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Huntingtin-lowering strategies are central to therapeutic approaches for Huntington's disease. Recent studies reported the induction of age- and cell type-specific phenotypes by conditional huntingtin knockout, but these experimental conditions did not precisely mimic huntingtin-lowering or gene-editing conditions in terms of the cells targeted and brain distribution, and no transcriptional profiles were provided. Here, we used the adeno-associated delivery system commonly used in CNS gene therapy programmes and the self-inactivating KamiCas9 gene-editing system to investigate the long-term consequences of wild-type mouse huntingtin inactivation in adult neurons and, thus, the feasibility and safety of huntingtin inactivation in these cells. Behavioural and neuropathological analyses and single-nuclei RNA sequencing indicated that huntingtin editing in 77% of striatal neurons and 16% of cortical projecting neurons in adult mice induced no behavioural deficits or cellular toxicity. Single-nuclei RNA sequencing in 11.5-month-old animals showed that huntingtin inactivation did not alter striatal-cell profiles or proportions. Few differentially expressed genes were identified and Augur analysis confirmed an extremely limited response to huntingtin inactivation in all cell types. Our results therefore indicate that wild-type huntingtin inactivation in adult striatal and projection neurons is well tolerated in the long term., Competing Interests: The authors have no conflicts of interest to declare., (© The Author(s) 2023. Published by Oxford University Press on behalf of the Guarantors of Brain.)

One obstacle to the development of gene therapies for the central nervous system is the lack of workflows for quantifying transduction efficiency in affected neural networks and ultimately predicting therapeutic potential. We integrated data from a brain cell atlas with 3D or 2D semi-automated quantification of transduced cells in segmented images to predict AAV transduction efficiency in multiple brain regions. We used this workflow to estimate the transduction efficiency of AAV2/rh.10 and AAV2.retro co-injection in the corticostriatal network affected in Huntington's disease. We then validated our pipeline in gene editing experiments targeting both human and mouse huntingtin genes in transgenic and wild-type mice, respectively. Our analysis predicted that 54% of striatal cells and 7% of cortical cells would be edited in highly transduced areas. Remarkably, in the treated animals, huntingtin gene inactivation reached 54.5% and 9.6%, respectively. These results demonstrate the power of this workflow to predict transduction efficiency and the therapeutic potential of gene therapies in the central nervous system., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Author(s).)

Gene transfer is a widely developed technique for studying and treating genetic diseases. However, the development of therapeutic strategies is challenging, due to the cellular and functional complexity of the central nervous system (CNS), its large size and restricted access. We explored two parameters for improving gene transfer efficacy and capacity for the selective targeting of subpopulations of cells with lentiviral vectors (LVs). We first developed a second-generation LV specifically targeting astrocytes for the efficient expression or silencing of genes of interest, and to better study the importance of cell subpopulations in neurological disorders. We then made use of the retrograde transport properties of a chimeric envelope to target brain circuits affected in CNS diseases and achieve a broad distribution. The combination of retrograde transport and specific tropism displayed by this LV provides opportunities for delivering therapeutic genes to specific cell populations and ensuring high levels of transduction in interconnected brain areas following local administration. This new LV and delivery strategy should be of greater therapeutic benefit and opens up new possibilities for the preclinical development of gene therapy for neurodegenerative diseases.

Development, physiological functions, and pathologies of the brain depend on tight interactions between neurons and different types of glial cells, such as astrocytes, microglia, oligodendrocytes, and oligodendrocyte precursor cells. Assessing the relative contribution of different glial cell types is required for the full understanding of brain function and dysfunction. Over the recent years, several technological breakthroughs were achieved, allowing "glio-scientists" to address new challenging biological questions. These technical developments make it possible to study the roles of specific cell types with medium or high-content workflows and perform fine analysis of their mutual interactions in a preserved environment. This review illustrates the potency of several cutting-edge experimental approaches (advanced cell cultures, induced pluripotent stem cell (iPSC)-derived human glial cells, viral vectors, in situ glia imaging, opto- and chemogenetic approaches, and high-content molecular analysis) to unravel the role of glial cells in specific brain functions or diseases. It also illustrates the translation of some techniques to the clinics, to monitor glial cells in patients, through specific brain imaging methods. The advantages, pitfalls, and future developments are discussed for each technique, and selected examples are provided to illustrate how specific "gliobiological" questions can now be tackled., (© 2020 Wiley Periodicals, Inc.)

We describe a rare case of pedunculated endobronchial hemangioma observed in a 60-years-old patient complaining of chronic productive cough and accessional dyspnea which had been progressively worsening over 20 years. The lesion was first noticed at fiberoptic bronchoscopy; then computed tomography scan was performed and integrated with tridimensional reconstruction techniques. Pathology showed the picture of a vascular neoplasm, compatible with capillary hemangioma. The lesion was submitted to laser-assisted endoscopic removal in order to relieve the obstruction, leading to remission of symptoms.

