A Narrative Review on Pharmacogenomics in Psychiatry: Scientific Definitions, Principles, and Practical Resources.

PURPOSE/BACKGROUND: Pharmacogenetics (PGx) studies the genetic factors underlying interindividual variability in drug response. Only a few countries around the world are already using PGx testing in psychiatric clinical practice, whereas others are still far from adopting it. The main barrier to the clinical adoption of PGx testing seems to be the limited knowledge among psychiatrists regarding the clinical relevance of specific genetic variants to personalize therapies and the accessibility of PGx data. This review aims at further highlighting the importance of PGx-driven clinical decision making for psychotropic medications and raising psychiatrists' awareness of the value of PGx testing in psychiatry. METHODS/PROCEDURES: We summarize the genes for which substantial evidence exists about the clinical utility of integrating their PGx testing in psychiatry. Specifically, we systematically describe the functional role of clinically relevant allelic variants, their frequency across different ethnic groups, and how they contribute to classify patients in relation to their capability in metabolizing psychotropic drugs. FINDINGS/RESULTS: Briefly, clinical guidelines recommend considering PGx testing of the cytochrome class 2 C9 (CYP2C9), C19 (CYP2C19), and D6 (CYP2D6) genes and the human leukocyte antigen (HLA)-A and -B genes for several psychotropic drugs. IMPLICATIONS/CONCLUSIONS: Extensive studies have been carried out to provide a solid rationale for the inclusion of PGx testing in psychiatry. Comprehensive clinical guidelines are readily accessible to support health care providers in tailoring the prescription of psychotropic drugs based on patient's genotype information. This approach presents a tangible opportunity to significantly improve individual responses to psychiatric medications.

Deficits in emotion recognition and processing in children with high callous-unemotional traits: the role of the MAOA gene.

Children with high Callous-Unemotional (CU) traits show deficits in recognizing and processing facial expressions. Alterations in emotion recognition have been linked to a higher synaptic concentration of monoaminergic neurotransmitters. The current study investigated the relationship between the MAOA-Low-activity alleles and the ability to recognize and process facial expressions in 97 male children (8-12 years old) diagnosed with disruptive behavior disorder. Participants completed a computerized emotion-recognition task while an eye-tracking system recorded the number (Fixation Count, FC) and length (Fixation Duration, FD) of fixations to the eye region of the emotional stimuli. Children with high CU traits exhibited lower scores in recognition of sadness and anger, and lower FC and FD for sadness and fear than children with low CU traits. Children carrying the MAOA-Low-activity alleles displayed lower FD for sadness, and FD and FC for fear than those carrying the MAOA-High-activity alleles. These genetic effects appeared even stronger in children with CU traits. Moderation analysis revealed that CU traits were associated with lower FC and FD for fear, and lower FD for sadness, probably due to the MAOA-Low-activity alleles. Our findings, although to be replicated, suggest MAOA-Low-activity alleles as potential genetic biomarkers to identify CU children in need of training focused on emotion processing.

Transgenic pigs expressing near infrared fluorescent protein-A novel tool for noninvasive imaging of islet xenotransplants.

