Soluble interleukin-6 receptor enhanced by oncostatin M induces major changes in gene expression profile of human hepatoma cells

Interleukin-6 (IL-6) binds to a receptor complex consisting of an 80 kDa binding unit (IL-6R) and gp130 responsible for signal transduction. Due to alternative splicing and/or proteolytic digestion IL-6R occurs in soluble form (sIL-6R), as well. Soluble IL-6R is able to bind to gp130 expressing on nucleated cells, thus sIL-6R makes most cells responsive to IL-6. In this study we found that oncostatin M (OSM), an other gp130 dependent cytokine with proliferation inhibitory potential, increases the expression of both membrane-bound IL-6R and sIL-6R generated by alternative splicing in hepatic and mammary carcinoma cell lines. Furthermore, we studied the functional relevance of the presence and binding of soluble IL-6R to HepG2 cells. Using a cDNA expression array, mRNA levels of about 580 human genes were tested by differential display analysis. Our findings suggest, that elevation of surface density of IL-6R by attachment of sIL-6R induces major modulation in gene expression profile of the hepatoma cells. Soluble IL-6R alone has minor effect, it rather decreases expression of some genes, while incubation with IL-6 and sIL-6R together induces major changes in the mRNA pattern of HepG2 cells. These data strongly suggest that presence and binding of soluble cytokine receptors are important elements of inter-cytokine cross talk and affects actual gene expression profile of responding cells.     

High circulating levels of biologically inactive IL-6/SIL-6 receptor complexes in systemic juvenile idiopathic arthritis: evidence for serum factors interfering with the binding to gp130

We previously demonstrated that high levels of IL-6/sIL-6R complexes are present in sera of patients with systemic juvenile idiopathic arthritis (s-JIA) and that the amount of IL-6 estimated in the IL-6/sIL-6R complexes is markedly higher than that measured by the B9 assay. Here, we show that two additional bioassays, employing human myeloma XG-1 cells and human hepatoma Hep3B cells, detected serum IL-6 levels similar to those measured by the B9 assay and approximately 10-fold lower than the IL-6 levels estimated to be present in the IL-6/sIL-6R complex. Using an assay for the measurement of the amount of circulating IL-6 complexed with the sIL-6R and available for binding to gp130 (gp130 binding activity), we show that the IL-6/gp130 binding activity is similar to that detected by the bioassays and again significantly lower than that estimated to be present in the IL-6/sIL-6R complex. Addition of recombinant human IL-6 (rhIL-6) to sera of patients or controls results in a markedly lower increase in the gp130 binding activity in patients than in controls. Moreover, sera from s-JIA patients inhibited in a dose dependent manner the gp130 binding activity assay. These results show that sera from patients with s-JIA contain a factor, or factors, that inhibit(s) the binding of the IL-6/sIL-6R complex to gp130. This inhibitory activity does not appear to be due to soluble gp130, C-reactive protein or autoantibodies to IL-6.     

IL-6, its receptors and its relationship with bcl-2 and bax proteins in infiltrating and in situ human breast carcinoma

AIMS: To characterize the expression pattern of IL-6 and its receptors (IL-6R(alpha) and gp130), to relate this pattern to bcl-2 and bax expression and to elucidate the effects on the proliferation/apoptosis equilibrium in benign conditions and in situ and infiltrating breast cancer. METHODS AND RESULTS: The immunoexpression of IL-6 and its receptors (IL-6R(alpha) and gp130), and their relationship with bcl-2 and bax proteins, were studied in in situ and infiltrating tumours and in benign breast lesions by means of Western blotting and immunohistochemistry. The percentages of samples positive for IL-6, bcl-2 and bax and their immunoreaction densities were higher in in situ carcinomas and infiltrating tumours than in benign lesions; although in in situ lesions were not so high as in infiltrating tumours, except for bax, whose immunoexpression was as weak as in benign conditions, resulting in a bcl-2/bax ratio higher than in infiltrating tumours. CONCLUSIONS: The high expression of IL-6 and its receptors in tumours might be related to the enhanced cell proliferation occurring in breast cancer. IL-6 could act by increasing bcl-2 expression and thus altering the proliferation/apoptosis balance toward neoplastic cell proliferation. The increased bax immunoreactivity observed only in infiltrating tumours, which was not so high as the increase in bcl-2 immunoreactivity, might be interpreted as an attempt to hinder cell proliferation.     

