© 2004 Blackwell Publishing Ltd 119 Parasite Immunology , 2004, 26, 119–125 Blackwell Publishing, Ltd. ORIGINAL ARTICLE MAbs against a proteinase of T. vaginalis decrease parasite cytoadherence Monoclonal antibodies against a 62 kDa proteinase of Trichomonas vaginalis decrease parasite cytoadherence to epithelial cells and confer protection in mice H. HERNÁNDEZ, 1 I. SARIEGO, 1 G. GARBER, 2 R. DELGADO, 3 O. LÓPEZ 3 & J. SARRACENT 1 1 Parasitology Department, ‘Pedro Kourí’ Tropical Medicine Institute, Havana, Cuba, 2 Ottawa General Hospital, Ottawa, Ontario, Canada and 3 Pharmacology Department, Pharmaceutical Chemistry Institute, Havana, Cuba SUMMARY Trichomonas vaginalis infects the epithelium of the genital tract. The mechanism by which it invades the tissue leading to the disease is not thoroughly understood. However, results of several studies seem to agree that parasite adhesion to epithelium cells is the initial step leading to infection in women. T. vaginalis is associated with high levels of proteolytic activity. The role of some of these proteinases in the development of infection has been demonstrated. The current study establishes the role of a 62 kDa excretion–secretion proteinase in parasite cytoadher- ence. Monoclonal antibodies (MAbs) against this enzyme were tested for their ability to inhibit this process. Three stable hybrid producers of IgG 1 class MAbs (4D8, 1A8, 3C11) against the 62 kDa proteinase were obtained. Two of them (4D8 and 1A8) showed parasite recognition by immunofluorescence. Parasite cytoadherence to a monolayer of HeLa cells was inhibited by the 4D8, 1A8 and 3C11 antibodies. MAb 4D8 administered 24 h before a challenge with T. vaginalis by the intraperitoneal route was able to protect the majority of mice. Nitric oxide levels in the serum of animals inoculated with MAb 4D8 and challenged with the parasite were significantly different from those recorded in mice treated with an unrelated MAb. These studies show that an appropriate antibody against 62 kDa proteinase can help the host resist a challenge by the intraperitoneal route with T. vaginalis. Keywords cytoadherence, monoclonal antibody, protection, proteinase INTRODUCTION Around 170 million people worldwide suffer Trichomonas vaginalis infection each year. This is one of the most frequent sexually transmitted diseases, widely spread in all continents (1). This parasite is a major cause of vaginitis, cervicitis and urethritis, and in men it may lead to a non-gonococci ure- thritis, prostatitis and to some syndromes of the lower genital tract (2). Trichomonal control is important due to the high incidence of acute infections, complications and sequelae, as well as the possible role of Trichomonas in the transmission of HIV (3). The pathogenic mechanisms of T. vaginalis are unclear. The parasite infects the epithelium of the genital tract. Tri- chomonal cytoadherence to epithelial cells is a highly specific, critical step during the initiation phase of the infection and subsequent pathogenesis (4,5). The mechanisms of cytoadher- ence have been studied, relying on genetic and biochemical approaches (6,7). T. vaginalis, as is the case of other protozoan organisms, possess high levels of proteolytic activity, mainly of the cysteine- proteinase type. At least 23 cysteine-proteinases have been identified by two-dimensional electrophoresis (8). The role of some of these in the onset of the infection has been demonstrated (9). The parasite’s cysteine-proteinase activity is necessary for recognition and adhesion of the parasite to the superficial epithelial cells of the host (7). In 1989, Garber and Lemchuk-Favel (10) purified and characterized several (23 kDa, 30 kDa, 40 kDa and 60 kDa) excretion–secretion proteinases of T. vaginalis. using hide powder azure and casein as substrate to measure enzyme activity. A proteinase with molecular weight around 60 kDa was purified in the current study. We show here that this protein- ase plays a major role in parasite cytoadherence. A mono- clonal antibody (MAb) against this enzyme was found to be able to protect mice against infection challenge. Correspondence: H. Hernández, Parasitology Department, ‘Pedro Kourí’ Tropical Medicine Institute, Havana, Cuba (e-mail: hilda@ipk.sld.cu). Accepted for publication: 12 May 2004