BACKGROUND: Islet xenotransplantation is a promising concept for beta-cell replacement therapy. Reporter genes for noninvasive monitoring of islet engraftment, graft mass changes, long-term survival, and graft failure support the optimization of transplantation strategies. Near-infrared fluorescent protein (iRFP) is ideal for fluorescence imaging (FI) in tissue, but also for multispectral optoacoustic tomography (MSOT) with an even higher imaging depth. Therefore, we generated reporter pigs ubiquitously expressing iRFP. METHODS: CAG-iRPF720 transgenic reporter pigs were generated by somatic cell nuclear transfer from FACS-selected stable transfected donor cells. Neonatal pig islets (NPIs) were transplanted into streptozotocin-diabetic immunodeficient NOD-scid IL2Rgnull (NSG) mice. FI and MSOT were performed to visualize different numbers of NPIs and to evaluate associations between signal intensity and glycemia. MSOT was also tested in a large animal model. RESULTS: CAG-iRFP transgenic NPIs were functionally equivalent with wild-type NPIs. Four weeks after transplantation under the kidney capsule, FI revealed a twofold higher signal for 4000-NPI compared to 1000-NPI grafts. Ten weeks after transplantation, the fluorescence intensity of the 4000-NPI graft was inversely correlated with glycemia. After intramuscular transplantation into diabetic NSG mice, MSOT revealed clear dose-dependent signals for grafts of 750, 1500, and 3000 NPIs. Dose-dependent MSOT signals were also revealed in a pig model, with stronger signals after subcutaneous (depth ∼6 mm) than after submuscular (depth ∼15 mm) placement of the NPIs. CONCLUSIONS: Islets from CAG-iRFP transgenic pigs are fully functional and accessible to long-term monitoring by state-of-the-art imaging modalities. The novel reporter pigs will support the development and preclinical testing of novel matrices and engraftment strategies for porcine xeno-islets.

Strategies to Improve the Safety of iPSC-Derived ß Cells for ß Cell Replacement in Diabetes.

Allogeneic islet transplantation allows for the re-establishment of glycemic control with the possibility of insulin independence, but is severely limited by the scarcity of organ donors. However, a new source of insulin-producing cells could enable the widespread use of cell therapy for diabetes treatment. Recent breakthroughs in stem cell biology, particularly pluripotent stem cell (PSC) techniques, have highlighted the therapeutic potential of stem cells in regenerative medicine. An understanding of the stages that regulate ß cell development has led to the establishment of protocols for PSC differentiation into ß cells, and PSC-derived ß cells are appearing in the first pioneering clinical trials. However, the safety of the final product prior to implantation remains crucial. Although PSC differentiate into functional ß cells in vitro, not all cells complete differentiation, and a fraction remain undifferentiated and at risk of teratoma formation upon transplantation. A single case of stem cell-derived tumors may set the field back years. Thus, this review discusses four approaches to increase the safety of PSC-derived ß cells: reprogramming of somatic cells into induced PSC, selection of pure differentiated pancreatic cells, depletion of contaminant PSC in the final cell product, and control or destruction of tumorigenic cells with engineered suicide genes.

A Comprehensive Characterization of Stemness in Cell Lines and Primary Cells of Pancreatic Ductal Adenocarcinoma.

The aim of this study is to provide a comprehensive characterization of stemness in pancreatic ductal adenocarcinoma (PDAC) cell lines. Seventeen cell lines were evaluated for the expression of cancer stem cell (CSC) markers. The two putative pancreatic CSC phenotypes were expressed heterogeneously ranging from 0 to 99.35% (median 3.46) for ESA+CD24+CD44+ and 0 to 1.94% (median 0.13) for CXCR4+CD133+. Cell lines were classified according to ESA+CD24+CD44+ expression as: Low-Stemness (LS; <5%, n = 9, median 0.31%); Medium-Stemness (MS; 6−20%, n = 4, median 12.4%); and High-Stemness (HS; >20%, n = 4, median 95.8%) cell lines. Higher degree of stemness was associated with in vivo tumorigenicity but not with in vitro growth kinetics, clonogenicity, and chemo-resistance. A wide characterization (chemokine receptors, factors involved in pancreatic organogenesis, markers of epithelial−mesenchymal transition, and secretome) revealed that the degree of stemness was associated with KRT19 and NKX2.2 mRNA expression, with CD49a and CA19.9/Tie2 protein expression, and with the secretion of VEGF, IL-7, IL-12p70, IL-6, CCL3, IL-10, and CXCL9. The expression of stem cell markers was also evaluated on primary tumor cells from 55 PDAC patients who underwent pancreatectomy with radical intent, revealing that CXCR4+/CD133+ and CD24+ cells, but not ESA+CD24+CD44+, are independent predictors of mortality.

Treating iPSC-Derived ß Cells with an Anti-CD30 Antibody-Drug Conjugate Eliminates the Risk of Teratoma Development upon Transplantation.

