Abstract
The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is associated with a hyperinflammatory state and lymphocytopenia, a hallmark that appears as both signature and ...prognosis of disease severity outcome. Although cytokine storm and a sustained inflammatory state are commonly associated with immune cell depletion, it is still unclear whether direct SARS-CoV-2 infection of immune cells could also play a role in this scenario by harboring viral replication. We found that monocytes, as well as both B and T lymphocytes, were susceptible to SARS-CoV-2 infection in vitro, accumulating double-stranded RNA consistent with viral RNA replication and ultimately leading to expressive T cell apoptosis. In addition, flow cytometry and immunofluorescence analysis revealed that SARS-CoV-2 was frequently detected in monocytes and B lymphocytes from coronavirus disease 2019 (COVID-19) patients. The rates of SARS-CoV-2-infected monocytes in peripheral blood mononuclear cells from COVID-19 patients increased over time from symptom onset, with SARS-CoV-2-positive monocytes, B cells, and CD4+ T lymphocytes also detected in postmortem lung tissue. These results indicated that SARS-CoV-2 infection of blood-circulating leukocytes in COVID-19 patients might have important implications for disease pathogenesis and progression, immune dysfunction, and virus spread within the host.
Summary
Background
Molecular biomarkers of maternal leptin resistance associated with infant weight are needed.
Objectives
To evaluate gene expression of leptin receptor (LEPR), suppressor of ...cytokine signalling 3 (SOCS3) and insulin receptor in peripheral blood mononuclear cells (PBMCs) of lactating women and their relationship with infant body weight and adiposity.
Methods
At day 10 postpartum, maternal gene expression in PBMCs as well as leptin and insulin concentrations in plasma and milk were assessed (
n
= 68). Infant weight and BMI
z
‐scores, skinfolds and arm circumference were obtained at 10 days and/or at 3 months old.
Results
In mothers with pre‐pregnancy overweight or obesity (OW/OB), LEPR expression was reduced (
p
= 0.013) whereas plasma and milk leptin and milk insulin concentrations were elevated. LEPR expression was positively related with infant weight
z
‐score (Beta (95% CI): 0.40 (0.17, 0.63),
p
= 0.001) but not with leptin concentrations. SOCS3 expression was positively related with infant weight
z
‐score (Beta (95% CI): 0.28 (0.04, 0.51),
p
= 0.024) and arm circumference (Beta (95% CI): 0.57 (0.32, 0.82),
p
< 0.001). Relationships remained significant after adjusting for maternal and infant confounders.
Conclusions
LEPR and SOCS3 gene expression in PBMCs are novel maternal molecular biomarkers that reflect leptin resistance and are associated with infant body weight and adiposity.
γδT cells function in the regulation of T-cell activation in cancer and have been identified as a novel target for cancer immunotherapy. Activated γδT cells release a series of cytotoxic ...molecules-including granulysin, perforin, Fas/Fas ligand (Fas-L), and granzymes A and B-to kill target cells. Our previous research has shown that high mobility group nucleosomal-binding domain 2 (HMGN2), which is expressed at a high level in activated CD8T cells, is an antitumor effector molecule of CD8T cells. In the present study, we examined the expression and antitumor effects of HMGN2 in γδT cells. Peripheral blood mononuclear cells (PBMCs) were isolated from healthy donors with a PBMC separation column. PMBCs were stimulated with isopentenyl pyrophosphate (IPP) and interleukin-2 (IL-2) for 10 days for activation and expansion. Activated γδT cells were isolated from IPP-pretreated PBMCs with a Moflo XDP flow cytometry sorter. The expression of HMGN2 in γδT cells was detected by flow cytometry and enzyme-linked immunosorbent assay. The cytotoxic effects of γδT cells and HMGN2 were analyzed by carboxyfluorescein succinimidyl ester labeling. IPP combined with IL-2 induced significant activation and expansion of γδT cells in vitro. HMGN2 was constitutively expressed in γδT cells. IPP-activated γδT cells expressed a high level of HMGN2 that could be detected intracellularly and in the supernatant. Moreover, supernatants of purified γδT cells were sufficient to kill tumor cells and could be blocked with anti-human HMGN2 antibody. This study suggests that HMGN2 is an antitumor effector molecule of γδT cells.
