High Blood Glucose Levels In Early Pregnancy May Deprive Embryo Of Oxygen And Lead To Birth Defects

"Until recently, it was not understood how diabetic pregnancy couldcause birth defects. My laboratory wanted to explore this researchbecause the more we know about the effects of the mother's diabetes onthe embryo, the more tools we have to identify therapies that mayprevent birth defects in diabetic pregnancy," says the study's leadinvestigator, Mary R. Loeken, Ph.D., an investigator in Joslin'sSection on Developmental and Stem Cell Biology and Assistant Professorof Medicine at Harvard Medical School.

Women with both type 1 and type 2 diabetes run a high risk ofhaving babies with birth defects, especially of the heart and spinalcord. Because these organs form during the first few weeks ofpregnancy, coinciding with the time that a woman may first learn she ispregnant, aggressive control of blood glucose levels just before andafter conception is critical. "Women with diabetes should be consultingwith their healthcare team to be sure they have good glycemic controlbefore becoming pregnant," says Dr. Loeken. Maintaining blood glucosecontrol continues to be important throughout the pregnancy, but it isparticularly important during the first eight weeks, when an embryo'sorgans are forming.

In addition to recommending that women with diabetes have goodcontrol of their glucose levels before becoming pregnant, Dr. Loekenrecommends that obese women who don't know if they have diabetes butwho are planning to become pregnant be tested for diabetes. There havebeen several recent reports of increased birth defects in thepregnancies of obese women. "Many obese individuals have type 2diabetes and do not know it, so it is a good idea to bring glucoselevels to within the normal range before becoming pregnant, and tomonitor women with pre-diabetes closely during pregnancy to make surethat they don't develop diabetes," Dr. Loeken says.

In the new study, Dr. Loeken and her colleagues examinedembryos of pregnant mice injected with glucose (the sugar that iselevated in the blood during diabetes) to mimic diabetic pregnancy. Theresearchers knew that oxygen is needed by cells to break down glucoseand produce energy, and that normally, when oxygen is consumed, moreoxygen is delivered to tissues by increasing blood flow to thosetissues. However, at the stage of embryonic development in which birthdefects in women with diabetes frequently are believed to occur, theembryo does not yet have a heart or blood supply, and so the scientiststheorized it might not be possible to replace oxygen as rapidly as itis consumed. This has the potential to cause hypoxic stress, or damageto cells caused by low oxygen (hypoxia).

Working in collaboration with Peter Smith, Ph.D., Director ofthe BioCurrents Research Center at the Marine Biological Laboratory atWoods Hole, Mass., Dr. Loeken found that the oxygen concentrations inembryos of mice injected with glucose were significantly lower than incontrol embryos. This demonstrated that breaking down higher amounts ofglucose caused oxygen to be used up faster than it could be delivered.

The researchers then injected pregnant mice with glucose, orexposed them to varying levels of oxygen to see if raising and loweringoxygen delivery to the embryos had the same effect as raising andlowering glucose. The scientists' goal was to see whether oxygendeprivation is what mediates the effects of high glucose on the embryoin pregnant diabetic mice. Dr. Loeken's lab had previously found thatinducing high blood glucose levels in pregnant mice suppressed Pax3expression in embryos. Pax3 is a gene required for healthy formation ofthe brain and spinal cord.

In the new study, the researchers found that restricting oxygendelivery (by housing pregnant mice for one day in cages containing 12percent oxygen -- a concentration that did not cause any stress to themothers, but which might significantly reduce the amount of oxygendelivered to the uterus -- instead of 20 percent oxygen contained inroom air at sea level) had the same effect as high glucose. In fact,embryos from pregnant mice with high blood glucose levels, oroxygen-restricted mice, had five-fold decreases in Pax3 expression andeight-fold increases in a severe type of birth defect called neuraltube defects. Conversely, increasing the oxygen delivery to pregnantdiabetic mice (by housing them in cages containing 30 percent oxygen)blocked the decrease in Pax3 expression and neural tube defects intheir embryos.

Neural tube defects occur when parts of the brain, spinal cord,or their protective coverings fail to develop properly. For example,spina bifida -- the most common neural tube defect in humans -- resultsfrom the incomplete closure of the spinal cord. Neural tube defects andheart abnormalities are the most common birth defects affecting babiesborn to women with diabetes.

Previous research by Dr. Loeken and others has shown that inpregnant mice, high blood glucose levels boost an embryo's productionof free radicals -- products of metabolism that cause oxidative stress,and that oxidative stress leads to birth defects. These new experimentsshowed that glucose also caused hypoxic stress in embryos. However,what Dr. Loeken and her colleagues did not know was whether glucosecaused two different disturbances, hypoxic stress and oxidative stress,each having separate effects on Pax3 expression, or whether the twodisturbances were linked. To their surprise, when they investigated theproduction of free radicals, they found that embryos ofoxygen-restricted mice showed the same three- to five-fold changes inmarkers of oxidative stress as did embryos of glucose-injected mice.Conversely, the markers of oxidative stress were suppressed in embryosof oxygen-supplemented diabetic mice. These results suggest that thelack of oxygen caused by increased glucose consumption triggers theproduction of free radicals, which then causes birth defects, Dr.Loeken explains.

The researchers also found that administration of high levelsof antioxidants -- which keep free radicals from wreaking cellularhavoc -- prevented the decrease in Pax3 expression and birth defects inembryos of diabetic mice. This lends weight to the theory that lack ofoxygen and the accompanying increase in free radical production thatoccurs in mice with high blood glucose levels are what ultimatelyincrease risk of birth defects. "We're trying to pinpoint all the stepsthat occur between oxygen deprivation and gene expression, and toidentify the signals and molecules that regulate Pax3," Dr. Loekensays.

Funding for this study was provided in part by the NationalInstitutes of Health. The citation for the article is Am J PhysiolEndocrinol Metab 289: E591-E599, 2005 (October).