Differences in muscle protein synthesis and anabolic signaling in the postabsorptive state and in response to food in 65-80 year old men and women

Women have less muscle than men but lose it more slowly during aging. To discover potential underlying mechanism(s) for this we evaluated the muscle protein synthesis process in postabsorptive conditions and during feeding in twenty-nine 65-80 year old men (n = 13) and women (n = 16). We discovered that the basal concentration of phosphorylated eEF2(Thr56) was approximately 40% less (P<0.05) and the basal rate of MPS was approximately 30% greater (P = 0.02) in women than in men; the basal concentrations of muscle phosphorylated Akt(Thr308), p70s6k(Thr389), eIF4E(Ser209), and eIF4E-BP1(Thr37/46) were not different between the sexes. Feeding increased (P<0.05) Akt(Thr308) and p70s6k(Thr389) phosphorylation to the same extent in men and women but increased (P<0.05) the phosphorylation of eIF4E(Ser209) and eIF4E-BP1(Thr37/46) in men only. Accordingly, feeding increased MPS in men (P<0.01) but not in women. The postabsorptive muscle mRNA concentrations for myoD and myostatin were not different between sexes; feeding doubled myoD mRNA (P<0.05) and halved that of myostatin (P<0.05) in both sexes. Thus, there is sexual dimorphism in MPS and its control in older adults; a greater basal rate of MPS, operating over most of the day may partially explain the slower loss of muscle in older women.     

Effects of resistance exercise with and without creatine supplementation on gene expression and cell signaling in human skeletal muscle.

To test the hypothesis that creatine supplementation would enhance the anabolic responses of muscle cell signaling and gene expression to exercise, we studied nine subjects who received either creatine or a placebo (maltodextrin) for 5 days in a double-blind fashion before undergoing muscle biopsies: at rest, immediately after exercise (10 x 10 repetitions of one-leg extension at 80% 1 repetition maximum), and 24 and 72 h later (all in the morning after fasting overnight). Creatine supplementation decreased the phosphorylation state of protein kinase B (PKB) on Thr308 at rest by 60% (P < 0.05) and that of eukaryotic initiation factor 4E-binding protein on Thr37/46 (4E-BP1) by 30% 24 h postexercise (P < 0.05). Creatine increased mRNA for collagen 1 (alpha(1)), glucose transporter-4 (GLUT-4), and myosin heavy chain I at rest by 250%, 45%, and 80%, respectively, and myosin heavy chain IIA (MHCIIA) mRNA immediately after exercise by 70% (all P < 0.05). Immediately after exercise, and independent of creatine, mRNA for muscle atrophy F-box (MAFbx), MHCIIA, peroxisome proliferator-activated receptor gamma coactivator-1alpha, and interleukin-6 were upregulated (60-350%; P < 0.05); the phosphorylation state of p38 both in the sarcoplasm and nucleus were increased (12- and 25-fold, respectively; both P < 0.05). Concurrently, the phosphorylation states of PKB (Thr308) and 4E-BP1 (Thr37/46) were decreased by 50% and 75%, respectively (P < 0.05). Twenty-four hours postexercise, MAFbx, myostatin, and GLUT-4 mRNA expression decreased below preexercise values (-35 to -50%; P < 0.05); calpain 1 mRNA increased 70% 72 h postexercise (P < 0.05) and at no other time. In conclusion, 5 days of creatine supplementation do not enhance anabolic signaling but increase the expression of certain targeted genes.     

Helicobacter pylori-induced homotypic phagosome fusion in human monocytes is independent of the bacterial vacA and cag status

