sIL-2R Alpha Research Articles

Serum Soluble Interleukin 2 Receptor Alpha vs. Urinary Parameter and Serum Immunological Markers as a Monitoring Tool of Treatment Response of Lupus Nephritis and their Relation to Lupus Nephritis Class on Renal Biopsy

Background: Lupus Nephritis (LN) is one of the most common and serious manifestations in SLE patients that causes significant morbidity and mortality. Current conventional biomarkers for LN are sometimes unable to predict treatment response of lupus nephritis. Recently serum soluble interleukin-2 receptor alpha (sIL-2 R alpha) is shown to be a good marker to predict treatment response of LN. Objective: To compare serum sIL-2R alpha with other commonly used markers as a marker of treatment response of LN and their relation to the LN class in renal biopsy. Methods: This prospective observational study was conducted in the Department of Nephrology, Bangabandhu Sheikh Mujib Medical University (BSMMU), Dhaka, Bangladesh from July 2018 to August 2019. Twenty seven patients who were diagnosed with lupus nephritis after kidney biopsy were included in this study. Serum soluble interleukin-2 receptor alpha, 24 hrs UTP, anti-dsDNA, complements level (C3 & C4) were measured in all patient at baseline, 3-months and 6- months after treatment. Serum soluble interleukin-2 receptor alpha level was measured by ELISA and is expressed as pg/mg. Serum sIL-2R alpha as well as conventional biomarker value were compared before and after treatment and in between treatment response and non-response group. Results: Serum sIL-2R alpha levels were significantly higher in patients of proliferative lupus nephritis (Class III & Class IV) than non-proliferative lupus nephritis (Class V) at baseline (3934.3 ± 1095.2, 3934.3 ± 1095.2 & 1801.98 ± 205.8 pg/L respectively) and levels were decreased significantly 6 months after treatment (p <0.001). Serum sIL-2R alpha levels were also significantly higher in non-remission proliferative group then remission proliferative group (4071.60 ± 769.91 vs 5169.20 ± 394.76) with p value 0.008 at baseline. In contrast no significant difference were observed for 24 hrs UTP levels at baseline between this group which suggest that serum sIL-2R alpha are more sensitive marker than 24 hrs UTP in predicting treatment response of lupus nephritis. Conclusions: Serum sIL-2R alpha might be a valuable serological biomarker to diagnose and to predict and monitor the treatment response of lupus nephritis.

Possible Role of Impaired Erk1,2 Phosphorilation and Increased sIL2r Alpha Plasma Levels in the Reduced Frequency of Circulating T Regulatory Cells of Patients with Primary Myelofibrosis

Background. Primary myelofibrosis (PMF) is a clonal, neoplastic disorder of the hematopoietic stem cells, characterized by an increased number of circulating CD34+ cells. Available evidence indicates that enhanced activation of JAK-STAT pathway is the main molecular mechanism of myeloproliferation due to somatic mutations in JAK2, CALR or MPL genes. A subset of patients display immune-related abnormalities that suggest an (auto)immune pathogenesis. In addition, it has been reported a significant association between personal history of a broad span of autoimmune diseases and subsequent risk of myeloproliferative neoplasm (Kristinsson SY Haematologica 2010; 95:1216-1220).CD4+ T cells maintain cancer surveillance and immune tolerance. Chronicinflammation has been proposed as a driver of clonal evolution in myeloproliferative neoplasms (MPN) (Hasselbalch HC et al. Leuk Res 2013;37:214-20), suggesting that T cells play an important role in their pathogenesis. A recent paper by Keohane C et al (BJH 2015; doi 10.1111/bjh 13519) shows that peripheral blood (PB) T regulatory cells (Tregs) are reduced in MPN patients compared to healthy subjects (CTRLs) and that this decrease is even more pronounced following JAKi therapy (Massa M et al Leukemia 2014;28:449-51).Unpublished data from our group further indicate a significant (p<0.001) decrease of PB Tregs in PMF (n=193, CD4+CD25highCD127low/- FOXP3+ cells expressed as % of CD4+ cells, median 0.87, 0-10.2) versus CTRLs (n=16, median 2.09, 0.54-6.5). Based on the distribution of Tregs in CTRLs, we established a cut-off point of 1.0% to define an actual reduction of Tregs.Study design. To investigate the possible factors favouring a decrease of Tregs, we examined in patients with PMF and <1% Tregs (n=15), and in CTRLs (n=10): 1) the percentage of PB CD4+, CD4+CD25+, and CD4+CD25++CD127low/- cells expressing membrane Annexin V at time of sampling as a measure of spontaneous apoptosis 2) the phosphorylation of Stat1, Stat3, Stat5, Stat6, Erk1,2 and p38-MAPK determined by using flow-based intracellular staining (BD Biosciences) at baseline and following stimulation with the appropriate cytokine in CD4+, CD4+CD25-, CD4+CD25+, and CD4+CD25++FOXP3+ cells 3) the levels of circulating soluble IL2 receptor alpha (sIL2rα) in 34 patients with PMF and Tregs <1% and 11 CTRLs by ELISA (R&D), due to the relevance of IL2 in the generation, function, and survival of Tregs and data reported by Tefferi A et al (J Clin Onc 2011; 29:1356-63).Results.1) The percentages of PB CD4+, CD4+CD25+, or CD4+CD25++CD127low/- cells expressing membrane Annexin V were comparable in patients with PMF and CTRLs (data not shown).2) The phosphorilation of Stat1, Stat3, Stat5, Stat6, and p38-MAPK (expressed as mean fluorescence intensity: MFI) on CD4+, CD4+CD25-, CD4+CD25+, and CD4+CD25++FOXP3+ cells before and after stimulation with IFNα, IL6, IL4, or PMA was comparable in patients and CTRLs. The Erk1,2 phosphorilation was comparable in patients and CTRLs at baseline condition, while in patients with PMF Erk1,2 phosphorilation following stimulation with PMA was significantly lower in both CD4+CD25+ cells (p=0.039; median MFI 5.4, range 0.3-40) and CD4+CD25++FOXP3+ cells (p=0.01; median MFI 4.3, range 1.4-9.3) than in CTRLs (CD4+CD25+ cells median MFI 8.1, range 4.9-20; CD4+CD25++FOXP3+ cells median MFI 7.3, 3.7-10.8).3) sIL2rα plasma levels were higher (p=0.01) in patients with PMF (1658 pg/ml, range 476-8968) than in CTRLs (1040 pg/ml, 482-1719); the sIL2rα plasma levels were significantly (R=-0.35, p=0.039) inversely correlated with the percentages of circulating Tregs. No significant correlation was found with the presence (n=13)/absence (n=14) of JAK2V617F mutation.Conclusions. These results indicate that in patients with PMF Tregs present an untuned Erk signaling; moreover, sIL2rα plasma levels are higher in patients than in CTRLs, and significantly inversely correlated with Treg percentages. These findings may partly explain the reduced frequency of Tregs in PMF since Erk signaling is pivotal in T cell differentiation, regulation of T cell homeostasis, and defects in Erk expression have been associated with autoimmunity (Chiung-Fang Chang J Immunol 2012; 189:721-731). Moreover, increased levels of sIL2rα may impact on IL-2-mediated functions and alterate the optimal activation of Erk1/2 that is IL-2 dependent (Lequn Li, Blood. 2005;106:3068-3073), DisclosuresNo relevant conflicts of interest to declare.

