Thursday, March 29, 2018

HPHS Journal Watch: Jan/Feb 2018

Am J Gastroenterol
The rate of publication of new guidelines is accelerating – the European Association of Liver Disease is about to publish their updated guidelines for alcoholic liver disease (ALD) later this year.  This paper includes guidance on the role of liver biopsies in the management of ALD. One of the key statements is that “Liver biopsy is not routinely recommended for diagnosis of alcoholic fatty liver disease. However, liver biopsy and non-invasive tools of fibrosis may be considered for diagnosis of steatohepatitis and/or liver fibrosis”. This cautiously worded indication is focused in a subsequent one: “In patients with suspected AH, a transjugular liver biopsy is recommended when the clinical diagnosis is confounded by another liver disease etiology or there is uncertainty on alcohol consumption history.”  Having said this, the authors require histology for a definitive diagnosis of alcoholic hepatitis. This paradox is not unique to this paper. Tellingly, they quote a recent paper (J Hepatol 2017 ; 66 : 610 – 8) which showed that one-third of patients with clinically early disease have advanced fibrosis/cirrhosis on histology.
The authors refer to the paper by Altamirano ( Gastroenterology2014; 146 : 1231 – 9 ), one of the most evidence based papers in this field, but do not elevate it to the level of a recommendation. There are a number of nuggets such as the inflammation is more marked in patients with the alcohol withdrawal syndrome than those without it (Alcohol Clin Exp Res 2004 ; 28 : 131 – 6)

Chromosomal instability (CIN) is the most common promoter of hepatocellular carcinoma (HCC) progression. It does so by increasing tumour heterogeneity, drug resistance and immune escape. The DNA damage response protein checkpoint kinase 2 (Chk2) expression was evaluated in an animal model (of diethylnitrosamine-induced HCC) and also in two cohorts of human HCC specimens. In the animal model it was shown that overexpression/phosphorylation and mislocalisation within the mitotic spindle of Chk2 contributes to the induction of lagging chromosomes. The forkhead-associated domain of Chk2 was required for Chk2 mislocalisation to mitotic structures. A cohort and independent validation cohort show a strong cytoplasm to nuclear Chk2 translocation in a subset of patients with HCC.
This study suggests Chk2 is a putative biomarker that can be used to detect DNA damage and chromosomal instability in HCC providing a valuable support, for prognosis, and for identifying patients who are likely to have a response to therapy.

Acute liver failure (ALF) is characterised by overwhelming hepatocyte death and liver inflammation and a  massive infiltration of myeloid cells into the  necrotic areas. This study investigated the impact of Mer tyrosine kinase (MerTK) during ALF and how the microenvironmental mediator, secretory leucocyte protease inhibitor (SLPI), governs this response. The human work demonstrated a significant expansion of resolution-like MerTK+HLA-DRhigh macrophages in circulatory and tissue compartments of patients with ALF. Compared with WT mice which show an increase of MerTK+MHCIIhigh macrophages during the resolution phase in ALF, APAP-treated Mer−/− mice exhibit persistent liver injury and inflammation. Both in vitro and in APAP-treated mice, SLPI reprogrammes myeloid cells towards resolution responses through induction of a MerTK+HLA-DRhigh phenotype which promotes neutrophil apoptosis and their subsequent clearance.
In this paper, the authors have clearly demonstrated the liver protective, MerTK+, macrophage phenotype that evolves during the resolution phase following ALF and that it represents a potential immunotherapeutic target to promote resolution responses following acute liver injury. For the histopathologist, the immunohistochemical identification of this subset of macrophages has the potential to identify patients more likely to have an inflammatory response that will resolve.  This may be important in a wide range of liver diseases.

The most recent evidence based official recommendations of the American Association for the Study of Liver Diseases (AASLD) on the surveillance, diagnosis, and treatment of hepatocellular carcinoma (HCC) occurring in the setting of adults with cirrhosis was recently released using for the first time the Grading of Recommendation Assessment, Development and Evaluation (GRADE) approach. The developers of the guidelines addressed 10 crucial scenarios that health care providers frequently encounters in the management of patients with HCC.  HCC is unique among malignancies as its diagnosis can be obtained noninvasively, particularly in a cirrhotic patient, and treatment can be initiated based on imaging alone, without assenting biopsy. Find out the latest AASLD recommendations addressing the role of biopsy in adults with cirrhosis and an indeterminate hepatic nodule; and other key clinical situations.

