ABSTRACT
Hydatidosis is a zoonosis disease caused by the larva of Echinococcus granulosus parasite in man and animals. The Proteases enzymes are necessary for nutrition, migration and evasion of immunity of parasite into host. The aim of this study was to determine the protease activity of hydatid cyst protoscolices [PSC], healthy and cystic liver tissues in order to compare of this biomarker for host and parasite in hydatid disease. In this study, PSC were collected from sheep liver tissue infected with hydatid cysts at an abattoir and washed 3 times with PBS buffer. PSC samples were freeze-thawed and sonicated while collected liver tissues were homogenized. Extract solution samples were centrifuged and stored at - 20degree C. Protease enzymes activity was measured in the extract solutions of PSC and sheep liver tissue samples [healthy and cystic livers]. Samples protein concentrations and protein bands were detected using Bradford and sodium dodecyl sulfate polyacrylamide gel electrophoresis [SDS-PAGE] methods respectively. To determine significant difference between two groups, statistical t-test was performed. The values of protease enzyme activities in healthy, cystic and PSC were determined 0.0028, 0.0087 and 0.50U/ml/mg respectively. Elevation of protease enzyme activity in cystic liver as compared with healthy was not significant. Statistical T-test showed higher protease enzyme activity for PSC as compared to healthy [p<0.05]. SDS-PAGE confirmed 24 kDa and 54 kDa bands for protease enzyme in PSC samples and 24 kDa band in liver samples. Protease enzyme activity and molecular weight as compared to healthy liver tissues could be concerned as a comparative metabolic biomarker for host and parasite in hydatidosis
ABSTRACT
Fascioliasis is an endemic disease in Iran and triclabendazole [TCBZ] is using for treatment of domestic animals and infected people. Excretory-secretory products [ESP] play an important role in the host biochemical defense by means of activities of detoxifying and antioxidant glutathione S-transferase [GST] and superoxide dismutase [SOD] enzymes respectively. Therefore, the aim of this comparative study was to evaluate fasciola protection against TCBZ drug by detection of enzymatic activities, GST and SOD, in TCBZ treated Fasciola hepatica / Fasciola gigantica and control ESP samples. F. gigantic and F. hepatica helminthes were collected and cultured within buffer media [TCBZ treated and untreated or control] for 4 h at 37[degree]C. Three TCBZ treated and 1 control ESP samples for each species were collected, centrifuged and supernatants were stored at-20°C. ESP samples protein concentrations were measured by Bradford method. SOD and GST enzymes activities of ESP samples were estimated photometrically. To determine the statistically significant difference between ESP of treated and control samples, t-test was conducted. ESP protein bands were detected by sodium dodecyl sulfate polyacrylamide gel electrophoresis [SDS-PAGE]. Protein concentrations in treated F. hepatica and F. gigantica ESP samples were estimated 204.88, 428, 130.4 and 288.2, 488.2, 308.2 micro g/ml respectively. Protein concentrations in control samples were estimated 488.18 and 124.8 micro g/ml respectively. SOD enzyme specific activities level in treated F.hepatica and F. gigantica ESP samples were determined 0.14, 0.31, 3.96 and 11.11, 13.54, 19.95 U/mg/protein respectively. SOD activities level in control samples were detected 70.69 and 10.92 U/mg/protein. GST specific activities level in treated F.hepatica and F. gigantica ESP samples were calculated 25.3, 85.5, 37.3 and 1823, 1314.3, 1320.8 U/mg respectively. GST activities levels in control samples were detected 98.6 and 1083.9 U/mg/protein respectively. Statistical analysis reveal the significant different between proteins concentrations, GST and SOD enzyme specific activities of TCBZ treated ESP samples of F. gigantic in comparison to the control samples [P<0.05], however, is not significant for treated F. hepatica and control ESP samples [P>0.05]. There is no difference between SDS-PAGE results of treated and control samples. Based on the results of the present work, significant increase of enzymatic activities of GST and SOD in TCBZ treated F. gigantica ESP, it seems, the protection of this species against drug is higher than F. hepatica