Interleukin-8 Protein Production Research Articles

Potential of Trilayered Gelatin/Polycaprolactone Nanofibers for Periodontal Regeneration: An In Vitro Study.

Over the past few years, biomaterial-based periodontal tissue engineering has gained popularity. An ideal biomaterial for treating periodontal defects is expected to stimulate periodontal-derived cells, allowing them to contribute most efficiently to tissue reconstruction. The present study focuses on evaluating the in vitro behavior of human periodontal ligament-derived stromal cells (hPDL-MSCs) when cultured on gelatin/Polycaprolactone prototype (GPP) and volume-stable collagen matrix (VSCM). Cells were cultured onto the GPP, VSCM, or tissue culture plate (TCP) for 3, 7, and 14 days. Cell morphology, adhesion, proliferation/viability, the gene expression of Collagen type I, alpha1 (COL1A1), Vascular endothelial growth factor A (VEGF-A), Periostin (POSTN), Cementum protein 1 (CEMP1), Cementum attachment protein (CAP), Interleukin 8 (IL-8) and Osteocalcin (OCN), and the levels of VEGF-A and IL-8 proteins were investigated. hPDL-MSCs attached to both biomaterials exhibited a different morphology compared to TCP. GPP exhibited stronger capabilities in enhancing cell viability and metabolic activity compared to VSCM. In most cases, the expression of all investigated genes, except POSTN, was stimulated by both materials, with GPP having a superior effect on COL1A1 and VEGF-A, and VSCM on OCN. The IL-8 protein production was slightly higher in cells grown on VSCM. GPP also exhibited the ability to absorb VEGF-A protein. The gene expression of POSTN was promoted by GPP and slightly suppressed by VSCM. In summary, our findings indicate that GPP electrospun nanofibers effectively promote the functional performance of PDLSCs in periodontal regeneration, particularly in the periodontal ligament and cementum compartment.

Hypervolemia in Dialysis Patients Impairs STAT3 Signaling and Upregulates miR-142-3p: Effects on IL-10 and IL-6.

Fluid overload in hemodialysis patients (HD) has been proven to be associated with inflammation. Elevated levels of the pro-inflammatory cytokine interleukin-6 (IL-6) appear to be inadequately counterbalanced by the anti-inflammatory cytokine interleukin-10 (IL-10). We initiated a cross-sectional study enrolling 40 HD patients who were categorized by a bioimpedance measurement in normovolemic (N; 23) and hypervolemic (H; 17) groups to test whether IL-10- and IL-6-related signal transduction pathways (signal transducer of transcript 3: STAT3) and/or a post-transcriptional regulating mechanism (miR-142) are impaired by hypervolemia. IL-10/IL-6 transcript and protein production by PBMCs (peripheral blood mononuclear cells) were determined. Phospho-flow cytometry was used to detect the phosphorylated forms of STAT3 (pY705 and pS727). miR-142-3p/5p levels were detected by qPCR. Hypervolemic patients were older, more frequently had diabetes, and showed higher CRP levels. IL-10 transcripts were elevated in H patients but not IL-10 protein levels. In spite of the elevated mRNA expression of the suppressor of cytokine expression 3 (SOCS3), IL-6 mRNA and protein expression were increased in immune cells of H patients. The percentage of cells staining positive for STAT3 (pY705) were comparable in both groups; in STAT3 (pS727), however, the signal needed for full transactivation was decreased in H patients. miR-142-3p, a proven target of IL-10 and IL-6, was significantly elevated in H patients. Insufficient phosphorylation of STAT3 may impair inflammatory and anti-inflammatory cytokine signaling. How far degradative mechanisms induced by elevated miR-142-3p levels contribute to an inefficient anti-inflammatory IL-10 signaling remains elusive.

Pigment epithelium-derived factor is an interleukin-6 antagonist in the RPE: Insight of structure-function relationships.

