Angiotensin II triggers knee joint lesions in experimental osteoarthritis.

OBJECTIVES: This study aimed to evaluate the involvement of Angiotensin II (Ang II) in joint lesions associated with osteoarthritis (OA) in vitro and in vivo. METHODS: Chondrocyte cultures were obtained from knee joints of neonatal rats and stimulated with Ang II/MIA/ACE inhibitors. In vivo, rats treated or not with the ACE inhibitor captopril, received daily injections of Ang II or sodium monoiodoacetate (MIA) in knee joints for evaluation of cartilage, bone, and synovial lesions. RESULTS: Cultured chondrocytes expressed the mRNA for Ace, Agtr1, Agtr2, and Mas1. Stimulating cells with Ang II reduced chondrocyte viability and metabolism. Accordingly, in vivo Ang II injection into the knees of rats triggered hyperalgesia, joint edema, increased the number of leukocytes in the joint cavity, and induced cartilage lesions associated with OA alterations. In further experiments, Ang II synthesis was prevented with the ACE inhibitor Captopril in the context of MIA-induced OA. Ang II inhibition with captopril improved the OARSI score, induced chondroprotection, and reduced the leukocyte recruitment from synovium after MIA. Additionally, captopril prevented MIA-induced bone resorption, by decreasing the number of osteoclasts and increasing the expression of IL-10 in the bone. In vitro, inhibiting Ang II synthesis decreased MIA-induced chondrocyte death and increased Col2a1 transcription. CONCLUSION: Ang II induces chondrocyte death and joint tissue damages associated with OA and its modulation can be a therapeutic strategy in osteoarthritis.

Postnatal development of skeletal muscle in pigs with intrauterine growth restriction: morphofunctional phenotype and molecular mechanisms.

Intrauterine growth restriction (IUGR) is a serious condition which impairs the achievement of the fetus' full growth potential and occurs in a natural and severe manner in pigs as a result of placental insufficiency. Reduced skeletal muscle mass in the fetus with IUGR persists into adulthood and may contribute to increased metabolic disease risk. To investigate skeletal muscle postnatal development, histomorphometrical patterns of the semitendinosus muscle, myosin heavy chain (MyHC; embryonic I, IIA, IIB and IIX isoforms) fiber composition and the relative expression of genes related to myogenesis, adipogenesis and growth during three specific periods: postnatal myogenesis (newborn to 100 days old), hypertrophy (100-150 days old), and postnatal development (newborn to 150 days old) were evaluated in female pigs with IUGR and normal birth weight (NW) female littermates. NW females presented higher body weights compared to their IUGR counterparts at all ages evaluated (P < 0.05). Moreover, growth restriction in utero affected the semitendinosus muscle weight, muscle fiber diameter, and muscle cross-sectional area, which were smaller in IUGR pigs at birth (P < 0.05). Notwithstanding the effects on muscle morphology, IUGR also affected muscle fiber composition, as the percentage of MyHC-I myofibers was higher at birth (P < 0.05), and, in 150-day-old gilts, a lower percentage of MyHC-IIX isoform (P < 0.05) and the presence of embryonic MyHC isoform were also observed. Regarding the pattern of gene expression in both the postnatal myogenesis and postnatal development periods, IUGR led to the downregulation of myogenic factors, which delayed skeletal muscle myogenesis (PAX7, MYOD, MYOG, MYF5 and DES). Altogether, growth restriction in utero affects muscle fiber number and size at birth and muscle fiber composition through the downregulation of myogenic factors, which determines the individual´s postnatal growth rate. This fact, associated with delayed myofiber development in growth-restricted animals, may affect meat quality characteristics in animal production. Hence, knowledge of the morphofunctional phenotype of the skeletal muscle throughout postnatal development in individuals with IUGR, and the mechanism that governs it, may provide a better understanding of the mechanisms that limit postnatal muscle growth, and help the establishment of potential strategies to improve muscle development and prevent the onset of later-life metabolic diseases.

Engrailed controls epaxial-hypaxial muscle innervation and the establishment of vertebrate three-dimensional mobility.

