Litter live weight at birth with varying levels of D’Man germplasm

A total of 208 litter birth weights, with live lambs produced in the northwest of Morocco was analyzed both by factorial analysis and by multiple regression analysis using least squares procedure. In the first model, breed group of dam and age of dam were included as fixed effects. In the second model, maternal additive and heterotic effects were included instead of breed group of dam. Results of the first analysis indicated that breed group of dam influenced litter live weight at birth (LLWB). The crossbred S x D dams (with the sire breed listed first), and the purebred D’Man dams produced the heaviest live litters, in average 4.25 and 4.08 kg, respectively. The purebred Beni Guil dams produced the lightest live litter, 3.32 kg in average. The second analysis showed that maternal additive effects were important for LLWB, and that maternal heterosis was of little importance. The ranking of maternal additive breed effects for LLWB was D’Man, Sardi, and Beni Guil in descending order

Cardiac thromboxane A2 receptor activation does not directly induce cardiomyocyte hypertrophy but does cause cell death that is prevented with gentamicin and 2-APB

Abstract Background We have previously shown that the thromboxane (TXA2) receptor agonist, U46619, can directly induce ventricular arrhythmias that were associated with increases in intracellular calcium in cardiomyocytes. Since TXA2 is an inflammatory mediator and induces direct calcium changes in cardiomyocytes, we hypothesized that TXA2 released during ischemia or inflammation could also cause cardiac remodeling. Methods U46619 (0.1-10 μM) was applied to isolated adult mouse ventricular primary cardiomyocytes, mouse ventricular cardiac muscle strips, and cultured HL-1 cardiomyocytes and markers of hypertrophy and cell death were measured. Results We found that TXA2 receptors were expressed in ventricular cardiomyocytes and were functional via calcium imaging. U46619 treatment for 24 h did not increase expression of pathological hypertrophy genes (atrial natriuretic peptide, β-myosin heavy chain, skeletal muscle α-actin) and it did not increase protein synthesis. There was also no increase in cardiomyocyte size after 48 h treatment with U46619 as measured by flow cytometry. However, U46619 (0.1-10 μM) caused a concentration-dependent increase in cardiomyocyte death (trypan blue, MTT assays, visual cell counts and TUNEL stain) after 24 h. Treatment of cells with the TXA2 receptor antagonist SQ29548 and inhibitors of the IP3 pathway, gentamicin and 2-APB, eliminated the increase in cell death induced by U46619. Conclusions Our data suggests that TXA2 does not induce cardiac hypertrophy, but does induce cell death that is mediated in part by IP3 signaling pathways. These findings may provide important therapeutic targets for inflammatory-induced cardiac apoptosis that can lead to heart failure.Peer Reviewe

Fibroblast Growth Factor 23 Does Not Directly Influence Skeletal Muscle Cell Proliferation and Differentiation or Ex Vivo Muscle Contractility

Skeletal muscle dysfunction accompanies the clinical disorders of chronic kidney disease (CKD) and hereditary hypophosphatemic rickets. In both disorders, fibroblast growth factor 23 (FGF23), a bone-derived hormone regulating phosphate and vitamin D metabolism, becomes chronically elevated. FGF23 has been shown to play a direct role in cardiac muscle dysfunction; however, it is unknown whether FGF23 signaling can also directly induce skeletal muscle dysfunction. We found expression of potential FGF23 receptors ( Fgfr1-4) and α-Klotho in muscles of two animal models (CD-1 and Cy/+ rat, a naturally occurring rat model of chronic kidney disease-mineral bone disorder) as well as C2C12 myoblasts and myotubes. C2C12 proliferation, myogenic gene expression, oxidative stress marker 8-OHdG, intracellular Ca2+ ([Ca2+]i), and ex vivo contractility of extensor digitorum longus (EDL) or soleus muscles were assessed after treatment with various amounts of FGF23. FGF23 (2-100 ng/ml) did not alter C2C12 proliferation, expression of myogenic genes, or oxidative stress after 24- to 72-h treatment. Acute or prolonged FGF23 treatment up to 6 days did not alter C2C12 [Ca2+]i handling, nor did acute treatment with FGF23 (9-100 ng/ml) affect EDL and soleus muscle contractility. In conclusion, although skeletal muscles express the receptors involved in FGF23-mediated signaling, in vitro FGF23 treatments failed to directly alter skeletal muscle development or function under the conditions tested. We hypothesize that other endogenous substances may be required to act in concert with FGF23 or apart from FGF23 to promote muscle dysfunction in hereditary hypophosphatemic rickets and CKD