Retinal and choroidal inflammatory lesions increase the levels of the pro-inflammatory cytokine interleukin-6 (IL-6). Pigment epithelium-derived factor (PEDF) has anti-inflammatory properties, but it is not known if it can prevent the production of IL-6 by the retinal pigment epithelium. To investigate the anti-inflammatory effects of PEDF in the RPE, we used human ARPE-19 cells stimulated with human recombinant tumor necrosis factor-alpha (TNF-α) to induce overexpression of the IL6 gene. We found that the viability of ARPE-19 cells decreased by 22% with TNF-α at 10ng/ml, being drastically decreased at ≥50ng/ml. TNF-α at 5-100ng/ml elevated the production and secretion of IL-6 protein, as measured by ELISA. To challenge the TNF-α-mediated stimulation of IL-6, we used recombinant human PEDF protein. PEDF at 100nM recovered the TNF-α-mediated loss of cell viability and repressed IL-6 gene expression as determined by RT-PCR. PEDF at 10-100nM attenuated the IL-6 protein secretion in a dose dependent fashion (IC50 = 65nM), being abolished with 100nM PEDF. To map the region that confers the IL-6 blocking effect to the PEDF polypeptide, we used chemically synthesized peptides designed from its biologically active domains, pro-death 34-mer, and pro-survival 44-mer and 17-mer (H105A), to challenge the IL-6 overproduction. The pro-survival peptides recovered the TNF-α-mediated cell viability loss, and inhibited IL-6 secretion, while the 34-mer did not have an effect, suggesting a role for the pro-survival domain in blocking TNF-α-mediated cell death and IL-6 stimulation. Our findings position PEDF as a novel antagonistic agent of IL-6 production in RPE cells, underscoring its use for the management of retinal disease-related inflammation.

Fluid shear stress-induced IL-8/CXCR signaling in human ovarian cancer cells

Background: Interleukin-8 (IL-8) released from mechanosensitive cancer cells plays a key role in the inflammation and metastasis of solid carcinomas. In this study, we have explored IL-8 and its receptors signal transduction process of human ovarian cancer cells under conditions of FSS. Methods: After the fluid shear stress was loaded, LightCycler™ system and quantitative sandwich ELISA were employed to assay the IL-8 mRNA expression and protein production, respectively. IL-8 reporter gene pEGFP1-IL8USCS was constructed for determining IL-8 gene transcriptional activation through gene transfer and flow cytometric analysis. NF-κB nuclear translocation was observed by immunocytofluorescent staining. Western blot was used to examine IκB phosphorylation and degradation. RT-PCR, Northern blot and immunofluorescence were used to determine the expression of a cell-surface chemokine receptor CXCR2 at mRNA and protein levels. Results: IL-8 mRNA expression and protein production had biphasic responses to low shear stress (1.5 dyne/cm 2 ), with the peaks at 1 and 2 h respectively. There was an increase in enhanced green fluorescent protein expression in pEGFP1-IL8USCS-transfected SKOV3 cells subjected to a fluid shear stress of 1.5 dyne/cm 2 for 2 h. Following the application of shear stress of 1.5 dyne/cm 2 , NF-κB p65 became detectable in the cell nuclei, and Phosphorylated IκB in cell lysates increased significantly. CXCR2, which was constitutively present on the surface of SKOV3 cells, increased following exposure to fluid shear stress for 60 min. Conclusions: Fluid shear stress triggered IL-8/CXCR2 signaling of SKOV3 cells is an early gene activation, and the activation can be mediated through NF-κB. This observation suggested that fluid shear stress-induced IL-8 activation and the downstream signal pathways may have an important contribution to the pathogenesis and development of both inflammation and metastasis of ovarian carcinomas.

Fluid shear stress-induced IL-8/CXCR signaling in human ovarian cancer cells.

