanced Proteome Coverage in Rat Vocal Fold Mucosa

The vocal fold mucosa is a biomechanically unique tissue comprised of a densely cellular epithelium, superficial to an extracellular matrix (ECM)-rich lamina propria. Such ECM-rich tissues are challenging to analyze using proteomic assays, primarily due to extensive crosslinking and glycosylation of the majority of high Mr ECM proteins. In this study, we implemented an LC-MS/MS-based strategy to characterize the rat vocal fold mucosa proteome. Our sample preparation protocol successfully solubilized both proteins and certain high Mr glycoconjugates and resulted in the identification of hundreds of mucosal proteins. A straightforward approach to the treatment of protein identifications attributed to single peptide hits allowed the retention of potentially important low abundance identifications (validated by a cross-sample match and de novo interpretation of relevant spectra) while still eliminating potentially spurious identifications (global single peptide hits with no cross-sample match). The resulting vocal fold mucosa proteome was characterized by a wide range of cellular and extracellular proteins spanning 12 functional categories

Identification of TGFβ-related genes regulated in murine osteoarthritis and chondrocyte hypertrophy by comparison of multiple microarray datasets

Objective: Osteoarthritis (OA) is a joint disease characterized by progressive degeneration of articular cartilage. Some features of OA, including chondrocyte hypertrophy and focal calcification of articular cartilage, resemble the endochondral ossification processes. Alterations in transforming growth factor β (TGFβ) signaling have been associated with OA as well as with chondrocyte hypertrophy. Our aim was to identify novel candidate genes implicated in chondrocyte hypertrophy during OA pathogenesis by determining which TGFβ-related genes are regulated during murine OA and endochondral ossification. Methods: A list of 580 TGFβ-related genes, including TGFβ signaling pathway components and TGFβ-target genes, was generated. Regulation of these TGFβ-related genes was a

Matrilin-3 from chicken cartilage

AbstractBy subtractive cDNA cloning we have identified a novel constituent of chicken cartilage termed matrilin-3. This constituent is encoded by a mRNA of 2.2 kbp whose expression is restricted to cartilaginous tissues. The predicted protein is composed of 452 amino acids with a molecular mass of 49 kDa. It contains a single von Willebrand factor A-like domain, four epidermal growth factor-like repeats and an α-helical region which may induce the formation of oligomers via a coiled-coil. The primary structure is similar to that of matrilin-1 which is also expressed in a cartilage-specific manner. This similarity suggests that the genes for the two proteins may have evolved from a common ancestor by gene duplication