Established diet-induced obesity in female rats leads to offspring hyperphagia, adiposity and insulin resistance.

AIMS/HYPOTHESIS: Accumulating evidence suggests that maternal obesity may increase the risk of metabolic disease in the offspring. We investigated the effects of established maternal diet-induced obesity on male and female offspring appetite, glucose homeostasis and body composition in rats. METHODS: Female Wistar rats were fed either a standard chow (3% fat, 7% sugar [wt/wt]) or a palatable obesogenic diet (11% fat, 43% sugar [wt/wt]) for 8 weeks before mating and throughout pregnancy and lactation. Male and female offspring of control and obese dams were weaned on to standard chow and assessed until 12 months of age. RESULTS: At mating, obese dams were heavier than control with associated hyperglycaemia and hyperinsulinaemia. Male and female offspring of obese dams were hyperphagic (p < 0.0001) and heavier than control (p < 0.0001) until 12 months of age. NEFA were raised at 2 months but not at 12 months. At 3 months, OGTT showed more pronounced alteration of glucose homeostasis in male than in female offspring of obese animals. Euglycaemic-hyperinsulinaemic clamps performed at 8 to 9 months in female and 10 to 11 months in male offspring revealed insulin resistance in male offspring of obese dams (p < 0.05 compared with control). Body compositional analysis at 12 months also showed increased fat pad weights in male and female offspring of obese animals. CONCLUSIONS/INTERPRETATION: Diet-induced obesity in female rats leads to a state of insulin resistance in male offspring, associated with development of obesity and increased adiposity. An increase in food intake may play a role.

Influence of the vitamin D-binding protein on the serum concentration of 1,25-dihydroxyvitamin D3. Significance of the free 1,25-dihydroxyvitamin D3 concentration.

The influence of the serum binding protein (DBP) for vitamin D and its metabolites on the concentration of its main ligands, 25-hydroxyvitamin D(3) (25-OHD(3)) and 1,25-dihydroxyvitamin D(3) (1,25-[OH](2)D(3)) was studied. The concentration of both 1,25-(OH)(2)D(3) and DBP in normal female subjects (45+/-14 ng/liter and 333+/-58 mg/liter, mean+/-SD, respectively; n = 58) increased during the intake of estro-progestogens (69+/-27 ng/liter and 488+/-90 mg/liter, respectively; n = 29), whereas the 25-OHD(3) concentration remained unchanged. A positive correlation was found between the concentrations of 1,25-(OH)(2)D(3) and DBP in these women. At the end of pregnancy, the total concentrations of 1,25-(OH)(2)D(3) (97+/-26 ng/liter, n = 40) and DBP (616+/-84 mg/liter) are both significantly higher than in nonpregnant females and paired cord serum samples (48+/-11 ng/liter and 266+/-41 mg/liter, respectively). A marked seasonal variation of 25-OHD(3) was observed in pregnant females and their infants, whereas in the same samples the concentrations of both DBP and 1,25-(OH)(2)D(3) remained constant throughout the year. The free 1,25-(OH)(2)D(3) index, calculated as the molar ratio of this steroid and DBP, remains normal in women taking estro-progestogens, however, and this might explain their normal intestinal calcium absorption despite a high total 1,25-(OH)(2)D(3) concentration. In pregnancy the free 1,25-(OH)(2)D(3) index remains normal up to 35 wk of gestation, but during the last weeks of gestation, the free 1,25-(OH)(2)D(3) index increases in both circulations. A highly significant correlation exists between the (total and free) 25-OHD(3) and 1,25-(OH)(2)D(3) concentrations in maternal and cord serum both at 35 and 40 wk of gestation.

Animal evidence for the transgenerational development of diabetes mellitus.

