Hyporesponsiveness to vaccination with Borrelia burgdorferi OspA in humans and in TLR1- and TLR2-deficient mice

The Lyme disease vaccine is based on the outer-surface lipoprotein (OspA) of the pathogen Borrelia burgdorferi, and 95% of vaccine recipients develop substantial titers of antibodies against OspA. Here, we identified seven individuals with very low antibody titers after vaccination (low responders). The macrophages of low responders produced less tumor necrosis factor-alpha and interleukin-6 after OspA stimulation and had lower cell-surface expression of Toll-like receptor (TLR) 1 as compared to normal cells, but normal expression of TLR2. TLRs activate innate responses to pathogens, and TLR2 recognizes lipoproteins and peptidoglycan (PGN). After OspA immunization, mice genetically deficient in either TLR2 (TLR2(-/-)) or TLR1 (TLR1(-/-)) produced low titers of antibodies against OspA. Notably, macrophages from TLR2(-/-) mice were unresponsive to OspA and PGN, whereas those from TLR1(-/-) mice responded normally to PGN but not to OspA. These data indicate that TLR1 and TLR2 are required for lipoprotein recognition and that defects in the TLR1/2 signaling pathway may account for human hyporesponsiveness to OspA vaccination.     

Programmed cell death in 2',3'-dideoxycytidine-resistant human monoblastoid U937 cells

2,3-Dideoxycytidine is a powerful in vitro inhibitor of human immunodeficiency virus and is currently used in the treatment of acquired immunodeficiency syndrome. A long-term exposure of U937 monoblastoid cells to dideoxycytidine induces the selection of drug-resistant cells (U937-R). In previous studies, we investigated some important biochemical properties and functional activities, such as basal respiration, protein kinase C activity, superoxide anion release, and the level of reduced glutathione, which were found to be higher in the drug-resistant cell line, compared to the parental one. In the present study, we evaluated the response of the two cell lines to the induction of apoptosis by treatment with staurosporine and okadaic acid, which interfere with the protein kinase and phosphatase pathways, respectively. Moreover, knowing that GSH plays a crucial role in the regulation of nitric oxide-dependent apoptosis, U937-R and parental lines have been treated with SIN-1, which is known to generate significant amounts of O₂ and nitric oxide.Resistant and parental cells have been analysed by light and electron microscopy and agarose gel electrophoresis of isolated DNA has been performed. The obtained results demonstrate a different susceptibility of U937-R cell line to apoptosis induced with the three triggers. U937-R cells show more advanced apoptotic features if compared with parental cells, after staurosporine treatment. Differently, the okadaic acid does not induce a different behaviour in the two models. On the contrary, the agent SIN-1 determines an increased number of apoptotic cells in the U937 line. The results suggest that a higher level of protein kinase C and glutathione could prevent programmed cell death in U937-R.     

