Increased Interleukin-8 Research Articles

Lysophospholipids Increase Interleukin-8 Expression in Ovarian Cancer Cells

Objectives. We have previously described that bioactive lysophospholipids—lysophosphatidic acid (LPA), sphingosine 1-phosphate (S1P), and sphingosylphosphorylcholine (SPC)—are present in ascitic fluids from patients with ovarian cancer. To understand the role of these lipids in ovarian cancer, we investigated the effects of these lipids on interleukin-8 (IL-8) production in ovarian cancer cells. IL-8 is a proinflammatory and proangiogenic factor, which is potentially involved in ovarian cancer development.Methods. The Clontech PCR-Select cDNA subtraction method (Clontech Laboratories, Inc., Palo Alto, CA) was used to identify genes potentially regulated by LPA in HEY and OCC1 ovarian cancer cell lines. Northern blot analysis was used to confirm and examine IL-8 mRNA regulation by lysolipids. Enzyme-linked immunosorbent assay (ELISA) was used for detecting secreted IL-8.Results. We describe here that LPA, S1P, and SPC increased mRNA levels (2- to 7-fold) and protein secretion (2- to 12-fold) of IL-8 from ovarian cancer cells (HEY, OCC1, and SKOV3) in vitro. These regulations were both dose- and time-dependent. All three lipids increased the stability IL-8 mRNA in HEY cells. In contrast to malignant ovarian cancer cells, immortalized human ovarian epithelial cells did not respond to any of these lipids to increase the secretion of IL-8, although these cells secreted similar basal levels of IL-8 (310 pg/ml/10,000 cells). Two breast cancer cell lines (MCF7 and T47D) secreted lower basal levels of IL-8 (48-80 pg/ml/10,000 cells), compared with ovarian cancer cells (200–500 pg/ml/10,000 cells). MCF7 cells responded to LPA, but not S1P and SPC, by increasing the secretion of IL-8. T47D and MCF10A, an immortalized breast cell line, did not respond to LPA, S1P, or SPC to increase IL-8 secretion.Conclusions. LPA, S1P, and SPC regulate the mRNA and protein levels of the proinflammatory and proangiogenic factor IL-8 in ovarian cancer cells. The pathological significance of these regulations in ovarian cancer remains to be further investigated.

Increased interleukin-8 (IL-8) expression is related to aseptic loosening of total hip replacement.

Aseptic loosening is an increasing problem in total hip replacement (THR). Chronic inflammatory reaction against implant wear particle results in collageno- and osteolysis, leading to loosening of the implant. Cytokines are known to play a major role in this particular inflammatory process. The aim of the present study was to examine interleukin-8 (IL-8) in the synovial-like interface membrane (SLIM) and pseudocapsular tissue of THRs and to compare it to normal knee synovial membrane. Eleven patients suffering from aseptically loosened THRs were included. All the SLIM and pseudocapsular tissue samples were obtained during revision operations. Ten control samples of normal synovium were collected per arthroscopy from the superior recessus of the knee. For immunohistochemical IL-8 detection, polyclonal mouse anti-human immunoglobulin (Ig)G1 IL-8-primary antibody was used with the alkaline phosphatase anti-alkaline phosphatase (APAAP) method. Results were quantitated using the Vidas image analysis system. The highest count levels (mean +/- SEM) were detected in SLIM tissue (386+/-82 cells/mm2). The difference was statistically significant compared with pseudocapsular tissue (193+/-36 cells/mm2) and control samples (18+/-5 cells/mm2). Count levels in control tissue were on average 5% of the SLIM tissues values. The present study determines for the first time the cellular origin of IL-8 in aseptically loosened THRs and also quantitates the IL-8-producing cells in the periprosthetic tissue. The results reveal a high rise in IL-8 concentration in SLIM and in synovial tissues. This finding moves us one step forward in solving the complex network of multiple factors affecting loosening of hip implants.

