752

46th FORUM

IN IMMUNOLOGY

IL6 and thrombocytopoiesis T. Ishibashi and S. Asano First Department of Internal Medicine, Fukushima Medical College, Fukushima, Japan, and Department of Hematology/Oncology, The Institute of Medical Science, The University of Tokyo, Tokyo

Platelets, the main blood components involved in haemostasis, are produced by megakaryocytes. Megakaryocytic lineage has a unique differential process which emcompasses the proliferation of megakaryocytic progenitor cells (CFU-MK) and cellular maturation that is composed of nuclear polyploidization and cytoplasmic development without cell division. Advances in cellular and molecular biology have facilitated the investigation of the involvement of cytokines in this process. Several humoral factors appear to promote this lineage, such as megakaryocyte colony-stimulating factors in vitro, or as maturational factors including megakaryocyte potentiators in vitro, and molecules that stimulate megakaryocyte maturation and platelet production in vivo (Hoffman, 1989; Williams, 1991). At least in vitro, interleukin-3 (IL3), granulocyte-macrophage colony-stimulating factor (GM-CSF), erythropoietin (Ep) and stem cell factor (SCF) have all been shown to stimulate not only the proliferation of megakaryocytic progenitors, but also the differentiation of megakaryocytes. In vitro studies have also shown that IL6, IL1 1 and leukaemia inhibitory factor (LIF) are synergistic with IL3, leading to an increase in the number of CFU-MKderived colonies (Ishibashi et al., 1989a ; Paul et al., 1990; Metcalf et al., 1991). Although IL3, GMCSF and Ep fail to induce a consistent increase in platelet count, the administration of IL6, IL1 1, LIF and SCF to animals has resulted in a steady increase in platelet count (Ishibashi et al., 1989b; Asano et al., 1990; Bree et al., 1991; Metcalf et al., 1990; Andrews et al., 1991). Of those, IL6 is the first molecule to elicit reproducible increases in megakaryocytic parameters and platelet counts. This cytokine has been purified to homogeneity as a Bcell stimulatory factor 2 which exhibits a wide range of biological activities. In the area of haematopoiesis, this factor has been shown to act on primitive haematopoietic progenitor cells and megakaryocytic lineage (Ikebuchi et al., 1987, Ishibashi et al., 1989a). In this paper, effects of IL6 on thrombocytopoiesis are described.

In vitro studies Marrow colony assays indicate the megakaryocyte potentiating activity of IL6 in vitro. These data show that IL6 alone does not stimulate megakaryocyte colony formation, but that it is synergistic with IL3, leading to an increase in the number of colonies induced by IL3 (Ishibashi et al., 1989a; Warren et al., 1989; Williams et al., 1990; Koike et al., 1990). Other groups have reported the megakaryocyte colonystimulating activity of IL6 using marrow cells of mice and humans (Lotem et al., 1989 ; Bruno and Hoffman, 1989; Quesenberry et al., 1991). Considering megakaryocytic maturation, IL6 alone induced marked increases in the size of megakaryocytes, the activity of relatively specific lineage markers (acetylcholinesterase in mice and glycoprotein IIb/IIIa in humans) and ploidy in liquid culture systems (Ishibashi et al., 1989a; Kimura et al., 1990a). Another group of investigators showed that IL6 stimulates the growth of immature megakaryocytes in liquid culture (Williams et al., 1990). These findings suggest that, in vitro, IL6 acts primarily on the maturational process of this lineage. The next issue is whether the in vitro effects of IL6 on megakaryocytes are direct or indirect. Our study of single cell cultures indicates that IL6 acts directly on the immature and mature megakaryocytes derived from murine megakaryocyte colonies, suggesting the presence of IL6 receptors on these cells (Ishibashi et al., 1989a). Recent evidence indicates that IL6 receptors are expressed by human megakaryocytes (Navarro et al., 1991). We also demonstrated that a single class of IL6 receptors is present on rat megakaryocyte, as detected by binding assays of purified megakaryocyte population (Shikama et al., 1990). In vivo studies In vitro observations led to the hypothesis that IL6 used alone may exert a potent thrombopoietic

