Endotoxin activates human vascular smooth muscle cells despite lack of expression of CD14 mRNA or endogenous membrane CD14

Abstract

During infection or inflammation, cells of the blood vessel wall, such as endothelial cells (EC) and smooth muscle cells (SMC), contribute to the regulation of the immune response by production of cytokines or expression of adhesion molecules. Little is known about the mechanism(s) involved in the stimulation of vascular cells by endotoxin (lipopolysaccharide [LPS]). As reported previously, LPS antagonists reduce LPS-induced cytokine production or adhesion in vitro specifically, suggesting a specific LPS recognition mechanism. We thus investigated the role of CD14 for stimulation of vascular SMC by LPS. Complement-fixing antibodies directed against CD14 (LeuM3, RoMo I, or Mo2) lysed monocytes but failed to mediate lysis of EC or SMC, indicating the lack of endogenous membrane CD14 in vascular cells. In addition, we did not detect expression of CD14 protein on EC and SMC in cell sorting analysis or cell immunoassay experiments. These observations are in line with our finding that a CD14 probe did not hybridize with mRNA of EC or SMC in Northern (RNA) blot experiments, although it hybridized well with monocyte-derived mRNA. We obtained the same results with the much more sensitive reverse transcription-PCR. Since the vascular SMC did not express endogenous CD14, we investigated the role of human serum-derived soluble CD14 (sCD14) for activation of SMC by LPS. In medium containing human serum, anti-CD14 antibodies inhibited activation of SMC by LPS. In contrast, the same antibodies did not inhibit activation of cells cultured in medium containing fetal calf serum. SMC cultured in sCD14-depleted medium responded 1,000-fold less to LPS than cells cultured in presence of sCD14. Reconstitution of sCD14-depleted serum or supplementation of serum-free medium with recombinant CD14 restored the capacity of the cells to respond to LPS. These results show that specific activation of vascular SMC by LPS does not involve binding to endogenous membrane CD14, but that the activation of vascular SMC by LPS is mediated to a great extent by serum-derived sCD14.

Heightened Levels of Antimicrobial Response Factors in Patients With Rheumatoid Arthritis

“…We observed a significant elevation of sCD14 levels in RA patients compared to healthy controls confirming previous reports (74)(75)(76). sCD14 acts as a co-receptor for endotoxin and facilitates the activation of those cells which are devoid of membrane bound CD14 via TLR4 transmembrane signaling (77)(78)(79)(80). Elevated levels of sCD14 could be caused by bacterial exposure, alterations in the microbiome, compromised gut integrity and increased levels of cytokines.…”

Evaluation of CD14 in host defence

“…The soluble form(s) of CD14 play an important role in the LPS-mediated activation of CD14-negative (endothelial, epithelial and smooth muscle) cells ( Fig. 1) [67,68]. There is a striking difference between sCD14 and mCD14 in the interaction with LBP and LPS.…”

cAMP induces CD14 expression in murine macrophages via increased transcription

“…Soluble CD14 lacking the GPI tail is shed from mononuclear phagocytes [18], or other non-myeloid cells such as hepatocytes [19], and circulates in plasma [20,21]. Soluble CD14 is required for LPS-induced activation in some membrane-bound CD14-negative cells such as endothelial [22,23], epithelial [24], and smooth muscle cells [25]. Binding of LPS to the cell surface via membrane-bound CD14 triggers protein tyrosine kinases [26,27], activation of mitogenactivated protein kinases [28][29][30], and NF-B translocation [31].…”