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Decreased Endothelial Cell Resistance Following Knock Down of Claudin 11
Author(s) -
Chitaley Kanchan,
Kim Bryan,
Thorner Daniel,
Engel Karen,
Tsubota Yoshi,
Wessells Hunter
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.1_supplement.964.15
Subject(s) - transfection , claudin , tight junction , western blot , endothelium , endothelial stem cell , barrier function , chemistry , in vitro , intracellular , microbiology and biotechnology , andrology , biology , medicine , gene , endocrinology , biochemistry
The corpus cavernosum is a specialized vascular organ in which endothelial cells (ECs) are exposed to a wide range of pressures (10–500 mmHg) during flaccidity and rigidity. The tight junction associated gene, claudin 11, is highly expressed in human cavernosal ECs (HCCECs) as compared to ECs from other blood vessels. We tested the hypothesis that claudin 11 plays an important role in barrier function in HCCECs by measuring transendothelial electrical resistance (TEER) following claudin 11 knock down. Ambion siRNA constructs for CLDN11 and a negative control scrambled sequence (SCR) were transfected into HCCECs using DharmaFECT‐1 (Dharmacon). Significant down‐regulation of claudin 11 was verified by Western blot at 2 and 7 days following transfection. Untransfected control (CNT), SCR siRNA, and CLDN11 siRNA transfected ECs were seeded in 6.5‐mm Transwell inserts and integrity of the endothelial monolayer assessed by daily measurements of TEER with an Endohm‐6 electrode chamber and an EVOM voltohmmeter (World Precision Instrument). HCCECs transfected with CLDN11 siRNA had significantly reduced TEER compared to both SCR and CNT groups (% of CNT at 75.5 and 95hr: 73±8.7 and 76±8.9, p<0.05). There was no significant difference in TEER in the SCR siRNA vs. CNT group (% of CNT at 75.5 and 95hr: 97.1±2.2 and 98.5±1.9). These data suggest that claudin 11 is important for cavernosal EC junctions and barrier function.