Chordates are characterised by contractile muscle on either side of the body that promotes movement by side-to-side undulation. In the lineage leading to modern jawed vertebrates (crown group gnathostomes), this system was refined: body muscle became segregated into distinct dorsal (epaxial) and ventral (hypaxial) components that are separately innervated by the medial and hypaxial motors column, respectively, via the dorsal and ventral ramus of the spinal nerves. This allows full three-dimensional mobility, which in turn was a key factor in their evolutionary success. How the new gnathostome system is established during embryogenesis and how it may have evolved in the ancestors of modern vertebrates is not known. Vertebrate Engrailed genes have a peculiar expression pattern as they temporarily demarcate a central domain of the developing musculature at the epaxial-hypaxial boundary. Moreover, they are the only genes known with this particular expression pattern. The aim of this study was to investigate whether Engrailed genes control epaxial-hypaxial muscle development and innervation. Investigating chick, mouse and zebrafish as major gnathostome model organisms, we found that the Engrailed expression domain was associated with the establishment of the epaxial-hypaxial boundary of muscle in all three species. Moreover, the outgrowing epaxial and hypaxial nerves orientated themselves with respect to this Engrailed domain. In the chicken, loss and gain of Engrailed function changed epaxial-hypaxial somite patterning. Importantly, in all animals studied, loss and gain of Engrailed function severely disrupted the pathfinding of the spinal motor axons, suggesting that Engrailed plays an evolutionarily conserved role in the separate innervation of vertebrate epaxial-hypaxial muscle.

The emergence of Pax7-expressing muscle stem cells during vertebrate head muscle development.

Pax7 expressing muscle stem cells accompany all skeletal muscles in the body and in healthy individuals, efficiently repair muscle after injury. Currently, the in vitro manipulation and culture of these cells is still in its infancy, yet muscle stem cells may be the most promising route toward the therapy of muscle diseases such as muscular dystrophies. It is often overlooked that muscular dystrophies affect head and body skeletal muscle differently. Moreover, these muscles develop differently. Specifically, head muscle and its stem cells develop from the non-somitic head mesoderm which also has cardiac competence. To which extent head muscle stem cells retain properties of the early head mesoderm and might even be able to switch between a skeletal muscle and cardiac fate is not known. This is due to the fact that the timing and mechanisms underlying head muscle stem cell development are still obscure. Consequently, it is not clear at which time point one should compare the properties of head mesodermal cells and head muscle stem cells. To shed light on this, we traced the emergence of head muscle stem cells in the key vertebrate models for myogenesis, chicken, mouse, frog and zebrafish, using Pax7 as key marker. Our study reveals a common theme of head muscle stem cell development that is quite different from the trunk. Unlike trunk muscle stem cells, head muscle stem cells do not have a previous history of Pax7 expression, instead Pax7 expression emerges de-novo. The cells develop late, and well after the head mesoderm has committed to myogenesis. We propose that this unique mechanism of muscle stem cell development is a legacy of the evolutionary history of the chordate head mesoderm.

Evolutionarily conserved morphogenetic movements at the vertebrate head-trunk interface coordinate the transport and assembly of hypopharyngeal structures.

The vertebrate head-trunk interface (occipital region) has been heavily remodelled during evolution, and its development is still poorly understood. In extant jawed vertebrates, this region provides muscle precursors for the throat and tongue (hypopharyngeal/hypobranchial/hypoglossal muscle precursors, HMP) that take a stereotype path rostrally along the pharynx and are thought to reach their target sites via active migration. Yet, this projection pattern emerged in jawless vertebrates before the evolution of migratory muscle precursors. This suggests that a so far elusive, more basic transport mechanism must have existed and may still be traceable today. Here we show for the first time that all occipital tissues participate in well-conserved cell movements. These cell movements are spearheaded by the occipital lateral mesoderm and ectoderm that split into two streams. The rostrally directed stream projects along the floor of the pharynx and reaches as far rostrally as the floor of the mandibular arch and outflow tract of the heart. Notably, this stream leads and engulfs the later emerging HMP, neural crest cells and hypoglossal nerve. When we (i) attempted to redirect hypobranchial/hypoglossal muscle precursors towards various attractants, (ii) placed non-migratory muscle precursors into the occipital environment or (iii) molecularly or (iv) genetically rendered muscle precursors non-migratory, they still followed the trajectory set by the occipital lateral mesoderm and ectoderm. Thus, we have discovered evolutionarily conserved morphogenetic movements, driven by the occipital lateral mesoderm and ectoderm, that ensure cell transport and organ assembly at the head-trunk interface.

