Expression of Slug is regulated by c-Myb and is required for invasion and bone marrow homing of cancer cells of different origin.

In metastatic cancer cells, the process of invasion is regulated by several transcription factors that induce changes required for migration and resistance to apoptosis. Slug (SNAI2, Snail2) is involved in epithelial mesenchymal transition in physiological and in pathological contexts. We show here that in embryonic kidney, colon carcinoma, chronic myeloid leukemia-blast crisis, and in neuroblastoma cells, expression of Slug is transcriptionally regulated by c-Myb via Myb binding sites in the 5'-flanking region and in the first intron of the slug gene. In embryonic kidney and neuroblastoma cells, c-Myb induced vimentin, fibronectin, and N-cadherin expression and membrane ruffling via actin polymerization consistent with the acquisition of a mesenchymal-like phenotype. Furthermore, down-regulation of endogenous c-Myb levels in colon carcinoma cells led to increased expression of E-cadherin and reduced levels of vimentin. Some of these changes are predominantly Slug-dependent as Slug silencing via RNA interference (RNAi) reverts the cells to a quasi-parental condition. Changes in gene expression and morphology induced by c-Myb-activated Slug correlated with increased ability to migrate (embryonic kidney) and to invade through a Matrigel membrane (embryonic kidney, colon carcinoma, neuroblastoma). c-Myb-dependent Slug expression was also essential for the homing of chronic myeloid leukemia K562 cells to the bone marrow. In summary, we show here that the proto-oncogene c-Myb controls Slug transcription in tumor cells of different origin. Such a regulatory pathway contributes to the acquisition of invasive properties that are important for the metastatic process.

Altered expression of innate immunity genes in different intestinal sites of children with ulcerative colitis.

BACKGROUND: Innate immunity has been very rarely investigated in ulcerative colitis and never in paediatrics. The present study was aimed at describing expression of innate immunity genes (NOD2, RIP2, α-defensins HD5 and HD6) in inflamed colon and in ileum of children with ulcerative colitis. Expression of TNFα and IL-1ß was also analyzed. METHODS: 15 children with ulcerative colitis (9 pancolitis, 6 left-sided colitis) and 10 control children were enrolled. mRNA and protein expressions were detected by real time PCR and western blot assays. RESULTS: NOD2, RIP2, IL-1ß, TNFα expression levels were significantly increased in colonic mucosa of patients compared to controls (p<0.01). These genes were also upregulated (p<0.01) in the ileum of both pancolitis and left-sided colitis children. HD5 and HD6 were significantly upregulated (p<0.01) in the inflamed colon of patients as well as in the ileum of those with pancolitis. CONCLUSIONS: An increased mucosal expression of innate immunity genes was found in the inflamed colon of children with ulcerative colitis, outlining the role of the innate immune response in disease pathogenesis. Involvement of the ileum in ulcerative colitis suggests that an immune activation can also be established in intestinal sites classically uninvolved by the inflammation, carrying implications for the treatment and course of the disease.

Interaction between glutathione-S-transferase polymorphisms, smoking habit, and HPV infection in cervical cancer risk.

PURPOSE: Human papillomavirus (HPV) infection is considered the major cause of cervical cancer (CC), but a number of infected women do not develop invasive lesions, suggesting the role of genetic susceptibility and environmental co-factors for cancer outbreak. The aim of this study was to investigate whether some GST polymorphisms could influence the risk to develop CC, either by themselves or in combination with smoking habit, in a cohort of high-risk HPV (HR-HPV) infected Italian women. METHODS: The study population comprises 192 Italian women including 81 HR-HPV infected women bearing cervical lesions and 111 healthy controls. The cases include: 26 low-grade squamous intraepithelial lesions (LSILs), 30 high-grade-SIL, and 25 CCs, while controls were all negative for HPV. DNA was extracted from peripheral blood samples or cytobrush and individuals were genotyped for GSTM1, GSTT1, and GSTP1 polymorphisms using PCR and PCR/RFLP techniques. RESULTS: On studying the association of GSTs gene polymorphisms with cervical cancer lesions, the combination of GSTM1 null, GSTT1 null and GSTP1 AA genotypes, independently on smoking habit, seems to be related to a 5.7-fold increased risk of developing CLs with a considerable statistical significance (P = 0.0091). CONCLUSIONS: We suggest that the investigation of multiple gene polymorphisms, versus single genes, could contribute to a better understanding of the effect of susceptibility genes on cancer risk.