Age-related changes in relative expression stability of commonly used housekeeping genes in selected porcine tissues

<p>Abstract</p> <p>Background</p> <p>Gene expression analysis using real-time RT-PCR (qRT-PCR) is increasingly important in biological research due to the high-throughput and accuracy of qRT-PCR. For accurate and reliable gene expression analysis, normalization of gene expression data against housekeeping genes or internal control genes is required. The stability of reference genes has a tremendous effect on the results of relative quantification of gene expression by qRT-PCR. The expression stability of reference genes could vary according to tissues, age of individuals and experimental conditions. In the pig however, very little information is available on the expression stability of reference genes. The aim of this research was therefore to develop a new set of reference genes which can be used for normalization of mRNA expression data of genes expressed in varieties of porcine tissues at different ages.</p> <p>Results</p> <p>The mRNA expression stability of nine commonly used reference genes (<it>B2M, BLM, GAPDH, HPRT1, PPIA, RPL4, SDHA, TBP </it>and <it>YWHAZ</it>) was determined in varieties of tissues collected from newborn, young and adult pigs. geNorm, NormFinder and BestKeeper software were used to rank the genes according to their stability. geNorm software revealed that <it>RPL4, PPIA </it>and <it>YWHAZ </it>showed high stability in newborn and adult pigs, while <it>B2M, YWHAZ </it>and <it>SDHA </it>showed high stability in young pigs. In all cases, <it>GAPDH </it>showed the least stability in geNorm. NormFinder revealed that <it>TBP </it>was the most stable gene in newborn and young pigs, while <it>PPIA </it>was most stable in adult pigs. Moreover, geNorm software suggested that the geometric mean of three most stable gene would be the suitable combination for accurate normalization of gene expression study.</p> <p>Conclusions</p> <p>Although, there was discrepancy in the ranking order of reference genes obtained by different analysing software methods, the geometric mean of the <it>RPL4, PPIA </it>and <it>YWHAZ </it>seems to be the most appropriate combination of housekeeping genes for accurate normalization of gene expression data in different porcine tissues at different ages.</p

Hyperthermia Induces Functional and Molecular Modifications in Cardiac, Smooth and Skeletal Muscle Cells

Comparative Medicine - OneHealth and Comparative Medicine Poster SessionHyperthermia is used for the treatment of a number of diseases, including muscle injuries, inflammations, tendinitis, and osteoarticular disorder. More recently, hyperthermia has been used as an adjuvant in cancer treatment. Only two studies have shown that hyperthermia leads to hypertrophy in in-vitro models of cardiac and skeletal muscle cells. Functional, biochemical and molecular mechanisms of hyperthermia-induced hypertrophy in muscles remain largely undiscovered. We investigated the effects of mild heat shock (HS) on C2C12 skeletal, HL-1 cardiac and AR-75 smooth muscle cells. Mild HS (20 min 43ºC) induced increases in the cell area in all muscle cells tested. C2C12 cells are a well-accepted model of skeletal muscle fibers, and were selected for complementary studies. First, to biochemically confirm an increase in protein synthesis we measured and found an increase of ~6% in total protein content 24 hrs after HS. Second, we examined potential modifications in calcium (Ca) homeostasis regulation by measuring intracellular Ca. We detected a lower resting level of intracellular Ca and smaller and longer caffeine-induced Ca transients in C2C12 muscle cells 24 hrs after HS. Next, to search for molecular mechanisms involved with HS-induced hypertrophy and calcium homeostasis modifications, mRNA from C2C12 muscle cells was analyzed at different time points after HS (0, 1, 2, and 24 hrs). We used an ABI Step One Plus RT2 PCR Array System and a custom-built 96 gene array. We report for the first time that the expression of key heat-shock, hypertrophy/ metabolic, and Ca+2 signaling genes were altered after HS. Hsp70 and Hsp72 genes were highly expressed (211-1829 fold change) after HS. Also, Myh7 (MHC-I), Myh6, Srf, Ppp3r1 and Pck1 were up-regulated by 2-6 fold change compared with control cells.. Furthermore, a reduction in the expression of RyR and Trdn genes was observed (2- 3.6 fold change) with an associated increase in the expression of IP3R genes (2-4 fold change). These results indicate that hyperthermia modulates not only heat-shock related and hypertrophy genes, but also genes involved with metabolism, apoptosis repression, calcium homeostasis and signaling, and cell homeostasis. Our studies offer an initial exploration of the functional, biochemical and molecular mechanisms that may help explain the beneficially adaptive effects of hyperthermia on muscle function. Our studies shall also prove useful for the refinement of a specific device (EM-Stim) to be employed for the treatment of muscle and bone diseases (See poster by Hatem et al). Importantly, our studies have potential translational applications. By learning how to more precisely use hyperthermia to control specific genes that can improve or treat muscle injuries, musculoskeletal, and cardiovascular diseases, the ensuing benefits shall be unmistakable. Our short and long-term goals are: i) optimize our protocols; ii) test HS in animal models; iii) manipulate expression of promising genes of interest in vitro and in in-vivo animal models; iv) initiate clinical studies to fully translate from the bench to the bed-side

