Dysplasia In Mice Research Articles

Effects of nanoscale electric fields on the histology of liver cell dysplasia.

The current study examined the effects of nanoscale electric fields generated by magneto-electric nanoparticles (MENs) on precancerous liver tissue. A total of30nm MENs synthesized by sol-gel method were tested in vitro on HepG2 cells and in vivo on liver cell dysplasia in mice, which were exposed to 50Hz 2mT for 2weeks, +/- MENs. MENs with alternating field (AF) reversed liver cells dysplastic features. In vitro cytotoxicity assay showed high lethal dose (LD 50) of 1.4mg/ml. We also report on the expression of alpha-fetoprotein and cytochrome C. MEN-generated nanoscale electric fields have significant biological effects on precancerous liver cells.

Defining a critical period in calvarial development for Hedgehog pathway antagonist-induced frontal bone dysplasia in mice

The Hedgehog (Hh) signalling pathway is essential for cellular proliferation and differentiation during embryonic development. Gain and loss of function of Hh signalling are known to result in an array of craniofacial malformations. To determine the critical period for Hh pathway antagonist-induced frontal bone hypoplasia, we examined patterns of dysmorphology caused by Hh signalling inhibition. Pregnant mice received a single oral administration of Hh signalling inhibitor GDC-0449 at 100 mg•kg−1 or 150 mg•kg−1 body weight at preselected time points between embryonic days (E)8.5 and 12.5. The optimal teratogenic concentration of GDC-0449 was determined to be 150 mg•kg−1. Exposure between E9.5 and E10.5 induced frontal bone dysplasia, micrognathia and limb defects, with administration at E10.5 producing the most pronounced effects. This model showed decreased ossification of the frontal bone with downregulation of Hh signalling. The osteoid thickness of the frontal bone was significantly reduced. The amount of neural crest-derived frontal bone primordium was reduced after GDC-0449 exposure owing to a decreased rate of cell proliferation and increased cell death.

Selective inactivation of LGI1 in neuronal precursor cells leads to cortical dysplasia in mice.

Constitutional mutations in Leucine-rich glioma inactivated 1 (LGI1) predispose to an autosomal dominant epilepsy syndrome in humans and germline inactivation of LGI1 in mice leads to early onset seizures. LGI1 is highly expressed in the regions involved in neuronal stem cell generation and migration and detailed analysis of the brains in these mice reveals a subtle cortical dysplasia characterized by hypercellularity in the outer cortical layers. To investigate the cellular origin for this cortical dysplasia, we created mice that allow cell-specific, conditional inactivation of LGI1. Exons 3-4, which contain critical motifs for LGI1 function, were targeted for deletion and, using a CMV-cre mouse strain, global inactivation of LGI1 led to early onset seizures and the same cortical dysplasia seen in the constitutionally null mice. Similarly, inactivation of LGI1 in cells expressing Nestin, expressed primarily in neuronal precursor cells, led to early onset seizures and cortical dysplasia. In contrast, targeting inactivation of LGI1 in cells expressing Gfap, Camk2a, and parvalbumin, did not lead to cortical dysplasia. This strain of mouse, therefore, allows for a more refined investigation of the cell types involved in the cortical dysplasia seen following inactivation of LGI1 and potentially a better understanding of the molecular mechanisms behind LGI1-induced epilepsy.

Integrated Analysis of Mouse and Human Gastric Neoplasms Identifies Conserved microRNA Networks in Gastric Carcinogenesis