DNA repair capacity and acute radiotherapy adverse effects in Italian breast cancer patients.

Therapeutic exposure to ionising radiation can induce normal tissue side effects which consistently differ among individuals suggesting a possible genetic control. One approach to elucidate the underlying mechanisms is to analyse the relation between genetic traits, biomarkers of in vitro DNA damage and side toxicity in vivo. 43 breast cancer (BC) patients receiving radiotherapy after a breast-conserving surgery were recruited together with 34 age- and sex-matched healthy controls. Adverse tissue reactions were recorded as indicators of radiotherapy susceptibility. All blood samples from both patients (35) and controls (34) were irradiated in vitro and DNA primary damage and repair kinetic were measured through Comet assay. All study subjects were genotyped for XRCC1, OGG1 and XRCC3 gene polymorphisms. In our small groups we found a positive association between XRCC1 variant allele (399Gln) and the occurrence of breast cancer [p=0.01; OR=2.41, 95%CI (1.24-4.66)]. BC patients showed a higher degree of basal (p<0.001) and X-ray induced DNA damage (p<0.01) when compared to healthy controls. A reduced repair ability was found in BC patients showing high degrees of tissue side effects as classified by Radiation Morbidity Scoring Scheme. BC patients showed an impairment of their DNA repair capacity associated with the development of radiation sensitivity but not with polymorphisms in any of the considered genes.

Activation of NOD2-mediated intestinal pathway in a pediatric population with Crohn's disease.

BACKGROUND: NOD2 is an intracellular protein involved in host recognition of specific bacterial molecules and is genetically associated with several inflammatory diseases, including Crohn's disease (CD). NOD2 stimulation activates the transcription factor, NF-kappaB, through RIP2, a caspase-recruitment domain-containing kinase. NOD2/RIP2 signaling also mediates the activation of antimicrobial peptides such as human alpha-defensin 5 (HD-5) and human alpha-defensin 6 (HD-6), both produced by Paneth cells. The present study is aimed at describing the downstream events triggered specifically by NOD2 induction in order to demonstrate that the protein, other than overexpressed, is also physiologically associated with RIP2 and Erbin in the bioptic intestinal inflamed specimens of children affected by CD. METHODS: Fifteen children with CD and 10 children used as controls were entered in the study. Mucosal biopsy specimens were taken during endoscopy and mRNA and protein expressions were detected by using real-time polymerase chain reaction and Western blot. RESULTS: NOD2 is able to form an immunocomplex with the kinase RIP2. As compared to controls, in the inflamed mucosa of patients both mRNA and protein expression levels of RIP2 are increased, and its active phosphorylated form is overexpressed. CONCLUSIONS: In this study we provide for the first time ex vivo evidence of physiologically relevant protein interactions with NOD2, which are able to trigger the innate immune response in intestinal mucosal specimens of children with CD.

Occupational exposure to antineoplastic agents induces a high level of chromosome damage. Lack of an effect of GST polymorphisms.

The aim of our study was to investigate whether occupational exposure to antineoplastic drugs (AND) resulted in genetic damage, possibly indicative of adverse health effects in the long term. We performed a chromosomal aberrations (CA) analysis in peripheral blood lymphocytes (PBL) of a group of 76 trained nurses occupationally exposed to AND. Furthermore, we analysed whether genetic polymorphisms in four metabolic genes of the glutathione S-transferase (GST) family involved in antineoplastic drugs detoxification (GSTM1, GSTT1, GSTP1, GSTA1) had any effect on the yield of chromosomal aberrations in nurses exposed to antineoplastic agents. The exposed group showed a very significant increase of genetic damage (p<0.0001) potentially indicative of an increased risk of cancer. Unexpectedly, besides the elevated level of chromatid-type aberrations usually related to exposure to chemical agents, we found also severe chromosome damages such as chromosome deletions and dicentric chromosomes, usually related to radiation exposure. No significant association was detected between all GSTs genotypes and chromosome damage. In conclusion, our data show how the occupational exposure to AND is associated to a potential cancer risk, suggesting that current prevention methods do not completely eliminate opportunities for exposure and supporting the need to improve the actual safety practices.