JoVE Visualize What is visualize?
Stop Reading. Start Watching.
Advanced Search
Stop Reading. Start Watching.
Regular Search
Find video protocols related to scientific articles indexed in Pubmed.
Nuclear Factor of Activated T-cell Activity Is Associated with Metastatic Capacity in Colon Cancer.
Cancer Res.
PUBLISHED: 10-15-2014
Show Abstract
Hide Abstract
Metastatic recurrence is the leading cause of cancer-related deaths in patients with colorectal carcinoma. To capture the molecular underpinnings for metastasis and tumor progression, we performed integrative network analysis on 11 independent human colorectal cancer gene expression datasets and applied expression data from an immunocompetent mouse model of metastasis as an additional filter for this biologic process. In silico analysis of one metastasis-related coexpression module predicted nuclear factor of activated T-cell (NFAT) transcription factors as potential regulators for the module. Cells selected for invasiveness and metastatic capability expressed higher levels of NFATc1 as compared with poorly metastatic and less invasive parental cells. We found that inhibition of NFATc1 in human and mouse colon cancer cells resulted in decreased invasiveness in culture and downregulation of metastasis-related network genes. Overexpression of NFATc1 significantly increased the metastatic potential of colon cancer cells, whereas inhibition of NFATc1 reduced metastasis growth in an immunocompetent mouse model. Finally, we found that an 8-gene signature comprising genes upregulated by NFATc1 significantly correlated with worse clinical outcomes in stage II and III colorectal cancer patients. Thus, NFATc1 regulates colon cancer cell behavior and its transcriptional targets constitute a novel, biologically anchored gene expression signature for the identification of colon cancers with high risk of metastatic recurrence. Cancer Res; 74(23); 1-11. ©2014 AACR.
Related JoVE Video
Concerted loss of TGF?-mediated proliferation control and E-cadherin disrupts epithelial homeostasis and causes oral squamous cell carcinoma.
Carcinogenesis
PUBLISHED: 09-18-2014
Show Abstract
Hide Abstract
Although the etiology of squamous cell carcinomas of the oral mucosa is well understood, the cellular origin and the exact molecular mechanisms leading to their formation are not. Previously, we observed the coordinated loss of E-cadherin (CDH1) and transforming growth factor beta receptor II (TGFBR2) in esophageal squamous tumors. To investigate if the coordinated loss of Cdh1 and Tgfbr2 is sufficient to induce tumorigenesis in vivo, we developed two mouse models targeting ablation of both genes constitutively or inducibly in the oral-esophageal epithelium. We show that the loss of both Cdh1 and Tgfbr2 in both models is sufficient to induce squamous cell carcinomas with animals succumbing to the invasive disease by 18 months of age. Advanced tumors have the ability to invade regional lymph nodes and to establish distant pulmonary metastasis. The mouse tumors showed molecular characteristics of human tumors such as overexpression of Cyclin D1. We addressed the question whether TGF? signaling may target known stem cell markers and thereby influence tumorigenesis. From our mouse and human models, we conclude that TGF? signaling regulates key aspects of stemness and quiescence in vitro and in vivo. This provides a new explanation for the importance of TGF? in mucosal homeostasis.
Related JoVE Video
The Impact of Tumor Deposits on Colonic Adenocarcinoma AJCC TNM Staging and Outcome.
Am. J. Surg. Pathol.
PUBLISHED: 09-18-2014
Show Abstract
Hide Abstract
The definition of tumor deposits (TDs) in colonic adenocarcinoma has been modified in different editions of the AJCC/TNM staging system. Studies have shown that the presence of TD is associated with advanced tumor growth and poor prognosis. Most of these data were obtained in patients with simultaneous lymph node (LN) metastases. Reports focusing on the impact of TD in patients without LN metastasis are limited. We retrospectively restaged all right-sided colonic adenocarcinomas over a 10-year period using criteria from the fifth, sixth, and seventh AJCC edition. We compared the number of tumor nodule interpreted as LN and TD in each edition and evaluated the stage migration caused by TD definition change. We then assessed clinical significance of TD in the AJCC seventh edition by comparing 5-year overall survival of N1c patients versus other N category (N0, N1, N2) patients with similar T and M status. We showed that the average number of tumor nodules interpreted as LNs per case and the number of cases with positive LNs were significantly decreased with the seventh edition compared with fifth/sixth; however, numbers of cases with TDs and <12 LNs were significantly increased with the seventh edition compared with fifth/sixth. These changes, however, resulted in minimal effects on the final stage grouping. Our survival analysis showed that N1c patients had significantly worse survival compared with N0 patients. Although not statistically significant, the hazard ratios indicated that the N1c group might have worse survival than the N1 group and better survival than the N2 group. Therefore, we conclude that TDs predict patient outcome at least similarly to positive LNs.
Related JoVE Video
Viral Suppression and Cirrhosis Regression with Tenofovir Disoproxil Fumarate in Asians with Chronic Hepatitis B.
Dig. Dis. Sci.
PUBLISHED: 09-02-2014
Show Abstract
Hide Abstract
Chronic hepatitis B (CHB) is a major public health concern, particularly in endemic areas like Asia-Pacific. Sustained virologic suppression correlates with regression of histologic fibrosis and cirrhosis.
Related JoVE Video
Activation of ?-catenin signalling by TFF1 loss promotes cell proliferation and gastric tumorigenesis.
Gut
PUBLISHED: 08-08-2014
Show Abstract
Hide Abstract
In this study, we investigated the role of Trefoil factor 1 (TFF1) in regulating cell proliferation and tumour development through ?-catenin signalling using in vivo and in vitro models of gastric tumorigenesis.
Related JoVE Video
Systems modeling of the role of interleukin-21 in the maintenance of effector CD4+ T cell responses during chronic Helicobacter pylori infection.
MBio
PUBLISHED: 07-24-2014
Show Abstract
Hide Abstract
The development of gastritis during Helicobacter pylori infection is dependent on an activated adaptive immune response orchestrated by T helper (Th) cells. However, the relative contributions of the Th1 and Th17 subsets to gastritis and control of infection are still under investigation. To investigate the role of interleukin-21 (IL-21) in the gastric mucosa during H. pylori infection, we combined mathematical modeling of CD4(+) T cell differentiation with in vivo mechanistic studies. We infected IL-21-deficient and wild-type mice with H. pylori strain SS1 and assessed colonization, gastric inflammation, cellular infiltration, and cytokine profiles. Chronically H. pylori-infected IL-21-deficient mice had higher H. pylori colonization, significantly less gastritis, and reduced expression of proinflammatory cytokines and chemokines compared to these parameters in infected wild-type littermates. These in vivo data were used to calibrate an H. pylori infection-dependent, CD4(+) T cell-specific computational model, which then described the mechanism by which IL-21 activates the production of interferon gamma (IFN-?) and IL-17 during chronic H. pylori infection. The model predicted activated expression of T-bet and ROR?t and the phosphorylation of STAT3 and STAT1 and suggested a potential role of IL-21 in the modulation of IL-10. Driven by our modeling-derived predictions, we found reduced levels of CD4(+) splenocyte-specific tbx21 and rorc expression, reduced phosphorylation of STAT1 and STAT3, and an increase in CD4(+) T cell-specific IL-10 expression in H. pylori-infected IL-21-deficient mice. Our results indicate that IL-21 regulates Th1 and Th17 effector responses during chronic H. pylori infection in a STAT1- and STAT3-dependent manner, therefore playing a major role controlling H. pylori infection and gastritis. Importance: Helicobacter pylori is the dominant member of the gastric microbiota in more than 50% of the world's population. H. pylori colonization has been implicated in gastritis and gastric cancer, as infection with H. pylori is the single most common risk factor for gastric cancer. Current data suggest that, in addition to bacterial virulence factors, the magnitude and types of immune responses influence the outcome of colonization and chronic infection. This study uses a combined computational and experimental approach to investigate how IL-21, a proinflammatory T cell-derived cytokine, maintains the chronic proinflammatory T cell immune response driving chronic gastritis during H. pylori infection. This research will also provide insight into a myriad of other infectious and immune disorders in which IL-21 is increasingly recognized to play a central role. The use of IL-21-related therapies may provide treatment options for individuals chronically colonized with H. pylori as an alternative to aggressive antibiotics.
Related JoVE Video
Fluorescence-based endoscopic imaging of Thomsen-Friedenreich antigen to improve early detection of colorectal cancer.
Int. J. Cancer
PUBLISHED: 05-20-2014
Show Abstract
Hide Abstract
Thomsen-Friedenreich (TF) antigen belongs to the mucin-type tumor-associated carbohydrate antigen. Notably, TF antigen is overexpressed in colorectal cancer (CRC) but is rarely expressed in normal colonic tissue. Increased TF antigen expression is associated with tumor invasion and metastasis. In this study, we sought to validate a novel nanobeacon for imaging TF-associated CRC in a preclinical animal model. We developed and characterized the nanobeacon for use with fluorescence colonoscopy. In vivo imaging was performed on an orthotopic rat model of CRC. Both white light and fluorescence colonoscopy methods were utilized to establish the ratio-imaging index for the probe. The nanobeacon exhibited specificity for TF-associated cancer. Fluorescence colonoscopy using the probe can detect lesions at the stage which is not readily confirmed by conventional visualization methods. Further, the probe can report the dynamic change of TF expression as tumor regresses during chemotherapy. Data from this study suggests that fluorescence colonoscopy can improve early CRC detection. Supplemented by the established ratio-imaging index, the probe can be used not only for early detection, but also for reporting tumor response during chemotherapy. Furthermore, since the data obtained through in vivo imaging confirmed that the probe was not absorbed by the colonic mucosa, no registered toxicity is associated with this nanobeacon. Taken together, these data demonstrate the potential of this novel probe for imaging TF antigen as a biomarker for the early detection and prediction of the progression of CRC at the molecular level.
Related JoVE Video
Controlled colonic insufflation by a remotely triggered capsule for improved mucosal visualization.
Endoscopy
PUBLISHED: 05-20-2014
Show Abstract
Hide Abstract
Capsule endoscopy is an attractive alternative to colorectal cancer screening by conventional colonoscopy, but is currently limited by compromised mucosal visibility because of the lack of safe, controlled colonic insufflation. We have therefore developed a novel system of untethered, wireless-controlled carbon dioxide (CO2) insufflation for use in colonic capsule endoscopy, which this study aims to assess in vivo.
Related JoVE Video
Inducible loss of one Apc allele in Lrig1-expressing progenitor cells results in multiple distal colonic tumors with features of familial adenomatous polyposis.
Am. J. Physiol. Gastrointest. Liver Physiol.
