Prognóstico de cães com tumor de mastócitos tratados cirurgicamente: análise de 18 casos

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Genistein blocks breast cancer cells in the G(2)M phase of the cell cycle.

Genistein, a natural isoflavone phytoestrogen present in soybeans, caused a dose-dependent growth inhibition of the two hormone-sensitive cell lines T47D and ZR75.1 and of the two hormone-independent cell lines MDAMB-231 and BT20. Flow cytometric analysis of cells treated for 4 days with 15 and 30 microM genistein showed a dose-dependent accumulation in the G(2)M phase of the cell cycle. At the highest tested concentration, there was a sevenfold increase in the percentage of cells in G(2)M (63%) with respect to the control (9%) in the case of T47D cells and a 2.4-fold increase in the case of BT20. An intermediate fourfold accumulation was observed in the case of MDAMB-231 and ZR75.1. The G(2)M arrest was coupled with a parallel depletion of the G(0)/G(1) phase. To understand the mechanism of action underlying the block in G(2)M induced by genistein, we investigated the expression and the activity of cyclins and of cyclin-dependent kinases specifically involved in the G(2)-->M transition. As expected, p34(cdc-2) expression, monitored by Western blotting, was unaffected by genistein treatment in all cell lines. With exception of the T47D cell line, we revealed an increase in the tyrosine phosphorylated form of p34, suggesting an inactivation of the p34(cdc-2) catalytic activity consequent to treatment of cells with genistein. In fact, immunoprecipitates from genistein-treated MDAMB-231 and BT20 cells displayed a fourfold decrease in kinase activity evaluated using the histone H1 as substrate. Conversely, no variation in kinase activity was observed between treated and untreated ZR75.1 cells despite the increase in p34 phosphorylation. In cells treated with 30 microM genistein, cyclin B(1) (p62) increased 2.8-,8-and 103-fold, respectively, in BT20, MDAMB-231, and ZR75.1 cells, suggesting an accumulation of the p62, which is instead rapidly degraded in cycling cells. No effects were observed on cyclin expression in T47D cells. We therefore conclude that genistein causes a G(2)M arrest in breast cancer cell lines, but that such growth arrest is not necessarily coupled with deregulation of the p34(cdc-2)/cyclin B(1) complex only in all of the studied cell lines.

Importance of acute Mycoplasma pneumoniae and Chlamydia pneumoniae infections in children with wheezing.

In order to evaluate the role of Mycoplasma pneumoniae and Chlamydia pneumoniae in reactive airway disease, 71 children aged 2-14 yrs with an acute episode of wheezing and 80 age-matched healthy children were studied. Sera for the determination of specific antibody levels and nasopharyngeal aspirates for the detection of M. pneumoniae and C. pneumoniae deoxyribonucleic acid were obtained on admission and after 4-6 weeks. All children with wheezing received a standard therapy with inhaled corticosteroids and bronchodilators for 5-7 days; when antibiotic was added on the basis of the judgement of the paediatrician in charge, clarithromycin 15 mg.kg body weight(-1).day(-1) for 10 days was used. Acute M. pneumoniae and C. pneumoniae infections were detected significantly more often in children with wheezing than in controls. In patients infected with one of the two pathogens, a history of recurrent wheezing was significantly more frequent than in those without either infection. During a 3-month follow-up period, among nonantibiotic-treated children, those with acute M. pneumoniae and/or C. pneumoniae infection showed a significantly higher recurrence of wheezing than those without acute M. pneumoniae and/or C. pneumoniae infection (p=0.03). These results highlight the apparently significant relationship of Mycoplasma pneumoniae and Chlamydia pneumoniae with wheezing in children, particularly in subjects with a history of recurrent episodes, and the possible improvement in the course of reactive airway disease within paediatric patients with acute Mycoplasma pneumoniae and/or Chlamydia pneumoniae infection.

The two phyto-oestrogens genistein and quercetin exert different effects on oestrogen receptor function.

