Identification and characterization of permissive cells to dengue virus infection in human hematopoietic stem and progenitor cells.

BACKGROUND: Dengue virus (DENV) is a significant threat to public health in tropical and subtropical regions, where the frequency of human migration is increasing. Transmission of DENV from donors to recipients after hematopoietic stem cell transplantation has been steadily described. However, the underlying mechanisms remain unclear. STUDY DESIGN AND METHODS: Freshly isolated bone marrow (BM) was subjected to DENV infection, followed by multicolor fluorescence-activated cell sorting (FACS) analysis. Virus in supernatants was collected and analyzed by plaque assay. RESULTS: DENV-1 to DENV-4 could effectively infect freshly obtained BM and produced infectious virus. DENV infection did not change the quantitative population of hematopoietic stem and progenitor cells (HSPCs), megakaryocytic progenitor cells (MkPs) and megakaryocytes. Additionally, DENV antigen, nonstructural protein 1, was enriched in HSPCs and MkPs of DENV infected marrow cells. CD34+, CD133+, or CD61+ cells sorted out from BM were not only the major contributing targets facilitating the DENV infection directly but also facilitated the spread of DENV into other cells when cocultured. CONCLUSION: Results suggest that DENV can efficiently infect HSPCs, which might jeopardize the recipients if DENV-infected cells were subsequently used. We therefore raise the need for DENV screening for both the donors and recipients of hematopoietic stem cell transplantation, especially for donors exposed to endemic areas, to mitigate DENV infection in immunocompromised recipients.

Pharmacophore anchor models of flaviviral NS3 proteases lead to drug repurposing for DENV infection.

BACKGROUND: Viruses of the flaviviridae family are responsible for some of the major infectious viral diseases around the world and there is an urgent need for drug development for these diseases. Most of the virtual screening methods in flaviviral drug discovery suffer from a low hit rate, strain-specific efficacy differences, and susceptibility to resistance. It is because they often fail to capture the key pharmacological features of the target active site critical for protein function inhibition. So in our current work, for the flaviviral NS3 protease, we summarized the pharmacophore features at the protease active site as anchors (subsite-moiety interactions). RESULTS: For each of the four flaviviral NS3 proteases (i.e., HCV, DENV, WNV, and JEV), the anchors were obtained and summarized into 'Pharmacophore anchor (PA) models'. To capture the conserved pharmacophore anchors across these proteases, were merged the four PA models. We identified five consensus core anchors (CEH1, CH3, CH7, CV1, CV3) in all PA models, represented as the "Core pharmacophore anchor (CPA) model" and also identified specific anchors unique to the PA models. Our PA/CPA models complied with 89 known NS3 protease inhibitors. Furthermore, we proposed an integrated anchor-based screening method using the anchors from our models for discovering inhibitors. This method was applied on the DENV NS3 protease to screen FDA drugs discovering boceprevir, telaprevir and asunaprevir as promising anti-DENV candidates. Experimental testing against DV2-NGC virus by in-vitro plaque assays showed that asunaprevir and telaprevir inhibited viral replication with EC50 values of 10.4 µM & 24.5 µM respectively. The structure-anchor-activity relationships (SAAR) showed that our PA/CPA model anchors explained the observed in-vitro activities of the candidates. Also, we observed that the CEH1 anchor engagement was critical for the activities of telaprevir and asunaprevir while the extent of inhibitor anchor occupation guided their efficacies. CONCLUSION: These results validate our NS3 protease PA/CPA models, anchors and the integrated anchor-based screening method to be useful in inhibitor discovery and lead optimization, thus accelerating flaviviral drug discovery.

Defective determination of synthetic cathinones in blood for forensic investigation.