PUBLISHED: 05-15-2014
Show Abstract
Hide Abstract
Individuals with familial adenomatous polyposis (FAP) harbor a germline mutation in adenomatous polyposis coli (APC). The major clinical manifestation is development of multiple colonic tumors at a young age due to stochastic loss of the remaining APC allele. Extracolonic features, including periampullary tumors, gastric abnormalities, and congenital hypertrophy of the retinal pigment epithelium, may occur. The objective of this study was to develop a mouse model that simulates these features of FAP. We combined our Lrig1-CreERT2/+ mice with Apcfl/+ mice, eliminated one copy of Apc in leucine-rich repeats and immunoglobulin-like domains protein 1 (Lrig1)-positive (Lrig1(+)) progenitor cells with tamoxifen injection, and monitored tumor formation in the colon by colonoscopy and PET. Initial loss of one Apc allele in Lrig1(+) cells results in a predictable pattern of preneoplastic changes, culminating in multiple distal colonic tumors within 50 days of induction, as well as the extracolonic manifestations of FAP mentioned above. We show that tumor formation can be monitored by noninvasive PET imaging. This inducible stem cell-driven model recapitulates features of FAP and offers a tractable platform on which therapeutic interventions can be monitored over time by colonoscopy and noninvasive imaging.
Related JoVE Video
A peptide-based positron emission tomography probe for in vivo detection of caspase activity in apoptotic cells.
Clin. Cancer Res.
PUBLISHED: 02-26-2014
Show Abstract
Hide Abstract
Apoptosis, or programmed cell death, can be leveraged as a surrogate measure of response to therapeutic interventions in medicine. Cysteine aspartic acid-specific proteases, or caspases, are essential determinants of apoptosis signaling cascades and represent promising targets for molecular imaging. Here, we report development and in vivo validation of [(18)F]4-fluorobenzylcarbonyl-Val-Ala-Asp(OMe)-fluoromethylketone ([(18)F]FB-VAD-FMK), a novel peptide-based molecular probe suitable for quantification of caspase activity in vivo using positron emission tomography (PET).
Related JoVE Video
Novel mechanistic insights into ectodomain shedding of EGFR Ligands Amphiregulin and TGF-?: impact on gastrointestinal cancers driven by secondary bile acids.
Cancer Res.
PUBLISHED: 02-11-2014
Show Abstract
Hide Abstract
Secondary bile acids (BA) such as deoxycholic acid (DCA) promote the development of several gastrointestinal malignancies, but how they mediate this effect is unclear. In this study, we offer evidence of a mechanism involving ectodomain shedding of the EGFR ligands amphiregulin (AREG) and TGF-?, which rely upon the cell surface protease TACE/ADAM-17. Specifically, we show that AREG participates in DCA-induced EGFR and STAT3 signaling, cell-cycle progression, and tumorigenicity in human colorectal cancer and pancreatic ductal adenocarcinoma (PDAC). TACE and AREG, but not TGF-?, were overexpressed in both colorectal cancer and PDAC tissues compared with normal tissues. Exposure of colorectal cancer and PDAC cells to DCA resulted in colocalization of Src and TACE to the cell membrane, resulting in AREG-dependent activation of EGFR, mitogen-activated protein kinase (MAPK), and STAT3 signaling. Src or TACE inhibition was sufficient to attenuate DCA-induced AREG, but not TGF-? shedding. We also examined a role for the BA transporter TGR5 in DCA-mediated EGFR and STAT3 signaling. RNA interference-mediated silencing of TGR5 or AREG inhibited DCA-induced EGFR, MAPK, and STAT3 signaling, blunted cyclin D1 expression and cell-cycle progression, and attenuated DCA-induced colorectal cancer or PDAC tumorigenicity. Together, our findings define an AREG-dependent signaling pathway that mediates the oncogenic effects of secondary BAs in gastrointestinal cancers, the targeting of which may enhance therapeutic responses in their treatment.
Related JoVE Video
Activation of EGFR and ERBB2 by Helicobacter pylori results in survival of gastric epithelial cells with DNA damage.
Gastroenterology
PUBLISHED: 02-06-2014
Show Abstract
Hide Abstract
The gastric cancer-causing pathogen Helicobacter pylori up-regulates spermine oxidase (SMOX) in gastric epithelial cells, causing oxidative stress-induced apoptosis and DNA damage. A subpopulation of SMOX(high) cells are resistant to apoptosis, despite their high levels of DNA damage. Because epidermal growth factor receptor (EGFR) activation can regulate apoptosis, we determined its role in SMOX-mediated effects.
Related JoVE Video
Excess PLAC8 promotes an unconventional ERK2-dependent EMT in colon cancer.
J. Clin. Invest.
PUBLISHED: 01-30-2014
Show Abstract
Hide Abstract
The epithelial-to-mesenchymal transition (EMT) transcriptional program is characterized by repression of E-cadherin (CDH1) and induction of N-cadherin (CDH2), and mesenchymal genes like vimentin (VIM). Placenta-specific 8 (PLAC8) has been implicated in colon cancer; however, how PLAC8 contributes to disease is unknown, and endogenous PLAC8 protein has not been studied. We analyzed zebrafish and human tissues and found that endogenous PLAC8 localizes to the apical domain of differentiated intestinal epithelium. Colon cancer cells with elevated PLAC8 levels exhibited EMT features, including increased expression of VIM and zinc finger E-box binding homeobox 1 (ZEB1), aberrant cell motility, and increased invasiveness. In contrast to classical EMT, PLAC8 overexpression reduced cell surface CDH1 and upregulated P-cadherin (CDH3) without affecting CDH2 expression. PLAC8-induced EMT was linked to increased phosphorylated ERK2 (p-ERK2), and ERK2 knockdown restored cell surface CDH1 and suppressed CDH3, VIM, and ZEB1 upregulation. In vitro, PLAC8 directly bound and inactivated the ERK2 phosphatase DUSP6, thereby increasing p-ERK2. In a murine xenograft model, knockdown of endogenous PLAC8 in colon cancer cells resulted in smaller tumors, reduced local invasion, and decreased p-ERK2. Using MultiOmyx, a multiplex immunofluorescence-based methodology, we observed coexpression of cytosolic PLAC8, CDH3, and VIM at the leading edge of a human colorectal tumor, supporting a role for PLAC8 in cancer invasion in vivo.
Related JoVE Video
Should mesenteric tumor deposits be included in staging of well-differentiated small intestine neuroendocrine tumors?
Mod. Pathol.
PUBLISHED: 01-24-2014
Show Abstract
Hide Abstract
Well-differentiated small intestine neuroendocrine tumors can give rise to mesenteric tumor deposits, which are not included in the current American Joint Committee on Cancer staging system for small intestine neuroendocrine tumors, and their impact on patient prognosis is unknown. Seventy-two small intestine neuroendocrine tumors resections were identified in our files with slides, reports, and follow-up data available. Cases were assessed for T-category and for the presence of mesenteric tumor deposits, lymph node metastases, lymphovascular invasion, and liver metastases. Mesenteric tumor deposits were defined as discrete mesenteric tumor nodules ?1?mm with an irregular growth profile. Similar lesions clearly resulting from extranodal extension or direct contiguous spread by the primary lesion were excluded. Forty-three of the 72 cases had mesenteric tumor deposits (60%). The deposits were significantly associated with lymphovascular invasion (P=0.001), pT3 or pT4 disease (P=0.001), nodal metastases (P=0.040), and liver metastases (P<0.001) at the time of surgery. In addition, four of six cases with tumor deposits and no nodal disease had liver disease. Tumor deposits were associated with an increased incidence of disease progression and death due to the disease (P=0.001). Finally, the presence of tumor deposits at the time of surgery was associated with an increase in hazard of progression or death due to disease (hazard ratio: 4.0; 95% confidence interval: 1.3, 12.5; P=0.016). Mesenteric tumor deposits are present in the majority of cases of small intestine neuroendocrine tumors and are indicators of poor prognosis for this disease. Therefore, they may have a place in staging of small intestine neuroendocrine tumors, perhaps as analogous to lymph node disease.
Related JoVE Video
Fibrogenesis in pancreatic cancer is a dynamic process regulated by macrophage-stellate cell interaction.
Lab. Invest.
PUBLISHED: 01-15-2014
Show Abstract
Hide Abstract
Pancreatic cancer occurs in the setting of a profound fibrotic microenvironment that often dwarfs the actual tumor. Although pancreatic fibrosis has been well studied in chronic pancreatitis, its development in pancreatic cancer is much less well understood. This article describes the dynamic remodeling that occurs from pancreatic precursors (pancreatic intraepithelial neoplasias (PanINs)) to pancreatic ductal adenocarcinoma, highlighting similarities and differences between benign and malignant disease. Although collagen matrix is a commonality throughout this process, early stage PanINs are virtually free of periostin while late stage PanIN and pancreatic cancer are surrounded by an increasing abundance of this extracellular matrix protein. Myofibroblasts also become increasingly abundant during progression from PanIN to cancer. From the earliest stages of fibrogenesis, macrophages are associated with this ongoing process. In vitro co-culture indicates there is cross-regulation between macrophages and pancreatic stellate cells (PaSCs), precursors to at least some of the fibrotic cell populations. When quiescent PaSCs were co-cultured with macrophage cell lines, the stellate cells became activated and the macrophages increased cytokine production. In summary, fibrosis in pancreatic cancer involves a complex interplay of cells and matrices that regulate not only the tumor epithelium but the composition of the microenvironment itself.
Related JoVE Video
Identification of pathologic features associated with "ulcerative colitis-like" Crohn's disease.
World J. Gastroenterol.
PUBLISHED: 01-04-2014
Show Abstract
Hide Abstract
To identify pathologic features associated with this "ulcerative colitis (UC)-like" subgroup of Crohn's disease (CD).
Related JoVE Video
Phospholipid ether analogs for the detection of colorectal tumors.
PLoS ONE
PUBLISHED: 01-01-2014
Show Abstract
Hide Abstract
The treatment of localized colorectal cancer (CRC) depends on resection of the primary tumor with adequate margins and sufficient lymph node sampling. A novel imaging agent that accumulates in CRCs and the associated lymph nodes is needed. Cellectar Biosciences has developed a phospholipid ether analog platform that is both diagnostic and therapeutic. CLR1502 is a near-infrared fluorescent molecule, whereas 124/131I-CLR1404 is under clinical investigation as a PET tracer/therapeutic agent imaged by SPECT. We investigated the use of CLR1502 for the detection of intestinal cancers in a murine model and 131I-CLR1404 in a patient with metastatic CRC. Mice that develop multiple intestinal tumors ranging from adenomas to locally advanced adenocarcinomas were utilized. After 96 hours post CLR1502 injection, the intestinal tumors were analyzed using a Spectrum IVIS (Perkin Elmer) and a Fluobeam (Fluoptics). The intensity of the fluorescent signal was correlated with the histological characteristics for each tumor. Colon adenocarcinomas demonstrated increased accumulation of CLR1502 compared to non-invasive lesions (total radiant efficiency: 1.76×1010 vs 3.27×109 respectively, p?=?0.006). Metastatic mesenteric tumors and uninvolved lymph nodes were detected with CLR1502. In addition, SPECT imaging with 131I-CLR1404 was performed as part of a clinical trial in patients with advanced solid tumors. 131I-CLR1404 was shown to accumulate in metastatic tumors in a patient with colorectal adenocarcinoma. Together, these compounds might enhance our ability to properly resect CRCs through better localization of the primary tumor and improved lymph node identification as well as detect distant disease.
