Decreased interleukin-11 levels in the semen of infertile males.

The relationship between the interleukin (IL)-11 levels and semen quality of infertile males remains unknown. In this study, 120 semen samples from 60 normozoospermic volunteers and 60 infertile males were examined. The semen pH of the normozoospermic group was not significantly different from that of the infertile group. The semen volume, semen density, forward movement percentage, activity, survival rate and normal morphology rate of the sperm of the infertile group were significantly lower than those of the normozoospermic group. The semen IL-11 levels of the infertile group were significantly lower than those of the normozoospermic group. In the infertile group, semen IL-11 levels were positively correlated with sperm motility, vitality, survival rate and normal sperm morphology rate and negatively correlated with IL-17 and IL-18 levels. Therefore, semen IL-11 levels could be used as an index of male infertility.

Expression, Purification, and Characterization of Interleukin-11 Orthologues.

Interleukin-11 (IL-11) is a multifunctional cytokine implicated in several normal and pathological processes. The decoding of IL-11 function and development of IL-11-targeted drugs dictate the use of laboratory animals and need of the better understanding of species specificity of IL-11 signaling. Here, we present a method for the recombinant interleukin-11 (rIL-11) production from the important model animals, mouse and macaque. The purified mouse and macaque rIL-11 interact with extracellular domain of human IL-11 receptor subunit α and activate STAT3 signaling in HEK293 cells co-expressing human IL-11 receptors with efficacies resembling those of human rIL-11. Hence, the evolutionary divergence does not impair IL-11 signaling. Furthermore, compared to human rIL-11 its macaque orthologue is 8-fold more effective STAT3 activator, which favors its use for treatment of thrombocytopenia as a potent substitute for human rIL-11. Compared to IL-6, IL-11 signaling exhibits lower species specificity, likely due to less conserved intrinsic disorder propensity within IL-6 orthologues. The developed express method for preparation of functionally active macaque/mouse rIL-11 samples is suited for exploration of the molecular mechanisms underlying IL-11 action and for development of the drug candidates for therapy of oncologic/hematologic/inflammatory diseases related to IL-11 signaling.

Bone-Conditioned Medium Changes Gene Expression in Bone-Derived Fibroblasts.

PURPOSE: Autologous bone is used for augmentation in the course of oral implant placement. Bone grafts release paracrine signals that can modulate mesenchymal cell differentiation in vitro. The detailed genetic response of the bone-derived fibroblasts to these paracrine signals has remained elusive. Paracrine signals accumulate in bone-conditioned medium (BCM) prepared from porcine cortical bone chips. MATERIALS AND METHODS: In this study, bone-derived fibroblasts were exposed to BCM followed by a whole genome expression profiling and downstream quantitative reverse transciptase polymerase chain reaction of the most strongly regulated genes. RESULTS: The data show that ADM, IL11, IL33, NOX4, PRG4, and PTX3 were differentially expressed in response to BCM in bone-derived fibroblasts. The transforming growth factor beta (TGF-ß) receptor 1 antagonist SB431542 blocked the effect of BCM on the expression of the gene panel, except for IL33. CONCLUSION: These in vitro results extend existing evidence that cortical bone chips release paracrine signals that provoke a robust genetic response in mesenchymal cells that is not exclusively mediated via the TGF-ß receptor. The present data provide further insights into the process of graft consolidation.

Interleukin-6 Family of Cytokines in Crevicular Fluid of Renal Transplant Recipients With and Without Cyclosporine A-Induced Gingival Overgrowth.