Role of Microenvironment-Associated Chemokines and Cytokines for Binet Stage A CLL Patients Included in a Prospective Trial (CLL1 trial) of the German CLL Study Group (GCLLSG): sIl2Ralpha Is An Independent Predictor of Progression-Free Survival (PFS),

Abstract 3869 Background:In CLL, proliferation of the leukemic cell clone occurs in the bone marrow and lymphatic tissues rather than peripheral blood. In this microenvironment, CLL cells interact with accessory cells, such as T cells and CD68+ nurselike cells (NLCs) and display signs of B cell receptor (BCR) activation, suggesting that CLL proliferation is T cell- and BCR-driven. In addition, cytokines and chemokines secreted by leukemic cells, stromal cells and T cells are essential in forming the disease-specific microenvironment. However, the exact biological role of cytokines and chemokines needs to be defined, especially in the light of distinct prognostic and biological subgroups of patients (pts). Methods:In order to address this issue, we measured serum levels of chemokines and cytokines in a prospective cohort of 157 previously untreated Binet stage A pts., a small subgroup of the risk-stratified CLL1 trial of the GCLLSG. Median follow-up time of that subgroup was 50.3 months. Median time to progression was 61.3 months. Median time from diagnosis to study entry was 1 year. Serum samples had been centrally collected at study entry and stored at −80°C. Sera were analyzed on a luminex-based multiplex platform, allowing simultaneous screening of multiple serum parameters. In a pilot phase, sera of 21 pts were pre-analytically screened for a total of 62 different chemokines. Median serum levels of 27 different chemokines and cytokines were found to differ from healthy controls in a significant manner. Those 27 chemokines and cytokines were subsequently analyzed in 157 pts. For all parameters univariate and multivariate analyses were performed for progression-free survival. Results:Serum levels of CCL3 and CCL4, chemokines known to be secreted by CLL cells upon BCR engagement, were elevated compared to healthy controls. High CCL3 levels correlated strongly with high CCL4 levels (p<0.001) and tended to be associated with unmutated IgHV status (p=0.06), supporting the role of BCR triggering in biologically selected CLL pts. Concerning CCL3 and CCL4, no significant difference in PFS was detected. Serum levels of CCL2 and CCL17, both binding chemokine receptor CCR4, were concordantly elevated compared to healthy controls (p<0.001), suggesting a possible role of chemoattraction of CCR4+ T cells towards these chemokines in CLL. CXCL12, CCL21, and CXCL10, chemokines associated with chemotaxis of CLL cells, were concordantly elevated compared to healthy controls. Pts with serum levels of CCL21 beyond the median also had higher levels of CXCL10 (p<0.001) and CXCL12 (p=0.02). CLL pts have been found to have abnormal neovascularization in the bone marrow and lymph nodes. Elevated VEGF levels were strongly associated with elevated EGF level (p<0.001). High VEGF levels correlated with high white blood cell counts (p=0.008) and showed a trend towards association with del(11q) (p=0.07) and lymphadenopathy (p=0.165). Concerning VEGF, no significant difference in PFS was detected. In contrast, pts with high levels of sIl2R alpha showed a significant shorter PFS. When confirmed by conventional ELISA, median PFS in pts within the highest quartile of sIl2Ralpha levels were 22 months compared to 72 months in the lower three quartiles (p<0.001). When we analyzed sIl2R alpha together with genetic abnormalities like del(11q), trisomy 12, del(13q), del(17p), the risk stratification model used in CLL1 (high risk versus low risk for disease progression), the hierarchical model as published by Döhner et al., and IgHV status in a multivariate Cox regression, we found that sIl2R alpha (OR 2.5, 95% CI 1.4–4.8, p=0.004) and IgHV mutational status (OR 4.0, 95% CI 2.2–7.3, p<0.001) were both independent prognostic variables. Conclusion:The assessment of sera in a small subgroup of the CLL1 study cohort of the GCLLSG revealed that the levels of several chemokines and cytokines were elevated when compared to healthy controls. Median serum levels of different chemokines correlated with distinct biological characteristics of CLL pts, like genetic abnormalities or clinical parameters. In addition, sIl2R alpha could be identified as an independent prognostic variable for PFS in early CLL. Disclosures:Eichhorst:Hoffmann La Roche: Honoraria, Research Funding, Travel Grants; Mundipharma: Research Funding, Travel Grants; Gilead: Consultancy. Stilgenbauer:Hoffmann La Roche: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Travel Grants. Hallek:Hoffmann La Roche: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees. Bergmann:Celgene: Honoraria.