The Turkish delight: a pliable graft for rhinoplasty

In nose surgery, carved or crushed cartilage used as a graft has some disadvantages, chiefly that it may be perceptible through the nasal skin after tissue resolution is complete. To overcome these problems and to obtain a smoother surface, the authors initiated the use of Surgicel-wrapped diced cartilage. This innovative technique has been used by the authors on 2365 patients over the past 10 years: in 165 patients with traumatic nasal deformity, in 350 patients with postrhinoplasty deformity, and in 1850 patients during primary rhinoplasty. The highlights of the surgical procedure include harvested cartilage (septal, alar, conchal, and sometimes costal) cut in pieces of 0.5 to 1 mm using a no. 11 blade. The fine-textured cartilage mass is then wrapped in one layer of Surgicel and moistened with an antibiotic (rifamycin). The graft is then molded into a cylindrical form and inserted under the dorsal nasal skin. In the lateral wall and tip of the nose, some overcorrection is performed depending on the type of deformity. When the mucosal stitching is complete, this graft can be externally molded, like plasticine, under the dorsal skin. In cases of mild-to-moderate nasal depression, septal and conchal cartilages are used in the same manner to augment the nasal dorsum with consistently effective and durable results. In cases with more severe defects of the nose, costal cartilage is necessary to correct both the length of the nose and the projection of the columella. In patients with recurrent deviation of the nasal bridge, this technique provided a simple solution to the problem. After overexcision of the dorsal part of deviated septal cartilage and insertion of Surgicel-wrapped diced cartilage, a straight nose was obtained in all patients with no recurrence (follow-up of 1 to 10 years). The technique also proved to be highly effective in primary rhinoplasties to camouflage bone irregularities after hump removal in patients with thin nasal skin and/or in cases when excessive hump removal was performed. As a complication, in six patients early postoperative swelling was more than usual. In 16 patients, overcorrection was persistent owing to fibrosis, and in 11 patients resorption was excessive beyond the expected amount. A histologic evaluation was possible in 16 patients, 3, 6, and 12 months postoperatively, by removing thin slices of excess cartilage from the dorsum of the nose during touch-up surgery. This graft showed a mosaic-type alignment of graft cartilage with fibrous tissue connection among the fragments. In conclusion, this type of graft is very easy to apply, because a plasticine-like material is obtained that can be molded with the fingers, giving a smooth surface with desirable form and long-lasting results in all cases. The favorable results obtained by this technique have led the authors to use Surgicel-wrapped diced cartilage routinely in all types of rhinoplasty.     

YPED:An Integrated Bioinformatics Suite and Database for Mass Spectrometry-based Proteomics Research

We report a significantly-enhanced bioinformatics suite and database for proteomics research called Yale Protein Expression Database(YPED) that is used by investigators at more than 300 institutions worldwide. YPED meets the data management, archival, and analysis needs of a high-throughput mass spectrometry-based proteomics research ranging from a singlelaboratory, group of laboratories within and beyond an institution, to the entire proteomics community. The current version is a significant improvement over the first version in that it contains new modules for liquid chromatography–tandem mass spectrometry(LC–MS/MS) database search results, label and label-free quantitative proteomic analysis, and several scoring outputs for phosphopeptide site localization. In addition, we have added both peptide and protein comparative analysis tools to enable pairwise analysis of distinct peptides/proteins in each sample and of overlapping peptides/proteins between all samples in multiple datasets. We have also implemented a targeted proteomics module for automated multiple reaction monitoring(MRM)/selective reaction monitoring(SRM) assay development. We have linked YPED's database search results and both label-based and label-free fold-change analysis to the Skyline Panorama repository for online spectra visualization. In addition, we have built enhanced functionality to curate peptide identifications into an MS/MS peptide spectral library for all of our protein database search identification results.     

Histopathological changes induced by maneb and carbaryl on some tissues of rainbow trout, Oncorhynchus mykiss

Acute toxicity of the pesticides, maneb and carbaryl, to juvenile rainbow trout were evaluated under static-renewal test conditions. Actual concentrations of maneb ranged from 0.10 mg/L to 2.00 mg/L and carbaryl ranged from 0.20 mg/L to 3.90 mg/L. The concentrations of maneb that killed 50% of the rainbow trout (3.27 ± 0.9 g) within 24-h (24-h; LC₅₀), 48-h, 72-h and 96-h were 1.19 ± 0.12, 1.04 ± 0.11, 0.92 ± 0.12 and 0.81 ± 0.14 mg/L (95% confidence limits), respectively. LC₅₀ values of carbaryl for 24-h, 48-h, 72-h and 96-h were 2.52 ± 0.71, 2.16 ± 0.63, 1.71 ± 0.46 and 1.39 ± 0.15 mg/L, respectively. None of the unexposed control fish died and the first fish died 6 h after exposure to maneb (≥1.30 mg/L), and carbaryl (≥2.60 mg/L). Lamellar edema, separation of epithelium from lamellae, lamellar fusion, swelling of the epithelial cells and epithelial cell necrosis were observed on maneb and carbaryl exposed fish. Gills also had scattered areas of focal lamellar hyperplasia. Fish exposed to pesticides had inflammation and focal necrosis in liver, trunk kidney and spleen. Maneb and carbaryl had similar histopathological lesions. In order, the most affected organs were gill, trunk kidney and liver.     

