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Fig. 3. ADI participates in the control of cell death, probably by altering VSP switching. (A) Cytotoxicity assays of G. lamblia trophozoites WB/1267, GS/H7, WB/1267 transfected with H7-HA or WB/1267 transfected with R-H7 (from left to right on x axis) analyzed after 24 hours post-addition of the anti-VSPH7-specific antibodies G10-4 and anti-GS-VSPs pAb by estimating the number of adherent viable parasites. Controls include cells without treatment (w/o mAb) and the use of a non-related mAb (8G8 mAb). Data represent the means ± s.d. for n=2 of three independent experiments. (B) Progenies were analyzed by addition of goat anti-mouse FITC-conjugated antibody in IFA. Positive cells correspond to trophozoites expressing VSPH7. DIC, differential interference contrast. Scale bars: 10 µm. (C) VSP expression is established in WB/9B10 wild-type and WB/9B10-ADI+ transgenic trophozoites after a short time exposure to specific VSP9B10 mAb. Controls had no exposure to the mAb (–). Data represent the means ± s.d. for n=3 of two independent experiments. (D) VSP9B10 citrullination is analyzed by western blotting in WB/9B10 and WB/9B10-ADI+ trophozoites after a short time exposure to the anti-VSP9B10 mAb. The membrane labeled with anti-citrulline pAb (right) was stripped and re-blotted with anti-VSP9B10 mAb (left). A similar amount of loaded VSP9B10 is observed. Molecular mass of VSP9B10 is indicated on the right.

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