Efficacy of gallium maltolate against Lawsonia intracellularis infection in a rabbit model.

Antimicrobial efficacy against Lawsonia intracellularis is difficult to evaluate in vitro, thus, the effects of gallium maltolate's (GaM) were investigated in a rabbit model for equine proliferative enteropathy (EPE). Juvenile (5-6-week-old) does were infected with 3.0 × 10(8) L. intracellularis/rabbit and allocated into three groups (n = 8). One week postinfection, one group was treated with GaM, 50 mg/kg; one, with doxycycline, 5 mg/kg; and one with a sham-treatment (control). Feces and blood were collected daily and weekly, respectively, to verify presence of L. intracellularis fecal shedding using qPCR, and seroconversion using immunoperoxidase monolayer assay. Rabbits were sacrificed after 1 week of treatment to collect intestinal tissues focusing on EPE-affected sections. Intestinal lesions were confirmed via immunohistochemistry. No difference was noted between treatments regarding EPE-lesions in jejunum (P = 0.51), ileum (P = 0.74), and cecum (P = 0.35), or in L. intracellularis fecal shedding (P = 0.64). GaM and doxycycline appear to have similar efficacy against EPE in infected rabbits.

Pharmacokinetics of gallium maltolate in Lawsonia intracellularis-infected and uninfected rabbits.

Oral gallium maltolate (GaM) pharmacokinetics (PK) and intestinal tissue (IT) concentrations of elemental gallium ([Ga]) and iron ([Fe]) were investigated in a rabbit model of equine proliferative enteropathy (EPE). New Zealand white does (uninfected controls and EPE-infected, n = 6/group) were given a single oral GaM dose (50 mg/kg). Serial blood samples were collected from 0 to 216 h post-treatment (PT) and IT samples after euthanasia. Serology, qPCR, and immunohistochemistry confirmed, or excluded, EPE. Blood and IT [Ga] and [Fe] were determined using inductively coupled plasma-mass spectrometry. PK parameters were estimated through noncompartmental approaches. For all statistical comparisons on [Ga] and [Fe] α = 5%. The Ga log-linear terminal phase rate constant was lower in EPE rabbits vs. uninfected controls [0.0116 ± 0.004 (SD) vs. 0.0171 ± 0.0028 per hour; P = 0.03]; but half-life (59.4 ± 24.0 vs. 39.4 ± 10.8 h; P = 0.12); Cmax (0.50 ± 0.21 vs. 0.59 ± 0.42 µg/mL; P = 0.45); tmax (1.75 ± 0.41 vs. 0.9 ± 0.37 h; P = 0.20); and oral clearance (6.743 ± 1.887 vs. 7.208 ± 2.565 L/h; P = 0.74) were not. IT's [Ga] and [Fe] were higher (P < 0.0001) in controls. In conclusion, although infection reduces IT [Ga] and [Fe], a 48 h GaM dosing interval is appropriate for multidose studies in EPE rabbits.

Skeletal muscle-specific overexpression of SIRT1 does not enhance whole-body energy expenditure or insulin sensitivity in young mice.

AIMS/HYPOTHESIS: The NAD(+)-dependent protein deacetylase sirtuin (SIRT)1 is thought to be a key regulator of skeletal muscle metabolism. However, its precise role in the regulation of insulin sensitivity is unclear. Accordingly, we sought to determine the effect of skeletal muscle-specific overexpression of SIRT1 on skeletal muscle insulin sensitivity and whole-body energy metabolism. METHODS: At 10 weeks of age, mice with muscle-specific overexpression of SIRT1 and their wild-type littermates were fed a standard diet with free access to chow or an energy-restricted (60% of standard) diet for 20 days. Energy expenditure and body composition were measured by indirect calorimetry and magnetic resonance imaging, respectively. Skeletal muscle insulin-stimulated glucose uptake was measured ex vivo in soleus and extensor digitorum longus muscles using a 2-deoxyglucose uptake technique with a physiological insulin concentration of 360 pmol/l (60 µU/ml). RESULTS: Sirt1 mRNA and SIRT1 protein levels were increased by approximately 100- and 150-fold, respectively, in skeletal muscle of mice with SIRT1 overexpression compared with wild-type mice. Despite this large-scale overexpression of SIRT1, body composition, whole-body energy expenditure, substrate oxidation and voluntary activity were comparable between genotypes. Similarly, skeletal muscle basal and insulin-stimulated glucose uptake were unaltered with SIRT1 overexpression. Finally, while 20 days of energy restriction enhanced insulin-stimulated glucose uptake in skeletal muscles of wild-type mice, no additional effect of SIRT1 overexpression was observed. CONCLUSIONS/INTERPRETATION: These results demonstrate that upregulation of SIRT1 activity in skeletal muscle does not affect whole-body energy expenditure or enhance skeletal muscle insulin sensitivity in young mice on a standard diet with free access to chow or in young mice on energy-restricted diets.