Insulin-producing cells derived from induced pluripotent stem cells (iPSCs) are promising candidates for ß cell replacement in type 1 diabetes. However, the risk of teratoma formation due to residual undifferentiated iPSCs contaminating the differentiated cells is still a critical concern for clinical application. Here, we hypothesized that pretreatment of iPSC-derived insulin-producing cells with an anti-CD30 antibody−drug conjugate could prevent in vivo teratoma formation by selectively killing residual undifferentiated cells. CD30 is expressed in all human iPSCs clones tested by flow cytometry (n = 7) but not in iPSC-derived ß cells (ißs). Concordantly, anti-CD30 treatment in vitro for 24 h induced a dose-dependent cell death (up to 90%) in human iPSCs while it did not kill ißs nor had an impact on iß identity and function, including capacity to secrete insulin in response to stimuli. In a model of teratoma assay associated with iß transplantation, the pretreatment of cells with anti-CD30 for 24 h before the implantation into NOD-SCID mice completely eliminated teratoma development (0/10 vs. 8/8, p < 0.01). These findings suggest that short-term in vitro treatment with clinical-grade anti-CD30, targeting residual undifferentiated cells, eliminates the tumorigenicity of iPSC-derived ß cells, potentially providing enhanced safety for iPSC-based ß cell replacement therapy in clinical scenarios.

Faecal microbiota transplantation halts progression of human new-onset type 1 diabetes in a randomised controlled trial.

OBJECTIVE: Type 1 diabetes (T1D) is characterised by islet autoimmunity and beta cell destruction. A gut microbiota-immunological interplay is involved in the pathophysiology of T1D. We studied microbiota-mediated effects on disease progression in patients with type 1 diabetes using faecal microbiota transplantation (FMT). DESIGN: Patients with recent-onset (<6 weeks) T1D (18-30 years of age) were randomised into two groups to receive three autologous or allogenic (healthy donor) FMTs over a period of 4 months. Our primary endpoint was preservation of stimulated C peptide release assessed by mixed-meal tests during 12 months. Secondary outcome parameters were changes in glycaemic control, fasting plasma metabolites, T cell autoimmunity, small intestinal gene expression profile and intestinal microbiota composition. RESULTS: Stimulated C peptide levels were significantly preserved in the autologous FMT group (n=10 subjects) compared with healthy donor FMT group (n=10 subjects) at 12 months. Small intestinal Prevotella was inversely related to residual beta cell function (r=-0.55, p=0.02), whereas plasma metabolites 1-arachidonoyl-GPC and 1-myristoyl-2-arachidonoyl-GPC levels linearly correlated with residual beta cell preservation (rho=0.56, p=0.01 and rho=0.46, p=0.042, respectively). Finally, baseline CD4 +CXCR3+T cell counts, levels of small intestinal Desulfovibrio piger and CCL22 and CCL5 gene expression in duodenal biopsies predicted preserved beta cell function following FMT irrespective of donor characteristics. CONCLUSION: FMT halts decline in endogenous insulin production in recently diagnosed patients with T1D in 12 months after disease onset. Several microbiota-derived plasma metabolites and bacterial strains were linked to preserved residual beta cell function. This study provides insight into the role of the intestinal gut microbiome in T1D. TRIAL REGISTRATION NUMBER: NTR3697.

Transcriptional dynamics of induced pluripotent stem cell differentiation into ß cells reveals full endodermal commitment and homology with human islets.

