Ying-King Wu

Dietary vitamin E prophylaxis and diabetic embryopathy: Morphologic and biochemical analysis ☆ ☆☆ ★ ★★

OBJECTIVE In this study we sought to determine whether dietary supplementation with vitamin E, a known antioxidant, would reduce the incidence of diabetic embryopathy in an in vivo rat model. STUDY DESIGN Eighty-day-old Sprague-Dawley rats were assigned to one of five groups: two control groups (groups 1 and 2) and three diabetic groups (groups 3, 4, and 5). One group of controls (group 2) and one group of diabetic rats (group 4) received dietary supplements of vitamin E (440 mg/day). The other three groups (groups 1, 3, and 5) received a normal diet only. Group 5 received insulin therapy to control glucose levels. On day 6 of gestation diabetes was induced in groups 3, 4, and 5 with streptozotocin (65 mg/kg). Animals were killed on day 12; embryos were examined for size, protein content, evidence of malformations, and superoxide dismutase activity. RESULTS In both groups (groups 3 and 4) of diabetic rats the mean blood glucose level than was significantly higher in controls. Insulin-treated animals (group 5) had glucose levels that were comparable to those of controls. The unsupplemented diabetic group had a neural tube defect rate of 21.48% +/- 9.6% (percentage of neural tube defects per rat) and a resorption rate of 21.37% +/- 20.39% (percentage of resorptions per rat) as compared with rates in the supplemented diabetic group of 6.92% +/- 4.08% and 2.17% +/- 3.74%, respectively (p < 0.01). Groups 1, 2, and 5 had similar neural tube defect rates (6.63% +/- 5.0%, 5.01% +/- 4.87%, and 3.55% +/- 5.92%, respectively. Vitamin E levels, measured by high-performance liquid chromatography, were significantly higher in maternal serum and embryos in the supplemented groups (p < 0.001) than in controls. Superoxide dismutase activity was reduced in the diabetes groups and was not affected by vitamin E therapy. CONCLUSIONS Supplementation with the antioxidant vitamin E confers a significant protective effect against diabetic embryopathy and may potentially serve as a dietary prophylaxis in the future. We postulate that this protective effect is mediated by a reduction in the oxidative load induced by hyperglycemia.

Prevention of diabetic embryopathy in offspring of diabetic rats with use of a cocktail of deficient substrates and an antioxidant

OBJECTIVE The current study sought to determine whether a dietary cocktail of deficient substrates and antioxidant could reduce the incidence of diabetic embryopathy to the background rate in an in vivo rat model. STUDY DESIGN Sprague-Dawley rats 70 to 90 days old were assigned to one of eight groups: two control groups (groups 1 and 2) and six diabetic groups (groups 3 to 8). One group of controls (group 2) received an oral supplemental cocktail of vitamin E ((alpha-tocopherol, 400 mg/day), safflower oil (arachidonic acid, 1 ml/day), and myo-inositol 0.08 mg per day. Four other diabetic groups (groups 5 to 8) received the cocktail in varying strengths. One diabetic group (group 3) received a normal diet only without insulin or cocktail, whereas another diabetic group (group 4) received insulin but no cocktail supplementation. On day 6 of gestation diabetes was induced in groups 3 to 8 with intravenous streptozotocin (65 mg/kg), and maternal glucose levels were monitored. Animals were killed on day 12; embryos were examined for size, somite number, and evidence of malformations and were biochemically evaluated for vitamin E and myo-inositol levels and superoxide dismutase activity. RESULTS The diabetic groups had a significantly higher (p < 0.05) mean blood glucose level than controls did. The insulin-treated group 4 had glucose levels that were comparable to those of controls. The unsupplemented diabetic group 3 had a neural tube defect rate of 23.7% versus 4.04% in controls and 3.55% in insulin-treated diabetics (p < 0.05). The rate of neural tube defects was significantly reduced to the background level in animals receiving half-strength cocktail or stronger doses (groups 6 to 8) compared with the diabetic unsupplemented controls (group 3). Diabetic animals in group 5 receiving only quarter-strength cocktail did not demonstrate a significant reduction in the malformation rate. Serum myo-inositol levels were not significantly different among the groups. However, serum levels of vitamin E were significantly higher in diabetics receiving half-strength cocktail than in nondiabetic controls, diabetics receiving no supplements, and diabetics receiving quarter-strength cocktail. Superoxide dismutase activity was also significantly increased in diabetic animals receiving supplementation versus animals not receiving the same, and the increases in vitamin E and superoxide dismutase were significantly correlated (r = 0.66, p < 0.05). CONCLUSION These data demonstrate that a cocktail containing deficient substrates and an antioxidant in varying strengths reduces the malformation rate to background in offspring of diabetic rats.

