When genetic load does not correlate with phenotypic spectrum: lessons from the GnRH receptor (GNRHR).

CONTEXT: A broad spectrum of GnRH-deficient phenotypes has been identified in individuals with both mono- and biallelic GNRHR mutations. OBJECTIVE: The objective of the study was to determine the correlation between the severity of the reproductive phenotype(s) and the number and functional severity of rare sequence variants in GNRHR. SUBJECTS: Eight hundred sixty-three probands with different forms of GnRH deficiency, 46 family members and 422 controls were screened for GNRHR mutations. The 70 subjects (32 patients and 38 family members) harboring mutations were divided into four groups (G1-G4) based on the functional severity of the mutations (complete or partial loss of function) and the number of affected alleles (monoallelic or biallelic) with mutations, and these classes were mapped on their clinical phenotypes. RESULTS: The prevalence of heterozygous rare sequence variants in GNRHR was significantly higher in probands vs. controls (P < 0.01). Among the G1-G3 groups (homozygous subjects with successively decreasing severity and number of mutations), the hypogonadotropic phenotype related to their genetic load. In contrast, subjects in G4, with only monoallelic mutations, demonstrated a greater diversity of clinical phenotypes. CONCLUSIONS: In patients with GnRH deficiency and biallelic mutations in GNRHR, genetic burden defined by severity and dose is associated with clinical phenotype. In contrast, for patients with monoallelic GNRHR mutations this correlation does not hold. Taken together, these data indicate that as-yet-unidentified genetic and/or environmental factors may combine with singly mutated GNRHR alleles to produce reproductive phenotypes.

Different PTH response to oral peptone load and oral calcium load in patients with normocalcemic primary hyperparathyroidism, primary hyperparathyroidism, and healthy subjects.

BACKGROUND: Normocalcemic primary hyperparathyroidism (PHPT-N) is a condition that may have similar long-term implications to primary hyperparathyroidism (PHPT); however, differential diagnosis and treatment for parathyroid disorders are not clearly defined. We investigated the effect of an oral peptone and an oral calcium load on calcium-regulating hormones in PHPT-N compared with PHPT and healthy controls to provide a new potential diagnostic tool. DESIGN: Case-control study. METHODS: We evaluated serum gastrin, PTH, ionized calcium, and phosphate responses to oral calcium (1 g) and peptone (10 g) load in 22 PHPT and 20 PHPT-N patients matched for PTH serum values. Moreover, 30 healthy subjects were enrolled as controls. In 12 patients for each group, we also performed the oral peptone test adding aluminum hydroxide (AH) to suppress phosphate absorption. RESULTS: In PHPT patients, PTH increased significantly 30 min after the oral peptone load, while no significant increase was found in PHPT-N and controls. After oral calcium load, PTH remained stable in PHPT while it decreased dramatically in PHPT-N patients, and ionized calcium increased significantly in each of the three groups. Peptones plus AH induced a blunted PTH increase in the three groups. CONCLUSIONS: Considering the marked difference in PTH response elicited by peptones in PHPT compared with PHPT-N, we suggest that the oral peptone test could be added to the diagnostic evaluation of PHPT patients. In case of absent response to peptones, patients should have their serum calcium levels assessed twice a year in accordance with recent guidelines.

Joint evolution of dispersal and inbreeding load.

Inbreeding avoidance is often invoked to explain observed patterns of dispersal, and theoretical models indeed point to a possibly important role. However, while inbreeding load is usually assumed constant in these models, it is actually bound to vary dynamically under the combined influences of mutation, drift, and selection and thus to evolve jointly with dispersal. Here we report the results of individual-based stochastic simulations allowing such a joint evolution. We show that strongly deleterious mutations should play no significant role, owing to the low genomic mutation rate for such mutations. Mildly deleterious mutations, by contrast, may create enough heterosis to affect the evolution of dispersal as an inbreeding-avoidance mechanism, but only provided that they are also strongly recessive. If slightly recessive, they will spread among demes and accumulate at the metapopulation level, thus contributing to mutational load, but not to heterosis. The resulting loss of viability may then combine with demographic stochasticity to promote population fluctuations, which foster indirect incentives for dispersal. Our simulations suggest that, under biologically realistic parameter values, deleterious mutations have a limited impact on the evolution of dispersal, which on average exceeds by only one-third the values expected from kin-competition avoidance.

