Distribution of androstenedione and its effects on total free fatty acids in pregnant rats.

Androstenedione, an anabolic steroid used to enhance athletic performance, was administered in corn oil by gastric intubation once daily in the morning to nonpregnant female rats at a dose of 5 or 60 mg/kg/day, beginning two weeks before mating and continuing through gestation day (GD) 19. On GD 20, the distribution of androstenedione and other steroid metabolites was investigated in the maternal plasma and target organs, including brain and liver. The concentration of estradiol in plasma approached a statistically significant increase after treatment as compared with the controls, whereas the levels of androstenedione, testosterone and progesterone were not significantly different from the controls. In the liver, the concentrations of androstenedione and estradiol only were increased in a dose-related manner. None of these steroids was detectable in the brain. Androstenedione treatment also produced changes in the level of selected free fatty acids (FFAs) in the maternal blood, brain, liver and fetal brain. The concentrations of palmitic acid (16:0) and stearic acid (18:0) in the plasma were not significantly different between the controls and treated rats. However, oleic acid (18:1), linoleic acid (18:2) and docosahexaenoic acid (DHA, 22:6) were 17.94 +/- 2.06 microg/ml, 24.23 +/- 2.42 microg/ml and 4.08 +/- 0.53 microg/ml, respectively, in the controls, and none of these fatty acids was detectable in the treated plasma. On the other hand, palmitic, stearic, oleic, linoleic and DHA were present in both control and treated livers. Among the FFAs in liver, linoleic and DHA were increased 87% and 169%, respectively, over controls. Palmitic, stearic and oleic acids were not significantly affected by the 60 mg/kg treatment. These were present in both control maternal and fetal brains, whereas linoleic acid was found only in fetal brain control. DHA was present only in the control maternal brain (0.02 +/- 0.02 microg/mg protein) and fetal brain (0.24 +/- 0.15 microg/mg protein). The results indicated that androstenedione exhibits significantly different effects on the FFA composition among target organs during pregnancy.

Effects of oral androstenedione on phospholipid fatty acids, ATP, caspase-3, prostaglandin E(2) and C-reactive protein in serum and livers of pregnant and non-pregnant female rats.

Androstenedione, a steroidal dietary supplement taken to enhance athletic performance, could affect serum and liver lipid metabolism, induce liver toxicity or alter inflammatory response depending on dose and duration of exposure. Pregnancy could further exaggerate these effects. To examine this, mature female rats were gavaged with 0, 5, 30 or 60 mg/kg/day androstenedione beginning two weeks prior to mating and continuing through gestation day 19. Non-pregnant female rats were gavaged over the same time frame with 0 or 60 mg/kg/day androstenedione. Serum was collected and livers were removed from dams on gestation day 20 and from non-pregnant rats after 5 weeks of treatment. Androstenedione had no effect on serum total cholesterol, triglycerides or HDL-cholesterol, but significantly decreased C-reactive protein in pregnant rats and prostaglandin E(2) in serum of both pregnant and non-pregnant rats. There were treatment related decreases in liver ATP and, to a lesser degree, caspase-3 and no change in alkaline phosphatase of pregnant female rats. Androstenedione decreased docosahexaenoic acid in both serum and liver phospholipids of pregnant female rats. In conclusion, oral androstenedione did not result in overt hepatotoxicity in pregnant female rats, but produced modest changes in lipid metabolism and may impair regeneration of injured hepatic cells or tissue.

Effects of oral androstenedione on steroid metabolism in liver of pregnant and non-pregnant female rats.

