Efficacy of silymarin in patients with non-alcoholic fatty liver disease — the Siliver trial: a study protocol for a randomized controlled clinical trial

Eligibility criteria for the Siliver trial include consenting adult patients aged between 20 and 60 years of both sexes. All patients had a medically confirmed diagnosis of NAFLD prior to participation in this trial.

Patients who meet any of the following criteria have not been included lactating or pregnant women, and women during their menacme, except for those who underwent definitive contraception such as hysterectomy or tubal ligation; patients with a previously established diagnosis of chronic disease including congestive heart failure, decompensated or severe lung disease, neoplasms, kidney disease, carriers of the human immunodeficiency virus (HIV), and advanced chronic liver diseases (Child-Pugh classification B and C) caused by any causal agent unrelated to NAFLD; recreational drug users; patients with an average intake of more than 20 g of alcohol per day and or a history of alcoholic disease in abstention for less than 6 months; use of prescribed medication including steroids, oestrogens, amiodarone, warfarin, anticonvulsants, antipsychotics, or chemotherapy drugs in the past 6 months; infections, surgery, trauma, or hospitalization in the past 30 days; chronic degenerative diseases of non-hepatic origin; and unavailability of previously obtained imaging tests with a diagnosis of NAFLD in its screening phase.

The following patients will be excluded patients who at the recruitment stage do not show confirmation of NAFLD by CT scan; patients who missed any phase of the trial; and patients who during the trial were diagnosed with a condition listed in the non-inclusion criteria above.

The primary outcome will be the assessment of NAFLD resolution, or change in its grade, as assessed by the difference in the attenuation coefficient between liver and spleen, obtained by CT of the upper abdomen, at the end (after) compared to beginning (before) the trial. In summary, the primary outcome is to investigate whether silymarin supplementation can reduce liver fat content in NAFLD patients, measured by a CT scan.

The secondary outcomes investigated in this trial include:

Patients who meet the eligibility criteria are referred to the screening consultation to receive information about the clinical trial and are given the opportunity to ask questions to the research team. Consenting participants are invited to sign the free and informed consent form. All participants are informed that they can withdraw at any time without giving reasons and that they will not be penalized for withdrawing. After signing the consent form, all participants answer questions on a standardized form filled in by the researcher. Information on sociodemographics, clinical history and clinical presentation, lifestyle data, diet intake, and feeding patterns are collected. The first anthropometric assessment is completed at that point.

At the end of the screening consultation, participants receive general nutritional guidelines and a study identification card with the contact details of the researchers. Participants are encouraged to make contact should they have any late questions about the trial. Following the first appointment, participants are referred to an upper abdomen CT scan before the intervention begins.

At this appointment, participants receive their CT scan and blood test results and are submitted to a second anthropometric assessment. At this consultation, a new form with nutritional and dietary data is completed by the researcher, and patients are given additional nutritional guidelines to complement the advice offered in the screening appointment. Subsequently, participants are sent to the clinical research pharmacy of the Hospital Complex for capsule collection according to their randomization.

All participants were followed up through face-to-face individual consultations in 4-week intervals during the 12-week supplementation period. The Return Visits were scheduled as detailed in Table 1. At these appointments, patients were asked about adherence to the study. They were asked to bring along to each consultation the flasks they receive containing the capsules from the previous 4-week period, so that the research team can more accurately estimate adherence to the protocol.

At each monthly follow-up visit participants had a nutritional and dietary assessment undertaken by a registered nutritionist member of the research team. The researcher completed a form with dietary intake and dietary habits over the last 4-week period and evaluated adherence to nutritional guidelines, ingestion of capsules as prescribed, tolerance, and the occurrence of any adverse effects.

except for the weeks when participants had their scheduled Return Visits, all participants were contacted weekly by telephone to monitor adherence to the protocol, clarify any questions, and report any occurrence or adverse reactions.

At Return Visit 3, which took place at the end of the 12^th week of intervention, patients were referred to a second CT scan and blood tests following the same protocols as the baseline tests. A final consultation was scheduled for the test results to be delivered (Table 1).

In the final consultation, participants who no longer showed fatty infiltration received guidance on how to prevent the recurrence of NAFLD. Participants who showed evidence of NAFLD, regardless of the degree of steatosis, were referred to the outpatient follow-up clinic for continuation of their care provision with a hepatologist consultant and a registered nutritionist (Fig. 1).