IRF1 Mediates Growth Arrest and the Induction of a Secretory Phenotype in Alveolar Epithelial Cells in Response to Inflammatory Cytokines IFNγ/TNFα.

In COVID-19, cytokine release syndrome can cause severe lung tissue damage leading to acute respiratory distress syndrome (ARDS). Here, we address the effects of IFNγ, TNFα, IL-1ß and IL-6 on the growth arrest of alveolar A549 cells, focusing on the role of the IFN regulatory factor 1 (IRF1) transcription factor. The efficacy of JAK1/2 inhibitor baricitinib has also been tested. A549 WT and IRF1 KO cells were exposed to cytokines for up to 72 h. Cell proliferation and death were evaluated with the resazurin assay, analysis of cell cycle and cycle-regulator proteins, LDH release and Annexin-V positivity; the induction of senescence and senescence-associated secretory phenotype (SASP) was evaluated through ß-galactosidase staining and the quantitation of secreted inflammatory mediators. While IL-1 and IL-6 proved ineffective, IFNγ plus TNFα caused a proliferative arrest in A549 WT cells with alterations in cell morphology, along with the acquisition of a secretory phenotype. These effects were STAT and IRF1-dependent since they were prevented by baricitinib and much less evident in IRF1 KO than in WT cells. In alveolar cells, STATs/IRF1 axis is required for cytokine-induced proliferative arrest and the induction of a secretory phenotype. Hence, baricitininb is a promising therapeutic strategy for the attenuation of senescence-associated inflammation.

Iron Bioavailability in the Extracellular Environment Is More Relevant Than the Intracellular One in Viability and Gene Expression: A Lesson from Oligodendroglioma Cells.

Oligodendroglioma (OG) is a brain tumor that contributes to <1% of brain tumor diagnoses in the pediatric population. Unfortunately, pediatric OG remains without definitive molecular characteristics to aid in diagnosis, and little is known about the tumor microenvironment. Tumor cells' metabolism and proliferation rate are generally higher than those of healthy cells, so their iron demand is also significantly higher. This consideration underlines the great importance of iron for tumor development and progression. In this context, this study aims to evaluate the effect of iron in a cellular in vitro model of human oligodendroglioma brain tumor. Cell morphology, the effect of siderotic medium on cell growth, iron uptake, and the expression of iron-metabolism-related genes were evaluated via optic microscopy, ICP-MS, confocal microscopy, and real-time PCR, respectively. This study underlines the great importance of iron for tumor development and progression and also the possibility of reducing the available iron concentration to determine an antiproliferative effect on OG. Therefore, every attempt can be promising to defeat OG for which there are currently no long-term curative therapies.

Cytokine-Induced iNOS in A549 Alveolar Epithelial Cells: A Potential Role in COVID-19 Lung Pathology.

BACKGROUND: In COVID-19, an uncontrolled inflammatory response might worsen lung damage, leading to acute respiratory distress syndrome (ARDS). Recent evidence points to the induction of inducible nitric oxide synthase (NOS2/iNOS) as a component of inflammatory response since NOS2 is upregulated in critical COVID-19 patients. Here, we explore the mechanisms underlying the modulation of iNOS expression in human alveolar cells. METHODS: A549 WT and IRF1 KO cells were exposed to a conditioned medium of macrophages treated with SARS-CoV-2 spike S1. Additionally, the effect of IFNγ, IL-1ß, IL-6, and TNFα, either alone or combined, was addressed. iNOS expression was assessed with RT-qPCR and Western blot. The effect of baricitinib and CAPE, inhibitors of JAK/STAT and NF-kB, respectively, was also investigated. RESULTS: Treatment with a conditioned medium caused a marked induction of iNOS in A549 WT and a weak stimulation in IRF1 KO. IFNγ induced NOS2 and synergistically cooperated with IL-1ß and TNFα. The inhibitory pattern of baricitinb and CAPE indicates that cytokines activate both IRF1 and NF-κB through the JAK/STAT1 pathway. CONCLUSIONS: Cytokines secreted by S1-activated macrophages markedly induce iNOS, whose expression is suppressed by baricitinib. Our findings sustain the therapeutic efficacy of this drug in COVID-19 since, besides limiting the cytokine storm, it also prevents NOS2 induction.

