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Genetic polymorphisms within the IFNL3/IFNL4genomic region, which encodes type III interferons, have been strongly associated with clearance of hepatitis C virus. We hypothesized that type III interferons might be important for the immune response to other pathogens as well. In a cohort of 914 Malian children, we genotyped functional variants IFNL4-rs368234815, IFNL4-rs117648444, and IFNL3-rs4803217 and analyzed episodes of malaria, gastrointestinal, and respiratory infections recorded at 30,626 clinic visits from birth up to 5 years of age. Compared to children with the rs368234815-TT/TT genotype (IFN-λ4-Null), rs368234815-dG allele was most strongly associated with an earlier time-to-first episode of gastrointestinal infections (p= 0.003). The risk of experiencing an infection episode during the follow-up was also significantly increased with rs368234815-dG allele, with OR = 1.53, 95%CI (1.13–2.07), p= 0.005 for gastrointestinal infections and OR = 1.30, 95%CI (1.02–1.65), p= 0.033 for malaria. All the associations for the moderately linked rs4803217 (r2= 0.78 in this set) were weaker and lost significance after adjusting for rs368234815. We also analyzed all outcomes in relation to IFN-λ4-P70S groups. Our results implicate IFN-λ4 and not IFN-λ3 as the primary functional cause of genetic associations with increased overall risk and younger age at first clinical episodes but not with recurrence or intensity of several common pediatric infections.

Pathologic immune hyperactivation is emerging as a key feature of critical illness in COVID-19, but the mechanisms involved remain poorly understood. We carried out proteomic profiling of plasma from cross-sectional and longitudinal cohorts of hospitalized patients with COVID-19 and analyzed clinical data from our health system database of more than 3300 patients. Using a machine learning algorithm, we identified a prominent signature of neutrophil activation, including resistin, lipocalin-2, hepatocyte growth factor, interleukin-8, and granulocyte colony-stimulating factor, which were the strongest predictors of critical illness. Evidence of neutrophil activation was present on the first day of hospitalization in patients who would only later require transfer to the intensive care unit, thus preceding the onset of critical illness and predicting increased mortality. In the health system database, early elevations in developing and mature neutrophil counts also predicted higher mortality rates. Altogether, these data suggest a central role for neutrophil activation in the pathogenesis of severe COVID-19 and identify molecular markers that distinguish patients at risk of future clinical decompensation.

Pathologic immune hyperactivation is emerging as a key feature of critical illness in COVID-19, but the mechanisms involved remain poorly understood. We carried out proteomic profiling of plasma from cross-sectional and longitudinal cohorts of hospitalized patients with COVID-19 and analyzed clinical data from our health system database of more than 3300 patients. Using a machine learning algorithm, we identified a prominent signature of neutrophil activation, including resistin, lipocalin-2, hepatocyte growth factor, interleukin-8, and granulocyte colony-stimulating factor, which were the strongest predictors of critical illness. Evidence of neutrophil activation was present on the first day of hospitalization in patients who would only later require transfer to the intensive care unit, thus preceding the onset of critical illness and predicting increased mortality. In the health system database, early elevations in developing and mature neutrophil counts also predicted higher mortality rates. Altogether, these data suggest a central role for neutrophil activation in the pathogenesis of severe COVID-19 and identify molecular markers that distinguish patients at risk of future clinical decompensation.

The core pathology of coronavirus disease 2019 (COVID-19) is infection of airway cells by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that results in excessive inflammation and respiratory disease, with cytokine storm and acute respiratory distress syndrome implicated in the most severe cases. Thrombotic complications are a major cause of morbidity and mortality in patients with COVID-19. Patients with pre-existing cardiovascular disease and/or traditional cardiovascular risk factors, including obesity, diabetes mellitus, hypertension and advanced age, are at the highest risk of death from COVID-19. In this Review, we summarize new lines of evidence that point to both platelet and endothelial dysfunction as essential components of COVID-19 pathology and describe the mechanisms that might account for the contribution of cardiovascular risk factors to the most severe outcomes in COVID-19. We highlight the distinct contributions of coagulopathy, thrombocytopathy and endotheliopathy to the pathogenesis of COVID-19 and discuss potential therapeutic strategies in the management of patients with COVD-19. Harnessing the expertise of the biomedical and clinical communities is imperative to expand the available therapeutics beyond anticoagulants and to target both thrombocytopathy and endotheliopathy. Only with such collaborative efforts can we better prepare for further waves and for future coronavirus-related pandemics.

