Identification and quantification of proteases in germinal cells of Echinococcus granulosus.

ALBANI CLARA MARÍA; REINDERS JOERG; FABBRI, JULIA; PENSEL, PATRICIA EUGENIA; ELISSONDO MARIA CELINA

Mar del Plata

Congreso; Reunión Conjunta SAIC-SAFE- SAB-SAP-NANOMED-ar- AACYTAL -2019; 2019

The cestode Echinococcus granulosus (Eg) is the causative agent of cystic echinococcosis, a severe neglected zoonosis which has important medical and economic impact. The inner layer of the hydatid cyst or ?germinative layer? contains a group of cells called germinative cells (GC). These cells are responsible for cell proliferation and differentiation. Eg has the capability to survive for long periods of time in many mammalian host species. The prolonged survival indicates that the parasite is able to evade host attack for example through the production and release of proteases to digest host proteins. The aim of the present work was to identify and to quantify proteases in Eg GC grown in different conditions using a proteomic strategy. Eg primary cell culture was obtained from hydatid cysts and maintained with weekly splitting during 1-4 month. Cells were culture in normal media and conditioned media (resembling host environment) for different time periods. Then, 50 µl of cells were homogenized in lysis buffer and the peptide content was estimated. Aliquots of 100 µg protein per sample were used for filter-aided sample preparation and the resulting peptides were subjected to nano-LC-MS/MS-analysis. A combined library was set-up by combining the different runs using the Protein Pilot-software and Uniprot database. We identified a total of 455 proteins in all the studied conditions. We identified and quantified 7 different proteases which were differentially represented in the studied conditions. For example Calpain A (Accession U6J063) and Mitochondrial processing peptidase beta subunit (Accession U6JE67) were found overrepresented in cells cultured in normal media with a fold change 12 and 2.8 respectively (p < 0.05). Through the methodology used, it was possible to describe the presence of several proteases in the GC of Eg, suggesting that some evasion mechanisms are present.