Role of extracellular matrix in regulation of staurosporine-induced apoptosis in breast cancer cells.

Autocrine and paracrine mechanisms modulate the synthesis and secretion of extracellular matrix (ECM); moreover, each component of the ECM is capable of modulating the synthesis and release of other ECM molecules. Therefore, the synthesis of ECM glycoprotein fibronectin and laminin was studied in the human breast cancer cell lines MCF7 and MDA MB 23, plated on different ECM. Our results showed that the cells plated on a fibronectin substrate increased laminin synthesis: this event correlated with an increase in alpha2 and alpha3 integrin subunits. Staurosporine-induced apoptosis was then analyzed in the cell lines plated on different ECM. Staurosporine treatment determined the apoptosis of 35 and 33% respectively of MDA MB 231 and MCF7; these values increased to 60 and 64% in cells plated on laminin, to 48 and 63% in cells plated on fibronectin and to 64 and 69% in cells plated on matrigel. Moreover, staurosporine treatment decreased bcl-2 expression in the cells plated on fibronectin and laminin. Yet, staurosporine treatment determined PARP cleavage and PARP partial disappearance when the cells were plated on matrigel. Finally, a partial loss of function mutant Ras protein that activated only Raf pathway, was expressed in MCF7, in order to identify whether the increase of apoptosis induced by extracellular matrix involved the Raf/MAP kinase pathway. The increase of apoptosis of the cells plated on matrigel suggested that the activation of the Raf pathway is probably involved in the decrease of survival on matrigel. These data demonstrate that the modification of ECM modulates the apoptotic process of breast cancer cells and suggest that it is worthwhile to dissect the role of ECM in the control of apoptotic process.

Modulation of integrin expression on mesotheliomas: the role of different histotypes in invasiveness.

The in vitro and in vivo integrin expression in human pleural malignant mesothelioma (MM) of three different histotypes was studied. Cell lines from MM of epithelioid (E1), fibrous (F1), byphasic histotype (B1) and normal mesothelial cells (NM) were analysed for the surface expression of alpha2, alpha3, alpha4, alpha5, alpha6, alphav, beta1, beta3, beta4 subunits and alphavbeta5 integrins. We found that alpha6, beta4 subunits and alphavbeta5, weakly detectable on NM cells, were expressed on MM cells. The beta3 subunit, well expressed on NM cells, was absent on MM cells. Differential expression among histotypes was observed, the MM-E1 was the least and the MM-B1 the most positive. Specimens for each MM histotype, were analysed by immunohistochemistry. The alpha6 and alphav subunits were more evident on the epithelioid histotype. Intense staining for beta3 and beta4 subunits, was found in all MM, particularly in invading cells, while the alpha5, and alphavbeta5 integrins were variously expressed. The different histotypes can affect the in vitro integrin expression and may indicate a preferential involvement of some subunits in vivo during MM tumor progression.

An alpha 5 beta 1-like integrin receptor mediates the binding of less pathogenic Candida species to fibronectin.

The present study was undertaken to investigate whether less pathogenic Candida species (C. tropicalis, C. stellatoidea, C. krusei and C. glabrata) express a fibronectin receptor (FNr) antigenically related to alpha 5 beta 1 integrin, which mediates their binding to fibronectin (FN). By flow cytometric analysis, a monoclonal antibody (MAb) directed against human alpha 5 integrin subunit (clone SAM-1) and two different antisera to FNr positively stained C. tropicalis, C. stellatoidea and C. glabrata, with the greatest expression observed for C. tropicalis. No or only marginal immunoreactivity was found on C. krusei. C. tropicalis, C. stellatoidea, C. glabrata, but not C. krusei yeasts specifically adhered to FN; higher levels of adhesion were found for C. tropicalis and C. stellatoidea with respect to C. glabrata. Less pathogenic Candida spp. bound to the Arg-Gly-Asp (RGD) containing 120-kDa fragment of FN and adhesion to intact FN was markedly inhibited by Gly-Arg-Gly-Asp-Ser-Pro (GRGDSP), but not by Gly-Arg-Gly-Glu-Ser-Pro (GRGESP) peptides. In addition, anti-alpha 5 SAM-1 MAb and both anti-FNr antisera strongly blocked binding of less pathogenic Candida spp. to FN. Overall, these results indicate that less pathogenic Candida spp., including C. tropicalis, C. stellatoidea and C. glabrata, express a receptor antigenically related to alpha 5 beta 1 integrin which mediates their adhesion to FN.

In vivo capsaicin treatment inhibits rat NK cell cytotoxic functions.

The direct and indirect interactions between the nervous system and its transmitters with NK cell cytotoxic functions has been evaluated in the rat by using the neurotoxin capsaicin (8-methyl-N-vanillyl-6-nonenamide). When administered to neonatal rats, capsaicin (50 mg/Kg in 10% ethanol and 10% tween 80 at 2 days of age) interferes with the synthesis and intraneuronal transport of peptides by causing irreversible degeneration of c fiber afferent nerves. Capsaicin treatment resulted in a marked inhibition of NK and ADCC activities both in the spleen and peripheral blood. Inhibition was already evident on day 15 after treatment and persisted until day 90 in the spleen; at this time NK cytotoxicity in the peripheral blood returned to control levels. The inhibitory effect of capsaicin treatment on peripheral blood NK and ADCC activities was associated with changes in NK cell number evaluated as percentage of cells with an LGL morphology and expressing the NK-RP1 cell surface receptor. LGL numbers did not always correlate with the percentage of NK-RP1+ cells suggesting that capsaicin may interfere with maturation of lytic effector cells. Overall these results indicate a direct influence of the nervous system on natural immune cytotoxic functions.