Complexity of ballooned hepatocyte feature recognition: Defining a training atlas for artificial intelligence-based imaging in NAFLD.

BACKGROUND & AIMS: Histologically assessed hepatocyte ballooning is a key feature discriminating non-alcoholic steatohepatitis (NASH) from steatosis (NAFL). Reliable identification underpins patient inclusion in clinical trials and serves as a key regulatory-approved surrogate endpoint for drug efficacy. High inter/intra-observer variation in ballooning measured using the NASH CRN semi-quantitative score has been reported yet no actionable solutions have been proposed. METHODS: A focused evaluation of hepatocyte ballooning recognition was conducted. Digitized slides were evaluated by 9 internationally recognized expert liver pathologists on 2 separate occasions: each pathologist independently marked every ballooned hepatocyte and later provided an overall non-NASH NAFL/NASH assessment. Interobserver variation was assessed and a 'concordance atlas' of ballooned hepatocytes generated to train second harmonic generation/two-photon excitation fluorescence imaging-based artificial intelligence (AI). RESULTS: The Fleiss kappa statistic for overall interobserver agreement for presence/absence of ballooning was 0.197 (95% CI 0.094-0.300), rising to 0.362 (0.258-0.465) with a ≥5-cell threshold. However, the intraclass correlation coefficient for consistency was higher (0.718 [0.511-0.900]), indicating 'moderate' agreement on ballooning burden. 133 ballooned cells were identified using a ≥5/9 majority to train AI ballooning detection (AI-pathologist pairwise concordance 19-42%, comparable to inter-pathologist pairwise concordance of between 8-75%). AI quantified change in ballooned cell burden in response to therapy in a separate slide set. CONCLUSIONS: The substantial divergence in hepatocyte ballooning identified amongst expert hepatopathologists suggests that ballooning is a spectrum, too subjective for its presence or complete absence to be unequivocally determined as a trial endpoint. A concordance atlas may be used to train AI assistive technologies to reproducibly quantify ballooned hepatocytes that standardize assessment of therapeutic efficacy. This atlas serves as a reference standard for ongoing work to refine how ballooning is classified by both pathologists and AI. LAY SUMMARY: For the first time, we show that, even amongst expert hepatopathologists, there is poor agreement regarding the number of ballooned hepatocytes seen on the same digitized histology images. This has important implications as the presence of ballooning is needed to establish the diagnosis of non-alcoholic steatohepatitis (NASH), and its unequivocal absence is one of the key requirements to show 'NASH resolution' to support drug efficacy in clinical trials. Artificial intelligence-based approaches may provide a more reliable way to assess the range of injury recorded as "hepatocyte ballooning".

Development and prognostic relevance of a histologic grading and staging system for alcohol-related liver disease.

BACKGROUND & AIMS: The SALVE Histopathology Group (SHG) developed and validated a grading and staging system for the clinical and full histological spectrum of alcohol-related liver disease (ALD) and evaluated its prognostic utility in a multinational cohort of 445 patients. METHODS: SALVE grade was described by semiquantitative scores for steatosis, activity (hepatocellular injury and lobular neutrophils) and cholestasis. The histological diagnosis of steatohepatitis due to ALD (histological ASH, hASH) was based on the presence of hepatocellular ballooning and lobular neutrophils. Fibrosis staging was adapted from the Clinical Research Network staging system for non-alcoholic fatty liver disease and the Laennec staging system and reflects the pattern and extent of ALD fibrosis. There are 7 SALVE fibrosis stages (SFS) ranging from no fibrosis to severe cirrhosis. RESULTS: Interobserver κ-value for each grading and staging parameter was >0.6. In the whole study cohort, long-term outcome was associated with activity grade and cholestasis, as well as cirrhosis with very broad septa (severe cirrhosis) (p <0.001 for all parameters). In decompensated ALD, adverse short-term outcome was associated with activity grade, hASH and cholestasis (p = 0.038, 0.012 and 0.001, respectively), whereas in compensated ALD, hASH and severe fibrosis/cirrhosis were associated with decompensation-free survival (p = 0.011 and 0.001, respectively). On multivariable analysis, severe cirrhosis emerged as an independent histological predictor of long-term survival in the whole study cohort. Severe cirrhosis and hASH were identified as independent predictors of short-term survival in decompensated ALD, and also as independent predictors of decompensation-free survival in compensated ALD. CONCLUSION: The SALVE grading and staging system is a reproducible and prognostically relevant method for the histological assessment of disease activity and fibrosis in ALD. LAY SUMMARY: Patients with alcohol-related liver disease (ALD) may undergo liver biopsy to assess disease severity. We developed a system to classify ALD under the microscope by grading ALD activity and staging the extent of liver scarring. We validated the prognostic performance of this system in 445 patients from 4 European centers.