INTERLEUKIN

activity in vivo and increase the platelet count. Our group was the first to show that IL6 stimulates the development of the megakaryocytes and increases the platelet count in mice and primates (Ishibashi et al., 1989b ; Asano et al., 1990). The thrombopoietic activity of IL6 in normal animals, as determined by the size and ploidy of marrow megakaryocytes, platelet counts and the incorporation of “S-selenomethionine, has been confirmed by several laboratories (Hill et al., 1990; Nagasawa et al., 1990; Pojda and Tsuboi, 1990; Hill et al., 1991; Mayer ef al., 1991; Stahl ef al., 1991). Data show that IL6 enhances the size of megakaryocy-tes in bone marrow and increases the platelet count with no increase in the frequency of megakaryocytes in bone marrow. However, the in vivo action of IL6 on committed CFU-MK remains controversial, as observed in in vitro studies. In addition to the responses to IL6 observed in normal animals, the enhancement of platelet production by IL6 in thrombocytopenic conditions has been studied. Data indicate that platelet recovery is accelerated in response to IL6 in mice treated with cytotoxic agents as well as in mice exposed to sublethal doses of irradiation (Takatsuki et al., 1990; Patchen et al., 1991).

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Future clinical applications No thrombopoietic factor specific to megakaryocytopoiesis and platelet production has yet been purified. IL6 may be one of the candidates for use in treating patients with thrombocytopaenia. The thrombopoietic activity of IL6 was accompanied by an elevation of acute phase reactive proteins in the sera of animals treated with IL6, since this pleiotropic cytokine is also capable of stimulating hepatocytes and other target cells (Asano et al., 1990; Mayer et al., 1991). However, the acute phase proteins were normalized soon after the termination of the injection of IL6. Additionally, it is not known whether the increase in acute phase proteins via a direct effect of IL6 on the liver implies the induction of inflammation. In inflammatory conditions, endogenous IL6 is released follotiing the initial elevation of cytokines such as IL1 and TNF (Content et al., 1985; Kohase et al., 1986). Therefore, its administration to humans may perhaps be tolerated, as suggested by our study in primates. The phase I clinical trial is now ongoing.

References Physiological

role of IL6 in thrombocytopoiesis

Reactive or secondary thrombocytosis occurs in various conditions such as inflammation, following surgery or trauma, and malignant disease. Holler et al. (1991) reported an elevation of serum levels of IL6 in patients with reactive thrombocytosis. Their serial assays showed that the highest elevation in serum IL6 followed a rise in platelet level. Moreover, Kimura et al. (1990b) demonstrated an elevation of platelets in mice injected with ILIP. The latter findings imply that the thrombocytosis induced by ILlp is mediated by the release of IL6. These observations suggest that IL6 plays a key role in the development of reactive or secondary thrombocytosis. However, the data showed no correlation between serum IL6 levels and platelet counts in reactive thrombocytosis (Holler ef al., 1991). Moreover, no elevation of serum IL6 levels has been observed in patients with myeloproliferative disorders and idiopathic thrombocytopenic purpura. Thus, serum IL6 levels may not reflect circulating platelet counts in this condition, since the balance of platelet production and destruction should be considered. To determine the further characteristics of IL6 in thrombocytopoiesis, it would be important to investigate the correlation between the production of IL6 in marrow stromal cells and the rate of platelet production. The physiological role of IL6 in thrombocytopoiesis requires further study.

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T., Taga, T., Nakano,

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IL6 and thrombocytopoiesis.

752 46th FORUM IN IMMUNOLOGY IL6 and thrombocytopoiesis T. Ishibashi and S. Asano First Department of Internal Medicine, Fukushima Medical College,...
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