Effects of Obesity and Physical Inactivity on Pregnancy Outcomes

Exercise during pregnancy is associated with the delivery of leaner, lighter, and healthier
infants1-7. Subsequently, high infant adiposity and birth weight are strong predictors of
childhood obesity and adult adiposity8-10. Therefore, maternal physical inactivity during
pregnancy may have significant ramifications for the child, the effects of which may extend
well into adulthood. Exercise during pregnancy also plays an important role in the health of
the mother. Active pregnant women tend to gain less weight during pregnancy4 and retain less
weight following pregnancy36. With excessive gestational weight gain being the strongest
risk factor for maternal overweight and obesity postpartum, as well as being associated with
many adverse maternal and neonatal metabolic outcomes such as adiposity and insulin
resistance11-13, the impact of exercise on maternal and neonatal outcomes could be
substantial. The mechanisms underlying these changes are poorly understood and studies which
strive to expose them are critical.

Habitual physical activity in non-gravid individuals has been shown to positively alter
lipid metabolism by increasing fatty acid oxidation14, but the effect of physical activity
on maternal lipid metabolism during pregnancy has not been studied. Due to previous research
suggesting that an altered intrauterine metabolic environment may play a significant role in
fetal programming15, it is reasonable to believe improvements in maternal lipid metabolism
may contribute to improved neonatal metabolic outcomes in exercising pregnant women16-17.
Preliminary data from our group found that in obese and lean pregnant women, lipid oxidation
rate was significantly correlated to offspring birth weight; suggesting maternal lipid
metabolism may contribute to neonatal metabolic outcomes. In inactive pregnant women,
impaired lipid oxidative capacity in conjunction with known increased physiologic adipose
tissue lipolysis that occurs during pregnancy and obesity18-19 would result in excess
un-oxidized plasma fatty acids that are likely to be re-esterified in adipose tissue and/or
delivered to the fetus. This series of events may contribute to increased maternal and
neonatal adiposity.

In addition, generation of excess reactive oxygen species, known byproducts of lipid
metabolism, may contribute to altered/abnormal oxidative stress profiles in obese pregnant
women. Reactive oxygen species are up-regulated during physiologic pregnancy as well as
non-gravid obesity, and research suggests oxidative stress may be related to poorer neonatal
outcomes10,14. In non-gravid individuals, long-term physical activity has been shown to
improve oxidative stress profiles37. Therefore, women who exercise during pregnancy may also
have higher antioxidant capacity and lower markers of oxidative stress; both of which may
contribute to favorable neonatal outcomes. However, this has not yet been studied.

Obesity is also believed to adversely influence lipid metabolism, oxidative stress, and
neonatal outcomes in pregnancy16-22. Therefore, we plan to compare these parameters in obese
active and obese inactive pregnant women. This study design will allow us to compare groups
in order to determine if unfavorable maternal lipid metabolism and oxidative stress
profiles, and neonatal metabolic outcomes (adiposity and insulin resistance) are more
attributable to physical inactivity or obesity. Previous research with non-gravid adults
suggests that the presence of comorbidity is more correlated with physical activity levels
than with body weight23-25. This finding is contrary to much of the previous literature on
pregnancy which suggests "obesity may be the most common health risk for the developing
fetus"15. Knowledge about maternal lipid metabolism and oxidative stress profiles and their
relationships in neonatal outcomes in active and inactive pregnant women can guide lifestyle
and medical interventions designed to target factors that may be contributing to poor
outcomes in obese pregnancy. Currently, we have collected data on lean, inactive pregnant
women that can be used for comparison at the end of all data collection.

This is the first study to examine the relationship between physical activity, lipid
metabolism and oxidative stress in obese pregnancy. We anticipate that results from the
proposed study will demonstrate the importance of a physically active lifestyle during
pregnancy (irrespective of body weight) in order to maximize the short and long-term health
of the neonate. In addition, we hope that these results will encourage obese and overweight
women of childbearing age to remain or become physically active by demonstrating that
physical inactivity has a greater effect on poor neonatal outcomes than obesity. These
findings are unique as much of the current literature focuses on the negative impact of
maternal obesity on neonatal outcomes. Blair et al. has consistently demonstrated in
non-gravid populations that physical inactivity is a stronger predictor of all-cause
mortality than obesity23-25. Similarly, we believe physical activity in pregnancy is more
important than simply maintaining a healthy body weight in improving neonatal outcomes. This
idea is novel and innovative as it is previously unexplored in pregnancy and pregnancy
outcomes.

