Toll-like receptor-4 message is up-regulated in lipopolysaccharide-exposed rat lung pericytes

Pericytes are multifunctional, polymorphic perivascular cells that lie within the microvessel basal lamina, are located on the abluminal side of endothelial cells, and are thought to play a regulatory role in capillary leak observed in sepsis. Toll-Like receptor 4 (TLR-4) has been implicated as the proximal transmembrane receptor for the LPS/CD 14 complex during the activation of lipopolysacharide (LPS)-induced sepsis. It is our hypothesis that TLR-4 is present on lung pericytes and is up-regulated in response to LPS.

Rat microvascular lung pericytes were isolated and cultured. Cells from passage 3-5 were used and treated with LPS (control, 10 ng/mL, and 100 ng/mL) for 18 h. Immunostaining and immunoblotting were performed to detect the presence of CD-14, TLR-2, and TLR-4. Real-time polymerase chain reaction was used to analyze the presence and quantity of mRNA for CD-14, TLR-2, and TLR-4.

Immunostaining and immunoblotting revealed the presence of CD-14, TLR-2, and TLR-4 in pericytes from each treatment group, and real-time polymerase chain reaction confirmed the presence of mRNA for CD-14, TLR-2, and TLR-4. An increase in the mRNA was observed in CD-14, TLR-2, and TLR-4 in the presence of increasing LPS 4 h after treatment. At 18 h after LPS treatment, a decrease in mRNA was noted.

The up-regulation of TLR-4 in the presence of increasing LPS suggests its importance in pericyte LPS-induced activation. Pericyte TLR-4 recognition of LPS could play a role in capillary leak seen in sepsis. These data also demonstrates that pericytes, once thought to be passive participants in the inflammatory cascade, may be active members.