Effect of induced airway inflammation in asthma : the expression of trefoil factors, secretoglobins and genes involved in histone acetylation

Asthma is an incapacitating disease of the respiratory system, which causes extensive morbidity and mortality worldwide. Asthma affects more than 300 million people globally(Masoli et al. 2004). In Australia, it affects 10.2% of the population (Masoli et al. 2004) and causes 60,000 people to be hospitalised annually. Health care expenditure due to asthma in Australia was $606 million in 2004–2005. There are four primary biological factors that function in the initiation and exacerbation of asthma. Airway inflammation is important as it is often the first response to an airway insult, initiating the three other components: bronchoconstriction, mucus hyper-secretion and hyper-reactivity. The mediators involved in asthma are still not well understood, and current anti-inflammatory corticosteroid treatments are not effective with all asthmatics. As there is currently no cure for asthma, and airway inflammation is the primary component of the disease, it is important that we understand and investigate the mediators of airway inflammation to look for a potential cure and to produce better therapeutics to treat the inflammation. Trefoil factors (TFFs) and secretoglobins (SCGBs) are small secreted proteins involved in the mediation of inflammation and epithelial restitution. TFFs are pro-inflammatory and SCGBs anti-inflammatory by nature. The hypothesis of this study is that in response to induced acute airway inflammation, the expression of TFF1 and TFF3 will increase and expression of SCGB1A1 and SCGB3A2 will decrease in non-asthmatics (N-A), asthmatics medicating with bronchodilators (A-BD) and asthmatics medicating with corticosteroids (A-ST). When comparing the three groups, we expect to see higher expression of the TFFs in the A-BD group compared to the N-A and A-ST groups, indicating that inflammation is mediated by TFFs in asthma and that corticosteroid medication controls their expression as part of the control of inflammation. We expect to see the opposite with SCGBs, with a greater decrease in the A-BD group compared to the other two groups, suggesting that the A-BD group has the least anti-inflammatory activity in response to inflammatory insult. Epigenetic modification plays a role in the regulation of genes that initiate disease states such as inflammatory conditions and cancers. Histone acetylation is one such modification, which involves the acetylation of histones in chromatin by histone acetyltransferases (HATs). This increases the transcription of genes involved with inflammation or enrols histone deacetylases (HDACs) to down-regulate the transcription of inflammatory genes. These HATs and HDACs work in a homeostatic fashion; however, in the event of inflammation, increased HAT activity can stimulate further inflammation, which is believed to be the mechanism involved in some inflammatory diseases. This study hypothesises that in response to inflammation, the expression of HDACs (HDAC1-5) will decrease and the expression of HATs (NCOA1-3, HAT-1 and CREBBP) will increase in all groups. When comparing the expression between the groups, it was expected that a greater decrease in HDACs and a greater increase in HATs will be seen in the A-BD group compared to the other two groups. This would identify histone acetylation as a mechanism involved in the inflammatory condition of asthma and indicate that corticosteroids may treat the inflammation in asthma at least in part by controlling histone acetylation. The aim of the project was to compare the expression of inflammatory genes TFF1, TFF3, SCGB1A1 and SCGB3A2, as well as to compare the gene expression of HDAC1-5, NCOA1-3, HAT-1 and CREBBP within and between N-A (n=15), A-BD (n=15) and A-ST (n=15) groups in response to inflammation. This was performed by collecting airway cells and proteins by sputum induction in three sessions. The sessions were coordinated into an initial baseline collection (SI-1), followed by a second session at least one week later (SI-2) and a third session, six hours after SI-2 to collect a sample containing the resultant acute inflammation caused in SI-2 (SI-3). Analysis of the SI-1 and SI-2 samples in all three groups had high amounts of variability between samples. The samples were taken at least one weak apart and the environmental stimuli on each participant outside of the testing sessions could not be controlled. For this reason, the SI-1 samples were not used for analysis; instead SI-2 and SI-3 samples were compared as they were same-day collections, reducing the probability of differences being due to anything other than the sputum induction. The gene expressions of the TFFs, SCGBs, HDACs and HATs were analysed using real-time PCR. Western blot analysis was performed to analyse the protein concentrations of the TFFs and SCGBs in secreted fractions of the sputum collection. Both the secreted and intracellular protein fractions collected from the sputum inductions for pre- and post-inflammation (SI-2, SI-3) samples of the N-A and A-BD groups were analysed using a proteomic method called iTRAQ. This allowed the comparison of the change in protein expression as a result of airway inflammation in each group. This technique was used as a discovery method to identify novel proteins that are modulated by induced acute airway inflammation. Any proteins of interest would then be further validated and used for future research. Inflammation was achieved in