BACKGROUND: Interleukin (IL)-6 family of cytokines, including IL-6, oncostatin M (OSM), leukemia inhibitory factor (LIF), and IL-11, have fibrogenic features. The current study determines gingival crevicular fluid (GCF) levels of fibrosis-related IL-6-type cytokines in cyclosporine A (CsA)-induced gingival overgrowth (GO). METHODS: Eighty non-smokers were included (40 CsA-medicated renal transplant patients with GO [GO+; n = 20] or without GO [GO-; n = 20], 20 individuals with gingivitis, and 20 healthy participants). Probing depth and plaque, papilla bleeding, and hyperplastic index scores were recorded. GCF samples were obtained from the mesio-buccal aspects of two teeth. GCF IL-6, IL-1ß, OSM, LIF, and IL-11 levels were analyzed by enzyme-linked immunosorbent assay. RESULTS: The GO+ and GO- groups had higher IL-6 total amounts than the healthy group (P <0.008). IL-1ß total amounts in the GO+ group were significantly higher than in both the healthy and GO- groups (P <0.008). OSM total amount was elevated in the GO+ and GO- groups compared with both the gingivitis and healthy groups (P <0.008). All groups had similar LIF and IL-11 total amounts (P >0.008). Moderate positive correlations were detected among IL-6, IL-1ß, OSM, and IL-11 total amount in GCF and clinical parameters (P <0.05). CONCLUSIONS: IL-6 and OSM increases in GCF as a result of CsA usage or an immunosuppressed state irrespective of the severity of inflammation and the presence of GO. The IL-6 family of cytokines might not be directly involved in biologic mechanisms associated with CsA-induced GO. Lack of an association between assessed IL-6 cytokines and CsA-induced GO might indicate distinct effects of these cytokines on fibrotic changes of different tissues.

Conditioned medium of demineralized freeze-dried bone activates gene expression in periodontal fibroblasts in vitro.

BACKGROUND: Demineralized bone matrix (DBM) is used for the treatment of osseous defects. Conditioned medium from native bone chips can activate transforming growth factor (TGF)-ß signaling in mesenchymal cells. The aim of this study is to determine whether processing of native bone into DBM affects the activity of the conditioned medium. METHODS: Porcine cortical bone blocks were subjected to defatting, different concentrations of hydrochloric acid, and various temperatures. DBM was lyophilized, ground, and placed into culture medium. Human gingiva and periodontal fibroblasts were exposed to the respective conditioned medium obtained from DBM (DBCM). Changes in the expression of TGF-ß target genes were determined. RESULTS: DBCM altered the expression of TGF-ß target genes (e.g., adrenomedullin, pentraxin 3, KN motif and ankyrin repeat domains 4, interleukin 11, NADPH oxidase 4, and BTB [POZ] domain containing 11) by at least five-fold. The response was observed in fibroblasts from both sources. Defatting lowered the activity of DBCM. The TGF-ß receptor type I kinase inhibitor SB431542 [4-(4-(benzo[d][1,3]dioxol-5-yl)-5-(pyridin-2-yl)-1H-imidazol-2-yl)benzamide] but not the inhibitor of bone morphogenetic protein receptor dorsomorphin, blocked the effects of DBCM on gene expression. Moreover, conditioned medium obtained from commercial human DBM modulated the expression of TGF-ß target genes. CONCLUSION: The findings suggest that the DBCM can activate TGF-ß signaling in oral fibroblasts.

A Major Human Oral Lysophosphatidic Acid Species, LPA 18:1, Regulates Novel Genes in Human Gingival Fibroblasts.

BACKGROUND: The small bioactive lipid lysophosphatidic acid (LPA) plays critical roles in both normal physiology and inflammation in many systems. However, its actions are just beginning to be defined in oral biology and pathophysiology. METHODS: Microarray analysis was used to test the hypothesis that human gingival fibroblasts (GFs) would show significant changes in wound-healing and inflammation-related gene transcripts in response to a major human salivary and gingival crevicular fluid LPA species, 18:1, and that they would express transcript for the major LPA-producing enzyme autotaxin. The microarray results were validated for three highly relevant upregulated inflammatory transcripts using quantitative reverse transcription-polymerase chain reaction (QRT-PCR). Liquid chromatography-tandem mass spectrometry was used to assay time-dependent LPA species production by GFs. RESULTS: LPA 18:1 significantly regulated 20 GF novel and 27 known genes linked to the control of inflammation (P ≤0.01). QRT-PCR validation of interleukin (IL)-8, IL-11, and suppressor of cytokine signaling 2 (SOCS2) messenger RNAs confirmed statistically significant differences from control (P ≤0.05). Autotaxin transcript was present, and GFs were found to produce multiple LPA species in a time-dependent manner. CONCLUSIONS: The upregulation of transcripts for known GF proinflammatory (IL-6, IL-8) and anti-inflammatory (IL-11) ILs, along with SOCS2, shows that LPA transiently regulates a complex set of GF genes critical to periodontal wound healing and inflammation. These results implicate LPA exerting actions on GFs that are compatible with functioning as a mediator in oral fibroblast biology and inflammatory responses. Therefore, LPA may potentially modulate/regulate periodontal inflammation.