BACKGROUND AIMS: Induced pluripotent stem cells (iPSCs) have the capacity to generate ß cells in vitro, but the differentiation is incomplete and generates a variable percentage of off-target cells. Single-cell RNA sequencing offers the possibility of characterizing the transcriptional dynamics throughout differentiation and determining the identity of the final differentiation product. METHODS: Single-cell transcriptomics data were obtained from four stages across differentiation of iPSCs into ß cells and from human donor islets. RESULTS: Clustering analysis revealed that iPSCs undertake a full endoderm commitment, and the obtained endocrine pancreatic cells have high homology with mature islets. The iPSC-derived ß cells were devoid of pluripotent residual cells, and the differentiation was pancreas-specific, as it did not generate ectodermal or mesodermal cells. Pseudotime trajectory identified a dichotomic endocrine/non-endocrine cell fate and distinct subgroups in the endocrine branch. CONCLUSIONS: Future efforts to produce ß cells from iPSCs must aim not only to improve the resulting endocrine cell but also to avoid differentiation into non-pancreatic endoderm cells.

CCL2/MCP-1 and CXCL12/SDF-1 blockade by L-aptamers improve pancreatic islet engraftment and survival in mouse.

The blockade of pro-inflammatory mediators is a successful approach to improve the engraftment after islet transplantation. L-aptamers are chemically synthesized, nonimmunogenic bio-stable oligonucleotides that bind and inhibit target molecules conceptually similar to antibodies. We aimed to evaluate if blockade-aptamer-based inhibitors of C-C Motif Chemokine Ligand 2/monocyte chemoattractant protein-1 (CCL2/MCP-1) and C-X-C Motif Chemokine Ligand 12/stromal cell-derived factor-1 (CXCL12/SDF-1) are able to favor islet survival in mouse models for islet transplantation and for type 1 diabetes. We evaluated the efficacy of the CCL2-specific mNOX-E36 and the CXCL12-specific NOX-A12 on islet survival in a syngeneic mouse model of intraportal islet transplantation and in a multiple low doses of streptozotocin (MLD-STZ) diabetes induction model. Moreover, we characterized intrahepatic infiltrated leukocytes by flow cytometry before and 3 days after islet infusion in presence or absence of these inhibitors. The administration for 14 days of mNOX-E36 and NOX-A12 significantly improved islet engraftment, either compound alone or in combination. Intrahepatic islet transplantation recruited CD45+ leucocytes and more specifically CD45+/CD11b+ mono/macrophages; mNOX-E36 and NOX-A12 treatments significantly decreased the recruitment of inflammatory monocytes, CD11b+ /Ly6Chigh /CCR2+ and CD11b+ /Ly6Chigh /CXCR4+ cells, respectively. Additionally, both L-aptamers significantly attenuated diabetes progression in the MLD-STZ model. In conclusion, CCL2/MCP-1 and CXCL12/SDF-1 blockade by L-aptamers is an efficient strategy to improve islet engraftment and survival.

Core Set of Patient-reported Outcomes in Pancreatic Cancer (COPRAC): An International Delphi Study Among Patients and Health Care Providers.

OBJECTIVE: To establish an international core set of patient-reported outcomes (PROs) selected by both patients and healthcare providers (HCPs) from the United States (US), Europe, and Asia. SUMMARY BACKGROUND DATA: PROs are increasingly recognized in pancreatic cancer studies. There is no consensus on which of the many available PROs are most important. METHODS: A multicenter Delphi study among patients with pancreatic cancer (curative- and palliative-setting) and HCPs in 6 pancreatic centers in the US (Baltimore, Boston), Europe (Amsterdam, Verona), and Asia (Mumbai, Seoul) was performed. In round 1, participants rated the importance of 56 PROs on a 1 to 9 Likert scale. PROs rated as very important (scores 7-9) by the majority (≥80%) of curative- and/or palliative-patients as well as HCPs were included in the core set. PROs not fulfilling these criteria were presented again in round 2, together with feedback on individual and group ratings. Remaining PROs were ranked based on the importance ratings. RESULTS: In total 731 patients and HCPs were invited, 501 completed round 1, and 420 completed both rounds. This included 204 patients in curative-setting, 74 patients in palliative-setting, and 142 HCPs. After 2 rounds, 8 PROs were included in the core set: general quality of life, general health, physical ability, ability to work/do usual activities, fear of recurrence, satisfaction with services/care organization, abdominal complaints, and relationship with partner/family. CONCLUSIONS: This international Delphi study among patients and HCPs established a core set of PROs in pancreatic cancer, which should facilitate the design of future pancreatic cancer trials and outcomes research.