Mouse hypothalamic GT1-7 cells demonstrate AMPK-dependent intrinsic glucose-sensing behaviour.

AIMS/HYPOTHESIS: Hypothalamic glucose-excited (GE) neurons contribute to whole-body glucose homeostasis and participate in the detection of hypoglycaemia. This system appears defective in type 1 diabetes, in which hypoglycaemia commonly occurs. Unfortunately, it is at present unclear which molecular components required for glucose sensing are produced in individual neurons and how these are functionally linked. We used the GT1-7 mouse hypothalamic cell line to address these issues. METHODS: Electrophysiological recordings, coupled with measurements of gene expression and protein levels and activity, were made from unmodified GT1-7 cells and cells in which AMP-activated protein kinase (AMPK) catalytic subunit gene expression and activity were reduced. RESULTS: Hypothalamic GT1-7 neurons express the genes encoding glucokinase and ATP-sensitive K(+) channel (K(ATP)) subunits K ( ir ) 6.2 and Sur1 and exhibit GE-type glucose-sensing behaviour. Lowered extracellular glucose concentration hyperpolarised the cells in a concentration-dependent manner, an outcome that was reversed by tolbutamide. Inhibition of glucose uptake or metabolism hyperpolarised cells, showing that energy metabolism is required to maintain their resting membrane potential. Short hairpin (sh)RNA directed to Ampkα2 (also known as Prkaa2) reduced GT1-7 cell AMPKα2, but not AMPKα1, activity and lowered the threshold for hypoglycaemia-induced hyperpolarisation. shAmpkα1 (also known as Prkaa1) had no effect on glucose-sensing or AMPKα2 activity. Decreased uncoupling protein 2 (Ucp2) mRNA was detected in AMPKα2-reduced cells, suggesting that AMPKα2 regulates UCP2 levels. CONCLUSIONS/INTERPRETATION: We have demonstrated that GT1-7 cells closely mimic GE neuron glucose-sensing behaviour, and reducing AMPKα2 blunts their responsiveness to hypoglycaemic challenge, possibly by altering UCP2 activity. These results show that suppression of AMPKα2 activity inhibits normal glucose-sensing behaviour and may contribute to defective detection of hypoglycaemia.

Investigating the correlation between bone density and fracture frequency in the mandibular condyle with micro-computed tomography.

Fractures of the mandibular condyle are common and include diacapitular fractures that affect the condylar head. The medial part of the condylar head is least commonly fractured, possibly due to decreased propensity for lines of force to run in the region. Micro-computed tomography (X-ray microtomography) of five temporomandibular joint specimens was conducted to explore whether trabecular bone structure correlates positively with fracture prevalence, which could reflect adaptation in response to lower exposure to physiological loads throughout life. Models of trabecular bone, and graphic representation of bone density indicated least dense bone medially, but a statistically significant ANOVA result was not obtained. Further study is required to verify whether a relationship between bone microstructure and fracture frequency exists, and whether or not this is the product of association between the directions of physiological and traumatic forces.

Leisure time sedentary behavior, physical activity and frequency of protein consumption on lower extremity strength and lean mass.

BACKGROUND/OBJECTIVES: The purpose of this study was to evaluate the independent and combined associations of moderate-to-vigorous physical activity (MVPA), leisure time sedentary behavior and daily protein consumption on lower extremity muscular strength and lean mass. SUBJECTS/METHODS: Data from the 1999-2002 NHANES were utilized (N=1080 adults 50-85 y). Leg lean mass was estimated from dual-energy x-ray absorptiometry scans. Knee extensor strength was assessed objectively using the Kin Com MP dynamometer. MVPA and leisure time sedentary behavior were assessed via questionnaire, with the number of meals per day of ⩾30 g of protein per meal assessed via a 'multiple pass' 24-h dietary interview. RESULTS: Meeting MVPA guidelines (ß=16.3, P=0.02) and consuming at least two meals per day of ⩾30 g of protein per meal (ß=28.8, P=0.02) were independently associated with greater lower extremity strength, whereas sedentary behavior was not (ß=11.6, P=0.23). Finally, there was no evidence of a three-way interaction of these behaviors on lower extremity strength (ß=-8.7; P=0.70) or lower extremity lean mass (ß=144.5; P=0.75). CONCLUSIONS: Although MVPA and frequency of protein consumption of ⩾30 g of protein per meal were independently associated with lower extremity lean mass and strength, the results of the present study do not provide evidence to suggest that there is a three-way interplay between MVPA, sedentary behavior and frequency of protein consumption ⩾30 g of protein per meal on lower extremity lean mass and strength.