BackgroundInterleukin-8 (IL-8) released from mechanosensitive cancer cells plays a key role in the inflammation and metastasis of solid carcinomas. In this study, we have explored IL-8 and its receptors signal transduction process of human ovarian cancer cells under conditions of FSS.MethodsAfter the fluid shear stress was loaded, LightCycler™ system and quantitative sandwich ELISA were employed to assay the IL-8 mRNA expression and protein production, respectively. IL-8 reporter gene pEGFP1-IL8USCS was constructed for determining IL-8 gene transcriptional activation through gene transfer and flow cytometric analysis. NF-κB nuclear translocation was observed by immunocytofluorescent staining. Western blot was used to examine IκB phosphorylation and degradation. RT-PCR, Northern blot and immunofluorescence were used to determine the expression of a cell-surface chemokine receptor CXCR2 at mRNA and protein levels.ResultsIL-8 mRNA expression and protein production had biphasic responses to low shear stress (1.5 dyne/cm2), with the peaks at 1 and 2 h respectively. There was an increase in enhanced green fluorescent protein expression in pEGFP1-IL8USCS-transfected SKOV3 cells subjected to a fluid shear stress of 1.5 dyne/cm2 for 2 h. Following the application of shear stress of 1.5 dyne/cm2, NF-κB p65 became detectable in the cell nuclei, and Phosphorylated IκB in cell lysates increased significantly. CXCR2, which was constitutively present on the surface of SKOV3 cells, increased following exposure to fluid shear stress for 60 min.ConclusionsFluid shear stress triggered IL-8/CXCR2 signaling of SKOV3 cells is an early gene activation, and the activation can be mediated through NF-κB. This observation suggested that fluid shear stress-induced IL-8 activation and the downstream signal pathways may have an important contribution to the pathogenesis and development of both inflammation and metastasis of ovarian carcinomas.

ZC3H12B/MCPIP2, a new active member of the ZC3H12 family.

ZC3H12B is the most enigmatic member of the ZC3H12 protein family. The founding member of this family, Regnase-1/MCPIP1/ZC3H12A, is a well-known modulator of inflammation and is involved in the degradation of inflammatory mRNAs. In this study, for the first time, we characterized the properties of the ZC3H12B protein. We show that the biological role of ZC3H12B depends on an intact NYN/PIN RNase domain. Using RNA immunoprecipitation, experiments utilizing actinomycin D and ELISA, we show that ZC3H12B binds interleukin-6 (IL-6) mRNA in vivo, regulates its turnover, and results in reduced production of IL-6 protein upon stimulation with IL-1β. We verified that regulation of IL-6 mRNA stability occurs via interaction of ZC3H12B with the stem–loop structure present in the IL-6 3′UTR. The IL-6 transcript is not the only target of ZC3H12B. ZC3H12B also interacts with other known substrates of Regnase-1 and ZC3H12D, such as the 3′UTRs of IER3 and Regnase-1, and binds IER3 mRNA in vivo. Using immunofluorescence, we examined the localization of ZC3H12B within the cell. ZC3H12B forms small, granule-like structures in the cytoplasm that are characteristic of proteins involved in mRNA turnover. The overexpression of ZC3H12B inhibits proliferation by stalling the cell cycle in the G2 phase. This effect of ZC3H12B is also NYN/PIN dependent. The analysis of the ZC3H12B mRNA level reveals its highest expression in the human brain and the neuroblastoma cell line SH-SY5Y, although the factors regulating its expression remain elusive. Down-regulation of ZC3H12B in SH-SY5Y cells by specific shRNAs results in up-regulation of ZC3H12B-target mRNAs.

Intrinsic anti-inflammatory properties in the serum of two species of deep-diving seal.

Weddell and elephant seals are deep-diving mammals, which rely on lung collapse to limit nitrogen absorption and prevent decompression injury. Repeated collapse and re-expansion exposes the lungs to multiple stressors, including ischemia-reperfusion, alveolar shear stress and inflammation. There is no evidence, however, that diving damages pulmonary function in these species. To investigate potential protective strategies in deep-diving seals, we examined the inflammatory response of seal whole blood exposed to lipopolysaccharide (LPS), a potent endotoxin. Interleukin-6 (IL6) cytokine production elicited by LPS exposure was 50 to 500 times lower in blood of healthy northern elephant seals and Weddell seals compared with that of healthy human blood. In contrast to the ∼6× increased production of IL6 protein from LPS-exposed Weddell seal whole blood, isolated Weddell seal peripheral blood mononuclear cells, under standard cell culture conditions using medium supplemented with fetal bovine serum (FBS), produced a robust LPS response (∼300×). Induction of Il6 mRNA expression as well as production of IL6, IL8, IL10, KC-like and TNFα were reduced by substituting FBS with an equivalent amount of autologous seal serum. Weddell seal serum also attenuated the inflammatory response of RAW 267.4 mouse macrophage cells exposed to LPS. Cortisol level and the addition of serum lipids did not impact the cytokine response in cultured cells. These data suggest that seal serum possesses anti-inflammatory properties, which may protect deep divers from naturally occurring inflammatory challenges such as dive-induced hypoxia-reoxygenation and lung collapse.