The mammalian fetus develops inside the uterus of its mother and is completely dependent on the nutrients supplied by its mother. Disturbances in the maternal metabolism that alter this nutrient supply from mother to fetus can induce structural and functional adaptations during fetal development, with lasting consequences for growth and metabolism of the offspring throughout life. This effect has been investigated, by several research groups, in different experimental models where the maternal metabolism during pregnancy was experimentally manipulated (maternal diabetes and maternal malnutrition) and the effect on the offspring was investigated. The altered maternal/fetal metabolism appears to be associated with a diabetogenic effect in the adult offspring, including gestational diabetes. This diabetic pregnancy in the offspring again induces a diabetogenic effect into the next generation, via adaptations during fetal development. These experimental data in laboratory animals are confirmed by epidemiological studies on infants of mothers suffering from diabetes or malnutrition during pregnancy. It can be concluded that fetal development in an abnormal intra-uterine milieu can induce alterations in the fetal metabolism, with lasting consequences for the glucose tolerance of the offspring in adult life. The most marked effect is the development of gestational diabetes, thereby transmitting the diabetogenic tendency to the next generation again. The concept of fetal origin of adult diabetes therefore is of major significance for public health in the immediate and the far future.

Long-term effect of diabetes and pregnancy in the rat.

Islet hyperplasia and B-cell degranulation were found in the fetuses of the third generation from mothers (second generation) born to a diabetic mother (first generation) regardless of the origin of the father, while pancreatic islets were normal in fetuses from control mothers, even when the father was an offspring of a diabetic mother. These data support the hypothesis that in our experimental model overstimulation of the fetal endocrine pancreas results in long-term consequences to the third generation.

Metabolic alterations in adulthood after intrauterine development in mothers with mild diabetes.

We studied the long-term effects of maternal diabetes mellitus on the offspring of experimentally induced diabetic Wistar rats. When stressed by an intravenous glucose load, the adult female offspring had impaired glucose tolerance and developed gestational diabetes mellitus when pregnant. Our results show that even mild diabetes mellitus induces an abnormal intrauterine milieu that causes morphological and functional changes in fetal development with consequences for later life.

Influence of lactation on morphometric and secretory variables in pancreatic beta-cell of mildly diabetic rats.

In nondiabetic rats, lactation accelerates the restoration of pancreatic beta-cell function after the period of increased secretory activity associated with pregnancy. To investigate whether a comparable situation prevails in mildly diabetic animals, streptozocin (22.5 mg/kg body wt) was administered to female rats at the onset of pregnancy. Plasma glucose and insulin concentrations, content and release of insulin in isolated islets, total mass and volume density of both the endocrine pancreas and granulated beta-cells, and ultrastructural prevalence of light and dark secretory granules were measured on the 20th day of pregnancy and in lactating and nonlactating animals 20 days after delivery. In the mildly diabetic animals, the changes in endocrine pancreatic function normally associated with pregnancy and lactation were greatly attenuated, albeit not completely eliminated. We propose that the increased biosynthetic and secretory activity imposed on surviving beta-cells after streptozocin administration tends to mask the adaptative changes in beta-cell function otherwise seen during the postpartum and lactation period.

Pancreatic endocrine cell fractions in erythroblastosis fetalis.

Pancreatic sections from 21 cases of rhesus disease and 20 control newborn infants of 30--40-wk gestational age were stained by the immunoperoxidase method for insulin, glucagon, somatostatin, and pancreatic polypeptide (PP). The fractional area occupied by each cell type was estimated, taking note of whether the gland contained PP-rich (ventral lobe) or PP-poor islets (dorsal lobe). In the PP-rich part of the pancreas, the volume fraction of all four endocrine cell types was significantly greater in the rhesus cases than in the controls. No difference was found between the two groups in the PP-poor part of the gland. The results show that abnormal development of the PP-rich part of the pancreas occurs in erythroblastosis fetalis. The localization of the changes to one part of the pancreas may explain some of the earlier conflicting reports on this topic.

In vivo glucose utilization by individual tissues in virgin and pregnant offspring of severely diabetic rats.

