Therapeutic manipulation of the microbiome in liver disease.

Myriad associations between the microbiome and various facets of liver physiology and pathology have been described in the literature. Building on descriptive and correlative sequencing studies, metagenomic studies are expanding our collective understanding of the functional and mechanistic role of the microbiome as mediators of the gut-liver axis. Based on these mechanisms, the functional activity of the microbiome represents an attractive, tractable, and precision medicine therapeutic target in several liver diseases. Indeed, several therapeutics have been used in liver disease even before their description as a microbiome dependent approach. To bring successful microbiome-targeted and -inspired therapies to the clinic, a comprehensive appreciation of the different approaches to influence, collaborate with, or engineer the gut microbiome to coopt a disease relevant function of interest in the right patient is key. Herein, we describe the various levels at which the microbiome can be targeted - from prebiotics, probiotics, synbiotics and antibiotics, to microbiome reconstitution and precision microbiome engineering. Assimilating data from preclinical animal models, human studies as well as clinical trials, we describe the potential for and rationale behind studying such therapies across several liver diseases, including metabolic dysfunction associated steatotic liver disease, alcohol associated liver disease, cirrhosis, and hepatic encephalopathy, as well as liver cancer. Lastly, we discuss lessons learnt from previous attempts at developing such therapies, the regulatory framework that needs to be navigated and the challenges that remain.

Macrophage RAGE activation is proinflammatory in NASH.

Intrahepatic macrophages in nonalcoholic steatohepatitis (NASH) are heterogenous and include proinflammatory recruited monocyte-derived macrophages. The receptor for advanced glycation endproducts (RAGE) is expressed on macrophages and can be activated by damage associated molecular patterns (DAMPs) upregulated in NASH, yet the role of macrophage-specific RAGE signaling in NASH is unclear. Therefore, we hypothesized that RAGE-expressing macrophages are proinflammatory and mediate liver inflammation in NASH. Compared with healthy controls, RAGE expression was increased in liver biopsies from patients with NASH. In a high-fat, -fructose, and -cholesterol-induced (FFC)-induced murine model of NASH, RAGE expression was increased, specifically on recruited macrophages. FFC mice that received a pharmacological inhibitor of RAGE (TTP488), and myeloid-specific RAGE KO mice (RAGE-MKO) had attenuated liver injury associated with a reduced accumulation of RAGE+ recruited macrophages. Transcriptomics analysis suggested that pathways of macrophage and T cell activation were upregulated by FFC diet, inhibited by TTP488 treatment, and reduced in RAGE-MKO mice. Correspondingly, the secretome of ligand-stimulated BM-derived macrophages from RAGE-MKO mice had an attenuated capacity to activate CD8+ T cells. Our data implicate RAGE as what we propose to be a novel and potentially targetable mediator of the proinflammatory signaling of recruited macrophages in NASH.

Extracellular Vesicles in Hepatobiliary Health and Disease.

Extracellular vesicles (EVs) are membrane-bound nanoparticles released by cells and are an important means of intercellular communication in physiological and pathological states. We provide an overview of recent advances in the understanding of EV biogenesis, cargo selection, recipient cell effects, and key considerations in isolation and characterization techniques. Studies on the physiological role of EVs have relied on cell-based model systems due to technical limitations of studying endogenous nanoparticles in vivo . Several recent studies have elucidated the mechanistic role of EVs in liver diseases, including nonalcoholic fatty liver disease, viral hepatitis, cholestatic liver disease, alcohol-associated liver disease, acute liver injury, and liver cancers. Employing disease models and human samples, the biogenesis of lipotoxic EVs downstream of endoplasmic reticulum stress and microvesicles via intracellular activation stress signaling are discussed in detail. The diverse cargoes of EVs including proteins, lipids, and nucleic acids can be enriched in a disease-specific manner. By carrying diverse cargo, EVs can directly confer pathogenic potential, for example, recruitment and activation of monocyte-derived macrophages in NASH and tumorigenicity and chemoresistance in hepatocellular carcinoma. We discuss the pathogenic role of EVs cargoes and the signaling pathways activated by EVs in recipient cells. We review the literature that EVs can serve as biomarkers in hepatobiliary diseases. Further, we describe novel approaches to engineer EVs to deliver regulatory signals to specific cell types, and thus use them as therapeutic shuttles in liver diseases. Lastly, we identify key lacunae and future directions in this promising field of discovery and development. © 2023 American Physiological Society. Compr Physiol 13:4631-4658, 2023.

Assessment of Lipotoxic Endoplasmic Reticulum (ER) Stress in Nonalcoholic Steatohepatitis (NASH).