Characterization of BoHV-4 ORF45.

Bovine herpesvirus 4 (BoHV-4) is a Gammaherpesvirus belonging to the Rhadinovirus genus. The bovine is BoHV-4's natural host, and the African buffalo is BoHV-4's natural reservoir. In any case, BoHV-4 infection is not associated with a specific disease. Genome structure and genes are well-conserved in Gammaherpesvirus, and the orf 45 gene and its product, ORF45, are one of those. BoHV-4 ORF45 has been suggested to be a tegument protein; however, its structure and function have not yet been experimentally characterized. The present study shows that BoHV-4 ORF45, despite its poor homology with other characterized Rhadinovirus ORF45s, is structurally related to Kaposi's sarcoma-associated herpesvirus (KSHV), is a phosphoprotein, and localizes in the host cell nuclei. Through the generation of an ORF45-null mutant BoHV-4 and its pararevertant, it was possible to demonstrate that ORF45 is essential for BoHV-4 lytic replication and is associated with the viral particles, as for the other characterized Rhadinovirus ORF45s. Finally, the impact of BoHV-4 ORF45 on cellular transcriptome was investigated, an aspect poorly explored or not at all for other Gammaherpesvirus. Many cellular transcriptional pathways were found to be altered, mainly those involving p90 ribosomal S6 kinase (RSK) and signal-regulated kinase (ERK) complex (RSK/ERK). It was concluded that BoHV-4 ORF45 has similar characteristics to those of KSHV ORF45, and its unique and incisive impact on the cell transcriptome paves the way for further investigations.

Notch-Jagged1 signaling and response to bevacizumab therapy in advanced colorectal cancer: A glance to radiomics or back to physiopathology?

Introduction: The Notch intracellular domain (NICD) and its ligands Jagged-1(Jag1), Delta-like ligand (DLL-3) and DLL4 play an important role in neoangiogenesis. Previous studies suggest a correlation between the tissue levels of NICD and response to therapy with bevacizumab in colorectal cancer (CRC). Another marker that may predict outcome in CRC is radiomics of liver metastases. The aim of this study was to investigate the expression of NICD and its ligands and the role of radiomics in the selection of treatment-naive metastatic CRC patients receiving bevacizumab. Methods: Immunohistochemistry (IHC) for NICD, Jag1 and E-cadherin was performed on the tissue microarrays (TMAs) of 111 patients with metastatic CRC treated with bevacizumab and chemotherapy. Both the intensity and the percentage of stained cells were evaluated. The absolute number of CD4+ and CD8+ lymphocytes was counted in three different high-power fields and the mean values obtained were used to determine the CD4/CD8 ratio. The positivity of tumor cells to DLL3 and DLL4 was studied. The microvascular density (MVD) was assessed in fifteen cases by counting the microvessels at 20x magnification and expressed as MVD score. Abdominal CT scans were retrieved and imported into a dedicated workstation for radiomic analysis. Manually drawn regions of interest (ROI) allowed the extraction of radiomic features (RFs) from the tumor. Results: A positive association was found between NICD and Jag1 expression (p < 0.001). Median PFS was significantly shorter in patients whose tumors expressed high NICD and Jag1 (6.43 months vs 11.53 months for negative cases; p = 0.001). Those with an MVD score ≥5 (CD31-high, NICD/Jag1 positive) experienced significantly poorer survival. The radiomic model developed to predict short and long-term survival and PFS yielded a ROC-AUC of 0.709; when integrated with clinical and histopathological data, the integrated model improved the predictive score (ROC-AUC of 0.823). Discussion: These results show that high NICD and Jag1 expression are associated with progressive disease and early disease progression to anti VEGF-based therapy; the preliminary radiomic analyses show that the integration of quantitative information with clinical and histological data display the highest performance in predicting the outcome of CRC patients.

Redox-Neutral Ru(0)-Catalyzed Alkenylation of 2-Carboxaldimine-heterocyclopentadienes.

A new Ru3(CO)12-catalyzed directed alkenylation of 2-carboxaldimine-heterocyclopentadienes has been accomplished. This process allows coupling of furan, pyrrole, indole, and thiophene 2-carboxaldimines with electron-poor alkenes such as acrylates, vinylsulfones, and styrenes. This regio- and chemoselective oxidative C-H coupling does not require the presence of an additional sacrificial oxidant. Density functional theory calculations allowed us to propose a mechanism and unveiled the nature of the H2 acceptor.