New strategies in sport nutrition to increase exercise performance.

Despite over 50 years of research, the field of sports nutrition continues to grow at a rapid rate. Whilst the traditional research focus was one that centred on strategies to maximise competition performance, emerging data in the last decade has demonstrated how both macronutrient and micronutrient availability can play a prominent role in regulating those cell signalling pathways that modulate skeletal muscle adaptations to endurance and resistance training. Nonetheless, in the context of exercise performance, it is clear that carbohydrate (but not fat) still remains king and that carefully chosen ergogenic aids (e.g. caffeine, creatine, sodium bicarbonate, beta-alanine, nitrates) can all promote performance in the correct exercise setting. In relation to exercise training, however, it is now thought that strategic periods of reduced carbohydrate and elevated dietary protein intake may enhance training adaptations whereas high carbohydrate availability and antioxidant supplementation may actually attenuate training adaptation. Emerging evidence also suggests that vitamin D may play a regulatory role in muscle regeneration and subsequent hypertrophy following damaging forms of exercise. Finally, novel compounds (albeit largely examined in rodent models) such as epicatechins, nicotinamide riboside, resveratrol, ß-hydroxy ß-methylbutyrate, phosphatidic acid and ursolic acid may also promote or attenuate skeletal muscle adaptations to endurance and strength training. When taken together, it is clear that sports nutrition is very much at the heart of the Olympic motto, Citius, Altius, Fortius (faster, higher, stronger).

Site-specific recombination by the bacteriophage P1 lox-Cre system. Cre-mediated synapsis of two lox sites.

The bacteriophage P1-encoded recombinase Cre forms a simple DNA-protein complex at the specific recognition site loxP. Furthermore, Cre is able to mediate a synaptic union of two loxP sites. When two loxP sites are on the same linear DNA molecule, Cre binds the two sites together to form a circular protein-DNA complex. These complexes can be resolved into a linear DNA molecule and a closed circular DNA molecule, the end products of site-specific recombination.

Inhibition of the type I restriction-modification enzymes EcoB and EcoK by the gene 0.3 protein of bacteriophage T7.

The gene 0.3 protein of bacteriophage T7 is a potent inhibitor of the restriction-modification enzymes EcoB and EcoK, both in vivo and in vitro. We have analyzed the ability of purified 0.3 protein to inhibit different steps in the reactions of EcoB and EcoK with DNA. Most of our experiments were done with EcoK, but selected tests with EcoB indicate that the two enzymes are affected by 0.3 protein in the same way. Purified 0.3 protein binds tightly to free enzyme, apparently to one of the small subunits, and prevents it from binding to DNA. If EcoK is allowed to form specific recognition complexes with unmodified DNA before 0.3 protein is added, relatively low levels of 0.3 protein prevent the nuclease activity that would otherwise appear upon addition of ATP, but considerably higher levels are needed to prevent formation of filter-binding complexes or ATPase activity. This, together with other results, suggests that the binding site for 0.3 protein is protected in recognition complexes and in the early stages of the ATP-stimulated reactions, but that it becomes accessible again before cleavage of the DNA, perhaps after the translocation step. If added after the nuclease reaction is substantially over, 0.3 protein has little effect on ATPase activity, and indeed, the subunit having the binding site for 0.3 protein apparently dissociates from the enzyme-DNA complex. The methylase activity of EcoK on hemi-methylated recognition sites is strongly inhibited by 0.3 protein added at any stage of the reaction.

The effect of the submandibular salivary gland on the erythropoietin response to hypoxia in mice with chronic renal failure.

The role of the submandibular salivary gland (SG) in the renal and extrarenal erythropoietin (Epo) response to hypoxia was evaluated in adult male mice with chronic renal failure from partial nephrectomy. A partial nephrectomy model for chronic renal failure was used in an attempt to evaluate erythropoiesis and Epo production in mice whose renal source of Epo may be compromised and thus more dependent on extrarenal sources. Mice with two-thirds of total renal mass excised developed a three-fold increase in serum creatinine concentration, polyuria, and polydipsia but not anemia. They responded to the combined challenge of hypobaric hypoxia (17 hours, 0.5 atm) and anemia from phenylhydrazine treatment (60 mg/kg intraperitoneally [i.p.]) with a consistent increase in serum Epo. This response was not affected by either acute or chronic bilateral submandibularectomy. However, bilateral submandibularectomy in mice with chronic renal failure was associated with a reduction in serum creatinine (p < 0.01). The possibility that reduction of renal mass might increase extrarenal secretion of Epo was examined in mice with chronic renal failure by removing the kidney remnant and exposing them to a severe hypoxic challenge. While acute bilateral submandibularectomy did not influence the extrarenal Epo response to severe hypoxia, mice with partial nephrectomies had a greater Epo response to hypoxia than control mice with a recent bilateral nephrectomy. In conclusion, the submandibular salivary glands do not appear to be an extrarenal source of Epo, nor do they appear to contribute to the enhanced extrarenal Epo response of mice with chronic renal failure.

