MRTF-A regulates myoblast commitment to differentiation by targeting PAX7 during muscle regeneration.

Myocardin-related transcription factor-A/serum response factor (MRTF-A/SRF), a well-known transcriptional programme, has been proposed to play crucial roles in skeletal muscle development and function. However, whether MRTF-A participates in muscle regeneration and the molecular mechanisms are not completely understood. Here, we show that MRTF-A levels are highly correlated with myogenic genes using a RNA-seq assay, which reveal that MRTF-A knockdown in C2C12 cells significantly reduces PAX7 expression. Subsequent in vitro and in vivo data show that MRTF-A and PAX7 present identical expression patterns during myoblast differentiation and CTX-induced muscle injury and repair. Remarkably, MRTF-A overexpression promotes myoblast proliferation, while inhibiting cell differentiation and the expression of MyoD and MyoG. MRTF-A loss of function produces the opposite effect. Moreover, mice with lentivirus (MRTF-A) injection possesses more PAX7+ satellite cells, but less differentiating MyoD+ and MyoG+ cells, leading subsequently to diminished muscle regeneration. Our mechanistic results reveal that MRTF-A contributes to PAX7-mediated myoblast self-renewal, proliferation, and differentiation by binding to its distal CArG box. Overall, we propose that MRTF-A functions as a novel PAX7 regulator upon myoblast commitment to differentiation, which could provide pathways for dictating muscle stem cell fate and open new avenues to explore stem cell-based therapy for muscle degenerative diseases.

Divergent patterns of genic copy number variation in KCNIP1 gene reveal risk locus of type 2 diabetes in Chinese population.

Copy number variation (CNV) has emerged as another important genetic marker in addition to SNP for understanding etiology of complex disease. Kv channel interacting protein 1 (KCNIP1) is a Ca2+-dependent transcriptional modulator that contributes to the regulation of insulin secretion. Previous genome-wide CNV assay identified the KCNIP1 gene encompassing a CNV region, however, its further effect and risk rate on type 2 diabetes (T2D) have rarely been addressed, especially in Chinese population. The current study aims to detect and excavate genetic distribution profile of KCNIP1 CNV in Chinese T2D and control populations, and further to investigate the associations with clinical characteristics. Divergent patterns of the KCNIP1 CNV were identified (p < 0.01), in which the copy number gain was predominant in T2D, while the copy number normal accounted for the most in control group. Consistently, the individuals with copy number gain showed significant risk on T2D (OR = 4.550, p < 0.01). The KCNIP1 copy numbers presented significantly positive correlations with fasting plasma glucose and glycated hemoglobin in T2D. For OGTT test, the T2D patients with copy number gain had remarkably elevated glucose contents (60, 120, 180-min, p < 0.05 or p < 0.01) and diminished insulin levels (60, 120-min, p < 0.05) than those with copy number loss and normal, which suggested that the KCNIP1 CNV was correlated with the glucose and insulin action. This is the first CNV association study of the KCNIP1 gene in Chinese population, and these data indicated that KCNIP1 might function as a T2D-susceptibility gene whose dysregulation alters insulin production.

Ethyl pyruvate downregulates tumor necrosis factor alpha and interleukin (IL)-6 and upregulates IL-10 in lipopolysaccharide-stimulated canine peripheral blood mononuclear cells.

We investigated the anti-inflammatory effects of ethyl pyruvate (EP) on LPS-stimulated canine PBMCs in vitro. We found that EP treatment inhibited the mRNA expressions of proinflammatory cytokines (TNF-α and IL-6), but induced mRNA expression of anti-inflammatory cytokines (IL-10). ELISA measurements revealed that EP also effectively downregulates the LPS-induced increase in proinflammatory cytokine release, while upregulating anti-inflammatory cytokine release. These data indicate that EP could be an effective anti-inflammatory agent in dogs.

Rotavirus-mediated alteration of gut microbiota and its correlation with physiological characteristics in neonatal calves.