Selective local irradiation improves islet engraftment and survival in intra-bone marrow islet transplantation.

BACKGROUND: Bone marrow (BM) is as an alternative site for islet transplantation, but it is not an immunoprotected microenvironment and allogeneic islets are rejected. However, the BM, for its structure and anatomic position, offers the possibility to modulate microenvironment by local interventions. We here investigate whether local irradiation is able to improve islet engraftment and prevent rejection in BM in the absence of immunosuppression. METHODS: A model of BM local irradiation was set up. Islets were transplanted in syngeneic and fully major histocompatibility complex-mismatched recipients in control and locally irradiated BM; gain of normoglycemia and time to rejection were evaluated. RESULTS: BM local irradiation proved to be a selective and safe procedure. Syngeneic islet transplantation into locally irradiated BM had better outcome compared with not irradiated recipients in terms of capacity to gain normoglycemia (100% versus 56% in irradiated versus not irradiated mice). In the allogenic setting, glycemia was significantly lower in the first days after transplantation in the group of irradiated mice and local irradiation also delayed time to graft rejection (from 4 ± 1 days for not irradiated to 11 ± 1 days for locally irradiated mice). DISCUSSION: These data indicate that local immunosuppression by irradiation before islet transplantation in BM favors islet engraftment and delays time to rejection.

Brain Hemodynamic Intermediate Phenotype Links Vitamin B12 to Cognitive Profile of Healthy and Mild Cognitive Impaired Subjects.

Vitamin B12, folate, and homocysteine are implicated in pivotal neurodegenerative mechanisms and partake in elders' mental decline. Findings on the association between vitamin-related biochemistry and cognitive abilities suggest that the structural and functional properties of the brain may represent an intermediate biomarker linking vitamin concentrations to cognition. Despite this, no previous study directly investigated whether vitamin B12, folate, and homocysteine levels are sufficient to explain individual neuropsychological profiles or, alternatively, whether the activity of brain regions modulated by these compounds better predicts cognition in elders. Here, we measured the relationship between vitamin blood concentrations, scores at seventeen neuropsychological tests, and brain activity of sixty-five elders spanning from normal to Mild Cognitive Impairment. We then evaluated whether task-related brain responses represent an intermediate phenotype, providing a better prediction of subjects' neuropsychological scores, as compared to the one obtained considering blood biochemistry only. We found that the hemodynamic activity of the right dorsal anterior cingulate cortex was positively associated (p value < 0.05 cluster corrected) with vitamin B12 concentrations, suggesting that elders with higher B12 levels had a more pronounced recruitment of this salience network region. Crucially, the activity of this area significantly predicted subjects' visual search and attention abilities (p value = 0.0023), whereas B12 levels per se failed to do so. Our results demonstrate that the relationship between blood biochemistry and elders' cognitive abilities is revealed when brain activity is included into the equation, thus highlighting the role of brain imaging as intermediate phenotype.

Gene expression analysis of embryonic pancreas development master regulators and terminal cell fate markers in resected pancreatic cancer: A correlation with clinical outcome.