Liver stiffness measurements by 2-dimensional shear wave elastography compared to histological and ultrasound parameters in primary biliary cholangitis.

BACKGROUND AND AIMS: Liver stiffness measurements (LSMs) by 2-dimensional-shear-wave elastography (LSM2D-SWE) are now widely used in hepatology. However, relevant information for primary biliary cholangitis (PBC) is scant. We compare LSM2D-SWE with liver biopsy (LB) in a cohort of PBC patients in Greece. METHODS: Data of 68 LBs from 53 PBC patients were retrospectively analyzed and fibrosis stage was compared to LSM2D-SWE. Forty-six patients (86.8%) were females and at the time of LBx median (IQR) age was 62.6 (53.2-72.1). Demographic, UDCA treatment, histological and B-mode ultrasound data were tested for their influence on LSM2D-SWE estimates. RESULTS: Liver fibrosis stages F0-F4 were found in 4, 19, 19, 16 and 10 cases, respectively. Across stages F0-F4, the LSM2D-SWE was 5.6 (5.1-6.1), 7.0 (5.8-7.7), 9.1 (7.3-11.5), 10.8 (9.9-12.2) and 14.5 (11.9-25.7) kPa, respectively, with highly significant difference (p<.001). The LSM2D-SWE differed also significantly between F0 vs. F1 (p=.027), F1 vs. F2 (p=.005) and F3 vs. F4 (p=.017). The discriminatory ability of LSM2D-SWE for mild, significant, severe fibrosis and cirrhosis was highly significant in all comparisons (p<.001), with AUC2D-SWE 95.3%, 87.4%, 85.3% and 95.3% and accuracy 89.7%, 85.3%, 80.9% and 86.8%, respectively. Among 21 parameters tested, significant predictors of LSM2D-SWE by multiple linear regression were fibrosis stage, portal inflammation and parenchymal heterogeneity. The portal inflammation grade accounted for 32.2% of LSM variation with adjusted R2 0.428. CONCLUSIONS: In patients with PBC, LSM measurements by 2D-SWE can reliably discriminate between mild, significant, severe fibrosis and cirrhosis. Measurements are significantly affected by portal inflammation grade.

Fatty Liver Disease and Hepatocellular Carcinoma: The Pathologist's View.

Chronic alcohol misuse and progressed nonalcoholic fatty liver disease (NAFLD) due to the metabolic syndrome and resulting to nonalcoholic steatohepatitis (NASH) are prime causes of hepatocellular carcinoma (HCC) in Western industrialized countries. The incidence of HCC in NASH-cirrhosis is lower than that of HCC occuring in HCV-related or alcoholic cirrhosis. Up to 20% of cases of alcohol-associated HCC may develop in pre-cirrhotic liver while HCC is also increasingly recognised in pre-cirrhotic NASH raising questions on appropriate surveillance measures for these patient populations. The recently described steatohepatitic subtype of HCC presents with higher frequency in NAFLD compared to alcoholic liver disease (ALD) patients. This review will mainly focus on histopathology and summarize current data on the epidemiology, pathogenesis, diagnosis and management of NAFLD- and ALD-related HCC.

Diagnosis and Assessment of NAFLD: Definitions and Histopathological Classification.