Activation of the B‐cell receptor successively activates NF‐κB and STAT3 in chronic lymphocytic leukemia cells

In chronic lymphocytic leukemia (CLL) cells, both interleukin-6 (IL-6) and the B-cell receptor (BCR) activate Janus kinase 2 (JAK2) and induce the phosphorylation of signal transduction and activator of transcription 3 (STAT3) on tyrosine 705 residues. However, whereas IL-6 phosphorylates STAT3 within 15 min, stimulation of the BCR with anti-immunoglobulin M (IgM) antibodies phosphorylates STAT3 in 2-4 hr. Here, we show that this process takes longer because it requires transcriptional activity of NF-κB. Using an electromobility shift assay, we found that incubation with IgM antibodies for 4 or 18 hr, but not 15 min, increased NF-κB DNA-binding of CLL cells and increased binding was translated to increased transcriptional activity. Hence, 42% of the 83 NF-κB target genes were constitutively expressed in all CLL cells prior to any inducible stimuli. However, activation of the BCR increased the number of NF-κB target genes with detectable expression by 23%. Remarkably, prolonged incubation with anti-IgM antibodies induced a time-dependent transcription, production and secretion of IL-6 protein. The IgM-induced production of IL-6 prompted the phosphorylation of STAT3 on tyrosine residues. This effect was inhibited by the JAK1/2 inhibitor of the JAK/STAT3 pathway ruxolitinib. Taken together, these results suggest that in CLL cells, constitutive tonic activation of NF-κB can be further enhanced by the BCR and that the BCR-induced activation of the JAK/STAT3 pathway depends on the NF-κB induced production of IL-6.

Hyaluronan carried by tumor-derived microvesicles induces IL-10 production in classical (CD14++CD16−) monocytes via PI3K/Akt/mTOR-dependent signalling pathway

Tumor-derived microvesicles (TMV) can mimic effects of tumor cells leading to an increased anti-inflammatory cytokine production, such as interleukin 10 (IL-10), by tumor-infiltrating monocytes and macrophages. Yet, the mechanism of IL-10 induction by TMV in monocytes remains unclear.The co-incubation of TMV derived from the human pancreas carcinoma cell line (HPC-4) with human monocytes resulted in a nearly 30-fold increase in IL-10 protein production. This effect operates at the level of transcription since monocytes transduced with an adenovirus containing IL-10-promoter luciferase reporter gene showed a 5-fold induction of luciferase activity after treatment with TMV. Since tumor cells can express hyaluronan (HA), which participates in tumor invasion and metastases, we have tested its effect on IL-10 expression. We showed that HA at the concentration of 100μg/ml induces IL-10 protein expression and the IL-10 promoter activation in monocytes. Moreover, hyaluronidase treatment of TMV reduced IL-10 protein production by 50% and promoter activity by 40%. Inhibitors of the PI3K/Akt/mTOR pathway reduced both, TMV-induced IL-10 promoter activity and protein production, and the same was observed in monocytes when stimulated by HPC-4 cells or HA. Inhibition of PI3K activity down-regulated phosphorylation of the Akt and (to a lesser extent) mTOR proteins in monocytes following TMV or HA stimulation. When comparing monocyte subsets, TMV induced IL-10 protein and mRNA synthesis only in classical CD14++CD16− but not in CD16-positive monocytes. Our data show that TMV induce IL-10 synthesis in human classical monocytes via HA, which, in turn, activates the PI3K/Akt/mTOR pathway.

Fungus induces the release of IL-8 in human corneal epithelial cells, via Dectin-1-mediated protein kinase C pathways.

