EAL

DLM: Elevated Triglycerides and Omega-3 Fatty Acids (2007)

Citation:

Study Design:

Class:

- Click here for explanation of classification scheme.

Quality Rating:

Research Purpose:

To investigate the effects of supplementation with n-3 fatty acids in a lipid-lowering diet in well-defined dyslipoproteinemias on plasma lipoproteins and hemorrheology.

Inclusion Criteria:

Primary hypertriglyeridemia (plasma triglycerides > 2.85 mmol/L)
- Familial dysbetalipoproteinemia type III hyperlipoproteinemia (FDL)
  - Diagnosis established by determination of apolipoprotein E phenotype 2/2
  - Familial combined hyperlipidemia was excluded by measuring the ratio of triglyceride and apolipoproteinemia B in VLDL
- Familial hypertriglyceridemia (FHTG)
  - Diagnosis established by obtaining the family history (at least one member of the family suffered from hypertriglyceridemia) and excluding FDL and familial combined hyperlipidemia.
No lipid lowering drugs for at least 6 weeks prior to the start of the study.

Exclusion Criteria:

Smoking
Diabetes mellitus
Obesity (BMI > 28.5)
Arterial hypertension (systolic blood pressure >180 mm Hg and diastolic blood pressure >100 mm Hg)
Chronic liver disease (gamma-glutamyl transpeptidase >70 U/L)
Renal disease (creatinine >2 mg/dL)
Hypothyroidism (thyrotropin >4.0 µU/mL or total thyroxine <5 µg/dL)
Chronic alcohol abuse
Unstable angina pectoris
History of myocardial or cerebral infarction
No antilipidemic or lipid affecting drugs allowed (ß-blockers, thiazides, corticoides, or estrogens)

Description of Study Protocol:

Recruitment not discussed

Design Non-randomized trial

Blinding used (if applicable)

Intervention (if applicable)

Week -4: start lipid lowering diet
Week 0: initiated acclimating dose of fish oil capsules for 2 weeks
- 3 gm/day containing 1.8 gm n-3 fatty acid ethyl esters (50% EPA, 33% DHA)
Week 2: Increased fish oil capsules to intervention dose for 6 weeks
- 6 gm/day containing 3.6 gm ethyl esters
Weeks 8-11: washout period (fish oil capsules discontinued)
Week 12: slow-release fenofibrate (250 mg) started daily for 8 weeks

Statistical Analysis

Results are reported as the mean ± SEM.
Statistical analyses were performed using the Wilcoxon matched-pairs signed-rank test to assess differences within groups under therapy.
- Blood values during fish oil therapy were compared with baseline levels (week 0)
- Values during fenofibrate therapy (week 20) were compared with values after the fish oil washout (week 12)
- No statistically significant changes were seen when values obtained with fenofibrate therapy were compared with baseline levels (week 0) instead of fish oil washout levels (week 12)
The Mann-Whitney U test was used to assess differences between groups (FDL vs FHTG).
P values <0.05 were considered to indicate statistical significance

Data Collection Summary:

Timing of Measurements

Performed at weeks 0 (baseline), 2, 4, 8, 12, and 20.

Dependent Variables

Lipid parameters (after an overnight fast)
- VLDL separated by ultracentrifugation
- HDL cholesterol was determined in the infranatant after heparin-manganese precipitation of LDL
- LDL cholesterol content was calculated by subtracting HDL-cholesterol from total infranatant cholesterol
- TG and cholesterol levels were masured enzymatically by an autoanalyzer
- Apoprotein levels were measured with a nephelometer
- Free EPA and DHA in plasma (deep frozen) were determined by HPLC
- Apolipoprotein E phenotype was determined by isoelectric focusing
- Lipoprotein analyses were performed or started on the same day.
- All measurements were performed in duplicate
Hemorrheology parameters
- Hematocritwas determined after centrifugation
- Viscosity and red blood cell aggregation measurements were made with 4 hours after blood sampling.
  - Plasma and blood viscosity
  - Red blood cell aggregation was determined photometrically (in triplicate)
  - Measurements were performed at stasis and low shear (3/s)
  - Transit time and clogging rate were determine with a blood filtrometer to assess red blood cell filterability
  - Fibrinogen
Safety parameters
- Creatinine, prothrombin index, blood glucose and gamma-glutamyl transpeptidase levels were measured by standard lab techniques
Compliance determined by measuring plasma concentrations of EPA and DHA

Independent Variables

Fish oil capsules
Fenofibrate

Control Variables

Description of Actual Data Sample:

Initial N: 25 (male/female breakdown not provided)

Attrition (final N):

N=23 (22 males, 1 female)
- 15 familial hypertriglyceridemia (FHTG)
- 8 familial dysbetalipoproteinemia (FDL)
Reasons for attrition
- Abdominal discomfort (3.6 gm/day)
- Pregnancy

