Induction of eotaxin expression and release from human airway smooth muscle cells by IL-1beta and TNFalpha: effects of IL-10 and corticosteroids.

Eotaxin is a novel C-C chemokine with selective chemoattractant activity for eosinophils. We determined whether eotaxin could be produced by human airway smooth muscle (HASM) cells in culture and examined its regulation by interleukin-10 (IL-10) and the corticosteroid, dexamethasone. Stimulation of the cells with interleukin-1beta (IL-1beta) or tumour necrosis factor (TNFalpha) each at 10 ng ml(-1) induced the release of eotaxin protein with maximal accumulation by 24 h. Interferon-gamma (IFNgamma) alone at 10 ng ml(-1) had no effect and there was no synergy between these cytokines on the release of eotaxin. Reverse phase high performance liquid chromatographic (HPLC) analysis of supernatents from cells treated with TNFalpha (10 ng ml(-1) for 96 h showed immunoreactivity to eotaxin which eluted with the expected retention time of 34.5-35 min. Both IL-1beta and TNFalpha-induced release of eotaxin was not inhibited by dexamethasone (1 microM), however IL-10 (10 ng ml(-1)) had a significant inhibitory effect. Dexamethasone and IL-10 did not inhibit the induction of eotaxin mRNA induced by IL-1beta or TNFalpha. Thus, human airway smooth muscle cells can release eotaxin and could be an important source of chemokine production during airway inflammatory events.

TNF-alpha, unlike other pro- and anti-inflammatory cytokines, induces rapid release of the IL-1 type II decoy receptor in human myelomonocytic cells.

The present study was designed to investigate the effect of a series of cytokines on the release of the type II IL-1 decoy receptor, which represents a unique pathway of negative regulation of the IL-1 system. After 20 min, IL-1, IL-2, IL-4, IL-6, IL-10, IL-12, IL-13, IFN-gamma, granulocyte-CSF, macrophage-CSF, and TGF-beta had little or no effect on IL-1 binding by human polymorphonuclear cells. In contrast granulocyte-macrophage-CSF and, to a greater extent, TNF markedly reduced IL-1 binding. The action of TNF was rapid, reaching 50% of its maximum (80%) at 2 min, and plateauing at 20 min with decrease in receptor number and no significant change in affinity. Loss of surface receptor was associated to rapid release of a 45-kDa IL-1-binding molecule identified as the decoy RII. TNF-induced release of the decoy RII was independent of protein synthesis and reactive oxygen intermediates. Monocytes showed a similar response to TNF, except for the size of the released molecule (approximately 60 kDa). TNF induced rapid release of its own receptors. In contrast IL-1beta affected neither its own receptors nor the TNF-R. TNF and, more efficiently, PMA caused release of the decoy RII in fibroblasts transfected with the full-length decoy RII or with a cytoplasmatic deletion mutant. TNF-induced decoy RII release represents an unidirectional pathway of communication in the interplay between the IL-1 and TNF system.

Detection of circulating prostate-specific antigen-positive cells in patients with prostate cancer by flow cytometry and reverse transcription polymerase chain reaction.

The presence of prostate-specific antigen (PSA)-positive cells has previously been demonstrated in the peripheral blood of prostate cancer patients by flow cytometry (FC), but the identity of these cells has not been established. In this study, the reverse transcriptase polymerase chain reaction (RT-PCR) was compared with analytical FC in an attempt to detect and characterise these cells. Peripheral blood was obtained from 12 patients with newly diagnosed and untreated prostate cancer and five controls. Nine of the 12 patients with prostate cancer (75%) had circulating PSA-positive cells as shown by FC. Only one of those patients (11.1%) was found to express PSA mRNA by RT-PCR. The absence of PSA mRNA in the majority of samples showing PSA-positive cells suggests that they do not represent haematogenous micrometastases. PSA-positive cells in the blood could represent monocytes that express PSA, either following binding/phagocytosis of free serum PSA or phagocytosis of tumour cells.

Reactive oxygen intermediates cause rapid release of the interleukin-1 decoy receptor from human myelomonocytic cells.