To identify whether Aspergillus fumigatus (A. fumigatus) hyphae antigens induced the release of interleukin-8 (IL-8) in anti-fungal innate immunity of cultured human corneal epithelial cells (HCECs) and determine the involvement of intracellular signalling pathways. HCECs were treated with A. fumigatus hyphae antigens with different concentrations and time. The cytoplasmic calcium of HCECs were assessed by fluorescence imaging. Western blot was used to detect the expression of Ca(2+)-dependent protein kinase C (PKC). The IL-8 levels were determined by specific human IL-8 enzyme-linked immunosorbent assay (ELISA) and reverse transcriptase polymerase chain reaction (RT-PCR). Using a series of pharmacological inhibitors, we examined the upstream signalling pathway responsible for IL-8 expression in response to A. fumigatus hyphae antigens. Cells exposed to A. fumigatus hyphae antigens showed higher level of IL-8 mRNA expression and protein production. We demonstrated here that stimulation of HCECs with A. fumigatus hyphae triggers an intracellular Ca(2+) flux and results in the activation of Ca(2+)-dependent PKC (α, βI and βII) which can be attenuated by pre-treatment of cells with laminarin, suggesting that Dectin-1 signals pathway induced cytoplasmic calcium and influence the activation of PKC in HCECs. Inhibitors of Ca(2+)-dependent PKC (Ro-31-8220 and Go6976) significantly abolished hyphae-induced expression of IL-8. Our findings suggest that A. fumigatus hyphae-induced IL-8 expression was regulated by the activation of Dectin-1-mediated Ca(2+)-dependent PKC in HCECs.

Lipopolysaccharide Attenuates CD40 Ligand-Induced Regulatory B10 Cell Expansion and IL-10 Production in Mouse Splenocytes.

Toll-like receptors (TLRs) play a key role in B cell-mediated innate and adaptive immunity. It has been shown that interleukin 10 (IL-10)-producing regulatory B cells (B10 cells) can negatively regulate cellular immune responses and inflammation in autoimmune diseases. In this study, we determined the effect of TLR4 signaling on the CD40-activated B10 cell competency. The results demonstrated that LPS and CD40L synergistically stimulated proliferation of mouse splenocytes. The percentage of B10 cells in cultured splenocytes was significantly increased after CD40L stimulation but such increase was diminished by the addition of LPS. Such effects by LPS were only observed in cells from WT but not TLR4−/− mice. IL-10 mRNA expression and protein production in B10 cells from cultured splenocytes were significantly up-regulated by CD40L stimulation but were inhibited after the addition of LPS in a TLR4-dependent manner. This study suggests that LPS-induced TLR4 signaling attenuate CD40L-activated regulatory B10 cell competency.

Effects of Separate and Concomitant TLR-2 and TLR-4 Activation in Peripheral Blood Mononuclear Cells of Newborn and Adult Horses

Deficient innate and adaptive immune responses cause newborn mammals to be more susceptible to bacterial infections than adult individuals. Toll-like receptors (TLRs) are known to play a pivotal role in bacterial recognition and subsequent immune responses. Several studies have indicated that activation of certain TLRs, in particular TLR-2, can result in suppression of inflammatory pathology. In this study, we isolated peripheral blood mononuclear cells (PBMCs) from adult and newborn horses to investigate the influence of TLR-2 activation on the inflammatory response mediated by TLR-4. Data were analysed in a Bayesian hierarchical linear regression model, accounting for variation between horses. In general, cytokine responses were lower in PBMCs derived from foals compared with PBMCs from adult horses. Whereas in foal PBMCs expression of TLR-2, TLR-4, and TLR-9 was not influenced by separate and concomitant TLR-2 and TLR-4 activation, in adult horse PBMCs, both TLR ligands caused significant up-regulation of TLR-2 and down-regulation of TLR-9. Moreover, in adult horse PBMCs, interleukin-10 protein production and mRNA expression increased significantly following concomitant TLR-2 and TLR-4 activation (compared with sole TLR-4 activation). In foal PBMCs, this effect was not observed. In both adult and foal PBMCs, the lipopolysaccharide-induced pro-inflammatory response was not influenced by pre-incubation and co-stimulation with the specific TLR-2 ligand Pam3-Cys-Ser-Lys4. This indicates that the published data on other species cannot be translated directly to the horse, and stresses the necessity to confirm results obtained in other species in target animals. Future research should aim to identify other methods or substances that enhance TLR functionality and bacterial defence in foals, thereby lowering susceptibility to life-threatening infections during the first period of life.

