Maternal diabetes adversely affects preovulatory oocyte maturation, development, and granulosa cell apoptosis.

Maternal diabetes adversely affects preimplantation embryo development and pregnancy outcomes. The objective of this study was to determine whether diabetes has an impact at an earlier stage of development, the preovulatory oocyte. Models of both acute and chronic insulin-dependent diabetes were used. Acute hyperglycemia was induced by a single streptozotocin injection. Akita mice, which harbor an autosomal dominant mutation causing them to be chronically hypoinsulinemic and hyperglycemic, were used. In both models, preovulatory oocytes were markedly smaller when compared with control animals. A significantly greater number of control oocytes had progressed to meiotic maturation before diabetic oocytes. Both models were found to have smaller, less developed ovarian follicles with a greater number of apoptotic foci by histological evaluation as well as by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling staining. Immunohistochemistry displayed a greater amount of TNF-related apoptosis-inducing ligand (TRAIL) and KILLER, a key murine ligand and receptor involved in the extrinsic pathway, expressed in cumulus cells from hyperglycemic mice compared with controls, suggesting that this apoptotic pathway may be up-regulated under diabetic stress. Elevated KILLER expression was also confirmed through Western blotting. Connexin-43 expression was found to be lower by immunohistochemistry and Western blot analysis in the diabetic samples. Both models of maternal hyperglycemia and hypoinsulinemia may have a detrimental effect on oocyte maturation and development as detailed by the smaller sizes of oocytes and developing ovarian follicles, the lowered percentage reaching germinal vesicle breakdown, and the greater amount of apoptosis. In addition, there may be dysfunctional or decreased communication in diabetic oocytes, as demonstrated by lower expression of connexin-43.

Malformations in infants of diabetic mothers occur before the seventh gestational week. Implications for treatment.

In the present study we used a developmental morphologic approach to fix the latest time in development at which the malformations commonly reported in infants of diabetic mothers could occur. Developmental morphologic dating shows that the significantly more common congenital malformations in infants of diabetic mothers occur before the seventh week of gestation. This suggests that any therapeutic intervention aimed at decreasing the incidence of congenital malformations must be instituted during the critical early period.

Antepartum fetal surveillance in gestational diabetes mellitus.

Pregnancy in patients with gestational diabetes mellitus (GDM) is associated with increased perinatal morbidity. Whether the perinatal mortality rate, particularly the fetal death rate, is greater in these patients remains controversial. The present study was undertaken to review the role of antepartum fetal monitoring in 69 patients with GDM controlled by diet only (class A) and 28 women requiring insulin therapy (class AB). Hypertension complicated 21.6% of these pregnancies. Antepartum fetal surveillance included outpatient nonstress testing, urinary estriol assays, maternal assessment of fetal activity, and clinical estimation of fetal weight. All insulin-requiring patients as well as fourteen class A patients with identifiable risk factors underwent testing. No perinatal deaths occurred. Only six patients required intervention for suspected fetal jeopardy and four of these women had hypertension. Macrosomia was correctly identified in only 6 of 16 infants weighing 4000 g or more. This study suggests that, in GDM, an outpatient program of fetal testing, using primarily the nonstress test and maternal assessment of fetal activity, can be employed in patients requiring insulin as well as class A patients with identifiable risk factors. This protocol resulted in a low rate of unnecessary intervention and good perinatal outcome. The risks for abnormal antepartum testing results appear increased in GDM with hypertension and prolonged pregnancy.

Gestational diabetes mellitus. Is further improvement necessary?

The maternal antepartum, intrapartum, and neonatal characteristics of 158 patients with gestational diabetes mellitus (GDM) attending a large teaching hospital between 1979 and 1983 were described and compared with a matched nondiabetic control group. The primary cesarean section rate in patients with GDM (18%) was significantly greater than in the control group (11%, P less than 0.04). Neonatal macrosomia, as reflected in mean birthweight (P less than 0.04), the number of neonates weighing greater than 4 kg (P less than 0.05) and large-for-gestational-age infants (P less than 0.05), and the birthweight adjusted for gestational age (K-score, P less than 0.01) was significantly increased in the diabetic group. The characteristics of patients with GDM treated with diet alone and diet and insulin together were examined. The insulin-therapy group was characterized by more patients older than 25 yr (P less than 0.01) and a higher mean birthweight (3743 +/- 677 g) (P less than 0.02) than the diet-alone group. This may reflect an increased magnitude of glucose intolerance in the insulin-treated group. Obese patients with GDM delivered heavier neonates than the nonobese patients with GDM (P less than 0.01). Although there was no difference between the groups, perinatal mortality was present in this study. These data indicate that the major perinatal morbidity in GDM included increased cesarean section for fetal macrosomia. Early diagnosis with strict diagnostic criteria and rigid antenatal surveillance may result in further improvements in outcome.

