Distribution of ACE2, CD147, CD26, and other SARS-CoV-2 associated molecules in tissues and immune cells in health and in asthma, COPD, obesity, hypertension, and COVID-19 risk factors.

BACKGROUND: Morbidity and mortality from COVID-19 caused by novel coronavirus SARS-CoV-2 is accelerating worldwide, and novel clinical presentations of COVID-19 are often reported. The range of human cells and tissues targeted by SARS-CoV-2, its potential receptors and associated regulating factors are still largely unknown. The aim of our study was to analyze the expression of known and potential SARS-CoV-2 receptors and related molecules in the extensive collection of primary human cells and tissues from healthy subjects of different age and from patients with risk factors and known comorbidities of COVID-19. METHODS: We performed RNA sequencing and explored available RNA-Seq databases to study gene expression and co-expression of ACE2, CD147 (BSG), and CD26 (DPP4) and their direct and indirect molecular partners in primary human bronchial epithelial cells, bronchial and skin biopsies, bronchoalveolar lavage fluid, whole blood, peripheral blood mononuclear cells (PBMCs), monocytes, neutrophils, DCs, NK cells, ILC1, ILC2, ILC3, CD4+ and CD8+ T cells, B cells, and plasmablasts. We analyzed the material from healthy children and adults, and from adults in relation to their disease or COVID-19 risk factor status. RESULTS: ACE2 and TMPRSS2 were coexpressed at the epithelial sites of the lung and skin, whereas CD147 (BSG), cyclophilins (PPIA andPPIB), CD26 (DPP4), and related molecules were expressed in both epithelium and in immune cells. We also observed a distinct age-related expression profile of these genes in the PBMCs and T cells from healthy children and adults. Asthma, COPD, hypertension, smoking, obesity, and male gender status generally led to the higher expression of ACE2- and CD147-related genes in the bronchial biopsy, BAL, or blood. Additionally, CD147-related genes correlated positively with age and BMI. Interestingly, we also observed higher expression of CD147-related genes in the lesional skin of patients with atopic dermatitis. CONCLUSIONS: Our data suggest different receptor repertoire potentially involved in the SARS-CoV-2 infection at the epithelial barriers and in the immune cells. Altered expression of these receptors related to age, gender, obesity and smoking, as well as with the disease status, might contribute to COVID-19 morbidity and severity patterns.

Differential T cell response against BK virus regulatory and structural antigens: A viral dynamics modelling approach.

BK virus (BKV) associated nephropathy affects 1-10% of kidney transplant recipients, leading to graft failure in about 50% of cases. Immune responses against different BKV antigens have been shown to have a prognostic value for disease development. Data currently suggest that the structural antigens and regulatory antigens of BKV might each trigger a different mode of action of the immune response. To study the influence of different modes of action of the cellular immune response on BKV clearance dynamics, we have analysed the kinetics of BKV plasma load and anti-BKV T cell response (Elispot) in six patients with BKV associated nephropathy using ODE modelling. The results show that only a small number of hypotheses on the mode of action are compatible with the empirical data. The hypothesis with the highest empirical support is that structural antigens trigger blocking of virus production from infected cells, whereas regulatory antigens trigger an acceleration of death of infected cells. These differential modes of action could be important for our understanding of BKV resolution, as according to the hypothesis, only regulatory antigens would trigger a fast and continuous clearance of the viral load. Other hypotheses showed a lower degree of empirical support, but could potentially explain the clearing mechanisms of individual patients. Our results highlight the heterogeneity of the dynamics, including the delay between immune response against structural versus regulatory antigens, and its relevance for BKV clearance. Our modelling approach is the first that studies the process of BKV clearance by bringing together viral and immune kinetics and can provide a framework for personalised hypotheses generation on the interrelations between cellular immunity and viral dynamics.

Pro-Inflammatory versus Immunomodulatory Effects of Silver Nanoparticles in the Lung: The Critical Role of Dose, Size and Surface Modification.

