Alpha and Beta Type 1 Interferon Signaling: Passage for Diverse Biologic Outcomes.

Type I interferon (IFN-I) elicits a complex cascade of events in response to microbial infection. Here, we review recent developments illuminating the large number of IFN-I species and describing their unique biologic functions.

Methylome-wide Analysis of Chronic HIV Infection Reveals Five-Year Increase in Biological Age and Epigenetic Targeting of HLA.

HIV-infected individuals are living longer on antiretroviral therapy, but many patients display signs that in some ways resemble premature aging. To investigate and quantify the impact of chronic HIV infection on aging, we report a global analysis of the whole-blood DNA methylomes of 137 HIV+ individuals under sustained therapy along with 44 matched HIV- individuals. First, we develop and validate epigenetic models of aging that are independent of blood cell composition. Using these models, we find that both chronic and recent HIV infection lead to an average aging advancement of 4.9 years, increasing expected mortality risk by 19%. In addition, sustained infection results in global deregulation of the methylome across >80,000 CpGs and specific hypomethylation of the region encoding the human leukocyte antigen locus (HLA). We find that decreased HLA methylation is predictive of lower CD4 / CD8 T cell ratio, linking molecular aging, epigenetic regulation, and disease progression.

Phenotypic and functional characterization of the CD6-ALCAM T-cell co-stimulatory pathway after allogeneic cell transplantation.

CD6 is a co-stimulatory receptor expressed on T cells that binds activated leukocyte cell adhesion molecule (ALCAM), expressed on antigen presenting cells, epithelial and endothelial tissues. The CD6-ALCAM pathway plays an integral role in modulating T-cell activation, proliferation, and trafficking. In this study we examined expression of CD6 by reconstituting T cells in 95 patients after allogeneic cell transplantation and evaluated the effects of itolizumab, an anti- CD6 monoclonal antibody, on T-cell activation. CD6 T cells reconstituted early after transplant with CD4 regulatory T cells (Treg)-expressing lower levels of CD6 compared to conventional CD4 T cells (Tcon) and CD8 T cells. After onset of acute graft-versus-host disease (aGvHD), CD6 expression was further reduced in Treg and CD8 T cells compared to healthy donors, while no difference was observed for Tcon. ALCAM expression was highest in plasmacytoid dendritic cells (pDC), lowest in myeloid dendritic cells (mDC) and intermediate in monocytes and was generally increased after aGvHD onset. Itolizumab inhibited CD4 and CD8 T-cell activation and proliferation in preGvHD samples, but inhibition was less prominent in samples collected after aGvHD onset, especially for CD8 T cells. Functional studies showed that itolizumab did not mediate direct cytolytic activity or antibody-dependent cytotoxicity in vitro. However, itolizumab efficiently abrogated the costimulatory activity of ALCAM on T-cell proliferation, activation and maturation. Our results identify the CD6-ALCAM pathway as a potential target for aGvHD control and a phase I/II study using itolizumab as first line treatment in combination with steroids for patients with aGvHD is currently ongoing (clinicaltrials gov. Identifier: NCT03763318).

Residues K465 and G467 within the Cytoplasmic Domain of GP2 Play a Critical Role in the Persistence of Lymphocytic Choriomeningitis Virus in Mice.