microRNAs (miRNAs) are small noncoding RNAs that bind to the 3' untranslated regions of mRNAs to promote their degradation or block their translation. Mice with disruption of the trefoil factor 1 gene (Tff1) develop gastric neoplasms. We studied these mice to identify conserved miRNA networks involved in gastric carcinogenesis. We performed next-generation miRNA sequencing analysis of normal gastric tissues (based on histology) from patients without evidence of gastric neoplasm (n= 64) and from TFF1-knockout mice (n= 22). We validated our findings using 270 normal gastric tissues (including 61 samples from patients without evidence of neoplastic lesions) and 234 gastric tumor tissues from 3 separate cohorts of patients and from mice. We performed molecular and functional assays using cell lines (MKN28, MKN45, STKM2, and AGS cells), gastric organoids, and mice with xenograft tumors. We identified 117 miRNAs that were significantly deregulated in mouse and human gastric tumor tissues compared with nontumor tissues. We validated changes in levels of 6 miRNAs by quantitative real-time polymerase chain reaction analyses of neoplastic gastric tissues from mice (n= 39) and 3 independent patient cohorts (n= 332 patients total). We found levels of MIR135B-5p, MIR196B-5p, and MIR92A-5p to be increased in tumor tissues, whereas levels of MIR143-3p, MIR204-5p, and MIR133-3p were decreased in tumor tissues. Levels of MIR143-3p were reduced not only in gastric cancer tissues but also in normal tissues adjacent to tumors in humans and low-grade dysplasia in mice. Transgenic expression of MIR143-3p in gastric cancer cell lines reduced their proliferation and restored their sensitivity to cisplatin. AGS cells with stable transgenic expression of MIR143-3p grew more slowly as xenograft tumors in mice thancontrol AGS cells; tumor growth from AGS cells that expressed MIR143-3p, but not control cells, was sensitive to cisplatin. We identified and validated bromodomain containing 2 (BRD2) as a direct target of MIR143-3p; increased levels of BRD2 in gastric tumors was associated with shorter survival times for patients. In an analysis of miRNA profiles of gastric tumors from mice and human patients, we identified a conserved signature associated with the early stages of gastric tumorigenesis. Strategies to restore MIR143-3p or inhibit BRD2 might be developed for treatment of gastric cancer.

Chronic inflammation triggered by the NLRP3 inflammasome in myeloid cells promotes growth plate dysplasia by mesenchymal cells

Skeletal complications are common features of neonatal-onset multisystem inflammatory disease (NOMID), a disorder caused by NLRP3-activating mutations. NOMID mice in which NLRP3 is activated globally exhibit several characteristics of the human disease, including systemic inflammation and cartilage dysplasia, but the mechanisms of skeletal manifestations remain unknown. In this study, we find that activation of NLRP3 in myeloid cells, but not mesenchymal cells triggers chronic inflammation, which ultimately, causes growth plate and epiphyseal dysplasia in mice. These responses are IL-1 signaling-dependent, but independent of PARP1, which also functions downstream of NLRP3 and regulates skeletal homeostasis. Mechanistically, inflammation causes severe anemia and hypoxia in the bone environment, yet down-regulates the HIF-1α pathway in chondrocytes, thereby promoting the demise of these cells. Thus, activation of NLRP3 in hematopoietic cells initiates IL-1β-driven paracrine cascades, which promote abnormal growth plate development in NOMID mice.

Fra‐1 Signaling both in Lung Resident and Myeloid Cells Is Required for Hyperoxia‐Induced Bronchopulmonary Dysplasia in Mice

RationaleBronchopulmonary dysplasia (BPD) is a chronic disease of preterm babies. While current clinical care has largely improved the survival of preterm babies, incidence of BPD and poor health outcomes associated with this disease remain major clinical problems. Lung cellular stress and inflammation have been linked to BPD pathogenesis in humans and experimental models. Fra‐1 (Fosl1) is a heterodimeric partner of Jun/AP‐1 transcription factor and is known to differentially regulate gene expression involved in various cellular processes, including cellular stress and inflammation, which are known to contribute to BPD pathogenesis. We examine whether Fra‐1/AP1 transcription factor is a critical modulator of BPD pathogenesis.MethodsTo test our hypothesis, we have used Fra‐1 floxed (Fra‐1FF or WT) and Fra‐1 null (Fra‐1−/−) mice, and mice with a conditional deletion of Fra‐1 in myeloid cells. Fra‐1 in myeloid cells was deleted by crossing Fra‐1 floxed mice with a transgenic mice bearing Cre under the control of the LyzM promoter (hereafter notated as Fra‐1−/−LysM mice). Fra‐1F/F, Fra‐1−/− and Fra‐1−/−LysM pups (1‐day‐old) (n = 3–5/group) with mothers were exposed to 95% O2 for 72 h and allowed to recover in room air for two weeks. Mice were sacrificed, the left lungs were fixed, sectioned, and stained with H&E to visualize alveolar simplification and to quantify mean chord length (MCL) by morphometry analysis. Fra‐1F/F, Fra‐1−/− and Fra‐1−/−LysM pups exposed to 95% O2 for 72 h were immediately sacrificed and the left lung was collected for RNA isolation, and right lobe was used for protein analysis. Expression levels of several pro‐inflammatory mediators and antioxidant genes in the lungs of WT (Fra‐1F/F) and Fra‐1−/− pups exposed to 95% O2 for 72 h were analyzed by qPCR.ResultsWe found increased expression of Fra‐1 in the lungs of pups exposed to hypeorixia. Fra‐1 null pups showed markedly reduced levels of hyperoxia‐induced alveoli simplification (or MCL) as compared to WT pups (33% vs 6%, WT vs Fra1−/−), suggesting that Fra‐1 signaling is required for BPD development. Alveoli simplification in the lungs of Fra‐1−/−LysM pups exposed to hyperoxia was 12% lower than WT counterparts (33% vs 21%, WT vs Fra‐1−/−LysM), suggesting that Fra‐1 signaling in myeloid cells partly contributes to BPD. The expression levels of inflammatory mediators (IL‐1β, TNF‐α, and IL‐6) were markedly lower in the lungs of Fra1−/− pups exposed to 72 h hyperoxia than in WT counterparts. Surprisingly, antioxidant gene expression in the lungs of hyperoxia exposed Fra‐1−/− pups was also markedly lower than in WT counterparts.ConclusionOur findings suggest that Fra‐1 signaling both in lung resident and inflammatory cells play an important role in promoting hyperoxia‐induced BPD, and that Fra‐1 regulates this process at least in part by modulating inflammatory cytokine and antioxidant gene expression in the lung.Support or Funding InformationPartly supported by NIH grants, HL66109 and ES11863 (to SPR)

