Effects of the Supplementation With Omega 3 Fatty Acids in Patients With Chronic Renal Disease
Phase: PHASE3
Conditions: Renal Insufficiency, Chronic
Sponsor: University of Chile

Official title of the Study: 
Omega 3 Fatty Acids in Patients with Chronic Kidney Disease 
NCT Number 
NCT03280615   
Date of the Document 
July 31, 2019 

1 
 
Material and methods: 
We invited to participate patients with Chronic Kidney Disease (CKD) who had a urine albumin 
excretion of 30 mg/g creatinine or over. All these participants were part of Non- communicable 
Chronic Disease programs at public primary health care clinics of East Metropolitan Santiago. 
Participants were invited to a first interview to assess their eligibility for the study. We excluded 
participants younger than 18 years and older than 85 years of age. We also excluded those who 
had chronic debilitating diseases such as active cancer, heart failure grade III or more according to 
the classification of the New York Heart Association 1, chronic debilitating infections or a history of 
drug or alcohol abuse. In the case of diabetic patients, we excluded those who were 
decompensated, defined as a fasting blood glucose over 150 mg/dl or a glycosylated hemoglobin 
over 8%. The study was approved by the ethics committee of East Metropolitan Santiago Health 
Service.  
Participants deemed eligible were cited after an overnight fast to the outpatient clinic of the 
Institute of Nutrition and Food Technology (INTA). After signing an informed consent, they were 
subjected to the following procedures: 
1. Weight, height and waist circumference were measured.  
2. A fasting blood sample was obtained to measure blood glucose, serum lipids, glycosylated 
hemoglobin, creatinine, cystatin C, β2 microglobulin, C reactive protein, interleukin-6 and 
fatty acid profile in red cell membranes. Cystatin C and interleukin 6 were measured using 
ELISA kits from R&D Systems. Red cell membrane fatty acids were measured by liquid 
chromatography after extracting lipids from erytrhocytes using Bligh y Dryer 2 method 
and subsequently derivatized. All other determinations were carried out using routine 
clinical laboratory methods in a certified clinical laboratory (Vida Integra). 

2 
 
3. A spot urine sample was obtained to measure albumin, creatinine and urea nitrogen at 
Vida Integra laboratory. 
4. Blood pressure and pulse wave velocity were measured using a Mobil-O-Graph® 
oscillometric device. Augmentation pressure and index and reflection magnitude of the 
pulse wave are also measured. The measurements were done in the sitting position after 
a five minutes rest in a quiet room. 
5. Measurement of carotid intima media thickness using a General Electric LogiQ 
ultrasonographic device with border detection software. A 14.2 x 47  mm  lineal GE 12L-
RS transducer was used. International measurement norms were used 3. 
6. Measurement of liver fat infiltration using the same ultrasonographic device but with a 
convex  multifrequency (2 – 5.5 MHz) 4C RS GE transducer. Fatty infiltration was 
ascertained using Hamaguchi score 4. We have previously validated this technique against 
liver fat measurement using magnetic resonance spectroscopy. 
Participant features were reassessed with the new laboratory determinations and those who 
continued to comply with inclusion criteria were randomized into two groups, balancing by 
gender, age, body mass index and urine albumin excretion expressed per mg of creatinine. 
Randomization was carried out using an iteration software based on random number generation. 
The output was an individual number for each patient. This number was sent to the oil 
manufacturer, who had the codes for each number and prepared the supplement to be used.  
1. Active supplement contained 282 mg of docosahexanoic and eicosapentanoic acids per 
ml. 
2. Placebo supplement: Contained corn oil 
Participants were instructed to drink 13 ml of the supplement per day in one or two doses. Both 
supplements had a lemon flavor and were indistinguishable due to deodorization technology used 

