Evaluation of Matrix Metalloproteinase Gene Expression in Infected Cells to Theileria annulata.

Theileria annulata, the causative agent of Bovine Tropical Theileriosis is a tick-born parasitic protozoan. The sporozoites of Theileria annulata invade bovine MHC II cells, where they differentiate into schizonts. Attenuated vaccines are an important means of controlling Theileria annulata infection of cattle. Vaccine production is by prolonged cultivation of macroschizont-infected cells. The mechanism of attenuation remains unclear. The intracellular apicomplexan parasite, Theileria annulata, manipulates its bovine host cell by over-riding the cells natural apoptotic response and results in their reversible transformation such that become immortalised and metastatic. The latter can immortalize and induce fundamental changes in their host cells. The mechanisms underlying this transformation are not understood. In this study, the evaluated cells were Sa Theileria annulata infected cell line vaccine strain and two newly isolated and 44 established Theileria annulata-infected cell lines "C1 and C2". The purposes of this work were to analyze that prolonged in vitro culture of Theileria-infected cell lines results in their attenuation and this process is associated with alterations in both host and parasite gene expression. Particularly, it was reported a loss in bovine MMP9 expression in high passage cultures suggests that this parasite-induced MMP9 is a virulence factor. Of three MMP2, MMP9 and MMP13 genes studied in this study, the results showed in high passage cultures and attenuated Sa cell line, the level of specific transcripts for MMP9 was decreased in attenuated cell line (245-283) and in high passage culture of C1 and C2 cell lines (25 and more). Whereas, it was detectable in passage number (25 and less). Tash HN nucleoprotein was recently identified with characteristics that indicate is involved in control of host cell gene expression. The analyses of three available cell lines indicated increase in Tash HN gene expression, with host nuclear reactivity particularly cell proliferation. In addition the level of transcription for pro-inflammatory cytokines (IL-1ƒز and TNFƒز) exhibited no noticeable correlation between specific cytokine transcripts and attenuation in low and high passage number of three available cell lines. Analysis of Tams1 gene expression in Sa vaccine, C1 and C2 cell lines determined the detectable transcripts for Tams1 gene in low passage of C1 and C2 but no specific transcripts was detectable for high passage culture of these two cell lines as well as Sa vaccine strain. In conclusion, the results of this work clearly showed that the level of MMP9 transcripts are in contrast with the amounts of Tams1 mRNAs in T.annulata schizont infected cell lines for virulence and attenuation respectively. Understanding the mechanisms of virulence, attenuation of infected cell line by using molecular biology methods and in vivo animal experiments could help to increase our knowledge about attenuation mechanisms and preparing and identifying appropriate cell lines in order to develop the new Theileria annulata vaccine cell lines.