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Anna Velcich

Publications and source records attributed to Anna Velcich.

13 recordsLinked to original sources
PubMed2007

Methotrexate-induced mucositis in mucin 2-deficient mice.

The mucin Muc2 or Mycin2 (Muc2), which is the main structural component of the protective mucus layer, has shown to be upregulated during chemotherapy-induced mucositis. As Muc2 has shown to have protective capacities, upregulation of Muc2 may be a counter reaction of the intestine protecting against mucositis. Therefore, increasing Muc2 protein levels could be a therapeutic target in mucositis prevention or reduction. Our aim was to determine the role of Muc2 in chemotherapy-induced mucositis. Mucositis was induced in Muc2 knockout (Muc2(-/-)) and wild type (Muc2(+/+)) mice by injecting methotrexate (MTX). Animals were weighed and sacrificed on Days 2-6 after MTX treatment and jejunal segments were analyzed. Before MTX treatment, the small intestine of Muc2(+/+) and Muc2(-/-) mice were similar with respect to epithelial morphology and proliferation. Moreover, sucrase-isomaltase and trefoil factor-3 protein expression levels were comparable between Muc2(+/+) and Muc2(-/-) mice. Up to Day 3 after MTX treatment, percentages of weight-loss did not differ. Thereafter, Muc2(+/+) mice showed a trend towards regaining weight, whereas Muc2(-/-) mice continued to lose weight. Surprisingly, MTX-induced intestinal damage of Muc2(-/-) and Muc2(+/+) mice was comparable. Prior to MTX-injection, tumor necrosis factor-alpha and interleukin-10 mRNAs were upregulated in Muc2(-/-) mice, probably due to continuous exposure of the intestine to luminal antigens. Muc2 deficiency does not lead to an increase in chemotherapy-induced mucositis. A possible explanation is the mechanism by which Muc2 deficiency may trigger the immune system to release interleukin-10, an anti-inflammatory cytokine before MTX-treatment.

Animals↗
PubMed2006

Histone deacetylase 3 (HDAC3) and other class I HDACs regulate colon cell maturation and p21 expression and are deregulated in human colon cancer.

Inhibitors of histone deacetylases (HDACs) induce growth arrest, differentiation, and apoptosis of colon cancer cell lines in vitro and have demonstrated anti-cancer efficacy in clinical trials. Whereas a role for HDAC1 and -2 in mediating components of the HDAC inhibitor response has been reported, the role of HDAC3 is unknown. Here we demonstrate increased protein expression of HDAC3 in human colon tumors and in duodenal adenomas from Apc1638(N/+) mice. HDAC3 was also maximally expressed in proliferating crypt cells in normal intestine. Silencing of HDAC3 expression in colon cancer cell lines resulted in growth inhibition, a decrease in cell survival, and increased apoptosis. Similar effects were observed for HDAC2 and, to a lesser extent, for HDAC1. HDAC3 silencing also selectively induced expression of alkaline phosphatase, a marker of colon cell maturation. Concurrent with its effect on cell growth, overexpression of HDAC3 and other Class I HDACs inhibited basal and butyrate-induced p21 transcription in a Sp1/Sp3-dependent manner, whereas silencing of HDAC3 stimulated p21 promoter activity and expression. However, the magnitude of the effects elicited by silencing of individual Class I HDACs was significantly less than that induced by HDAC inhibitors. These findings identify HDAC3 as a gene deregulated in human colon cancer and as a novel regulator of colon cell maturation and p21 expression. These findings also demonstrate that multiple Class I HDACs are involved in repressing p21 and suggest that the growth-inhibitory and apoptotic effects induced by HDAC inhibitors are probably mediated through the inhibition of multiple HDACs.

Apoptosis↗
PubMed2006

Muc2-deficient mice spontaneously develop colitis, indicating that MUC2 is critical for colonic protection.