Adult offspring of diabetic rats or SDF rats are characterized by insulin resistance in the liver and extrahepatic tissues; this insulin resistance does not worsen during pregnancy. In this study, we determined the glucose metabolic index in tissues of anesthetized virgin and pregnant control and SDF rats in basal conditions and during a euglycemic hyperinsulinemic clamp. Tissues comprised insulin-sensitive tissues (five skeletal muscles, diaphragm, and periovarian white adipose tissue) and control tissues (duodenum and cerebrum). In addition, this study measured the GMI of placenta and fetuses. In basal conditions, SDF rats showed a slight decrease (9-29%) in the GMI of skeletal muscles compared with control rats; it was not altered by pregnancy in any of the tissues. During physiological hyperinsulinemia, virgin SDF rats exhibited a 25-70% decrease in the GMI of skeletal muscles compared with control rats; this decrease was not observed in diaphragm, or in adipose tissue in which the GMI was found to be increased. During pregnancy, SDF rats did not show an additional drop in the GMI of skeletal muscles, whereas the GMI of both skeletal muscles and adipose tissue was clearly diminished (25-60%) in control rats. The GMI of skeletal muscles was therefore comparable in pregnant control rats and SDF rats. The placental, but not fetal, GMI was increased by 24% during hyperinsulinemia in control rats; the placental and fetal GMIs, in basal and hyperinsulinemic conditions, were similar in control rats and SDF rats. In conclusion, skeletal muscles, but not white adipose tissue, are involved in the peripheral insulin resistance of the SDF rats.(ABSTRACT TRUNCATED AT 250 WORDS)

[Scientific education in the training of specialists from the European perspective]. / Wetenschappelijke vorming in de opleiding van de specialisten in Europees perspectief.

An overview is given on the scientific education of the Trainees (Specialists in Training in Europe). European legislation is clear and insists on the scientific formation in the undergraduate as well as in the Postgraduate education. It is the task of the national (or regional) authorities to create possibilities for research during clinical training. In most of the national training programmes Research activities are included. Moreover in the European visiting system Research facilities are an important item. Examples are shown how the Royal College of Obstetricians and Gynaecologists (U.K.) and the European Board and College of Obstetrics and Gynaecology have implemented these Research activities in the training programme.

Vitamin D and bone mineral homeostasis during pregnancy in the diabetic BB rat.

Vitamin D and bone mineral metabolism during pregnancy were studied in 17 diabetic and 13 control BB rats. On day 21 of pregnancy, reduced mean levels of 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3; 56.9 vs. 97.9 pg/ml; P less than 0.0001] and vitamin D-binding protein (304 vs. 482 micrograms/ml; P less than 0.0001) were found in the diabetic rats, while the free 1,25-(OH)2D3 concentration was not different from the control level. Total plasma calcium and total plasma protein concentrations were also significantly decreased in the diabetic group, but the calculated diffusible calcium was not significantly lower. Calcium and phosphorus urinary excretion were increased in the diabetic rats. There was no difference in bone mineral content. The fetuses of the diabetic BB rat had a lower body weight and were hypoinsulinemic. Both 1,25-(OH)2D3 (41.3 vs. 54.7 pg/ml; P less than 0.01) and vitamin D-binding protein (80 vs. 123 micrograms/ml; P less than 0.001) were decreased in the fetuses of diabetic rats, but the free 1,25-(OH)2D3 concentration was slightly but significantly (6.96 vs. 5.54; P less than 0.05) increased. We observed that the fetuses of diabetic rats had fewer ossification centers, counted with the Alizarin Red S staining method. The fetal ash weight was lower in the diabetic group (16.7 vs. 26.9 mg; P less than 0.0001). In addition, the relative calcium and phosphorus, but not magnesium, content of ash was lower in the fetuses of diabetic rats. This reduced mineral content in fetuses of diabetic mothers could be implicated in the pathogenesis of early neonatal hypocalcemia in infants of diabetic mothers.

Measurement of individual plasma angiotensins in normal pregnancy and pregnancy-induced hypertension.

