The Role of Epigenetic Control of Mitochondrial (Dys)Function in MASLD Onset and Progression.

Metabolic dysfunction-associated steatotic fatty liver disease (MASLD), a novel definition for NAFLD, represents one of the most common causes of liver disease, and its incidence is increasing worldwide. It is characterized by a complex etiopathogenesis in which mitochondrial dysfunction exerts a pivotal role together with alteration of lipid metabolism, inflammation, and oxidative stress. Nutrients and bioactive compounds can influence such mechanisms so that changes in diet and lifestyle are regarded as important treatment strategies. Notably, natural compounds can exert their influence through changes of the epigenetic landscape, overall resulting in rewiring of molecular networks involved in cell and tissue homeostasis. Considering such information, the present review aims at providing evidence of epigenetic modifications occurring at mitochondria in response to natural and bioactive compounds in the context of liver (dys)function. For this purpose, recent studies reporting effects of compounds on mitochondria in the context of NAFLD/MASLD, as well as research showing alteration of DNA methylation and non-coding RNAs-related circuits occurring at liver mitochondria, will be illustrated. Overall, the present review will highlight the importance of understanding the bioactive compounds-dependent epigenetic modulation of mitochondria for improving the knowledge of MASLD and identifying biomarkers to be employed for effective preventative strategies or treatment protocols.

Artificial Intelligence and liver: Opportunities and barriers.

Artificial Intelligence (AI) has recently been shown as an excellent tool for the study of the liver; however, many obstacles still have to be overcome for the digitalization of real-world hepatology. The authors present an overview of the current state of the art on the use of innovative technologies in different areas (big data, translational hepatology, imaging, and transplant setting). In clinical practice, physicians must integrate a vast array of data modalities (medical history, clinical data, laboratory tests, imaging, and pathology slides) to achieve a diagnostic or therapeutic decision. Unfortunately, machine learning and deep learning are still far from really supporting clinicians in real life. In fact, the accuracy of any technological support has no value in medicine without the support of clinicians. To make better use of new technologies, it is essential to improve clinicians' knowledge about them. To this end, the authors propose that collaborative networks for multidisciplinary approaches will improve the rapid implementation of AI systems for developing disease-customized AI-powered clinical decision support tools. The authors also discuss ethical, educational, and legal challenges that must be overcome to build robust bridges and deploy potentially effective AI in real-world clinical settings.

Sarcopenia, a condition shared by various diseases: can we alleviate or delay the progression?

Sarcopenia is a severe condition common to various chronic diseases and it is reckoned as a major health problem. It encompasses many different molecular mechanisms that have been for a while discovered but not definitely clarified. Although sarcopenia is a disability status that leads to serious health consequences, the scarcity of suitable animal models has curtailed research addressing this disorder. Another limitation in the field of clinical investigation of sarcopenic patients is the lack of a generally accepted definition coupled with the difficulty of adopting common diagnostic criteria. In fact, both do not permit to clarify the exact prevalence rate and consequently limit physicians to establish any kind of therapeutical approach or, when possible, to adopt preventive measures. Unfortunately, there is no standardized cure, apart from doing more physical activity and embracing a balanced diet, but newly discovered substances start being considered. In this review, authors try to give an overview addressing principal pathways of sarcopenia and offer critical features of various possible interventions.

Application of machine learning approach in emergency department to support clinical decision making for SARS-CoV-2 infected patients.

To support physicians in clinical decision process on patients affected by Coronavirus Disease 2019 (COVID-19) in areas with a low vaccination rate, we devised and evaluated the performances of several machine learning (ML) classifiers fed with readily available clinical and laboratory data. Our observational retrospective study collected data from a cohort of 779 COVID-19 patients presenting to three hospitals of the Lazio-Abruzzo area (Italy). Based on a different selection of clinical and respiratory (ROX index and PaO2/FiO2 ratio) variables, we devised an AI-driven tool to predict safe discharge from ED, disease severity and mortality during hospitalization. To predict safe discharge our best classifier is an RF integrated with ROX index that reached AUC of 0.96. To predict disease severity the best classifier was an RF integrated with ROX index that reached an AUC of 0.91. For mortality prediction the best classifier was an RF integrated with ROX index, that reached an AUC of 0.91. The results obtained thanks to our algorithms are consistent with the scientific literature an accomplish significant performances to forecast safe discharge from ED and severe clinical course of COVID-19.