the SI-3 samples of the N-A group with a 21% unit increase in % neutrophils compared to SI-2 (p=0.01). The N-A group had a marked 5.5-fold decrease in HDAC1 gene expression in SI-3 compared to SI-2 (p=0.03). No differences were seen in any of the TFFs, SCGBs or any of the rest of the HDACs and HATs. Western blot data did not display any significant changes in the protein levels of the TFFs and SCGBs analysed. However, non-significant analysis of the data displayed increases in TFF1 and TFF3, and decreases in SCGB1A1 and SCGB3A2 for the majority of SI-3 samples compared to SI-2. The A-BD group also presented a marked increase in neutrophils in the SI-3 samples compared to SI-2 (27% unit increase, p=0.04). The A-BD group had a significant increase in TFF3 and SCGB1A1 gene expression concomitant with induced acute airway inflammation. A 7.3-fold increase in TFF3 (p=0.05) in SI-3 indicated that TFF3 is linked to inflammation in asthmatics. A 2.8-fold increase in SCGB1A1 (p=0.03) indicated that this gene is also up-regulated, suggesting that this SCGB is expressed to try to combat induced acute airway inflammation. No significant changes were seen in any of the other genes analysed. Western blot data did not display any significant changes in the protein levels of the TFFs and SCGBs analysed. However, non-significant analysis of the data displayed an increase in TFF1 and TFF3, and a decrease in SCGB1A1 and SCGB3A2 in SI-3, similar to that seen in the N-A group. The A-ST group was different from the A-BD group, characterised by the use of inhaled corticosteroid medication to treat asthma symptoms. Inhaled corticosteroids are known to treat asthma symptoms through the control of inflammation. Therefore, it was expected that corticosteroid medication would also control the expression of TFFs, SCGBs, HATs and HDACs. Gene expression results only identified a 7.6-fold decrease in HDAC2 expression in SI-3 (p=0.001), which is proposed to be due to the up-regulation of HDAC2 protein that is known to be a function of corticosteroid use. Western blot data did not display any significant changes in the protein levels of the TFFs and SCGBs analysed. The gene expression in SI-2 and SI-3 in each group was compared. When comparing the A-BD group to the N-A group, a 9-fold increase in TFF3 (p=0.008) and a 34-fold increase in SCGB1A1 (p=0.03) were seen in the SI-3 samples. Comparisons of the A-ST group to the N-A group had an increased expression in SI-2 samples for HDAC5 (3.6-fold, p=0.04), NCOA2 (8.5-fold, p=0.04), NCOA3 (17-fold, p=0.01), HAT-1 (36-fold, p=0.003) and CREBBP (13-fold, p=0.001). The SI-3 samples in the A-ST group compared to the N-A group had increased expression for HDAC1 (6.4-fold, p=0.04), HDAC5 (5.2-fold, p=0.008), NCOA2 (9.6-fold, p=0.03), NCOA3 (16-fold, p=0.06), HAT-1 (41-fold, p<0.001) and CREBBP (31-fold, p=0.001). Comparisons of the A-ST group to the A-BD group had SI-2 increases in HDAC1 (3.8-fold, p=0.03), NCOA3 (4.5-fold, p=0.03), HAT-1 (5.3-fold, p=0.01) and CREBBP (23-fold, p=0.001), while SI-3 comparisons saw a decrease in HDAC2 (41-fold, p=0.008) and increases in HAT-1 (4.3-fold, p=0.003) and CREBBP (40-fold, p=0.001). Results showed that TFF3 and SCGB1A1 expression is higher in asthmatics than non-asthmatics and that histone acetylation is more active in the A-ST group than either the N-A or A-BD group, which suggests that histone acetylation activity may be positively correlated with asthma severity. The iTRAQ proteomic analysis of the secreted protein samples identified the SCGB1A1 protein and found it to be decreased in both the N-A and A-BD groups post-inflammation, but significantly so only in the A-BD group. Although no significant results were obtained from the western blot data, both groups displayed a decrease in SCGB1A1 concentration in SI-3 samples, suggesting a correlation with the proteomic data. Only 31 peptides were identified from the secreted samples. The intracellular iTRAQ analysis successfully identified 664 peptides, eight of which had differential expression in association with induced acute airway inflammation. Significant increases were seen in the A-BD group in SI-3 compared to SI-2 than in the N-A group in chloride intracellular channel protein 1, keratin-19, eosinophil cationic protein, calnexin, peroxiredoxin-5, and ATP-synthase delta subunit, while decreases were seen in cystatin-A and mucin-5AC. The iTRAQ analysis was only a discovery measure and further validation must be performed. In summary, the expression of TFFs and SCGBs differed between non-asthmatics and asthmatics. It is clear that TFF3 is active in the airway inflammation associated with asthma as indicated by an increase associated with inflammation in the A-BD group compared to the N-A group. Results for HDAC and HAT genes showed high HAT expression in the A-ST group compared to the N-A and A-BD groups, suggesting that histone acetyltransferases may be responsible for the characteristic unregulated inflammatory symptoms of asthmatics taking corticosteroids. Interestingly, corticosteroid medication did not seem to silence the expression of the analysed HAT genes, which indicates that corticosteroids may not control inflammation by direct regulation of HATs, but instead by competition, most probably with HDAC2 protein. As a discovery tool, iTRAQ is a potent method to both identify and compare the concentration of proteins between samples. The method is a powerful first step into the identification of novel proteins that are regulated in response to different treatments.