Effects of common origin and common rearing environment on variance in ectoparasite load and phenotype of nestling Alpine swifts

Knowledge of the quantitative genetics of resistance to parasitism is key to appraise host evolutionary responses to parasite selection. Here, we studied effects of common origin (i.e. genetic and pre-hatching parental effects) and common rearing environment (i.e. post-hatching parental effects and other environment effects) on variance in ectoparasite load in nestling Alpine swifts (Apus melba). This colonial bird is intensely parasitized by blood sucking louse-flies that impair nestling development and survival. By cross-fostering half of the hatchlings between pairs of nests, we show strong significant effect of common rearing environment on variance (90.7% in 2002 and 90.9% in 2003) in the number of louse-flies per nestling and no significant effect of common origin on variance in the number of louse-flies per nestling. In contrast, significant effects of common origin were found for all the nestling morphological traits (i.e. body mass, wing length, tail length, fork length and sternum length) under investigation. Hence, our study suggests that genetic and pre-hatching parental effects play little role in the distribution of parasites among nestling Alpine swifts, and thus that nestlings have only limited scope for evolutionary responses against parasites. Our results highlight the need to take into consideration environmental factors, including the evolution of post-hatching parental effects such as nest sanitation, in our understanding of host-parasite relationships.

Bystander-Activated Memory CD8 T Cells Control Early Pathogen Load in an Innate-like, NKG2D-Dependent Manner.

During an infection the antigen-nonspecific memory CD8 T cell compartment is not simply an inert pool of cells, but becomes activated and cytotoxic. It is unknown how these cells contribute to the clearance of an infection. We measured the strength of T cell receptor (TCR) signals that bystander-activated, cytotoxic CD8 T cells (BA-CTLs) receive in vivo and found evidence of limited TCR signaling. Given this marginal contribution of the TCR, we asked how BA-CTLs identify infected target cells. We show that target cells express NKG2D ligands following bacterial infection and demonstrate that BA-CTLs directly eliminate these target cells in an innate-like, NKG2D-dependent manner. Selective inhibition of BA-CTL-mediated killing led to a significant defect in pathogen clearance. Together, these data suggest an innate role for memory CD8 T cells in the early immune response before the onset of a de novo generated, antigen-specific CD8 T cell response.

Inefficacité de la gestion de l'hyperglycémie du patient hospitalisé: origines et remèdes [The inefficacy of glycemic control in the hospitalized patient: origins and solutions]

La prévalence du diabète peut être estimée entre 20 et 30% parmi les patients en hôpital aigu. Il a été démontré que l'hyperglycémie, même modérée, est associée à une augmentation de la morbi-mortalité hospitalière, tandis que le contrôle glycémique efficace a un impact favorable sur celle-ci. La prise en charge de l'hyperglycémie demeure pourtant largement inefficace hors des soins intensifs, en raison de la persistance d'une pratique inadaptée. Nous développons actuellement un projet de soins destiné à faire changer les pratiques. Pour un contrôle glycémique efficace, une formation des soignants à une gestion basée sur le concept de couverture des besoins en insuline du patient est nécessaire. La démarche doit être intégrée à une approche de type systémique, prenant en compte le contexte dans lequel les soignants évoluent. The hospital inpatient prevalence of diabetes mellitus can be estimated between 20 and 30%. Even moderate hyperglycemia is associated with increased morbidity and mortality in the acute care setting, whereas efficient glycemic control has been shown to improve both of them significantly. Glycemic control however remains largely inefficient outside of the intensive care unit due to the persistance of an inadequate glycemic management practice. We are currently developing a clinical care project aimed at changing this practice. For an efficient glycemic control, a training programme for health care professionals based on the concept of covering the insulin needs of the patient is mandatory. This programme needs to be integrated in a systemic approach, which takes the professionals' context in account.

How reliable is an undetectable viral load?

