IL32 downregulation lowers triglycerides and type I collagen in di-lineage human primary liver organoids.

Steatotic liver disease (SLD) prevails as the most common chronic liver disease yet lack approved treatments due to incomplete understanding of pathogenesis. Recently, elevated hepatic and circulating interleukin 32 (IL-32) levels were found in individuals with severe SLD. However, the mechanistic link between IL-32 and intracellular triglyceride metabolism remains to be elucidated. We demonstrate in vitro that incubation with IL-32ß protein leads to an increase in intracellular triglyceride synthesis, while downregulation of IL32 by small interfering RNA leads to lower triglyceride synthesis and secretion in organoids from human primary hepatocytes. This reduction requires the upregulation of Phospholipase A2 group IIA (PLA2G2A). Furthermore, downregulation of IL32 results in lower intracellular type I collagen levels in di-lineage human primary hepatic organoids. Finally, we identify a genetic variant of IL32 (rs76580947) associated with lower circulating IL-32 and protection against SLD measured by non-invasive tests. These data suggest that IL32 downregulation may be beneficial against SLD.

Predictors of controlled attenuation parameter in metabolic dysfunction.

BACKGROUND & AIMS: Hepatic fat content can be non-invasively estimated by controlled attenuation parameter (CAP) during transient elastography. The aim of this study was to examine the determinants and predictors of CAP values in individuals with metabolic dysfunction. METHODS: We enrolled 1230 consecutive apparently healthy individuals (Liver-Bible-2022 cohort) with ≥3 metabolic dysfunction features. CAP was measured by Fibroscan. CAP determinants and predictors were identified using backward stepwise analysis and introduced in generalized linear models. RESULTS: Participants were predominantly males (82.9%), mean age was 53.8 ± 6.4 years, 600 (48.8%) had steatosis (CAP ≥ 275 dB/m), and 27 had liver stiffness measurement (LSM) ≥ 8 kPa. CAP values correlated with LSM (p < 10-22). In multivariable analysis, fasting insulin and abdominal circumference (AC) were the main determinants of CAP (p < 10-6), together with body mass index (BMI; p < 10-4), age, diabetes, triglycerides, ferritin, and lower HDL and thyroid stimulating hormone (TSH; p < 0.05 for all). In a subset of 592 participants with thyroid hormone measurement, we found an association between higher free triiodothyronine levels, correlating with lower TSH, and CAP values, independent of TSH and of levothyroxine treatment (p = 0.0025). A clinical CAP score based on age, BMI, AC, HbA1c, ALT, and HDL predicted CAP ≥ 275 dB/m with moderate accuracy (AUROC = 0.73), which was better than that of the Fatty Liver Index and of ALT (AUROC = 0.70/0.61, respectively) and validated it in multiple cohorts. CONCLUSION: Abdominal adiposity and insulin resistance severity were the main determinants of CAP in individuals with metabolic dysfunction and may improve steatotic liver disease risk stratification. CAP values were modulated by the hypophysis-thyroid axis.

Liver Involvement in Patients with Rare MBOAT7 Variants and Intellectual Disability: A Case Report and Literature Review.

The membrane-bound O-acyltransferase domain-containing 7 (MBOAT7) protein is an acyltransferase catalyzing arachidonic acid incorporation into lysophosphatidylinositol. Patients with rare, biallelic loss-of-function variants of the MBOAT7 gene display intellectual disability with neurodevelopmental defects. The rs641738 inherited variant associated with reduced hepatic MBOAT7 expression has been linked to steatotic liver disease susceptibility. However, the impact of biallelic loss-of-function MBOAT7 variants on liver disease is not known. We report on a 2-year-old girl with MBOAT7-related intellectual disability and steatotic liver disease, confirming that MBOAT7 loss-of-function predisposes to liver disease.

Validation of a targeted gene panel sequencing for the diagnosis of hereditary chronic liver diseases.