Soluble interleukin-6 receptor strongly increases the production of acute-phase protein by hepatoma cells but exerts minimal changes on human primary hepatocytes

Interleukin-6 (IL-6) interacts with a system of receptors, which include a 80-kDa IL-6-binding subunit (IL-6R) and a transducing element (gp130). The soluble form of IL-6R (sIL-6R) can bind its ligand and induce cellular responses by association with gp130, thus acting as an IL-6 agonist. We and others have previously shown that the responsiveness to IL-6 is different in hepatoma and human primary hepatocytes. We therefore compared the effects of sIL-6R on the two types of cells, and on the B9 hybridoma, another IL-6-sensitive cell line. Human primary hepatocytes, hepatoma cells PLC/PRF/5, and B9 cells were incubated with different concentrations of IL-6, sIL-6-R, or both. The hepatocyte culture supernatants were tested for their content of acute-phase proteins (APP). The proliferation of B9 cells was assessed by a colorimetric method. Results showed that sIL-6R alone markedly increases the production of APP by hepatoma cells in a dose-dependent manner, but affects only minimally primary hepatocytes and the proliferation of B9 cells. The combinations of IL-6R and its ligand enhanced the effects of IL-6 alone in both PLC/PRF/5 and B9 cells, but had no effect on primary hepatocytes. An immunohistochemical study indicated that the cell-surface expression of IL-6R was dramatically lower in hepatoma cells than in primary hepatocytes. In conclusion, our results show that the expression of IL-6R is low in the hepatoma cell PLC/PRF/5 when compared with primary hepatocytes and that this difference can, at least partly, explain their deficient responsiveness to IL-6. On the other hand, it appears that IL-6R expression by primary hepatocytes is sufficient and that circulating sIL-6R is unlikely to play a significant role in the modulation of IL-6 effects.     

A combination of interleukin-6 and its soluble receptor impairs sperm motility: implications in infertility associated with endometriosis

BACKGROUND: We previously reported that the level of interleukin (IL)-6 is increased in the peritoneal fluid of women with endometriosis. This study was undertaken to assess the effects of IL-6 and soluble IL-6 receptor (sIL-6R) on in vitro sperm motility. METHODS: Sperm (n = 20) were cultured with IL-6 or sIL-6R, or with a combination of both. After 24 h cultures, sperm motility was evaluated using a computer-assisted semen analysis system. Gene and protein expressions of IL-6, IL-6 receptor (IL-6R), and glycoprotein 130 (gp130) were examined in sperm by RT-PCR analysis and western blot analysis. RESULTS: Addition of IL-6 or sIL-6R individually to the culture media had no affect on sperm motion. However, adding a combination of IL-6 and sIL-6R dose-dependently reduced the percentage of motile and rapidly moving sperm. Adding anti-IL-6R antibody abolished these adverse effects. Sperm expressed the gp130 gene and protein, but not IL-6 or IL-6R. CONCLUSIONS: A combination of IL-6 and sIL-6R may be associated with gp130 expressed in the sperm and reduce sperm motility. IL-6 and sIL-6R may contribute to the pathogenesis of endometriosis-associated infertility.     

Retinoic acid modulates the in vivo and in vitro growth of IL-6 autocrine human myeloma cell lines via induction of apoptosis

We previously showed that IL-6 is an autocrine growth factor for two human myeloma cell lines, RPMI 8226 and U266. We investigated here the in vitro and in vivo effects of all-trans retinoic acid (RA) on the growth and survival of these two cell lines. RA induced a dramatic dose- and time-dependent inhibition of the proliferation of both cell lines. This inhibition was correlated with a down-modulation of the cell surface expression of the IL-6 binding chain (gp80) and the transducing chain (gp130) of the IL-6 receptor (IL-6R). Long-term culture experiments showed that down-modulation of gp80 expression was complete at days 15 and 30 in the presence of 10^–5 and 10^–7 mol/l of RA, respectively. Gp130 expression was greatly decreased, albeit still detectable, in similar culture conditions. RA-mediated interruption of the IL-6 autocrine loop was associated with a decrease of bcl-2 oncoprotein expression and apoptosis of the myeloma cells which was RA concentration- and time-dependent. The in vivo relevance of the effects of RA was studied on tumours which developed in nude mice inoculated with a subclone of RPMI 8226. Whereas tumours grew in all control mice, 40% of tumours regressed within 20 days in RA-treated mice. Cells from regressing tumours featured characteristics of apoptosis and exhibited low gp80 and gp 130 expression. Our study indicate that long-term RA treatment interferes in vivo and in vitro with IL-6 autocrine growth of myeloma cell lines, leading to apoptosis.     

