SYSTEMATIC LITERATURE REVIEW: MATERNAL CHOLESTEROL LEVELS DURING PREGNANCY By Mallory Taby Doan A THESIS Submitted to Michigan State University in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE Epidemiology 2011 ABSTRACT SYSTEMATIC LITERATURE REVIEW: MATERNAL CHOLESTEROL LEVELS DURING PREGNANCY By Mallory Taby Doan Elevations in the cholesterol profiles of non-pregnant individuals are a cause for medical attention due to the health complications associated with high cholesterol. Current research suggests that during a normal pregnancy cholesterol levels increase, but the nature and extent of the increase are not well described. In pregnancy, this elevation is considered normal and does not alarm medical personnel or require medical intervention. Although this rise in lipids does not yet warrant great concern, research has yet to agree on the reasons for, and effects of, this change. This systematic analysis reviews the literature on maternal cholesterol and cholesterol sub-fractions and their changes in pregnancy, focusing on total cholesterol, low density lipoprotein cholesterol and high density lipoprotein cholesterol. In 21 studies, the average increase in total cholesterol from the first to third trimester was 46% and in low density lipoprotein cholesterol a 60% increase was observed, both peaking in the third trimester. However, high density lipoprotein cholesterol levels only increased 10% on average and peaked during the second trimester. Theories about this increase in cholesterol, and areas for future research are also discussed. There is insufficient understanding of the role of cholesterol in pregnancy, making it difficult to define target cholesterol levels during pregnancy. Further research is needed to help researchers better understand the mechanism behind pregnancy cholesterol changes and health complications that may result from abnormal levels. ACKNOWLEDGEMENTS First and foremost, I would like to thank each of my committee members for their support and guidance, Dr. Ronald Horowitz, Dr. Qing Lu, Dr. Joel Maurer, with a special thanks to my advisor Dr. Nigel Paneth. To my fellow T-32 fellowship trainees, current and past members, thank you for your guidance and advice. Secondly, my family, especially my mom and grandma, are deserving of a special thanks, as they have all been my support system from the beginning. Their prayers and encouragement have helped in keeping me on track. In addition, I would also like to thank my friends for their inspiring words and unconditional love. Without them I would not have made it this far. Lastly, I would like to individually thank Lynette Biery, Cortney Davis, McKenzie Maynor, Yasaman Back, and Leslee Wilkins for going above and beyond the call of duty and taking time to look over, critique, and comment on all the work that went into the completion of this degree. iii TABLE OF CONTENTS LIST OF TABLES…………………………………………………………………………...........v LIST OF FIGURES………………………………………………………………………............vi LIST OF SYMBOLS or ABBREVIATIONS……………………………………………...........vii CHAPTER 1 BACKGROUND………………………………………………………………………….............1 Cholesterol………………………………………………………………………...........…1 Cholesterol...............................................................................................................1 Cholesterol’s Chemical Structure and Function......................................................2 Cholesterol Measurements...................................................... ................................3 Health Complications Associated with a High Risk Cholesterol Profile................4 CHAPTER 2 METHODS……………………………………………………………………..............................6 Search Criteria.....................................................................................................................6 Inclusion Criteria.................................................................................................................7 Statistical Methodology.......................................................................................................7 CHAPTER 3 RESULTS……………………………………………………………………................................8 Description of Selected Studies...........................................................................................8 Cholesterol Trends During Pregnancy.................................................................................8 Total Cholesterol......................................................................................................8 Low Density Lipoprotein Cholesterol.....................................................................9 High Density Lipoprotein Cholesterol...................................................................10 CHAPTER 4 DISCUSSION………………………………………………………………………………........11 Summary of Findings.........................................................................................................11 Speculations as to Why Cholesterol Increases...................................................................12 Further Research................................................................................................................15 Strengths and Limitations..................................................................................................17 APPENDICES...............................................................................................................................19 REFERENCES……………………………………………………………………………..........36 iv LIST OF TABLES Table 1: National Cholesterol Education Program cholesterol guidelines for the non-pregnant population………………………………………………………………………….....20 Table 2: Google search terms used to search the literature….......................................................21 Table 3: PubMed search terms used to search the literature..........................................................22 Table 4: 14 inclusion criteria for the quantitative analysis on maternal lipid profiles during pregnancy………………………………………………………………………….......23 Table 5: Summary statistics for total cholesterol by longitudinal study........................................25 Table 6: Summary statistics for total cholesterol by cross-sectional study...................................26 Table 7: Summary statistics for low density lipoprotein cholesterol by longitudinal study.........27 Table 8: Summary statistics for low density lipoprotein cholesterol by cross-sectional study.....28 Table 9: Summary statistics for high density lipoprotein cholesterol by longitudinal study.........29 Table 10: Summary statistics for high density lipoprotein cholesterol by cross-sectional study..30 v LIST OF FIGURES Figure 1: Total Cholesterol levels during and after gestation....……………………………........31 Figure 2: Total Cholesterol ………...................................………………………………………32 Figure 3: Total Cholesterol levels by study type……………………………………...................33 Figure 4: Low Density Lipoprotein Cholesterol............................................................................34 Figure 5: High Density Lipoprotein Cholesterol...........................................................................35 vi KEY TO SYMBOLS OR ABBREVIATIONS g / ml Grams per milliliter HDL High Density Lipoprotein HDL-C High Density Lipoprotein- Cholesterol LDL Low Density Lipoprotein LDL-C High Density Lipoprotein- Cholesterol mg / dL Milligrams per deciliter NCEP National Cholesterol Education Program TC Total Cholesterol TG Triglycerides vii CHAPTER 1: BACKGROUND The 1999- 2006 National Health and Nutrition Examination Survey reported that 47% of the United States population surveyed between the ages of 20 and 47 was at risk of cardiovascular health complications resulting from total cholesterol (TC) levels greater than or equal to 200 mg/dL [1]. Interestingly, pregnant women have high cholesterol levels during the second and third trimester of pregnancy, often to levels that are considered quite dangerous in the non-pregnant population [2]. Elevations in cholesterol lipid levels during pregnancy do not gain medical attention, often go untreated, and little is known as to why these changes occur. The present quantitative analysis explored and examined current research on cholesterol levels during pregnancy, focusing on TC, low density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol (HDL-C) molecules. To begin, basic descriptions of cholesterol and its two major subtypes will be provided. Information will then be provided on optimal cholesterol levels and the health complications associated with high risk cholesterol levels. A. Cholesterol I. Cholesterol Cholesterol is an essential waxy lipid molecule that is required for proper cell development and structure [3-7], and is necessary for the production of steroid hormones and bile acids [3-8]. This lipid is a precursor to sex hormones, including estrogen, progesterone and testosterone [5, 8]. Additionally, cholesterol molecules have a role in maintaining membrane permeability and fluidity [2]. 