Background: The cause of chronic liver diseases (CLD) remains undiagnosed in up to 30% of adult patients. Whole-Exome Sequencing (WES) can improve the diagnostic rate of genetic conditions, but it is not yet widely available, due to the costs and the difficulties in results interpretation. Targeted panel sequencing (TS) represents an alternative more focused diagnostic approach. Aims: To validate a customized TS for hereditary CLD diagnosis. Methods: We designed a customized panel including 82 CLD-associated genes (iron overload, lipid metabolism, cholestatic diseases, storage diseases, specific hereditary CLD and susceptibility to liver diseases). DNA samples from 19 unrelated adult patients with undiagnosed CLD were analyzed by both TS (HaloPlex) and WES (SureSelect Human All Exon kit v5) and the diagnostic performances were compared. Results: The mean depth of coverage of TS-targeted regions was higher with TS than WES (300x vs. 102x; p < 0.0001). Moreover, TS yielded a higher average coverage per gene and lower fraction of exons with low coverage (p < 0.0001). Overall, 374 unique variants were identified across all samples, 98 of which were classified as "Pathogenic" or "Likely Pathogenic" with a high functional impact (HFI). The majority of HFI variants (91%) were detected by both methods; 6 were uniquely identified by TS and 3 by WES. Discrepancies in variant calling were mainly due to variability in read depth and insufficient coverage in the corresponding target regions. All variants were confirmed by Sanger sequencing except two uniquely detected by TS. Detection rate and specificity for variants in TS-targeted regions of TS were 96.9% and 97.9% respectively, whereas those of WES were 95.8% and 100%, respectively. Conclusion: TS was confirmed to be a valid first-tier genetic test, with an average mean depth per gene higher than WES and a comparable detection rate and specificity.

Programmed cell death 1 genetic variant and liver damage in nonalcoholic fatty liver disease.

BACKGROUND AND AIMS: Programmed cell death 1/programmed cell death-ligand 1 (PD-1/PDL-1) axis has been reported to modulate liver inflammation and progression to hepatocellular carcinoma (HCC) in patients with nonalcoholic fatty liver disease (NAFLD). Here, we examined whether the PDCD1 variation is associated with NAFLD severity in individuals with liver biopsy. METHODS: We examined the impact of PDCD1 gene variants on HCC, as robust severe liver disease phenotype in UK Biobank participants. The strongest genetic association with the rs13023138 G>C variation was subsequently tested for association with liver damage in 2889 individuals who underwent liver biopsy for suspected nonalcoholic steatohepatitis (NASH). Hepatic transcriptome was examined by RNA-Seq in a subset of NAFLD individuals (n = 121). Transcriptomic and deconvolution analyses were performed to identify biological pathways modulated by the risk allele. RESULTS: The rs13023138 C>G showed the most robust association with HCC in UK Biobank (p = 5.28E-4, OR = 1.32, 95% CI [1.1, 1.5]). In the liver biopsy cohort, rs13023138 G allele was independently associated with severe steatosis (OR 1.17, 95% CI 1.02-1.34; p = .01), NASH (OR 1.22, 95% CI 1.09-1.37; p < .001) and advanced fibrosis (OR 1.26, 95% CI 1.06-1.50; p = .007). At deconvolution analysis, rs13023138 G>C allele was linked to higher hepatic representation of M1 macrophages, paralleled by upregulation of pathways related to inflammation and higher expression of CXCR6. CONCLUSIONS: The PDCD1 rs13023138 G allele was associated with HCC development in the general population and with liver disease severity in patients at high risk of NASH.

Metabolic dysfunction outperforms ultrasonographic steatosis to stratify hepatocellular carcinoma risk in patients with advanced hepatitis C cured with direct-acting antivirals.

