Obesity and Gestational DiabetesMellitus Pathways for ProgramminginMouse,Monkey,andMandWhereDo We Go Next? The 2014 NorbertFreinkel Award LectureDiabetes Care 2015;38:1402–1411 | DOI: 10.2337/dc15-0628

Obesity and gestational diabetes mellitus continue to increase worldwide and spanthe spectrum of age, race, ethnicity, and socioeconomic status. Alarmingly, 1 in 10infants and toddlers is obese, and 1 in 5 youths is both obese and at risk formetabolicsyndrome prior to puberty. The mechanisms underlying how poor maternal healthimparts risk for future metabolic disease in the offspring are beginning to emerge indeeply phenotyped human and nonhuman primate models. Maternal diet and obe-sity impact fuels, hormones, and inflammation with powerful effects on fetal met-abolic systems. These are accompanied by persistent changes in the infantmicrobiome and epigenome and in offspring behavior. These results suggest thatgestational and lactational dietary exposures are driving health risks in the nextgeneration. Whether maternal diet can prevent changes in the womb to alterinfant life-course disease risk is still unknown. Controlled, mechanistic studies toidentify interventions are sorely needed for a healthier next generation.

In the late 1980s, Dr. David Barker used historical birth records from Hertfordshire,England, to pioneer the concept that the origins of adult disease could be stronglyassociated with fetal environmental exposures in pregnancy that resulted in lowbirth weight and predicted increased risk for cardiovascular morbidity and mortalityin the offspring (1,2). Over the past several decades, the Developmental Origins ofHealth and Disease (DOHaD) hypothesis has been validated epidemiologically andmechanistically in both human and animal models. Data from animal models ofnutritional constraint and uteroplacental insufficiency suggest that the gestationalmilieu influences the postnatal phenotype to cause susceptibility to childhood obe-sity and metabolic disorders through changes in tissue and organ development andmetabolic reprogramming (3–7). DOHaD has taught us about the role of a mismatchbetween a constrained prenatal and a plentiful postnatal environment in the path-ogenesis of obesity, i.e., the “thrifty” pathway, likely operating in populations un-dergoing rapid transition.Another developmental pathway to obesity and its comorbidities, likely more

important in Western societies, is developmental overnutrition. In fact, NorbertFreinkel, in his Banting Lecture in 1980 (8) may have been the first to conceptualizethe overnutrition hypothesis in utero as a contributor to long-term offspring met-abolic complications. He discussed this in the context of diabetes in pregnancy

Departments of Pediatrics, Biochemistry andMolecular Genetics, Endocrinology, Metabolism& Diabetes, and Basic Reproductive Sciences,University of Colorado School of Medicine; Colo-rado Program for Nutrition and Healthy Devel-opment, Children’s Hospital Colorado ResearchInstitute; University of Colorado Nutrition andObesity Research Center, Aurora, CO

Corresponding author: Jacob E. (Jed) Friedman,jed.friedman@ucdenver.edu.

© 2015 by the American Diabetes Association.Readers may use this article as long as the workis properly cited, the use is educational and notfor profit, and the work is not altered.

Jacob E. (Jed) Friedman

1402 Diabetes Care Volume 38, August 2015

NORBER

TFR

EINKEL

AWARDLECTU

RE

leading to excess fuel exposure to thefetal-placental unit, not only from glu-cose but also from excess lipids andamino acids (AAs). He termed this“fuel-mediated teratogenesis,” definedas alterations occurring subsequent toorganogenesis during the differentia-tion and proliferation of fetal cells, andwrote “such changes could cause long-range effects upon behavioral, anthro-pometric, and metabolic functions” (8).This pathway reflects the effects of hy-pernutrition during fetal life and sets thestage for further amplification of thepathophysiological effects of encoun-tering an obesogenic environment post-natally. More than 30 years later,emerging data suggest that in fact ma-ternal diet, potentially amenable to ma-nipulation, may be a key determinant ofthe fetal epigenotype and resulting phe-notype (9–17).Animal models, including nonhuman

primates (NHP), sheep, rodents, andeven flies and zebrafish, have shownthat fetal growth and development arevulnerable to changes in nutrition in allthree phases of development. These in-clude 1) early gestation during implan-tation, placentation, and subsequentembryogenesis when placental nutrienttransport may be set; 2) midgestationwhen number, growth, and function ofcritical organs like pancreas, brain, kid-ney, and skeletal muscle develop; and3) during the late third trimester whenfetal growth (adiposity) acceleratesand regulatory set points in the brain,neuronal-metabolic feedback loops,and mitochondrial function may be im-pacted. While animal models may havevastly different patterns of gestationand placental structure, they havebeen critical in the discovery of poten-tial molecules and mechanisms leadingto early-onset obesity. Despite thesediscoveries, there is lack of knowledgeabout the impact of environmental ex-posures on biochemical and molecularprocesses that govern metabolic risk,particularly in utero on infants born tomothers who are obese or have gesta-tional diabetes mellitus (GDM).The focus of our research group is on

understanding the role of maternal nu-trition and the intrauterine environ-ment on molecular, endocrine, andepigenetic origins of childhood obesity.This involves developing novel animalmodels of GDM and obesity (transgenic

mice, NHP) together with longitudinal in-vasive human clinical investigation in vivoand in vitro utilizing myocytes, adiposetissue, and, more recently, umbilical-derived mesenchymal stem cells (MSCs)obtained from infants born to obesewomen with and without GDM. More re-cently, we have begun investigating therole of the microbiome (MB) in mothersand their infants with a goal of under-standing how maternal nutrition in preg-nancy and breast milk composition caninfluence infant microbial function andthe trajectory of weight gain, adiposity,and development of nonalcoholic fattyliver disease (NAFLD), the leading causeof liver disorders in children and adults.The work of myself and my colleaguesover the past 20 years has been due inno small part to my association and col-laboration with Dr. Patrick Catalano(Case Western Reserve University) andDr. Linda Barbour (University of Colo-rado Denver), who continue to teachme about the important clinical prob-lems encountered in the diagnosis,treatment, and underlying pathophysi-ology of normal, obese, and GDM preg-nancies. They have also taught meabout the power of collaboration andthe value of true friendship.

