Clinical Trial Details
— Status: Completed
Administrative data
| NCT number |
NCT05905185 |
| Other study ID # |
GUP 2018-052 |
| Secondary ID |
|
| Status |
Completed |
| Phase |
Phase 4
|
| First received |
|
| Last updated |
|
| Start date |
January 1, 2020 |
| Est. completion date |
January 21, 2022 |
Study information
| Verified date |
June 2023 |
| Source |
National University of Malaysia |
| Contact |
n/a |
| Is FDA regulated |
No |
| Health authority |
|
| Study type |
Interventional
|
Clinical Trial Summary
The goal of this clinical trial is to investigate the effects of tocotrienol-rich fraction
vitamin E supplementation on liver enzymes in overweight and obese children with
non-alcoholic fatty liver disease as compared to placebo.
The main question[s] it aims to answer are:
1. Does supplementation of tocotrienol-rich fraction vitamin E reduce the level of liver
enzymes and improve liver steatosis in non-alcoholic fatty liver disease among
overweight and obese children?
2. Does tocotrienol-rich fraction vitamin E supplementation improve the level of liver
steatosis by reducing the level of DNA damage?
Participants will :
1. consume daily either a dose of 50 mg of tocotrienol-rich fraction (TRF) vitamin E or a
placebo for 6 months.
2. Routine clinical assessments include weight, height, waist circumference, and BMI.
Fasting glucose, and fasting serum lipid.
3. The following investigations were performed upon recruitment and following 6 months of
intervention: (i) liver biomarker and enzymes; (ii) DNA damage; (iii) TNFα, IL-6 and
IFN-gamma genes; (iv) Fibroscan.
Description:
Methodology This study enrolled a diverse group of participants aged 10 to 18, regardless of
gender, who were diagnosed with fatty liver disease detected via ultrasonography and
abnormally high alanine transaminase levels (at least two-fold higher than the upper limits
for their respective genders). The trial consisted of 29 patients, with 15 receiving a daily
oral dose of 50 mg TRF and 14 receiving a placebo for a duration of six months. Various
clinical parameters, LIVERFASt in vitro diagnostic test, fibroscan, biochemical parameters,
DNA damage, and gene expression, both at the outset and at the end of the study were
monitored.
The amount of blood sample taken was 13mls each at entry and on completion at 6 months.
Details of these tests are as shown below:
Transient Elastography (FibroScan):
This is a non-invasive test to measure liver stiffness, kPa (indicator of liver fibrosis) and
to detect degree of fat accumulation (CAP range value of 100-400 dB/m), liver stiffness of >
8kPa indicates advanced fibrosis while CAP of > 263 indicates fatty liver. During the
screening, the participant lied down on the examination bed with his/her right hand behind
his/her head. A probe was placed by the investigator between the right ribs of the patient.
Measurements were recorded into the machine (FibroScan® 502 Touch). The screening is quick
and easy, usually taking less than 15 minutes. The scanned steatosis was scored and graded.
Determination of DNA Damage The DNA Damage pre and post intervention were conducted by using
CometAssay Kit (Trevigen, Gaithersburg, USA) following the manufacturer's protocol. Briefly,
blood cells in 5 μL medium was suspended in 70 μL warm 0.6% low-melting point (LMA) agarose
(Boehringer Mannheim, Germany) (DNAse-free, RNAsefree). Slides were allowed to sit in the
alkaline buffer for 20 min to allow unwinding of DNA strands and expression of alkali-labile
damage. Electrophoresis was performed for 20 min at 300 mA and 25 V. Following dropwise
neutralization (TrisHCl, pH 7.5) for 5 min, cells were stained by applying 30 μL 1X ethidium
bromide. The slides were examined and the tail length was measured with a fluorescence
microscope (Carl Zeiss, Germany). The DNA migration of 100 randomly selected cells were
examined for each sample. A total damage score was determined by multiplying the number of
cells assigned to each grade of damage by the numeric value of the grade according to methods
described by Heaton et al. [19]. Total DNA damage score was calculated as follows. Total DNA
damage = [(0 × n0) + (1 × n1) + (2 × n2) + (3 × n3) + (4 × n4)] where n0 =cells with Score 0,
n1 = cells with Score 1, n2 = cells with Score 2, n3 = cells with Score 3, and n4 = cells
with Score 4.
Determination of liver biomarker and enzyme The levels of liver enzymes including alanine
aminotransferase, ALT, aspartate aminotransferase, AST, gamma glutamyl transferase, GGT, and
alkaline phosphatase, ALP, pre and post intervention were determined from the blood samples
of the patients. Other biomarkers such as lipid profile, α-2 macroglobulin and haptoglobin
were also be analysed. The blood samples were collected in a BD Vacutainer and centrifuge at
1500 xg for 15 mins for the serum separation. The serum was stored in -80°C if it is not
processed immediately. The levels of the liver enzymes were determined by using ADVIA 1800
(Siemens, USA).
Quantitative Polymerase Chain Reaction (qPCR):
Total RNA was extracted from blood samples of NAFLD patients with or without tocotrienol.
Primers and probes for TNFα, IL-6 and IFN-gamma genes were designed. 1 ug/ul of total RNA was
converted to cDNA which then be used for PCR reaction. All samples were analysed in
duplicate. Expression of genes was determined after normalised with the housekeeping genes.
