An integrated in silico-in vitro investigation to assess the skin sensitization potential of 4-Octylphenol.

One of the major challenges in chemical toxicity testing is the possibility to protect human health against adverse effects with non-animal methods. In this paper, 4-Octylphenol (OP) was tested for skin sensitization and immunomodulatory effects using an integrated in silico-in vitro test approach. In silico tools (QSAR TOOLBOX 4.5, ToxTree and VEGA) were used together with several in vitro tests including HaCaT cells (quantification of IL-6; IL-8; IL-1α and IL-18 by ELISA and expression of genes TNF, IL1A, IL6 and IL8 by RT- qPCR), RHE model (quantification of IL-6; IL-8; IL-1α and IL-18 by ELISA) and THP-1 activation assay (CD86/CD54 expression and IL-8 release). Additionally, the immunomodulatory effect of OP was investigated using lncRNAs MALAT1 and NEAT1 expression and LPS-induced THP-1 activation (CD86/CD54 expression and IL-8 release). The in silico tools predicted OP as a sensitizer. In vitro tests are also concordant with the in silico prediction. OP increased IL-6 expression (HaCaT cells); IL-18 and IL-8 expressions (RHE model). An irritant potential was also shown by a great expression of IL-1α (RHE model); and increased expression of CD54 marker and IL-8 in THP-1 cells. Immunomodulatory effects of OP were demonstrated by the downregulation of NEAT1, MALAT1 (epigenetic markers), IL6 and IL8; and an increase in LPS-induced CD54 and IL-8 expressions. Overall, results indicate that OP is a skin sensitizer, being positive in three key events of the AOP for skin sensitization, also showing immunomodulatory effects.

Characterization of polysaccharides extracted from Platycodon grandiflorus (Jacq.) A.DC. affecting activation of chicken peritoneal macrophages.

Polysaccharides were isolated from Platycodon grandiflorus (Jacq.) A.DC. (PG) and the effects of three polysaccharides (PGPS80, PGPS60, PGPSt) on their immunological activities were studied. The structure identification of PGPSs was assessed using physicochemical and spectral methods. Results showed that PGPSt(2.67×105Da) compared to PGPS80(1.01×105Da) and PGPS60(1.12×105Da) has relatively higher average molecular weight(Mw) at the first peak with a narrower molecular weight distribution and all consisted of glucose, mannose, arabinose, galactose, xylose and rhamnose in different mass percentages. PGPS80 and PGPSt linked mainly by 1,3-and 1,6-ß-d-Galp residues. The immunological efficacy of PGPSs was performed on chicken peritoneal macrophages. Results showed that PGPSt significantly increased phagocytic rates, proliferation and NO production, stimulated macrophages to produce cytokines, including TNF-α, IL-1ß and IL-6 as well as stimulated macrophages to express the maturation markers CD80 and CD86. These findings suggest that PGPSt exerted significant immunological activity and might be associated with special characters.

Use of B7 costimulatory molecules as adjuvants in a prime-boost vaccination against Visna/Maedi ovine lentivirus.

RNA transcripts of the B7 family molecule (CD80) are diminished in blood leukocytes from animals clinically affected with Visna/Maedi virus (VMV) infection. This work investigates whether the use of B7 genes enhances immune responses and protection in immunization-challenge approaches. Sheep were primed by particle-mediated epidermal bombardment with VMV gag and env gene recombinant plasmids together with plasmids encoding both CD80 and CD86 or CD80 alone, boosted with gag and env gene recombinant modified vaccinia Ankara virus and challenged intratracheally with VMV. Immunization in the presence of one or both of the B7 genes resulted in CD4+ T cell activation and antibody production (before and after challenge, respectively), but only immunization with CD80 and CD86 genes together, and not CD80 alone, resulted in a reduced number of infected animals and increased early transient cytotoxic T lymphocytes (CTL) responses. Post-mortem analysis showed an immune activation of lymphoid tissue in challenge-target organs in those animals that had received B7 genes compared to unvaccinated animals. Thus, the inclusion of B7 genes helped to enhance early cellular responses and protection (diminished proportion of infected animals) against VMV infection.

B7 blockade alters the balance between regulatory T cells and tumor-reactive T cells for immunotherapy of cancer.

PURPOSE: In prostate cancer-bearing host, regulatory T (Treg) cells restrain activity of tumor antigen-specific T cells. Because B7:CD28 interactions are needed for both function of CD4+CD25+ Treg cells and CD8+ effective T cells, targeting this pathway may help to overcome the immunotherapy barriers. EXPERIMENTAL DESIGN: The anti-B7-1/B7-2 monoclonal antibodies were administered to a transgenic mouse model of prostate cancer (TRAMP) ectopically expressing SV40 large T antigen in different tumor development stages for prevention and therapy of prostate cancer. The treatment was also tested in treating transplanted MC38 colon adenocarcinoma in mice. RESULTS: Here, we showed that short-term administration of anti-B7-1/B7-2 monoclonal antibodies in TRAMP mice leads to significant inhibited primary tumor growth and the size of metastatic lesions. The treatment is effective to inhibit MC38 colon cancer growth. Correspondingly, this treatment results in a transient reduction of Treg in both thymus and the periphery. In vivo cytotoxicity assay revealed T antigen-specific CTL effectors in anti-B7-treated but not control IgG-treated TRAMP mice. CONCLUSIONS: Transient blockade of B7-1/B7-2 alters the balance between Treg and cancer-reactive T cells to enhance cancer immunotherapy.

Characterization of oligomers induced by inverse agonists of CTLA-4.

Although cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) inhibits T cell activation when ligated by B7 molecules on antigen-presenting cells, it can also act as an activating receptor when binding certain soluble recombinant ligands known as inverse agonists. Following ligation with an inverse agonist, we observed CTLA-4 microclusters evenly distributed on the T cell surface over a 60-min period. We have previously shown that the inverse agonist properties of these ligands correlate with their capacity to induce the formation of large CTLA-4 oligomers that are distinctly different from those resulting by CTLA-4 engagement with membrane-bound B7. These oligomers are composed of CTLA-4 molecules expressed on the cell surface and decrease from both the soluble cell lysate and lipid rafts upon cellular fractionation. Formation of these inverse agonist-induced CTLA-4 oligomers does not require an intact actin cytoskeleton. However, modulation of these oligomers was partially blocked upon actin depolymerization. Retention of CTLA-4 oligomers on the cell surface correlated with enhanced T cell signaling. Together, our data further characterize the structural basis of inverse agonist properties for CTLA-4 ligands that may be used in the design and screening of therapeutic biologicals targeting this receptor.