Fall of viral and bacterial pneumonia hospitalizations following COVID-19 pandemic mitigation strategies: a central Italian Region retrospective study.

Community-Acquired Pneumonia (CAP) represents one of the first causes of hospitalization and death in the elderly all over the world and weighs heavily on public health system. Since the beginning of the COVID-19 (CoronaVirus Disease-19) pandemic, everybody's behavior was forced to change, as the result of a global lockdown strategy and the obligation of using personal protection equipment (PPE). We aimed to evaluate how the mitigation strategies adopted to fight SARS-CoV-2 (Severe Acute Respiratory Coronavirus Syndrome 2) infection have influenced hospitalizations due to CAP in two different Local Health Boards (LHBs) of central Italy. We considered two main periods of observation: before and after the national start of lockdown, in two Abruzzo's LHBs. We analyzed 19,558 hospital discharge records of bacterial and viral CAP. Excluding SARS-CoV2 infection, a significant decrease in CAP hospitalizations was observed. Through the analysis of Diagnosis Related Group (DRG) values, we highlighted a significant saving of founds for the Regional Health Service. The enactment of social distancing measures to contain COVID-19 spread, brought down admissions for bacterial and viral pneumonia. Our study emphasizes that costs for hospitalizations due to CAP could be drastically reduced by mask wearing and social distancing.

Profiling of cell-free DNA methylation and histone signatures in pediatric NAFLD: A pilot study.

Nonalcoholic fatty liver disease (NAFLD) has become the most common chronic liver disease in children and adolescents, increasing the risk of its progression toward nonalcoholic steatohepatitis (NASH), cirrhosis, and cancer. There is an urgent need for noninvasive early diagnostic and prognostic tools such as epigenetic marks (epimarks), which would replace liver biopsy in the future. We used plasma samples from 67 children with biopsy-proven NAFLD, and as controls we used samples from 20 children negative for steatosis by ultrasound. All patients were genotyped for patatin-like phospholipase domain containing 3 (PNPLA3), transmembrane 6 superfamily member 2 (TM6SF2), membrane bound O-acyltransferase domain containing 7 (MBOAT7), and klotho-ß (KLB) gene variants, and data on anthropometric and biochemical parameters were collected. Furthermore, plasma cell-free DNA (cfDNA) methylation was quantified using a commercially available kit, and ImageStream(X) was used for the detection of free circulating histone complexes and variants. We found a significant enrichment of the levels of histone macroH2A1.2 in the plasma of children with NAFLD compared to controls, and a strong correlation between cfDNA methylation levels and NASH. Receiver operating characteristic curve analysis demonstrated that combination of cfDNA methylation, PNPLA3 rs738409 variant, coupled with either high-density lipoprotein cholesterol or alanine aminotransferase levels can strongly predict the progression of pediatric NAFLD to NASH with area under the curve >0.87. Conclusion: Our pilot study combined epimarks and genetic and metabolic markers for a robust risk assessment of NAFLD development and progression in children, offering a promising noninvasive tool for the consistent diagnosis and prognosis of pediatric NAFLD. Further studies are necessary to identify their pathogenic origin and function.

Alteration of Inflammatory Parameters and Psychological Post-Traumatic Syndrome in Long-COVID Patients.

The aim of our study is to evaluate the correlation between the psychological status of patients recovered from SARS-CoV-2 (Severe Acute Respiratory Syndrome Coronavirus 2) infection (long-COVID patients) and their inflammatory status. Three months after hospital discharge, ninety-three patients were recruited and categorized into two distinct populations: control and long-COVID (COrona VIrus Disease) group. Patients belonging to the control group presented with an entering diagnosis of cardiovascular, metabolic, or respiratory disease and a negative history of SARS-CoV-2 infection, whereas the long-COVID population presented with a severe SARS-CoV-2 infection treated in the sub-intensive Care Unit. Psychological evaluation was performed through the administration of the Symptom Checklist-90 (SCL90) and LDH (Lactate dehydrogenase), ferritin, CRPhs (C-high sensitivity Reactive Protein), NLR (Neutrophil-to-lymphocyte ratio), PLR (Platelet-to-lymphocyte ratio), and SII (systemic immune-inflammation index) were investigated. We highlighted that beyond the first three months after contagion, patients recovered from SARS-CoV-2 infection are characterized by the persistence of a systemic inflammatory state and are at high risk for developing somatization, depression, anxiety, and sleep disturbances. Interestingly, ferritin value was strongly correlated with sleep disorders (p < 0.05). Our study emphasizes how COVID-19 strategies for risk stratification, prognosis, and therapy management of patients should be implemented with a psychological follow-up.