Nonalcoholic fatty liver disease (NAFLD) comprises a spectrum of clinical and histopathological changes including "simple" steatosis, steatosis with inflammation, steatohepatitis, cirrhosis, and hepatocellular carcinoma. It was initially described in the context of drug-induced liver injury and acute liver disease following jejunoileal bypass surgery, but since the early 1980s it has been widely acknowledged as the hepatic manifestation of metabolic syndrome. It now represents a burgeoning public health crisis and is fast becoming the main indication for liver transplantation in some parts of the world. Its true incidence and prevalence are unknown, although estimates have been made from large imaging studies. Liver biopsy interpretation is still regarded as the gold standard for making accurate diagnoses in NAFLD, but sampling limitations are recognized. Furthermore, clear definitions for key histopathological components have been lacking, resulting in significant interobserver variations in making a diagnosis of steatohepatitis. In this review the authors consider some aspects of classification and variant forms of NAFLD such as that occurring in children. They provide an update on grading and staging systems and histopathological prognostic factors, and address the role of liver biopsy in contemporary clinical care of patients with NAFLD.

Ubiquitin C-terminal hydrolase 1: A novel functional marker for liver myofibroblasts and a therapeutic target in chronic liver disease.

BACKGROUND & AIMS: Ubiquitination is a reversible protein modification involved in the major cellular processes that define cell phenotype and behaviour. Ubiquitin modifications are removed by a large family of proteases named deubiquitinases. The role of deubiquitinases in hepatic stellate cell (HSC) activation and their contribution to fibrogenesis are poorly defined. We have identified that the deubiquitinase ubiquitin C-terminal hydrolase 1 (UCHL1) is highly induced following HSC activation, determined its function in activated HSC and its potential as a therapeutic target for fibrosis. METHODS: Deubiquitinase expression was determined in day 0 and day 10 HSC. Increased UCHL1 expression was confirmed in human HSC and in an alcoholic liver disease (ALD) patient liver. The importance of UCHL1 in hepatic fibrosis was investigated in CCl4 and bile duct ligation injured mice using a pharmacological inhibitor (LDN 57444). The effects of UCHL1 inhibition on HSC proliferation were confirmed by Western blot and 3H thymidine incorporation. RESULTS: Here we report that pharmacological inhibition of UCHL1 blocks progression of established fibrosis in CCl4 injured mice. UCHL1 siRNA knockdown, LDN 57444 treatment, or HSC isolated from UCHL1(-/-) mice show attenuated proliferation in response to the mitogen, platelet-derived growth factor. Additionally, we observed changes in the phosphorylation of the cell cycle regulator retinoblastoma protein (Rb) in the absence of UCHL1 highlighting a potential mechanism for the reduced proliferative response. CONCLUSIONS: UCHL1 expression is highly upregulated upon HSC activation and is involved in the regulation of HSC proliferation. This study highlights therapeutic opportunities for pharmacological targeting of UCHL1 in chronic liver disease.

Liver innervation and hepatic function: new insights.

The hepatic nervous system has a well-known impact on the regulation of liver function and organism homeostasis. The aim of this review is to summarize the new available data regarding the role of hepatic nerves. In the last decade, studies have shown that hepatic nerves exert subtle but significant modifications on the regulation of glucose and lipid metabolism, food intake, and liver regeneration. They also play a role in liver disease pathogenesis, and hepatic denervation has beneficial results to liver graft ischemia-reperfusion injury. Available data are still limited, and further research toward neural pathways involving the liver that can modify response to disease is required.

Role of Histopathology in Autoimmune Hepatitis.

The diagnosis of autoimmune hepatitis (AIH) is based on a combination of biochemical, immunological and histological features and exclusion of other causes of liver disease. Typical histological features include a chronic hepatitis pattern of injury with portal inflammation and interface activity, predominance of plasma cells in the portal infiltrate, emperipolesis, and hepatocellular rosette formation. Centrilobular injury with prominent hepatocellular necrosis and mononuclear inflammation is now recognised in the histological spectrum of AIH and may represent an early stage of the disease. Liver histology plays a major role in clinical diagnostic scoring systems and is important to confirm or support the clinical diagnosis of AIH. This review focuses on the role of histopathology in AIH and highlights the contribution of histological interpretation to the diagnosis of AIH, differential diagnosis from other entities, recognition of concurrent liver disease, and identification of the so-called overlap or variant syndromes, and addresses the importance of liver biopsy in the management and prognosis of patients with AIH.

Pathology of the liver sinusoids.

