|Knowles, Donald - Don|
Submitted to: Veterinary Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/10/2007
Publication Date: 11/15/2007
Citation: Lawrence, P.K., Dassanayake, R.P., Knowles Jr, D.P., Srikumaran, S. 2007. Transfection of non-susceptible cells with Ovis aries recombinant lymphocyte function-associated antigen 1 renders susceptibility to Mannheimia haemolytica leukotoxin. Veterinary Microbiology. 125(1-2):91-99. Interpretive Summary: Mannheimia haemolytica is one bacterial component of pneumonia in sheep. The pathways by which this bacterium leads to pneumonia is being characterized. These data show that one of the components found on sheep cells (phagocytes) that are involved in controlling bacteria (Leukocyte functional antigen - 1) serves as a receptor for a major component (leukotoxin) of the infecting bacterium (Mannheimia haemolytica) involved in disease symptoms. A detailed understanding of how M. haemolytica causes disease is one pathway toward prevention and treatment.
Technical Abstract: Mannheimia haemolytica is an important etiological agent of pneumonia in domestic sheep (DS, Ovis aries). Leukotoxin (Lkt) produced by this organism is the principal virulence factor responsible for the acute inflammation and lung injury characteristic of M. haemolytica caused disease. Previously, we have shown that the leukocyte-specific integrins, beta(2) integrins, serve as the receptor for Lkt. Although it is certain that CD18, the beta subunit of beta(2) integrins, mediates Lkt-induced cytolysis of leukocytes, it is not clear whether CD18 of all three beta(2) integrins, LFA-1, Mac-1 and CR4, mediates Lkt-induced cytolysis of DS leukocytes. Since polymorphonuclear leukocytes, which express all three beta(2) integrins, are the leukocyte subset that is most susceptible to Lkt, we hypothesized that all three beta(2) integrins serve as the receptor for Lkt. The objective of this study was to determine whether DS LFA-1 serves as a receptor for M. haemolytica Lkt. We cloned the cDNA for DS CD11a, the alpha subunit of LFA-1, and co-transfected it along with the previously cloned cDNA for DS CD18, into a Lkt-non-suceptible cell line. Transfectants stably expressing DS LFA-1 were bound by Lkt. More importantly, Lkt lysed the DS LFA-1 transfectants in a concentration-dependent manner. Pre-incubation of Lkt with a Lkt-neutralizing monoclonal antibody (MAb), or pre-incubation of transfectants with MAbs specific for DS CD11a or CD18, inhibited Lkt-induced cytolysis of the transfectants. Exposure of LFA-1 transfectants to low concentrations of Lkt resulted in elevation of intracellular [Ca(2+)](i). Taken together, these results indicate that DS LFA-1 serves as a receptor for M. haemolytica Lkt.