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The soluble TNF-like weak inducer of apoptosis (TWEAK, TNFSF12) binds to the fibroblast growth factor-inducible 14 receptor (FN14, TNFRSF12A) on the cell membrane and induces multiple biological responses, such as proliferation, migration, differentiation, angiogenesis and apoptosis. Previous reports show that TWEAK, which does not contain a death domain in its cytoplasmic tail, induces the apoptosis of tumor cell lines through the induction of TNFα secretion. TWEAK induces apoptosis in human keratinocytes. Our experiments clearly demonstrate that TWEAK does not induce the secretion of TNFα or TRAIL proteins. The use of specific inhibitors and the absence of procaspase-3 cleavage suggest that the apoptosis of keratinocytes follows a caspase- and cathepsin B-independent pathway. Further investigation showed that TWEAK induces a decrease in the mitochondrial membrane potential of keratinocytes. Confocal microscopy showed that TWEAK induces the cleavage and the translocation of apoptosis inducing factor (AIF) from the mitochondria to the nucleus, thus initiating caspase-independent apoptosis. Moreover, TWEAK induces FOXO3 and GADD45 expression, cdc2 phosphorylation and cdc2 and cyclinB1 degradation, resulting in the arrest of cell growth at the G2/M phase. Finally, we report that TWEAK and FN14 are normally expressed in the basal layer of the physiological epidermis and are greatly enhanced in benign (psoriasis) and malignant (squamous cell carcinoma) skin pathologies that are characterized by an inflammatory component. TWEAK might play an essential role in skin homeostasis and pathology.

In recent years, our knowledge on estrogen receptors (ER) has been modified profoundly with the identification and the deciphering of the role of its protein effectors, as well as with the deeper insight of its molecular structure/function dynamics, characteristics associated with its nucleo-cytoplasmic-membrane shuttling properties. Also, significant progress has been made concerning its turn-over and associated final proteasomal degradation processes. These advances could lead in the near future to the design and the synthesis of novel receptor-interacting drugs. Recently, a number of receptor-related peptides acting as specific ER ligands have been identified and extensively studied with respect to their estrogenic/antiestrogenic activities. Among them, ERα17p, a synthetic analog of the P(295)-T(311) sequence of ERα, has been shown to exert pseudo-estrogenic effects by interacting in the close vicinity of its hinge region (BF3 domain). Remarkably, this sequence appears as the epicenter of a number of post-transcriptional modifications as well as of the recruitment of co-regulators, suggesting that it would play a key role in ERα functions. Here, we provide evidence that ERα17p induces apoptosis in ERα-positive (MCF-7, T47D) and -negative (MDA-MB-231, SK-BR-3) breast cancer cells by an ERα-independent membrane mechanism, triggering major pro-apoptotic signaling cascades. Finally, ERα17p induces the regression of breast ERα-negative cancer tumor xenografts, without apparent toxicity, suggesting that it could represent a new attractive tool for the development of future promising therapeutic approaches, and providing a novel insight to ER regulation of cell fate., (Copyright © 2010 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.)

Toxic epidermal necrolysis (TEN) is characterized by an acute detachment and destruction of keratinocytes, affecting large areas of the skin. It is often related to adverse drug reactions. Previous studies have shown that effector CD8+ T cells, which accumulate in the blister fluid, are functionally cytotoxic and act through a classical perforin/granzyme B pathway. It has recently been shown that these cytotoxic T cells also secrete granulysin peptide, which is lethal to keratinocytes. These cytotoxic T cells exert their killer activity against autologous keratinocytes in the presence of the drug. However, they are unlikely to be the only effectors of TEN. We therefore searched for soluble death factors in the blister fluids that might kill keratinocytes. We found that the amounts of interferon-γ, TRAIL and TNF-α proteins were significantly greater in TEN blister fluids than in all controls (normal sera, TEN sera, burns and Eosinophilic pustular folliculitis blister fluids) and TNF-like weak inducer of apoptosis (TWEAK) amounts are also greater in all controls except burns. We showed that these proteins acted in synergy to induce the death of keratinocytes in vitro. We also found that TRAIL and TWEAK were secreted by CD1a+ and CD14+ cells present in the blister fluids. Thus, in addition to MHC class I-restricted cytotoxic T lymphocytes (CTLs), which lyse keratinocytes, ligands secreted by non-lymphoid cells capable of inducing keratinocyte death in an MHC class I-independent manner, also seem to be present in the blister fluids of patients with TEN., (© 2011 John Wiley & Sons A/S.)