Apheresis Platelet Rich-Plasma for Regenerative Medicine: An In Vitro Study on Osteogenic Potential.

Background: Platelet-Rich Plasma (PRP) induces bone regeneration; however, there is low evidence supporting its efficacy in bone healing. The lack of a standardized protocol of administration represents the main obstacle to its use in the clinical routine for bone defects' treatment. The purpose of this study was to characterize PRP and elucidate its osteogenic potential. Methods: Platelet count, fibrinogen levels, and growth factors concentration were measured in PRP obtained by four apheresis procedures. HOB-01-C1, a pre-osteocytic cell line, was used to examine the effects of different PRP dilutions (from 1% to 50%) on cell viability, growth, and differentiation. Gene expression of RUNX2, PHEX, COL1A1, and OCN was also assayed. Results: PRP showed a mean 4.6-fold increase of platelets amount compared to whole blood. Among the 36 proteins evaluated, we found the highest concentrations for PDGF isoforms, EGF, TGF-ß and VEGF-D. PDGF-AA positively correlated with platelet counts. In three of the four tested units, 25% PRP induced a growth rate comparable to the positive control (10% FBS); whereas, for all the tested units, 10% PRP treatment sustained differentiation. Conclusions: This study showed that PRP from apheresis stimulates proliferation and differentiation of pre-osteocyte cells through the release of growth factors from platelets.

Photobiomodulation with a 645 nm Diode Laser of Saos-2 Cells and Platelet-Rich Plasma: The Potential for a New Mechanism of Action.

Objective: The main focus of this in vitro study was to highlight possible differences between outcomes of photobiomodulation performed in the presence or absence of growth factors derived from platelet-rich plasma. Background: Photobiomodulation has garnered increasing attention, thanks to a large number of controlled clinical trials that have proven its efficacy in various oral pathologies. Nevertheless, the mechanism of action is still a matter of debate. Materials and methods: The cell model used was Saos-2ATTC HTB-85, a human osteosarcoma cell line that retains an osteogenic potential matching that of osteoblastic cells. Photobiomodulation was performed with a 645 nm diode laser; we investigated three different fluence values (2, 5, and 10 J/cm2) delivered with 3 different irradiation times (1, 2, and 4 min). The design of the study included a case-control structure. Cell viability was assessed by resazurin reduction assay before laser irradiation. We assessed cell differentiation by Alizarin-red Sigma Aldrich assay 48 h after the last laser irradiation. Results: Results show that the combination of photobiomodulation and platelet-rich plasma can lead to a statistically significant increase in both proliferation and differentiation rates. Conclusions: Only a defined amount of energy, that is, a fluence of 5 J/cm2 delivered in 2 min and of 10 J/cm2 in 4 min, was proven to be the most effective in the presence of platelet-rich plasma to induce cell proliferation and calcium deposition.

Standard treatment-refractory cytomegalovirus encephalitis unmasked by immune reconstitution inflammatory syndrome and successfully treated with virus-specific hyperimmune globulin.

OBJECTIVES: Cytomegalovirus (CMV)-related encephalitis is a rare but potentially life-threatening complication of CMV infection in immunocompromised patients. The high mortality rate is associated with deficient immune system reconstitution after hematopoietic stem cell transplant (HSCT) and poor bioavailability of antiviral drugs in cerebrospinal fluid (CSF). CMV-related central nervous system (CNS) infection may occur with aspecific symptoms, without evidence of either blood viral load or magnetic resonance imaging (MRI) signs of encephalitis. METHODS: Here, we describe a 10-year-old girl who underwent an allogeneic HSCT and subsequently developed CMV encephalitis. Because of the absence of CMV antigen in the blood, the diagnosis of encephalitis was proposed only after a delay, following the onset of immune reconstitution inflammatory syndrome (IRIS). Two months of combined dual antiviral therapy with ganciclovir and foscarnet proved ineffective against CMV and caused significant bone marrow and renal toxicity. To avoid further toxicity, the girl was given daily treatment with CMV-hyperimmune globulins alone. RESULTS: After three weeks, the CSF viral load dropped significantly and was undetectable within three more weeks. In the meantime, the renal impairment resolved, and there was a complete bone marrow recovery. CONCLUSION: We suggest that this patient succeeded in achieving CMV CSF clearance with high dose of CMV-hyperimmune globulin, given alone, because of the ability of immunoglobulins to penetrate the blood-brain barrier (BBB).