Soluble interleukin-6 receptor (sIL-6R) in cerebrospinal fluid of patients with inflammatory and non inflammatory neurological diseases

IL-6 acts on target cells via the ligand-binding protein interleukin-6 receptor (IL-6R) and the affinity-converting and signal-transducing glycoprotein 130 (gp130). Soluble interleukin-6 receptor (sIL-6R) has an agonistic role because the soluble complex (IL-6/sIL-6R) can activate cells that do not express IL-6R and an antagonistic role as it enhances the inhibitory activity of sgp130. Soluble forms of both receptors, sIL-6R and sgp130, regulate the action of IL-6. sIL-6R was measured by a sensitive enzyme-linked immunosorbent assay in paired sera and cerebrospinal fluid (CSF) from 46 patients with inflammatory neurological diseases (IND), 45 patients with relapsing-remitting multiple sclerosis (RR-MS), 13 patients with primary progressive multiple sclerosis (PP-MS), 17 patients with other non inflammatory neurological diseases (NIND) and 13 mentally healthy individuals--healthy controls (HC). Patients with RR-MS had CSF sIL-6R levels comparable to those from patients with IND, but higher than patients with NIND and HC. A positive correlation between the CSF/serum albumin (QAlb) and CSF sIL-6R levels was observed in IND but not in RR-MS patients indicating that CSF sIL-6R levels in IND patients could be influenced by serum sIL-6R and blood brain barrier (BBB) permeability properties. RR-MS patients had higher values of [CSF/serum sIL-6R:CSF/serum albumin] (sIL-6R index) than IND patients suggesting that in multiple sclerosis (MS), the increase in CSF sIL-6R could be due to intrathecal synthesis of sIL-6R. The finding of increased CSF sIL-6R concentrations (>979 pg/ml) with sIL-6R index (>4.66), in correlation with positive oligoclonal bands in RR-MS patients, suggests that values of sIL-6R index > 4.66 indicate intrathecal increase of sIL-6R and might be used as an indicator of neuroimmunoregulatory and inflammatory processes in the central nervous system (CNS).     

Association of recombinant soluble IL-6-signal transducer, gp130, with a complex of IL 6 and soluble IL-6 receptor, and establishment of an ELISA for soluble gp130.

IL-6 mediates its pleiotropic functions through two membrane proteins, a ligand-binding molecule (IL-6 receptor, IL-6R) and a non-ligand-binding signal transducer (gp130). Starting with a previously isolated cDNA clone encoding human gp130, recombinant soluble gp130 (sgp130) lacking the transmembrane and cytoplasmic regions was expressed in COS7 cells or CHO cells. sgp130 could associate with a complex of IL-6 and soluble IL-6R (sIL-6R), also lacking transmembrane and cytoplasmic regions. This indicated that extracellular region of gp130 was responsible for the association with IL-6R which was occupied by IL-6. An enzyme-linked immunosorbent assay (ELISA) for the quantitation of sgp130 was established, which was based on the interaction of sgp130 with the complex of IL-6 and sIL-6R and could detect sgp130 as low as 1 ng/ml.     

The importance of IL-6 protein expression in primary human renal cell carcinoma: an immunohistochemical study

AIMS: Interleukin 6 (IL-6) is produced by some renal carcinoma cell lines in vitro. This might be biologically important because IL-6 is a cytokine of particular interest, owing to its involvement in the growth of renal cell carcinoma. In this study, the expression of IL-6 protein in tissue samples from primary renal cell carcinoma was analysed, and then its clinical importance was examined. METHODS: The distribution of IL-6 in renal cell carcinoma was examined by means of an immunohistochemical method in 47 untreated primary renal cell carcinoma samples. The search for a significant difference between histological patterns, Furhman's grading system, TNM classification, and IL-6 protein expression was carried out. RESULTS: Immunohistochemistry demonstrated that IL-6 is expressed in 70% of primary tumours. There was no significant difference in the tumour size and grade between renal cell carcinomas with or without IL-6 expression. However, a relatively large number of high grade tumours expressed IL-6. CONCLUSION: The importance of IL-6 expression with regard to tumour size/local growth is questionable because IL-6 has been correlated with the development of metastatic disease. These data suggest that the production of IL-6 could exert a growth inhibitory effect on primary renal cell carcinoma.     

