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Summary Natural killer (NK) cell effector functions are dependent on metabolic regulation of cellular function; however, less is known about in vivo metabolic pathways required for NK cell antiviral function. Mice with an inducible NK-specific deletion of Cox10, which encodes a component of electron transport chain complex IV, were generated to investigate the role of oxidative phosphorylation in NK cells during murine cytomegalovirus (MCMV) infection. Ncr1-Cox10Δ/Δ mice had normal numbers of NK cells but impaired expansion of antigen-specific Ly49H+ NK cells and impaired NK cell memory formation. Proliferation in vitro and homeostatic expansion were intact, indicating a specific metabolic requirement for antigen-driven proliferation. Cox10-deficient NK cells upregulated glycolysis, associated with increased AMP-activated protein kinase (AMPK) and mammalian target of rapamycin (mTOR) activation, although this was insufficient to protect the host. These data demonstrate that oxidative metabolism is required for NK cell antiviral responses in vivo.

BackgroundHigh-intensity glucocorticoid regimens are commonly used to induce and maintain remission in Juvenile Dermatomyositis but are associated with several adverse side-effects. Evidence-based treatment guidelines from North American and European pediatric rheumatology research societies both advocate induction with intravenous pulse steroids followed by high dose oral steroids (2 mg/kg/day), which are then tapered. This study reports the time to disease control with reduced glucocorticoid dosing.MethodsWe retrospectively reviewed the records at a single tertiary-care children’s hospital of patients diagnosed with Juvenile Dermatomyositis between 2000 and 2014 who had a minimum of 2 years of follow-up. The primary outcome measure was time to control of muscle and skin disease. Additional outcome measures included glucocorticoid dosing, effect of treatment on height, frequency of calcinosis, and complications from treatment.ResultsOf the 69 patients followed during the study period, 31 fulfilled inclusion criteria. Median length of follow-up was 4.58 years, (IQR 3–7.5). Myositis control was achieved in a median of 7.1 months (IQR 0.9–63.4). Cutaneous disease control was achieved in a median of 16.7 months (IQR 4.3–89.5). The median starting dose of glucocorticoids was 0.85 mg/kg/day, (IQR 0.5–1.74). The median duration of steroid treatment was 9.1 months, (IQR 4.7–17.4), while the median duration of any pharmacotherapy was 29.2 months (IQR 10.4 to 121.3). Sustained disease control off medications was achieved in 21/31 (68%) patients by the end of review. Persistent calcinosis was identified in only one patient (3%).ConclusionCurrent accepted treatment paradigms for Juvenile Dermatomyositis include oral glucocorticoids beginning at 2 mg/kg/day and reduced over a prolonged time period. However, our results suggest that treatment using reduced doses and duration with early use of steroid-sparing agents is comparably effective in achieving favorable outcomes in Juvenile Dermatomyositis.

An unbridled host immune response to SARS-CoV-2 infection likely underlies severe cases of the disease and has been labeled a “Cytokine Storm Syndrome”. We emphasize that categorization of syndromes triggered by a completely novel pathogen based on other seemingly similar, but potentially distinct, known entities is an inherently risky endeavor., Highlights • An uncontrolled host immune response following SARS-CoV-2 infection and a cytokine storm contribute to severe COVID-19. • Among the myriad clinical syndromes identified in COVID-19 patients, Multisystem Inflammatory Syndrome in Children (MIS-C) is emerging as a severe complication in pediatric patients. • Cohorts of pediatric patients with MIS-C reveal overlapping but distinct pictures from SARS-CoV-2 associated Cytokine Storm Syndrome with respect to their clinical and laboratory profile. • While candidates for targeted therapies based on understanding of the pathogenesis of SARS-CoV-2-associated Cytokine Storm Syndromes are emerging, and there may be overlap in the pathogenesis and therapy, it is clear that syndrome-specific approaches will be necessary.

OBJECTIVE The incidence and prevalence of granulomatosis with polyangiitis (GPA) in the US is not well characterized. Owing to its rarity, outcomes data in pediatric-onset GPA are also lacking. The aims of this study were to describe the epidemiology of GPA and outcomes in GPA patients in the US, and to compare outcomes between pediatric and working-age adult patients. METHODS A retrospective cohort study using the 2006-2014 Truven Health Analytics MarketScan Commercial Claims and Encounters Database was conducted. The incidence and prevalence rates of pediatric and adult GPA (age

NKG2D is a key component of cytotoxic antitumor and antiviral responses. Multiple viruses evade NKG2D recognition by blocking NKG2D ligand expression on infected cells. In contrast, cowpox virus targets NKG2D directly by encoding a secreted antagonist, Orthopoxvirus MHC Class I-like Protein (OMCP). We have previously reported that OMCP also binds to the orphan receptor FcRL5 on innate B cells. Here, we demonstrate that mammalian-derived, glycosylated OMCP binds NKG2D but not FcRL5. Cowpox viruses either lacking OMCP, or expressing an NKG2D-binding deficient mutant, are significantly attenuated in wild type and FcRL5-deficient mice but not NKG2D-deficient mice, demonstrating that OMCP is critical in subverting NKG2D-mediated immunityin vivo. Next we determined the structure of OMCP bound to human NKG2D. Despite a structure similar to that of host NKG2D ligands, OMCP uses a drastically different orientation for NKG2D binding. The re-orientation of OMCP is associated with dramatically higher affinity for human NKG2D and the targeted interface is highly conserved in mammalian NKG2Ds, increasing the zoonotic potential of cowpox virus. We also show that cell surface presented OMCP can trigger NKG2D effector functions equivalently to host NKG2D ligands, demonstrating that NKG2D-mediated signaling requires clustering but is insensitive to binding orientation. Thus, in contrast to TCR/MHC interactions, the docking topology of NKG2D with its ligands does not appear to regulate its activation.Author SummaryVirally infected or tumor-transformed cells display NKG2D ligands (NKG2DLs) on their cell surface, which activates NKG2D-bearing lymphocytes to kill the transformed cell. Pathogens are known to counter this by blocking NKG2DL expression and/or surface display. In contrast, some tumor cells cleave endogenous NKG2DLs creating soluble NKG2D antagonists. Unlike other viral pathogens, cowpox virus uses a strategy analogous to cancer cells by targeting NKG2D directly with a soluble, high affinity NKG2D-antagonist named OMCP. We determined that OMCP’s virulencein vivois attributed to blocking NKG2D-mediated NK cell responses with no apparent effect due to binding to other receptors or cell types. We have also determined the crystal structure of cowpox OMCP bound to human NKG2D, revealing that despite conservation of the ligand scaffolding with host NKG2DLs, the viral protein is engaged with a radically altered orientation compared to all host NKG2DLs. Our structure provides key insight into how OMCP binds with an ∼5,000-fold increased affinity compared to human NKG2DLs and show that the OMCP binding site is exceptionally conserved among primates and rodents, suggesting that the ability of OMCP to recognize this conserved interface contributes to the broad zoonotic potential of cowpox virus. Finally, we show that cell membrane-anchored OMCP can trigger equivalent NKG2D-mediated killing as host NKG2DLs, demonstrating that NKG2D signaling is insensitive to ligand binding orientation.