1 II. Cholesterol’s Chemical Structure and Function Cholesterol can be found in two forms, cholesterol ester and free cholesterol [8]. Free cholesterol is embedded in the surface layer of the lipid molecule [8]. Cholesterol ester molecules are cholesterol molecules with an attached fatty acid molecule [8]. Cholesterol ester is highly hydrophobic; therefore, it is found in the center of a lipid molecule, shielded from the water-based blood [8]. Free cholesterol and cholesterol ester can be found on low density lipoprotein (LDL) and high density lipoprotein (HDL) molecules in varying concentrations [8]. LDL is low in density but relatively large in size in comparison to HDL molecules. The density of an LDL molecule ranges from 1.019 – 1.063 g/ml [8]. LDL molecules contain 10% free cholesterol and 40% cholesterol ester [8], and carry approximately 60% - 70% of an individual’s cholesterol (LDL-C) in the blood [2, 9]. The structure of this relatively large lipoprotein is unique in that most of the time it only consists of one apolipoprotein, apo B-100 [9]. Occasionally, small amounts of apolipoproteins C and E can be found in LDL molecules [8]. Given the large size of an LDL molecule and its movement to the peripheral tissues, it is hypothesized that LDL-C builds up on the artery walls thereby narrowing the arteries [8]. The negative effects associated with increased levels of LDL-C have generated an alias for LDL-C as the “bad cholesterol” [10]. HDL molecules are small lipoprotein molecules with a high density between 1.063 – 1.21 g/ml [8]. Unlike LDL, which contains only one apolipoprotein, HDL is comprised of two different apolipoproteins, apo A-I and apo A-II [2]. HDL contributes 20%-30% of the body’s cholesterol supply, measured as HDL-C [2]. HDL molecules are thought to transport cholesterol molecules from the artery walls back to the liver. The transport of HDL-C from the arteries to the liver decreases an individual’s risk of heart attack or stroke; therefore, high HDL-C levels 2 reduce ones risk of cardiovascular complications by reducing the cholesterol plaque buildup on the artery walls [8]. Due to HDL-C’s purported role in ridding the arteries of cholesterol buildup, HDL-C is known as the “good cholesterol” [10]. III. Cholesterol Measurements The National Cholesterol Education Program (NCEP), launched in 1985 by the National Heart, Lung, and Blood Institute, has developed cholesterol guidelines and standards for testing each component of cholesterol [11]. TC levels are used as overall predictors of coronary health complications and are equal to the sum of LDL-C, HDL-C and very low density lipoprotein cholesterol levels. The NCEP recommends non-pregnant TC levels measure less than 200 mg/dL to minimize risk of potential atherosclerosis and coronary heart disease [2]. Cholesterol is routinely measured in fasting subjects, although Weiss et al. have shown that TC measurements are not statistically different in fasting and non-fasting subjects [12]. Higher levels of HDL-C are associated with a decreased risk of cardiovascular complications. Optimal HDL-C levels, listed by the NCEP, in men and non-pregnant women, are greater than 60 mg/dL [2]. The primary focus of public health initiatives is the lowering of LDL-C levels [2]. LDLC levels less than 100 mg/dL are categorized as reflecting a low risk for cardiovascular health complications [2]. In past years, direct measurements of LDL-C were obtained through time consuming and costly testing. As a result, direct measurements of LDL-C are not routinely used in clinical care; rather, calculated measurements using the Friedwald equation are used more often [13]. LDL-C levels are calculated from total cholesterol, HDL-C and triglyceride (TG) measurements. Equation 1 shows how to calculate LDL-C levels for measurements with mg/dL units [13]. 3 Equation 1 (mg/dL): Given the relationship between TG and LDL-C, LDL-C measurements using the Friedwald equation are unreliable in non-fasting participants. The Friedwald equation also proves to be inaccurate when TG levels are above 250 mg/dL [14]. Current research has developed less expensive methods for determining LDL-C levels directly. These direct measurements use electrophoresis and can provide more reliable, accurate results when completed on both fasting and non-fasting samples [15]. NCEP guidelines are listed in Table 1 and have been used as a reference in this study [2]. IV. Health Complications Associated with a High Risk Cholesterol Profile Recommendations for non-pregnant individuals from the NCEP are in place to address health complications that can occur with abnormally high TC and LDL-C levels. High TC and LDL-C concentrations in men and non-pregnant women have been linked to an increased risk of atherosclerosis, cardiovascular disease, coronary heart disease, myocardial infarctions, stroke and peripheral vascular disease [2]. Non-pregnant individuals with elevated TC and LDL-C levels are advised to seek treatment to help lower TC and LDL-C to optimal levels, while taking into consideration all other risk factors for coronary heart disease [2]. Cholesterol predicts the risk of cardiovascular complications in women differently than in men [2, 16, 17], although little is known about these differences. As such, there is a need for additional research in this area. Patients with high levels of TC and LDL-C, and low levels of HDL-C are initially advised to alter their diet and exercise [2]. If diet and exercise do not help improve lipid values, 4 non-pregnant patients are often prescribed cholesterol lowering medications to help control cholesterol lipid levels and decrease the risk of health complications associated with hypercholesterolemia [2]. 5 CHAPTER 2: METHODS A. Search Criteria Both Google Scholar and PubMed were used to search the literature for publications on maternal lipid levels during pregnancy. 32 different search terms were used when searching Google Scholar. These search terms are listed in Table 2. Results varied from 4,750 results upwards to 49,900 results depending on the search term used. Regardless of the search term, the first 100 results were evaluated for all 32 different searches. If no publications, relevant to the proposed research topic, were found within the first 100 results, results beyond this were not considered. For search terms that provided pertinent literature, results were evaluated until 200 consecutive results provided no additionally applicable publications. 