Hepatocyte lipotoxicity is a hallmark of nonalcoholic steatohepatitis (NASH), and lipid induced liver injury occurs, in part, via activation of endoplasmic reticulum (ER) stress. Consequently, the unfolded protein response (UPR) is initiated, driven by three key ER transmembrane proteins, resulting in downstream responses that are dynamic and interconnected. Thus, careful interrogation of these pathways is required to investigate the complex role of ER stress in NASH. Herein, we describe different mechanisms of, and in vitro assays for assessment of lipotoxic ER stress in mouse hepatocytes.

Circulating extracellular vesicles are a biomarker for NAFLD resolution and response to weight loss surgery.

There is increasing interest in the development of minimally invasive biomarkers for the diagnosis and prognosis of NAFLD via extracellular vesicles (EV). Plasma EVs were isolated by differential ultracentrifugation and quantified by nanoparticle tracking analysis from pre (n = 28) and post (n = 28) weight loss patients. In the pre weight loss group 22 had NAFLD. Nanoplasmon enhanced scattering (nPES) of gold nanoparticles conjugated to hepatocyte-specific antibodies was employed to identify hepatocyte-specific EVs. Complex lipid panel and targeted sphingolipids were performed. Logistic regression analysis was used to identify predictors of NAFLD. Plasma levels of EVs and hepatocyte-derived EVs are dynamic and decrease following NAFLD resolution due to weight loss surgery. Hepatocyte-derived EVs correlate with steatosis in NAFLD patients and steatosis and inflammation in NASH patients. Plasma levels of small EVs correlate with EV sphingolipids in patients with NASH. Hepatocyte-derived EVs measured by the nPES assay could serve as a point-of-care test for NAFLD.

Hepatic steatosis and steatohepatitis: a functional meta-analysis of sex-based differences in transcriptomic studies.

BACKGROUND: Previous studies have described sex-based differences in the epidemiological and clinical patterns of non-alcoholic fatty liver disease (NAFLD); however, we understand relatively little regarding the underlying molecular mechanisms. Herein, we present the first systematic review and meta-analysis of NAFLD transcriptomic studies to identify sex-based differences in the molecular mechanisms involved during the steatosis (NAFL) and steatohepatitis (NASH) stages of the disease. METHODS: Transcriptomic studies in the Gene Expression Omnibus database were systematically reviewed following the PRISMA statement guidelines. For each study, NAFL and NASH in premenopausal women and men were compared using a dual strategy: gene-set analysis and pathway activity analysis. Finally, the functional results of all studies were integrated into a meta-analysis. RESULTS: We reviewed a total of 114 abstracts and analyzed seven studies that included 323 eligible patients. The meta-analyses identified significantly altered molecular mechanisms between premenopausal women and men, including the overrepresentation of genes associated with DNA regulation, vinculin binding, interleukin-2 responses, negative regulation of neuronal death, and the transport of ions and cations in premenopausal women. In men, we discovered the overrepresentation of genes associated with the negative regulation of interleukin-6 and the establishment of planar polarity involved in neural tube closure. CONCLUSIONS: Our meta-analysis of transcriptomic data provides a powerful approach to identify sex-based differences in NAFLD. We detected differences in relevant biological functions and molecular terms between premenopausal women and men. Differences in immune responsiveness between men and premenopausal women with NAFLD suggest that women possess a more immune tolerant milieu, while men display an impaired liver regenerative response.

A natural language-based tool for diagnosis of serrated polyposis syndrome.

BACKGROUND AND AIMS: Serrated polyposis syndrome (SPS) is common but under-recognized and is associated with an increased risk of colorectal cancer. The diagnosis is based on the World Health Organization (WHO) criteria and is inclusive of the cumulative number of lifetime serrated polyps. We used natural language processing (NLP) to extract colonoscopy and pathology data from the electronic medical record (EMR). The aim of this study was to assess feasibility of using an NLP-based SPS tool to identify patients with SPS. METHODS: NLP was used to extract data from 323,494 colonoscopies performed in 255,074 distinct patients between August 1998 and March 2016 to identify individuals who met SPS criteria. The accuracy of diagnosis of SPS was assessed by manual review of the EMR. RESULTS: Of 255,074 patients, 71 were identified as meeting 1 WHO criteria for SPS. Manual review confirmed the diagnosis of SPS to be accurate in 66 cases (93%). Erroneous diagnosis in the remaining 5 cases occurred because of duplicate polyp data by NLP extraction. Only 25 of 66 patients (38%) were diagnosed with SPS by a clinician in the EMR. Of these, SPS was diagnosed by NLP at least 2 years before the clinician in 5 of 25 patients (20%). CONCLUSIONS: In a large cohort, NLP accurately identified SPS in over 90% of cases, most of which were not previously recognized. NLP can assist in collating colonoscopy and pathology data across multiple procedures in the same patient to make an accurate and earlier diagnosis of SPS.