Am J Clin Pathol
"Whole Slide Imaging diagnosis is feasible and accurate" concludes French investigators, Villa et al., in a concordance study between glass slide and digital slide diagnosis published in the latest issue of AJCP.  Concordance was reported in 87% of cases, minor discordance in 10%, and major discordance with therapeutic impact in 2.5%.  Causes of major discordance were attributed to technical issues including poor image quality and logistics issues.  Recent works have shown these factors were no longer a problem indicating rapid improvement of the technology.  Clearly, standardization and quality control need to be implemented in order for whole slide imaging diagnosis to be successfully incorporated in the laboratory workflow.

Mod Pathol
The authors address the role of bile ductular reaction as a prognostic factor in NASH progression, based on periportal or centrizonal location. They studied liver biopsies from 90 patients (over 17 years of age) with primary diagnosis of NASH. 47 patients, who received no treatments, had paired liver biopsies, at least 12 months apart. They excluded cases with other superimposed chronic liver diseases such as HCV, alcohol, VOD, cirrhosis and transplant. Centrilobular ductular reaction was identified in 90% of patients and 38% of centrilobular zones. The prevalence of centrilobular ductular reaction increased in correlation with NASH grade and fibrosis stage. The frequency of centrilobular ductular reaction in the initial biopsies was significantly higher in the patients who showed fibrosis progression (P=0.02). They concluded that presence of centrilobular ductular reaction correlates significantly with increasing necroinflammatory activity and fibrosis stage and can predict progression of fibrosis in subsequent biopsies.

Recently, fibrolamellar carcinomas were found to harbor a characteristic somatic gene fusion, DNAJB1-PRKACA. They developed a break-apart FISH assay to detect this fusion and examine its diagnostic performance. 104 cases that were classified histologically as typical of fibrolamellar carcinoma, 12 cases as 'possible fibrolamellar carcinoma' and 8 cases as 'unlikely to be fibrolamellar carcinoma' were used. PRKACA FISH was positive for rearrangement in 102 of 103 (99%) typical fibrolamellar carcinomas, 9 of 12 'possible fibrolamellar carcinomas' and 0 of 8 cases 'unlikely to be fibrolamellar carcinomas'. Within the morphologically typical group of fibrolamellar carcinomas, they also identified two tumors with unusual FISH patterns in the morphologically typical fibrolamellar carcinoma. Both had DNAJB1-PRKACA fusion but one had an additional amplification of the fusion gene and one had heterozygous deletion of the normal PRKACA locus. PRKACA FISH was negative in 88 conventional HCC. They conclude that PRKACA rearrangement is a clinically useful tool to confirm the diagnosis of fibrolamellar carcinoma, with high sensitivity and specificity.

J Gastroenterol Hepatol
The authors screened the effectiveness of hepatic steatosis index (HSI) and fatty liver index (FLI), as well as their link to metabolic syndrome in type 1 diabetes in an observational pilot study. FLI and HSI were calculated for 201 patients with type 1 diabetes. 40 patients with FLI/HSI values corresponding to different risk of liver steatosis were invited for liver MRI. Twelve (30.0%) patients had liver steatosis. For FLI, sensitivity was 90%; specificity, 74%; positive likelihood ratio, 3.46; negative likelihood ratio, 0.14; PPV, 0.64; and NPV, 0.93. For HSI, sensitivity was 86%; specificity, 66%; positive likelihood ratio, 1.95; negative likelihood ratio, 0.21; PPV, 0.50; and NPV, 0.92. HSI correlated with waist circumference and C-reactive protein. FLI ≥ 60 and HSI ≥ 36 were significantly associated with metabolic syndrome and nephropathy. They concluded that FLI, can serve as surrogate markers for liver fat content and metabolic syndrome in type 1 diabetes.

2017 update on the guidelines for the assessment and management of non-alcoholic fatty liver disease (NAFLD) by the Asia–Pacific Working Party on NAFLD.

2017 update on the guidelines for the assessment and management of non-alcoholic fatty liver disease (NAFLD) by the Asia–Pacific Working Party on NAFLD

Liver macrophages are activated in chronic hepatitis B virus (CHB) infection and play a pivotal role in hepatic inflammation and fibrosis. However, their role during antiviral treatment is unclear. The soluble (s) macrophage activation markers, sCD163 and mannose receptor (sMR), are released during liver damage, and their serum levels reflect liver disease severity and portal hypertension. The authors investigated the associations between sCD163 (IHC and serum level) and sMR (ELISA) and histopathological activity (using ISHAK score) and fibrosis in pre and post antiviral treated CHB patients.The had 254 liver biopsies, 71 patientswith pre and post treatment biopsies. They found elevated sCD163 and sMR correlated with histologic inflammatory activity and fibrosis. Both were reduced with antiviral treatment.

