High CD169 Monocyte/Lymphocyte Ratio Reflects Immunophenotype Disruption and Oxygen Need in COVID-19 Patients

Background: Sialoadhesin (CD169) has been found to be overexpressed in the blood of COVID-19 patients and identified as a biomarker in early disease. We analyzed CD169 in the blood cells of COVID-19 patients to assess its role as a predictive marker of disease progression and clinical outcomes. Methods: The ratio of the median fluorescence intensity of CD169 between monocytes and lymphocytes (CD169 RMFI) was analyzed by flow cytometry in blood samples of COVID-19 patients (COV) and healthy donors (HDs) and correlated with immunophenotyping, inflammatory markers, cytokine mRNA expression, pulmonary involvement, and disease progression. Results: CD169 RMFI was high in COV but not in HDs, and it correlated with CD8 T-cell senescence and exhaustion markers, as well as with B-cell maturation and differentiation in COV. CD169 RMFI correlated with blood cytokine mRNA levels, inflammatory markers, and pneumonia severity in patients who were untreated at sampling, and was associated with the respiratory outcome throughout hospitalization. Finally, we also report the first evidence of the specific ability of the spike protein of SARS-CoV-2 to trigger CD169 RMFI in a dose-dependent manner in parallel with IL-6 and IL-10 gene transcription in HD PBMCs stimulated in vitro. Conclusion: CD169 is induced by the spike protein and should be considered as an early biomarker for evaluating immune dysfunction and respiratory outcomes in COVID-19 patients.

Ganoderma formosanum Exopolysaccharides Inhibit Tumor Growth via Immunomodulation

Ganoderma formosanum (GF) is a medicinal mushroom endemic to Taiwan. Previous research established the optimal culture conditions to produce exopolysaccharide rich in β-glucan (GF-EPS) from submerged fermentation of GF. The present study investigated the antitumor effects of GF-EPS in a Lewis lung carcinoma cell (LLC1) tumor-bearing mice model. In the preventive model, GF-EPS was orally administered to mice before LLC1 injection. In the therapeutic model, GF-EPS oral administration was initiated five days after tumor cell injection. The tumor size and body weight of the mice were recorded. After sacrifice, the lymphocyte subpopulation was analyzed using flow cytometry. Spleen tissues were used to analyze cytokine mRNA expression. The results showed that GF-EPS (80 mg/kg) effectively suppressed LLC1 tumor growth in both the preventive and therapeutic models. GF-EPS administration increased the proportion of natural killer cells in the spleen and activated gene expression of several cytokines. Our results provide evidence that GF-EPS promotes tumor inhibition through immunomodulation in tumor-bearing mice.

PDL1 expression on monocytes is associated with plasma cytokines in Tuberculosis and HIV

Introduction PDL1 and its interaction with PD1 is implicated in immune dysfunction in TB and HIV. The expression of PDL1 on multiple subsets of monocytes as well as their associations with cytokines and microbial products have not been well studied. Method HIV (TB-HIV+), TB (TB+HIV-) and TB/HIV co-infected (TB+HIV+) patients as well as apparently healthy controls (TB-HIV-) were recruited. TB and HIV patients were treatment naïve while TB/HIV patients were both ART naïve and experienced but not yet started TB therapy. Monocyte subsets were evaluated for PDL1 expression by flow cytometry; plasma TNFα, IL6, IP10, IFNγ and IL10 were measured by Luminex; and cytokine mRNA from purified monocytes quantitated by qPCR. The association of PDL1 with cytokines, clinical and microbial indices, including HIV viral load, TB smear microscopy and TB urinary lipoarabinomannan (LAM) were assessed. Results Monocyte expression of PDL1 was significantly higher in TB, HIV and TB/HIV co-infected patients compared with healthy controls (p = 0.0001), with the highest levels in TB/HIV co-infected patients. The highest expression of PDL1 was on intermediate (CD14+CD16+) monocytes in all participant groups. PDL1 strongly correlated with HIV viral load in TB/HIV while weakly correlated in HIV. PDL1 levels moderately correlated with plasma TNFα, IL6, IP10, IFNγ and IL10 level in TB subjects whereas weakly correlated with TNFα and IP10 in HIV patients. However, cytokine mRNA from purified monocytes showed no association with either plasma cytokines or monocyte PDL1 expression, implying that if cytokines modulate PDL1, they are likely not originating from circulating monocytes themselves. These results underscore the importance of further characterization of multiple monocyte subsets and their phenotypic and functional differences in different disease states.

Viral manipulation of a mechanoresponsive signaling axis disassembles processing bodies

Processing bodies (PBs) are ribonucleoprotein granules important for cytokine mRNA decay that are targeted for disassembly by many viruses. Kaposi’s sarcoma-associated herpesvirus is the etiological agent of the inflammatory endothelial cancer, Kaposi’s sarcoma, and a PB-regulating virus. The virus encodes Kaposin B (KapB), which induces actin stress fibres (SFs) and cell spindling as well as PB disassembly. We now show that KapB-mediated PB disassembly requires actin rearrangements, RhoA effectors and the mechanoresponsive transcription activator, YAP. Moreover, ectopic expression of active YAP or exposure of ECs to mechanical forces caused PB disassembly in the absence of KapB. We propose that the viral protein KapB activates a mechanoresponsive signaling axis and links changes in cell shape and cytoskeletal structures to enhanced inflammatory molecule expression using PB disassembly. Our work implies that cytoskeletal changes in other pathologies may similarly impact the inflammatory environment.

