Characterization of the Intrahepatic Inflammatory Microenvironment in Patients With Non-alcoholic Steatohepatitis

Characterization of the Intrahepatic Inflammatory Microenvironment in Patients With Non-alcoholic Steatohepatitis by Transcriptomics, Immunophenotyping and Functional Proteomics: Profil-NASH

Non-alcoholic fatty liver disease (NAFLD) is a nosological entity that groups together non-alcoholic fatty liver (NAFL) and non-alcoholic steatohepatitis (NASH). Unlike NAFL, NASH is characterized by intrahepatic inflammation, and is solely at risk of progression to cirrhosis and hepatocellular carcinoma (HCC).

It is currently estimated that NAFLD affects approximately 25% of the world's adult population, and its incidence is rising in all regions of the world. Nevertheless, of all patients with NAFLD, only ~25% have NASH.

Identifying patients with NASH is therefore crucial, determining the need for follow-up to detect the onset of fibrosis and/or HCC, and eventual access to therapeutic trials. Furthermore, intrahepatic inflammation, the initial driver of NASH, appears to play an important role in the development of fibrosis and HCC, which can occur in the absence of cirrhosis in these patients. However, few studies have been carried out in humans to date, with data mainly coming from mouse models.

An innovative technique, Fine-Needle Aspiration (FNA), enables to obtain cells from the liver compartment, including large numbers of immune cells. In participants with NAFLD and indication of liver biopsy, a FNA will also be performed. Forty patients will be included, with ~75% of NASH and ~25% of NAFL expected. The investigators will study the phenotypic and functional characteristics of human intrahepatic inflammatory cells obtained by the FNA with different innovative techniques (RNAseq, multiparameter immunophenotyping, single-cell secretome and phosphoproteome). Peripheral Blood Mononuclear Cells and circulating microRNAs, known to regulate immune responses, will also be analysed.

The hypothesis of Profile-NASH is that intrahepatic inflammatory profiles differ between NASH and NAFL, and is associated with fibrosis progression and carcinogenesis.

This pilot study, based on high-definition technologies, will provide precise new insights into the quality of intrahepatic inflammation and the mechanisms favoring the transition from NAFL to NASH and its progression. Precise analysis of the intrahepatic inflammatory microenvironment will enable the investigators to identify new players in the pathogenesis of NASH, and potential future therapeutic targets.

Inclusion Criteria:

• patient with a clinical diagnosis of NAFLD: steatosis detected on imaging and exclusion of secondary causes of steatosis (drugs, genetics, alcohol consumption >30 g/d in men and 20 g/d in women, chronic viral infection), in the absence or presence of an associated metabolic syndrome
• and with significant liver fibrosis (≥ F2) on at least one non-invasive test (FibroScan®, Fibrometer®, NAFLD Fibrosis Score);
• Patient of legal age (age ≥ 18 years);
• Patient willing to undergo liver biopsy ;
• Patient consenting to inclusion in the study after being informed and obtaining written consent;
• Patient affiliated to a social security scheme.

Exclusion Criteria:

• Decompensated cirrhosis or clinically significant portal hypertension (clinical, radiological or endoscopic signs of portal hypertension; presence of hepatocellular insufficiency);
• Secondary causes of steatosis, including chronic viral hepatitis, drugs, excessive alcohol consumption according to World Health Organization (WHO) criteria (> 30 g/d in men and 20 g/d in women), genetic mutations;
• Any other cause of liver disease: genetic hemochromatosis, autoimmune liver disease, etc. (non-exhaustive list);
• Presence of HCC at the time of inclusion;
• Contraindications to liver biopsy (identical to those for FNA): coagulation disorders, biliary tract dilatation, intrahepatic tumor;
• Pregnant, parturient or breast-feeding women;
• Persons deprived of their liberty by judicial or administrative decision;
• adults under legal protection (guardianship, curators).