Reprogramming of HUVECs into Induced Pluripotent Stem Cells (HiPSCs), Generation and Characterization of HiPSC-Derived Neurons and Astrocytes

Neurodegenerative diseases are characterized by chronic and progressive structural or functional loss of neurons. Limitations related to the animal models of these human diseases have impeded the development of effective drugs. This emphasizes the need to establish disease models using human-derived cells. The discovery of induced pluripotent stem cell (iPSC) technology has provided novel opportunities in disease modeling, drug development, screening, and the potential for “patient-matched” cellular therapies in neurodegenerative diseases. In this study, with the objective of establishing reliable tools to study neurodegenerative diseases, we reprogrammed human umbilical vein endothelial cells (HUVECs) into iPSCs (HiPSCs). Using a novel and direct approach, HiPSCs were differentiated into cells of central nervous system (CNS) lineage, including neuronal, astrocyte and glial cells, with high efficiency. HiPSCs expressed embryonic genes such as nanog, sox2 and Oct-3/4, and formed embryoid bodies that expressed markers of the 3 germ layers. Expression of endothelial-specific genes was not detected in HiPSCs at RNA or protein levels. HiPSC-derived neurons possess similar morphology but significantly longer neurites compared to primary human fetal neurons. These stem cell-derived neurons are susceptible to inflammatory cell-mediated neuronal injury. HiPSC-derived neurons express various amino acids that are important for normal function in the CNS. They have functional receptors for a variety of neurotransmitters such as glutamate and acetylcholine. HiPSC-derived astrocytes respond to ATP and acetylcholine by elevating cytosolic Ca²⁺ concentrations. In summary, this study presents a novel technique to generate differentiated and functional HiPSC-derived neurons and astrocytes. These cells are appropriate tools for studying the development of the nervous system, the pathophysiology of various neurodegenerative diseases and the development of potential drugs for their treatments.     

A Cross-Sectional Study of ‘Yaws’ in Districts of Ghana Which Have Previously Undertaken Azithromycin Mass Drug Administration for Trachoma Control

Yaws, caused by Treponema pallidum ssp. pertenue, is reportedly endemic in Ghana. Mass distribution of azithromycin is now the cornerstone of the WHO yaws eradication campaign. Mass distribution of azithromycin at a lower target dose was previously undertaken in two regions of Ghana for the control of trachoma. Ongoing reporting of yaws raises the possibility that resistance may have emerged in T. pallidum pertenue, or that alternative infections may be responsible for some of the reported cases. We conducted a cross-sectional survey in thirty communities in two districts of Ghana where MDA for trachoma had previously been conducted. Children aged 5–17 years with ulcerative lesions compatible with yaws were enrolled. Samples for treponemal serology and lesion PCR were collected from all children. 90 children with 98 lesions were enrolled. Syphilis serology was negative in all of them. PCR for T. pallidum ssp pertenue was negative in all children, but Haemophilus ducreyi DNA was detected in 9 lesions. In these communities, previously treated for trachoma, we found no evidence of ongoing transmission of yaws. H. ducreyi was associated with a proportion of skin lesions, but the majority of lesions remain unexplained. Integration of diagnostic testing into both pre and post-MDA surveillance systems is required to better inform yaws control programmes.     

PrEP uptake,persistence, adherence,and effect of retrospective drug level feedback on PrEP adherence among young women in southern Africa: Results from HPTN 082, a randomized controlled trial

BackgroundPre-exposure prophylaxis (PrEP) is highly effective and an important prevention tool for African adolescent girls and young women (AGYW), but adherence and persistence are challenging. PrEP adherence support strategies for African AGYW were studied in an implementation study.Methods and findingsHIV Prevention Trials Network (HPTN) 082 was conducted in Cape Town, Johannesburg (South Africa) and Harare (Zimbabwe) from October 2016 to October 2018 to evaluate PrEP uptake, persistence, and the effect of drug level feedback on adherence. Sexually active HIV–negative women ages 16–25 were offered PrEP and followed for 12 months; women who accepted PrEP were randomized to standard adherence support (counseling, 2-way SMS, and adherence clubs) or enhanced adherence support with adherence feedback from intracellular tenofovir-diphosphate (TFV-DP) levels in dried blood spots (DBS). PrEP uptake, persistence through 12 months (no PrEP hold or missed visits), and adherence were assessed. The primary outcome was high adherence (TFV-DP ≥700 fmol/punch) at 6 months, compared by study arm. Of 451 women enrolled, median age was 21 years, and 39% had curable sexually transmitted infections (STIs). Most (95%) started PrEP, of whom 55% had uninterrupted PrEP refills through 12 months. Of those with DBS, 84% had detectable TFV-DP levels at month 3, 57% at month 6, and 31% at month 12. At 6 months, 36/179 (21%) of AGYW in the enhanced arm had high adherence and 40/184 (22%) in the standard adherence support arm (adjusted odds ratio [OR] of 0.92; 95% confidence interval [CI] 0.55, 1.34; p = 0.76). Four women acquired HIV (incidence 1.0/100 person-years), with low or undetectable TFV-DP levels at or prior to seroconversion, and none of whom had tenofovir or emtricitabine resistance mutations. The study had limited power to detect a modest effect of drug level feedback on adherence, and there was limited awareness of PrEP at the time the study was conducted.ConclusionsIn this study, PrEP initiation was high, over half of study participants persisted with PrEP through month 12, and the majority of young African women had detectable TFV-DP levels through month 6 with one-fifth having high adherence. Drug level feedback in the first 3 months of PrEP use did not increase the proportion with high adherence at month 6. HIV incidence was 1% in this cohort with 39% prevalence of curable STIs and moderate PrEP adherence. Strategies to support PrEP use and less adherence-dependent formulations are needed for this population.Trial registrationClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT02732730","term_id":"NCT02732730"}}NCT02732730.

