title: Ultrastructural features of lymphocyte suppression induced by anthrax lethal toxin and treated with chloroquine.
abstract: Antibacterial therapy does not fully protect against anthrax because of severe systemic intoxication. Lysosomal processing of anthrax lethal toxin (LTX) is a key event in the disease pathogenesis, and agents interfering with this process, like chloroquine (CQ), may have practical applications. Although LTX is known to induce T-cell suppression, precise mechanisms of this phenomenon are not completely characterized. In the present study, we investigated alterations of lymphocyte ultrastructure caused by LTX and associated with favorable effect of CQ on the LTX-related dysfunction. Peripheral blood lymphocytes were activated via CD3 crosslinking in the presence or absence of LTX and CQ, and examined by transmission electron microscopy, flow cytometry and immunoblotting. Crosslinking of CD3 induced ultrastructural signs of lymphocyte activation, mostly disappeared after LTX treatment. The cell ultrastructure was well preserved in LTX-treated cells, despite dose- and time-dependent inhibition of T-cell function associated with impaired activation of mitogen-activated protein kinase. Regardless of intracellular signaling abnormalities, LTX did not decrease T-cell viability. CQ restored expression of CD69 (P<0.001) and improved phosphorylation of p38 (P=0.022) in LTX-exposed T lymphocytes. The exposure of cells to CQ, with or without LTX, led to appearance of many phagolysosomes with heterogeneous content, possibly representing unprocessed internalized material. In conclusion, LTX suppressed T-cell functions, but did not affect the viability and caused no ultrastructural damage. Ultrastructural observations indicated that CQ reduced harmful effects of LTX, possibly by interfering with lysosomal activity.
authors.affiliation: Laboratory for Shock and Trauma Research, Ruth and Bruce Rappaport Faculty of Medicine, Technion, Haifa, Israel. hirsh@tx.technion.ac.il
authors.forename: Mark I
authors.initials: MI
authors.lastname: Hirsh
authors.forename: Irena
authors.initials: I
authors.lastname: Manov
authors.forename: Victoria
authors.initials: V
authors.lastname: Cohen-Kaplan
authors.forename: Theodore C
authors.initials: TC
authors.lastname: Iancu
mesh_terms: D000704:Analysis of Variance; D000942:Antigens, Bacterial / Q000633:toxicity*; D001427:Bacterial Toxins / Q000633:toxicity*; D002738:Chloroquine / Q000494:pharmacology*; D005434:Flow Cytometry; D006801:Humans; D015151:Immunoblotting; D046529:Microscopy, Electron, Transmission; D013601:T-Lymphocytes / Q000187:drug effects* / Q000648:ultrastructure*
chemical_list: D000942:Antigens, Bacterial; D001427:Bacterial Toxins; C030325:anthrax toxin; D002738:Chloroquine