title: [Regulation of sphingosine kinase 1 in the TNF-α-induced expression of MUC5AC in airway epithelial cells].
abstract: OBJECTIVE
  To explore the effects of sphingosine kinase 1 (SphK1) on mucin (MUC)5AC overexpression under the induction of tumor necrosis factor-α (TNF-α).
  
  
  METHODS
  The HBE16 airway epithelial cells were cultured, pretreated with SphK1-specific inhibitor DMS or transfected with SphK1 siRNA. Then each group was stimulated by a certain concentration of TNF-α. The relative contents of SphK1, COX-2, p-P38, p-ERK and IκBα were detected by Western blot while the levels of MUC5AC, PGE2 and cAMP analyzed by enzyme linked immunoassay. The transcription activities of cAMP-response-element-binding protein and nuclear factor κB (NF-κB) were detected by luciferase reporter gene detection system. The mRNA expressions of SphK1 and MUC5AC were detected by RT-PCR (reverse transcription-polymerase chain reaction). And the intracellular MUC5AC protein was detected by immunofluorescence.
  
  
  RESULTS
  The levels of protein and mRNA expression of SphK1 and MUC5AC were obviously elevated [(0.69 ± 0.12, 0.86 ± 0.07), (0.60 ± 0.09, 0.79 ± 0.05)]. They were all significantly higher than those in the control group (0.26 ± 0.03, 0.33 ± 0.04, 0.18 ± 0.06, 0.22 ± 0.10). There was an elevation of COX-2, PGE2, cAMP, p-P38, p-ERK, CREB and NF-κB activity (all P < 0.01); the production of IκBα protein was lower than that in the control group (P = 0.003). DMS decreased the levels of MUC5AC mRNA expression (0.37 ± 0.06) and protein production (0.27 ± 0.04), lowered the production of COX-2, PGE2, cAMP, p-P38 and p-ERK, inhibited the activity of CREB and NF-κB and increased the IκBα production (all P < 0.01). Transfection of SphK1 siRNA showed the similar effects as DMS (all P < 0.01).
  
  
  CONCLUSIONS
  SphK1 plays an important role in regulating the MUC5AC expression under the induction of TNF-α. And the activation of critical signal factors in that process is dependent on SphK1.
authors.affiliation: Department of Respiratory Medicine, Second Affiliated Hospital, Chongqing Medical University, Chongqing 400010, China.
authors.forename: Hong-Mei
authors.initials: HM
authors.lastname: Yu
authors.forename: Qi
authors.initials: Q
authors.lastname: Li
authors.forename: Juliy M
authors.initials: JM
authors.lastname: Perelman
authors.forename: Victor P
authors.initials: VP
authors.lastname: Kolosov
authors.forename: Xiang-Dong
authors.initials: XD
authors.lastname: Zhou
mesh_terms: D002478:Cells, Cultured; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D055271:Mucin 5AC / Q000378:metabolism*; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012137:Respiratory System / Q000166:cytology; D015398:Signal Transduction; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D000099157:Sphingosine Kinase
chemical_list: D055271:Mucin 5AC; D017853:Phosphotransferases (Alcohol Group Acceptor); D013110:Sphingosine; D014409:Tumor Necrosis Factor-alpha; D000099157:Sphingosine Kinase; C524678:MUC5AC protein, human; C061800:N,N-dimethylsphingosine