The effect of bilateral removal of the submandibulary salivary glands on the erythropoietic response of mice.

The role of the submandibular salivary gland in erythropoiesis in the male mouse (MRC TO strain) was evaluated by subjecting mice without submandibular salivary glands (SX) and control (C) sham-operated mice to a variety of stimuli intended to stress the erythropoietic system. In SX mice, after removal of the submandibular glands at age 4 weeks and observation for 8 weeks, mean hematocrit was the same as in C mice, but mean body weight was less. Bilateral removal of the submandibular glands at age 6 weeks neither affected the rate of fall and subsequent recovery of hematocrit which followed treatment with phenylhydrazine (80 mg/kg intraperitoneally [i.p.] 9 days after operation) nor altered the rate of increase in hematocrit or change in body weight which occurred during hypobaric hypoxia (0.5 atm, > 23 hours/day) for 23 days. Mean (SEM) estimates of serum immunoreactive erythropoietin after 17 hours' continuous hypobaric (0.5 atm) exposure were not significantly different between SX [186 (30) mU/mL, n = 7] and C mice [232 (17) mU/mL, n = 7]. In mice given bilateral nephrectomies at age 6 weeks--2 weeks after SX or C surgery--and then both treated with phenylhydrazine (60 mg/kg i.p.) and exposed for 17 hours to hypobaric (0.5 atm) hypoxia, mean estimates of serum immunoreactive erythropoietin were 22.6 (10.6) mU/mL and 22.3 (5.4) mU/mL in SX (n = 5) and C (n = 5) mice. Results of the study do not support the premise that the submandibular salivary glands either contribute to the erythropoietic response or are a source of extrarenal erythropoietin.

Perceiving persons and groups.

This article analyzes the similarities and differences in forming impressions of individuals and in developing conceptions of groups. In both cases, the perceiver develops a mental conception of the target (individual or group) on the basis of available information and uses that information to make judgments about that person or group. However, a review of existing evidence reveals differences in the outcomes of impressions formed of individual and group targets, even when those impressions are based on the very same behavioral information. A model is proposed to account for these differences. The model emphasizes the role of differing expectancies of unity and coherence in individual and group targets, which in turn engage different mechanisms for processing information and making judgments. Implications of the model are discussed.

Self-stereotyping and social context: the effects of relative in-group size and in-group status.

In 2 laboratory experiments, the tendency to stereotype oneself in terms of one's group membership as a function of the social context was examined. Experiment 1 examined the effects of relative in-group size on self-stereotyping. The results confirmed the prediction that minority members are more likely than majority members to stereotype themselves. Experiment 2 examined the interactive impact of relative in-group size and in-group status. As predicted, a high (relative to a low) status of the in-group increased self-stereotyping primarily for minority members, but not for majority members. Moreover, analyses of the differences in perceived in-group and out-group homogeneity suggest that the in-group homogeneity effect should also be interpreted in terms of self-stereotyping processes. Finally, the interplay between cognitive and motivational determinants of self-stereotyping is discussed as well as a possible distinction between self-stereotyping effects on individual level versus group level self-representations.

Resolving the apparent discrepancy between the incongruency effect and the expectancy-based illusory correlation effect: the TRAP model.

The incongruency effect and the expectancy-based illusory correlation effect seem contradictory because they describe apparently contrasting consequences of previously held expectancies: better recall of incongruent than congruent items but overestimation of congruent items. This article resolves this dilemma by presenting a model that is able to simultaneously predict both of these effects. The Twofold Retrieval by Associative Pathways (TRAP) model adopts the encoding assumptions of person memory models but distinguishes between two different retrieval processes, exhaustive and heuristic, hypothesized to underlie recall and frequency estimation, respectively. Experiment 1 showed that expectancy-based illusory correlation effects and incongruency effects are compatible in that they were produced simultaneously. Experiments 2 and 3 tested and rejected alternative explanations for the obtained pattern of results.