High expression of interleukin-11 is an independent indicator of poor prognosis in clear-cell renal cell carcinoma.

Interleukin-11 (IL-11), a member of the IL-6 family of cytokines, exerts pleiotropic oncogenic activities by stimulating angiogenesis and metastasis in many cancer types. The present study aims to evaluate the impact of IL-11 expression on recurrence and mortality of patients with clear-cell renal cell carcinoma (ccRCC). We retrospectively enrolled 193 ccRCC patients undergoing nephrectomy at a single center. Clinicopathologic features, recurrence-free survival (RFS) and overall survival (OS) were recorded. IL-11 intensity was assessed by immunohistochemistry in tumor specimens. The Kaplan-Meier method was applied to compare survival curves. Cox regression models were used to analyze the impact of prognostic factors on RFS and OS. The concordance index (C-index) was calculated to assess predictive accuracy. High IL-11 expression is associated with increased risk of recurrence and poor survival for ccRCC patients (P < 0.001 and P < 0.001, respectively), especially those with early-stage disease (TNM stage I + II). Multivariate analyses confirmed that IL-11 expression was an independent prognostic factor for RFS and OS (P = 0.006 and P = 0.008, respectively). The predictive accuracy of well-established prognostic models was improved when IL-11 expression was integrated. In conclusion, high IL-11 expression is an independent predictor of poor prognosis in ccRCC patients. It may help identify patients who could benefit from additional treatments and closer follow up.

Stability-indicating capillary zone electrophoresis method for the assessment of recombinant human interleukin-11 and its correlation with reversed-phase liquid chromatography and biossay.

A stability-indicating capillary zone electrophoresis (CZE) method was validated for the analysis of recombinant human interleukin-11(rhIL-11) using rupatadine fumarate, as internal standard (IS). A fused-silica capillary, (50 µm i.d.; effective length, 40 cm) was used at 25°C; the applied voltage was 20 kV. The background electrolyte solution consisted of 50 mmol L(-1) sodium dihydrogen phosphate solution at pH 3.0. Injections were performed using a pressure mode at 50 mbar for 45 s, with detection by photodiode array detector set at 196 nm. Specificity and stability-indicating capability were established in degradation studies, which also showed that there was no interference of the excipients. The method was linear over the concentration range of 1.0-300 µg mL(-1) (r(2)=0.9992) and the limit of detection (LOD) and limit of quantitation (LOQ) were 0.2 µg mL(-1) and 1.0 µg mL(-1), respectively. The accuracy was 100.4% with bias lower than 1.1%. Moreover, the in vitro cytotoxicity test of the degraded products showed significant differences (p<0.05). The method was applied for the content/potency assessment of rhIL-11 in biopharmaceutical formulations, and the results were correlated to those of a validated reversed-phase LC method (RP-LC) and an TF-1 cell culture assay, showing non-significant differences (p>0.05). In addition the CZE and RP-LC methods were applied for the analysis of rhIL-11 in human plasma. Therefore, the proposed alternative method can be applied to monitor stability, to assure the batch-to-batch consistency and quality of the bulk and finished biotechnology-derived medicine.

IL-17 and IL-11 GCF levels in aggressive and chronic periodontitis patients: relation to PCR bacterial detection.