16 search terms were used in PubMed, listed in Table 3. Although the search used “lipids” or “cholesterol” in Google Scholar, the word “lipids” was not used when searching PubMed. This change was made after conducting preliminary searches on lipids and cholesterol in PubMed. Cholesterol, which is a type of lipid, is a more specific term to use when searching the literature. As such, applicable search results did not differ between “lipids” and “cholesterol”. Unlike Google Scholar, PubMed did not differentiate between the terms “pregnancy” and “gestation” or “during” and “in”. As a result, PubMed provided four different searches. Three out of the four searches resulted in 410 results or less. The three searches were: “maternal cholesterol changes in (during) pregnancy (gestation)”, “cholesterol changes in (during) pregnancy (gestation)”, and “maternal cholesterol in (during) pregnancy (gestation)”. The fourth search term, “cholesterol in (during) pregnancy (gestation)” produced 1,381 results. In PubMed, all results from each of the four search algorithms were evaluated for relevant 6 literature. In addition to searching Google Scholar and PubMed, references from the included literature were checked in an effort to reduce the chances of missing relevant research. B. Inclusion Criteria Prior to searching the literature, 14 inclusion criteria were created to help search the literature for relevant studies. The search was restricted to healthy human subjects in developed countries. A developed country was categorized as a country classified as very high or high human development by the United Nations Human Development Program [18]. If explicit information was not provided on a single inclusion criterion, that study was included in the analysis. For example, if a study did not clearly state the average weight of study subjects, but abided by all other study criteria, the given study was included. Abstracts were required to help sort through the results. The criteria were selected in an effort to provide the most generalizable findings. These search criteria, and justifications for each, are listed in detail in Table 4. C. Statistical Methodology 21 papers (14 longitudinal; seven cross-sectional) were used for the quantitative analysis. A weighted average cholesterol level was calculated for each trimester based on the information obtained from the included studies [19]. For each trimester, an average cholesterol level was obtained from each individual study. This average was then weighted by the sample size of the study to calculate an overall weighted average. Publications with multiple measurements per trimester were given a greater weight than those with a single time point per trimester. In order to give a greater weight to studies with multiple measurements, the cholesterol data was multiplied by the sample size at each time point during the given trimester. This estimate was then combined with those from the remaining studies to produce an overall weight for the trimester. 7 CHAPTER 3: RESULTS A. Description of Selected Studies 14 longitudinal studies, published between 1985 and 2010, and seven cross-sectional studies, published between 1964 and 2007 were included in this review. The average age of women within the selected studies was 29 years. 19 of the 21 studies were conducted in European countries [20-38]. Of the remaining, one was in the United States [39] and the other in Japan [40]. One of the selected studies stratified results on pre-pregnancy body mass index [39]. Data from this study was used for the normal weight group only, although significant changes in cholesterol were also seen in the overweight/obese group. Dietary intervention was studied in one of the included studies [27]. Cholesterol results were stratified on the assigned dietary group. For this review, data was included from the control group only, despite significant increases in cholesterol levels in the intervention group. None of the studies provided results stratified on demographic variables, such as race, income and education. B. Cholesterol Trends During Pregnancy I. Total Cholesterol This review found total cholesterol increased by 46%, on average, during a term pregnancy [20-38, 40, 41]. Until the 12 th week of gestation, TC levels in the pregnant population tend to be similar to non-pregnant levels [23, 24, 30]; however, these findings are not consistent [29]. Although there are inconsistencies in the literature as to exactly when TC levels begin to increase in the first trimester, an initial rise in TC from late first to early-mid second trimester was observed in all studies. This rate of increase, however, differed by study. For example, Vahratian et al. found a small subset of women experienced total cholesterol 8 levels over 300 mg/dL as early as 20 weeks gestation [39], whereas Ogura et al. reported an average TC of 202 mg/dL at 20 weeks [40], despite similar averages in TC during the first trimester. In 14 longitudinal studies, involving 566 women, TC levels increased 28% on average from 184 mg/dL in the first trimester to 223 mg/dL in the second trimester [22-33, 40, 41]. In these same studies, TC levels continued to increase into the third trimester by an additional 16%. Less information is provided, from the selected studies, on the changes in lipid levels after pregnancy. Nine studies, based on linked follow-up, showed that total cholesterol levels begin to decline after pregnancy; however, as seen in Figure 1, the rate of decrease is less than the rate of increase during pregnancy [21, 22, 25, 26, 31, 33, 35, 38, 40]. Results did not differ between fasting and non-fasting samples from either serum and plasma, as seen in Figure 2. As seen in Figure 3, cross-sectional studies reported slightly higher average TC levels per trimester than longitudinal studies; however, the percent increase across trimesters did not differ. These study results are summarized in Table 5 (longitudinal) and Table 6 (cross-sectional). II. Low Density Lipoprotein Cholesterol As with TC, LDL-C levels increase during pregnancy and peak in the third trimester. In 437 women from ten longitudinal studies, LDL-C levels increased 59% on average from the first to the third trimester [22-24, 26, 27, 30-32, 40, 41]. Cross-sectional studies showed an overall increase from first to third trimester of 60% [35-38]. These studies, both longitudinal and cross-sectional, showed LDL-C levels in the first trimester had an average of 100 mg/dL and an average third trimester measurement of 158 mg/dL. Second trimester levels increased 35% over first trimester levels and averaged 133 mg/dL. 9 Minimal information on postpartum LDL-C levels is available. Among the included studies, there were only six data points on LDL-C levels, limited to first eight weeks postpartum [22, 26, 31, 35, 38, 40]. Given the lack of information of postpartum measurements for LDL-C, these data are not shown. The study results are summarized in Table 7 and Table 8, and Figure 4. III. High Density Lipoprotein Cholesterol Unlike trends in TC and LDL-C, HDL-C levels increase at a reduced rate and information is conflicting on when these levels begin to increase as well as when they peak. Berge et al. and Tranquilli et al. show HDL-C levels in the first trimester are significantly higher than the non-pregnant population [21, 29], Fahraeus et al. report HDL-C levels are significantly lower in the first trimester [31], whereas four other studies show first trimester measurements to be insignificantly different from the non-pregnant control [23, 30, 36, 38]. 75% of referenced studies, both cross-sectional and longitudinal, reported HDL-C levels peaked in the second trimester [21-24, 26, 27, 29, 35-38, 41]. Winkler et al. found HDL-C levels highest in the first trimester [30]. Fahraeus and Munoz both described peaks around 28 th weeks gestation [31, 32], and Ogura et al. found HDL-C levels to peak in the 37 week of gestation [40]. In 16 studies (11 longitudinal, 5 cross-sectional), HDL-C levels increased 18% on average from the first trimester to the second [21-24, 26, 27, 29-32, 35-38, 40, 41]. In these same studies, following the increase in the second trimester, HDL-C decreased 7% in the third trimester to 67 mg/dL. Third trimester levels were 10% greater than levels in the first trimester. Thus unlike TC and LDL-C, HDL-C tends to decline in the third trimester, although the decline is small. 