Children and Adolescents Dietary Habits and Lifestyle Changes during COVID-19 Lockdown in Italy.

BACKGROUND: We evaluated the changes in lifestyle during the COVID-19 pandemic lockdown in a sample of children and adolescents in order to assess any increase in risk factors for the onset of cardiovascular diseases in later ages. METHODS: We conducted a cross-sectional study involving 965 parents who completed an online survey about dietary habits and lifestyle during the first lockdown in Italy (from 9 March 2020 to 18 May 2020) and compared their findings with the period before the pandemic. The inclusion criteria were parents (or caregivers) with Italian residency and with children aged between 5 and 18 years. RESULTS: We identified 563 adolescents and 402 children. The mean age was 12.28 years (SD 3.754). The pandemic was associated with an increase in the consumption of high-calorie snack foods. The total amount of food in homes during lockdown compared with before the pandemic increased 50%. Relating to the parent-perceived child weight status, more parents reported obesity in their children after lockdown (+0.6% in the 5-11 age group and +0.2% in the 12-18 age group). We reported a reduction of physical activity, an increase of sedentary lifestyle and sleep habits changes. CONCLUSION: The COVID-19 pandemic was associated with changes in the lifestyles of children and adolescents; this could cause an increase in the incidence of obesity and of cardiovascular and metabolic diseases in adulthood.

Copper-catalyzed dicarbonyl stress in NAFLD mice: protective effects of Oleuropein treatment on liver damage.

BACKGROUND: Nonalcoholic fatty liver disease (NAFLD) or more appropriately, metabolic associated fatty liver disease (MAFLD), is the hepatic manifestation of metabolic syndrome. An imbalance of copper homeostasis has been described in the progression of NAFLD/MAFLD toward NASH/MASH. We were interested in understanding whether the chelating activity of Oleuropein (Ole) was able to improve the copper accumulation and the related pro-oxidant and glycative damage in the liver of mice fed HFD. METHODS: Twelve C57BL/6J mice fed normal diet (ND) or high-fat diet (HFD) for 16 weeks and then thirty two female and male mice fed ND or HFD for 8 weeks adding Ole for the following 8 weeks were studied. RESULTS: Altered expression of copper-trafficking genes and proteins (CTR1, CTR2, ATP7B, COX17, CCS, and ATOX1) induced imbalance of copper homeostasis combined with an increase in dicarbonyl stress in the liver of HFD fed mice. Interestingly enough, glyoxalase system was improved by Ole administration and the Ole related protective effects differ in the two sexes of mice. CONCLUSIONS: Our study highlights the role of the dicarbonyl stress in the pathogenesis of NAFLD and suggests Ole as a natural copper chelator to prevent the liver damage induced by methyglyoxal pathway derangement.

PaO2/FiO2 ratio forecasts COVID-19 patients' outcome regardless of age: a cross-sectional, monocentric study.

We studied the predictive value of the PaO2/FiO2 ratio for classifying COVID-19-positive patients who will develop severe clinical outcomes. One hundred fifty patients were recruited and categorized into two distinct populations ("A" and "B"), according to the indications given by the World Health Organization. Patients belonging the population "A" presented with mild disease not requiring oxygen support, whereas population "B" presented with a severe disease needing oxygen support. The AUC curve of PaO2/FiO2 in the discovery cohort was 0.838 (95% CI 0.771-0.908). The optimal cut-off value for distinguishing population "A" from the "B" one, calculated by Youden's index, with sensitivity of 71.79% and specificity 85.25%, LR+4.866, LR-0.339, was < 274 mmHg. The AUC in the validation cohort of 170 patients overlapped the previous one, i.e., 0.826 (95% CI 0.760-0.891). PaO2/FiO2 ratio < 274 mmHg was a good predictive index test to forecast the development of a severe respiratory failure in SARS-CoV-2-infected patients. Moreover, our work highlights that PaO2/FiO2 ratio, compared to inflammatory scores (hs-CRP, NLR, PLR and LDH) indicated to be useful in clinical managements, results to be the most reliable parameter to identify patients who require closer respiratory monitoring and more aggressive supportive therapies. Clinical trial registration: Prognostic Score in COVID-19, prot. NCT04780373 https://clinicaltrials.gov/ct2/show/NCT04780373 (retrospectively registered).