Pathogenesis of Nonalcoholic Steatohepatitis: An Overview.

Nonalcoholic fatty liver disease (NAFLD) is a heterogeneous group of liver diseases characterized by the accumulation of fat in the liver. The heterogeneity of NAFLD is reflected in a clinical and histologic spectrum where some patients develop isolated steatosis of the liver, termed nonalcoholic fatty liver, whereas others develop hepatocyte injury, ballooning, inflammation, and consequent fibrosis, termed nonalcoholic steatohepatitis (NASH). Systemic insulin resistance is a major driver of hepatic steatosis in NAFLD. Lipotoxicity of accumulated lipids along with activation of the innate immune system are major drivers of NASH. Lipid-induced sublethal and lethal stress culminates in the activation of inflammatory processes, such as the release of proinflammatory extracellular vesicles and cell death. Innate and adaptive immune mechanisms involving macrophages, dendritic cells, and lymphocytes are central drivers of inflammation that recognize damage- and pathogen-associated molecular patterns and contribute to the progression of the inflammatory cascade. While the activation of the innate immune system and the recruitment of proinflammatory monocytes into the liver in NASH are well known, the exact signals that lead to this remain less well defined. Further, the contribution of other immune cell types, such as neutrophils and B cells, is an area of intense research. Many host factors, such as the microbiome and gut-liver axis, modify individual susceptibility to NASH. In this review, we discuss lipotoxicity, inflammation, and the contribution of interorgan crosstalk in NASH pathogenesis.

Development of an Automated Algorithm to Generate Guideline-based Recommendations for Follow-up Colonoscopy.

BACKGROUND AND AIMS: Physician adherence to published colonoscopy surveillance guidelines varies. We aimed to develop and validate an automated clinical decision support algorithm that can extract procedure and pathology data from the electronic medical record (EMR) and generate surveillance intervals congruent with guidelines, which might increase physician adherence. METHODS: We constructed a clinical decision support (CDS) algorithm based on guidelines from the United States Multi-Society Task Force on Colorectal Cancer. We used a randomly generated validation dataset of 300 outpatient colonoscopies performed at the Cleveland Clinic from 2012 through 2016 to evaluate the accuracy of extracting data from reports stored in the EMR using natural language processing (NLP). We compared colonoscopy follow-up recommendations from the CDS algorithm, endoscopists, and task force guidelines. Using a testing dataset of 2439 colonoscopies, we compared endoscopist recommendations with those of the algorithm. RESULTS: Manual review of the validation dataset confirmed the NLP program accurately extracted procedure and pathology data for all cases. Recommendations made by endoscopists and the CDS algorithm were guideline-concordant in 62% and 99% of cases, respectively. Discrepant recommendations by endoscopists were earlier than recommended in 94% of the cases. In the testing dataset, 69% of endoscopist and NLP-CDS algorithm recommendations were concordant. Discrepant recommendations by endoscopists were earlier than guidelines in 91% of cases. CONCLUSIONS: We constructed and tested an automated CDS algorithm that can use NLP-extracted data from the EMR to generate follow-up colonoscopy surveillance recommendations based on published guidelines.

The time required to diagnose and treat lung cancer in Delhi, India: an updated experience of a public referral center

Background: Delays that postpone the evaluation and management of malignancy may lead to considerable morbidity. The primary objective of this study was to assess the time required to diagnose and treat lung cancer at an Indian public referral center that predominantly serves lower-income patients. Methods: A review of patients diagnosed with lung cancer between January 2008 and December 2016 was completed. We computed the median time intervals and inter-quartile ranges between symptom onset, definitive diagnostic investigation, confirmed histologic diagnosis, and chemotherapy initiation. Median intervals were correlated with baseline demographics and disease characteristics using Kruskal-Wallis test. Results: One thousand, three hundred and-seventy patients were selected. A majority (94.5%) with non-small cell lung cancer were diagnosed with advanced disease. After developing symptoms, patients required 101 [56­168] days to undergo a definitive diagnostic study, 107 [60­173] days to confirm a diagnosis, and 126 [85­196.8] days to initiate treatment. Patients who were previously treated for tuberculosis required more time to receive chemotherapy compared to those who were not (187 [134­261.5] days vs. 113 [75­180] days, p < 0.0001). A specialty Lung Cancer Clinic was implemented in 2012, and the mean referrals per month increased nearly four-fold (p < 0.0001), but the time required to administer treatment was not shortened. Conclusion: Among lower-income Indian patients, the most prominent delays occur prior to diagnosis. Efforts should be directed toward encouraging physicians to maintain a high index of clinical suspicion and educating patients to report concerning symptoms as early as possible.