Polyarticular juvenile idiopathic arthritis (JIA) is among the most challenging of the JIA subtypes to treat. Even with current biologic therapies, the disease remains difficult to control in a substantial subset of patients, highlighting the need for new therapies. The aim of this study was to use the high dimensionality afforded by mass cytometry with phospho-specific antibodies to delineate signaling abnormalities in immune cells from treatment-naive polyarticular JIA patients. Peripheral blood mononuclear cells were isolated from 17 treatment-naive polyarticular JIA patients, 10 of the patients after achieving clinical remission, and 19 healthy controls. Samples were stimulated for 15 minutes with IL-6 or IFN-γ and analyzed by mass cytometry. Following IFN-γ stimulation, increased STAT1 and/or STAT3 phosphorylation was observed in subsets of CD4 T cells and classical monocytes from treatment-naive patients. The enhanced IFN-γ signaling was associated with increased expression of JAK1 and SOCS1 in CD4 T cells. Furthermore, substantial heterogeneity in surface marker expression was observed among the subsets of CD4 T cells and classical monocytes with increased IFN-γ responsiveness. The identification of enhanced IFN-γ signaling in CD4 T cells and classical monocytes from treatment-naive polyarticular JIA patients provides mechanistic support for investigations into therapies that attenuate IFN-γ signaling in this disease.

Lupus mesenteric vasculitis (LMV) is a severe and potentially fatal complication of systemic lupus erythematosus (SLE). Although LMV is always a consideration in adolescents and adult patients with SLE, who present with acute abdominal pain, diagnosis and management remain a great challenge. We describe the cases of five patients age 14 to 21 years old diagnosed with LMV. All five patients had active SLE and typical clinical presentation suspicious of LMV. Abdominal CT was the preferred imaging modality and was useful in four patients. Corticosteroids were the mainstay of treatment for all five patients. All five patients survived and complete remission of symptoms was achieved in four out of five patients with the addition of cyclophosphamide and in one out of five with rituximab. A review of the literature was performed including a systemic review of the case reports and case series published in the English literature over the last 20 years.

SummaryThe host immune system functions constantly to maintain chronic commensal and pathogenic organisms in check. The consequences of these immune responses on host physiology are as yet unexplored, and may have long-term implications in health and disease. We show that chronic viral infection increases epithelial turnover in multiple tissues, and the antiviral cytokines type I interferons (IFNs) mediate this response. Using a murine model with persistently elevated type I IFNs in the absence of exogenous viral infection, the Irgm1−/− mouse, we demonstrate that type I IFNs act through nonepithelial cells, including macrophages, to promote increased epithelial turnover and wound repair. Downstream of type I IFN signaling, the highly related IFN-stimulated genes Apolipoprotein L9a and b activate epithelial proliferation through ERK activation. Our findings demonstrate that the host immune response to chronic viral infection has systemic effects on epithelial turnover through a myeloid-epithelial circuit.

NK cell activation has been shown to be metabolically regulated in vitro; however, the role of metabolism during in vivo NK cell responses to infection is unknown. We examined the role of glycolysis in NK cell function during murine cytomegalovirus (MCMV) infection and the ability of IL-15 to prime NK cells during CMV infection. The glucose metabolism inhibitor 2-deoxy-ᴅ-glucose (2DG) impaired both mouse and human NK cell cytotoxicity following priming in vitro. Similarly, MCMV-infected mice treated with 2DG had impaired clearance of NK-specific targets in vivo, which was associated with higher viral burden and susceptibility to infection on the C57BL/6 background. IL-15 priming is known to alter NK cell metabolism and metabolic requirements for activation. Treatment with the IL-15 superagonist ALT-803 rescued mice from otherwise lethal infection in an NK-dependent manner. Consistent with this, treatment of a patient with ALT-803 for recurrent CMV reactivation after hematopoietic cell transplant was associated with clearance of viremia. These studies demonstrate that NK cell-mediated control of viral infection requires glucose metabolism and that IL-15 treatment in vivo can reduce this requirement and may be effective as an antiviral therapy.

Objective.Accumulating evidence links juvenile idiopathic arthritis (JIA) to nonhost factors such as gut microbes. We hypothesize that children with new-onset JIA have increased intestinal bacterial translocation and circulating lipopolysaccharide (LPS).Methods.We studied systemic treatment-naive patients with JIA [polyarticular JIA, n = 22, oligoarticular JIA, n = 31, and spondyloarthropathies (SpA), n = 16], patients with established inflammatory bowel disease–related arthritis (IBD-RA, n = 11), and 34 healthy controls. We determined circulating IgG reactivity against LPS, LPS-binding protein (LBP), α-1-acid glycoprotein (α-1AGP), and C-reactive protein (CRP) in plasma or serum from these patients and controls. Juvenile Arthritis Disease Activity Score (JADAS-27) was calculated for patients with JIA.Results.Circulating anticore LPS antibody concentrations in patients with polyarticular JIA (p = 0.001), oligoarticular JIA (p = 0.024), and SpA (p = 0.001) were significantly greater than in controls, but there were no significant intergroup differences. Circulating LBP concentrations were also significantly greater in patients with polyarticular JIA (p = 0.001), oligoarticular JIA (p = 0.002), and SpA (p = 0.006) than controls, as were α-1AGP concentrations (p = 0.001, 0.001, and 0.003, respectively). No differences were observed between controls and patients with IBD-RA in any of the assays. Circulating concentrations of LBP and α-1AGP correlated strongly with CRP concentrations (r = 0.78 and r = 0.66, respectively). Anticore LPS antibody levels and CRP (r = 0.26), LBP (r = 0.24), and α-AGP (r = 0.22) concentrations had weaker correlations. JADAS-27 scores correlated with LBP (r = 0.66) and α-1AGP concentrations (r = 0.58).Conclusion.Children with polyarticular JIA, oligoarticular JIA, and SpA have evidence of increased exposure to gut bacterial products. These data reinforce the concept that the intestine is a source of immune stimulation in JIA.