The serum levels of soluble interleukin-2 receptor α and lactate dehydrogenase but not of B2-microglobulin correlate with selected clinico-pathological prognostic factors and response to therapy in childhood soft tissue sarcomas

To establish the clinical utility of serum soluble IL-2 receptor (sIL-2R alpha), lactate dehydrogenase (LDH) [corrected] and B2-microglobulin [corrected] (B2-M) as markers for diagnosis, prognosis and treatment monitoring in childhood soft tissue sarcomas (STS). The markers[corrected] were measured prospectively before treatment, in complete remission (CR) during and after therapy and at relapse [corrected] in 35 children with STS and in 50 healthy children [corrected] (once). Serum sIL-2R alpha and LDH [corrected] correlated with age thus they were [corrected] presented as multiplications [corrected] of the upper normal ranges [corrected] for age. Pre-treatment levels [corrected] of sIL-2R alpha and LDH [corrected] but not of B2-M exceeded significantly those of controls. [corrected] Elevated [corrected] sIL-2R alpha levels correlated with more [corrected] advanced stages, poor-risk histology and poor response to chemotherapy, higher LDH with incomplete primary tumour [corrected] resection and increased B2-M with poor-risk histology. [corrected] Patients' age >10 years, male gender and unfavourable tumour localisation were not accompanied by the markers' elevation. [corrected] None of the markers predicted EFS and OS. [corrected] Good response to chemotherapy was paralleled by significant decline of sIL-2R alpha and LDH pre-treatment levels while relapse--by sIL-2R alpha and LDH increase to values similar to those at diagnosis. [corrected] Monitoring of B2-M did not reflect the disease course. [corrected] sIL-2R alpha and LDH were [corrected] proven to be promising markers [corrected] for diagnosis and treatment monitoring in children with STS. The markers [corrected] correlated also with some [corrected] important prognostic clinico-pathological factors for childhood [corrected] STS; however, they [corrected] failed to predict EFS and OS. Measurements of serum [corrected] B2-M were shown [corrected] to have no clinical value in the diagnostics, prognostics and treatment monitoring in paediatric STS.

HIV vector-mediated targeted suicide gene therapy for adult T-cell leukemia

We investigated the potential efficacy of treating adult T-cell leukemia (ATL) using a gene therapeutic approach involving the use of a herpes simplex virus-thymidine kinase (HSV-TK)-mediated suicide system. Human immunodeficiency virus (HIV)-based vectors containing the HSV-TK gene were constructed to achieve targeted gene transfer into CD4-positive ATL cells, after which the transduced cells were selectively killed by treatment with ganciclovir (GCV). To examine the utility of HIV vectors in vivo, ATL-NOD-SCID mice were prepared by intraperitoneal injection of 1 x 10(7) MT2 cells into NK-depleted nonobese diabetic/severely compromised immunodeficient (NOD-SCID) mice. Thereafter, 1 ml of concentrated HIV vector expressing HSV-TK (HXCTKN) or GFP (HXGFP) stock was injected into the intraperitoneal cavity, and GCV was administered twice a day for 5 days. Fluorescence-activated cell sorting (FACS) analysis showed that 7-11% of MT2 or HUT102 cells recovered from the peritoneal cavity were transduced with the HXGFP. After 3 weeks, plasma sIL2-R alpha levels were significantly lower in mice administered HXCTKN than in those administered HXGFP. Moreover, HXCTKN-injected mice survived significantly longer than HXGFP-injected mice. Taken together, these findings suggest that HIV vectors could be used for in vivo targeted gene transfer into ATL cells and could thus serve as the basis for the development of effective new therapies for the treatment of ATL.

