Intestinal ribosomal p70(S6K) signaling is increased in piglet rotavirus enteritis

Recent identification of the mammalian target of rapamycin (mTOR) pathway as an amino acid-sensing mechanism that regulates protein synthesis led us to investigate its role in rotavirus diarrhea. We hypothesized that malnutrition would reduce the jejunal protein synthetic rate and mTOR signaling via its target, ribosomal p70 S6 kinase (p70(S6K)). Newborn piglets were artificially fed from birth and infected with porcine rotavirus on day 5 of life. Study groups included infected (fully fed and 50% protein calorie malnourished) and noninfected fully fed controls. Initially, in "worst-case scenario studies," malnourished infected piglets were killed on days 1, 3, 5, and 11 postinoculation, and jejunal samples were compared with controls to determine the time course of injury and p70(S6K) activation. Using a 2 x 2 factorial design, we subsequently determined if infection and/or malnutrition affected mTOR activation on day 3. Western blot analysis and immunohistochemistry were used to measure total and phosphorylated p70(S6K); [(3)H]phenylalanine incorporation was used to measure protein synthesis; and lactase specific activity and villus-crypt dimensions were used to quantify injury. At the peak of diarrhea, the in vitro jejunal protein synthetic rate increased twofold (compared with the rate in the uninfected pig jejunum), concomitant with increased jejunal p70(S6K) phosphorylation (4-fold) and an increased p70(S6K) level (3-fold, P < 0.05). Malnutrition did not alter the magnitude of p70(S6K) activation. Immunolocalization revealed that infection produced a major induction of cytoplasmic p70(S6K) and nuclear phospho-p70(S6K), mainly in the crypt. A downregulation of semitendinosus muscle p70(S6K) phosphorylation was seen at days 1-3 postinoculation. In conclusion, intestinal activation of p70(S6K) was not inhibited by malnutrition but was strongly activated during an active state of mucosal regeneration.     

Autosomal-Dominant Multiple Pterygium Syndrome Is Caused by Mutations in MYH3

Multiple pterygium syndrome (MPS) is a phenotypically and genetically heterogeneous group of rare Mendelian conditions characterized by multiple pterygia, scoliosis, and congenital contractures of the limbs. MPS typically segregates as an autosomal-recessive disorder, but rare instances of autosomal-dominant transmission have been reported. Whereas several mutations causing recessive MPS have been identified, the genetic basis of dominant MPS remains unknown. We identified four families affected by dominantly transmitted MPS characterized by pterygia, camptodactyly of the hands, vertebral fusions, and scoliosis. Exome sequencing identified predicted protein-altering mutations in embryonic myosin heavy chain (MYH3) in three families. MYH3 mutations underlie distal arthrogryposis types 1, 2A, and 2B, but all mutations reported to date occur in the head and neck domains. In contrast, two of the mutations found to cause MPS in this study occurred in the tail domain. The phenotypic overlap among persons with MPS, coupled with physical findings distinct from other conditions caused by mutations in MYH3, suggests that the developmental mechanism underlying MPS differs from that of other conditions and/or that certain functions of embryonic myosin might be perturbed by disruption of specific residues and/or domains. Moreover, the vertebral fusions in persons with MPS, coupled with evidence of MYH3 expression in bone, suggest that embryonic myosin plays a role in skeletal development.     

Potent bradykinin B1 receptor antagonists: 4-substituted phenyl cyclohexanes

Selective bradykinin (BK) B(1) receptor antagonists have been shown to be antinociceptive in animal models and could be novel therapeutic agents for the treatment of pain and inflammation. Elucidation of the structure-activity relationships of the biphenyl moiety of the lead compound 1 provided a potent new structural class of BK B(1) receptor antagonists.     