BACKGROUND & AIMS: Expression of mucin MUC2, the structural component of the colonic mucus layer, is lowered in inflammatory bowel disease. Our aim was to obtain insight in the role of Muc2 in epithelial protection. METHODS: Muc2 knockout (Muc2(-/-)) and Muc2 heterozygous (Muc2(+/-)) mice were characterized and challenged by a colitis-inducing agent, dextran sulfate sodium (DSS). We monitored clinical symptoms, intestinal morphology, and differences in intestine-specific protein and messenger RNA levels. RESULTS: The Muc2(-/-) mice showed clinical signs of colitis (as of 5 weeks), aggravating as the mice aged. Microscopic analysis of the colon of Muc2(-/-) mice showed mucosal thickening, increased proliferation, and superficial erosions. Colonic goblet cells in the Muc2(-/-) mice were negative for Muc2, but trefoil factor 3 was still detectable. In Muc2(-/-) mice, transient de novo expression of Muc6 messenger RNA was observed in the distal colon. On day 2 of DSS treatment, the histologic damage was more severe in Muc2(+/-) versus wild-type (Muc2(+/+)) mice, but the disease activity index was not yet different. By day 7, the disease activity index and histologic score were significantly elevated in Muc2(+/-) versus Muc2(+/+) mice. The disease activity index of the Muc2(-/-) mice was higher (versus both Muc2(+/+) and Muc2(+/-) mice) throughout DSS treatment. The histologic damage in the DSS-treated Muc2(-/-) mice was different compared with Muc2(+/+) and Muc2(+/-) mice, with many crypt abscesses instead of mucosal ulcerations. CONCLUSIONS: This study shows that Muc2 deficiency leads to inflammation of the colon and contributes to the onset and perpetuation of experimental colitis.

Animals↗
PubMed2005

Inactivation of p21WAF1/cip1 enhances intestinal tumor formation in Muc2-/- mice.

In the Apc1638(+/-) mouse model of intestinal tumorigenesis, targeted inactivation of the cyclin-dependent kinase inhibitor p21(WAF1/cip1) is highly effective in enhancing Apc-initiated tumor formation in the intestine. Because p21(WAF1/cip1) plays a critical role in regulating intestinal cell proliferation, maturation, and tumorigenesis, we examined whether its inactivation would enhance tumor formation in a different mouse model of colon cancer. Therefore, we mated p21(-/-) mice with mice carrying a genetic deficiency of the Muc2 gene, which encodes the major gastrointestinal mucin. Muc2(-/-) mice develop tumors in the small and large intestine and the rectum, but in contrast to tumors in Apc1638(+/-) mice, this does not involve increased expression or nuclear localization of beta-catenin. We found that inactivation of p21(WAF1/cip1) significantly increased the frequency and size of intestinal tumors in Muc2 knockout mice and also led to development of more invasive adenocarcinomas. This enhanced tumorigenesis significantly decreased mouse life span. Further, inactivation of p21(WAF1/cip1) increased cell proliferation, decreased apoptosis, and decreased intestinal trefoil factor expression in the mucosa of both the small and large intestine. Surprisingly, reduced expression of p27(kip1) was also observed in the Muc2(-/-), p21(+/-), and p21(-/-) mice. In contrast, the expression of c-myc was significantly elevated. Thus, p21 modulates the formation of tumors whose initiation does (Apc) or does not (Muc2) involve altered beta-catenin-Tcf4 signaling, but which may converge on common elements downstream of this signaling pathway.

Adenocarcinoma↗
PubMed2005

Dietary components modify gene expression: implications for carcinogenesis.

Mouse genetic models that probe important pathways in intestinal cell maturation, such as cell-cycle regulation, apoptosis, and, especially, lineage specific differentiation, have provided profound insight into the underlying mechanisms of intestinal tumor formation and progression. However, a wealth of epidemiological and experimental data indicates that environment, especially the diet, is a principal determinant of relative risk for tumor development. We have demonstrated that even in mouse models in which tumor incidence is strongly initiated by genetic manipulation of genes, such as Apc, p21(WAF1/cip1), and p27(Kip1), a Western-style diet that is high in fat and low in calcium and vitamin D can dramatically increase and accelerate tumor formation. Moreover, experiments show that modulation of calcium and vitamin D levels can substantially influence tumor formation in both the mouse genetic models, as well as in a new dietary model that appears to mimic the development of sporadic colon cancer. Finally, analysis of gene expression profiles provides important insights into how diets may alter metabolic profiles and regulatory pathways that influence probability of tumor formation in the histologically and physiologically normal intestinal mucosa.