We compared the oestrogenic and anti-oestrogenic properties of the two well-known phyto-oestrogens, genistein and quercetin, on the oestrogen-sensitive breast cancer cell line MCF-7. Genistein exerted a biphasic effect on growth of MCF-7 cells, stimulating at low and inhibiting at high concentrations, whereas quercetin was only growth inhibitory. At doses which did not inhibit cell growth, respectively 5 and 1 microM, genistein and quercetin counteracted oestrogen- and transforming growth factor-alpha-promoted cell growth stimulation. Furthermore, genistein promoted transcription of the oestrogen-regulated genes pS2 and cathepsin-D, whereas quercetin interfered with the oestrogen-induced expression of the proteins. In in vitro binding experiments, genistein competed with oestradiol for binding to the oestrogen receptor (ER), but quercetin did not. Quercetin and genistein down-regulated cytoplasmic ER levels and promoted a tighter nuclear association of the ER, but only genistein was able to up-regulate progesterone receptor protein levels. In gel mobility assays, ER preincubation with oestradiol or with the two phyto-oestrogens led to the appearance of the same retarded band, excluding differences between the various complexes in binding to the consensus sequence. The data allowed us to conclude that quercetin acts like a pure anti-oestrogen, whereas genistein displays mixed agonist/antagonist properties, and to formulate a hypothesis on the possible mechanism of action of such phyto-oestrogens.

Genistein in the control of breast cancer cell growth: insights into the mechanism of action in vitro.

Genistein significantly inhibited cell growth (IC50 around 10 microM) of MCF-7, MDAMB-231 and HBL-100 cell lines, but not of skin-derived fibroblasts and counteracted the growth-stimulatory effects exerted by estradiol and growth factors. It abolished the paracrine stimulation observed in MCF-7 cells in co-culture with MDAMB-231 or fibroblasts. Genistein-treated cells accumulated in the S and G2/M phases of the cell cycle and underwent apoptosis. Genistein decreased tyrosine phosphorylation induced upon treatment with transforming growth factor-alpha. Finally, genistein bound the estrogen receptor (ER) (relative affinity constant Kd = 4 nM), induced pS2 and cathepsin-D transcription and increased nuclear ER levels.

Synthetic analogs of vitamin D3 have inhibitory effects on breast cancer cell lines.

BACKGROUND: 1,25-dihydroxycholecalciferol has been previously reported to negatively regulate human breast cancer cell growth. MATERIAL AND METHODS: The antiproliferative effect of 1,25-dihydroxycholecalciferol (Ro 21-5535) and of the two non hypercalcemic analogs (1a,25-dihydroxy-16-ene-23-yne-26,27-hexafluorocholecalciferol+ ++, Ro 24-5531 and 1a,25-dihydroxy-16-ene-23-yne-26,27-hexafluoro-19-nor-cholec alciferol, Ro 25-6760) was studied in MCF-7 and MDAMB-468 human breast cancer cell lines. Cell cycle distribution and apoptosis were evaluated by flow cytometry. Steroid receptor modulation was investigated by radioligand assay. RESULTS: The most effective drug was the Ro 25-6760 which at concentrations ranging between 1-100 nM caused a dose dependent growth inhibition apparently due to accumulation in G0/G1. Vitamin D3 analogs (10 nM) significantly counteracted the growth stimulation induced by TGF-a and IGF-I as well as the paracrine stimulation observed in co-cultures. They antagonized estradiol-promoted growth stimulation and progestrone receptor induction in MCF-7 cells. CONCLUSION: Vitamin D3 analogs represent a class of clinically attractive drugs for treatment of breast cancer due to their ability to counteract estradiol and growth factor-induced growth stimulation.

int-2 oncogene amplification and prognosis in node-negative breast carcinoma.