Diarrhea is a fatal disease to neonatal calves, and rotavirus is the main pathogen associated with neonatal calf diarrhea. Although previous studies have reported that the gut microbiota is changed in calves during diarrhea, less is known about whether rotavirus infection alters the structure of the gut microbiota. Here, we characterized fecal microbial communities and identified possible relationships between the gut microbiota profiles and physiological parameters. Five fecal specimens of rotavirus-infected calves from 1 to 30 days after birth and five fecal specimens of age-matched healthy calves were used for the microbial community analysis using the Illumina MiSeq sequencer. Rotavirus infection was associated with reduced rotavirus infection significantly reduced the richness and diversity of the bacterial community. Weighted unique fraction metric analysis exhibited significant differences in community membership and structure between healthy and rotavirus-infected calves. Based on relative abundance analysis and linear discriminant analysis effect size, we found that the representative genera from Lactobacillus, Subdoligranulum, Blautia, and Bacteroides were closely related to healthy calves, while the genera Escherichia and Clostridium were closely affiliated to rotavirus-infected calves. Furthermore, canonical correlation analysis and Pearson correlation coefficient results revealed that the increased relative abundances of Lactobacillus, Subdoligranulum, and Bacteroides were correlated with normal levels of physiological characteristics such as white blood cells, blood urea nitrogen, serum amyloid protein A, and glucose concentration in serum. These results suggest that rotavirus infection alters the structure of the gut microbiota, correlating changes in physiological parameters. This study provides new information on the relationship between gut microbiota and the physiological parameters of rotavirus-mediated diarrheic calves.

CREB1 functional polymorphisms modulating promoter transcriptional activity are associated with type 2 diabetes mellitus risk in Chinese population.

The cAMP responsive element binding protein 1 (CREB1) is a ubiquitous transcription factor that contributes to the regulation of gluconeogenesis. The mechanisms of the CREB1 function remain largely unknown. In this study, we aimed to explore genetic variations in CREB1 promoter region and determine whether these loci affect transcriptional activity and risk on type 2 diabetes (T2D). Three polymorphisms were identified and designated as MU1, MU2 and MU3, respectively. Genotypic distribution analysis revealed that MU1 genotypes presented similar distribution between T2D and healthy controls (P > 0.05), while the MU2 and MU3 showed significant differences (P < 0.05). Haplotypic blocks of the three loci were constructed, and H1-TGA, H2-TTT and H3-ATT had higher frequencies in T2D patients than those in controls. Association studies revealed that the three loci significantly affected plasma glucose, glycated hemoglobin and insulin secretion. Disequilibrium analysis identified that the MU2 and MU3 variants were strongly linked in T2D (r2 = 0.348, D' = 1.0). Further analysis indicated that MU2 (TT vs GG, OR = 2.38, 95%CI = 1.19-4.77, P = 0.01) and MU3 (AA vs TT, OR = 1.16, 95%CI = 1.19-4.77, P = 0.04) were significantly associated with T2D in dominant genotypes. Luciferase assay showed that T-A haplotype from the highly linked MU2 and MU3 exhibited maximal promoter activity, which was consistent with the correlation results. We concluded that the TT genotype of MU2 and the AA genotype of MU3 could be used as molecular markers for evaluating the risk on T2D.

Hyperoside and let-7a-5p synergistically inhibits lung cancer cell proliferation via inducing G1/S phase arrest.

Lung cancer remains one of the most aggressive human malignancies with a low survival rate. Hyperoside (quercetin-3-O-ß-d-galactopyranoside) is a flavonol glycoside with an anti-cancer activity. The microRNA-let-7 was widely regarded as a tumor suppressor in human tumors. Here, we investigated the role of hyperoside and let-7a-5p on the lung cancer cell proliferation, cell cycle and apoptosis in A549 cells in vitro. Our results showed that hyperoside could inhibit the proliferation of A549 cells through inducing apoptosis and G1/S phase arrest. Let-7a-5p could inhibit the proliferation of A549 cells via inhibiting the process of G1/S phase. Additionally, hyperoside and let-7a-5p had a synergetic effect on suppressing the proliferation of A549 cells; microRNA-let-7a-5p directly regulated the expression of CCND1 in A549 cells. Our study illustrated that hyperoside and microRNA-let7a-5p might provide a synergistic effect on anti-cancer, which may provide a new idea for lung cancer treatment.