3 
 
to prepare them. Therefore the investigators and the participants were blinded to the treatment 
allocation. 
The intervention period lasted 3 months. Participants were cited every two weeks to provide them 
with a new bottle of supplement. They returned the used bottle and the residual content was 
measured to determine compliance. They were also interrogated about adverse events or other 
eventualities.  
At the sixth week and at the end of the intervention at 12 weeks, participants were subjected to 
the same assessments done at baseline.  
All participants could call the investigators in any moment if they an intercurrent problem. If a 
subject failed to attend an appointment, he or she was called by phone or visited at their houses 
to find out the reason for not showing. I a subject declined to continue in the study, he or she was 
considered as a loss from follow up. 
Data analysis: 
Until the end of the intervention period for all participants, they were only identified with their 
allocation number. Once the intervention ended, each participant was classified as pertaining to 
group 1 or 2, without knowing which was the group receiving the active supplement. All red cell 
membrane fatty acid composition measurements, was analyzed when all statistical analysis of the 
rest of the variables were done, to maintain the blinding. An intention to treat analysis was 
performed. 
Normality of variable distribution was analyzed using Shapiro Wilk test. Since most variables had a 
non-normal distribution, results are expressed as median (interquartile range). By the same token, 
non-parametric statistical methods were used to compare variables. Chi square or fisher tests 
were used to compare proportions. Differences between groups were compared using Kruskal 

4 
 
Wallis test. To assess changes in repeated measures of a single parameter a mixed linear 
regression model for repeated measures was used 5. Significance level was set at 0.05 
The main outcome of the study was set as a reduction in urine albumin excretion by 20% or more. 
We anticipated that 35% more participants receiving the active product than those receiving 
placebo would attain this reduction. Thus, the sample size to obtain results with an α of 0.05 and a 
power of 0.8 was 38 participants per group. 
Results: 
Participant flow is shown in figure 1. We randomized 50 participants per group and lost five in corn 
oil group  and four in Ω-3 groupl The demographic features, body mass index and baseline albumin 
excretion of participants randomized to each group are shown in table 1. No differences between 
groups were observed, therefore the premises of randomization were correctly attained. Red 
blood cell membrane phospholipids fatty acid composition during the study are shown in table 2. 
Patients receiving the active supplement had significant increases in eicosapentanoic and 
docosahexanoic fatty acid levels.  
A 20% reduction in urine albumin excretion was attained in 13 participants of group 1 and 19 
participants of group 2 (fisher p= 0.274). The clinical and laboratory parameters in the three 
assessment periods for groups 1 and 2 are observed in table 3.  
Summarizing,  participants receiving corn oil maintained their pulse wave velocity during the 
follow up while controls experienced an increase in this parameter. Triacylglycerol levels 
decreased significantly in the Ω-3 group and glycosylated hemoglobin increased significantly in 
both groups 
The compliance with the study product for groups 1 and 2, calculated measuring the leftovers was 
88.7 (74-97) and 89.6 (77-98) %, respectively (p= 0.99). Eighty nine adverse events were recorded 
(48 in group 1 and 27 in group 2, p = NS). Of these, two were severe (a severe hyperkalemia and 

5 
 
the discovery of a cervical mass that resulted in a medullary thyroid carcinoma, both in 
participants of group 1). Mild gastrointestinal symptoms were reported by 12 and 14 participants 
in groups 1 and 2 respectively.  
Preliminary conclusions: 
1. The primary outcome measure of the study was not attained, since urine albumin 
excretion decreased by 20% or more in a similar proportion of participants in each 
intervention group. 
2. The striking reduction in serum triacylglycerol observed in participants receiving Ω-3 fatty 
acids is a known pharmacological effect of Ω-3 fatty acids.   
3. The deterioration of glycosylated hemoglobin in both groups, may be associated with the 
fact that participants received 110 Kcal/day of extra calories with the supplement. 
4. The lack of increase in pulse wave velocity in group 2 may indicate that the supplement 
that this group received had a favorable effect on arterial stiffness. 
5. The supplement was well tolerated. The lack of differences between groups in compliance 
and gastrointestinal complains indicates that the fish oil effectively almost completely 
deodorized.  
 
