IRCCS San Raffaele Scientific Institute Showcase

Supplementary Fig 1. Workflow illustrating tumors distribution according to dermoscopic, confocal and histologic diagnoses. Supplementary Fig 2. A) Dermoscopy showing brown pseudonetwork with defined borders, interpreted as solar lentigo. B-C) Reflectance confocal microscopy (RCM) revealing LM features, specifically: B) RCM at epidermal layer showing perifollicular and interfollicular dendritic cells. C) RCM at dermal-epidermal layer showing medusa-head like structure. Supplementary Fig 3. Double negative LM case: A) Dermoscopy showing multiple grey dots. B) RCM at epidermal layer showing honeycombed pattern with focal mottled pigmentation. C) RCM at dermal-epidermal layer showing abundant melanophages. Supplementary Fig 4. Double negative BCC case: A) Dermoscopy showing brown-red background. B) RCM at epidermal layer showing honeycombed pattern with focal mottled pigmentation. C) RCM at dermal-epidermal layer showing aspecific structures.

BACKGROUND: Major noncardiac surgery is associated with high rates of postoperative myocardial injury and other complications. Remote ischemic preconditioning (RIPC) was reported to decrease these complication rates. However, such supportive evidence lacks robustness. METHODS: In a multinational, double-blind trial, we randomly assigned adult high-risk patients undergoing noncardiac surgical procedures to receive RIPC or sham RIPC after the induction of general anesthesia and before surgery. RIPC involved three 5-minute ischemic cycles, each followed by 5 minutes of reperfusion, using a blood pressure cuff inflated to 200 mm Hg. The primary end point was the rate of myocardial injury, defined by an increase in postoperative troponin levels above the highest 99th percentile of reference values. Secondary outcomes included myocardial infarction, stroke, acute kidney injury, need for intensive care unit, length of hospital stay, and 30-day all-cause mortality. RESULTS: We recruited 1213 patients in 25 hospitals and 8 countries. We randomly assigned 599 patients to RIPC and 614 to sham RIPC. The most frequent surgical procedures were abdominal or intrathoracic surgeries (406 patients [33.6%]). RIPC was applied to the upper limb in 1014 patients (84.8%) and to the lower limb in 182 patients (15.2%). Postoperative myocardial injury occurred in 215 of 566 patients (38.0%) in the RIPC group and in 223 of 596 patients (37.4%) in the sham RIPC group (relative risk, 1.02 [95% CI, 0.88–1.18; P=0.84). There were no significant differences in the rate of any secondary outcomes. We observed 11 episodes of limb petechiae (10 [1.7%] in the RIPC group versus one [0.2%] in the sham RIPC group) and 34 (6.0%) hospital readmissions in the RIPC group versus 20 (3.5%) in the sham RIPC group. CONCLUSIONS: Among adult patients undergoing noncardiac surgery, RIPC did not reduce myocardial injury or other postoperative complications. REGISTRATION: URL: https://www.clinicaltrials.gov; Unique identifier: NCT02427867.

BACKGROUND Patients undergoing cardiac surgery often receive red-cell transfusions, along with the associated risks and costs. Early intraoperative normovolemic hemodilution (i.e., acute normovolemic hemodilution [ANH]) is a blood-conservation technique that entails autologous blood collection before initiation of cardiopulmonary bypass and reinfusion of the collected blood after bypass weaning. More data are needed on whether ANH reduces the number of patients receiving allogeneic red-cell transfusion. METHODS In a multinational, single-blind trial, we randomly assigned adults from 32 centers and 11 countries who were undergoing cardiac surgery with cardiopulmonary bypass to receive ANH (withdrawal of ≥650 ml of whole blood with crystalloids replacement if needed) or usual care. The primary outcome was the transfusion of at least one unit of allogeneic red cells during the hospital stay. Secondary outcomes were death from any cause within 30 days after surgery or during the hospitalization for surgery, bleeding complications, ischemic complications, and acute kidney injury. RESULTS A total of 2010 patients underwent randomization; 1010 were assigned to ANH and 1000 to usual care. Among patients with available data, 274 of 1005 (27.3%) in the ANH group and 291 of 997 (29.2%) in the usual-care group received at least one allogeneic red-cell transfusion (relative risk, 0.93; 95% confidence interval, 0.81 to 1.07; P = 0.34). Surgery for postoperative bleeding was performed in 38 of 1004 patients (3.8%) in the ANH group and 26 of 995 patients (2.6%) in the usual-care group. Death within 30 days or during hospitalization occurred in 14 of 1008 patients (1.4%) in the ANH group and 16 of 997 patients (1.6%) in the usual care group. Safety outcomes were similar in the two groups. CONCLUSIONS Among adults undergoing cardiac surgery, ANH did not reduce the number of patients receiving allogeneic red-cell transfusion. (Funded by the Italian Ministry of Health; ANH ClinicalTrials.gov number, NCT03913481.)

