Fibroblast activation and senescence in oral cancer.

There is now compelling evidence that the tumour stroma plays an important role in the pathogenesis of cancers of epithelial origin. The pre-eminent cell type of the stroma is carcinoma-associated fibroblasts. These cells demonstrate remarkable heterogeneity with activation and senescence being common stress responses. In this review, we summarise the part that these cells play in cancer, particularly oral cancer, and present evidence to show that activation and senescence reflect a unified programme of fibroblast differentiation. We report advances concerning the senescent fibroblast metabolome, mechanisms of gene regulation in these cells and ways in which epithelial cell adhesion is dysregulated by the fibroblast secretome. We suggest that the identification of fibroblast stress responses may be a valuable diagnostic tool in the determination of tumour behaviour and patient outcome. Further, the fact that stromal fibroblasts are a genetically stable diploid cell population suggests that they may be ideal therapeutic targets and early work in this context is encouraging.

Pleiotropic actions of miR-21 highlight the critical role of deregulated stromal microRNAs during colorectal cancer progression.

The oncogene microRNA-21 (miRNA; miR-21) is overexpressed in most solid organ tumours; however, a recent examination of stage II colorectal cancer (CRC) specimens suggests this may be a stromal phenomenon and not only a feature of cancer cells. In vitro and in vivo studies show that miR-21 has potent pro-metastatic effects in various malignant carcinoma cell lines. The tumour microenvironment has also been identified as a key actor during the metastatic cascade; however to date the significance of deregulated miR-21 expression within the cancer-associated stroma has not been examined. In the present study, a quantitative RT-PCR-based analysis of laser microdissected tissue confirmed that miR-21 expression is associated with a four-fold mean increase in CRC stroma compared with normal tissue. In situ hybridisation using locked nucleic acid probes localised miR-21 expression predominantly to fibroblasts within tumour-associated stroma. To study the molecular and biological impact of deregulated stromal miR-21 in CRC, stable ectopic expression was induced in immortalised fibroblasts. This resulted in upregulated α-smooth muscle actin expression implying miR-21 overexpression is driving the fibroblast-to-myofibroblast transdifferentiation. Conditioned medium from miR-21-overexpressing fibroblasts protected CRC cells from oxaliplatin-induced apoptosis and increased their proliferative capacity. 3D organotypic co-cultures containing fibroblasts and CRC cells revealed that ectopic stromal miR-21 expression was associated with increased epithelial invasiveness. Reversion-inducing cysteine-rich protein with kazal motifs, an inhibitor of matrix-remodelling enzyme MMP2, was significantly downregulated by ectopic miR-21 in established and primary colorectal fibroblasts with a reciprocal rise in MMP2 activity. Inhibition of MMP2 abrogated the invasion-promoting effects of ectopic miR-21. This data, which characterises a novel pro-metastatic mechanism mediated by miR-21 in the CRC stroma, highlights the importance of miRNA deregulation within the tumour microenvironment and identifies a potential application for stromal miRNAs as biomarkers in cancer.

PIR2/Rnf144B regulates epithelial homeostasis by mediating degradation of p21WAF1 and p63.

ΔNp63 is a transcription factor that is critical for the development of stratified epithelia and is overexpressed or amplified in >80% of squamous cell carcinomas (SCCs). We identified the RING finger E3 ubiquitin ligase PIR2/Rnf144b as a direct transcriptional target of ΔNp63α and showed that its expression parallels that of ΔNp63α in keratinocytes, SCC cell lines and SCCs. We used primary keratinocytes as a model system to investigate the function of PIR2/Rnf144b in stratified epithelia. Depletion of PIR2/Rnf144b severely impaired keratinocyte proliferation and differentiation, associated with accumulation of p21(WAF1/CIP1); a known target of PIR2/Rnf144b. More importantly, we found that PIR2/Rnf144b binds and mediates proteasomal degradation of ΔNp63α, generating a hitherto unknown auto-regulatory feedback loop. These findings substantiate PIR2/Rnf144b as a potentially critical component of epithelial homeostasis, acting downstream of ΔNp63α to regulate cellular levels of p21(WAF1/CIP1) and ΔNp63α.

HMGA molecules in neuroblastic tumors.

The high mobility group A (HMGA) proteins are thought to work as ancillary transcription factors and to regulate the expression of a growing number of genes through direct binding to DNA or via protein-protein interactions. Both HMGA1 and HMGA2 are important regulators of basic biological processes, including cell growth, differentiation and transformation. Their qualitatively or quantitatively altered expression has been described in a number of human tumors. We studied and review here their expression in neuroblastic tumors. HMGA2 is expressed only in a subset of ex vivo neuroblastoma (NB) tumors and in the embryonic adrenal gland, but it is undetectable in the adult adrenal gland, suggesting that its anomalous expression might be associated with NB tumorigenesis and/or tumor progression. In vitro, its expression is easily detectable in retinoic acid (RA)-resistant cell lines. The exogenous expression of HMGA2 is sufficient to convert RA-sensitive SY5Y NB cells into RA-resistant cells, thus suggesting that HMGA2 might be a relevant player in determining NB cell responses to endogenous or therapeutically important growth inhibitory substances. In contrast, HMGA1 expression is readily detectable in all NB cell lines and tumors, but its expression is consistently higher in less differentiated NBs compared with ganglioneuromas and ganglioneuroblastomas. Interestingly, RA increases HMGA1 expression in RA-resistant NB cells but inhibits it in cells undergoing RA-induced growth inhibition and neuronal differentiation. Our studies indicate that HMGA molecules might be biologically and pathologically relevant factors in neuroblastic tumor development and progression.