Increased interleukin-8 (IL-8) in rectal dialysate from patients with ulcerative colitis: evidence for a biological role for IL-8 in inflammation of the colon

OBJECTIVE: Infiltration of neutrophils and their release of toxic reactive oxygen species (ROS) in the colonic mucosa are associated with tissue damage in ulcerative colitis (UC). This neutrophil migration may be induced by chemoattractants, such as cytokines, in the colonic milieu. One such chemoattractant is interleukin-8 (IL-8), a neutrophil chemokine that is present at high concentrations in inflamed mucosa. However, the functional significance of IL-8 in neutrophil attraction and activation in UC has not been established. We hypothesized that IL-8 in the colonic lumen of patients with UC primes neutrophils, leading to their attraction and activation. METHODS: The colonic milieu was sampled by rectal dialysis. Using a semi-permeable membrane with a molecular weight cut-off of 12 kDa, dialysis solution was placed in the rectum and allowed to equilibrate over a 4-h period with the colonic milieu of controls or of patients with UC. IL-8 concentrations were measured by ELISA. Two functions of healthy neutrophils (PMN) were measured: expression of CD11-b surface adhesion molecules (by flow cytometry), and production of ROS (by both chemiluminescence and cytochrome C reduction assays). Neutrophil functions after exposure to rectal dialysates or n-formyl-methionyl-leucyl-phenylalanine (fMLP) were assessed before and after adding anti–IL-8 antibody or the fMLP blocker BMLP. RESULTS: IL-8 concentrations in dialysates from patients with active UC were significantly higher than in controls and correlated with disease activity. UC dialysates significantly increased ROS production and CD11-b expression by neutrophils and anti–IL-8 antibody partially (50%) inhibited these stimulatory effects of UC dialysates. Preincubation of neutrophils with UC dialysates significantly potentiated the fMLP-induced rise in ROS and anti–IL-8 antibody completely abolished this priming effect. CONCLUSIONS: The colonic milieu, sampled by rectal dialysis, from patients with active UC can both activate and prime neutrophils in vitro. High concentrations of IL-8 in the colonic lumen of UC patients are partially responsible for the activating effects of rectal dialysates, and account for all of its priming effects. These findings provide direct evidence for a role for IL-8 in inflammatory bowel disease.

Increased interleukin-8 (IL-8) in rectal dialysate from patients with ulcerative colitis: evidence for a biological role for IL-8 in inflammation of the colon.

Infiltration of neutrophils and their release of toxic reactive oxygen species (ROS) in the colonic mucosa are associated with tissue damage in ulcerative colitis (UC). This neutrophil migration may be induced by chemoattractants, such as cytokines, in the colonic milieu. One such chemoattractant is interleukin-8 (IL-8), a neutrophil chemokine that is present at high concentrations in inflamed mucosa. However, the functional significance of IL-8 in neutrophil attraction and activation in UC has not been established. We hypothesized that IL-8 in the colonic lumen of patients with UC primes neutrophils, leading to their attraction and activation. The colonic milieu was sampled by rectal dialysis. Using a semi-permeable membrane with a molecular weight cut-off of 12 kDa, dialysis solution was placed in the rectum and allowed to equilibrate over a 4-h period with the colonic milieu of controls or of patients with UC. IL-8 concentrations were measured by ELISA. Two functions of healthy neutrophils (PMN) were measured: expression of CD11-b surface adhesion molecules (by flow cytometry), and production of ROS (by both chemiluminescence and cytochrome C reduction assays). Neutrophil functions after exposure to rectal dialysates or n-formyl-methionyl-leucyl-phenylalanine (fMLP) were assessed before and after adding anti-IL-8 antibody or the fMLP blocker BMLP. IL-8 concentrations in dialysates from patients with active UC were significantly higher than in controls and correlated with disease activity. UC dialysates significantly increased ROS production and CD11-b expression by neutrophils and anti-IL-8 antibody partially (50%) inhibited these stimulatory effects of UC dialysates. Preincubation of neutrophils with UC dialysates significantly potentiated the fMLP-induced rise in ROS and anti-IL-8 antibody completely abolished this priming effect. The colonic milieu, sampled by rectal dialysis, from patients with active UC can both activate and prime neutrophils in vitro. High concentrations of IL-8 in the colonic lumen of UC patients are partially responsible for the activating effects of rectal dialysates, and account for all of its priming effects. These findings provide direct evidence for a role for IL-8 in inflammatory bowel disease.