MATERNAL INSULIN RESISTANCEIN OBESITY AND GDM: CELLULARMECHANISMS THAT IMPARTINSULIN RESISTANCE,CONSEQUENCES OF EXCESS FETALNUTRIENT EXPOSURE, ANDPOSTPARTUM RISK FORPROGRESSION TO TYPE 2DIABETES

Maternal insulin resistance (IR) is a normalpart of human pregnancy and is criticallyimportant to maintain the maternal fuelsupply to support the growing fetus, par-ticularly during the third trimester. How-ever, women with obesity or a history ofGDM enter pregnancy with preexisting IRthat worsens with advancing gestation(18). In type 2 diabetes and GDM, thema-ternal metabolic environment is charac-terized by IR and inflammation, and bothconditions shunt excess fuels to facilitatefat accretion and fetal growth (19). Emerg-ing data suggest that excess maternal IRmay be a more important predictor ofneonatal adiposity than prepregnancyBMI (20,21). In a study of 301 infantsof women with GDM, mild glucose in-tolerance, or normal glucose tolerance,

maternal IR by the Matsuda index pre-dicted offspring weight gain and adiposityfrom 0–12months, but prepregnancy BMIdid not (20). Maternal IR has the capacityto shunt not only glucose but all nutrientsto the fetus, including glucose, triglycer-ides (TGs), free fatty acids (FFAs), andAAs, all of which can be used for fetalfat accretion and excess fetal growth.Furthermore, the placental transcriptomeis a target of the altered environment ofdiabetic and obese pregnancies. Genes forlipids and AA transport are upregulated inthe placenta of women with GDM morethan those for glucose, as are genes forinflammatory pathways (22,23).

Mechanistically, we demonstratedthat insulin-stimulated glucose trans-port in human skeletal muscle fibersfrom obese women is suppressed inlate pregnancy and more so in skeletalmuscle of women who develop GDM(24). In transgenic mice, and later in hu-man pregnancy, we showed that humanplacental growth hormone is a majordriver of the normal IR of pregnancyby suppressing the IRS1-associated PI3-kinase insulin signaling cascade in skeletalmuscle (25,26). Further, we showed inGDM subjects that reduced IRS1 in skel-etal muscle and adipose tissue, alongwith decreased insulin receptor activa-tion, may be responsive to overnutritionas well as inflammatory changes duringlate pregnancy (27). In addition, usingproteomic analysis in skeletal muscle ofobese and GDM subjects, we found re-duced mitochondrial function and in-creased oxidative stress in both obeseand GDM patients, controlled in part bythe mitochondria deacetylase SIRT3(28). In GDM subjects, there are addi-tional unique changes in calcium bindingproteins associated with reduced mito-chondrial function, suggesting thesemight underlie the reduced oxidativecapacity and ultimately lower exercisetolerance, particularly evident in womenwith GDM (29). Last, when we followeda series of obese women with GDM lon-gitudinally postpartum we found thatspecific defects in IRS1 and p70S6K inhuman skeletal muscle may persist upto a year later and may underlie thechronic IR and future risk for type 2 di-abetes (19,30), beyond simple obesity.While there are complex mechanisms un-derlying the IR in skeletal muscle and ad-ipose tissue, given that fetal growthand adiposity increases with even

care.diabetesjournals.org Friedman 1403

modest changes in glucose (31), FFAs,and TGs (32), the IR pathways uncoveredhere may be extremely important. Fur-thermore, they are highly relevant inexplaining why obese women, whom wehave shown to have elevated 24-h gly-cemic profiles as well as FFAs and TGscompared with normal-weight women(32), are at an increased risk for deliver-ing infants with increased adiposity.

IMPACT OF A WESTERN-STYLEDIET AND MATERNAL OBESITY ONMETABOLIC SYSTEMS IN THENEXT GENERATION: LESSONSLEARNED FROM NHP

Maternal obesity and GDM are impor-tant risk factors for obesity in the nextgeneration. Whether this is due to spe-cific changes in maternal diet, the ma-ternal phenotype (IR), or a combinationof the two has remained poorly under-stood. In order to study the impact ofspecific changes in maternal diet andobesity on fetal pathophysiology and fu-ture health risks, in 2007 Dr. Kevin Groveat the Oregon National Primate Re-search Center and I began a collaborationto develop an NHP model for studyingmaternal obesity and its impact on thedevelopment of fetal metabolic systems.Our group has spent the past decade de-veloping and studying a sophisticatedNHP JapaneseMacaque model of chronicconsumption of a maternal high-fat/calorie-dense Western-style diet (WSD)starting early in the reproductive yearsthat has critically important develop-mental and physiological similarities tohumans. The importance of the NHPmodel is that the placenta, brain, liver,skeletal muscle, and pancreas structuresare similar to humans, and it is the onlynatural model that develops the full spec-trum of metabolic diseases as in humans.Consequentially, the developmentalchanges in vital organs are highly sim-ilar to humans, including complex psy-chosocial behaviors that can be studiedlongitudinally in the offspring. Thesequalities make the NHPmodel uniquelypowerful and critically important.We have shown that consumption

of a maternal WSD causes placental dys-function, tissue-specific changes in theoffspring mitochondria, widespreadinflammation, hepatic steatosis, and broaddevelopmental changes in the liver, skele-talmuscle, brain, andpancreas, as outlinedbelow. These alterations are accompanied

by significant andpersistent changes in theepigenome, the MB, and offspring behav-ior. Importantly, many of these abnormal-ities persist even when NHP offspring areweaned to a healthy diet after lactation,suggesting that gestational and lactationaldietary exposures are significant and pos-sibly permanent contributors in the initia-tion and development of pathways thatdrive health risks in the next generation.

One of the most striking early findingswe discovered in the NHP model is thedevelopment of fetal NAFLD. NAFLD is ageneral term used to describe a broadspectrum of liver abnormalities, rangingfrom simple, uncomplicated hepaticsteatosis to nonalcoholic steatohepa-titis (NASH), with different degrees ofinflammation and fibrosis. NAFLD is themost common liver disease in childrenand adults (33,34) and the leading causeof liver transplantation. Prevalence esti-mates of NAFLD in children range from3–10% in Western societies, approaching55% in children who are obese (35).When we performed cesarean sectionsin the early third trimester (G130) in

mothers fed a WSD, we discovered thefetuses had a dramatic increase in liverlipids (Fig. 1) (13). Importantly, we foundthat all fetuses had elevated liver lipidswhether their mothers developed obe-sity or not on the WSD. Fetuses at thisstage of development have very little ad-ipose tissue to store lipids, suggestingthat exposure to a WSD resulted in ex-cess lipid availability to the fetus thataccumulates in the liver during gesta-tion. The livers also stained for a markerof oxidative stress (4-hydroxynonenal),suggesting lipid overload may have ef-fects on protein functions, or so-calledlipotoxicity.

During the next breeding season, weperformed a diet reversal, wherebyobese mothers were switched from aWSD to a healthy control diet. Althoughthemothers remained obese, theyman-ifested less IR. In theseG130 fetuses, liverTGs were significantly improved buthad not returned to control levels, sug-gesting that maternal obesity, evenon a healthy diet, contributes to fetalhepatic steatosis. To examine if these

Figure 1—Liver steatosis in the fetal liver during the early third trimester of NPH on a high-fat diet.Female Japanese Macaques were maintained on high-fat or chow diet for a minimum of 2 yearsprior to mating. In a subsequent pregnancy, obese females were switched to a chow diet priorto mating and cesarean sections (C-section) were performed. Livers were stained for neutral lipid(Oil-Red-O) and 4-hydroxynonenal (4-HNE), a marker of oxidative stress (data are fromMcCurdyet al. [13]).