The hepatic sinusoids comprise a complex of vascular conduits to transport blood from the porta hepatis to the inferior vena cava through the liver. Under normal conditions, portal venous and hepatic artery pressures are equalized within the sinusoids, oxygen and nutrients from the systemic circulation are delivered to the parenchymal cells and differentially distributed throughout the liver acini, and proteins of liver derivation are carried into the cardiac/systemic circulation. Liver sinusoid structures are lined by endothelial cells unique to their location, and Kupffer cells. Multifunctional hepatic stellate cells and various immune active cells are localized within the space of Disse between the sinusoid and the adjacent hepatocytes. Flow within the sinusoids can be compromised by physical or pressure blockage in their lumina as well as obstructive processes within the space of Disse. The intimate relationship of the liver sinusoids to neighbouring hepatocytes is a significant factor affecting the health of hepatocytes, or transmission of the effects of injury within the sinusoidal space. Pathologists should recognize several patterns of injury involving the sinusoids and surrounding hepatocytes. In this review, injury, alterations and accumulations within the liver sinusoids are illustrated and discussed.

Liver regeneration: immunohistochemical study of intrinsic hepatic innervation after partial hepatectomy in rats.

BACKGROUND: We examined the intrinsic hepatic innervation after partial hepatectomy (PH) in rats and the presence and pattern of neural sprouting in regenerating liver. METHODS: Male Wistar rats (age 9-13 weeks-w, weight 204-356 g), were submitted to two-thirds PH. Rats were sacrificed at postoperative days (d) 1, 3, 5, 7, at 2 and 4 w, and at 3 and 6 months (m) (6-7 animals/group, control group n = 4). Immunohistochemistry for the pan-neural marker protein gene product 9.5 (PGP9.5) and growth-associated protein 43 (GAP-43), a marker of regenerating nerve axons, was performed on tissue sections from the R1 lobe of the regenerating liver. Portal tracts (PTs) with immunoreactive fibers were counted in each section and computer-assisted morphometric analysis (Image Pro Plus) was used to measure nerve fiber density (number of immuno-positive nerve fibers/mm2 (40x)). RESULTS: Immunoreactivity for PGP9.5 was positive in all groups. The number of PGP9.5 (+) nerve fibers decreased from 0.32 +/- 0.12 (control group) to 0.18 +/- 0.09 (1d post-PH group), and gradually increased reaching pre-PH levels at 6 m (0.3 +/- 0.01). In contrast, immunoreactivity for GAP-43 was observed at 5d post-PH, and GAP-43 (+) PTs percentage increased thereafter with a peak at 3 m post-PH. GAP-43 (+) nerve fiber density increased gradually from 5d (0.05 +/- 0.06) with a peak at 3 m post-PH (0.21 +/- 0.027). At 6 m post-PH, immunoreactivity for GAP-43 was not detectable. CONCLUSIONS: Following PH in rats: 1) nerve fiber density in portal tracts decreases temporarily, and 2) neural sprouting in the regenerating liver lobes starts at 5d, reaches peak levels at 3 m and disappears at 6 m post-PH, indicating that the increase in hepatic mass after PH provides an adequate stimulus for the sprouting process.

Increased density of cutaneous nerve fibres in the affected dermatomes after herpes zoster therapy.

Herpes zoster neural injury was assessed by determining cutaneous nerve density in skin biopsies from the affected dermatomes of 35 adult patients with herpes zoster in the acute phase and 3 months post-treatment, using protein gene product 9.5 immunohistochemistry. In contrast to the significant increase in subepidermal nerve fibre density (11.77 ± 4.88/mm vs. 13.29 ± 5.74/mm, p = 0.045) after 3 months, no differences were found in epidermal free nerve endings (2.43 ± 2.35/mm and 2.8 ± 2.86/mm, p = 0.168). Patients with post-herpetic neuralgia had significantly lower subepidermal nerve fibre densities (9.7 ± 2.05/mm vs. 14.72 ± 6.13/mm, p = 0.011) compared with non-post-herpetic neuralgia patients. No differences in cutaneous nerve density were found in relation to antiviral therapy. In conclusion, 3 months after acute infection, no sign of epidermal innervation recovery is observed, while the increased subepidermal nerve fibre density in the affected dermatomes probably reflects nerve regeneration that is not affected by antiviral agent type. Subepidermal nerve fibre density is decreased in patients with post-herpetic neuralgia 3-months post-acute herpes zoster infection.