Free radicals play an important role in inflammation. We found that reactive oxygen intermediates (ROI) inhibit interleukin-1beta (IL-1beta) binding on human myelomonocytes. Production of superoxide anion (O2-) by Xanthine (X) and Xanthine-Oxidase (XO) or NADPH caused a reduction (48% +/- 15% in 25 experiments) in the IL-1beta binding of polymorphonuclear cells (PMN) and monocytes that was inhibited by superoxide dismutase (SOD). Hydrogen peroxide (H2O2) was only active on monocytes and this effect was prevented by catalase. O2(-)-induced loss of IL-1beta binding on PMN reached half maximum at 5 minutes and peaked after 30 minutes. The reduction of IL-1beta binding was due to reduction of IL-1beta receptors (R) on PMN surface without any change in affinity. ROI-induced reduction of surface IL-1R was not caused by receptor internalization, but rather by the release of a soluble form (45 kD) of the type II decoy R. The action of ROI on IL-1 binding was selective because major histocompatibility complex class I, CD18 and CD16 were unaffected. The O2(-)-induced release of IL-1 decoy R was not affected by protein synthesis inhibitors, but was partially blocked by protease inhibitors. Release of the IL-1 type II decoy R might represent one mechanism by which ROI antagonize and limit the proinflammatory effects of IL-1.

Chemoattractants induce rapid release of the interleukin 1 type II decoy receptor in human polymorphonuclear cells.

Molecules representative of different classes of chemotactic agents, including formyl-Met-Leu-Phe (FMLP), C5a, leukotriene B4, platelet-activating factor, and interleukin (IL)-8, caused a rapid reduction in the IL-1 binding capacity by human polymorphonuclear leukocytes (PMN), a cell type expressing predominantly the IL-1 type II decoy receptor (IL-1 decoy RII). N-t-Boc-Met-Leu-Phe, an antagonist for the FMLP receptor, inhibited the loss of IL-1 binding capacity induced by FMLP. Monocyte chemotactic protein 1, a chemokine related to IL-8 but inactive on PMN, had no effect on IL-1 binding in this cell type. FMLP was selected for further detailed analysis of chemoattractant-induced loss of IL-1 binding by PMN. The action of FMLP was rapid, reaching 50% of its maximum (80%) at 30 s, the earliest measurable time point, and plateauing between 10 and 30 min. Dose-response analysis revealed that maximal reduction of IL-1 binding was reached at FMLP concentrations that were also optimal for chemotaxis (50% effective dose = 5 x 10(-9) M). The loss of IL-1 binding capacity caused by FMLP was determined by a reduction in receptor number with no change in their affinity. The effect of FMLP on IL-1 receptor (IL-1R) was selective in that the PMN surface structures IL-8R, CD16, CD18, and major histocompatibility complex class I antigens were unaffected under these conditions. Loss of surface IL-1R was not due to an augumented rate of internalization. FMLP caused rapid release of a 45-kD IL-1-binding molecule identified as the IL-1 decoy RII. After FMLP-induced release, PMN reexpressed newly synthesized receptors, reaching basal levels by 4 h. FMLP-induced release of the IL-1 decoy RII did not impair the responsiveness of PMN to IL-1 in terms of promotion of survival and cytokine production. FMLP-induced release of the IL-1 decoy RII was unaffected by protein synthesis inhibitors, was blocked by certain protease inhibitors, and was mimicked by agents (the Ca++ ionophore A23187 and phorbol myristate acetate) that recapitulate elements in the signal transduction pathway of chemoattractant receptors. The time frame and concentration range of chemoattractant-induced rapid release of the IL-1 decoy RII are consistent with the view that IL-1 decoy RII release is an early event in the multistep process of leukocyte recruitment.(ABSTRACT TRUNCATED AT 400 WORDS)

Matrix metalloproteinase 9 expression in primary human prostatic adenocarcinoma and benign prostatic hyperplasia.

Matrix metalloproteinase (MMP) expression was investigated in patients with prostatic adenocarcinoma and benign prostatic hyperplasia (BPH). Forty-one men were studied: 26 had histologically proven prostate cancer, with 14 (54%) showing metastatic disease; 15 patients had BPH. Prostatic tissue was obtained from transurethral resection and needle core biopsies; gelatinolytic activity was determined by zymography. Seven gelatinolytic bands were detected, with molecular weights ranging from > 100 kilodalton (kDa) to 29 kDa. Nine of 14 patients (64%) with skeletal metastases had 92 kDa activity, present in only two of 12 patients (17%) with a negative bone scan, and absent in BPH. The 92 kDa gelatinolytic activity was expressed in 73% of aneuploid tumours compared with 20% of diploid tumours. A 97 kDa gelatinase was expressed in 80% of BPH samples and 23% of carcinoma patients. Enzyme bands of 72, 66 and 45 kDa were equally expressed in malignant tissue, irrespective of metastatic status, but were expressed in fewer BPH patients. The 97, 92, 66 and 45 kDa enzymes were identified as being pro-MMP-9 sequences by Western blotting, using a specific antibody directed against the pro sequence of the mature protein. MMP activity appeared to be increased in malignant prostatic tissue compared with BPH. Pro-MMP-9, in its 92 kDa form, was shown to be exclusively expressed by malignant prostatic tissue, and in particular by tumours that exhibited the aggressive and metastatic phenotype.