1404 Norbert Freinkel Award Lecture Diabetes Care Volume 38, August 2015

changes persisted postnatally, a sepa-rate cohort of obese mothers on a WSDwere allowed to give birth and the off-spring were weaned to a healthy diet at6–7 months of age. At 12–14 months ofage, the livers from the juvenile ani-mals showed increased TGs andincreased liver macrophage cell num-bers and the liver macrophages werehyperresponsive to fatty acids in vitro,producing high levels of cytokines (36).Overall, a WSD coupled with maternalobesity was associated with long-termconsequences, increasing the risk forprogression from uncomplicated he-patic steatosis to NASH. This includesinflammation via Toll-like receptors onthe resident macrophages (Kupffercells), immune cell activation, andpriming for recruitment of additionalimmune cells from the bone marrow(Fig. 2). Importantly, these changes oc-curred prior to the development of ju-venile obesity in offspring born toobese mothers with high IR on a WSD.The placentas from the obese motherson aWSD showed evidence of oxidative

stress, inflammation, reduced placentalblood flow, and hypoxia (9). These find-ings suggest that chronic WSD and ma-ternal obesity may be the “first hit” inthe progression of simple steatosis toinflammation and NASH in later life.

To determine whether this early liversteatosis is present in human neonateswho have more subcutaneous fat tostore lipid than NHP, we used magneticresonance spectroscopy to show thatmaternal obesity and GDM predictneonatal hepatic fat storage (37). Wemeasured intrahepatocellular lipid in2-week-old newborns born to bothnormal-weight and obese mothers withGDM and showed a 68% increase in liverfat in the infants of obese GDMmothers.In another cohort, Modi et al. (38) re-ported an 8.6% increase in intrahepato-cellular lipid content for each 1-pointincrease in prepregnancy BMI. The posi-tive correlation between offspring in-trahepatocellular lipid and maternalprepregnancy BMI held for the entirepopulation of obese and normal-weightmothers. Other variables, such as

gestational weight gain and early post-natal weight change, did not influenceintrahepatic fat storage in this cohort of25 mother-infant pairs. Importantly, in-creased fetal hepatic fat storage appearedto be independent of overall adiposity inthe newborn, suggesting that the driversof hepatic fat storage and subcutaneousadipose fat storagemay be different duringfetal life. In another infant cohort, liver fatdoubled between birth and 2 months, re-gardless of breast-feeding (39), raising thequestion of whether excess liver fatstorage at birth may have long-term con-sequences, particularly in obese teenagers(40,41). Notably, NAFLD increases therisk of cardiovascular events eightfoldand type 2 diabetes threefold and is astrong risk factor for hepatocellular carci-noma (42).

A second striking finding in the NHPstudies is the long-term behavioralchanges noted in the offspring exposedto maternal WSD during pregnancy andweaning. When these animals wereswitched to a healthy diet at weaning(6–7 months of age) and studied at12–14 months of age as juveniles, themale animals displayed an increase in re-petitive behaviors (pacing stereotypy),while the females displayed increasedanxiety-like behavior (10). Offspring ofboth sexes displayed impairments insocial behavior. A potential mechanismfor these behavioral changes is the sup-pression of serotonin synthesis by theraphe nuclei noted in these juveniles (10),suggesting that the systems that governanxiety and depression may be hardwiredby the maternal diet.

There is strong evidence that obese chil-dren tend to consume more dietary fatthan nonobese children as a function oftotal calories (43), thus contributing tohigher BMI. In addition, fats tend to de-regulate control mechanisms for bodyweight regulation (appetite and activity)associated with changes in the hypothala-mus, dopaminergic pathways, and otherareas of the brain for reasons that arenot completely understood (44). Whethermaternal obesity and/or diet can modifythe neurocircuitry in utero and whether itundergoes furthermodification during lac-tation and early feeding resulting in foodpreferences that occur prior to the devel-opmentof obesity remainunknown. In ourstudies of juvenile NHP offspring exposedto a maternal WSD during pregnancy andlactation and thenweaned to healthy diets

Figure 2—Consequences of maternal overfeeding on fetal liver and the evolution of NAFLD.Exposure to excess lipids triggers placental inflammation, oxidative stress, and liver steatosisprior to birth. After weaning, a combination of recruited hepatic macrophages and alterna-tively activated resident immune cells, together with IR and obesity, is a significant risk factorfor inflammation and progression of NAFLD. Prevalence of NAFLD is shown. WBC, white bloodcell.

care.diabetesjournals.org Friedman 1405

postnatally, there was already a strongpreference for selection of high-energy(high-fat) foods over all other sources ofcalories. Of note, these changes took placeprior to the emergence of obesity in theanimals. Although not all animals wereequally affected, when combined withthe social/behavioral abnormalities notedabove, the results suggest that earlychanges in brain development and behav-ior may be one of the most compellingconcerns for the DOHaD hypothesis andits impact on children in modern society.Whether these changes are governed byexposure in utero to higher fuels, cyto-kines, or oxidative stress noted in the fe-tuses of WSD-fed mothers; postnatally byhigher breast milk n-6:n-3 fatty acids (12);or by other bioactive and appetite regula-tory components such as leptin or insulinremain to be determined. Interventionalstudies in NHP obese mothers and thestudy of longer-term brain epigeneticmodifications in their 3-year-old offspringare currently under way.

MATERNAL OBESITY AND THEINFANT MB: THE UNDISCOVEREDCOMMUNITY WITHIN

The gut MB plays a significant role in bothmaternal and infant gut health and childdevelopment. The intestinal microbiota isreferred to as our “second genome” and isacknowledged as 10 times the size of ourown genetic repertoire, involved in thesusceptibility of many disorders as diverseas obesity, type 1 diabetes, inflammatorybowel disease, allergies, autism, cancer,and asthma. The term “gut microbiota”represents a complex bacterial commu-nity within the small and large intestine,capable of affecting health by contributingto energy retention and appetite, pre-venting colonization of the host by patho-gens, and influencing the developmentand maintenance of the immune system.In human pregnancy, there are temporalshifts in both the diversity within a micro-bial community, known as a-diversity,which can be measured by phylogeneticdiversity and species richness, and in thediversity shared among different commu-nities, known asb-diversity, which can bemeasured by changes in microbial abun-dance (45). The microbiota in late preg-nancy has reduced a-diversity (richness)but higher b-diversity (abundance ofcertain species) comparedwith nonpreg-nant women or women in early preg-nancy (46).

