Loss of RACK1 Promotes Metastasis of Gastric Cancer by Inducing a miR-302c/IL8 Signaling Loop.

Gastric cancer remains the third leading cause of cancer-related mortality worldwide, and invasion and metastasis of gastric cancer represent the major reason for its poor prognosis. In this study, we found that loss of the receptor for activated C-kinase 1 (RACK1) promoted the metastasis of gastric cancer by enhancing the autocrine expression of IL8 in vitro and in vivo. microRNA (miRNA; miR) array identified that RACK1 modulated the expression of a series of miRNAs, including the miR-302 cluster, and RACK1 modulated the IL8 expression and tumor invasion through miRNA-302c. Moreover, upregulation of IL8 in turn decreased the level of miRNA-302c and induced IL8 expression in a feedback manner. Tissue microarray also indicated that RACK1 was correlated with invasion/metastasis phenotype, IL8 expression, as well as 5-year survival in clinical cases of gastric cancer. Together, our results imply that loss of RACK1 in gastric cancer links epigenetics to inflammatory cytokines to promote tumor metastasis.

Targeted delivery of pulmonary arterial endothelial cells overexpressing interleukin-8 receptors attenuates monocrotaline-induced pulmonary vascular remodeling.

OBJECTIVE: Interleukin-8 (IL-8) receptors IL8RA and IL8RB (IL8RA/B) on neutrophil membranes bind to IL-8 with high affinity and play a critical role in neutrophil recruitment to sites of injury and inflammation. This study tested the hypothesis that administration of rat pulmonary arterial endothelial cells (ECs) overexpressing IL8RA/B can accelerate the adhesion of ECs to the injured lung and inhibit monocrotaline-induced pulmonary inflammation, arterial thickening and hypertension, and right ventricular hypertrophy. APPROACH AND RESULTS: The treatment groups included 10-week-old ovariectomized Sprague-Dawley rats that received subcutaneous injection of PBS (vehicle), a single injection of monocrotaline (monocrotaline alone, 60 mg/kg, SC), monocrotaline followed by intravenous transfusion of ECs transduced with the empty adenoviral vector (null-EC), and monocrotaline followed by intravenous transfusion of ECs overexpressing IL8RA/B (1.5 × 10(6) cells/rat). Two days or 4 weeks after monocrotaline treatment, endothelial nitric oxide synthase, inducible nitric oxide synthase, cytokine-induced neutrophil chemoattractant-2ß (IL-8 equivalent in rat), and monocyte chemoattractant protein-1 expression, neutrophil and macrophage infiltration into pulmonary arterioles, and arteriolar and alveolar morphology were measured by histological and immunohistochemical techniques. Proinflammatory cytokine/chemokine protein levels were measured by Multiplex rat-specific magnetic bead-based sandwich immunoassay in total lung homogenates. Transfusion of ECs overexpressing IL8RA/B significantly reduced monocrotaline-induced neutrophil infiltration and proinflammatory mediator (IL-8, monocyte chemoattractant protein-1, inducible nitric oxide synthase, cytokine-induced neutrophil chemoattractant, and macrophage inflammatory protein-2) expression in lungs and pulmonary arterioles and alveoli, pulmonary arterial pressure, and pulmonary arterial and right ventricular hypertrophy and remodeling. CONCLUSIONS: These provocative findings suggest that targeted delivery of ECs overexpressing IL8RA/B is effective in repairing the injured pulmonary vasculature.

Effect of dietary supplementation on the antimicrobial activity of blood leukocytes isolated from Holstein heifers.

The purpose of this investigation was to evaluate the effect of an immunostimulating feed supplement (OmniGen-AF®) on the antimicrobial properties of blood leukocytes in dairy heifers in an attempt to prevent mastitis. Blood leukocytes from supplemented and unsupplemented controls were used. Phagocytic activity and reactive oxygen species (ROS) production were studied on d 0 (prior to feed supplementation) and on days 30 and 60 after supplementation. L-selectin and IL-8R mRNA expressions on blood leukocytes were evaluated on d 0 (prior to feed supplementation) and monthly thereafter for 15 mo. On d 30 after supplementation, neutrophils from treated heifers exhibited greater binding and internalization of Escherichia coli and greater ROS production compared with unsupplemented controls. L-selectin mRNA expression was increased in supplemented heifers vs. controls; however, IL-8R mRNA expression was not different. Results support the continued study of dietary supplementation as an additional management tool to enhance udder health in dairy heifers.

Endothelial cells overexpressing IL-8 receptor reduce cardiac remodeling and dysfunction following myocardial infarction.