Increased glucose oxidation in H-FABP null soleus muscle is associated with defective triacylglycerol accumulation and mobilization, but not with the defect of exogenous fatty acid oxidation

Heart-type fatty acid-binding protein (H-FABP) is a major fatty acid-binding factor in skeletal muscles. Genetic lack of H-FABP severely impairs the esterification and oxidation of exogenous fatty acids in soleus muscles isolated from chow-fed mice (CHOW-solei) and high fat diet-fed mice (HFD-solei), and prevents the HFD-induced accumulation of muscle triacylglycerols (TAGs). Here, we examined the impact of H-FABP deficiency on the relationship between fatty acid utilization and glucose oxidation. Glucose oxidation was measured in isolated soleus muscles in the presence or absence of 1 mM palmitate (simple protocol) or in the absence of fatty acid after preincubation with 1 mM palmitate (complex protocol). With the simple protocol, the mutation slightly reduced glucose oxidation in CHOW-muscles, but markedly increased it in HFD-muscles; unexpectedly, this pattern was not altered by the addition of palmitate, which reduced glucose oxidation in both CHOW- and HFD-solei irrespective of the mutation. In the complex protocol, the mutation first inhibited the synthesis and accumulation of TAGs and then their mobilization; with this protocol, the mutation increased glucose oxidation in both CHOW- and HFD-solei. We conclude: (i) H-FABP mediates a non-acute inhibition of muscle glucose oxidation by fatty acids, likely by enabling both the accumulation and mobilization of a critical mass of muscle TAGs; (ii) H-FABP does not mediate the acute inhibitory effect of extracellular fatty acids on muscle glucose oxidation; (iii) H-FABP affects muscle glucose oxidation in opposing ways, with inhibition prevailing at high muscle TAG contents.     

Anatomical and functional plasticity of pancreatic beta-cells and type 2 diabetes

The most common form of diabetes, type 2 diabetes (T2D) is a major Public Health issue which is receiving a great deal of attention both in industrial and public research, in order to develop new and more effective drugs. The hyperglycaemia of T2D is the result of two interdependent defects : decreased biological efficacy of insulin in target tissues (insulin resistance), and a decreased capacity for beta cells to secrete insulin in response to glucose. Furthermore, hyperglycaemia evolves with time and even with rigorous treatment there is a progressive deterioration of glucose homeostasis. Seventy five percent of DT2 patients are obese and show a perturbed lipid profile. beta-cell plasticity is a unique property of these cells to adapt their number and volume (beta-cell mass) and their function to the increased secretory demand linked to insulin resistance. This is well documented in physiological (pregnancy) as well in pathophysiological conditions (obesity, acromegaly). Although the lack of reliable techniques makes it very difficult to document it in humans, this property is likely altered in DT2, mainly as a consequence of the prolonged exposure of islet cells to high plasma levels of glucose and free fatty acids (gluco-lipotoxicity). The mechanisms by which hyperglycaemia and hyperlipidemia exert their deleterious effects on the beta-cell include the generation of Reactive Oxygen Species (ROS) and Reactive Nitrogen Species (RNS) and Advanced Glycosylation End Products (AGE). Altogether the prevailing clinical and experimental data urge us to consider that the pathophysiology of DT2 lies, at least in part, the inability of beta-cells to adapt their functional mass to the prevailing insulin demand. This re-evaluation of the pathophysiology of DT2 stimulates the research of new therapeutic approaches aimed at maintaining and/or restoring the functional beta-cell mass by targeting the mechanisms responsible for its decrease.