Several arenaviruses, chiefly Lassa virus (LASV), cause hemorrhagic fever disease in humans and pose serious public health concerns in their regions of endemicity. Moreover, mounting evidence indicates that the worldwide-distributed prototypic arenavirus, lymphocytic choriomeningitis virus (LCMV), is a neglected human pathogen of clinical significance. We have documented that a recombinant LCMV containing the glycoprotein (GPC) gene of LASV within the backbone of the immunosuppressive clone 13 (Cl-13) variant of the Armstrong strain of LCMV (rCl-13/LASV-GPC) exhibited Cl-13-like growth properties in cultured cells, but in contrast to Cl-13, rCl-13/LASV-GPC was unable to establish persistence in immunocompetent adult mice, which prevented its use for some in vivo experiments. Recently, V459K and K461G mutations within the GP2 cytoplasmic domain (CD) of rCl-13/LASV-GPC were shown to increase rCl-13/LASV-GPC infectivity in mice. Here, we generated rCl-13(GPC/VGKS) by introducing the corresponding revertant mutations K465V and G467K within GP2 of rCl-13 and we show that rCl-13(GPC/VGKS) was unable to persist in mice. K465V and G467K mutations did not affect GPC processing, virus RNA replication, or gene expression. In addition, rCl-13(GPC/VGKS) grew to high titers in cultured cell lines and in immunodeficient mice. Further analysis revealed that rCl-13(GPC/VGKS) infected fewer splenic plasmacytoid dendritic cells than rCl-13, yet the two viruses induced similar type I interferon responses in mice. Our findings have identified novel viral determinants of Cl-13 persistence and also revealed that virus GPC-host interactions yet to be elucidated critically contribute to Cl-13 persistence. IMPORTANCE: The prototypic arenavirus, lymphocytic choriomeningitis virus (LCMV), provides investigators with a superb experimental model system to investigate virus-host interactions. The Armstrong strain (ARM) of LCMV causes an acute infection, whereas its derivative, clone 13 (Cl-13), causes a persistent infection. Mutations F260L and K1079Q within GP1 and L polymerase, respectively, have been shown to play critical roles in Cl-13's ability to persist in mice. However, there is an overall lack of knowledge about other viral determinants required for Cl-13's persistence. Here, we report that mutations K465V and G467K within the cytoplasmic domain of Cl-13 GP2 resulted in a virus, rCl-13(GPC/VGKS), that failed to persist in mice despite exhibiting Cl-13 wild-type-like fitness in cultured cells and immunocompromised mice. This finding has uncovered novel viral determinants of viral persistence, and a detailed characterization of rCl-13(GPC/VGKS) can provide novel insights into the mechanisms underlying persistent viral infection.

Type I interferon is a therapeutic target for virus-induced lethal vascular damage.

The outcome of a viral infection reflects the balance between virus virulence and host susceptibility. The clone 13 (Cl13) variant of lymphocytic choriomeningitis virus--a prototype of Old World arenaviruses closely related to Lassa fever virus--elicits in C57BL/6 and BALB/c mice abundant negative immunoregulatory molecules, associated with T-cell exhaustion, negligible T-cell-mediated injury, and high virus titers that persist. Conversely, here we report that in NZB mice, despite the efficient induction of immunoregulatory molecules and high viremia, Cl13 generated a robust cytotoxic T-cell response, resulting in thrombocytopenia, pulmonary endothelial cell loss, vascular leakage, and death within 6-8 d. These pathogenic events required type I IFN (IFN-I) signaling on nonhematopoietic cells and were completely abrogated by IFN-I receptor blockade. Thus, IFN-I may play a prominent role in hemorrhagic fevers and other acute virus infections associated with severe vascular pathology, and targeting IFN-I or downstream effector molecules may be an effective therapeutic approach.

IL-10: achieving balance during persistent viral infection.

The clearance of viral infections is reliant on the coordination and balance of inflammatory factors necessary for viral destruction and immunoregulatory mechanisms necessary to prevent host pathology. In the case of persistent viral infections, immunoregulatory pathways prevent the immune response from clearing the virus, resulting in a long-term equilibrium between host and pathogen. Consequently, negative immune regulators are being considered as a therapeutic target to treat persistent and chronic viral infections. In this review, we will highlight the current understanding of the important negative immune regulator interleukin-10 (IL-10) in persistent viral infection. Though its main role for the host is to limit immune-mediated pathology, IL-10 is a multifunctional cytokine that differentially regulates a number of different hematopoietic cell types. IL-10 has been shown to play a role in a number of infectious diseases and many viral pathogens specifically exploit the IL-10 pathway to help evade host immunity. Recent advances have demonstrated that manipulation of IL-10 signaling during persistent viral infection can alter T cell responses in vivo and that this manipulation can lead to the clearance of persistent viral infection. Furthermore, there have been crucial advances in the understanding of factors that induce IL-10. We summarize lessons learned about IL-10 in model organisms and human persistent infections and conclude with the potential use of IL-10 to treat persistent viral infections.