The case of a 43-year-old woman with intralobar pulmonary sequestration, Pryce type one, is presented. The medical history was characterised by recurrent bronchopneumonia, productive cough with purulent sputum and hemoptysis in the last three years. Diagnosis was made by CT angiography: multiplanar, maximum intensity projection and volume rendering reconstructions were visualised. A volume reduction of middle and lower lobe with multiple cyst-like bronchiectasis was detected and no evident relationship with tracheobronchial tree was pointed out. Reconstructions aimed at evaluating bronchial structures demonstrated no patency of middle and lower lobar bronchi. The study carried out after contrast medium infusion in arterial phase showed a vascular disorder characterised by an accessory arterial branch arising from the upper portion of thoracic aorta which, after moving caudally to pulmonary hilus with a tortuous course, supplied the atelectatic parenchyma. No anomalous venous drainage was detected. The patient underwent surgery with resection of two pulmonary lobes. CT compares favourably with other alternative imaging technique for pulmonary sequestration as multiplanar reconstructions allow not only the detection of supplying vessel, but also the accurate description of heterogeneous characteristics of the mass and adjacent structures. Finally an imaging-based diagnostic algorhithm is proposed.

We report eight cases of death or decerebration occurred during major cardiothoracic surgery. The technical aspects that contributed to the failure are analyzed on the basis of the Italian Criminal Code (art. 43). When this type of event occurs, a case review frequently identifies failure to take action or imprudent behavior. The informed consent regulation currently in force may be helpful in identifying and following the most prudent conduct for the patient. The "death conference" should be a formalized and binding instrument for hospital accreditation.

234 patients with lung cancer and operated in Thoracic and Cardiovascular Surgery Department of Careggi Hospital in Florence have been evaluated in order to examine surgical staging accuracy in comparison with pathological staging. There is a statistically significative difference between surgical and pathological staging as a datum point. Surgeon is inclined to over-estimate the lymph-nodes involvement and the primitive tumor extension. It is important to bear in mind this bent whenever decisions of surgical strategy have to be taken.

The follow-up of 240 N2 lung cancer cases operated in Thoracic and Cardiovascular Surgery Department of Florence is examined. The analysis is performed in compliance with global survival, "T", histology, therapeutic choices. Global survival is 81% after 6 months, 60% after 1 year, 37% after 2 years, 26% after 3 years, 23% after 4 years, 23% after 5 years. Significative difference on survival does not exist between principal histologic types (squamous, adenocarcinoma, adenosquamous). Raising the "T" survival decreases, but only for adenocarcinoma. Different therapeutic options (only surgery, surgery+radiotherapy, surgery+chemotherapy, surgery+radio and chemotherapy) do not influence the survival in a way statistically significative. From the literature, any certainty about radiotherapy and chemotherapy associated to surgery for N2 lung cancer treatment does not exist at the moment. Thus radical surgery is essential.

We examined in this perspective study 30 patients (15 males and 15 females) who undergone classic thoracotomy for lung or mediastinal diseases, without postoperative complications. Intensity and different qualitative components of postoperative pain have been evaluated in the 1st, 3rd, 5th, the day after last pleural drainage removal and 2 months after the operation. All patients answered a schedule adherent to Questionario Italiano del Dolore (QUID). Pain became less intense chiefly with drainage removal with the same time trend for both sex (parallel curves). Pain is probably stronger in women. From a qualitative point of view, the sensorial component is the same in both sexes. Affective and evaluative component is greater in women: in other words, women realize more than men the painful stimulus and are more troubled. A computerized analysis of answers to a questionnaire like QUID or, better, its evolution, may be helpful for a more effective pharmacological choice between pure analgesics, sedative analgesics and ataractic drugs.

In order to evaluate the reliability of thoraco-mediastinal CT in the preoperative evaluation of primary lung cancer, regarding "N" (lymph nodes) parameter, we compared CT data with those obtained at histopathology of mediastinal lymph nodes. We re-examined 130 patients who had undergone lobectomy or pneumonectomy combined with mediastinal node dissection. CT criterion of neoplastic nodal involvement is morphological, based on size of the node as related to its location. CT is very sensitive in evaluating both normal and pathological nodes but not likewise specific; in fact, it does not allow differential diagnosis between neoplastic and phlogistic causes. This limitation must be kept in mind in the preoperative evaluation of the "N" parameter. Moreover, CT findings of mediastinal involvement on the opposite side (N3) must be confirmed with mediastinoscopy or CT-guided biopsy before ruling radical surgery out.