The role of int-2 oncogene amplification on the prognosis of breast cancer patients was investigated in 128 patients with node-negative primary breast cancers given first-line local-regional treatments until relapse and with a median follow-up of 65 months. Tumours had been previously characterised for oestrogen (ER) and progesterone receptor (PgR) status and proliferative activity (3H-thymidine labelling index). Amplification of the int-2 oncogene occurred in 18% of cases and was significantly related to the presence of hormone receptors and to menopausal status or age, but not to proliferative status. Patients with tumours exhibiting int-2 amplification had a lower probability of disease-free survival than patients with non-amplified tumours and frequently developed local-regional recurrence. Disease-free survival analysis, adjusted for the prognostic contribution provided by tumour size, steroid receptors and proliferative rate, indicated that the association between int-2 amplification and risk of relapse was maintained and remained constant even in the presence of the other co-variates. Interestingly, int-2 amplification was a further prognostic discriminant within subsets of patients with a putatively good (i.e., tumour size <20 mm, ER+ and PgR+) or poor prognosis (i.e., high labelling index). Our exploratory study suggests that within node-negative patients, int-2 amplification could be a valuable and independent prognosticator, useful to identify patients at high risk of local-regional recurrence.

Modulation of cathepsin-D and pS2 protein levels in human breast cancer cell lines.

Cathepsin-D and pS2 are two estrogen-regulated proteins in human breast cancer cell lines. They have been considered possible prognostic factors in breast cancer, but results have been contradictory. To better understand the regulation of these proteins, we investigated the role of estradiol (E2), serum, and growth factors in hormone-dependent (MCF-7, ZR75.1) and hormone-independent (MDAMB-231, BT20) breast cancer cell lines. E2 treatment in serum-free conditions increased intracellular and secreted levels of pS2 in ZR75.1 and in MCF-7, secreted levels only of cathepsin-D in MCF-7, and both levels of cathepsin-D in ZR75.1. Insulin-like growth factor I (IGF-I) and progesterone receptors were also stimulated by E2, whereas the estrogen receptor was down-regulated. Following treatment with epidermal growth factor (EGF), secreted pS2 levels doubled only in MCF-7 cells. IGF-I did not modify cathepsin-D or pS2 levels in either cell line, but caused an increase in its own receptor. Cathepsin-D and pS2 doubled in MCF-7 cells grown in medium supplemented with denaturated serum, but estrogen regulation of these proteins was still maintained. Cathepsin-D was expressed in MDAMB-231 and BT20, but its levels were modified by neither E2 nor growth factor treatment. Conversely, neither cell line expressed detectable levels of pS2 before or after treatment. In conclusion, our results show that in different types of breast cancer cells, some estrogen-regulated proteins (e.g. pS2) are also regulated by growth factors-such as EGF and other unknown serum factors. This may account for the contradictory results obtained regarding the prognostic relevance of cathepsin-D and pS2.

Effect of progestin treatment on estradiol-and growth factor-stimulated breast cancer cell lines.

BACKGROUND: Reports about the effects of progestins on cell proliferation are contradictory. We investigated the effect of progesterone, medroxyprogesterone acetate, megestrol acetate, ORG 2058 and the antiprogestin RU 486 on two hormone-dependent cell lines, T47D and MCF-7 (characterized by a different content of PgR). The aim of the study was to understand the eventual ability of progestins to interfere with cell proliferation stimulated by estradiol and various growth factors (TGF-a, IGF-I, IGF-II). MATERIAL AND METHODS: MCF-7 and T47D cells were maintained in DMEM/F12 medium supplemented with 2% FCS while experiments were carried out in the same culture medium using DCC-stripped FCS. RESULTS: In the absence of estradiol, all tested progestins generally tended to stimulate cell growth in the T47D cell line, but in the MCF-7 cell line only the highest concentrations (10(-6) M and 10(-7) M) were found to be stimulatory. In contrast, in the presence of 10(-8) M estradiol, progestins tended to inhibit cell growth stimulation in MCF-7 and T47D cell lines. The antiprogestin RU 486 exerted a stimulatory effect similar to that promoted by estradiol itself in MCF-7 cells. Instead, in T47D cells, RU 486 did not modify cell growth in the absence of estradiol, but tended to counteract the estradiol-promoted cell proliferation. In MCF-7 cells, medroxyprogesterone acetate and megestrol acetate were also able to effectively counteract the cell growth induced by TGF-alpha. However, none of these progestins was able to abolish cell proliferation promoted by IGF-I or IGF-II. CONCLUSION: We therefore concluded that failure to respond to progestin treatment may be due to the very heterogeneous nature of human breast tumors and to the inability of these molecules to interfere with the IGF-R pathway.

