Early emergence of T central memory precursors programs clonal dominance during chronic viral infection.

Chronic cytomegalovirus (CMV) infection leads to long-term maintenance of extraordinarily large CMV-specific T cell populations. The magnitude of this so-called 'memory inflation' is thought to mainly depend on antigenic stimulation during the chronic phase of infection. However, by mapping the long-term development of CD8+ T cell families derived from single naive precursors, we find that fate decisions made during the acute phase of murine CMV infection can alter the level of memory inflation by more than 1,000-fold. Counterintuitively, a T cell family's capacity for memory inflation is not determined by its initial expansion. Instead, those rare T cell families that dominate the chronic phase of infection show an early transcriptomic signature akin to that of established T central memory cells. Accordingly, a T cell family's long-term dominance is best predicted by its early content of T central memory precursors, which later serve as a stem-cell-like source for memory inflation.

Fate mapping of single NK cells identifies a type 1 innate lymphoid-like lineage that bridges innate and adaptive recognition of viral infection.

Upon viral infection, natural killer (NK) cells expressing certain germline-encoded receptors are selected, expanded, and maintained in an adaptive-like manner. Currently, these are thought to differentiate along a common pathway. However, by fate mapping of single NK cells upon murine cytomegalovirus (MCMV) infection, we identified two distinct NK cell lineages that contributed to adaptive-like responses. One was equivalent to conventional NK (cNK) cells while the other was transcriptionally similar to type 1 innate lymphoid cells (ILC1s). ILC1-like NK cells showed splenic residency and strong cytokine production but also recognized and killed MCMV-infected cells, guided by activating receptor Ly49H. Moreover, they induced clustering of conventional type 1 dendritic cells and facilitated antigen-specific T cell priming early during MCMV infection, which depended on Ly49H and the NK cell-intrinsic expression of transcription factor Batf3. Thereby, ILC1-like NK cells bridge innate and adaptive viral recognition and unite critical features of cNK cells and ILC1s.

NFAT signaling is indispensable for persistent memory responses of MCMV-specific CD8+ T cells.

Cytomegalovirus (CMV) induces a unique T cell response, where antigen-specific populations do not contract, but rather inflate during viral latency. It has been proposed that subclinical episodes of virus reactivation feed the inflation of CMV-specific memory cells by intermittently engaging T cell receptors (TCRs), but evidence of TCR engagement has remained lacking. Nuclear factor of activated T cells (NFAT) is a family of transcription factors, where NFATc1 and NFATc2 signal downstream of TCR in mature T lymphocytes. We show selective impacts of NFATc1 and/or NFATc2 genetic ablations on the long-term inflation of MCMV-specific CD8+ T cell responses despite largely maintained responses to acute infection. NFATc1 ablation elicited robust phenotypes in isolation, but the strongest effects were observed when both NFAT genes were missing. CMV control was impaired only when both NFATs were deleted in CD8+ T cells used in adoptive immunotherapy of immunodeficient mice. Transcriptome analyses revealed that T cell intrinsic NFAT is not necessary for CD8+ T cell priming, but rather for their maturation towards effector-memory and in particular the effector cells, which dominate the pool of inflationary cells.

Rapid SARS-CoV-2 Adaptation to Available Cellular Proteases.