Clinicopathological Bird's-Eye View of Left Atrial Myocardial Fibrosis in 121 Patients With Persistent Atrial Fibrillation: Developing Architecture and Main Cellular Players.

BACKGROUND: Scientific research on atrial fibrosis in atrial fibrillation (AF) has mainly focused on quantitative or molecular features. The purpose of this study was to perform a clinicoarchitectural/structural investigation of fibrosis to provide one key to understanding the electrophysiological/clinical aspects of AF. METHODS: We characterized the fibrosis (amount, architecture, cellular components, and ultrastructure) in left atrial biopsies from 121 patients with persistent/long-lasting persistent AF (group 1; 59 males; 60±11 years; 91 mitral disease-related AF, 30 nonmitral disease-related AF) and from 39 patients in sinus rhythm with mitral valve regurgitation (group 2; 32 males; 59±12 years). Ten autopsy hearts served as controls. RESULTS: Qualitatively, the fibrosis exhibited the same characteristics in all cases and displayed particular architectural scenarios (which we arbitrarily subdivided into 4 stages) ranging from isolated foci to confluent sclerotic areas. The percentage of fibrosis was larger and at a more advanced stage in group 1 versus group 2 and, within group 1, in patients with rheumatic disease versus nonrheumatic cases. In patients with AF with mitral disease and no rheumatic disease, the percentage of fibrosis and the fibrosis stages correlated with both left atrial volume index and AF duration. The fibrotic areas mainly consisted of type I collagen with only a minor cellular component (especially fibroblasts/myofibroblasts; average value range 69-150 cells/mm2, depending on the areas in AF biopsies). A few fibrocytes-circulating and bone marrow-derived mesenchymal cells-were also detectable. The fibrosis-entrapped cardiomyocytes showed sarcolemmal damage and connexin 43 redistribution/internalization. CONCLUSIONS: Atrial fibrosis is an evolving and inhomogeneous histological/architectural change that progresses through different stages ranging from isolated foci to confluent sclerotic zones which-seemingly-constrain impulse conduction across restricted regions of electrotonically coupled cardiomyocytes. The fibrotic areas mainly consist of type I collagen extracellular matrix and, only to a lesser extent, mesenchymal cells.

Intramolecular Aminoazidation of Unactivated Terminal Alkenes by Palladium-Catalyzed Reactions with Hydrogen Peroxide as the Oxidant.

The palladium-catalyzed aminoazidation of aminoalkenes yielding azidomethyl-substituted nitrogen-containing heterocycles was developed. The procedure requires oxidative conditions and occurs at room temperature in the presence of hydrogen peroxide and NaN3 as the azide source. These conditions provide selective exo-cyclization/azidation of the carbon-carbon double bond, furnishing a versatile approach toward five-, six-, and seven-membered heterocyclic rings.

High levels of Notch intracellular cleaved domain are associated with stemness and reduced bevacizumab efficacy in patients with advanced colon cancer.