Age:

FHTG: 44.07±2.51 yr
FDL: 48.63±3.33

Ethnicity: not described

Other relevant demographics: not described

Anthropometrics

	FHTG (N=15)	FDL (N=8)
Weight (kg)	78.0±2.6	74.9±2.2
Total cholesterol (mmol/L)	6.67±0.41	9.76±1.32
HDL choleterol (mmol/L)	0.92±0.05	1.00±0.06
HDL₂ cholesterol (mmol/L)	0.23±0.05	0.19±0.02
HDL₃cholesterol (mmol/L)	0.68±0.04	0.80±0.07
LDL cholesterol (mmol/L)	2.75±0.28	2.69±0.32
Total TG (mmol/L)	7.44±1.50	6.90±1.70
VLDL cholesterol (mmol/L)	3.00±0.40	5.88±1.22
VLDL TG (mmol/L)	5.82±1.13	5.89±1.52
VLDL-C:VLDL-T	0.25±0.02	0.48±0.04
Apo A-1 (mg/dl)	141.0±9.7	146.0±10.5
Apo B (mg/dl)	176.2±19.9	172.3±31.6

Location: Medical Department, Klinikum Großhadern, University of Munich, Munich, Germany

Summary of Results:

Safety Parameters

Safety parameters did not differ significantly during therapy with fish oil or fenofibrate, with the exception of a slight increase in creatinine during fenofibrate therapy (1.03±0.05 at baseline to 1.10±0.05 mg/dl, P<0.05) in FHTG subjects

Compliance

EPA (µmol/L)			DHA (µmol/L)
Baseline	1.8 g/day	3.6 g/day	Baseline	1.8 gm/day	3.6 gm/day
1.23±0.18	3.50±0.47¹	4.77±0.74 ²	1.09±0.14	1.96±0.38 ³	1.88±0.20 ¹

¹ P<0.005, ² P<0.001, ³ P<0.05

Lipid Parameters in Familial Hypertriglyceridemia (N=15)

	Week 0 (Baseline)	Week 2	Week 4	Week 8	Week 12 (washout)	Week 20 (Fenofibrate)
Total cholesterol (mmol/L)	6.67±0.41	6.69±0.35	6.43±0.24	6.77±0.33	7.31±0.44	6.46±0.26 ³
HDL₂ cholesterol (mmol/L)	0.23±0.05	0.15±0.02	0.12±0.01	0.12±0.01 ¹	0.14±0.01	0.14±0.01
HDL₃ cholesterol (mmol/L)	0.68±0.04	0.72±0.04	0.72±0.04	0.71±0.03	0.71±0.03	0.82±0.04 ³
LDL cholesterol (mmol/L)	2.75±0.28	3.18±0.36	3.59±0.19 ¹	3.97±0.35 ²	2.94±0.28	3.74±0.37 ³
Total TG (mmol/L)	7.44±1.50	6.13±1.30	4.30±0.65 ¹	4.15±0.55 ¹	8.16±1.58	5.25±2.42³
VLDL cholesterol (mmol/L)	3.00±0.04	2.69±0.35	2.07±0.27	2.00±0.23 ²	3.53±0.51	1.78±0.38 ⁴
LDL TG (mmol/L)	5.82±1.13	5.54±1.19	4.01±0.67	3.57±0.50 ¹	7.31±1.39	4.56±2.06 ⁴
Apo B (mg/dl)	176.2±19.9	187.1±15.5	165.4±9.6	159.6±7.0	190.9±23.0	163.7±14.5 ³

¹ P<0.05, ² P<0.01: vs baseline

³ P<0.05, ⁴ P<0.01: vs washout period

In subjects with FHTG, total TG decreased by 44% (P<0.05), LDL cholesterol increased by 44% (P<0.01), and VLDL cholesterol significantly declined by 33% (P<0.01), and VLDL TG by 39% (P<0.05) with n-3 fatty acid therapy.
Comparable results were observed with fenofibrate therapy (significant reduction in total and VLDL TG and VLDL cholesterol and significant increase in LDL cholesterol).
With fenofibrate therapy, total cholesterol and Apo B decreased while HDL₃ cholesterol increased significantly.
With both n-3 fatty acid and fenofibrate therapy, total TG decreased in 12/15 subjects; LDL cholesterol increased in 13 subjects with fish oil and in 12 with fenofibrate therapy.