10 Similar to LDL-C, data on postpartum HDL-C levels are limited; therefore these data have been omitted from my results [21, 22, 26, 31, 35, 38, 40]. These study results are summarized in Table 9, Table 10, and Figure 5. 11 CHAPTER 4: DISCUSSION A. Summary of Findings The presented literature review shows levels of TC, as well as LDL-C, and HDL-C significantly increase during pregnancy, but with different time courses and magnitudes. TC and LDL-C both increase to levels that are looked upon as high risk in the non-pregnant population. The overall rate of increase is greatest from first to second trimester in all three cholesterol subgroups measured. Rates of change for HDL-C differ from those of LDL-C and TC from the second to third trimester, in that they begin to decrease, although, as shown, these findings are variable in the literature. Cross-sectional and longitudinal study results were comparable; therefore, cross-sectional studies may provide as much valuable information as longitudinal studies for population trends. Cross-sectional studies, however, cannot be used when analyzing individual percent changes in cholesterol as the main exposure. Based on the presented studies, elevations in cholesterol levels during pregnancy occur regardless of geographic location, diet during pregnancy [27] and pre-pregnancy body mass index [39]. In the non-pregnant population, both diet and body mass index have been shown to be positively associated with cholesterol levels [2]. A similar relationship between diet and cholesterol is seen in the pregnant population; however, controlling for diet, at least in two studies, does not fully explain the significant changes in cholesterol levels during pregnancy [27, 42]. A cholesterol lowering diet was introduced to 141 Norwegian pregnant women and was found to significantly reduce cholesterol levels, although, the 21% increase in TC from second to third trimester in the intervention group was only 4% lower than the average reported rates in the control group [27]. An inverse relationship between pre-pregnancy body mass index and 12 changes in cholesterol levels from second to third trimester was found in a predominantly white cohort of 142 women [39]. Vahratian et al. found significantly lower rates of change in cholesterol levels in the overweight and obese population when compared to women of normal weight, although controlling for pre-pregnancy body mass index does not eliminate the significant changes in cholesterol levels [39]. Reduced rates of change in overweight and obese women led to lower absolute cholesterol levels in the third trimester, compared to third trimester levels in normal weight women [39, 41]. It will be interesting to see if there is a correlation between relatively lower cholesterol levels in overweight and obese women compared to normal weight women in the third trimester and health complications in the offspring. None of the included studies stratified on race or ethnicity. A study on a small group of women, not included in this review, showed significantly lower levels of TC and LDL-C concentrations in black (n=15) women compared to white (n=15) women throughout pregnancy [43]. Conversely, in the non-pregnant population, a study of 5,116 men and women in the United States found black women have significantly higher LDL-C levels than white women [44]; however, the NCEP reports, on average, there are no significant differences in cholesterol levels between races [2]. B. Speculations as to Why Cholesterol Increases The elevation in lipids in pregnancy may be an adaptive change that takes place for proper fetal development and for the general safeguarding of the pregnancy [3, 5-7, 29]. Early in pregnancy, the fetus does not produce its own cholesterol, but rather, is dependent upon the transport of maternal cholesterol across the placenta [6, 45]. Maternal cholesterol levels may rise during the first two trimesters of pregnancy to address the needs of the fetus. Roux et al. show developmental complications and deficiencies in the rat fetus, resulting from insufficient 13 maternal cholesterol production [46] and Porter et al. review these deficiencies in human fetus’ [47]. As shown in this systematic review, the rate of change from the second to third trimester is less than that from first to third, perhaps as a result of the development of an endogenous source of cholesterol for the fetus [7]. Napoli et al. found fetal cholesterol levels to be highly correlated with maternal levels until the sixth month of pregnancy, at which time the fetus begins to produce its own cholesterol [48]. As the fetus begins to produce its own cholesterol, the demand for maternal cholesterol lessens thereby resulting in a decrease in the rate of increase in maternal cholesterol. Fetuses who fail to produce their own cholesterol, or produce cholesterol at a reduced rate are often diagnosed with Smith-Lemi-Opitz Syndrome and congenital defects [3, 47], thereby showing the importance of cholesterol in development. It would be interesting to evaluate the health of a human fetus born to mothers lacking the early to mid-pregnancy increase in cholesterol levels to confirm this hypothesis. White pregnant women with low TC levels (< 159 mg/dL) during the second trimester have been shown in the study to be at an increased risk of preterm delivery [49]. In the same study, this relationship was not found in black women; rather an increased risk of preterm delivery was associated with high maternal TC levels (≥ 159 mg/dL). A study looking at th Michigan women during their pregnancy found women with cholesterol levels in the bottom 10 percentile had an increased risk of medically indicated preterm delivery, where women in the top th 30 percentile had an increased risk of spontaneous preterm delivery, irrespective of race.[50] The biological pathway explaining the potential relationship between maternal cholesterol levels and preterm delivery is in need of additional research. 14 Cholesterol during pregnancy may also be a substrate for the increases in hormone levels, as cholesterol molecules are precursors to hormones, including those with an irreplaceable role during pregnancy [5, 8, 51, 52] and in the placenta [6, 53, 54]. In addition, there is a positive relationship between hormones, such as estrogens and progesterone, and cholesterol [37, 55, 56]. Without increases in cholesterol during pregnancy, hormone levels may not be able to adequately increase and thereby support the pregnancy [3]. However, the opposite causal relationship between cholesterol and hormones may be true. Cholesterol levels have been found to vary 5% - 8% depending upon where a woman is in her menstrual cycle and this change in lipids is thought to be positively correlated with changes in hormone levels [55]. Increases in hormonal levels, such as during a woman’s menstrual cycle, may influence or cause the changes in cholesterol levels. Just as changes in hormonal and cholesterol levels are correlated in the non-pregnant woman, perhaps this same type of relationship is seen within pregnancy with a greater magnitude. Increases in cholesterol are found within all pregnant women, even those with familial hypercholesterolemia [42, 57]. In a group of 19 women with this genetic disorder, the average th level of TC in the 36 week of pregnancy was 449 mg/dL which increased 28% from 352 mg/dL at the end of the second trimester [57]. A rise in cholesterol levels in this subset of women, who already have high levels of cholesterol prior to pregnancy, poses an interesting question. If cholesterol is needed to maintain a healthy pregnancy, these women already have a relatively large absolute amount of cholesterol. An increase in cholesterol, if any, should be significantly less in these women compared to their healthy counterpart; however, this is not seen. Rather, there appears