Merkel cell carcinoma is a tumor with aggressive biological behavior and limited response to chemotherapy. The present study investigated the effect of interferon (IFN)-alpha on growth and apoptosis of Merkel carcinoma cells in vitro. Proliferation of MCC-1 cell line was reduced dose-dependently by IFN-alpha and diminished when higher IFN-alpha concentrations were used. Additionally, IFN-alpha potently decreased DNA-synthesis and Ki67/MIB-1 proliferation index of MCC-1 cultures. Furthermore, IFN-alpha induced dose-dependently apoptosis of MCC-1 cells as shown by caspase-3 activation, and detection of apoptotic DNA strand breaks and fragmented nuclei. These findings suggest that IFN-alpha may have antitumor activity against Merkel cell carcinoma.

Classical steroid mode of action involves binding to intracellular receptors, the later acting as ligand-activated nuclear transcription factors. Recently, membrane sites for different steroids have been also identified, mediating rapid, non-genomic, steroid actions. Membrane sites for estrogen and androgen have been found in a number of different cell types, bearing or not classical intracellular receptors. In the present study, with the use of radioligand binding, flow cytometry and confocal laser microscopy, we report that T47D human breast cancer cells express specific and saturable membrane receptors for both estrogen (K(D) 4.06 +/- 3.31 nM) and androgen (K(D) 7.64 +/- 3.15 nM). Upon activation with BSA-conjugated, non-permeable ligands (E(2)-BSA and testosterone-BSA), membrane estrogen receptors protect cells from serum-deprivation-induced apoptosis, while androgen receptors induce apoptosis in serum-supplemented T47D cells. In addition, co-incubation of cells with a fixed concentration of one steroid and varying concentrations of the other reversed the abovementioned effect (apoptosis for androgen, and anti-apoptosis for E(2)), suggesting that the fate of the cell depends on the relative concentration of either steroid in the culture medium. We also report the identification of membrane receptors for E(2) and androgen in biopsy slides from breast cancer patients. Both sites are expressed, with the staining for membrane E(2) being strongly present in ER-negative, less differentiated, more aggressive tumors. These findings suggest that aromatase inhibitors may exert their beneficial effects on breast cancer by also propagating the metabolism of local steroids towards androgen, inducing thus cell apoptosis through membrane androgen receptor activation.

Nongenomic androgen actions imply mechanisms different from the classical intracellular androgen receptor (iAR) activation. We have recently reported the identification of a membrane androgen receptor (mAR) on LNCaP human prostate cancer cells, mediating testosterone signal transduction within minutes. In the present study we provide evidence that activation of mAR by nonpermeable, BSA-coupled testosterone results in 1) inhibition of LNCaP cell growth (with a 50% inhibitory concentration of 5.08 nM, similar to the affinity of testosterone for membrane sites); 2) induction in LNCaP cells of both apoptosis and the proapoptotic Fas protein; and 3) a significant decrease in migration, adhesion, and invasion of iAR-negative DU145 human prostate cancer cells. These actions persisted in the presence of antiandrogen flutamide or after decreasing the content of iAR in LNCaP cells by iAR antisense oligonucleotides. Testosterone-BSA was also effective in inducing apoptosis of DU145 human prostate cancer cells, negative for iAR, but expressing mAR sites. In LNCaP cell-inoculated nude mice, treatment with testosterone-BSA (4.8 mg/kg body weight) for 1 month resulted in a 60% reduction of tumor size compared with that in control animals receiving only BSA, an effect that was not affected by the antiandrogen flutamide. Our findings suggest that activators of mAR may represent a new class of antitumoral agents of prostate cancer.