The growing use of silver nanoparticles (Ag-NPs) in consumer products raises concerns about their toxicological potential. The purpose of the study was to investigate the size- and coating-dependent pulmonary toxicity of Ag-NPs in vitro and in vivo, using an ovalbumin (OVA)-mouse allergy model. Supernatants from (5.6-45 µg/mL) Ag50-PVP, Ag200-PVP or Ag50-citrate-treated NR8383 alveolar macrophages were tested for lactate dehydrogenase and glucuronidase activity, tumor necrosis factor (TNF)-α release and reactive oxygen species (ROS) production. For the in vivo study, NPs were intratracheally instilled in non-sensitized (NS) and OVA-sensitized (S) mice (1-50 µg/mouse) prior to OVA-challenge and bronchoalveolar lavage fluid (BALF) inflammatory infiltrate was evaluated five days after challenge. In vitro results showed a dose-dependent cytotoxicity of Ag-NPs, which was highest for Ag50-polyvinilpyrrolidone (PVP), followed by Ag50-citrate, and lowest for Ag200-PVP. In vivo 10-50 µg Ag50-PVP triggered a dose-dependent pulmonary inflammatory milieu in NS and S mice, which was significantly higher in S mice and was dampened upon instillation of Ag200-PVP. Surprisingly, instillation of 1 µg Ag50-PVP significantly reduced OVA-induced inflammatory infiltrate in S mice and had no adverse effect in NS mice. Ag50-citrate showed similar beneficial effects at low concentrations and attenuated pro-inflammatory effects at high concentrations. The lung microbiome was altered by NPs instillation dependent on coating and/or mouse batch, showing the most pronounced effects upon instillation of 50 µg Ag50-citrate, which caused an increased abundance of operational taxonomic units assigned to Actinobacteria, Bacteroidetes, Firmicutes and Proteobacteria. However, no correlation with the biphasic effect of low and high Ag-NPs dose was found. Altogether, both in vitro and in vivo data on the pulmonary effects of Ag-NPs suggest the critical role of the size, dose and surface functionalization of Ag-NPs, especially in susceptible allergic individuals. From the perspective of occupational health, care should be taken by the production of Ag-NPs-containing consumer products.

A revised strategy for monitoring BKV-specific cellular immunity in kidney transplant patients.

Reactivation of Polyomavirus BKV is a severe complication in kidney transplant patients. Current treatment requires close monitoring, and modification of immunosuppressive drugs. As an important additional tool, the monitoring of BKV immunity has been based on detection of cytokine-secreting T cells upon BKV-antigen challenge. However, low frequent BKV-specific T cells are often barely detectable and their roles in BKV clearance remain unclear. Here, we analyzed the effects of immunosuppressive agents on BKV-specific T cells in vitro. Significant reductions in expression of several markers, and reduced killing functions upon treatment with calcineurin but not mTOR inhibitors were detected. However, effects of these drugs on expression of surface markers and GranzymeB were substantially less striking than effects on cytokine expression. Consequently, we applied a novel detection strategy for BKV-specific T cells in immunosuppressed kidney transplant patients using these more robust markers, and showed significantly improved sensitivity compared with the conventional IFNγ-based method. Using this strategy and 17-color flow cytometry, we found BKV-specific helper and cytolytic CD4+ T-cell subsets that differed in their memory phenotype, which corresponded with BKV clearance in kidney transplant patients. Thus, our results offer an improved detection strategy for BKV-specific T cells in kidney transplant patients, and shed light on the contributions of these cells to BKV clearance.

Contrast-enhanced spectral mammography: Does mammography provide additional clinical benefits or can some radiation exposure be avoided?