It is unknown whether androstenedione, a steroidal dietary supplement taken to enhance athletic performance, can affect physiological hormone levels by altering liver enzyme activities that metabolize steroid hormones. Altered hormone levels could be especially devastating during pregnancy. Mature female rats were gavaged with 0, 5, 30 or 60 mg/kg/day androstenedione beginning two weeks prior to mating and continuing through gestation day 19. Non-pregnant female rats were gavaged over the same time frame with 0 or 60 mg/kg/day androstenedione. Livers were removed from dams on gestation day 20 and from non-pregnant rats after five weeks' treatment. Liver microsomes were incubated with 200 microM testosterone, and the reaction products were isolated and analyzed by HPLC. In pregnant rats, formation of 6alpha-, 15beta-, 7alpha-, 16beta-, and 2beta-hydroxytestosterone was increased significantly vs. control at the highest dose level only. Formation of 6beta-hydroxytestosterone increased significantly at both the 30 and 60 mg/kg/day dose levels. In non-pregnant rats, 60 mg/kg/day androstenedione significantly increased formation of 15beta-, 6beta-, 16beta-, and 2beta-hydroxytestosterone. The data suggest that high oral doses of androstenedione can induce some female rat liver cytochromes P450 that metabolize steroid hormones and that the response to androstenedione does not differ between pregnant and non-pregnant female rats.

Developmental effects of serum from flaxseed-fed rats on cultured rat embryos.

Gestation day 9.5 rat embryos were cultured for 45 h in serum obtained from pregnant rats that had been fed throughout gestation with either a control diet (based on the AIN-93 formulation), a diet supplemented with flaxseed (20% or 40%, w/w), or a diet supplemented with de-fatted flaxseed ("flaxseed meal", 13 or 26%, w/w). The embryos were fixed in neutral formalin at the end of culture. Overall growth and development was assessed, and the presence of abnormalities was noted. A significant inhibition of growth (as determined by crown-rump length) relative to control was observed in embryos cultured in serum from rats fed the 20% flaxseed diet. The incidence of spontaneous heart inversions was increased significantly in the embryos cultured in serum from the 20% flaxseed and 26% flaxseed meal fed rats. The incidence of flexion defects was increased significantly in embryos cultured in serum from 20% flaxseed-fed rats. The lack of an apparent dose response in any of the statistically significant effects suggests that the observed anomalies were chance occurrences unrelated to the treatment group from which serum was obtained. It is therefore concluded that diets high in flaxseed or flaxseed meal do not result in serum factors that are directly embryotoxic to organogenesis-staged rat embryos. This finding is consistent with the findings of a parallel in vivo rat teratology study where no significant embryotoxicity attributable to flaxseed exposure was observed.

Flaxseed increased alpha-linolenic and eicosapentaenoic acid and decreased arachidonic acid in serum and tissues of rat dams and offspring.

The effects of dietary flaxseed (FS), and defatted flaxseed meal (FLM) on serum and tissue fatty acid profiles were investigated. Pregnant Sprague-Dawley rats were fed AIN-93 based diets balanced in calories, fat, nitrogen, and fiber. Diets contained 0, 20%, 40% FS or 13% or 26% FLM by weight. The control, FS and FLM diets differed in linoleic acid to alpha-linolenic acid (ALA) fatty acid ratio. These diets were fed continuously during gestation, suckling period and 8 weeks post-weaning (F(1)). FS fatty acids were bioavailable and metabolized by pregnant and F(1) rats. ALA and eicosapentaenoic acid increased; linoleic and arachidonic acid decreased; and docosahexaeonic acid was unchanged in serum, 'gastric milk' and liver of FS and FLM-fed pregnant and F(1) rats. FS more than FLM, changed fatty acids profiles, but FLM and 40% FS significantly reduced serum cholesterol. Dietary 40% FS may have increased oxidative stress as evidenced by a reduction in liver vitamin E.

Impact of high flaxseed diet on mitogen-induced proliferation, IL-2 production, cell subsets and fatty acid composition of spleen cells from pregnant and F1 generation Sprague-Dawley rats.