OBJECTIVES: This study evaluated IL-17 and IL-11 in gingival crevicular fluid (GCF) of generalized chronic periodontitis (GCP) and generalized aggressive periodontitis (GAgP) patients in relation to periodontopathic bacteria. SUBJECTS AND METHODS: GCF samples were collected from 65 subjects including 25 CP, 25 GAgP, and 15 controls (C) and analyzed for IL-17 and IL-11 by an enzyme-linked immunosorbent assay. Molecular detection of bacteria in the dental plaque was determined by polymerase chain reaction. RESULTS: The total amount of IL-17 was significantly higher in GAgP group than in GCP and C groups (P < 0.001). The IL-11 concentration was significantly higher in C and GCP groups than GAgP group (P < 0.001). The IL-11/IL-17 ratio was significantly higher in the C group than in GCP and GAgP groups (P < 0.05). Moreover, GAgP group showed lower ratios of IL-11/IL-17 when compared to GCP group. The high positivity of P. gingivalis in the dental plaque was associated with significantly increased GCF levels of IL-17 in GCP and GAgP patients. CONCLUSIONS: The increased IL-17 level in GCF of GAgP suggests a potential role in the aetiopathogenesis. Meanwhile, the decreased ratio of IL-11/IL-17 might reflect an imbalance between the proinflammatory and anti-inflammatory cytokines in different periodontal diseases.

Validation of a stability-indicating RP-LC method for the assessment of recombinant human interleukin-11 and its correlation with bioassay.

A stability-indicating reversed-phase liquid chromatography (RP-LC) method was validated for the assessment of recombinant human interleukin-11 (rhIL-11), based on the ICH guidelines. The method was carried out on a Jupiter C(4) column (250 mm × 4.6 mm i.d.), maintained at 25°C. The mobile phase A consisted of 0.1% trifluoroacetic acid (TFA) and the mobile phase B was acetonitrile with 0.1% TFA, run at a flow rate of 1 mL/min, and using a photodiode array (PDA) detection at 214 nm. Separation was obtained with a retention time of 27.6 min, and was linear over the concentration range of 1-200 µg/mL (r(2) = 0.9995). Specificity was established in degradation studies, which also showed that there was no interference of the excipients. The accuracy was 100.22% with bias lower than 1.25%. Moreover, the in vitro cytotoxicity test of the degraded products showed non-significant differences (p > 0.05). The method was applied to the assessment of rhIL-11 and related proteins in biopharmaceutical dosage forms, and the results were correlated to those of a bioassay.

The gingival crevicular fluid levels of interleukin-11 and interleukin-17 in patients with aggressive periodontitis.

BACKGROUND: The balance (ratio) of anti-inflammatory and proinflammatory cytokines is thought to play an important role in the pathogenesis of chronic periodontitis. Moreover, the imbalance of anti-inflammatory/proinflammatory cytokines may modulate disease progression in aggressive periodontitis (AgP). This study aims to investigate the levels of interleukin (IL)-11 and IL-17 and their ratio in gingival crevicular fluid (GCF) in patients with AgP. METHODS: This study included 20 patients with generalized AgP (GAgP) and 18 healthy controls (HC). For each patient, the values of clinical parameters, such as gingival index, plaque index, probing depth, and clinical attachment level, were recorded. Levels of IL-11 and IL-17 in GCF samples were evaluated using enzyme-linked immunosorbent assay. The values of clinical parameters, cytokine levels, and the ratios of cytokines were evaluated. RESULTS: The values of all the clinical parameters were significantly higher in the GAgP group than in the HC group (P < 0.001). The total amount and concentration of IL-11 and the concentration of the IL-17 and IL-11/IL-17 ratio were significantly lower in the GAgP group than in the HC group (P < 0.001). The total amount of IL-17 was not significantly different between the groups (P = 0.317). CONCLUSIONS: The IL-11/IL-17 ratio was decreased in the GAgP group because of the decreased IL-11 levels. The IL-11/IL-17 axis and the link between IL-17 and neutrophil function disorders in AgP should be investigated to clarify the role of the IL-11/IL-17 axis and its balance and imbalance in the pathogenesis of AgP.

Gingival crevicular fluid and plasma acute-phase cytokine levels in different periodontal diseases.