Lipid Parameters in Familial Dysbetalipoproteinemia (N=8)

	Week 0 (Baseline)	Week 2	Week 4	Week 8	Week 12 (Washout)	Week 20 (Fenofibrate)
Total cholesterol (mmol/L)	9.76±1.32	8.00±1.01 ¹	7.52±0.92 ¹	7.34±1.07 ¹	8.65±1.07	6.77±0.76 ²
Total triglycerides (mmol/L)	6.90±1.70	4.79±1.03	4.11±0.84	3.61±0.78 ¹	5.12±0.85	3.33±0.65 ²
VLDL cholesterol (mmol/L)	5.88±1.22	3.37±0.88 ¹	3.31±0.70 ¹	2.69±0.50 ¹	4.13±0.93	2.99±0.65 ²
VLDL triglycerides (mmol/L)	5.89±1.52	3.54±0.76 ¹	3.22±0.78 ¹	2.32±0.37 ¹	3.72±0.76	2.58±0.44 ²
Apo B (mg/dl)	172.3±31.6	133.3±28.8 ¹	130.8±21.5 ¹	114.7±25.1 ¹	137.0±27.2	102.8±14.9 ²

¹ P<0.05 vs baseline; ² P<0.05 vs washout period

Significant decreases in VLDL cholesterol of 54% (P<0.05) and VLDL TG of 61% (P<0.05) were observed with n-3 fatty acid therapy.
Total cholesterol (-25%, P<0.05) and Apo B (-34%, P<0.05) were reduced significantly.
With fenofibrate therapy, significant reductions were seen in total and VLDL cholesterol (-22%, -28%, respectively), total and VLDL TG (-35%, -31%, respectively) and Apo B (-26%).
Total TG decreased in 7/8 subjects with n-3 fatty acid therapy and in all subjects with fenofibrate therapy.

Hemorrheological Parameters in Familial Hypertriglyceridemia

No significant changes were observed in fibrinogen concentration, plasma viscosity, standard blood viscosity, or parameters of red blood cell filterability with n-3 fatty acid or fenofibrate therapy.
Red blood cell aggregation was significantly decreased with n-3 fatty acid therapy for stasis (from 5.47±0.34 to 4.81±0.029 U, P<0.01) and for shear rate of 3/s [from 10.5±0.43 to 9.47±0.045 U, P<0.5 (typo?)].
Decreasing red cell aggregation was observed in 13/15 subjects.

Hemorrheological Paramters in Familial Dysbetalipoproteinemia

Plasma viscosity was higher for FDL compared with FHTG (P<0.005).
Whole blood viscosity showed a trend to higher values in FDL compared with FHTG, which did not reach significance (P=0.11).
n-3 fatty acid therapy did not alter hemorrheological parameters.
With fenofibrate therapy, fibrinogen concentration (-20%, P<0.05), plasma viscosity (-6%, P<0.05), and standard blood viscosity for all shear rates (P<0.05) were significantly reduced.
No statistically significant changes were observed for red blood cell filterability.
Red blood cell aggregation was not determined because aggregometer was not available at the beginning of the study.

Author Conclusion:

n-3 fatty acid and fenofibrate had comparable effects on lipid parameters in subjects with familial dysbetalipoproteinemia and familial hypertriglyceridemia.
Because of additional beneficial effects on hemorrheological parameters, fenofibrate may be preferable in familial dysbetalipoproteinemia.

Funding Source:

Reviewer Comments:

Small sample size, no sample size calculation.

No information provided about the lipid lowering diet. Did subjects adhere to diet during study?

Did subjects take any supplements that could have affected results?

Wonder whether capsule count should have been conducted in addition to measuring EPA and DHA levels to monitor compliance.

How was compliance measured with fenofibrate?

Quality Criteria Checklist: Primary Research
Relevance Questions
	1.	Would implementing the studied intervention or procedure (if found successful) result in improved outcomes for the patients/clients/population group? (Not Applicable for some epidemiological studies)	Yes
	2.	Did the authors study an outcome (dependent variable) or topic that the patients/clients/population group would care about?	Yes
	3.	Is the focus of the intervention or procedure (independent variable) or topic of study a common issue of concern to dieteticspractice?	Yes
	4.	Is the intervention or procedure feasible? (NA for some epidemiological studies)	Yes