Association of IGF1 and KDM5A polymorphisms with performance, fatness and carcass traits in chickens.

Two functional and positional candidate genes were selected in a region of chicken chromosome 1 (GGA1), based on their biological roles, and also where several quantitative trait loci (QTL) have been mapped and associated with performance, fatness and carcass traits in chickens. The insulin-like growth factor 1 (IGF1) gene has been associated with several physiological functions related to growth. The lysine (K)-specific demethylase 5A (KDM5A) gene participates in the epigenetic regulation of genes involved with the cell cycle. Our objective was to find associations of selected single-nucleotide polymorphisms (SNPs) in these genes with performance, fatness and carcass traits in 165 F(2) chickens from a resource population. In the IGF1 gene, 17 SNPs were detected, and in the KDM5A gene, nine SNPs were detected. IGF1 SNP c.47673G > A was associated with body weight and haematocrit percentage, and also with feed intake and percentages of abdominal fat and gizzard genotype × sex interactions. KDM5A SNP c.34208C > T genotype × sex interaction affected body weight, feed intake, percentages of abdominal fat (p = 0.0001), carcass, gizzard and haematocrit. A strong association of the diplotype × sex interaction (p < 0.0001) with abdominal fat was observed, and also associations with body weight, feed intake, percentages of carcass, drums and thighs, gizzard and haematocrit. Our findings suggest that the KDM5A gene might play an important role in the abdominal fat deposition in chickens. The IGF1 and KDM5A genes are strong candidates to explain the QTL mapped in this region of GGA1.

Permanent genetic resources added to molecular ecology resources database 1 October 2012-30 November 2012.

This article documents the addition of 153 microsatellite marker loci to the Molecular Ecology Resources Database. Loci were developed for the following species: Brassica oleracea, Brycon amazonicus, Dimorphandra wilsonii, Eupallasella percnurus, Helleborus foetidus, Ipomoea purpurea, Phrynops geoffroanus, Prochilodus argenteus, Pyura sp., Sylvia atricapilla, Teratosphaeria suttonii, Trialeurodes vaporariorum and Trypanosoma brucei. These loci were cross-tested on the following species: Dimorphandra coccicinea, Dimorphandra cuprea, Dimorphandra gardneriana, Dimorphandra jorgei, Dimorphandra macrostachya, Dimorphandra mollis, Dimorphandra parviflora and Dimorphandra pennigera.

Development of microsatellite primers for Jatropha curcas (Euphorbiaceae) and transferability to congeners.

PREMISE OF THE STUDY: Microsatellite primers were developed for Jatropha curcas (Euphorbiaceae), a tree species with large potential for biofuel production, to investigate its natural genetic diversity and mating system to facilitate the establishment of tree improvement and conservation programs. METHODS AND RESULTS: Using a protocol for genomic library enrichment, 104 clones containing 195 repeat motifs were identified. Primer pairs were developed for 40 microsatellite loci and validated in 41 accessions of J. curcas from six provenances. Nine loci were polymorphic revealing from two to eight alleles per locus, and six primers were able to amplify alleles in the congeners J. podagrica, J. pohliana, and J. gossypifolia, but not in other Euphorbiaceae species, such as Hevea brasiliensis, Manihot esculenta, or Ricinus communis. CONCLUSIONS: The primers developed here revealed polymorphic loci that are suitable for genetic diversity and structure, mating system, and gene flow studies in J. curcas, and some congeners.

EST analysis of mRNAs expressed during embryogenesis in Gallus gallus.

Chicken Expressed Sequence Tags (ESTs) were analyzed to identify genes associated with myogenesis during embryonic development. A total of 6,184 ESTs were generated from three cDNA libraries constructed from whole embryos (Stage 26), somites associated with neural tube (Stage 15), and limb buds (Stages 21, 24 and 26). Clustering and assembly of 4,998 valid ESTs resulted in 2,329 unique sequences with 902 clusters (38.7%) and 1,427 singletons (61.3%). There are more than 400,000 chicken ESTs available at GenBank and we were able to identify 143 novel sequences. From these, 45 sequences found either a human EST homolog or a match with conserved regions among proteins. Most of these sequences were found to be expressed in somites, an important tissue for muscle development and not characterized before. This study revealed the value of micro dissected embryonic libraries for describing gene expression profiles associated with myogenesis and gene discovery.