IL-6 and soluble IL-6 receptors (sIL-6R and sgp130) in human pleural effusions: massive IL-6 production independently of underlying diseases

IL-6, soluble IL-6 receptor (sIL-6R) and soluble gp130 (sgp130) levels were measured in sera and pleural effusions from 42 patients with metastatic carcinoma, non-Hodgkin's lymphoma, tuberculosis, cardiac failure and miscellaneous diseases. Pleural IL-6 levels measured by ELISA were very high in all patient groups (mean 34.8 ± 15.3 ng/ml) without significant difference according to diseases. IL-6 was shown to be biologically active in a proliferative assay. Serum IL-6 levels were low (0.049 ± 0.014 ng/ml) and did not correlate with pleural fluid levels. Pleural IL-6 levels correlated with the number of polymorphonuclear cells in pleural fluid (P< 0.03). Pleural sIL-6R levels (76 ± 8 ng/ml) were always lower than serum levels (196 ± 12 ng/ml; P< 0.0001) but correlated with them (P< 0.01). Pleural sIL-6R and albumin levels correlated (P< 0.01), suggesting a transudation of sIL-6R from the serum. Pleural sgp130 levels (10.9 ± 1.0 ng/ml) were lower than serum levels (24.6 ± 2.8 ng/ml; P< 0.002). After gel filtration of pleural fluid, the bulk of IL-6 (>90%) was recovered in a 15 000–30 000 fraction, corresponding to the expected mol. wt of free IL-6. These results suggest a production and a sequestration of IL-6 in the pleural cavity in all studied conditions.     

Development of a monoclonal antibody-based enzyme-linked immunoabsorbent assay for the binding of gp130 to the IL-6/IL-6R complex and its competitive inhibition

The proinflammatory cytokine IL-6 binds to the membrane bound or soluble IL-6 receptor (IL-6R) and activates an intracellular signaling cascade after complex formation with two gp130 molecules. These mediate general homeostasis and orchestrates the immune response during disease. Trans-signalling via the soluble IL-6R has importance for the development and maintenance of human diseases like Crohn's disease, peritonitis and rheumatoid arthritis. We have developed an enzyme-linked immunoabsorbent assay (ELISA) that detects the binding of gp130 to the IL-6/sIL-6R complex. Competitive binding of sgp130-Fc, viral IL-6, and the inhibitory drug Suramin to gp130 has been demonstrated. The assay can be used for high-throughput screening of peptide and chemical compound libraries for the identification of new agonists and antagonists of the binding between gp130 and IL-6/sIL-6R.     

Major role of the soluble interleukin-6/interleukin-6 receptor complex for the proliferation of interleukin-6-dependent human myeloma cell lines

Interleukin-6 (IL-6) is a proinflammatory cytokine which possesses a central growth factor activity for certain tumor cells such as plasma cells in multiple myeloma (MM). Upon binding of IL-6, soluble IL-6 receptor (sIL-6R) has been shown to retain its affinity for IL-6 and to associate with the signal-transducing gp130 chain. Therefore, contrary to the majority of soluble cytokine receptors, it plays an agonist role in IL-6 signaling. In order to test its physiological importance as compared to that of its membrane counterpart, we studied cells from two myeloma cell lines which need exogenous IL-6 to proliferate and release sIL-6R into their culture supernatant. Using a new culture system where the supernatant recirculated permanently through an anti-IL-6R affinity column, all sIL-6R was removed from the culture medium throughout the culture period. Under these conditions IL-6-dependent cells were unable to grow in the presence of physiological concentrations of IL-6, showing the major role of the sIL-6R for sustaining the proliferation of these cell lines. Increasing IL-6 concentrations well over the physiological values allowed the cells to proliferate again. No effect was seen when sIL-6R was removed from the supernatant of an IL-6-independent myeloma cell line. These results show that the levels of circulating sIL-6R (and thus those of IL-6/sIL-6R complex) are worth looking at in pathologies involving IL-6 hyperactivity.     