Individual angiotensin peptides were measured by a high pressure liquid chromatography-RIA (HPLC-RIA) technique in the plasma of 20 nonpregnant women, 17 women with normal pregnancy, and 49 with pregnancy-induced hypertension. Immunoreactive angiotensin-II (ANG-II) consisted mainly of ANG-(1-8) octapeptide (greater than 65%), variable proportions (15-25%) of ANG-(4-8) pentapeptide, and small to negligible proportions of ANG-(2-8) heptapeptide and ANG-(3-8) hexapeptide. Levels of ANG-(1-8) were significantly higher in women with normal pregnancy than in both nonpregnant women (P less than 0.0006) and women with pregnancy-induced hypertension (P less than 0.008); in the latter, levels were lower with increasing severity of disease. Levels of ANG-(4-8) were higher in women with normal pregnancy than in women with pregnancy-induced hypertension or nonpregnant women. When expressed as a proportion of ANG-(1-8) levels, however, ANG-(4-8) levels were not higher in normal pregnancy than in women with pregnancy-induced hypertension or nonpregnant women. It is concluded that the well known increase in ANG-II levels in normal pregnancy relates predominantly to the active ANG-(1-8) octapeptide and to a far lesser extent to the smaller ANG peptides. Similarly, lower ANG-II levels in pregnancy-induced hypertension relate predominantly to lower ANG-(1-8) levels.

Fibrinolytic response to venous occlusion and fibrin fragment D-dimer levels in normal and complicated pregnancy.

The fibrinolytic response to venous occlusion was assessed in 29 women with normal or complicated pregnancy, by measurements of total t-PA and free t-PA with specific ELISAs. The release of t-PA from the vessel wall was 11 +/- 9 ng/ml in non-pregnant women (mean +/- SD, n = 6) but was markedly reduced throughout pregnancy. Following venous occlusion, free t-PA increased by 12 +/- 11 ng/ml in non-pregnant women but remained below the detection limit of 2 ng/ml towards the end of pregnancy. A markedly reduced t-PA release with absence of free t-PA was also observed during late pregnancy in patients with insulin-dependent diabetes mellitus, intra-uterine growth retardation and pre-eclampsia. Plasma levels of fragment D-dimer of cross-linked fibrin were measured with a specific ELISA in 79 pregnant women. D-dimer levels were 129 +/- 36 ng/ml (mean +/- SD, n = 8) in non-pregnant women and increased to 400 +/- 170 ng/ml (n = 25) and 440 +/- 220 ng/ml (n = 22) during the second and third trimester of pregnancy respectively. Significantly higher levels than observed in uncomplicated third trimester pregnancies were found in 3 out of 6 diabetic and in 2 out of 7 pre-eclamptic women. It is concluded that the t-PA release after venous occlusion is significantly reduced during pregnancy. In addition, released t-PA is rapidly inhibited. The levels of fragment D-dimer increase during pregnancy, suggesting that, notwithstanding the marked impairment of the fibrinolytic response to venous occlusion, the fibrinolytic system remains functionally active.

Severe systemic hypertension during pregnancy.

Severe hypertension during pregnancy remains an important cause of maternal mortality. Cerebral complications are frequently responsible for these deaths. Early detection of preeclampsia and preventive treatment are certainly goals for the future. In the management of severe pregnancy-induced hypertension, central issues are delivery and antihypertensive and anticonvulsive treatment. Cerebral disease consists mainly of bleeding and edema. Antihypertensive therapy can be dangerous and it is essential that hemodynamic control is provided.

Endocrine pancreas in the offspring of rats with experimentally induced diabetes.

At birth newborn rats from mothers with experimentally induced diabetes show hypertrophy and degranulation of the pancreatic islets. With birth the maternal hyperglycaemic stimulus is removed and during the lactation period the overstimulated B cells can restore their normal secretory activity. The increase of B-cell mass, however, remains retarded for several weeks. By adulthood the endocrine pancreas of offspring from mildly diabetic mothers seems to have recovered from the influence of the abnormal intra-uterine milieu, at least as judged by morphometric examination. In offspring from severely diabetic mothers an increased secretory activity of the individual B cells might be responsible for their sustained hypoglycaemia.

Rat foetal endocrine pancreas in experimental diabetes.

The endocrine pancreas of foetuses and newborn rats of experimental diabetic mothers showed morphological and ultrastructural changes. Islet hypertrophy and beta cell hyperplasia were constantly present, but the beta cells of foetuses of severely diabetic mothers were degranulated. The ultrastructural changes indicated hyperfunction in the beta cells of foetuses of experimental diabetic mothers. The morphological changes mentioned were similar to those seen in human diabetic pregnancy.

Absence of pregnancy-induced alterations in tissue insulin sensitivity in the offspring of diabetic rats.