BACKGROUND: The aim of the present study is to investigate gingival crevicular fluid (GCF) and plasma acute-phase cytokines, interleukin-1ß (IL-1ß), interleukin-6 (IL-6), interleukin-11 (IL-11), oncostatin M (OSM), and leukemia inhibitory factor (LIF) levels in patients with different periodontal diseases. METHODS: Eighty individuals were included in this study; 20 with chronic periodontitis (CP), 20 with generalized aggressive periodontitis (GAgP), 20 with gingivitis, and 20 classified as healthy (H). Probing depth, clinical attachment level, plaque index, and papilla bleeding index were recorded. Plasma and GCF IL-1ß, IL-6, IL-11, OSM, and LIF levels were analyzed by enzyme-linked immunosorbent assay. RESULTS: CP and GAgP groups had significantly higher GCF IL-1ß, IL-6, and IL-11 levels when compared with the H group (P <0.05). Conversely, GCF LIF levels of the CP and GAgP groups were lower than those of the H group (P <0.05). GCF OSM levels did not differ significantly among study groups. Plasma levels of all the cytokines studied were not significantly different among the study groups. CONCLUSIONS: Based on the present data, elevated IL-1ß, IL-6, and IL-11 GCF levels, but not plasma levels, are suggested as reliable inflammatory biomarkers in periodontal diseases. Decreased LIF levels in diseased groups might reflect the possible beneficial effects of LIF in the modulation of inflammatory response in gingiva.

Expression of osteoclastogenesis inducers in a tissue model of periodontal ligament under compression.

UNLABELLED: There is increasing interest in the development of new in vitro tissue models. In this study, a tissue model of periodontal ligament (PDL) was established by 3-D-culturing human PDL cells in a thin sheet of porous poly (lactic-co-glycolic acid) scaffold. Growth of the model was evidenced by MTT assay and various microscopies. After being subjected to static compression of 5 ~ 35 g/cm(2) for 6 hrs, the RANKL mRNA expression was significantly up-regulated by force ≥ 25 g/cm(2) in the model. After being subjected to static compression of 25 g/cm(2) for 6 ~ 72 hrs, the mRNA expression of PTHrP, IL-11, IL-8, and FGF-2, potential osteoclastogenesis inducers, was significantly up-regulated in the model, which was further verified by the compression of human PDL in vivo. However, when human gingival fibroblasts were substituted for PDL cells in the model, almost no osteoclastogenesis inducers were up-regulated by compression. This tissue model can serve as an effective tool for the study of PDL mechanoresponse. ABBREVIATIONS: periodontal ligament, PDL; periodontal ligament cells, PDLCs; poly (lactic-co-glycolic acid), PLGA; orthodontic tooth movement, OTM; extracellular matrix, ECM.

The impact of the IL-11:IL-17 ratio on the chronic periodontitis pathogenesis: a preliminary report.

OBJECTIVE: An imbalance in the pro- and anti-inflammatory cytokines may be responsible for periodontal breakdown through immune responses. This study aimed to determine the total amount, concentration and ratio of interleukin (IL)-11 and IL-17 in gingival crevicular fluid (GCF) of chronic periodontitis (CP) patients. MATERIALS AND METHODS: Forty CP patients and 20 healthy controls (C) were included. The CP group was divided into two subgroups in line with the probing depth (PD) in GCF-sampling sites (CPa: PD >or= 5 mm, CPb: PD

Profiling of inflammatory cytokines produced by gingival fibroblasts after human cytomegalovirus infection.