The purpose of this study was to compare contrast-enhanced spectral mammography (CESM) with mammography (MG) and combined CESM + MG in terms of detection and size estimation of histologically proven breast cancers in order to assess the potential to reduce radiation exposure. A total of 118 patients underwent MG and CESM and had final histological results. CESM was performed as a bilateral examination starting 2 min after injection of iodinated contrast medium. Three independent blinded radiologists read the CESM, MG, and CESM + MG images with an interval of at least 4 weeks to avoid case memorization. Sensitivity and size measurement correlation and differences were calculated, average glandular dose (AGD) levels were compared, and breast densities were reported. Fisher's exact and Wilcoxon tests were performed. A total of 107 imaging pairs were available for analysis. Densities were ACR1: 2, ACR2: 45, ACR3: 42, and ACR4: 18. Mean AGD was 1.89 mGy for CESM alone, 1.78 mGy for MG, and 3.67 mGy for the combination. In very dense breasts, AGD of CESM was significantly lower than MG. Sensitivity across readers was 77.9 % for MG alone, 94.7 % for CESM, and 95 % for CESM + MG. Average tumor size measurement error compared to postsurgical pathology was -0.6 mm for MG, +0.6 mm for CESM, and +4.5 mm for CESM + MG (p < 0.001 for CESM + MG vs. both modalities). CESM alone has the same sensitivity and better size assessment as CESM + MG and was significantly better than MG with only 6.2 % increase in AGD. The combination of CESM + MG led to systematic size overestimation. When a CESM examination is planned, additional MG can be avoided, with the possibility of saving up to 61 % of radiation dose, especially in patients with dense breasts.

The genetic predisposition of natural killer cell to BK virus-associated nephropathy in renal transplant patients.

BK virus (BKV) infection represents a serious complication in renal transplant patients resulting in BKV-associated nephropathy and subsequent allograft loss. Natural killer cells are crucial in the antiviral immune response; however, an understanding of the role of natural killer cells in protection against BKV is limited. To elucidate whether killer-cell immunoglobulin-like receptors and their interaction between donor-/recipient-related ligands have a role in BKV infection, we performed genotyping analysis in 48 kidney transplant recipients with a history of severe BKV infection/BKV-associated nephropathy and 110 recipients with stable renal function and no BKV reactivation. Of interest, we found that telomeric gene content motif was significantly associated with severe course of BKV infection/BKV-associated nephropathy and detected significantly higher percentage of patients with BKV-associated nephropathy carrying low numbers of activating receptors compared with the control group. Detailed analysis of each single receptor revealed significantly lower frequencies of the activating receptor KIR3DS1 in patients with BKV infection/nephropathy as compared with the controls. Thus, our study supports protective effects of activating receptors in BKV infection and suggest natural killer-cell-related genetic predisposition to the development of BKV-associated nephropathy.

Impact of soluble CD26 on treatment outcome and hepatitis C virus-specific T cells in chronic hepatitis C virus genotype 1 infection.

BACKGROUND: Interferon and ribavirin therapy for chronic hepatitis C virus (HCV) infection yields sustained virological response (SVR) rates of 50-80%. Several factors such as non-1 genotype, beneficial IL28B genetic variants, low baseline IP-10, and the functionality of HCV-specific T cells predict SVR. With the pending introduction of new therapies for HCV entailing very rapid clearance of plasma HCV RNA, the importance of baseline biomarkers likely will increase in order to tailor therapy. CD26 (DPPIV) truncates the chemokine IP-10 into a shorter antagonistic form, and this truncation of IP-10 has been suggested to influence treatment outcome in patients with chronic HCV infection patients. In addition, previous reports have shown CD26 to be a co-stimulator for T cells. The aim of the present study was to assess the utility of CD26 as a biomarker for treatment outcome in chronic hepatitis C and to define its association with HCV-specific T cells. METHODS: Baseline plasma from 153 genotype 1 and 58 genotype 2/3 infected patients enrolled in an international multicenter phase III trial (DITTO-HCV) and 36 genotype 1 infected patients participating in a Swedish trial (TTG1) were evaluated regarding baseline soluble CD26 (sCD26) and the functionality of HCV-specific CD8(+) T cells. RESULTS: Genotype 1 infected patients achieving SVR in the DITTO (P = 0.002) and the TTG1 (P = 0.02) studies had lower pretreatment sCD26 concentrations compared with non-SVR patients. Sixty-five percent of patients with sCD26 concentrations below 600 ng/mL achieved SVR compared with 39% of the patients with sCD26 exceeding 600 ng/mL (P = 0.01). Patients with sCD26 concentrations below 600 ng/mL had significantly higher frequencies of HCV-specific CD8(+) T cells (P = 0.02). CONCLUSIONS: Low baseline systemic concentrations of sCD26 predict favorable treatment outcome in chronic HCV infection and may be associated with higher blood counts of HCV-specific CD8(+) T cells.