Essential role of p38 MAPK for activation of skeletal muscle glucose transport by lithium

Lithium increases glucose transport and glycogen synthesis in insulin-sensitive cell lines and rat skeletal muscle, and has been used as a non-selective inhibitor of glycogen synthase kinase-3 (GSK-3). However, the molecular mechanisms underlying lithium action on glucose transport in mammalian skeletal muscle are unknown. Therefore, we examined the effects of lithium on glucose transport activity, glycogen synthesis, insulin signaling elements (insulin receptor (IR), Akt, and GSK-3beta), and the stress-activated p38 mitogen-activated protein kinase (p38 MAPK) in the absence or presence of insulin in isolated soleus muscle from lean Zucker rats. Lithium (10 mM LiCl) enhanced basal glucose transport by 62% (p < 0.05) and augmented net glycogen synthesis by 112% (p < 0.05). Whereas lithium did not affect basal IR tyrosine phosphorylation or Akt ser(473) phosphorylation, it did enhance (41%, p < 0.05) basal GSK-3beta ser(9) phosphorylation. Lithium further enhanced (p < 0.05) the stimulatory effects of insulin on glucose transport (43%), glycogen synthesis (44%), and GSK-3beta ser(9) phosphorylation (13%). Lithium increased (p < 0.05) p38 MAPK phosphorylation both in the absence (37%) and presence (41%) of insulin. Importantly, selective inhibition of p38 MAPK (using 10 microM A304000) completely prevented the basal activation of glucose transport by lithium, and also significantly reduced (52%, p < 0.05) the lithium-induced enhancement of insulin-stimulated glucose transport. Theses results demonstrate that lithium enhances basal and insulin-stimulated glucose transport activity and glycogen synthesis in insulin-sensitive rat skeletal muscle, and that these effects are associated with a significant enhancement of GSK-3beta phosphorylation. Importantly, we have documented an essential role of p38 MAPK phosphorylation in the action lithium on the glucose transport system in isolated mammalian skeletal muscle.     

The Influence of Physical Activity,Socioeconomic Status,and Ethnicity on the Weight Status of Adolescents

Objective: This study examined the effects of physical activity, television viewing, video game play, socioeconomic status (SES), and ethnicity on body mass index (BMI). Research Methods and Procedures: The sample was 2389 adolescents, 10 to 16 years of age (12.7 ± 1.0 years); 1240 (52%) females and 1149 (48%) males; 77% white and 23% African American; from rural (77%) and urban (23%) settings. BMI and skinfolds were directly assessed. All other data were obtained from questionnaires. Results: Watching television on non‐school days was related to being overweight (p < 0.005). However, when BMI analyses were adjusted for ethnicity and SES, there were no significant effects of television viewing on BMI (p > 0.061). Increased hours of video game play enhanced the risk of being overweight for both genders when analyses were adjusted for ethnicity and SES (p < 0.019). In males, participation in as little as one high‐intensity physical activity 3 to 5 days a week decreased the ethnic‐ and SES‐adjusted relative risk of being overweight (RR = 0.646; CI: 0.427 to 0.977). For females, the ethnic‐ and SES‐adjusted relative risk for being overweight was not significantly altered by physical activity. The logistic analyses further indicated the influence of low SES and African American ethnicity overshadowed any direct effect of television or videos. Discussion: Because weight status of male adolescents appears to be more related to exercise habits than to television or video game habits, increased participation in high‐intensity exercise appears to be important. For females, neither videos nor exercise habits appear to be related to risk of being overweight. However, ethnicity and SES may be important factors that can influence body weight status, while television viewing may be of some importance. Thus, programs to reduce obesity in female adolescent should focus their efforts in lower SES communities.     

Laser capture microdissection and metagenomic analysis of intact mucosa-associated microbial communities of human colon

Metagenomic analysis of colonic mucosa-associated microbes has been complicated by technical challenges that disrupt or alter community structure and function. In the present study, we determined the feasibility of laser capture microdissection (LCM) of intact regional human colonic mucosa-associated microbes followed by phi29 multiple displacement amplification (MDA) and massively parallel sequencing for metagenomic analysis. Samples were obtained from the healthy human subject without bowel preparation and frozen sections immediately prepared. Regional mucosa-associated microbes were successfully dissected using LCM with minimal contamination by host cells, their DNA extracted and subjected to phi29 MDA with a high fidelity, prior to shotgun sequencing using the GS-FLX DNA sequencer. Metagenomic analysis of approximately 67 million base pairs of DNA sequences from two samples revealed that the metabolic functional profiles in mucosa-associated microbes were as diverse as those reported in feces, specifically the representation of functional genes associated with carbohydrate, protein, and nucleic acid utilization. In summary, these studies demonstrate the feasibility of the approach to study the structure and metagenomic profiles of human intestinal mucosa-associated microbial communities at small spatial scales.