Differentiation of Human Umbilical Cord Derived Mesenchymal Stem Cells Into Neural Stem Cells Induced by hPRDX5

Background: The objectives of this study were to investigate the characteristics and capacity of human umbilical cord‑derived mesenchymal stem cells (hUC-MSCs) differentiation into neural stem cells (NSCs) and whether this event enhanced by hPRDX5. Methods and Results: The adherent cells were obtained from umbilical cord of normal full-term newborn by caesarean section under aseptic condition, and cultivated by tissue block culture method. The surface antigen expression profiles of hUC-MSCs were monitored and the multi-directional differentiation potential was identified. Following amplification, the cells of the 4th passage were divided into 5 groups (groups A-E). The morphology was observed under inverted microscope, and the positive expression rate of markers of neural stem cell was detected by immunocytochemical and western blot. Flow cytometry revealed that the hUC-MSCs expressed CD29, CD73, CD90 and CD105, but not CD19, CD34, CD45 or HLA-DR. Treatment with hPRDX5 led to the surface markers of neural stem cells which were positive for Nestin, but negative for NSE and GFAP expression. Conclusions: Thus, the findings of the present study demonstrate that hPRDX5 effectively promotes hUC-MSCs to differentiate into neural stem cells possibly through TLR4 signaling pathway.

A simple and intuitive methodology for estimating red blood cell surface antigen expression variation using optical cell-detachment technique

The analysis of surface antigens on cells, especially red blood cells (RBCs), has attracted increasing attention due to the recognition of antigenic variation that can facilitate early diagnoses. This paper presents an alternative methodology to estimate the variation of surface antigen expressions using an optical cell-detachment technique to validate the binding of individual RBCs stuck on corresponding antibody-coated surfaces. The detachment tests were implemented by an optical tweezers with gradually decreasing laser powers associated with serial antibody dilutions. Then, the antigen expression variation was estimated based on the known antibody dilution folds. The B- and B3-types of RBCs were selected for the demonstration subjects. With the semi-quantitative analysis, the proposed methodology was successfully verified for evaluating the variation of the RBC surface antigen expressions. The analysis result shows good consistency with the literature’s findings.

Strategies to Inhibit Hepatitis B Virus Gene Transcription

Approximately 240 million people are chronically infected with hepatitis B virus (HBV), despite four decades of an effective HBV vaccine. During chronic infection, HBV forms two distinct templates responsible for viral gene transcription: (1) episomal covalently closed circular (ccc)DNA and (2) host-genome integrated viral templates. Multiple ubiquitous and liver-specific transcription factors are recruited onto these templates and modulate viral gene transcription. This review details the latest developments in antivirals that inhibit HBV gene transcription, and their impact on the stability of viral transcripts. Notably, nuclear receptor agonists exhibit potent inhibition of viral gene transcription from cccDNA, small molecule inhibitors repress HBV X protein-mediated transcription from cccDNA and small interfering RNAs and single-stranded oligonucleotides result in transcript degradation from both cccDNA and integrant templates. These antivirals mediate their effects by reducing viral transcripts abundance, eventually leading to loss of surface antigen expression, and can potentially be added to the arsenal of drugs with demonstrable anti-HBV activity. Thus, these candidates deserve special attention for future repurposing or further development as anti-HBV therapeutics.

Time-Resolved scRNA-Seq Tracks the Adaptation of a Sensitive MCL Cell Line to Ibrutinib Treatment

Since the approval of ibrutinib for relapsed/refractory mantle cell lymphoma (MCL), the treatment of this rare mature B-cell neoplasm has taken a great leap forward. Despite promising efficacy of the Bruton tyrosine kinase inhibitor, resistance arises inevitably and the underlying mechanisms remain to be elucidated. Here, we aimed to decipher the response of a sensitive MCL cell line treated with ibrutinib using time-resolved single-cell RNA sequencing. The analysis uncovered five subpopulations and their individual responses to the treatment. The effects on the B cell receptor pathway, cell cycle, surface antigen expression, and metabolism were revealed by the computational analysis and were validated by molecular biological methods. The observed upregulation of B cell receptor signaling, crosstalk with the microenvironment, upregulation of CD52, and metabolic reprogramming towards dependence on oxidative phosphorylation favor resistance to ibrutinib treatment. Targeting these cellular responses provide new therapy options in MCL.