Fibrosis is the abnormal deposition of extracellular matrix, which can lead to organ dysfunction, morbidity, and death. The disease burden caused by fibrosis is substantial, and there are currently no therapies that can prevent or reverse fibrosis. Metabolic alterations are increasingly recognized as an important pathogenic process that underlies fibrosis across many organ types. As a result, metabolically targeted therapies could become important strategies for fibrosis reduction. Indeed, some of the pathways targeted by antifibrotic drugs in development — such as the activation of transforming growth factor-β and the deposition of extracellular matrix — have metabolic implications. This Review summarizes the evidence to date and describes novel opportunities for the discovery and development of drugs for metabolic reprogramming, their associated challenges, and their utility in reducing fibrosis. Fibrotic therapies are potentially relevant to numerous common diseases such as cirrhosis, non-alcoholic steatohepatitis, chronic renal disease, heart failure, diabetes, idiopathic pulmonary fibrosis, and scleroderma.

We present the first reported case showing metastatic giant bone cell tumor being treated successfully with pembrolizumab after failing prior tyrosine kinase inhibitor therapy. Of note, the patient developed multiple systemic effects associated with checkpoint inhibitor use. One year after starting the checkpoint inhibitor (ICI), the patient also developed hepatitis that was confirmed by liver biopsy and pathology to be, in part, due to drug-mediated toxicity similar to prior ICI toxicity cases that have been reported. Additionally, although the patient had vascular risk factors (hypertension, diabetes and smoking), it was notable from a cardiology perspective that the patient developed 2 subsequent non–ST-elevation myocardial infarctions, with rapid progression of stenosis of the left circumflex artery 2 months apart. The first left heart catheterization showing minimal disease of the left circumflex, but 2 months later, presenting with chest pain, a repeat left heart catheterization showed significant stenosis of the left proximal circumflex, raising the possibilities that either ICI can promote plaque rupture and/or accelerated atherosclerosis; both phenomena have been shown to occur in animal models. The patient also developed thyroiditis with subsequent hypothyroidism, now on thyroid replacement from checkpoint inhibitor use. This case demonstrates the multiorgan adverse effects this new antioncologic agent can have and yet also its promising antitumor effects. Awareness of the side effects among primary care doctors and all specialists will be helpful in managing these potential side effects and research will help elucidate ways to prevent the adverse effects.

Renal transplant outcomes have been shown to be impacted by ethnicity. Prior studies have evaluated the disparate transplant outcomes of Black recipients and recipients of Black donors. However, it has remained unclear whether other donor ethnicities independent of medical comorbidities can influence transplant outcomes. Utilizing the Scientific Registry of Transplant Recipients (SRTR) (with greater than 100,000 patients), we evaluated the effect of each ethnicity, Black, American Indian, Hispanic, Native Hawaiian or other Pacific Islander, and Asian as compared to White recipients on adverse kidney transplant outcomes, assessing for delayed graft function, positive urine protein, acute rejection, and graft failure. Additionally, we assessed the interplay of donor ethnicity on recipient transplant outcomes, which has not previously been comprehensively examined. Logistic regression analysis that took into consideration gender, age, comorbidities, graft type, donor ethnicity, body mass index (BMI), HLA mismatch, ever been on hemodialysis, and time on dialysis indicates that Black recipients have worse outcomes compared to Whites in all outcomes assessed. A logistic regression analysis showed that recipient ethnicity was an independent risk factor for adverse outcomes. Notably, we found that donor ethnicity also independently affects graft outcomes. Hispanic donors lead to better outcomes in Whites and Blacks, while Asian donors have the best outcomes amongst Asian recipients. Recipients of Black donors fared the worst of all ethnicity donors. These data suggest the potential importance of risk stratification for the donor allograft and developing risk calculators that include both donor and recipient ethnicity may be useful, which the current Kidney Donor Profile Index (KDPI) does not currently take into account as they give black donors a different weight, but the same score is assigned to Whites, Asians, and Hispanics.