Decreased renal excretion of soluble interleukin-2 receptor α after treatment with daclizumab

Daclizumab (+/-150 kD), a humanized monoclonal antibody (mAb) against the alpha-chain of the membrane-bound interleukin-2 (IL-2) receptor (IL-2R) also binds soluble interleukin-2R alpha (sIL-2R alpha; +/-45 kD), and thus may influence the glomerular filtration of sIL-2R alpha. We have studied the influence of daclizumab on the renal excretion of sIL-2R alpha in 38 recipients of a renal transplant (32 treated with daclizumab and six controls). sIL-2R alpha was measured every 2 weeks after transplantation in serum and urine with Immulite IL-2R, a solid-phase enzyme-linked immunosorbent assay (ELISA). In the control population, the fractional excretion of sIL-2R alpha was relatively constant with a median value of 1.7%+/- 0.5%. In daclizumab-treated patients, sIL-2R alpha was not detectable in the urine immediately after the administration of daclizumab. sIL-2R alpha became detectable in the urine at a mean of 8 +/- 3 weeks after transplantation. In additional experiments, serum compounds were separated by size-exclusion chromatography and sIL-2R alpha was measured in the collected fractions. In the control patients, sIL-2R alpha was only present in the low-molecular-weight fractions of serum. In contrast, in daclizumab-treated patients evaluated several weeks after transplantation, sIL-2R alpha was merely detected in the high-molecular-weight fractions of serum. During follow-up there was a relative shift of sIL-2R alpha from the high- to the low-molecular-weight fractions and this coincided with normalization of sIL-2R alpha excretion. Daclizumab inhibits the renal excretion of sIL-2R alpha by the formation of a complex with sIL-2R alpha in serum, which is too large for glomerular filtration. Measurement of urinary sIL-2R alpha may provide information on the concentration of anti-IL-2R alpha mAb in serum.

Cytokine serum levels in soft tissue sarcoma patients: correlations with clinico-pathological features and prognosis.

We investigated the correlations between serum levels of selected proinflammatory, hematopoietic and angiogenic cytokines and soluble cytokine receptors with the clinico-pathological features and prognosis in soft tissue sarcoma patients. Serum levels of 9 cytokines (TNFalpha, IL-1ra, IL-6, IL-8, IL-10, M-CSF, G-CSF, VEGF, bFGF) and 4 free cytokine receptors (sIL-2R alpha, sIL-6R, TNFRI, TNFRII) were measured by means of an enzyme-linked immunoadsorbent assay kit in 156 soft tissue sarcoma patients before the treatment and in 50 healthy controls. Serum levels of 10 cytokines and cytokine receptors were also assayed during patients' follow-up after the treatment. Significantly elevated pretreatment serum levels of 11/13 cytokines and cytokine receptors were found in sarcoma patients, as compared to healthy controls. In 40.4% of patients 6 or more cytokines and cytokine receptors (most frequently: TNF RI, IL-6, IL-8) were elevated in parallel. Serum levels of IL-6, sIL-2R, VEGF, M-CSF and TNF RI correlated significantly with tumor size and serum levels of IL-8 and IL-6 were significantly higher in patients with Grade 2/3 vs. Grade 1 tumors. We did not observe any significant differences in cytokine serum levels between patients with primary and recurrent tumors and patients with and without distant metastases. Using univariate analysis, overall survival (OS) in all patients was affected by tumor size (<5 cm vs. 5-10 cm vs. >10 cm), tumor grade (G1 vs. G2/3), presence of metastases, pretreatment serum levels of 8 cytokines (IL-6, IL-8, IL-10, sIL-2R, TNF RI, TNF RII, M-CSF, VEGF) and the number of cytokines increased (0-1 vs. 2-5 vs. < or = 6). Elevated serum levels of IL-6, IL-8, IL-10 and sIL-2R alpha, high tumor grade and larger tumor size strongly correlated with shorter disease-free survival (DFS). Multivariate analysis identified G2/3 tumor grade (p = 0.001), the presence of metastases (p = 0.004), elevated IL-6 serum level (p = 0.02), elevated IL-8 serum level (p = 0.048) and the number of cytokine serum levels above upper cut-off values (p = 0.01) as the independent prognostic factors related to OS, and G2/3 tumor grade (p = 0.005) and increased IL-6 serum level (p = 0.035) as independent prognostic factors related to DFS. In a group of patients without metastases (M0) higher tumor grade, elevated serum level of IL-6 and TNF RII, and the number of elevated cytokine serum levels correlated independently with poor survival. We found a significant decrease of several cytokine serum levels in patients after treatment (IL-1ra, IL-6, IL-8, IL-10, TNF RII, M-CSF) [p < 0.05]. Persistently elevated serum level of IL-6 after the treatment has also shown negative prognostic significance for OS (univariate analysis). Serum levels of some proinflammatory, hematopoietic and angiogenic cytokines and cytokine receptors are elevated, frequently in parallel, in a large percentage of soft tissue sarcoma patients. Significant correlations of serum cytokine levels with tumor size and grade suggest that some of these cytokines may be directly or indirectly involved in the progression of soft tissue sarcomas. Serum assays of IL-6, IL-8 and TNF RII before or after the treatment may be useful in establishing soft tissue sarcoma patients prognosis.

The significance of soluble interleukin-2 receptor in monitoring disease relapse in patients with nasopharyngeal cancer.

