B cells require DOCK8 to elicit and integrate T cell help when antigen is limiting

Dedicator of cytokinesis 8 (DOCK8) immunodeficiency syndrome is characterized by a failure of the germinal center response, a process involving the proliferation and positive selection of antigen-specific B cells. Here, we describe how DOCK8-deficient B cells are blocked at a light-zone checkpoint in the germinal centers of immunized mice, where they are unable to respond to T cell–dependent survival and selection signals and consequently differentiate into plasma cells or memory B cells. Although DOCK8-deficient B cells can acquire and present antigen to initiate activation of cognate T cells, integrin up-regulation, B cell–T cell conjugate formation, and costimulation are insufficient for sustained B cell and T cell activation when antigen availability is limited. Our findings provide an explanation for the failure of the humoral response in DOCK8 immunodeficiency syndrome and insight into how the level of available antigen modulates B cell–T cell cross-talk to fine-tune humoral immune responses and immunological memory

Oxytocin enhances basolateral amygdala activation and functional connectivity while processing emotional faces: preliminary findings in autistic versus non-autistic women

Oxytocin is hypothesized to promote social interactions by enhancing the salience of social stimuli. While previous neuroimaging studies have reported that oxytocin enhances amygdala activation to face stimuli in autistic men, effects in autistic women remain unclear. In this study, the influence of intranasal oxytocin on activation and functional connectivity of the basolateral amygdala – the brain’s “salience detector” – while processing emotional faces vs. shapes was tested in 16 autistic and 21 non-autistic women by fMRI in a placebo-controlled, within-subjects, cross-over design. In the placebo condition, minimal activation differences were observed between autistic and non-autistic women. However, significant drug × group interactions were observed for both basolateral amygdala activation and functional connectivity. Oxytocin increased left basolateral amygdala activation among autistic women (35 voxel cluster, MNI coordinates of peak voxel= -22 -10 -28; mean change=+0.079%, t=3.159, ptukey=0.0166), but not non-autistic women (mean change =+0.003%, t=0.153, ptukey=0.999). Furthermore, oxytocin increased functional connectivity of the right basolateral amygdala with brain regions associated with socio-emotional information processing in autistic women, but not non-autistic women, attenuating group differences in the placebo condition. Taken together, these findings extend evidence of oxytocin’s effects on the amygdala to specifically include autistic women and specify the subregion of the effect.TLP was supported by the Autism Research Trust, Cambridge Trust, and Natural Sciences and Engineering Research Council of Canada. MVL was supported by an ERC Starting Grant (ERC-2017-STG; 755816). MCL was supported by a Canadian Institutes of Health Research (CIHR) Sex and Gender Science Chair (GSB 171373), the O’Brien Scholars Program within the Child and Youth Mental Health Collaborative at the Centre for Addiction and Mental Health (CAMH) and The Hospital for Sick Children, Toronto, the Academic Scholars Award from the Department of Psychiatry, University of Toronto, the CAMH Foundation, and the Ontario Brain Institute. SBC received funding from the Wellcome Trust 214322\Z\18\Z. For the purpose of Open Access, the author has applied a CC BY public copyright licence to any Author Accepted Manuscript version arising from this submission. SBC also received funding from the Autism Centre of Excellence, SFARI, the Templeton World Charitable Fund, the MRC, and the National Institute for Health Research (NIHR). Any views expressed are those of the author(s) and not necessarily those of the funder. RB was supported by the MRC UK, Pinsent Darwin Trust and British Academy post-doctoral fellowship

NDRG1 is induced by antigen-receptor signaling but dispensable for B and T cell self-tolerance.

Peripheral tolerance prevents the initiation of damaging immune responses by autoreactive lymphocytes. While tolerogenic mechanisms are tightly regulated by antigen-dependent and independent signals, downstream pathways are incompletely understood. N-myc downstream-regulated gene 1 (NDRG1), an anti-cancer therapeutic target, has previously been implicated as a CD4+ T cell clonal anergy factor. By RNA-sequencing, we identified Ndrg1 as the third most upregulated gene in anergic, compared to naïve follicular, B cells. Ndrg1 is upregulated by B cell receptor activation (signal one) and suppressed by co-stimulation (signal two), suggesting that NDRG1 may be important in B cell tolerance. However, though Ndrg1-/- mice have a neurological defect mimicking NDRG1-associated Charcot-Marie-Tooth (CMT4d) disease, primary and secondary immune responses were normal. We find that B cell tolerance is maintained, and NDRG1 does not play a role in downstream responses during re-stimulation of in vivo antigen-experienced CD4+ T cells, demonstrating that NDGR1 is functionally redundant for lymphocyte anergy

Aberrant mucin assembly in mice causes endoplasmic reticulum stress and spontaneous inflammation resembling ulcerative colitis