Dietary polyunsaturated fatty acid prevents malformations in offspring of diabetic rats.

OBJECTIVE The purpose of the current study was to determine whether a dietary source of arachidonic acid could serve as a pharmacologic prophylaxis to obviate the teratogenic effects of hyperglycemia. STUDY DESIGN Eighty-day-old Sprague-Dawley rats were mated, and after conception were randomly allocated to five groups: two groups were nondiabetic normal controls and three groups had diabetes experimentally induced with streptozocin. Of the two control groups, one was fed a normal diet (group 1) and the other group (group 2) received a normal diet and 1.0 ml of safflower oil, a polyunsaturated fatty acid known to increase serum arachidonic acid levels. In the three diabetic groups (groups 3, 4, and 5) glucose levels were allowed to remain > 350 mg/dl by withholding daily insulin therapy. Group 3 received a normal diet without supplementation; group 4 received a normal diet plus normal saline solution sham feedings, whereas group 5 received a normal diet supplemented with 1.0 ml of safflower oil. The oral agents (normal saline solution and polyunsaturated fatty acid) were administered with a tuberculin syringe. RESULTS Diabetic rats not receiving insulin therapy and receiving normal diets produced offspring with malformation rates of 20% compared with control rates of 4.8%. Supplemental normal saline solution or safflower oil given orally to controls did not alter the growth or malformation rates. These rates were similarly unaffected in the diabetic rats receiving oral supplementation of normal saline solution. However, with safflower oil supplementation to diabetic rats the incidence of neural tube defects was decreased from 20.0% to 7.6% (p < 0.0001). An inverse relationship was observed between the malformation rate and the serum arachidonic acid level: 17.83 (SD 5.84 micrograms/ml) in the nondiabetic controls, with a malformation rate of 4.8%, versus 14.18 (SD 2.58 micrograms/ml) in the diabetic rats, with a malformation rate of 20.0% (p < 0.05). With safflower oil supplementation serum levels of arachidonic increased from 14.18 +/- 2.58 micrograms/ml to 19.99 +/- 7.99 micrograms/ml (p < 0.05); this was associated with a concomitant decline in the malformation rate. CONCLUSION These data demonstrate that diabetic embryopathy is associated with a deficiency state in essential fatty acid, corroborating our previous in vitro findings. Furthermore, the use of a dietary polyunsaturated fatty acid that specifically increases arachidonic levels significantly reduced the incidence of diabetic embryopathy. These findings may serve as a basis for developing strategies of pharmacologic prophylaxis against diabetes-induced congenital malformations.

Dietary intake of myo-inositol and neural tube defects in offspring of diabetic rats

OBJECTIVE Embryopathy in diabetic mothers occurs at a rate four to five times higher than that observed in the general population. The current investigation was undertaken to assess the use of dietary myo-inositol supplementation as a pharmacologic prophylaxis to obviate the teratogenic effects of hyperglycemia in an in vivo study. STUDY DESIGN Seventy Sprague-Dawley rats were mated and after conception were randomly divided into five groups: one group was nondiabetic normal controls and four groups had diabetes experimentally induced with streptozotocin. Of the diabetic groups, one received the usual diet, whereas the others received, respectively, 0.08, 0.16, and 0.5 mg/day supplemental myo-inositol orally. RESULTS With the myo-inositol supplementation (0.08 mg/day), the incidence of neural tube defects was significantly reduced from 20.4% to 9.5% (p < 0.01). The most effective dosage of myo-inositol was 0.08 mg/day. Increasing the dose of myo-inositol beyond that level did not significantly reduce the rate of neural tube defects. However, the resorption rate was increased to 29.8%. CONCLUSION These data demonstrate that myo-inositol supplementation reduces the incidence of diabetic embryopathy and may serve as a pharmacologic prophylaxis against diabetes-induced congenital malformations.

The Role of Free Radicals and Membrane Lipids in Diabetes-Induced Congenital Malformations

