Studies On Morphogenesis Of PRRSV And Ultrastructure Changes And Apoptosis Of Marc-145 Cells Induced By PRRSV In Vitro

Morphogenesis of PRRSV, ultrastructural changes and apoptosis were studied in Marc-145 cell culture which was infected by porcine reproductive and respiratory syndrome virus Sichuan isolates strain (PRRSV-SCl). The study included two aspects: (1) morphogenesis of PRRSV and ultrastructural changes of PRRSV-infected Marc-145 cells were studied by electron microscopy and ultrathin section techniques; (2) morphologic changes and dynamic changing rule of apoptosis of PRRSV-infected Marc-145 cells were studied by optical microscopy, DNA Ladder. ELISA (Enzyme-Linked Immuno Sorbent Assay) and TUNEL(terminal deoxy nucleotidyltrans ferase-mediated digoxigenin-UTP nick end labeling).1. The rusults of electron microscopy and ultrathin section techniques indicated that PRRSV is an enveloped, global virion, 45 to 65 nm in diameter, with a nucleocapsid approximately 25 to 30 nm in diameter. PRRSV enter into the cell by endocytosis. nucleic acid of virus release out of vesicle after intemalization and copy in cytoplasm. Some nucleocapsids bud into the lumen of the smooth endoplasmic reticulum (SER) and obtain their tegument from the membrance of SER. then accumulate in SER. which move to the cytoplasmic membrane where they fuse to release virions by exocytosis. Other nucleocapsids release to extracellular by budding and obtain their tegument from the cytoplasmic membrance.2. Ultrastructural changes of PRRSV-infected Marc-145 cells. In the infected prophase, cells had no obvious ultrastructural changes. Along with infected time extended, the smooth endoplasmic reticulum (SER) dilated and became vaculization. the rough endoplasmic reticulum (RER) slightly dilated, and some ribosome particles dropped from it. Plastosomes in the cytoplasm were hyperplastic and their cristae collapsed and lysed. Some cells presented characteristic morphological changes of apoptosis: the cytoplasm concentrated, the chromatin condensed into lunate or orbicular mass, karyotheca of some cell disappeared and nucleus lysed. then the membrance became sunken and the cell wasdivide up apoptotic bodies with membrance enwraped. In the infected anaphase, organelles of whole cell became vacuolization. some high electron density paticles. lysosome and lipochrome presented in the cytoplasm. Finally, the cell lysed and fell to pieces.3. Apoptosis of PRRSV-infected Marc-145 cells. (1 Characteristic morphological changes of apoptosis were observed by optical and electron microscopy in PRRSV-infected Marc-145 cells, such as extensive condensation of chromatin into lunate or orbicular masses and concentration of cytoplasm associated with lots of cytoplasmic vacuolization, and aopototic bodies come into being. (2) The results of DNA Ladder, ELISA and TU'NEL indicated that the level of apoptosis of PRRSV-infected Marc-145 cells was variational with the infected time extend. The level of apoptosis of infected group cells was unconspicuous compared with control group (P>0.05) at 24h p.i. The level of apoptosis of infected group cells was very conspicuous(PO.Ol) compared with control group at 48h , 72h, 96h, 120h p.i. The peak value of apoptosis appeared at 72h p.i.. the level of apoptosis began to drop at 96h and 120h p.i. At 144h p.i., DNA Ladder couldn't detect apoptosis. the level of apoptosis detected by ELISA was lower than control group, and TUNEL aslo detected a little apoptosis. The dynamic changing rules of apoptosis detected by three biochemic mothods were almost identical.