Remarkably, when germ-free (gnoto-biotic) mice were transplanted with fe-cal microbiota from healthy women intheir first or third trimester of preg-nancy, those receiving the third trimes-ter microbiota had increased intestinalcytokines, gained more weight, andwere more glucose-intolerant thanthose receiving the first trimestermicro-biota (46). This striking result impliesthat the third trimester microbiotahave the ability to induce an alternativemetabolic state in mothers, associatedwith greater energy extraction and in-flammation. Such a transformationwould serve to potentiate maternal IRfor fuel transfer to the fetus and ulti-mately to transfer the capacity forincreased energy retention to the new-born, ensuring the survival of the nextgeneration. For example, in the third tri-mester, there is greater represen-tation of lactic acid bacteria, which arehighly prevalent in the infant gut,whereas butyrate-producing bacteria(Faecalibacterium, Blautia, and Rumino-coccus), which dominate the gut inadulthood, are enriched in early preg-nancy (46). The acquisition of increasedmaternal lactic acid bacteria by the thirdtrimester may be an adaptation to trans-fer these organisms during the perinatalperiod to the infant to take maximumadvantage of the main energy sourcefor the child, lactose in mother’s milk.Although the newborn acquires its mi-crobiota from the mother during deliv-ery, there is some limited evidence formicrobial presence in the placenta, am-niotic fluid, and meconium in full-termpregnancies without overt infection(47–51). However, if and to what extentearly tolerance of the fetus by the ante-partum acquisition of any bacterial spe-cies toward colonization of the mouthand gastrointestinal tract may impactthe health and development of the new-born remain to be seen.

In newborn infants, gastrointestinalmicrobes introduced through dietaryexposures are noted for their ability toserve as direct inducers/regulators ofthe infant immune system duringbreast-feeding through alterations ofthe infant gut microbiota (52,53). Dietis a powerful driver of the MB; how-ever, the specific molecular factorsand mechanisms by which the mother’sdiet influence development of the infantmicrobiota in the offspring are unknown.

Further, the role of the MB in obesityand GDM or how these states alter theneonatal MB remains largely unknown.Differences in the gut MB in infantsexperiencing different life events,such as breast-feeding versus formulafeeding, cesarean versus vaginal birth,and environmental exposure to antibi-otics, have been well documented. Ep-idemiological studies have shown thatantibiotic treatment during the first6 months of life (54,55) or disruptedcolonization from cesarean delivery(56,57) may increase the risk of beingoverweight later in life. These two in-terventions have no direct contributionto host caloric intake or metabolism(58) but have large effects on the MB(59,60). The mechanisms by which theMB may affect newborn weight gain oradipose tissue development remainsunknown but could be due to immune sig-naling, toxin release, nutrient utilization,or regulation of appetite. Our ground-breaking studies in NHP have shownthat a WSD in obese mothers leads todecreased diversity of offspring intestinalMB at 1 year and increased liver steato-sis and inflammation (36,61). Impor-tantly, these deleterious outcomesoccur prior to the onset of obesity andare not reversed by switching to ahealthy diet after weaning. This suggeststhat the maternal influence to modifythe microbial ecosystem in infants maybe driven by the pioneering bacteria ac-quired at birth and during lactation. Inhumans, we recently discovered that aproinflammatory profile of increasedn-6 relative to n-3 fatty acid in humanmilk, along with increased human milkinsulin and leptin at 2 weeks of breast-feeding, predicts changes in the infantmicrobiota and the infant bacterial meta-genome (B.E. Young and M.C. Rudolph,unpublished data). Importantly, the nor-malization of the n-6:n-3 ratio in trans-genic mice fed a high-fat diet duringpregnancy has been shown to preventexcess weight gain and fatty liver in thenext generation (62).

Despite these suggestive data, thereare no published studies directly linkingmaternal diet to altered microbiota-derived metabolites and durable changesin the developing infant immune systemor adipose tissue development. The com-position and metabolism of the adult gutMB are known to be rapidly influenced bydiet; however, these changes are often

1406 Norbert Freinkel Award Lecture Diabetes Care Volume 38, August 2015

transient (63–65), and there is tremen-dous interindividual variation (so-called“responders and nonresponders” [66]),suggesting that host genetics can shapethe composition of the gut MB (67).Conversely, the gut MB can also modifydietary exposures in ways that are bene-ficial or detrimental to the human host.For example, the SCFAs acetate, butyrate,and propionate, which are formed by mi-crobial metabolism of fiber, resistantstarches, and nonstarch polysaccharidesin the distal colon, may be metabolicallyconsequential both at the site of produc-tion and at distal tissues. The major prod-uct of microbial fermentation, acetate,is a substrate for hepatic cholesterol andTG synthesis and increases hepatic ex-pression of genes involved in fatty acidmetabolism and lipogenesis (68). Con-versely, exposure of adipose tissue topro-pionate suppresses the expression ofproinflammatory cytokines, upregulatesGLUT4 expression, and stimulates leptin(69–71). In a study of pregnant womenwith obesity, serum acetate levels wereassociatedwithmaternal weight gain andmaternal adiponectin levels (72). Propio-nate, on the other hand, was inverselycorrelated with maternal leptin (72). Bu-tyrate largely serves to support the en-ergy needs of the colonic epithelial cellsto proliferate and differentiate (73). How-ever, butyrate is also an epigenetic regu-lator (a histonedeacetylase inhibitor) thatpromotes early anti-inflammatory im-mune cell development (74) as well asadipose tissue metabolism (75) throughthe SCFA receptors GPR43 and GPR41 inadipose tissue and liver.While the core human gut microbiota

may contribute to the developmentalorigins of disease by modifying metabolicpathways in maternal and infant tissues,it may also participate as an epigeneticmodifier. Recently, Kumar et al. (76) clas-sified eight well-matched pregnantwomen into two groups based on theirdominantmicrobiota, i.e., Bacteroidetes,Firmicutes, and Proteobacteria. Deep se-quencing of DNAmethylomes revealed aclear association between bacterial pre-dominance and epigenetic profiles. Thegenes with differentially methylatedpromoters in the Firmicutes group werelinked to genes specifically involved inlipid metabolism, obesity, and the in-flammatory response. This is one of thefirst studies that highlights the associa-tion of the predominant bacterial phyla

in the gut with methylation patterns.While these studies are simple corre-lations, longitudinal studies identifyingmicrobial species or metabolites priorto health consequences may give us adeeper insight into the molecular mech-anism of such epigenetic modifications.It should be noted, however, that despitethe wealth of 16S sequencing data andmetatranscriptomic data emerging basedon shotgun sequencing methods, thefunctions of most bacterial genes fromthe mammalian microbiota remain poorlyunderstood. Whereas 16S sequencing hasproven extremely useful in terms of typingand determination of the number of phyla,metagenomic sequencing has provided anincreasingly high depth of data based onhomology with known microbial genesand may shed light on how microbiotaaffect metabolic pathways. There is anincontrovertible need to identify the func-tional roles of these bacteria and the bio-active molecules that affect human health.

FUTURE OF DOHAD RESEARCH:PLAUSIBLE INTERVENTIONS TOINTERRUPT THE VICIOUS CYCLE

While maternal genetic, epigenetic, and di-etary factors contribute to the development

of obesity and metabolic syndrome in thenext generation, the determination of howand when to intervene during pregnancyand postnatally in human infants is a com-plex problem. The lack of compelling evi-dence necessary to justify the huge effortsthat would be required to modify the ma-ternal diet of high-risk women with obesityand GDM is a significant challenge to thisfield. Our own data in NHP demonstratethat maternal diet has a significant impacton gene expression in the fetus and 1 yearlater prior to the development of obesity inthe offspring (16,17,36). Recent humanstudies suggest that maternal nutrition andthe intrauterine environment can alter DNAmethylation in umbilical cord blood (77,78),umbilical cord tissue (79,80), andbuccal cells(81). However, most clinical studies examin-ing the role of maternal nutrition and infantepigenetics are retrospective; there are nostudies investigating maternal nutritionalinventions and epigenetic changes in theinfant. Furthermore, the tissue-specificnature of epigenetic control adds to thecomplexity of such studies and remains aconcern.