Effects of selenium and zinc supplementation on nutritional status in patients with cancer of digestive tract.

OBJECTIVE: To evaluate the effect of oral administration of selenium and zinc tablets in patients with cancer of the digestive tract during chemotherapy. DESIGN: A case-control, randomized study. SETTING: Medical Oncology, II University of Naples, Naples, Italy. SUBJECTS: A total of 60 patients (median age 55 y, range 46-61 y) with diagnosis of gut cancer were randomized in 1999. Patients were treated for 60 days with chemotherapy. INTERVENTIONS: Trace elements were measured by atomic absorption spectroscopy. The nutritional status of the patients was assessed by biochemical and bio-impedance analysis (BIA) parameters in basal condition and after 60 days of treatment. Oral administration of selenium and zinc in oral tablet form for 50 days was Se 200 microg/day (50 microg/tablet) and Zn 21 mg/day (7 mg/tablet). RESULTS: Both in the basal condition and at 60 days all patients were malnourished. Selenium and zinc concentrations were significantly lower (P < 0.01) whereas copper concentration was significantly higher (P < 0.01) in cancer patients than in control subjects. However, 21/30 (70%) of those treated with Se and Zn did not showed a further worsening of nutritional status and experienced a significant decrease of asthenia with an increase of appetite. On the other hand, 24/30 (80%) untreated patients had a significant decline of all parameters studied after 60 days (prealbumin, cholesterol, transferrin, P < 0.05 vs 0 time; total proteins, albumin/globulin ratio, P < 0.01 vs 0 time; fat-free mass, fat mass, Na+/K+ ratio, body mass index P < 0.05 vs 0 time; fat free mass/fat mass, total body water, extra cellular/intra cellular water, basal metabolic rate: P < 0.01 vs 0 time). CONCLUSIONS: Data indicate that Se and Zn supplementation may improve the clinical course of general conditions in patients with gut cancer. These effects of Se and Zn require confirmation in an independent trial of appropriate design before new public health recommendations regarding Se and Zn supplementation can be made.

Analysis of metal-regulated metallothionein and heat shock gene expression in HeLa-derived cadmium-resistant cells.

The expression of metallothionein (MT) and heat shock protein gene families was investigated in normal and in HeLa-derived cadmium-resistant cells, named H454. In the absence of amplification of MT genes H454 cells accumulated elevated concentrations of cadmium ions and synthesized higher levels of MT proteins than unselected HeLa cells. Northern blot analyses revealed higher levels of MT mRNAs in the resistant cells than in wild-type cells after Cd2+ and Zn2+ exposure. Evaluation of the cytotoxic potential of the different metals confirmed the high resistance to cadmium of the H454 cells. Two proteins of the heat shock family, hsp70 and GRP78, were synthesized in Cd(2+)-exposed H454 cells at levels comparable to the ones present in Cd(2+)-treated normal cells. Northern blot analyses of the mRNA levels corresponding to these proteins revealed elevated expression of both hsp70 and GRP78 mRNAs in H454 cells upon exposure to cadmium ions and no response to zinc induction. These data suggest the existence in the H454 cells of a cadmium-specific pathway of regulation of MT and heat shock genes.

Transglutaminase changes in intestinal mucosa after experimental small bowel resection in the rat.

The low serum transglutaminase found in various intestinal disorders (celiac disease and IBD) suggested to us to study the serum and mucosal transglutaminase behaviour in an experimental model of small intestine resection in rats to reduce cellular mass and induce enterocyte hyperproliferation in the proximal part left in continuity. Transglutaminase activity in the intestinal mucosa was significantly higher in resected rats than in control and sham operated animals from days 4 (121 +/- 10 v basal 94 +/- 3 mU/g protein, p < 0.01) to 10 (165 +/- 37 mU/g protein, p < 0.05) after surgery; no significant difference was observed at days 12 and 15 (110 +/- 15 and 105 +/- 23 respectively). Both serum alkaline phosphatase activity (partly produced in enterocytes) and serum transglutaminase were significantly lower in resected rats at each time-point beginning at day 6 (208 +/- 34 v 557 +/- 125 UI and 1.55 +/- 0.11 v 3.78 +/- 0.70 mU/ml, p < 0.001 respectively). These data suggest an involvement of transglutaminase in enterocyte proliferation and confirm the association between reduced intestinal mass and low levels of the enzyme in serum.