Flaxseed (FS) being rich in alpha-linolenic acid may alter the immune parameters. Therefore, we assessed the impact of FS and defatted flaxseed meal (FLM) on fatty acid composition, cell subsets, proliferation and IL-2 production by splenic lymphocytes. Pregnant female Sprague-Dawley rats were fed diets containing 0% FS and FLM, 20 or 40% FS, 13 or 26% FLM during gestation or gestation, lactation and 8 week post-weaning period. FS and FLM resulted in up to 8.3 fold and 4.6 fold increase in splenic ALA among pregnant rats, 4.5 fold and 1.2 fold increase in splenic ALA among F(1) generation rats. Splenic linoleic acid (LA) and arachidonic acid (AA) were 18 and 40% lower in 40% FS fed pregnant rats, and AA was 15% lower in all the other groups. Among F(1) rats, splenic LA and AA were 16 and 48% lower in 40% FS group, and AA was 18% lower in 20% FS and 26% FLM groups. Concanavalin A and phytohemagglutinin mediated proliferation of spleen cells were 60 and 52% lower in 40% FS fed pregnant and F(1) generation rats, respectively. No significant changes were observed in the cell subsets or IL-2 production by splenic cells from different groups.

Effect of long-chain fatty acids in the culture medium on fatty acid composition of WEHI-3 and J774A.1 cells.

As a first step in determining the mechanism of action of specific fatty acids on immunological function of macrophages, a comparative study of the effect of long-chain polyunsaturated fatty acids (PUFA) in the medium was conducted in two macrophage cell lines, J774A.1 and WEHI-3. The baseline fatty-acid profiles of the two cell lines differed in the % distribution of saturated (SFA) and unsaturated fatty acids (UFA). J774A.1 cells had a higher % of SFA (primarily palmitic acid) than WEHI-3 cells. Conversely, WEHI-3 cells had a higher % of UFA (primarily oleic acid) than J774A.1 cells. Neither cell line had detectable amounts of alpha-linolenic acid (ALA) or eicosapentaenoic acid (EPA). The most abundant polyunsaturated fatty acid in both cells lines was arachidonic acid (AA). The efficiency of transport of fatty acids from the medium to the macrophages by two delivery vehicles (BSA complexes and ethanolic suspensions) was compared. Overall, fatty acids were transported satisfactorily by both delivery systems. Alpha-linolenic acid and doscosahexenoic acid (DHA) were transported more efficiently by the ethanolic suspension system. Linoleic acid (LA) was taken up more completely than ALA, and DHA was taken up more completely than EPA by both cell cultures and delivery systems. A dose-response effect was demonstrated for LA, ALA, EPA and DHA in both J774A.1 and WEHI-3 cells. Addition of polyunsaturated fatty acids (PUFA) to the cell cultures modified the total lipid fatty acid composition of the cells. The presence of ALA in the culture medium resulted in a significant decrease in AA in both cell lines. The omega-3/omega-6 fatty acid ratio (omega-3/omega-6), polyunsaturated/saturated fatty acid ratio (P/S), and unsaturation index (UI) increased directly with the amount of PUFA and omega-3 fatty acid provided in the medium. The results indicate that the macrophage cell lines have similar, but not identical, fatty acid profiles that may be the result of differences in fatty acid metabolism. These distinctions could in turn produce differences in immunological function. The ethanol fatty-acid delivery system, when compared with the fatty acid-BSA complex system, is preferable for measurement of dose-response effects, because the cellular fatty acid content increased in proportion to the amount of fatty acid provided in the medium. Similar dose-response results were observed in a previous in vivo study using flaxseed, rich in ALA, as a source of PUFA.

Testing the potential of flaxseed to affect spermatogenesis: morphometry.