Recent emergence of SARS-CoV-1 variants demonstrates the potential of this virus for targeted evolution, despite its overall genomic stability. Here we show the dynamics and the mechanisms behind the rapid adaptation of SARS-CoV-2 to growth in Vero E6 cells. The selective advantage for growth in Vero E6 cells is due to increased cleavage efficiency by cathepsins at the mutated S1/S2 site. S1/S2 site also constitutes a heparan sulfate (HS) binding motif that influenced virus growth in Vero E6 cells, but HS antagonist did not inhibit virus adaptation in these cells. The entry of Vero E6-adapted virus into human cells is defective because the mutated spike variants are poorly processed by furin or TMPRSS2. Minor subpopulation that lack the furin cleavage motif in the spike protein rapidly become dominant upon passaging through Vero E6 cells, but wild type sequences are maintained at low percentage in the virus swarm and mediate a rapid reverse adaptation if the virus is passaged again on TMPRSS2+ human cells. Our data show that the spike protein of SARS-CoV-2 can rapidly adapt itself to available proteases and argue for deep sequence surveillance to identify the emergence of novel variants. IMPORTANCE Recently emerging SARS-CoV-2 variants B.1.1.7 (alpha variant), B.1.617.2 (delta variant), and B.1.1.529 (omicron variant) harbor spike mutations and have been linked to increased virus pathogenesis. The emergence of these novel variants highlights coronavirus adaptation and evolution potential, despite the stable consensus genotype of clinical isolates. We show that subdominant variants maintained in the virus population enable the virus to rapidly adapt to selection pressure. Although these adaptations lead to genotype change, the change is not absolute and genomes with original genotype are maintained in the virus swarm. Thus, our results imply that the relative stability of SARS-CoV-2 in numerous independent clinical isolates belies its potential for rapid adaptation to new conditions.

Cytomegalovirus inhibition of extrinsic apoptosis determines fitness and resistance to cytotoxic CD8 T cells.

Viral immune evasion is currently understood to focus on deflecting CD8 T cell recognition of infected cells by disrupting antigen presentation pathways. We evaluated viral interference with the ultimate step in cytotoxic T cell function, the death of infected cells. The viral inhibitor of caspase-8 activation (vICA) conserved in human cytomegalovirus (HCMV) and murine CMV (MCMV) prevents the activation of caspase-8 and proapoptotic signaling. We demonstrate the key role of vICA from either virus, in deflecting antigen-specific CD8 T cell-killing of infected cells. vICA-deficient mutants, lacking either UL36 or M36, exhibit greater susceptibility to CD8 T cell control than mutants lacking the set of immunoevasins known to disrupt antigen presentation via MHC class I. This difference is evident during infection in the natural mouse host infected with MCMV, in settings where virus-specific CD8 T cells are adoptively transferred. Finally, we identify the molecular mechanism through which vICA acts, demonstrating the central contribution of caspase-8 signaling at a point of convergence of death receptor-induced apoptosis and perforin/granzyme-dependent cytotoxicity.

Paradigm shift in heart failure treatment: are cardiologists ready to use gliflozins?

Despite recent advances in chronic heart failure (HF) therapy, the prognosis of HF patients remains poor, with high rates of HF rehospitalizations and death in the early months after discharge. This emphasizes the need for incorporating novel HF drugs, beyond the current approach (that of modulating the neurohumoral response). Recently, new antidiabetic oral medications (sodium-glucose cotransporter 2 inhibitors (SGLT2i)) have been shown to improve prognosis in diabetic patients with previous cardiovascular (CV) events or high CV risk profile. Data from DAPA-HF study showed that dapaglifozin is associated with a significant reduction in mortality and HF hospitalization as compared with placebo regardless of diabetes status. Recently, results from EMPEROR-Reduced HF trial were consistent with DAPA-HF trial findings, showing significant beneficial effect associated with empagliflozin use in a high-risk HF population with markedly reduced ejection fraction. Results from the HF with preserved ejection fraction trials using these same agents are eagerly awaited. This review summarizes the evidence for the use of gliflozins in HF treatment.

Knowledge and Attitude on Eye Donation among Undergraduate Medical Students of Kathmandu University School of Medical Sciences, Nepal.