Following reports that a VacA+cag+ toxigenic but not a VacA-cag- non-toxigenic Helicobacter pylori strain induced homotypic phagosome fusion in murine macrophages, we addressed that phenomenon in human cells. Mononuclear phagocytes and epitheloid cells were challenged with H. pylori strains of different vacA and cag genotypes and with VacA- and Cag- isogenic mutants, and chased in the absence or presence of signal transduction modulators. Electron microscopy revealed that, in monocytes: (i) homotypic phagosome fusion was frequently induced by all live H. pylori strains investigated but not by exogenous VacA; (ii) phagosomes containing bacteria fused, but not those containing latex beads; (iii) fusion resulted in communal compartments resembling giant multivesicular bodies; and (iv) formation of these compartments was blocked by inhibiting the host cell regulators PI 3-kinase, phospholipase C and p42 MAP kinase. Whereas some internalized bacteria remained viable 1 h after uptake, none survived a 24 h period. In contrast to monocytes, infected epitheloid cells rarely developed communal compartments. In combination, these results demonstrate that, in human monocytes, the H. pylori-induced homotypic phagosome fusion depends on neither the vacuolating cytotoxin VacA nor the cag pathogenicity island of H. pylori and does not result in prolonged intracellular survival.     

Genetic and molecular characterization of embryonic mutants identified following seed transformation in Arabidopsis

Over 5000 transgenic families of Arabidopsis thaliana produced following seed transformation with Agrobacterium tumefaciens were screened for embryonic lethals, defectives, and pattern mutants. One hundred and seventy-eight mutants with a wide range of developmental abnormalities were identified. Forty-one mutants appear from genetic studies to be tagged (36% of the 115 mutants examined in detail). Mapping with visible markers demonstrated that mutant genes were randomly distributed throughout the genome. Seven mutant families appeared to contain chromosomal translocations because the mutant genes exhibited linkage to visible markers on two different chromosomes. Chromosomal rearrangements may therefore be widespread following seed transformation. DNA gel blot hybridizations with 34 tagged mutants and three T-DNA probes revealed a wide range of insertion patterns. Models of T-DNA structure at each mutant locus were constructed to facilitate gene isolation. The value of such models was demonstrated by using plasmid rescue to clone flanking plant DNA from four tagged mutants. Further analysis of genes isolated from these insertional mutants should help to elucidate the relationship between gene function and plant embryogenesis.     

Genetic and molecular characterization of embryonic mutants identified following seed transformation in Arabidopsis

Over 5000 transgenic families of Arabidopsis thaliana produced following seed transformation with Agrobacterium tumefaciens were screened for embryonic lethals, defectives, and pattern mutants. One hundred and seventy-eight mutants with a wide range of developmental abnormalities were identified. Forty-one mutants appear from genetic studies to be tagged (36% of the 115 mutants examined in detail). Mapping with visible markers demonstrated that mutant genes were randomly distributed throughout the genome. Seven mutant families appeared to contain chromosomal translocations because the mutant genes exhibited linkage to visible markers on two different chromosomes. Chromosomal rearrangements may therefore be widespread following seed transformation. DNA gel blot hybridizations with 34 tagged mutants and three T-DNA probes revealed a wide range of insertion patterns. Models of T-DNA structure at each mutant locus were constructed to facilitate gene isolation. The value of such models was demonstrated by using plasmid rescue to clone flanking plant DNA from four tagged mutants. Further analysis of genes isolated from these insertional mutants should help to elucidate the relationship between gene function and plant embryogenesis.     

Genetic relatedness of the genomes of equine herpesvirus types 1, 2, and 3.

Genomic DNAs of equine herpesvirus type 1 (EHV-1), EHV-2 (equine cytomegalovirus), and EHV-3 were examined by reassociation kinetic and thermal denaturation analyses to determine the extent and degree of homology among the three viral DNAs. Results of reassociation analyses indicated a limited homology among the three EHV genomes. Homologous DNA sequences equivalent to 1.8 to 3.7 megadaltons between EHV-1 and equine cytomegalovirus, 7.6 to 8.2 megadaltons between EHV-1 and EHV-3, and 1.3 to 1.9 megadaltons between equine cytomegalovirus and EHV-3 were detected. Examination by thermal denaturation of the DNA homoduplexes and heteroduplexes formed during reassociation revealed a high degree of base pairing within the duplexes, suggesting that closely related sequences may be conserved among the genomes of EHV.     