BACKGROUND AND AIMS: Metabolic dysfunction (MD)-associated fatty liver disease has been proposed to identify individuals at risk of liver events irrespectively of the contemporary presence of other liver diseases. The aim of this study was to examine the impact of MD in patients cured of chronic hepatis C (CHC). PATIENTS AND METHODS: We analysed data from a real-life cohort of 2611 Italian patients cured of CHC with direct antiviral agents and advanced liver fibrosis, without HBV/HIV, transplantation and negative for hepatocellular carcinoma (HCC) history (age 61.4 ± 11.8 years, 63.9% males, median follow-up 34, 24-40 months). Information about ultrasonographic steatosis (US) after sustained virological response was available in 1978. RESULTS: MD affected 58% of patients, diagnosed due to the presence of diabetes (MD-diabetes, 19%), overweight without diabetes (MD-overweight, 37%) or multiple metabolic abnormalities without overweight and diabetes (MD-metabolic, 2%). MD was more frequent than and not coincident with US (32% MD-only, 23% MD-US and 13% US-only). MD was associated with higher liver stiffness (p < 0.05), particularly in patients with MD-diabetes and MD-only subgroups, comprising older individuals with more advanced metabolic and liver disease (p < 0.05). At Cox proportional hazard multivariable analysis, MD was associated with increased risk of HCC (HR 1.97, 95% CI 1.27-3.04; p = 0.0023). Further classification according to diagnostic criteria improved risk stratification (p < 0.0001), with the highest risk observed in patients with MD-diabetes. Patients with MD-only appeared at highest risk since the sustained virological response achievement (p = 0.008), with a later catch-up of those with combined MD-US, whereas US-only was not associated with HCC. CONCLUSIONS: MD is more prevalent than US in patients cured of CHC with advanced fibrosis and identifies more accurately individuals at risk of developing HCC.

Circulating Interlukin-32 and Altered Blood Pressure Control in Individuals with Metabolic Dysfunction.

Fatty liver disease is most frequently related to metabolic dysfunction (MAFLD) and associated comorbidities, heightening the risk of cardiovascular disease, and is associated with higher hepatic production of IL32, a cytokine linked with lipotoxicity and endothelial activation. The aim of this study was to examine the relationship between circulating IL32 concentration and blood pressure control in individuals with metabolic dysfunction at high risk of MAFLD. IL32 plasma levels were measured by ELISA in 948 individuals with metabolic dysfunction enrolled in the Liver-Bible-2021 cohort. Higher circulating IL32 levels were independently associated with systolic blood pressure (estimate +0.008 log10 per 1 mmHg increase, 95% c.i. 0.002-0.015; p = 0.016), and inversely correlated with antihypertensive medications (estimate -0.189, 95% c.i. -0.291--0.088, p = 0.0002). Through multivariable analysis, IL32 levels predicted both systolic blood pressure (estimate 0.746, 95% c.i 0.173-1.318; p = 0.010) and impaired blood pressure control (OR 1.22, 95% c.i. 1.09-1.38; p = 0.0009) independently of demographic and metabolic confounders and of treatment. This study reveals that circulating IL32 levels are associated with impaired blood pressure control in individuals at risk of cardiovascular disease.

Adverse effect of PNPLA3 p.I148M genetic variant on kidney function in middle-aged individuals with metabolic dysfunction.

BACKGROUND: The PNPLA3 p.I148M variant is the main genetic determinant of nonalcoholic fatty liver disease, and PNPLA3 silencing is being evaluated to treat this liver condition. Data suggest that the p.I148M variant predisposes to kidney damage, but the relative contribution to kidney function, compared to overall genetic susceptibility, is not defined. AIMS: We aimed to assess the effect of PNPLA3 p.I148M on the estimated glomerular filtration rate (eGFR) in individuals with metabolic dysfunction. METHODS: We included 1144 middle-aged individuals from the Liver-Bible-2022 cohort. Glomerular filtration rate (eGFR) was estimated using the Chronic Kidney Disease Epidemiology Collaboration equation. The effect of PNPLA3 p.I148M on eGFRCKD-EPI levels was tested under additive genetic models adjusted for clinical predictors, ethnicity and a polygenic risk score of chronic kidney disease (PRS-CKD). In a subset of 144 individuals, we examined the effect of PNPLA3 p.I148M on eGFRCKD-EPI over a median follow-up of 17 months. RESULTS: The p.I148M variant was associated with lower eGFRCKD-EPI levels (-1.24 mL/min/1.73 m2 per allele, 95% CI: -2.32 to -0.17; p = 0.023), independent of age, sex, height, waist circumference, systolic blood pressure, LDL-cholesterol, transaminases, fasting insulin, albuminuria, lipid-lowering drugs, ethnicity and PRS-CKD score. In the prospective evaluation, the p.I148M variant was independently associated with faster eGFRCKD-EPI decline (ΔeGFRCKD-EPI -3.57 mL/min/1.73 m2 per allele, 95% CI: -6.94 to -0.21; p = 0.037). CONCLUSIONS: We found a detrimental impact of the PNPLA3 p.I148M variant on eGFRCKD-EPI levels in middle-aged individuals with metabolic dysfunction. This association was independent of established risk factors, ethnicity and genetic predisposition to CKD. PNPLA3 p.I148M silencing may protect against kidney damage progression in carriers.

