Perifosine as a Potential Novel Anti-Cancer Agent Inhibits EGFR/MET-AKT Axis in Malignant Pleural Mesothelioma

PI3K/AKT signalling pathway is aberrantly active and plays a critical role for cell cycle progression of human malignant pleural mesothelioma (MMe) cells. AKT is one of the important cellular targets of perifosine, a novel bio-available alkylphospholipid that has displayed significant anti-proliferative activity in vitro and in vivo in several human tumour model systems and is currently being tested in clinical trials.We tested Perifosine activity on human mesothelial cells and different mesothelioma cell lines, in order to provide evidence of its efficacy as single agent and combined therapy.We demonstrate here that perifosine, currently being evaluated as an anti-cancer agent in phase 1 and 2 clinical trials, caused a dose-dependent reduction of AKT activation, at concentrations causing MMe cell growth arrest. In this study we firstly describe that MMe cells express aside from AKT1 also AKT3 and that either the myristoylated, constitutively active, forms of the two proteins, abrogated perifosine-mediated cell growth inhibition. Moreover, we describe here a novel mechanism of perifosine that interferes, upstream of AKT, affecting EGFR and MET phosphorylation. Finally, we demonstrate a significant increase in cell toxicity when MMe cells were treated with perifosine in combination with cisplatin.This study provides a novel mechanism of action of perifosine, directly inhibiting EGFR/MET-AKT1/3 axis, providing a rationale for a novel translational approach to the treatment of MMe

First update of the living European guideline (EuroGuiDerm) on atopic eczema

First update of the living European guideline (EuroGuiDerm) on atopic eczem

Essential versus accessory aspects of cell death: recommendations of the NCCD 2015

Cells exposed to extreme physicochemical or mechanical stimuli die in an uncontrollable manner, as a result of their immediate structural breakdown. Such an unavoidable variant of cellular demise is generally referred to as ‘accidental cell death’ (ACD). In most settings, however, cell death is initiated by a genetically encoded apparatus, correlating with the fact that its course can be altered by pharmacologic or genetic interventions. ‘Regulated cell death’ (RCD) can occur as part of physiologic programs or can be activated once adaptive responses to perturbations of the extracellular or intracellular microenvironment fail. The biochemical phenomena that accompany RCD may be harnessed to classify it into a few subtypes, which often (but not always) exhibit stereotyped morphologic features. Nonetheless, efficiently inhibiting the processes that are commonly thought to cause RCD, such as the activation of executioner caspases in the course of apoptosis, does not exert true cytoprotective effects in the mammalian system, but simply alters the kinetics of cellular demise as it shifts its morphologic and biochemical correlates. Conversely, bona fide cytoprotection can be achieved by inhibiting the transduction of lethal signals in the early phases of the process, when adaptive responses are still operational. Thus, the mechanisms that truly execute RCD may be less understood, less inhibitable and perhaps more homogeneous than previously thought. Here, the Nomenclature Committee on Cell Death formulates a set of recommendations to help scientists and researchers to discriminate between essential and accessory aspects of cell death

Optimum porphyrin accumulation in epithelial skin tumours and psoriatic lesions after topical application of δ-aminolaevulinic acid

Photodynamic therapy with topically applied δ-aminolaevulinic acid is used to treat skin tumours by employing endogenously formed porphyrins as photosensitizers. This study examines the time course of porphyrin metabolite formation after topical application of δ-aminolaevulinic acid. Porphyrin biosynthesis in human skin tumours (basal cell carcinoma, squamous cell carcinoma), in psoriatic lesions, and in normal skin was investigated. Skin areas were treated with δ-aminolaevulinic acid, and levels of total porphyrins, porphyrin metabolites and proteins were measured in samples excised after 1, 2, 4, 6, 9, 12 and 24 h. There was an increase in porphyrin biosynthesis in all tissues with maximum porphyrin levels in tumours between 2 and 6 h and in psoriatic lesions 6 h after treatment. The pattern of porphyrins showed no significant difference between normal and neoplastic skin, protoporphyrin being the predominant metabolite. The results suggest that optimum irradiation time for superficial epithelial skin tumours may be as soon as 2 h after application of δ-aminolaevulinic acid, whereas for treatment of psoriatic lesions an application time of 6 h is more suitable. © 1999 Cancer Research Campaig

