Toll-Like Receptor-4 Message Is Up-Regulated in Lipopolysaccharide-Exposed Rat Lung Pericytes

Background: 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. Methods: 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. Results: 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. Conclusions: 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. [Copyright &y& Elsevier]