Genetic aspects of susceptibility, severity, and clinical expression in ankylosing spondylitis

While twin studies have previously demonstrated high heritability of susceptibility to ankylosing spondylitis (AS), it is only recently that the involvement of genetic factors in determining the severity of the disease has been demonstrated. The genes involved in determining the rate of ankylosis in AS are likely to be different from those involved in the underlying immunologic events, and represent important potential targets for treatment of AS. This article will describe the progress that has been made in the genetic epidemiology of AS, and in identifying the genes involved.

Matrix metalloproteinases 2 and 9 (gelatinases A and B) expression in malignant mesothelioma and benign pleura

Matrix metalloproteinases (MMPs), in particular the gelatinases (MMP-2 and -9), play a significant role in tumour invasion and angiogenesis. The expression and activities of MMPs have not been characterised in malignant mesothelioma (MM) tumour samples. In a prospective study, gelatinase activity was evaluated in homogenised supernatants of snap frozen MM (n = 35), inflamed pleura (IP, n = 12) and uninflammed pleura (UP, n = 14) tissue specimens by semiquantitative gelatin zymography. Matrix metalloproteinases were correlated with clinicopathological factors and with survival using Kaplan-Meier and Cox proportional hazard models. In MM, pro- and active MMP-2 levels were significantly greater than for MMP-9 (P = 0.006, P<0.001). Active MMP-2 was significantly greater in MM than in UP (P=0.04). MMP-2 activity was equivalent between IP and MM, but both pro- and active MMP-9 activities were greater in IP (P=0.02, P=0.009). While there were trends towards poor survival with increasing total and pro-MMP-2 activity (P=0.08) in univariate analysis, they were both independent poor prognostic factors in multivariate analysis in conjunction with weight loss (pro-MMP-2 P = 0.03, total MMP-2 P = 0.04). Total and pro-MMP-2 also contributed to the Cancer and Leukemia Group B prognostic groups. MMP-9 activities were not prognostic. Matrix metalloproteinases, and in particular MMP-2, the most abundant gelatinase, may play an important role in MM tumour growth and metastasis. Agents that reduce MMP synthesis and/or activity may have a role to play in the management of MM. © 2003 Cancer Research UK.

