Lipopolysaccharide activation of pericyte's Toll-like receptor-4 regulates co-culture permeability.
BACKGROUND
Pericytes (PCs) have a synergistic relationship with endothelial cells (MVEC) in regulating capillary permeability. PCs express Toll-like receptor-4 (TLR-4). We hypothesize one mechanism of MVEC/PC co-culture permeability is regulated through lipopolysaccharide (LPS) activation of pericyte TLR-4.


METHODS
Rat PCs were harvested and cultured. PCs were transfected with siRNA targeted to TLR-4. Western blotting was used to confirm gene silencing of TLR-4. A previously described co-culture permeability assay was performed after LPS treatment.


RESULTS
Western blot confirmed successful silencing of TLR-4 in PCs, which was sustained for 7 days. A dose- and time-dependent effect of LPS on albumin clearance was seen in MVEC/PC co-cultures. Co-cultures with TLR-4 silenced in PCs eliminated the LPS dose-dependent increase in albumin clearance.


CONCLUSIONS
TLR-4 regulates pericyte mediated capillary leak seen with LPS exposure. Our TLR-4 silencing model can be used to further investigate TLR-4's role in pericyte mediated capillary leak.
Edelman
David A
DA
Department of Surgery, Wayne State University, 3900 John R, Suite 400 POB, Detroit, MI 48201, USA.
Jiang
Yang
Y
Tyburski
James G
JG
Wilson
Robert F
RF
Steffes
Christopher P
CP
D000418:Albumins / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D002463:Cell Membrane Permeability / Q000187:drug effects / Q000235:genetics; D002478:Cells, Cultured; D018920:Coculture Techniques; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D020868:Gene Silencing / Q000187:drug effects*; D008168:Lung / Q000098:blood supply / Q000166:cytology; D020286:Pericytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D011134:Polysaccharides / Q000494:pharmacology*; D034741:RNA, Small Interfering / Q000235:genetics*; D051381:Rats; D017207:Rats, Sprague-Dawley; D051197:Toll-Like Receptor 4 / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism
D000418:Albumins; D011134:Polysaccharides; D034741:RNA, Small Interfering; C495347:Tlr4 protein, rat; D051197:Toll-Like Receptor 4