Soluble interleukin-2 receptor alpha (sIL-2R alpha) is a well-known indicator of T-cell activation noted to be increasing in nasopharyngeal cancer. However, the significance of sIL-2R alpha in monitoring disease relapse is unclear. This study was initiated to address this issue. Serum of 56 patients with NPC, which underwent either primary, salvage, or palliative treatments, from 1992 to 1993 at the Cancer Center, Veterans General Hospital-Taipei, were collected from our serum bank. According to their disease status at the time of study, at least two years after last treatments, the 56 patients were divided into four groups. The remission group represented those in remission at the time of study (n = 24). The metastasis group represented those with distant metastasis present at the time of study (n = 17). The recurrence group represented those with locoregional recurrence present at the time of study (n = 11). The combined group represented those with locoregional recurrence as well as distant metastasis (n = 4). The seral sIL-2R alpha concentrations of the 56 NPC patients were determined with enzyme-linked immunoabsorbent assay. The combined group was excluded in our statistical analysis. We performed statistical analysis on the differences of paired serum sIL-2R alpha concentrations between different periods of the diseases. The first analysis was on the differences of sIL-2R alpha concentrations between diagnosis and post-radiotherapy periods for 13 out of 24 patients in the remission group and 7 out of 11 patients in the recurrence group. The second analysis was on the differences of sIL-2R alpha concentration between follow-up before detection-of-relapse and after detection-of-relapse for 5 out of 17 patients in the metastasis group and six out of 11 patients in the recurrence group. The first statistical analysis revealed no significant differences of sIL-2R alpha concentrations for the remission group (P = 0.946) and the recurrence group (P = 0.156) between diagnosis and post-radiotherapy periods. The second statistical analysis revealed no significant differences of sIL-2R alpha concentrations between before and after detection-of-relapse for the recurrence group, neither (P = 0.438). The results for the metastasis group were different. The sIL-2R alpha concentrations were shown to increase after the detection of metastasis for the 5 paired samples from the metastasis group, although the Wilcoxon signed ranks test on the differences only showed borderline significance (P = 0.063). Our findings show that sIL-2R alpha would be of no value in monitoring the development of locoregional recurrence but might be useful in monitoring distant metastasis. Although our current limited data did not provide strong support for the role of sIL-2R alpha in monitoring metastasis, it might be delineated in the future by collecting more data.

Serum cytokine levels after HLA-identical bone marrow transplantation.

Altered profiles of cytokine production are observed after bone marrow transplantation (BMT). The presence of certain cytokines in serum can be indicative for BMT-related complications, such as graft-versus-host disease (GVHD) and infections. The putative correlation between abnormal serum cytokine levels and BMT-related complications was further analyzed in this retrospective study. Serum levels of a panel of cytokines and cytokine-associated molecules (i.e., interleukin [IL]-1alpha, IL-1beta, IL-4, IL-6, IL-10, IL-12, IL-1 receptor antagonist, and the soluble a chain of the IL-2 receptor [sIL-2R alpha]) were assessed in 329 sera of 46 patients who had undergone HLA-identical or autologous BMT. Serum cytokine levels of the BMT donor and of the patients before BMT and at different time points during the post-BMT period were measured. The results were correlated with relapse, acute GVHD, chronic GVHD, and infections. Serum levels of IL-1alpha, IL-1beta, IL-4, and IL-12 were undetectable. The transplantation regimen itself causes a significant rise in IL-10 and sIL-2R alpha levels in patients receiving allogeneic bone marrow. In the post-BMT period, increased IL-6 serum levels were significantly correlated with infections. Increased IL-10 levels were significantly correlated with acute graft-versus-host disease, chronic GVHD, and infections. Increased sIL-2R alpha levels were correlated with chronic GVHD, as were IL-1 receptor antagonist levels. During the post-HLA-identical BMT period, the serum cytokine levels of IL-6 were enhanced during infections, whereas the sIL-2R alpha levels were increased during chronic GVHD. The serum levels of IL-10 and of the cytokine-related molecule IL-1ra were enhanced during both infections and chronic GVHD. These results further substantiate the complex cytokine cascade that is initiated by the conditioning regimen and that evolves further in reaction to BMT-related complications and their treatments.

The circulating common gamma chain (CD132) in inflammatory bowel disease

Inflammatory bowel disease (IBD) is characterized by T cell activation. Activated T cells shed interleukin-2 receptors (IL-2R) in a soluble form. A positive correlation between sIL-2Ralpha (CD25) and disease activity is well documented in IBD, whereas IL-2Rgamma (CD132) has not been investigated in this respect. Sera from 42 patients with ulcerative colitis (UC), 34 with Crohn's disease (CD), 31 healthy volunteers, and 12 patients with infectious enterocolitis were obtained.Disease activity was scored according to a semiquantitative score for UC and CD. sIL-2R alpha chain and gamma chain were assessed by sandwich ELISA techniques using monoclonal antibodies specific for CD25 and CD132, respectively.The concentration of IL-2Ralpha chain (CD25) was found to be median 3.8 ng/ml in healthy volunteers versus 7.0 ng/ml in UC patients (p < 0.001), and 9.6 ng/ml in CD patients (p < 0.001). With respect to IL-2Rgamma (CD132), significantly higher amounts were found in CD patients: 6.6 ng/ml as compared with healthy controls <1.0 ng/ml (p < 0.004). A Kruskal-Wallis test revealed a significant correlation between alpha chain and disease activity in CD (p < 0.001), and further significantly higher gamma chain levels were found in active CD (p = 0.03). For UC patients, a statistically significant increase of the alpha chain with increasing disease activity (p < 0.01) was observed, whereas no significant changes of the gamma chain levels were found (p > 0.05). A difference of gamma chain levels were found between CD and UC in moderate and severe disease activity (p < 0.05). Further analyses revealed that mesalazine did not influence the IL-2Ralpha or -gamma concentration either in UC or in CD patients.An increased circulating level of the soluble common gamma chain (CD132) seems to be found in CD, and an overlap exists between CD and UC.

Cytokines of the human reproductive tract.