Waning immunity and emerging variants necessitate continued vaccination against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Improvements in vaccine safety, tolerability, and ease of manufacturing would benefit these efforts. Here, we develop a potent and easily manufactured nanoparticle vaccine displaying the spike receptor-binding domain (RBD). Computational design to stabilize the RBD, eliminate glycosylation, and focus the immune response to neutralizing epitopes results in an RBD immunogen that resolves issues hindering the efficient nanoparticle display of the native RBD. This non-glycosylated RBD can be genetically fused to diverse single-component nanoparticle platforms, maximizing manufacturing ease and flexibility. All engineered RBD nanoparticles elicit potently neutralizing antibodies in mice that far exceed monomeric RBDs. A 60-copy particle (noNAG-RBD-E2p) also elicits potently neutralizing antibodies in non-human primates. The neutralizing antibody titers elicited by noNAG-RBD-E2p are comparable to a benchmark stabilized spike antigen and reach levels against Omicron BA.5 that suggest that it would provide protection against emerging variants.

The Learning from Experience: Improving the Process of Internationally Educated Nurses’ Application for Registrationresearch project, which was led by the College and Association of Registered Nurses of Alberta (CARNA), was the first study to use quantitative data to determine which characteristics of internationally educated nurse (IEN) applicants lead to successful nursing licensure. The analysis from the project led to the development of registration policies and processes that can bridge the gaps between IEN competencies and those competencies required by the licensing body in Alberta, Canada. This article describes those policies and processes—which include an assessment criteria checklist and new timelines for IEN applications—and how they were developed. These changes align with the Pan-Canadian Framework for the Assessment and Recognition of Foreign Qualifications.

Intro:Angiotensin-II (Ang-II) plays an important role in the pathophysiology of various cardiovascular disorders, including cardiomyopathy and hypertension. Angiotensin II type 1 receptor (AT1R) mediates the deleterious effects of Ang-II and can cause vasoconstriction, vascular and cardiac remodeling, and cell death. Animal models have shown that the AT1R signal is obligatory for the development of virus-induced myocardial injury through the proinflammatory action of Ang-II. Data from animal studies and human case studies showed AT1R autoantibodies (AT1R A) were a novel mediator of vasomotor changes and LV dysfunction related to acute myocarditis and that blocking AT1R can be effective.Hypothesis:Little is known whether patients on immune checkpoint inhibitors (ICI) develop AT1R A and whether this can worsen the severity of ICI myocarditis. We measured AT1R A in 9 hospitalized patients who were diagnosed with ICI induced myocarditis. We describe this cohort, the course of their illness and their response to steroids.Methods:This was a prospective study. Quantitative antibodies to AT1R at the time of hospitalization were measured by an Enzyme Linked Immunosorbent Assay (ELISA) (One Lambda) and classified as positive (>17U/mL); at risk (10-17 U/mL) and negative (<10 U/mL).Results:Demographic and co-morbidity details are shown (Table 1). 5/9 patients (55%) were either positive or at risk for AT1R A elevation. 4/9 patients (45%) were AT1R A negative. 4/5 (80%) of the AT1R A positive/at risk required increased immunosuppression whereas 1/4 (25%) of the AT1R A negative required increased immunosuppression for myocarditis.Conclusions:AT1R autoantibodies may worsen the severity of ICI myocarditis. A larger cohort study and comparisons of AT1R A levels in non-ICI patients are underway. If trends hold true, testing for AT1R A and treating with an angiotensin receptor blocker may be a potential strategy to reduce ICI myocarditis severity.

While arginine deiminase (ADI) from Mycoplasma arginini is a well-known anticancer agent, very little is known about any such role of ADI from other bacteria. Additionally, M. arginini ADI is believed to exert its anticancer activity due to depletion of arginine from cancer cells. In this report, we demonstrate anticancer activity of ADI from Pseudomonas aeruginosa. We have also cloned and expressed a truncated form from the P. aeruginosa ADI which harbors a caspase recruitment domain (CARD). This polypeptide, called Pa-CARD, demonstrates anticancer activity without exhibiting any ADI enzymatic activity by inducing apoptosis in cancer cells but not in normal cells. Microarray experiments suggest that Pa-CARD modulates NF-kB signaling pathway genes to exert its anticancer activity.