Background Corneal blindness accounts for a significant proportion of avoidable visual impairment in developing countries. Eye donation is voluntary and awareness in undergraduate medical students being a future practitioner in any field are expected to be linked to patients during death in hospitals. Objective To assess the awareness of medical students on eye donation at Kathmandu University School of Medical Sciences (KUSMS). Method This was a cross-sectional study conducted among undergraduate medical students of KUSMS. Students' responses were recorded using a predesigned, pretested, semistructured questionnaire inquiring knowledge and attitude of eye donation, sources of information, their willingness to donate eyes as well as the reasons for donating/ not donating eyes. Result Less than half of the medical students (45.6%) were aware of eye donation only after death. Newspapers (72.2%) were the major source of information. The final year medical students were more aware (Average knowledge score = 11.56 ± 2.05) than their juniors. 80.7% of the students were willing to donate their eyes. The adjudged reasons for willingness to donate were that eye donation is a noble work and pleasure in helping a blind person while the reasons for unwillingness to donate were lack of awareness followed by family objection to eye donation. Conclusion Future medical practitioners possessed satisfactory knowledge about eye donation. Educating this cadre of human resources to sensitize them towards the need for eye donation would be a crucial step towards reducing the global burden of corneal blindness.

Does the gradual increase in dietary zinc oxide supplementation can affect egg quality, serum indices, and productive performance of laying hens?

This study was conducted to evaluate the influence of increased supplementation of zinc oxide (ZnO) on performance, quality of egg, blood chemistry, and antioxidant ability in serum of laying chickens (Hisex Brown) reared from 22 to 34 weeks of age. Seventy-two 22-week-old laying hens (Hisex Brown) were haphazardly separated into 3 handling collections of 24 chickens (6 replicates per treatment and four laying hens per replicate). Dietary treatments included basal diet without zinc addition for control group while the 2nd and 3rd groups contained basal diet with 25 or 75 mg ZnO/kg diet. Results showed that the higher level of ZnO (75 mg ZnO/kg diet) elevated (P < 0.01) feed intake during all studied periods compared with the control group and other groups that contained ZnO. The handling groups supplied with 75 mg ZnO/kg diet gave the worst feed: egg ratio within the whole period and the intervals compared with the control and other ZnO levels. Supplementation of zinc decreased egg number and egg output when compared with the control groups. Egg quality traits were statistically differed due to dietary ZnO supplementation except egg shape index, yolk %, and albumin %. Supplementation of zinc decreased triglyceride (P = 0.001) of laying hens. The low-density lipoproteins (LDL) cholesterol level in serum was decreased with 75 mg ZnO/kg in comparison with all treatment groups. Zinc supplementation increased the level of serum zinc without differences in supplemented zinc levels. Dietary supplemental zinc did not affect antioxidant parameters in the serum. It is concluded that dietary zinc supplementation up to 75 mg/kg used as effective supplement to enhance zinc status and antioxidant ability and activities in laying hens.

Evidence based treatment for unstable slipped upper femoral epiphysis: Systematic review and exploratory patient level analysis.

The management of slipped upper femoral epiphysis is controversial and evolving as insight into the condition develops. Loder introduced the concept of slip stability and demonstrated a strong association between poor outcome and instability. Almost half of patients with unstable slip developed femoral head osteonecrosis. This has been influential in surgeons' choice of treatments. Some surgeons have adopted a minimal intervention approach such as pinning in situ or gentle reduction and pinning whereas others advocated an urgent open reduction and stabilisation of slip using various surgical techniques. In this review we analysed the influence of various interventions, timing of surgery and severity of the slip on the outcome of unstable slip.

The efficacy of using exogenous enzymes cocktail on production, egg quality, egg nutrients and blood metabolites of laying hens fed distiller's dried grains with solubles.