Tu1928 - Peptostreptococcus Anaerobius Induces Intracellular Cholesterol Biosynthesis in Colon Cells to Induce Proliferation and Causes Dysplasia in Mice

Tu1928 - Peptostreptococcus Anaerobius Induces Intracellular Cholesterol Biosynthesis in Colon Cells to Induce Proliferation and Causes Dysplasia in Mice

Peptostreptococcus anaerobius Induces Intracellular Cholesterol Biosynthesis in Colon Cells to Induce Proliferation and Causes Dysplasia in Mice

Stool samples from patients with colorectal cancer (CRC) have a higher abundance of Peptostreptococcus anaerobius than stool from individuals without CRC, based on metagenome sequencing. We investigated whether P anaerobius contributes to colon tumor formation in mice and its possible mechanisms of carcinogenesis. We performed quantitative polymerase chain reaction analyses to measure P anaerobius in 112 stool samples and 255 colon biopsies from patients with CRC or advanced adenoma and from healthy individuals (controls) undergoing colonoscopy examination at hospitals in Hong Kong and Beijing. C57BL/6 mice were given broad-spectrum antibiotics, followed by a single dose of azoxymethane, to induce colon tumor formation. Three days later, mice were given P anaerobius or Esherichia coli MG1655 (control bacteria), via gavage, for 6 weeks. Some mice were also given the nicotinamide adenine dinucleotide phosphate oxidase inhibitor apocynin. Intestine tissues were collected and analyzed histologically. The colon epithelial cell line NCM460 and colon cancer cell lines HT-29 and Caco-2 were exposed to P anaerobius or control bacteria; cells were analyzed by immunoblot, proliferation, and bacterial attachment analyses and compared in gene expression profiling studies. Gene expression was knocked down in these cell lines with small interfering RNAs. P anaerobius was significantly enriched in stool samples from patients with CRC and in biopsies from patients with colorectal adenoma or CRC compared with controls. Mice depleted of bacteria and exposed to azoxymethane and P anaerobius had a higher incidence of intestinal dysplasia (63%) compared with mice not given the bacteria (8.3%; P < .01). P anaerobius mainly colonized the colon compared with the rest of the intestine. Colon cells exposed to P anaerobius had significantly higher levels of proliferation than control cells. We found genes that regulate cholesterol biosynthesis, Toll-like receptor (TLR) signaling, and AMP-activated protein kinase signaling to be significantly up-regulated in cells exposed to P anaerobius. Total cholesterol levels were significantly increased in colon cell lines exposed to P anaerobius via activation of sterol regulatory element-binding protein 2. P anaerobius interacted with TLR2 and TLR4 to increase intracellular levels of reactive oxidative species, which promoted cholesterol synthesis and cell proliferation. Depletion of reactive oxidative species by knockdown of TLR2 or TLR4, or incubation of cells with an antioxidant, prevented P anaerobius from inducing cholesterol biosynthesis and proliferation. Levels of P anaerobius are increased in human colon tumor tissues and adenomas compared with non-tumor tissues; this bacteria increases colon dysplasia in a mouse model of CRC. Panaerobius interacts with TLR2 and TLR4 on colon cells to increase levels of reactive oxidative species, which promotes cholesterol synthesis and cell proliferation.

