CD101 genetic variants modify regulatory and conventional T cell phenotypes and functions.

We recently reported that the risk of sexually acquired HIV-1 infection is increased significantly by variants in the gene encoding CD101, a protein thought to modify inflammatory responses. Using blood samples from individuals with and without these variants, we demonstrate that CD101 variants modify the prevalence of circulating inflammatory cell types and show that CD101 variants are associated with increased proinflammatory cytokine production by circulating T cells. One category of CD101 variants is associated with a reduced capacity of regulatory T cells to suppress T cell cytokine production, resulting in a reduction in the baseline level of immune quiescence. These data are supported by transcriptomics data revealing alterations in the intrinsic regulation of antiviral pathways and HIV resistance genes in individuals with CD101 variants. Our data support the hypothesis that CD101 contributes to homeostatic regulation of bystander inflammation, with CD101 variants altering heterosexual HIV-1 acquisition by facilitating increased prevalence and altered function of T cell subsets.

IgG Subclass Determines Suppression Versus Enhancement of Humoral Alloimmunity to Kell RBC Antigens in Mice.

It has long been appreciated that immunoglobulins are not just the effector endpoint of humoral immunity, but rather have a complex role in regulating antibody responses themselves. Donor derived anti-RhD IgG has been used for over 50 years as an immunoprophylactic to prevent maternal alloimmunization to RhD. Although anti-RhD has dramatically decreased rates of hemolytic disease of the fetus and newborn (for the RhD alloantigen), anti-RhD also fails in some cases, and can even paradoxically enhance immune responses in some circumstances. Attempts to generate a monoclonal anti-RhD have largely failed, with some monoclonals suppressing less than donor derived anti-RhD and others enhancing immunity. These difficulties likely result, in part, because the mechanism of anti-RhD remains unclear. However, substantial evidence exists to reject the common explanations of simple clearance of RhD + RBCs or masking of antigen. Donor derived anti-RhD is a mixture of 4 different IgG subtypes. To the best of our knowledge an analysis of the role different IgG subtypes play in immunoregulation has not been carried out; and, only IgG1 and IgG3 have been tested as monoclonals. Multiple attempts to elicit alloimmune responses to human RhD epitopes in mice have failed. To circumvent this limitation, we utilize a tractable animal model of RBC alloimmunization using the human Kell glycoprotein as an antigen to test the effect of IgG subtype on immunoregulation by antibodies to RBC alloantigens. We report that the ability of an anti-RBC IgG to enhance, suppress (at the level of IgM responses), or have no effect is a function of the IgG subclass in this model system.

Understanding how combinatorial targeting of TLRs and TNFR family costimulatory members promote enhanced T cell responses.

INTRODUCTION: Due to the ability of pathogen-associated molecular patters and tumor necrosis factor receptor (TNFR) family costimulatory agonists to boost T cell responses, studies have combined Toll-like receptor (TLR) ligands with TNFR family costimulatory receptor agonists to induce impressive and long-lasting T cell responses. Although some studies have determined how these combinatorial vaccines promote enhanced T cell responses, much remains unknown about the mechanism used by these combinations to promote synergistic T cell responses - especially in settings of infectious diseases or cancer. AREAS COVERED: In this review, we look in detail at the signaling pathways induced by combinatorial targeting of TLR and TNFR family costimulatory members that help them promote synergistic T cell responses. Understanding this can greatly aid the development of novel vaccine regimens that promote cellular immune responses, which is essential for treating certain infectious diseases and cancer. EXPERT OPINION: Vaccines against some infectious diseases as well as therapeutic cancer vaccines require cellular immunity. Therefore, we evaluate here how signaling pathways induced by TLR ligand and costimulatory agonist combinations promote enhanced T cell responses during immunization with model antigens, viral pathogens, or tumor antigens. Once pathways that drive these combinatorial vaccines to boost T cell activation are identified, they can be incorporated in vaccines designed to target pathogens or cancer.

Optimal CD4 T cell priming after LPS-based adjuvanticity with CD134 costimulation relies on CXCL9 production.

LPS is a powerful adjuvant, and although LPS-mediated TLR4 signaling has been exquisitely delineated, the in vivo mechanism of how TLR4 responses impact T cell priming is far less clear. Besides costimulation, TNF and type 1 IFN are dominant cytokines released after TLR4 activation and can shape T cell responses, but other downstream factors have not been examined extensively. Depending on context, we show that IFNαR1 blockade resulted in minor to major effects on specific CD4 T cell clonal expansion. To help explain these differences, it was hypothesized that IFNαR1 blockade would inhibit specific T cell migration by reducing chemokine receptor signaling, but specific CD4 T cells from IFNαR1-blocked mice were readily able to migrate in response to specific chemokines. Next, we examined downstream factors and found that type 1 IFN signaling was necessary for chemokine production, even when mice were immunized with specific Ag with LPS and CD134 costimulation. IFNαR1 signaling promoted CXCL9 and CXCL10 synthesis, suggesting that these chemokines might be involved in the LPS and CD134 costimulation response. After immunization, we show that CXCL9 blockade inhibited CD4 T cell accumulation in the liver but also in LNs, even in the presence of elevated serum IFN-ß levels. Thus, whereas type 1 IFN might have direct effects on primed CD4 T cells, the downstream chemokines that play a role during migration also impact accumulation. In sum, CXCL9 production is a key benchmark for productive CD4 T cell vaccination strategies.

Ca2+ oscillations, Ca2+ sensitization, and contraction activated by protein kinase C in small airway smooth muscle.

Protein kinase C (PKC) has been implicated in the regulation of smooth muscle cell (SMC) contraction and may contribute to airway hyperresponsiveness. Here, we combined optical and biochemical analyses of mouse lung slices to determine the effects of PKC activation on Ca(2+) signaling, Ca(2+) sensitivity, protein phosphorylation, and contraction in SMCs of small intrapulmonary airways. We found that 10 µM phorbol-12-myristate-13-acetate or 1 µM phorbol 12,13-dibutyrate induced repetitive, unsynchronized, and transient contractions of the SMCs lining the airway lumen. These contractions were associated with low frequency Ca(2+) oscillations in airway SMCs that resulted from Ca(2+) influx through L-type voltage-gated Ca(2+) channels and the subsequent release of Ca(2+) from intracellular stores through ryanodine receptors. Phorbol ester stimulation of lung slices in which SMC intracellular Ca(2+) concentration ([Ca(2+)](i)) was "clamped" at a high concentration induced strong airway contraction, indicating that PKC mediated sensitization of the contractile response to [Ca(2+)](i). This Ca(2+) sensitization was accompanied by phosphorylation of both the PKC-potentiated PP1 inhibitory protein of 17 kD (CPI-17) and the regulatory myosin light chain. Thrombin, like the phorbol esters, induced a strong Ca(2+) sensitization that was inhibited by the PKC inhibitor GF-109203X and also potentiated airway contraction to membrane depolarization with KCl. In conclusion, we suggest that PKC activation in small airways leads to both the generation of Ca(2+) oscillations and strong Ca(2+) sensitization; agents associated with airway inflammation, such as thrombin, may activate this pathway to sensitize airway smooth muscle to agonists that cause membrane depolarization and Ca(2+) entry and induce airway hyperresponsiveness.