Objective: The current study evaluates the therapeutic efficacy of Jasminum sambac Linn. essential oil (JEO) by investigating its pharmacological potentials, specifically its anticancer (using MCF-7 ...cells), anti-inflammatory, antioxidant and antimicrobial activity. Methods: With the anti-inflammatory activity assessed as red blood cell membrane stabilization and protein denaturation assay), antioxidant properties tested in DPPH and ABTS assay, and antimicrobial activity screened through disc diffusion method, anti-cancer functions were probed using NCF-7 breast cancer cells. The treatment of these cells with acridine orange–ethidium bromide (AOEtBr) stain helped checking the incidence of apoptotic body formation in the nuclear compartments. Results: The trial treating JEO on peripheral blood mononuclear cells (PBMCs) evinced no signs of cytotoxic changes revealing that JEO is exclusively cancer-cell-specific in action provides added value to this study. Free-radical scavenging activity confirms the antioxidant properties as the anti-inflamatory action is vouched in respective assays. Additionally, JEO is found to exert antimicrobial activity against Gram-positive Staphylococcus aureus, Gram-negative Escherichia coli, and the fungal yeast Candida albicans in disc diffusion assay stretching its manifold utility. Conclusion: Although the present results are convincing, it must be acknowledged that further research focusing on cellular and molecular mechanisms is necessary. Nevertheless, the positive cues gathered on the therapeutic attributes of JEO lend credence to the folk notion that jasmine flowers can be used as an effective medicament for breast ailments.
Daphne mezereum L., an important medicinal plant in Scandinavian folk medicine, was used to treat ailments such as diarrhea, swelling and stomach pain. A range of natural compounds have been ...isolated, but little attention has been given to the polysaccharides in this plant. Previous work in our group have shown that a polysaccharide enriched fraction from the bark of D. mezereum exhibited pro-inflammatory effects. To pursue this further, the aim of the present work was to isolate and characterize these polysaccharides. From the ethanol-precipitate of a water extract, one neutral (DMP-NF) and one acidic (DMP-AF) fraction was isolated by anion-exchange chromatography. GC, GC-MS and 1D- and 2D-NMR were used to characterize the polysaccharide structures. DMP-NF appeared to be a mixture of arabinan, arabinogalactan and hemicelluloses such as xyloglucan, mannan and xylan. DMP-AF contained a pectic polysaccharide mainly consisting of an unusually long homogalacturonan backbone. Enzymatic treatment by pectinase of DMP-AF yielded DMP-ED, which contained a rhamnogalacturonan-I backbone with arabinan, galactan and arabinogalactan side chains. Both DMP-NF and DMP-ED induced IFN-γ and TNF-α secretion in peripheral blood mononuclear cells (PBMCs), DMP-ED being the most potent fraction. DMP-AF was less active, which might be due to a less sterically available rhamnogalacturonan-I domain.
Data on cellular immunity mediators in the early phase of human leishmaniasis are still limited and controversial. In order to mimic the changes of humoral mediators during the early phase of human ...natural infection, some Th1, Th2, Treg, and Breg cytokines, MCP-1, and the nitric oxide (NO) from human PBMC, stimulated by Leishmania infantum, Leishmania major, Leishmania donovani and Leishmania tropica infective metacyclic promastigotes, were determined. After 4 h of L. major, L. donovani, and L. tropica challenge, TNFα, IL-1β, IL-6 levels were significantly higher than negative control cultures with saline (SF) instead of Leishmania promastigotes, unlike L. infantum-stimulated TNFα and L. major-stimulated IL-1β. We obtained higher levels of IL-4 and IL-10 cytokines after stimulation of human PBMCs by L. infantum and L. donovani, compared to those observed after the challenge of PBMCs by L. major and L. tropica. Regarding IL-35, such cytokine levels were significantly increased following infection with L. infantum and L. donovani, in contrast to L. major and L. tropica. Up to our knowledge, we are the first to study the effect of four different species of Leishmania on IL-35 levels in human cells. Our study highlights how several Leishmania species can up-regulate different groups of cytokines (Th1, Th2, Treg and Breg) and modulate NO release in a different way. This original aspect can be explained by different Leishmania cell products, such as LPG, obtained from different strains/species of live parasites. Our findings would contribute to the development of new therapeutics or vaccination strategies.
Background: Endometriosis is believed to be associated with dysfunction of the lymphocyte population and cytotoxicity of natural killer (NK) cells, induced by the production of interleukin-2 (IL-2).
...Objective: This study aimed to investigate T lymphocytes and NK cell activity in the peripheral blood mononuclear cells (PBMCs) of women with endometriosis.
Materials and Methods: PBMCs were obtained from the peripheral venous blood samples of 14 women with and without endometriosis (n = 7 for each group). Then, the PBMCs were co-cultured for 4 days and were treated with recombinant IL-2 for cytotoxic activity toward target cells (Daudi and K562 cells). The cytotoxicity activity was determined using the 51 chromium release assay before and after stimulation. Flow cytometry measurement was used to examine the expression of T lymphocytes and NK cells before and after being treated with IL-2.
Results: The concentration of CD3+CD28+ (co-stimulatory) was significantly lower in the endometriosis group (65.62 ± 5.38) compared to in its counterpart (50.24 ± 4.22) (p = 0.04) before stimulation. However, no significant differences were observed in any other T lymphocytes and NK cells. It was also found that there was a significant increase of CD3-CD28+ after treatment with IL-2 only in the healthy control but not in women with endometriosis.