Significance of liver histology in HBsAg-positive, IgM anti-HBc-negative acute hepatitis B virus-related hepatitis.

AIMS: The natural course of HBsAg-positive, IgM anti-HBc-negative acute hepatitis B virus (HBV)-related hepatitis is unclear. The aim of this study was to evaluate the prognostic significance of histological features and hepatic expression of HBV antigens in such patients. METHODS AND RESULTS: Fifty patients with HBsAg-positive, IgM anti-HBc-negative acute hepatitis B who underwent liver biopsy during the acute hepatitis episode were studied [HBeAg seroconversion (n = 16), persistently positive for HBeAg (n = 9), and persistently negative for HBeAg (n = 25)]. Twenty-six cases had features of typical acute hepatitis only (group A), and 24 cases had changes suggesting pre-existing chronic hepatitis (group B). HBcAg and/or HBsAg immunoreactivity was detected less frequently in group A than in group B (31% versus 79%, P = 0.01). HBsAg clearance was observed in 24% of patients, almost exclusively in cases with HBeAg seroconversion. HBsAg loss was significantly more frequent in group A than in group B (52% versus 0%, P < 0.001), and in cases without rather than with immunohistochemical expression of HBV antigens (55% versus 0%, P < 0.001). In group A, HBsAg clearance was observed in 80%, 54% and 0% of patients with mild, moderate or severe acute hepatitis, respectively (P < 0.034). CONCLUSIONS: Histological information is very important for the prognosis of HBsAg-positive, IgM anti-HBc-negative acute hepatitis B. HBeAg seroconversion with underlying typical acute hepatitis changes of mild to moderate severity without hepatic expression of HBV antigens strongly predicts subsequent HBsAg loss.

Development and Application of a Semi quantitative Scoring Method for Ultrastructural Assessment of Acute Stress in Pancreatic Islets.

BACKGROUND: Pancreas and islet transplantation outcomes are negatively impacted by injury to the endocrine cells from acute stress during donor death, organ procurement, processing, and transplant procedures. Here, we report a novel electron microscopy scoring system, the Newcastle Pancreas Endocrine Stress Score (NPESS). METHODS: NPESS was adapted and expanded from our previously validated method for scoring pancreatic exocrine acinar cells, yielding a 4-point scale (0-3) classifying ultrastructural pathology in endocrine cell nuclei, mitochondria, endoplasmic reticulum, cytoplasmic vacuolization, and secretory granule depletion, with a maximum additive score of 15. We applied NPESS in a cohort of deceased organ donors after brainstem (DBD) and circulatory (DCD) death with a wide range of cold ischemic times (3.6-35.9 h) including 3 donors with type 1 and 3 with type 2 diabetes to assess islets in situ (n = 30) in addition to pancreata (n = 3) pre- and postislet isolation. RESULTS: In DBD pancreata, NPESS correlated with cold ischemic time (head: r = 0.55; P = 0.02) and mirrored exocrine score (r = 0.48; P = 0.01). When stratified by endocrine phenotype, cells with granules of heterogeneous morphology had higher scores than α, ß, and δ cells (P < 0.0001). Cells of mixed endocrine-exocrine morphology were observed in association with increased NPESS (P = 0.02). Islet isolation was associated with improved NPESS (in situ: 8.39 ± 0.77 [Mean ± SD]; postisolation: 5.44 ± 0.31; P = 0.04). CONCLUSIONS: NPESS provides a robust method for semiquantitative scoring of subcellular ultrastructural changes in human pancreatic endocrine cells in situ and following islet isolation with utility for unbiased evaluation of acute stress in organ transplantation research.

Hepatitis B surface antigen: relation to hepatitis B replication parameters in HBeAg-negative chronic hepatitis B.