Validity Questions
1.	Was the research question clearly stated?		Yes
	1.1.	Was (were) the specific intervention(s) or procedure(s) [independent variable(s)] identified?	Yes
	1.2.	Was (were) the outcome(s) [dependent variable(s)] clearly indicated?	Yes
	1.3.	Were the target population and setting specified?	Yes
2.	Was the selection of study subjects/patients free from bias?		Yes
	2.1.	Were inclusion/exclusion criteria specified (e.g., risk, point in disease progression, diagnostic or prognosis criteria), and with sufficient detail and without omitting criteria critical to the study?	Yes
	2.2.	Were criteria applied equally to all study groups?	Yes
	2.3.	Were health, demographics, and other characteristics of subjects described?	Yes
	2.4.	Were the subjects/patients a representative sample of the relevant population?	???
3.	Were study groups comparable?		Yes
	3.1.	Was the method of assigning subjects/patients to groups described and unbiased? (Method of randomization identified if RCT)	Yes
	3.2.	Were distribution of disease status, prognostic factors, and other factors (e.g., demographics) similar across study groups at baseline?	Yes
	3.3.	Were concurrent controls or comparisons used? (Concurrent preferred over historical control or comparison groups.)	Yes
	3.4.	If cohort study or cross-sectional study, were groups comparable on important confounding factors and/or were preexisting differences accounted for by using appropriate adjustments in statistical analysis?	N/A
	3.5.	If case control study, were potential confounding factors comparable for cases and controls? (If case series or trial with subjects serving as own control, this criterion is not applicable.)	N/A
	3.6.	If diagnostic test, was there an independent blind comparison with an appropriate reference standard (e.g., "gold standard")?	N/A
4.	Was method of handling withdrawals described?		Yes
	4.1.	Were follow-up methods described and the same for all groups?	Yes
	4.2.	Was the number, characteristics of withdrawals (i.e., dropouts, lost to follow up, attrition rate) and/or response rate (cross-sectional studies) described for each group? (Follow up goal for a strong study is 80%.)	Yes
	4.3.	Were all enrolled subjects/patients (in the original sample) accounted for?	Yes
	4.4.	Were reasons for withdrawals similar across groups?	No
	4.5.	If diagnostic test, was decision to perform reference test not dependent on results of test under study?	N/A
5.	Was blinding used to prevent introduction of bias?		Yes
	5.1.	In intervention study, were subjects, clinicians/practitioners, and investigators blinded to treatment group, as appropriate?	No
	5.2.	Were data collectors blinded for outcomes assessment? (If outcome is measured using an objective test, such as a lab value, this criterion is assumed to be met.)	Yes
	5.3.	In cohort study or cross-sectional study, were measurements of outcomes and risk factors blinded?	N/A
	5.4.	In case control study, was case definition explicit and case ascertainment not influenced by exposure status?	N/A
	5.5.	In diagnostic study, were test results blinded to patient history and other test results?	N/A
6.	Were intervention/therapeutic regimens/exposure factor or procedure and any comparison(s) described in detail? Were interveningfactors described?		Yes
	6.1.	In RCT or other intervention trial, were protocols described for all regimens studied?	Yes
	6.2.	In observational study, were interventions, study settings, and clinicians/provider described?	N/A
	6.3.	Was the intensity and duration of the intervention or exposure factor sufficient to produce a meaningful effect?	Yes
	6.4.	Was the amount of exposure and, if relevant, subject/patient compliance measured?	No
	6.5.	Were co-interventions (e.g., ancillary treatments, other therapies) described?	No
	6.6.	Were extra or unplanned treatments described?	N/A
	6.7.	Was the information for 6.4, 6.5, and 6.6 assessed the same way for all groups?	Yes
	6.8.	In diagnostic study, were details of test administration and replication sufficient?	N/A
7.	Were outcomes clearly defined and the measurements valid and reliable?		Yes
	7.1.	Were primary and secondary endpoints described and relevant to the question?	No
	7.2.	Were nutrition measures appropriate to question and outcomes of concern?	Yes
	7.3.	Was the period of follow-up long enough for important outcome(s) to occur?	Yes
	7.4.	Were the observations and measurements based on standard, valid, and reliable data collection instruments/tests/procedures?	Yes
	7.5.	Was the measurement of effect at an appropriate level of precision?	Yes
	7.6.	Were other factors accounted for (measured) that could affect outcomes?	No
	7.7.	Were the measurements conducted consistently across groups?	Yes
8.	Was the statistical analysis appropriate for the study design and type of outcome indicators?		Yes
	8.1.	Were statistical analyses adequately described and the results reported appropriately?	Yes
	8.2.	Were correct statistical tests used and assumptions of test not violated?	Yes
	8.3.	Were statistics reported with levels of significance and/or confidence intervals?	Yes
	8.4.	Was "intent to treat" analysis of outcomes done (and as appropriate, was there an analysis of outcomes for those maximally exposed or a dose-response analysis)?	No
	8.5.	Were adequate adjustments made for effects of confounding factors that might have affected the outcomes (e.g., multivariate analyses)?	N/A
	8.6.	Was clinical significance as well as statistical significance reported?	Yes
	8.7.	If negative findings, was a power calculation reported to address type 2 error?	No
9.	Are conclusions supported by results with biases and limitations taken into consideration?		Yes
	9.1.	Is there a discussion of findings?	Yes
	9.2.	Are biases and study limitations identified and discussed?	Yes
10.	Is bias due to study's funding or sponsorship unlikely?		Yes
	10.1.	Were sources of funding and investigators' affiliations described?	Yes
	10.2.	Was the study free from apparent conflict of interest?	Yes