 

6 
 
Figure 1 
 
 
 
 

7 
 
Table 1: Initial features of randomized participants expressed as median (interquartile range)  
Gender (F/M) 
Age (years) 
Body mass index (kg/m2) Urine albumin (mg/g creatinine) 
Corn oil 
Group 
26/24 
66 (58-74) 
31 (27.2-35.3) 
604.5 (212-1234) 
Ω-3 group 
23/27 
67 (56-74) 
33.5 (30-37.3) 
365.5 (157-1079) 
Difference 
NS 
NS 
NS 
 
 
 

8 
 
Table 2: Changes in red blood cell membrane phospholipids fatty acid composition. Values expressed as median (interquartile 
range)  in mg/100 mg of total fatty acids 
 
Corn oil group (n=45) 
Ω-3 Group (n=46) 
p¶ 
 
Basal  
Six weeks  
Final  
Basal  
Six weeks 
Final 
 
Pentadecanoic 
Acid (C15:0) 
0.2 
(0-3) 
2.3 
(0-3.2) 
2 
(0-3) 
0.2 
(0-2.9) 
1.2 
(0-3.2) 
2.3 
(0-3.3) 
b 
Palmitic  
Acid (C16:0) 
27.4 
(25.2-29.9) 
28.9 
(27-31.1) 
25.7 
(23.7-28.3) 
27.8 
(25.5-29.2) 
28.8 
(26.7-30.8) 
27.7 
(24.8-28.9) 
Heptadecanoic 
Acid (C17:0) 
2.4 
(0.4-4.2) 
3.1 
(0.8-4.7) 
2.3 
(0.3-4.7) 
1.7 
(0.4-3.6) 
3.1 
(0.5-4.5) 
2.7 
(0.5-4.3) 
b 
Estearic 
Acid (C18:0) 
15.6 
(12.9-18.4) 
15.3 
(13.7-17.1) 
17.5 
(13.6-18.8) 
16.3 
(13.8-17.9) 
15.6 
(14-17.9) 
16.9 
(14.1-18.1) 
Arachidic  
Acid (C20:0) 
0 
(0-0.3) 
0 
(0-0.3) 
0 
(0-0.3) 
0 
(0-0.3) 
0 
(0-0.3) 
0.2 
(0-0.3) 
Eneicosanoic 
Acid (C21:0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
Behenic 
Acid (C22:0) 
0.9 
(0.6-1.2) 
1.1 
(0.8-1.2) 
0.8 
(0.3-1) 
1 
(0.7-1.2) 
1.1 
(0.9-1.3) 
0.9 
(0.4-1.2) 
Lignocerico  
Acid (C24:0) 
2.6 
(1.4-3.4) 
2.8 
(2.4-3.4) 
2.2 
(1-3.4) 
2.6 
(1.3-3.3) 
2.8 
(2.2-3.3) 
2.6 
(0.8-3.3) 
Total Saturated 
54.9 
(50.7-58.3) 
55 
(51.9-57.7) 
55.8 
(51.7-59.8) 
54.1 
(49.7-58.2) 
55.2 
(50.1-57.8) 
55.9 
(52.9-62.2) 
b 
Miristoleic 
Acid (C14:1) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
Pentadecanoico  
Acid (C15:1) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
Palmitoleic  
Acid (C16:1) 
0 
(0-0.4) 
0 
(0-0.4) 
0 
(0-0.4) 
0 
(0-0.4) 
0 
(0-0.4) 
0 
(0-0.3) 
Cis 10-Heptadecanoic A 
Acid (C17:1) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
Oleico  
Acid (C18:1 ) 
14.3 
(13.3-15.7) 
14.7 
(13.7-15.7) 
13.7 
(12.1-14.5) 
14.7 
(13.3-15.9) 
14.8 
(14-16.1) 
14.2 
(13-15) 
Gadoleic  
Acid (C20:1) 
0 
(0-0.2) 
0 
(0-0.2) 
0 
(0-0.2) 
0 
(0-0.2) 
0 
(0-0) 
0 
(0-0.2) 
Erucic  
Acid (C22:1) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
Nervonic Acid 
(C24:1) 
2.5 
(1.5-3.1) 
2.7 
(2.3-3.2) 
2.2 
(1.7-2.7) 
2.6 
(1.7-3) 
2.8 
(2.4-3.2) 
2.5 
(0.9-2.8) 
Total monounsaturated 
17.4 
(16.2-19) 
17.5 
(16.9-19) 
16.6 
(15.3-18.5) 
17.6 
(16.3-19) 
17.7 
(17.2-19) 
17.4 
(15.3-18.1) 
Linoleic  
Acid (C18:2 n6 c) 
11.8 
(9.3-13.9) 
12.2 
(10-13.9) 
11.8 
(8.6-13.4) 
10.9 
(9.4-12.6) 
11 
(9.5-12.3) 
9.9 
(8.5-11.5) 
Gama linolenic  
Acid (C18:3 n6 c) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
Alfa linolenic  
Acid 
(C18:3 n-3) 
0 
(0-0.2) 
0 
(0-0) 
0 
(0-0.1) 
0 
(0-0.1) 
0 
(0-0) 
0 
(0-0.1) 
Eicosadienoico 
Acid (C20:2) 
0 
0-0.3) 
0 
(0-0.3) 
0.1 
(0-0.3) 
0 
(0-0.3) 
0 
(0-0.3) 
0 
(0-0.2) 
 
