Impairment of organ-specific T cell negative selection by diabetes susceptibility genes: genomic analysis by mRNA profiling

BACKGROUND T cells in the thymus undergo opposing positive and negative selection processes so that the only T cells entering circulation are those bearing a T cell receptor (TCR) with a low affinity for self. The mechanism differentiating negative from positive selection is poorly understood, despite the fact that inherited defects in negative selection underlie organ-specific autoimmune disease in AIRE-deficient people and the non-obese diabetic (NOD) mouse strain RESULTS Here we use homogeneous populations of T cells undergoing either positive or negative selection in vivo together with genome-wide transcription profiling on microarrays to identify the gene expression differences underlying negative selection to an Aire-dependent organ-specific antigen, including the upregulation of a genomic cluster in the cytogenetic band 2F. Analysis of defective negative selection in the autoimmune-prone NOD strain demonstrates a global impairment in the induction of the negative selection response gene set, but little difference in positive selection response genes. Combining expression differences with genetic linkage data, we identify differentially expressed candidate genes, including Bim, Bnip3, Smox, Pdrg1, Id1, Pdcd1, Ly6c, Pdia3, Trim30 and Trim12. CONCLUSION The data provide a molecular map of the negative selection response in vivo and, by analysis of deviations from this pathway in the autoimmune susceptible NOD strain, suggest that susceptibility arises from small expression differences in genes acting at multiple points in the pathway between the TCR and cell death.This work was supported by grants from the NHMRC and the Juvenile Diabetes Research Foundation

Adsorption models of hybridization and post-hybridisation behaviour on oligonucleotide microarrays

Analysis of data from an Affymetrix Latin Square spike-in experiment indicates that measured fluorescence intensities of features on an oligonucleotide microarray are related to spike-in RNA target concentrations via a hyperbolic response function, generally identified as a Langmuir adsorption isotherm. Furthermore the asymptotic signal at high spike-in concentrations is almost invariably lower for a mismatch feature than for its partner perfect match feature. We survey a number of theoretical adsorption models of hybridization at the microarray surface and find that in general they are unable to explain the differing saturation responses of perfect and mismatch features. On the other hand, we find that a simple and consistent explanation can be found in a model in which equilibrium hybridization followed by partial dissociation of duplexes during the post-hybridization washing phase.Comment: 26 pages, 6 figures, some rearrangement of sections and some additions. To appear in J.Phys.(condensed matter

Effective affinities in microarray data

In the past couple of years several studies have shown that hybridization in Affymetrix DNA microarrays can be rather well understood on the basis of simple models of physical chemistry. In the majority of the cases a Langmuir isotherm was used to fit experimental data. Although there is a general consensus about this approach, some discrepancies between different studies are evident. For instance, some authors have fitted the hybridization affinities from the microarray fluorescent intensities, while others used affinities obtained from melting experiments in solution. The former approach yields fitted affinities that at first sight are only partially consistent with solution values. In this paper we show that this discrepancy exists only superficially: a sufficiently complete model provides effective affinities which are fully consistent with those fitted to experimental data. This link provides new insight on the relevant processes underlying the functioning of DNA microarrays.Comment: 8 pages, 6 figure

Modelling the human epidermis in vitro: tools for basic and applied research

Culture models of tissues and organs are valuable tools developed by basic research that help investigation of the body functions. Modelling is aimed at simplifying experimental procedures in order to better understand biological phenomena, and consequently, when sufficiently characterized, culture models can also be utilized with high potential in applied research. In skin biology and pathology, the development of cultures of keratinocytes as monolayers has allowed the elucidation of most functional and structural characteristics of the cell type. Beside the multiple great successes that have been obtained with this type of culture, this review draws attention on several neglected characteristics of monolayer cultures. The more sophisticated models created in order to reconstruct the fully differentiated epidermis have followed the monolayers. The epidermal reconstruction produces all typical layers found in vivo and thus makes the model much less simple, but only this kind of model allows the study of full differentiation in keratinocyte and production of the cornified barrier. In addition to its interest in basic research, the reconstructed epidermis is currently gaining a lot of interest for applied research, particularly as an alternative to laboratory animals in the chemical and cosmetic industry. Today several commercial providers propose reconstructed skin or epidermis, but in vitro assays on these materials are still under development. In order to be beneficial at long term, the validation of assays must be performed on a material whose availability will not be interrupted. We warn here providers and customers that the longevity of in vitro assays will be guaranteed only if these assays are done with well-described models, prepared according to published procedures, and must consider having a minimum of two independent simultaneous producers of similar material

Evaluation of an Extended-duration Chemoprophylaxis Regimen for Venous Thromboembolism after Microsurgical Breast Reconstruction