Related JoVE Video
3'-Deoxy-3'-[18F]-Fluorothymidine PET imaging reflects PI3K-mTOR-mediated pro-survival response to targeted therapy in colorectal cancer.
PLoS ONE
PUBLISHED: 01-01-2014
Show Abstract
Hide Abstract
Biomarkers that predict response to targeted therapy in oncology are an essential component of personalized medicine. In preclinical treatment response studies that featured models of wild-type KRAS or mutant BRAF colorectal cancer treated with either cetuximab or vemurafenib, respectively, we illustrate that [(18)F]-FLT PET, a non-invasive molecular imaging readout of thymidine salvage, closely reflects pro-survival responses to targeted therapy that are mediated by PI3K-mTOR activity. Activation of pro-survival mechanisms forms the basis of numerous modes of resistance. Therefore, we conclude that [(18)F]-FLT PET may serve a novel and potentially critical role to predict tumors that exhibit molecular features that tend to reflect recalcitrance to MAPK-targeted therapy. Though these studies focused on colorectal cancer, we envision that the results may be applicable to other solid tumors as well.
Related JoVE Video
Endoscopic mucosal resection of Barretts esophagus detects high prevalence of subsquamous intestinal metaplasia.
World J Gastrointest Endosc
PUBLISHED: 10-01-2013
Show Abstract
Hide Abstract
To report the prevalence of Subsquamous intestinal metaplasia (SSIM) in patients undergoing endoscopic mucosal resection (EMR) for staging of Barretts esophagus (BE).
Related JoVE Video
Malignant transformation of colonic epithelial cells by a colon-derived long noncoding RNA.
Biochem. Biophys. Res. Commun.
PUBLISHED: 09-05-2013
Show Abstract
Hide Abstract
Recent progress has been made in the identification of protein-coding genes and miRNAs that are expressed in and alter the behavior of colonic epithelia. However, the role of long non-coding RNAs (lncRNAs) in colonic homeostasis is just beginning to be explored. By gene expression profiling of post-mitotic, differentiated tops and proliferative, progenitor-compartment bottoms of microdissected adult mouse colonic crypts, we identified several lncRNAs more highly expressed in crypt bottoms. One identified lncRNA, designated non-coding Nras functional RNA (ncNRFR), resides within the Nras locus but appears to be independent of the Nras coding transcript. Stable overexpression of ncNRFR in non-transformed, conditionally immortalized mouse colonocytes results in malignant transformation, as determined by growth in soft agar and formation of highly invasive tumors in nude mice. Moreover, ncNRFR appears to inhibit the function of the tumor suppressor let-7. These results suggest precise regulation of ncNRFR is necessary for proper cell growth in the colonic crypt, and its misregulation results in neoplastic transformation.
Related JoVE Video
A Call to Standardize Preanalytic Data Elements for Biospecimens.
Arch. Pathol. Lab. Med.
PUBLISHED: 08-12-2013
Show Abstract
Hide Abstract
Context.-Biospecimens must have appropriate clinical annotation (data) to ensure optimal quality for both patient care and research. Clinical preanalytic variables are the focus of this study. Objectives.-To define the essential preanalytic variables (data fields) that should be attached to every collected biospecimen and to provide a complete list of such variables, along with their relative importance, which can vary, depending on downstream use, institutional needs, and information technology capabilities. Design.-The College of American Pathologists Diagnostic Intelligence and Health Information Technology Committee sponsored a Biorepository Working Group to develop a ranked list of the preanalytic variables for annotating biospecimens. Members of the working group were experts in anatomic, clinical, and molecular pathology; biobanking; medical informatics; and accreditation. Several members had experience with federal government programs, such as the National Cancer Institutes Biospecimens and Biorepository Branch and National Cancer Institutes Community Cancer Center Program. Potential preanalytic variables were identified and ranked along with available supporting evidence, definitions, and potential negative effects if the variable was not attached to the biospecimen. Additional national and international stakeholders reviewed the draft manuscript. Results.-The ranked listing of 170 preanalytic variables produced can be used as a guide for site-specific implementation into patient care and/or research biorepository processes. Conclusions.-In our collective experience, it is often difficult to choose which of the many preanalytic variables to attach to any specific set of biospecimens used for patient care and/or research. The attached ranked list should aid in the selection of preanalytic variables for a given biospecimen collection.
Related JoVE Video
Data set for reporting of lung carcinomas: recommendations from International Collaboration on Cancer Reporting.
Arch. Pathol. Lab. Med.
PUBLISHED: 08-01-2013
Show Abstract
Hide Abstract
The International Collaboration on Cancer Reporting (ICCR) is a quadripartite alliance formed by the Royal College of Pathologists of Australasia, the Royal College of Pathologists of the United Kingdom, the College of American Pathologists, and the Canadian Partnership Against Cancer. The ICCR was formed with a view to reducing the global burden of cancer data set development and reduplication of effort by different international institutions that commission, publish, and maintain standardized cancer-reporting data sets. The resultant standardization of cancer reporting would be expected to benefit not only those countries directly involved in the collaboration but also others not in a position to develop their own data sets.
Related JoVE Video
Debating Deposits: An Interobserver Variability Study of Lymph Nodes and Pericolonic Tumor Deposits in Colonic Adenocarcinoma.
Arch. Pathol. Lab. Med.
PUBLISHED: 07-31-2013
Show Abstract
Hide Abstract
Context.- The American Joint Committee on Cancers Cancer Staging Manual 7th edition defines pericolonic tumor deposits (TDs) as discrete tumor foci in pericolic fat showing no evidence of residual lymph node (LN). This definition relies on subjective features rather than size (5th edition) or shape (6th edition) and introduced the category N1c. Although typically straightforward, metastases are encountered for which the distinction between LNs and TDs is unclear. For data to be meaningful, agreement on distinguishing features between positive LNs and TDs is needed. Objectives.-To assess agreement among gastrointestinal pathologists evaluating difficult metastases and to report the distinguishing features they found helpful. Design.-Twenty-five tumor metastases from right-sided colonic adenocarcinomas were selected in which the distinction between positive LNs and TDs was challenging. Virtual slides were reviewed by 7 gastrointestinal pathologists. A list of features potentially helpful in differentiating positive LNs and TDs was ranked for usefulness by each pathologist. Every metastasis was diagnosed as positive LN or TD. For each case diagnosed as positive LN, reviewers were asked to list every feature used in diagnosis. Results.-Complete agreement was found for 11 of 25 metastases, 5 positive LNs and 6 TDs (? statistic, 0.48; 95% confidence interval, 0.28-0.67). Top-ranked features included round shape, peripheral lymphocyte rim, peripheral lymphoid follicles, possible subcapsular sinus, residual LN in surrounding fibroadipose tissue, and thick capsule. The top used features were similar among reviewers. Conclusions.-Significant agreement on positive LNs and TDs in difficult colonic adenocarcinoma metastases was found among evaluators, but inconsistency remains. Round shape, peripheral lymphocyte rim, peripheral lymphoid follicles, possible subcapsular sinus, residual LN in surrounding fibroadipose tissue, and thick capsule were most often used to aid in diagnosis.
Related JoVE Video
Pattern of breast cancer susceptibility gene 1 expression is a potential prognostic biomarker in resectable pancreatic ductal adenocarcinoma.
Pancreas
PUBLISHED: 07-16-2013
Show Abstract
Hide Abstract
The tumor-suppressor breast cancer susceptibility gene 1 (BRCA1) is a nuclear-cytoplasmic shuttling protein that when in the nucleus is required for DNA repair whereas when in the cytoplasm is important in activating cell death processes. Although BRCA1 mutations have been shown to be associated with an increased risk of pancreatic ductal adenocarcinoma (PDAC), its role in disease progression is yet to be determined. We hypothesized that BRCA1 expression pattern could be used as a prognostic biomarker.
Related JoVE Video
Transformation of epithelial cells through recruitment leads to polyclonal intestinal tumors.
Proc. Natl. Acad. Sci. U.S.A.
PUBLISHED: 06-24-2013
Show Abstract
Hide Abstract
Intestinal tumors from mice and humans can have a polyclonal origin. Statistical analyses indicate that the best explanation for this source of intratumoral heterogeneity is the presence of interactions among multiple progenitors. We sought to better understand the nature of these interactions. An initial progenitor could recruit others by facilitating the transformation of one or more neighboring cells. Alternatively, two progenitors that are independently initiated could simply cooperate to form a single tumor. These possibilities were tested by analyzing tumors from aggregation chimeras that were generated by fusing together embryos with unequal predispositions to tumor development. Strikingly, numerous polyclonal tumors were observed even when one genetic component was highly, if not completely, resistant to spontaneous tumorigenesis in the intestine. Moreover, the observed number of polyclonal tumors could be explained by the facilitated transformation of a single neighbor within 144 ?m of an initial progenitor. These findings strongly support recruitment instead of cooperation. Thus, it is conceivable that these interactions are necessary for tumors to thrive, so blocking them might be a highly effective method for preventing the formation of tumors in the intestine and other tissues.
Related JoVE Video
Deletion of cationic amino acid transporter 2 exacerbates dextran sulfate sodium colitis and leads to an IL-17-predominant T cell response.
Am. J. Physiol. Gastrointest. Liver Physiol.
