Expression of integrin α5 and β1 in osteoblast in the process of gingipains-induced apoptosis
- Author:
- No author available
- Journal Title:
- Chinese Journal of Stomatology
- Issue:
- 11
- DOI:
- 10.3760/cma.j.issn.1002-0098.2013.11.004
- Key Word:
- 细胞凋亡;整合素α5;整合素β1;牙龈蛋白酶;Apoptosis;Integrin alpha5;Integrin beta1;Gingipains
Abstract： Objective To investigate the regulatory mechanisms of integrin α5 and β1 in osteoblast in the process of gingipains-induced apoptosis.Methods Gingipains were isolated and purified from supernatants of Porphyromonas gingivalis W83 which was cultured under standard anaerobic conditions.MC3T3-E1 was challenged with or without 8.3480 U/L gingipains for 48 h and apoptosis was examined by transferase-mediated deoxyuridine triphosphate-biotin nick end labeling-(4-Amidinophenyl)-6-indolecarbamidine dihydrochloride (TUNEL-DAPI) staining.The expression of integrin α5 and β1 was analyzed by Western blotting after MC3T3-E1 was treated under different conditions.Results Argininespecific proteinases (Rgp) activity was (41.74 ± 2.11) U/L and lysine-specific proteinase (Kgp) was (1.02 ±0.25) U/L.Gingipains induced MC3T3-E1 cells apoptosis after 48 h.Compared with control group,expression of integrin α5 and β1 was down-regulated by gingipains in a time-dependent manner within short periods (≤72 h),integrin o5 and β1 relative expression was (0.485 ±0.039),(0.504 ±0.002) at 48 h,(0.398-±0.058),(0.179 ±0.001) at 72 h respectively (P ＜0.05).After 72 h,integrin α5 expression in MC3T3-E1 cells was stable compared with control group while integrin β1 was still lower(control group:1.000 ± 0.000,96 h:0.604 ± 0.003,120 h:0.357 ± 0.002) (P ＜ 0.05).Proteinase inhibitor tosyl-L-lysine-chloromethyl-ketone(TLCK) effectively blocked the activity of gingipain and inhibited down-regulation of integrin α5 and β1 induced by gingipains from (0.398 ± 0.058,0.179 ± 0.001) to (0.781 ± 0.012,0.857-±0.060) (P ＜ 0.05).TLCK alone did not have any effect on integrin α5 and β1 (P ＞ 0.05).Gingipains also decreased integrin α5 and β1 in a dose-dependent manner.When cells were treated with 20.8700 U/L gingipains,integrin α5 and β1 relative expression reached to the lowest (0.105 ± 0.004,0.020 ± 0.000) (P ＜ 0.05).Conclusions Gingipains inhibited the expression of integrin o5 and β1 in a time-and dose-dependent manner in osteoblasts in the process of apoptosis,which may not be mediated by direct proteolytic effect.
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