Infected CD8α- dendritic cells are the predominant source of IL-10 during establishment of persistent viral infection.

Interleukin-10 (IL-10) is an important factor involved in T-cell dysfunction during persistent viral infection. Although several factors can negatively regulate T-cell activity, targeting of the IL-10 pathway alone is sufficient to regenerate T-cell activity and increase viral control. How IL-10 mediates these effects is unclear. Here, we investigated the cellular source of IL-10 necessary for establishing T-cell exhaustion and viral persistence, using IL-10 reporter mice (VertX), cell-type-specific IL-10 and IL-10 receptor deletion mice, and bone marrow chimeric mice. During establishment of viral persistence, the cellular subset with the most prevalent expression of IL-10 was CD8α(-)CD4(+) dendritic cells (DCs), which produced IL-10 with increasing kinetics until 9 d postinfection. After this time point, DCs exhibited a modest decline in percentage of IL-10(+) cells whereas B cells and CD4(+) T cells increased minimally. Further analysis of the DC population demonstrated that IL-10 was primarily expressed in infected DCs. These DCs were a notable source of IL-10 as mutant mice with a DC-specific deletion of IL-10 had significantly decreased serum levels. Interestingly, viral infection was not directly causative of IL-10 expression; rather, IL-10 production appeared to be linked to type I IFN signaling. Our findings further illuminate the contribution of DCs to the production of IL-10 and to viral persistence.

Immortalized clones of fibroblastic reticular cells activate virus-specific T cells during virus infection.

Fibroblastic reticular cells (FRCs) are lymphoid stromal cells essential to T-cell migration and survival. Although FRCs are targets of multiple viral infections, little is known about their role during infection due to the cells' scarcity and difficulty in isolating in vivo. To initiate studies of interactions among FRCs, viruses, and immune cells, we isolated and immortalized CD45(-)gp38(+)CD35(-)CD31(-)CD44(+)VCAM1(+) cell lines from C57BL/6 mice designated as immortalized FRC. Using these cloned cell lines, we have established that FRCs express the major histocompatibility complex (MHC) II molecule, a factor necessary for stimulation of CD4(+) T cells thought to be expressed primarily by antigen-presenting cells, along with other T-cell stimulatory ligands in an IFN-γ-dependent manner. In this environment, lymphocytic choriomeningitis virus (LCMV)-infected iFRCs activated naive LCMV-specific CD4(+) and CD8(+) T cells while limiting expansion of effector LCMV-specific T cells. Thus, FRCs effectively presented antigen along with activating signals during viral infection using both MHC I and MHC II molecules, illustrating a previously undescribed interaction with CD4(+) T cells and indicating a unique role for FRCs.

131 I Uptake in a Left Cerebral Infarct on Posttherapy Whole-Body Scan.

ABSTRACT: A 75-year-old woman with papillary thyroid carcinoma who underwent 131 I radioiodine treatment was incidentally found to have an established left cerebral infarct demonstrating 131 I uptake on posttherapy whole-body scan. False-positive iodine accumulation can occur in benign processes and other malignancies, necessitating awareness among nuclear medicine physicians to avoid misdiagnosing metastatic disease. SPECT/CT can be utilized to enhance diagnostic accuracy when needed.

Safety and Efficacy of 90Y Selective Internal Radiation Therapy Using Glass Microspheres in Hepatocellular Carcinoma: A Southeast Asian Single-Institution Initial Experience.