Natural killer (NK) cells are cytotoxic type 1 innate lymphoid cells (ILCs) that defend against viruses and mediate anti-tumor responses, yet mechanisms controlling their development and function remain incompletely understood. We hypothesized that the abundantly expressed microRNA-142 (miR-142) is a critical regulator of type 1 ILC biology. Interleukin-15 (IL-15) signaling induced miR-142 expression, whereas global and ILC-specific miR-142-deficient mice exhibited a cell-intrinsic loss of NK cells. Death of NK cells resulted from diminished IL-15 receptor signaling within miR-142-deficient mice, likely via reduced suppressor of cytokine signaling-1 (Socs1) regulation by miR-142-5p. ILCs persisting in Mir142-/- mice demonstrated increased expression of the miR-142-3p target αV integrin, which supported their survival. Global miR-142-deficient mice exhibited an expansion of ILC1-like cells concurrent with increased transforming growth factor-β (TGF-β) signaling. Further, miR-142-deficient mice had reduced NK-cell-dependent function and increased susceptibility to murine cytomegalovirus (MCMV) infection. Thus, miR-142 critically integrates environmental cues for proper type 1 ILC homeostasis and defense against viral infection.

NK cells are innate lymphocytes important for host defense against viral infections and malignancy. However, the molecular programs orchestrating NK cell activation are incompletely understood. MicroRNA-155 (miR-155) is markedly upregulated following cytokine activation of human and mouse NK cells. Surprisingly, mature human and mouse NK cells transduced to overexpress miR-155, NK cells from mice with NK cell–specific miR-155 overexpression, and miR-155−/− NK cells all secreted more IFN-γ compared with controls. Investigating further, we found that activated NK cells with miR-155 overexpression had increased per-cell IFN-γ with normal IFN-γ+ percentages, whereas greater percentages of miR-155−/− NK cells were IFN-γ+. In vivo murine CMV–induced IFN-γ expression by NK cells in these miR-155 models recapitulated the in vitro phenotypes. We performed unbiased RNA-induced silencing complex sequencing on wild-type and miR-155−/− NK cells and found that mRNAs targeted by miR-155 were enriched in NK cell activation signaling pathways. Using specific inhibitors, we confirmed these pathways were mechanistically involved in regulating IFN-γ production by miR-155−/− NK cells. These data indicate that miR-155 regulation of NK cell activation is complex and that miR-155 functions as a dynamic tuner for NK cell activation via both setting the activation threshold as well as controlling the extent of activation in mature NK cells. In summary, miR-155−/− NK cells are more easily activated, through increased expression of proteins in the PI3K, NF-κB, and calcineurin pathways, and miR-155−/− and 155-overexpressing NK cells exhibit increased IFN-γ production through distinct cellular mechanisms.

Objective Systemic lupus erythematosus (SLE) is a systemic autoimmune disease typically associated with elevated serum immunoglobulin G (IgG). Hypogammaglobulinemia in SLE patients has been attributed to immunosuppressive treatment or a transient effect associated with nephrotic syndrome. We retrospectively reviewed pediatric SLE patients from a single institution to identify patients with hypogammaglobulinemia and risk factors for hypogammaglobulinemia. Methods A total of 116 pediatric SLE cases from 1997 to 2011 were reviewed and patients with hypogammaglobulinemia (IgG Results Eighty-six patients were included in our study, with a median age of 15 years and a median follow-up of 39.5 months. Seven percent (six of 86) of patients had hypogammaglobulinemia with a median onset of 27 months (0–72 months) after SLE diagnosis. Significant associations were noted for white race ( p value 0.029), male sex ( p value 0.009), and the presence of lupus nephritis at SLE diagnosis ( p value 0.004). Use of immunosuppressive treatment did not show a statistical association with hypogammaglobulinemia, although two of the patients with hypogammaglobulinemia did receive rituximab. Most patients with hypogammaglobulinemia received intravenous immunoglobulin (IVIG) replacement therapy because of infections and/or concern for infection. Conclusion Measurement of immunoglobulin levels during treatment in SLE could help identify patients with hypogammaglobulinemia who might require more aggressive follow-up to monitor for increased risk of infection and need for IVIG treatment. A prospective study is needed to validate associated risk factors identified in this study.

NK cell activation is controlled by the integration of signals from cytokine receptors and germline–encoded activation and inhibitory receptors. NK cells undergo two distinct phases of activation during murine CMV (MCMV) infection: a nonselective phase mediated by proinflammatory cytokines and a specific phase driven by signaling through Ly49H, an NK cell activation receptor that recognizes infected cells. We sought to delineate cell surface markers that could distinguish NK cells that had been activated nonselectively from those that had been specifically activated through NK cell receptors. We demonstrated that stem cell Ag 1 (Sca-1) is highly upregulated during viral infections (to an even greater extent than CD69) and serves as a novel marker of early, nonselective NK cell activation. Indeed, a greater proportion of Sca-1+ NK cells produced IFN-γ compared with Sca-1− NK cells during MCMV infection. In contrast to the universal upregulation of Sca-1 (as well as KLRG1) on NK cells early during MCMV infection, differential expression of Sca-1, as well as CD27 and KLRG1, was observed on Ly49H+ and Ly49H− NK cells late during MCMV infection. Persistently elevated levels of KLRG1 in the context of downregulation of Sca-1 and CD27 were observed on NK cells that expressed Ly49H. Furthermore, the differential expression patterns of these cell surface markers were dependent on Ly49H recognition of its ligand and did not occur solely as a result of cellular proliferation. These findings demonstrate that a combination of Sca-1, CD27, and KLRG1 can distinguish NK cells nonselectively activated by cytokines from those specifically stimulated through activation receptors.

Natural killer (NK) cells have important functions in cancer immunosurveillance, BM allograft rejection, fighting infections, tissue homeostasis, and reproduction. NK cell–based therapies are promising treatments for blood cancers. Overcoming their currently limited efficacy requires a better understanding of the molecular mechanisms controlling NK cell development and dampening their effector functions. NK cells recognize the loss of self-antigens or up-regulation of stress-induced ligands on pathogen-infected or tumor cells through invariant NK cell receptors (NKRs), and then kill such stressed cells. Two second-messenger pathways downstream of NKRs are required for NK cell maturation and effector responses: PIP3 generation by PI3K and generation of diacylglycerol and IP3 by phospholipase-Cγ (PLCγ). In the present study, we identify a novel role for the phosphorylated IP3 metabolite inositol (1,3,4,5)tetrakisphosphate (IP4) in NK cells. IP4 promotes NK cell terminal differentiation and acquisition of a mature NKR repertoire. However, in mature NK cells, IP4 limits NKR-induced IFNγ secretion, granule exocytosis, and target-cell killing, in part by inhibiting the PIP3 effector-kinase Akt. This identifies IP4 as an important novel regulator of NK cell development and function and expands our understanding of the therapeutically important mechanisms dampening NK cell responses. Our results further suggest that PI3K regulation by soluble IP4 is a broadly important signaling paradigm.