Development and Validation of a Score for Fibrotic Nonalcoholic Steatohepatitis.

BACKGROUND & AIMS: Noninvasive assessment of histological features of nonalcoholic fatty liver disease (NAFLD) has been an intensive research area over the last decade. Herein, we aimed to develop a simple noninvasive score using routine laboratory tests to identify, among individuals at high risk for NAFLD, those with fibrotic nonalcoholic steatohepatitis (NASH) defined as NASH, NAFLD activity score ≥4, and fibrosis stage ≥2. METHODS: The derivation cohort included 264 morbidly obese individuals undergoing intraoperative liver biopsy in Rome, Italy. The best predictive model was developed and internally validated using a bootstrapping stepwise logistic regression analysis (2000 bootstrap samples). Performance was estimated by the area under the receiver operating characteristic curve (AUROC). External validation was assessed in 3 independent European cohorts (Finland, n = 370; Italy, n = 947; England, n = 5368) of individuals at high risk for NAFLD. RESULTS: The final predictive model, designated as Fibrotic NASH Index (FNI), combined aspartate aminotransferase, high-density lipoprotein cholesterol, and hemoglobin A1c. The performance of FNI for fibrotic NASH was satisfactory in both derivation and external validation cohorts (AUROC = 0.78 and AUROC = 0.80-0.95, respectively). In the derivation cohort, rule-out and rule-in cutoffs were 0.10 for sensitivity ≥0.89 (negative predictive value, 0.93) and 0.33 for specificity ≥0.90 (positive predictive value, 0.57), respectively. In the external validation cohorts, sensitivity ranged from 0.87 to 1 (negative predictive value, 0.99-1) and specificity from 0.73 to 0.94 (positive predictive value, 0.12-0.49) for rule-out and rule-in cutoff, respectively. CONCLUSION: FNI is an accurate, simple, and affordable noninvasive score which can be used to screen for fibrotic NASH in individuals with dysmetabolism in primary health care.

Clinical and genetic determinants of the fatty liver-coagulation balance interplay in individuals with metabolic dysfunction.

