The Cohen diabetic rat as a model for fetal growth restriction: vitamins C and E reduce fetal oxidative stress but do not restore normal growth.

UNLABELLED: Fetal growth restriction (FGR) describes newborns that were born small for gestational age. The etiology of FGR is unknown, but it is assumed that it is the consequences of both genetic and environmental factors, and that one of the important environmental factors is oxidative stress. In this study we used the Cohen diabetic (CD) rats (sensitive and resistant strains) and the original Sabra strain fed either high sucrose low copper diet-HSD or regular diet-RD to evaluate the genetic and environmental factors contributing to FGR. In addition, we treated the pregnant rats with antioxidants (vitamins C and E added to their food) to evaluate the effects of antioxidants in the prevention of FGR and in changing the redox state of the fetuses. METHODS: The study was performed on term 21-day-old fetuses of the three strains fed RD or HSD. Fetal and placental weight and fetal crown rump length were measured. Heart, kidneys, brain and liver were also weighted and studied. The fetal and placental redox status was investigated by studying the levels of Malondialdehyde (MDA) to determine the lipid peroxidation damage and by measuring the activity of catalase (CAT) and superoxide dismutase (SOD) enzymes. Similar studies were performed following the addition of 0.1% of vitamins C and E to the diet. RESULTS: FGR in the Cohen diabetic rats is a consequence of genetic (6-20% reduction in fetal weight in the CDr and CDs compared to Sabra) and environmental (11-36% reduction in fetal weight while on HSD) factors, with greater susceptibility in the CDs diabetic rats. Increased lipid peroxidation was observed in some of the organs only in HSD, however not only in the sensitive strain. In each organ, different patterns of anti oxidant capacity were observed. The addition of antioxidants to the food significantly reduced the signs of enhanced oxidative stress in all animals but partially restored normal fetal growth only in the diabetic CDs rats. This may imply that in this model oxidative stress is apparently not a major contributor to FGR. CONCLUSIONS: Cohen diabetic rats are a good model for the study of the interaction of genetic and environmental factors in the development of FGR. Maternal nutrition can influence the antioxidant capacity of the fetal organs which is modified by antioxidants. However, FGR in our model does not seem to result primarily from enhanced oxidative stress, as it is only partially affected by the antioxidant treatment. Thus, the repeated observations of oxidative stress in SGA infants may be a resulting metabolic alteration of FGR and not the main cause.

Beta-glycosphingolipids improve glucose intolerance and hepatic steatosis of the Cohen diabetic rat.

A link between altered levels of various gangliosides and the development of insulin resistance was described in transgenic mice. Naturally occurring glycosphingolipids were shown to exert immunomodulatory effects in a natural killer T (NKT) cell-dependent manner. This study examined whether glycosphingolipid-induced modulation of the immune system may reduce pancreatic and liver steatosis and stimulate insulin secretion in the Cohen diabetes-sensitive (CDS) rat, a lean model of non-insulin-resistant, nutritionally induced diabetes. Four groups of CDS rats fed a diabetogenic diet were treated with daily intraperitoneal injections of glycosphingolipids beta-glucosylceramide, beta-lactosylceramide, a combination of both (IGL), or vehicle (PBS) for up to 45 days. Immune modulation was assessed by fluorescence-activated cell sorting analysis of intrahepatic and intrasplenic lymphocytes. Steatosis was assessed by MRI imaging and histological examination of liver and pancreas, Blood glucose and plasma insulin concentrations were assessed during an oral glucose tolerance test. Administration of glycosphingolipids, particularly IGL, increased intrahepatic trapping of CD8 T and NKT lymphocytes. Pancreatic and liver histology were markedly improved and steatosis was reduced in all treated groups compared with vehicle-treated rats. Insulin secretion was restored after glycosphingolipid treatment, resulting in improved glucose tolerance. The immunomodulatory effect of beta-glycosphingolipids improved the beta-cell function of the hyperglycemic CDS rat. Thus our results suggest a role for the immune system in the pathogenesis of diabetes in this model.

Decreased response of osteoblasts obtained from aged Cohen diabetic sensitive rats to sex steroid hormones and 1,25OH2D3 in culture.