BACKGROUND & AIMS: Translation of HBsAg depends on transcription of the appropriate mRNAs from cccDNA, but its relation to other hepatitis B virus (HBV) replication parameters is not known, inasmuch as integrated sequences of HBV-DNA may also contribute to its serum levels, especially in HBeAg-negative chronic hepatitis B (CHB) patients. METHODS: We investigated HBsAg serum levels, its hepatocellular expression, and their relationship to HBV replicative- and host-response parameters before treatment in 54 HBeAg-negative CHB patients and in 15 of them after 40.1±33.3months of virological response on oral antiviral (NUC) therapy also. Liver cccDNA and HBV-DNA quantitation, HBsAg- and HBcAg-immunostaining were performed in the same needle biopsy material, while serum HBsAg and HBV-DNA levels were measured in samples drawn on the day of liver biopsy. RESULTS: In untreated patients, serum HBsAg correlated positively with HBsAg-positive hepatocytes/mm(2) (p=0.003) and weakly with serum HBV-DNA, but not with cccDNA, liver HBV-DNA, HBcAg-positive hepatocytes/mm(2), or ALT. cccDNA correlated significantly with liver HBV-DNA (p<0.00001), ALT (p=0.001), and serum HBV-DNA levels (p=0.012) but not with liver HBsAg or HBcAg. Antiviral therapy decreased serum HBsAg levels by 79.6% (p=0.012) and liver HBV-DNA by 84.4% (p=0.026) in paired comparisons and, as expected, significantly decreased serum HBV-DNA and ALT levels, but not cccDNA. CONCLUSIONS: In untreated HBeAg-negative CHB, serum HBsAg levels reflect liver HBsAg, but not cccDNA or liver HBV-DNA, suggesting that they are not solely dependent on the replicative cycle of HBV. Effective NUC therapy for 3.34 years significantly lowers serum HBsAg and liver HBV-DNA, but not cccDNA.

Editorial: Exercise for NAFLD: does intensity matter?

Currently, there are no established means for the prevention or treatment of non-alcoholic steatohepatitis (NASH) despite our increasing understanding of nonalcoholic fatty liver disease (NAFLD) pathogenesis implicating insulin resistance (IR) as a key factor and highlighting the central role of lipotoxic liver injury in the development of NASH. Lifestyle interventions aiming at decreasing IR and preventing lipotoxicity, including weight loss, diet, and physical exercise, are in the frontline for NASH patient management. Physical activity may ameliorate IR, maintain weight loss, and improve liver histology in NASH patients. However, there are no recognized criteria for the optimal intensity, duration, or total volume of exercise needed to obtain these beneficial effects. In this issue of the American Journal of Gastroenterology, Kistler and colleagues show that vigorous, but not moderate exercise, nor total duration or volume of physical activity, is related to decreased odds of having NASH or advanced fibrosis. Prospective studies using the objective criteria of physical activity, addressing the role of concurrent weight loss, and assessing individual histological features are needed to further clarify the effects of exercise intensity on NAFLD histology.

Inflammatory myoglandular polyp as a cause of recurrent abdominal pain in a patient with quiescent Crohn's disease.

Chronic inflammation of the intestinal mucosa has been associated with the appearance of inflammatory polyps or pseudopolyps. Among the distinct categories of inflammatory polyps are inflammatory myoglandular polyps (IMGP) usually found in the colorectum. Only one case of IMGP in the terminal ileus has been described since their first description. We report the first case of an inflammatory polyp with both hyperplastic and myoglandular histological characteristics, in the terminal ileum of a patient with quiescent Crohn's disease, causing recurrent intussusception.

Tumor mutation burden testing: a survey of the International Quality Network for Pathology (IQN Path).

While tumour mutation burden (TMB) is emerging as a possible biomarker for immune-checkpoint inhibitors (ICI), methods for testing have not been standardised as yet. In April 2019, the International Quality Network for Pathology (IQN Path) launched a survey to assess the current practice of TMB testing. Of the 127 laboratories that replied, 69 (54.3%) had already introduced TMB analysis for research purposes and/or clinical applications. Fifty laboratories (72.5%) used targeted sequencing, although a number of different panels were employed. Most laboratories tested formalin-fixed paraffin-embedded material (94.2%), while 18/69 (26%) tested also cell-free DNA. Fifty-five laboratories used both single nucleotide variants and indels for TMB calculation; 20 centers included only non-synonymous variants. In conclusion, the data from this survey indicate that multiple global laboratories were capable of rapidly introducing routine clinical TMB testing. However, the variability of testing methods raises concerns about the reproducibility of results among centers.

