Screening And Identity Of The OPIDN-related Proteins In The Spinal Cord Tissue Of TOCP-treated Hens

Objective: To screen the proteins of differential expression in the spinal cordtissue of TOCP-treated hens which were protected by phenylmethylsulfonyl fluride(PMSF). Two-Dimensional electrophoresis and Mass Spectrometry were used to screenand idengtify the proteins. To use “PMSF and TOCP dual receptor impact hypothesis”explore the mechanism of TOCP induced OPIDN.Methods:56white and roman hens were randomly divided to four groups and14hens in one group. these four groups were designated as the control group, low doseTOCP (750mg/kg) exposure group, high dose of TOCP (1000mg/kg) exposuregroup and PMSF intervention group (this group first were given40mg/kg PMSF andthen treated in1000mg/kg TOCP after24hours) On the five days and twenty daysafter exposure, the animals were sacrificed,8hens spinal cord tissue from1000mg/kgTOCP group were used for pathological morphology observation, Two-Dimensionalelectrophoresis and Mass Spectrometry were used to screen and idengtify the proteinsfrom1000mg/kg TOCP group. The remaining hens from each group were used toobserve the OPIDN syndromes in the term of23days Total proteins of the spinal cordtissue tissues were separated by isoelectric focusing as the first dimension andSDS-PAGE as the second dimension. The2-DE maps were visualized and analyzed byImageMaster2D software.Results: Morphological observation: the two groups of TOCP exposure hensbegan to appear mild OPIDN on the five days and the symptoms gradually increasedwith the time going on. The average score value of OPIDN symptom was basicallyreached its peak on the20day and there was no changing in the last. In particular, thehigh dose group hens OPIDN symptoms were more serious; Histopathology observation:neurites disappear, myelin swelling and demyelination, neuronal necrosis, and collagenfiber hyperplasia were observerd in the23days groups of TOCP exposure hens spinal cord tissue. According to the PMSF features, there were23up and113down proteinmatch spots in TOCP group, which were significant difference compared with controlgroup and no significant difference compared with PMSF intervention group, therewere15proteins identified by using the mass spectrometry from spinal cord tissue inthe differences expression of the protein spots, which may be associated with OPIDN.The15proteins were Stathmin、THAP5、Pancreatic protein kinase A、cofilin-1-B、phosphotyrosine protein phosphatase、myelin basic protein1、Vesicle amine transportprotein1、Arginine kinase、malate dehydrogenase、HSP beta-1、electron transferflavoprotein、BMP、proteasome alpha1、Neurofilament protein.Conclusion: we got the stable OPIDN model and PMSF intervention model.According to the PMSF features; there were23up and113down protein match spots inTOCP group, there were14proteins identified by using the mass spectrometry fromspinal cord tissue in the differences expression of the protein spots; There were fivenerve tissue proteins、4protease-like proteins and four other related proteins. There wasone raised protein that was neurofilament protein.