Effect of Dietary Supplementation With Fish Oil in Alleviating Cardiopulmonary Hazards Associated With Air Pollution
Phase: NA
Conditions: Oxidative Stress, Inflammation, Blood Pressure, Vasoconstriction, Coagulation, Pulmonary Function
Sponsor: Fudan University

A cover page 
 
The Official Title of the study: Effect of dietary supplementation with fish oil in 
alleviating cardiopulmonary hazards associated with air pollution 
 
NCT number: 03255187 
 
Date of the document: December 15, 2017 
 

Study protocol 
This trial will be conducted between September 2017 and January 2018. We plan 
to recruit 70 healthy college students on Fenglin campus of Fudan University in 
Shanghai, China. We will exclude students who do not live on campus, or have a history 
of tobacco smoking or alcohol addition, clinically diagnosed cardiovascular and 
respiratory diseases, or recent infections, or are allergic to omega-3 fatty acids or fish. 
Eligible participants will be randomly assigned to receive dietary supplementation of 
either marine-derived fish oil (2.5 g/day) or placebo (sunflower-seed oil), and will be 
monitored in a double-blinded fashion throughout the study period. We plan to schedule 
4 rounds of follow-up visits with an interval of 2 weeks in the last 2 months of the 
intervention. To expand the daily variation range of PM2.5 exposures, in each round we 
will divide each group to 2 subgroups and arranged health examinations on either 
Saturday for placebo group or Sunday for fish-oil group at adjacent weeks. There will 
have identical working conditions and technical procedures between these two 
assessment days. We will collect data on individual characteristics (i.e., age, gender, 
height, and weight), time-location information and physical activity at baseline and 
during the follow-ups. We also plan to ask participants to complete a validated food 
frequency questionnaire at the start, middle and the end of study period to record the 
profile of daily dietary nutrients intake. Health measurements will include blood 
pressure (BP) and lung function, blood samples and skin tape samplings in each round 
of follow-up. 
Statistical analysis Plan 
Environmental and health data will be linked by the time of health measurements 
(rounded to the integer hour).  
T-test or chi-square test will be applied to compare the difference of characteristics 
and outcome variables of participants between the fish-oil and placebo groups. Linear 
mixed-effect models will be applied to evaluate short-term exposure to PM2.5 in relation 
to each of cardiovascular outcomes within each treatment group. We will apply multiple 
averaging periods preceding the health measurements [i.e., 0–6 hours (h), 0–12 h, 0–24 
h, and 0–48 h] to explore the lag patterns that the acute effects of PM2.5 might have on 
individual biomarkers. We will further test the statistical differences between the two 
treatment groups in their effect estimates of PM2.5 on individual biomarkers and 
calculated the 95% confidence intervals (CI) as  
(𝑄̂1-𝑄̂2) ± 1.96√SÊ1
2 + SÊ2
2, 
where 𝑄̂ 1 and 𝑄̂ 2 are the effect estimates of PM2.5 for the two treatment groups, 
SÊ1and SÊ2 are their respective standard errors.  
To test the robustness of our results, we will perform several sensitivity analyses. 
First, we will repeat analyses by fitting two-pollutant models by adding the present-day 
concentrations of 4 gaseous pollutants (sulfur dioxide, nitrogen dioxide, carbon 
monoxide, and ozone) to regression models one at a time. Second, in the basic models, 
we will further adjust for baseline dietary intakes of nutrients that will show 
significantly different between the two groups. Finally, to explore the possibly 
diminishing effects of PM2.5, we will apply longer lag periods (i.e., 0-72 h and 0-96 h) 

in supplementary analyses. Additionally, we will adjust for present-day ultraviolet 
radiation to exclude its confounding effects on skin. 
All analyses will be conduct using the “lme4” package of R software. In the 
within treatment group analyses, we will present the absolute changes in BP (in 
millimeters mercury) and the percentage changes in other biomarkers and their 95% 
CIs for each 10-μg/m3 increment in PM2.5 exposure. To demonstrate potential benefits 
of fish-oil supplementation, the differences in the effect sizes of the PM2.5-biomarker 
associations between the fish-oil and placebo groups will also be presented.