PUBLISHED: 05-23-2013
Show Abstract
Hide Abstract
L-Arginine (L-Arg) is a semiessential amino acid that has altered availability in human ulcerative colitis (UC), a form of inflammatory bowel disease, and is beneficial in murine colitis induced by dextran sulfate sodium (DSS), a model with similarity to UC. We assessed the role of cationic amino acid transporter 2 (CAT2), the inducible transporter of L-Arg, in DSS colitis. Expression of CAT2 was upregulated in tissues from colitic mice and localized predominantly to colonic macrophages. CAT2-deficient (CAT2-/-) mice exposed to DSS exhibited worsening of survival, body weight loss, colon weight, and histological injury. These effects were associated with increased serum L-Arg and decreased tissue L-Arg uptake and inducible nitric oxide synthase protein expression. Clinical benefits of L-Arg supplementation in wild-type mice were lost in CAT2-/- mice. There was increased infiltration of macrophages, dendritic cells, granulocytes, and T cells in colitic CAT2-/- compared with wild-type mice. Cytokine profiling revealed increases in proinflammatory granulocyte colony-stimulating factor, macrophage inflammatory protein-1?, IL-15, and regulated and normal T cell-expressed and -secreted and a shift from an IFN-?- to an IL-17-predominant T cell response, as well as an increase in IL-13, in tissues from colitic CAT2-/- mice. However, there were no increases in other T helper cell type 2 cytokines, nor was there a global increase in macrophage-derived proinflammatory cytokines. The increase in IL-17 derived from both CD4 and ?? T cells and was associated with colonic IL-6 expression. Thus CAT2 plays an important role in controlling inflammation and IL-17 activation in an injury model of colitis, and impaired L-Arg availability may contribute to UC pathogenesis.
Related JoVE Video
Gastric cancer, version 2.2013: featured updates to the NCCN Guidelines.
J Natl Compr Canc Netw
PUBLISHED: 05-14-2013
Show Abstract
Hide Abstract
The NCCN Clinical Practice Guidelines in Oncology for Gastric Cancer provide evidence- and consensus-based recommendations for a multidisciplinary approach for the management of patients with gastric cancer. For patients with resectable locoregional cancer, the guidelines recommend gastrectomy with a D1+ or a modified D2 lymph node dissection (performed by experienced surgeons in high-volume centers). Postoperative chemoradiation is the preferred option after complete gastric resection for patients with T3-T4 tumors and node-positive T1-T2 tumors. Postoperative chemotherapy is included as an option after a modified D2 lymph node dissection for this group of patients. Trastuzumab with chemotherapy is recommended as first-line therapy for patients with HER2-positive advanced or metastatic cancer, confirmed by immunohistochemistry and, if needed, by fluorescence in situ hybridization for IHC 2+.
Related JoVE Video
Transformation of polarized epithelial cells by apical mistrafficking of epiregulin.
Proc. Natl. Acad. Sci. U.S.A.
PUBLISHED: 05-13-2013
Show Abstract
Hide Abstract
Establishment and maintenance of apico-basolateral trafficking pathways are critical to epithelial homeostasis. Loss of polarity and trafficking fidelity are thought to occur as a consequence of transformation; however, here we report that selective mistrafficking of the epidermal growth factor receptor (EGFR) ligand epiregulin (EREG) from the basolateral to the apical cell surface drives transformation. Normally, EREG is preferentially delivered to the basolateral surface of polarized Madin-Darby canine kidney cells. EREG basolateral trafficking is regulated by a conserved tyrosine-based basolateral sorting motif in its cytoplasmic domain (YXX?: Y(156)ERV). Both Y156 and V159 are required for basolateral sorting of EREG, because Y156A and V159G substitutions redirect EREG to the apical cell surface. We also show that basolateral sorting of EREG is adaptor protein 1B-independent. Apical mistrafficking of EREG has a distinctive phenotype. In contrast to transient EGFR tyrosine phosphorylation after basolateral EREG stimulation, apical EREG leads to prolonged EGFR tyrosine phosphorylation, which may be related, at least in part, to a lack of negative regulatory Y1045 phosphorylation and subsequent ubiquitylation. Notably, Madin-Darby canine kidney cells stably expressing apically mistrafficked EREG form significantly larger, hyperproliferative, poorly differentiated, and locally invasive tumors in nude mice compared with WT EREG-expressing cells.
Related JoVE Video
Epidermal growth factor receptor signaling pathway is frequently altered in ampullary carcinoma at protein and genetic levels.
Mod. Pathol.
PUBLISHED: 04-15-2013
Show Abstract
Hide Abstract
Our objective was to explore alteration of the epidermal growth factor receptor (EGFR) signaling pathway in ampullary carcinoma. Immunohistochemical studies were employed to evaluate expression of amphiregulin as well as expression and activation of EGFR. A lab-developed assay was used to identify mutations in the EGFR pathway genes, including KRAS, BRAF, PIK3CA, PTEN, and AKT1. A total of 52 ampullary carcinomas were identified, including 25 intestinal-type and 24 pancreatobiliary-type tumors, with the intestinal type being associated with a younger age at diagnosis (P=0.03) and a better prognosis (P<0.01). Expression of amphiregulin correlated with better differentiation (P<0.01), but no difference was observed between two major histologic types. Expression and activation of EGFR was more commonly seen in the pancreatobiliary type (P<0.01). Mutations were detected in 50% of the pancreatobiliary type and 60% of the intestinal type. KRAS was the most common gene mutated in the pancreatobiliary type (42%) as well as the intestinal type (52%). Other mutations detected included PIK3CA, SMAD4 and BRAF. KRAS mutations at codons 12 and 13 did not adversely affect overall survival. In conclusion, EGFR expression and activation were different between intestinal- and pancreatobiliary-type ampullary carcinoma. KRAS mutation was common in both histologic types; however, the incidence appeared to be lower in the pancreatobiliary type compared with its pancreatic counterpart, pancreatic ductal adenocarcinoma. Mutational analysis of the EGFR pathway genes may provide important insights into personalized treatment for patients with ampullary carcinoma.Modern Pathology advance online publication, 1 November 2013; doi:10.1038/modpathol.2013.185.
Related JoVE Video
High dietary salt intake exacerbates Helicobacter pylori-induced gastric carcinogenesis.
Infect. Immun.
PUBLISHED: 04-08-2013
Show Abstract
Hide Abstract
Persistent colonization of the human stomach with Helicobacter pylori is a risk factor for gastric adenocarcinoma, and H. pylori-induced carcinogenesis is dependent on the actions of a bacterial oncoprotein known as CagA. Epidemiological studies have shown that high dietary salt intake is also a risk factor for gastric cancer. To investigate the effects of a high-salt diet, we infected Mongolian gerbils with a wild-type (WT) cagA(+) H. pylori strain or an isogenic cagA mutant strain and maintained the animals on a regular diet or a high-salt diet. At 4 months postinfection, gastric adenocarcinoma was detected in 100% of the WT-infected/high-salt-diet animals, 58% of WT-infected/regular-diet animals, and none of the animals infected with the cagA mutant strain (P < 0.0001). Among animals infected with the WT strain, those fed a high-salt diet had more severe gastric inflammation, higher gastric pH, increased parietal cell loss, increased gastric expression of interleukin 1? (IL-1?), and decreased gastric expression of hepcidin and hydrogen potassium ATPase (H,K-ATPase) compared to those on a regular diet. Previous studies have detected upregulation of CagA synthesis in response to increased salt concentrations in the bacterial culture medium, and, concordant with the in vitro results, we detected increased cagA transcription in vivo in animals fed a high-salt diet compared to those on a regular diet. Animals infected with the cagA mutant strain had low levels of gastric inflammation and did not develop hypochlorhydria. These results indicate that a high-salt diet potentiates the carcinogenic effects of cagA(+) H. pylori strains.
Related JoVE Video
Identification and manipulation of biliary metaplasia in pancreatic tumors.
Gastroenterology
PUBLISHED: 03-25-2013
Show Abstract
Hide Abstract
Metaplasias often have characteristics of developmentally related tissues. Pancreatic metaplastic ducts are usually associated with pancreatitis and pancreatic ductal adenocarcinoma. The tuft cell is a chemosensory cell that responds to signals in the extracellular environment via effector molecules. Commonly found in the biliary tract, tuft cells are absent from normal murine pancreas. Using the aberrant appearance of tuft cells as an indicator, we tested if pancreatic metaplasia represents transdifferentiation to a biliary phenotype and what effect this has on pancreatic tumorigenesis.
Related JoVE Video
53BP1 expression is a modifier of the prognostic value of lymph node ratio and CA 19-9 in pancreatic adenocarcinoma.
BMC Cancer
PUBLISHED: 03-08-2013
Show Abstract
Hide Abstract
53BP1 binds to the tumor suppressor p53 and has a key role in DNA damage response and repair. Low 53BP1 expression has been associated with decreased survival in breast cancer and has been shown to interact with several prognostic factors in non-small cell lung cancer. The role of 53BP1 in pancreatic ductal adenocarcinoma (PDAC) has yet to be determined. We aimed to investigate whether 53BP1 levels interact with established prognostic factors in PDAC.
Related JoVE Video
The interleukin-17 receptor B subunit is essential for the Th2 response to Helicobacter pylori, but not for control of bacterial burden.
PLoS ONE
PUBLISHED: 02-26-2013
Show Abstract
Hide Abstract
Helicobacter pylori infection leads to an inflammatory response in 100% of infected individuals. The inflammatory cells which are recruited to the gastric mucosa during infection produce several pro- and anti-inflammatory cytokines including several cytokines in the interleukin-17 family. The anti-inflammatory cytokine, interleukin 25 (IL-25, also known as IL-17E), signals through a receptor, which is a heterotrimeric receptor comprised of two IL-17 receptor A subunits and an IL-17 receptor B subunit. Previous studies in our laboratory demonstrated that IL-17RA is required to control infection with Helicobacter pylori in the mouse model. Moreover, the absence of IL-17 receptor A leads to a significant B cell infiltrate and a remarkable increase in lymphoid follicle formation in response to infection compared to infection in wild-type mice. We hypothesized that IL-25, which requires both IL-17 receptor A and IL-17 receptor B for signaling, may play a role in control of inflammation in the mouse model of Helicobacter pylori infection. IL-17 receptor B deficient mice, IL-17 receptor A deficient mice and wild-type mice were infected with Helicobacter pylori (strains SS1 and PMSS1). At several time points H. pylori-infected mice were sacrificed to investigate their ability to control infection and inflammation. Moreover, the effects of IL-17 receptor B deficiency on T helper cytokine expression and H. pylori- specific serum antibody responses were measured. IL-17 receptor B-/- mice (unlike IL-17 receptor A-/- mice) exhibited similar or modest changes in gastric colonization, inflammation, and Th1 and Th17 helper cytokine responses to wild-type mice infected with Helicobacter pylori. However, H. pylori-infected IL-17 receptor B-/- mice have reduced expression of IL-4 and lower serum IgG1 and IgG2a levels compared to infected IL-17 receptor A-/- and wild-type mice. These data indicate that signaling through the IL-17 receptor B subunit is not necessary for control of Helicobacter pylori in our model.
Related JoVE Video
Aurora kinase a promotes inflammation and tumorigenesis in mice and human gastric neoplasia.