Several biological phenomena depend on differential methylation of chromosomal strands. While understanding the role of these processes requires information on allele-specific methylation, the available methodologies are not quantitative or labor-intensive. We describe a novel, rapid method to quantitate allele-specific DNA methylation based on the combination of bisulfite PCR and Pyrosequencing™. In this method, DNA is first treated with sodium bisulfite, which converts cytosine but not 5-methylcytosine to uracil. Genes of interest are subsequently amplified using PCR. Allele-specific methylation can then be determined by pyrosequencing each allele individually using sequencing primers that incorporate single nucleotide polymorphisms (SNPs) that allow differentiation between the two parental alleles. This allele-specific methylation methodology can potentially afford quantitative analyses relevant to the regulation of X chromosome inactivation, allele-specific expression of genes in the immune system, repetitive elements, and genomic imprinting. As an illustration of our new method, we quantitated allele-specific methylation of the differentially methylated region of the H19gene, which is imprinted. Although we could reliably determine allele-specific methylation with our technique, additional studies will be required to confirm the ability of our assay to measure loss of imprinting.

Increase in thrombotic and microvascular complications is emerging to be a key feature of patients with critical illness associated with COVID-19 infection. While endotheliopathy is thought to be a key factor of COVID-19-associated coagulopathy, markers indicative of this process that are prognostic of disease severity have not been well-established in this patient population. Using plasma profiling of patients with COVID-19, we identified circulating markers that segregated with disease severity: markers of angiogenesis (VEGF-A, PDGF-AA and PDGF-AB/BB) were elevated in hospitalized patients with non-critical COVID-19 infection, while markers of endothelial injury (angiopoietin-2, FLT-3L, PAI-1) were elevated in patients with critical COVID-19 infection. In survival analysis, elevated markers of endothelial injury (angiopoietin-2, follistatin, PAI-1) were strongly predictive of in-hospital mortality. Our findings demonstrate that non-critical and critical phases of COVID-19 disease may be driven by distinct mechanisms involving key aspects of endothelial cell function, and identify drivers of COVID-19 pathogenesis and potential targets for future therapies.

During pregnancy, Plasmodium falciparum-infected erythrocytes (IE) accumulate in the intervillous spaces of the placenta by binding to chondroitin sulfate A (CSA) and elicit inflammatory responses that are associated with poor pregnancy outcomes. Primigravidae lack immunity to IE that sequester in the placenta and thus are susceptible to placental malaria (PM).

MicroRNAs (miRNAs) are small non-coding RNA molecules that have key regulatory roles in cancer, acting as both oncogenes and tumor suppressors. Due to the potential roles of miRNAs in improving cancer prognostic, predictive, diagnostic and therapeutic approaches, they have become an area of intense research focus in recent years. MiRNAs harbor attractive features allowing for translation to the clinical world, such as relatively simple extraction methods, resistance to molecular degradation, and ability to be quantified. Numerous prognostic, predictive and diagnostic miRNA signatures have been developed. To date however, miRNA analysis has not been adopted for routine clinical use. The objectives of this article are to provide an overview of miRNA research and review a selection of miRNA studies in breast cancer, cervical cancer, sarcoma, and nasopharyngeal carcinoma to highlight advances and challenges in miRNA cancer research.