A relatively new research tool in pre-gnancy is the study of infant-derivedMSCs. In utero, MSCs give rise to specific

Figure 3—Fetal adaptations to maternal obesity and GDM. Maternal obesity, estimated toaffect about one in four pregnant women, and GDM and type 2 diabetes in pregnancy, therates of which are rapidly growing, increase circulating glucose and lipids due to increasedlipolysis, hepatic glucose production, and dietary (chylomicron-derived) lipids. Newborns showincreased hepatic lipids, subcutaneous fat, and adipogenesis in infant-derived umbilical cordMSCs. MB changes in infants born to obese and GDMmothers are under study. MRS, magneticresonance spectroscopy.

care.diabetesjournals.org Friedman 1407

tissue types, including fat, skeletal mus-cle, and bone. In amniotic-derivedMSCsfrom obese women, there was a shift inMSC commitment to the adipocyte lin-eage during fetal development (82). Ourown preliminary studies point to new-born adiposity as a stronger phenotypicbiomarker of adipocyte differentiationand lower mitochondrial energy metab-olism in the MSCs from infants born toobese women, suggesting both factorscould contribute to lineage changes ininfant adiposity and low energy expen-diture. More mechanistic epigeneticand metabolomic analysis of these cellsand longitudinal analysis of these in-fants are under way as part of theHealthy Start Study (83) and may givecritical translational insight into path-ways underlying developmental originsof obesity.Although the transmission of meta-

bolic risk in infants born to motherswith obesity or GDM may be mediatedby excess fuels, inflammation, oxidativestress, and other metabolic risk factors,altering the maternal diet is one of thefew modifiable and potentially potentmanipulations. Dietary intake in humanpregnancy is often poorly measured, ifat all, and may be a very fundamentalsource of variability in metabolic path-ways influencing maternal and fetal fataccretion, requiring further investiga-tion. Recently, we have shown in highlycontrolled diet studies that womenwith GDM randomized to a eucalorichigher–complex carbohydrate/low-fat(CHOICE) diet compared with the con-ventional low-carbohydrate/higher-fatdiet resulted in normoglycemia and ap-peared to reduce maternal IR (84). Fur-thermore, after extending the diets todelivery with all meals provided, thesubjects on the CHOICE diet showed de-creased adipose cytokine expression, in-creased maternal adipose tissue insulinsensitivity, and a trend for lower infantadiposity (85) compared with those onthe conventional diet. These outcomessuggest the metabolic actions of lower-ing fat intake on maternal tissues are apromising target for reducing IR andcontrolling excess fuel transfer to thefetus. Our current studies are aimed atexploring how this diet alters maternalglycemic profiles and lipids, adipose tis-sue metabolism, placental nutrient trans-port and inflammation, and mechanismsunderlying infant growth, including

changes in infant liver steatosis and theMB (Fig. 3).

SUMMARY AND FUTUREDIRECTIONS

Fundamentally, fetal developmentalprogramming by maternal nutritionmay occur in two ways: first, by gene-environment interactions, such as diet,that may produce persistent epigeneticevents and second, by impacting normalorgan development to impart risk fordeveloping chronic disease(s). Most ofthe changes in adiposity in offspringborn to obese or GDM women occurwithin the normal range in birth weight,suggesting that programming likely in-volves subtle effects on metabolic regu-lation during development. The fact thatmany outcomes are modifiable by dietin the first 1,000 days of life suggeststhat maternal diet can be a powerful in-tervention to modify the transgenera-tional risk of obesity by modifying

organ growth and development, fat ac-quisition, appetite/behavior, and epi-genetic risk in the offspring, as shownin Fig. 4. For example, it is possible that al-tered maternal microbiota and mucosalimmunitymight directly influencematernalmetabolism and as a result influence thepathogenesis of IR and perhaps placentalfunction. While these observations ofchanges in the MB and inflammationare highly provocative and there is evi-dence for increased circulating lipopoly-saccharide levels in pregnant obesepatients (86), they suggest a hypothesisfor future experiments, including the de-velopment of novel pre/probiotics forpregnancy. Tying together the transmis-sion of the maternal MB from an obeseor GDM mother with antibiotic expo-sure, mode of delivery, infant energy re-tention, and immune function that maycontribute to predisposition to NAFLDand other immunologic diseases alsodeserves greater attention. The

Figure 4—The impact of maternal diet on changes in obesity risk includes epigenetic and MBchanges during the first 1,000 days of life. Metabolomic changes in mothers and babies may beassociated with exposure to changes in gut microbes, dietary fatty acids, inflammation, excessinsulin and leptin, along with antibiotics, and they may impact the host epigenome to increaseinfant weight gain. B/F, bacteroidetes/firmicutes; C-section, cesarean section; GWG, gestationalweight gain; LPS, lipopolysaccharide; miRNA, microRNA; ncRNA, noncoding RNA.

1408 Norbert Freinkel Award Lecture Diabetes Care Volume 38, August 2015

development of comprehensive ad-vanced techniques, including deep se-quencing of bacteria genomes (themetagenome), epigenetic platforms(methylation, histone modification,microRNAs, and noncoding RNAs),and metabolomic discovery tools, mayfundamentally inform our research direc-tions.While human investigation is inher-

ently variable and large sample sizes areoften needed to sort out trends fromnoise in such data sets, random samplingunder controlled conditions can increasetheir predictive accuracy. Metabolomicintermediates and biomarkers, includingAAs, lipids, and carbohydrate intermedi-ates, can influence the offspring in utero,and data on these biomarkers are beingapplied to human pregnancy (87–89).However, our understanding of the me-tabolomic signatures that influence in-fant growth and development duringthe first 2 years of life is minimal andgravely needed. Human studies outsideof pregnancy suggest that reduced die-tary fat intake and improved exercise canslow down the progression of diabetesfrom GDM (90,91). However, random-ized interventional trials in pregnancythat attempt to modify lifestyle in orderto favorably affect infant outcomes havebeen largely disappointing (92). Whileefforts to address the environmental fac-tors influencing infant health (e.g., exer-cise, limiting maternal weight gain,promoting exclusive breast-feeding,and administrating pre/probiotics) areunder way, sizable dietary changes maylead to better outcomes. Although thereremains a considerable challenge to de-liver these interventions in low-resourcesettings, the genetic and epidemiologicallife-course data showing long-term ef-fects of maternal obesity or GDM ex-posure on the metabolic health ofoffspring commands our efforts to dis-cern how gestational exposures in themodern environment can be specificallytargeted to reduce childhood obesityrisk. Primary intervention in utero andin early life that attenuates obesity poten-tial may be one of the most importantpublic health efforts to enhance popula-tion health worldwide.