Gastroenterology
PUBLISHED: 02-20-2013
Show Abstract
Hide Abstract
Chronic inflammation contributes to the pathogenesis of gastric tumorigenesis. The aurora kinase A (AURKA) gene is frequently amplified and overexpressed in gastrointestinal cancers. We investigated the roles of AURKA in inflammation and gastric tumorigenesis.
Related JoVE Video
Limits of [18F]-FLT PET as a biomarker of proliferation in oncology.
PLoS ONE
PUBLISHED: 02-08-2013
Show Abstract
Hide Abstract
Non-invasive imaging biomarkers of cellular proliferation hold great promise for quantifying response to personalized medicine in oncology. An emerging approach to assess tumor proliferation utilizes the positron emission tomography (PET) tracer 3-deoxy-3[(18)F]-fluorothymidine, [(18)F]-FLT. Though several studies have associated serial changes in [(18)F]-FLT-PET with elements of therapeutic response, the degree to which [(18)F]-FLT-PET quantitatively reflects proliferative index has been continuously debated for more that a decade. The goal of this study was to elucidate quantitative relationships between [(18)F]-FLT-PET and cellular metrics of proliferation in treatment naïve human cell line xenografts commonly employed in cancer research.
Related JoVE Video
Pathology of rodent models of intestinal cancer: progress report and recommendations.
Gastroenterology
PUBLISHED: 01-26-2013
Show Abstract
Hide Abstract
In October 2010, a pathology review of rodent models of intestinal neoplasia was held at The Jackson Laboratory. This review complemented 2 other concurrent events: a workshop on methods of modeling colon cancer in rodents and a conference on current issues in murine and human colon cancer. We summarize the results of the pathology review and the committees recommendations for tumor nomenclature. A virtual high-resolution image slide box of these models has been developed. This report discusses significant recent developments in rodent modeling of intestinal neoplasia, including the role of stem cells in cancer and the creation of models of metastatic intestinal cancer.
Related JoVE Video
3-Deoxy-3-18F-fluorothymidine PET predicts response to (V600E)BRAF-targeted therapy in preclinical models of colorectal cancer.
J. Nucl. Med.
PUBLISHED: 01-22-2013
Show Abstract
Hide Abstract
Selective inhibition of oncogenic targets and associated signaling pathways forms the basis of personalized cancer medicine. The clinical success of (V600E)BRAF inhibition in melanoma, coupled with the emergence of acquired resistance, underscores the importance of rigorously validating quantitative biomarkers of treatment response in this and similar settings. Because constitutive activation of BRAF leads to proliferation in tumors, we explored 3-deoxy-3-(18)F-fluorothymidine ((18)F-FLT) PET to noninvasively quantify changes in tumor proliferation that are associated with pharmacologic inhibition of (V600E)BRAF downstream effectors and that precede changes in tumor volume.
Related JoVE Video
SPDEF functions as a colorectal tumor suppressor by inhibiting ?-catenin activity.
Gastroenterology
PUBLISHED: 01-17-2013
Show Abstract
Hide Abstract
Expression of the SAM pointed domain containing ETS transcription factor (SPDEF or prostate-derived ETS factor) is regulated by Atoh1 and is required for the differentiation of goblet and Paneth cells. SPDEF has been reported to suppress the development of breast, prostate, and colon tumors. We analyzed levels of SPDEF in colorectal tumor samples from patients and its tumor-suppressive functions in mouse models of colorectal cancer (CRC).
Related JoVE Video
Expert pathology review and endoscopic mucosal resection alters the diagnosis of patients referred to undergo therapy for Barretts esophagus.
Surg Endosc
PUBLISHED: 01-11-2013
Show Abstract
Hide Abstract
Endoscopic therapy has emerged as an alternative to surgical esophagectomy for the management of Barretts esophagus (BE)-associated neoplasia. Accurate pretreatment staging is essential to ensure an appropriate choice of therapy and optimal long-term outcomes. This study aimed to assess the frequency with which expert histopathologic review of biopsies combined with endoscopic mucosal resection (EMR) would alter the pretreatment diagnosis of BE-associated neoplasia.
Related JoVE Video
STAT6 deficiency ameliorates severity of oxazolone colitis by decreasing expression of claudin-2 and Th2-inducing cytokines.
J. Immunol.
PUBLISHED: 01-09-2013
Show Abstract
Hide Abstract
Patients suffering from ulcerative colitis (UC) exhibit chronic colonic inflammation caused by a dysregulated mucosal immune response and epithelial barrier disruption. Th2 cytokines, including IL-13, have been implicated in the pathogenesis of UC. IL-13 induces phosphorylation of STAT6, and we previously demonstrated increased epithelial p-STAT6 in children with UC. In this study, we investigated the role of STAT6 in oxazolone colitis, a murine model of UC, by inducing colitis in STAT6-deficient (STAT6(-/-)) and wild type (WT) mice. We observed increased epithelial cell, T cell, macrophage, and NKT cell STAT6 phosphorylation, as well as increased p-STAT6(+) IL-13-producing NKT cells, in colitic WT mice. Colitis was attenuated in STAT6(-/-) mice, with improvements in weight, colon length, and histopathology. There was decreased induction of the pore-forming tight junction protein claudin-2 in STAT6(-/-) mice. Similarly, short hairpin RNA STAT6 knockdown reduced claudin-2 induction and transepithelial resistance decrease in IL-13-treated human T84 cells. Tissue expression of IL-13, IFN-?, IL-17, and IL-10 mRNA was similarly induced in WT and STAT6(-/-) colitic mice; however, we observed increased mRNA expression for the Th2-inducing cytokines IL-33 and thymic stromal lymphopoietin in WT mice with colitis, which was abrogated in STAT6(-/-) mice. Mesenteric lymph node cells from STAT6(-/-) mice with colitis exhibited reduced secretion of IL-4, IL-5, IL-13, and IFN-?. IL-33 augmented mesenteric lymph node cell secretion of IL-5, IL-13, IL-6, and IFN-?. These data implicate STAT6 in the pathogenesis of colitis in vivo with important roles in altering epithelial barrier function and regulating Th2-inducing cytokine production.
Related JoVE Video
Gastric adenocarcinoma has a unique microRNA signature not present in esophageal adenocarcinoma.
Cancer
PUBLISHED: 01-02-2013
Show Abstract
Hide Abstract
MicroRNAs (miRNAs) play critical roles in tumor development and progression. The finding that a single miRNA can regulate hundreds of genes places miRNAs at critical hubs of signaling pathways. For the current study, the authors investigated the miRNA expression profile of gastric adenocarcinomas and compared it with esophageal adenocarcinomas to better identify a unique miRNA signature of gastric adenocarcinoma.
Related JoVE Video
Diagnostic and therapeutic implications of a novel immunohistochemical panel detecting duodenal mucosal invasion by pancreatic ductal adenocarcinoma.
Int J Clin Exp Pathol
PUBLISHED: 01-01-2013
Show Abstract
Hide Abstract
We investigated a series of pancreaticoduodenectomy and duodenal biopsies with a panel of immunohistochemical markers to identify duodenal mucosal invasion by pancreatic ductal adenocarcinoma (PDAC), including markers of poor prognosis and targets of promising novel immunotherapies.
Related JoVE Video
Four jointed box 1 promotes angiogenesis and is associated with poor patient survival in colorectal carcinoma.
PLoS ONE
PUBLISHED: 01-01-2013
Show Abstract
Hide Abstract
Angiogenesis, the recruitment and re-configuration of pre-existing vasculature, is essential for tumor growth and metastasis. Increased tumor vascularization often correlates with poor patient outcomes in a broad spectrum of carcinomas. We identified four jointed box 1 (FJX1) as a candidate regulator of tumor angiogenesis in colorectal cancer. FJX1 mRNA and protein are upregulated in human colorectal tumor epithelium as compared with normal epithelium and colorectal adenomas, and high expression of FJX1 is associated with poor patient prognosis. FJX1 mRNA expression in colorectal cancer tissues is significantly correlated with changes in known angiogenesis genes. Augmented expression of FJX1 in colon cancer cells promotes growth of xenografts in athymic mice and is associated with increased tumor cell proliferation and vascularization. Furthermore, FJX1 null mice develop significantly fewer colonic polyps than wild-type littermates after combined dextran sodium sulfate (DSS) and azoxymethane (AOM) treatment. In vitro, conditioned media from FJX1 expressing cells promoted endothelial cell capillary tube formation in a HIF1-? dependent manner. Taken together our results support the conclusion that FJX1 is a novel regulator of tumor progression, due in part, to its effect on tumor vascularization.
Related JoVE Video
Deciphering the unique microRNA signature in human esophageal adenocarcinoma.
PLoS ONE
PUBLISHED: 01-01-2013
Show Abstract
Hide Abstract
Esophageal adenocarcinoma (EAC) is characterized by a steep rise in incidence rates in the Western population. The unique miRNA signature that distinguishes EAC from other upper gastrointestinal cancers remains unclear. Herein, we performed a comprehensive microarray profiling for the specific miRNA signature associated with EAC. We validated this signature by qRT-PCR.
Related JoVE Video
Proinflammatory cytokines and bile acids upregulate ?Np73 protein, an inhibitor of p53 and p73 tumor suppressors.
PLoS ONE
PUBLISHED: 01-01-2013
Show Abstract
Hide Abstract
Gastroesophageal reflux disease (GERD) is the main etiological factor behind the recent rapid increase in the incidence of esophageal adenocarcinoma. During reflux, esophageal cells are exposed to bile at low pH resulting in cellular damage and inflammation, which are known to facilitate cancer development. In this study, we investigated the regulation of p73 isoform, ?Np73?, in the reflux condition. Previous studies have reported that ?Np73 exhibits anti-apoptotic and oncogenic properties through inhibition of p53 and p73 proteins. We found that direct exposure of esophageal cells to bile acids in an acidic environment alters the phosphorylation of ?Np73, its subcellular localization and increases ?Np73 protein levels. Upregulation of ?Np73 was also observed in esophageal tissues collected from patients with GERD and Barretts metaplasia, a precancerous lesion in the esophagus associated with gastric reflux. c-Abl, p38 MAPK, and IKK protein kinases were identified to interact in the regulation of ?Np73. Their inhibition with chemotherapeutic agents and siRNA suppresses ?Np73. We also found that pro-inflammatory cytokines, IL-1? and TNF?, are potent inducers of ?Np73?, which further enhance the bile acids/acid effect. Combined, our studies provide evidence that gastroesophageal reflux alters the regulation of oncogenic ?Np73 isoform that may facilitate tumorigenic transformation of esophageal metaplastic epithelium.
Related JoVE Video
mTOR inhibition elicits a dramatic response in PI3K-dependent colon cancers.