Metabolic and therapeutic assessment of gestational diabetes by two-hour and twenty-four-hour isocaloric meal tolerance tests.

Lean and obese women with gestational diabetes (GDM) were given two different isocaloric meal challenge tests to assess glucose and insulin responses. Forty-three pregnant women received a 400-kcal isocaloric breakfast meal tolerance test (mini-MTT). Twenty of the subjects were also given a 2000-kcal isocaloric diet with three meals and three snacks during a 24-h period (maxi-MTT). This was the first study to utilize the physiologic challenge of mixed meals to compare insulin and glucose responses of both obese and lean normal pregnant women and women with GDM around the clock. Normal obese pregnant women had higher integrated glucose and insulin values around the 24-h clock (P less than 0.003 and less than 0.03, respectively) than lean pregnant women. Lean and obese women with GDM also responded differently to the physiologic challenge of mixed meals. Some, but not all, obese diabetic subjects were markedly hyperinsulinemic in contrast to lean diabetic women who were relatively insulin deficient. One normoglycemic massively obese 18-yr-old pregnant woman was unexpectedly found to be severely hyperinsulinemic. The two meal tolerance tests clearly defined a delay in the release of insulin in women with GDM (lean and obese) and markedly different quantitative insulin responses to identical meal challenges in obese diabetic subjects. Maternal hyperinsulinemia was positively correlated with prepregnancy body mass index (kg/m2) and heavier infants, but not with plasma glucose levels. These studies provide evidence that GDM is a heterogeneous syndrome.(ABSTRACT TRUNCATED AT 250 WORDS)

The possibility of an early growth delay in White's class A diabetic pregnancy.

Ultrasound scanning has revealed that some fetuses of women with insulin-dependent diabetes are smaller than normal in early pregnancy as judged by the crown-rump length. This early growth delay is negatively correlated to the quality of diabetes regulation. Nine White's Class A diabetic women had fetuses that were on average 4.5 days (range, 0-11 days) smaller than expected from the menstrual history. Apparently, the expectedly modest metabolic disturbances in early pregnancy of Class A diabetic women are able to interfere with normal embryonic growth.

Neonatal morbidities in infants of mothers with glucose intolerance in pregnancy.

Of 1839 pregnant women screened prospectively, 52 were identified to have glucose intolerance. Ten additional pregnant women identified as having glucose intolerance before the universal screening were also included in the study cohort. These 62 patients were followed in a perinatal high-risk clinic with weekly plasma glucose determinations. The patients were treated with diet and, in addition, 21 of 62 were treated with insulin therapeutically. By observational cohort design, the infants and a comparable number of matched controls were evaluated for evidence of neonatal morbidities and classified into percentile for birth weight. Compared with the control group, the operative mode of delivery, the mean birth weight, the birth-weight percentile, the male/female ratio, the frequency of low Apgar score (less than or equal to 6 at 1 min), and the number of infants with congenital anomalies were significantly higher in the infants born to the glucose-intolerant mothers. Although the mean maternal blood sugar was maintained within a reasonably euglycemic range, the usual neonatal morbidities were not eliminated entirely. Further understanding and management of glucose intolerance in pregnancy is necessary to further diminish or eliminate neonatal morbidities.

Pregnancy in diabetes complicated by vascular disease.

Over 50 years of experience with pregnancy in diabetic women is reviewed. In particular, the maternal and fetal survival in mothers with either microvascular or macrovascular disease is considered. This includes White classes E, F, R, RF, H, and T. In this group of patients with vascular disease, maternal survival during pregnancy is virtually 100 per cent with the exception of class H (ischemic heart disease). Fetal survival has steadily improved throughout the time period examined, but is still considerably below that of pregnancies occurring in women without vascular disease. Long-term maternal survival is adversely affected by the first decade after delivery.

Evidence for multifactorial origin of diabetes-induced embryopathies.