An evaluation of tumor oxygenation and gene expression in patients with early stage non-small cell lung cancers

Background: To directly assess tumor oxygenation in resectable non - small cell lung cancers (NSCLC) and to correlate tumor pO2 and the selected gene and protein expression to treatment outcomes. Methods: Twenty patients with resectable NSCLC were enrolled. Intraoperative measurements of normal lung and tumor pO2 were done with the Eppendorf polarographic electrode. All patients had plasma osteopontin measurements by ELISA. Carbonic anhydrase-IX (CA IX) staining of tumor sections was done in the majority of patients (n = 16), as was gene expression profiling (n = 12) using cDNA microarrays. Tumor pO2 was correlated with CA IX staining, osteopontin levels, and treatment outcomes. Results: The median tumor pO2 ranged from 0.7 to 46 mm Hg (median, 16.6) and was lower than normal lung pO2 in all but one patient. Because both variables were affected by the completeness of lung deflation during measurement, we used the ratio of tumor/normal lung (T/L) pO2 as a reflection of tumor oxygenation. The median T/L pO 2 was 0.13. T/L pO2 correlated significantly with plasma osteopontin levels (r = 0.53, P = 0.02) and CA IX expression (P = 0.006). Gene expression profiling showed that high CD44 expression was a predictor for relapse, which was confirmed by tissue staining of CD44 variant 6 protein. Other variables associated with the risk of relapse were T stage (P = 0.02), T/L pO2 (P = 0.04), and osteopontin levels (P = 0.001). Conclusions: Tumor hypoxia exists in resectable NSCLC and is associated with elevated expression of osteopontin and CA IX. Tumor hypoxia and elevated osteopontin levels and CD44 expression correlated with poor prognosis. A larger study is needed to confirm the prognostic significance of these factors. © 2006 American Association for Cancer Research.

Mediating orientation and self-expression in the world of consumption : Australian and German lifestyle journalists' professional views

Despite having experienced rapid popularity over the past two decades, lifestyle journalism is still somewhat neglected by academic researchers. So far mostly explored as either part of wider lifestyle programming, particularly on television, or in terms of individual sub-fields, such as travel, fashion or food journalism, lifestyle journalism is in need of scholarly analysis particularly in the area of production, based on the increasing importance which the field has in influencing audiences’ ways of life. This study explores the professional views of 89 Australian and German lifestyle journalists through in-depth interviews in order to explore the ways in which they engage in processes of influencing audiences’ self-expression, identities and consumption behaviors. The article argues that through its work, lifestyle journalism is a significant shaper of identities in today’s consumer societies.

Vascular remodeling in the internal mammary artery graft and association with in situ endothelin-1 and receptor expression