Repository of data related to the publications of UO2 (San Raffaele Hospital) pertaining to the PNRR-MAD-2022-12375859 project "Inflammation and depression: a multidisciplinary approach to dissect pathogenetic mechanisms in the onset, comorbidity, and treatment response".

Post-COVID-19 syndrome has unveiled intricate connections between inflammation, depressive psychopathology, and cognitive impairment. This study investigates these relationships in 101 COVID-19 survivors, focusing on sex-specific variations. Utilizing path modelling techniques, we analysed the interplay of one-month 48-biomarker inflammatory panel, on three-month depressive symptoms and cognitive performance. The findings indicate that cognitive impairment is influenced by both inflammation and depression in the overall cohort. However, sex-specific differences emerged prominently. In females, residual dysregulated immune-inflammatory response significantly affects cognitive functioning also showing a trend of association with depressive burden thus suggesting that a mixed anti- and pro-inflammatory profile could foster these outcomes. Conversely, in males, inflammation was inversely associated with depression severity, with protective effects from regulatory mediators (IL-2, IL-4, IL-6, IL-15, LIF, TNF-α, β-NGF) on depression. Cognitive impairment in males was primarily influenced by depression, not inflammation. These results highlight distinct sex-specific pathways in immune and inflammatory responses post-COVID-19, potentially shaped by endocrine mechanisms. The findings emphasize the persistent impact of inflammation on the brain and underscore the need for sex-tailored therapeutic strategies to mitigate the long-term neuropsychiatric burden of COVID-19.

Multiple sclerosis (MS) is a chronic, autoimmune, inflammatory disease of the central nervous system (CNS) characterized by demyelination and neurodegeneration. Among the various pathogenic mechanisms, oxidative stress plays a critical role in driving both inflammation and neuronal damage, and emerging evidence suggests that mitochondrial dysfunction may significantly contribute to disease progression. Whole mtDNA was sequenced from blood-derived DNA using long-range PCR and Illumina® Nextera XT kit. Somatic mutations were defined based on heteroplasmy levels between 1–5%. Linear regression models were used to assess the association between age and mutation rate. We observed a significant age-dependent increase in low-frequency nonsynonymous mtDNA mutations in MS. Analyses stratified by disease course revealed this effect was driven by PPMS patients, while no association was seen in RRMS, suggesting course-specific mitochondrial trajectories. Furthermore, fast-progressing patients showed a positive linear relationship between age and mtDNA mutations rate, while slow-progressing ones showed a negative trend. These findings support the existence of a differential age-related accumulation of somatic mtDNA mutations across MS courses and underline the importance of mitochondrial genome instability in disease progression.

Introduction: Depressive disorders are a leading cause of global disease burden, particularly with the challenge of treatment-resistant depression (TRD). Research points to a complex bidirectional relationship between cardiovascular (CV) risk factors and TRD, with CV risk negatively impacting brain structure and potentially influencing antidepressant resistance. Moreover, the association between depression and the genetic vulnerability to cardiovascular disease suggests a shared pathophysiological process between the two. This study investigates the mediating role of brain structural alterations in the relationship between CV and cerebrovascular (CeV) risk and treatment resistance in depression. Methods: We assessed 165 inpatients with Major depressive disorder. Each patient's CV risk was assessed via the QRISK 3 calculator. For a subset of patients, CV and CeV disease polygenic risk scores (PRS) were obtained. All patients underwent a 3 T MRI scan, and white matter hyperintensities estimates and indicators of brain trophic state were obtained. Results: Both CV risk and CV disease PRSs are associated with treatment resistance status, white matter hyperintensities, and indicators of brain atrophy. Mediation analyses suggested that CV-induced brain alterations might underlie the relation between CV genetic and phenotypic risk and antidepressant treatment resistance. Conclusion: These results underscore the need to explore cardiovascular risk management as part of treatment strategies for depression, pointing toward a shared pathophysiological process linking heart and brain health in treatment-resistant depression.

