Load-dependent effects of duodenal glucose on glycemia, gastrointestinal hormones, antropyloroduodenal motility, and energy intake in healthy men

Gastric emptying is a major determinant of glycemia, gastrointestinal hormone release, and appetite. We determined the effects of different intraduodenal glucose loads on glycemia, insulinemia, glucagon-like peptide-1 (GLP-1), glucose-dependent insulinotropic polypeptide (GIP) and cholecystokinin (CCK), antropyloroduodenal motility, and energy intake in healthy subjects. Blood glucose, plasma hormone, and antropyloroduodenal motor responses to 120-min intraduodenal infusions of glucose at 1) 1 ("G1"), 2) 2 ("G2"), and 3) 4 ("G4") kcal/min or of 4) saline ("control") were measured in 10 healthy males in double-blind, randomized fashion. Immediately after each infusion, energy intake at a buffet meal was quantified. Blood glucose rose in response to all glucose infusions (P < 0.05 vs. control), with the effect of G4 and G2 being greater than that of G1 (P < 0.05) but with no difference between G2 and G4. The rises in insulin, GLP-1, GIP, and CCK were related to the glucose load (r > 0.82, P < 0.05). All glucose infusions suppressed antral (P < 0.05), but only G4 decreased duodenal, pressure waves (P < 0.01), resulted in a sustained stimulation of basal pyloric pressure (P < 0.01), and decreased energy intake (P < 0.05). In conclusion, variations in duodenal glucose loads have differential effects on blood glucose, plasma insulin, GLP-1, GIP and CCK, antropyloroduodenal motility, and energy intake in healthy subjects. These observations have implications for strategies to minimize postprandial glycemic excursions in type 2 diabetes.     

Viral interference with antigen presentation does not alter acute or chronic CD8 T cell immunodominance in murine cytomegalovirus infection

Both human CMV and murine CMV (MCMV) elicit large CD8 T cell responses, despite the potent effects of viral genes that interfere with the MHC class I (MHC I) pathway of Ag presentation. To investigate the impact of immune evasion on CD8 T cell priming, we infected mice with wild-type (wt) MCMV or a mutant lacking its MHC I immune evasion genes, Deltam4+m6+m152 MCMV. In acute infection, the two viruses elicited a CD8 T cell response to 26 peptide epitopes that was virtually identical in total size, kinetics, and immunodominance hierarchy. This occurred despite results demonstrating that primary DCs are susceptible to the effects of MCMV's MHC I immune evasion genes. Eight months later, responses to both wt and mutant MCMV displayed the same CD8 T cell "memory inflation" and altered immunodominance that characterize the transition to chronic MCMV infection in C57BL/6 mice. Taken together, these findings suggest either that cross-priming dominates over direct CD8 T cell priming in both acute and chronic MCMV infection, or else that the MHC I immune evasion genes of MCMV are unable to alter direct CD8 T cell priming in vivo. At 2 years postinfection, differences in CD8 T cell immunodominance emerged between individual mice, but on average there were only slight differences between wt and mutant virus infections. Overall, the data indicate that the presence or absence of MHC I immune evasion genes has remarkably little impact on the size or specificity of the MCMV-specific CD8 T cell response over an entire lifetime of infection.     

A genomewide association study of skin pigmentation in a South Asian population

We have conducted a multistage genomewide association study, using 1,620,742 single-nucleotide polymorphisms to systematically investigate the genetic factors influencing intrinsic skin pigmentation in a population of South Asian descent. Polymorphisms in three genes—SLC24A5, TYR, and SLC45A2—yielded highly significant replicated associations with skin-reflectance measurements, an indirect measure of melanin content in the skin. The associations detected in these three genes, in an additive manner, collectively account for a large fraction of the natural variation of skin pigmentation in a South Asian population. Our study is the first to interrogate polymorphisms across the genome, to find genetic determinants of the natural variation of skin pigmentation within a human population.     

