High Doses of Ascorbate Kill Y79 Retinoblastoma Cells In vitro

Objectives: To tests the sensitivity of Y79 retinoblastoma cell lines to high doses of ascorbate, in vitro, and compare its effects with those of some chemotherapeutic agents routinely employed in the treatment of retinoblastoma. Methods: Y79 retinoblastoma cells have been exposed to increasing doses of either sodium ascorbate (SA) or Melphalan (MEL), to define a dose-response curve around the peak plasma concentrations reached by both chemicals when administered according to the existing therapeutic procedures and protocols. The assessment of cell number and viability was performed, before and after exposure, with both the manual (Trypan Blue Exclusion Test) and automated (flow cytometry) methods. Fluorescence microscopy and direct observation of cells in culture, with inverted microscope, were also performed. Results: Y79 cells are highly sensitive to the cytotoxic effect of SA, with cell viability reduced of over 90% in some experiments. As reported in the literature, this effect is directly cytotoxic and most probably mediated by acute oxidative stress on different cellular components. The same does not apply to Melphalan which, at the doses commonly used for therapeutic purposes, did not show any significant effect on cell viability, in vitro. Conclusion: To our knowledge, this is the first report showing that high doses of SA can actively kill retinoblastoma cells in vitro. While it is not surprising for SA, to show direct cytotoxic effect on tumor cells, the data reported herein represent the first evidence in favor of the possible clinical use of high doses of intravenous SA, to treat children affected by retinoblastoma. Given the many advantages of SA over the chemotherapeutic agents commonly employed to treat cancer (including its almost total absence of toxic or side effects, and its exclusive specificity for cancer cells), it is reasonable to assume, from the data reported herein, that the high doses of intravenous ascorbate, have the potential to represent a real revolution in the treatment of retinoblastoma

Alternative Pathways of Cancer Cell Death by Rottlerin: Apoptosis versus Autophagy

Since the ability of cancer cells to evade apoptosis often limits the efficacy of radiotherapy and chemotherapy, autophagy is emerging as an alternative target to promote cell death. Therefore, we wondered whether Rottlerin, a natural polyphenolic compound with antiproliferative effects in several cell types, can induce cell death in MCF-7 breast cancer cells. The MCF-7 cell line is a good model of chemo/radio resistance, being both apoptosis and autophagy resistant, due to deletion of caspase 3 gene, high expression of the antiapoptotic protein Bcl-2, and low expression of the autophagic Beclin-1 protein. The contribution of autophagy and apoptosis to the cytotoxic effects of Rottlerin was examined by light, fluorescence, and electron microscopic examination and by western blotting analysis of apoptotic and autophagic markers. By comparing caspases-3-deficient (MCF-73def) and caspases-3-transfected MCF-7 cells (MCF-73trans), we found that Rottlerin induced a noncanonical, Bcl-2-, Beclin 1-, Akt-, and ERK-independent autophagic death in the former- and the caspases-mediated apoptosis in the latter, in not starved conditions and in the absence of any other treatment. These findings suggest that Rottlerin could be cytotoxic for different cancer cell types, both apoptosis competent and apoptosis resistant

Muscle pathology patterns in possibly adjuvant related autoimmune/inflammatory syndrome (ASIA)

Growing evidence shows a link for biologically inert molecules, such as vaccine adjuvants and silicone implants, with the occurrence of autoimmunity-related disorders, defined as autoimmune/inflammatory syndrome induced by adjuvant-ASIA (1). Clinical conditions encompass siliconosis, the Gulf war syndrome, the macrophagic myofasciitis syndrome (MMF), post-vaccination phenomena and the spectrum of related syndromes is expanding (2). Involvement of skeletal muscle in ASIA is acknowledged in MMF, defined by long-term persistence of vaccine alum adjuvants within macrophages at sites of previous immunization. A few reports describe vaccine and silicone implants related autoimmune inflammatory myopathies (3). We carried out an immunopathological analysis of skeletal muscle biopsy in a case of MMF and two cases of possible ASIA myositis, chronologically subsequent to breast silicone implant. MMF showed the typical fascial/ perimysial macrophagic invasion, with no endomysial mononuclear infiltrates and fibral neolocalization of MHC-I complex restricted to the adjacency of macrophage deposits. The first myositis case presented with a subacute onset twenty years after an uneventful additive breast silicone implant. Endomysial inflammation, microangiopathy and multifocal fibral localization of MHC-II complex were observed. In the second patient, the onset of proximal weakness, myalgiae and a tenfold increase of creatinkinase levels occurred seven years after an unsuccessful additive mastoplasty, with rupture of prostheses and re-implantation three years later. Muscle biopsy, besides inflammation changes, showed peculiar myofibrillar disruption, with MHC-I reactive sarcoplasmic inclusions expressing several structural muscle proteins. Molecular pathogenesis of ASIA is yet undefined: genetical susceptibility is currently investigated (1,2). Due to the role of vaccines in medicine and the wide use of silicon medical devices, identification of their cause/effect link with autoimmunity is of great interest