Effect of linoleic acid on endothelial cell inflammatory mediators

Selected lipids may influence the inflammatory cascade within the vascular endothelium. To test this hypothesis, endothelial cells were treated with linoleic acid (18:2, n − 6) for 12 hours and/or tumor necrosis factor-α (TNF) for 4 hours. For a combined exposure to 18:2 and TNF (18:2 + TNF), cells were first preenriched with 18:2 for 8 hours before exposure to TNF for an additional 4 hours. Exposure to 18:2 increased cellular oxidative stress, activated nuclear factor-κB (NF-κB), increased interleukin-8 (IL-8) production, and elevated intercellular adhesion molecule-1 (ICAM-1) levels. A combined exposure to 18:2 + TNF resulted in decreased NF-κB activation compared with TNF treatment alone. In addition, preexposure to 18:2 altered TNF-mediated IκB-α signaling. Within the first 15 minutes of a 90-minute period, cytoplasmic levels of IκB-α decreased more rapidly in cells treated with 18:2 + TNF compared with TNF, suggesting translocation and activation of NF-κB in cultures that were pretreated with 18:2 before TNF exposure. A combined exposure to 18:2 + TNF had various effects on IL-8 production and ICAM-1 levels depending on the time of exposure. For example, 18:2 + TNF treatment increased ICAM-1 levels at 12 hours but decreased ICAM-1 levels at 24 hours compared with treatment with TNF alone. These data suggest that selected fatty acids such as 18:2 can exert proinflammatory effects and, in addition, may markedly alter TNF-mediated inflammatory events.

Elevation of interleukin-8 and interleukin-6 precedes the influx of neutrophils in tracheal aspirates from preterm infants who develop bronchopulmonary dysplasia.

The influx of inflammatory mediators and cells into the tracheobronchial effluent of preterm infants with respiratory distress syndrome (RDS) appears to be important in signaling the development of bronchopulmonary dysplasia (BPD). The mechanism that initiates this early inflammatory response is not well understood. The purpose of this study was to test the hypothesis whether increased interleukin-8 (IL-8), a potent chemoattractant for human neutrophils, appears in the airways of preterm infants with RDS in whom BPD develops before the influx of neutrophils. In addition, airway secretions were analyzed for the cytokine interleukin-6 (IL-6) to test the hypothesis whether this pro-inflammatory cytokine is an early marker of inflammation in preterm infants with RDS who progress to BPD. Sixty-five infants less than 32 weeks gestation with RDS were enrolled on the first day of life and 56 infants completed the study, with 31 recovering from RDS (Non-BPD) and 25 infants progressing to BPD. Infants were excluded from enrollment in the presence of maternal chorioamnionitis, infection at birth, or infection within the first week of life. There were no significant differences in birthweight, gestational age, or prolonged rupture of membranes between the two groups. Serial tracheal aspirates (TA) were collected on days 1, 3, 5, and 7 while the infants remained intubated. Significant elevations of TA neutrophil counts were detected in the BPD group on days 5 and 7. Cell-free TA revealed marked elevations of IL-8 in the BPD group compared to the Non-BPD group [median (25th percentile, 75th percentile), ng/ml epithelial lining fluid (ELF)] on day 1 [BPD 485 (195, 840); Non-BPD 63.1 (28.3, 197), P < 0.05] and day 3 [BPD 740 (319, 1310); Non-BPD 111 (54.3, 337); P < 0.05], while on days 5 and 7, the differences were not statistically significant. Interleukin-6 (IL-6) was measured as a marker of acute inflammation and was not different in the two groups on day 1, but was significantly elevated on day 3 [median (25th percentile, 75th percentile), ng/ml ELF; BPD 297 (62.1, 702); Non-BPD 72 (32.8, 266), P < 0.05] and on day 5 [BPD 270 (136, 672); Non-BPD 86.4 (57.8, 138), P < 0.05]. These studies demonstrate that elevation of IL-8 and IL-6 levels precedes the marked neutrophil influx seen in the TA of preterm infants in whom BPD develop. The presence of IL-8 and IL-6 in TA from these infants suggests that these cytokines either initiate the acute inflammatory cascade in the lungs, or they are early markers of the inflammatory process that places preterm infants at high risk for BPD.

