Effect On Apoptosis And Antioxide Capacity Of Piglets Lymphocyte Inoculated Of PCV2 In Vitro

Porcine circovirus2(PCV2) ties up with post weaning multisystemic wastingsyndrome(PMWS), which has recently emerged as an important disease of pigs in theworld. PCV2 has nearly relationship with many diseases of pigs, including PMWS,porcine dermatitis and nephropathy syndrome(PDNS), reproductive failure, congenitaltremors(CT) , porcine respiratory disease complex(PRDC), porcine proliferate andnecrotizing pneumonia(PNP). It has been studied that PCV2 has an effect on immunesystem. But it hasn't been resolved that how the virus has an effect on the immune system.And the mechanism of PCV2 causing disease has not been made sure. The purpose of ourstudy is to make sure in vitro whether PCV-2 can induce piglets lymphocyte apoptosis,and analyze the function of oxygen free radical (OFR) in this process.In this report 4 conventional piglets free of PCV-2 antibody were used. Lymphocytesof spleen separated with pinhead of injectors were distributed in 2 groups: control andPCV2 inoculated. Apoptosis was qualitative analyzed by agarose gel electrophoresis DNAand eletronic microscope at different time. We mensurated cell cycle, the rate of apoptosisand the expression of CASPASE-3 by flow cytometry(FCM). The result of agarose gelelectrophoresis DNA show the special DNA ladder of apoptosis; we can see morphologicalchanges of apoptosisatdifferentphase. At 0, 2, 4, 6, 12, 24, 36, 48 hours after cellculturing, the average apoptotic rate(percent) of control is 0.13±0.02, 2.10±0.56, 5.01±1.22, 6.70±1.62, 11.60±2.62, 15.64±3.81, 30.19±4.1, 37.08±4.26; and the PCV2group is 0.13±0.02, 3.33±0.66, 15.46±3.21, 17.76±3.41, 23.21±4.62, 27.88±4.98,41.23±5.45, 49.53±6.55 respectively. The data are prominent higher than that of thecontrol group. The proliferation index at each time, control is 38.53±4.39, 29.28±3.71,29.13±3.41, 24.88±4.65, 17.88±3.63, 12.97±1.70, 12.43±2.66, 10.04±0.35;thePCV2 group is 38.53±4.39, 14.95±3.05, 14.33±2.31, 14.30±3.36, 12.79±2.85, 12.13±1.30, 7.44±0.10, 7.04±0.26. Compared the two group, there is no difference at 0h,at 2, 4, 6, 12, 24h there is significance at 5%, but little difference at 36, 48h (P＞0.05 ).So we can say that PCV2 may restrain lymphocyte in vtro culturing from proliferation at acertain time. The percentage of cell expressed CASPASE-3 which plays a key protein in the progress ofapoptosis, control is2.12±0.69, 3.60±0.38, 5.33±0.89, and the PCV2 groupis 2.12±0.69, 7.75±0.93, 17.42±3.70 at 0, 6, 48h, the data of PCV2 group is prominenthigher than that of the control group(P＜0.05) at 6 and 48h. The result indicate thatapoptosis rate of lymphocytes in vitro culturing increasing and proliferation capacitydeclining induced by PCV2. CASPASE-3 is an important conditioner in this apoptoticprogress induced by PCV2.For farther finding out the mechanism of lymphocyte apoptosis induced by PCV2,we mensurated the antioxide capacity of lymphocyte in vitro culturing infected by PCV2.The total antioxygen capability, antisuperoxide anion capability and superoxide dismutaseof culture supematant at different time were examined. At 0, 2, 6, 12, 24, 48h afterculturing, total antioxygen capability of PCV2 group is 1.05±0.053 , 1.82±0.188,3.89±0.109, 4.65±0.242, 6.03±0.188 (U/ml), and that of control is 1.04±0.039,1.50±0.053, 2.29±0.062, 3.25±0.295, 4.60±0.435 (U/ml), there is no prominentdiscrepancy between the two group; the antisuperoxide anion capability of PCV2 group is79.89±2.43, 106.72±2.35, 168.16±5.61, 249.41±4.88, 471.93±4.23 (U/L), that ofcontrol is 70.12±1.18, 95.02±3.05, 133.61±2.32, 202.24±6.29, 261.44±3.47 (U/L),the data of the two groups indicate significance at 5% except Oh. Superoxide dismutase ofPCV2 and control is 19.13±1.86, 18.88±1.48, 18.58±1.45, 18.03±1.02, 19.30±1.59(U/ml) and 19.14±0.67, 18.82±1.27, 19.09±0.87, 18.90±1.77, 18.27±1.21 (U/ml),which all holding a level of 19U/ml show unnotable difference. The results indicated thatPCV2 inoculated has certain effect on the antioxide capability of lymphocyte in vitro.All in all, PCV2 infection can induce apoptosis and inhibit proliferation of lymphocytein vitro culture. In this progress, CASPASE-3 is an important conditioner, andantisuperoxide anion has effect on too.