title: High glucose alters Cx43 expression and gap junction intercellular communication in retinal Müller cells: promotes Müller cell and pericyte apoptosis.
abstract: OBJECTIVE
  To investigate whether high glucose (HG) alters connexin 43 (Cx43) expression and gap junction intercellular communication (GJIC) activity in retinal Müller cells, and promotes Müller cell and pericyte loss.
  
  
  METHODS
  Retinal Müller cells (rMC-1) and cocultures of rMC-1 and retinal pericytes were grown in normal (N) or HG (30 mM glucose) medium. Additionally, rMC-1 transfected with Cx43 small interfering RNA (siRNA) were grown as cocultures with pericytes, and rMC-1 transfected with Cx43 plasmid were grown in HG. Expression of Cx43 was determined by Western blotting and immunostaining and GJIC was assessed by scrape-loading dye transfer (SLDT) technique. Apoptosis was analyzed by TUNEL or differential staining assay, and Akt activation by assessing Akt phosphorylation.
  
  
  RESULTS
  In monocultures of rMC-1 and cocultures of rMC-1 and pericytes, Cx43 protein level, number of Cx43 plaques, GJIC, and Akt phosphorylation were significantly reduced in HG medium. Number of TUNEL-positive cells was also significantly increased in rMC-1 monocultures and in rMC-1 and pericyte cocultures grown in HG medium. Importantly, when rMC-1 transfected with Cx43 siRNA were grown as cocultures with pericytes, a significant decrease in GJIC, and increase in TUNEL-positive cells was observed, concomitant with decreased Akt phosphorylation. Upregulation of Cx43 rescued rMC-1 from HG-induced apoptosis.
  
  
  CONCLUSIONS
  Gap junction communication between Müller cells and pericytes is essential for their survival. Downregulation of Cx43 that is HG induced and impairment of GJIC activity in Müller cells contributes to loss of glial and vascular cells associated with the pathogenesis of diabetic retinopathy.
authors.affiliation: Department of Medicine, Boston University School of Medicine, Boston, Massachusetts, United States Department of Ophthalmology, Boston University School of Medicine, Boston, Massachusetts, United States.
authors.forename: Tetsuya
authors.initials: T
authors.lastname: Muto
authors.affiliation: Department of Medicine, Boston University School of Medicine, Boston, Massachusetts, United States Department of Ophthalmology, Boston University School of Medicine, Boston, Massachusetts, United States.
authors.forename: Thomas
authors.initials: T
authors.lastname: Tien
authors.affiliation: Department of Medicine, Boston University School of Medicine, Boston, Massachusetts, United States Department of Ophthalmology, Boston University School of Medicine, Boston, Massachusetts, United States.
authors.forename: Dongjoon
authors.initials: D
authors.lastname: Kim
authors.affiliation: Department of Ophthalmology, Northwestern University Feinberg School of Medicine, Chicago, Illinois, United States.
authors.forename: Vijay P
authors.initials: VP
authors.lastname: Sarthy
authors.affiliation: Department of Medicine, Boston University School of Medicine, Boston, Massachusetts, United States Department of Ophthalmology, Boston University School of Medicine, Boston, Massachusetts, United States.
authors.forename: Sayon
authors.initials: S
authors.lastname: Roy
mesh_terms: D000818:Animals; D017209:Apoptosis; D015153:Blotting, Western; D002450:Cell Communication; D018031:Connexin 43 / Q000096:biosynthesis / Q000235:genetics*; D004247:DNA / Q000235:genetics*; D003921:Diabetes Mellitus, Experimental / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D003930:Diabetic Retinopathy / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D004730:Endothelium, Vascular / Q000378:metabolism / Q000473:pathology; D063928:Ependymoglial Cells / Q000378:metabolism / Q000473:pathology*; D017629:Gap Junctions; D005786:Gene Expression Regulation; D020287:In Situ Nick-End Labeling; D020286:Pericytes / Q000378:metabolism / Q000473:pathology; D051381:Rats
chemical_list: D018031:Connexin 43; D004247:DNA
keywords: Müller cells; connexin 43; gap junctions; high glucose; pericytes