Background & Aims: The aim of this study was to examine the determinants of the interplay between liver damage and the coagulation balance in individuals at risk of non-alcoholic fatty liver disease (NAFLD). Methods: We considered 581 healthy participants with ≥3 metabolic alterations undergoing clinical and genomic evaluation, measurement of liver stiffness (LSM) and controlled attenuation parameter (CAP) by Fibroscan, Pro-C3, coagulation balance (von Willebrand factor [vWF], factor VIII/protein C ratio [F8/PC] as the main outcome, D-dimer as marker of coagulation/fibrinolysis activation). Results: Liver fibrosis indices (both Fibrosis-4 [FIB-4] and liver stiffness measurement [LSM]), but not liver fat (CAP), were independently associated with higher F8/PC ratio (p <0.01), triggering D-dimer formation (p = 2E-21). In keeping with a causal role of liver damage in determining a procoagulant status, the main fatty liver inherited risk variant PNPLA3 p.I148M was independently associated with the F8/PC ratio (p = 0.048). Vice versa, the main determinant of the coagulation balance was ABO locus variation (p = 1E-16), through the impact on vWF (p = 8E-26). Both rs687289 ABO and factor V Leiden were independently associated with higher Pro-C3 (p <0.025), with the effect of ABO being mediated by the impact on vWF (p = 5E-10 for association with Pro-C3). Mendelian randomisation analysis was consistent with a causal association of procoagulant imbalance with heightened fibrogenesis (p = 0.001 at robust MR-Egger for Pro-C3), but not with fibrosis (for LSM; p = not significant). Conclusions: In individuals with metabolic dysfunction, liver damage severity and possibly the PNPLA3 p.I148M variant were associated with procoagulant status. Vice versa, evaluation of inherited variants in ABO and other genes influencing coagulation was consistent with a causal role of procoagulant imbalance in activation of early stages of fibrogenesis. Lay summary: In individuals with metabolic alterations at risk of metabolic fatty liver disease, there is a tendency toward heightened blood coagulation (clotting), but the cause and the impact on the progression of liver disease remain unclear. Here we show that liver damage severity and metabolic alterations, but not hepatic fat, are mainly responsible for heightened coagulation in patients with metabolic fatty liver disease. By using genetic approaches, we showed that hepatic inflammation due to lipotoxicity may favour heightened coagulation, which in turn can trigger liver fibrosis, igniting a vicious cycle that leads to progressive liver disease.

The Long Non-Coding BC200 Is a Novel Circulating Biomarker of Parathyroid Carcinoma.

Long non-coding RNAs (lncRNAs) are an important class of epigenetic regulators involved in both physiological processes and cancer development. Preliminary evidence suggested that lncRNAs could act as accurate prognostic and diagnostic biomarkers. Parathyroid cancer is a rare endocrine neoplasia, whose management represents a clinical challenge due to the lack of accurate molecular biomarkers. Our previous findings showed that human parathyroid tumors are characterized by a different lncRNAs signature, suggesting heterogeneity through the different histotypes. Particularly, we found that the lncRNA BC200/BCYRN1 could represent a candidate biomarker for parathyroid carcinomas (PCas). Here we aimed to extend our preliminary data evaluating whether BC200 could be an accurate non-invasive biomarker of PCas to support the clinical management of patients affected by parathyroid tumors at diagnosis, prognosis and follow-up. To provide a non-invasive point-of-care for parathyroid carcinoma diagnosis and follow-up, we analyzed BC200 expression in patients' serum through digital PCR. Our results show that BC200 counts are higher in serum from patients harboring PCa (n=4) compared to patients with parathyroid adenoma (PAd; n=27). Further, in PAd patients circulating BC200 levels are positively correlated with serum total calcium. Then, we found that BC200 is overexpressed in metastatic PCas (n=4) compared to non-metastatic ones (n=9). Finally, the lncRNA expression in PCa patients' serum drops are reduced after parathyroidectomy, suggesting its possible use in the post-operative setting for patients follow-up. Overall, these findings extend the knowledge on BC200 in parathyroid tumors, supporting its role as a useful biomarker for management of PCa.

Rare ATG7 genetic variants predispose patients to severe fatty liver disease.