δ­like ligand 4 (DLL4)­Notch signaling is associated with tumor resistance to anti­vascular endothelial growth factor (VEGF) therapy. Furthermore, Notch signaling is critical for the maintenance of colon cancer stem cells (CSCs), which are relevant in drug resistance and tumor angiogenesis. CD44 is a transmembrane glycoprotein and is considered a putative marker of CSCs. To assess the association of Notch intracellular cleaved domain (NICD), DLL4 and CD44 expression with the efficacy of anti­angiogenic drugs, a series of samples derived from patients with advanced colon cancer enrolled in prospective clinical trials were analyzed. Histological samples from 51 primary tumors that originated from patients treated with bevacizumab­based first­line chemotherapy were analyzed by immunohistochemistry for NICD, DLL4 and CD44 expression, and CD31 for microvessel count. The expression levels of genes relevant for angiogenesis [angiopoietin (ANGPT)1, ANGPT2, fibroblast growth factor (FGF)1, FGF2, epidermal growth factor, placental growth factor, VEGFA and DLL4] were detected by reverse transcription­quantitative PCR using RNA extracted from the frozen tissues of four tumors with low and four tumors with high NICD expression. Strong NICD levels were observed in 12/51 (24%) of the patients, whereas 16/51 (31%) of the colon cancer subjects exhibited high CD44 expression. Strong CD44 staining was associated with high NICD levels compared with the CD44 expression levels noted in samples with low NICD levels (67 vs. 20%, P=0.005). No association was observed with regards to the expression levels of NICD, CD44 and the other aforementioned biomarkers. High expression levels of NICD and CD44 predicted reduced progression­free survival (P<0.001) and overall survival (P=0.002). No significant differences in the expression of angiogenesis­related genes were detected between low and high NICD­expressing tumors. In conclusion, NICD and CD44 tissue levels exhibited an association and may be related to a reduced survival rate in patients with advanced colon cancer treated with bevacizumab.

Effects of zoledronic acid and dexamethasone on early phases of socket healing after tooth extraction in rats: A preliminary macroscopic and microscopic quantitative study

Background: The exact pathogenesis of medication-related osteonecrosis of the jaw (MRONJ) is still unknown. The aim of this paper was to investigate the effects of zoledronic acid and dexamethasone on the early phases of socket healing in rats subjected to tooth extractions. Material and Methods: Thirty male Sprague-Dawley rats were divided into 2 groups: pharmacologically treated group (T, n = 20) and non-pharmacologically treated group (C, n=10). T group rats received 0.1 mg/Kg of zoledronic acid (ZOL) and 1 mg/Kg of dexamethasone (DEX) three times a week for 10 consecutive weeks. C group rats were infused with vehicle. After 9 weeks from the first infusion, first maxillary molars were extracted in each of the rats. Quantitative macroscopic and microscopic analysis was performed to evaluate socket healing 8 days after extraction. Results: Pharmacologically treated rats showed significant inhibition of bone remodeling. Connective tissue/al-eolar bone ratio, osteoclast number and woven bone deposition were significantly reduced in group T compared to group C. Conversely, the proportion of necrotic bone was higher in group T compared to group C (0.8% and 0.3%, respectively. P = 0.031). ZOL plus DEX do not cause gross effects on socket healing at a macroscopic level. Conclusions: Our findings confirmed that exposure to ZOL plus DEX impairs alveolar wound repair. Inhibition of osteoclastic resorption of socket walls after tooth extraction and the inability to dispose of the necrotic bone may be considered the initial steps of MRONJ onset

No disponible

Monocyte-predominant engraftment, cytokine levels and early transplant-related complications in pediatric hematopoietic stem cell recipients.

Myeloablative conditioning is a well-established procedure that precedes hematopoietic stem cell transplantation (HSCT), particularly in pediatric patients. In the period directly following transplantation, several factors may contribute to complications that lead to the activation or damage of endothelial cells, involved in the pathogenesis of vascular endothelial syndromes (VES). However, to date, sufficiently specific and sensitive diagnostic markers for the various forms of VES have not been identified. This was a retrospective single-center study of patients who underwent allogeneic HSCT. For this cohort of patients, parameters including type of engraftment, donor characteristics, and cytokine production were measured and correlated with a high prevalence of short-term complications after HSCT. The aim of this study was to identify specific parameters useful for improving diagnostics and predicting adverse effects in VES. We confirmed that monocyte-predominant engraftment was related to a higher risk for an early transplant-related complication termed sinusoidal obstruction syndrome (SOS). The increased production of specific cytokines, in particular RANTES, represents a marker associated with prevalent engraftment. In addition, patients undergoing prophylaxis with defibrotide had "classical" engraftment, a common cytokine profile and a lower incidence of life-threatening transplant-related complications. The beneficial effect of defibrotide might be a starting point for developing selective prophylaxis for patients with monocyte engraftment to prevent severe early transplant-related complications.