The regulation of the late viral gene expression in betaherpesviruses is largely undefined. We have previously shown that the murine cytomegalovirus proteins pM79 and pM92 are required for late gene transcription. Here, we provide insight into the mechanism of pM79 and pM92 activity by determining their interaction partners during infection. Co-immunoprecipitation-coupled MS studies demonstrate that pM79 and pM92 interact with an array of cellular and viral proteins involved in transcription. Specifically, we identify RNA polymerase II as a cellular target for both pM79 and pM92. We use inter-protein coevolution analysis to show how pM79 and pM92 likely assemble into a late transcription complex composed of late transcription regulators pM49, pM87 and pM95. Combining proteomic methods with coevolution computational analysis provides novel insights into the relationship between pM79, pM92 and RNA polymerase II and allows the generation of a model of the multi-component viral complex that regulates late gene transcription.

Author(s): Fotis, Lampros; Shaikh, Nur; Baszis, Kevin; French, Anthony; Tarr, Phillip; Grevich, Sriharsha; Lee, Peggy; Ringold, Sarah; Leroux, Brian; Leahey, Hannah; Yuasa, Megan; Foster, Jessica; Sokolove, Jeremy; Lahey, Lauren; Robinson, William; Newsom, Joshua; Stevens, Anne; Karasawa, Rie; Tamaki, Mayumi; Tanaka, Megumi; Sato, Toshiko; Yudoh, Kazuo; Jarvis, James N; Moncrieffe, Halima; Bennett, Mark F; Tsoras, Monica; Luyrink, Lorie; Xu, Huan; Prahalad, Sampath; Morris, Paula; Dare, Jason; Nigrovic, Peter A; Rosenkranz, Margalit; Becker, Mara; O’Neil, Kathleen M; Griffin, Thomas; Lovell, Daniel J; Grom, Alexei A; Medvedovic, Mario; Thompson, Susan D; Zhu, Lisha; Jiang, Kaiyu; Wong, Laiping; Buck, Michael J; Chen, Yanmin; Moncrieffe, Halima; Brungs, Laura; Liu, Tao; Wang, Ting; Jarvis, James N; Alsaeid, Khaled; Alfailakawi, Jasim; Alenezi, Hamid; Alsaeed, Hazim; Beukelman, Tim; Natter, Marc; Ilowite, Norm; Mieszkalski, Kelly; Burrell, Grendel; Best, Brian; Bristow, Helen; Carr, Shannon; Dennos, Anne; Kaufmann, Rachel; Kimura, Yukiko; Schanberg, Laura; Blier, Peter R; Boneparth, Alexis; Wenderfer, Scott E; Moorthy, L Nandini; Radhakrishna, Suhas M; Sagcal-Gironella, Anna Carmela P; von Scheven, Emily; Gedik, Kader Cetin; Siddique, Salma; Aguiar, Cassyanne L; Erkan, Doruk; Cohen, Ezra; Lee, Yvonne; Dossett, Michelle; Mehta, Darshan; Davis, Roger; Gilbert, Mileka; Goilav, Beatrice; Meidan, Esra

Catatonia is a rare manifestation in patients with systemic lupus erythematosus (SLE). As catatonia can be associated with both psychiatric and organic conditions, this could create a diagnostic dilemma once this occurs in SLE patients. The report describes a 15-year-old female with SLE who developed catatonia three days after the diagnosis of SLE was made. Her catatonia was refractory to the treatment with immunosuppressive therapy, which included pulse methylprednisolone, intravenous cyclophosphamide, rituximab, intravenous immunoglobulin (IVIG) and plasmapheresis. Given her persistent catatonia, electroconvulsive therapy (ECT) was initiated three months after the onset of her symptoms. After the third ECT treatment, her mental status dramatically improved and returned nearly to baseline while she was continued on the immunosuppression. This is the first report of a successful ECT therapy in catatonic lupus in children.

NK cells are innate lymphocytes that mediate early host defense against viruses, such as cytomegalovirus. IL-15 is upregulated during viral infections and drives the expansion of NK cells. However, the influence of IL-15 on murine NK cell division and death rates has not been quantitatively studied. Therefore, we developed a series of two-compartment (representing quiescent and dividing NK cell subpopulations) mathematical models, incorporating different assumptions about the kinetic parameters regulating NK cell expansion. Using experimentally derived division and death rates, we tested each model’s assumptions by comparing predictions of NK cell numbers with independent experimental results and demonstrated that the kinetic parameters are distinct for nondividing and dividing NK cell subpopulations. IL-15 influenced NK cell expansion by modulating recruitment and division rates to a greater extent than death rates. The observed time delay to first division could be accounted for by differences in the kinetic parameters of nondividing and dividing subsets of NK cells. Although the duration of the time delay to first division was not significantly influenced by IL-15, the recruitment of nondividing NK cells into the replicating subpopulation increased with greater IL-15 concentrations. Our model quantitatively predicted changes in NK cell accumulation when IL-15 stimulation was reduced, demonstrating that NK cell divisional commitment was interrupted when cytokine stimulation was removed. In summary, this quantitative analysis reveals novel insights into the in vitro regulation of NK cell proliferation and provides a foundation for modeling in vivo NK cell responses to viral infections.

Parotitis is a common condition seen in the pediatric population, usually as an isolated occurrence associated with viral or bacterial infection. The differential diagnosis expands when recurrent parotitis is encountered. One etiology is primary pediatric Sjögren syndrome (SS), an autoimmune condition typically associated with dryness of the eyes and mouth in adults. Pediatric patients often present with isolated recurrent bilateral parotitis, however, and we describe 4 such cases in children aged 9 to 17 years at presentation. Despite lack of ocular complaints, 3 of these patients had ocular findings on ophthalmologic exam. Our patients also exhibited classic laboratory abnormalities, including positive antinuclear antibody, SS A, and SS B antibodies; presence of rheumatoid factor; and hypergammaglobulinemia. Consideration of SS in the child with recurrent parotitis is important for timely and appropriate referral and treatment. We review the differential diagnosis of parotitis in children as well as the salient features of pediatric SS.