Acknowledgments. The author would like tothank Dr. Linda A. Barbour and Rachel C. Janssen,both of the University of Colorado Denver, forhelp in the preparation of the manuscript.

Funding. This work was supported by NationalInstitutes of Health grants (5R01-DK078645, 5R01-DK076648-03, R21-DK088324, R24-DK90964, NIH-P30DK048520), theColoradoNutritionandObesityResearch Center, American Diabetes Association/GlaxoSmithKline Targeted Research Award 1-13-GSK-13, and National Institutes of Health NationalCenter for Advancing Translational Sciences Colo-rado Clinical and Translational Science Award grantUL1 TR001082.Duality of Interest. No potential conflicts ofinterest relevant to this article were reported.

References1. Barker DJ,Winter PD, Osmond C, Margetts B,Simmonds SJ. Weight in infancy and death fromischaemic heart disease. Lancet 1989;2:577–5802. Barker DJ. The origins of the developmentalorigins theory. J Intern Med 2007;261:412–4173. Bhargava SK, Sachdev HS, Fall CH, et al. Re-lation of serial changes in childhood body-massindex to impaired glucose tolerance in youngadulthood. N Engl J Med 2004;350:865–8754. Eriksson JG, Forsen T, Tuomilehto J, OsmondC, Barker DJ. Early growth and coronary heartdisease in later life: longitudinal study. BMJ2001;322:949–9535. Knight B, Shields BM, Hill A, Powell RJ,WrightD, Hattersley AT. The impact of maternal glyce-mia and obesity on early postnatal growth in anondiabetic Caucasian population. DiabetesCare 2007;30:777–7836. Metzger BE, Lowe LP, Dyer AR, et al; HAPOStudy Cooperative Research Group. Hyperglyce-mia and adverse pregnancy outcomes. N Engl JMed 2008;358:1991–20027. Symonds ME, Gopalakrishnan G, Bispham J,et al. Maternal nutrient restriction during pla-cental growth, programming of fetal adiposityand juvenile blood pressure control. Arch Phys-iol Biochem 2003;111:45–528. Freinkel N. Banting Lecture 1980. Of pregnancyand progeny. Diabetes 1980;29:1023–10359. Frias AE, Morgan TK, Evans AE, et al. Maternalhigh-fat diet disturbs uteroplacental hemo-dynamics and increases the frequency of stillbirthin a nonhuman primatemodel of excess nutrition.Endocrinology 2011;152:2456–246410. Sullivan EL, Grayson B, Takahashi D, et al.Chronic consumption of a high-fat diet duringpregnancy causes perturbations in the seroto-nergic system and increased anxiety-like behav-ior in nonhuman primate offspring. J Neurosci2010;30:3826–383011. Sullivan EL, Smith MS, Grove KL. Perinatalexposure to high-fat diet programs energy bal-ance, metabolism and behavior in adulthood.Neuroendocrinology 2011;93:1–812. Grant WF, Gillingham MB, Batra AK, et al.Maternal high fat diet is associated with de-creased plasma n-3 fatty acids and fetal hepaticapoptosis in nonhuman primates. PLoS One2011;6:e1726113. McCurdy CE, Bishop JM, Williams SM, et al.Maternal high-fat diet triggers lipotoxicity in thefetal livers of nonhuman primates. J Clin Invest2009;119:323–33514. Aagaard-Tillery KM, Grove K, Bishop J, et al.Developmental origins of disease and determi-nants of chromatin structure: maternal diet

modifies the primate fetal epigenome. J MolEndocrinol 2008;41:91–10215. Cox J, Williams S, Grove K, Lane RH,Aagaard-Tillery KM. A maternal high-fat diet isaccompanied by alterations in the fetal primatemetabolome. Am J Obstet Gynecol 2009;201:281.e1–916. Suter MA, Chen A, Burdine MS, et al. A ma-ternal high-fat diet modulates fetal SIRT1 histoneand protein deacetylase activity in nonhumanprimates. FASEB J 2012;26:5106–511417. Suter MA, Takahashi D, Grove KL, AagaardKM. Postweaning exposure to a high-fat diet isassociated with alterations to the hepatic his-tone code in Japanese macaques. Pediatr Res2013;74:252–25818. Catalano PM, Huston L, Amini SB, KalhanSC. Longitudinal changes in glucose metabolismduring pregnancy in obese women with normalglucose tolerance and gestational diabetes mel-litus. Am J Obstet Gynecol 1999;180:903–91619. Barbour LA, McCurdy CE, Hernandez TL,Friedman JE. Chronically increased S6K1 is asso-ciated with impaired IRS1 signaling in skeletalmuscle of GDM women with impaired glucosetolerance postpartum. J Clin Endocrinol Metab2011;96:1431–144120. Hamilton JK, Odrobina E, Yin J, Hanley AJ,Zinman B, Retnakaran R. Maternal insulin sen-sitivity during pregnancy predicts infant weightgain and adiposity at 1 year of age. Obesity (Sil-ver Spring) 2010;18:340–34621. Shapiro AL, Schmiege SJ, Brinton JT, et al. Test-ing the fuel-mediated hypothesis: maternal insulinresistance and glucosemediate the association be-tween maternal and neonatal adiposity, theHealthy Start study. Diabetologia 2015;58:937–94122. Radaelli T, Lepercq J, Varastehpour A, BasuS, Catalano PM, Hauguel-De Mouzon S. Differ-ential regulation of genes for fetoplacental lipidpathways in pregnancy with gestational andtype 1 diabetes mellitus. Am J Obstet Gynecol2009;201:209.e1–209.e1023. Aye IL, Jansson T, Powell TL. Interleukin-1binhibits insulin signaling and prevents insulin-stimulated system A amino acid transport inprimary human trophoblasts. Mol Cell Endocri-nol 2013;381:46–5524. Friedman JE, Ishizuka T, Shao J, Huston L,HighmanT, Catalano P. Impaired glucose transportand insulin receptor tyrosine phosphorylation inskeletal muscle from obese women with gesta-tional diabetes. Diabetes 1999;48:1807–181425. Barbour LA, Mizanoor Rahman S, GurevichI, et al. Increased P85alpha is a potent negativeregulator of skeletal muscle insulin signalingand induces in vivo insulin resistance associatedwith growth hormone excess. J Biol Chem 2005;280:37489–3749426. del Rincon JP, Iida K, Gaylinn BD, et al.Growth hormone regulation of p85alpha ex-pression and phosphoinositide 3-kinase activityin adipose tissue:mechanism for growth hormone-mediated insulin resistance. Diabetes 2007;56:1638–164627. Barbour LA, McCurdy CE, Hernandez TL,Kirwan JP, Catalano PM, Friedman JE. Cellularmechanisms for insulin resistance in normalpregnancy and gestational diabetes. DiabetesCare 2007;30(Suppl. 2):S112–S11928. Boyle KE, Newsom SA, Janssen RC, LappasM, Friedman JE. Skeletal muscle MnSOD,