The first genomewide interaction and locus-heterogeneity linkage scan in bipolar affective disorder: strong evidence of epistatic effects between loci on chromosomes 2q and 6q

We present the first genomewide interaction and locus-heterogeneity linkage scan in bipolar affective disorder (BPAD), using a large linkage data set (52 families of European descent; 448 participants and 259 affected individuals). Our results provide the strongest interaction evidence between BPAD genes on chromosomes 2q22-q24 and 6q23-q24, which was observed symmetrically in both directions (nonparametric LOD [NPL] scores of 7.55 on 2q and 7.63 on 6q; P<.0001 and P=.0001, respectively, after a genomewide permutation procedure). The second-best BPAD interaction evidence was observed between chromosomes 2q22-q24 and 15q26. Here, we also observed a symmetrical interaction (NPL scores of 6.26 on 2q and 4.59 on 15q; P=.0057 and .0022, respectively). We covered the implicated regions by genotyping additional marker sets and performed a detailed interaction linkage analysis, which narrowed the susceptibility intervals. Although the heterogeneity analysis produced less impressive results (highest NPL score of 3.32) and a less consistent picture, we achieved evidence of locus heterogeneity at chromosomes 2q, 6p, 11p, 13q, and 22q, which was supported by adjacent markers within each region and by previously reported BPAD linkage findings. Our results provide systematic insights in the framework of BPAD epistasis and locus heterogeneity, which should facilitate gene identification by the use of more-comprehensive cloning strategies.     

A canine model of septic shock: balancing animal welfare and scientific relevance

A shock canine pneumonia model that permitted relief of discomfort with the use of objective criteria was developed and validated. After intrabronchial Staphylococcus aureus challenge, mechanical ventilation, antibiotics, fluids, vasopressors, sedatives, and analgesics were titrated based on algorithms for 96 h. Increasing S. aureus (1 to 8 x 10(9) colony-forming units/kg) produced decreasing survival rates (P = 0.04). From 4 to 96 h, changes in arterial-alveolar oxygen gradients, mean pulmonary artery pressure, IL-1, serum sodium levels, mechanical ventilation, and vasopressor support were ordered based on survival time [acute nonsurvivors (< or =24 h until death, n = 8) > or = subacute nonsurvivors (>24 to 96 h until death, n = 8) > or = survivors (> or =96 h until death, n = 22) (all P < 0.05)]. In the first 12 h, increases in lactate and renal abnormalities were greatest in acute nonsurvivors (all P < 0.05). Compared with survivors, subacute nonsurvivors had greater rises in cytokines and liver enzymes and greater falls in platelets, white cell counts, pH, and urine output from 24 to 96 h (all P < 0.05). Importantly, these changes were not attributable to dosages of sedation, which decreased in nonsurvivors [survivors vs. nonsurvivors: 5.0 +/- 1.0 vs. 3.8 +/- 0.7 ml x h(-1) x (fentanyl/midazolam/ medetomidine)(-1); P = 0.02]. In this model, the pain control regimen did not mask changes in metabolic function and lung injury or the need for more hemodynamic and pulmonary support related to increasing severity of sepsis. The integration into this model of both specific and supportive titrated therapies routinely used in septic patients may provide a more realistic setting to evaluate therapies for sepsis.     

A cross-strand Trp Trp pair stabilizes the hPin1 WW domain at the expense of function

Using the human Pin1 WW domain (hPin1 WW), we show that replacement of two nearest neighbor non-hydrogen-bonded residues on adjacent beta-strands with tryptophan (Trp) residues increases beta-sheet thermodynamic stability by 4.8 kJ mol(-1) at physiological temperature. One-dimensional NMR studies confirmed that introduction of the Trp-Trp pair does not globally perturb the structure of the triple-stranded beta-sheet, while circular dichroism studies suggest that the engineered cross-strand Trp-Trp pair adopts a side-chain conformation similar to that first reported for a designed "Trp-zipper" beta-hairpin peptide, wherein the indole side chains stack perpendicular to each other. Even though the mutated side chains in wild-type hPin1 WW are not conserved among WW domains and compose the beta-sheet surface opposite to that responsible for ligand binding, introduction of the cross-strand Trp-Trp pair effectively eliminates hPin1 WW function as assessed by the loss of binding affinity toward a natural peptide ligand. Maximizing both thermodynamic stability and the domain function of hPin1 WW by the above mentioned approach appears to be difficult, analogous to the situation with loop 1 optimization explored previously. That introduction of a non-hydrogen-bonded cross-strand Trp-Trp pair within the hPin1 WW domain eliminates function may provide a rationale for why this energetically favorable pairwise interaction has not yet been identified in WW domains or any other biologically evolved protein with known three-dimensional structure.     