Plecanatide-mediated activation of guanylate cyclase-C suppresses inflammation-induced colorectal carcinogenesis in Apc+/Min-FCCC mice.

AIMTo evaluate the effect of orally administered plecanatide on colorectal dysplasia in Apc+/Min-FCCC mice with dextran sodium sulfate (DSS)-induced inflammation.METHODSInflammation driven colorectal carcinogenesis was induced in Apc+/Min-FCCC mice by administering DSS in their drinking water. Mice were fed a diet supplemented with plecanatide (0-20 ppm) and its effect on the multiplicity of histopathologically confirmed polypoid, flat and indeterminate dysplasia was evaluated. Plecanatide-mediated activation of guanylate cyclase-C (GC-C) signaling was assessed in colon tissues by measuring cyclic guanosine monophosphate (cGMP) by ELISA, protein kinase G-II and vasodilator stimulated phosphoprotein by immunoblotting. Ki-67, c-myc and cyclin D1 were used as markers of proliferation. Cellular levels and localization of β-catenin in colon tissues were assessed by immunoblotting and immunohistochemistry, respectively. Uroguanylin (UG) and GC-C transcript levels were measured by quantitative reverse transcription polymerase chain reaction (RT-PCR). A mouse cytokine array panel was used to detect cytokines in the supernatant of colon explant cultures.RESULTSOral treatment of Apc+/MinFCCC mice with plecanatide produced a statistically significant reduction in the formation of inflammation-driven polypoid, flat and indeterminate dysplasias. This anti-carcinogenic activity of plecanatide was accompanied by activation of cGMP/GC-C signaling mediated inhibition of Wnt/β-catenin signaling and reduced proliferation. Plecanatide also decreased secretion of pro-inflammatory cytokines (IL-6, IL1 TNF), chemokines (MIP-1, IP-10) and growth factors (GCSF and GMCSF) from colon explants derived from mice with acute DSS-induced inflammation. The effect of plecanatide-mediated inhibition of inflammation/dysplasia on endogenous expression of UG and GC-C transcripts was measured in intestinal tissues. Although GC-C expression was not altered appreciably, a statistically significant increase in the level of UG transcripts was detected in the proximal small intestine and colon, potentially due to a reduction in intestinal inflammation and/or neoplasia. Taken together, these results suggest that reductions in endogenous UG, accompanied by dysregulation in GC-C signaling, may be an early event in inflammation-promoted colorectal neoplasia; an event that can potentially be ameliorated by prophylactic intervention with plecanatide.CONCLUSIONThis study provides the first evidence that orally administered plecanatide reduces the multiplicity of inflammation-driven colonic dysplasia in mice, demonstrating the utility for developing GC-C agonists as chemopreventive agents.

Deletion of IQGAP1 promotes Helicobacter pylori-induced gastric dysplasia in mice and acquisition of cancer stem cell properties in vitro.

Helicobacter pylori infection is responsible for gastric carcinogenesis but host factors are also implicated. IQGAP1, a scaffolding protein of the adherens junctions interacting with E-cadherin, regulates cellular plasticity and proliferation. In mice, IQGAP1 deficiency leads to gastric hyperplasia. The aim of this study was to elucidate the consequences of IQGAP1 deletion on H. pylori-induced gastric carcinogenesis.Transgenic mice deleted for iqgap1 and WT littermates were infected with Helicobacter sp., and histopathological analyses of the gastric mucosa were performed. IQGAP1 and E-cadherin expression was evaluated in gastric tissues and in gastric epithelial cell lines in response to H. pylori infection. The consequences of IQGAP1 deletion on gastric epithelial cell behaviour and on the acquisition of cancer stem cell (CSC)-like properties were evaluated. After one year of infection, iqgap1+/- mice developed more preneoplastic lesions and up to 8 times more gastro-intestinal neoplasia (GIN) than WT littermates. H. pylori infection induced IQGAP1 and E-cadherin delocalization from cell-cell junctions. In vitro, knock-down of IQGAP1 favoured the acquisition of a mesenchymal phenotype and CSC-like properties induced by H. pylori infection.Our results indicate that alterations in IQGAP1 signalling promote the emergence of CSCs and gastric adenocarcinoma development in the context of an H. pylori infection.