PLoS ONE
PUBLISHED: 01-01-2013
Show Abstract
Hide Abstract
The phosphatidylinositide-3-kinase (PI3K) signaling pathway is critical for multiple cellular functions including metabolism, proliferation, angiogenesis, and apoptosis, and is the most commonly altered pathway in human cancers. Recently, we developed a novel mouse model of colon cancer in which tumors are initiated by a dominant active PI3K (FC PIK3ca). The cancers in these mice are moderately differentiated invasive mucinous adenocarcinomas of the proximal colon that develop by 50 days of age. Interestingly, these cancers form without a benign intermediary or aberrant WNT signaling, indicating a non-canonical mechanism of tumorigenesis. Since these tumors are dependent upon the PI3K pathway, we investigated the potential for tumor response by the targeting of this pathway with rapamycin, an mTOR inhibitor. A cohort of FC PIK3ca mice were treated with rapamycin at a dose of 6 mg/kg/day or placebo for 14 days. FDG dual hybrid PET/CT imaging demonstrated a dramatic tumor response in the rapamycin arm and this was confirmed on necropsy. The tumor tissue remaining after treatment with rapamycin demonstrated increased pERK1/2 or persistent phosphorylated ribosomal protein S6 (pS6), indicating potential resistance mechanisms. This unique model will further our understanding of human disease and facilitate the development of therapeutics through pharmacologic screening and biomarker identification.
Related JoVE Video
Berberine promotes recovery of colitis and inhibits inflammatory responses in colonic macrophages and epithelial cells in DSS-treated mice.
Am. J. Physiol. Gastrointest. Liver Physiol.
PUBLISHED: 12-15-2011
Show Abstract
Hide Abstract
Inflammatory bowel disease (IBD) results from dysregulation of intestinal mucosal immune responses to microflora in genetically susceptible hosts. A major challenge for IBD research is to develop new strategies for treating this disease. Berberine, an alkaloid derived from plants, is an alternative medicine for treating bacterial diarrhea and intestinal parasite infections. Recent studies suggest that berberine exerts several other beneficial effects, including inducing anti-inflammatory responses. This study determined the effect of berberine on treating dextran sulfate sodium (DSS)-induced intestinal injury and colitis in mice. Berberine was administered through gavage to mice with established DSS-induced intestinal injury and colitis. Clinical parameters, intestinal integrity, proinflammatory cytokine production, and signaling pathways in colonic macrophages and epithelial cells were determined. Berberine ameliorated DSS-induced body weight loss, myeloperoxidase activity, shortening of the colon, injury, and inflammation scores. DSS-upregulated proinflammatory cytokine levels in the colon, including TNF, IFN-?, KC, and IL-17 were reduced by berberine. Berberine decreased DSS-induced disruption of barrier function and apoptosis in the colon epithelium. Furthermore, berberine inhibited proinflammatory cytokine production in colonic macrophages and epithelial cells in DSS-treated mice and promoted apoptosis of colonic macrophages. Activation of signaling pathways involved in stimulation of proinflammatory cytokine production, including MAPK and NF-?B, in colonic macrophages and epithelial cells from DSS-treated mice was decreased by berberine. In summary, berberine promotes recovery of DSS-induced colitis and exerts inhibitory effects on proinflammatory responses in colonic macrophages and epithelial cells. Thus berberine may represent a new therapeutic approach for treating gastrointestinal inflammatory disorders.
Related JoVE Video
ErbB2 and ErbB3 regulate recovery from dextran sulfate sodium-induced colitis by promoting mouse colon epithelial cell survival.
Lab. Invest.
PUBLISHED: 12-12-2011
Show Abstract
Hide Abstract
ErbB2 and ErbB3 receptor tyrosine kinases are key regulators of proliferation, migration, differentiation and cell survival; however, their roles in gastrointestinal biology remain poorly defined. We hypothesized that ErbB2 and ErbB3 promote colon epithelial cell survival in the context of the wound-healing response following colitis. In this study, mice bearing intestinal epithelial-specific deletion of ErbB2 or ErbB3 were treated with dextran sulfate sodium (DSS). Colon sections were examined for injury, cytokine expression, epithelial cell proliferation and apoptosis. Deletion of epithelial ErbB2 did not affect the extent of intestinal injury in response to DSS, whereas deletion of ErbB3 slightly increased injury. However, the roles of both receptors were more apparent during recovery from DSS colitis, in which ErbB2 or ErbB3 epithelial deletion resulted in greater inflammation and crypt damage during the early reparative period. Moreover, loss of ErbB3 prevented normal epithelial regeneration in the long term, with damage persisting for at least 6 weeks following a single round of DSS. Delayed recovery in mice with epithelial deletion of ErbB2 or ErbB3 was associated with increased colonic expression of tumor necrosis factor alpha and increased epithelial apoptosis. Furthermore, epithelial ErbB3 deletion increased apoptosis at baseline and during DSS injury. Additionally, epithelial cell hyperproliferation during recovery was exacerbated by deletion of either ErbB2 or ErbB3. These results suggest that ErbB2 and ErbB3 have important cytoprotective and reparative roles in the colonic epithelium following injury, by promoting colon epithelial cell survival.
Related JoVE Video
DNA methylation profiling in Barretts esophagus and esophageal adenocarcinoma reveals unique methylation signatures and molecular subclasses.
Epigenetics
PUBLISHED: 12-06-2011
Show Abstract
Hide Abstract
Barretts esophagus (BE) is a metaplastic process whereby the normal stratified, squamous esophageal epithelium is replaced by specialized intestinal epithelium. Barretts is the only accepted precursor lesion for esophageal adenocarcinoma (EAC), a solid tumor that is rapidly increasing in incidence in western countries. BE evolves into EAC through intermediate steps that involve increasing degrees of dysplasia. Current histologic criteria are quite subjective and the clinical behavior of BE is highly variable and difficult to predict using these standards. It is widely believed that molecular alterations present in BE and EAC will provide more precise prognostic and predictive markers for these conditions than the current clinical and histologic features in use. In order to further define molecular alterations that can classify unique groups of BE and EAC, we utilized methylation microarrays to compare the global gene methylation status of a collection of normal squamous, BE, BE + high-grade dysplasia (HGD), and EAC cases. We found distinct global methylation signatures, as well as differential methylation of specific genes, that discriminated these histological groups. We also noted high and low methylation epigenotypes among the BE and EAC cases. Additional validation of those CpG sites that distinguished BE from BE + HGD and EAC may lead to the discovery of useful biomarkers with potential clinical applications in the diagnosis and prognosis of BE and EAC.
Related JoVE Video
Smad4-mediated signaling inhibits intestinal neoplasia by inhibiting expression of ?-catenin.
Gastroenterology
PUBLISHED: 10-18-2011
Show Abstract
Hide Abstract
Mutational inactivation of adenomatous polyposis coli (APC) is an early event in colorectal cancer (CRC) progression that affects the stability and increases the activity of ?-catenin, a mediator of Wnt signaling. Progression of CRC also involves inactivation of signaling via transforming growth factor ? and bone morphogenetic protein (BMP), which are tumor suppressors. However, the interactions between these pathways are not clear. We investigated the effects of loss of the transcription factor Smad4 on levels of ?-catenin messenger RNA (mRNA) and Wnt signaling.
Related JoVE Video
Endoscopic therapy of esophageal premalignancy and early malignancy.
J Natl Compr Canc Netw
PUBLISHED: 09-09-2011
Show Abstract
Hide Abstract
Esophageal adenocarcinoma (EAC) is an often deadly cancer with a rising incidence in Western countries. Chronic gastroesophageal reflux disease is associated with the metaplastic transformation of normal squamous epithelium to premalignant specialized intestinal metaplasia within the esophagus (Barretts esophagus). Barretts esophagus may progress to low-grade dysplasia (LGD), high-grade dysplasia (HGD), or even EAC. Although nondysplastic Barretts esophagus progresses to EAC at a rate of 0.5% per year, rates of progression for true LGD and HGD are significantly higher. Treatment is mandatory for HGD and may be appropriate in select patients with nondysplastic Barretts esophagus and many with LGD. Thus, accurate pathologic assessment is necessary before considering endoscopic therapy. Previously, only esophagectomy was offered to patients with HGD or EAC. However, esophagectomy has significant morbidity and mortality, and therefore endoscopic therapies have been advocated for early Barretts neoplasia. These methods include endoscopic mucosal resection (EMR) and ablative techniques. Ablation techniques include argon plasma coagulation, multipolar electrocoagulation, laser therapy, photodynamic therapy, radiofrequency ablation, and cryotherapy. Of these, radiofrequency ablation has experienced the greatest adoption for the treatment of dysplastic Barretts esophagus because of excellent published outcomes. The use of EMR to resect suspicious areas or raised lesions is mandatory to provide histology. In contrast, ablation techniques such as radiofrequency ablation have been shown to effectively eradicate large areas of dysplastic tissue with relative ease but do not allow for histologic assessment of the treated area. Combination EMR with radiofrequency ablation is thus advocated to resect visible lesions via EMR (providing histology) and ablate the remainder of the Barretts esophagus. As always, the appropriate treatment is best determined after careful discussion with patients in a multidisciplinary environment. However, endoscopic therapy offers an attractive alternative to esophagectomy for early Barretts neoplasia.
Related JoVE Video
CD44 upregulation in E-cadherin-negative esophageal cancers results in cell invasion.
PLoS ONE
PUBLISHED: 07-29-2011
Show Abstract
Hide Abstract
E-cadherin is frequently lost during epithelial-mesenchymal transition and the progression of epithelial tumorigenesis. We found a marker of epithelial-mesenchymal transition, CD44, upregulated in response to functional loss of E-cadherin in esophageal cell lines and cancer. Loss of E-cadherin expression correlates with increased expression of CD44 standard isoform. Using an organotypic reconstruct model, we show increased CD44 expression in areas of cell invasion is associated with MMP-9 at the leading edge. Moreover, Activin A increases cell invasion through CD44 upregulation after E-cadherin loss. Taken together, our results provide functional evidence of CD44 upregulation in esophageal cancer invasion.
Related JoVE Video
BVES regulates EMT in human corneal and colon cancer cells and is silenced via promoter methylation in human colorectal carcinoma.