Stanozolol promotes osteogenic gene expression and apposition of bone mineral in vitro

Abstract Stanozolol (ST) is a synthetic androgen with high anabolic potential. Although it is known that androgens play a positive role in bone metabolism, ST action on bone cells has not been sufficiently tested to support its clinical use for bone augmentation procedures. Objective: This study aimed to assess the effects of ST on osteogenic activity and gene expression in SaOS-2 cells. Material and Methods: SaOS-2 deposition of mineralizing matrix in response to increasing doses of ST (0-1000 nM) was evaluated through Alizarin Red S and Calcein Green staining techniques at 6, 12 and 24 days. Gene expression of runt-related transcription factor 2 (RUNX2), vitamin D receptor (VDR), osteopontin (SPP1) and osteonectin (ON) was analyzed by RT-PCR. Results: ST significantly influenced SaOS-2 osteogenic activity: stainings showed the presence of rounded calcified nodules, which increased both in number and in size over time and depending on ST dose. RT-PCR highlighted ST modulation of genes related to osteogenic differentiation. Conclusions: This study provided encouraging results, showing ST promoted the osteogenic commitment of SaOS-2 cells. Further studies are required to validate these data in primary osteoblasts and to investigate ST molecular pathway of action.

Stanozolol promotes osteogenic gene expression and apposition of bone mineral in vitro.

Stanozolol (ST) is a synthetic androgen with high anabolic potential. Although it is known that androgens play a positive role in bone metabolism, ST action on bone cells has not been sufficiently tested to support its clinical use for bone augmentation procedures. OBJECTIVE: This study aimed to assess the effects of ST on osteogenic activity and gene expression in SaOS-2 cells. MATERIAL AND METHODS: SaOS-2 deposition of mineralizing matrix in response to increasing doses of ST (0-1000 nM) was evaluated through Alizarin Red S and Calcein Green staining techniques at 6, 12 and 24 days. Gene expression of runt-related transcription factor 2 (RUNX2), vitamin D receptor (VDR), osteopontin (SPP1) and osteonectin (ON) was analyzed by RT-PCR. RESULTS: ST significantly influenced SaOS-2 osteogenic activity: stainings showed the presence of rounded calcified nodules, which increased both in number and in size over time and depending on ST dose. RT-PCR highlighted ST modulation of genes related to osteogenic differentiation. CONCLUSIONS: This study provided encouraging results, showing ST promoted the osteogenic commitment of SaOS-2 cells. Further studies are required to validate these data in primary osteoblasts and to investigate ST molecular pathway of action.

Benchmarking Heart Rate Variability to Overcome Sex-Related Bias.

Since the seminal studies by Sayers (Ergonomics 16:17-32, 1973) and Akselrod et al. (Science 213:220-222, 1981) a few decades ago, it became clear that beat-by-beat oscillations in RR interval length (i.e. heart-rate variability [HRV]) contain information on underlying neural-control mechanisms based on the instantaneous balance between parasympathetic and sympathetic innervation. Over the years, the number of studies addressing HRV has increased markedly and now outnumbers 23,000. Despite such a large interest, there is still a continuing debate about interpretation of indices produced by computer analysis of HRV.The main part of studies relies on spectral techniques to extract parameters that are linked to hidden information. The general idea is that these proxies of autonomic regulation can be useful to clinical applications in various conditions in which autonomic dysregulation may play a role. There are, however, serious shortcomings related to algorithms, interpretation, and the hidden value of individual indices. In particular, it appears that specific training is necessary to interpret the hidden informational value of HRV. This technical complexity represents a severe barrier to large-scale clinical applications. Moreover, important differences in HRV separate the sexes, and age plays an additional confounding role.We present here a preliminary application of a novel unitary index of RR variability (Autonomic Nervous System Index of cardiac regulation) capable of providing information on the performance of autonomic regulation using a percentile rank position as projected on a large benchmark population. A summary of the underlying sympatho-vagal model is also presented.

Can the use of a single integrated unitary autonomic index provide early clues for eventual eligibility for olympic games?