Quantitative information was collected on male reproductive effects of maternal and postnatal dietary exposure to flaxseed (20 or 40%), flaxmeal (13 or 26%) or standard NIH AIN-93 feed (0% flaxseed control). Measurements were made on the testes of F1 generation males rats (1) whose mothers were exposed to the diets designated above, and (2) who, after weaning, were placed on the same diet as their mothers for an additional 70 days. The seminiferous tubules comprised 86%, 84%, 84%, 84% and 85% of the total testis volume while the interstitial space comprised 12%, 14%, 14%, 14%, 13% of the total testis volume for the 0% flaxseed/flaxmeal, 20% flaxseed, 13% flaxmeal, 40% flaxseed and 26% flaxmeal groups, respectively. Statistically significant decreases in the absolute volume of the seminiferous tubules were observed in the 20% and 40% flaxseed-treated groups when these groups were compared to controls. Borderline statistically significant differences were also observed when Sertoli cell nucleolar number per tubular cross-section were compared in the 13% flaxmeal and 20% flaxseed treatment groups. These effects were not considered biologically significant because other parameters of male reproductive function appeared normal. Overall, the quantitative information obtained suggests that exposure to flaxseed/flaxmeal at the doses used in the present study does not adversely affect testis structure or spermatogenesis in the rat.

Nutritional and hematological impact of dietary flaxseed and defatted flaxseed meal in rats.

An 8-week study was conducted to determine the impact of dietary ground flaxseed (FS) or defatted flaxseed meal (FLM) on plasma lipids, minerals, hematological parameters and vitamin E status of weanling female Sprague-Dawley rats. These rats were fed isocaloric modified AIN-76 diets supplemented with 0.0, 5.0, 10.0% (w/w) FS or 6.2% (w/w) FLM for 56 days. Total and HDL cholesterol were not influenced by any of the dietary treatments. Plasma triglyceride was significantly increased by FLM, but not affected by FS. Total RBC counts and hematocrit were significantly higher in FS groups than in the control group; however, hemoglobin was not affected by FS. Dietary FLM had no effect on any of the above hematological parameters. Plasma alkaline phosphatase, an indicator of Zn status and a marker of bone formation, was significantly lower in the FS and FLM groups than in the control group. Plasma vitamin E content was not influenced by dietary treatment. Liver vitamin E was significantly higher in groups fed 10% FS and 6.2% FLM. In summary, moderate amounts of dietary FS may have the potential to increase liver vitamin E level and improve iron status. However, FS/FLM consumption may have a negative effect on zinc status, as indicated by decreased alkaline phosphatase levels.

A potential mechanism for fumonisin B(1)-mediated hepatocarcinogenesis: cyclin D1 stabilization associated with activation of Akt and inhibition of GSK-3beta activity.

Fumonisin B(1) (FB(1)) is a worldwide corn contaminant and has been epidemiologically linked to the high incidence of human esophageal cancer in South Africa and China. FB(1) is hepatocarcinogenic in rats by an unknown mechanism. Inhibition of ceramide synthase and disruption of membrane phospholipids have been shown to be mechanisms of toxicity. Here we show overexpression of cyclin D1 protein in both preneoplastic and neoplastic liver specimens obtained from a long-term feeding study of FB(1) in rats. In rats fed FB(1) short-term, cyclin D1 protein levels in liver were increased up to five-fold in a dose-responsive manner. Northern blot analysis demonstrated no increase in mRNA levels of cyclin D1. 2D electrophoresis of cyclin D1 protein in FB(1)-treated samples showed a distinct pattern of migration (presence of less negatively charged form of the protein) that differed from controls. Recently, it has been shown that phosphorylation of cyclin D1 by glycogen synthase kinase 3beta (GSK-3beta) on a single threonine residue (Thr-286) positively regulates proteosomal degradation of cyclin D1. In FB(1)-treated samples we detected GSK-3beta phosphorylated on serine 9; activated protein kinase B (Akt) appears to be responsible for this activity-inhibiting phosphorylation. These findings suggest that overexpression of cyclin D1 results from stabilization due to a lack of phosphorylation mediated by GSK-3beta. We also observed an increase in cyclin dependent kinase 4 (Cdk4) complexes with cyclin D1 in FB(1)-treated samples; additionally, elevated Cdk4 activity was shown by increased phosphorylation of the retinoblastoma protein. In summary, the activation of Akt leads to increased survival, inhibition of GSK-3beta activity and post-translational stabilization of cyclin D1, all events responsible for disruption of the cell cycle G(1)/S restriction point in hepatocytes. This is the first report suggesting the mechanism by which FB(1) acts as a carcinogen.