to be some biological process driving cholesterol levels to increase during pregnancy regardless of the baseline measurement. It follows from the preceding that the 15 percent change in cholesterol levels may hold more information than absolute values at given time points during pregnancy. More research is needed to compare health complications in the mother and fetus between women with a large percent change in lipids and women with a lesser percent change. C. Further Research Research examining the physiological basis for changes in cholesterol in pregnancy is needed. At the same time, research on potential health complications associated with variation in lipid levels is also needed. Limited data exists on the potential role racial and socioeconomic disparities play in pregnancy related cholesterol levels. It would be interesting to further explore data looking for differences in the changes of maternal cholesterol stratified on race as well as socioeconomic status and compare these findings to those reported by Patrick et al. [43]. Research as it pertains to black women in the United States is needed, as out of the selected studies, only one study was conducted within the United States [41] and none of the selected studies stratified on race. Elevations in cholesterol levels seemed to be inevitable in the women referenced, although the degree of change depended upon the population being studied. Research shows maternal hypercholesterolemia (TC > 240 mg/ dL) during pregnancy increases the risk and size of plaque build-up on preterm fetal arteries [48, 58, 59], although the biological mechanism is currently unclear. It is also unclear as to when and for how long hypercholesterolemia must occur before adverse effects are seen. Women with chronic hypercholesterolemia (before, during and after pregnancy) are thought to place their preterm fetuses at an even great risk of atherosclerosis [48, 58, 59]. However, as fetuses begin producing their own cholesterol, at roughly 6 months gestation, they become less dependent on maternal cholesterol and the 16 correlation between the two cholesterol levels begins to decline, with lowest levels of association seen at term [48]. Additional research is needed to analyze the long term effects of fetal plaque build-up and the risk of subsequent cardiovascular health complications in both term and preterm infants. When studying this relationship one must also take into consideration the genetic aspect of hypercholesterolemia compared to pregnancy induced hypercholesterolemia as well as the environmental effects (i.e. diet and exercise) on cholesterol levels within the newborn and developing child. Increased parity has been shown to have a protective effect against certain health complications, such as breast and ovarian cancer, but a negative effect against others including cardiovascular disease [60-63]. TC levels have been found to be significantly lower in multiparous women [63, 64], although is finding is not consistent [63]. LDL-C levels are not significantly associated with increases in parity, and HDL-C levels and parity tend to be inversely related [61, 64, 65]. Additional research is needed to evaluate the relationship between low HDL-C levels, which are a risk factor for atherosclerosis and cardiovascular disease, and multiparity. In addition, atherosclerotic plaques were positively correlated with parity in a homogeneous cohort of women from the Netherlands [64]. Ness et al. found a significant relationship between cardiovascular disease and parity in women from the United States, with six or more pregnancies [63]. Hinkula et al. studied causes of mortality in Finnish grand multiparous women (≥ 5 deliveries) and grand grand multiparous women (≥ 10 deliveries) [62]. This study found mortality rates from cardiovascular complications were higher in grand grand multiparous women when compared to grand multiparous women, and both rates were higher than the national average in Finland. Hinkula speculates changes in body mass index and decreased HDL-C levels from multiparity as two potential reasons for these women being at an 17 increased risk. Lawlor et al. attribute the positive relationship between cardiovascular disease and parity to a combination of both biological and life-style changes resulting from multiple pregnancies [61]. Life-style changes may play a role in this association between parity and cardiovascular health, yet, a similar relationship has been found between cardiovascular disease and the use of contraceptives, which also causes changes in hormones and cholesterol levels [66, 67]. This similarity in health complications between parity and oral contraceptive usage is interesting, warranting additional research on the role hormones play in cardiovascular health and if this relationship is confounded by cholesterol levels. The literature is also lacking studies looking at repetitive insults to the cardiovascular system in multiparous women resulting from multiple changes in cholesterol levels during pregnancy and if this relationship is affected by pregnancy spacing. Additional research is also needed in high risk pregnancies, such as those affected by diabetes [68-71] and hypertension [33, 40, 72, 73], as current research remains inconclusive with regards to rates of cholesterol elevation in these conditions. D. Strengths and Limitations The presented systematic analysis is the first of its kind evaluating trends in maternal cholesterol during pregnancy. This thesis combines 21 studies, all looking at gestational cholesterol levels in the mom, and creates weighted averages for TC and the two main cholesterol subsets during each trimester. The literature was exhaustively searched, thereby minimizing the chance of missing qualified publications. Another strength of this review was the inclusion of studies with multiple time points during pregnancy. This criterion allowed for the analysis of change within the individual studies. Lastly, the large cumulative sample size of the 18 21 studies strengthens the presented results allowing for comparison with the findings of future studies. Despite strengths, the main limitation of this review was the exclusion of studies that did not provide quantitative values for their findings. Further research is needed to analyze cholesterol trends during pregnancy in the United States, as only one of the included studies looked at cholesterol levels in this location. In addition, this review did not adjust for racial differences in cholesterol levels, as this information was not provided in the included studies. 19 APPENDICES 20 TABLE 1: National Cholesterol Education Program cholesterol guidelines for the nonpregnant population [2] TOTAL CHOLESTEROL < 200 mg/dL 200-239 mg/dL ≥ 240 mg/dL LDL CHOLESTEROL < 100 mg/dL 100-129 mg/dL 130-159 mg/dL 160-189 mg/dL ≥ 190 mg/dL HDL CHOLESTEROL < 40 mg/dL ≥ 60 mg/dL Desirable Borderline High High Optimal Near optimal/above optimal Borderline high High Very High Low High 21 TABLE 2: Google search terms used to search the literature SEARCH TERM Cholesterol during gestation Cholesterol during pregnancy Cholesterol in gestation Cholesterol in pregnancy Lipids during gestation Lipids during pregnancy Lipids in gestation Lipids in pregnancy Maternal cholesterol during gestation Maternal cholesterol during pregnancy Maternal cholesterol in gestation Maternal cholesterol in pregnancy Maternal lipids during gestation Maternal lipids during pregnancy Maternal lipids in gestation Maternal lipids in pregnancy Cholesterol changes during gestation Cholesterol changes during pregnancy Cholesterol changes in gestation Cholesterol changes in pregnancy Lipids changes during gestation Lipids changes during pregnancy Lipids changes in gestation Lipids changes in pregnancy Maternal cholesterol changes during gestation Maternal cholesterol changes during pregnancy Maternal cholesterol changes in gestation Maternal cholesterol