Paracrine interaction in co-culture of hormone-dependent and independent breast cancer cells.

A permeable solid support (Transwell Coll.) was used to develop serum-free co-cultures allowing paracrine interactions between hormone-dependent (MCF-7, ZR75.1) and hormone-independent (MDAMB-231, BT20) breast cancer cell lines. Both hormone-independent cell lines were able to stimulate the growth of the hormone-dependent lines, whereas the opposite was observed only in the case of BT20 co-culture with ZR75.1 cells. The cell growth stimulation observed in co-cultures could be abolished by the addition to the culture medium of an excess of transforming growth factor alpha (TGF alpha) or insulin-like growth factor I (IGF-I). Similarly, treatment with a neutralizing anti TGF alpha antibody impaired the growth stimulation exerted by hormone-independent cells on hormone-dependent cells. These results confirm the important role of paracrine interactions in control of the growth of human heterogeneous breast tumors and suggest that the main growth factors involved in such interactions are TGF alpha and probably some growth factors from the insulin-like growth factor family rather than IGF-I itself.

[Prosthetic rehabilitation in patients with malignant tumors of the soft palate. A clinical case report]. / Riabilitazione protesica nei pazienti con tumori maligni del palato molle. Presentazione di un caso clinico.

Soft palate neoplasms are relatively rare and their care requires not only an adequate surgical removal but also a post-treatment rehabilitation. In this article the authors after having considered the importance of the soft palate function on the individual's ability to shallow and speak, describe one case of squamous cell carcinoma of the soft palate and treated with surgery followed by prosthetic rehabilitation. Especially it is pointed out the utility of a palatal obturator in the care of deleterious esthetic and psychological aspect of the residual anatomical defect, considering the possibility of the re-establishment the integrity of the tissues and structures in the oral cavity by hand-made prosthesis more and more exact and functional.

[Neuropsychiatric approach to a child with spinal muscular atrophy. A study of relational problems]. / Approccio neuropsichiatrico al bambino con amiotrofia spinale. Studio delle problematiche relazionali.

Various years' experience of an extensive series of children suffering from progressive chronic spinal muscular atrophy have made it possible to elaborate a neuropsychiatric approach model conducted on the basis of a viewpoint capable of integrating the clinico-medical aspect and aptitude to empathic listening aimed at establishing a therapeutic alliance with the patient. With advancing age and the course of the disease, in most cases very different difficulties and problems are encountered. These change continuously in relation to the stage reached in emotional development. Entry to school and socialisation mark a particular moment in the confrontation with peers and present the ill child with his diversity. With the emergence of adolescent problems, there is also a dramatic new impact between the body image of a wounded, deteriorated self and the evolutionary effort aimed at identifying with the peer group, the autonomisation of parental figures, the integration of a sexed image of the self. Room can be found in this framework for the child neuropsychiatrist who is able to offer himself as a point of reference and set himself up as a model for elaborating disease anxieties. From personal experience it also emerges that in children suffering from a chronic, often fatal disease there is often the need for a physician to coordinate and integrate needs of both a clinical and sociorelational nature for the purpose of allowing the family to get closer to the needs of their child and prevent him from falling prey to destructive type anxieties.