Our objective was to demonstrate primarily the safety and secondarily the efficacy of 90Y glass microspheres in selective internal radiation therapy (SIRT) for hepatocellular carcinoma (HCC) in a local Southeast Asian hospital. Methods: Eleven consecutive patients with small, unresectable, nonmetastatic HCC and referred for locoregional therapy with SIRT with a curative intention were followed up for 6 mo after the procedure by way of interviews, blood tests, and anatomic scans. Results: Although 5 patients had deranged liver function tests after the procedure, in only 1 patient did this constitute a grade 1 toxicity (in alkaline phosphatase) by the Common Terminology Criteria for Adverse Events. Half the patients showed a reduction in serum α-fetoprotein measurements, and 6 of 11 patients demonstrated an objective response (complete or partial) on imaging. Conclusion: SIRT with 90Y glass microspheres is a safe and efficacious locoregional therapy for unresectable HCC. There are similar articles published in the West; however, the patient population there comprises far fewer Asians and the underlying cause for HCC is different from that in the Asian population. Despite these differences, SIRT is an equally effective and safe option for such patients.

Optimization of Bayesian penalized likelihood reconstruction for 68 Ga-prostate-specific membrane antigen-11 PET/computed tomography.

OBJECTIVE: The objective of this study is to determine the optimal ß value for clinical use in digital 68 Ga-prostate-specific membrane antigen (PSMA-11) PET/computed tomography (CT) imaging. METHODS: 68 Ga PSMA PET/CT of 21 patients with prostate cancer were reconstructed using block-sequential regularized expectation maximization ( ß value of 400-1600) and ordered subsets expectation maximization. Nine independent blinded readers evaluated each reconstruction for overall image quality, noise level and lesion detectability. Maximum standardized uptake value (SUVmax) of the most intense lesion, liver SUVmean and liver SUV SD were recorded. Lesions were then subdivided according to uptake and size; the SUVmax of these lesions were analyzed. RESULTS: There is a statistically significant correlation between improvement in image quality and ß value, with the best being ß 1400. This trend was also seen in image noise ( P  < 0.001), with the least image noise reported with ß 1400. Lesion detectability was not significantly different between the different ß values ( P  =  0.6452). There was no statistically significant difference in SUVmax of the most intense lesion ( P  = 0.9966) and SUVmean of liver background between the different ß values ( P  = 0.9999); however, the SUV SD of the liver background showed a clear trend, with the lowest with ß 1400 ( P  = 0.0008). There was a decreasing trend observed in SUVmax when ß values increased from 800 to 1400 for all four subgroups, and this decrease was greatest in small and low uptake lesions. CONCLUSION: Bayesian penalized likelihood reconstruction algorithms improve image quality without affecting lesion detectability. A ß value of 1400 is optimal.

Differential effects of team-based learning on clinical reasoning.

BACKGROUND: Clinical reasoning (CR) is the ability to integrate information, knowledge and contextual factors for patient care. Few studies have explored effects of team-based learning (TBL) on neurological CR. This study compared simplified TBL (sTBL) against interactive lectures (IL) for teaching CR in neuroanatomical localisation (NL) and neurological emergencies (NE), assessed using a validated Script Concordance Test (SCT). METHODS: A crossover study was conducted with third- and fifth-year undergraduates, randomly assigned to two groups, from the Yong Loo Lin School of Medicine in Singapore. Group 1 was taught NE with sTBL and NL with IL, whereas Group 2 was taught NL with sTBL and NE with IL. Teaching was conducted sequentially over 3 h followed immediately by the SCT. The primary outcome was the difference in mean SCT scores of NE and NL taught with sTBL versus IL. FINDINGS: A total of 179 students (Group 1, n = 81; Group 2, n = 98) participated. Mean NL SCT scores for students taught with sTBL were significantly higher compared with IL (64.8% vs. 61.7%, mean difference 3.1%, 95% confidence interval [CI] 0.6%-5.5%, p = 0.013); effect size was 0.38. Mean NE SCT scores were similar between students taught with sTBL or IL (66.6% vs. 67.0%, mean difference -0.4%, 95% CI -2.2% to 3.1%, p = 0.75). CONCLUSIONS: sTBL was superior to IL for teaching NL, whereas both methods were comparable for teaching NE. TBL may be suitable for teaching more complex neurological topics involving diagnostic reasoning through development of problem representation, hypothesis generation and illness script selection.

The CD6/ALCAM pathway promotes lupus nephritis via T cell-mediated responses.