Connie Celum and co-workers report on use of pre-exposure prophylaxis by young women in Southern Africa.     

Nanomiemgel - A Novel Drug Delivery System for Topical Application - In Vitro and In Vivo Evaluation

AimThe objective of this study was to formulate and evaluate a unique matrix mixture (nanomiemgel) of nanomicelle and nanoemulsion containing aceclofenac and capsaicin using in vitro and in vivo analyses and to compare it to a marketed formulation (Aceproxyvon).MethodsNanomicelles were prepared using Vitamin E TPGS by solvent evaporation method and nanoemulsion was prepared by high-pressure homogenization method. In vitro drug release and human skin permeation studies were performed and analyzed using HPLC. The efficiency of nanomiemgel as a delivery system was investigated using an imiquimod-induced psoriatic like plaque model developed in C57BL/6 mice.ResultsAtomic Force Microscopy images of the samples exhibited a globular morphology with an average diameter of 200, 250 and 220 nm for NMI, NEM and NMG, respectively. Nanomiemgel demonstrated a controlled release drug pattern and induced 2.02 and 1.97-fold more permeation of aceclofenac and capsaicin, respectively than Aceproxyvon through dermatomed human skin. Nanomiemgel also showed 2.94 and 2.09-fold greater Cmax of aceclofenac and capsaicin, respectively than Aceproxyvon in skin microdialysis study in rats. The PASI score, ear thickness and spleen weight of the imiquimod-induced psoriatic-like plaque model were significantly (p<0.05) reduced in NMG treated mice compared to free drug, NEM, NMI & Aceproxyvon.ConclusionUsing a new combination of two different drug delivery systems (NEM+NMI), the absorption of the combined system (NMG) was found to be better than either of the individual drug delivery systems due to the utilization of the maximum possible paths of absorption available for that particular drug.     

An amino acidic adjuvant to augment cryoinjury of MCF-7 breast cancer cells

One of the major challenges in cryosurgery is to minimize incomplete cryodestruction near the edge of the iceball. In the present study, the feasibility and effectiveness of an amino acidic adjuvant, glycine was investigated to enhance the cryodestruction of MCF-7 human breast cancer cell at mild freezing/thawing conditions via eutectic solidification. The effects of glycine addition on the phase change characteristics of NaCl–water binary mixture were investigated with a differential scanning calorimeter and cryo-macro/microscope. The results confirmed that a NaCl–glycine–water mixture has two distinct eutectic phase change events – binary eutectic solidification of water–glycine, and ternary eutectic solidification of NaCl–glycine–water. In addition, its effects on the cryoinjury of MCF-7 cells were investigated by assessing the post-thaw cellular viability after a single freezing/thawing cycle with various eutectic solidification conditions due to different glycine concentrations, end temperatures and hold times. The viability of MCF-7 cells in isotonic saline supplemented with 10% or 20% glycine without freezing/thawing remained higher than 90% (n = 9), indicating no apparent toxicity was induced by the addition of glycine. With 10% glycine supplement, the viability of the cells frozen to −8.5 °C decreased from 85.9 ± 1.8% to 38.5 ± 1.0% on the occurrence of binary eutectic solidification of glycine–water (n = 3 for each group). With 20% glycine supplement, the viability of the cells frozen to −8.5 °C showed similar trends to those with 10% supplement. However, as the end temperature was lowered to −15 °C, the viability drastically decreased from 62.5 ± 2.0% to 3.6 ± 0.7% (n = 3 for each group). The influences of eutectic kinetics such as nucleation temperature, hold time and method were less significant. These results imply that the binary eutectic solidification of water–glycine can augment the cryoinjury of MCF-7 cells, and the extent of the eutectic solidification is significant.     

Functional Diversity as a New Framework for Understanding the Ecology of an Emerging Generalist Pathogen

Emerging infectious disease outbreaks are increasingly suspected to be a consequence of human pressures exerted on natural ecosystems. Previously, host taxonomic communities have been used as indicators of infectious disease emergence, and the loss of their diversity has been implicated as a driver of increased presence. The mechanistic details in how such pathogen–host systems function, however, may not always be explained by taxonomic variation or loss. Here we used machine learning and methods based on Gower’s dissimilarity to quantify metrics of invertebrate functional diversity, in addition to functional groups and their taxonomic diversity at sites endemic and non-endemic for the model generalist pathogen Mycobacterium ulcerans, the causative agent of Buruli ulcer. Changes in these metrics allowed the rapid categorisation of the ecological niche of the mycobacterium’s hosts and the ability to relate specific host traits to its presence in aquatic ecosystems. We found that taxonomic diversity of hosts and overall functional diversity loss and evenness had no bearing on the mycobacterium’s presence, or whether the site was in an endemic area. These findings, however, provide strong evidence that generalist environmentally persistent bacteria such as M. ulcerans can be associated with specific functional traits rather than taxonomic groups of organisms, increasing our understanding of emerging disease ecology and origin.