Objective To estimate the length of time to disease flare and the likelihood of achieving clinical remission after discontinuation of treatment with tumor necrosis factor α (TNFα) blockers in patients with juvenile idiopathic arthritis (JIA). Methods We conducted a retrospective chart review in a cohort of patients with JIA treated with TNFα inhibitors between January 1, 1998 and November 1, 2009. Demographic information, laboratory data, and medication exposure were extracted using a standardized tool. Outcomes of interest were based on preliminary criteria for remission in JIA. Results One hundred seventy-one patients with 255 discrete episodes of anti-TNFα treatment were reviewed. The median duration of patient observation was 59.7 months (range 5.8-211.2 months). Among patients in whom disease was inactive after discontinuation of anti-TNFα therapy, 50% had persistently inactive disease at 6 months, and 33% had clinical remission at 12 months. The median duration of anti-TNFα therapy after inactive disease was obtained was 6.1 months (range 0-67.9 months). No significant association was observed between the time to disease flare after cessation of treatment with TNFα antagonists and the length of time from the diagnosis of JIA to the initiation of anti-TNFα therapy, the duration of therapy following the onset of inactive disease, or the total duration of treatment with TNFα antagonists prior to discontinuation. The category of JIA, sex, and age at diagnosis were not associated with the risk of relapse. Conclusion One-third of patients with JIA can successfully undergo withdrawal of treatment with TNFα antagonists and be spared the cost and potential morbidity of treatment for at least 12 months. Further studies are needed to identify factors to accurately identify these patients.

Under selective pressure from host immunity, viruses have retained genes encoding immunoevasins, molecules interfering with host viral recognition and clearance. Due to their binding specificities, immunoevasins can be exploited as affinity labels to identify host-encoded molecules of previously unsuspected importance in defense against the relevant class of virus. We previously described an orthopoxvirus MHC class I-like protein (OMCP) that binds with high affinity to the activating receptor NKG2D on NK and T cell subsets, implicating NKG2D in antiorthopoxvirus immunity. In this study, we report that OMCP also binds in an NKG2D-independent manner to B cells and monocytes/macrophages. We identify murine FcR-like 5 (FCRL5), an orphan immunoregulatory protein highly expressed by innate B lymphocytes, as a specific receptor for OMCP. The three N-terminal Ig domains of FCRL5 are required for OMCP binding. The targeting of FCRL5 by an orthopoxvirus immunoevasin strongly implicates it in contributing to host defense against zoonotic orthopoxviruses.

Studies in diverse cell types have rediscovered the importance of basic cellular metabolism in regulating immune responses. For example, glycolytic and oxidative metabolism are counter-regulated in T cells to develop inflammatory effectors vs. memory and regulatory cells, respectively. NK cells, which are first responders to viral infection and malignant transformation, may be similarly regulated. We recently showed that NK cell cytotoxic activity during infection with murine cytomegalovirus (MCMV) required glucose metabolism, whereas other NK cell responses such as cytokine production did not. We hypothesized that oxidative metabolism would be required for some of these glycolysis-independent responses. To test this, we examined responses to MCMV in mice with an NK-specific deletion of Cox10, which is required for assembly of electron transport chain complex IV. These “Ncr1-Cox10” mice had normal NK cell development and numbers, and stimulated Cox10-deficient NK cells survived and proliferated normally in vitro. However, Ncr1-Cox10 mice were more susceptible to MCMV infection, with impaired expansion of MCMV-specific Ly49H+ NK cells and increased apoptosis of splenic NK cells. Thus, we find that Cox10 is required by NK cells to expand and survive in response to MCMV infection. These deficiencies were not found in vitro, emphasizing the importance of using NK-specific genetic models to investigate metabolism in physiological contexts.

NK cells vigorously proliferate during viral infections, resulting in an expanded pool of innate lymphocytes that are able to participate in early host defense. The relative contributions of cytokines and activation receptors in stimulating NK cell proliferation during viral infections are not well characterized. In this study, we demonstrated that signaling through the NK cell activation receptor Ly49H was able to compensate for the absence of cytokine stimulation in the preferential phase of viral-induced proliferation during murine cytomegalovirus infection. In the absence of type I IFN stimulation, NK cell proliferation was strongly biased toward cells expressing the Ly49H receptor, even at early time points when minimal preferential Ly49H-mediated proliferation was observed in wild-type mice. In the absence of effective Ly49H signaling or following infection with virus that did not express the ligand for Ly49H, no difference was observed in the proliferation of subsets of NK cells that either express or lack expression of Ly49H, although the overall proliferation of NK cells in IFNαβR−/− mice was substantially reduced. These results highlight the contribution of NK cell activation receptors in stimulating proliferation and subsequent expansion of NK cells that are able to recognize virally infected cells.

Cytotoxic lymphocytes use the granule exocytosis pathway to kill pathogen-infected cells and tumor cells. Although many genes in this pathway have been extensively characterized (e.g., perforin, granzymes A and B), the role of granzyme C is less clear. We therefore developed a granzyme C-specific mAb and used flow cytometry to examine the expression of granzyme B and C in the lymphocyte compartments of wild-type and mutant GzmB−/− cre mice, which have a small deletion in the granzyme B gene. We detected granzyme B and C expression in CD4+ and CD8+ T cells activated with CD3/CD28 beads or MLRs. Stimulation of NK cells in vitro with IL-15 also induced expression of both granzymes. Granzyme C up-regulation was delayed relative to granzyme B in wild-type lymphocytes, whereas GzmB−/− cre cells expressed granzyme C earlier and more abundantly on a per-cell basis, suggesting that the deleted 350-bp region in the granzyme B gene is important for the regulation of both granzymes B and C. Quantitative RT-PCR revealed that granzyme C protein levels were regulated by mRNA abundance. In vivo, a population of wild-type CD8αα+ intraepithelial lymphocytes constitutively expressed granzyme B and GzmB−/− cre intraepithelial lymphocytes likewise expressed granzyme C. Using a model of a persistent murine CMV infection, we detected delayed expression of granzyme C in NK cells from infected hosts. Taken together, these findings suggest that granzyme C is activated with persistent antigenic stimulation, providing nonredundant backup protection for the host when granzyme B fails.

Background The development of natural killer (NK) cells in the bone marrow is not well characterized. We recently described a mouse (referred to as an NK cell–deficient [NKD] mouse) with a selective deficiency in NK cells caused by the insertion of a transgene construct into the genetic locus for the basic leucine zipper transcription factor ATF-2. NK cells in this mouse were both phenotypically and functionally immature and accumulated in the bone marrow at a stage at which constitutive NK cell proliferation occurs in wild-type mice. Objective We hypothesized that excess IL-15 could potentially overcome this developmental block, allowing normal emigration of mature NK cells from the bone marrow to the periphery. Methods Double-transgenic mice were generated by crossing the NKD mice with transgenic mice overexpressing IL-15. Results The double-transgenic mice had a dramatic accumulation of phenotypically immature NK cells in the bone marrow and subsequently in the blood, liver, and spleen. NK cells from these double-transgenic mice manifested functional deficits similar to those observed in NK cells from NKD mice, as assessed by decreased cytokine production and cytotoxicity. Conclusion Rather than bypass the observed developmental defect in NKD mice, excess IL-15 drove a massive accumulation of phenotypically and functionally immature NK cells in the bone marrow and periphery. Clinical implications We propose that these double-transgenic mice will serve as a murine model of chronic NK cell lymphocytosis in human patients.