T cells are central to the pathogenesis of lupus nephritis (LN), a common complication of systemic lupus erythematosus (SLE). CD6 and its ligand, activated leukocyte cell adhesion molecule (ALCAM), are involved in T cell activation and trafficking. Previously, we showed that soluble ALCAM is increased in urine (uALCAM) of patients with LN, suggesting that this pathway contributes to disease. To investigate, uALCAM was examined in 1038 patients with SLE and LN from 5 ethnically diverse cohorts; CD6 and ALCAM expression was assessed in LN kidney cells; and disease contribution was tested via antibody blockade of CD6 in murine models of SLE and acute glomerulonephritis. Extended cohort analysis offered resounding validation of uALCAM as a biomarker that distinguishes active renal involvement in SLE, irrespective of ethnicity. ALCAM was expressed by renal structural cells whereas CD6 expression was exclusive to T cells, with elevated numbers of CD6+ and ALCAM+ cells in patients with LN. CD6 blockade in models of spontaneous lupus and immune-complex glomerulonephritis revealed significant decreases in immune cells, inflammatory markers, and disease measures. Our data demonstrate the contribution of the CD6/ALCAM pathway to LN and SLE, supporting its use as a disease biomarker and therapeutic target.

Translation of Genotype to Phenotype by a Hierarchy of Cell Subsystems.

Accurately translating genotype to phenotype requires accounting for the functional impact of genetic variation at many biological scales. Here we present a strategy for genotype-phenotype reasoning based on existing knowledge of cellular subsystems. These subsystems and their hierarchical organization are defined by the Gene Ontology or a complementary ontology inferred directly from previously published datasets. Guided by the ontology's hierarchical structure, we organize genotype data into an "ontotype," that is, a hierarchy of perturbations representing the effects of genetic variation at multiple cellular scales. The ontotype is then interpreted using logical rules generated by machine learning to predict phenotype. This approach substantially outperforms previous, non-hierarchical methods for translating yeast genotype to cell growth phenotype, and it accurately predicts the growth outcomes of two new screens of 2,503 double gene knockouts impacting DNA repair or nuclear lumen. Ontotypes also generalize to larger knockout combinations, setting the stage for interpreting the complex genetics of disease.

Evaluation of policies regarding physicians infected with blood-borne pathogens.

OBJECTIVE: Formulating an effective approach to preventing surgeon-to-patient transmission of blood-borne pathogens has been controversial. The objective of our study was to evaluate current community hospital policies, if any, regarding restrictions on surgeons (general surgeons and obstetricians and gynecologists) infected with blood-borne pathogens operating on patients. DESIGN: A survey on hospital policies regarding surgeons infected with blood-borne pathogens was sent to infection control officers at Northern California community hospitals (n = 113). RESULTS: Forty-five hospitals responded to the survey. Of these, only 6 (13.3%) had a policy. Of the 39 (86.7%) that did not have a policy, only 3 hospitals were planning on implementing one. CONCLUSIONS: Many community hospitals are uninterested in instituting a policy regarding the practice of surgeons infected with blood-borne pathogens. Possible reasons include the lack of concern on the individual level, difficulty in defining exposure-prone procedures, and the nature of the relationship between medical staff and community hospitals.

Short report: asymptomatic Cryptosporidium hominis infection among human immunodeficiency virus-infected patients in Tanzania.

Few data exist on the relative importance of individual Cryptosporidium species in acquired immunodeficiency syndrome cryptosporidiosis. We characterized 127 inpatients infected with human immunodeficiency virus (HIV) in Tanzania for their CD4 cell count and by stool analysis, including Cryptosporidium immunofluorescence and polymerase chain reaction-restriction fragment length polymorphism. Cryptosporidium was detected in patients both with and without diarrheal symptoms (defined as > or = 3 liquid stools/day, 11 of 61 versus 11 of 66; P = not significant) and was a marker for low CD4 cell count (median = 124/microL versus 212/microL in Cryptosporidium-negative patients; P < 0.04). Cryptosporidium hominis was the predominant species in this region and was associated with a longer duration of symptoms, a higher rate of asymptomatic infection, and a lower CD4 cell count versus C. parvum-infected patients (P < 0.05). This study suggests there may be important differences in the natural history of Cryptosporidium infection in HIV-infected persons depending on parasite species.