抽动秽语综合征患者血清自身抗体与可溶性白介素-6受体表达增高

目的探讨抽动秽语综合征(TS)病程中自身免疫损伤相关机制。方法用ELISA法检测TS患者血清中抗脑抗体(ABAb)、抗核抗体(ANAb)、可溶性白介素-6受体(sIL-6R)及可溶性gp130(sgp130)的含量,用胶乳增强免疫比浊法检测抗链球菌溶血素O抗体(ASO)水平。结果TS组血清sIL-6R和sgp130含量显著高于对照组[(44.1±15.8)ng/mLvs(30.3±9.0)ng/mL和(69.0±24.6)ng/mLvs(47.3±14.1)ng/mL,P<0.01];血清ABAb和ANAb检出率显著高于对照组(66%vs4%和53%vs25%,P<0.01),并且ASO增高的比例高于对照组(P<0.01);TS组ABAb水平与sgp130呈负相关(r=0.375,P<0.05)。结论TS患者依赖IL-6信号传导的生理功能上调和反馈抑制的网络机制启动;自身免疫相关的损伤机制均可能参与了疾病发生发展的病理生理过程。     

Soluble IL-6 receptor induces calcium flux and selectively modulates chemokine expression in human dermal fibroblasts.

Truncated forms of cytokine receptors have been regarded as modulators of the activity of their cognate ligands. In addition to inhibiting effects of their respective ligands, soluble receptors can also facilitate ligand-mediated signaling. Several studies have demonstrated that exogenous IL-6 in association with the soluble IL-6 receptor alpha (sIL-6Ralpha) can activate cells expressing the gp130 signal transducer lacking the specific, membrane-bound IL-6Ralpha. Since cell cultures of human dermal fibroblasts express high amounts of IL-6, we examined whether the addition of sIL-6Ralpha in association with endogenous IL-6 would be sufficient to stimulate these cells via gp130. As an early rapid signal we analyzed changes in intracellular free calcium concentrations ([Ca2+]i). Addition of sIL-6Ralpha induced an acute and transient increase in cytosolic free calcium concentrations in a dose-dependent fashion. This Ca2+-signal was abolished when cells were pretreated with anti-IL-6 or anti-gp130 antibodies. Using flow cytometric analysis we could demonstrate membrane-associated IL-6 and gp130, but not IL-6Ralpha on fibroblasts. We also analyzed MCP-1 and IL-8 expression as a response involved in the more recently recognized chemoattractant functions of fibroblasts, and found MCP-1 to be up-regulated, but not IL-8. These data suggest that sIL-6Ralpha binds to cell-associated, endogenous IL-6 produced by fibroblasts and this complex then activates the cells via gp130. This pathway of fibroblast activation by sIL-6Ralpha adds another dimension to the role of fibroblasts in the cytokine network.     

Effects of IL-6 and Soluble IL-6 Receptor on the Expression of Cartilage Matrix Proteins in Human Chondrocytes

Elevated concentrations of interleukin (IL)-6 and soluble IL-6 receptor (sIL-6Rα) in synovial fluid have been implicated in joint cartilage destruction. We examined the effect of IL-6 and sIL-6Rα on cell growth, alkaline phosphatase (ALPase) activity, and the expression of Sox-9, type II collagen, aggrecan core, link protein, BMP-7, and BMP receptors in human chondrocytes. Cell proliferation increased slightly in the presence of both IL-6 and sIL-6Rα, whereas ALPase activity decreased markedly. The expression of Sox-9 and aggrecan core did not change in the presence or absence of IL-6 and sIL-6Rα, whereas the expression of type II collagen, link protein, BMP-7, and BMP receptors increased in the presence of both IL-6 and sIL-6Rα. These results suggest that IL-6 and sIL-6Rα suppress the differentiation of chondrocytes and induce the repair of arthrodial cartilage through an increase in the expression of cartilage matrix proteins, BMP-7, and BMP receptors in the cells.     

Therapeutic targeting of interleukin-6 trans-signaling does not affect the outcome of experimental tuberculosis

