Background: A protective response against tetanus toxin and toxoid demands efficient specific T cell and B cell responses. Tetanus neurotoxin (TeNT), a 150 kDa polypeptide, is the main cause of tetanus disease. TeNT consists of two structurally distinct chains, a 50 kDa N-terminal light (L) and a 100 kDa C-terminal heavy (H) chain. C-terminal heavy (H) chain (fragment C) has two sub-domains named as proximal HCN and carboxy sub-domain or HCC. Beside neural binding property, HCC has been recently found as an immunodominant module of TeNT. In the present study, we investigated the effects of recombinant HCC (rHCC) on the expression of lineage specific transcription factors and secretion of a panel of functional cytokines including IFN-?, IL-4, and IL-17 from purified human T cells.
Methods: The heparinized peripheral blood samples were collected from 10 adult healthy volunteers with no history of tetanus after obtaining their informed consent. Peripheral blood mononuclear cells (PBMCs) were isolated by density gradient centrifugation on Ficoll–Paque from whole blood. T cells were isolated by MACS. Purified T cells were co-incubated with recombinant Hcc. Then, we evaluated the expression level of transcription factors (T-bet, GATA3 and ROR?T), the expression and secretion of cytokines (IFN-?, IL-4 and IL-17A) using Real-time PCR and ELISA. The capability of recombinant Hcc fragments in the activation of T cells was assessed by CD69 expression.
Results: The results showed that rHcc enhanced expression of CD69 on the surface of T-cells and promoted differentiation of CD4+ T-lymphocytes toward a T-helper1 (TH1) phenotype and up-regulation of interferon (IFN)-? secretion.
Conclusion: These results indicated that rHcc stimulates human Tcells to secrete IFN-? that maybe considered as a promising condidat for tetanus vaccine design in near future.