HIF-1α Plays a Critical Role in the Gestational Sidestream Smoke-Induced Bronchopulmonary Dysplasia in Mice.

RationaleSmoking during pregnancy increases the risk of bronchopulmonary dysplasia (BPD) and, in mice, gestational exposure to sidestream cigarette smoke (SS) induces BPD-like condition characterized by alveolar simplification, impaired angiogenesis, and suppressed surfactant protein production. Normal fetal development occurs in a hypoxic environment and nicotinic acetylcholine receptors (nAChRs) regulate the hypoxia-inducible factor (HIF)-1α that controls apoptosis and angiogenesis. To understand SS-induced BPD, we hypothesized that gestational SS affected alveolar development through HIF-1α.MethodsPregnant BALB/c mice were exposed to air (control) or SS throughout the gestational period and the 7-day-old lungs of the progeny were examined.ResultsGestational SS increased apoptosis of alveolar and airway epithelial cells. This response was associated with increased alveolar volumes, higher levels of proapoptotic factors (FOXO3a, HIPK2, p53, BIM, BIK, and BAX) and the antiangiogenic factor (GAX), and lower levels of antiapoptotic factors (Akt-PI3K, NF-κB, HIF-1α, and Bcl-2) in the lung. Although gestational SS increased the cells containing the proangiogenic bombesin-like-peptide, it markedly decreased the expression of its receptor GRPR in the lung. The effects of SS on apoptosis were attenuated by the nAChR antagonist mecamylamine.ConclusionsGestational SS-induced BPD is potentially regulated by nAChRs and associated with downregulation of HIF-1α, increased apoptosis of epithelial cells, and increased alveolar volumes. Thus, in mice, exposure to sidestream tobacco smoke during pregnancy promotes BPD-like condition that is potentially mediated through the nAChR/HIF-1α pathway.

Bak deletion stimulates gastric epithelial proliferation and enhances Helicobacter felis-induced gastric atrophy and dysplasia in mice.

Helicobacter infection causes a chronic superficial gastritis that in some cases progresses via atrophic gastritis to adenocarcinoma. Proapoptotic bak has been shown to regulate radiation-induced apoptosis in the stomach and colon and also susceptibility to colorectal carcinogenesis in vivo. Therefore we investigated the gastric mucosal pathology following H. felis infection in bak-null mice at 6 or 48 wk postinfection. Primary gastric gland culture from bak-null mice was also used to assess the effects of bak deletion on IFN-γ-, TNF-α-, or IL-1β-induced apoptosis. bak-null gastric corpus glands were longer, had increased epithelial Ki-67 expression, and contained fewer parietal and enteroendocrine cells compared with the wild type (wt). In wt mice, bak was expressed at the luminal surface of gastric corpus glands, and this increased 2 wk post-H. felis infection. Apoptotic cell numbers were decreased in bak-null corpus 6 and 48 wk following infection and in primary gland cultures following cytokine administration. Increased gastric epithelial Ki-67 labeling index was observed in C57BL/6 mice after H. felis infection, whereas no such increase was detected in bak-null mice. More severe gastric atrophy was observed in bak-null compared with C57BL/6 mice 6 and 48 wk postinfection, and 76% of bak-null compared with 25% of C57BL/6 mice showed evidence of gastric dysplasia following long-term infection. Collectively, bak therefore regulates gastric epithelial cell apoptosis, proliferation, differentiation, mucosal thickness, and susceptibility to gastric atrophy and dysplasia following H. felis infection.