We have previously demonstrated insulin resistance in the liver and peripheral tissues of the adult offspring of rats made diabetic with streptozotocin (SDF rats). In this study, a euglycaemic hyperinsulinaemic clamp was used to test the hypothesis that insulin resistance is further aggravated during pregnancy in SDF rats. Normal pregnancy was accompanied by a decrease in the sensitivity of the liver and peripheral tissues to insulin, with a normal responsiveness to insulin. In SDF rats no further decrease in the sensitivity of peripheral tissues to insulin occurred during pregnancy when compared with non-pregnant rats, and the dose-response curves of the glucose metabolic clearance rate during hyperinsulinaemia were similar in pregnant control and pregnant SDF rats. There was, however, a modest decrease in the sensitivity of the liver to insulin during pregnancy in SDF rats. The normal increase in plasma insulin levels during pregnancy was blunted in SDF rats: this resulted in increased glucose levels in maternal and fetal rats and increased fetal insulin concentrations, features compatible with mild 'gestation diabetes'. In conclusion, gestational diabetes develops in pregnant SDF rats, although there is no further deterioration in peripheral insulin resistance.

Morphological changes in the endocrine pancreas in pregnant rats with experimental diabetes.

This present study has demonstrated that during normal pregnancy in the rat the number of beta-cells is increased (hyperplasia) and the volume of the individual beta-cells is increased (hypertrophy). During experimental diabetes, however, the endocrine pancreas has an impaired capacity to compensate during pregnancy. In the experimental diabetic pregnant rat the beta-cells cannot replicate due to the unfavourable metabolic environment. This could reflect the complications caused by diabetes during human pregnancy.

Effects of treatment with progesterone and oestradiol-17 beta on the endocrine pancreas in ovariectomized rats: ultrastructural variations in the B cells.

The effects of progesterone and/or oestradiol treatment on the ultrastructural appearance of the pancreatic B cells has been studied in ovariectomized Wistar rats. A morphometric examination of the numberical density of dark and high granules in the B cells was therefore performed in each group of experimental rats as well as in control (olive oil-injected) rats. In the oestradiol-treated rats, and especially in the rats with combined oestradiol/progesterone treatment, the proportions of light and dark granules in the pancreatic B cells changed, compared with control values, in favour of the light granules. This increase in light granule content was comparable to changes in B cells during a pregnancy and it is suggested that the secretory activity of the B cells increases during pregnancy in a manner similar to that seen during oestradiol treatment.

Osteocalcin during the reproductive cycle in normal and diabetic rats.

Concentrations of osteocalcin were measured in plasma and bone of normal and diabetic rats during the reproductive cycle and compared with plasma 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) concentrations. The age-dependence of osteocalcin was also examined. Plasma concentrations of osteocalcin levels were low but detectable in 21-day-old fetuses (3.7 +/- 0.3 nmol/l); osteocalcin concentrations were highest in weaning rats (104 +/- 9 nmol/l) and decreased thereafter. In adult rats, plasma concentrations of both osteocalcin and 1,25-(OH)2D3 increased during the last days of normal pregnancy, and even more so in rats fed a diet low in calcium and phosphate. After an early post-partum decline, osteocalcin concentrations in plasma remained at non-pregnant levels in lactating rats fed a high calcium/phosphate diet while their 1,25-(OH)2D3 concentrations were higher than in non-pregnant rats; however, lactating rats fed a low calcium/phosphate diet showed increasing osteocalcin concentrations. In spontaneously diabetic BB rats, plasma osteocalcin concentrations were severely decreased compared with those in non-diabetic rats, more than would have been expected from their decreased 1,25-(OH)2D3 concentrations. Moreover, plasma osteocalcin did not increase during pregnancy or lactation in diabetic rats, even when fed a low calcium/phosphate diet. Fetuses of diabetic rats also had lower plasma osteocalcin levels than fetuses from non-diabetic rats or than weight-matched fetuses from semistarved rats. In contrast to plasma osteocalcin concentrations, bone osteocalcin concentrations and content were not altered by pregnancy, lactation, low calcium/phosphate diet or diabetes.(ABSTRACT TRUNCATED AT 250 WORDS)