OBJECTIVES: An article by the Swiss AIDS Commission states that patients with stably suppressed viraemia [i.e. several successive HIV-1 RNA plasma concentrations (viral loads, VL) below the limits of detection during 6 months or more of highly active antiretroviral therapy (HAART)] are unlikely to be infectious. Questions then arise: how reliable is the undetectability of the VL, given the history of measures? What factors determine reliability? METHODS: We assessed the probability (henceforth termed reliability) that the n+1 VL would exceed 50 or 1000 HIV-1 RNA copies/mL when the nth one had been <50 copies/mL in 6168 patients of the Swiss HIV Cohort Study who were continuing to take HAART between 2003 and 2007. General estimating equations were used to analyse potential factors of reliability. RESULTS: With a cut-off at 50 copies/mL, reliability was 84.5% (n=1), increasing to 94.5% (n=5). Compliance, the current type of HAART and the first antiretroviral therapy (ART) received (HAART or not) were predictive factors of reliability. With a cut-off at 1000 copies/mL, reliability was 97.5% (n=1), increasing to 99.1% (n=4). Chart review revealed that patients had stopped their treatment, admitted to major problems with compliance or were taking non-HAART ART in 72.2% of these cases. Viral escape caused by resistance was found in 5.6%. No explanation was found in the charts of 22.2% of cases. CONCLUSIONS: After several successive VLs at <50 copies/mL, reliability reaches approximately 94% with a cut-off of 50 copies/mL and approximately 99% with a cut-off at 1000 copies/mL. Compliance is the most important factor predicting reliability.

Polymorphisms in toll-like receptor 4 and toll-like receptor 9 influence viral load in a seroincident cohort of HIV-1-infected individuals.

OBJECTIVES: Toll-like receptors (TLRs) are innate immune sensors that are integral to resisting chronic and opportunistic infections. Mounting evidence implicates TLR polymorphisms in susceptibilities to various infectious diseases, including HIV-1. We investigated the impact of TLR single nucleotide polymorphisms (SNPs) on clinical outcome in a seroincident cohort of HIV-1-infected volunteers. DESIGN: We analyzed TLR SNPs in 201 antiretroviral treatment-naive HIV-1-infected volunteers from a longitudinal seroincident cohort with regular follow-up intervals (median follow-up 4.2 years, interquartile range 4.4). Participants were stratified into two groups according to either disease progression, defined as peripheral blood CD4(+) T-cell decline over time, or peak and setpoint viral load. METHODS: Haplotype tagging SNPs from TLR2, TLR3, TLR4, and TLR9 were detected by mass array genotyping, and CD4(+) T-cell counts and viral load measurements were determined prior to antiretroviral therapy initiation. The association of TLR haplotypes with viral load and rapid progression was assessed by multivariate regression models using age and sex as covariates. RESULTS: Two TLR4 SNPs in strong linkage disequilibrium [1063 A/G (D299G) and 1363 C/T (T399I)] were more frequent among individuals with high peak viral load compared with low/moderate peak viral load (odds ratio 6.65, 95% confidence interval 2.19-20.46, P < 0.001; adjusted P = 0.002 for 1063 A/G). In addition, a TLR9 SNP previously associated with slow progression was found less frequently among individuals with high viral setpoint compared with low/moderate setpoint (odds ratio 0.29, 95% confidence interval 0.13-0.65, P = 0.003, adjusted P = 0.04). CONCLUSION: This study suggests a potentially new role for TLR4 polymorphisms in HIV-1 peak viral load and confirms a role for TLR9 polymorphisms in disease progression.

Impact of tight glycemic control on cerebral glucose metabolism after severe brain injury: a microdialysis study.