Human cytomegalovirus kinetics following institution of artesunate after hematopoietic stem cell transplantation.

The anti-malaria drug artesunate has been shown to be an effective inhibitor of cytomegalovirus (CMV) in vitro, in an experimental animal model, and in a recent single-case clinical use. In this first case-series of 6 stem cell transplant recipients who received preemptive artesunate treatment for CMV infection, we have examined the viral kinetics following institution of artesunate, and employed first-phase viral kinetics studies to calculate its antiviral effectiveness. Two patients demonstrated a rapid 0.8-2.1 log viral load decline by 7 days, with a viral decay half-live of 0.9-1.9 days. Four patients demonstrated a continued yet stalled viral growth slope during treatment. No adverse events were noted in treatment courses of up to 28 days. Overall, a divergent antiviral efficacy was revealed, ranging from 43% to 90%, which appeared to be primarily dependent on the virus baseline growth dynamics. Further dose escalation studies are needed to examine the role of artesunate in the treatment of CMV infection in the transplantation setting.

Novel mechanism of antibodies to hepatitis B virus in blocking viral particle release from cells.

UNLABELLED: Antibodies are thought to exert antiviral activities by blocking viral entry into cells and/or accelerating viral clearance from circulation. In particular, antibodies to hepatitis B virus (HBV) surface antigen (HBsAg) confer protection, by binding circulating virus. Here, we used mathematical modeling to gain information about viral dynamics during and after single or multiple infusions of a combination of two human monoclonal anti-HBs (HepeX-B) antibodies in patients with chronic hepatitis B. The antibody HBV-17 recognizes a conformational epitope, whereas antibody HBV-19 recognizes a linear epitope on the HBsAg. The kinetic profiles of the decline of serum HBV DNA and HBsAg revealed partial blocking of virion release from infected cells as a new antiviral mechanism, in addition to acceleration of HBV clearance from the circulation. We then replicated this approach in vitro, using cells secreting HBsAg, and compared the prediction of the mathematical modeling obtained from the in vivo kinetics. In vitro, HepeX-B treatment of HBsAg-producing cells showed cellular uptake of antibodies, resulting in intracellular accumulation of viral particles. Blocking of HBsAg secretion also continued after HepeX-B was removed from the cell culture supernatants. CONCLUSION: These results identify a novel antiviral mechanism of antibodies to HBsAg (anti-HBs) involving prolonged blocking of the HBV and HBsAg subviral particles release from infected cells. This may have implications in designing new therapies for patients with chronic HBV infection and may also be relevant in other viral infections.

Positive and negative prediction of sustained virologic response at weeks 2 and 4 of treatment with albinterferon alfa-2b or peginterferon alfa-2a in treatment-naïve patients with genotype 1, chronic hepatitis C.

BACKGROUND/AIMS: Albinterferon alfa-2b is a novel, long-acting, fusion polypeptide that is dosed q2wk or q4wk. The predictive value of early virologic response during albinterferon alfa-2b or peginterferon alfa-2a treatment was investigated in interferon-naïve patients with genotype 1, chronic hepatitis C. METHODS: Four hundred and fifty-eight patients were randomized to: albinterferon 900 or 1200 microg q2wk, or 1200 microg q4wk, or peginterferon 180 microg qwk. HCV RNA was measured by real-time PCR. A linear exhaustive search algorithm was used to determine the best SVR prediction algorithm in the per-protocol population (n=368), with inclusion of key ITT analyses to assess impact. RESULTS: SVR rate: 54-67% (P=NS between arms). Rapid initial virologic response rate at week 2 (RIVR; viral decline >2 log(10)IU/mL) was 32-50% and gave rise to positive predictive value of 88-97% for SVR. No initial virologic response at week 4 (NIVR; viral decline <2 log(10)IU/mL; viral load >5.5 log(10)IU/mL) demonstrated a 100% negative predictive value for SVR. A sequential prediction algorithm based on viral kinetics at weeks 2 and 4 identified four prediction groups that reliably predicted SVR, positively or negatively, in 65-72% of patients. CONCLUSIONS: Improved SVR prediction was obtained by integrating absolute levels and reduction of HCV RNA at treatment week 2 and 4. Patients with RIVR had a high likelihood of achieving SVR.