BACKGROUND: The vasoconstricting peptide endothelin-1 (ET-1) has been associated with atherosclerotic cardiovascular disease, vascular smooth muscle cell (VSMC) growth stimulation, and intimal thickening. ET-1 binds 2 receptor subtypes, endothelin A and B, and the ETA receptor mediates vasoconstriction and VSMC growth. This study aims to quantitatively assess arterial remodeling variables and compare them with changes in ET-1, ETA, and ETB expression in the internal mammary artery (IMA). METHODS AND RESULTS: Specimens from 55 coronary artery disease (CAD) patients (45 men, 10 women; mean age 65 years) and 14 control IMA specimens (from 7 men and 7 women; mean age 45 years) were collected. IMA cross sections were assessed by histochemical and immunohistochemical staining methods to quantify the levels of medionecrosis, fibrosis, VSMC growth, ET-1, ETA, ETB, and macrophage infiltration. The percentage area of medionecrosis in the patients was almost double that in the controls (31.85+/-14.52% versus 17.10+/-9.96%, P=0.0006). Total and type 1 collagen was significantly increased compared with controls (65.8+/-18.3% versus 33.7+/-13.7%, P=0.07, and 14.2+/-10.0% versus 4.8+/-2.8%, P=0.01, respectively). Despite ACE and/or statin therapy, ET-1 expression and cell cycling were significantly elevated in the patient IMAs relative to the controls (46.27+/-18.46 versus 8.56+/-8.42, P=0.0001, and 37.29+/-12.88 versus 11.06+/-8.18, P=0.0001, respectively). ETA and ETB staining was elevated in the patient vessels (46.88+/-11.52% versus 18.58+/-7.65%, P=0.0001, and 42.98+/-7.08% versus 34.73+/-5.20%, P=0.0067, respectively). A mild presence of macrophages was noted in all sections. CONCLUSIONS: Elevated distribution of collagen indicative of fibrosis coupled with increased cell cycling and high levels of ET-1 and ETA expression in the absence of chronic inflammation suggests altered IMA VSMC regulation is fundamental to the remodeling process.

Single nucleotide polymorphisms and mRNA expression of VEGF-A in papillary thyroid carcinoma: Potential markers for aggressive phenotypes

BACKGROUND AND OBJECTIVES Polymorphisms of the VEGF gene are known to affect the biological behaviour of cancers but have seldom been studied in thyroid cancer. The aim of the current study is to evaluate the prevalence and relevance of VEGF-A polymorphisms and mRNA expression in papillary thyroid carcinoma (PTC). MATERIALS AND METHODS Genomic DNA and total RNA were isolated from paraffin-embedded tissue from 91 PTC (51 conventional PTC and 40 follicular variant) and 78 control thyroid tissues. Three DNA polymorphisms (+936C > T, +405C > G and -141A > C) in the 3' and 5' untranslated region (3'-UTR, 5'-UTR) of VEGF-A were studied using PCR and RFLP. Also, the mRNA expression of VEGF-A in these tissues was studied by real-time PCR. RESULTS Distribution of polymorphisms in the 5'-UTR (VEGF-VEGF -141A > C and +405C > G) and 3'-UTR (VEGF +936C > T) were all significantly different in PTC and benign thyroid tissue (p = 0.0001, 0.001 and 0.028 respectively). The VEGF -141 C allele was more common in PTC with lymph node metastases (p = 0.026). VEGF + 405 Galleles andVEGF +936 CC genotype were more common in PTC of advanced pathological staging (p = 0.018 and 0.017 respectively). Also, increased expression of VEGF-A mRNA was noted in PTC compared to control (p = 0.009). Within the group of patients with conventional PTC, those with lymph nodal metastases had a higher level of VEGF-A mRNA expression than other patients (p = 0.0003). CONCLUSION These findings suggest that VEGF polymorphisms and mRNA expression may predict the aggressiveness behaviour of thyroid cancer.

Transcriptional regulation of IRS5/DOK4 expression in non-small-cell lung cancer cells

The insulin-receptor substrate family plays important roles in cellular growth, signaling, and survival. Two new members of this family have recently been isolated: IRS5/Dok4 and IRS6/Dok5. This study examines the expression of IRS5/DOK4 in a panel of lung cancer cell lines and tumor specimens. The results demonstrate that expression of IRS5/DOK4 is frequently altered with both elevated and decreased expression in non-small-cell lung cancer (NSCLC) tumor specimens. The altered expression of IRS5/DOK4 observed in tumor samples is not due to aberrant methylation. In vitro cell culture studies demonstrate that treatment of NSCLC cell lines with the histone deacetylase inhibitor trichostatin A (TSA) upregulates IRS5/DOK4. This finding indicates that expression is regulated epigenetically at the level of chromatin remodeling. Chromatin immunoprecipitation experiments confirm that the IRS5/DOK4 promoter has enhanced histone hyperacetylation following treatments with TSA. Finally, hypoxia was demonstrated to downregulate IRS5/DOK4 expression. This expression was restored by TSA. The clinical relevance of altered IRS5/DOK4 expression in NSCLC requires fur ther evaluation.