BACKGROUND: MUC2 mucin produced by intestinal goblet cells is the major component of the intestinal mucus barrier. The inflammatory bowel disease ulcerative colitis is characterized by depleted goblet cells and a reduced mucus layer, but the aetiology remains obscure. In this study we used random mutagenesis to produce two murine models of inflammatory bowel disease, characterised the basis and nature of the inflammation in these mice, and compared the pathology with human ulcerative colitis. METHODS AND FINDINGS: By murine N-ethyl-N-nitrosourea mutagenesis we identified two distinct noncomplementing missense mutations in Muc2 causing an ulcerative colitis-like phenotype. 100% of mice of both strains developed mild spontaneous distal intestinal inflammation by 6 wk (histological colitis scores versus wild-type mice, p , 0.01) and chronic diarrhoea. Monitoring over 300 mice of each strain demonstrated that 25% and 40% of each strain, respectively, developed severe clinical signs of colitis by age 1 y. Mutant mice showed aberrant Muc2 biosynthesis, less stored mucin in goblet cells, a diminished mucus barrier, and increased susceptibility to colitis induced by a luminal toxin. Enhanced local production of IL-1b, TNF-a, and IFN-c was seen in the distal colon, and intestinal permeability increased 2-fold. The number of leukocytes within mesenteric lymph nodes increased 5-fold and leukocytes cultured in vitro produced more Th1 and Th2 cytokines (IFN-c, TNF-a, and IL-13). This pathology was accompanied by accumulation of the Muc2 precursor and ultrastructural and biochemical evidence of endoplasmic reticulum (ER) stress in goblet cells, activation of the unfolded protein response, and altered intestinal expression of genes involved in ER stress, inflammation, apoptosis, and wound repair. Expression of mutated Muc2 oligomerisation domains in vitro demonstrated that aberrant Muc2 oligomerisation underlies the ER stress. In human ulcerative colitis we demonstrate similar accumulation of nonglycosylated MUC2 precursor in goblet cells together with ultrastructural and biochemical evidence of ER stress even in noninflamed intestinal tissue. Although our study demonstrates that mucin misfolding and ER stress initiate colitis in mice, it does not ascertain the genetic or environmental drivers of ER stress in human colitis. CONCLUSIONS: Characterisation of the mouse models we created and comparison with human disease suggest that ER stress-related mucin depletion could be a fundamental component of the pathogenesis of human colitis and that clinical studies combining genetics, ER stress-related pathology and relevant environmental epidemiology are warranted. The Editors’ Summary of this article follows the references

Prolidase deficiency causes spontaneous T cell activation and lupus-like autoimmunity

Prolidase deficiency (PD) is a multisystem disorder caused by mutations in the PEPD gene, which encodes a ubiquitously expressed metallopeptidase essential for the hydrolysis of dipeptides containing C-terminal proline or hydroxyproline. PD typically presents in childhood with developmental delay, skin ulcers, recurrent infections, and, in some patients, autoimmune features that can mimic systemic lupus erythematosus. The basis for the autoimmune association is uncertain, but might be due to self-antigen exposure with tissue damage, or indirectly driven by chronic infection and microbial burden. In this study, we address the question of causation and show that Pepd-null mice have increased antinuclear autoantibodies and raised serum IgA, accompanied by kidney immune complex deposition, consistent with a systemic lupus erythematosus–like disease. These features are associated with an accumulation of CD4 and CD8 effector T cells in the spleen and liver. Pepd deficiency leads to spontaneous T cell activation and proliferation into the effector subset, which is cell intrinsic and independent of Ag receptor specificity or antigenic stimulation. However, an increase in KLRG1+ effector CD8 cells is not observed in mixed chimeras, in which the autoimmune phenotype is also absent. Our findings link autoimmune susceptibility in PD to spontaneous T cell dysfunction, likely to be acting in combination with immune activators that lie outside the hemopoietic system but result from the abnormal metabolism or loss of nonenzymatic prolidase function. This knowledge provides insight into the role of prolidase in the maintenance of self-tolerance and highlights the importance of treatment to control T cell activation

Natural HLA Class I Polymorphism Controls the Pathway of Antigen Presentation and Susceptibility to Viral Evasion

HLA class I polymorphism creates diversity in epitope specificity and T cell repertoire. We show that HLA polymorphism also controls the choice of Ag presentation pathway. A single amino acid polymorphism that distinguishes HLA-B*4402 (Asp116) from B*4405 (Tyr116) permits B*4405 to constitutively acquire peptides without any detectable incorporation into the transporter associated with Ag presentation (TAP)-associated peptide loading complex even under conditions of extreme peptide starvation. This mode of peptide capture is less susceptible to viral interference than the conventional loading pathway used by HLA-B*4402 that involves assembly of class I molecules within the peptide loading complex. Thus, B*4402 and B*4405 are at opposite extremes of a natural spectrum in HLA class I dependence on the PLC for Ag presentation. These findings unveil a new layer of MHC polymorphism that affects the generic pathway of Ag loading, revealing an unsuspected evolutionary trade-off in selection for optimal HLA class I loading versus effective pathogen evasion