BACKGROUND & AIMS: Non-alcoholic fatty liver disease (NAFLD) is the leading cause of liver disorders and has a strong heritable component. The aim of this study was to identify new loci that contribute to severe NAFLD by examining rare variants. METHODS: We performed whole-exome sequencing in individuals with NAFLD and advanced fibrosis or hepatocellular carcinoma (n = 301) and examined the enrichment of likely pathogenic rare variants vs. the general population. This was followed by validation at the gene level. RESULTS: In patients with severe NAFLD, we observed an enrichment of the p.P426L variant (rs143545741 C>T; odds ratio [OR] 5.26, 95% CI 2.1-12.6; p = 0.003) of autophagy-related 7 (ATG7), which we characterized as a loss-of-function, vs. the general population, and an enrichment in rare variants affecting the catalytic domain (OR 13.9; 95% CI 1.9-612; p = 0.002). In the UK Biobank cohort, loss-of-function ATG7 variants increased the risk of cirrhosis and hepatocellular carcinoma (OR 3.30; 95% CI 1.1-7.5 and OR 12.30, 95% CI 2.6-36, respectively; p <0.001 for both). The low-frequency loss-of-function p.V471A variant (rs36117895 T>C) was also associated with severe NAFLD in the clinical cohort (OR 1.7; 95% CI 1.2-2.5; p = 0.003), predisposed to hepatocellular ballooning (p = 0.007) evolving to fibrosis in the Liver biopsy cohort (n = 2,268), and was associated with liver injury in the UK Biobank (aspartate aminotransferase levels, p <0.001), with a larger effect in severely obese individuals in whom it was linked to hepatocellular carcinoma (p = 0.009). ATG7 protein localized to periportal hepatocytes, particularly in the presence of ballooning. In the Liver Transcriptomic cohort (n = 125), ATG7 expression correlated with suppression of the TNFα pathway, which was conversely upregulated in p.V471A carriers. CONCLUSIONS: We identified rare and low-frequency ATG7 loss-of-function variants that promote NAFLD progression by impairing autophagy and facilitating ballooning and inflammation. LAY SUMMARY: We found that rare mutations in a gene called autophagy-related 7 (ATG7) increase the risk of developing severe liver disease in individuals with dysmetabolism. These mutations cause an alteration in protein function and impairment of self-renewal of cellular content, leading to liver damage and inflammation.

Clinical exome sequencing for diagnosing severe cryptogenic liver disease in adults: A case series.

Liver diseases remain unexplained in up to 30% of adult patients; genetic analysis could help establish the correct diagnosis. In six adult patients with cryptogenic liver disease, we performed whole-exome sequencing (WES) and evaluated the individual predisposition to progressive fatty liver disease by polygenic risk scores (PRS). In one patient, WES was allowed to diagnose the Hermansky-Pudlak syndrome. In the other two patients, genetic variants in LDLRAP1/MSH6 and ALDOB genes were identified, contributing to explaining the clinical presentation and disease pathogenesis (50% diagnostic uptake). In the other three patients, rare variants with a high likelihood of disrupting protein function in APOB, ATP7B, ABCB4 and ATP8B1 were identified. One patient who developed hepatocellular carcinoma during the follow-up had a high PRS value. The study supports the role of WES, combined with risk stratification by PRS and accurate clinical assessment in improving the diagnosis and informed management in patients with cryptogenic liver disease, a positive family history or severe fatty liver not fully accounted for by environmental triggers.

Dysmetabolism, Diabetes and Clinical Outcomes in Patients Cured of Chronic Hepatitis C: A Real-Life Cohort Study.

The aim of this study was to examine the impact of features of dysmetabolism on liver disease severity, evolution, and clinical outcomes in a real-life cohort of patients treated with direct acting antivirals for chronic hepatitis C virus (HCV) infection. To this end, we considered 7,007 patients treated between 2014 and 2018, 65.3% with advanced fibrosis, of whom 97.7% achieved viral eradication (NAVIGATORE-Lombardia registry). In a subset (n = 748), liver stiffness measurement (LSM) was available at baseline and follow-up. Higher body mass index (BMI; odds ratio [OR] 1.06 per kg/m2 , 1.03-1.09) and diabetes (OR 2.01 [1.65-2.46]) were independently associated with advanced fibrosis at baseline, whereas statin use was protective (OR 0.46 [0.35-0.60]; P < 0.0001 for all). The impact of BMI was greater in those without diabetes (P = 0.003). Diabetes was independently associated with less pronounced LSM improvement after viral eradication (P = 0.001) and in patients with advanced fibrosis was an independent predictor of the most frequent clinical events, namely de novo hepatocellular carcinoma (HCC; hazard ratio [HR] 2.09 [1.20-3.63]; P = 0.009) and cardiovascular events (HR 2.73 [1.16-6.43]; P = 0.021). Metformin showed a protective association against HCC (HR 0.32 [0.11-0.96]; P = 0.043), which was confirmed after adjustment for propensity score (P = 0.038). Diabetes diagnosis further refined HCC prediction in patients with compensated advanced chronic liver disease at high baseline risk (P = 0.024). Conclusion: Metabolic comorbidities were associated with advanced liver fibrosis at baseline, whereas statins were protective. In patients with advanced fibrosis, diabetes increased the risk of de novo HCC and of cardiovascular events. Optimization of metabolic comorbidities treatment by a multi-disciplinary management approach may improve cardiovascular and possibly liver-related outcomes.