care.diabetesjournals.org Friedman 1409

mitochondrial complex II, and SIRT3 enzyme ac-tivities are decreased inmaternal obesity duringhuman pregnancy and gestational diabetesmel-litus. J Clin Endocrinol Metab 2013;98:E1601–E160929. Boyle KE, Hwang H, Janssen RC, et al. Ges-tational diabetes is characterized by reducedmitochondrial protein expression and alteredcalcium signaling proteins in skeletal muscle.PLoS One 2014;9:e10687230. Friedman JE, Kirwan JP, Jing M, Presley L,Catalano PM. Increased skeletal muscle tumornecrosis factor-alpha and impaired insulin sig-naling persist in obese women with gestationaldiabetes mellitus 1 year postpartum. Diabetes2008;57:606–61331. HAPO Study Cooperative Research Group.Hyperglycemia and Adverse Pregnancy Outcome(HAPO) Study: associations with neonatal anthro-pometrics. Diabetes 2009;58:453–45932. Harmon KA, Gerard L, Jensen DR, et al. Con-tinuous glucose profiles in obese and normal-weight pregnant women on a controlled diet:metabolic determinants of fetal growth. Diabe-tes Care 2011;34:2198–220433. Pacifico L, Cantisani V, Ricci P, et al. Nonalco-holic fatty liver disease and carotid atherosclerosisin children. Pediatr Res 2008;63:423–42734. Welsh JA, Karpen S, VosMB. Increasing prev-alence of nonalcoholic fatty liver disease amongUnited States adolescents, 1988-1994 to 2007-2010. J Pediatr 2013;162:496–500.e135. Alisi A, Manco M, Vania A, Nobili V. Pediat-ric nonalcoholic fatty liver disease in 2009.J Pediatr 2009;155:469–47436. Thorn SR, Baquero KC, Newsom SA, et al.Early life exposure to maternal insulin resis-tance has persistent effects on hepatic NAFLDin juvenile nonhuman primates. Diabetes 2014;63:2702–271337. Brumbaugh DE, Friedman JE. Developmen-tal origins of nonalcoholic fatty liver disease.Pediatr Res 2014;75:140–14738. Modi N, Murgasova D, Ruager-Martin R,et al. The influence of maternal bodymass indexon infant adiposity and hepatic lipid content.Pediatr Res 2011;70:287–29139. Gale C, Thomas EL, Jeffries S, et al. Adiposityand hepatic lipid in healthy full-term, breastfed,and formula-fed human infants: a prospectiveshort-term longitudinal cohort study. Am J ClinNutr 2014;99:1034–104040. Anderson EL, Howe LD, Fraser A, et al.Weight trajectories through infancy and child-hood and risk of non-alcoholic fatty liver diseasein adolescence: the ALSPAC study. J Hepatol2014;61:626–63241. Breij LM, Kerkhof GF, Hokken-Koelega AC.Accelerated infant weight gain and risk for non-alcoholic fatty liver disease in early adulthood.J Clin Endocrinol Metab 2014;99:1189–119542. ArmstrongMJ, Adams LA, Canbay A, SynWK.Extrahepatic complications of nonalcoholic fattyliver disease. Hepatology 2014;59:1174–119743. Berkowitz RI, Moore RH, Faith MS, StallingsVA, Kral TV, Stunkard AJ. Identification of anobese eating style in 4-year-old children bornat high and low risk for obesity. Obesity (SilverSpring) 2010;18:505–51244. Begg DP, Woods SC. Hedonic and homeo-static overlap following fat ingestion. CellMetab 2013;18:459–460

45. Lozupone CA, Knight R. Species divergenceand the measurement of microbial diversity.FEMS Microbiol Rev 2008;32:557–57846. Koren O, Goodrich JK, Cullender TC, et al.Host remodeling of the gut microbiome andmetabolic changes during pregnancy. Cell2012;150:470–48047. BearfieldC,Davenport ES, SivapathasundaramV, Allaker RP. Possible association between amni-otic fluid micro-organism infection and microflorain the mouth. BJOG 2002;109:527–53348. Jimenez E, Fernandez L, Marın ML, et al.Isolation of commensal bacteria from umbilicalcord blood of healthy neonates born by cesar-ean section. Curr Microbiol 2005;51:270–27449. Jimenez E, Marın ML, Martın R, et al. Is me-conium from healthy newborns actually sterile?Res Microbiol 2008;159:187–19350. Stout MJ, Conlon B, Landeau M, et al. Identi-fication of intracellular bacteria in the basal plateof the human placenta in term and preterm ges-tations. Am J Obstet Gynecol 2013;208:226.e1–751. Aagaard K, Ma J, Antony KM, Ganu R,Petrosino J, Versalovic J. The placenta harbors aunique microbiome. Sci Transl Med 2014;6:237ra26552. Maynard CL, Elson CO, Hatton RD, WeaverCT. Reciprocal interactions of the intestinal mi-crobiota and immune system. Nature 2012;489:231–24153. Nauta AJ, Ben Amor K, Knol J, Garssen J, vander Beek EM. Relevance of pre- and postnatal nu-trition to development and interplay between themicrobiota and metabolic and immune systems.Am J Clin Nutr 2013;98:586S–593S54. Ajslev TA, Andersen CS, Gamborg M,Sørensen TI, Jess T. Childhood overweight afterestablishment of the gut microbiota: the role ofdelivery mode, pre-pregnancy weight and earlyadministration of antibiotics. Int J Obes (Lond)2011;35:522–52955. Trasande L, Blustein J, LiuM, Corwin E, Cox LM,Blaser MJ. Infant antibiotic exposures and early-lifebody mass. Int J Obes (Lond) 2013;37:16–2356. Blustein J, Attina T, Liu M, et al. Associationof caesarean delivery with child adiposity fromage 6 weeks to 15 years. Int J Obes (Lond) 2013;37:900–90657. Huh SY, Rifas-Shiman SL, Zera CA, et al. De-livery by caesarean section and risk of obesity inpreschool age children: a prospective cohortstudy. Arch Dis Child 2012;97:610–61658. CoatesME, Fuller R,HarrisonGF, LevM, SuffolkSF. A comparison of the growth of chicks in theGustafsson germ-free apparatus and in a conven-tional environment, with and without dietary sup-plements of penicillin. Br J Nutr 1963;17:141–15059. Dethlefsen L, Huse S, Sogin ML, Relman DA.The pervasive effects of an antibiotic on thehuman gut microbiota, as revealed by deep16S rRNA sequencing. PLoS Biol 2008;6:e28060. Dominguez-Bello MG, Costello EK,Contreras M, et al. Delivery mode shapes theacquisition and structure of the initialmicrobiotaacross multiple body habitats in newborns. ProcNatl Acad Sci U S A 2010;107:11971–1197561. Ma J, Prince AL, Bader D, et al. High-fatmaternal diet during pregnancy persistently al-ters the offspring microbiome in a primatemodel. Nat Commun 2014;5:388962. HeerwagenMJ, Stewart MS, de la HoussayeBA, Janssen RC, Friedman JE. Transgenic

