49.    Modification of antibody specificity by cytokines inhibition

Introduction: Lactate Dehydrogenase Elevating Virus (LDV) is a single stranded positive sense RNA enveloped arterivirus, is persistent, non pathogenic and induces NK, macrophages and B- cell activation in mice. It was found that LDV- infection modified Ab specificity to different antigens, depending on the genetic background
of the host. Objectives: The purpose of this work was to study, through of anticytokine auto-vaccination and monoclonal antibody (MAb) treatment, the effect in the change of specificity of antibodies to an model antigen (OVA). Methods: i) C57BL/6 mice were infected with 2×107 50% infectious doses of LDV in saline. Subsequently, the animals were inoculated in the footpads, four times at 2-wk intervals, with 2μg of IL-17/OVA complex emulsified in Gerbu adjuvant. Ten days after the last inoculation the animals were bled. ii) C57BL/6 mice were inoculated subcutaneously with 25 μg of OVA emulsified in phosphate-buffered saline (PBS) and Complete Freund’s Adjuvant. At day 15, the mice were boosted with the Ag in Incomplete Freund’s Adjuvant. At days 4, 7 and 11, half of the animals were inoculated intraperitoneally with 150 μg of anti-IL-17A MAb (MM17F3) in 200 μl of PBS. Another mouse group were treated as before but infected with 2×107 50% infectious doses of LDV in saline at day -1. Bleeding was performed at days 8, 21, and 30. IL-17 concentration was measured by Sandwich ELISA. The titer of Ab anti- OVA was determinated by Indirect ELISA whereas the proportion of Ab directed to
native OVA epitopes was calculated by ELISA competition assays. Results: Vaccination with IL-17/OVA decreased titers of anti-OVA Ab LDV-infected (1/290,000 ± 6,000) and non-infected animals (1/280,000 ± 17,000), respect to control mince only inoculated with OVA (790,000 ± 70,000, P<0.001). The proportion of native anti-OVA Ab in control C57BL/6 mice vaccinated with IL-17/OVA developed low titers of anti native OVA epitopes (38%) that increased to 71%, in LDV-infected animals. Furthermore, vaccination with IL-17/OVA augmented plasmatic IL-17 levels in LDV-infected animals in comparison with control (150±12 pg/ml and 24±6 pg/ml P<0.001, respectively). Whereas that in the MAb to IL-17A treatment titers of anti-OVA Ab in LDV-infected animals decreased by MAb
treatment (1/167,000 ± 1/13,000 to 1/68000±1/5,000, for control and MAb treated mice, respectively, P<0.001). Besides, percent of native anti-OVA Ab in non-infected mice increased by treatment with the MAb (48 to 61%, respectively). Same effect was shown in LDV- infected mice (65 to 87%, for control and treated animals, respectively) Conclusions: This work suggests that inhibition of a cytokine (IL-17A) induce a modification in the specificity of antibodies against conformational determinants to a given antigen.