5-azacytidine (azacitidine, Vidaza) and 5-aza-2′deoxycytidine (decitabine, Dacogen) are cytidine analogs currently used to treat myelodysplastic syndrome and as an investigational agent to treat acute myelogenous leukemia, sickle cell disease and other cancers. Decitabine is phosphorylated to a triphosphate form and incorporated into DNA where it can covalently trap DNA methyltransferase. Azacitidine is also phosphorylated to a triphosphate form, but requires reduction from a ribonucleotide to a deoxyribonucleotide by ribonucleotide reductase (RR) before incorporation into DNA. Hydroxyurea (Hydrea) is a drug routinely used to treat patients with leukemia and sickle cell disease. Hydroxyurea inhibits cell proliferation by inhibiting RR. Inhibition of RR leads to a depletion of nucleotide pools and cell cycle arrest in S-phase. Therefore, we hypothesized that the combination of hydroxyurea with azacitidine would be antagonistic to the ability of azacitidine to inhibit DNA methylation, and synergistic with decitabine in inhibiting DNA methylation. Hydroxyurea inhibits RR, which could block the conversion of azacitidine from a ribonucleotide to a deoxyribonucleotide, and therefore prevent its incorporation into DNA. In contrast, decitabine could increase its incorporation into DNA by taking advantage of a depletion of other deoxynucleotides induced RR inhibition by hydroxyurea. In order to test this hypothesis, we treated HL-60 (leukemia) and T24 (bladder cancer) cell lines with hydroxyurea in combination with either decitabine or azacitidine. DNA methylation was quantitated using Bisulfite-PCR Pyrosequencing of LINE-1 DNA repetitive elements, the MAGE-A1 gene and the p16 gene promoter region. We clearly showed that hydroxyurea at concentrations above 0.5 mM completely blocked the ability of azacitidine to inhibit DNA methylation (IC50=0.36 ± 0.04 mM). Surprisingly, we found that hydroxyurea at concentrations above 0.5mM also completely inhibited the effect of decitabine on DNA methylation (IC50=0.24 ± 0.06 mM). Further investigation showed that inhibition of the ability to block DNA methylation correlated with inhibition of the cell cycle. Using flow cytometry we clearly show that the antagonistic effect of hydroxyurea on DNA methylation inhibitors correlates with the arresting of cell division in S-phase. Aphidicolin, which inhibits DNA polymerase, also blocked the cell cycle and the ability of azacitidine and decitabine to inhibit DNA methylation. Thus, azacitidine and decitabine are S-phase specific drugs and are antagonized when used in combination with drugs that arrest the cell cycle. However, we also show that sequential treatment of hydroxyurea followed by azacitidine or decitabine could be used to avoid this antagonism. Therefore, due to the antagonistic effect of hydroxyurea with the mechanistic inhibitors of DNA methylation, azacitidine and decitabine, we recommend sequential combination treatment strategy.

5-azacytidine (azacitidine, Vidaza) and 5-aza-2′deoxycytidine (decitabine, Dacogen) are cytidine analogs currently used to treat myelodysplastic syndrome and as an investigational agent to treat acute myelogenous leukemia, sickle cell disease and other cancers. Decitabine is phosphorylated to a triphosphate form and incorporated into DNA where it can covalently trap DNA methyltransferase. Azacitidine is also phosphorylated to a triphosphate form, but requires reduction from a ribonucleotide to a deoxyribonucleotide by ribonucleotide reductase (RR) before incorporation into DNA. Hydroxyurea (Hydrea) is a drug routinely used to treat patients with leukemia and sickle cell disease. Hydroxyurea inhibits cell proliferation by inhibiting RR. Inhibition of RR leads to a depletion of nucleotide pools and cell cycle arrest in S-phase. Therefore, we hypothesized that the combination of hydroxyurea with azacitidine would be antagonistic to the ability of azacitidine to inhibit DNA methylation, and synergistic with decitabine in inhibiting DNA methylation. Hydroxyurea inhibits RR, which could block the conversion of azacitidine from a ribonucleotide to a deoxyribonucleotide, and therefore prevent its incorporation into DNA. In contrast, decitabine could increase its incorporation into DNA by taking advantage of a depletion of other deoxynucleotides induced RR inhibition by hydroxyurea. In order to test this hypothesis, we treated HL-60 (leukemia) and T24 (bladder cancer) cell lines with hydroxyurea in combination with either decitabine or azacitidine. DNA methylation was quantitated using Bisulfite-PCR Pyrosequencing of LINE-1 DNA repetitive elements, the MAGE-A1 gene and the p16 gene promoter region. We clearly showed that hydroxyurea at concentrations above 0.5 mM completely blocked the ability of azacitidine to inhibit DNA methylation (IC50=0.36 ± 0.04 mM). Surprisingly, we found that hydroxyurea at concentrations above 0.5mM also completely inhibited the effect of decitabine on DNA methylation (IC50=0.24 ± 0.06 mM). Further investigation showed that inhibition of the ability to block DNA methylation correlated with inhibition of the cell cycle. Using flow cytometry we clearly show that the antagonistic effect of hydroxyurea on DNA methylation inhibitors correlates with the arresting of cell division in S-phase. Aphidicolin, which inhibits DNA polymerase, also blocked the cell cycle and the ability of azacitidine and decitabine to inhibit DNA methylation. Thus, azacitidine and decitabine are S-phase specific drugs and are antagonized when used in combination with drugs that arrest the cell cycle. However, we also show that sequential treatment of hydroxyurea followed by azacitidine or decitabine could be used to avoid this antagonism. Therefore, due to the antagonistic effect of hydroxyurea with the mechanistic inhibitors of DNA methylation, azacitidine and decitabine, we recommend sequential combination treatment strategy.