Differential expression of follistatin and FLRG in human breast proliferative disorders

<p>Abstract</p> <p>Background</p> <p>Activins are growth factors acting on cell growth and differentiation. Activins are expressed in high grade breast tumors and they display an antiproliferative effect inducing G0/G1 cell cycle arrest in breast cancer cell lines. Follistatin and follistatin- related gene (FLRG) bind and neutralize activins. In order to establish if these activin binding proteins are involved in breast tumor progression, the present study evaluated follistatin and FLRG pattern of mRNA and protein expression in normal human breast tissue and in different breast proliferative diseases.</p> <p>Methods</p> <p>Paraffin embedded specimens of normal breast (NB - n = 8); florid hyperplasia without atypia (FH - n = 17); fibroadenoma (FIB - n = 17); ductal carcinoma <it>in situ </it>(DCIS - n = 10) and infiltrating ductal carcinoma (IDC - n = 15) were processed for follistatin and FLRG immunohistochemistry and <it>in situ </it>hybridization. The area and intensity of chromogen epithelial and stromal staining were analyzed semi-quantitatively.</p> <p>Results</p> <p>Follistatin and FLRG were expressed both in normal tissue and in all the breast diseases investigated. Follistatin staining was detected in the epithelial cytoplasm and nucleus in normal, benign and malignant breast tissue, with a stronger staining intensity in the peri-alveolar stromal cells of FIB at both mRNA and protein levels. Conversely, FLRG area and intensity of mRNA and protein staining were higher both in the cytoplasm and in the nucleus of IDC epithelial cells when compared to NB, while no significant changes in the stromal intensity were observed in all the proliferative diseases analyzed.</p> <p>Conclusion</p> <p>The present findings suggest a role for follistatin in breast benign disease, particularly in FIB, where its expression was increased in stromal cells. The up regulation of FLRG in IDC suggests a role for this protein in the progression of breast malignancy. As activin displays an anti-proliferative effect in human mammary cells, the present findings indicate that an increased FST and FLRG expression in breast proliferative diseases might counteract the anti-proliferative effects of activin in human breast cancer.</p

The physiological interferon response. V. Antiviral activity present in rat lymph is neutralized by anti-mouse interferon-gamma antibodies

By neutralization tests using anti-rat and mouse interferon (IFN) antibodies, we tested whether the antiviral activity present in abdominal lymph but absent in plasma of healthy rats, could be ascribed to IFN. Antimouse IFN-gamma antibodies neutralized the inhibitor completely while anti-rat IFN-alpha/beta antibodies did not. We conclude that rat lymph contains traces of IFN-gamma and that the antiviral activity is not due to low-density lipoproteins, immunoglobulins, or to cell-produced viral inhibitors. This finding extends previous observations on rabbit lymph and further supports the existence of a physiological low-level interferon response

Athletic humans and horses: Comparative analysis of interleukin-6 (IL-6) and IL-6 receptor (IL-6R) expression in peripheral blood mononuclear cells in trained and untrained subjects at rest

Background Horses and humans share a natural proclivity for athletic performance. In this respect, horses can be considered a reference species in studies designed to optimize physical training and disease prevention. In both species, interleukin-6 (IL-6) plays a major role in regulating the inflammatory process induced during exercise as part of an integrated metabolic regulatory network. The aim of this study was to compare IL-6 and IL-6 receptor (IL-6R) mRNA expression in peripheral blood mononuclear cells (PBMCs) in trained and untrained humans and horses.Results Nine highly trained male swimmers (training volume: 21.6 ± 1.7 h/wk in 10-12 sessions) were compared with two age-matched control groups represented by eight lightly trained runners (training volume: 6.4 ± 2.6 h/wk in 3-5 sessions) and nine untrained subjects. In addition, eight trained horses (training volume: 8.0 ± 2.1 h/wk in 3-4 sessions) were compared with eight age-matched sedentary mares. In humans, IL-6 mRNA levels in PBMCs determined by quantitative reverse transcription-polymerase chain reaction were significantly higher in highly trained subjects, whereas IL-6R expression did not differ among groups. In horses, transcripts of both IL-6 and IL-6R were significantly up-regulated in the trained group.Conclusions Up-regulation of IL-6R expression in PBMCs in horses could reflect a mechanism that maintains an adequate anti-inflammatory environment at rest through ubiquitous production of anti-inflammatory cytokines throughout the body. These findings suggest that the system that controls the inflammatory response in horses is better adapted to respond to exercise than that in humans