title: [Expression of integrin α5 and β1 in osteoblast in the process of gingipains-induced apoptosis].
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
  Arginine-specific 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 α5 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 α5 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.
authors: Department of Periodontology, Guanghua School of Stomatology, Hospital of Stomatology, Sun Yet-sen University & Guangdong Provincial Key Laboratory of Stomatology, Guangzhou 510055, China.
authors: Jian-ying
authors: JY
authors: Zhang
authors: Department of Periodontology, Guanghua School of Stomatology, Hospital of Stomatology, Sun Yet-sen University & Guangdong Provincial Key Laboratory of Stomatology, Guangzhou 510055, China.
authors: Yun
authors: Y
authors: Fu
authors: Department of Periodontology, Guanghua School of Stomatology, Hospital of Stomatology, Sun Yet-sen University & Guangdong Provincial Key Laboratory of Stomatology, Guangzhou 510055, China.
authors: Xiang-chen
authors: XC
authors: Song
authors: Department of Periodontology, Guanghua School of Stomatology, Hospital of Stomatology, Sun Yet-sen University & Guangdong Provincial Key Laboratory of Stomatology, Guangzhou 510055, China. Email: liangm8@mail.sysu.edu.cn.
authors: Min
authors: M
authors: Liang
mesh_terms: D018829:Adhesins, Bacterial / Q000008:administration & dosage / Q000302:isolation & purification / Q000494:pharmacology*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D003546:Cysteine Endopeptidases / Q000008:administration & dosage / Q000302:isolation & purification / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D000080867:Gingipain Cysteine Endopeptidases; D039482:Integrin alpha5 / Q000378:metabolism*; D019012:Integrin beta1 / Q000378:metabolism*; D051379:Mice; D010006:Osteoblasts / Q000166:cytology / Q000378:metabolism; D016966:Porphyromonas gingivalis / Q000737:chemistry; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology; D013997:Time Factors; D014107:Tosyllysine Chloromethyl Ketone / Q000494:pharmacology
chemical_list: D018829:Adhesins, Bacterial; D000080867:Gingipain Cysteine Endopeptidases; D039482:Integrin alpha5; D019012:Integrin beta1; D015842:Serine Proteinase Inhibitors; D014107:Tosyllysine Chloromethyl Ketone; D003546:Cysteine Endopeptidases