changes in pregnancy Maternal lipid changes during gestation Maternal lipid changes during pregnancy Maternal lipid changes in gestation Maternal lipid changes in pregnancy 22 SEARCH RESULTS 49,900 33,900 31,000 38,100 19,200 20,500 6,030 23,500 12,900 11,500 7,070 12,700 7,130 6,340 4,750 6,890 20,200 26,300 11,800 28,000 21,000 27,600 16,700 26,600 10,600 8,670 5,740 9,130 13,600 10,900 8,660 11,600 TABLE 3: PubMed search terms used to search the literature SEARCH TERM Cholesterol in pregnancy Maternal cholesterol in pregnancy Cholesterol changes in pregnancy Maternal cholesterol changes in pregnancy SEARCH RESULTS 1,381 406 238 75 23 TABLE 4: 14 inclusion criteria for the quantitative analysis on maternal lipid profiles during pregnancy INCLUSION CRITERIA Abstract available Average age of population younger than 35 years of age Data collected from developed world English publication Healthy mother Healthy term pregnancy Human model Minimum of 2 time points in consecutive trimesters Mothers of a healthy weight REASONING Used to determine eligibility of the study Over the age of 35 women are classified as having a high risk pregnancy. In under developed countries diets differ from those in developed countries [18]. In the non-pregnant state, diet effects cholesterol. Inability to interpret non-English writings. Inconclusive evidence exists as to the influence gestational diabetes and preeclampsia has on maternal cholesterol levels [33, 40, 68-72]. There is some research that reports maternal differences in lipid levels between term and pre-term deliveries [49]. The review summarized cholesterol during human pregnancy, therefore animal models were excluded. Looking at trends of cholesterol across pregnancy, cross-sectional and longitudinal studies with only cholesterol measurements during one trimester in pregnancy were eliminated. In addition, studies that provided data from the first and third trimesters only were also excluded. These studies were excluded in order to capture the second trimester peak in HDL-C. In the non-pregnant state weight is often used as a proxy for health. Those that are overweight prior to pregnancy maybe less healthy and may have elevated cholesterol levels. To reduce this potential for bias, studies with an average sample BMI ≥ 25 were excluded. Studies that provided separate results for normal weight and overweight populations were included, keeping only data from the normal weight stratum [41]. In addition, there is some evidence that lipids levels do indeed differ in pregnancy when stratified on maternal BMI [39]. 24 TABLE 4 (cont’d) INCLUSION CRITERIA No familial hypercholesterolemia Observational study on lipids -or data on a control group (if available) Report data values (not just graphics) Sample size greater than or equal to 15 Singleton pregnancy REASONING These women were excluded to eliminate the influence of familial hypercholesterolemia. In order to obtain a natural reflection of cholesterol levels during pregnancy, non-observational studies were excluded (unless data from a control group with no intervention was provided) [27]. In order to summarize research findings in the literature, studies that did not include data values were not included. The change in cholesterol levels during pregnancy differs between women. In hopes of incorporating representative averages from each selected study, studies with small (n < 15) sample sizes were excluded. For cross-sectional studies, each individual group of women studied had to have at least 15 women in order for the study to be included. Pregnancies with multiples are known to have differences from singleton pregnancies. 25 TABLE 5: Summary statistics for total cholesterol by longitudinal study st nd First Author (Year) Average 1 trimester mg/dL Average 2 trimester mg/dL Alvarez (1996) Belo (2004) Brizzi (1999) Darmady* (1992) Fahraeus* (1985) Jimenez* (1988) Khoury* (2005) Misra* (2011) Munoz (1999) Ogura (2002) Rosing (1989) Sanchez-Vera* (2005) Tranquilli (2004) Winkler (2000) 170 178 178 200 161 182 172 206 169 206 191 st Weighted 1 trimester average mg/dL 234 255 257 253 195 218 222 225 211 202 228 179 276 253 nd Weighted 2 trimester average mg/dL rd Average 3 trimester mg/dL 254 283 282 307 244 260 256 260 246 265 249 229 279 277 rd Weighted 3 trimester average mg/dL st 1 trimesternd 2 trimester % change 38% 43% 44% 27% 21% 20% 31% 2% 20% 34% 33% st Average 1 nd trimester-2 trimester % change nd 2 st trimester 1 trimester- -3 trimester % change 9% 11% 10% 21% 25% 19% 16% 16% 17% 31% 9% 28% 1% 10% nd Average 2 3 trimester % change 49% 59% 58% 53% 52% 43% 51% 19% 57% 35% 45% st Average 1 rd rd trimester -3 trimester % change rd rd trimester -3 trimester % change Summary Statistics 184 223 264 28% 16% 47% *Studies with multiple total cholesterol measurements per trimester. An average of these multiple measurements is reported. These studies were given a greater weight in the weighted average than those with one total cholesterol measurement per trimester. 26 TABLE 6: Summary statistics for total cholesterol by cross-sectional study st nd rd First Author (Year) Average 1 trimester mg/dL Average 2 trimester mg/dL Belo (2002) Berge (1996) Gratacos (1996) Lippi (2007) Martin (1999) Ordovas (1984) Piechota (1992) 176 226 179 173 224 175 192 st Weighted 1 trimester average mg/dL 252 274 227 243 266 218 244 nd Weighted 2 trimester average mg/dL 285 321 252 267 315 259 285 rd Weighted 3 trimester average mg/dL 195 243 277 Summary Statistics Average 3 trimester mg/dL st 1 trimesternd 2 trimester % change 43% 21% 27% 41% 19% 25% 27% st Average 1 nd nd 2 st trimester 1 trimester- -3 trimester % change 13% 17% 11% 10% 18% 19% 17% nd Average 2 3 trimester % change 62% 42% 41% 54% 41% 48% 48% st Average 1 rd rd rd rd trimester -3 trimester % change trimester -3 trimester % change 28% 27 trimester-2 trimester % change 13% 45% TABLE 7: Summary statistics for low density lipoprotein cholesterol by longitudinal study st nd First Author (Year) Average 1 trimester mg/dL Average 2 trimester mg/dL Alvarez (1996) Belo (2004) Brizzi (1999) Fahraeus* (1985) Jimenez* (1988) Khoury* (2005) Misra* (2011) Munoz (1999) Ogura (2002) Winkler (2000) 89 85 97 98 105 98 108 89 94 st Weighted 1 trimester average mg/dL 136 128 153 112 129 125 136 100 106 121 nd Weighted 2 trimester average mg/dL Average 3 trimester mg/dL rd 153 145 168 147 169 152 166 131 146 137 rd Weighted 3 trimester average mg/dL st 1 trimesternd 2 trimester % change 53% 51% 58% 14% 23% 38% -7% 19% 29% st Average 1 nd trimester-2 trimester % change nd 2 st trimester 1 trimester- -3 trimester % change 13% 13% 10% 31% 31% 21% 23% 31% 38% 13% nd Average 2 3 trimester % change 72% 71% 73% 50% 61% 69% 21% 64% 46% st Average 1 rd rd trimester -3 trimester % change rd rd trimester -3 trimester % change Summary Statistics 98 127 154 31% 22% 59% *Studies with multiple total cholesterol measurements per trimester. An average of these multiple measurements is reported. These studies were given a greater weight in the weighted average than those with one total cholesterol measurement per trimester. 