Modulation of Langerhans cell phenotype, migration and maturation by agents known to cause herpes simplex virus reactivation in a mouse model.

The factors which control whether an asymptomatic or symptomatic infection in the skin follows reactivation of herpes simplex virus (HSV) in the sensory ganglia remain unclear. Xylene, retinoic acid and dimethylsuphoxide (DMSO) all stimulate similar levels of virus reactivation in the ganglia of latently infected mice, yet give rise to high, moderate and very low incidences of clinical skin disease, respectively. This observation suggests that the chemicals may be capable of affecting the local microenvironment of the skin. In the present study we have investigated the effects of xylene, retinoic acid and DMSO on Langerhans cell (LC) phenotype and function. The results show that none of the chemicals inhibit the phenotypic maturation of Langerhans cells in vitro. However, DMSO induced a dramatic elevation in class II MHC expression on Langerhans cells. In addition, fewer antigen-bearing dendritic cells (DC) were evident in the lymph nodes if xylene was administered to the skin prior to challenge with the contact sensitizer, FITC. None of the chemicals inhibited the accumulation of DC in the lymph nodes after such an antigen-challenge, suggesting that xylene was affecting not migration, but antigen uptake. The inhibition of antigen uptake by xylene together with the activation of LC in terms of class II expression by DMSO may explain, in part, the relative abilities of these chemicals to allow the establishment of recurrent HSV disease in the skin.

Kinetics of peptide uptake and tissue distribution following a single intranasal dose of peptide.

We have previously used intranasal (i.n.) peptide application to induce mucosal tolerance in experimental autoimmune encephalomyelitis (EAE). This strategy, however, appeared to give rise to similar phenomena of tolerance observed as a result of systemic administration of soluble antigenic peptide. We were interested, therefore, in the uptake and tissue distribution of peptide following i.n. treatment. In the H-2u mouse model of EAE, the highly tolerogenic peptide analogue Ac1-9[4Y] of myelin basic protein (MBP) displays high affinity binding to Au MHC class II. For the purpose of the present study this peptide was synthesised to contain a tritiated acetyl group and a protocol was developed to recover radioactivity in solubilised tissues taken at various times after [3H]Ac1-9[4Y] i.n.. Radiolabel loads of the lung and gastro-intestinal tract were initially high but declined rapidly. Radiolabel uptake by blood and lymphoid tissues followed similar kinetics with peak levels around 2.5-4 hours after i.n. administration. Concentrations were high in the draining cervical lymph nodes (CLN) but also reached significant levels in the spleen and 'nondraining' inguinal lymph nodes. The presence of intact antigenic peptide was demonstrated in spleens and CLN from Ac1-9[4Y] i.n. treated mice. Cell suspensions prepared from these tissues at selected time points after peptide i.n. were able to stimulate peptide-specific T cell lines up to at least one day after peptide i.n., suggesting long lasting formation of stable Au-Ac1-9[4Y] complexes in vivo.

Qualitative and quantitative effects of CD28/B7-mediated costimulation on naive T cells in vitro.

The CD28/B7 system provides costimulatory signals necessary for optimal T cell activation. We have examined the effects of blocking B7.1 and/or B7.2 in an in vitro system using TCR transgenic T cells specific for myelin basic protein. Activation of naive T cells was found to be B7.2 dependent and not dependent on the presence of B7.1 molecules. However, increasing the strength of signal through the TCR using peptide analogues with higher affinity for MHC compensated for blockade of B7.2 molecules, suggesting that signal 1 alone can be sufficient for the activation of naive T cells. The role of B7 molecules in the differentiation of T cells was further investigated by restimulating T cells with fresh APC and peptide in B7-sufficient conditions. A down-regulation of IL-2 and IFN-gamma production by T cells primed in the presence of anti-B7.2 mAb was partially overcome when high affinity peptide analogues were used to restimulate T cells. In contrast, a significant down-regulation of the differentiation of cells producing Th-2 cytokines was observed in the presence of anti-B7 Abs. Differentiation of IL-4-secreting cells was influenced by both B7.1 and B7.2, while IL-5 secretion was totally dependent on B7.2. These results suggest that B7-mediated costimulation is essential for the development of Th-2-associated cytokines, the absence of which cannot be overcome by increasing the strength of the signal through the TCR.

Fine specificity of the myelin-reactive T cell repertoire: implications for TCR antagonism in autoimmunity.

The use of altered peptide ligands (APL) to modulate T cell responses has been suggested as a means of treating T cell-mediated autoimmune disorders. We have assessed the therapeutic potential of TCR antagonist peptides in autoimmunity using murine experimental autoimmune encephalomyelitis (EAE) as a model. The Tg4 transgenic mouse expresses an MHC class II-restricted TCR specific for the immunodominant encephalitogenic epitope of myelin basic protein, Ac1-9 (acetylated N-terminal nonamer). We have used T cell lines derived from Tg4 mice to define the TCR contact residues within Ac1-9. APL with appropriate substitutions at the primary TCR contact residue were effective antagonists of Tg4 T cells. These antagonist APL, however, were found to induce EAE in susceptible, nontransgenic strains of mice. Underlying this, the Ac1-9-specific T cell repertoire of normal mice, rather than reflecting the Tg4 phenotype, showed considerable diversity in fine specificity and was able to respond to the Tg4 antagonist APL. Defining antagonist APL in vitro using T cell clones, therefore, was not a reliable approach for the identification of APL with EAE-suppressing potential in vivo. Our findings highlight the complexities of the autoreactive T cell repertoire and have major implications for the use of APL in autoimmune diseases.