Treatment of autoreactive inflammatory diseases such as rheumatoid arthritis with anti-inflammatory drugs is associated with an increased rate of reactivation tuberculosis (TB). Interleukin-6 (IL-6) plays a pivotal role in inflammation and protection against various infectious diseases. IL-6 signals by two mechanisms via the ubiquitous transmembrane protein gp130: 'classic' signaling using the membrane-bound IL-6 receptor (IL-6R), which is expressed mainly on hepatocytes and some leukocytes, and trans-signaling using soluble IL-6R (sIL-6R). Trans-signaling by the IL-6/sIL-6R complex is selectively inhibited by natural soluble gp130 (sgp130) and by sgp130 designer proteins. As specific blockade of IL-6 trans-signaling represents a promising approach for the therapy of inflammatory diseases, we evaluated the potential risk of interfering with this alternative pathway and analyzed the outcome of experimental TB after treatment with an IgG1-Fc fusion protein of soluble gp130 (sgp130Fc) and in sgp130Fc-overexpressing transgenic (sgp130Fc(tg)) mice. In contrast to treatment with anti-tumor necrosis factor (TNF) antibodies, administration of sgp130Fc did not interfere with protective immune responses after infection with Mycobacterium tuberculosis (Mtb). Moreover, Mtb-infected sgp130Fc(tg) mice were capable of controlling mycobacterial growth. Our finding that IL-6 trans-signaling plays no role for protective immune responses against Mtb supports the superior safety of therapeutic targeting of IL-6 trans-signaling compared to anti-TNF treatment.     

Interleukin-6 and its receptor: from bench to bedside

Interleukin-6 (IL-6) is an inflammatory cytokine with a well-documented role in inflammation and cancer. The cytokine binds to a membrane bound IL-6 receptor (IL-6R) and this complex associates with two molecules of the signal transducing protein gp130 thereby initiating intracellular signaling. While gp130 is present on most if not all cells of the body, the IL-6R is only present on some cells, mainly hepatocytes and several leukocytes. Cells, which only express gp130 and no IL-6R are refractory to IL-6 signals. We have shown earlier that the IL-6R can exist as a soluble protein generated by limited proteolysis of the membrane bound receptor or by translation from an alternatively spliced mRNA. This soluble IL-6R (sIL-6R) can bind the ligand IL-6 and the soluble complex of sIL-6R and IL-6 can bind to gp130 on cells which lack the membrane bound IL-6R and trigger gp130 signaling. We have named this process ‘trans-signaling’. We will review data, which clearly show that IL-6 uses classical signaling via the membrane bound receptor and trans-signaling via the soluble receptor in various physiological and pathophysiological situations. Furthermore, we have developed designer cytokines, which can specifically enhance or inhibit IL-6 trans-signaling. These designer cytokines have been shown to be extremely useful to in therapeutic applications ranging from the long-term culture of stem cells and enhancing liver regeneration up to the blockade of chronic inflammation and cancer.There are inclusions in this text from a number of different articles, which are cited in the reference section.     

Preeclampsia Status Controls Interleukin-6 and Soluble IL-6 Receptor Release from Neutrophils and Endothelial Cells: Relevance to Increased Inflammatory Responses

Increased neutrophil–endothelial binding and inflammatory responses are significant pathophysiological events in the maternal vascular system in preeclampsia, a hypertensive disorder in human pregnancy. Interleukin 6 (IL-6) and its soluble receptors (soluble IL-6R (sIL-6R) and soluble gp130 (sgp130)) are critical inflammatory mediators. During pregnancy, maternal IL-6 and sgp130 levels were increased, but sIL-6R levels were decreased, in women with preeclampsia compared to normotensive pregnant women. However, little is known about differences in IL-6, sIL-6R, and sgp130 production by neutrophils and endothelial cells between normal pregnancy and preeclampsia. To study this, we isolated neutrophils and cultured human umbilical vein endothelial cells (HUVECs) from normal and preeclamptic pregnancies. Production of IL-6, sIL-6R, and sgp130 was measured. The role of placental factor(s)-mediated neutrophil production of IL-6, sIL-6R, and sgp130 was also determined by pretreating neutrophils with placental conditioned medium generated from placental villous cultures. We found that IL-6 and sgp130 were mainly produced by endothelial cells, while sIL-6R was mainly produced by neutrophils. Endothelial cells from preeclampsia produced significantly more IL-6 and sgp130, and neutrophils from preeclampsia produced significantly less sIL-6R than normal pregnancy cells. Interestingly, production of IL-6, sIL-6R, and sgp130 were time-dependently increased when neutrophils and endothelial cells were co-cultured. We also found that neutrophils from normal pregnancies produced more IL-6, but less sIL-6R, after being primed by preeclamptic-placental conditioned medium. These results demonstrated that neutrophils and endothelial cells have different capacities in producing IL-6, sIL-6R, and sgp130 between normal pregnancy and preeclampsia. These results also provide evidence that the placenta plays a role in inducing neutrophil activation in preeclampsia.