The application of artificial intelligence in hepatology: A systematic review.

The integration of human and artificial intelligence (AI) in medicine has only recently begun but it has already become obvious that intelligent systems can dramatically improve the management of liver diseases. Big data made it possible to envisage transformative developments of the use of AI for diagnosing, predicting prognosis and treating liver diseases, but there is still a lot of work to do. If we want to achieve the 21st century digital revolution, there is an urgent need for specific national and international rules, and to adhere to bioethical parameters when collecting data. Avoiding misleading results is essential for the effective use of AI. A crucial question is whether it is possible to sustain, technically and morally, the process of integration between man and machine. We present a systematic review on the applications of AI to hepatology, highlighting the current challenges and crucial issues related to the use of such technologies.

Using the Social Robot NAO for Emotional Support to Children at a Pediatric Emergency Department: Randomized Clinical Trial.

BACKGROUND: Social robots (SRs) have been used for improving anxiety in children in stressful clinical situations, such as during painful procedures. However, no studies have yet been performed to assess their effect in children while waiting for emergency room consultations. OBJECTIVE: This study aims to assess the impact of SRs on managing stress in children waiting for an emergency room procedure through the assessment of salivary cortisol levels. METHODS: This was an open randomized clinical trial in children attending a pediatric emergency department. Children accessing the emergency room were randomized to 1 of 3 groups: (1) playing with a NAO SR, (2) playing with a study nurse, or (3) waiting with parents. The salivary cortisol levels of all children were measured through a swab. Salivary cortisol levels before and after the intervention were compared in the 3 groups. We calculated the effect size of our interventions through the Cohen d-based effect size correlation (r). RESULTS: A total of 109 children aged 3-10 years were enrolled in the study, and 94 (86.2%) had complete data for the analyses. Salivary cortisol levels significantly decreased more in the group exposed to robot interaction than in the other two groups (r=0.75). Cortisol levels decreased more in girls (r=0.92) than in boys (r=0.57). CONCLUSIONS: SRs are efficacious in decreasing stress in children accessing the emergency room and may be considered a tool for improving emotional perceptions of children and their families in such a critical setting. TRIAL REGISTRATION: ClinicalTrials.gov NCT04627909; https://clinicaltrials.gov/ct2/show/study/NCT04627909.

It Is High Time Physicians Thought of Natural Products for Alleviating NAFLD. Is There Sufficient Evidence to Use Them?

Non-alcoholic fatty liver disease (NAFLD) is the most common form of liver disease all over the world due to the obesity pandemic; currently, therapeutic options for NAFLD are scarce, except for diet recommendations and physical activity. NAFLD is characterized by excessive accumulation of fat deposits (>5%) in the liver with subsequent inflammation and fibrosis. Studies in the literature show that insulin resistance (IR) may be considered as the key mechanism in the onset and progression of NAFLD. Recently, using natural products as an alternative approach in the treatment of NAFLD has drawn growing attention among physicians. In this review, the authors present the most recent randomized controlled trials (RCTs) and lines of evidence from animal models about the efficacy of nutraceutics in alleviating NAFLD. Among the most studied substances in the literature, the following molecules were chosen because of their presence in the literature of both clinical and preclinical studies: spirulina, oleuropein, garlic, berberine, resveratrol, curcumin, ginseng, glycyrrhizin, coffee, cocoa powder, epigallocatechin-3-gallate, and bromelain.