BACKGROUND: Despite the recent introduction of new drugs and the development of innovative multi-target treatments, the prognosis of pancreatic ductal adenocarcinoma (PDAC) remains very poor. Even when PDAC is resectable, the rate of local or widespread disease recurrence remains particularly high. Currently, reliable prognostic biomarkers of recurrence are lacking. We decided to explore the potential usefulness of pancreatic developmental regulators as biomarkers of PDAC relapse. METHODS: We analyzed by quantitative real-time PCR the mRNA of selected factors involved either in pancreatic organogenesis (ISL1, NEUROD1, NGN3, NKX2.2, NKX6.1, PAX4, PAX6, PDX1 and PTF1α) or associated with terminally committed pancreatic cells (CHGA, CHGB, GAD2, GCG, HNF6α, INS, KRT19, SYP) in 17 PDAC cell lines and in frozen tumor samples from 41 PDAC patients. RESULTS: High baseline levels of the ISL1, KRT19, PAX6 and PDX1 mRNAs in PDAC cell lines, were risk factors for time-dependent xenograft appearance after subcutaneous injection in CD1-Nude mice. Consistently, in human PDAC samples, high levels of KRT19 mRNA were associated with reduced overall survival and earlier recurrence. Higher levels of PDX1 or PAX6 mRNAs were instead associated with a higher frequency of local recurrence. CONCLUSIONS: Our findings suggest that selected factors associated with pancreas development or its terminal differentiation might be implicated in mechanisms of PDAC progression and/or metastatic spread and that the measurement of their mRNA in tumors might be potentially used to improve patient prognostic stratification and prediction of the relapse site.

Establishment, characterization and long-term culture of human endocrine pancreas-derived microvascular endothelial cells.

BACKGROUND: In vitro primary cultures of microvascular endothelial cells from endocrine pancreas are difficult to obtain, but can be a very helpful tool for studies of islet biology, transplantation and regenerative medicine. METHODS: We applied a protocol recently described for the isolation and culture of brain microvascular endothelial cells (EC) on human pancreatic islets. EC obtained were characterized in terms of morphological (light and transmission electron microscopy), phenotypical (by immunofluorescence and flow cytometry) and functional (cord formation assay and protein secretion by multiplex bead-based assay) characteristics. RESULTS: EC were obtained from 25% of islet preparations processed. Two primary endothelial cell lines showed high proliferative potential and were deeply characterized: they presented endothelial cell morphology and expressed CD31, CD49a, CD49e, CD34, von Willebrand Factor (vWF), Vascular Endothelial CAdherin (VE-CAD), Tyrosine Kinase with Ig and EGF Homology Domains-2 (TIE2), Vascular Endothelial Growth Factor Receptor 1 (VEGFR1), Ulex lectin and the endothelium endocrine-specific marker nephrin. Besides, they were able to form cordons in vitro and secreted factors involved in the process of angiogenesis such as Vascular Endothelial Growth Factor (VEGF), Monocyte Chemotactic Protein 1 (MCP-1), interleukin (IL)-8 and Melanoma Growth Stimulatory Activity Alpha (GROα). These cell lines were termed Human Islet Microvascular Endothelial Cells (HIMEC). DISCUSSION: This study establishes a simple and effective strategy for isolation and long-term culture of EC derived from human pancreatic islet. HIMEC in culture preserve phenotype and functional properties and are, therefore, a useful tool for future experiments of in vitro pancreas modelling, co-transplantation with pancreatic islets, re-vascularization of scaffold or matrix for regenerative medicine purposes.

Immunological Issues After Stem Cell-Based ß Cell Replacement.

PURPOSE OF REVIEW: Islet and pancreas transplantation prove that ß cell replacement can cure the glycemic derangements in type 1 diabetes (T1D). Induced pluripotent stem cells (iPSCs) can differentiate into functional insulin-producing cells, able to restore normoglycemia in diabetic animal models. iPSCs in particular can be derived from the somatic cells of a person with T1D. This review aims to clarify if it is possible to transplant autologous iPSC-derived ß cells without immunosuppression or which are the alternative approaches. RECENT FINDINGS: Several lines of evidence show that autologous iPSC and their derivatives can be immune rejected, and this immunogenicity depends on the reprogramming, the type of cells generated, the transplantation site, and the genetic/epigenetic modifications induced by reprogramming and differentiation. Besides, cell replacement in T1D should keep in consideration also the possibility of autoimmune reaction against autologous stem cell-derived ß cells. Autologous iPSC-derived ß cells could be immunogenic upon transplantation, eliciting both auto and allogeneic immune response. A strategy to protect cells from immune rejection is still needed. This strategy should be efficacious in protecting the grafted cells, but also avoid toxicity and the risk of tumor formation.

