Fatty Liver and Pancreatic Steatosis

The purpose of this study is to evaluate the association between non-alcoholic fatty liver disease and non-alcoholic pancreatic steatosis and to describe cytokine levels in the context of a global epidemic of obesity and metabolic syndrome, which could represent a new treatment target in previously underdiagnosed patients with pancreatic insufficiency and pancreatic steatosis.

Patients over 18 years old with a diagnosis of MAFLD (Metabolic Dysfunction-Associated Fatty Liver Disease) will undergo the following assessments:

* Exocrine Pancreatic Insufficiency Questionnaire (PEI-Q)
* Measurement of weight, height, and abdominal circumference
* Fecal elastase (Fel-1)
* Pro-inflammatory and anti-inflammatory cytokines in serum
* Hydrogen and methane breath tests to evaluate small intestinal bacterial overgrowth (SIBO)
* Endoscopic ultrasound with sedation
* Fibroscan Exocrine pancreatic insufficiency (EPI) will be defined as a fecal elastase-1 (Fel-1) concentration of < 100 µg/g or Fel-1 between 100 and 200 µg/g with alterations in additional pancreatic pathology tests, such as serum albumin, vitamin E, vitamin D, vitamin A, folic acid, iron, transferrin, calcium, magnesium, and/or malnutrition identified through anthropometric measurements conducted by an expert nutritionist. Fel-1 ≥ 200 µg/g will be considered normal.

Additionally, for those patients with fecal elastase levels below 200 µg/g, the following measurements will be conducted:

* Proteinogram
* Vitamin E, vitamin D, vitamin A, vitamin K
* Folic acid, B12
* Calcium, magnesium, zinc
* Iron profile
* Nutritional assessment with anthropometry

For those with exocrine pancreatic insufficiency (EPI):

* IgG4
* Alpha-1 antitrypsin
* Endoscopic ultrasound (EUS) with biopsies

• Fecal elastase (Diagnostic test): **Stool Samples** To investigate exocrine pancreatic insufficiency (EPI), a stool sample will be requested from all patients for fecal elastase (Fel-1) analysis. Participants will be instructed to collect their stool sample in a sterile, disposable plastic container and submit it to the Gastroenterology Chemistry Laboratory (Litwin Laboratory) for processing and analysis. Based on previously published reports, samples will be stored refrigerated at 4-8 °C for no more than 48 hours. The concentrations of Fel-1 in all samples will be measured using a commercially available enzyme-linked immunosorbent assay (ELISA) kit (ScheBo-Pancreatic Elastase 1™, Giessen, Germany), and fecal elastase will be extracted and analyzed according to the manufacturer's instructions.
• Ecoendoscopy (Diagnostic test): **Diagnosis of Pancreatic Steatosis** The diagnosis of pancreatic steatosis will be performed using endoscopic ultrasound. A Pentax EG-3870UTK endoscope will be used in conjunction with Hitachi Avius ultrasound equipment. The procedure is performed under anesthesia using propofol.
• Fibroscan (Diagnostic test): **Fibroscan Procedure:** This is a new technique based on the evaluation of liver elasticity or stiffness that allows for the measurement of liver hardness and quantification of liver fibrosis in a simple and completely painless manner using ultrasound. The results are obtained immediately and can be safely repeated periodically.
• Serum cytokines (Diagnostic test): Peripheral blood will be drawn from all participants in the Gastroenterology Division of the Hospital de Clínicas. Samples will be collected in EDTA tubes, centrifuged, and the serum will be frozen at -80 °C until processing. Serum cytokine levels will be measured using a commercial kit (Bio-Plex Pro human cytokine, Bio-Rad Lab., Inc.), which includes a panel of 27 cytokines: FGF basic, Eotaxin, G-CSF, GM-CSF, IFN-γ, IL-1β, IL-1ra, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, MIP-1α, IL-12 (p70), MIP-1β, IL-13, PDGF-BB, IL-15, RANTES, IL-17, TNF-α, IP-10, VEGF, MCP-1 (MCAF).

Primary outcomes

• Prevalence Non-Alcoholic Fatty Pancreas Disease [At baseline]: Determine the prevalence of Non-Alcoholic Fatty Pancreas Disease (NAFPD) in patients with Metabolic Dysfunction-Associated Fatty Liver Disease (MAFLD).
• Prevalence of exocrine pancreatic insufficiency [At baseline]: Determine the prevalence of exocrine pancreatic insufficiency (EPI) in patients with Non-Alcoholic Fatty Pancreas Disease (NAFPD).
• Prevalence of IPE in MAFLD [At baseline]: Determine the prevalence of exocrine pancreatic insufficiency (EPI) in patients with Metabolic Dysfunction-Associated Fatty Liver Disease (MAFLD).