INTRODUCTION: The purpose of this study was to determine the profile of inflammatory cytokines that are produced after in vitro infection of gingival fibroblasts with human cytomegalovirus (HCMV). MATERIALS AND METHODS: Gingival fibroblasts were infected with the Towne strain of HCMV and the cytokine profile in the supernatant was studied using a human inflammation antibody array. Expression of messenger RNA (mRNA) using reverse transcription-polymerase chain reaction for interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) was also analyzed in infected gingival fibroblasts and gingival specimens from subjects with and without periodontitis according to HCMV detection. HCMV was determined in subgingival samples by nested polymerase chain reaction. RESULTS: Gingival fibroblasts produced mainly IL-1alpha, IL-12p40, IL-12p70, IL-6, TNF-alpha, and IL-1beta after HCMV infection. Expression of mRNA for IL-1beta and TNF-alpha was increased after HCMV infection. Production of IL-1beta and TNF-alpha was increased in HCMV-positive periodontitis specimens. In addition, infected gingival fibroblasts produced more IL-8, monocyte chemoattractant protein 1, macrophage inflammatory proteins 1alpha, and 1beta over time postinfection in comparison to baseline. The lowest production of all cytokines studied corresponded to IL-2, IL-4, IL-13, and interferon-gamma. A decreasing production pattern was observed for granulocyte-macrophage colony-stimulating factor, IL-7, and IL-17 while IL-11 and macrophage colony-stimulating factor were increased at 72 h postinfection. CONCLUSIONS: HCMV infection in gingival fibroblasts upregulated the production of proinflammatory-related cytokines and chemokines. The expression of IL-1beta and TNF-alpha was increased both in vitro and in specimens from HCMV-positive subjects with periodontitis. The overproduction of proinflammatory cytokines and chemokines as a result of viral infection should be considered an important pathogenic mechanism linking HCMV to periodontitis in vivo.

Interleukin-11, interleukin-1beta, interleukin-12 and the pathogenesis of inflammatory periodontal diseases.

OBJECTIVES: The balance between pro-inflammatory and anti-inflammatory cytokines may be crucial for determining the immunopathology of gingivitis (G) and periodontitis. This study aimed to analyse interleukin-1beta (IL-1beta), IL-11 and IL-12 levels in gingival crevicular fluid (GCF) of patients with G and chronic periodontitis (CP). MATERIAL AND METHODS: Fourty subjects including 12 CP, 14 G and 14 controls (C) were enrolled. GCF samples were collected from six maxillary sites per patient and analysed for IL-1beta, IL-11 and IL-12 by an enzyme-linked immunosorbent assay. RESULTS: Significantly lower concentrations of IL-11 were detected in CP compared with both G and C groups (p<0.05). The CP group had a significantly higher total amount of IL-12 and IL-1beta compared with the C group (p<0.05). The IL-11:IL-1beta cytokine ratio was higher in both G and C groups compared with the CP group. The IL-11:IL-1beta ratio became progressively lower with increasing probing depth (p<0.01). CONCLUSIONS: Our data showed that IL-11 levels are significantly decreased in GCF from sites with periodontitis compared with G and healthy sites. Because of the possible preventive effect of IL-11 on inflammation, IL-11 may be an important factor in the therapeutic modulation of periodontal disease.

Intrathecal levels of IL-6, IL-11 and LIF in Alzheimer's disease and frontotemporal lobar degeneration.

Cerebrospinal fluid (CSF) levels of interleukin (IL)-6, IL-11 and leukaemia inhibitory factor (LIF) were evaluated in 43 patients with Alzheimer's disease (AD) and 24 patients with frontotemporal lobar degeneration (FTLD) as compared with 30 agematched controls (CON), and correlated with clinical and demographic data and with CSF biomarkers amyloid beta (A beta)42, total tau and tau phosphorylated at position 181 (P-tau). CSF IL-11 mean levels were significantly increased in AD and FTLD as compared with CON (6.5 +/- 4.6 and 6.6 +/- 5.1 versus 3.1 +/- 3.3 pg/ml, P = 0.009). IL-6 mean levels did not differ between patients and CON (P > 0.05),whereas LIF levels were not detectable in patients or in CON. In AD patients, a significantly positive correlation between MMSE scores and IL-11 CSF concentration was observed (r = 0.344, P = 0.028). No correlations with CSF A beta 42, total tau and P-tau were found. IL-11, but not IL-6 levels are increased in AD and FTLD, and the highest peaks were observed in patients with a less severe degree of cognitive deterioration, therefore suggesting a role of this cytokine in early phases of neurodegeneration.

Increased hyperoxia-induced mortality and acute lung injury in IL-13 null mice.

