title: The influence of actin depolymerization induced by Cytochalasin D and mechanical stretch on interleukin-8 expression and JNK phosphorylation levels in human retinal pigment epithelial cells.
abstract: BACKGROUND
  This study explores the role of actin cytoskeleton depolymerization induced by Cytochalasin D and mechanical stretch on the interleukin-8 (IL-8) expression and c-jun N-terminal kinase (JNK) phosphorylation levels in human retinal pigment epithelial (RPE) cells.
  
  
  METHODS
  A Flexcell FX-5000 Tension system was used to apply cyclic stretch to cultured human RPE cells (ARPE-19) at 0.33 Hz with 20% elongation for 0 h, 6 h or 24 h. The cells were stretched alone or pre-treated with Cytochalasin D. The redistribution of the actin cytoskeleton was evaluated using phalloidin immunofluorescence staining. The protein expression levels of IL-8 and JNK in the RPE cells were determined via Western blotting.
  
  
  RESULTS
  The cells in the control groups displayed abundant and uniform phalloidin staining. After exposure to mechanical stretch for 24 h, phalloidin staining revealed an unclear and irregular actin cytoskeleton. In all Cytochalasin D-treated cells, the shrinkage and disruption of the cytoskeletal structure was observed regardless of mechanical stress. The stimulation of the RPE cells with cyclic stretch alone did not induce a significant increase in IL-8 expression and JNK phosphorylation levels, which were similar to those of the control groups. After pre-treatment with Cytochalasin D alone, IL-8 expression and JNK phosphorylation levels were not significantly different at 6 h but were significantly increased by approximately 1.2-fold (1.18 ± 0.05; P<0.01) and 3.0-fold (3.01 ± 0.02; P<0.01) at 24 h, respectively. After the pre-incubation of the RPE cells with Cytochalasin D followed by exposure to cyclic stretch, IL-8 expression and JNK phosphorylation levels increased by approximately 1.3-fold (1.31 ± 0.02; P<0.01) and 1.3-fold (1.31 ± 0.02; P<0.01) at 6 h, respectively, and by 1.7-fold (1.69 ± 0.06; P<0.01) and 3.2-fold (3.21 ± 0.12; P<0.01) at 24 h, respectively.
  
  
  CONCLUSIONS
  This study demonstrates that disruption of actin polymerization by cytochalasin D and mechanical stretch upregulates interleukin-8 expression and JNK phosphorylation levels in human RPE cells, which indicates that the integrity of the actin cytoskeleton may play important roles in the pro-inflammatory processes in RPE cells.
authors.affiliation: Beijing Tongren Eye Center, Beijing Tongren Hospital, Capital Medical University; Beijing Ophthalmology and Visual Sciences Key Laboratory, Beijing, China.
authors.forename: Meng
authors.initials: M
authors.lastname: Gao
authors.affiliation: Beijing Institute of Ophthalmology, Beijing Tongren Eye Center, Beijing Tongren Hospital, Capital Medical University; Beijing Ophthalmology and Visual Sciences Key Laboratory, Beijing, China.
authors.forename: Shen
authors.initials: S
authors.lastname: Wu
authors.affiliation: Key Laboratory for Biomechanics and Mechanobiology of Ministry of Education, School of Biological Science and Medical Engineering, Beihang University, Beijing, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Ji
authors.affiliation: Beijing Institute of Ophthalmology, Beijing Tongren Eye Center, Beijing Tongren Hospital, Capital Medical University; Beijing Ophthalmology and Visual Sciences Key Laboratory, Beijing, China.
authors.forename: JingXue
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Beijing Institute of Ophthalmology, Beijing Tongren Eye Center, Beijing Tongren Hospital, Capital Medical University; Beijing Ophthalmology and Visual Sciences Key Laboratory, Beijing, China.
authors.forename: Qian
authors.initials: Q
authors.lastname: Liu
authors.affiliation: Department of Ophthalmology, Beijing Fuxing Hospital, Capital Medical University, Beijing, China.
authors.forename: YanKun
authors.initials: Y
authors.lastname: Yue
authors.affiliation: Department of Ophthalmology, Beijing Fuxing Hospital, Capital Medical University, Beijing, China.
authors.forename: Lu
authors.initials: L
authors.lastname: Liu
authors.affiliation: Department of Ophthalmology, Kailuan General Hospital, Tangshan, China.
authors.forename: XinXin
authors.initials: X
authors.lastname: Liu
authors.affiliation: Beijing Tongren Eye Center, Beijing Tongren Hospital, Capital Medical University; Beijing Ophthalmology and Visual Sciences Key Laboratory, Beijing, China. wuliubj@sina.com.
authors.forename: Wu
authors.initials: W
authors.lastname: Liu
mesh_terms: D000199:Actins / Q000378:metabolism*; D015153:Blotting, Western; D002478:Cells, Cultured; D015638:Cytochalasin D / Q000494:pharmacology*; D003599:Cytoskeleton / Q000473:pathology; D006801:Humans; D016209:Interleukin-8 / Q000096:biosynthesis*; D048670:MAP Kinase Kinase 4 / Q000378:metabolism*; D018613:Microscopy, Confocal; D010766:Phosphorylation; D055213:Retinal Pigment Epithelium / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D013314:Stress, Mechanical
chemical_list: D000199:Actins; C508606:CXCL8 protein, human; D016209:Interleukin-8; D015638:Cytochalasin D; D048670:MAP Kinase Kinase 4
keywords: Cytochalasin; Cytokines; Cytoskeleton; JNK Mitogen-activated protein Kinases; Mechanical; Retinal pigment epithelium; Stress