Patients undergoing free flap breast reconstruction are at a high risk for venous thromboembolism based upon Caprini scores. Guidelines for venous thromboembolism prophylaxis recommend high-risk groups receive extended chemoprophylaxis for several weeks after gynecological, orthopedic, and surgical oncology cases. Extended prophylaxis has not been studied in free flap breast reconstruction. The purpose of this study was to compare outcomes of free flap breast reconstruction patients who received extended venous thromboembolism (VTE) prophylaxis with those who received standard inpatient-only prophylaxis. Methods: Patients undergoing microsurgical breast reconstruction were divided into two groups: standard VTE prophylaxis (Group I) and extended prophylaxis (Group II). Both groups received prophylactic subcutaneous heparin or enoxaparin preoperatively and enoxaparin 40 mg daily postoperatively while inpatient. Group II was discharged with a home regimen of enoxaparin 40 mg daily for an additional 14 days. Results: In total, 103 patients met inclusion criteria (36 patients in Group I, 67 patients in Group II). The incidence of VTE was 1.5% in Group II compared with 2.8% in Group I (P = 0.6). There was no difference in reoperative hematoma between Group I (n = 0) and Group II (n = 1) (P = 0.7). Total flap loss was 2.2%. Conclusions: Although this retrospective pilot study did not show statistical significance in VTE between those receiving extended home chemoprophylaxis (1.5% incidence) compared with inpatient-only chemoprophylaxis (2.8%), the risk of bleeding complications was similar. These results indicate that a larger, higher powered study is justified to assess if an extended home chemoprophylaxis protocol should be standard of care post free flap breast reconstruction

Eficiência de inseticidas para o controle da mosca-branca Bemisia tabaci biótipo B (Hemiptera: Aleyrodidae) em soja nas safras 2017/2018 e 2018/2019: resultados sumarizados dos ensaios cooperativos.

bitstream/item/212145/1/CT-158.pd

The adaptive capacity of maize-based conservation agriculture systems to climate stress in tropical and subtropical environments: A meta-regression of yields

Conservation agriculture is widely promoted across sub-Saharan Africa as a sustainable farming practice that enhances adaptive capacity to climate change. The interactions between climate stress, management, and soil are critical to understanding the adaptive capacity of conservation agriculture. Yet conservation agriculture syntheses to date have largely neglected climate, especially the effects of extreme heat. For the sub-tropics and tropics, we use meta-regression, in combination with global soil and climate datasets, to test four hypotheses: (1) that relative yield performance of conservation agriculture improves with increasing drought and temperature stress; (2) that the effects of moisture and temperature stress exposure interact; (3) that the effects of moisture and temperature stress are modified by soil texture; and (4) that crop diversification, fertilizer application rate, or the time since no-till implementation will enhance conservation agriculture performance under climate stress. Our results support the hypothesis that the relative maize yield performance of conservation agriculture improves with increasing drought severity or exposure to high temperatures. Further, there is an interaction of moisture and heat stress on conservation agriculture performance and their combined effect is both non-additive and modified by soil clay content, supporting our second and third hypotheses. Finally, we found only limited support for our fourth hypothesis as (1) increasing nitrogen application rates did not improve the relative performance of conservation agriculture under high heat stress; (2) crop diversification did not notably improve conservation agriculture performance, but did increase its stability with heat stress; and (3) a statistically robust effect of the time since no-till implementation was not evident. Our meta-regression supports the narrative that conservation agriculture enhances the adaptive capacity of maize production in sub-Saharan Africa under drought and/or heat stress. However, in very wet seasons and on clay-rich soils, conservation agriculture yields less compared to conventional practices

Toward allele-specific targeting therapy and pharmacodynamic marker for spinocerebellar ataxia type 3

Spinocerebellar ataxia type 3 (SCA3), caused by a CAG repeat expansion in the ataxin-3 gene (ATXN3), is characterized by neuronal polyglutamine (polyQ) ATXN3 protein aggregates. Although there is no cure for SCA3, gene-silencing approaches to reduce toxic polyQ ATXN3 showed promise in preclinical models. However, a major limitation in translating putative treatments for this rare disease to the clinic is the lack of pharmacodynamic markers for use in clinical trials. Here, we developed an immunoassay that readily detects polyQ ATXN3 proteins in human biological fluids and discriminates patients with SCA3 from healthy controls and individuals with other ataxias. We show that polyQ ATXN3 serves as a marker of target engagement in human fibroblasts, which may bode well for its use in clinical trials. Last, we identified a single-nucleotide polymorphism that strongly associates with the expanded allele, thus providing an exciting drug target to abrogate detrimental events initiated by mutant ATXN3. Gene-silencing strategies for several repeat diseases are well under way, and our results are expected to improve clinical trial preparedness for SCA3 therapies