Implications of noncoding regulatory functions in the development of insulinomas.

Insulinomas are rare neuroendocrine tumors arising from pancreatic ß cells, characterized by aberrant proliferation and altered insulin secretion, leading to glucose homeostasis failure. With the aim of uncovering the role of noncoding regulatory regions and their aberrations in the development of these tumors, we coupled epigenetic and transcriptome profiling with whole-genome sequencing. As a result, we unraveled somatic mutations associated with changes in regulatory functions. Critically, these regions impact insulin secretion, tumor development, and epigenetic modifying genes, including polycomb complex components. Chromatin remodeling is apparent in insulinoma-selective domains shared across patients, containing a specific set of regulatory sequences dominated by the SOX17 binding motif. Moreover, many of these regions are H3K27me3 repressed in ß cells, suggesting that tumoral transition involves derepression of polycomb-targeted domains. Our work provides a compendium of aberrant cis-regulatory elements affecting the function and fate of ß cells in their progression to insulinomas and a framework to identify coding and noncoding driver mutations.

Interpreting the gene expression microarray results: a user-based experience.

In recent years many tools have been developed to cope with the interpretation of gene expression results from microarray experiments. The effectiveness of these tools largely depends on their ease of use by biomedical researchers. Tools based on effective computational methods, indeed, cannot be fully exploited by users if they are not supported by an intuitive interface, a large set of utilities and effective outputs. In this paper, 10 tools for the interpretation of gene expression microarray results have been tested on 11 microarray datasets and evaluated according to eight assessment criteria: 1. interface design and usability, 2. easiness of input submission, 3. effectiveness of output representation and 4. of the downloaded outputs, 5. possibility to submit multiple gene IDs, 6. sources of information, 7. provision of different statistical tests and 8. of multiple test correction methods. Strengths and weaknesses of each tool are highlighted to: a. provide useful tips to users dealing with the biological interpretation of microarray results; b. draw the attention of software developers on the usability of their tools.

Interactions between immune, stress-related hormonal and cardiovascular systems following strenuous physical exercise.

Physical exercise represents a eustress condition that promotes rapid coordinated adjustments in the immune, stress-related hormonal and cardiovascular systems, for maintaining homeostasis in response to increased metabolic demands. Compared to the tight multisystem coordination during exercise, evidence of between-systems cross talk in the early post exercise is still lacking. This study was aimed at identifying possible interactions between multiple systems following strenuous physical exercise (Ironman race) performed by twenty well-trained triathletes. Cardiac hemodynamics, left ventricle systolic and diastolic function and heart rate variability were measured along with plasma concentrations of immune messengers (cytokines and C-reactive protein) and stress-related hormones (catecholamines and cortisol) both 24h before and within 20 min after the race. Observed changes in antiinflammatory pathways, stress-related hormones and cardiovascular function were in line with previous findings; moreover, correlating parameters' changes (post versus pre-race) highlighted a dependence of cardiovascular function on the post-race biohumoral milieu: in particular, individual post-race variations of heart rate and diastolic function were strongly correlated with individual variations of anti-inflammatory cytokines, while individual baroreflex sensitivity changes were linked to IL-8 increase. Multiple correlations between anti-inflammatory cytokines and catecholamines were also found according with the autonomic regulation of immune function. Observed post-race cytokine and hormone levels were presumptively representative of the increases reached at the effort end while the cardiovascular parameters after the race were measured during the cardiovascular recovery; thus, results suggest that sustained strenuous exercise produced a stereotyped cardiovascular early recovery, whose speed could be conditioned by the immune and stress-related hormonal milieu.

Does denosumab exert a protective effect against COVID-19? Results of a large cohort study.

