Dott. Marcello Dallio Specialista in formazione in Malattie dell’Apparato Digerente

1. Dott. Marcello Dallio Specialista in formazione in Malattie dell’Apparato Digerente Tutor: Prof. Alessandro Federico

2. Prevalenzadella NAFLD • 20-33% in adulti-anziani: • Incrementa con l’età; • Più alta negli uomini rispetto alle donne; • Più alta nei soggetti di etnia caucasica rispetto agli ispanici; • Non è significativamente più alta nei pazienti con alterazione della ALT rispetto a quelli senza tale alterazione

3. La prevalenza della NAFLD è più alta in: • Soggetti obesi (36-78%) • Pazienti con iperglicemia o diabete (43-62%) • Pazienti con iperlipemia (45-65%) • Pazienti con ipertensione (35-45%) • Pazienti con sindrome metabolica: il rischio di progressione verso stadi più gravi di malattia è significativamente maggiore • Pazienti con infezione da HCV (55%)

4. L’eziologia del dannoepaticostacambiando: passato, presente e futuro %

5. Storia naturale Attenzione: l’HCC insorge su epatopatia metabolica spesso anche prima che essa giunga allo stadio di cirrosi epatica, quindi su “semplice” epatite cronica. Tale condizione sarebbe dovuta allo stimolo procancerogeno sostenuto dalle condizioni costituenti la stessa sindrome: il diabete in primis!

6. Come si fa la diagnosi di NAFLD? Ecografia Biopsia epatica Marcatori ematici RMN Fibroscan CAP?

7. Diagnosi • Anamnesi, EO, misurazione indici antropometrici • Prelievi ematochimici: insulinemia, The Homeostasis Model Assessment (HOMA), colesterolemia, trigliceridemia, glicemia a digiuno, ecc • Ecografia epatica: metodica non proprio eccezionale a causa della scarsa sensibilità e dell’operatore dipendenza • RM: consente una definizione precisa della presenza del grasso epatico ma non differenzia la steatosi dalla steatoepatite • Analisi CAP: software annesso ad un apparecchio chiamato “Fibroscan”. Consente un’analisi specifica per la presenza del grasso epatico • Biopsia: attualmente l’unica possibilità reale per porre diagnosi di steatoepatite

8. MODELLI PREDITTIVI SEMPLICI DI PROGRESSIONE • AST/ALT ratio • AST/ALT > 1 suggeriscefibrosiavanzata • non distingue trasteatosisemplice e NASH • BMI, AST/ALT Ratio e Diabete- BARD score • combinazione di 3 variabili (BMI ≥ 28: 1 punto; AST/ALT ratio ≥ 0.8: 2 punti; diabete: 1 punto) • solo 43% di sensibilità • NAFLD fibrosis score (NFS) • basatosuetà, iperglicemia, BMI, piastrine, livello di albumina e AST/ALT ratio • unagrandepercentuale cade nellacategoriaindeterminata • FIB4 index • basatosuetà, piastrine e livelli di AST e ALT

9. MODELLI PREDITTIVI COMPLESSI DI PROGRESSIONE • Enhanced Liver Fibrosis – ELF panel • INCLUDE 3 biomarkers (acidoialuronico, tissue inhibitor della metalloproteinasi-1 e N teleopeptide del collagenoIII) • ELF panel + NAFLD score aumental’accuratezzadiagnostica • Fibrotest • INCLUDE 5 biomarkers (GGT, bilirubina, aptoglobina, α2-macroglobulina e apo-lipoproteinaA) • sensibilità e specificità: 77%

11. Liver Biopsy IL RUOLO DELL’ ISTOLOGIA

12. Ruolo della biopsia epatica nella NAFLD A favore Contro Prognosi generalmente buona Mancanza di terapia specifica Rischio e costi associati • Esclude altre cause di malattia epatica • Distingue la NAFL dalla NASH • Stima la prognosi • Valuta la progressione della malattia

13. US is the preferred first-line diagnostic procedure for imaging of NAFLD, asitprovidesadditionaldiagnostic information (A1) • Wheneverimagingtools are notavailable or feasible(e.g. large epidemiologicalstudies), serumbiomarkers and scores are an acceptable alternative for the diagnosisof steatosis (B2) • A quantitative estimation of liverfat can only be obtained by H-MRS. Thistechniqueis of value in clinical trials and experimentalstudies, butisexpensive and notrecommended in the clinicalsetting(A1) • NASH has to be diagnosed by a liverbiopsyshowingsteatosis, hepatocyteballooning and lobularinflammation(A1) • Biomarkers and scores of fibrosis, aswellastransientelastography, are acceptable non-invasive procedures for the identification of casesatlowrisk of advancedfibrosis/cirrhosis(A2). The combination of biomarkers/ scores and transientelastographymightconferadditionaldiagnosticaccuracy and mightsave a number of diagnosticliverbiopsies(B2)