J. Clin. Invest.
PUBLISHED: 07-27-2011
Show Abstract
Hide Abstract
The acquisition of a mesenchymal phenotype is a critical step in the metastatic progression of epithelial carcinomas. Adherens junctions (AJs) are required for suppressing this epithelial-mesenchymal transition (EMT) but less is known about the role of tight junctions (TJs) in this process. Here, we investigated the functions of blood vessel epicardial substance (BVES, also known as POPDC1 and POP1), an integral membrane protein that regulates TJ formation. BVES was found to be underexpressed in all stages of human colorectal carcinoma (CRC) and in adenomatous polyps, indicating its suppression occurs early in transformation. Similarly, the majority of CRC cell lines tested exhibited decreased BVES expression and promoter DNA hypermethylation, a modification associated with transcriptional silencing. Treatment with a DNA-demethylating agent restored BVES expression in CRC cell lines, indicating that methylation represses BVES expression. Reexpression of BVES in CRC cell lines promoted an epithelial phenotype, featuring decreased proliferation, migration, invasion, and anchorage-independent growth; impaired growth of an orthotopic xenograft; and blocked metastasis. Conversely, interfering with BVES function by expressing a dominant-negative mutant in human corneal epithelial cells induced mesenchymal features. These biological outcomes were associated with changes in AJ and TJ composition and related signaling. Therefore, BVES prevents EMT, and its epigenetic silencing may be an important step in promoting EMT programs during colon carcinogenesis.
Related JoVE Video
Targeting notch pathway enhances rapamycin antitumor activity in pancreas cancers through PTEN phosphorylation.
Mol. Cancer
PUBLISHED: 05-03-2011
Show Abstract
Hide Abstract
Pancreas cancer is one of most aggressive human cancers with the survival rate for patients with metastatic pancreas cancer at 5-6 months. The poor survival demonstrates a clear need for better target identification, drug development and new therapeutic strategies. Recent discoveries have shown that the role for Notch pathway is important in both development and cancer. Its contribution to oncogenesis also involves crosstalks with other growth factor pathways, such as Akt and its modulator, PTEN. The mounting evidence supporting a role for Notch in cancer promotion and survival suggests that targeting this pathway alone or in combination with other therapeutics represents a promising therapeutic strategy.
Related JoVE Video
Location-specific epigenetic regulation of the metallothionein 3 gene in esophageal adenocarcinomas.
PLoS ONE
PUBLISHED: 04-15-2011
Show Abstract
Hide Abstract
Metallothionein 3 (MT3) maintains intracellular metal homeostasis and protects against reactive oxygen species (ROS)-induced DNA damage. In this study, we investigated the epigenetic alterations and gene expression of the MT3 gene in esophageal adenocarcinomas (EACs).
Related JoVE Video
Colon-specific delivery of a probiotic-derived soluble protein ameliorates intestinal inflammation in mice through an EGFR-dependent mechanism.
J. Clin. Invest.
PUBLISHED: 03-09-2011
Show Abstract
Hide Abstract
Probiotic bacteria can potentially have beneficial effects on the clinical course of several intestinal disorders, but our understanding of probiotic action is limited. We have identified a probiotic bacteria-derived soluble protein, p40, from Lactobacillus rhamnosus GG (LGG), which prevents cytokine-induced apoptosis in intestinal epithelial cells. In the current study, we analyzed the mechanisms by which p40 regulates cellular responses in intestinal epithelial cells and p40s effects on experimental colitis using mouse models. We show that the recombinant p40 protein activated EGFR, leading to Akt activation. Activation of EGFR by p40 was required for inhibition of cytokine-induced apoptosis in intestinal epithelial cells in vitro and ex vivo. Furthermore, we developed a pectin/zein hydrogel bead system to specifically deliver p40 to the mouse colon, which activated EGFR in colon epithelial cells. Administration of p40-containing beads reduced intestinal epithelial apoptosis and disruption of barrier function in the colon epithelium in an EGFR-dependent manner, thereby preventing and treating DSS-induced intestinal injury and acute colitis. Furthermore, p40 activation of EGFR was required for ameliorating colon epithelial cell apoptosis and chronic inflammation in oxazolone-induced colitis. These data define what we believe to be a previously unrecognized mechanism of probiotic-derived soluble proteins in protecting the intestine from injury and inflammation.
Related JoVE Video
The concept of hepatic artery-bile duct parallelism in the diagnosis of ductopenia in liver biopsy samples.
Am. J. Surg. Pathol.
PUBLISHED: 02-15-2011
Show Abstract
Hide Abstract
Absence of bile ducts (BDs) in >50% of portal tracts is currently the most widely accepted criterion for the diagnosis of ductopenia. In this study, we describe an alternative method for the quantitative assessment of BDs based on the percentage of portal tracts containing unpaired hepatic arteries (HAs). Diagnostic criteria for ductopenia were defined as follows: 1. presence of at least 1 unpaired HA in >10% of all portal tracts; 2. at least 2 unpaired HAs present in different portal tracts in a given sample. In liver biopsies from patients with primary biliary cirrhosis and suspected chronic allograft rejection (n = 32), loss of BD was detected in 59.4% of patients using the unpaired HA method compared with 43.7% (P = 0.31), 21.9% (P = 0.005), and 12.5% (P = 0.001) by the traditional method, depending on specific adequacy criteria used (no adequacy criteria, >10 portal tracts, or >5 complete portal tracts per biopsy, respectively). The percentage of portal tracts containing BD(s) was significantly affected by the degree of portal inflammation, fibrosis stage, percentage of complete portal tracts, and biopsy width, whereas none of these factors influenced the prevalence of unpaired arteries. The unpaired HA method showed higher sensitivity for the detection of mild degrees of loss of BD compared with the traditional method, and was not influenced by factors that affected the percentage of portal tracts containing BDs.
Related JoVE Video
STAT6 activation in ulcerative colitis: a new target for prevention of IL-13-induced colon epithelial cell dysfunction.
Inflamm. Bowel Dis.
PUBLISHED: 02-09-2011
Show Abstract
Hide Abstract
Interleukin 13 (IL-13) is upregulated in ulcerative colitis (UC) and increases colon epithelial permeability by inducing apoptosis and expression of the pore-forming tight junction protein claudin-2. IL-13 induces activation of signal transducer and activator of transcription 6 (STAT6). However, the STAT6 phosphorylation status in patients with UC is unknown, as is the effect of STAT6 inhibition on colonic epithelium exposed to IL-13. The study aims were to determine if mucosal STAT6 phosphorylation is increased in patients with UC, and if STAT6 inhibition attenuates IL-13-induced colon epithelial cell dysfunction.
Related JoVE Video
Loss of TFF1 is associated with activation of NF-?B-mediated inflammation and gastric neoplasia in mice and humans.
J. Clin. Invest.
PUBLISHED: 01-26-2011
Show Abstract
Hide Abstract
Trefoil factor 1 (TFF1) is a tumor suppressor gene that encodes a peptide belonging to the trefoil factor family of protease-resistant peptides. Although TFF1 expression is frequently lost in gastric carcinomas, the tumorigenic pathways this affects have not been determined. Here we show that Tff1-knockout mice exhibit age-dependent carcinogenic histological changes in the pyloric antrum of the gastric mucosa, progressing from gastritis to hyperplasia, low-grade dysplasia, high-grade dysplasia, and ultimately malignant adenocarcinoma. The histology and molecular signatures of gastric lesions in the Tff1-knockout mice were consistent with an inflammatory phenotype. In vivo, ex-vivo, and in vitro studies showed that TFF1 expression suppressed TNF-?-mediated NF-?B activation through the TNF receptor 1 (TNFR1)/I?B kinase (IKK) pathway. Consistent with these mouse data, human gastric tissue samples displayed a progressive decrease in TFF1 expression and an increase in NF-?B activation along the multi-step carcinogenesis cascade. Collectively, these results provide evidence that loss of TFF1 leads to activation of IKK complex-regulated NF-?B transcription factors and is an important event in shaping the NF-?B-mediated inflammatory response during the progression to gastric tumorigenesis.
Related JoVE Video
Combined blockade of Src kinase and epidermal growth factor receptor with gemcitabine overcomes STAT3-mediated resistance of inhibition of pancreatic tumor growth.
Clin. Cancer Res.
PUBLISHED: 01-25-2011
Show Abstract
Hide Abstract
We previously established a mechanistic rationale for Src inhibition as a novel therapeutic target in pancreatic cancer and have identified activated STAT3 as a potential biomarker of resistance to Src inhibition. The purpose of this study was to translate the current understanding of complementary activated tyrosine kinase signaling pathways by targeting Src kinase and epidermal growth factor receptor (EGFR).
Related JoVE Video
Adenoma formation following limited ablation of p120-catenin in the mouse intestine.
PLoS ONE
PUBLISHED: 01-13-2011
Show Abstract
Hide Abstract
p120 loss destabilizes E-cadherin and could therefore result in tumor and/or metastasis-promoting activities similar to those caused by E-cadherin downregulation. Previously, we reported that p120 is essential in the intestine for barrier function, epithelial homeostasis and survival. Conditional p120 ablation in the mouse intestine induced severe inflammatory bowel disease, but long-term cancer-related studies were impossible because none of the animals survived longer than 21 days. Here, we used a tamoxifen-inducible mouse model (Vil-Cre-ER(T2);p120(fl/fl)) to limit the extent of p120 ablation and thereby enable long-term studies. Reducing p120 KO to ?10% of the intestinal epithelium produced long-lived animals outwardly indistinguishable from controls. Effects of prolonged p120 absence were then evaluated at intervals spanning 2 to 18 months. At all time points, immunostaining revealed microdomains of p120-null epithelium interspersed with normal epithelium. Thus, stochastic p120 ablation is compatible with crypt progenitor cell function and permitted lifelong renewal of the p120-null cells. Consistent with previous observations, a barrier defect and frequent infiltration of neutrophils was observed, suggesting that focal p120 loss generates a microenvironment disposed to chronic inflammation. We report that 45% of these animals developed tumors within 18 months of tamoxifen induction. Interestingly, ?-catenin was upregulated in the majority, but none of the tumors were p120 null. Although further work is required to directly establish mechanism, we conclude that limited p120 ablation can promote tumorigenesis by an indirect non-cell autonomous mechanism. Given that byproducts of inflammation are known to be highly mutagenic, we suggest that tumorigenesis in this model is ultimately driven by the lifelong inability to heal chronic wounds and the substantially increased rates of stochastic gene mutation in tissue microenvironments subjected to chronic inflammation. Indeed, although technical issues precluded direct identification of mutations, ?-catenin upregulation in human colon cancer almost invariably reflects mutations in APC and/or ?-catenin.
Related JoVE Video
Epithelial tissues have varying degrees of susceptibility to Kras(G12D)-initiated tumorigenesis in a mouse model.