Vulnerability of Polarised Intestinal Porcine Epithelial Cells to Mycotoxin Deoxynivalenol Depends on the Route of Application

BACKGROUND AND AIMS: Deoxynivalenol (DON) is a Fusarium derived mycotoxin, often occurring on cereals used for human and animal nutrition. The intestine, as prominent barrier for nutritional toxins, has to handle the mycotoxin from the mucosa protected luminal side (apical exposure), as well as already absorbed toxin, reaching the cells from basolateral side via the blood stream. In the present study, the impact of the direction of DON exposure on epithelial cell behaviour and intestinal barrier integrity was elucidated. METHODS: A non-transformed intestinal porcine epithelial cell line (IPEC-J2), cultured in membrane inserts, serving as a polarised in vitro model to determine the effects of deoxynivalenol (DON) on cellular viability and tight junction integrity. RESULTS: Application of DON in concentrations up to 4000 ng/mL for 24, 48 and 72 hours on the basolateral side of membrane cultured polarised IPEC-J2 cells resulted in a breakdown of the integrity of cell connections measured by transepithelial electrical resistance (TEER), as well as a reduced expression of the tight junction proteins ZO-1 and claudin 3. Epithelial cell number decreased and nuclei size was enlarged after 72 h incubation of 4000 ng/mL DON from basolateral. Although necrosis or caspase 3 mediated apoptosis was not detectable after basolateral DON application, cell cycle analysis revealed a significant increase in DNA fragmentation, decrease in G0/G1 phase and slight increase in G2/M phase after 72 hours incubation with DON 2000 ng/mL. CONCLUSIONS: Severity of impact of the mycotoxin deoxynivalenol on the intestinal epithelial barrier is dependent on route of application. The epithelium appears to be rather resistant towards apical (luminal) DON application whereas the same toxin dose from basolateral severely undermines barrier integrity

Apoptosis-Like Death in Bacteria Induced by HAMLET, a Human Milk Lipid-Protein Complex

Background: Apoptosis is the primary means for eliminating unwanted cells in multicellular organisms in order to preserve tissue homeostasis and function. It is characterized by distinct changes in the morphology of the dying cell that are orchestrated by a series of discrete biochemical events. Although there is evidence of primitive forms of programmed cell death also in prokaryotes, no information is available to suggest that prokaryotic death displays mechanistic similarities to the highly regulated programmed death of eukaryotic cells. In this study we compared the characteristics of tumor and bacterial cell death induced by HAMLET, a human milk complex of alpha-lactalbumin and oleic acid. Methodology/Principal Findings: We show that HAMLET-treated bacteria undergo cell death with mechanistic and morphologic similarities to apoptotic death of tumor cells. In Jurkat cells and Streptococcus pneumoniae death was accompanied by apoptosis-like morphology such as cell shrinkage, DNA condensation, and DNA degradation into high molecular weight fragments of similar sizes, detected by field inverse gel electrophoresis. HAMLET was internalized into tumor cells and associated with mitochondria, causing a rapid depolarization of the mitochondrial membrane and bound to and induced depolarization of the pneumococcal membrane with similar kinetic and magnitude as in mitochondria. Membrane depolarization in both systems required calcium transport, and both tumor cells and bacteria were found to require serine protease activity (but not caspase activity) to execute cell death. Conclusions/Significance: Our results suggest that many of the morphological changes and biochemical responses associated with apoptosis are present in prokaryotes. Identifying the mechanisms of bacterial cell death has the potential to reveal novel targets for future antimicrobial therapy and to further our understanding of core activation mechanisms of cell death in eukaryote cells