Conclusion: Increased expression of CD160 and decreased CD28 play a role in inhibiting NK cell activation and T cell response in women with endometriosis.
Key words: CD28, CD160, Cytotoxic, Endometriosis, PBMC.
Lung cancer (LC) is the leading cause of cancer-related death worldwide and miRNAs play a key role in LC development. To better diagnose LC and to predict drug treatment responses we evaluated 228 ...articles encompassing 16,697 patients and 12,582 healthy controls. Based on the criteria of ≥3 independent studies and a sensitivity and specificity of >0.8 we found blood-borne miR-20a, miR-10b, miR-150, and miR-223 to be excellent diagnostic biomarkers for non–small cell LC whereas miR-205 is specific for squamous cell carcinoma. The systematic review also revealed 38 commonly regulated miRNAs in tumor tissue and the circulation, thus enabling the prediction of histological subtypes of LC. Moreover, theranostic biomarker candidates with proven responsiveness to checkpoint inhibitor treatments were identified, notably miR-34a, miR-93, miR-106b, miR-181a, miR-193a-3p, and miR-375. Conversely, miR-103a-3p, miR-152, miR-152-3p, miR-15b, miR-16, miR-194, miR-34b, and miR-506 influence programmed cell death-ligand 1 and programmed cell death-1 receptor expression, therefore providing a rationale for the development of molecularly targeted therapies. Furthermore, miR-21, miR-25, miR-27b, miR-19b, miR-125b, miR-146a, and miR-210 predicted response to platinum-based treatments. We also highlight controversial reports on specific miRNAs. In conclusion, we report diagnostic miRNA biomarkers for in-depth clinical evaluation. Furthermore, in an effort to avoid unnecessary toxicity we propose predictive biomarkers. The biomarker candidates support personalized treatment decisions of LC patients and await their confirmation in randomized clinical trials.
Cu and Fe/Mn dual doped ZnO magnetic nanoparticles (MNPs) had been synthesized by sol-gel method. Characterization of synthesized MNPs showed the fine and significant size of MNPs, it had ...ferromagnetism property at room temperature, and (Cu, Fe) dual doped MNPs were found to have maximum saturation magnetization with larger hysteresis area. In-vitro cytotoxic assessment of these MNPs revealed that 20 ng of Cu doped ZnO MNPs are nontoxic to human PBMC with 60% viable cell count, whereas Fe dopant increases cell viability by 5% in the same concentration. These MNPs showed a significant (20–25 mm ZOI) antimicrobial activity against Pseudomonas putida and Bacillus subtilis bacteria. Particularly Fe dopant showed increased antimicrobial activity against gram-positive (B. subtilis), whereas Mn dopant doesn't have considerable antimicrobial activity. Having known that these MNPs are biocompatible with the ferromagnetic property they may be considered for magnetic guided immunotherapy and hyperthermia applications.
•All the NPs had ferromagnetic nature at room temperature.•The hysteresis loss was high for Cu, Fe co-doped magnetic nanoparticles (MNPs).•The In-vitro cytotoxicity had been reduced by Fe dopant.•The cytotoxicity of MNPs is Dosage independent.•Suitable candidate materials for magnetic hyperthermia applications.
Immunoassays that quantitate cytokines and other surrogate markers of immunity from peripheral blood mononuclear cells (PBMCs), such as flow cytometry or Enzyme-Linked Immunosorbent Spot (ELIspot), ...allow highly sensitive measurements of immune effector function. However, those assays consume relatively high numbers of cells and expensive reagents, precluding comprehensive analyses and high-throughput screening (HTS). To address this issue, we developed a sensitive and specific reverse transcription-quantitative PCR (RT-qPCR)-based HTS assay, specifically designed to quantify surrogate markers of immunity from very low numbers of PBMCs. We systematically evaluated the volumes and concentrations of critical reagents within the RT-qPCR protocol, miniaturizing the assay and ultimately reducing the cost by almost 90% compared to current standard practice. We assessed the suitability of this cost-optimized RT-qPCR protocol as an HTS tool and determined the assay exceeds HTS uniformity and signal variance testing standards. Furthermore, we demonstrate this technique can effectively delineate a hierarchy of responses from as little as 50,000 PBMCs stimulated with CD4
+
or CD8
+
T cell peptide epitopes. Finally, we establish that this HTS-optimized protocol has single-cell analytical sensitivity and a diagnostic sensitivity equivalent to detecting 1:10,000 responding cells (
i.e.
, 100 Spot Forming Cells/10
6
PBMCs by ELIspot) with over 90% accuracy. We anticipate this assay will have widespread applicability in preclinical and clinical studies, especially when samples are limited, and cost is an important consideration.