28 TABLE 8: Summary statistics for low density lipoprotein cholesterol by cross-sectional study First Author (Year) Belo (2002) Lippi (2007) Ordovas (1984) Piechota (1992) Summary Statistics st nd Average 3 trimester mg/dL rd Average 1 trimester mg/dL Average 2 trimester mg/dL 84 90 97 110 st Weighted 1 trimester average mg/dL 123 130 126 146 nd Weighted 2 trimester average mg/dL 146 136 159 167 rd Weighted 3 trimester average mg/dL 102 138 162 st 1 trimesternd 2 trimester % change 46% 44% 30% 33% st Average 1 nd nd 2 trimester rd st 1 trimesterrd -3 trimester 3 trimester % change % change 19% 74% 5% 51% 26% 64% 14% 52% nd st Average 2 Average 1 rd rd trimester -3 trimester % change trimester -3 trimester % change 38% 29 trimester-2 trimester % change 16% 60% TABLE 9: Summary statistics for high density lipoprotein cholesterol by longitudinal study st nd First Author (Year) Average 1 trimester mg/dL Average 2 trimester mg/dL Alvarez (1996) Belo (2004) Brizzi (1999) Fahraeus* (1985) Jimenez* (1988) Khoury* (2005) Misra* (2011) Munoz (1999) Ogura (2002) Tranquilli (2004) Winkler (2000) 68 54 68 54 63 68 67 62 55 83 st Weighted 1 trimester average mg/dL 82 63 73 72 69 72 78 70 72 72 76 nd Weighted 2 trimester average mg/dL Average 3 trimester mg/dL rd 71 56 68 74 66 67 76 71 85 53 75 rd Weighted 3 trimester average mg/dL st 1 trimesternd 2 trimester % change 21% 17% 7% 33% 10% 16% 5% 16% 31% -8% st Average 1 nd trimester-2 trimester % change nd 2 trimester - rd 3 trimester % change -13% -11% -7% 3% -5% -7% -3% 1% 17% -26% -1% nd Average 2 rd trimester -3 trimester % change st 1 trimesterrd 3 trimester % change 4% 4% 0% 37% 4% 13% 6% 36% -4% -10% st Average 1 rd trimester -3 trimester % change Summary Statistics 65 73 69 15% -5% 10% *Studies with multiple total cholesterol measurements per trimester. An average of these multiple measurements is reported. These studies were given a greater weight in the weighted average than those with one total cholesterol measurement per trimester. 30 TABLE 10: Summary statistics for high density lipoprotein cholesterol by cross-sectional study st nd rd st First Author (Year) Average 1 trimester mg/dL Average 2 trimester mg/dL Belo (2002) Berge (1996) Lippi (2007) Ordovas (1984) Piechota (1992) 55 61 67 62 60 st Weighted 1 trimester average mg/dL 70 76 83 69 71 nd Weighted 2 trimester average mg/dL 62 63 81 65 63 rd Weighted 3 trimester average mg/dL 60 72 64 Summary Statistics Average 3 trimester mg/dL 1 trimesternd 2 trimester % change 27% 25% 24% 11% 18% st Average 1 nd nd 2 st trimester 1 trimester- -3 trimester % change -11% -17% -2% -5% -11% nd Average 2 3 trimester % change 13% 3% 21% 5% 5% st Average 1 rd rd rd rd trimester -3 trimester % change trimester -3 trimester % change 21% 31 trimester-2 trimester % change -9% 10% FIGURE 1: * Scatter plot of average TC levels through pregnancy and postpartum for both longitudinal and cross-sectional studies. For interpretation of the references to color in this and all other figures, the reader is referred to the electronic version of this thesis. 32 FIGURE 2: * Scatter plot of average TC levels through pregnancy reported by longitudinal studies stratified on the type of sample used for cholesterol assays (fasting, non-fasting; plasma, serum). 33 FIGURE 3: * Bar plot of the average TC levels through pregnancy stratified on the type of th th observational study used (cross-sectional, longitudinal). Error bars mark the 25 and 75 percentiles. 34 FIGURE 4: * Scatter plot of average LDL-C levels through pregnancy by study stratified on the study type. 35 FIGURE 5: * Scatter plot of average HDL-C levels through pregnancy by study stratified on the study type. 36 REFERENCES 37 REFERENCES 1. Cohen, J.D., et al., 30-Year Trends in Serum Lipids Among United States Adults: Results from the National Health and Nutrition Examination Surveys II, III, and 1999-2006. The American Journal of Cardiology, 2010. 106(7): p. 969-975. 2. Third Report of the National Cholesterol Education Program (NCEP) Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults (Adult Treatment Panel III) Final Report. Circulation, 2002. 106(25): p. 3143- 3421. 3. Woollett, L.A., Where does fetal and embryonic cholesterol originate and what does it do? Annual Review of Nutrition, 2008. 28: p. 97-114. 4. Cooper, G.M. and R.E. Hausman, The Cell: A molecular Approach 4th ed. 2007: Sinauer Associates Inc. 820. 5. Woollett, L.A., Maternal cholesterol in fetal development: transport of cholesterol from the maternal to the fetal circulation. Am J Clin Nutr, 2005. 82(6): p. 1155-1161. 6. Palinski, W., Maternal-Fetal Cholesterol Transport in the Placenta: Good, Bad, and Target for Modulation. Circ Res, 2009. 104(5): p. 569-571. 7. Herrera, E., Lipid metabolism in pregnancy and its consequences in the fetus and newborn. Endocrine, 2002. 19(1): p. 43-55. 8. Schaefer, E.J., Introduction to High-Density Lipoprotein, Dyslipidemia, and Coronary Heart Disease, in High Density Lipoproteins, Dyslipidemia, and Coronary Heart Disease. 2010, Springer New York. p. 1-14. 9. Schaefer, E.J., Lipoproteins, nutrition, and heart disease. Am J Clin Nutr, 2002. 75(2): p. 191-212. 10. NHLBI. Diseases and Conditions Index. High Blood Cholesterol 2011 [cited April 11, 2011]; Available from: http://www.nhlbi.nih.gov/health/dci/Diseases/Hbc/HBC_WhatIs.html. 11. NHLBI. National Cholesterol Education Program. 2011 [cited April 6, 2011]; Available from: http://www.nhlbi.nih.gov/about/ncep/. 12. Weiss, R., M. Harder, and J. Rowe, The relationship between nonfasting and fasting lipid measurements in patients with or without type 2 diabetes mellitus receiving treatment with 3-hydroxy-3-methylglutaryl-coenzyme A reductase inhibitors. Clinical Therapeutics, 2003. 25(5): p. 1490-1497. 38 13. Friedewald, W., R. Levy, and D. Fredrickson, Estimation of the Concentration of LowDensity Lipoprotein Cholesterol in Plasma, without the use of the preparative ultracentrifuge. Chemical Chemistry, 1972. 18(6): p. 499-502. 14. McNamara, J.R., et al., Calculated values for low-density lipoprotein cholesterol in the assessment of lipid abnormalities and coronary disease risk. Clin Chem, 1990. 36(1): p. 36-42. 15. Schaefer, E.J., et al., Comparison of fasting and postprandial plasma lipoproteins in subjects with and without coronary heart disease. The American Journal of Cardiology, 2001. 88(10): p. 1129-1133. 16. LaRosa, J.C., Lipids and cardiovascular disease: Do the findings and therapy apply equally to men and women? Women's Health Issues, 1992. 2(2): p. 102-113. 17. Pilote, L., et al., A comprehensive view of sex-specific issues related to cardiovascular disease. CMAJ, 2007. 176(6): p. S1-44. 18. Programme, U.N.D. Human Development Index. 2010 [cited; Available from: http://hdr.undp.org/en/media/HDR_2010_EN_Table1.pdf. 19. Bland, J.M. and S.M. Kerry, Weighted comparison of means. BMJ, 1998. 316(7125): p. 129. 20. Gratacos, E., et al., Variation in lipid levels during pregnancy in women with different types of hypertension. Acta Obstetricia et Gynecologica Scandinavica, 1996. 75(10): p. 896-901. 21. Berge, L.N., E. Arnesen, and A. Forsdahl, Pregnancy related changes in some cardiovascular risk factors. Acta Obstetricia et Gynecologica Scandinavica, 1996. 75(5): p. 439-442. 22. Alvarez, J.J., et al., Longitudinal study on lipoprotein profile, high density lipoprotein subclass, and postheparin lipases during gestation in women. Journal of Lipid Research, 1996. 37(2): p. 299-308. 23. Belo, L., et al., LDL size, total antioxidant status and oxidised LDL in normal human pregnancy: a longitudinal study. Atherosclerosis, 2004. 177(2): p. 391-399. 24. Brizzi, P., et al., Lipoprotein metabolism during normal pregnancy. American Journal of Obstetrics and Gynecology, 1999. 181(2): p. 430-434. 25. Darmady, J.m. and A.d. Postle, Lipid metabolism in pregnancy. BJOG: An International Journal of Obstetrics & Gynaecology, 1982. 89(3): p. 211-215. 39 26. Jimenez, D.M., et al., Longitudinal Study of Plasma Lipids and Lipoprotein Cholesterol in Normal Pregnancy and Puerperium. Gynecologic and Obstetric Investigation, 1988. 25(3): p. 158-164. 27. Khoury, J., et al., Effect of a cholesterol-lowering diet on maternal, cord, and neonatal lipids, and pregnancy outcome: A randomized clinical trial. American Journal of Obstetrics and Gynecology, 2005. 193(4): p. 1292-1301. 28. Sanchez-Vera, I., et al., Increased Low-Density Lipoprotein Susceptibility to Oxidation in Pregnancies and Fetal Growth Restriction. Obstetrics & Gynecology, 2005. 106(2): p. 345-351 10.1097/01.AOG.0000171112.95083.86. 29. Tranquilli, A.L., et al., Plasma lipids and physicochemical properties of the erythrocyte plasma membrane throughout pregnancy. Acta Obstetricia et Gynecologica Scandinavica, 2004. 83(5): p. 443-448. 