The soluble TNF-like weak inducer of apoptosis (TWEAK, TNFSF12) binds to the fibroblast growth factor-inducible 14 receptor (FN14, TNFRSF12A) on the cell membrane and induces multiple biological responses, such as proliferation, migration, differentiation, angiogenesis and apoptosis. Previous reports show that TWEAK, which does not contain a death domain in its cytoplasmic tail, induces the apoptosis of tumor cell lines through the induction of TNFα secretion. TWEAK induces apoptosis in human keratinocytes. Our experiments clearly demonstrate that TWEAK does not induce the secretion of TNFα or TRAIL proteins. The use of specific inhibitors and the absence of procaspase-3 cleavage suggest that the apoptosis of keratinocytes follows a caspase- and cathepsin B-independent pathway. Further investigation showed that TWEAK induces a decrease in the mitochondrial membrane potential of keratinocytes. Confocal microscopy showed that TWEAK induces the cleavage and the translocation of apoptosis inducing factor (AIF) from the mitochondria to the nucleus, thus initiating caspase-independent apoptosis. Moreover, TWEAK induces FOXO3 and GADD45 expression, cdc2 phosphorylation and cdc2 and cyclinB1 degradation, resulting in the arrest of cell growth at the G2/M phase. Finally, we report that TWEAK and FN14 are normally expressed in the basal layer of the physiological epidermis and are greatly enhanced in benign (psoriasis) and malignant (squamous cell carcinoma) skin pathologies that are characterized by an inflammatory component. TWEAK might play an essential role in skin homeostasis and pathology. [ABSTRACT FROM AUTHOR]

Abstract: In recent years, our knowledge on estrogen receptors (ER) has been modified profoundly with the identification and the deciphering of the role of its protein effectors, as well as with the deeper insight of its molecular structure/function dynamics, characteristics associated with its nucleo-cytoplasmic-membrane shuttling properties. Also, significant progress has been made concerning its turn-over and associated final proteasomal degradation processes. These advances could lead in the near future to the design and the synthesis of novel receptor-interacting drugs. Recently, a number of receptor-related peptides acting as specific ER ligands have been identified and extensively studied with respect to their estrogenic/antiestrogenic activities. Among them, ERα17p, a synthetic analog of the P295-T311 sequence of ERα, has been shown to exert pseudo-estrogenic effects by interacting in the close vicinity of its hinge region (BF3 domain). Remarkably, this sequence appears as the epicenter of a number of post-transcriptional modifications as well as of the recruitment of co-regulators, suggesting that it would play a key role in ERα functions. Here, we provide evidence that ERα17p induces apoptosis in ERα-positive (MCF-7, T47D) and -negative (MDA-MB-231, SK-BR-3) breast cancer cells by an ERα-independent membrane mechanism, triggering major pro-apoptotic signaling cascades. Finally, ERα17p induces the regression of breast ERα-negative cancer tumor xenografts, without apparent toxicity, suggesting that it could represent a new attractive tool for the development of future promising therapeutic approaches, and providing a novel insight to ER regulation of cell fate. [ABSTRACT FROM AUTHOR]

Background/Aims: KIT receptor has been implicated in the pathogenesis of cancer, either by mutation or autocrine activation. Merkel cell carcinoma (MCC) is a rare KIT-positive cutaneous tumor. We investigated the co-expression of KIT and its ligand stem cell factor (SCF) in MCC. Methods: Sixteen specimens from 13 MCC patients of various tumor stages were examined by immunohistochemistry for SCF, KIT, Ki67/MIB-1 and cleaved caspase 3 expression, and for apoptosis by TUNEL. Results: KIT was expressed in 13 of 16 tumors, and SCF in 15 of 16 specimens. Co-expression of KIT and SCF was detected in 12 of 16 tumors. KIT and SCF immunoreactivity scores were independent of tumor stage. Ki67/MIB-1 proliferation rates were high, whereas apoptosis rates were low, and did not depend on KIT or SCF expression. Conclusion: Co-expression of KIT and SCF in a high percentage of MCC tumors hints to an autocrine mechanism. KIT and SCF expression in primary tumors and in metastases suggests an early event in Merkel cell transformation. Copyright © 2008 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Discusses the findings of a study which investigated the antiapoptotic effect of estrogen. Mechanisms underlying the neuroprotective role of estrogen; Detection of intracellular steroid receptors in pheochromocytoma cell line PC12; Analysis of the presence of membrane binding sites for estrogen.