The endothelium is a dynamic component of the cardiovascular system that plays an important role in health and disease. This study tested the hypothesis that targeted delivery of endothelial cells (ECs) overexpressing neutrophil membrane IL-8 receptors IL8RA and IL8RB reduces acute myocardial infarction (MI)-induced left ventricular (LV) remodeling and dysfunction and increases neovascularization in the area at risk surrounding the infarcted tissue. MI was created by ligating the left anterior descending coronary artery in 12-wk-old male Sprague-Dawley rats. Four groups of rats were studied: group 1: sham-operated rats without MI or EC transfusion; group 2: MI rats with intravenous vehicle; group 3: MI rats with transfused ECs transduced with empty adenoviral vector (Null-EC); and group 4: MI rats with transfused ECs overexpressing IL8RA/RB (1.5 × 106 cells post-MI). Two weeks after MI, LV function was assessed by echocardiography; infarct size was assessed by triphenyltetrazolium chloride (live tissue) and picrosirus red (collagen) staining, and capillary density and neutrophil infiltration in the area at risk were measured by CD31 and MPO immunohistochemical staining, respectively. When compared with the MI + vehicle and MI-Null-EC groups, transfusion of IL8RA/RB-ECs decreased neutrophil infiltration and pro-inflammatory cytokine expression and increased capillary density in the area at risk, decreased infarct size, and reduced MI-induced LV dysfunction. These findings provide proof of principle that targeted delivery of ECs is effective in repairing injured cardiac tissue. Targeted delivery of ECs to infarcted hearts provides a potential novel strategy for the treatment of acute MI in humans.

Differential expression of IL-8 and IL-8 receptors in benign, borderline and malignant ovarian epithelial tumours.

INTRODUCTION: Ovarian Cancer is the leading cause of death from gynecological malignancy. The poor prognosis is mainly due to presentation at a late stage and poor response to therapy. Much research is needed to identify diagnostic and prognostic biomarkers as well as therapeutic targets for ovarian cancer. Interleukin-8 is expressed by many tumour types and is known to have mitogenic, motogenic and angiogenic effects on tumour cells. AIMS: The aim of this study was to investigate the expression of IL-8 and IL-8 receptors (IL-8RA and IL-8RB) in different histological subtypes of ovarian tumours, as potential prognostic biomarkers in ovarian tumours. MATERIALS AND METHODS: Immunohitochemistry was used to study the expression of IL-8 and IL-8 receptors in 115 ovarian tumours including 21 benign tumours, 25 borderline tumours and 69 carcinomas of serous, clear cell, endometrioid and mucinous types. The correlation of expression profile, tumour type, stage, and progression free survival and overall survival was statistically analysed. RESULTS: IL-8 and IL-8 receptors were expressed in all types of tumours with variable intensity and subcellular distribution. There was a statistically significant correlation between levels of expression and tumour stage and tumour type, being mostly significant in serous tumours. No correlation with patient progression free survival or overall survival was found. CONCLUSION: This is the first study investigating the expression of IL-8 and IL-8 receptors using immunohistochemistry in different types of ovarian tumours, including benign and borderline tumours. IL-8 and IL-8RA are potential prognostic biomarkers and therapeutic targets in ovarian cancer, particularly in ovarian serous carcinoma.

Knockdown of aberrantly expressed nuclear localized decorin attenuates tumour angiogenesis related mediators in oral cancer progression model in vitro.

BACKGROUND: Oral cancer accounts for roughly 3% of cancer cases in the world with about 350,000 newly reported cases annually and a 5-year survival rate of only 50%. Majority of oral cancers are squamous cell carcinomas that originate in the oral mucosal epithelial linings. We have previously shown that in human malignant squamous cells carcinoma (SCC-25) as well as in dysplastic oral keratinocytes (DOK), a small leucine-rich multifunctional proteoglycan decorin is aberrantly expressed and localized in the nucleus where it interacts with nuclear epidermal growth factor receptor (EGFR). Post-transcriptional silencing of nuclear decorin significantly reduced IL-8 and IL8-dependent migration and invasion in these dysplastic and malignant oral epithelia. The objective of this study was to further examine the effects of nuclear decorin silencing on angiogenesis and angiogenesis related mediators in this oral cancer progression cell line model. METHODS: We have used multiplex PCR, western blotting, and in vitro endothelial tube formation assay to study angiogenesis and related pathways in nuclear decorin silenced (stable knockdown) DOK and SCC-25 cells. RESULTS: Nuclear decorin knockdown resulted in significant down regulation of IL-8 expression, however IL-10, and TGF-ß expression was not affected in either DOK or SCC25 cells as measured by multiplex RT PCR. IL-8 receptor CXCR 1 and 2 expression was slightly lower in nuclear decorin silenced cells indicating a contributing mechanism in previously shown reduced IL-8 mediated migration and invasion phenotype in these cells. IL-8 is known to induce Matrix metalloproteinase 9 (MMP9) which not only plays a role in tumour migration and invasion but also induces angiogenic switch. We found MMP9 to be significantly reduced in nuclear decorin silenced dysplastic and malignant oral epithelia. Other potent angiogenic mediators, VEGF189 and ANG-1 were either significantly reduced or completely abrogated in these cells. Angiogenesis as measured by endothelial tube-like formations of HUVEC cells was reduced by almost 50 percent when HUVECs were incubated in the presence of conditioned medium form nuclear decorin silenced dysplastic and malignant cell lines as compared to respective controls. CONCLUSIONS: Together these results indicate that aberrantly expressed nuclear localized decorin strongly influences angiogenic potential of dysplastic and malignant oral epithelial cells.