Focal adhesion kinase inhibitor TAE226 combined with Sorafenib slows down hepatocellular carcinoma by multiple epigenetic effects.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most common and lethal malignant tumours worldwide. Sorafenib (SOR) is one of the most effective single-drug systemic therapy against advanced HCC, but the identification of novel combination regimens for a continued improvement in overall survival is a big challenge. Recent studies highlighted the crucial role of focal adhesion kinase (FAK) in HCC growth. The aim of this study was to investigate the antitumor effects of three different FAK inhibitors (FAKi), alone or in combination with SOR, using in vitro and in vivo models of HCC. METHODS: The effect of PND1186, PF431396, TAE226 on cell viability was compared to SOR. Among them TAE226, emerging as the most effective FAKi, was tested alone or in combination with SOR using 2D/3D human HCC cell line cultures and HCC xenograft murine models. The mechanisms of action were assessed by gene/protein expression and imaging approaches, combined with high-throughput methods. RESULTS: TAE226 was the more effective FAKi to be combined with SOR against HCC. Combined TAE226 and SOR treatment reduced HCC growth both in vitro and in vivo by affecting tumour-promoting gene expression and inducing epigenetic changes via dysregulation of FAK nuclear interactome. We characterized a novel nuclear functional interaction between FAK and the NuRD complex. TAE226-mediated FAK depletion and SOR-promoted MAPK down-modulation caused a decrease in the nuclear amount of HDAC1/2 and a consequent increase of the histone H3 lysine 27 acetylation, thus counteracting histone H3 lysine 27 trimethylation. CONCLUSIONS: Altogether, our findings provide the first evidence that TAE226 combined with SOR efficiently reduces HCC growth in vitro and in vivo. Also, our data highlight that deep analysis of FAK nuclear interactome may lead to the identification of new promising targets for HCC therapy.

Laboratory parameters related to severe disease and death in SARS-CoV-2 pneumonia: Retrospective analysis.

The clinical evolution of coronavirus disease 2019 (COVID-19) is highly variable and hospitalized patients can rapidly develop conditions requiring oxygen support, intensive care unit (ICU) or high dependency unit (HDU) care. Early identification of high-risk patients is mandatory. We retrospectively collected the medical history, symptoms, radiological, and laboratory findings of COVID-19 patients hospitalized between February and April 2020. Laboratory data were collected at the first, last, and middle times of hospitalization. We used arterial oxygen partial pressure and fractional inspired oxygen ratio (P/F) to evaluate respiratory status. Outcomes considered were death and ICU/HDU admission. We used the χ2 or Fisher's exact test to examine differences between categorical variables. Continuous variables were analyzed using the Wilcoxon matched pairs signed-ranks test and Mann-Whitney test sample test. Of 71 patients admitted, 92% had interstitial pneumonia, and 17% an unfavorable outcome. Negative predictors were age, cerebrovascular disease, obesity, and chronic obstructive pulmonary disease. Baseline P/F was strongly associated with all outcomes. Markers linked to immunological dysregulation like elevated neutrophil-to-lymphocyte ratio exhibited prognostic significance over time. A validated prognostic score comprehensive of all these conditions for early staging and management of COVID-19 patients is urgently needed. Further studies are desirable to evaluate whether laboratory tests can target early treatment in high-risk patients.

Age and Interleukin-15 Levels Are Independently Associated With Intima-Media Thickness in Obesity-Related NAFLD Patients.

Common carotid intima-media thickness (IMT) represents a functional and structural marker of early, precocious, and subclinical atherosclerosis, independently from the carotid plaque. Macrophage cells, which have been detected in adipose tissue and atherosclerotic plaques, are regulated by interleukin-15 (IL-15). At the light of the conflicting results concerning the role of IL-15 in atherosclerosis, the aim of the study was to retrospectively evaluate in a population of 80 obese patients, with median age of 46 years (IQR 34-53 years), with a low rate of comorbidities but with non-alcoholic fatty liver disease (NAFLD) or hepatic steatosis (HS), the relationship between IMT and serum concentrations of IL-15. Anthropometric measures, metabolic profile, and serum inflammatory markers, as well as the levels of IL-15, MCP-1, b FGF, and GM-CSF, were analyzed by a bead-based assay. IMT, HS, subcutaneous, and visceral adipose tissues were detected by ultrasonography. The IL-15 levels of the obese patients were increased with respect to those of 44 young healthy subjects, i.e., 2.77 (1.21-4.8) vs. 1.55 (1-2.4) pg/mL (P = 0.002). In the univariate analysis, IL-15 levels were associated to IMT and to those of MCP-1, b FGF, and GM-CSF, without any relation to other inflammatory markers such as CRP and ferritin, except fibrinogen. In the multivariate analysis, after adjusting the HS severity for the extent of visceral adiposity, a dramatic change in prediction of IMT by HS was shown (ß from 0.29 to 0.10, P from 0.008 to 0.37). When the visceral adipose tissue was combined with IL-15, on the one hand, and the well-known coronary artery disease (CAD) risk factors-i.e., age, gender, smoking status, HDL-cholesterol concentrations, triglycerides levels, and HOMA-on the other, only age and IL-15 remained the predictors of IMT (ß = 0.60, P = 0.0001 and ß = 0.25, P = 0.024, respectively). There was no association of IL-15 with various anthropometric parameters nor with body fat distribution and severity of HS, also after adjusting for age. Age is resulted to be the main factor in the prediction of IMT and thus of early atherosclerosis. The prediction of IMT by IL-15 coupled with the lack of prediction by the well-known CAD risks is in agreement with recent data, which emphasizes the main role of the immune system in the onset/worsening of atherosclerosis, even though the role of visceral adiposity should be further deepened. Age and IL-15 levels were both predictors of early atherosclerosis in this population of obese patients with NAFLD, suggesting a possible role of this cytokine in the atherosclerosis process.

