Case Study: Mouse Parvovirus Outbreak Likely Caused by a Contaminated Commercial Lyophilized Antibody Powder

An MPV-contaminated lyophilized antibody product obtained from a commercial vendor was the probable cause of an outbreak of mouse parvovirus (MPV) in an academic research institution. The outbreak was initially discovered by the seroconversion of the mouse sentinels receiving soiled bedding from the affected cage(s). After further investigation, a suspected antibody product was submitted to a diagnostic laboratory and the sample tested positive for MPV via polymerase chain reaction (PCR). To confirm administration of this product to mice could produce MPV infection, we inoculated the MPV-positive antibody product into experimental mice (n=5). We collected faecal pellets at Days 0, 5, 9, 12, and 14 post-inoculation. At the end of the experimental period, we collected mesenteric lymph nodes (mLN) and submitted both mLN and faecal pellets for MPV analysis via PCR. While all faecal pellets were negative for MPV, we were able to detect MPV in mLN from one of the five mice, thus replicating the likely method of transmission and the cause of the MPV outbreaks

A colitogenic memory CD4+ T cell population mediates gastrointestinal graft-versus-host disease

Damage to the gastrointestinal tract is a major cause of morbidity and mortality in graft-versus-host disease (GVHD) and is attributable to T cell–mediated inflammation. In this work, we identified a unique CD4+ T cell population that constitutively expresses the β2 integrin CD11c and displays a biased central memory phenotype and memory T cell transcriptional profile, innate-like properties, and increased expression of the gut-homing molecules α4β7 and CCR9. Using several complementary murine GVHD models, we determined that adoptive transfer and early accumulation of β2 integrin–expressing CD4+ T cells in the gastrointestinal tract initiated Th1-mediated proinflammatory cytokine production, augmented pathological damage in the colon, and increased mortality. The pathogenic effect of this CD4+ T cell population critically depended on coexpression of the IL-23 receptor, which was required for maximal inflammatory effects. Non–Foxp3-expressing CD4+ T cells produced IL-10, which regulated colonic inflammation and attenuated lethality in the absence of functional CD4+Foxp3+ T cells. Thus, the coordinate expression of CD11c and the IL-23 receptor defines an IL-10–regulated, colitogenic memory CD4+ T cell subset that is poised to initiate inflammation when there is loss of tolerance and breakdown of mucosal barriers

Blockade of Interleukin 27 Signaling Attenuates Graft Versus Host Disease By Augmenting CD4+ and CD8+ Regulatory T Cell Reconstitution

Reestablishment of competent regulatory pathways has emerged as a strategy to reduce the severity of graft-versus-host disease (GVHD), and recalibrate the effector and regulatory arms of the immune system. However, clinically feasible, cost-effective strategies that do not require extensive ex vivo cellular manipulation have remained elusive. In the current study, we demonstrate that inhibition of the interleukin-27p28 (IL-27p28) signaling pathway through antibody blockade or genetic ablation prevented lethal GVHD in multiple murine transplant models. Moreover, protection from GVHD was attributable to augmented global reconstitution of CD4+ natural regulatory T cells (nTregs), CD4+ induced Tregs (iTregs), and CD8+ iTregs, and was more potent than temporally concordant blockade of IL-6 signaling. Inhibition of IL-27p28 also enhanced the suppressive capacity of adoptively transferred CD4+ nTregs by increasing the stability of Foxp3 expression. Notably, blockade of IL-27p28 signaling reduced T-cell–derived-IL-10 production in conventional T cells; however, there was no corresponding effect in CD4+ or CD8+ Tregs, indicating that IL-27 inhibition had differential effects on IL-10 production and preserved a mechanistic pathway by which Tregs are known to suppress GVHD. Targeting of IL-27 therefore represents a novel strategy for the in vivo expansion of Tregs and subsequent prevention of GVHD without the requirement for ex vivo cellular manipulation, and provides additional support for the critical proinflammatory role that members of the IL-6 and IL-12 cytokine families play in GVHD biology

Calcium Mobilization Triggered by the Chemokine CXCL12 Regulates Migration in Wounded Intestinal Epithelial Monolayers*

Restitution of intestinal epithelial barrier damage involves the coordinated remodeling of focal adhesions in actively migrating enterocytes. Defining the extracellular mediators and the intracellular signaling pathways regulating those dynamic processes is a key step in developing restitution-targeted therapies. Previously we have determined that activation of the chemokine receptor CXCR4 by the cognate ligand CXCL12 enhances intestinal epithelial restitution through reorganization of the actin cytoskeleton. The aim of these studies was to investigate the role of calcium effectors in CXCL12-mediated restitution. CXCL12 stimulated release of intracellular calcium in a dose-dependent manner. Inhibition of intracellular calcium flux impaired CXCL12-mediated migration of IEC-6 and CaCo2 cells. Pharmacological blockade and specific shRNA depletion of the phospholipase-C (PLCβ3) isoform attenuated CXCL12-enhanced migration, linking receptor activation with intracellular calcium flux. Immunoblot analyses demonstrated CXCL12 activated the calcium-regulated focal adhesion protein proline-rich tyrosine kinase-2 (Pyk2) and the effector proteins paxillin and p130Cas. Interruption of Pyk2 signaling potently blocked CXCL12-induced wound closure. CXCL12-stimulated epithelial cell migration was enhanced on laminin and abrogated by intracellular calcium chelation. These results suggest CXCL12 regulates restitution through calcium-activated Pyk2 localized to active focal adhesions. Calcium signaling pathways may therefore provide a novel avenue for enhancing barrier repair