Secondary outcomes

• Clinical characteristics [At baseline]: Analyze the causes of exocrine pancreatic insufficiency (EPI) and Non-Alcoholic Fatty Pancreas Disease (NAFPD) in this population.
• Cytokine profile [At baseline]: Evaluate the cytokine phenotypic profile in this population.

Sex: All, ages: 18 Years to 65 Years

Inclusion Criteria:

* Patients over 18 years old with a diagnosis of Metabolic Dysfunction-Associated Fatty Liver Disease (MAFLD).

Exclusion Criteria:

* \- Alcohol consumption >20 g/day in women, >30 g/day in men
* Chronic hepatitis B or C infection
* Autoimmune liver diseases: autoimmune hepatitis, primary biliary cholangitis, primary sclerosing cholangitis
* Hereditary hemochromatosis
* Wilson's disease
* Alpha-1 antitrypsin deficiency
* Celiac disease
* Uncontrolled thyroid disease
* Active or chronic infectious disease
* Active cancer or ongoing treatment
* Chronic renal insufficiency
* Pregnancy/lactation
* Insufficient data
* Patients who do not complete follow-up

• Results publication: Olmos JI, Piskorz MM, Litwin N, Schaab S, Tevez A, Bravo-Velez G, Uehara T, Hashimoto H, Rey E, Sorda JA, Olmos JA. Exocrine Pancreatic Insufficiency is Undiagnosed in Some Patients with Diarrhea-Predominant Irritable Bowel Syndrome Using the Rome IV Criteria. Dig Dis Sci. 2022 Dec;67(12):5666-5675. doi: 10.1007/s10620-022-07568-8. Epub 2022 Jun 15. PMID 35704255
• Results publication: Bellentani S. The epidemiology of non-alcoholic fatty liver disease. Liver Int. 2017 Jan;37 Suppl 1:81-84. doi: 10.1111/liv.13299. PMID 28052624
• Results publication: Bedogni G, Miglioli L, Masutti F, Castiglione A, Croce LS, Tiribelli C, Bellentani S. Incidence and natural course of fatty liver in the general population: the Dionysos study. Hepatology. 2007 Nov;46(5):1387-91. doi: 10.1002/hep.21827. PMID 17685472
• Results publication: Weiss J, Rau M, Geier A. Non-alcoholic fatty liver disease: epidemiology, clinical course, investigation, and treatment. Dtsch Arztebl Int. 2014 Jun 27;111(26):447-52. doi: 10.3238/arztebl.2014.0447. PMID 25019921
• Results publication: Eslam M, Sanyal AJ, George J; International Consensus Panel. MAFLD: A Consensus-Driven Proposed Nomenclature for Metabolic Associated Fatty Liver Disease. Gastroenterology. 2020 May;158(7):1999-2014.e1. doi: 10.1053/j.gastro.2019.11.312. Epub 2020 Feb 8. PMID 32044314
• Results publication: Eslam M, Newsome PN, Sarin SK, Anstee QM, Targher G, Romero-Gomez M, Zelber-Sagi S, Wai-Sun Wong V, Dufour JF, Schattenberg JM, Kawaguchi T, Arrese M, Valenti L, Shiha G, Tiribelli C, Yki-Jarvinen H, Fan JG, Gronbaek H, Yilmaz Y, Cortez-Pinto H, Oliveira CP, Bedossa P, Adams LA, Zheng MH, Fouad Y, Chan WK, Mendez-Sanchez N, Ahn SH, Castera L, Bugianesi E, Ratziu V, George J. A new definition for metabolic dysfunction-associated fatty liver disease: An international expert consensus statement. J Hepatol. 2020 Jul;73(1):202-209. doi: 10.1016/j.jhep.2020.03.039. Epub 2020 Apr 8. PMID 32278004
• Results publication: Dominguez-Munoz JE. Diagnosis and treatment of pancreatic exocrine insufficiency. Curr Opin Gastroenterol. 2018 Sep;34(5):349-354. doi: 10.1097/MOG.0000000000000459. PMID 29889111