Serum factors characteristically altered in the diabetic state, e.g., glucose, ketone bodies (beta-hydroxybutyrate), and somatomedin inhibitors, induce dysmorphogenesis with or without growth retardation in rodent embryos in whole-embryo culture. Furthermore, serum from diabetic animals, which contains combinations of altered factors, is teratogenic, thereby implying that the diabetic embryopathy is of a multifactorial origin. However, a detailed investigation with various combinations of factors at known concentrations to test this hypothesis has not been conducted. Therefore, we employed combinations of hyperglycemia (300 and 600 mg/dl; 16.6 and 33.3 mM), hyperketonemia (8 and 16 mM D-beta-hydroxybutyrate), and an 800- to 1000-Mr somatomedin-inhibitor serum fraction (at an amount equal to that found in 0.05 and 0.1 ml of serum from a diabetic rat per deciliter culture medium), which represented doses with low teratogenic potential, to determine if interactions of the factors could occur that would increase the risk of malformations in mouse embryos exposed in whole-embryo culture during gastrulation and neurulation. The results demonstrate that glucose and D-beta-hydroxybutyrate can act synergistically to produce growth retardation and additively to induce malformations. The addition of the somatomedin inhibitor exacerbates the induction of malformations produced by the ketone body and glucose. Thus, the origin of the diabetic embryopathy may be multifactorial, involving several altered maternal factors.

Growth retardation during early organogenesis in embryos of experimentally diabetic rats.

Embryos from rats rendered diabetic with streptozotocin for 1 wk or more before conception were examined on day 11.5 of gestation (i.e., at the 26-29-somite stage of normal rat embryonic development). The studies were designed to assess whether poorly regulated maternal diabetes is associated with demonstrable abnormalities even during this early phase of embryogenesis. We found that manifest retardations in growth and development were invariably present as judged by significant reductions in crown-rump length and somite number, respectively. Total protein and DNA content of the embryos were also reduced, although not symmetrically, so that protein/DNA ratios were increased. Gross dysmorphogenic lesions in neural tissue disproportional to the overall growth retardation at 11.5 days could not be demonstrated. The findings suggest that maternal diabetes can compromise intra-uterine growth and development during the period preceding and coinciding with the establishment of circulation in the allantoic placenta. The possible multifactorial determinants remain to be elucidated. It also remains to be established whether the early embryotoxicity provides a setting conducive to the increased dysmorphogenesis that is traditionally recognized during the later stages of pregnancy complicated by diabetes.

Maternal diabetes increases the risk of caudal regression caused by retinoic acid.

Maternal diabetes increases the risk of congenital malformations in the offspring of affected pregnancies. This increase arises from the teratogenic effect of the maternal diabetic milieu on the developing embryo, although the mechanism of this action is poorly understood. In the present study, we examined whether the vitamin A metabolite retinoic acid (RA), a common drug with well-known teratogenic properties, may interact with maternal diabetes to alter the incidence of congenital malformations in mice. Our results show that when treated with RA, embryos of diabetic mice are significantly more prone than embryos of nondiabetic mice to develop caudal regression, a defect that is highly associated with diabetic pregnancy in humans. By studying the vestigial tail (Wnt-3a(vt)) mutant, we provide evidence that Wnt-3a, a gene that controls the development of the caudal region, is directly involved in the pathogenic pathway of RA-induced caudal regression. We further show that the molecular basis of the increased susceptibility of embryos of diabetic mice to RA involves enhanced downregulation of Wnt-3a expression. This positive interaction between RA and maternal diabetes may have implications for humans in suggesting increased susceptibility to environmental teratogens during diabetic pregnancy.

Polymorphic susceptibility to the molecular causes of neural tube defects during diabetic embryopathy.

Previously, we demonstrated that neural tube defects (NTDs) are significantly increased in a mouse model of diabetic pregnancy. In addition, expression of Pax-3, a gene encoding a transcription factor required for neural tube development, is significantly decreased. This suggests that diabetic embryopathy results from impaired expression of genes regulating essential morphogenetic processes. Here, we report that in one mouse strain, C57Bl/6J, embryos are resistant to the effects of maternal diabetes on NTDs and Pax-3 expression, in contrast to a susceptible strain, FVB, in which maternal diabetes significantly increases NTDs (P = 0.02) and inhibits Pax-3 expression (P = 0.01). Resistance to NTDs caused by diabetic pregnancy is a dominant trait, as demonstrated by heterozygous embryos of diabetic or nondiabetic mothers of either strain. There was no significant difference between strains in expression of genes that regulate free radical scavenging pathways, suggesting that susceptibility to oxidative stress does not account for the genetic differences. Understanding the genetic bases for differential susceptibility to altered gene expression and NTDs in diabetic mice may be important in delineating the mechanisms by which maternal hyperglycemia interferes with embryo gene expression. Moreover, if susceptibility to diabetic embryopathy is variable in humans as well as in mice, it may be possible to screen individuals at increased risk for this complication of diabetes.

Gestational diabetes. Incidence, maternal characteristics, and perinatal outcome.

