Apoptotic Bodies: Particular Extracellular Vesicles Involved in Intercellular Communication.

In the last decade, a new method of cell-cell communication mediated by membranous extracellular vesicles (EVs) has emerged. EVs, including exosomes, microvesicles, and apoptotic bodies (ApoBDs), represent a new and important topic, because they are a means of communication between cells and they can also be involved in removing cellular contents. EVs are characterized by differences in size, origin, and content and different types have different functions. They appear as membranous sacs released by a variety of cells, in different physiological and patho-physiological conditions. Intringuingly, exosomes and microvesicles are a potent source of genetic information carriers between different cell types both within a species and even across a species barrier. New, and therefore still relatively poorly known vesicles are apoptotic bodies, on which numerous in-depth studies are needed in order to understand their role and possible function. In this review we would like to analyze their morpho-functional characteristics

Liquid Biopsy: A Family of Possible Diagnostic Tools

none1noLiquid biopsies could be considered an excellent diagnostic tool, in different physiological or pathological conditions. The possibility of using liquid biopsies for non-invasive clinical purposes is quite an old idea: indeed many years ago it was already being used in the field of non-invasive prenatal tests (NIPT) for autosomal fetal aneuploidy evaluation. In 1997 Lo et al. had identified fetal DNA in maternal plasma and serum, showing that about 10-15% of cfDNA in maternal plasma is derived from the placenta, and biologic fluid represents an important and non-invasive technique to evaluate state diseases and possible therapies. Nowadays, several body fluids, such as blood, urine, saliva and other patient samples, could be used as liquid biopsy for clinical non-invasive evaluation. These fluids contain numerous and various biomarkers and could be used for the evaluation of pathological and non-pathological conditions. In this review we will analyze the different types of liquid biopsy, their potential role in clinical diagnosis and the functional involvement of extracellular vesicles in these fluids as carriers.openMichela, BattistelliBattistelli, Michel

Further considerations on in vitro skeletal muscle cell death

The present review discusses the apoptotic behavior induced by chemical and physical triggers in C2C12 skeletal muscle cells, comparing myoblast to myotube sensitivity, and investigating it by means of morphological, biochemical and cytofluorimetric analyses. After all treatments, myotubes, differently from myoblasts, showed a poor sensitivity to cell death. Intriguingly, in cells exposed to staurosporine, etoposide and UVB radiation, apoptotic and normal nuclei within the same fibercould be revealed. The presence of nuclear-dependent “territorial” death domains in the syncytium could explain a delayed cell death of myotubes compared to mononucleated cells. Moreover, autophagic granules abundantly appeared in myotubes after each treatment. Autophagy could protect muscle cell integrity against chemical and physical stimuli, making C2C12 myotubes, more resistant to cell death inductio

Prevention of UVB radiation-induced cell death: “in vitro” studies

The ultraviolet component of sun light consists of UVA, UVB and UVC rays. UVB radiation represents an environmental hazard because of its role in skin aging, cancer and infection exacerbation. UVB stimulate the production of reactive oxygen species (ROS) in epidermal cells, resulting in skin lesions, accelerating aging and eliciting malignancies. At least 50% of UVB-induced damage is attributable to the formation of reactive ROS which cause cellular lesions if antioxidant defence mechanisms are down-regulated. Thus, exogenous supplementation of antioxidants may be an effective strategy to reduce or prevent skin damage. In the last years, we demonstrated the antioxidant effects of melatonin (Mel) (Luchetti et al., 2006) and, more recently of hydroxytyrosol (HyT) and its derivatives (Burattini et al., 2013) in hemopoietic human cells exposed to pro-oxidants. Therefore, in this project we propose to evaluate the antioxidant and/or anti-apoptotic effect of Mel and HyT in HaCaT human keratinocytes exposed to UVB. Keratinocytes in the non-irradiated condition are morphologically similar in Mel- and HyT-treated and untreated group. TUNEL reaction appears negative in both conditions, as well as in control. UVB radiation induces a significant decrease in cell confluence, with a diffuse cell detachment and the appearance of rounding and blebbed cells. TUNEL reaction evidences several nuclei with DNA fragmentation in UVB treated keratinocytes. In addition, cell viability evaluated by means of supravital propidium iodide (PI) evidences a diffuse staining positivity. Pre-treatment with Mel or HyT before UVB exposure is able to reduce cell death. In conclusion, HyT and Mel evidence an intringuing capability to prevent cell death in keratinocytes too. They could so represent a potential tool in skin protection from UVB radiation