This is a review article on the significance of microvascular invasion in surgical specimen and correlation with high post-operative HCC recurrence.

Am J Surg Pathol
The investigators reviewed 36 hepatobiliary mucinous cystic neoplasms (MCNs) with the requirement that ovarian type stroma (OS) be present to better determine its frequency among hepatic cysts, clinicopathologic characteristics, and risk for malignant transformation.  MCNs accounted for 36 of 241 cases (15%).  100% of cases occurred in women, 72% were located in the left lobe, mean size was 11.6 cm, mean age of patients was 51 years old, and median follow-up time was 18.5 months.  Only 2 of the 36 cases (5.5%) harbored high-grade dysplasia/carcinoma in situ (HGD/CIS); both cases of HGD/CIS also had associated foci of invasive carcinoma that measured 7 and 8 mm.  This study also found that the presence of OS and mucinous epithelium can be very focal.  In conclusion, this study adds further data to our understanding of hepatobiliary MCNs and the rarity of malignant transformation in these lesions.

This resident short review summarizes clinical, imaging, and histologic features, differential diagnosis, and treatment options for hepatic epithelioid hemangioendothelioma. 

Am J Pathol
The authors evaluated mucin 13 (MUC13) in hepatocellular carcinoma (HCC). Overexpression of MUC13 was detected in 74 of 168 HCCs (44%), and was significantly associated with tumor size, stage, encapsulation, venous invasion, and poor outcome. MUC13 promoted cellular G1/S phase transition by activating Wnt signaling, promoting nuclear translocation of β-catenin and up-regulation of downstream target genes.

Clin Gastroenterol Hepatol
This is a systemic review and meta-analysis aimed at characterizing racial and ethnic differences in the prevalence of NAFLD. It also looked at the differences in severity and prognosis of NAFLD among different ethnic groups. This article used NAFLD as diagnosed by biochemical, radiological, or histologic criteria per AASLD guidelines and assessed severity as presence of steatohepatitis and advances (stage F3-F4) fibrosis. Thirty-four studies totaling 368,569 patients showed that NAFLD is highest in Hispanics and lowest in Blacks but fibrosis was not significantly different among racial groups. Nearly 1 in 6 Americans have NAFLD and 30% had evidence of NASH with nearly 20% having evidence of advanced fibrosis. Clear differences in prognosis among ethnic groups were not identified. It is unclear why disparities exist, as diabetes and obesity are higher in black vs white, but white subjects have significantly higher risk of NAFLD. Similarly, further studies are needed to characterize the contribution of genetic factors, in particular PNPLA3, TM6SF2, and MBOAT single nucleotide polymorphisms vs. contribution of environmental factors.

The authors performed a competing-risk analysis to evaluate factors associated with survival of patients with HCC and developed a prognostic model based on features of HCC patients before liver transplantation. The training set comprised 1018 patients (western cohort) and a validation set of 341 consecutive patients (eastern cohort) who underwent liver transplantation for HCC.  In the competing-risk regression, the sum of tumor number and size and of log10 level of AFP were significantly associated with HCC-specific death (P < .001). Five-year cumulative incidence of non-HCC-related death was 8.6% in HCV-negative patients and 18.1% in HCV-positive patients. For patients with HCC to have a 70% chance of HCC-specific survival 5 years after transplantation, their level of AFP should be <200 ng/mL and the sum of number and size of tumors (in centimeters) should not exceed 7; if the level of AFP was 200-400 ng/mL, the sum of the number and size of tumors should be </= 5; if their level of AFP was 400-1000 ng/mL, the sum of the number and size of tumors should be </=4. In the validation set, the model identified patients who survived 5 years after liver transplantation with 0.721 accuracy (95% confidence interval, 0.648%-0.793%). The authors proposed that their model, based on patients’ level of AFP and HCC number and size, outperformed the Milan; University of California, San Francisco; Shanghai-Fudan; Up-to-7 criteria (P < .001); and AFP French model (P ¼ .044) to predict which patients will survive for 5 years after liver transplantation. To predict 5-year survival and risk of HCC-related death using an online calculator, please see

