title: Tanshinone IIA increases aquaporins expression in human amniotic epithelial WISH cells by stimulating GSK-3β phosphorylation.
abstract: BACKGROUND
  A recent study found that Salvia miltiorrhiza extract substantially increased amniotic fluid index in women with oligohydramnios. Abnormal aquaporin (AQP) expression is associated with oligohydramnios. This study aims to investigate the effect of tanshinone IIA, the major lipophilic component in the root of Salvia miltiorrhiza, on the function and expression of AQPs in human amniotic epithelial WISH cells and the molecular mechanism underlying the effect.
  
  
  METHODS
  WISH cells were treated with tanshinone IIA. Cell proliferation and swelling were determined by cell counting kit-8 assay and measurement of cell size, respectively. A human phospho-MAPK antibody array was used to screen tanshinone IIA-associated signaling molecules. The protein expression of AQPs, glycogen synthase kinase 3β (GSK-3β), and phosphorylated GSK-3β (p-GSK-3β) was analyzed by Western blot.
  
  
  RESULTS
  Tanshinone IIA significantly inhibited WISH cell proliferation at ≥50μM (P<0.05) and the IC50was 35μM. Cell size was significantly increased by the treatment with 35μM tanshinone IIA for 24h (P<0.05). WISH cells expressed AQP1, AQP3, AQP8, and AQP9 predominantly in cytoplasm. The dose of tanshinone IIA to maximally induce AQP1 and AQP3 expression was 35μM; for AQP8 and AQP9, the dose of maximal stimulation was 5μM (All P<0.05). Tanshinone IIA (35μM)-mediated up-regulation of AQP8, AQP9, AQP1, and AQP3 peaked at 6, 12, 24, and 24hours of treatment, respectively. Tanshinone IIA increased p-GSK-3β level significantly (P<0.05). Blockade of GSK-3β expression by siRNA and inhibition of GSK-3β activity by lithium chloride significantly reduced tanshinone IIA-induced AQP1 and AQP3 expression (All P<0.05).
  
  
  CONCLUSIONS
  Tanshinone IIA increases AQPs expression in WISH cells by stimulating p-GSK-3β. AQP8 responds to tanshinone IIA the most rapidly compared with AQP9, AQP1, and AQP3 in WISH cells.
authors.affiliation: Department of Obstetrics and Gynecology, The Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University, Wenzhou 325027, Zhejiang Province, China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Hua
authors.affiliation: Department of Obstetrics and Gynecology, The Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University, Wenzhou 325027, Zhejiang Province, China.
authors.forename: Shengdi
authors.initials: S
authors.lastname: Ding
authors.affiliation: Department of Obstetrics and Gynecology, The Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University, Wenzhou 325027, Zhejiang Province, China.
authors.forename: Huihui
authors.initials: H
authors.lastname: Cheng
authors.affiliation: Department of Obstetrics and Gynecology, The Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University, Wenzhou 325027, Zhejiang Province, China.
authors.forename: Hui
authors.initials: H
authors.lastname: Luo
authors.affiliation: Department of Obstetrics and Gynecology, The Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University, Wenzhou 325027, Zhejiang Province, China. Electronic address: zjwzzxq@163.com.
authors.forename: Xueqiong
authors.initials: X
authors.lastname: Zhu
mesh_terms: D045784:Abietanes / Q000494:pharmacology*; D000653:Amniotic Fluid / Q000166:cytology; D020346:Aquaporins / Q000378:metabolism*; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D000071679:Glycogen Synthase Kinase 3 beta / Q000378:metabolism*; D006801:Humans; D010766:Phosphorylation / Q000187:drug effects; D015854:Up-Regulation / Q000187:drug effects*
chemical_list: D045784:Abietanes; D020346:Aquaporins; C021751:tanshinone; D000071679:Glycogen Synthase Kinase 3 beta
keywords: Aquaporins; Cell size; Glycogen synthetic kinase 3β; Human amniotic epithelial WISH cells; Oligohydramnios; Tanshinone IIA