Clinical impacts of mammalian target of rapamycin expression in human colorectal cancers

This study investigated the clinicopathologic roles of mammalian target of rapamycin (mTOR) expression and its relationship to carcinogenesis and tumor progression in a colorectal adenoma-adenocarcinoma model. Two colon cancer cell lines with different pathologic stages (SW480 and SW48) and 1 normal colonic epithelial cell line (FHC) were used, in addition to 119 colorectal adenocarcinomas and 32 adenomas. mTOR expression profiles at messenger RNA (mRNA) and protein levels were investigated in the cells and tissues using real-time quantification polymerase chain reaction and immunohistochemistry. The findings were correlated with the clinicopathologic features of the tumors. The colon cell line from stage III cancer (SW48) showed higher expression of mTOR mRNA than that from stage II cancer (SW480). At the tissue level, mTOR showed higher mRNA and protein expression in colorectal carcinoma than in adenoma. The mRNA and protein expression was correlated with each other in approximately one-third of the carcinomas and adenomas. High levels of mTOR mRNA expression were noted more in carcinoma or adenoma arising from the distal portion of the large intestine (P = .025 and .019, respectively). Within the colorectal cancer population, a high level of expression of mTOR mRNA was related to the presence of lymph node metastases (P = .031), advanced pathologic stage (P = .05), and presence of persistent disease or tumor recurrence (P = .035). To conclude, the study has indicated that mTOR is likely to be involved in the development and progression of colorectal cancer and is linked to cancer initiation, invasiveness, and progression.

Stem cell regulatory gene expression in human adult dental pulp and periodontal ligament cells undergoing odontogenic/osteogenic differentiation

Introduction During development and regeneration, odontogenesis and osteogenesis are initiated by a cascade of signals driven by several master regulatory genes. Methods In this study, we investigated the differential expression of 84 stem cell–related genes in dental pulp cells (DPCs) and periodontal ligament cells (PDLCs) undergoing odontogenic/osteogenic differentiation. Results Our results showed that, although there was considerable overlap, certain genes had more differential expression in PDLCs than in DPCs. CCND2, DLL1, and MME were the major upregulated genes in both PDLCs and DPCs, whereas KRT15 was the only gene significantly downregulated in PDLCs and DPCs in both odontogenic and osteogenic differentiation. Interestingly, a large number of regulatory genes in odontogenic and osteogenic differentiation interact or crosstalk via Notch, Wnt, transforming growth factor β (TGF-β)/bone morphogenic protein (BMP), and cadherin signaling pathways, such as the regulation of APC, DLL1, CCND2, BMP2, and CDH1. Using a rat dental pulp and periodontal defect model, the expression and distribution of both BMP2 and CDH1 have been verified for their spatial localization in dental pulp and periodontal tissue regeneration. Conclusions This study has generated an overview of stem cell–related gene expression in DPCs and PDLCs during odontogenic/osteogenic differentiation and revealed that these genes may interact through the Notch, Wnt, TGF-β/BMP, and cadherin signalling pathways to play a crucial role in determining the fate of dental derived cell and dental tissue regeneration. These findings provided a new insight into the molecular mechanisms of the dental tissue mineralization and regeneration

Stromal upregulation of lateral epithelial adhesions : gene expression analysis of signalling pathways in prostate epithelium