228Th retention and dosimetry in beagles

Total-body and skeletal retention of 228Th were determined in a group of 104 young adult male and female beagles for about the first 7 years after the injection of 0.00159 to 2.76 muCi/kg. Ratios of 224Ra / 228Th , 212Pb / 228Th , and 212Bi / 228Th in the skeleton and in soft tissues of 20 beagles were measured as a function of time after injection. A humerus, femur, and ulna from 20 dogs dying 7 to 554 days after injection were sectioned, and the 228Th concentration was obtained for each piece. Percentage biological retention in the skeleton of 228Th at t days after injection could be described as 68.1 e-0. 000180t . Ratios of daughter-to-parent activity in soft tissue showed no definite trend with dose level or time and averaged Ra/Th = 0.56, Pb/Th = 0.83, and Bi/Th = 0.91, whereas the ratios for the skeleton varied with both dose level and time. Ratios of activity in the skeleton from lowest to highest dose level after 2 years following injection ranged between Ra/Th = 0.88 to 0.95, Pb/Th = 0.78 to 0.92, and Bi/Th = 0.77 to 0.90. Retained 228Th was deposited most heavily in parts of the skeleton with much trabecular bone, much bone surface area, and high bone remodeling rates. No changes in this deposition pattern could be discerned during the 554 days over which the measurements of sectioned long bones were made.     

Protection of mice from herpes simplex virus-induced retinitis by in vitro-activated immune cells.

A form of acute retinal necrosis occurred in the contralateral eyes of susceptible mice 1 week after each received a uniocular injection of live herpes simplex virus type 1 (HSV-1) in the anterior chamber. Although these mice did not develop systemic delayed hypersensitivity to virus antigens, their sera contained virus-specific antibodies at the time contralateral retinitis occurred. These findings suggest that systemic immunity might not be able to protect against contralateral retinitis. To explore this possibility further, we examined lymph nodes and spleens of intraocularly infected mice to determine whether their lymphoid tissues contained primed HSV-1-specific cytotoxic T cells. Virus-specific cytotoxic T cells were readily identified in these mice. We wondered why successful immune priming did not confer protection against HSV-1 retinitis. We examined this issue by evaluating the capacity of in vitro-generated, HSV-1-specific effector T cells to prevent retinitis by infusing these cells by various routes and at various times into mice that received an intracameral injection of HSV-1. The results revealed that virus-specific effector cells could prevent contralateral retinitis if injected intravenously or into the anterior chamber of the contralateral eye at the same time that virus was injected into one eye. However, the effector cells failed to prevent retinitis if they were injected into the same eye that received HSV-1 or if their intravenous administration was delayed until 24 h after the HSV-1 injection into the eye. We concluded that immune T cells can protect against contralateral retinal necrosis caused by uniocular injection of HSV-1 into the anterior chamber but only if they are administered during the first 24 h after virus infection. We propose that a retinitis-inducing process is set in motion during this early time interval postinfection. Once the process has been initiated and established, it is no longer susceptible to immune intervention. It would appear that mice that are susceptible to contralateral retinitis fail to mobilize a protective response quickly enough to ward off the establishment of the retinitis-inducing process and its disastrous eventuality.     

9OSr +90Y bremsstrahlung efficiency predicted for humans

Summary Bremsstrahlung x-rays are produced when energetic-particles are emitted and absorbed. Measurements of total-body bremsstrahlung efficiency (x-ray photon output per Ci⁹⁰Sr in the body, relative to that in water) have been made in the intact mouse, rat, rabbit, and dog sacrificed 2 weeks after the injection of⁹⁰Sr +⁸⁵Sr. Efficiencies were determined by a comparison of the bremsstrahlung output from⁹⁰Sr + its daughter⁹⁰Y and the gamma-ray emission of⁸⁵Sr. Results were checked by a-assay of the ashed animals. Bremsstrahlung efficiencies averaged 1.10 in a 0.04 kg mouse, 1.14 in a 0.13 kg rat, 1.23 in a 2.6 kg rabbit, and 1.32 in an 8.5 kg beagle. Extrapolating to a 70 kg human, a relative bremsstrahlung efficiency of about 1.4 is predicted. An estimate was made of the⁹⁰Sr body content in a former dial painter based on in vivo counting and a bremsstrahlung efficiency of 1.39 predicted for a 55 kg human female by these animal data. Our value of 1.42 ± 0.08 Ci⁹⁰Sr was in good agreement with corresponding results reported for this subject by 8 other laboratories.Supported by USERDA Contract E (11-1)-119