Objective: The incidence of major congenital malformations is approximately 6-9% in pregnancies complicated by diabetes mellitus. This incidence is 3-4-fold higher than that in the general population. Congenital malformations are now ranked as the leading cause of death in the offspring of women with diabetes. The precise mechanism(s) by which these anomalies are induced is unknown. It is also not clear what predisposes women to deliver malformed infants, which infants are at risk, and why some are spared even when exposed to presumably high risk conditions. The purpose of this report is to determine, from the literature, the primary etiologic factors associated with diabetes-induced embryopathy and its prevention. Methods: A review of the current literature regarding congenital malformations in diabetic pregnancies was conducted to elucidate dominant concepts in the pathogenic mechanism(s) of these anomalies and to discuss current and future strategies for their prevention. Results: Numerous investigators have demonstrated that hyperglycemia has a teratogenic effect during organogenesis. However, the exact mechanisms involved have not been completely elucidated. Dietary supplementation of deficient substrates (arachidonic acid or myo-inositol), either in vitro or in vivo, has been shown to reduce the incidence of diabetes-related malformations in offspring of diabetic pregnant animals. In addition, free oxygen radical-scavenging enzymes and antioxidants aimed at reducing the excess load of radicals also result in a reduced malformation rate. Clinical evidence has demonstrated that the teratogenic effects of hyperglycemia may be obviated by maintaining euglycemia throughout organogenesis. Numerous studies have demonstrated that participation in a preconception care program can reduce the incidence of malformations in women with diabetes to the background rate. Unfortunately, less than 10% of women with diabetes currently enter these programs. Conclusions: Diabetic embryopathy remains the single most common lethal problem affecting diabetic pregnancies today. Although preconception planning and glycemic control can reduce the incidence of malformations, it is often difficult to get women to attend such programsand to achieve and maintain euglycemia. The use of dietary supplements, which presumably would override the teratogenic effects of aberrant metabolic fuels, holds great promise for the future as a prophylaxis against diabetic embryopathy.

Review Article : The Yolk Sac Theory Closing the Circle on Why Diabetes-Associated Malformations Occur

The purpose of this article is to examine the role of yolk sac failure during organogenesis in the development of diabetes-associated embryopathy. The current literature regarding congenital malformations in diabetic pregnancies was reviewed to elucidate the precise role of the yolk sac in embryonic development and the relation between yolk sac injury and embryopathy. We and others have demonstrated that hyperglycemia produces a teratogenic effect during organogenesis. In addition, we have shown that the yolk sac appears to be the target site of injury induced by hyperglycemia. We have also presented evidence that cell membrane dysfunction leads to failed vitelline vessel formation and that arachidonic acid supplementation prevents many of the morphologic and biochemical alterations observed under hyperglycemic conditions. These data strongly support the teratogenic effect of hyperglycemia, the arachidonic acid deficiency state, the resultant maldevelopment of vitelline vessels, and the ability to prevent these changes by arachidonic acid supplementation. These studies have made significant inroads in explaining why diabetes-associated anomaties occur, and suggest a potential future role for prophylaxis against these organogenetic malformations using dietary polyunsaturated fatty acid supplementation. (J Soc Gynecol Invest 1994;1:3–13)OBJECTIVE: The purpose of this article is to examine the role of yolk sac failure during organogenesis in the development of diabetes-associated embryopathy. METHODS: The current literature regarding congenital malformations in diabetic pregnancies was reviewed to elucidate the precise role of the yolk sac in embryonic development and the relation between yolk sac injury and embryopathy. RESULTS: We and others have demonstrated that hyperglycemia produces a teratogenic effect during organogenesis. In addition, we have shown that the yolk sac appears to be the target site of injury induced by hyperglycemia. We have also presented evidence that cell membrane dysfunc tion leads to failed vitelline vessel formation and that arachidonic acid supplementation prevents many of the morphologic and biochemical alterations observed under hyperglycemic conditions. CONCLUSIONS: These data strongly support the teratogenic effect of hyperglycemia, the arachi donic acid deficiency state, the resultant maldevelopment of vitelline vessels, and the ability to prevent these changes by arachidonic acid supplementation. These studies have made significant inroads in explaining why diabetes-associated anomalies occur, and suggest a potential future role for prophylaxis against these organogenetic malformations using dietary polyunsaturated fatty acid supplementation. (J Soc Gynecol Invest 1994;1:3-13)

Demonstration of the Essential Role of Protein Kinase C Isoforms in Hyperglycemia-Induced Embryonic Malformations

To address the role of PKC isoforms in hyperglycemia-induced apoptosis and malformations in the embryos of diabetic pregnancies, expression of PKCα, β1, β 2, γ, δ, ε, and ζ was examined in the neural tube of rat embryos and showed to overlap with the regions of increased apoptosis. Levels of activated (phosphorylated) PKCα , β2, and δ were increased in the embryos of diabetic dams whereas those of PKCε and ζ were decreased when compared with those in control groups. Cytosolic phospholipase A2 (cPLA2) was also activated. Blocking the activity of PKCα , β2, and δ using isoform-specific inhibitors in embryos cultured in hyperglycemia (40 mM) reduced malformation rates when compared with those in untreated hyperglycemic and euglycemic (8.3 mM) groups. These observations demonstrate that PKCα, β2, and δ play an essential role in diabetic embryopathy. Activation of cPLA2 was also decreased, suggesting that PKCs mediate the hyperglycemic effects through the cPLA2-phospholipid peroxidation pathway.