Patients with diabetes mellitus are known to develop osteopenia and osteoporosis, apparently as a reduction in the process of bone formation. In order to evaluate whether bone-modulating hormones--estradiol, testosterone, and 1,25(OH)(2)D(3)--have different effects on osteoblasts derived from diabetic and from normal non-diabetic rats, we studied the specific effects of these hormones on the differentiation and function of cultured osteoblasts derived from 1-year-old Cohen diabetic rats. (The Cohen diabetic model consists of a diabetic-sensitive strain [CDs; diabetic] and a diabetic-resistant strain [CDr; normal]). The CDs and CDr male and female rats were fed on a regular diet (RD) or a high-sucrose low-copper diet (HSD; diabetogenic). On the HSD diet, only CD rats develop type 2 diabetes, while CDr do not. Bones were removed for primary osteoblast cultures, and osteoblastic responses to the bone-modulation hormones--estradiol, testosterone, and 1,25(OH)(2)D(3)--were studied. In male rats fed RD, primary cultures of osteoblasts without hormone addition to the culture medium showed that alkaline phosphatase (ALP) activity was similar in the Cohen diabetic rats (both CDr and CDs) to that of the original Sabra strain. However, collagen synthesis was reduced in the CDr and CDs compared to the Sabra strain. The addition of the hormones to the culture medium did not change ALP activity or collagen synthesis in the male-derived osteoblasts, but increased mineralization in all strains. In female rats (studied only in CDs and CDr animals) there were no differences between animals fed the RD. HSD increased the basal activity of ALP in the CDr but not in the CDs rats, and decreased the rate of collagen synthesis in both CDr and CDs (diabetic) animals. The addition of the bone-modulation hormones to the culture medium further increased ALP activity in the osteoblasts derived from the CDr animals, while decreasing ALP activity in the CDs. These hormones also decreased collagen synthesis in both strains and increased mineralization in all osteoblasts. In conclusion, the metabolic status (HSD and diabetes) in rats prior to culture affected the phenotype of cultured osteoblasts, decreasing their response to bone-modulation hormones. This decreased response, especially to estradiol, may be a major cause of the osteopenia observed in diabetes.

Alterations in the expression of antioxidant genes and the levels of transcription factor NF-kappa B in relation to diabetic embryopathy in the Cohen diabetic rat model.

BACKGROUND: We have previously shown that oxidative stress is important in the pathogenesis of diabetes-induced anomalies in Cohen Diabetic sensitive (CDs) rat embryos and seems to interplay with genetic factors. We investigated the role of genetic factors related to the antioxidant defense mechanism in CDs rat embryos. METHODS: We studied 11.5- and 12.5-day embryos of Cohen Diabetic resistant (CDr) and CDs rats that were fed a regular diet (RD), and hence not diabetic, compared to rats fed a high-sucrose low-copper diet (HSD) where only the CDs animals became diabetic. Embryos were monitored for growth and congenital anomalies. mRNA of catalase (CAT), glutathione peroxidase (GSHpx), CuZn-SOD (SOD-superoxide dismutase), and Mn-SOD and the extent of nuclear factor kappa B (NF-kappaB) activation were assessed. RESULTS: Embryos of CDs dams fed RD were significantly smaller and had an increased rate of NTDs compared to embryos of CDr dams fed RD. When CDs dams were fed HSD, >50% of the CDs embryos were dead and 44% of the live embryos had NTDs. Live 11.5-day old embryos of CDs dams fed RD had a statistically significant increase in CAT, CuZn-SOD, and GSHpx mRNA levels compared with the levels in the CDr embryos from dams fed RD. CDs embryos from dams fed HSD showed significant overactivation of NF-kappaB compared with CDr embryos from dams fed HSD (in which activation was decreased), without any increase in the expression of SOD, CAT, and GSHpx. CONCLUSIONS: This study demonstrates that one of the genetic differences between the CDr and CDs strains fed RD is an increased expression of genes encoding for antioxidant enzymes in the CDs but inability for upregulation in diabetes. In addition, while activation of NF-kappaB is decreased in CDr on HSD, it is increased in the CDs. These differences may play a role in the increased sensitivity of the CDs embryos to diabetic-induced teratogenicity.

The role of reactive oxygen species in diabetes-induced anomalies in embryos of Cohen diabetic rats.

The role of the antioxidant defense mechanism in diabetes-induced anomalies was studied in the Cohen diabetes-sensitive (CDs) and -resistant (CDr) rats, a genetic model of nutritionally induced type 2 diabetes mellitus. Embryos, 12.5-day-old, of CDs and CDr rats fed regular diet (RD) or a diabetogenic high-sucrose diet (HSD) were monitored for growth retardation and congenital anomalies. Activity of superoxide dismutase (SOD) and catalase-like enzymes and levels of ascorbic acid (AA), uric acid (UA), and dehydroascorbic acid (DHAA) were measured in embryonic homogenates. When fed RD, CDs rats had a decreased rate of pregnancy, and an increased embryonic resorption. CDs embryos were smaller than CDr embryos; 46% were maldeveloped and 7% exhibited neural tube defects (NTDs). When fed HSD, rate of pregnancy was reduced, resorption rate was greatly increased (56%; P < .001), 47.6% of the embryos were retrieved without heart beats, and 27% exhibited NTD. In contrast, all the CDr embryos were normal when fed RD or HSD. Activity of SOD and catalase was not different in embryos of CDs and CDr rats fed RD. When fed HSD, levels of AA were significantly reduced, the ratio DHAA/AA was significantly increased, and SOD activity was not sufficiently increased when compared to embryos of CDr. The reduced fertility of the CDs rats, the growth retardation, and NTD seem to be genetically determined. Maternal hyperglycemia seems to result in environmentally induced embryonic oxidative stress, resulting in further embryonic damage.