Identification and characterization of non-alcoholic fatty liver disease in primary care.

Non-alcoholic fatty liver disease (NAFLD) is with 25% the most prevalent liver
disorder in Western society and is associated with overweight, obesity,
metabolic syndrome (MetS), type 2 diabetes mellitus (T2DM), cardiovascular
diseases (CVD) and increased risk of cancer development. NAFLD is defined by a
hepatic fat accumulation of more than 5% in the absence of classical causes of
steatogenesis (e.g. alcohol and steatogenic drugs). It represents a broad
spectrum of clinical entities from non-alcoholic fatty liver (NAFL) to advanced
liver disease with hepatic failure. Most of the patients have simple steatosis,
however in about 15-30% non-alcoholic steatohepatitis (NASH) develops, which
leads to an overall increase in morbidity and mortality due to the progression
to fibrosis, cirrhosis and hepatocellular carcinoma (HCC). Patients with NAFLD
have no or few, mainly a specific symptoms; and generally there is a silent
progression of simple steatosis to NASH and in the end liver-related morbidity
and mortality.
Despite the clinical importance and the potential impact on healthcare
resources, there is a striking lack of awareness on all levels of NAFLD.
Furthermore, little to no data are available concerning the quality of life of
NAFLD patients. Additionally, the majority of NAFLD patients are currently not
detected due to the lack of non-invasive methods to diagnose NAFLD. Most of
these patients, as a first contact in the healthcare system, will be found in
the outpatient clinic of the general practitioner (GP). To date, it is not
clear what the burden is of NAFLD and related diseases in at risk subjects in
primary care. Therefore, identification of NAFLD patients in this cohort will
give information on the prevalence in the group of uncomplicated overweight and
obesity and those with concomitant cardiometabolic diseases. By early detecting
these patients at risk to develop progressive liver diseases and extrahepatic
manifestations, it will be possible to intervene and improve health.

The primary objective of this cohort study is to determine the prevalence of
simple and progressed NAFLD in subjects at risk followed by the general
practitioner
The secondary objectives are:
1. To study the factors linked to the development of NAFLD (e.g. overweight,
obesity, metabolic syndrome, T2DM and CVD) in a population of at risk primary
care subjects.
2. To investigate non-invasive tools, both blood sample derived scores and
FibroScan® with Controlled Attenuation Parameter (CAP)TM, to identify patients
with more progressed NAFLD e.g. liver fibrosis.
3. To assess the quality of life and social-economic consequences of NAFLD
patients with validated questionnaires.
4. To compare the clinical characteristics of a matched control group of the
Maastricht Study with the cohort of NAFLD patients found in primary care.
5. Assess the performance of a benzylalcohol, 2-pentanol, 2-butanol, nonanal
and limonene breath levels (after administration of a standard dose) in
discriminating patients with progressed NALFD (FibroScan >=7.5 kPa) from
patients with early NAFLD (FibroScan <= 7 kPa and CAP score <= S1).
6. Explore correlations between hepatic metabolism of benzylalcohol,
2-pentanol, 2-butanol, nonanal and limonene and NAFLD co-morbidities.
7. Investigate correlations between breath benzylalcohol, 2-pentanol,
2-butanol, nonanal end limonene and blood biomarkers of liver
performance/damage or fibrosis.

A multicentre cohort study in primary care.
Eligible subjects will be included by the GP practices. Subjects will undergo a
FibroScan® with CAPTM measurement to diagnose the stage of NAFLD. All
participants will be asked to complete several questionnaires (i.e.
demographics, clinical data, SF-36, GAD-7 and PHQ-9, Baecke, WPAI-SHP, MDS and
sleep questionnaire) and to undergo anthropometric measurements. Furthermore,
blood samples and exhaled air will be collected. The urine of the patients will
be tested for proteinuria and glycemia. After analysis the blood, urine and
exhaled air samples are destroyed. Additionally, patients are asked to test
their hand grip strength with a hand dynamometer. This will be the standard to
be collected data set.
The subjects identified with NAFLD in primary care will be compared to a
healthy, matched control group from the Maastricht Study.
Subjects that will be recruited from the NAFLD primary care cohort will be
allocated to the sample or control group based on their FibroScan result.
The sample group (also referred to as progressed NAFLD) will be represented by
subjects with FibroScan >=7.5 kPa. The control group (also referred to as early
NAFLD), will be represented by subjects with FibroScan <= 7 kPa and CAP score <=
S1. (Fig. 2.1) The FibroScan test is performed on the day of the GP visit.
Ideally, on that day the subject will be invited to participate to the food
additives and limonene part of the study. Subjects for which the FibroScan
fails will not be considered for breath testing.
The metabolism of benzylalcohol, 2-pentanol, 2-butanol, nonanal and limonene
will be evaluated by collecting breath samples using the ReCIVA breath
collector, developed by Owlstone Medical and classified as research use only
device, before and after administration of food additives and limonene(Fig. 1.1
and Table 1.1). Information about diet, smoking status, alcohol intake and
utilization of medication will be collected by a questionnaire.

The measurements, data and sample collections will be planned with the
participant. Anthropometric data collection, hand grip strength, blood
collection, urine dipstick, exhaled air (VOC), and FibroScan® with CAPTM will
be performed during study visit 1 after signing the informed consent. The
participant can fill out the questionnaires at home and bring the forms when
visiting the research physician or fill out the questionnaires via an online
survey.
No side effects of the investigations are expected, apart from a small bruise
of taking the blood samples. Three tubes (21 ml) will be collected for research
purposes; if possible this will be combined with regular blood sampling. If
participants are not regular patients at the outpatient clinic, a maximum of 4
extra tubes (max.17,5 ml in total) will be collected to determine (part of) the
standard clinical laboratory investigations (e.g. liver function tests,
cholesterol) for research purposes only.
The time burden associated with participating in the research is: 45 minutes to
fill out the questionnaires, 30 min for the study visit to collect samples,
anthropometry data and perform a FibroScan®.
In case of unexpected findings, awareness of normally unknown pathology may
affect a person*s perception of his own health condition negatively. On the
other hand, early detection is likely to have favourable effects on disease
progression and enable early intervention.