SummaryAlthough activated murine NK cells can use the granule exocytosis pathway to kill target cells immediately upon recognition, resting murine NK cells are minimally cytotoxic for unknown reasons. Here, we showed that resting NK cells contained abundant granzyme A, but little granzyme B or perforin; in contrast, the mRNAs for all three genes were abundant. Cytokine-induced in vitro activation of NK cells resulted in potent cytotoxicity associated with a dramatic increase in granzyme B and perforin, but only minimal changes in mRNA abundance for these genes. The same pattern of regulation was found in vivo with murine cytomegalovirus infection as a physiologic model of NK cell activation. These data suggest that resting murine NK cells are minimally cytotoxic because of a block in perforin and granzyme B mRNA translation that is released by NK cell activation.

Natural killer (NK) cells play a crucial role in the initial host defense against pathogens such as murine cytomegalovirus (MCMV). They respond rapidly and effectively control pathogen replication while the adaptive immune system is being activated. However, in the absence of an adaptive immune system, an effective initial NK cell response is not sufficient for long-term pathogen control as demonstrated by the late recrudescence of disease and mortality in immunodeficient mice infected with MCMV. In this setting, NK cells suppress the initial infection but exert enough selective pressure to drive the outgrowth of MCMV mutants that escape recognition by NK cells. Herein, we characterize the rapid emergence of escape mutants following infection with a plaque-purified MCMV isolate and demonstrate that these mutant viruses are no longer effectively controlled by NK cells. These findings suggest that late recrudescence of viral infections in certain clinical settings may also be due to viral escape from NK cells or other components of innate immunity.

Objective To assess mortality in a population-based cohort of adults with a history of juvenile rheumatoid arthritis (JRA). Methods The Rochester Epidemiology Project database was used to identify all cases of JRA diagnosed among Rochester, Minnesota residents under the age of 16 between January 1, 1960 and December 31, 1993. Fifty-seven patients in this cohort are now adults (ages 18–53 years, mean age 34.3 years), and this subgroup was contacted for a long-term followup study. The average length of followup from the time of diagnosis was 25.6 years. Results Four deaths occurred in this cohort of 57 adults with a history of JRA. All 4 deceased patients had other autoimmune illnesses and died of complications of these diseases. The observed frequency of 4 deaths was significantly greater (P < 0.0026 by one-sample log-rank test) than the 1 death that would be expected among Minnesota whites of similar age and sex, and corresponds to a mortality rate of 0.27 deaths per 100 years of patient followup compared with an expected mortality rate of 0.068 deaths per 100 years of followup in the general population. Conclusion The results indicate a significant, unexpected increase in mortality in this population-based cohort of adults with a history of JRA in comparison with the rate in the general population. The deaths in this group were all associated with other autoimmune disorders, suggesting that special emphasis should be given to the diagnosis and treatment of other autoimmune diseases, including immunodeficiencies, in JRA patients. The frequency of deaths in this cohort suggests that JRA patients are at substantial risk for mortality, and highlights the need for longitudinal followup and care into adulthood.

Receptor-mediated endocytosis is the process by which cells internalize ligands that have specifically interacted with cell surface receptors. Within intracellular endosomal compartments, receptor/ligand complexes can be targeted to lysosomes for degradation, recycled back to the plasma membrane, or sorted separately to these destinations. We have developed a mechanistic mathematical model that can account for the spectrum of experimentally observed endosomal sorting outcomes. The central hypothesis of this model is that receptors may be selectively retained by putative endosomal retention components and that this process may be modulated by receptor occupancy. This hypothesis is supported by the recent discovery of an endosomal retention component involved in targeting epidermal growth factor receptors to lysosomes. In this paper, we use the model to predict how changes in key cellular and molecular parameters alter sorting outcomes. This analysis provides guidance for rationally modulating the sorting process in a variety of biomedical applications, either by the manipulation of cellular parameters or the design of ligand properties.

Autoimmune diseases are thought to be caused by a loss of self-tolerance of the immune system. One candidate marker of immune dysregulation in autoimmune disease is the presence of increased double negative T cells (DNTs) in the periphery. DNTs are characteristically elevated in autoimmune lymphoproliferative syndrome, a systemic autoimmune disease caused by defective lymphocyte apoptosis due to Fas pathway defects. DNTs have also been found in the peripheral blood of adult patients with systemic lupus erythematosus (SLE), where they may be pathogenic. DNTs in children with autoimmune disease have not been investigated.We evaluated DNTs in pediatric patients with SLE, mixed connective tissue disease (MCTD), juvenile idiopathic arthritis (JIA), or elevated antinuclear antibody (ANA) but no systemic disease. DNTs (CD3(+)CD56(-)TCRαβ(+)CD4(-)CD8(-)) from peripheral blood mononuclear cells were analyzed by flow cytometry from 54 pediatric patients including: 23 SLE, 15 JIA, 11 ANA and 5 MCTD compared to 28 healthy controls.Sixteen cases (29.6 %) had elevated DNTs (≥2.5 % of CD3(+)CD56(-)TCRαβ(+) cells) compared to 1 (3.6 %) control. Medication usage including cytotoxic drugs and absolute lymphocyte count were not associated with DNT levels, and percentages of DNTs were stable over time. Analysis of multiple phenotypic and activation markers showed increased CD45RA expression on DNTs from patients with autoimmune disease compared to controls.DNTs are elevated in a subset of pediatric patients with autoimmune disease and additional investigations are required to determine their precise role in autoimmunity.

In this study, we report that murine cytomegalovirus (MCMV) protein pM92 regulates viral late gene expression during virus infection. Previously, we have shown that MCMV protein pM79 and its human cytomegalovirus (HCMV) homologue pUL79 are required for late viral gene transcription. Identification of additional factors involved is critical to dissecting the mechanism of this regulation. We show here that pM92 accumulated abundantly at late times of infection in a DNA synthesis-dependent manner and localized to nuclear viral replication compartments. To investigate the role of pM92, we constructed a recombinant virus SM in 92, in which pM92 expression was disrupted by an insertional/frameshift mutation. During infection, SM in 92 accumulated representative viral immediate-early gene products, early gene products, and viral DNA sufficiently but had severe reduction in the accumulation of late gene products and was thus unable to produce infectious progeny. Coimmunoprecipitation and mass spectrometry analysis revealed an interaction between pM92 and pM79, as well as between their HCMV homologues pUL92 and pUL79. Importantly, we showed that the growth defect of pUL92-deficient HCMV could be rescued in trans by pM92. This study indicates that pM92 is an additional viral regulator of late gene expression, that these regulators (represented by pM92 and pM79) may need to complex with each other for their activity, and that pM92 and pUL92 share a conserved function in CMV infection. pM92 represents a potential new target for therapeutic intervention in CMV disease, and a gateway into studying a largely uncharted viral process that is critical to the viral life cycle.