Modified-Live Feline Calicivirus Vaccination Elicits Cellular Immunity against a Current Feline Calicivirus Field Strain in an Experimental Feline Challenge Study

Feline calicivirus (FCV) is a common cat virus associated with oral ulcerations and virulent-systemic disease. Efficacious FCV vaccines protect against severe disease but not against infection. The high genetic diversity of FCV poses a challenge in vaccine design. Protection against FCV has been related to humoral and cellular immunity; the latter has not been studied in detail. This study investigates the cellular and humoral immune response of specified pathogen-free (SPF) cats after modified-live FCV F9 vaccinations and two heterologous FCV challenges by the analysis of lymphocyte subsets, cytokine mRNA transcription levels, interferon (IFN)-γ release assays in peripheral blood mononuclear cells (PBMCs), anti-FCV antibodies, and neutralisation activity. Vaccinated cats developed a Th1 cytokine response after vaccination. Vaccination resulted in antibodies with neutralising activity against the vaccine but not the challenge viruses. Remarkably, IFN-γ-releasing PBMCs were detected in vaccinated cats upon stimulation with the vaccine strain and the first heterologous FCV challenge strain. After the first experimental infection, the mRNA transcription levels of perforin, granzyme B, INF-γ, and antiviral factor MX1 and the number of IFN-γ-releasing PBMCs when stimulated with the first challenge virus were higher in vaccinated cats compared to control cats. The first FCV challenge induced crossneutralising antibodies in all cats against the second challenge virus. Before the second challenge, vaccinated cats had a higher number of IFN-γ-releasing PBMCs when stimulated with the second challenge virus than control cats. After the second FCV challenge, there were less significant differences detected between the groups regarding lymphocyte subsets and cytokine mRNA transcription levels. In conclusion, modified-live FCV vaccination induced cellular but not humoral crossimmunity in SPF cats; innate immune mechanisms, secretory and membranolytic pathways, and IFN-γ-releasing PBMCs seem to be important in the host immune defence against FCV.

ACE2-like carboxypeptidase B38-CAP protects from SARS-CoV-2-induced lung injury

Angiotensin-converting enzyme 2 (ACE2) is a receptor for cell entry of SARS-CoV-2, and recombinant soluble ACE2 protein inhibits SARS-CoV-2 infection as a decoy. ACE2 is the carboxypeptidase to degrade angiotensin II (Ang II) to angiotensin 1-7 and improves the pathologies of cardiovascular disease and acute lung injury. To address whether the carboxypeptidase enzymatic activity of ACE2 is protective against COVID-19, we investigated the effects of B38-CAP, an ACE2-like enzyme, on SARS-CoV-2-induced lung injury. Expression of ACE2 protein was significantly downregulated in the lungs of SARS-CoV-2-infected hamsters. Recombinant S1 domain or receptor-binding domain (RBD) of the SARS-CoV-2 Spike protein also directly downregulated ACE2 expression and elevated Ang II levels and considerably worsened acid-induced lung injury in hamsters. Treatment with B38-CAP downregulated Spike RBD-induced high Ang II levels, severe inflammation and pulmonary edema through its ACE2-like enzymatic activity. Consistently, elevated cytokine mRNA levels and impaired lung functions were improved by B38-CAP treatment. Moreover, in SARS-CoV-2-infected humanized ACE2 transgenic mice, B38-CAP significantly improved the pathologies of lung injury, alleviated the cytokine storms and downregulated viral RNA levels. These results provide the first experimental in vivo evidence that increasing ACE2-like enzymatic activity is a potential and powerful therapeutic strategy for lung pathologies in COVID-19.

Dietary Choline-Enhanced Skin Immune Response of Juvenile Grass Carp Might Be Related to the STAT3 and NF-kB Signaling Pathway (Ctenopharyngodon idella)