Adiponectin, Leptin, and IGF-1 Are Useful Diagnostic and Stratification Biomarkers of NAFLD.

Background: Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disease where liver biopsy remains the gold standard for diagnosis. Here we aimed to evaluate the role of circulating adiponectin, leptin, and insulin-like growth factor 1 (IGF-1) levels as non-invasive NAFLD biomarkers and assess their correlation with the metabolome. Materials and Methods: Leptin, adiponectin, and IGF-1 serum levels were measured by ELISA in two independent cohorts of biopsy-proven obese NAFLD patients and healthy-liver controls (discovery: 38 NAFLD, 13 controls; validation: 194 NAFLD, 31 controls) and correlated with clinical data, histology, genetic parameters, and serum metabolomics. Results: In both cohorts, leptin increased in NAFLD vs. controls (discovery: AUROC 0.88; validation: AUROC 0.83; p < 0.0001). The leptin levels were similar between obese and non-obese healthy controls, suggesting that obesity is not a confounding factor. In the discovery cohort, adiponectin was lower in non-alcoholic steatohepatitis (NASH) vs. non-alcoholic fatty liver (AUROC 0.87; p < 0.0001). For the validation cohort, significance was attained for homozygous for PNPLA3 allele c.444C (AUROC 0.63; p < 0.05). Combining adiponectin with specific serum lipids improved the assay performance (AUROC 0.80; p < 0.0001). For the validation cohort, IGF-1 was lower with advanced fibrosis (AUROC 0.67, p < 0.05), but combination with international normalized ratio (INR) and ferritin increased the assay performance (AUROC 0.81; p < 0.01). Conclusion: Serum leptin discriminates NAFLD, and adiponectin combined with specific lipids stratifies NASH. IGF-1, INR, and ferritin distinguish advanced fibrosis.

Tityus: a forgotten myth of liver regeneration.

The ancient Greek myth of Tityus is related to liver regeneration in the same way as the well known myth of Prometheus is. Depictions of the punishment of Prometheus are frequently used by lecturers on liver regeneration; however, Tityus remains unknown despite the fact that he received the same punishment and his myth could also be used as a paradigm for the organ's extraordinary ability to regenerate. Nevertheless, there is no convincing evidence that ancient Greeks had any specific knowledge about liver regeneration, a concept introduced in the early 19th century. We describe and analyze the myth of Tityus and compare it to the myth of Prometheus. We also explore artistic and literary links and summarize recent scientific data on the mechanisms of liver regeneration. Finally, we highlight links of the legend of Tityus with other sciences.

Serum apoptotic caspase activity in chronic hepatitis C and nonalcoholic Fatty liver disease.

BACKGROUND: Apoptotic caspases are substantially activated in liver and serum caspase activity has been suggested as a marker of early liver injury. AIM: To investigate whether serum levels of caspase-generated fragments of cytokeratin-18 are associated with the severity of histologic lesions in chronic hepatitis C virus (HCV) infection and nonalcoholic fatty liver disease (NAFLD). METHODS: We included 134 patients with chronic HCV infection and 58 patients with NAFLD, who consecutively underwent liver biopsy, and 40 healthy controls. Caspase-generated cytokeratin-18 fragment levels were blindly measured in stored serum samples. RESULTS: Median cytokeratin-18 fragment levels were lower in HCV-positive patients with minimal/mild than patients with moderate/severe histologic lesions (174 U/L vs. 223 U/L, P<0.001) offering moderate accuracy for differentiation between the 2 groups (c-statistic: 0.74). Cytokeratin-18 fragments levels were lower in healthy subjects (148 U/L) than patients with simple fatty liver (174 U/L, P=0.013) than patients with nonalcoholic steatohepatitis (355 U/L, P<0.001) offering excellent diagnostic accuracy for differentiation between the 2 latter groups (c-statistic: 0.87). CONCLUSIONS: Serum apoptotic caspase activity is associated with the severity of liver histologic lesions in both chronic HCV infection and NAFLD, but it has excellent diagnostic accuracy in NAFLD and moderate accuracy in chronic HCV patients.