Animals↗
PubMed2004

Quantitative rather than qualitative differences in gene expression predominate in intestinal cell maturation along distinct cell lineages.

Several cell types are present in the intestinal epithelium that likely arise from a common precursor, the stem cell, and each mature cell type expresses a unique set of genes that characterizes its functional phenotype. Although the process of differentiation is intimately linked to the cessation of proliferation, the mechanisms that dictate intestinal cell fate determination are not well characterized. To investigate the reprogramming of gene expression during the cell lineage allocation/differentiation process, we took advantage of a unique system of two clonal derivatives of HT29 cells, Cl16E and Cl19A cells, which spontaneously differentiate as mucus producing goblet and chloride-secreting cells, respectively, as a function of time. By profiling gene expression, we found that these two cell lines show remarkably similar kinetics of change in gene expression and common clusters of coordinately regulated genes. This demonstrates that lineage-specific differentiation of intestinal epithelial cells is characterized overall by the sequential recruitment of functionally similar gene sets independent of the final phenotype of the mature cells.

Animals↗
PubMed2003

Repression of MUC2 gene expression by butyrate, a physiological regulator of intestinal cell maturation.

Sodium butyrate (NaB) inhibits proliferation, stimulates apoptosis, and promotes differentiation of human colon cancer cells along the absorptive phenotype. In vitro, butyrate induces a switch from cells with a secretory to an absorptive phenotype. Here, we report that NaB specifically represses the expression of the MUC2 gene, a differentiation marker of the secretory goblet cell lineage, in forskolin- and 12-O-tetradecanoylphorbol 13-acetate-induced HT29 cells, and Cl.16E cells, a clonal derivative of HT29 cells that spontaneously differentiates into goblet cells. Thus, NaB repression is independent of the nature of the stimulus that triggers MUC2 expression. Further, repression was independent of new protein synthesis. Our results suggest that inhibition of MUC2 is linked to the ability of butyrate to repress histone deacetylase activity, since trichostatin A, another inhibitor of histone deacetylases, also inhibited MUC2 expression in induced HT29 cells. Finally, we demonstrate that the NaB effect is specific for this marker of the secretory cell lineage, since carcinoembryonic antigen, which is expressed in both the secretory and absorptive cells, is induced by NaB. Thus, the NaB repression of a definitive function of the secretory cell lineage is a further mechanism, in addition to the effects on proliferation and apoptotic pathways, through which butyrate can regulate intestinal homeostasis.

Butyrates↗
PubMed2003

Application of gene expression profiling to colon cell maturation, transformation and chemoprevention.

Methods for high-throughput analysis of profiles of gene expression that assay thousands of genes simultaneously are powerful approaches for understanding and classifying cell and tissue phenotype. This includes analysis of normal pathways of cell maturation and their perturbation in transformation, the sensitivity and mechanism of response of normal and tumor cells to physiological and pharmacological agents, and modulation of tumor risk and progression by nutritional factors. However, the complex data generated by such approaches raise difficulties in analysis. We will describe some of the methods we have used in analyzing databases generated in a number of projects in our laboratories. These include: the role of k-ras mutations in colon cell transformation; the role of p21(WAF1/cip1) in intestinal tumor formation and response to sulindac; the development of the absorptive and goblet cell lineages; sensitivity of colonic cells to chemotherapeutic agents; mechanisms that regulate c-myc expression utilizing novel methods of transcriptional imaging; and interaction of nutritional and genetic factors in modulation of intestinal tumor formation.

Animals↗
PubMed2002

Novel detection and differential utilization of a c-myc transcriptional block in colon cancer chemoprevention.

Mutations in the adenomatous polyposis coli (APC) gene, which initiate almost all human colon cancers, directly target the proto-oncogene, c-myc, by elevating beta-catenin/T-cell factor (TCF) signaling. We have shown that agents ascribed chemopreventive activity for colon cancer in fact also stimulate beta-catenin/TCF activity in vitro. Their effects on c-myc transcription were assayed using a novel variant of fluorescence in situ hybridization that detects c-myc transcription sites in intact nuclei. Increased transcriptional initiation of c-myc induced by the short-chain fatty acid, butyrate, consistent with elevated beta-catenin/TCF activity, was efficiently abrogated by a block to transcriptional elongation, resulting in decreased c-myc expression. 1alpha,25-Dihydroxyvitamin D(3) also induced transcriptional blockage. In contrast, the nonsteroidal anti-inflammatory drug, sulindac, increased c-myc expression, an effect attributable at least in part to its failure to induce transcriptional blockage. We have described a novel approach for evaluating the effects of chemopreventive agents on the expression of a gene critical in colonic tumorigenesis.