Changes in gene expression of tumor necrosis factor alpha and interleukin 6 in a canine model of caerulein-induced pancreatitis.

Acute pancreatitis is an inflammatory process that frequently involves peripancreatic tissues and remote organ systems. It has high morbidity and mortality rates in both human and veterinary patients. The severity of pancreatitis is generally determined by events that occur after acinar cell injury in the pancreas, resulting in elevated levels of various proinflammatory mediators, such as interleukin (IL) 1ß and 6, as well as tumor necrosis factor alpha (TNF-α). When these mediators are excessively released into the systemic circulation, severe pancreatitis occurs with systemic complications. This pathophysiological process is similar to that of sepsis; thus, there are many striking clinical similarities between patients with septic shock and those with severe acute pancreatitis. We induced acute pancreatitis using caerulein in dogs and measured the change in the gene expression of proinflammatory cytokines. The levels of TNF-α and IL-6 mRNA peaked at 3 h, at twice the baseline levels, and the serum concentrations of amylase and lipase also increased. Histopathological examination revealed severe hyperemia of the pancreas and hyperemia in the duodenal villi and the hepatic sinusoid. Thus, pancreatitis can be considered an appropriate model to better understand the development of naturally occurring sepsis and to assist in the effective treatment and management of septic patients.

La pancréatite aigüe est un processus inflammatoire qui implique fréquemment les tissus péri-pancréatiques et des systèmes organiques éloignés. Elle a des taux de morbidité et de mortalité élevés autant chez les humains que chez les animaux. La sévérité de la pancréatite est généralement déterminée par des évènements qui se produisent suite à des dommages aux cellules acinaires dans le pancréas, et qui induisent des niveaux élevés de différents médiateurs pro-inflammatoires, tels que l'interleukine (IL) 1ß et 6, ainsi que le facteur nécrosant des tumeurs alpha (TNFα). Lorsque ces médiateurs sont libérés de manière excessive dans la circulation systémique, une pancréatite sévère se produit avec des complications systémiques. Ce processus pathophysiologique est similaire à celui d'un sepsis; donc, il y a plusieurs similarités cliniques entre des patients avec un choc septique et ceux avec une pancréatite aigüe sévère. Nous avons induit une pancréatite aigüe en utilisant de la caeruléine chez des chiens et avons mesuré le changement dans l'expression des gènes des cytokines pro-inflammatoires. Les niveaux d'ARNm de TNFα et d'IL-6 ont culminé après 3 h, atteignant le double des niveaux de base, et les concentrations sériques d'amylase et de lipase augmentèrent également. Un examen histopathologique a révélé une hyperémie sévère du pancréas et une hyperémie dans les villosités duodénales et les sinusoïdes hépatiques. Ainsi, la pancréatite peut être considérée un modèle approprié pour mieux comprendre le développement d'un sepsis naturel et aider dans le traitement efficace et la gestion de patients septiques.(Traduit par Docteur Serge Messier).

Valproic acid attenuates the expression of pro-inflammatory cytokines lipopolysaccharide-treated canine peripheral blood mononuclear cells (in vitro) and in a canine endotoxemia model (in vivo).

Valproic acid (VPA), a known histone deacetylase inhibitor, has been used as an anticonvulsant in dogs. VPA also has anti-inflammatory properties, but there are no reports on the immunomodulatory effects of VPA in canine endotoxemia. In the present study, we demonstrate that the use of VPA significantly reduces the production of early-phase pro-inflammatory cytokines (TNF-α, IL-6) caused by lipopolysaccharide (LPS) stimulation both in vitro and in vivo. For the in vitro study, VPA was evaluated for 24h on LPS (100 ng/ml)-treated canine peripheral blood mononuclear cells (PBMCs) which isolated from 5 healthy Beagle dogs. VPA significantly decreased the mRNA expression of TNF-α and IL-6 in a dose-dependent manner (p<0.05 for IL-6; p<0.01 for TNF-α). Fourteen adult Beagles were studied for in vivo study; nine dogs received a low dose of LPS (10 µg/kg/h) via continuous IV infusion for 12h to induce endotoxemia whereas 5 dogs received normal saline as controls. Four out of 9 endotoxemic dogs were administered VPA (50mg/kg, IV) at 1h and 12h along with the LPS infusion. Three hours after the first administration of VPA, IL-6 mRNA expressions in PBMCs significantly decreased (p=0.033 vs. LPS group). VPA also significantly decreased the circulating TNF-α (p=0.044 vs. LPS group at 3h) and IL-6 protein at 3h (p=0.034 vs. LPS group) and 6h (p=0.026 vs. LPS group) post-treatment. Our study suggests that VPA attenuates the expression of pro-inflammatory cytokines in a canine endotoxemia model in vitro and in vivo. We speculate that valproic acid may be useful for reducing inflammatory cytokine levels in dogs with sepsis.