In addition to determining the effect of regular physical activity on neonatal outcomes,
measuring maternal lipid metabolism and oxidative stress profiles will provide valuable
knowledge about mechanisms responsible for improved outcomes in physically active pregnant
women. Also, measuring maternal lipid metabolism and oxidative stress profiles during
exercise is novel and clinically insightful. This paradigm holds the potential to reveal
alterations in maternal metabolism and/or oxidative stress profiles that may not be detected
when measuring these factors at rest. Additionally, measuring metabolism and oxidative
stress during exercise is clinically useful by providing information about maternal
metabolism during activities that will mimic daily lifestyle tasks such as childcare,
household chores, etc. (~3-5 METS). Thus, this study design will provide us with valuable
insight and enhanced understanding of maternal lipid metabolism and oxidative stress
profiles during everyday lifestyle activities.

METHODS

Subjects:

All women who seek pre-natal care at the Women's Health Clinic at Barnes Jewish
Hospital/Washington University will be screened for inclusion BMI by history at the clinic.
Subjects will be recruited late in their 2nd trimester at the women's health clinic after
asking about their exercise habits. All patients who meet criterion with on-going
pregnancies will be approached for enrollment in the study. This study will compare 2 groups
of pregnant women between 30 and 35 weeks gestation. The first group will inactive obese
women and the other will be active obese women. We will recruit 15 subjects per group
(N=30). Groups will be race-matched.

Sample Size Calculation Data from Pomeroy et al.in 2012 stated that when using an
accelerometer to measure physical activity and air displacement plethysmography to measure
neonatal body composition (the same measurements we are proposing to use), the Spearman
correlation coefficient showing the association between maternal physical activity level and
neonatal fat free mass is r=0.5226. Using this R-value and an alpha of 0.05, 30 total
participants (15 per group) are needed to adequately power our study at .85 (beta=.15).

Study Procedures:

All study procedures will be performed at the Washington University School of Medicine
(WUSM) Institute for Clinical and Translational Sciences Clinical Research Unit (CRU).

CRU Visit #1 of 2: Body composition and Fitness Assessment (32-37 weeks gestation):

Maternal Body Composition:

A skin fold measurement will be performed to determine maternal body composition (% body
fat). This will be done by pressing folds of the skin at 7 sites with a caliper and
recording its thickness as previously described28.

Maternal Physical Fitness Levels:

Maternal fitness levels will be assessed using a submaximal cycle test on a recumbent
bicycle. Subjects will sit comfortably on the bicycle while the pedals are properly adjusted
so there is a slight bend in the knee when legs are extended. They will then complete the
YMCA submaximal multistage cycle ergometer test according ACSM's guidelines for exercise
testing and prescription27. Sady and colleagues concluded that the VO2-Heart Rate
extrapolation method is the most precise way to predict VO2max in pregnancy29, and the YMCA
test utilizes this method. A 3-lead ECG will be applied to monitor heart rate during the
exercise test.

Maternal Physical Activity Levels:

Daily maternal physical activity will be assessed in the week following these tests using
the ActiGraph GT3X+ accelerometer (ActiGraph LLC, Pensacola, FL) in order to objectively
measure daily physical activity levels. ActiGraph data will be collected for seven
consecutive days on the non-dominant wrist at 30 Hertz. Time spent in sedentary and active
(light, lifestyle, moderate, vigorous, and very vigorous) activities will be calculated
using algorithms from Freedson and colleagues using ActiGraph software30. We will also
measurement maternal physical activity levels subjectively using the Pregnancy Physical
Activity Questionnaire (PPAQ). The PPAQ is a valid and reliable instrument to measure
physical activity levels during pregnancy31. Not only will the PPAQ provide us with
additional details about their activity levels, but it will allow us to account for
activities that the actigraph may be unable to detect (i.e. riding a stationary bicycle).

Dietary Intake and Composition:

In order to account for differences in diet, subjects will complete the National Institutes
of Health's Dietary History Questionnaire II. This dietary assessment has been rigorously
validated32 and is widely used among many different populations. Previous literature also
demonstrates that dietary history questionnaires are valid and reproducible among pregnant
populations33.