To investigate the effects and potential mechanisms of dietary choline on immune function in the skin of juvenile grass carp (Ctenopharyngodon idella), fish were fed different diets containing different levels of choline (142. 2, 407.4, 821.6, 1215.8, 1589.3, and 1996.6 mg/kg) for 70 d and then sampled after a 6-d challenge test. The results exhibited that dietary choline (1) advanced the contents of phosphatidylcholine (PC), betaine, and choline in grass carp skin (P < 0.05) and upregulated the mRNA abundance of choline transporter high-affinity choline transporter 1 (CHT1), choline transporter-like protein 1 (CTL1), and choline transporter-like protein 5 (CTL5), indicating that dietary choline could increase the contents of choline which might be connected with choline transporters in the grass carp skin; (2) receded skin rot symptom after infection with A. hydrophila (Aeromonas hydrophila), increased the levels of IgM, C4, and C3 and the activities of acid phosphatase (ACP) and lysozyme (LZ), raised mucin2, β-defensin, hepcidin, and LEAP-2B mRNA abundance (rather than LEAP-2A), downregulated pro-inflammatory cytokine mRNA abundance (IFN-γ2, IL-15, TNF-α, IL-6, IL-12P40, and IL-1β) in skin of juvenile grass carp (P < 0.05), and upregulated anti-inflammatory cytokine mRNA abundance (IL-10, IL-4/13A, TGF-β1, IL-11, and IL-4/13B) in grass carp skin (P < 0.05), demonstrating that choline enhanced the skin immune function; and (3) downregulated the mRNA abundance of IKKγ, NF-κBp52, IKKβ, c-Rel, NF-κBp65, STAT3b2, STAT3b1, JAK1, and JAK2 as well as protein level of NF-κBp65, p-STAT3 Tyr705, and p-STAT3 Ser727 in nucleus and inhibited the mRNA and protein level of IkBα (P < 0.05), indicating that choline-enhanced immune function might be relevant to the JAK1, 2 /STAT3, and NF-κB signaling pathway in fish skin. In conclusion, choline enhanced the skin immune function which might be related to JAK1, 2/STAT3, and NF-κB signaling molecules in fish. Furthermore, based on immune indices of grass carp (9.28–108.97 g) skin (C3 and IgM contents as well as ACP activities), the choline requirements were estimated to be 1475.81, 1364.24, and 1574.37 mg/kg diet, respectively.

Comparative Analysis of Cytokine mRNA Expressions in Human Tissues with Mycobacterium tuberculosis Infection

In the present study, we aimed to investigate whether an automated molecular diagnostic method based on PCR-reverse blot hybridization assay can discriminate between human Mycobacterium tuberculosis (MTB)-positive and -negative FFPE tissues and to compare the relative mRNA expression levels of various host immune markers between MTB-infected and uninfected human tissues using quantitative reverse transcription (qRT) PCR. A total of 52 human FFPE tissue samples from various regions of the body, including the lungs, lymph nodes, tendons, colon, and appendix, were collected and used for the molecular identification of Mycobacterium species and analysis of cytokine mRNA expression. As a result, IFN-γ, TNF-α, IP-10, CXCL9, CXCL11, and GM-CSF mRNA expression levels in MTB-infected tissues were significantly higher than those in uninfected samples. Additionally, the differences in the mRNA expression levels of IFN-γ, CXCL9, and GM-CSF between MTB-infected and uninfected tissues were statistically significant were statistically significant (p < 0.05). Correlation curve analysis indicated that the mRNA expression of IFN-γ was inversely proportional to that of IP-10 and that the mRNA expression levels of IFN-γ, TNF-α, CXCL9, CXCL11, GM-CSF, and TNFR were proportional and well-correlated. Furthermore, to establish marker profiles for detecting MTB infection, the statistically significant expression levels of three markers were combined. We confirmed that the combined profile of IFN-γ, CXCL9, and GM-CSF expression levels was statistically significant (P < 0.001). Although the mRNA expression patterns of host immune markers may vary according to MTB infection status, these patterns may be highly correlated and can be simultaneously used as an additional indicator for diagnosing TB.

Differential Expression of Proinflammatory Cytokines IFN-γ and TNF-α in CKD Patients from South India.

Objective: To explore the proinflammatory cytokine expression from chronic kidney disease patients (CKD) with its secondary complications.Methods: A total of 133 CKD patients and 149 healthy controls were evaluated for TNF-α and IFN-γ cytokine mRNA expression by qRT- PCR methods.Results: We found upregulated expression for TNF-α (FC: 2.5) and IFN-γ (FC:1.76) in pooled CKD patients when compared to controls. The expression profile for TNF-α and IFN-γ was 2.6 and 1.71 fold respectively for dialysis patients. However, in Non-Dialysis patients, a down regulated expression for TNF-α (FC: 0.19) and upregulated expression for IFN-γ (FC:1.6) were noticed. The IFN-γ and TNF-α expression level was 2.02 and 1.79 fold respectively for CKD patients with diabetes. Whereas in non-diabetic CKD patients, the IFN-γ and TNF-α expressions were 1.79 and 1.87 fold respectively. When we grouped the data based on with and without complications, we found a significant upregulated expression for IFN-γ (FC:1.64) and down regulated expression for TNF- α (FC:0.65) were observed in without complications. A significant upregulated expression were observed for IFN-γ (FC:1.82) and TNF- α (FC:2.27) in with complications. We have observed a significant positive correlation between TNF-α and IFN-γ (R=0.620; p< 0.0001). Analysis of data showed negative correlation between eGFR and Creatinine in Dialysis and Non-Dialysis group of patients. The disease severity progression showed that, 84.2% (n=112) of individuals fall under <15 eGFR.Conclusion: Increased TNF-α and IFN-γ expression suggests that Th1 cells are involved in CKD inflammation and its disease pathogenesis.