Characterization of a bovine WC1(+) gammadelta T cell memory population

One of the key aspects of the immune system is the ability to prime cells by vaccination. The γδ T lymphocytes represent a significant population of cells in the peripheral blood of cattle that have largely been ignored in this regard. Here we have explored the potential for WC1+ γδ T cells to be primed by vaccination. We hypothesize that the γδ T cells that exhibit a recall response in vitro following in vivo priming represent a unique population. Utilizing a Leptospira borgpetersenii vaccine, we examined the recall responses of WC1+ and CD4+ T cells to leptospiral antigen. The WC1+ γδ cells were the major responding population to antigen for the first few weeks following in vivo priming, with the CD4 T cell response only began to overtake them after a booster dose was administered. The primed WC1+ γδ T cells displayed a unique pattern of surface marker expression when stimulated with antigen compared to mitogen-stimulated cells, and which paralleled that observed on the CD4 T cells that responded to antigen. Additionally, chemokine receptor expression was assessed in both ex vivo and antigen-stimulated WC1 and CD4T cells. Ex vivo CD4+ and WC1+ T cells differed with regard to chemokine receptor transcript expression while the antigen-activated cells had very similar patterns of expression. Both subsets expressed genes typical of TH1-polarized cells, but differed with regard to transcripts for co-stimulatory molecules expressed. TCR usage by the antigen-responsive WC1+ γδ T cells from vaccinated animals was evaluated. The antigen-responsive cells had transcripts for several different Vγ and Vδ gene segments with very limited usage of J genes and highly variable CDR3 sequences. These results did not differ greatly from those obtained with nondividing cells or with ex vivo peripheral blood mononuclear cells suggesting little if any enrichment for a specific TCR type. These results support the hypothesis that WC1 + γδ T cells and CD4 αβ T cells that respond to leptospira antigen are likely to differ mainly with regard to how they are activated and the pathogen molecules that activate them, rather than with regard to their effector functions

Adjuvant Immunotherapy of Experimental Autoimmune Encephalomyelitis: Immature Myeloid Cells Expressing CXCL10 and CXCL16 Attract CXCR3(+)CXCR6(+) and Myelin-Specific T Cells to the Draining Lymph Nodes Rather Than the Central Nervous System

CFA is a strong adjuvant capable of stimulating cellular immune responses. Paradoxically, adjuvant immunotherapy by prior exposure to CFA or live mycobacteria suppresses the severity of EAE and spontaneous diabetes in rodents. Here we investigated immune responses during adjuvant immunotherapy of experimental autoimmune encephalomyelitis (EAE). Induction of EAE in CFA-pretreated mice resulted in a rapid influx into the draining lymph nodes (dLNs) of large numbers of CD11b(+)Gr-1(+) myeloid cells, consisting of immature cells with ring-shaped nuclei, macrophages, and neutrophils. Concurrently, a population of mycobacteria-specific IFN-γ-producing T cells appeared in the dLNs. Immature myeloid cells in dLNs expressed the chemokines CXCL10 and CXCL16 in an IFN-γ-dependent manner. Subsequently, CD4(+) T cells co-expressing the cognate chemokine receptors, CXCR3 and CXCR6, and myelin oligodendrocyte glycoprotein (MOG)-specific CD4(+) T cells accumulated within the chemokine-expressing dLNs, rather than within the CNS. Migration of CD4(+) T cells toward dLN cells was abolished by depleting the CD11b(+) cells and was also mediated by the CD11b(+) cells alone. In addition to altering the distribution of MOG-specific T cells, adjuvant-treatment suppressed development of MOG-specific IL-17. Thus, CFA-adjuvant immunotherapy of EAE requires IFN-γ, which suppresses development of the Th17-response, and diverts autoreactive T cells away from the CNS towards immature myeloid cells expressing CXCL10 and CXCL16 in the lymph nodes

Evaluation of Type 1 Immune Response in Naïve and Vaccinated Animals following Challenge with Leptospira borgpetersenii Serovar Hardjo: Involvement of WC1(+) γδ and CD4 T Cells