Melatonin role in skeletal muscle disorders

Abstract. – OBJECTIVE: This review discusses the impact of the neuro-hormone melatonin on skeletal muscle disorders based on recent literature data with the aim to clarify the utility of the melatonin therapy in patients affected by muscle diseases. MATERIALS AND METHODS: It has been pointed out the possible role of melatonin as a food supplement to cure muscular disorders characterized by muscle wasting. Oxidative damage has been proposed as one of the major contributors of the skeletal muscle decline occurring both in physiological and pathological conditions. It is known that excessive oxidant levels lead to mitochondrial damage, and in turn, contribute to apoptotic signaling activation and autophagic impairment. This condition is common in a variety of skeletal muscle disorders. RESULTS: The scientific evidence enhances the antioxidant effect of melatonin, that has been demonstrated by several studies both in vitro and in vivo. This effect counteracts mitochondrial impairments and reduces oxidative stress and autophagic alterations in muscle fibers. Its beneficial role in restoring muscle decline, takes place mainly in atrophic conditions correlated to muscle aging. CONCLUSIONS: The findings of the research suggest that melatonin may be considered as a valid dietary supplement, useful to prevent muscle wasting, in particular, in sarcopenia-associated diseases

Oxidative stress to promote cell death or survival

Editoria

Protective effect of different antioxidant agents in UVB-irradiated keratinocytes

Skin cells can respond to UVB-induced damage either by tolerating it, or restoring it through antioxidant activation and DNA repair mechanisms or, ultimately, undergoing programmed cell death, when damage is massive. Nutritional factors, in particular, food antioxidants, have attracted much interest because of their potential use in new preventive, protective, and therapeutic strategies for chronic degenerative diseases, including skin inflammation and cancer. Some polyphenols, present in virgin olive oil, well tolerated by organism after oral administration, show a variety of pharmacological and clinical benefits such as anti-oxidant, anti-cancer, anti-inflammatory, and neuro-protective activities. Here, the protective effects of antioxidant compounds against UV-induced apoptosis have been described in HaCat cell line. Human keratinocytes were pre-treated with antioxidants before UVB exposure and their effects have been evaluated by means of ultrastructural analyses. After UVB radiation, a known cell death trigger, typical apoptotic features, absent in control condition and in antioxidant alone-treated cells, appear. An evident numerical decrease of ultrastructural apoptotic patterns and TUNEL positive nuclei can be observed when natural antioxidants were supplied before cell death induction. These data have been confirmed by molecular investigation of caspase activity. In conclusion, this paper highlights antioxidant compound ability to prevent apoptotic cell death in human keratinocytes exposed to UVB, suggesting, for these molecules, a potential role in preventing skin damage

Melatonin effects in normal and tumoral skeletal muscle cells: a preliminary study