Comparative Responsiveness of HL-60, HL-60R, and HL-60R+ (LRARSN) Cells to Retinoic Acid, Calcitriol, 9 cis-Retinoic Acid, and Sodium Butyrate

In HL-60 cells, retinoic acid (RA) and 9 cis-RA induce granulocytic differentiation, and calcitriol and sodium butyrate induce monocytic differentiation. To study the role of retinoid resistance on the response to these agents, we investigated their effects in HL-60 cells, retinoid-resistant HL-60R cells, and HL-60R+ cells in which retinoid sensitivity has been restored. In HL-60 cells, cathepsin D (ctsd) mRNA levels are increased by these agents and by cholera toxin after pretreatment with each agent. Calcitriol, 9 cis-RA, and sodium butyrate increase interleukin-8 (IL-8) mRNA expression, and pretreatment with these agents or RA potentiates the stimulation of IL-8 by phorbol ester (TPA). Pretreatment of HL-60 cells with all of the agents confers inducibility of cathepsin L (ctsl) mRNA by TPA in previously unresponsive cells. In HL-60R cells, none of the agents alone or in combination significantly enhances the expression of the ctsd, IL-8, or ctsl mRNAs. Retinoid stimulation (either alone or in combination with the other agents) of the three mRNAs is partially restored in the HL-60R+ cells. Calcitriol does not alter the expression of any of these mRNAs, and only the stimulation of IL-8 mRNA by sodium butyrate is recovered. Treatment with all of the agents inhibits proliferation and stimulates differentiation of the HL-60 cells. RA and calcitriol are unable to inhibit proliferation of the HL-60R cells, whereas only calcitriol fails to inhibit proliferation of the HL-60R+ cells. None of the agents induces differentiation in either the HL-60R or HL-60R+ cells. Therefore, the mutation of the RA receptor alpha is insufficient to account for the altered responses of the HL-60R cells, and there are likely defects in other signaling pathways in these cells. These cells may prove useful in examining the mechanism of cross-resistance between various differentiating agents.

Comparative responsiveness of HL-60, HL-60R, and HL-60R+ (LRARSN) cells to retinoic acid, calcitriol, 9 cis-retinoic acid, and sodium butyrate

In HL-60 cells, retinoic acid (RA) and 9 cis-RA induce granulocytic differentiation, and calcitriol and sodium butyrate induce monocytic differentiation. To study the role of retinoid resistance on the response to these agents, we investigated their effects in HL-60 cells, retinoid-resistant HL-60R cells, and HL-60R+ cells in which retinoid sensitivity has been restored. In HL-60 cells, cathepsin D (ctsd) mRNA levels are increased by these agents and by cholera toxin after pretreatment with each agent. Calcitriol, 9 cis-RA, and sodium butyrate increase interleukin-8 (IL-8) mRNA expression, and pretreatment with these agents or RA potentiates the stimulation of IL-8 by phorbol ester (TPA). Pretreatment of HL-60 cells with all of the agents confers inducibility of cathepsin L (ctsl) mRNA by TPA in previously unresponsive cells. In HL-60R cells, none of the agents alone or in combination significantly enhances the expression of the ctsd, IL-8, or ctsl mRNAs. Retinoid stimulation (either alone or in combination with the other agents) of the three mRNAs is partially restored in the HL-60R+ cells. Calcitriol does not alter the expression of any of these mRNAs, and only the stimulation of IL-8 mRNA by sodium butyrate is recovered. Treatment with all of the agents inhibits proliferation and stimulates differentiation of the HL-60 cells. RA and calcitriol are unable to inhibit proliferation of the HL-60R cells, whereas only calcitriol fails to inhibit proliferation of the HL-60R+ cells. None of the agents induces differentiation in either the HL-60R or HL-60R+ cells. Therefore, the mutation of the RA receptor alpha is insufficient to account for the altered responses of the HL-60R cells, and there are likely defects in other signaling pathways in these cells. These cells may prove useful in examining the mechanism of cross-resistance between various differentiating agents.