Dental implants stimulate expression of Interleukin-8 and its receptor in human blood--an in vitro approach.

Interleukin (IL)-8 secreted from osteoblasts and peripheral blood monocytes increases in patients with aseptic hip-implant loss and in patients with mucositis after dental implant insertion. We explored in vitro the possibility of an IL-8-mediated inflammatory response as a consequence of contact between different dental implant surfaces and human blood. Titanium and zirconia implants were incubated in human blood. Nonstimulated blood served as negative, while blood stimulated with bacterial lipopolysaccharides (LPS) served as positive control. After depyrogenization, to examine the possible role of LPS, implants were again submerged in blood. Gene-expression of IL-8 and its receptor was measured by real-time quantitative polymerase chain reaction. In a receptor mediated, but LPS-independent manner, titanium implants led to a more pronounced increase in IL-8 gene expression when compared with zirconia implants. Depyrogenization resulted after 24 h in zirconia implants in decreased IL-8 gene expression. Altered IL-8 expression could indicate aseptic, at least LPS-independent implant loss, which may be an additional feature in the manifestation of peri-implantitis, possibly triggered by microscopically small implant-particles. Hence, opening a new field of investigations to further understand the possible mechanism underlying the manifestation of implant failure.

IL-8 signaling plays a critical role in the epithelial-mesenchymal transition of human carcinoma cells.

The switch of tumor cells from an epithelial to a mesenchymal-like phenotype [designated as epithelial-to-mesenchymal transition (EMT)] is known to induce tumor cell motility and invasiveness, therefore promoting metastasis of solid carcinomas. Although multiple studies have focused on elucidating the signaling events that initiate this phenotypic switch, there has been so far no characterization of the pattern of soluble mediators released by tumor cells undergoing EMT, and the potential impact that this phenotypic switch could have on the remodeling of the tumor microenvironment. Here we show that induction of EMT in human carcinoma cells via overexpression of the transcription factor Brachyury is associated with enhanced secretion of multiple cytokines, chemokines, and angiogenic factors and, in particular, with the induction of the IL-8/IL-8R axis. Our results also indicate the essential role of interleukin 8 (IL-8) signaling for the acquisition and/or maintenance of the mesenchymal and invasive features of Brachyury-overexpressing tumor cells and show that IL-8 secreted by tumor cells undergoing EMT could potentiate tumor progression by inducing adjacent epithelial tumor cells into EMT. Altogether, our results emphasize the potential role of EMT in the modulation of the tumor microenvironment via secretion of multiple soluble mediators and suggest that IL-8 signaling blockade may provide a means of targeting mesenchymal-like, invasive tumor cells.

Short-term heat exposure inhibits inflammation by abrogating recruitment of and nuclear factor-{kappa}B activation in neutrophils exposed to chemotactic cytokines.

Cytokines, such as granulocyte macrophage-colony stimulating factor (GM-CSF) and interleukin (IL)-8 attract neutrophils into inflammatory sites. During emigration from the blood neutrophils interact with extracellular matrix proteins such as fibronectin. Fibronectin provides beta2-integrin co-stimulation, allowing GM-CSF and IL-8 to activate nuclear factor (NF)-kappaB, an effect that does not occur in suspension. We tested the hypothesis that exposure of mice to fever-like temperatures abrogates neutrophil recruitment and NF-kappaB activation in a mouse model of skin inflammation. Mice that were exposed to 40 degrees C for 1 hour showed strongly reduced GM-CSF- and IL-8-induced neutrophilic skin inflammation. In vitro heat exposure did not interfere with neutrophil adhesion or spreading on fibronectin but strongly inhibited migration toward both cytokines. Using specific inhibitors, we found that PI3-K/Akt was pivotal for neutrophil migration and that heat down-regulated this pathway. Furthermore, neutrophils on fibronectin showed abrogated NF-kappaB activation in response to GM-CSF and IL-8 after heat. In vivo heat exposure of mice followed by ex vivo stimulation of isolated bone marrow neutrophils confirmed these results. Finally, less NF-kappaB activation was seen in the inflammatory lesions of mice exposed to fever-like temperatures as demonstrated by in situ hybridization for IkappaBalpha mRNA. These new findings suggest that heat may have anti-inflammatory effects in neutrophil-dependent inflammation.