Introduction: Denosumab is a monoclonal antibody blocking the receptor activator of nuclear factor kappa-B/receptor activator of nuclear factor kappa-B ligand (RANK/RANKL) pathway, thus inhibiting osteoclastogenesis. Since RANK and RANKL are also involved in the immune system activation, denosumab might interfere with the response against infections. Our study aimed to explore the relationship between denosumab treatment and coronavirus disease 2019 (COVID-19). Design and methods: The occurrence and severity of COVID-19 were recorded in consecutive patients referred to the Endocrinology Department of Papa Giovanni XXIII Hospital, Bergamo, from 1 January 2020 to 1 January 2021. Patients treated with denosumab were compared to outpatient controls. Patients' features were summarized by descriptive statistics. Multivariate logistic regression assessed the relationship between denosumab and COVID-19, adjusting for potential confounders. Subgroup analyses according to age, sex, body mass index (BMI), smoking status, and vitamin D levels were performed. Results: The final population included 331 patients treated with denosumab and 357 controls. COVID-19 incidence was lower in the denosumab group (7.6% vs. 14.6%, p = 0.004). COVID-19 severity was similar in both groups. Multiple logistic regression confirmed an association between denosumab and a reduced occurrence of symptomatic COVID-19 [odds ratio (OR) 0.46, 95% CI 0.21-0.98, p = 0.049]. Subgroup analyses suggested a potential protective effect of denosumab in patients over 75 years (OR 0.12, 95% CI 0.02-0.6, p = 0.011), with a significant interaction between denosumab and age categories (p = 0.047). Conclusion: Our study confirms that denosumab may be safely continued in COVID-19 patients. RANK/RANKL inhibition seems associated with a reduced incidence of symptomatic COVID-19, particularly among the elderly.

Post hoc analysis of a randomized, double-blind, prospective trial evaluating a CXCR1/2 inhibitor in new-onset type 1 diabetes: endo-metabolic features at baseline identify a subgroup of responders.

Aim: In a recent randomized, multicenter trial (NCT02814838) a short-term anti-inflammatory treatment with ladarixin (LDX; an inhibitor of the CXCR1/2 chemokine receptors) did not show benefit on preserving residual beta cell function in new-onset type 1 diabetes. We present a post hoc analysis of trial patients in the predefined subgroup analysis developed according to baseline daily insulin requirement (DIR) tertiles. Method: A double-blind, randomized (2:1), placebo-controlled study was conducted in 45 men and 31 women (aged 18-46 years) within 100 days of the first insulin administration. Patients received LDX (400 mg twice daily) for three cycles of 14 days on/14 days off, or placebo. The primary endpoint was the area under the curve for C-peptide [AUC (0-120 min)] in response to a 2-h mixed meal tolerance test (MMTT) at week 13 ± 1. Seventy-five patients completed the week 13 MMTT and were divided into three groups according to the DIR tertiles: lower, ≤ 0.23U/kg/die (n = 25); middle, 0.24-0.40 U/kg/die (n = 24); upper, ≥ 0.41 U/kg/die (n = 26). Results: When considering the patients in the upper tertile (HIGH-DIR), C-peptide AUC (0-120 min) at 13 weeks was higher in the LDX group (n = 16) than in the placebo (n = 10) group [difference: 0.72 nmol/L (95% CI 0.9-1.34), p = 0.027]. This difference reduced over time (0.71 nmol/L at 26 weeks, p = 0.04; 0.42 nmol/L at 52 weeks, p = 0.29), while it has never been significant at any time in patients in the lower and/or middle tertile (LOW-DIR). We characterized at baseline the HIGH-DIR and found that endo-metabolic (HOMA-B, adiponectin, and glucagon-to-C-peptide ratio) and immunologic (chemokine (C-C motif) ligand 2 (CCL2)/monocyte chemoattractant protein 1 (MCP1) and Vascular Endothelial Growth Factor (VEGF)) features distinguished this group from LOW-DIR. Conclusion: While LDX did not prevent the progressive loss of beta-cell function in the majority of treated subjects, the post hoc analysis suggests that it could work in subjects with HIGH-DIR at baseline. As we found differences in endo-metabolic and immunologic parameters within this subgroup, this generates the hypothesis that the interactions between host factors and drug action can contribute to its efficacy. Further research is needed to evaluate this hypothesis.