14. Monitoring of fibrosisprogression in clinicalpracticemayrely on a combination of biomarkers/scores and transientelastography, althoughthisstrategyrequiresvalidation (C2) • The identification of advancedfibrosis or cirrhosis by serumbiomarkers/scores and/or elastographyisless accurate and needs to be confirmed by liverbiopsy, according to the clinicalcontext (B2) • In selectedpatientsat high risk of liverdiseaseprogression, monitoringshould include a repeatliverbiopsyafteratleast 5-year follow-up (C2) • In children, predictors of fibrosis, includingelastometry, acousticradiation force impulse (ARFI) imaging and serumbiomarkersmight help reduce the number of biopsies (B2)

15. EASL, EASD, EASO. J Hepatol. 2016

16. …e se la biopsia non fosse solo una metodica diagnostica?

17. Fibrosis stage butnot NASH predictsmortality and time to developmentof severe liverdisease in biopsy-provenNAFLD. HannesHagström, PatrikNasr, MattiasEkstedt, UlfHammar, Per Stål, RolfHultcrantz, StergiosKechagias. J Hepatol. 2017 Aug 10. pii: S0168-8278(17)32202-X. doi: 10.1016/j.jhep.2017.07.027. Aim To investigate the long-termprognosis in a large cohort of NAFLD patients with a prolonged follow-up time, and to study the specificeffect of NASH on the outcomes of mortality and liver-specificmorbidity.

18. Materials and methods Patients: retrospectivecohortstudyincludingallpatientsdiagnosed with biopsy-proven NAFLD, at the Karolinska University Hospital, Huddinge and LinköpingUniversity Hospital, during 1971 to 2009. Patients with othercauses for steatosisthan NAFLD or diagnosed with anyconcurrentliverdiseaseduring follow-up wereexcluded. Patients on treatment with drugsassociated with hepaticsteatosis or hepatotoxicityat the time of biopsywerealsoexcluded. Furthermore, tenpatients with a diagnosis of either HCC or decompensatedliverdiseaseat or withinsixmonths from baseline wereexcluded.

19. Materials and methods • Histopathologicalevaluation:Lobularinflammation and steatosiswerescored on a 0-3 scale and ballooning and portalinflammation on a 0-2 scale. The FLIP algorithmwasused to definepresence of NASH. Fibrosis stage wasscoredaccording to the Kleinerclassification on a 5-point scale (F0-F4). • Baseline characteristics: assessment of type2 diabetesmellitus, hypertension; cardiovasculardisease; hyperlipidaemia; smoking habits and weight. • Biochemicalvariables: routine biochemicalvariableswithinonemonth of liverbiopsywereregistered and included alanine and aspartateaminotransferase, albumin, bilirubin, alkalinephosphatase, and gamma-glutamyltransferaselevels, complete bloodcount, fastingcholesterol and triglycerides, fastingglucose, autoantibodies and α1-antitrypsin levels. In cases with missing data, multiple imputationwasused. Analysis for detection of hepatitis B surfaceantigenwasperformed in allcases and anti-hepatitis C virus (HCV) antibodieswereanalysed in casesevaluatedafter 1991 whentestingbecameavailable.

20. Materials and methods • Control group and follow up: The personal identificationnumber (PIN) is a uniqueten-digit code provided to allSwedishcitizens.ThePIN was first used to create a control populationderived from StatisticsSwedenusingtencontrols per case (N=6345 afterexclusions). For the follow up theyusedoutcome data from the National PatientRegister of Hospital Discharges (NPR), from the Cause of Death Register (CDR) and from the SwedishCancerRegister (SCR). • Severe liverdiseasewasdefinedas an ICD-code for liverfailure, cirrhosis, HCC or decompensatedliverdisease. Decompensatedliverdiseasewas in turn definedas an ICD-code for esophagealvarices (bleeding or notbleeding), ascites or hepaticencephalopathy, whileliverfailurewasdefinedashaving a specific ICD-code for liverfailureonly.