Copper concentrations are prevalently associated with antithrombin III, but also with prothrombin time and fibrinogen in patients with liver cirrhosis: A cross-sectional retrospective study.

BACKGROUND: Concerning the link between copper excess and the pathogenesis of chronic liver diseases, its retention is reckoned to develop as a complication of cholestasis. Recently, it has been found that cholestatic liver injury involves largely inflammatory cell-mediated liver cell necrosis, with consequent reduced hepatic mass, more than occurring through direct bile acid-induced apoptosis. On the other hand, interference with protein synthesis could be expected to result, ending in an altered ability of the liver to retain copper. Little is known about the association between serum copper and clotting factors in cirrhotics. We aimed at studying a possible relationship between increased levels of copper and an aspect of the haemostatic process in liver cirrhosis patients, assessing an index of protein synthesis (albumin) and parameters of protein synthesis/coagulation/fibrinolysis, such as prothrombin time (PT), antithrombin (AT) III and fibrinogen. METHODS: Records from 85 patients suffering from liver cirrhosis of various aetiology and different severity were retrospectively examined. Serum concentrations of copper were determined by atomic absorption spectrophotometer. An index of protein synthesis, such as albumin and parameters of both synthesis and coagulation/hypercoagulation such as PT %, AT III%, levels of fibrinogen were taken into account to study possible correlations to serum copper. The severity of cirrhosis was evaluated by the Child-Pugh (C-P) classification. The relationship among variables were studied by linear regression. RESULTS: Copper levels of patients suffering from liver cirrhosis were increased respect to those of controls, 102.7+/-28.7 versus 80.4+/-19.5 mcg/dL, (P = .0009), independently from disease severity, and were positively predicted by PT% (P = 0. 017), fibrinogen (P = 0.007) and AT III% (P = 0.000), at linear regression. Among the previous parameters, to which serum albumin was added, the unique predictor of copper levels was AT III%, at multiple regression (P = 0. 010); AT III% was negatively predicted by the C-P classification (P = 0.000); copper levels, adjusted for C-P classification, were predicted by AT III% (P = 0.020) and fibrinogen concentrations, but not by PT% (P = 0.09). CONCLUSION: The copper concentration is reckoned as responsible for production of the hydroxyl radicals. On the basis that oxidants may enhance the activity of the extrinsic coagulation cascade, ultimately leading to thrombin formation, via their combined effects on stimulation of tissue factor activity and inhibition of fibrinolytic pathways, the positive relationship of copper to coagulation/hypercoagulation parameters (mainly AT III) in our research could find a plausible interpretation.

Liver-spleen axis in nonalcoholic fatty liver disease.

Introduction: NAFLD is often under-diagnosed, even though rates of its co-morbidities such as obesity and type2 diabetes mellitus, prominent statuses of inflammation, are significantly high. The spleen-liver axis is gaining much credit in the last years like other well-known organ axes.Areas covered: PubMed/MEDLINE was searched for relevant articles related to concomitant occurrence of NAFLD and spleen. Areas covered in this review include: (1) updated findings of spleen dimensions at ultrasonography, (2) discussion of current data on pathophysiological connections between obesity-related NAFLD and increased volume of the spleen, and (3) analysis of current immune-mediated mechanisms characterizing the so.called chronic low-grade inflammation leading to insulin resistance.Expert opinion: The advances in explaining mechanisms underlying the spleen involvement in immune regulation, coupled with research about the role of spleen in NAFLD, could impact real world outcomes through establishing better tools for a precocious diagnosis. Using both liver and spleen ultrasonography, technique largely dealt with in this review, could expand the possibility to cover an adequate diagnostic path toward NAFLD, reaching a good sensibility and specificity.