Affected sib-pair analysis of the GLUT1 glucose transporter gene locus in non-insulin-dependent diabetes mellitus (NIDDM): evidence for no linkage

Despite the strong evidence for a major role played by genetic factors in the aetiology of non-insulin-dependent diabetes mellitus (NIDDM), the genes involved are still unknown. Association studies of candidate genes for the inheritance of NIDDM have so far yielded inconclusive results. Some evidence exists for an association between NIDDM and the glucose transporter gene GLUT1, involved in basal glucose transport, although this has not been confirmed. In the present study we have tested the hypothesis of linkage between NIDDM and the GLUT1 gene, using affected sib-pairs. With this method the concordance observed for a given gene marker is compared with that expected under the assumption of no linkage between that marker and the disease. Fifty-four pedigrees (22 Italians and 32 British), for a total of 82 sibpairs were studied by the affected sib-pair method proposed by Weeks and Lange, using two restriction fragment length polymorphisms (RFLPs) at the GLUT1 locus, the MspI RFLP, at an estimated 0.171 recombination frequency from the GLUT1 gene, and the XbaI RFLP, located within the GLUT1 gene and previously shown to be associated with the disease. Results showed that the MspI marker and NIDDM segregate independently; for the XbaI RFLP, linkage could be shown only if the results were weighted by the allele frequency [f(p) = 1/p], and only in the Italian and the combined (Italian and British) sib-pair groups. Multilocus analysis with both markers was also negative. We conclude that the GLUT1 gene is very unlikely to play a major role in the aetiology of NIDDM, although an accessory role cannot be excluded, and studies of the gene sequence should help to clarify this question.     

Lethal and sublethal effects of insecticides on the survival and reproduction of Brevipalpus yothersi (Acari: Tenuipalpidae)

The overuse of insecticides to control vector insects such as Diaphorina citri Kuwayama in citrus groves has altered the population dynamics of pest mites. Among phytophagous mites, population outbreaks of citrus leprosis mite, Brevipalpus yothersi Baker, have been increasingly intense and frequent in Brazilian citrus groves. Despite the great importance of the B. yothersi mite for citrus production, the lethal and sublethal effects of insecticides on this mite have not yet been studied. Therefore, in this study, the effects of insecticides commonly used for D. citri control on B. yothersi mortality, reproduction, and instantaneous growth rate were assessed. For this, two experiments were carried out, one under controlled conditions and another in a greenhouse. The insecticides tested were beta-cyfluthrin, bifenthrin, buprofezin, chlorpyrifos, dimethoate, pyriproxyfen, and thiamethoxam at 0 (control), 0.0625, 0.125, 0.25, 0.5, 1, and twofold the recommended insecticide concentration for D. citri control. The pyriproxyfen insecticide provided high mortality of B. yothersi even at low concentrations. Furthermore, this insecticide negatively interfered with the reproduction of this mite. Beta-cyfluthrin, bifenthrin, buprofezin, chlorpyrifos, dimethoate, and thiamethoxam, in the tested concentrations, showed low impact on citrus leprosis mite. Regarding the reproduction of the mite, no significant increase in fecundity was observed on B. yothersi females exposed to insecticide residues, regardless of the concentration tested. Therefore, the application of these insecticides in the management of pest insects is unlikely to promote an increase in the citrus leprosis mite population.

     

Dynamic changes in microbiota and mycobiota during spontaneous ‘Vino Santo Trentino’ fermentation

Vino Santo is a sweet wine produced from late harvesting and pressing of Nosiola grapes in a small, well‐defined geographical area in the Italian Alps. We used metagenomics to characterize the dynamics of microbial communities in the products of three wineries, resulting from spontaneous fermentation with almost the same timing and procedure. Comparing fermentation dynamics and grape microbial composition, we show a rapid increase in a small number of wine yeast species, with a parallel decrease in complexity. Despite the application of similar protocols, slight changes in the procedures led to significant differences in the microbiota in the three cases of fermentation: (i) fungal content of the must varied significantly in the different wineries, (ii) Pichia membranifaciens persisted in only one of the wineries, (iii) one fermentation was characterized by the balanced presence of Saccharomyces cerevisiae and Hanseniaspora osmophila during the later phases. We suggest the existence of a highly winery‐specific ‘microbial‐terroir’ contributing significantly to the final product rather than a regional ‘terroir’. Analysis of changes in abundance during fermentation showed evident correlations between different species, suggesting that fermentation is the result of a continuum of interaction between different species and physical–chemical parameters.     

Hypoxia-induced acute lung injury in murine models of sickle cell disease

Vaso-occlusive events are the major source of morbidity and mortality in sickle cell disease (SCD); however, the pathogenic mechanisms driving these events remain unclear. Using hypoxia to induce pulmonary injury, we investigated mechanisms by which sickle hemoglobin increases susceptibility to lung injury in a murine model of SCD, where mice either exclusively express the human alpha/sickle beta-globin (halphabetaS) transgene (SCD mice) or are heterozygous for the normal murine beta-globin gene and express the halphabetaS transgene (mbeta+/-, halphabetaS+/-; heterozygote SCD mice). Under normoxia, lungs from the SCD mice contained higher levels of xanthine oxidase (XO), nitrotyrosine, and cGMP than controls (C57BL/6 mice). Hypoxia increased XO and nitrotyrosine and decreased cGMP content in the lungs of all mice. After hypoxia, vascular congestion was increased in lungs with a greater content of XO and nitrotyrosine. Under normoxia, the association of heat shock protein 90 (HSP90) with endothelial nitric oxide synthase (eNOS) in lungs of SCD and heterozygote SCD mice was decreased compared with the levels of association in lungs of controls. Hypoxia further decreased association of HSP90 with eNOS in lungs of SCD and heterozygote SCD mice, but not in the control lungs. Pretreatment of rat pulmonary microvascular endothelial cells in vitro with xanthine/XO decreased A-23187-stimulated nitrite + nitrate production and HSP90 interactions with eNOS. These data support the hypotheses that hypoxia increases XO release from ischemic tissues and that the local increase in XO-induced oxidative stress can then inhibit HSP90 interactions with eNOS, decreasing *NO generation and predisposing the lung to vaso-occlusion.     