Development of hepatocellular carcinoma (HCC) is associated with alterations in the transforming growth factor-beta (TGF-β) signaling pathway, which regulates liver inflammation and can have tumor suppressor or promoter activities. Little is known about the roles of specific members of this pathway at specific of HCC development. The authors performed transcriptome analyses for a total of 488 HCCs that include data from The Cancer Genome Atlas. In addition, they screened 301 HCCs reported in the Catalogue of Somatic Mutations in Cancer and 202 from Cancer Genome Atlas for mutations in genome sequences. They found somatic mutations in at least 1 gene whose product is a member of TGF-β signaling pathway in 38% of HCC samples. SPTBN1 was mutated in the largest proportion of samples (12 of 202, 6%). Unsupervised clustering of transcriptome data identified a group of HCCs with activation of the TGF-β signaling pathway (increased transcription of genes in the pathway) and a group of HCCs with inactivation of TGF-β signaling (reduced expression of genes in this pathway). Patients with tumors with inactivation of TGF-β signaling had shorter survival times than patients with tumors with activation of TGF-β signaling (P = .0129). Patterns of TGF-β signaling correlated with activation of the DNA damage response and sirtuin signaling pathways. HepG2, SNU398, and SNU475 cells that expressed the D1089Y mutant or with knockdown of SPTBN1 had increased sensitivity to DNA crosslinking agents and reduced survival compared with cells that expressed normal SPTBN1 (controls). In summary, in genome and transcriptome analyses of HCC samples, they found mutations in genes in the TGF-b signaling pathway in almost 40% of samples. These correlated with changes in expression of genes in the pathways; up-regulation of genes in this pathway would contribute to inflammation and fibrosis, whereas downregulation would indicate loss of TGF-b tumor suppressor activity. Their findings indicate that therapeutic agents for HCCs can be effective, based on genetic features of the TGF-b pathway; agents that block TGF-b should be used only in patients with specific types of HCCs.

Hum Pathol
The role of BRCA1 and BRCA2 genes is mainly to maintain genome integrity in response to DNA damage through different mechanisms. Deregulation of BRCA1 and BRCA2 is associated with the development of tumor and altered sensitivity to chemotherapeutic agents. In this study, the authors determined protein expression of BRCA1 and BRCA2 in 4 digestive system cancers (gastric cancer, colorectal cancer, hepatocellular carcinoma, and pancreatic cancer) by immunohistochemistry on tissue microarrays. A total of 1546 samples of 4 types of cancer tissues, their matched adjacent nontumor tissues, and corresponding benign tissues were studied, respectively. Immunohistochemistry expression patterns of the 2 proteins and their correlation with patients' clinical parameters and overall survival were analyzed. The results showed that low expression of cytoplasmic BRCA1 and BRCA2 was commonly associated with advanced tumor–lymph node–metastasis stage, whereas high expression of nuclear BRCA1 was generally correlated with advanced tumor stages in these cancers. High expression of cytoplasmic BRCA1 and BRCA2 had significantly favorable overall survival in digestive system cancers; in contrast, BRCA1 nuclear expression usually predicted poor outcomes. They conclude that BRCA1 and BRCA2 could be used as clinicopathological biomarkers to evaluate the prognosis of digestive system cancers.

Patients with hepatocellular carcinoma (HCC) frequently have multiple anatomically distinct tumors. In these patients, multifocal HCC could represent intrahepatic metastases (IMs) of a single cancer or multicentric carcinogenesis (MC) with multiple independent neoplasms. To determine the frequency and clinical implications of these 2 possibilities, the authors performed histological and molecular analysis of 70 anatomically distinct HCCs from 24 patients. They assayed mutations in the TERT promoter region by Sanger sequencing and used next-generation sequencing to analyze the entire coding regions of 7 well-characterized HCC driver genes—based on shared or discordant mutations in these genes, we classified the HCCs in each patient as IM, MC, or indeterminate. Mutations in the TERT promoter were the most common alteration in their cohort, present in 71% of tumors analyzed. Mutations in the remaining genes occurred in less than 20% of analyzed tumors. They were able to determine the relatedness in 58% of the patients analyzed: MC occurred in 41% of patients, with 33% with exclusively MC and 8% with both MC and IM. IM occurred exclusively in 17% of patients, whereas the remainder were indeterminate. This study highlights the utility of molecular analyses to determine relatedness in multifocal HCC; however, targeted sequencing can only resolve this distinction in approximately 60% of patients with multifocal HCC.