IL-13 is a critical effector at sites of Th2 inflammation and remodeling. As a result, anti-IL-13-based therapies are being actively developed to treat a variety of diseases and disorders. However, the beneficial effects of endogenous IL-13 in the normal and diseased lung have not been adequately defined. We hypothesized that endogenous IL-13 is an important regulator of oxidant-induced lung injury and inflammation. To test this hypothesis, we compared the effects of 100% O(2) in mice with wild-type and null IL-13 loci. In this study, we demonstrate that hyperoxia significantly augments the expression of the components of the IL-13R, IL-13Ralpha1, and IL-4Ralpha. We also demonstrate that, in the absence of IL-13, hyperoxia-induced tissue inflammation is decreased. In contrast, in the IL-13 null mice, DNA injury, cell death, caspase expression, and activation and mortality are augmented. Interestingly, the levels of the cytoprotective cytokines vascular endothelial cell growth factor, IL-6, and IL-11 were decreased in the bronchoalveolar lavage fluid. These studies demonstrate that the expression of the IL-13R is augmented and that the endogenous IL-13-IL-13R pathway contributes to the induction of inflammation and the inhibition of injury in hyperoxic acute lung injury.

Expression of interleukin-11 (IL-11) and IL-11 receptor alpha in human gastric carcinoma and IL-11 upregulates the invasive activity of human gastric carcinoma cells.

Previous investigations have shown that interleukin-11 (IL-11) and the IL-11 receptor (IL-11R) have been correlated with the regulation of tumor progression, cellular growth and differentiation in several malignant tumors. The objectives of this study were to clarify the role of IL-11 and IL-11Ralpha in human gastric carcinoma. IL-11 and IL-11Ralpha were studied in 73 cases of surgically resected human gastric adenocarcinomas by immunohistochemistry. The invasive activity and cell signaling pathway of gastric carcinoma cell lines were also examined. Among the 73 cases of adenocarcinoma, 53 (72.6%) and 47 cases (64.4%) showed positive staining in carcinoma cells for the IL-11 and IL-11Ralpha proteins, respectively. Histologically, IL-11 expression correlated only with Lauren's classification (p<0.05). The expression of IL-11Ralpha correlated with the grade of tumor invasion (p<0.05) and vessel infiltration (p<0.01). All of the four gastric carcinoma cell lines expressed both IL-11 and IL-11Ralpha proteins in western blot analysis. Recombinant human IL-11 (rhIL-11) promoted the migration of SCH cells by the activation of the phosphatidylinositol-3 kinase pathway. Wortmannin diminished the promotion of chemotactic motility and invasive activity by rhIL-11. These findings suggest that the IL-11/IL-11R pathway plays an important role in the progression and the differentiation of human gastric carcinomas.

Development of an in vitro potency assay for therapeutic TGFbeta antagonists: the A549 cell bioassay.

A bioassay was developed to assess the potency of TGFbeta antagonists by measuring IL-11 production in TGFbeta-1 treated human lung epithelial cells (A549). The production of IL-11 by A549 cells, measured by ELISA, was shown to be proportional to the TGFbeta-1 concentration. The A549 cells were responsive to all three isoforms of TGFbeta in the range of 3.0 ng/mL to 1.4 pg/mL, with an 18 to 24 h exposure time found to be within the linear portion of the bioassay response range. The Effective Dose at 80% of the maximal response (ED80) of TGFbeta-1 determined for the assay was 0.3 ng/mL. With this level of TGFbeta-1, a human anti-TGFbeta-1 antibody (CAT-192) yielded an approximate median Inhibitory Concentration (IC50) value of 3 microg/mL. To investigate assay specificity, alternate members of the TGFbeta superfamily were evaluated. Recombinant human activin B, inhibin A and BMP-2 (Bone Morphogenetic Protein-2) did not elicit a significant IL-11 response from the A549 cell line. Bioassay qualification was performed to obtain estimates of precision and accuracy, as well as to establish plate validity criteria. Assay precision was estimated at 30%, while the accuracy was +/-13%. Additionally, the ability of the A549 cell potency assay to detect potency differences in structurally modified samples was investigated.