In Vitro N-Acetyl-l-Cysteine Promotes Proliferation and Suppresses Interleukin-8 Expression in Adipose-Derived Stem Cells

Adipose-derived stem cells (ADSCs) hold great promise for repair and regeneration of burn wounds by producing growth factors, but proinflammatory cytokines such as interleukin-8 (IL-8) released by ADSCs would potentially deepen the wound and inhibit healing. The reported research aimed to identify the effects of N-acetyl-L-cysteine (NAC) on the proliferation, death, and IL-8 production of ADSCs. In the presence or absence of NAC, ADSC proliferation was examined using a CCK-8 Kit, and cell death was evaluated by flow cytometry analysis. Subsequently, IL-8 mRNA expression was detected by reverse transcriptase-polymerase chain reaction and protein production by enzyme-linked immunoassay. Findings showed that cell proliferation in the NAC-treated group was a significant 1.53-fold greater than in the control group, that the apoptosis rate of ADSCs decreased by 55.4 % compared with the control group, and that the necrosis rate decreased by 48.8 %. Additionally, the IL-8 mRNA expression decreased to 46.2 ± 8.7 % that of the control group, and the IL-8 protein production decreased to 9.98 ± 0.57 %. The authors believe that NAC might be helpful in burn wound repair and regeneration by stimulating the proliferation of ADSCs, inhibiting cell death, and suppressing IL-8 production. This journal requires that authors assign a level of evidence to each article.

Silicone Nanoparticles Do Not Induce Immune Responses by Naïve Human Peripheral Blood Mononuclear Cells

Several studies have reported adverse immunological effects of silicone due to their ability to induce proinflammatory molecules, such as tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6). In recent years, use of nanoparticles has been under fast development for therapeutic drug targeting, diagnostic imaging, and immune response in various fields of nanomedicine. The authors hypothesize that immune responses induced by in vivo use of silicone materials can be reduced or eliminated by the use of nanosilicone. Peripheral blood mononuclear cells obtained from naïve normal subjects were cultured with different concentrations of silicone nanoparticles and microparticles for 24 hours. The culture supernatants were quantitated for TNF-α, IL-6, and interferon-γ (IFN-γ) secretion by enzyme-linked immunosorbent assay. The pellets were used for specific IL-6, TNF-α, and IFN-γ gene expression by real-time polymerase chain reaction, respectively. Cytotoxicity was evaluated by XTT viability assay. Results were compared between silicone nanoparticles and microparticles and untreated controls. Silicone nanoparticles up to 100 μg/ml did not induce any detectable levels of specific TNF-α, IFN-γ, and IL-6 gene expression and protein production and the results were comparable to those for untreated controls. Silicone microparticles at 100 μg/ml, however, significantly induced the production and gene expression of TNF-α, IL-6, and IFN-γ by peripheral blood mononuclear cells. XTT viability assay showed that silicone nanoparticles or microparticles, even at the highest concentration used, were not cytotoxic to cells. The results suggest that silicone nanoparticles can be engineered to avoid immune recognition and subsequent silicone microparticle-related adverse effects and thus may be of therapeutic significance in the cosmetic industry, plastic surgery, and aesthetic medicine.