How is it possible that the female genital tract immunologically does not reject spermatooa not the preimplantation and nidating embryo? Four fluids of the human reproductive tract, i.e., human oviductal fluid (hOF), follicular fluid (FF), amniotic fluid (AF), and seminal plasma (SP) were investigated by specific ELISA for 18 cytokines. The concentrations, presence or absence of these compounds were evaluated for their possible role in the immunology of the reproductive process. Stem cell factor and IL-11 were detected in all reproductive tract fluids examined whereas large amounts of IL-1 beta and IL-1RA was found in AF and hOF. Follicular fluid revealed IL-2. HOF contained IL 2, IL-6, IL-8, TNF-alpha, MIP-1 alpha, IFN-gamma, and high levels of IL-1 beta, IL-10, IL-1RA, and sIL-2R. Amniotic fluid contained sIL-2R, IL-8, IL-1 beta, IL-1RA, IL-6, TNF-alpha, and MIP-1 alpha. No IL-12 or IL-13 was detected in hOF follicular fluid or amniotic fluid. Almost no free TGF-beta 1 or TGF-beta 2 was found in any reproductive tract fluid except seminal plasma. Seminal plasma contained large quantities of free TGF-beta 1 (9,220 +/- 3,635 pg/mL) in addition to large quantities of latent TGF-beta 2 (2,933 +/- 2,169 pg/mL) and TGF-beta 1 (71,000 +/- 3,240 pg/mL). Furthermore, considerable concentrations of IL-8 (1900 +/- 374 pg/mL) and sIL-2R (350 mu/mL) exist in seminal plasma. HOF contains a high level of IL-10 (588 +/- 304 pg/mL), a powerful immune suppressor which probably plays a role in regulating immune responses in the fallopian tube and possibly in the endometrial cavity. Our observations suggest that seminal plasma with its huge content of TGF beta provides immune protection for sperm. Unfortunately, such high concentrations of TGF beta may also inhibit an immune defense in any organ in which semen is deposited.

Increased mucosal concentrations of soluble intercellular adhesion molecule-1 (sICAM-1), sE-selectin, and interleukin-8 in active ulcerative colitis.

Cell surface adhesion molecules (CAM) are important promotors of the immunoinflammatory cascade. The circulating levels of soluble intercellular adhesion molecule 1 (ICAM-1) have previously been shown to correlate with disease activity in inflammatory bowel disease. The primary aim of this study was consequently to investigate if this also applies to mucosal levels of soluble ICAM-1. We measured soluble ICAM-1 levels in intestinal biopsy specimens and the endoscopic activity of 69 patients with ulcerative colitis (UC) and 14 controls and found that the median concentration of soluble ICAM-1 was significantly higher in patients with moderately or very active UC (15.0 ng/ml) as compared to slightly active (9.8 ng/ml) and inactive UC (9.5 ng/ml) as well as controls (6.5 ng/ml) (P < 0.005). To further elucidate the interactions, two other CAM [E-selectin and vascular cellular adhesion molecule 1 (VCAM-1)], together with interleukin-8 (IL-8), IL-2 receptor (IL-2R) alpha and beta chains, were also measured. A significant trend towards higher soluble E-selectin levels in biopsies with active UC (1.8 pg/ml) as compared to inactive UC (1.3 pg/ml) and to controls (< 1.0 pg/ml) (P < 0.01) was also found. In contrast, soluble VCAM-1 was barely detectable in biopsies from two UC patients. A significant correlation was found between soluble ICAM-1 and IL-8 concentrations (r = 0.46; P < 0.0001), and between sICAM-1 and sIL-2R alpha concentrations (r = 0.69; P < 0.0001), while sIL-2R beta was not detected. This study shows that intestinal ICAM-1 and E-selectin correlate with endoscopic activity of UC and with IL-8 and IL-2R alpha levels. These mediators may be useful in monitoring mucosal inflammation in studies exploring the therapeutical potential of targeting CAM. The lack of detectable VCAM-1, which is induced only in venous endothelium is interesting. It may suggest that intestinal inflammation mainly affects arterial endothelial cells and support the theory that intestinal vasculitis is involved in the pathogenesis of inflammatory bowel disease.

Metabolism of Tac (IL2Ralpha): physiology of cell surface shedding and renal catabolism, and suppression of catabolism by antibody binding.

The interleukin 2 receptor alpha (IL2Ralpha; CD25; Tac) is the prototypic model for soluble receptor studies. It exists in vivo as a transmembrane complete molecule (TM-Tac) on cell surfaces and as a truncated soluble form (sTac; sIL2R alpha). sTac has been used as a serum marker of T cell activation in immune disorders and of tumor burden in Tac-expressing malignancies. In vivo, serum levels of all soluble proteins depend on the balance between production and catabolism, but little is known about the metabolic features of this class of molecules. We have developed a model for Tac metabolism that incorporates new insights in its production and catabolism. Tac was shed from the surface of malignant and activated human T cells with a model half-life (t1/2) of 2-6h, but which was prolonged under certain circumstances. The rate of shedding is first order overall and nonsaturable over a two order of magnitude range of substrate (TM-Tac) expression. Once shed from cells Tac is subject to catabolic activities in the host. In vivo studies in mice showed that 90% of Tac was catabolized by the kidney with a t1/2 of 1 h and a filtration fraction of 0.11 relative to creatinine. The remaining 10% of catabolism was mediated by other tissues with a t1/2 of 10 h. Approximately 1-3% of sTac is excreted intact as proteinuria with the remaining 97-99% catabolized to amino acids. Antibody to the receptor induced a marked delay in sTac catabolism by preventing filtration of the smaller protein through the renal glomerulus and additionally suppressing other nonrenal catabolic mechanisms. A discrepancy between the catabolic rats for Tac and anti-Tac in the same complex was interpreted as a previously unrecognized differential catabolic mechanism, suggesting features of the Brambell hypothesis and immunoglobulin G transport and catabolism, in which the antigen-in-complex in intracellular vesicles is relatively less protected from catabolism than the associated antibody. In light of the pivotal role played by the kidney in sTac catabolism and the impact of administered antibody, the serum concentration of Tac in the settings of renal dysfunction or antibody therapy is not a suitable surrogate of activated T cells or of the body burden of tumor. These results provide parameters for assessing soluble receptor-ligand interactions generally.

