title: High glucose mediates endothelial-to-chondrocyte transition in human aortic endothelial cells.
abstract: BACKGROUND
  Vascular calcification is one of the common complications in diabetes mellitus. Many studies have shown that high glucose (HG) caused cardiovascular calcification, but its underlying mechanism is not fully understood. Recently, medial calcification has been most commonly described in the vessels of patients with diabetes. Chondrocytes were involved in the medial calcification. Recent studies have shown that the conversion into mesenchymal stem cells (MSCs) via the endothelial-to-mesenchymal transition (EndMT) could be triggered in chondrocytes. Our previous research has indicated that HG induced EndMT in human aortic endothelial cells (HAECs). Therefore, we addressed the question of whether HG-induced EndMT could be transitioned into MSCs and differentiated into chondrocytes.
  
  
  METHODS
  HAECs were divided into three groups: a normal glucose (NG) group, HG group (30 mmol/L), and mannitol (5.5 mmol/L NG + 24.5 mmol/L) group. Pathological changes were investigated using fluorescence microscopy and electron microscopy. Immunofluorescence staining was performed to detect the co-expression of endothelial markers, such as CD31, and fibroblast markers, such as fibroblast-specific protein 1 (FSP-1). The expression of FSP-1 was detected by real time-PCR and western blots. Endothelial-derived MSCs were grown in MSC medium for one week. The expression of the MSCs markers STRO-1, CD44, CD10 and the chondrocyte marker SOX9 was detected by immunofluorescence staining and western blots. Chondrocyte expression was detected by alcian blue staining. Calcium deposits were analyzed by alizarin red staining.
  
  
  RESULTS
  The incubation of HAECs exposed to HG resulted in a fibroblast-like phenotype. Double staining of the HAECs indicated a co-localization of CD31 and FSP-1. The expression of FSP-1 was significantly increased in the HG group, and the cells undergoing EndMT also expressed STRO-1, CD44 and SOX9 compared with the controls (P < 0.05). Additionally, alcian blue staining in the HG group was positive compared to the NG group. Consistent with the evaluation of SOX9 expression, calcium deposits analyzed by alizarin red staining were also enhanced by the HG treatment. Specifically, we showed that HG-induced EndMT is accompanied by the activation of the canonical Snail pathway.
  
  
  CONCLUSIONS
  Our study demonstrated that HG could induce endothelial cells transdifferentiation into chondrocyte-like cells via the EndMT, which is mediated in part by the activation of the Snail signaling pathway.
authors.affiliation: Institute of Nephrology, ZhongDa Hospital, School of Medicine, Southeast University, 210009 Nanjing, China.
authors.forename: Rining
authors.initials: R
authors.lastname: Tang
authors.forename: Min
authors.initials: M
authors.lastname: Gao
authors.forename: Min
authors.initials: M
authors.lastname: Wu
authors.forename: Hong
authors.initials: H
authors.lastname: Liu
authors.forename: Xiaoliang
authors.initials: X
authors.lastname: Zhang
authors.forename: Bicheng
authors.initials: B
authors.lastname: Liu
mesh_terms: D000954:Antigens, Surface / Q000378:metabolism; D001011:Aorta / Q000378:metabolism* / Q000648:ultrastructure; D015415:Biomarkers / Q000378:metabolism; D015153:Blotting, Western; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism; D002478:Cells, Cultured; D019902:Chondrocytes / Q000378:metabolism* / Q000648:ultrastructure; D042783:Endothelial Cells / Q000378:metabolism* / Q000648:ultrastructure; D058750:Epithelial-Mesenchymal Transition; D005947:Glucose / Q000378:metabolism*; D006801:Humans; D018960:Hyaluronan Receptors / Q000235:genetics / Q000378:metabolism; D059630:Mesenchymal Stem Cells / Q000378:metabolism* / Q000648:ultrastructure; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D015260:Neprilysin / Q000235:genetics / Q000378:metabolism; D010641:Phenotype; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000235:genetics / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D000071999:S100 Calcium-Binding Protein A4; D055758:SOX9 Transcription Factor / Q000235:genetics / Q000378:metabolism; D000071250:Snail Family Transcription Factors; D014157:Transcription Factors / Q000378:metabolism; D061205:Vascular Calcification / Q000235:genetics / Q000378:metabolism* / Q000473:pathology
chemical_list: D000954:Antigens, Surface; D015415:Biomarkers; C497483:CD44 protein, human; D002135:Calcium-Binding Proteins; D018960:Hyaluronan Receptors; D019408:Platelet Endothelial Cell Adhesion Molecule-1; D000071999:S100 Calcium-Binding Protein A4; D055758:SOX9 Transcription Factor; C527932:SOX9 protein, human; C525591:STRO-1 antigen, human; D000071250:Snail Family Transcription Factors; D014157:Transcription Factors; C077591:S100A4 protein, human; D015260:Neprilysin; D005947:Glucose