 
 
 
 
 
 
 
Table 2 continuation 
 
 
 
 
 
Corn oil group (n=45) 
Ω-3 Group (n=46) 
p¶ 
 
Basal  
Six weeks  
Basal  
Six weeks  
Basal  
Six weeks  
 

9 
 
Dihomo gamalinolenic 
Acid (C20:3 n-6 ) 
1.5 
(1.1-1.9) 
1.4 
(1.1-1.7) 
1.4 
(1-1.9) 
1.3 
(0.8-1.9) 
1.1 
(0.8-1.5) 
1.2 
(0.7-1.6) 
Cis-11-14 icosatrienoico 
Acid (C20:3 n-3) 
0 (0-0) 
0 (0-0) 
0 (0-0) 
0 (0-0) 
0 (0-0) 
0 (0-0) 
Arachidonic 
Acid (C20:4 n-6) 
9.4 
(6.1-12.5) 
10.1 
(6.2-12.1) 
9.1 
(6.4-12.6) 
8.9 
(5.5-11.7) 
8.8 
(6.6-11.1) 
8.1 
(5.1-10) 
Eicosapentanoic 
Acid (C 20:5 n-3) 
0.4 
(0-0.7) 
0.4 
(0-0.7) 
0.5 
(0.2-0.8) 
1.3 
(0.4-3) 
2.8 
(1.4-3.3) 
2.6 
(1.1-3.5) 
c 
Docosadienoico  
Acid (C22:2) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
0 
(0-0) 
Docosahexanoic  
Acid (C22:6 n-3) 
1.9 
(1.3-2.8) 
1.8  
(1.4-2.6) 
1.6 
(1-2.4) 
2.7 
(2-3.7) 
2.9 
(2.2-3.8) 
2.8 
(2.2-4.2) 
c 
Total polyusaturated 
27.5 
(23.4-30.8) 
26.6 
(24.1-30.3) 
26.6 
(22.4-29.9) 
27.6 
(23.2-31.5) 
27 
(24.3-31.4) 
26.1 
(22.1-29.9) 
b 
 
¶ = significance for differences between groups determined by a mixed effects lineal regression model for 
repeated measures. a = difference between groups without intervention effect, b= intervention effects 
without differences between groups  c= intervention effect with differences between groups (Ω-3 fatty acid 
effect) 
 
 

10 
 
Table 3: Measured parameters in both treatment groups, expressed as median (interquartile range) 
 
 
Corn oil group (n=45) 
Ω-3 Group (n=46) 
p¶ 
Basal  
Six weeks  
Final  
Basal  
Six weeks 
Final 
 
Clinical parameters 
Systolic blood 
pressure (mm Hg) 
129 
(121-142) 
134 
(123-140) 
133 
(123-150) 
135.5 
(126-153) 
137 
(122-150) 
138.5 
(128-149) 
  
Diastolic blood 
pressure (mm Hg) 
81 
(73-87) 
80 
(73-86) 
83 
(74-90) 
81.5 
(73-95) 
80.5 
(75-92) 
84.5 
(74-93) 
Weight (kg) 
76.4 
(69.7-91.5) 
77.4 
(71.7-90.5) 
77.6 
(70.1-93.3) 
90.4 
(79.1-101.2) 
91.05 
(77.5-100.5) 
91.2 
(77.6-101.9) 
Waist circumference 
(cm) 
104 
(97-113.2) 
104 
(97.6-114) 
105 
(96-113) 
110.5 
(103.5-122.4) 
112 
(104.2-120) 
111.25 
(104-120.6) 
Carotid intima media thickness: 
 