−251 T/A polymorphism of the interleukin-8 gene and cancer risk: a HuGE review and meta-analysis based on 42 case–control studies

The -251T/A (rs4073), a single nucleotide polymorphism, has been identified in the promoter region of the interleukin-8 (IL-8) gene. It's presence could influence the production of IL-8 protein by regulating the transcriptional activity of the gene. A large number of studies have been performed to evaluate the role of -251T/A polymorphism on various cancers, with inconsistent results being reported. In this paper, we summarized 13,189 cases and 16,828 controls from 42 case-control studies and attempted to assess the susceptibility of -251T/A polymorphism to cancers by a comprehensive meta-analysis. Pooled odds ratios and 95% confidence intervals were calculated by using the random-effects model. Publication bias, subgroup, and sensitivity analysis were also performed. Results showed that the carriers of the -251A allele had about a 12-21% increased risk for the reviewed cancer, in total. The carriers of -251A had an elevated risk to breast cancer, gastric cancer and nasopharyngeal cancer and a reduced risk to prostate cancer, but no evidence was found to indicate that the -251A allele predisposed its carriers to colorectal and lung cancers. When stratified separately by 'racial descent' and 'study design', it was found that the carriers of the -251A allele among the African group, Asian group and hospital-based case-control study group were at a higher risk for cancer, but not in European group and population-based case-control study. These results show that -251A allele is susceptible in the development of low-penetrance cancers.

Gomisin N has anti-allergic effect and inhibits inflammatory cytokine expression in mouse bone marrow-derived mast cells

Gomisin N is a bioactive compound and a prominent anti-allergic agent found in the fruits of tree Schizandra chinensis. However, its effects on the bone marrow-derived mast cell (BMMC)-mediated allergy and inflammation mechanism remain unknown. In this study, the biological effects of gomisin were evaluated while focusing on its effects on the allergic mediator in PMA + A23187-stimulated BMMCs. The anti-allergic effect of gomisin has shown that inhibited PMA + A23187-induced interleukin-6 (IL-6) production. An investigation was also conducted to determine its effects on the production of several allergic mediators including prostaglandin D2 (PGD2), leukotriene C4 (LTC4), β-hexosaminidase (β-Hex), and cyclooxygenase-2 (COX-2) protein. The results revealed that gomisin inhibited the PMA + A23187-induced production of IL-6, PGD2, LTC4, β-Hex, and COX-2 protein. Taken together, these findings indicate that gomisin N has the potential for use in the treatment of allergy.

Stress hormones increase cell proliferation and regulates interleukin-6 secretion in human oral squamous cell carcinoma cells

Patients with oral cancer can have high psychological distress levels, but the effects of stress-related hormones on oral cancer cells and possible mechanisms underlying these relationships are unknown. In this study, we have investigated the effects of stress-related hormones on interleukin-6 (IL-6) secretion and proliferation of oral squamous cell carcinoma (OSCC) cells. The effects of norepinephrine (NE), and cortisol were studied in SCC9, SCC15, and SCC25 cells and effects of isoproterenol in SCC9 and SCC25 cells. Real-time PCR studies revealed constitutive β1- and β2-adrenergic receptors (β-ARs) expression in the SCC9, SCC15, and SCC25 cells. The results showed that NE and isoproterenol significantly enhanced IL-6 mRNA expression and protein production in supernatants of SCC9 and SCC25 cells. Physiological stress levels of NE and isoproterenol (10μM) at 1h elicited the most robust IL-6 increase. Regarding IL-6 secretion, 10μM NE induced a 5-fold increase at 1h, 3.7-fold increase at 6h, and 3.2-fold at 24h in SCC9 cells. These effects were blocked by the β-adrenergic antagonist propranolol, supporting a role for β-ARs in IL-6 secretion. The effects of cortisol varied according to the hormone concentration. Pharmacological concentrations of cortisol (1000nM) inhibited IL-6 production by SCC9 and SCC25 cells. Cortisol dose that simulates stress conditions (10nM) tended to increase IL-6 expression in SCC9 cells. Hormonal doses that simulate stress conditions (10μM NE, at 6h in SCC9 and SCC15 cells and 10nM cortisol, at 48h in SCC15 cells) stimulated increased cell proliferation. Treatment of SCC9 cells with IL-6 neutralizing ab (10μg/mL) partially inhibited NE-induced proliferation. Finally, 20 OSCC biopsies were shown to express β1- and β2-ARs. These findings suggest that stress hormones can affect oral cancer cells behavior.