Tracheal Dysplasia Precedes Bronchial Dysplasia in Mouse Model of N-Nitroso Trischloroethylurea Induced Squamous Cell Lung Cancer

Squamous cell lung cancer (SCC) is the second leading cause of lung cancer death in the US and has a 5-year survival rate of only 16%. Histological changes in the bronchial epithelium termed dysplasia are precursors to invasive SCC. However, the cellular mechanisms that cause dysplasia are unknown. To fill this knowledge gap, we used topical application of N-nitroso-tris chloroethylurea (NTCU) for 32 weeks to induce squamous dysplasia and SCC in mice. At 32 weeks the predominant cell type in the dysplastic airways was Keratin (K) 5 and K14 expressing basal cells. Notably, basal cells are extremely rare in the normal mouse bronchial epithelium but are abundant in the trachea. We therefore evaluated time-dependent changes in tracheal and bronchial histopathology after NTCU exposure (4, 8, 12, 16, 25 and 32 weeks). We show that tracheal dysplasia occurs significantly earlier than that of the bronchial epithelium (12 weeks vs. 25 weeks). This was associated with increased numbers of K5+/K14+ tracheal basal cells and a complete loss of secretory (Club cell secretory protein expressing CCSP+) and ciliated cells. TUNEL staining of NTCU treated tissues confirmed that the loss of CCSP+ and ciliated cells was not due to apoptosis. However, mitotic index (measured by bromodeoxyuridine incorporation) showed that NTCU treatment increased proliferation of K5+ basal cells in the trachea, and altered bronchial mitotic population from CCSP+ to K5+ basal cells. Thus, we demonstrate that NTCU-induced lung epithelial dysplasia starts in the tracheal epithelium, and is followed by basal cell metaplasia of the bronchial epithelium. This analysis extends our knowledge of the NTCU-SCC model by defining the early changes in epithelial cell phenotypes in distinct airway locations, and this may assist in identifying new targets for future chemoprevention studies.

GlcUAβ1–3Galβ1–3Galβ1–4Xyl(2-O-phosphate) Is the Preferred Substrate for Chondroitin N-Acetylgalactosaminyltransferase-1

A deficiency in chondroitin N-acetylgalactosaminyltransferase-1 (ChGn-1) was previously shown to reduce the number of chondroitin sulfate (CS) chains, leading to skeletal dysplasias in mice, suggesting that ChGn-1 regulates the number of CS chains for normal cartilage development. Recently, we demonstrated that 2-phosphoxylose phosphatase (XYLP) regulates the number of CS chains by dephosphorylating the Xyl residue in the glycosaminoglycan-protein linkage region of proteoglycans. However, the relationship between ChGn-1 and XYLP in controlling the number of CS chains is not clear. In this study, we for the first time detected a phosphorylated tetrasaccharide linkage structure, GlcUAβ1-3Galβ1-3Galβ1-4Xyl(2-O-phosphate), in ChGn-1(-/-) growth plate cartilage but not in ChGn-2(-/-) or wild-type growth plate cartilage. In contrast, the truncated linkage tetrasaccharide GlcUAβ1-3Galβ1-3Galβ1-4Xyl was detected in wild-type, ChGn-1(-/-), and ChGn-2(-/-) growth plate cartilage. Consistent with the findings, ChGn-1 preferentially transferred N-acetylgalactosamine to the phosphorylated tetrasaccharide linkage in vitro. Moreover, ChGn-1 and XYLP interacted with each other, and ChGn-1-mediated addition of N-acetylgalactosamine was accompanied by rapid XYLP-dependent dephosphorylation during formation of the CS linkage region. Taken together, we conclude that the phosphorylated tetrasaccharide linkage is the preferred substrate for ChGn-1 and that ChGn-1 and XYLP cooperatively regulate the number of CS chains in growth plate cartilage.

Abstract 4270: The use of mutational signatures in identifying carcinogen exposure