Left (mm) 
0.66 
(0.6-0.83) 
0.67 
(0.57-0.84) 
0.71 
(0.58-0.83) 
0.71 
(0.61-0.88) 
0.735 
(0.62-0.85) 
0.695 
(0.59-0.82) 
Right  (mm) 
0.62 
(0.55-0.715) 
0.62 
(0.54-0.73) 
0.645 
(0.555-0.755) 
0.68 
(0.6-0.78) 
0.7 
(0.61-0.78) 
0.71 
(0.61-0.79) 
Arterial stiffness measures 
Augmentation 
pressure (mm Hg) 
8 
(5-12) 
9 
(5-15) 
12 
(5-19) 
9 
(5-16) 
11 
(6-20) 
12 
(4-18) 
b 
Augmentation index 
(%) 
18 
(9-29) 
23 
(11-35) 
25 
(14-39) 
20 
(8-31) 
27 
(12-38) 
23 
(9-34) 
b 
Reflection 
magnitude (%) 
66 
(61-73) 
70 
(59-75) 
69 
(57-75) 
67 
(61-73) 
67 
(63-73) 
66 
(61-74) 
Pulse wave velocity 
(m/s) 
9.3 
(8.5-10.7) 
9.7 
(8.3-11) 
9.7 
(8.3-11.2) 
9.75 
(8.1-10.6) 
10 
(7.6-10.8) 
10.1 
(8-10.9) 
c 
Liver ultrasound 
Liver steatosis 
score (units) 
3 
(2-4) 
3 
(2-4) 
3 
(3-4) 
3 
(2-4) 
4 
(2-4) 
3 
(3-4) 
Laboratory values 
Fasting glucose 
(mg/dl) 
94 
(86-115) 
97 
(89-120) 
95 
(86-132) 
99 
(83-118) 
107.5 
(90-143) 
103.5 
(90-137) 
Total cholesterol 
(mg/dl) 
165 
(138-203) 
162 
(135-195) 
160.5 
(134-195) 
170 
(140-204) 
167.5 
(141-203) 
154.5 
(127-176) 
Hdl cholesterol 
(mg/dl) 
43.1 
(37.1-52.3) 
44.2 
(36-51.6) 
41.7 
(35.3-53.2) 
41.3 
(35.2-54.4) 
42.7 
(35.6-53.8) 
41.55 
(37.8-51.8) 
Triacylglycerol 
(mg/dl) 
147 
(117-219) 
133 
(96-220) 
150  
(106-219) 
163.5 
(111-238) 
125.5 
(90-173) 
117.5 
(90-167) 
c 
Serum creatinine 
(mg/dl) 
1 
(0.82-1.76) 
1.09 
(0.79-1.78) 
1.08 (0.81-
2.06) 
1.01 
(0.79-1.33) 
0.98 
(0.75-1.57) 
1.04 
(0.8-1.37) 
Urine albumin 
excretion (mg/L) 
370.6 
(140.9-756.2) 
344.3 
(132.1-1096.5) 
513.9 
(185.4-978.7) 
375.35 
(146.5-584.4) 
360.85 
(162.9-1099.6) 
467.9 
(144.6-1005.2)  
Blood urea nitrogen 
(mg/dl) 
22.3 
(16.7-36.7) 
20.5 
(16.3-37.5) 
20.7 
(16.1-36.9) 
18.95 
(15-25.5) 
19.6 
(15.4-28.6) 
19.15 
(13.6-28) 
Urine creatinine 
excretion (mg/dl) 
78 
(56-100) 
95 
(69.5-113.5) 
80 
(57-118) 
91.5 
(62-135) 
99.5 
(62-126) 
90.5 
(68-121) 
Glycosylated 
hemoglobin (%) 
5.9 
(5.4-6.7) 
6.2 
(5.4-6.8) 
6.2 
(5.6-6.9) 
6.35 
(5.7-7.2) 
6.45 
(5.8-7.2) 
6.5 
(5.9-7.4) 
b 
Urine 
albumin/creatinine 
ratio (mg/g) 
541 
(173-1125) 
561.5 
(129-1063) 
678 
(149-1217) 
363 
(157-1030) 
435 
(164-950) 
492 
(129-1071) 

