Isothermal Amplification Using a Chemical Heating Device for Point-of-Care Detection of HIV-1

Background: To date, the use of traditional nucleic acid amplification tests (NAAT) for detection of HIV-1 DNA or RNA has been restricted to laboratory settings due to time, equipment, and technical expertise requirements. The availability of a rapid NAAT with applicability for resource-limited or point-of-care (POC) settings would fill a great need in HIV diagnostics, allowing for timely diagnosis or confirmation of infection status, as well as facilitating the diagnosis of acute infection, screening and evaluation of infants born to HIV-infected mothers. Isothermal amplification methods, such as reversetranscription, loop-mediated isothermal amplification (RT-LAMP), exhibit characteristics that are ideal for POC settings, since they are typically quicker, easier to perform, and allow for integration into low-tech, portable heating devices. Methodology/Significant Findings: In this study, we evaluated the HIV-1 RT-LAMP assay using portable, non-instrumented nucleic acid amplification (NINA) heating devices that generate heat from the exothermic reaction of calcium oxide and water. The NINA heating devices exhibited stable temperatures throughout the amplification reaction and consistent amplification results between three separate devices and a thermalcycler. The performance of the NINA heaters was validated using whole blood specimens from HIV-1 infected patients. Conclusion: The RT-LAMP isothermal amplification method used in conjunction with a chemical heating device provides

Elevated apoptosis impairs epithelial cell turnover and shortens villi in TNF-driven intestinal inflammation

The intestinal epithelial monolayer, at the boundary between microbes and the host immune system, plays an important role in the development of inflammatory bowel disease (IBD), particularly as a target and producer of pro-inflammatory TNF. Chronic overexpression of TNF leads to IBD-like pathology over time, but the mechanisms driving early pathogenesis events are not clear. We studied the epithelial response to inflammation by combining mathematical models with in vivo experimental models resembling acute and chronic TNF-mediated injury. We found significant villus atrophy with increased epithelial cell death along the crypt-villus axis, most dramatically at the villus tips, in both acute and chronic inflammation. In the acute model, we observed overexpression of TNF receptor I in the villus tip rapidly after TNF injection and concurrent with elevated levels of intracellular TNF and rapid shedding at the tip. In the chronic model, sustained villus atrophy was accompanied by a reduction in absolute epithelial cell turnover. Mathematical modelling demonstrated that increased cell apoptosis on the villus body explains the reduction in epithelial cell turnover along the crypt-villus axis observed in chronic inflammation. Cell destruction in the villus was not accompanied by changes in proliferative cell number or division rate within the crypt. Epithelial morphology and immunological changes in the chronic setting suggest a repair response to cell damage although the villus length is not recovered. A better understanding of how this state is further destabilised and results in clinical pathology resembling IBD will help identify suitable pathways for therapeutic intervention

Candidate HIV-1 Tat vaccine development: from basic science to clinical trials.

Abstract not availabl

Genetic and phylogenetic analyses of HIV-1 corroborate the transmission link hypothesis

Background: Phylogenetic and genetic analyses have proven a valuable tool to infer epidemiological links between human immunodeficiency virus type-1 (HIV-1) isolates. These methods were applied in the present report for studying the genetic relatedness of the viral strains involved in two episodes of suspected HIV-1 transmission. Objectives: Provide any evidence that may help establish or refute the transmission link. Study design: In the first case, a leukemic patient became HIV-1 positive following the transfusion of platelets from a donor who was subsequently found to have tested false HIV-seronegative and to be sexual partner to an infected woman. In the second, a wife claimed to have acquired the infection from her husband who had concealed his infected status. Results and conclusions: The viral pairs detected in each of the suspected transmission cases exhibited common amino acid signatures and low genetic distances and segregated together in phylogenetic trees, thus showing a level of genetic relatedness similar to reference pairs known with certainty to be epidemiologically linked. These findings corroborated the existence of a direct transmission link in both the episodes with a high level of confidence. (C) 2003 Elsevier B.V. All rights reserved

Tailor-made core-shell nanospheres for antisense oligonucleotide delivery: IV. Adsorption/release behaviour

The adsorption/release behaviour of oligodeoxynucleotides (ODNs) on double functional core-shell polymethylmethacrylate nanospheres, with a narrow size distribution, is described. The outer shell consists of alkyl or glycolic chains containing permanently-charged quaternary ammonium groups. Ion pair formation between negatively-charged ODN phosphate groups and positively-charged groups, present on the nanosphere surface, is the main mechanism of interaction. The amount of adsorbed ODN depends on both the ODN concentration and the nanosphere surface charge density. An adsorption-induced swelling mechanism is proposed in which a modification of the charged diffuse layer around the nanospheres increases the ODN binding site accessibility with increasing ODN concentration. Adsorption on the nanosphere surface prevents serum degradation of the ODNs. ODN release is negligible in the presence of culture medium but occurs gradually in the presence of serum. No significant cytotoxicity of the free nanoparticles was found in PBMC and CEM cells after 24 h at ODN concentrations required for antisense activity