PLoS ONE
PUBLISHED: 01-11-2011
Show Abstract
Hide Abstract
Activating mutations in the Kras gene are commonly found in some but not all epithelial cancers. In order to understand the susceptibility of different epithelial tissues to Kras-induced tumorigenesis, we introduced one of the most common Kras mutations, Kras(G12D), broadly in epithelial tissues. We used a mouse model in which the G12D mutation is placed in the endogenous Kras locus controlled by inducible, Cre-mediated recombination in tissues expressing cytokeratin 19 including the oral cavity, GI tract, lungs, and ducts of the liver, kidney, and the pancreas. Introduction of the Kras(G12D) mutation in adult mouse tissues led to neoplastic changes in some but not all of these tissues. Notably, many hyperplasias, metaplasias and adenomas were observed in the oral cavity, stomach, colon and lungs, suggesting that exposure to products of the outside environment promotes Kras(G12D)-initiated tumorigenesis. However, environmental exposure did not consistently correlate with tumor formation, such as in the small intestine, suggesting that there are also intrinsic differences in susceptibility to Kras activation. The pancreas developed small numbers of mucinous metaplasias with characteristics of early stage pancreatic intraepithelial neoplasms (PanINs), supporting the hypothesis that pancreatic ducts have the potential to give rise pancreatic cancer.
Related JoVE Video
The apolipoprotein E-mimetic peptide COG112 inhibits NF-kappaB signaling, proinflammatory cytokine expression, and disease activity in murine models of colitis.
J. Biol. Chem.
PUBLISHED: 11-29-2010
Show Abstract
Hide Abstract
Inflammatory bowel disease (IBD), consisting of Crohns disease and ulcerative colitis, is a source of substantial morbidity and remains difficult to treat. New strategies for beneficial anti-inflammatory therapies would be highly desirable. Apolipoprotein (apo) E has immunomodulatory effects and synthetically derived apoE-mimetic peptides are beneficial in models of sepsis and neuroinflammation. We have reported that the antennapedia-linked apoE-mimetic peptide COG112 inhibits the inflammatory response to the colitis-inducing pathogen Citrobacter rodentium in vitro by inhibiting NF-?B activation. We now determined the effect of COG112 in mouse models of colitis. Using C. rodentium as an infection model, and dextran sulfate sodium (DSS) as an injury model, mice were treated with COG112 by intraperitoneal injection. With C. rodentium, COG112 improved the clinical parameters of survival, body weight, colon weight, and histologic injury. With DSS, COG112 ameliorated the loss of body weight, reduction in colon length, and histologic injury, whether administered concurrently with induction of colitis, during induction plus recovery, or only during the recovery phase of disease. In both colitis models, COG112 inhibited colon tissue inducible nitric-oxide synthase (iNOS), KC, TNF-?, IFN-?, and IL-17 mRNA expression, and reduced nuclear translocation of NF-?B, as determined by immunoblot and immunofluorescence confocal microscopy. I?B kinase (IKK) activity was also reduced, which is necessary for activation of the canonical NF-?B pathway. Isolated colonic epithelial cells exhibited marked attenuation of expression of iNOS and the CXC chemokines KC and MIP-2. These studies indicate that apoE-mimetic peptides such as COG112 are novel potential therapies for IBD.
Related JoVE Video
Targeted inhibition of SRC kinase signaling attenuates pancreatic tumorigenesis.
Mol. Cancer Ther.
PUBLISHED: 08-03-2010
Show Abstract
Hide Abstract
Elevated Src expression correlates with malignant potential and metastatic disease in many tumors including pancreatic cancer. We sought to characterize the molecular effects of Src kinase inhibition with dasatinib (BMS-354825), a novel, multitargeted kinase inhibitor that targets Src family kinases in pancreatic ductal adenocarcinoma (PDA). We identified sensitive and resistant PDA cell lines to dasatinib treatment and tested the molecular effects of Src inhibition in vitro and in vivo. We show for the first time that cellular localization of Src expression affects survival in patients with PDA. Pancreatic tumors with increased membranous expression of Src resulted in decreased survival compared with tumors that had increased cytoplasmic Src expression. Src kinase inhibition with dasatinib markedly inhibits cell proliferation, migration, invasion, cell cycle progression and anchorage-independent growth, and stimulates apoptosis. This was accompanied by decreased phosphorylation of Src, focal adhesion kinase, paxillin, AKT, signal transducers and activators of transcription 3 (STAT3), extracellular signal-regulated kinase, and mitogen-activated protein kinase (MAPK), as well as decreased cyclin D1 expression in a time- and concentration-dependent manner. Furthermore, small interfering RNA to Src results in a significant decrease in cell proliferation, invasion, and migration of pancreatic cancer cells. Dasatinib treatment also inhibits in vivo pancreatic tumor growth. Mechanisms of resistance to Src inhibition seem to be related to a lack of inhibition of STAT3 and MAPK signaling. These results establish a mechanistic rationale for Src inhibition with dasatinib as a therapeutic target in the treatment of pancreatic cancer and identify potential biomarkers of resistance to Src inhibition.
Related JoVE Video
The deployment of a tissue request tracking system for the CHTN: a case study in managing change in informatics for biobanking operations.
BMC Med Inform Decis Mak
PUBLISHED: 06-02-2010
Show Abstract
Hide Abstract
Managing change has not only been recognized as an important topic in medical informatics, but it has become increasingly important in translational informatics. The move to share data, together with the increasing complexity and volume of the data, has precipitated a transition from locally stored worksheet and flat files to relational data bases with object oriented interfaces for data storage and retrieval. While the transition from simple to complex data structures, mirroring the transition from simple to complex experimental technologies, seems natural, the human factor often fails to be adequately addressed leading to failures in managing change.
Related JoVE Video
KSR1 protects from interleukin-10 deficiency-induced colitis in mice by suppressing T-lymphocyte interferon-? production.
Gastroenterology
PUBLISHED: 05-26-2010
Show Abstract
Hide Abstract
Immunological disorders of the gastrointestinal tract such as inflammatory bowel disease often result in recurrent and persistently elevated levels of proinflammatory cytokines. Kinase suppressor of Ras 1 (KSR1) is involved in tumor necrosis factor-mediated colon epithelial cell survival, yet its role in chronic inflammation has not been defined. In this study, we tested the hypothesis that KSR1 is protective against spontaneous experimental colitis.
Related JoVE Video
Detailed in vivo analysis of the role of Helicobacter pylori Fur in colonization and disease.
Infect. Immun.
PUBLISHED: 04-26-2010
Show Abstract
Hide Abstract
Helicobacter pylori persistently colonizes the harsh and dynamic environment of the stomach in over one-half of the worlds population and has been identified as a causal agent in a spectrum of pathologies that range from gastritis to invasive adenocarcinoma. The ferric uptake regulator (Fur) is one of the few regulatory proteins that has been identified in H. pylori. Fur regulates genes important for acid acclimation and oxidative stress and has been shown to be important for colonization of H. pylori in both murine and Mongolian gerbil models of infection. To more thoroughly define the role of Fur in vivo, we conducted an extensive temporal analysis of the location of, competitive ability of, and resultant pathology induced by a Deltafur strain in the Mongolian gerbil model of infection and compared the results to results for its wild-type parent. We found that at the earliest time points postinfection, significantly more Deltafur bacteria than wild-type bacteria were recovered. However, this trend was reversed by day 3, when there was significantly increased recovery of the wild-type strain. The increased recovery of the Deltafur strain at 1 day postinfection reflected increased recovery from both the corpus and the antrum of the stomach. When the wild-type strain was allowed to colonize first, the Deltafur strain was unable to compete for colonization at any time postinfection. However, when the Deltafur strain was allowed to colonize first, the wild type efficiently outcompeted the Deltafur strain only at early times postinfection. Finally, we demonstrated that there was a delay in the development and severity of inflammation and pathology of the Deltafur strain in the gastric mucosa even after comparable levels of colonization occurred. Together, these data indicate that H. pylori Fur is most important at early stages of infection and illustrate the importance of the ability of H. pylori to adapt to its constantly fluctuating environment when it is establishing infection, inflammation, and disease.
Related JoVE Video
p120-catenin is essential for maintenance of barrier function and intestinal homeostasis in mice.
J. Clin. Invest.
PUBLISHED: 03-24-2010
Show Abstract
Hide Abstract
Epithelial-cadherin (E-cadherin) is a master organizer of the epithelial phenotype. Its function is regulated in part by p120-catenin (referred to herein as p120), a cytoplasmic binding partner that directly regulates cadherin stability. As it has been suggested that cadherins have a role in inflammatory bowel disease (IBD), we sought to investigate this further by assessing the effect of p120 deficiency in mouse small intestine and colon. p120 conditional KO mice were superficially normal at birth but declined rapidly and died within 21 days. Cell-cell adhesion defects and inflammation led to progressive mucosal erosion and terminal bleeding, similar to what is observed in a dominant-negative cadherin mouse model of IBD. Additionally, selective loss of adherens junctions and accumulation of atypical COX-2-expressing neutrophils in p120-null areas of the colon were observed. To elucidate the mechanism, direct effects of p120 deficiency were assessed in vitro in a polarizing colon cancer cell line. Notably, transepithelial electrical resistance was dramatically reduced, neutrophil binding was increased 30 fold, and levels of COX-2, an enzyme associated with IBD, were markedly increased in neutrophils. Our data suggest that p120 loss disrupts the neonatal intestinal barrier and amplifies neutrophil engagement and that these changes lead to catastrophic inflammation during colonization of the neonatal gut with bacteria and other luminal antigens. Thus, we conclude that p120 has an essential role in barrier function and epithelial homeostasis and survival in the intestine.
Related JoVE Video
Loss of FOXA1/2 is essential for the epithelial-to-mesenchymal transition in pancreatic cancer.
Cancer Res.
PUBLISHED: 02-16-2010
Show Abstract
Hide Abstract
FOXA1 and FOXA2, members of the forkhead transcription factor family, are critical for epithelial differentiation in many endoderm-derived organs, including the pancreas. However, their role in tumor progression is largely unknown. Here, we identified FOXA1 and FOXA2 as important antagonists of the epithelial-to-mesenchymal transition (EMT) in pancreatic ductal adenocarcinoma (PDA) through their positive regulation of E-cadherin and maintenance of the epithelial phenotype. In human PDA samples, FOXA1/2 are expressed in all epithelia from normal to well-differentiated cancer cells, but are lost in undifferentiated cancer cells. In PDA cell lines, FOXA1/2 expression is consistently suppressed in experimental EMT models and RNAi silencing of FOXA1/2 alone is sufficient to induce EMT. Conversely, ectopic FOXA1/2 expression can potently neutralize several EMT-related E-cadherin repressive mechanisms. Finally, ectopic FOXA2 expression could reactivate E-cadherin expression in a PDA cell line with extensive promoter hypermethylation. In fact, demethylation-mediated reactivation of E-cadherin expression in these cells required concurrent reactivation of endogenous FOXA2 expression. We conclude that suppression of FOXA1/2 expression is both necessary and sufficient for EMT during PDA malignant progression.
Related JoVE Video

What is Visualize?

JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

How does it work?

We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.

Video X seems to be unrelated to Abstract Y...

In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.