Anticarcinogenic Agents↗
PubMed2002

Colorectal cancer in mice genetically deficient in the mucin Muc2.

The gastrointestinal tract is lined by a layer of mucus comprised of highly glycosylated proteins called mucins. To evaluate the importance of mucin in intestinal carcinogenesis, we constructed mice genetically deficient in Muc2, the most abundant secreted gastrointestinal mucin. Muc2-/- mice displayed aberrant intestinal crypt morphology and altered cell maturation and migration. Most notably, the mice frequently developed adenomas in the small intestine that progressed to invasive adenocarcinoma, as well as rectal tumors. Thus, Muc2 is involved in the suppression of colorectal cancer.

Adenocarcinoma↗
PubMed2002

Short chain fatty acids and colon cancer.

The development of intestinal cancer involves complex genetic and epigenetic alterations in the intestinal mucosa. The principal signaling pathway responsible for the initiation of tumor formation, the APC-beta-catenin-TCF4 pathway, regulates both cell proliferation and colonic cell differentiation, but many other intrinsic and extrinsic signals also modulate these cell maturation pathways. The challenge is to understand how signaling and cell maturation are also modulated by nutritional agents. Through gene expression profiling, we have gained insight into the mechanisms by which short chain fatty acids regulate these pathways and the differences in response of gene programs, and of the specific regulation of the c-myc gene, to physiological regulators of intestinal cell maturation, such as butyrate, compared with pharmacological regulators such as the nonsteroidal antiinflammatory drug sulindac. Moreover, we used a combination of gene expression profiling of the response of cells in culture to sulindac and the response of the human mucosa in subjects treated with sulindac for 1 month, coupled with a mouse genetic model approach, to identify the cyclin dependent kinase inhibitor p21(WAF1/Cip1) as an important suppressor of Apc-initiated intestinal tumor formation and a necessary component for tumor inhibition by sulindac. Finally, the mucous barrier, secreted by intestinal goblet cells, is the interface between the luminal contents and the intestinal mucosa. We generated a mouse genetic model with a targeted inactivation of the Muc2 gene that encodes the major intestinal mucin. These mice have no recognizable goblet cells due to the failure of cells to synthesize and store mucin. This leads to perturbations in intestinal crypt architecture, increased cellular proliferation and rates of cell migration, decreased apoptosis and development of adenomas and adenocarcinomas in the small and large intestine and the rectum.

Animals↗
PubMed2002

Environment-gene interactions in intestinal cancer.

Only 5-10% of all colorectal cancer in the United States can be directly attributed to inheritance of genetic predisposition for tumor development. Thus, the vast majority of colorectal cancer is classified as sporadic, and in these patients environmental factors--particularly the diet--play a major role in determining the probability of tumor formation and its progression. Investigations of how dietary components interact with genetic factors in cancer development have been extremely productive in terms of understanding the subtle and complex mechanisms that maintain homeostasis of the intestinal mucosa, and how perturbations in these mechanisms cause disease. We have found that the cyclin-dependent kinase inhibitor p21(WAF1/cip1) plays a major role in regulating several aspects of mucosal homeostasis and the response to dietary and pharmacologic modulators of tumorigenesis; that disruption of lineages of differentiation of intestinal epithelial cells are intimately involved in tumor formation; and that important pathways that contribute to normal homeostasis and cancer development may be coordinately regulated by mitochondrial function, with the mitochondrial membrane potential playing a key role. Several lines of evidence from our work have also suggested that intestinal epithelial cells have adapted to the environment that they usually encounter. This renders the cells competent to efficiently utilize factors in the intestinal lumen in normal metabolic and signaling pathways that contribute to homeostasis.

Animals↗