Natural killer (NK) cells are innate lymphocytes that provide early host defense against intracellular pathogens, such as viruses. Although NK cell development, homeostasis, and proliferation are regulated by IL-15, the influence of IL-15 receptor (IL-15R)-mediated signaling at the cellular level has not been quantitatively characterized. We developed a mathematical model to analyze the kinetic interactions that control the formation and localization of IL-15/IL-15R complexes. Our computational results demonstrated that IL-15/IL-15R complexes on the cell surface were a key determinant of the magnitude of the IL-15 proliferative signal and that IL-15R occupancy functioned as an effective surrogate measure of receptor signaling. Ligand binding and receptor internalization modulated IL-15R occupancy. Our work supports the hypothesis that the total number and duration of IL-15/IL-15R complexes on the cell surface crosses a quantitative threshold prior to the initiation of NK cell division. Furthermore, our model predicted that the upregulation of IL-15Rα on NK cells substantially increased IL-15R complex formation and accelerated the expansion of dividing NK cells with the greatest impact at low IL-15 concentrations. Model predictions of the threshold requirement for NK cell recruitment to the cell cycle and the subsequent exponential proliferation correlated well with experimental data. In summary, our modeling analysis provides quantitative insight into the regulation of NK cell proliferation at the receptor level and provides a framework for the development of IL-15 based immunotherapies to modulate NK cell proliferation., Author Summary Natural killer (NK) cells are innate immune cells that are important in our bodies' initial defenses against pathogens, like viruses. NK cells rapidly proliferate early during viral infections to provide an expanded pool of effector cells to suppress the infection. This proliferative response is driven by a cytokine called interleukin-15 (IL-15); however, the influence of IL-15 and its receptor (IL-15R) in stimulating NK cell proliferation has not been quantitatively characterized at the cellular level. To better understand the factors controlling the vigorous expansion of NK cells during infections, we developed a mathematical model incorporating IL-15R binding and trafficking parameters that regulate the number of cell-surface IL-15/IL-15R signaling complexes. The analysis of this model provided us with insight on how IL-15-driven NK cell expansion can be modulated through changes in receptor kinetics and expression. Based on model predictions, we were able to draw inferences about NK cell population dynamics and to compare these conclusions with quantitative experimental results. Our results and model have applicability to studies designed to manipulate cell responses in the context of immunotherapies.

Peptide growth factors and other receptor-binding cytokine ligands are of interest in contemporary molecular health care approaches in applications such as wound healing, tissue regeneration, and gene therapy. Development of effective technologies based on operation of these regulatory molecules requires an ability to deliver the ligands to target cells in a reliable and wellcharacterizable manner. Quantitative information concerning the fate of peptide ligands within tissues is necessary for adequate interpretation of experimental observations at the tissue level and for truly rational engineering design of ligand-based therapies. To address this need, we are undertaking efforts to elucidate effects of key molecular and cellular parameters on temporal and spatial distribution of cytokines in cell population and cell/matrix systems. In this article we summarize some of our recent findings on dynamics of growth factor depletion by cellular endocytic trafficking, growth factor transport through cellular matrices, and growth factor production and release by autocrine cell systems.

Endocytosed molecules are sorted in endosomes to different cellular destinations (e.g., to lysosomes or to the plasma membrane). Diverse endosomal sorting results have been reported for different ligands and receptors in a variety of cell types, but the general principles governing these sorting outcomes are not well understood. For example, we observed a wide range of sorting outcomes with the epidermal growth factor (EGF)/receptor system in fibroblasts using several members of the EGF family and site-directed ligand and receptor mutants. In this article we describe a mechanistic mathematical model of endosomal sorting based on the hypothesis that receptors may be selectively retained by the endosomal sorting apparatus and that this process may be modulated by receptor occupancy. Our results show that this single mechanism can account for the wide variety of observed sorting outcomes. By providing a conceptual framework for understanding endosomal sorting, this model not only helps interpret our experimental results for the EGF/receptor system, but also provides some insight into the principles governing sorting. For example, the model predicts that the influence of selective endosomal retention of receptor/ligand complexes is seen in deviations of ligand sorting outcomes from pure fluid phase sorting behavior. Furthermore, the model suggests that selective endosomal retention of complexes within endosomes gives rise to three sorting regimes characterized by distinguishable qualitative trends in the dependence of ligand sorting fractions on intracellular ligand concentrations.

Individuals with robust natural killer (NK) cell function incur lower rates of malignancies. To expand our understanding of genetic factors contributing to this phenomenon, we analyzed NK cells from cancer resistant and susceptible strains of mice. We identified a correlation between NK levels of the X-chromosome-located adaptor protein SLy1 and immunologic susceptibility to cancer. Unlike the case for T or B lymphocytes, where SLy1 shuttles between the cytoplasm and nucleus to facilitate signal transduction, in NK cells SLy1 functions as a ribosomal protein and is located solely in the cytoplasm. In its absence, ribosomal instability results in p53-mediated NK cell senescence and decreased clearance of malignancies. NK defects are reversible under inflammatory conditions and viral clearance is not impacted by SLy1 deficiency. Our work defines a previously unappreciated X-linked ribosomopathy that results in a specific and subtle NK cell dysfunction leading to immunologic susceptibility to cancer.

Using members of the epidermal growth factor (EGF) family as well as site-directed recombinant human EGF mutants, we investigated how ligand binding properties influence endosomal sorting. Mouse EGF (mEGF), human EGF (hEGF), and transforming growth factor alpha (TGF alpha) bind to the human EGF receptor (EGFR) with similar affinities at pH 7.4. However, the binding properties of these ligands have substantially different pH sensitivities resulting in varying degrees of dissociation from the receptors at lower pH levels characteristic of endosomes. We employed a steady-state sorting assay to determine the fraction of ligand sorted to recycling versus degradation as a function of the number of intracellular ligand molecules in mouse B82 fibroblasts. mEGF, hEGF, and TGF alpha display significantly different steady-state endosomal sorting patterns which correspond to the extent of their dissociation at endosomal pH. Moreover, several recombinant hEGF mutants with differing affinities exhibit altered endosomal sorting compared to hEGF, demonstrating a similar direct relationship between ligand binding properties and endosomal sorting outcomes. Intracellular trafficking of the EGF ligands was also monitored by measuring the observed degradation rate constants. These likewise show marked differences that correlate with the differing pH sensitivities of the ligands' binding properties.