The histone methyltransferase G9a (EHMT2) is a key enzyme for dimethylation of lysine 9 at histone 3 (H3K9me2), a suppressive epigenetic mark. G9a is over-expressed in tumor cells and contributes to cancer aggressiveness. The authors analyzed G9a expression in 68 BTC specimens and correlated the data with clinicopathological and survival data. They measured G9a expression in a panel of BTC cell lines and evaluated the cytotoxic effect of G9a inhibition in BTC cells using established small-molecule G9a inhibitors. G9a was considerably expressed in about half of BTC cases and was significantly associated with grading and tumor size. There were significant differences of G9a expression between growth type and tumor localization groups. G9a expression diametrically correlated with Vimentin (positive) and E-Cadherin (negative) expression. Importantly, survival analysis revealed G9a as a significant prognostic factor of poor survival in patients with BTC. In BTC cells, G9a and H3K9me2 were detectable in a cell line–dependent manner on mRNA and/or protein level, respectively. Treatment of BTC cells with established small-molecule G9a inhibitors resulted in reduction of cell viability as well as reduced G9a and H3K9me2 protein levels. The present study strongly suggests that G9a contributes to BTC carcinogenesis and may represent a potential prognostic factor as well as a therapeutic target.

Liver Transplantation
A growing body of literature is illuminating the relationship between platelets and tumor growth and metastasis. The authors of this paper stratify living donor liver transplant recipients into two groups, one with a low platelet count (<75 x 109 /L) and one with a platelet count of >75 x 109 /L. The platelet count was incorporated into the Milan criteria. Incorporation of the preoperative platelet count into the Milan criteria significantly improved HCC recurrence predictive power. HCC recurrence risk was higher in the high platelet group on both univariate and multivariate analysis. The authors conclude that the preoperative platelet count is an important host factor affecting HCC recurrence following living donor liver transplantation.

Calcineurin inhibitors (CNIs; cyclosporine and tacrolimus) are common agents used for immunosuppression following orthotopic liver transplantation (OLT). Neurotoxic side effects are well known to occur within the first week following OLT, but the long term effects have not been well documented. These authors studied 85 OLT patients and grouped them into CNI-free, CNI low-dose, and CNI standard-dose categories. 33 healthy patients adjusted for age, gender and education served as controls. Subjects underwent psychometric testing and cerebral MRI approximately 10 years following OLT. Patients receiving CNIs showed significantly worse visuospatial/constructional ability compared with controls. Patients with low-dose CNI therapy had an overall impaired cognitive function compared with controls. Patients treated with CNIs showed significantly more white matter hyperintensities (WMH) than CNI-free patients and controls. The authors conclude that OLT patients on longterm CNI therapy carry a significant risk of cognitive dysfunction, and patients may benefit from a change to CNI-free therapies.

This well-respected author provides an update and overview of the recent NAFLD guidance documents provided from the American Association for the Study of Liver Disease (AASLD) and the European Association for the Study of the Liver (EASL) with regards to issues pertaining to liver transplantation.

As the population ages and incidence of obesity rises, the projected rates of deceased donor liver utilization are projected to fall from the current rate of 78% to 44% by 2030. Judicious donor selection will become increasingly critical to avoid discarding potentially adequate grafts. The authors of this manuscript evaluate the safety of and rate of liver utilization with preretrieval (or prerecovery) liver biopsy (PLB) in extended criteria liver donors. Three organ procurement organizations (OPOs) were included and the outcomes measured included complications, preempted liver recovery, and successful liver transplantation. The PLB group had fewer complications. In the liver-only and propensity score-matched multiorgan donor subgroups, the PLB group had fewer complications and the preempted liver recovery rate was significantly higher without a decrease in liver transplantation rate. The authors conclude that in extended criteria liver donors, PLB is safe and decreases the rate of futile liver recovery without decreasing the overall liver transplantation rate. PLB, especially in liver-only donors, is likely to save costs to OPOs and transplant centers as well as improve efficiencies in organ allocation.

This is a nice review article on the usefulness of next generation sequencing in pediatric patients with suspected monogenic diseases.

The authors present and illustrate a great case to remind us that not all “infiltrating” perineural and intraneural glands are malignant.

The liver is the leading site of colorectal carcinoma metastasis. This review summarizes a detailed protocol for specimen handling and for the reporting of important histopathological features in liver resections for metastatic colorectal carcinoma.

Prepared by:
Daniela Allende MD (Editor), Cleveland Clinic
Robert Goldin MD; Imperial College, London
Cynthia Guy MD; Duke University
Grace Guzman MD;  University of Illinois
Mojgan Hosseini MD; University of California San Diego
Nafis Shafizadeh MD; Southern California Permanente Medical Group
Eric Yee MD; University of Arkansas for Medical Sciences
Maria Westerhoff MD; University of Michigan 

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