OBJECTIVES: To analyze the effect of tight glycemic control with the use of intensive insulin therapy on cerebral glucose metabolism in patients with severe brain injury. DESIGN: Retrospective analysis of a prospective observational cohort. SETTING: University hospital neurologic intensive care unit. PATIENTS: Twenty patients (median age 59 yrs) monitored with cerebral microdialysis as part of their clinical care. INTERVENTIONS: Intensive insulin therapy (systemic glucose target: 4.4-6.7 mmol/L [80-120 mg/dL]). MEASUREMENTS AND MAIN RESULTS: Brain tissue markers of glucose metabolism (cerebral microdialysis glucose and lactate/pyruvate ratio) and systemic glucose were collected hourly. Systemic glucose levels were categorized as within the target "tight" (4.4-6.7 mmol/L [80-120 mg/dL]) vs. "intermediate" (6.8-10.0 mmol/L [121-180 mg/dL]) range. Brain energy crisis was defined as a cerebral microdialysis glucose <0.7 mmol/L with a lactate/pyruvate ratio >40. We analyzed 2131 cerebral microdialysis samples: tight systemic glucose levels were associated with a greater prevalence of low cerebral microdialysis glucose (65% vs. 36%, p < 0.01) and brain energy crisis (25% vs.17%, p < 0.01) than intermediate levels. Using multivariable analysis, and adjusting for intracranial pressure and cerebral perfusion pressure, systemic glucose concentration (adjusted odds ratio 1.23, 95% confidence interval [CI] 1.10-1.37, for each 1 mmol/L decrease, p < 0.001) and insulin dose (adjusted odds ratio 1.10, 95% CI 1.04-1.17, for each 1 U/hr increase, p = 0.02) independently predicted brain energy crisis. Cerebral microdialysis glucose was lower in nonsurvivors than in survivors (0.46 +/- 0.23 vs. 1.04 +/- 0.56 mmol/L, p < 0.05). Brain energy crisis was associated with increased mortality at hospital discharge (adjusted odds ratio 7.36, 95% CI 1.37-39.51, p = 0.02). CONCLUSIONS: In patients with severe brain injury, tight systemic glucose control is associated with reduced cerebral extracellular glucose availability and increased prevalence of brain energy crisis, which in turn correlates with increased mortality. Intensive insulin therapy may impair cerebral glucose metabolism after severe brain injury.

Predicting the deleterious effects of mutation load in fragmented populations.

Human-induced habitat fragmentation constitutes a major threat to biodiversity. Both genetic and demographic factors combine to drive small and isolated populations into extinction vortices. Nevertheless, the deleterious effects of inbreeding and drift load may depend on population structure, migration patterns, and mating systems and are difficult to predict in the absence of crossing experiments. We performed stochastic individual-based simulations aimed at predicting the effects of deleterious mutations on population fitness (offspring viability and median time to extinction) under a variety of settings (landscape configurations, migration models, and mating systems) on the basis of easy-to-collect demographic and genetic information. Pooling all simulations, a large part (70%) of variance in offspring viability was explained by a combination of genetic structure (F(ST)) and within-deme heterozygosity (H(S)). A similar part of variance in median time to extinction was explained by a combination of local population size (N) and heterozygosity (H(S)). In both cases the predictive power increased above 80% when information on mating systems was available. These results provide robust predictive models to evaluate the viability prospects of fragmented populations.

Large-scale association analyses identify new loci influencing glycemic traits and provide insight into the underlying biological pathways.

Through genome-wide association meta-analyses of up to 133,010 individuals of European ancestry without diabetes, including individuals newly genotyped using the Metabochip, we have increased the number of confirmed loci influencing glycemic traits to 53, of which 33 also increase type 2 diabetes risk (q < 0.05). Loci influencing fasting insulin concentration showed association with lipid levels and fat distribution, suggesting impact on insulin resistance. Gene-based analyses identified further biologically plausible loci, suggesting that additional loci beyond those reaching genome-wide significance are likely to represent real associations. This conclusion is supported by an excess of directionally consistent and nominally significant signals between discovery and follow-up studies. Functional analysis of these newly discovered loci will further improve our understanding of glycemic control.

Decreased thermogenic response to an oral glucose load in older subjects.

The thermogenic response to a 100 g oral glucose load was studied by indirect calorimetry in 13 older persons (age range, 38-68 years) and compared with that of 16 young matched controls of similar body weight (age range, 19-30 years). The glucose-induced thermogenesis measured over 180 min and expressed as a per cent of the energy content of the glucose load was found to be reduced in the older subjects, i.e., 5.8 +/- 0.3 per cent vs 8.6 +/- 0.7 per cent, P less than 0.002). This was also accompanied by a significant decrease in the glucose oxidation rate when averaged over the same three-hour period following the glucose load, i.e., 153 mg/min vs 213 mg/min in the control subjects (P less than 0.001) despite a similar time course of glycemia. This study suggests that the thermogenic response to an oral glucose load is blunted in older people, and this may represent an additional factor that contributes to the decreased energy requirement with age and therefore to the increased propensity to obesity if energy intake is not adjusted.

Effects of regular insulin or insulin LISPRO on glucose metabolism after an oral glucose load in patients with type 2 diabetes mellitus.