Natural killer (NK) cells play a critical role in the host defense against herpesviruses. Although herpesviruses are ubiquitous in human populations, only a minority of people experience severe recurrent infections. We hypothesize that uncharacterized NK cell functional deficits predispose individuals to more significant or frequent herpesvirus infections and reactivations. To investigate this hypothesis, we broadly analyzed NK cell phenotype and functional responses in a cohort of predominantly pediatric patients with recurrent and/or severe herpesvirus infections and compared them to a healthy control population. Our results identified no global differences in cytolysis, degranulation, interferon-γ production, or surface receptor upregulation following cytokine stimulation. However, abnormal NK cell functional responses were observed in nearly one-third of patients (including 3 with hyporesponsiveness to activating signals and 1 with markedly decreased CD11b expression associated with reduced cytotoxicity and degranulation), which might contribute to those individuals’ susceptibility to herpesvirus infections.

NK cells are innate immune lymphocytes important for early host defense against infectious pathogens and malignant transformation. MicroRNAs (miRNAs) are small RNA molecules that regulate a wide variety of cellular processes, typically by specific complementary targeting of the 3′UTR of mRNAs. The Dicer1 gene encodes a conserved enzyme essential for miRNA processing, and Dicer1 deficiency leads to a global defect in miRNA biogenesis. In this study, we report a mouse model of lymphocyte-restricted Dicer1 disruption to evaluate the role of Dicer1-dependent miRNAs in the development and function of NK cells. As expected, Dicer1-deficient NK cells had decreased total miRNA content. Furthermore, miRNA-deficient NK cells exhibited reduced survival and impaired maturation defined by cell surface phenotypic markers. However, Dicer1-deficient NK cells exhibited enhanced degranulation and IFN-γ production in vitro in response to cytokines, tumor target cells, and activating NK cell receptor ligation. Moreover, a similar phenotype of increased IFN-γ was evident during acute MCMV infection in vivo. miRs-15a/15b/16 were identified as abundant miRNAs in NK cells that directly target the murine IFN-γ 3′UTR, thereby providing a potential mechanism for enhanced IFN-γ production. These data suggest that the function of miRNAs in NK cell biology is complex, with an important role in NK cell development, survival, or homeostasis, while tempering peripheral NK cell activation. Further study of individual miRNAs in an NK cell specific fashion will provide insight into these complex miRNA regulatory effects in NK cell biology.

Intracellular trafficking of the epidermal growth factor receptor (EGF-R) is regulated by receptor occupancy. To investigate this, we developed an assay to study endosomal sorting under steady-state conditions. Using a cell line transfected with EGF-R variants, we found that the fraction of internalized EGF.EGF-R complexes sorted to lysosomes was a function of the number of intracellular complexes and required sequences in the cytoplasmic domain of the receptor. As the number of intracellular occupied wild-type receptors increased from 3 x 10(2) to 2 x 10(5)/cell, the fraction of internalized EGF that was degraded dropped from 70 to 20%. Transforming growth factor-alpha, which dissociates from the EGF-R at endosomal pH, was degraded to a uniform extent of approximately 50% at all intracellular ligand concentrations. EGF internalized by receptors lacking a cytoplasmic domain (c'647) was degraded to an extent of only 5-10% independent of the number of intracellular complexes. Mutant receptors truncated either at residues 1022 or 973 displayed sorting patterns intermediate between wild-type and c'647 receptors. Despite large differences in their internalization rates, the fractional sorting patterns of c'1022 and c'973 receptors were indistinguishable. Receptor tyrosine kinase activity appeared to have a small effect on sorting pattern, but only in the context of full-length receptors. Our results indicate that the default pathway of internalized receptors is rapidly recycling and that lysosomal targeting of occupied EGF-R is due to endosomal retention that is both specific and saturable. In addition, internalization and endosomal retention of EGF-R appear to be mediated by distinct structural elements.

Increased susceptibility to autoimmunity in females is often viewed as the consequence of enhanced immunoreactivity providing superior protection against infections. We paradoxically observed greater mortality in female compared to male mice during systemic viral infections with three large double-stranded DNA viruses (herpes simplex virus type I [HSV], murine cytomegalovirus [MCMV], and vaccinia virus [VV]). Indeed, female mice were 27-fold more susceptible to infection with HSV than male mice. Elimination of estrogen by ovariectomy in female mice or addition of estrogen to castrated male mice only partially eliminated the observed sex differences following HSV infection. However, the differences observed in survival between female and male mice were nearly abrogated in the absence of type I interferon receptor signaling and substantially mitigated in absence of DAP12 signaling. Interestingly, the sex-specific impact of type I interferon receptor and DAP12 signaling differentially influenced survival during systemic viral infections with type I interferon receptor signaling enhancing male survival and DAP12 signaling increasing the susceptibility of female mice. These results have potential implications for the sex disparities observed in human autoimmune disorders.

Objective Juvenile dermatomyositis (DM) is a chronic inflammatory myopathy of childhood primarily affecting the muscles and skin. Treatment for juvenile DM is often difficult, and conventional therapies include corticosteroids and other immune suppressants. We reviewed the records of 4 patients with juvenile DM who received the B cell–depleting anti-CD20 monoclonal antibody rituximab to determine whether this therapy resulted in improved control of their juvenile DM. Methods This is a retrospective review of 4 pediatric patients ages 10–17 years with juvenile DM who were treated with rituximab. All patients were tested for myositis autoantibodies and received weekly rituximab infusions for a total of 4 doses. Two patients received repeat courses of rituximab 1 year after their first dose. Patients were followed up between 12 and 24 months after their first course of rituximab, and their strength, muscle enzymes, and rash were reviewed. Results One patient was positive for a myositis-specific antibody, anti–Mi-2, and demonstrated striking reductions in her muscle enzyme levels for 1 year after rituximab therapy. Following a second course of rituximab, this patient remained disease free for 14 months before requiring a third course of rituximab. Two myositis antibody–negative patients showed clinical improvement and tolerated lower doses of corticosteroids following treatment with rituximab. Finally, 1 patient had worsening of her disease following rituximab. Conclusion These cases highlight the potential for anti–B cell therapies in the treatment of juvenile DM in both myositis-specific autoantibody–positive and –negative patients.