Gene-environment correlations (rGE) may drive the clinical heterogeneity of major depressive disorder (MDD) through their effects on brain structure. However, previous literature focused on isolated components of these interplays. Here, we jointly investigate how rGE shape neurobiological profiles in MDD, and whether rGE-driven brain signatures can disentangle depression subtypes. In 5951 MDD patients with genetic, trauma-related and neuroimaging data from the UK Biobank, cross-validated sparse canonical correlation analysis was employed to assess multivariate associations between polygenic scores (PGSs) for mental health conditions and adverse childhood experiences (ACEs). Linear regressions tested the impact of the shared PGSs–ACEs dimensions on gray matter (GM) measures. Consensus clustering was applied to the neuroimaging features significantly associated with PGSs or ACEs to identify latent biotypes of MDD, and the emerged clusters were compared for depressive symptomatology and organic comorbidities. We found a significant canonical correlation between PGSs and ACEs (r = 0.11, p < 0.001). The most contributing PGSs were schizophrenia, attention deficit-hyperactivity disorder, autism (positive weights) and education (negative weight). Canonical variates of PGSs and ACEs associated with reduced GM in frontal, temporal, cingulate, parietal and subcortical regions (b = [-0.041; -0.021], pFDR<0.05). Such rGE-sensitive brain regions underpinned two clusters of patients, with one showing higher genetic/environmental risk, reduced GM integrity, and a worse clinical profile, including atypical symptoms, anhedonia, lethargy, sleep alterations and diabetes comorbidity. These findings indicate that rGE-driven neurobiological signatures contribute to the clinical heterogeneity of depression, supporting biologically informed subtyping in MDD.

Targeted genomic integration of gene-sized cassettes into hematopoietic stem and progenitor cells (HSPCs) for genetic disease treatment is constrained by the low efficiency of homology-directed repair (HDR) and frequent unintended genetic changes at the editing site. To overcome these challenges, we introduce Selection by Means of Artificial Transactivators (SMArT), which transiently implements AND reporter gates to achieve templated integration of a functional cassette at the target site. HDR-edited HSPCs were enriched to very high purity through transient selector expression, whereas cells carrying undesired and potentially genotoxic on-target edits were preferentially depleted. Xenotransplantation of SMArT-enriched HSPCs in immunodeficient mice resulted in fully HDR-edited human grafts with the selector no longer detectable. SMArT strategies were implemented through clinically compliant manufacturing and selectors. They support both safe harbor integration and gene correction, can preserve physiological transcriptional regulation, and are portable across loci also with polyfunctional editors. Overall, SMArT strategies may broaden therapeutic applicability of gene-sized editing while reducing its genotoxic burden.

Objectives: The role of neutrophils in systemic sclerosis (SSc) remains incompletely understood. Methods: To address this, blood samples from 39 SSc patients, 39 healthy controls, and 22 systemic lupus erythematosus (SLE) patients were analyzed. Results: In SSc, neutrophils exhibited substantial activation, evidenced by granule mobilization, elevated plasma levels of Neutrophil Extracellular Trap (NET) byproducts, and upregulated TIE2 expression. In parallel, they underwent metabolic reprogramming, characterized by increased autophagy, likely to support the heightened energy demands of activation. By contrast, neutrophils from SLE patients displayed minimal autophagy, lacked TIE2 expression, and shifted toward low-density granulocytes. Neutrophil reprogramming in SSc correlated with plasma levels of HMGB1+ EVs. Mechanistically, EVs purified from the plasma of patients with SSc adhered to neutrophils when injected in immunodeficient NSG mice, inducing autophagy, TIE2 expression, and promoting lung inflammation and fibrosis. These effects were abrogated by HMGB1 inhibitors and required the HMGB1 receptor, RAGE. Recombinant HMGB1 recapitulated EV-induced effects, while neutrophil targeting by liposome-encapsulated clodronate prevented them. Conclusions: In summary, neutrophils in SSc exhibit a dual phenotype of autophagy and activation driven by HMGB1+ EVs, representing a pathogenic mechanism with therapeutic potential in SSc. This mechanism operates similarly in male and female mice and is sufficient to induce neutrophil-driven lung injury in vivo.