PURPOSE: Optimal autonomic regulation and stress resilience might be considered critical elements of athletic performance. We hypothesize that a novel unitary autonomic index for sports (ANSIs), together with a somatic stress related symptom score (4SQ) might help characterize athletes who were eventually selected for the Rio 2016 Olympic Games Italian team (Rio +). METHODS: In this retrospective study we examined 778 athletes (age 24.4 ± 6.7 yrs) who underwent a planned yearly pre-participation screening. All athletes underwent clinical, autonomic and exercise ECG evaluation. The combination of vagal and sympathetic indices from RR variability into ANSIs was performed by radar plot and percent ranking of index variables. We assessed (Rio +) versus (Rio -) athletes also after subdivision into three sport intensity groups (low, mid and high intensity). RESULTS: Overall there were no significant differences between (Rio +) and (Rio -) athletes when considering individual spectral derived variables. Conversely, the unitary Index ANSIs was significantly higher in (Rio +) compared to (Rio -) athletes (respectively 54.5 ± 29.5 and 47.9 ± 28.4 p = 0.014). This difference was particularly evident (p = 0.017) in the group of athletes characterized by both high static and dynamic components. 4SQ was smaller in the (Rio +) group, particularly in the groups of athletes characterized by both low-medium static and dynamic components. CONCLUSIONS: ANSIs, a proxy of integrated cardiac autonomic regulation and simple assessment of resilience to stress, may differentiate Italian athletes who were eventually selected for participation in the 2016 Rio Olympic Games from those who were not, suggesting the possibility of a "winning functional phenotype".

Heart rate variability to monitor performance in elite athletes: Criticalities and avoidable pitfalls.

BACKGROUND: Spectral analysis of Heart Rate Variability (HRV) is a simple, non-invasive technique that is widely used in sport to assess sympatho-vagal regulation of the heart. Its employment is increasing partly due to the rising usage of wearable devices. However data acquisition using these devices may be suboptimal because they cannot discriminate between sinus and non-sinus beats and do not record any data regarding respiratory frequency. This information is mandatory for a correct clinical interpretation. METHODS: This study involved 974 elite athletes, all of them underwent a complete autonomic assessment, by way of Autoregressive HRV analysis. RESULTS: In 91 subjects (9% of the total population) we observed criticalities of either cardiac rhythm or respiration. Through perusal of one-lead ECG analysis we observed that 77 subjects had atrial or ventricular ectopy, i.e. conditions which impair stationarity and sinus rhythm. Running anyway autonomic nervous system analysis in this population, we observed that RR variance and raw values of LF and HF regions are significantly higher in arrhythmic subjects. In addition 14 subjects had slow (about 6 breath/min, 0.1Hz) respiration. This condition clouds the separation between LF from HF spectral regions of RR interval variability, respectively markers of the prevalent sympathetic and vagal modulation of SA node and of their synergistic interaction. CONCLUSIONS: Caution must be payed when assessing HRV with non-ECG wearable devices. Recording ECG signal and ensuring that respiratory rate is higher than 10 breath/min are both prerequisites for a more reliable analysis of HRV particularly in athletes.

May a unitary autonomic index help assess autonomic cardiac regulation in elite athletes? Preliminary observations on the national Italian Olympic committee team.

BACKGROUND: Long term endurance training, as occurring in elite athletes, is associated to cardiac neural remodeling in favor of cardioprotective vagal mechanisms, resulting in resting bradycardia and augmented contribution of cardiac parasympathetic nerve activity. Autonomic assessment can be performed by way of heart rate variability. This technique however provides multiple indices, and there is not yet complete agreement on their specific significance. Purpose of the study was to assess whether a rank transformation and radar plot could provide a unitary autonomic index, capable to show a correlation between intensity of individual work and quality of autonomic regulation. METHODS: We studied 711 (23.6±6.2 years) elite athletes that took part in the selection procedure for the 2016 Rio Olympic Games for the National Italian Olympic Committee (CONI). Indices from Heart Rate Variability HRV obtained at rest, during standing up and during recovery from an exercise test were used to compute a percent ranked unitary autonomic index for sport (ANSIs), taken as proxy of quality of autonomic regulation. RESULTS: Within the observed wide range of energy expenditure, the unitary autonomic index ANSIs appears significantly correlated to individual and discipline specific training workloads (r=0.25, P<0.001 and r=0.78, P<0.001, respectively), correcting for possible age and gender bias. ANSIs also positively correlates to lipid profile. CONCLUSIONS: Estimated intensity of physical activity correlates with quality of cardiac autonomic regulation, as expressed by a novel unitary index of cardiac autonomic regulation. ANSIs could provide a novel and convenient approach to individual autonomic evaluation in athletes.