Circulating interleukin-2 receptors are a group of multimeric proteins with immunoreactivity for interleukin-2 receptor alpha, beta, and gamma chains.

Soluble interleukin-2 receptor (sIL-2R) alpha chain serum levels as determined by enzyme-linked immunosorbant assay (ELISA) are commonly used to monitor various inflammatory and neoplastic disorders associated with lymphocytic proliferation and activation. The in vivo structure of this soluble receptor species, however, is not characterized. We investigated sera with elevated sIL-2R serum levels of patients with histologically proven cutaneous T cell lymphoma (CTCL) and high-dose IL-2-treated melanoma patients and healthy donors. Purified recombinant IL-2R alpha and beta chain molecules, produced by transfection of NIH/3T3 fibroblasts and CHO cells, served as positive controls for purification and detection procedures. For selective enrichment of IL-2R molecules from supernatants and sera, affinity columns were prepared by coupling recombinant IL-2 or monoclonal antibodies against the alpha and the beta chain of the IL-2R complex to cyanogen bromide-activated Sepharose. Western blotting with affinity-purified fractions under reducing and nonreducing conditions revealed proteins that showed immunoreactivity for IL-2R alpha, beta, and gamma chain using several detection antibodies against these molecules. We conclude that the composition of sIL-2R in vivo is more complex than that of recombinant sIL-2R and can include all three IL-2R chains.

Targeting Human IL-2 Receptors for Diagnosis and Therapy

The high-affinity interleukin-2 receptor (IL-2R) is a multichain receptor with at least three IL-2 binding chains: IL-2R alpha (55 kDa) bound by the monoclonal antibody anti-Tac, IL-2R beta (75 kDa) and Il-2R gamma (64 kDa). The IL-2R alpha also exists as a naturally occurring soluble molecule (sIL-2R alpha). We target the IL-2R for immune intervention since resting normal cells do not express the high-affinity IL-2R, whereas this receptor is on some cells in certain lymphoid neoplasias, select autoimmune disorders, and in individuals rejecting organ allografts. Treatments have included unmodified murine anti-Tac and radioisotopes conjugated to murine anti-Tac. Our emerging understanding of the IL-2R system continues to open possibilities for more specific immune intervention.

Serum soluble interleukin-2 receptor alpha levels in patients with gynecologic cancers: early effect of surgery.

To determine the early effect of abdominal cytoreductive surgery on serum soluble interleukin-2 receptor alpha (sIL-2R alpha) levels in patients with ovarian and cervical cancer, and to determine if the extent of cytoreduction correlated with the changes in serum sIL-2R alpha. Thirty-nine patients with gynecologic cancer had serum sIL-2R alpha measured by enzyme-linked immunosorbent assay before abdominal cytoreductive surgery and once in the early postoperative period. Only patients with advanced epithelial ovarian cancer had elevated preoperative serum sIL-2R alpha levels. In 20 of 25 ovarian cancer cases (80%) and 10 of 14 cervical cancer cases (71.4%) the postoperative serum sIL-2R alpha levels exceeded the preoperative level (P = .003 and P = .011, respectively). Overall, the mean postoperative serum sIL-2R alpha level was greater than the preoperative level (P = .0001). Patients with early stage gynecologic cancer did not have elevated serum sIL-2R alpha levels before surgery. In the early postoperative period the serum sIL-2R alpha level was increased, which may be a nonspecific response to the trauma of surgery. Soluble IL-2R alpha may be one of the factors responsible for the immunosuppression in the early postoperative period, but may also herald a surge of activated T cells.

Cytokines predict coronary aneurysm formation in Kawasaki disease patients

In this study, we measured serially the serum levels of cytokines including interleukin-6 (IL-6), IL-8, soluble IL-2 receptor (sIL-2R) and tumour necrosis factor alpha (TNF-alpha) in 60 patients with Kawasaki disease (KD) and evaluated the clinical significance of these cytokines in predicting coronary aneurysm formation. Of the 60 patients, 12 were complicated with coronary aneurysm. Blood samples were collected within the 1st week after onset of fever, then once a week for the 1st month, and once a month for another 5 months. The serum levels of IL-6, IL-8, sIL-2R and TNF alpha were measured using an ELISA or RIA method. Our results show that the changes in serum IL-6 and IL-8 were faster than those of sIL-2R and TNF alpha. Within the 1st week, the serum levels of IL-6 and IL-8 were significantly higher in the patients with than in those without coronary aneurysm (P < 0.001). In addition, the serum levels of IL-6 and IL-8 obtained in the 1st week were highly correlated (P < 0.001) with those of C-reactive protein and erythrocyte sedimentation rate, and the serum levels of sIL-2R and TNF alpha were also increased at the 1st week reaching the highest level in the 2nd week. In the 2nd week, the serum levels of sIL-2R and TNF alpha were significantly higher in the patients with than in those without coronary aneurysm (P < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