An experiment was performed using 120 Hisex Brown laying hens for evaluating the effects of different inclusion levels of corn distiller's dried grains with solubles (DDGS) as a replacement of soybean meal (SBM) with or without enzyme cocktail on performance, egg quality, egg nutrients and blood metabolites in laying hens through 22-42 weeks of age. A 4 × 2 factorial design experiment was performed including four substitution levels of DDGS (0, 250, 500 and 750 g/kg respectively) and two enzyme cocktail levels (0 and 250 mg/kg diet). The used enzyme in this study "Gallazyme" composed of xylanase, Trichoderma longibrachiatum (600 units/g), protease, Bacillus subtilis (8,000 units/g) and amylase and Bacillus amyloliquofaciens (800 units/g). The control diet showed the best feed efficiency followed by the intermediate levels of DDGS. The lowest value of feed efficiency was found in the group fed the highest level of DDGS. Enzyme addition improved feed efficiency and decreased laying rate. Increasing DDGS levels was associated with albumin and shell thickness increases. Dietary DDGS depressed all egg components except the organic matter which maximised in enzyme-treated groups. Increasing DDGS level was accompanied by increase in yolk cholesterol and total lipids. No significant impacts were detected with enzymes supplementation on yolk lipids profile. Excepting serum calcium and phosphorous, all serum constituents increased with increasing level of DDGS. Using enzyme markedly depressed serum ammonia by 15.02% and increased calcium by 6.44% compared with enzyme-free diets. Interaction between DDGS and enzyme was significant on most of studied parameters. It could be concluded that using enzyme cocktail in DDGS-based diets may improve feed efficiency and egg quality, in addition to lowering blood ammonia and increasing blood calcium. It is recommended to substitute SBM by DDGS up to 500 g/kg diet.

Effects of phyto-oestrogen quercetin on productive performance, hormones, reproductive organs and apoptotic genes in laying hens.

Quercetin, a polyphenolic flavonoid with diverse biological activities including anti-inflammatory and antiviral, inhibits lipid peroxidation, prevents oxidative injury and cell death. The purpose of the research was to investigate the effect of quercetin on productive performance, reproductive organs, hormones and apoptotic genes in laying hens between 37 and 45 weeks of age, because of the structure and oestrogenic activities similar to 17ß-oestradiol. The trial was conducted using 240 Hessian laying hens (37 weeks old), housed in wire cages with two hens in each cage. These hens were randomly allotted to four treatments with six replicates, 10 hens in each replicate and fed with diets containing quercetin as 0, 0.2, 0.4 and 0.6 g/kg feed for 8 weeks. The results showed that dietary quercetin significantly increased (p < .05) the laying rate and was higher in group supplemented with 0.4 g/kg, and feed-egg ratio was decreased (p < .05) by quercetin. Dietary quercetin has no effect (p > .05) on average egg weight and average daily feed intake. Compared with control, secretion of hormones, oestradiol (E2 ), progesterone (P4), follicle-stimulating hormone (FSH), luteinizing hormone (LH), insulin-like growth factors-1 (IGF-1) and growth hormone (GH), was found to be significantly higher (p < .05) in quercetin-supplemented groups. Also ovary index, uterus index and oviduct index were not significantly influenced (p > .05) by quercetin, whereas magnum index, isthmus index, magnum length, isthmus length and follicle numbers were significantly increased (p < .05) with quercetin supplementation. Additionally, expression of apoptotic genes was significantly (p < .05) up-regulated or down-regulated by quercetin. These results indicated that quercetin improved productive performance, and its mechanism may be due to the oestrogen-like activities of quercetin.

Histological Evolution of BK Virus-Associated Nephropathy: Importance of Integrating Clinical and Pathological Findings.

Long-term clinicopathological studies of BK-associated nephropathy (PyVAN) are not available. We studied 206 biopsies (71 patients), followed 3.09 ± 1.46 years after immunosuppression reduction. The biopsy features (% immunostain for PyV large T ag + staining and inflammation ± acute rejection) were correlated with viral load dynamics and serum creatinine to define the clinicopathological status (PyVCPS). Incidence of acute rejection was 28% in the second biopsy and 50% subsequently (25% mixed T cell-mediated allograft rejection (TCMR) + antibody-mediated allograft rejection (AMR); rejection overall affected 38% of patients (>50% AMR). Graft loss was 15.4% (0.8-5.3 years after PyVAN); 76% had complete viral clearance (mean 28 weeks). The only predictors of graft loss were acute rejection (TCMR p = 0.008, any type p = 0.07), and increased "t" and "ci" in the second biopsy (p = 0.006 and 0.048). Higher peak viremia correlated with poorer viral clearance (p = 0.002). Presumptive and proven PyVAN had similar presentation, evolution, and outcome. Late PyVAN (>2 years, 9.8%) justifies BK viremia evaluation at any point with graft dysfunction and/or biopsy evaluation. This study describes the histological evolution of PyVAN and corresponding clinicopathological correlations. Although the pathological features overall reflect the viral and immunological interactions, the PyVAN course remains difficult to predict based on any single feature. Appropriate clinical management requires repeat biopsies and determination of the PyVCPS at relevant time points, for corresponding personalized immunosuppression adjustment.

