title: Norcantharidin inhibits lymphangiogenesis by downregulating the expression of VEGF-C and VEGF-D in human dermal lymphatic endothelial cells in vitro.
abstract: OBJECTIVE
  To investigate the effects of norcantharidin on the growth and migration of human dermal lymphatic endothelial cells (HDLECs) and further characterize its effect on lymphangiogenesis.
  
  
  METHODS
  A 3-dimensional fibrin gel lymphangiogenesis model was built. Flow cytometry was used to analyze the rate of apoptosis and necrosis. RT-PCR, immunohistochemistry and immunoblotting assays were used to examine the effect of norcantharidin on vascular endothelial growth factor C (VEGF-C), VEGF-D and VEGF receptor 3 during in vitro lymphangiogenesis.
  
  
  RESULTS
  Norcantharidin caused a marked dose and time-dependent inhibition of the growth of HDLECs with an IC50 of 40 nmol/l. The apoptotic rate of HDLECs was 13.21 ± 1.60% 24 h after treatment with 7.5 nmol/l norcantharidin and 42.34 ± 3.80% with 90 nmol/l norcantharidin (p < 0.01 vs. controls in both). Fibrin gel assays showed that norcantharidin (15 nmol/l) reduced the number of tubular structures from 68.4 ± 5.2 in untreated controls to 10.9 ± 2.3 (p = 0.000). RT-PCR, immunohistochemistry and immunoblotting assays showed norcantharidin markedly reduced the expression of VEGF-C and VEGF-D.
  
  
  CONCLUSIONS
  Norcantharidin inhibits lymphangiogenesis by downregulating the expression of VEGF-C and VEGF-D, suggesting that norcantharidin could be an effective agent for targeting neolymphangiogenesis.
authors.affiliation: Department of General Surgery, Maternity and Child Health Hospital, Jiaxing, PR China.
authors.forename: Xiaojian
authors.initials: X
authors.lastname: Yuan
authors.forename: Yi
authors.initials: Y
authors.lastname: Chen
authors.forename: Xianming
authors.initials: X
authors.lastname: Li
authors.forename: Gongnian
authors.initials: G
authors.lastname: Zhang
authors.forename: Donghui
authors.initials: D
authors.lastname: Jin
authors.forename: Hengfei
authors.initials: H
authors.lastname: Zhao
authors.forename: Zhiwei
authors.initials: Z
authors.lastname: Su
authors.forename: Yuezu
authors.initials: Y
authors.lastname: Fan
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000494:pharmacology*; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D015536:Down-Regulation; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology; D006801:Humans; D042583:Lymphangiogenesis / Q000187:drug effects*; D012867:Skin / Q000166:cytology; D042582:Vascular Endothelial Growth Factor C / Q000235:genetics / Q000378:metabolism; D042643:Vascular Endothelial Growth Factor D / Q000235:genetics / Q000378:metabolism; D040321:Vascular Endothelial Growth Factor Receptor-3 / Q000235:genetics / Q000378:metabolism
chemical_list: D019086:Bridged Bicyclo Compounds, Heterocyclic; C467486:VEGFC protein, human; C579459:VEGFD protein, human; D042582:Vascular Endothelial Growth Factor C; D042643:Vascular Endothelial Growth Factor D; C069741:norcantharidin; C581309:FLT4 protein, human; D040321:Vascular Endothelial Growth Factor Receptor-3