Search

Abstract Background Liquid biopsy based on cell-free DNA (cfDNA) analysis holds significant promise as a minimally invasive approach for the diagnosis, genotyping, and monitoring of solid malignancies. Human tumors release cfDNA in the bloodstream through a combination of events, including cell death, active and passive release. However, the precise mechanisms leading to cfDNA shedding remain to be characterized. Addressing this question in patients is confounded by several factors, such as tumor burden extent, anatomical and vasculature barriers, and release of nucleic acids from normal cells. In this work, we exploited cancer models to dissect basic mechanisms of DNA release. Methods We measured cell loss ratio, doubling time, and cfDNA release in the supernatant of a colorectal cancer (CRC) cell line collection (N = 76) representative of the molecular subtypes previously identified in cancer patients. Association analyses between quantitative parameters of cfDNA release, cell proliferation, and molecular features were evaluated. Functional experiments were performed to test the impact of modulating DNA methylation on cfDNA release. Results Higher levels of supernatant cfDNA were significantly associated with slower cell cycling and increased cell death. In addition, a higher cfDNA shedding was found in non-CpG Island Methylator Phenotype (CIMP) models. These results indicate a positive correlation between lower methylation and increased cfDNA levels. To explore this further, we exploited methylation microarrays to identify a subset of probes significantly associated with cfDNA shedding and derive a methylation signature capable of discriminating high from low cfDNA releasers. We applied this signature to an independent set of 176 CRC cell lines and patient derived organoids to select 14 models predicted to be low or high releasers. The methylation profile successfully predicted the amount of cfDNA released in the supernatant. At the functional level, genetic ablation of DNA methyl-transferases increased chromatin accessibility and DNA fragmentation, leading to increased cfDNA release in isogenic CRC cell lines. Furthermore, in vitro treatment of five low releaser CRC cells with a demethylating agent was able to induce a significant increase in cfDNA shedding. Conclusions Methylation status of cancer cell lines contributes to the variability of cfDNA shedding in vitro. Changes in methylation pattern are associated with cfDNA release levels and might be exploited to increase sensitivity of liquid biopsy assays.

Introduction There are many stressors that lead to burn out and decrease the quality of life of health professionals as a whole and it occurs also to trainee psychiatrists. Training programs rarely include specific interest in the personal self of students even if they begin to deal with severe human suffering. Authors present a model of experiential group training in psychiatry that is centred on the person/trainee at the very most. Objectives The aim is to focus on unsolved emotional needs of students to allow them to achieve the capacity of relationship with patients. It is not a mere application of empathy but a truth overcoming, for trainees, of major risks of collusion due to reflection of individual conflicts into the patients and/or due to the encounter with strong emotion during clinical training. Methods The model is Experiential because it is the space for personal expression and it is Strategic because it is born inside the strategic group training in psychotherapy (Battuello et al. Psichiatria e Psicoterapia 2022; 41, 2, 65-82). The conductor of the group carries on her/his skin the experience of own training group, to be brought into the trainees’ one. This is an original approach because the epistemology of the model came directly from the experience. The group is led by a psychotherapist that plays an active part inside the process instead of being only a facilitator. The main focus is to allow students to express themselves that includes various steps such as: tuning with their own experiences/emotions, freedom of expressing them to the group, active listening to the other and response to the same other even when feelings don’t resonate but instead are divergent. This phase is related to the conductor’s participation that is totally immersed into the group bringing personal feelings, stories and emotions to create an undifferentiated space, free from hierarchical roles. During a second period, students can access a more mature relational capacity that carries the group to a phase of individuation of the self that also engages professional issues. Results Students in the group question themselves: it is the root of every health professional that has to explore and overcome personal relational issues. Only after the expressiveness phase, as authors name this part of the training, an individuation phase is truly possible that leads to the definition of the professional. Conclusions The training group is necessary for students to explore the wider range of emotions, expressing personal ones, accepting others’, experiencing the tolerance to their frustration, and emerging as professional, that is, in few words, professional of the relationship, the key and the basement of the psychiatrist. The training in mental health should include, at first, the taking care of the person/student as it is proposed by authors inside the group model. Disclosure of Interest None Declared

Among non-native copepods, the calanoid Pseudodiaptomus marinus Sato, 1913 is the species probably spreading at the fastest pace in European and neighbouring waters since its first record in the Adriatic Sea in 2007. In this contribution, we provide an update on the distribution of P. marinus in the Mediterranean and Black Seas, along the Atlantic coasts of Europe, in the English Channel and in the southern North Sea. Starting from a previous distribution overview, we include here original and recently (2019–2023) published data to show the novel introduction of this species in different geographical areas, and its secondary spreading in already colonised regions. The picture drawn in this work confirms the strong ability of P. marinus to settle in environments characterised by extremely diverse abiotic conditions, and to take advantage of different vectors of introduction. The data presented allow speculations on realistic future introductions of P. marinus and on the potential extension of its distribution range.

The Mediterranean Sea is subjected to a high anthropic pressure, which determines direct or indirect discharges of persistent organic pollutants deriving from intensive industrial activities. These compounds could easily enter and contaminate the whole marine compartment, with possible transfers (and contamination) among water, sediment and biota. Based on the above-mentioned assumptions, in this work we studied the presence of 16 polycyclic aromatic hydrocarbons (PAHs) and 14 dioxin and non-dioxin-like polychlorinated biphenyls (PCBs) in the neritic protected marine area of the Southern Ligurian Sea, affected by the impact of human activities. The study was focused on the possible partition of micropollutants within seawater, sediment and zooplankton. Results showed that both seasonal and anthropic causes strongly affect contaminant transfer behaviors, with summertime periods more impacted by PAH and PCB contamination. Regarding the PAH contamination, low molecular weight congeners were mainly detected in the target matrices, revealing concentrations up to 1 µg/L in seawater (anthracene), 250 µg/Kg in sediments (benzo[b]fluoranthene) and 2.3 mg/Kg in carnivorous copepods. Concerning PCBs, only few congeners were detected in the matrices studied. To better understand the occurrence of preferential bioaccumulation pathways in zooplankton, partition studies were also performed in several taxa (hyperbenthic Isopoda, holoplanktonic crustacean copepods and ichthyoplankton) through the calculation of BAF values, observing that both living and feeding habits could influence the bioaccumulation process.