The cytokines Interleukin-1 (IL-1) and Tumor Necrosis Factor (TNF, also known as cachectin) exhibit multiple effects on circulating blood cells and cells of the blood vessel wall. For example, these mediators elicit a coordinated Drogram of functions of endothelial cells (EC) that promotes blood coagulation and thrombosis, and lead to clot stabilization. Furthermore, IL-1 and TNF promote adherence to vascular endothelium of leukocytes of many classes.Thus, these cytokines are likely to be involved in signaling the pathologic changes in blood vessels that characterize a number of inflammatory or infectious processes. These two cytokines were originally isolated frcm activated human mononuclear phagocytes, hence their comnon designation as monokines and the terminology "interleukin". However, recent findings have broadened this concept considerably. It is now clear that many cell types can produce IL-1-1ike activity.Several groups showed that human vascular EC can secrete material that stimulates proliferation of thymocytes incubated with suboptima1 doses of the mitogenic lectin phytohemagglutinin, a typical acitivty of IL-1 (thymocyte costimulation).Two related but distinct genes cloned frcm human peripheral blood monocytes encode IL-1 molecules. In human blood monocytes stimulated with bacterial lioopolysaccharide (LPS) IL-1 beta (pi ∼ 7) is the major form expressed while IL-1 alpha (pi ∼ 5) is the less abundant species secreted by human monocytes under these conditions. We found that EC and smooth muscle cells (SMC) isolated from adult human vessels can express these same IL-1 genes. LPS, a standard stimulus to IL-1 secretion in the monocyte, caused accumulation of IL-1 beta mRNA in both vascular cell types. Endothelial cells frcm adult human vessels also contained IL-1 alpha mRNA when treated with LPS in the presence of cycloheximide and LPS-stimulated smooth muscle cells contained RNA that hybridized with an IL-1 alpha cDNA probe as well. Although both vascular cell types can transcribe these IL-1 genes, the time course of this response differs. LPS induced IL-1 beta mRNA production by SMC maximally at 4-6 hr., whereas maximal IL-1 induction by LPS in EC occured 1 day after initiation of the exposure. Actinanycin D (1 ug/ml) blocked 3H-uridine incorporation into macromolecules by > 95% in both EC & SMC, and prevented the LPS-induced increases in IL-1 mRNA levels in these cells. This result suggests that this potentially injurious stimulus causes IL-1 mRNA accumulation by an increase in rates of transcription. These LPS-induced increases in IL-1 mRNA levels corresponded to production of biologically active IL-1 determined as thymocyte costimulation activity. Interestingly, gel filtration experiments revealed a molecular weight of around 22kD for both SMC and EC-derived IL-1 secreted into culture medium in response to LPS. This molecular weight contrasts with the 17 kD species which is the fully processed product secreted frcm activated human monocytes. A possible explanation for this disparity is that the vascular cells secrete a partially processed intermediate form of mature IL-1. Other stimuli for IL-1 mRNA accumulation and secretion of biological activity include TNF and IL-1 itself. Recombinant human INF (≥ 10 ng/ml) increased IL-1 beta mRNA levels in EC & SMC, and caused the EC & SMC to release IL-1-1 ike thymocyte costimulation activity. Of interest is the recent observation that IL-1 itself can stimulate expression of IL-rl genes in vascular wall cells. Both IL-1 aloha and beta can increase IL-1 beta mRNA content in EC & SMC. Hris observation was confirmed with homogenous IL-1 prepared by recombinant DNA technologies (rIL-1). These findings raise the possibility of a novel positive feedback loop in vascular pathophysiology. We also found that rIL-1 alpha or beta also induced the production of prostaglandin E2 (PGE2) by both vascular SMC & EC. This prostanoid, induced by IL-1, inhibits thymocyte _ proliferation. Thus, IL-1 not only induced its own expression but increased production of this immunosuppressive prostanoid. This mechanism provides a potential negative control loop in regulation of the local immune response in blood vessels. Vie conclude that these cells of the blood vessel wall are a source of the potent vasoregulatory and immune mediators IL-1 alpha and beta. Since IL-1 influences the thrombotic, hemostatic, and fibrinolytic functions of endothelium, as well as other responses to acute injury, our findings suggest novel local control mechanisms that may be important in a variety of pathologic states.
Downregulation of Endothelial Plexin A4 Under Inflammatory Conditions Impairs Vascular Integrity