Abstract Aristolochic acid (AA), a natural product of Aristolochia plants found in herbal remedies and health supplements, is a Group 1 carcinogen that can cause nephrotoxicity and upper urinary tract urothelial cell carcinoma (UCC). Whole genome and exome analysis of 9 AA-associated UCCs revealed a strikingly high somatic mutation rate (150 mutations/Mb), exceeding smoking-associated lung cancer (8 mutations/Mb) and ultraviolet radiation-associated melanoma (111 mutations/Mb). The AA-UCC mutational signature was characterized by A:T to T:A transversions at the sequence motif A[C|T]AGG, located primarily on non-transcribed strands. AA-induced mutations were also significantly enriched at splice sites, suggesting a role for splice site mutations in UCC pathogenesis. RNA sequencing of AA-UCC confirmed a general up-regulation of nonsense-mediated decay machinery components and aberrant splicing events associated with splice site mutations. We observed a high frequency of somatic mutations in chromatin modifiers, particularly KDM6A, in AA-UCC, and demonstrated the sufficiency of AA to induce renal dysplasia in mice and reproduced the AA mutational signature in experimentally treated human renal tubular cells. Finally, exploring other malignancies not previously associated with AA, we screened 93 hepatocellular carcinoma genomes/exomes and identified AA-like mutational signatures in eleven. Our study highlights a unique genome-wide AA mutational signature, and the potential use of mutation signatures as “molecular fingerprints” for interrogating high-throughput cancer genome data, to infer previous carcinogen exposures. Citation Format: Song Ling Poon, See-Tong Pang, John R. McPherson, Steven G. Rozen, Patrick Tan, Bin Tean Teh. The use of mutational signatures in identifying carcinogen exposure. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4270. doi:10.1158/1538-7445.AM2014-4270

Lactoferrin deficiency promotes colitis-associated colorectal dysplasia in mice.

Nonresolving inflammatory processes affect all stages of carcinogenesis. Lactoferrin, a member of the transferrin family, is involved in the innate immune response and anti-inflammatory, anti-microbial, and anti-tumor activities. We previously found that lactoferrin is significantly down-regulated in specimens of nasopharyngeal carcinoma (NPC) and negatively associated with tumor progression, metastasis, and prognosis of patients with NPC. Additionally, lactoferrin expression levels are decreased in colorectal cancer as compared with normal tissue. Lactoferrin levels are also increased in the various phases of inflammation and dysplasia in an azoxymethane–dextran sulfate sodium (AOM-DSS) model of colitis-associated colon cancer (CAC). We thus hypothesized that the anti-inflammatory function of lactoferrin may contribute to its anti-tumor activity. Here we generated a new Lactoferrin knockout mouse model in which the mice are fertile, develop normally, and display no gross morphological abnormalities. We then challenged these mice with chemically induced intestinal inflammation to investigate the role of lactoferrin in inflammation and cancer development. Lactoferrin knockout mice demonstrated a great susceptibility to inflammation-induced colorectal dysplasia, and this characteristic may be related to inhibition of NF-κB and AKT/mTOR signaling as well as regulation of cell apoptosis and proliferation. Our results suggest that the protective roles of lactoferrin in colorectal mucosal immunity and inflammation-related malignant transformation, along with a deficiency in certain components of the innate immune system, may lead to serious consequences under conditions of inflammatory insult.

Artemisinin and curcumin inhibit Drosophila brain tumor, prolong life span, and restore locomotor activity.

Deletion of tumor suppressor gene, lethal(2)giant larvae [l(2)gl], leads to brain tumor in Drosophila melanogaster at larval stage of development and severe brain dysplasia in mice. We have studied the effect of two potential antitumor drugs artemisinin and curcumin in the perspective of inhibiting l(2)gl brain tumor. Efficacies of these drugs are characterized morphologically by measuring brain sizes of untreated and treated larvae on the basis of tumor inhibition and anatomically by looking at the cellular patterning via antibody staining of the third instar Drosophila larval brains. Behavioral experiments were done in form of locomotion to correlate tumor inhibition with the revival of brain function and longevity assays to assess general health span. It was observed that both drugs show antitumor properties individually and in combination when larvae were treated with these drugs. We also found evidence for reactive oxygen species-mediated action of these drugs. Both the drugs when treated individually or together show better median life span and locomotory response. Although the efficacies of various treatments varied, overall, the positive effects of artemisinin and curcumin demonstrate a potential applicability of these drugs against brain tumor in higher organisms. It also paves a way for a simpler model system for screening such natural products for antitumor property.

Su1114 Photodynamic Diagnosis of Colitis-Associated Dysplasia in Patients With Ulcerative Colitis by Visualization Following Oral 5-Aminolevulinic Acid Sensitization, Compared With the Findings in a Model of Colitis-Associated Dysplasia in Mice

Su1114 Photodynamic Diagnosis of Colitis-Associated Dysplasia in Patients With Ulcerative Colitis by Visualization Following Oral 5-Aminolevulinic Acid Sensitization, Compared With the Findings in a Model of Colitis-Associated Dysplasia in Mice

Tu1174 Attenuated Progression of Gastric Dysplasia by Administration of Green Tea Catechins in INS-GAS Mice