Melatonin (MEL), also chemically known as N-acetyl-5-methoxytryptamine, is a hormone found in animals, plants, and microbes. It exhibits strong antioxidant effects and thanks to its structure it is able to diffuse through all the biological membranes, also overcoming the blood-brain barrier and the placenta (Salucci et al., 2014). Numerous in vitro and in vivo studies have documented Mel ability to induce apoptosis in tumor cells while inhibiting it in the normal ones (Cristofanos et al, 2009; Lanoix et al., 2011). In this study MEL activity has been investigated in vitro both in murine skeletal muscle (C2C12) and in alveolar rhabdomyosarcoma (RH30) cell lines by means of morpho-functional approaches. If MEL low concentrations are well tolerated by normal skeletal muscle cells, its effect appears completely different in tumor cells, where MEL can be considered a powerful apoptotic trigger. In RH30 cells, blebbing, chromatin condensation and margination, apoptotic bodies occur as well as necrotic cell death features. The latter appeared after prolonged exposure to MEL. In conclusion, the neuro-hormone shows a strong dose and time dependent pro-apoptotic activity and it could represent a potential tool in association with the current chemotherapeutic compounds to resolve alveolar rhabdomyosarcoma, the most common pediatric skeletal muscle tissue malignancy

Skeletal muscle cell death induced by physical agents

Apoptosis plays a pivotal role in the deletion of unwanted, damaged, or infected cells in multicellular organisms, as well as in development and tissue homeostasis, cell differentiation, and proliferation. In skeletal muscle cells it is unique for several reasons. First, skeletal muscle fibre is multinucleated. So muscle cell death is correlated to a loss of gene expression within the local myonuclear domain, potentially leading to muscle atrophy. In addition, skeletal muscle is a plastic tissue capable of changing its mitochondrial content and/or composition in response to chronic alterations in muscle use or disuse (Siu et al., 2009). Most of the research evidenced that many of the external apoptotic stimuli activate signaling pathways that converge on the mitochondria, determining cell death (Adhietty et al., 2008). Physical triggers such as UVB (D’Emilio et al., 2010), hyperthermia (Lee et al., 2011) and hypothermia (Pizanis et al., 2011) induced cell death by mitochondrial pathways in various cell types. In addition also low pH usually induced DNA damage in other cell lines (Xiao et al., 2003). The aim of this work is to investigate in vitro skeletal muscle cell death appearing after exposure to physical triggers, by means of TUNEL reaction, analysed at confocal microscope, and of electron microscopy. C2C12 myoblasts and myotubes, grown as previously reported (D’Emilio et al., 2010), were exposed to UV-B (312nm) for 30 min, hyperthermia 45°C for 1h and hypothermia (2-6°C) and low pH (5) for 4h. All treatments were followed by 2h recovery. Control cell evidentiated a good morphology and appeared negative to TUNEL reaction. UVB - treated sample presented nuclear features suggest apoptosis both at electron and confocal microscopy and in undifferentiated and differentiated conditions. Hyperthermia induced both apoptosis and necrosis with cell rounding and a certain positivity to TUNEL reaction both in myoblasts and myotubes. After hypothermia apoptosis was observed in some cells, but the majority appeared similar to the control, so evidentiating a scarce response. Cells treated with low had swollen nuclei, sometimes showing a thin film of condensed chromatin, and occasionally TUNEL-positive. In all conditions cytoplasm vacuolisaton and autophagic vacuole increase appeared. These findings suggest that skeletal muscle cells seem to be sensitive to physical agents induced cell death

Cell damage induced by asbestos similar particles

The presence, in nature, of asbestos similar particles, highly toxic and potentially cancerogenic for human healthy is well known (1). Inhalation of the fibrous form of erionite, has been shown to cause effects compared to those observed with mineral fibers classified as ‘‘asbestos,’’ including malignant mesothelioma, a disease typically associated with occupational and environmental exposures to asbestos (2). In this work various zeolite materials have been considered because of their suspected carcinogenic activity and, the possible interactions occurring between asbestiform fibers and U937 cell, a human hemopoietic cell line, have been evaluated. Chemical and morpho-functional analyses have been carried out, both to characterize fiber structure and cell response. Cells showed the ability to internalize the minerals, as observed after TEM analyses. With zeolite exposure time increasing, a diffuse cell damage with features of apoptotic and necrotic death can be evidenced (3). These findings suggest that the fibrous form of scolecite or offretite too can be considered potentially toxic for cell culture in vitro