21. Results The cohortwasfollowed for a meanperiod of 19.9 years Therewas a strong collinearitybetweenhigherstages of fibrosis and higherNAS and presence of NASH, with 82% of patients with stage 4 having a NAS of 5-8 and NASH in 94% compared to a NAS of 5-8 in 24% and NASH in 35% in patients with stage 0 (p<0.001, respectively). Mortality:

22. Results • Compared to controls • The entiregroup of NAFLD patientsshowed a trend towardshigherrisk for mortalitythancontrols (HR 1.14, 95%CI 0.99-1.32, p=0.07): Patients with fibrosis stage 0 (HR 0.86, 95%CI 0.65-1.16, p=0.33) and stage 1 (HR 0.88, 95%CI 0.70-1.12, p=0.30) didnothave an increasedmortality, whilepatients with stage 2 (HR 1.36, 95%CI 1.02- 1.80, p=0.03), stage 3 (HR 2.54, 95%CI 1.79-3.60, p<0.001) and stage 4 (HR 5.19, 95%CI 3.06-8.79, p<0.001) did with increasing HR:s per stage of fibrosis. Patients with NASH had a slight increase in overall mortality (1.22, 95%CI 1.02-1.46, p=0.03). • Using NAFLD cases with fibrosis stage 0 as controls, and adjusting for age, sex and T2DM, a significant increase in mortality was seen for patients with fibrosis stage 3 (HR 1.76, 95%CI 1.02-3.06, p=0.04) and stage 4 (HR 3.75, 95%CI 1.81-7.73, p<0.001). Adding presence of NASH to the model with fibrosis stage as the independent variable did not add to the predictive capacity of the model for any stage of fibrosis (LR test p > 0.05 for each stage of fibrosis), while adding information on fibrosis stage significantly improved the model with NASH as an independent variable (LR test p<0.001).

23. Results Overallmortalitystratified on fibrosis stage compared to matchedcontrols. Log-rank test p\0.001

24. Results Development of severe liverdiseasestratified on fibrosis stage compared to matchedcontrols. Log-rank test p \0.001.

25. Results Laplace regression revealed that the time until the first 10% of the patients had developed severe liver disease was 30.5 years in F0 (95%CI 21.5-39.6), 35.6 years in F1 (95%CI 25.6-45.4), 19.4 years in F2 (95%CI 9.3- 29.5) and 6.0 in F3 (95%CI 2.3-9.6). The time until 10% of the patients had developed liver decompensation was 33.4 years for F0 (95%CI 24.2-42.6), 34.1 years for F1 (95%CI 25.1-43.2), 22.7 years for F2 (95%CI 13.7-31.7), 11.8 years for F3 (95%CI 4.3-19.4) and 5.6 years for F4 (95%CI 0.9-10.3). Therewas no significantdifference in the number of severe liverdiseasecases in patientswithout and with NASH (9.8% vs. 12.8%, p=0.29). Cases with NAFLD weresignificantly more likelythancontrols to develop severe liverdisease (HR 4.25, 95%CI 3.09-5.84, p<0.001), and fibrosis stage washighlypredictive of this.

26. Conclusions • Aftera follow-up time of up to 40 years, presence of NASH didnotaffect the long-termoutcome of NAFLD significantly. • Patientswith lowerstages of fibrosis (stage 0-1) had the potential to progress to severe liverdisease and thisoccurred in roughly 7% of thesepatients, independent of presence of NASH. • The lower end of the 95% CI for the 10th percentile of liverdecompensationwas 0.9 years in manifestcirrhosis (5.6, 95%CI 0.9-10.3). The lower end of the 95% CI for the 10th percentile of time to severe liverdiseasewas 2.3 years in F3 (6.0, 95%CI 2.3-9.6), 9.3 years in F2 (19.4, 95%CI 9.3- 29.5) and around 22-26 years in F0-1 (30.5 years in F0, 95%CI 21.5-39.6, and 35.6 years in F1, 95%CI 25.6-45.4). • Althoughpatients with NASH seemed to have a small increase in risk for overallmortality and liver-specificmorbidity in univariateanalysescompared to a referencepopulation, thiswasgenerallynotsignificantwhenadjusting for confoundersincludingfibrosis stage, and no addedriskwasseen for presence of NASH afteradjusting for confounders. Furthermore, in patients with similarstages of fibrosis, presence of NASH didnotaffect the estimates in anyspecificmanner.

27. Punti di forza e limitazioni • Pro: • Ampia popolazione studiata; • Diagnosi ottenuta con biopsia esaminata dallo stesso patologo esperto per ridurre i bias di interpretazione dei quadri istologici; • Ampio follow-up; • Elevata affidabilità delle informazioni ottenute dai registri svedesi.

28. Punti di forza e limitazioni • Contro: • Il 12% della popolazione studiata aveva una fibrosi avanzata (F3-4), percentuale superiore rispetto a quella della popolazione generale; • Non sono stati ottenuti dati individuali dei pazienti attraverso un’analisi diretta ed inoltre le variabili valutate al baseline non sono state prese in considerazione durante il follow-up.