BACKGROUND: Stromal signalling increases the lateral cell adhesions of prostate epithelial cells grown in 3D culture. The aim of this study was to use microarray analysis to identify significant epithelial signalling pathways and genes in this process. METHODS: Microarray analysis was used to identify genes that were differentially expressed when epithelial cells were grown in 3D Matrigel culture with stromal co-culture compared to without stroma. Two culture models were employed: primary epithelial cells (ten samples) and an epithelial cell line (three experiments). A separate microarray analysis was performed on each model system and then compared to identify tissue-relevant genes in a cell line model. RESULTS: TGF beta signalling was significantly ranked for both model systems and in both models the TGF beta signalling gene SOX4 was significantly down regulated. Analysis of all differentially expressed genes to identify genes that were common to both models found several morphology related gene clusters; actin binding (DIAPH2, FHOD3, ABLIM1, TMOD4, MYH10), GTPase activator activity (BCR, MYH10), cytoskeleton (MAP2, MYH10, TMOD4, FHOD3), protein binding (ITGA6, CD44), proteinaceous extracellular matrix (NID2, CILP2), ion channel/ ion transporter activity (CACNA1C, CACNB2, KCNH2, SLC8A1, SLC39A9) and genes associated with developmental pathways (POFUT1, FZD2, HOXA5, IRX2, FGF11, SOX4, SMARCC1). CONCLUSIONS: In 3D prostate cultures, stromal cells increase lateral epithelial cell adhesions. We show that this morphological effect is associated with gene expression changes to TGF beta signalling, cytoskeleton and anion activity.

Gene expression profile of primary prostate epithelial and stromal cells in response to sulforaphane or iberin exposure

BACKGROUND: Broccoli consumption has been associated with a reduced risk of prostate cancer. Isothiocyanates (ITCs) derived from glucosinolates that accumulate in broccoli are dietary compounds that may mediate these health effects. Sulforaphane (SF, 4-methylsulphinylbutyl ITC) derives from heading broccoli (calabrese) and iberin (IB, 3-methylsulphinypropyl ITC) from sprouting broccoli. While there are many studies regarding the biological activity of SF, mainly undertaken with cancerous cells, there are few studies associated with IB. METHODS: Primary epithelial and stromal cells were derived from benign prostatic hyperplasia tissue. Affymetrix U133 Plus 2.0 whole genome arrays were used to compare global gene expression between these cells, and to quantify changes in gene expression following exposure to physiologically appropriate concentrations of SF and IB. Ontology and pathway analyses were used to interpret results. Changes in expression of a subset of genes were confirmed by real-time RT-PCR. RESULTS: Global gene expression profiling identified epithelial and stromal-specific gene expression profiles. SF induced more changes in epithelial cells, whereas IB was more effective in stromal cells. Although IB and SF induced different changes in gene expression in both epithelial and stromal cells, these were associated with similar pathways, such as cell cycle and detoxification. Both ITCs increased expression of PLAGL1, a tumor suppressor gene, in stromal cells and suppressed expression of the putative tumor promoting genes IFITM1, CSPG2, and VIM in epithelial cells. CONCLUSION: These data suggest that IB and SF both alter genes associated with cancer prevention, and IB should be investigated further as a potential chemopreventative agent.

Small RNA viruses of insects : expression in plants and RNA silencing

Interest in insect small RNA viruses (SRVs) has grown slowly but steadily. A number of new viruses have been analyzed at the sequence level, adding to our knowledge of their diversity at the level of both individual virus species and families. In particular, a number of possible new virus families have emerged. This research has largely been driven by interest in their potential for pest control, as well as in their importance as the causal agents of disease in beneficial arthropods. At the same time, research into known viruses has made valuable contributions to our understanding of an emerging new field of central importance to molecular biology-the existence of RNA-based gene silencing, developmental control, and adaptive immune systems in eukaryotes. Subject to RNA-based adaptive immune responses in their hosts, viruses have evolved a variety of genes encoding proteins capable of suppressing the immune response. Such genes were first identified in plant viruses, but the first examples known from animal viruses were identified in insect RNA viruses. This chapter will address the diversity of insect SRVs, and attempts to harness their simplicity in the engineering of transgenic plants expressing viruses for resistance to insect pests. We also describe RNA interference and antiviral pathways identified in plants and animals, how they have led viruses to evolve genes capable of suppressing such adaptive immunity, and the problems presented by these pathways for the strategy of expressing viruses in transgenic plants. Approaches for countering these problems are also discussed. © 2006 Elsevier Inc. All rights reserved.