Valentina Tirelli

Higher technical and professional education is increasingly discussed in relation to workforce readiness, innovation, and societal inclusion. In Italy, the PNRR-funded Rome Technopole operates as a multi-institutional ecosystem in which universities, research organizations, industry, and public bodies interact through a Hub & Spoke model to support training and innovation activities. Among its components, Spoke 4 addresses professional higher technical education through the co-development of modular learning initiatives involving multiple stakeholders. This commentary examines the role and activities of the Italian National Institute of Health (ISS) within this context, with particular reference to the development of two pilot modules: one on Artificial Intelligence and Algorethics, and one on Accessibility and Assistive Technologies, including applications supported by AI. The paper combines a conceptual discussion of the approach with selected empirical insights derived from pilot implementation, including stakeholder engagement processes, structured evaluations, and thematic prioritization exercises. The findings suggest the perceived relevance of multi-stakeholder co-design, the use of flexible and modular learning formats, and the integration of technical and ethical dimensions in higher technical education. At the same time, they point to challenges related to coordination, scalability, and alignment across institutional actors. Rather than proposing a definitive model, the Spoke 4 experience is discussed as a context-specific case that may offer insights contributing to ongoing debates on the design and implementation of higher technical education in complex, multi-institutional settings.

Higher technical education plays a strategic role in equipping the workforce to navigate rapid technological advancements and evolving labor market demands. Within the Rome Technopole framework, Spoke 4 targets ITS Academies, promoting the development of flexible, modular programs that integrate advanced technical skills with ethical, legal, and societal perspectives. This study reports on a pilot training initiative on Artificial Intelligence (AI) co-designed by the Istituto Superiore di Sanità (ISS), aimed at exploring the ethical, practical, and educational relevance of AI in higher technical education. The module was developed and tested through a multi-stakeholder collaboration involving educators, institutional actors, and learners. A four-phase approach was adopted: (1) initial stakeholder consultation to identify needs and content directions, (2) collaborative design of the training module, (3) online delivery and engagement using a CAWI-based focus group, and (4) mixed-method evaluation, combining quantitative assessments and open-ended qualitative feedback. This design facilitated asynchronous participation and encouraged critical reflection on the real-world implications of AI. Through the four-phase approach, the pilot module was developed, delivered, and assessed with 37 participants. Quantitative analysis revealed high ratings for clarity, relevance, and perceived utility in terms of employability. Qualitative feedback highlighted the interdisciplinary design, the integration of ethical reasoning, and the module’s broad applicability across sectors—particularly Healthcare and Industry. Participants suggested including more real-world case studies and collaborative learning activities to enhance engagement. The findings support the feasibility and added value of embedding ethically informed, interdisciplinary AI education in professional technical training pathways. Developed within the Rome Technopole ecosystem, the pilot module offers a promising approach to fostering critical digital literacy and preparing learners for responsible engagement with emerging technologies.

In recent years, there has been a notable increasing interest surrounding the identification and quantification of nano-sized particles, including extracellular vesicles (EVs) and viruses. The challenge posed by the nano-sized dimension of these particles makes precise examination a significant undertaking. Among the different techniques for the accurate study of EVs, flow cytometry stands out as the ideal method. It is characterized by high sensitivity, low time consumption, non-destructive sampling, and high throughput. In this article, we propose the optimization of flow cytometry procedures to identify, quantify, and purify EVs and virus-like particles. The protocol aims to reduce artefacts and errors in nano-sized particles counting, overall caused by the swarming effect. Different threshold strategies were compared to ensure result specificity. Additionally, the critical parameters to consider when using conventional flow cytometry outside of the common experimental context of use have also been identified. Finally, fluorescent-EVs sorting protocol was also developed with highly reliable results using a conventional cell sorter.

In recent years, there has been a notable increasing interest surrounding the identification and quantification of nanosized particles, including extracellular vesicles (EVs) and viruses. The challenge posed by the nano-sized dimension of these particles makes precise examination a significant undertaking. Among the different techniques for the accurate study of EVs, Flow cytometry (FCM) stands out as the ideal method. It is characterized by high sensitivity, low time consumption, non-destructive sampling, and high throughput. In this article, we propose the optimization of FCM procedures to identify, quantify, and purify EVs and virus like particles (VLPs). The protocol aims to reduce artifacts and errors in nano-sized particles counting, overall caused by the swarming effect. Different threshold strategies were compared to ensure result specificity. Additionally, the critical parameters to consider when using conventional FCM outside of the common experimental context of use have also been identified. Finally, fluorescent-EVs sorting protocol was also developed with highly reliable results using a conventional cell sorter.

