ISOLATION AND PURIFICATION OF GLUTATHIONE-S-TRANSFERASE (GST) AND ITS ROLE ON OXIDATIVE STRESS IN HEPATIC TISSUES OF MAREK’S DISEASE AFFECTED CHICKEN
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Date
2017-01
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SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA
Abstract
ABSTRACT:
A study was conducted to isolate and purify Glutathione-S-transferase
(GST) and to determine its role on oxidative stress in liver tissues of Marek’s
disease (MD) affected chicken. Ten healthy and MD suspected layer birds of
35-40 weeks age for each group were procured from poultry farms in Chittoor
district of A.P. The postmortem examination of MD affected birds revealed
enlarged liver with discrete grayish nodules of various sizes. Histopathological
examination of liver revealed extensive proliferation and infiltration of
pleomorphic cells. The purified DNA samples from liver tissue of MD
suspected birds were further used for PCR analysis which was found to be
positive yielding a 314 bp product indicating the presence of two copies of
132 bp tandem repeat and a 50 bp primer sequence. The liver samples positive
for MD were used for further analysis.
The results showed a significant increase of TBARS (179.82%) in liver
of MD affected chicken compared to control. Glutathione (GSH) levels were
significantly reduced (52%) in liver of MD affected chicken compared to
control. A significant decrease in SOD (32%), catalase (43%), GPx (55%) and
GST (51%) activities were observed in MD affected chicken compared to
control. The GST enzyme was isolated, purified and characterized from liver
tissue of both healthy and MD affected layer chicken using glutathione
agarose affinity chromatography. The enzyme was purified to 2.38 fold with a
yield of 32.27% in healthy chicken whereas in MD affected chicken it was
purified to 4.54 fold with a yield of 41.98%. The analysis of active fractions
using SDS-PAGE showed GST-α expression with a molecular weight of
25.6kDa in both healthy and MD affected liver. In addition GST-π was
expressed in MD affected chicken with a MW of 27 kDa. These results
confirm that Marek’s disease in chicken induces oxidative stress which in turn
leads to alterations in antioxidant enzymes. The oxidative stress induced in
liver is responsible for GST π expression as a defense mechanism in MD
affected chicken.
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