11 
 
Table 3 continuation 
 
 
 
Corn oil group (n=45) 
Ω-3 Group (n=46) 
p¶ 
 
Basal  
Six weeks  
Final  
Basal  
Six weeks 
Final 
 
Urine urea nitrogen 
(mg/dl) 
569.6 
(439.3-784.2) 
570.5 
(467.95-783.4) 
615.6 
(433.6-803.3) 
629.45 
(526.6-824.9) 
659.4 
(444.6-881.3) 
594.1 
(488.3-763.6) 
 
C Reactive protein 
(mg/L) 
1.68 
(1.11-4.11) 
1.81 
(0.99-4.49) 
1.91 
(0.99-5.22) 
3.265 
(1.54-5.27) 
2.5 
(1.31-5.35) 
2.14 
(1.15-5.85) 
β2 microglobulin 
(mg/L) 
2.7 
(1.9-5.1) 
2.7 
(2-5) 
2.4 
(2-5.1) 
2.35 
(1.8-3.4) 
2.3 
(1.8-3.8) 
2.2 
(1.7-3) 
Interleukin-6  
(pg/ml) 
0.3 
(0.2-0.4) 
0.3 
(0.2-0.3) 
0.2 
(0.2-0.4) 
0.3 
(0.2-0.4) 
0.3 
(0.2-0.4) 
0.3 
(0.2-0.4) 
Cistatin-C (mg/L) 
1 
(1-1.9) 
1.1 
(1-2.1) 
1.2 
(1-1.8) 
1.1 
(0.9-1.6) 
1.2 
(1-1.8) 
1.2 
(0.9-1.6) 
Estimated 
glomerular filtration 
rate  
(ml/min/1.73 m2) 
68 
(31.2-80) 
67 
(28.6-75.9) 
60.1 
(34.6-76.5) 
60.7 
(42.4-81.9) 
58.5 
(33.7-73.9) 
60.6 
(39.6-78.9) 
 
¶ = significance for differences between groups determined by a mixed effects lineal regression model for 
repeated measures. a = difference between groups without intervention effect, b= intervention effects 
without differences between groups  c= intervention effect with differences between groups (Ω-3 fatty acid 
effect  
 

12 
 
References: 
1  
WRITING COMMITTEE MEMBERS, Yancy CW, Jessup M, Bozkurt B, Butler J, Casey DE Jr, 
Drazner MH, Fonarow GC, Geraci SA, Horwich T, Januzzi JL, Johnson MR, Kasper EK, Levy 
WC, Masoudi FA, McBride PE, McMurray JJ, Mitchell JE, Peterson PN, Riegel B, Sam F, 
Stevenson LW, Tang WH, Tsai EJ, Wilkoff BL; American College of Cardiology 
Foundation/American Heart Association Task Force on Practice Guidelines. 2013 ACCF/AHA 
guideline for the management of heart failure: a report of the American College of 
Cardiology Foundation/American Heart Association Task Force on practice guidelines. 
Circulation. 2013 Oct 15;128(16):e240-327 
 
2  
Bligh EG, Dyer WJ. A rapid method of total lipid extraction and purification. Can J Biochem 
Physiol 1959;37:911e7. 
 
3  
Stein JH, Korcarz CE, Post WS. Use of carotid ultrasound to identify subclinical vascular 
disease and evaluate cardiovascular disease risk: summary and discussion of the American 
Society of Echocardiography consensus statement. Prev Cardiol. 2009 ;12(1):34-8 
 
4  
Hamaguchi M, Kojima T, Itoh Y, Harano Y, Fujii K, Nakajima T, Kato T, Takeda N, Okuda J, Ida 
K, Kawahito Y, Yoshikawa T, Okanoue T. The severity of ultrasonographic findings in 
nonalcoholic fatty liver disease reflects the metabolic syndrome and visceral fat 
accumulation. Am J Gastroenterol. 2007;102:2708-15. 
 
5  
Davivian M, Giltinan DM. Analysis of repeated measurement dat using the non linear mixed 
effects model. Chemometrics and Intelligent Laboratory Systems. 1993;20:1-24