Phosphatidylinositol 3,5‐bisphosphate: Low abundance, high significance

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/102155/1/bies201300012.pd

The TNF-α antagonist etanerceptreverses age-related decreases in colonic SERT expression and faecal output in mice

Treatment for chronic constipation in older people is challenging and the condition has a major impact on quality of life. A lack of understanding about the causes of this condition has hampered the development of effective treatments. 5-HT is an important pro-kinetic agent in the colon. We examined whether alterations in colonic 5-HT signalling underlie age–related changes in faecal output in mice and whether these changes were due to an increase in TNF-α. Components of the 5-HT signalling system (5-HT, 5-HIAA, SERT) and TNF-α expression were examined in the distal colon of 3, 12, 18 and 24- month old mice and faecal output and water content monitored under control conditions and following the administration of etanercept (TNF-α inhibitor; 1 mg Kg-1). Faecal output and water content were reduced in aged animals. Age increased mucosal 5-HT availability and TNF-α expression and decreased mucosal SERT expression and 5-HIAA. Etanercept treatment of old mice reversed these changes, suggesting that age-related changes in TNFα expression are an important regulator of mucosal 5-HT signalling and pellet output and water content in old mice. These data point to “anti-TNFα” drugs as potential treatments for age-related chronic constipation

The importance of the cellular stress response in the pathogenesis and treatment of type 2 diabetes

Organisms have evolved to survive rigorous environments and are not prepared to thrive in a world of caloric excess and sedentary behavior. A realization that physical exercise (or lack of it) plays a pivotal role in both the pathogenesis and therapy of type 2 diabetes mellitus (t2DM) has led to the provocative concept of therapeutic exercise mimetics. A decade ago, we attempted to simulate the beneficial effects of exercise by treating t2DM patients with 3 weeks of daily hyperthermia, induced by hot tub immersion. The short-term intervention had remarkable success, with a 1 % drop in HbA1, a trend toward weight loss, and improvement in diabetic neuropathic symptoms. An explanation for the beneficial effects of exercise and hyperthermia centers upon their ability to induce the cellular stress response (the heat shock response) and restore cellular homeostasis. Impaired stress response precedes major metabolic defects associated with t2DM and may be a near seminal event in the pathogenesis of the disease, tipping the balance from health into disease. Heat shock protein inducers share metabolic pathways associated with exercise with activation of AMPK, PGC1-a, and sirtuins. Diabetic therapies that induce the stress response, whether via heat, bioactive compounds, or genetic manipulation, improve or prevent all of the morbidities and comorbidities associated with the disease. The agents reduce insulin resistance, inflammatory cytokines, visceral adiposity, and body weight while increasing mitochondrial activity, normalizing membrane structure and lipid composition, and preserving organ function. Therapies restoring the stress response can re-tip the balance from disease into health and address the multifaceted defects associated with the disease