High serum and ascitic soluble interleukin-2 receptor alpha levels in advanced epithelial ovarian cancer [see comments

This study was undertaken to determine if advanced epithelial ovarian cancer was associated with increased serum and ascitic levels of soluble interleukin-2 receptor alpha (sIL-2R alpha). Serum and ascitic fluid samples from 23 ovarian cancer patients were analyzed for sIL-2R alpha using an enzyme-linked immunosorbent assay and compared with the serum and peritoneal levels in 18 normal females. The samples were analyzed for CA-125 levels using a radioimmunoassay and the total protein was also measured. Normal individuals had low serum levels of sIL-2R alpha (367.5 +/- 44.6 U/mL), with similar levels of sIL-2R alpha in the normal peritoneal fluid (438.6 +/- 48.8 U/mL). In contrast, the serum and ascitic fluid levels in ovarian cancer patients were significantly higher (746.7 +/- 82.9 U/mL, P = .0006; 2,656.7 +/- 373.7 U/mL, P = .00002, respectively). The results for sIL-2R alpha were also significant when the levels were expressed per milligram of total protein. More importantly, in almost every ovarian cancer patient the ascitic sIL-2R alpha level far exceeded the serum level, a pattern also observed for CA-125. There was no correlation between the serum and ascitic sIL-2R alpha levels, or between the serum and ascitic CA-125 levels. Although the serum levels of sIL-2R alpha and CA-125 were elevated in the same patient, overall there was no correlation between the serum sIL-2R alpha and serum CA-125 levels, either when the levels were expressed in absolute units or per milligram of total protein. Similarly, there was no correlation between sIL-2R alpha and CA-125 levels in individual ascitic samples. While CA-125 levels may reflect an independent index of tumor burden, these results suggest that selective accumulation of sIL-2R alpha in the ascites may be one of the factors associated with the known nonresponsiveness of the infiltrating lymphocytes against ovarian carcinoma cells.

The expression and biological activity of IL-2 receptor on a human pancreas cancer cell line

To ascertain whether the tumor cells can regulate the host immune systems through the production of the cytokines or their receptors, we examined the expressions of tumor necrosis factor alpha (TNF alpha), tumor necrosis factor beta (TNF beta), interleukin 2 (IL-2) and interleukin 2 receptor alpha chain (IL-2R alpha) on the human cancer cell lines by Northern blot analysis. We used K562 (leukemia cell line), MCF-7 (breast cancer cell line), LS180, HT29 (colon cancer cell lines), SH101 (gastric cancer cell line) and PH101 (pancreas cancer cell line). Expressions of TNF alpha, TNF beta and IL-2 mRNA were not detected in any of the tumor cell lines. However, 1.4 and 3.5 kilobases of the IL-2R alpha mRNA were expressed in the PH101 cells, but not in the other five cell lines. Furthermore, IL-2R alpha was detected on the cell surface of the PH101 cells by the flow-cytometric analysis with an anti-IL-2R alpha monoclonal antibody. Interestingly, the soluble IL-2R alpha (sIL-2R alpha) was found in the conditioned media obtained from the PH101 cell culture with a sandwich enzyme immunoassay. Moreover, the sIL-2R alpha secreted from the PH101 cells blocked the IL-2 dependent lymphocyte proliferation. These results indicate that the expression of IL-2R alpha on PH101 might suppress the IL-2 induced lymphocyte proliferation.

Increased levels of soluble tumor necrosis factor receptors in the sera and synovial fluid of patients with rheumatic diseases.

Recently, 2 classes of cytokine inhibitors have been defined at the molecular level. The largest group comprises the extracellular domains of cell surface cytokine receptors, and includes both tumor necrosis factor receptors (TNF-R). The present study was conducted to investigate the role of TNF inhibitors in arthritis. We measured p55 and p75 soluble TNF-R (sTNF-R) in serum and synovial fluid (SF) samples from patients with rheumatic diseases and compared their levels with levels of soluble interleukin-2 receptors (sIL-2R). Sensitive enzyme-linked immunosorbent assays (ELISA), specific for p55 and p75 sTNF-R and for sIL-2R, were used. Serum levels of p75 sTNF-R were 3-4-fold higher than levels of p55 sTNF-R, and both were significantly elevated in patients with osteoarthritis (OA) and rheumatoid arthritis (RA) compared with healthy controls. RA SF levels of sTNF-R were 4-5-fold higher than levels in serum, suggesting local production in the joint, and were significantly higher than levels in the SF of patients with seronegative arthropathy or OA. Furthermore, levels of p55 and p75 sTNF-R, but not sIL-2R or TNF alpha measured by ELISA, were increased in the SF of patients with clinically active RA. The soluble TNF-R in RA and OA SF were functional since they inhibited TNF activity in a cytotoxicity assay in proportion to the levels of inhibitor present. Evaluation of serially obtained serum samples suggested that sTNF-R may be a useful parameter for monitoring RA disease activity. Biologically active soluble TNF-R are up-regulated in patients with rheumatic disease and are produced locally in the joints. Measurement of serum levels of TNF-R may be useful for monitoring of disease, and determination of SF levels could be of diagnostic value.