Trends and risk factors of SARS-CoV-2 infection in asymptomatic blood donors.

BACKGROUND: A large proportion of SARS-CoV-2-infected individuals does not develop severe symptoms. Serological tests help in evaluating the spread of infection and disease immunization. The aim of this study was to prospectively examine the trends and risk factors of SARS-CoV-2 infection in blood donors. STUDY DESIGN AND METHODS: We screened 8798 asymptomatic donors presenting in Milan from July 2020 to February 2021 (10,680 presentations) before the vaccination campaign for anti-nucleoprotein (NP) antibodies, and for anti-spike receptor-binding domain (RBD) antibodies and nasopharyngeal swab PCR in those who tested positive. RESULTS: The prevalence of anti-NP+/RBD+ tests increased progressively with time up to ~15% (p < .0001), preceded by a peak of PCR+ tests. Anti-RBD titers were higher in anti-NP IgG+/IgM+ than in IgG+/IgM- individuals and in those with a history of infection (p < .0001); of these 197/630 (31.2%) displayed high titers (>80 AU/ml). Anti-RBD titers declined during follow-up, depending on baseline titers (p < .0001) and time (p = .025). Risk factors for seroconversion were a later presentation date and non-O ABO blood group (p < .001). A positive PCR was detected in 0.7% of participants in the absence of SARS-CoV-2 viremia. CONCLUSIONS: During the second wave of SARS-CoV-2 infection in Northern Italy, we detected an increase in seroprevalence in healthy blood donors from ~4% to ~15%, with a trend paralleling that observed in the general population. Seroconversion was more frequent in carriers of non-O blood groups. The persistence of anti-RBD antibodies was short-lived.

Genetic risk scores and personalization of care in fatty liver disease.

Nonalcoholic fatty liver disease is the leading cause of chronic liver disease. Genetic predisposition, lifestyle, and metabolic comorbidities concur to nonalcoholic fatty liver disease development and progression to nonalcoholic steatohepatitis, cirrhosis, and hepatocellular carcinoma. Improvement in risk stratification and development of effective therapies for nonalcoholic steatohepatitis are key unmet clinical needs. Knowledge emerging from genomics could meet this need. A polygenic risk score (PRS) is calculated by summing the number of trait-associated alleles carried by an individual, which can be weighted by their effect size on the trait and captures the individual's genetic risk to develop a disease. In this review, we focalize on the potential use of PRSs for disease detection at an early stage and stratification of the risk of progression to severe forms. PRSs may represent robust instruments to implement targeted prevention programs, hepatocellular carcinoma surveillance in at-risk individuals, and to develop precision medicine therapeutic approaches.

Definition of Healthy Ranges for Alanine Aminotransferase Levels: A 2021 Update.