Organisms within the Hardjo serovar of Leptospira species are harbored in cattle throughout the world, causing abortion in pregnant animals as well as being shed in the urine, thereby providing sources of zoonotic infection for humans. We recently showed that sterile immunity in vaccinated cattle is associated with induction of a type 1 (Th1) cell-mediated immune response. Here naïve and previously vaccinated pregnant cattle were challenged with a virulent strain of serovar Hardjo and subsequently evaluated for expression of a type 1 immune response. Lymphocytes that responded in a recall response to antigen by undergoing blast transformation were evident in cultures of peripheral blood mononuclear cells (PBMC) from vaccinated cattle throughout the postchallenge test period while those from naïve cattle were evident at one time point only. Nevertheless, beginning at 2 weeks after challenge, gamma interferon (IFN-γ) was measured in supernatants of antigen-stimulated PBMC cultures from nonvaccinated animals although the amount produced was always less than that in cultures of PBMC from vaccinated animals. IFN-γ(+) cells were also evident in antigen-stimulated cultures of PBMC from vaccinated but not from nonvaccinated animals throughout the postchallenge period. The IFN-γ(+) cells included CD4(+) and WC1(+) γδ T cells, and a similar proportion of these two subpopulations were found among the dividing cells in antigen-stimulated cultures as ascertained by carboxyfluorescein succinimidyl ester loading. Finally, while naïve and vaccinated animals had similar levels of antigen-specific immunoglobulin G1 (IgG1) following challenge, vaccinated animals had twofold-more IgG2. In conclusion, while infection may induce a type 1 response we suggest that it is too weak to prevent establishment of chronic infection

Adjuvant Immunotherapy of Experimental Autoimmune Encephalomyelitis: Immature Myeloid Cells Expressing CXCL10 and CXCL16 Attract CXCR3 +

CFA is a strong adjuvant capable of stimulating cellular immune responses. Paradoxically, adjuvant immunotherapy by prior exposure to CFA or live mycobacteria suppresses the severity of EAE and spontaneous diabetes in rodents. Here we investigated immune responses during adjuvant immunotherapy of experimental autoimmune encephalomyelitis (EAE). Induction of EAE in CFA-pretreated mice resulted in a rapid influx into the draining lymph nodes (dLNs) of large numbers of CD11b(+)Gr-1(+) myeloid cells, consisting of immature cells with ring-shaped nuclei, macrophages, and neutrophils. Concurrently, a population of mycobacteria-specific IFN-γ-producing T cells appeared in the dLNs. Immature myeloid cells in dLNs expressed the chemokines CXCL10 and CXCL16 in an IFN-γ-dependent manner. Subsequently, CD4(+) T cells co-expressing the cognate chemokine receptors, CXCR3 and CXCR6, and myelin oligodendrocyte glycoprotein (MOG)-specific CD4(+) T cells accumulated within the chemokine-expressing dLNs, rather than within the CNS. Migration of CD4(+) T cells toward dLN cells was abolished by depleting the CD11b(+) cells and was also mediated by the CD11b(+) cells alone. In addition to altering the distribution of MOG-specific T cells, adjuvant-treatment suppressed development of MOG-specific IL-17. Thus, CFA-adjuvant immunotherapy of EAE requires IFN-γ, which suppresses development of the Th17-response, and diverts autoreactive T cells away from the CNS towards immature myeloid cells expressing CXCL10 and CXCL16 in the lymph nodes

Modulation of Responses of Vibrio parahaemolyticus O3:K6 to pH and Temperature Stresses by Growth at Different Salt Concentrations ▿

Vibrio parahaemolyticus inhabits marine, brackish, and estuarine waters worldwide, where fluctuations in salinity pose a constant challenge to the osmotic stress response of the organism. Vibrio parahaemolyticus is a moderate halophile, having an absolute requirement for salt for survival, and is capable of growth at 1 to 9% NaCl. It is the leading cause of seafood-related bacterial gastroenteritis in the United States and much of Asia. We determined whether growth in differing NaCl concentrations alters the susceptibility of V. parahaemolyticus O3:K6 to other environmental stresses. Vibrio parahaemolyticus was grown at a 1% or 3% NaCl concentration, and the growth and survival of the organism were examined under acid or temperature stress conditions. Growth of V. parahaemolyticus in 3% NaCl versus that in 1% NaCl increased survival under both inorganic (HCl) and organic (acetic acid) acid conditions. In addition, at 42°C and −20°C, 1% NaCl had a detrimental effect on growth. The expression of lysine decarboxylase (encoded by cadA), the organism's main acid stress response system, was induced by both NaCl and acid conditions. To begin to address the mechanism of regulation of the stress response, we constructed a knockout mutation in rpoS, which encodes the alternative stress sigma factor, and in toxRS, a two-component regulator common to many Vibrio species. Both mutant strains had significantly reduced survival under acid stress conditions. The effect of V. parahaemolyticus growth in 1% or 3% NaCl was examined using a cytotoxicity assay, and we found that V. parahaemolyticus grown in 1% NaCl was significantly more toxic than that grown in 3% NaCl