Role of Hyperkalemia in Heart Failure and the Therapeutic Use of Potassium Binders.

Hyperkalemia can be a life-threatening disorder, especially for at-risk patients with heart failure, chronic kidney disease, with diabetes, and patients on certain drugs like renin-angiotensin-aldosterone system antagonists and mineralocorticoid receptor antagonists. There are limited therapeutic options available for hyperkalemia, and they have narrow effectiveness because of their unfavorable side effects profile in long-term and high cost utilization requiring inpatient care. Patiromersorbitex calcium and sodium zirconium cyclosilicate are novel potassium-lowering compounds for the treatment and prevention of hyperkalemia in at-risk population. These therapeutic agents have shown encouraging results in early phase II and phase III clinical trials. However, there is need to further study their efficacy and safety in heart failure population in order to establish their clinical use. The focus of this chapter will be to promote better understanding of potassium homeostasis in heart failure patients and the mechanistic overview of novel drugs, with emphasis on heart failure population.

Impact of Site Selection and Study Conduct on Outcomes in Global Clinical Trials.

PURPOSE OF REVIEW: There are over 25 million patients living with heart failure globally. Overall, and especially post-discharge, clinical outcomes have remained poor in heart failure despite multiple trials, with both successes and failures over the last two decades. Matching therapies to the right patient population, identifying high-quality sites, and ensuring optimal trial design and execution represent important considerations in the development of novel therapeutics in this space. RECENT FINDINGS: While clinical trials have undergone rapid globalization, this has come with regional variation in comorbidities, clinical parameters, and even clinical outcomes and treatment effects across international sites. These issues have now highlighted knowledge gaps about the conduct of trials, selection of study sites, and an unmet need to develop and identify "ideal" sites. There is a need for all stakeholders, including academia, investigators, healthcare organizations, patient advocacy groups, industry sponsors, research organizations, and regulatory authorities, to work as a multidisciplinary group to address these problems and develop practical solutions to improve trial conduct, efficiency, and execution. We review these trial-level issues using examples from contemporary studies to inform and optimize the design of future global clinical trials in heart failure.

Utility of positron emission tomography for drug development for heart failure.

Only about 1 in 5,000 investigational agents in a preclinical stage acquires Food and Drug Administration approval. Among many reasons for this includes an inefficient transition from preclinical to clinical phases, which exponentially increase the cost and the delays the process of drug development. Positron emission tomography (PET) is a nuclear imaging technique that has been used for the diagnosis, risk stratification, and guidance of therapy. However, lately with the advance of radiochemistry and of molecular imaging technology, it became evident that PET could help novel drug development process. By using a PET radioligand to report on receptor occupancy during novel agent therapy, it may help assess the effectiveness, efficacy, and safety of such a new medication in an early preclinical stage and help design successful clinical trials even at a later phase. In this article, we explore the potential implications of PET in the development of new heart failure therapies and review PET's application in the respective pathophysiologic pathways such as myocardial perfusion, metabolism, innervation, inflammation, apoptosis, and cardiac remodeling.

Bartonella henselae infection-associated vasculitis and crescentic glomerulonephritis leading to renal allograft loss.