Anti-Allergic Activity of a Platycodon Root Ethanol Extract

Platycodon grandiflorum (Campanulaceae) is used as traditional medicine in Asian countries. In Korean traditional medicine, Platycodon root has been widely used since ancient times as a traditional drug to treat cold, cough and asthma. However, its effects on bone marrow-derived mast cell (BMMC)-mediated allergy and inflammation mechanisms remain unknown. In this study, the biological effect of Platycodon root ethanol extract (PE) was evaluated in BMMC after induction of allergic mediators by phorbol 12-myristate 13-acetate (PMA) plus calcium ionophore A23187 (A23187) stimulation. The effect of PE on the production of several allergic mediators, such as interleukin-6 (IL-6), prostaglandin D2 (PGD2), leukotriene C4 (LTC4), β-Hexosaminidase (β-Hex) and cyclooxygenase-2 (COX-2) protein, was investigated. The results demonstrate that PE inhibits PMA + A23187 induced production of IL-6, PGD2, LTC4, β-Hexosaminidase and COX-2 protein. Taken together, these results indicate that PE has the potential for use in the treatment of allergy.

Targeted Knockdown of EGR-1 Inhibits IL-8 Production and IL-8-mediated Invasion of Prostate Cancer Cells through Suppressing EGR-1/NF-κB Synergy

IL-8 produced by prostate cancer cells may be responsible for the androgen-independent growth of advanced prostate cancers. Accumulating evidence from microarray analyses and animal genetic models highlights the central involvement of the transcription factor early growth response-1 (EGR-1) in prostate carcinoma progression. It is unknown, however, whether knockdown of EGR-1 inhibits IL-8 production and IL-8-mediated tumor metastasis. Here we show that EGR-1 knockdown by a specific shRNA-Egr1 inhibited gene transcription and production of IL-8 by the human prostate cancer cell line DU145. Conversely, enforced expression of EGR-1 in EGR-1-lacking PC3 prostate cancer cells markedly enhanced IL-8 transcription and secretion. By using wild type and a series of mutant IL-8 promoter luciferase constructs, we found that the NF-kappaB binding site is important for EGR-1 regulation of IL-8. Furthermore, silencing EGR-1 suppressed a synergistically functional interaction between EGR-1 and NF-kappaB. Consequently, knockdown of EGR-1 inhibited IL-8-mediated tumor colony formation and invasion. Thus, targeted knockdown of EGR-1 could be an effective therapeutic approach against prostate cancer.

The effect of endothelin‐1 on the production of interleukin‐6 in cultured human detrusor smooth muscle cells, and the effect of interleukin‐6 on the contractile response of bladder smooth muscle strips from rats

To determine the effect of endothelin-1 (ET-1) on the production of interleukin-6 (IL-6) in cultured human detrusor smooth muscle cells (HDSMCs) and the effect of IL-6 on the contractile response of bladder smooth muscle strips from rats. The expression of IL-6 mRNA and production of IL-6 protein in cultured HDSMCs treated with the different concentrations of ET-1 were assayed by reverse transcriptase-polymerase chain reaction and specific enzyme-linked immunosorbent assay, respectively. The strips from Sprague-Dawley rats were either untreated or treated with 1 ng/mL of IL-6 for 60 min. Using increasing cumulative concentrations of acetylcholine (ACh), bethanechol (BCh), or sodium nitroprusside, we assessed the concentration-contraction or the relaxation responses. In cystitis-induced strips, change of contractions induced by noradrenaline (NA) with or without treatment of IL-6 were assessed. The IL-6-treated strips were incubated for 30 min in the presence or absence indomethacin or SQ29548, and then the effects on ACh- or BCh-induced contractions were measured. The expression of IL-6 mRNA and the production of IL-6 protein on the cultured HDSMCs pretreated by ET-1 were significantly higher than in the control (P<0.05). The ACh- and BCh-induced contractions were increased in the IL-6 pretreated strips from both dome and trigone, regardless of the presence of urothelium (P<0.05). The presence of cystitis augmented the NA-induced contractions (P<0.05). The contractions induced by ACh and BCh were inhibited by indomethacin and SQ29548. ET-1 induces expression of IL-6 mRNA and production of IL-6 protein on HDSMCs. IL-6 enhances detrusor smooth muscle contractility via the muscarinic or adrenergic receptor pathway.