The changing epidemiology of liver disease, and modifications in the recommended analytical methodology call for a re-evaluation of the upper reference limits (URLs) of alanine aminotransferase (ALT) levels. Using the same approach consolidated 20 years ago to define the healthy population, we defined the URL for the newly recommended International Federation of Clinical Chemistry (IFCC) standardized test. In a cross-sectional study, we examined 21,296 apparently healthy blood donors (age 18-65 years) and calculated the sex-specific URL by the 95th percentile in individuals without risk factors for liver disease. These were tested for the ability to predict liver damage in a subset of 745 participants with dysmetabolism, in an independent cohort of 977 unselected donors, and in 899 patients with chronic liver disease. ALT levels were measured by the IFCC test. Male sex, body mass index, glucose, lipids, ferritin, hypertension, and younger age were independent ALT predictors (P < 0.001). Updated URLs were identified at 42/30 U/L in males/females, approximately 30% lower than those currently recommended by the IFCC. Due to improved sensitivity, they conferred the ability to detect steatosis and significant fibrosis in individuals with dysmetabolism (odds ratio [OR] = 2.31, range 1.40-3.80, P = 0.001; and OR = 3.35, range 1.19-9.42, P = 0.021; respectively), although with a limited accuracy, and significant fibrosis in unselected donors (OR = 2.32, 1.02-5.31, P = 0.045). Updated URLs had a moderate to high accuracy to discriminate liver conditions (area under the receiver operating characteristic curve = 0.81, range 0.78-0.91). Conclusion: Updated URLs by the IFCC method were lower than those calculated in initial studies, but higher than those in use with the recommended old, nonstandardized method, and were able to better predict liver disease. The limited awareness that different techniques are still in use should be regarded as a possible source of medical errors.

Genetic predisposition similarities between NASH and ASH: Identification of new therapeutic targets.

Fatty liver disease can be triggered by a combination of excess alcohol, dysmetabolism and other environmental cues, which can lead to steatohepatitis and can evolve to acute/chronic liver failure and hepatocellular carcinoma, especially in the presence of shared inherited determinants. The recent identification of the genetic causes of steatohepatitis is revealing new avenues for more effective risk stratification. Discovery of the mechanisms underpinning the detrimental effect of causal mutations has led to some breakthroughs in the comprehension of the pathophysiology of steatohepatitis. Thanks to this approach, hepatocellular fat accumulation, altered lipid droplet remodelling and lipotoxicity have now taken centre stage, while the role of adiposity and gut-liver axis alterations have been independently validated. This process could ignite a virtuous research cycle that, starting from human genomics, through omics approaches, molecular genetics and disease models, may lead to the development of new therapeutics targeted to patients at higher risk. Herein, we also review how this knowledge has been applied to: a) the study of the main PNPLA3 I148M risk variant, up to the stage of the first in-human therapeutic trials; b) highlight a role of MBOAT7 downregulation and lysophosphatidyl-inositol in steatohepatitis; c) identify IL-32 as a candidate mediator linking lipotoxicity to inflammation and liver disease. Although this precision medicine drug discovery pipeline is mainly being applied to non-alcoholic steatohepatitis, there is hope that successful products could be repurposed to treat alcohol-related liver disease as well.

A PDCD1 Role in the Genetic Predisposition to NAFLD-HCC?

Obesity and non-alcoholic fatty liver disease (NAFLD) are contributing to the global rise in deaths from hepatocellular carcinoma (HCC). The pathogenesis of NAFLD-HCC is not well understood. The severity of hepatic steatosis, steatohepatitis and fibrosis are key pathogenic mechanisms, but animal studies suggest altered immune responses are also involved. Genetic studies have so far highlighted a major role of gene variants promoting fat deposition in the liver (PNPLA3 rs738409; TM6SF2 rs58542926). Here, we have considered single-nucleotide polymorphisms (SNPs) in candidate immunoregulatory genes (MICA rs2596542; CD44 rs187115; PDCD1 rs7421861 and rs10204525), in 594 patients with NAFLD and 391 with NAFLD-HCC, from three European centres. Associations between age, body mass index, diabetes, cirrhosis and SNPs with HCC development were explored. PNPLA3 and TM6SF2 SNPs were associated with both progression to cirrhosis and NAFLD-HCC development, while PDCD1 SNPs were specifically associated with NAFLD-HCC risk, regardless of cirrhosis. PDCD1 rs7421861 was independently associated with NAFLD-HCC development, while PDCD1 rs10204525 acquired significance after adjusting for other risks, being most notable in the smaller numbers of women with NAFLD-HCC. The study highlights the potential impact of inter individual variation in immune tolerance induction in patients with NAFLD, both in the presence and absence of cirrhosis.