Bartonella henselae (BH) is the main cause of cat scratch disease (CSD), which more typically presents as a self-limited localized suppurative lymphadenopathy in immunocompetent individuals. In contrast, immunocompromised patients commonly have systemic disease with life-threatening complications. In addition to the angioproliferative lesions, such as bacillary angiomatosis, an increasing number of immune post-infectious complications are being recognized with BH infections, including glomerulonephritis, vasculitis, hemophagocytic syndrome, and neurological problems. We report the case of a renal transplant recipient who developed CSD in the second year post transplantation. In addition to prolonged fever and generalized lymphadenopathy and splenomegaly requiring differentiation from a post-transplant lymphoproliferative disorder, the course was complicated by the development of dermal leukocytoclastic vasculitis and pauci-immune necrotizing and crescentic glomerulonephritis, which led to failure of the renal graft. Glomerulonephritis as a complication of CSD has never been described in a kidney allograft, to our knowledge. Awareness of the diverse clinical symptoms associated with BH, including granulomatous/suppurative lesions and other less common complications can lead to more rapid and accurate diagnosis. Also, as recommended by the current guidelines, a thorough history of pet ownership should be part of the clinical evaluation before and after transplantation for all transplant recipients.

Radiation-induced microRNA: discovery, functional analysis, and cancer radiotherapy.

MicroRNAs (miRNAs) are small non-protein coding RNA that play an important role in gene regulation. These RNA molecules function as post-transcriptional regulators. miRNAs bind to complementary sequences on target messenger RNA transcripts, usually resulting in translational repression or target mRNA degradation and gene silencing. miRNA are abundantly present in all human cells, target approximately 60% of all genes, and are able to repress hundreds of targets each. Since their discovery in 1993 miRNA are emerging as important modulators in cellular pathways such as growth and proliferation, apoptosis, carcinogenesis, timing of cell-fate decision, and metabolic pathways. A large number of studies have examined the general and specific effects of miRNAs perturbation in radiation-exposed cells. These studies include expression profiling of miRNA, functional analysis, the role of specific miRNAs in tumor radiosensitivity, and targeting miRNA for improved cancer radiotherapy. Other studies have explored the involvement of miRNA in radiobiological phenomenon like bystander effect. Emerging evidence is establishing that miRNA are involved in regulating radiation-induced cellular processes, can be exploited to improve cancer radiation therapy, and could serve as biomarkers of human radiation exposure.

Risk surveillance with spatial distribution of Organochlorine Pesticides (OCPs) from sedimentary samples of Chenab River.

A pioneering study employed a holistic geostatistical approach to predict the spatial variability of a non sampled area in the Chenab River, Pakistan, using kriging interpolation for organochlorine pesticide (OCP)-polluted risk zones. The Present research intended to investigate the carcinogenic and non-carcinogenic human health risks, contamination levels, and spatial variation of OCPs in the Chenab River, Pakistan. The residual OCP content in sediment samples (n = 120) ranged from 0.056 to 32.14 ng/g. DDE and α-HCH were prevalent among all the samples analyzed, with mean concentrations of 15.84 ± 8.02 and 12.45 ± 6.72 ng/g, respectively. The order of magnitude of OCPs in sediment samples was DDTs > α-HCH > chlorothalonil > heptachlor > endosulfan > aldrin > dieldrin. The findings of the single (SPI) and Nemerow (Nel) pollution index of α-HCH, heptachlor, and aldrin depicted the Chenab River as a serious pollution risk zone. The outcomes of the Pearson correlation coefficient analysis represent the positive correlation among all OCPs, revealing the common origin. Distribution trends showed substantially higher (p < 0.05) contents of analyzed OCPs along the downstream zone. With regards to USEPA human health hazard assessment model, the estimated non-carcinogenic (ΣHI) and non-carcinogenic (ΣTCR) risk ranged from 1.1 × 10-5 to 1.0 × 10-1, 4.0 × 10-8 to 3.2 × 10-4 respectively. TCR >10-4 illustrated a substantial cancer health risk posed by α-HCH, heptachlor, aldrin, and dieldrin in the downstream zone. We recommend the urgent cessation of the ongoing discharge of OCPs into the Chenab River, which needs to be highlighted owing to the significant cancer risk to public health to ensure the good health and wellbeings.