increase in n-3/n-6 fatty acid ratio reduces ma-ternal obesity-associated inflammation and lim-its adverse developmental programming inmice. PLoS One 2013;8:e6779163. Russell WR, Gratz SW, Duncan SH, et al.High-protein, reduced-carbohydrate weight-loss diets promote metabolite profiles likely tobe detrimental to colonic health. Am J Clin Nutr2011;93:1062–107264. Smith SC, Choy R, Johnson SK, Hall RS,Wildeboer-Veloo AC, Welling GW. Lupin kernelfiber consumption modifies fecal microbiota inhealthy men as determined by rRNA gene fluo-rescent in situ hybridization. Eur J Nutr 2006;45:335–34165. Finley JW, Burrell JB, Reeves PG. Pinto beanconsumption changes SCFA profiles in fecal fer-mentations, bacterial populations of the lowerbowel, and lipid profiles in blood of humans. JNutr 2007;137:2391–239866. Korpela K, Flint HJ, Johnstone AM, et al. Gutmicrobiota signatures predict host and micro-biota responses to dietary interventions inobese individuals. PLoS One 2014;9:e9070267. Goodrich JK, Waters JL, Poole AC, et al. Hu-man genetics shape the gut microbiome. Cell2014;159:789–79968. Wong JM, de Souza R, Kendall CW, EmamA,Jenkins DJ. Colonic health: fermentation andshort chain fatty acids. J Clin Gastroenterol2006;40:235–24369. Al-Lahham S, Roelofsen H, Rezaee F, et al.Propionic acid affects immune status and me-tabolism in adipose tissue from overweight sub-jects. Eur J Clin Invest 2012;42:357–36470. Al-Lahham SH, Roelofsen H, Priebe M, et al.Regulation of adipokine production in humanadipose tissue by propionic acid. Eur J Clin Invest2010;40:401–40771. Rumberger JM, Arch JR, Green A. Butyrateand other short-chain fatty acids increase therate of lipolysis in 3T3-L1 adipocytes. PeerJ2014;2:e61172. Priyadarshini M, Thomas A, Reisetter AC,et al. Maternal short-chain fatty acids are asso-ciated with metabolic parameters in mothersand newborns. Transl Res 2014;164:153–15773. Gamet L, Daviaud D, Denis-Pouxviel C,Remesy C, Murat JC. Effects of short-chain fattyacids on growth and differentiation of the hu-man colon-cancer cell line HT29. Int J Cancer1992;52:286–28974. Park J, Kim M, Kang SG, et al. Short-chainfatty acids induce both effector and regulatory Tcells by suppression of histone deacetylases andregulation of the mTOR-S6K pathway. MucosalImmunol 2015;8:80–9375. Gao Z, Yin J, Zhang J, et al. Butyrate improvesinsulin sensitivity and increases energy expendi-ture in mice. Diabetes 2009;58:1509–151776. Kumar H, Lund R, Laiho A, et al. Gut micro-biota as an epigenetic regulator: pilot studybased on whole-genome methylation analysis.MBio 2014;5:e02113-1477. Relton CL, Groom A, St Pourcain B, et al. DNAmethylation patterns in cord blood DNA and bodysize in childhood. PLoS One 2012;7:e3182178. Morales E, Groom A, Lawlor DA, Relton CL.DNA methylation signatures in cord blood asso-ciated with maternal gestational weight gain:results from the ALSPAC cohort. BMC Res Notes2014;7:278

1410 Norbert Freinkel Award Lecture Diabetes Care Volume 38, August 2015

79. Godfrey KM, Sheppard A, Gluckman PD,et al. Epigenetic gene promoter methylation atbirth is associated with child’s later adiposity.Diabetes 2011;60:1528–153480. Teh AL, Pan H, Chen L, et al. The effect ofgenotype and in utero environment on interin-dividual variation in neonate DNA methylomes.Genome Res 2014;24:1064–107481. Ollikainen M, Smith KR, Joo EJ, et al. DNAmethylation analysis of multiple tissues fromnewborn twins reveals both genetic and intra-uterine components to variation in the humanneonatal epigenome. Hum Mol Genet 2010;19:4176–418882. Iaffaldano L, Nardelli C, Raia M, et al. Highaminopeptidase N/CD13 levels characterize hu-man amniotic mesenchymal stem cells and drivetheir increased adipogenic potential in obesewomen. Stem Cells Dev 2013;22:2287–229783. Crume TL, Shapiro AL, Brinton JT, et al. Ma-ternal fuels and metabolic measures duringpregnancy and neonatal body composition:

the Healthy Start Study. J Clin Endocrinol Metab2015;100:1672–168084. Hernandez TL, Van Pelt RE, Anderson MA,et al. A higher-complex carbohydrate diet ingestational diabetes mellitus achieves glucosetargets and lowers postprandial lipids: a ran-domized crossover study. Diabetes Care 2014;37:1254–126285. Hernandez TL, Van Pelt RE, Anderson MA,et al. Women with gestational diabetes random-ized to a higher complex carbohydrate/low fat dietmanifest lower adipose tissue insulin resistance,inflammation, glucose, and free fatty acids. Diabe-tes Care 2014;37:1254–126286. Basu S, HaghiacM, Surace P, et al. Pregravidobesity associates with increased maternal en-dotoxemia and metabolic inflammation. Obe-sity (Silver Spring) 2011;19:476–48287. SachseD, Sletner L,Mørkrid K, et al.Metabolicchanges in urine during and after pregnancy in alarge, multiethnic population-based cohort studyof gestational diabetes. PLoS One 2012;7:e52399

88. Kuc S, Koster MP, Pennings JL, et al. Metab-olomics profiling for identification of novelpotential markers in early prediction of pre-eclampsia. PLoS One 2014;9:e9854089. Austdal M, Skrastad RB, Gundersen AS,Austgulen R, Iversen AC, Bathen TF. Metabolomicbiomarkers in serum and urine in women withpreeclampsia. PLoS One 2014;9:e9192390. JohnsDJ, Hartmann-Boyce J, Jebb SA, AveyardP; Behavioural Weight Management ReviewGroup. Diet or exercise interventions vs combinedbehavioral weight management programs: a sys-tematic review and meta-analysis of direct com-parisons. J Acad Nutr Diet 2014;114:1557–156891. Schellenberg ES, Dryden DM, VandermeerB, Ha C, Korownyk C. Lifestyle interventions forpatients with and at risk for type 2 diabetes: asystematic review and meta-analysis. Ann In-tern Med 2013;159:543–55192. Tanvig M. Offspring body size and metabolicprofiledeffects of lifestyle intervention in obesepregnant women. Dan Med J 2014;61:B4893

care.diabetesjournals.org Friedman 1411