The cell cycle and the transcriptome dynamics of yeast exposed to extracellular self-DNA during an aerobic batch culture on glucose have been investigated using cytofluorimetric and RNA-seq analyses. In parallel, the same study was conducted on yeast cells growing in the presence of (heterologous) nonself-DNA. The self-DNA treatment determined a reduction in the growth rate and a major elongation of the diauxic lag phase, as well as a significant delay in the achievement of the stationary phase. This was associated with significant changes in the cell cycle dynamics, with slower exit from the G0 phase, followed by an increased level of cell percentage in the S phase, during the cultivation. Comparatively, the exposure to heterologous DNA did not affect the growth curve and the cell cycle dynamics. The transcriptomic analysis showed that self-DNA exposure produced a generalized downregulation of transmembrane transport and an upregulation of genes associated with sulfur compounds and the pentose phosphate pathway. Instead, in the case of the nonself treatment, a clear response to nutrient deprivation was detected. Overall, the presented findings represent further insights into the complex functional mechanisms of self-DNA inhibition.

Eosinophils are a rare immune cell subset with important roles in Th2 immunity and, recently, in cancer. Interleukin IL-33 (IL-33) is well recognized for its important roles in the activation of eosinophils in Th2 immunity. On the other hand, IL-33 has been recently discovered to play central roles in cancer, in particular by activating eosinophils and increase their degranulation consequent to an intrinsic tumor cell killing function. We propose a dual approach methodology to extrapolate functional interactions of eosinophils with tumor cells, as a result of eosinophil stimulation. Human eosinophils (Eos) isolated from the blood of healthy donors by dextran sedimentation followed by magnetic sorting are exposed to IL-33 (Eos33) or IL-5 (Eos5, control) for 18 h. These pre-conditioned cells are then co-cultured with A375P melanoma cells to monitor cell-cell interactions. Acoustic focusing flow cytometry analysis is employed to evaluate the presence of Eos-tumor cell conjugates after 1h incubation of human eosinophils and A375P melanoma cells. Moreover, a 24 h time-lapse video recording approach is employed to obtain single cell tracking Eos profiles. This allows to quantitatively determine the interaction extent of Eos33, as opposed to Eos5 (control condition), with tumor cells. In conclusion, our protocols easily and quickly allow the extrapolation of relevant kinematic and biologically relevant parameters for tumor reactive eosinophils. Furthermore, these methods are adaptable to various models with other types of immune cell subsets and cancer cells and can be implemented on different video microscopy platforms and advanced flow cytometry systems.

Basophils constitute a rare population of granulocytes with key functions in allergies, immunodeficiencies and cancer. The scarcity of basophils in the human blood and tissues constitutes a considerable limit for the study of these cells. Interleukin-3 (IL-3) is a cytokine that stimulates both the differentiation and the expansion of basophils from bone marrow (BM) precursors by positively regulating the expression of the IL3Ra receptor. We have standardized an in vitro differentiation protocol of mouse basophils (mBaso) from BM precursors through culture in presence of IL-3 for 10 days followed by cell sorting. At the end of the 10-day differentiation, a considerable number of mBaso can be obtained and cell sorting procedures further improved the isolation of an extraordinarily pure (>98%) and vital FcεR1 ⁺ CD11c ^- c-kit ^- mBaso population. Phenotypic analysis revealed that terminally differentiated (day 10) unsorted mBaso cultured for 24 hours showed a decrease in basophilic lineage (c-kit ^- ) and an increase of mastocytic lineage (c-kit ⁺ ) and reduced the expression of basophil markers FcεRI, CD49b and CD200R either in absence of stimuli or following activation with the alarmin IL-33, indicating cell dedifferentiation. In contrast, terminally differentiated and FcεR1 ⁺ CD11c ^- c-kit ^- sorted mBaso do not dedifferentiate in mast cells when placed in culture, and responded to IL-33 stimulation by up-regulating the activation marker CD63 without down-modulation of FcεRI and CD200R3. These evidences highlight that in vitro differentiation followed by cell sorting is a useful method to obtain elevated numbers of highly pure mBaso that preserve their lineage markers and thus are suitable for conducting the desired functional studies.

We developed a new class of inhibitors of protein-protein interactions of the SHP2 phosphatase, which is pivotal in multiple signaling pathways and a central target in the therapy of cancer and rare diseases. Currently available SHP2 inhibitors target the catalytic site or an allosteric pocket but lack specificity or are ineffective on disease-associated SHP2 mutants. Based on the consideration that pathogenic lesions cause signaling hyperactivation due to increased SHP2 association with cognate proteins, we developed peptide-based molecules with low nM affinity for the N-terminal Src homology domain of SHP2, good selectivity, stability to degradation and an affinity for pathogenic variants of SHP2 up to 20 times higher than for the wild-type protein. The best peptide reverted the effects of a pathogenic variant (D61G) in zebrafish embryos. Our results provide a novel route for SHP2-targeted therapies and a tool to investigate the role of protein-protein interactions in the function of SHP2.