Blockade of interferon Beta, but not interferon alpha, signaling controls persistent viral infection.

Although type I interferon (IFN-I) is thought to be beneficial against microbial infections, persistent viral infections are characterized by high interferon signatures suggesting that IFN-I signaling may promote disease pathogenesis. During persistent lymphocytic choriomeningitis virus (LCMV) infection, IFNα and IFNß are highly induced early after infection, and blocking IFN-I receptor (IFNAR) signaling promotes virus clearance. We assessed the specific roles of IFNß versus IFNα in controlling LCMV infection. While blockade of IFNß alone does not alter early viral dissemination, it is important in determining lymphoid structure, lymphocyte migration, and anti-viral T cell responses that lead to accelerated virus clearance, approximating what occurs during attenuation of IFNAR signaling. Comparatively, blockade of IFNα was not associated with improved viral control, but with early dissemination of virus. Thus, despite their use of the same receptor, IFNß and IFNα have unique and distinguishable biologic functions, with IFNß being mainly responsible for promoting viral persistence.

Treatment outcomes after highly active antiretroviral therapy: a meta-analysis of randomised controlled trials.

This systematic review summarises the evidence for treatment efficacy and tolerability of highly active antiretroviral therapies containing two nucleoside reverse transcriptase inhibitors (NRTI) with a protease inhibitor (PI), compared with two NRTIs alone for the treatment of HIV-1 infection in randomised controlled trials. Three electronic databases (Medline, Embase, and the Cochrane Library) were searched up to December 2003. 16 randomised controlled trials met the inclusion criteria and were included in the analysis from 328 articles screened. The pooled analysis indicated that treatment with two NRTIs with a PI is more effective in achieving viral suppression than two NRTIs alone (relative risk [RR] 3.44, 95% confidence interval [CI] 2.43-4.87). However, the RR for discontinuation of treatment due to adverse events of treatment with two NRTIs with a PI compared with two NRTIs alone was 1.81 (95% CI 1.17-2.79). The benefits of treatment with two NRTIs and a PI are substantial among those who can tolerate the regimen in comparison with treatment with two NRTIs alone.

Networking at the level of host immunity: immune cell interactions during persistent viral infections.

Persistent viral infections are the result of a series of connected events that culminate in diminished immunity and the inability to eliminate infection. By building our understanding of how distinct components of the immune system function both individually and collectively in productive versus abortive responses, new potential therapeutic targets can be developed to overcome immune dysfunction and thus fight persistent infections. Using lymphocytic choriomeningitis virus (LCMV) as a model of a persistent virus infection and drawing parallels to persistent human viral infections such as human immunodeficiency virus (HIV) and hepatitis C virus (HCV), we describe the cellular relationships and interactions that determine the outcome of initial infection and highlight immune targets for therapeutic intervention to prevent or treat persistent infections. Ultimately, these findings will further our understanding of the immunologic basis of persistent viral infection and likely lead to strategies to treat human viral infections.

Persistent LCMV infection is controlled by blockade of type I interferon signaling.

During persistent viral infections, chronic immune activation, negative immune regulator expression, an elevated interferon signature, and lymphoid tissue destruction correlate with disease progression. We demonstrated that blockade of type I interferon (IFN-I) signaling using an IFN-I receptor neutralizing antibody reduced immune system activation, decreased expression of negative immune regulatory molecules, and restored lymphoid architecture in mice persistently infected with lymphocytic choriomeningitis virus. IFN-I blockade before and after establishment of persistent virus infection resulted in enhanced virus clearance and was CD4 T cell-dependent. Hence, we demonstrate a direct causal link between IFN-I signaling, immune activation, negative immune regulator expression, lymphoid tissue disorganization, and virus persistence. Our results suggest that therapies targeting IFN-I may help control persistent virus infections.