An alternative approach of TUNEL assay to specifically characterize DNA fragmentation in cell model systems.

DNA damage is one of the most important effects induced by chemical agents. We report a comparative analysis of DNA fragmentation on three different cell lines using terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay, generally applied to detect apoptosis. Our approach combines cytogenetic techniques and investigation in detached cellular structures, recovered from the culture medium with the aim to compare the DNA fragmentation of three different cell line even beyond the cells adherent to substrate. Consequently, we detect any fragmentation points on single chromosomes, whole nuclei and other cellular structures. Cells were exposed to resveratrol (RSV) and doxorubicin (Doxo), in single and combined treatments. Control and treated astrocytes showed DNA damage in condensed nuclei and detached structures. Caco-2 cells showed fragmented DNA only after Doxo-treatment, while controls showed fragmented chromosomes, indicating DNA damage in replicating cells. MDA-MB-231 cells showed nuclear condensation and DNA fragmentation above all after RSV-treatment and related to detached structures. This model proved to perform a grading of genomic instability (GI). Astrocytes show a hybrid level of GI. Caco-2 cells showed fragmented metaphase chromosomes, proving that the DNA damage was transmitted to the daughter cells probably due to an absence of DNA repair mechanisms. Instead, MDA-MB-231 cells showed few or no fragmented metaphase, suggesting a probable activation of DNA repair mechanisms. By applying this alternative approach of TUNEL test, we obtained data that can more specifically characterize DNA fragmentation for a suitable application in various fields.

An In Vitro Model of Glioma Development.

Gliomas are the prevalent forms of brain cancer and derive from glial cells. Among them, astrocytomas are the most frequent. Astrocytes are fundamental for most brain functions, as they contribute to neuronal metabolism and neurotransmission. When they acquire cancer properties, their functions are altered, and, in addition, they start invading the brain parenchyma. Thus, a better knowledge of transformed astrocyte molecular properties is essential. With this aim, we previously developed rat astrocyte clones with increasing cancer properties. In this study, we used proteomic analysis to compare the most transformed clone (A-FC6) with normal primary astrocytes. We found that 154 proteins are downregulated and 101 upregulated in the clone. Moreover, 46 proteins are only expressed in the clone and 82 only in the normal cells. Notably, only 11 upregulated/unique proteins are encoded in the duplicated q arm of isochromosome 8 (i(8q)), which cytogenetically characterizes the clone. Since both normal and transformed brain cells release extracellular vesicles (EVs), which might induce epigenetic modifications in the neighboring cells, we also compared EVs released from transformed and normal astrocytes. Interestingly, we found that the clone releases EVs containing proteins, such as matrix metalloproteinase 3 (MMP3), that can modify the extracellular matrix, thus allowing invasion.

Addiction disorders: a need for change. Proposal for a new management. Position paper of SIA, Italian Society on Alcohol.

Various epidemiological and biological evaluations and the recent publication of the DSM-V (diagnostic and statistical manual of mental disorders) has imposed on the scientific community a period of reflection on the diagnosis and treatment of what in the DSM-IV was defined as "addiction". To date, the term "addiction" has been replaced by the DSM-5, because there is no global scientific consensus that has unequivocally characterized its clinical characteristics. This, we will talk about substance/alcohol use disorders (SUDs/AUDs) and disorders related to behavioral alterations (DBA) that can generate organic diseases, mental disorders, and social problems. In the first psychotic episode 40-70% of subjects meet the criteria of a SUDs/AUDs, excluding tobacco dependence. Substances can not only be the cause of a psychotic onset, but they can also disrupt a psychotic picture or interfere with drug therapy. The pharmacodynamic profiles of many substances are able to provoke the phenomenology of the main psychotic symptoms in a way that can be superimposed onto those presented by psychotic subjects without a history of SUDs/AUDs. The Department of Addictions (DAs) must not be absorbed by or incorporated into the Departments of Mental Health (DMH), with which, however, precise operational cooperation protocols will have to be defined and maintained, but it will have to maintain its own autonomy and independent connotation. Addiction Medicine is a discipline that brings together elements of public health, prevention, internal medicine, clinical pharmacology, neurology, and even psychiatry. The inclusion of the DAs in those of DMH refers purely to a problem of pathology that has to do with lifestyle, choices, and behaviors. These, over time, show their dysfunctionality and only then do related problems emerge. Moreover, epidemiological, social, and clinical motivations impose the creation of alcohological teams dedicated to alcohol-related activities. The collaboration with self-help-groups (SHGs) is mandatory. The action of SHGs is accredited in numerous international recommendations both on the basis of consensus and evidence in the literature.