Kinetics of IL-6 and TNF-α changes in a canine model of sepsis induced by endotoxin.

Sepsis is a major cause of death in veterinary medicine, although a better prognosis can result from an early diagnosis. To speed the diagnosis, the biomarkers TNF-α and IL-6 can provide valuable information regarding systemic inflammatory response. The purpose of this study was to investigate the changes in cytokine levels in an experimental model of sepsis using ELISA and real-time PCR. Ten adult Beagles were studied; seven received an IV bolus of high dose lipopolysaccharide solution (1mg/kg) to induce sepsis. The remaining three beagles were the control group. Blood samples were collected before and 1, 3, 6, 12, 24 and 48 h after administering LPS. Serum IL-6 level peaked at 3h (1.89 ± 0.10 ng/ml) and serum TNF-α peaked at 1h (1.11 ± 0.01 ng/ml). The expression of IL-6 mRNA in peripheral blood mononuclear cells (PBMC) increased 62-fold compared to the control group at 1h; TNF-α mRNA increased by 4.5-fold at 1h. The expressions of IL-6 and TNF-α mRNA in PBMCs changed more rapidly than serum IL-6 and TNF-α concentrations. In addition, TNF-α mRNA levels in PBMCs remained elevated longer than serum TNF-α. Our study establishes the basis for future work aimed at a better understanding of the systemic inflammatory response to infection and sepsis in canine patients.

High-mobility group box 1 as a surrogate prognostic marker in dogs with systemic inflammatory response syndrome.

OBJECTIVE: To evaluate various surrogate markers associated with the inflammatory and counter-inflammatory responses with respect to mortality in dogs with systemic inflammatory response syndrome (SIRS). DESIGN: Prospective observational study. SETTING: Veterinary Teaching Hospital. ANIMALS: Twenty-eight dogs with naturally occurring diseases and SIRS from January 2007 to May 2009. INTERVENTIONS: Upon admission to the veterinary hospital, history and baseline data from the physical examination, including parameters previously defined for meeting SIRS criteria, were documented. Heparinized blood samples were collected and plasma cytokines interleukin-6 (IL-6), IL-10, and high-mobility group box 1 (HMGB1) were measured by sandwich ELISA. MEASUREMENTS AND MAIN RESULTS: In nonsurvivors, median plasma HMGB1 concentrations (0.718 microg/L, interquartile range [IQR]; 0.300-1.626 microg/L) and the ratio of HMGB1 to IL-10 (2.236, IQR; 0.972-5.367) were significantly increased as compared with those found in survivors (0.300 microg/L, IQR; 0.300-0.312 microg/L for HMGB1; 1.017, IQR; 0.862-1.126 for the ratio of HMGB1 to IL-10, P=0.007 and 0.024, respectively). Plasma IL-6, IL-10, and the ratio of IL-6 to IL-10 were not significantly different between groups. Among the parameters studied, HMGB1 and the ratio of HMGB1 to IL-10 performed the best in discriminating outcome in dogs with SIRS according to receiver operator characteristic curve analysis. CONCLUSIONS: Increases in plasma HMGB1 concentration and the ratio of HMGB1 to IL-10 may predict poorer outcomes in dogs with SIRS. The approach described may lead to reliable prognostic biomarkers and new therapeutic concepts in the study of SIRS in dogs.