Backgrounds: Green tea catechins (GTCs) have been shown to provide anti-carcinogenic effects in several laboratory studies. Although the effect of green tea consumption on the risk of gastric cancer has been studied in epidemiological studies, the results remain controversial. The anti-carcinogenic mechanisms of GTCs have not been fully evaluated in rodent models of gastric cancer. Previous reports indicate that transgenic INS-GASmice are hypergastrinemic and induced corpus gastritis and eventually developed gastric cancer at over 20 months old. Aim: The aim of our study was to investigate the molecular effects of GTCs on gastritis and premalignant lesions in INS-GAS mice. Methods: Thirty eight, male INSGAS mice on a FVB background were used in this study. Mice were divided into two groups: A GTCs group and control. Mice in the GTCs group were allowed free access to drinking water containing 2000ppm of GTCs from the age of 9 weeks. Mice were euthanized at the age of 13 or 37 weeks. Histological scores (inflammation and dysplasia) were graded on a blinded basis by pathologist on a scale from 0 to 4. The mRNA expressions of IFN-gamma and TNF-alpha in the corpus of stomachs were evaluated using quantitative PCR. Statistical analysis was performed using a Mann-Whitney U test. Results: Body weights in mice supplemented with GTCs were significantly lower than that of controls at both 13 and 37 weeks of age (p < 0.05). Treated and control mice developed minimal to mild corpus gastritis, although dysplasia was evident. The histological score of dysplasia in mice supplemented with GTCs was significantly lower than that in controls at 37 weeks of age (p < 0.05). GTCs supplementation did not affect the inflammation score. The mRNA level of IFN-gamma in mice administrated GTCs was significantly lower than that of controls at 13 weeks of age (p < 0.05), although it did not reach statistical significance at 34 weeks of age (p = 0.056). GTCs supplementation did not alter TNF-alpha mRNA expression levels between both groups. Conclusions: Administration of GTCs attenuated progression of gastric dysplasia in INS-GAS male mice. These results demonstrated that IFN-gamma dependent mechanisms may be involved in the protective role of GTCs in development of gastritis and premalignant lesions in this rodent model.

Tu1175 The Protective Influence of Vitamin D in Hepatocellular Carcinoma

Backgrounds: Green tea catechins (GTCs) have been shown to provide anti-carcinogenic effects in several laboratory studies. Although the effect of green tea consumption on the risk of gastric cancer has been studied in epidemiological studies, the results remain controversial. The anti-carcinogenic mechanisms of GTCs have not been fully evaluated in rodent models of gastric cancer. Previous reports indicate that transgenic INS-GASmice are hypergastrinemic and induced corpus gastritis and eventually developed gastric cancer at over 20 months old. Aim: The aim of our study was to investigate the molecular effects of GTCs on gastritis and premalignant lesions in INS-GAS mice. Methods: Thirty eight, male INSGAS mice on a FVB background were used in this study. Mice were divided into two groups: A GTCs group and control. Mice in the GTCs group were allowed free access to drinking water containing 2000ppm of GTCs from the age of 9 weeks. Mice were euthanized at the age of 13 or 37 weeks. Histological scores (inflammation and dysplasia) were graded on a blinded basis by pathologist on a scale from 0 to 4. The mRNA expressions of IFN-gamma and TNF-alpha in the corpus of stomachs were evaluated using quantitative PCR. Statistical analysis was performed using a Mann-Whitney U test. Results: Body weights in mice supplemented with GTCs were significantly lower than that of controls at both 13 and 37 weeks of age (p < 0.05). Treated and control mice developed minimal to mild corpus gastritis, although dysplasia was evident. The histological score of dysplasia in mice supplemented with GTCs was significantly lower than that in controls at 37 weeks of age (p < 0.05). GTCs supplementation did not affect the inflammation score. The mRNA level of IFN-gamma in mice administrated GTCs was significantly lower than that of controls at 13 weeks of age (p < 0.05), although it did not reach statistical significance at 34 weeks of age (p = 0.056). GTCs supplementation did not alter TNF-alpha mRNA expression levels between both groups. Conclusions: Administration of GTCs attenuated progression of gastric dysplasia in INS-GAS male mice. These results demonstrated that IFN-gamma dependent mechanisms may be involved in the protective role of GTCs in development of gastritis and premalignant lesions in this rodent model.