Therapeutic Rationale for Endotoxin Removal with Polymyxin B Immobilized Fiber Column (PMX) for Septic Shock

Endotoxin removal therapy with polymyxin B immobilized fiber column (PMX) has been clinically applied for sepsis and septic shock patients since 1994. The effectiveness and usefulness of this therapy have been demonstrated for more than a quarter of a century. However, a documented survival benefit has not yet been demonstrable in a large, multicenter, randomized and controlled trial. Following the findings derived from a large sepsis clinical trial with PMX in North America, a new trial is ongoing to determine if PMX has a long-term survival benefit when administered to septic patients. Another approach to support a survival benefit from intervention with PMX is to utilize a detailed analysis available from a large clinical data base. The endotoxin adsorption capacity of PMX columns in vitro and the effectiveness of PMX columns can be further demonstrable in animal models. The capability of PMX and details of its mechanism of action to intervene in the sepsis cascade and impede organ dysfunction in septic patients is not fully understood. The surface antigen expression in monocytes and neutrophils are improved after PMX therapy. Immunomodulatory effects as a result of endotoxin removal and/or other mechanisms of action have been suggested. These effects and other potential immune effects may explain some of the improved effects upon organ dysfunction of sepsis and septic shock patients. Endotoxemia may be involved in the pathophysiology of other diseases than sepsis. A rapid diagnostic method to detect and target endotoxemia could allow us to practice precision medicine and expand the clinical indications of endotoxin removal therapy.

A quantitative methodology to detect surface antigens on red blood cells using optical cell-detachment technique

The quantitative analysis of surface antigens on cells, especially red blood cells (RBCs), has attracted increasing attention due to the recognition of antigenic changes that can facilitate early diagnoses. This paper presents an alternative methodology developed using the optical cell-detachment technique to evaluate antibody-antigen interactions and quantitatively analyze the RBC surface antigen expression. RBC subtyping was used to verify the proposed detection principle based on a comparison of the bonding strengths between individual RBCs and antibody coatings. The bonding strengths were measured with serial antibody dilutions with gradually decreasing laser powers, for which a single cell was optically detached from the corresponding antibody-coated surface. With the quantitatively analysis, the proposed alternative methodology was verified as a highly sensitive technique for detecting antigen expression on the RBC surface.

Differences in pulmonary innate lymphoid cells are dependent on mouse age, sex and strain

Innate lymphoid cells (ILC) are resident in the lung and are involved in both the maintenance of homeostasis and the pathogenesis of respiratory diseases. In this study, murine lung ILC were characterised using flow cytometry and the impact of mouse age, sex and strain were assessed. Lung ILC were found as early as postnatal day 4 and numbers peaked at 2 weeks, and then decreased as the lung matured. During postnatal lung development, ILC expressed differential amounts of ILC2-associated cell surface antigens including ST2, CD90.2 and ICOS. Using Il5venusIl13td-tomato dual reporter mice, neonates were found to have increased constitutive IL-13 expression compared to adult mice. Neonates and adults had similar ratios of IL-5+CD45+ leukocytes, however, these cells were mostly composed of ILC in neonates and T cells in adults. Sex-specific differences in ILC numbers were also observed, with females having greater numbers of lung ILC than males in both neonatal and adult mice. Female lung ILC also expressed higher levels of ICOS and decreased KLRG1. Mouse strain also impacted on lung ILC with BALB/c mice having more ILC in the lung and increased expression of ST2 and ICOS compared with C57BL/6J mice. Collectively, these data show that lung ILC numbers, cell surface antigen expression, IL-5 and IL-13 levels differed between neonatal and adult lung ILC. Additionally, cell surface antigens commonly used for ILC2 quantification, such as ST2, CD90.2, and ICOS, differ depending on age, sex and strain and these are important considerations for consistent universal identification of lung ILC2.

Subpopulations of miniature pig mesenchymal stromal cells with different differentiation potentials differ in the expression of octamer-binding transcription factor 4 and sex determining region Y-box 2

Objective: Human mesenchymal stromal cells (MSCs) exhibit variable differentiation potential and can be divided accordingly into distinct subpopulations whose ratios vary with donor age. However, it is unknown whether the same is true in pigs. This study investigated MSC subpopulations in miniature pig and compared their characteristics in young (2 to 3 months) and adult (27 to 35 months) pigs.Methods: Osteogenic, chondrogenic, and adipogenic capacity of isolated MSCs was evaluated by von Kossa, Alcian blue, and oil red O staining, respectively. Cell surface antigen expression was determined by flow cytometry. Proliferative capacity was assessed with the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Expression of marker genes was detected by quantitative real-time polymerase chain reaction.Results: Porcine MSCs comprised cells with trilineage and bilineage differentiation potential (tMSCs and bMSCs, respectively) and non-differentiating stromal cells (NDSCs). The tMSC and bMSC fractions were smaller in adult than in young pigs (63.0% vs 71.2% and 11.6% vs 24.0%, respectively, p<0.05); NDSCs showed the opposite trend (25.4% vs 4.8%; p<0.05). Subpopulations showed no differences in morphology, cell surface antigen expression, or proliferative capacity, but octamer-binding transcription factor 4 (OCT4) expression was higher in tMSCs than in bMSCs and NDSCs (p<0.05), whereas sex determining region Y-box 2 (SOX2) expression was higher in tMSCs and bMSCs than in NDSCs (p<0.05). Aging had no effect on these trends.Conclusion: Porcine MSCs comprise distinct subpopulations that differ in their differentiation potential and OCT4 and SOX2 expression. Aging does not affect the characteristics of each subpopulation but alters their ratios.