The Targeting of Plasmalemmal Ceramide to Mitochondria during Apoptosis

Ceramide is a key lipid mediator of cellular processes such as differentiation, proliferation, growth arrest and apoptosis. During apoptosis, ceramide is produced within the plasma membrane. Although recent data suggest that the generation of intracellular ceramide increases mitochondrial permeability, the source of mitochondrial ceramide remains unknown. Here, we determine whether a stress-mediated plasmalemmal pool of ceramide might become available to the mitochondria of apoptotic cells. We have previously established annexin A1—a member of a family of Ca2+ and membrane-binding proteins—to be a marker of ceramide platforms. Using fluorescently tagged annexin A1, we show that, upon its generation within the plasma membrane, ceramide self-associates into platforms that subsequently invaginate and fuse with mitochondria. An accumulation of ceramide within the mitochondria of apoptotic cells was also confirmed using a ceramide-specific antibody. Electron microscopic tomography confirmed that upon the formation of ceramide platforms, the invaginated regions of the plasma membrane extend deep into the cytoplasm forming direct physical contacts with mitochondrial outer membranes. Ceramide might thus be directly transferred from the plasma membrane to the mitochondrial outer membrane. It is conceivable that this “kiss-of-death” increases the permeability of the mitochondrial outer membrane thereby triggering apoptosis

Mitochondrial dysfunction and biogenesis: do ICU patients die from mitochondrial failure?

Mitochondrial functions include production of energy, activation of programmed cell death, and a number of cell specific tasks, e.g., cell signaling, control of Ca2+ metabolism, and synthesis of a number of important biomolecules. As proper mitochondrial function is critical for normal performance and survival of cells, mitochondrial dysfunction often leads to pathological conditions resulting in various human diseases. Recently mitochondrial dysfunction has been linked to multiple organ failure (MOF) often leading to the death of critical care patients. However, there are two main reasons why this insight did not generate an adequate resonance in clinical settings. First, most data regarding mitochondrial dysfunction in organs susceptible to failure in critical care diseases (liver, kidney, heart, lung, intestine, brain) were collected using animal models. Second, there is no clear therapeutic strategy how acquired mitochondrial dysfunction can be improved. Only the benefit of such therapies will confirm the critical role of mitochondrial dysfunction in clinical settings. Here we summarized data on mitochondrial dysfunction obtained in diverse experimental systems, which are related to conditions seen in intensive care unit (ICU) patients. Particular attention is given to mechanisms that cause cell death and organ dysfunction and to prospective therapeutic strategies, directed to recover mitochondrial function. Collectively the data discussed in this review suggest that appropriate diagnosis and specific treatment of mitochondrial dysfunction in ICU patients may significantly improve the clinical outcome

Molecular mechanisms of cell death: recommendations of the Nomenclature Committee on Cell Death 2018.

Over the past decade, the Nomenclature Committee on Cell Death (NCCD) has formulated guidelines for the definition and interpretation of cell death from morphological, biochemical, and functional perspectives. Since the field continues to expand and novel mechanisms that orchestrate multiple cell death pathways are unveiled, we propose an updated classification of cell death subroutines focusing on mechanistic and essential (as opposed to correlative and dispensable) aspects of the process. As we provide molecularly oriented definitions of terms including intrinsic apoptosis, extrinsic apoptosis, mitochondrial permeability transition (MPT)-driven necrosis, necroptosis, ferroptosis, pyroptosis, parthanatos, entotic cell death, NETotic cell death, lysosome-dependent cell death, autophagy-dependent cell death, immunogenic cell death, cellular senescence, and mitotic catastrophe, we discuss the utility of neologisms that refer to highly specialized instances of these processes. The mission of the NCCD is to provide a widely accepted nomenclature on cell death in support of the continued development of the field