Seven obese Type 2 diabetic patients were studied for two 4-h periods after ingestion of a glucose load to determine the effects of preprandial subcutaneous injection of Insulin Lispro (5 min before the meal) or regular insulin (20 min before the meal) on glucose metabolism. Glucose production and utilisation were measured using a dual isotope method. After Lispro, the mean postprandial increase in plasma glucose was 29% lower and the increase in insulin concentration 25% higher than after regular insulin (p < 0.05). Suppression of endogenous glucose production was similar with both types of insulin. Thus, preprandial injection of Lispro reduced postprandial glucose increments in Type 2 diabetic patients as compared to regular insulin. This effect is best explained by the increased postprandial bioavailability of Lispro.

Plasma PCSK9 concentrations during an oral fat load and after short term high-fat, high-fat high-protein and high-fructose diets.

BACKGROUND: PCSK9 (Proprotein Convertase Subtilisin Kexin type 9) is a circulating protein that promotes hypercholesterolemia by decreasing hepatic LDL receptor protein. Under non interventional conditions, its expression is driven by sterol response element binding protein 2 (SREBP2) and follows a diurnal rhythm synchronous with cholesterol synthesis. Plasma PCSK9 is associated to LDL-C and to a lesser extent plasma triglycerides and insulin resistance. We aimed to verify the effect on plasma PCSK9 concentrations of dietary interventions that affect these parameters. METHODS: We performed nutritional interventions in young healthy male volunteers and offspring of type 2 diabetic (OffT2D) patients that are more prone to develop insulin resistance, including: i) acute post-prandial hyperlipidemic challenge (n=10), ii) 4 days of high-fat (HF) or high-fat/high-protein (HFHP) (n=10), iii) 7 (HFruc1, n=16) or 6 (HFruc2, n=9) days of hypercaloric high-fructose diets. An acute oral fat load was also performed in two patients bearing the R104C-V114A loss-of-function (LOF) PCSK9 mutation. Plasma PCSK9 concentrations were measured by ELISA. For the HFruc1 study, intrahepatocellular (IHCL) and intramyocellular lipids were measured by 1H magnetic resonance spectroscopy. Hepatic and whole-body insulin sensitivity was assessed with a two-step hyperinsulinemic-euglycemic clamp (0.3 and 1.0 mU.kg-1.min-1). FINDINGS: HF and HFHP short-term diets, as well as an acute hyperlipidemic oral load, did not significantly change PCSK9 concentrations. In addition, post-prandial plasma triglyceride excursion was not altered in two carriers of PCSK9 LOF mutation compared with non carriers. In contrast, hypercaloric 7-day HFruc1 diet increased plasma PCSK9 concentrations by 28% (p=0.05) in healthy volunteers and by 34% (p=0.001) in OffT2D patients. In another independent study, 6-day HFruc2 diet increased plasma PCSK9 levels by 93% (p<0.0001) in young healthy male volunteers. Spearman's correlations revealed that plasma PCSK9 concentrations upon 7-day HFruc1 diet were positively associated with plasma triglycerides (r=0.54, p=0.01) and IHCL (r=0.56, p=0.001), and inversely correlated with hepatic (r=0.54, p=0.014) and whole-body (r=-0.59, p=0.0065) insulin sensitivity. CONCLUSIONS: Plasma PCSK9 concentrations vary minimally in response to a short term high-fat diet and they are not accompanied with changes in cholesterolemia upon high-fructose diet. Short-term high-fructose intake increased plasma PCSK9 levels, independent on cholesterol synthesis, suggesting a regulation independent of SREBP-2. Upon this diet, PCSK9 is associated with insulin resistance, hepatic steatosis and plasma triglycerides.

Inbreeding load, bet hedging, and the evolution of sex-biased dispersal

Inbreeding load affects not only the average fecundity of philopatric individuals but also its variance. From bet-hedging theory, this should add further dispersal pressures to those stemming from the mere avoidance of inbreeding. Pressures on both sexes are identical under monogamy or promiscuity. Under polygyny, by contrast, the variance in reproductive output decreases with dispersal rate in females but increases in males, which should induce a female-biased dispersal. To test this prediction, we performed individual-based simulations. From our results, a female-biased dispersal indeed emerges as both polygyny and inbreeding load increase. We conclude that sex-biased dispersal may be selected for as a bet-hedging strategy.