Effects Of Dexamethasone In The Action Of The Proliferation And Apoptosis Of Swine Kidney Fibroblast Induced By TNF-α Or TRAIL

Resently, many studies have showed that tumor necrosis factor-α(Tumor necrosis factor-α, TNF-α) not only could kill tumor cells selectively, but also play an important role in inflammation, autoimmune, shock and the others. In addition, TNF-αcan also effects the proliferation, apoptosis, collagen synthesis and catabolism on fibroblast.TNF-related apoptosis-inducing ligand (Tumor necrosis factor (TNF)-related apoptosis-inducing ligand, TRAIL) can induce apoptosis selectively in many transformed cells, the tumor cells and cells infected virus but not the normal cells.However, under some pathological or physiological state, TRAIL could also play some role on normal cells. In recent years, many studies found that TRAIL could induce RA (rheumatoid arthritis) FLS (fibroblast-like synovial cells) proliferation in vitro. In conclude, TRAIL plays an important role on the proliferation and apoptosis of fibroblasts.Dexamethasone is a kind of glucocorticoids. Under the physiological and pathological state dexamethasone have many effects on various cells. This study showed that dexamethasone could inhibite the constitutive OPG mRNA steady-state level in a dose dependent fashion and induced the osteoblasts to apoptosis. OPG is a decoy receptor for TRAIL. But it has poor homology with the other four receptor for TRAIL (DcR1, DcR2, DcR4, DcR5).OPG have an affinity with RANKL. It can inhibit the survival, differentiation, integration and mature of osteoclast cell. And it can also inhibit the action of osteoclast and increase the bone density. Due to the lack of death domain, the cells can't transduct the apoptotic signal.But what was the role of the OPG played on the balance of proliferation and apoptosis of swine kidney fibroblast, and how did the dexamethasone regulated the expression of OPG and RANKL and effected on TNF-αand TRAIL in FRSs had't been found.We adopted the methods of culture in viro and 3-(4,5-dimethylthiazol-2-yl)-2,5-dipheny- ltetraz-olium bromide (MTT) assay, The technique of flow cytometry (FCM), cell immunohistochemical and RT-RCR to research the role of TNF-αand TRAIL in the regulations of swine kidney fibroblast (FRSs) survival, proliferation or apoptosis and the effects of dexamethasone (DEX) related with them, and to explore the relevant gene pathway. From the investigation, main results were obtained as following:1. Primary FRSs isolated from swine kidney and were sub-cultured and cryopreservated. The result of Immunohistochemistry showed that: the cultured cells could express vimentin positively and factorⅧrelated antigen negatively. After the cells were cryopreservated, the morphology and cell viability still lies in good without bacteria and fungi contamination. The FRSs can be used to study the swine renal fibroblast in vitro.2. Tumor necrosis factor-α(TNF-α) and Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) prompt not only the growth but also the apoptosis of swine kidney fibroblast which depends on the concentrations of TRAIL. Under the lower concentrations of TNF-αand TRAIL can promote FRSs from the G0/G1 phase to the S and G2/M phase and promote the DNA synthesis in cell. Proliferation index (Prl.=S+G2/M) increased and the cells had high proliferation; Under the higher concentration, TNF-αand TRAIL could inhibit the proliferation and the process of G0/G1 phase to S phase on FRSs.3. Dexamethasone with TNF-αand TRAIL (lower concentrations) could promote FRSs proliferation and makes the G0/G1 phase decrease further and the Prl Increased more. These meant that dexamethasone can make more cells through the G1/S Checkpoint and regulate the cell cycle so as to promote cells proliferation.Dexamethasone with TNF-αand TRAIL (higher concentrations) could promote FRSs apoptosis and make the cell cycle arrest at G0/G1 so as to inhibit the FRSs proliferation or even made them apoptosis.4. Dexamethasone combined with TNF-αcould regulate the constitutive of OPG and RANKL mRANA steady-state in a dose-dependent fashion. TNF-αcould increase the expression of OPG but dexamethasone could inhibit it and up-regulate RANKL. Dexamethasone could offset the effect of TNF-αto regulate OPG with certain concentrations...