CRU Visit #2 of 2: Lipid metabolism during exercise study (32-37 weeks gestation):

After obtaining height, weight, and vitals, a catheter (IV) will be placed in a hand vein
and heated in a warming box prior to each blood draw. Participants will rest for
approximately 30 minutes prior to measuring lipid oxidation rate using indirect calorimetry
(True One 2400, Parvo Medics, Sandy, UT). Participants will lay supine while a hood device
is placed over their head for 15 minutes to measure oxygen consumption and carbon dioxide
production in order to determine lipid oxidation rate34. After the initial indirect
calorimetry measurement is taken, basal blood collection will be obtained. Basal insulin and
glucose levels will be used to calculate maternal insulin resistance via a homeostatic model
assessment-insulin resistance (HOMA-IR). After this blood draw, participants will exercise
at approximately 50% of their predicted VO2max (based on the YMCA submaximal cycle test) for
30 minutes on the recumbent cycle ergometer (Lode Corvial, InMed, New South Wales,
Australia). Blood will be collected at various time points during exercise. Indirect
calorimetry (using a mouthpiece, nose clip, and exercise version of the software) will also
be performed for 2 minutes at a time to measure lipid oxidation and total body oxygen
consumption during low-level exercise. After exercise termination, participants will return
to a supine position. Recovery blood draws will be taken and indirect calorimetry will be
performed. See figure below for outline of visit 2 procedures.

Blood drawn at different time points will be used to measure glucose, insulin, free fatty
acids, reactive oxygen species (F2- isoprostanes by mass spectrometry (also referred to as
8-iso-PGF2α)35), and total antioxidant capacity (Total Antioxidant Capacity Assay (TAC),
Cell Biolabs, Inc., San Diego, CA). All of these measurements will help us to better
understand insulin resistance, oxidative stress, and mechanisms that could be contributing
to either condition.

Parturition:

At parturition, maternal weight will be measured and gestational weight gain will be
determined. Neonatal weight, length, and head circumference will also be obtained. Infant
HOMA-IR and fatty acid delivery to the fetus will be determined by measuring umbilical cord
plasma glucose, insulin, and fatty acid concentrations at parturition. Within 48 hours of
delivery, neonatal body composition (fat and lean mass) will be measured by skin fold
thickness measurement and by air displacement plethysmography (Pea Pod, Life Measurement,
Inc., Concord, CA) in the CRU at WUSM. A summary of measurements can be found in Appendix
III. A summary of our overall study design can be found in Appendix IV.

Statistical Analysis: Repeated measures ANOVA (group x time) will be used to compare lipid
oxidation rates and oxidative stress profiles between the 2 groups during pregnancy before,
during, and after exercise. Pearson product moment correlation coefficients for normally
distributed variables and Spearmen's rank order coefficients for non-normally distributed
variables will be used to examine the relationships between maternal lipid oxidation rate,
plasma oxidative stress markers, and neonatal metabolic outcomes. We may also use a
regression analysis to examine the relationship between maternal physical activity levels in
obese women and neonatal body composition and/or insulin resistance (similar to what has
been done in normal weight pregnant women by Pomeroy et al. 2012)26.

Inclusion Criteria:

- . Age 18-44 2. Confirmed singleton viable pregnancy with no fetal abnormalities at
routine 18-22 ultrasonography 3. Obese: Pre-pregnancy BMI between 30 and 45 kg/m2 4.
Receipt of prenatal care and plans to deliver at Barnes-Jewish Hospital 5. Inactive:
< 30min of low intensity activity (>1.5 METS) all or most days of the week Physically
Active: >150 minutes/week of moderate to high intensity activity 6. Completion of a
normal routine, standard of care 1 hour 50 gram gestational diabetes screen

Exclusion Criteria:

1. Multiple gestation pregnancy

2. Inability to provide voluntary informed consent

3. Current use of illegal drugs (cocaine, methamphetamine, opiates, etc…)

4. Current smoker who does not consent to cessation

5. Current usage of daily medications by class: corticosteroids, anti-psychotics (known
to alter insulin resistance and metabolic profiles)

6. History of gestational diabetes, pre-pregnancy diabetes or prior macrosomic (>4500g)
infant (each elevate the risk for gestational diabetes in the current pregnancy, or
undiagnosed gestational diabetes)

7. History of heart disease.