Antemortem specimens are sometimes the sole sources available for forensic investigation, and samples collected in nonideal ways are inevitably employed to achieve toxicological analysis. It is essential to assess the effects of blood collection tubes on the recoveries of emerging synthetic cathinones (SC) to estimate actual drug concentrations, and no such systematic investigations have been previously carried out. Seventy-one SC with various LogP values were employed to examine commonly used blood collection tubes, including plasma tubes, serum tubes and gel-containing tubes in recoveries which determined by a reliable LC-MS/MS method. Significantly poor recoveries for hydrophobic SC were obtained using serum separating tubes (SST). Notably, the suppressed recoveries in SST can be reversed by adding anticoagulants. Adding a procoagulant to a plasma separating tube (PST) considerably reduced recoveries, which indicated that clotting processes in the presence of polymeric gels contributed to poor recoveries of these hydrophobic drugs. In this study, we find that clotting formation in the presence of polymeric gels could significantly affect the determination of hydrophobic drugs. However, in real-world scenarios, nonideal collection methods are inevitably employed for antemortem specimens. Thus, it is important to rigorously interpret forensic toxicological results, especially for susceptible species.

Nutritional geometry framework of sleep.

AIMS: Sleep is a fundamental physiological function and is essential for all animals. Sleep is affected by diet compositions including protein (P) and carbohydrates (C), but there has not been a systematic investigation on the effect of dietary macronutrient balance on sleep. MAIN METHODS: We used the nutritional geometry framework (NGF) to explore the interactive effects on sleep of protein (P) and carbohydrates (C) in the model organism Drosophila. Both female and male flies were fed various diets containing seven ratios of protein-to-carbohydrates at different energetic levels for 5 days and sleep was monitored by the Drosophila Activity Monitor (DAM) system. KEY FINDINGS: Our results showed that the combination of low protein and high carbohydrates (LPHC) prolonged sleep time and sleep quality, with fewer sleep episodes and longer sleep duration. We further found that the effects of macronutrients on sleep mirrored levels of hemolymph glucose and whole-body glycogen. Moreover, transcriptomic analyses revealed that a high-protein, low-carbohydrate (HPLC) diet significantly elevated the gene expression of metabolic pathways when compared to the LPHC diet, with the glycine, serine, and threonine metabolism pathway being most strongly elevated. Further studies confirmed that the contents of glycine, serine, and threonine affected sleep. SIGNIFICANCE: Our results demonstrate that sleep is affected by the dietary balance of protein and carbohydrates possibly mediated by the change in glucose, glycogen, glycine, serine, and threonine.

An automated workflow on data processing (AutoDP) for semiquantitative analysis of urine organic acids with GC-MS to facilitate diagnosis of inborn errors of metabolism.

Determination of urine organic acids (UOAs) is essential to understand the disease progress of inborn errors of metabolism (IEM) and often relies on GC-MS analysis. However, the efficiency of analytical reports is sometimes restricted by data processing due to labor-intensive work if no proper tool is employed. Herein, we present a simple and rapid workflow with an R-based script for automated data processing (AutoDP) of GC-MS raw files to quantitatively analyze essential UOAs. AutoDP features automatic quality checks, compound identification and confirmation with specific fragment ions, retention time correction from analytical batches, and visualization of abnormal UOAs with age-matched references on chromatograms. Compared with manual processing, AutoDP greatly reduces analytical time and increases the number of identifications. Speeding up data processing is expected to shorten the waiting time for clinical diagnosis, which could greatly benefit clinicians and patients with IEM. In addition, with quantitative results obtained from AutoDP, it would be more feasible to perform retrospective analysis of specific UOAs in IEM and could provide new perspectives for studying IEM.

PGC1α Controls Sucrose Taste Sensitization in Drosophila.

Perceived palatability of food controls caloric intake. Sweet taste is the primary means of detecting the carbohydrate content of food. Surprisingly, sweet taste sensitivity is responsive to extrinsic factors like diet, and this occurs by unknown mechanisms. Here, we describe an unbiased proteomic investigation into sweet taste sensitivity in the fruit fly. We identify a dopamine/cyclic AMP (cAMP)/CREB axis acting within sweet taste neurons that controls taste perception but is largely dispensable for acute taste transduction. This pathway modulates sweet taste perception in response to both sensory- and nutrient-restricted diets and converges on PGC1α, a critical regulator of metabolic health and lifespan. By electrophysiology, we found that enhanced sucrose taste sensitivity was the result of heightened sweet taste intensity and that PGC1α was both necessary and sufficient for this effect. Together, we provide the first molecular insight into how diet-induced taste perception is regulated within the sweet taste neuron.