Accurate estimates of the incidence of abnormal glucose tolerance during pregnancy are virtually nonexistent. Screening select populations of women with risk factors for the condition and the nonrandom, non-population-based nature of most studies have given rise to wide variances in reported incidence. We analyzed data from the states of Mississippi and Washington and from the National Natality and Fetal Mortality Surveys conducted in 1980 in an attempt to provide more accurate population-based estimates of the incidence of gestational diabetes mellitus (GDM). In the national surveys GDM was noted (screening and diagnostic criteria were unavailable) as a complication in 0.38% of all sampled pregnancies; overt (type I and type II) diabetes was noted in 0.78%. Mean maternal age for the GDM group was 28.4 yr; 85% were white (81% controls) and 15% non-white (19% controls). Prepregnancy weights were higher in the GDM group by an average of 20 lb. However, mean weight gain was less in this group than in controls (23 versus 29 lb). Perinatal mortality was noted in approximately 2.8% (1.3% in controls) of the offspring in GDM-complicated pregnancies and congenital malformations in 6.4% (7.9% in controls). Methodologic problems were encountered and included lack of screening and diagnostic criteria, underreporting, and underrecording.

Diabetes in pregnancy. Skeletal malformations in the offspring of diabetic rats after intermittent withdrawal of insulin in early gestation.

Precise timing of the teratogenic period in diabetic pregnancy is of clinical importance since correction of the glucose intolerance during this period may protect the offspring from malformations. An experimental approach to elucidate this problem with regard to skeletal development was made in groups of pregnant streptozotocin-diabetic rats (MDI), which were treated with daily insulin injections except for a 2-day period in the first half of pregnancy. The degree of metabolic derangement was estimated by measurements of serum glucose concentrations. During the insulin-free period, the rats showed severe hyperglycemia (greater than 20 mM) while during ongoing insulin treatment, only brief periods of hyper- or hypoglycemia were observed. Insulin treatment was withdrawn successively between gestational days 3 and 12. Control groups consisted of normal pregnant rats (N) or pregnant rats with manifest diabetes (MD) without insulin treatment. The serum glucose levels of the N animals were below 6 mM while those of the MD animals were above 25 mM throughout pregnancy. Skeletal malformations in the viable offspring were recorded on gestational day 20 after Alizarin staining of calcified ossification centers, which also allowed an estimate of skeletal development as a whole. Untreated diabetes in the MD rats induced a high rate of fetal resorptions, a decrease in fetal weight and viability, as well as retardation of skeletal development. Intermittent insulin treatment in the MDI rats ameliorated, but did not abolish, these changes. In the MD group 9 of 48 viable fetuses showed severe malformations of either the lower jaw (micrognathia) or of the lumbosacral region (caudal dysgenesis).(ABSTRACT TRUNCATED AT 250 WORDS)

Type 2 diabetes and low birth weight: the role of paternal inheritance in the association of low birth weight and diabetes.

Lower birth weight is associated with an increased occurrence of type 2 diabetes in later life. Whether this relationship is explained by environmental or genetic factors is unknown. We have examined the potential for genetic influences by determining whether parental diabetes is associated with lower birth weight in 1,608 children of known birth weight and gestational age born between 1941 and 1993 in the Gila River Indian Community in Arizona. The previously described relationships of maternal diabetes to increased birth weight and offspring diabetes were observed. In contrast to this we have determined novel relationships between low birth weight and paternal diabetes. The offspring of diabetic fathers were, on average, 78 g lighter than the offspring of nondiabetic fathers. For fathers, lower birth weight in their offspring was associated with an increased risk of later diabetes, i.e., fathers of offspring in the lowest quintile of birth weight, who were not diabetic at the time of birth of their child, had a 1.8-fold increased risk of developing diabetes later in life (95% CI 1.2-2.7; P = 0.004). For children, lower birth weight predicted diabetes in the offspring if paternal but not maternal diabetes was present, but it was not associated with higher plasma glucose if neither parent had diabetes. We conclude that the risk of diabetes associated with low birth weight is strongly related to the development of paternal diabetes, suggesting a genetic link between lower birth weight and later diabetes.

Therapeutic results of insulin therapy in gestational diabetes mellitus.