Unlocking the bottleneck in forward genetics using whole-genome sequencing and identity by descent to isolate causative mutations

Forward genetics screens with N-ethyl-N-nitrosourea (ENU) provide a powerful way to illuminate gene function and generate mouse models of human disease; however, the identification of causative mutations remains a limiting step. Current strategies depend on conventional mapping, so the propagation of affected mice requires non-lethal screens; accurate tracking of phenotypes through pedigrees is complex and uncertain; out-crossing can introduce unexpected modifiers; and Sanger sequencing of candidate genes is inefficient. Here we show how these problems can be efficiently overcome using whole-genome sequencing (WGS) to detect the ENU mutations and then identify regions that are identical by descent (IBD) in multiple affected mice. In this strategy, we use a modification of the Lander-Green algorithm to isolate causative recessive and dominant mutations, even at low coverage, on a pure strain background. Analysis of the IBD regions also allows us to calculate the ENU mutation rate (1.54 mutations per Mb) and to model future strategies for genetic screens in mice. The introduction of this approach will accelerate the discovery of causal variants, permit broader and more informative lethal screens to be used, reduce animal costs, and herald a new era for ENU mutagenesis.The High-Throughput Genomics Group at the Wellcome Trust Centre for Human Genetics is funded by Wellcome Trust grant reference 090532/Z/09/Z and MRC Hub grant G0900747 91070. This study was supported by Wellcome Trust Strategic Award 082030 (CCG), Wellcome Trust Studentship 094446/Z/10/Z (KRB), the Oxford NIHR Biomedical Research Centre, and the MRC Human Immunology Unit (RJC). AJR and GL were supported by Wellcome Trust grant 090532/Z/ 09/Z, CCG and AE by a Major initiative Award from the Clive and Vera Ramaciotti Foundation, and AE by an NHMRC Career Development Award. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Effects of oxytocin administration on salivary sex hormone levels in autistic and neurotypical women

Abstract: Background: Oxytocin administration, which may be of therapeutic value for individuals with social difficulties, is likely to affect endogenous levels of other socially relevant hormones. However, to date, the effects of oxytocin administration on endogenous hormones have only been examined in neurotypical individuals. The need to consider multi-hormone interactions is particularly warranted in oxytocin trials for autism due to evidence of irregularities in both oxytocin and sex steroid systems. Methods: In this double-blind cross-over study, saliva samples were collected from 16 autistic and 29 neurotypical women before and after intranasal administration of 24 IU oxytocin or placebo. Oestradiol, testosterone, and oxytocin levels were quantified in saliva samples. Participants also completed the Autism-Spectrum Quotient (AQ) and Empathy Quotient (EQ) questionnaires. Results: Distinct patterns of change in testosterone and oestradiol levels pre- to-post-administration were observed in autistic relative to neurotypical women (ANCOVA, p < 0.05 main effect of Group), controlling for sample collection time. The mean percent change oestradiol was + 8.8% for the autism group and − 13.0% for the neurotypical group (t = 1.81, p = 0.08), while the mean percent change testosterone was + 1.1% in the autism group and − 12.6% in the neurotypical group (t = 1.26, p = 0.22). In the oxytocin condition, the mean percent change oestradiol was + 12.6% in the autism group and − 6.9% in the neurotypical group (t = 1.78, p = 0.08), while the mean percent change testosterone was + 14.4% in the autism group and − 15.2% in the neurotypical group (t = 3.00, p = 0.006). Robust regression confirmed that group differences in percent change hormone levels were not driven by a small number of influential individuals. Baseline hormone levels did not differ between groups when considered individually. However, baseline testosterone relative to oestradiol (T:E2 ratio) was higher in autistic women (p = 0.023, Cohen’s d = 0.63), and this ratio correlated positively and negatively with AQ and EQ scores, respectively, in the combined sample. Limitations: Further studies with larger and more diverse autistic sample are warranted to confirm these effects. Conclusions: This study provides the first evidence that oxytocin influences endogenous testosterone levels in autistic individuals, with autistic women showing increases similar to previous reports of neurotypical men. These findings highlight the need to consider sex steroid hormones as a variable in future oxytocin trials

Aberrant Mucin Assembly in Mice Causes Endoplasmic Reticulum Stress and Spontaneous Inflammation Resembling Ulcerative Colitis

Michael McGuckin and colleagues identify two mutations that cause aberrant mucin oligomerization in mice. The resulting phenotype, including endoplasmic reticulum stress, resembles clinical and pathologic features of human ulcerative colitis