30. Winkler, K., et al., Low Density Lipoprotein (LDL) Subfractions during Pregnancy: Accumulation of Buoyant LDL with Advancing Gestation. J Clin Endocrinol Metab, 2000. 85(12): p. 4543-4550. 31. Fahraeus, L., U.L.F. Larsson-Cohn, and L. Wallentin, Plasma Lipoproteins Including High Density Lipoprotein Subfractions During Normal Pregnancy. Obstetrics & Gynecology, 1985. 66(4): p. 468-472. 32. Munoz, A., et al., Relationship of blood rheology to lipoprotein profile during normal pregnancies and those with intrauterine growth retardation. Journal of Clinical Pathology, 1995. 48(6): p. 571-574. 33. Rosing, U., et al., Serum Levels of Apolipoprotein A-I, A-II and HDL-Cholesterol in Second Half of Normal Pregnancy and in Pregnancy Complicated by Pre-Eclampsia. Horm Metab Res, 1989. 21(07): p. 376,382. 34. Martin, U., et al., Is normal pregnancy atherogenic? Clinical Science, 1999. 96(4): p. 421-425. 35. Belo, L.Ì.s., et al., Changes in LDL size and HDL concentration in normal and preeclamptic pregnancies. Atherosclerosis, 2002. 162(2): p. 425-432. 36. Lippi, G., et al., Lipid and lipoprotein profile in physiological pregnancy. Clinical Laboratory, 2007. 53(3-4): p. 173-177. 37. Piechota, W. and A. Staszewski, Reference ranges of lipids and apolipoproteins in pregnancy. 1992. 45(1): p. 27-35. 38. Ordovas, J.M., M. Pocovi, and F. Grande, Plasma Lipids and Cholesterol Esterification Rate During Pregnancy. Obstetrics & Gynecology, 1984. 63(1): p. 20-25. 40 39. Vahratian, A., et al., Prepregnancy Body Mass Index and Gestational Age-Dependent Changes in Lipid Levels During Pregnancy. Obstetrics & Gynecology, 2010. 116(1): p. 107-113 10.1097/AOG.0b013e3181e45d23. 40. Ogura, K., et al., Low-Density Lipoprotein Particle Diameter in Normal Pregnancy and Preeclampsia. Journal of Atherosclerosis and Thrombosis, 2002. 9(1): p. 42-47. 41. Misra, V.K. and D.P. Misra, Average cholesterol measurements per follow-up for GROW study participants stratified on maternal BMI. Unpublished data. 2011: Detroit, MI. 42. McMurry, M.P., W.E. Connor, and C.P. Goplerud, The effects of dietary cholesterol upon the hypercholesterolemia of pregnancy. Metabolism, 1981. 30(9): p. 869-879. 43. Patrick, T.E., C.A. Hubel, and J.M. Roberts, Evidence of Increased Oxidative Stress, Unexplained by Lipid Changes, Is Present in Nulliparous Black Women from Early Gestation. Hypertension in Pregnancy, 2004. 23(1): p. 91-100. 44. Donahue, R.P., et al., Distribution of lipoproteins and apolipoproteins in young adults The CARDIA Study. Arterioscler Thromb Vasc Biol, 1989. 9(5): p. 656-664. 45. Herrera, E., et al., Maternal lipid metabolism and placental lipid transfer. Hormone Research, 2006. 65: p. 59-64. 46. Roux, C., et al., Role of cholesterol in embryonic development. The American Journal of Clinical Nutrition, 2000. 71(5): p. 1270S-1279S. 47. Porter, F.D. and G.E. Herman, Malformation syndromes caused by disorders of cholesterol synthesis. Journal of Lipid Research. 52(1): p. 6-34. 48. Napoli, C., et al., Fatty streak formation occurs in human fetal aortas and is greatly enhanced by maternal hypercholesterolemia. Intimal accumulation of low density lipoprotein and its oxidation precede monocyte recruitment into early atherosclerotic lesions. The Journal of Clinical Investigation, 1997. 100(11): p. 2680-2690. 49. Edison, R.J., et al., Adverse birth outcome among mothers with low serum cholesterol. Pediatrics, 2007. 120(4): p. 723-733. 50. Mudd, L., et al., Maternal lipids at midpregnancy and risk of preterm delivery. 2010, MSU: East Lansing. 51. Elmes, M.J., et al., The effects of a high-fat, high-cholesterol diet on markers of uterine contractility during parturition in the rat. Reproduction. 141(2): p. 283-290. 52. Winkel, C.A., et al., Regulation of Cholesterol and Progesterone Synthesis in Human Placental Cells in Culture by Serum Lipoproteins. Endocrinology, 1980. 106(4): p. 10541060. 41 53. Knopp, R.H., et al., Population-based Lipoprotein Lipid Reference Values for PregnantWomen Compared to Nonpregnant Women Classified by Sex-hormone Usage. American Journal of Obstetrics and Gynecology, 1982. 143(6): p. 626-637. 54. Cedard, L., The Endocrine Function of the Placenta: Interactions Between Trophic Peptides and Hormones, in Placental Function and Fetal Nutrition, F.C. Battaglia, Editor. 1997, Vevey/Lippincott-Raven Publishers: Philadelphia. p. 59-74. 55. Mumford, S.L., et al., A Longitudinal Study of Serum Lipoproteins in Relation to Endogenous Reproductive Hormones during the Menstrual Cycle: Findings from the BioCycle Study. J Clin Endocrinol Metab, 2010. 95(9): p. E80-85. 56. Desoye, G., et al., Correlation of Hormones with Lipid and Lipoprotein Levels During Normal Pregnancy and Postpartum. J Clin Endocrinol Metab, 1987. 64(4): p. 704-712. 57. Amundsen, A., et al., Marked changes in plasma lipids and lipoproteins during pregnancy in women with familial hypercholesterolemia. Atherosclerosis, 2006. 189(2): p. 451-457. 58. Palinski, W. and C. Napoli, The fetal origins of atherosclerosis: maternal hypercholesterolemia, and cholesterol-lowering or antioxidant treatment during pregnancy influence in utero programming and postnatal susceptibility to atherogenesis. The FASEB Journal, 2002. 16(11): p. 1348-1360. 59. Napoli, C., et al., Influence of maternal hypercholesterolaemia during pregnancy on progression of early atherosclerotic lesions in childhood: Fate of Early Lesions in Children (FELIC) study. The Lancet, 1999. 354(9186): p. 1234-1241. 60. Beral, V., Long term effects of childbearing on health. Journal of Epidemiology and Community Health, 1985. 39(4): p. 343-346. 61. Lawlor, D.A., et al., Is the Association Between Parity and Coronary Heart Disease Due to Biological Effects of Pregnancy or Adverse Lifestyle Risk Factors Associated With Child-Rearing?: Findings From the British Women's Heart and Health Study and the British Regional Heart Study. Circulation, 2003. 107(9): p. 1260-1264. 62. Hinkula, M., et al., Cause-specific Mortality of Grand Multiparous Women in Finland. American Journal of Epidemiology, 2006. 163(4): p. 367-373. 63. Ness, R.B., et al., Number of Pregnancies and the Subsequent Risk of Cardiovascular Disease. New England Journal of Medicine, 1993. 328(21): p. 1528-1533. 64. Humphries, K.H., et al., Parity and Carotid Artery Atherosclerosis in Elderly Women: The Rotterdam Study. Stroke, 2001. 32(10): p. 2259-2264. 42 65. Mankuta, D., et al., Lipid profile in consecutive pregnancies. Lipids in Health and Disease. 9: p. 4. 66. Wynn, V., et al., Fasting Serum Triglyceride, Cholesterol, and Lipoprotein Levels during Oral-contraceptive Therapy. The Lancet, 1969. 294(7624): p. 756-760. 67. Wahl, P., et al., Effect of Estrogen/Progestin Potency on Lipid/Lipoprotein Cholesterol. New England Journal of Medicine, 1983. 308(15): p. 862-867. 68. Koukkou, E., G.F. Watts, and C. Lowy, Serum lipid, lipoprotein and apolipoprotein changes in gestational diabetes mellitus: a cross-sectional and prospective study. Journal of Clinical Pathology, 1996. 49(8): p. 634-637. 69. Enquobahrie, D.A., et al., Early pregnancy lipid concentrations and the risk of gestational diabetes mellitus. Diabetes research and clinical practice, 2005. 70(2): p. 134142. 70. Sanchez-Vera, I., et al., Changes in plasma lipids and increased low-density lipoprotein susceptibility to oxidation in pregnancies complicated by gestational diabetes: consequences of obesity. Metabolism: clinical and experimental, 2007. 56(11): p. 15271533. 71. Knopp, R.H., et al., Lipoprotein Metabolism in Pregnancy, Fat Transport to the Fetus, and the Effects of Diabetes. Neonatology, 1986. 50(6): p. 297-317. 72. Elzen, H.J.v.d., et al., Serum lipids in early pregnancy and risk of pre-eclampsia. British Journal of Obstertrics and Gynaecology, 1996. 103: p. 117-122. 73. Sattar, N., et al., Lipoprotein Subfraction Concentrations in Preeclampsia: Pathogenic Parallels to Atherosclerosis. Obstetrics & Gynecology, 1997. 89(3): p. 403-408. 43