Long-term study on symptomless human metapneumovirus infection in hematopoietic stem cell transplant recipients

From October 2004 through October 2006 a study was performed to evaluate the prevalence of human Metapneumovirus (hMPV) infection in adult hematopoietic stem cell transplant (HSCT) recipients. Sequential nasopharyngeal aspirates (NPA) were collected independently from respiratory symptoms and evaluated for hMPV-RNA by polymerase chain reaction (PCR) and sequence analysis. Results indicate epidemiological and molecular differences between the 2004-2005 and 2005-2006 periods and that hMPV seems not to symptomatically affect HSCT patients or cause late respiratory sequelae. In addition, data collected suggest a hospital origin of hMPV infection in most HSCT patients during the 2004-2005 period.     

Effect of progestins on serum hormones,semen production,and agonistic behavior in the gerenuk (Litocranius walleri walleri)

The use of progestins to suppress endogenous testosterone production to reduce agonistic behavior and prevent semen production was studied in gerenuk. Five male gerenuk (20 months to 3 years of age), housed as a bachelor group, were treated with 3 monthly injections of medroxyprogesterone acetate (MPA; 2.5–20 mg/kg), followed by a melengestrol acetate implant (MGA; 0.3 g/kg) for 2 months. Blood samples collected monthly were assayed for serum testosterone and cortisol using enzyme‐linked immunoassays. Quantitative behavioral data were collected for 30 min 3/week starting 1 month before treatment. Body weight, testes volume, and semen traits were measured before treatment, after MPA treatment, and after MGA treatment. Results showed lower (P<0.05) mean serum testosterone concentrations after MPA (4.34 pg/ml) and MGA (5.02 pg/ml) treatment compared to pre‐treatment values (65.9 pg/ml) in four of five gerenuk. The remaining sub‐adult gerenuk had low testosterone initially (4.9 pg/ml) that did not decrease further with treatment (1.4 and 7.8 pg/ml for MPA and MGA, respectively). Mean serum cortisol concentrations decreased markedly after treatment with MPA (6.0±3.7 ng/ml) and MGA (0.8±0.3 ng/ml). Cortisol concentrations were regained rapidly post‐treatment (42.8±4.8 ng/ml) and were not significantly different from the pre‐treatment value (60.6±12.6 ng/ml; P>0.05). The mean incidence of combined aggressive/dominant behaviors (horning, sparring, supplanting, threat) was not different before and after treatment. Body weight, total numbers of spermatozoa produced per ejaculate, percent motility, and percent normal spermatozoa declined maximally 8 months after treatment. Mean testes volume decreased (P<0.05) after MGA treatment (10.53 cm³vs. 11.96 cm³ pre‐treatment). Elevated hepatic enzymes and bile acids were seen in three of five animals after progestin treatment and anorexia was noted in two males after MGA implant removal, however two of three males had elevated liver enzymes before progestin treatment began. Results show that reducing serum testosterone concentration does not seem to modify agonistic behavior in bachelor gerenuk groups. Zoo Biol 26:245–257, 2007. © 2007 Wiley‐Liss, Inc.     

Shp2 in PC12 cells: NGF versus EGF signalling

The balance between specific signals from different growth factors dictates the biological response of mammalian cells including cell proliferation, differentiation and survival. PC12 cells represent a model of choice to compare the signalling of differentiative growth factors, as NGF, and of mitogenic growth factors, as EGF. In these cells the prolonged activity of the ERK kinase dictates the decision of cells to differentiate. Here we focused on the cytosolic tyrosine phosphatase Shp2 as an established regulator of the Ras-ERK cascade, to elucidate its involvement in determining the stimulation-dependent PC12 cell fate. To this end, we generated PC12 derived cell lines that express the interfering mutant of Shp2 under a tetracycline-inducible promoter. Our findings show that Shp2 participates to the opposite effects induced in PC12 cells by EGF and NGF and that the interactions with the multidocking Gab2 protein mediate such effects.