The spatial distribution of bacteria in Grana-cheese during ripening

The microbial composition and its spatial distribution of Grana Trentino, a hard Parmesan-like cheese, was determined, from vat milk to cheese. After cutting along the vertical axis of the cheese wheels, three layers were sampled diagonally across the cheese: under the cheese rind, an intermediate section and the cheese core. After two different ripening periods (9 and 18 months), the cheese samples were analysed using traditional culture dependent and culture independent methods. Milk samples were dominated by mesophilic and psychrophilic bacterial counts. Thermophilic bacteria (Lactobacillus helveticus) were found in high amounts in cooked whey and natural whey starter cultures. After 9 months of ripening, lactic acid bacteria (LAB) counts were higher than those after 18 months. Furthermore, the LAB numbers in the cheese core was lower than those under the rind or in the intermediate section. The main LAB species isolated from milk (Lactococcus lactis, Pediococcus pentosaceus, Streptococcus uberis and Lactococcus garvieae) were not found in the corresponding cheeses. Some differences were observed in the species composition among the three cheese sections. Microbiota under the rind and in the intermediate section was similar and dominated by Lactobacillus paracasei and Lactobacillus rhamnosus. The core, after 18 months of ripening, was characterized by a total absence of LAB. In each sample, all LAB were genotypically grouped and the different biotypes were subjected to several technological tests indicating that some non-starter LAB (NSLAB) displayed technological features that are favorable for the production of Grana Trentino cheese.     

Effects on breathing of carotid body denervation in neonatal piglets.

The purpose of these studies was to test the hypothesis that carotid chemoreceptor activity is necessary for postnatal maturation of the ventilatory control system. By using a lateral surgical access, 17 piglets were carotid body denervated (CBD) and 14 were sham denervated at 3-25 days of age. After surgery, there was no irregular breathing in any group. There was no significant hypoventilation when CBD was performed at less than 5 days of age (n = 5) and only a mild (arterial PCO(2) 5 Torr; P < 0.05) to moderate, transient (arterial PCO(2) 8 Torr; P < 0.5) hypoventilation in piglets denervated at 10-15 (n = 6) and 20-25 (n = 6) days of age, respectively. Three weeks after surgery, both breathing of a hypoxic gas mixture and jugular venous NaCN injections elicited a hyperpnea in the CBD piglets that was attenuated compared with that in sham CBD piglets. In the CBD piglets, there was no response to injections of NaCN in the carotid arteries, but there was a response to NaCN injected into the proximal descending aorta, suggesting the residual peripheral chemosensitivity was of aortic origin. Carotid chemoreceptor-intact piglets had carotid and aortic NaCN chemosensitivity by 2 days of age. The carotid response persisted for the 40 days of the study, but the aortic reflex persisted only until approximately 8 days of age. We conclude that 1) the major effect of CBD per se in neonatal piglets is age-dependent hypoventilation and 2) there is a high degree of plasticity in peripheral chemosensitivity in neonates that may contribute to minimizing the changes in breathing after CBD.     

Acipimox reduces circulating levels of insulin and associated neutrophilic inflammation in metabolic syndrome

Metabolic syndrome is a proatherosclerotic condition clustering cardiovascular risk factors, including glucose and lipid profile alterations. The pathophysiological mechanisms favoring atherosclerotic inflammation in the metabolic syndrome remain elusive. Here, we investigated the potential role of the antilipolytic drug acipimox on neutrophil- and monocyte-mediated inflammation in the metabolic syndrome. Acipimox (500 mg) was orally administered to metabolic syndrome patients (n = 11) or healthy controls (n = 8). Serum and plasma was collected before acipimox administration (time 0) as well as 2-5 h afterward to assess metabolic and hematologic parameters. In vitro, the effects of the incubation with metabolic syndrome serum were assessed on human neutrophil and monocyte migration toward the proatherosclerotic chemokine CCL3. Two to five hours after acipimox administration, a significant reduction in circulating levels of insulin and nonesterified fatty acid (NEFA) was shown in metabolic syndrome patients. At time 0 and 2 h after acipimox administration, metabolic syndrome serum increased neutrophil migration to CCL3 compared with healthy controls. No effect was shown in human monocytes. At these time points, serum-induced neutrophil migration positively correlated with serum levels of insulin and NEFA. Metabolic syndrome serum or recombinant insulin did not upregulate CCR5 expression on neutrophil surface membrane, but it increased intracellular JNK1/2 phosphorylation. Insulin immunodepletion blocked serum-induced neutrophil migration and associated JNK1/2 phosphorylation. Although mRNA expression of acipimox receptor (GPR109) was shown in human neutrophils, 5-500 μM acipimox did not affect insulin-induced neutrophil migration. In conclusion, results suggest that acipimox inhibited neutrophil proatherosclerotic functions in the metabolic syndrome through the reduction in circulating levels of insulin.