Most studies of gestational diabetes mellitus (GDM) have reported a marked reduction in perinatal mortality with appropriate dietary regimens and good medical and obstetrical surveillance. Nevertheless, fetal morbidity, including macrosomia, has remained high and appears to be linked to factors other than plasma glucose control. In a review of six investigations in which insulin therapy was combined with an appropriate diet, the incidence of fetal macrosomia was reduced in five studies as compared with diet-only treatments. Again, the improvement did not always correlate with altered plasma glucose profiles. Other studies suggest that maternal plasma substrate disturbances other than glucose may contribute to the development of fetal macrosomia. To what extent insulin administration reduces morbidity by containing circulating maternal fuels, such as lipids and amino acids, in a more normal range remains to be determined. Moreover, the role of diet, maternal obesity, and weight gain during pregnancy adds to the complexity of factors influencing obstetrical outcome in gestational diabetes. Until the relative importance of all of these variables is adequately assessed, criteria for selection of women with pregnancy-onset diabetes for insulin therapy are most likely to be based on fasting and postprandial plasma glucose concentrations.

Uteroplacental hemodynamic disturbances in establishment of fetal growth retardation in streptozocin-induced diabetic rats.

This study examined the relationship between uteroplacental blood flow and fetal hypotrophy in streptozocin-induced diabetic rats (40 mg/kg body wt i.v.). Our results showed that, in diabetic rats, fetal hypotrophy was associated with a significant reduction in arterial blood velocity in the uterine artery (P less than 0.001), placenta (P less than 0.01), umbilical artery (P less than 0.01), and fetal aorta (P less than 0.05). This was not observed when diabetic rats were treated with insulin. Treatment of rats with the alpha 1-blocking vasodilator nicergoline restored fetal growth and arterial blood velocity to control values without affecting the degree of hyperglycemia. Nicergoline in control rats did not change fetal weight and caused only minor hemodynamic changes on presumably already maximally vasodilated arteries. We concluded that the uteroplacental hemodynamic disturbances observed in diabetic rats play a major role in the establishment of fetal growth retardation.

Neural tube defects in embryos of diabetic mice: role of the Pax-3 gene and apoptosis.

Neural tube defects are among the most common of the malformations associated with diabetic embryopathy. To study the molecular mechanisms by which neural tube defects occur during diabetic pregnancy, we have developed a new experimental system using pregnant diabetic mice. In this system, the rate of neural tube defects is about three times higher in embryos of diabetic mice than in embryos of nondiabetic mice. Most of the defects affected presumptive midbrain and hindbrain structures and included open defects (i.e., exencephaly) and gross maldevelopment. By semiquantitative reverse transcription-polymerase chain reaction and in situ hybridization, we found that expression of Pax-3, a gene required for neural tube closure in the area of the midbrain and hindbrain, is significantly reduced in the embryos of diabetic mice. The same regions of the neural tube where Pax-3 had been underexpressed were found subsequently to contain high concentrations of cells undergoing apoptosis. Reduced expression of Pax-3 appears to be responsible for this apoptosis because apoptotic cells were also found at sites of neural tube defects in embryos carrying null mutation of the Pax-3 gene. Finally, mouse strains that carry null mutations in Pax-3 develop neural tube defects that resemble the malformations that occur in embryos of diabetic mice. These results suggest that Pax-3 is an important developmental control gene, expression of which is disturbed in embryos of diabetic mice, and that as a consequence, apoptosis of the neural tube occurs. This pathway may be responsible for many of the neural tube defects resulting from diabetic pregnancy.

Risk factors for conotruncal cardiac defects in Atlanta.

Because the causes of conotruncal cardiac defects are poorly understood, a case-control study was conducted to investigate maternal risk factors for conotruncal cardiac defects. Eligible cases included all infants who were born from 1976 through 1980 to residents of the five county metropolitan Atlanta area and diagnosed with truncus arteriosus, transposition of the great arteries or tetralogy of Fallot. Eligible control infants were a sample of comparable infants without birth defects. Maternal interviews were conducted for 73% (83 of 114) of eligible cases and 72% (1,303 of 1,804) of eligible control infants. The results showed increased risks associated with maternal diabetes (odds ratio 5.6; 90% confidence interval 2.5 to 15.6), maternal stress related to job loss, divorce, separation or death of a close friend or relative (odds ratio 2.4; 90% confidence interval 1.4 to 4.2) and a history of a sibling with a cardiac defect (odds ratio 4.8; 90% confidence interval 2.2 to 10.5). The statistical power of the data was adequate to rule out threefold or greater increases in risk for a wide variety of other exposures, including maternal illnesses other than diabetes, contraceptive use, nonmedicinal drugs (for example, coffee, tea, alcohol, cigarettes, street drugs), employment and education. This population-based study offers no clues that could explain either the high rate of transposition of the great arteries or the temporal trend of an increasing rate of tetralogy of Fallot in Atlanta.