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Background Vulvodynia is a remarkably prevalent chronic pain condition of unknown etiology. Epidemiologic studies associate the risk of vulvodynia with a history of atopic disease. We used an established model of hapten-driven contact hypersensitivity to investigate the underlying mechanisms of allergy-provoked prolonged sensitivity to pressure. Methods We sensitized female ND4 Swiss mice to the hapten oxazolone on their flanks, and subsequently challenged them four days later with oxazolone or vehicle for ten consecutive days on the labia. We evaluated labiar sensitivity to touch, local mast cell accumulation, and hyperinnervation after ten challenges. Results Oxazolone-challenged mice developed significant tactile sensitivity that persisted for over three weeks after labiar allergen exposures ceased. Allergic sites were characterized by mast cell accumulation, sensory hyper-innervation and infiltration of regulatory CD4+CD25+FoxP3+ T cells as well as localized early increases in transcripts encoding Nerve Growth Factor and nerve-mast cell synapse marker Cell Adhesion Molecule 1. Local depletion of mast cells by intra-labiar administration of secretagogue compound 48/80 led to a reduction in both nerve density and tactile sensitivity. Conclusions Mast cells regulate allergy-provoked persistent sensitivity to touch. Mast cell-targeted therapeutic strategies may provide novel means to manage and limit chronic pain conditions associated with atopic disease.

Programmed death-1 (PD-1) is a T cell inhibitory receptor important for tolerance maintenance. PD-1 deficiency or blockade accelerates autoimmune diabetes in non-obese diabetic (NOD) mice, but the mechanism remains unclear. Autoantibody production is a hallmark of autoimmunity, and has also been reported in patients treated with PD-1 blockade, suggesting that PD-1 might regulate this process. Autoantibody production results from B cell:CD4 T cell interactions in the germinal center of the lymph node. The dynamics and regulation of the germinal center response in spontaneous autoimmunity and in the face of PD-1 deprivation are not well understood, primarily due to an inability to track self-specific lymphocytes. To bridge this knowledge gap, we used tetramers to phenotype islet-specific CD4 T cells and B cells in mice. PD-1-deficient mice, and NOD mice treated with anti-PD-1, had increased insulin autoantibodies, as well as increased insulin-specific germinal center T follicular helper CD4 T cells and germinal center B cells compared to controls. This work provides a mechanistic explanation for autoantibody onset following PD-1 blockade in the clinic, and has important implications for cancer immunotherapy and autoimmunity.

Cellular and molecular mediators of immune responses are increasingly implicated in acute and chronic pain pathophysiologies. Here we demonstrate that passive cutaneous IgE/Ag anaphylaxis provokes increased thermal sensitivity in the hind paw tissue of mice. The murine anti-DNP IgE antibodies SPE-7 and ɛ26 are known to induce differential cytokine production in bone marrow cultured mast cells in vitro without antigen challenge. We found a novel, antigen-dependent heterogeneity in the thermal pain responses elicited in the hind paws between SPE-7 and ɛ26 sensitized DNP-challenged mice. Mice experienced pronounced hind paw thermal sensitivity lasting 6 h after DNP challenge when sensitized with SPE-7 but not ɛ26 IgE. The two IgE clones induced equivalent hind paw edema, neutrophil influx, cytokine production, and reduction in tissue histamine content in vivo , and bound to the same or overlapping epitopes on the DNP antigen in vitro . Therefore IgE antibodies against the same antigen can induce comparable inflammation, yet contribute to markedly different anaphylaxis-associated pain within an allergic response, suggesting that non-canonical IgE binding partners such as sensory neurons may play a role in allergy-related pain responses.

Epidemiological evidence suggests a link between a history of environmental allergies and the risk of developing vulvodynia - a chronic provoked vulvar pain condition affecting up to 20% of women in populates assessed. We have previously shown that acute labiar exposure to a hapten allergen provokes increased mechanical sensitivity in the vulvar tissue of sensitized mice. Here, we show repeated challenge with contact hypersensitivity allergen oxazolone produces sustained mechanical hyperalgesia in the vulvar region of previously sensitized female mice for 10-14 days after final allergen exposure. Acute inflammatory responses including labiar neutrophil influx, T and B cell amplification in the draining iliac lymph nodes, labiar expression of inflammatory cytokine genes (IL-6, IL-1β and Cxcl-2) are all resolved within 5 days after final allergen exposure while increased densities of both peptidergic nerve fibers and mast cells in the labiar tissue are observed. Our findings provide the first evidence of the induction of measurable and sustained pain persisting after the resolution of overt inflammatory events following an allergic response in the labiar/vulvar tissue in mice. We have established the first model of chronic allergy-provoked vulvar pain in mice that can be used to elucidate the underlying etiology of subsets of vulvar pain that present without accompanying inflammation and are associated with histories of allergic skin responses.

The interplay among pain, allergy and dysregulated inflammation promises to yield significant conceptual advances in immunology and chronic pain. Hapten-mediated contact hypersensitivity reactions are used to model skin allergies in rodents but have not been utilized to study associated changes in pain perception in the affected skin. Here we characterized changes in mechanical hyperalgesia in oxazolone-sensitized female mice challenged with single and repeated labiar skin exposure to oxazolone. Female mice were sensitized with topical oxazolone on their flanks and challenged 1-3 times on the labia. We then measured mechanical sensitivity of the vulvar region with an electronic pressure meter and evaluated expression of inflammatory genes, leukocyte influx and levels of innervation in the labiar tissue. Oxazolone-sensitized mice developed vulvar mechanical hyperalgesia after a single labiar oxazolone challenge. Hyperalgesia lasted up to 24 hours along with local influx of neutrophils, upregulation of inflammatory cytokine gene expression, and increased density of cutaneous labiar nerve fibers. Three daily oxazolone challenges produced vulvar mechanical hyperalgesic responses and increases in nerve density that were detectable up to 5 days post-challenge even after overt inflammation resolved. This persistent vulvar hyperalgesia is resonant with vulvodynia, an understudied chronic pain condition that is remarkably prevalent in 18-60 year-old women. An elevated risk for vulvodynia has been associated with a history of environmental allergies. Our pre-clinical model can be readily adapted to regimens of chronic exposures and long-term assessment of vulvar pain with and without concurrent inflammation to improve our understanding of mechanisms underlying subsets of vulvodynia and to develop new therapeutics for this condition.