Early prediction of sustained virological response at day 3 of treatment with albinterferon-alpha-2b in patients with genotype 2/3 chronic hepatitis C.

BACKGROUND: Albinterferon-alpha-2b (albIFN) is a long-acting fusion polypeptide composed of albumin and IFN-alpha-2b. In a phase 2 study of albIFN 1500 mug q2wk or q4wk in patients with genotype 2/3 chronic hepatitis C, albIFN demonstrated sustained virological response (SVR) rates of 62-77% (intent-to-treat population). AIMS: To assess the association of initial viral kinetics during albIFN therapy with baseline factors and SVR prediction. METHODS: In all, 43 patients were treated with albIFN 1500 mug (q2wk/q4wk) plus ribavirin (RBV) 800 mg/day for 24 weeks. Hepatitis C virus (HCV)-RNA levels were measured by real-time polymerase chain reaction, insulin resistance by homeostasis model assessment of insulin resistance (HOMA-IR) and serum albIFN levels by enyzme-linked immunosorbent assay. Prediction analysis was performed in a per protocol 28-patient subset who were > or =80% adherent to albIFN/RBV and had HCV-RNA levels measured at treatment day 3. RESULTS: Day-3 HCV-RNA level and first-phase viral decline as well as second-phase slope of viral decline were significantly associated with SVR. In adherent patients, 82.1% had a day-3 viral load <4.2 log(10) IU/ml or first-phase decline >1.25 log(10) IU/ml, which was predictive of SVR, both positively (95.7%; sensitivity: 100%) and negatively (100%; specificity: 83.3%). As low first-phase decline was associated with a high pretreatment HOMA-IR index (P=0.004) and a low day-3 serum albIFN level (P=0.01). CONCLUSIONS: First-phase viral decline with albIFN/RBV was predictive of SVR in this study and may be modulated in part by IR.

Artesunate as a potent antiviral agent in a patient with late drug-resistant cytomegalovirus infection after hematopoietic stem cell transplantation.

This is the first report of treatment of cytomegalovirus infection with artesunate, for a stem cell transplant recipient with a newly identified foscarnet-resistant and ganciclovir-resistant DNA polymerase L776M mutation. Artesunate treatment resulted in a 1.7-2.1-log reduction in viral load by treatment day 7, with a viral half-life of 0.9-1.9 days, indicating a highly effective block in viral replication.

Clinical utility of total HCV core antigen quantification: a new indirect marker of HCV replication.

Hepatitis C virus (HCV) RNA detection, viral load quantification, and HCV genotyping are widely used in clinical practice. Recently, the availability of an anticore antigen (Ag) monoclonal antibody allowed development of an enzyme-linked immunosorbent assay (ELISA) detecting and quantifying total HCV core Ag in peripheral blood of HCV-infected patients. The aims of the present study were to investigate the biologic significance of this new marker in HCV infection, to establish the intrinsic performance of the current assay, and to determine its potential utility in the management of HCV-infected patients. A panel of infected sera calibrated to the World Health Organization International Standard and 657 serum samples from infected patients receiving antiviral treatment were studied. We showed that total HCV core Ag quantification is an accurate, precise, and specific indirect marker of HCV replication. We estimated that 1 pg/mL of total HCV core Ag is equivalent to approximately 8,000 HCV RNA international units (IU)/mL, although minor between-patient differences may exist. In conclusion, total HCV core Ag quantification can be used in the various indications of viral load monitoring, including the evaluation of baseline viral load before therapy, the assessment of the virologic response to antiviral treatment, and the study of early viral kinetics during therapy. Nevertheless, the total HCV core Ag assay cannot be used as a marker of viral replication for HCV RNA values below 20,000 IU/mL, limiting its use in the monitoring of late events during and after antiviral treatment.