The effect of maternal exposure to flaxseed on spermatogenesis in F(1) generation rats.

Pregnant Sprague-Dawley rats were exposed to a flaxseed (20 or 40%), flaxmeal (13 or 26%) or standard NIH AIN-93 (0% flaxseed control) diet throughout gestation and until their offspring were weaned. After weaning, F(1) generation males were placed in the same diet treatment groups as their mothers for 70 days. Statistically significant differences were not observed between either low-dose or high-dose flaxseed and flaxmeal-treated animals and the 0% flaxseed control animals for testis weights, homogenization resistant spermatid counts, daily sperm production rates, epididymal weights, seminal vesicle weights, seminiferous tubule fluid testosterone concentrations and the percentage of sperm abnormalities. The following statistically significant differences were observed when treated groups and the 0% flaxseed control groups were compared: (1) increases in serum LH in the 20% and 40% flaxseed treatment groups and in serum LH and testosterone in the 26% flaxmeal treatment group; (2) increases in the cauda epididymal weight from the 20% and 40% flaxseed groups; (3) increases in cauda epididymal sperm numbers/g epididymis from the 20% and 40% flaxseed and the 13% and 26% flaxmeal treatment groups; (4) a decrease in prostatic weight from the 20% flaxseed and 13% and 26% flaxmeal treatment groups. Prostate weight in the 40% flaxseed treatment group was lower but not statistically significantly different than the 0% flaxseed control group. Histological effects on spermatogenesis were not observed in either the control group, flaxmeal or the flaxseed treated groups.

Effect of dietary rosemary extract on cell-mediated immunity of young rats.

The impact of rosemary extract on splenic mononuclear cell proliferation was determined. Weanling male Sprague-Dawley rats were fed diets containing 0, 100, 200 or 400 ppm rosemary extract or 400 ppm butylated hydroxytoluene (BHT) in combination with 10 or 20% casein enriched diets for 8 weeks. Splenic mononuclear cells were isolated from these animals and mitogenic response to Concanavalin A (Con A), Phytohemagglutinin (PHA) and lipopolysaccharide was determined. Con A and PHA-stimulated proliferation of spleen cells from rats fed 10% casein and 200 ppm rosemary extract was significantly higher than that of cells from the corresponding control animals. However, other levels of rosemary at 10% dietary casein or rosemary at any concentration fed along with 20% dietary casein had no impact on the mitogenic stimulation of splenic mononuclear cells. Thus, these results suggest that the use of rosemary might not have a generalized immunoenhancing effect, and will probably be effective in some stressed conditions, such as protein or antioxidant deficiency.

Effect of dietary flaxseed on fatty acid composition, superoxide, nitric oxide generation and antilisterial activity of peritoneal macrophages from female Sprague-Dawley rats.

The impact of ground flaxseed (FS) or flaxseed meal (FSM) diets on the fatty acid composition and functions of rat peritoneal exudate cells (PEC) was determined. Female weanling Sprague-Dawley rats (10/group) were fed isocaloric AIN-76 diets supplemented with 0.0, 10.0% (w/w) FS or 6.2% (w/w) FSM. At the end of 56-days, rat serum and thioglycollate-elicited PEC were analyzed for total lipid fatty acids. Production of nitric oxide (NO) and superoxide (O2-), Listeria monocytogenes (LM) phagocytic index and antilisterial activity of resident PEC were also assessed. A significant increase in alpha-linolenic (C18:3), eicosapentanoic (C20:5) and docosahexanoic (C22:6) acids, as well as a significant reduction in arachidonic acid (C20:4) was observed in the serum of rats fed 10% FS. Dietary FS caused a significant reduction in palmitic acid (C16:0) and an increase in stearic acid (C18:0) of PEC. Defatted FSM produced a significant increase in long chain fatty acids, which included eicosadienoic acid (C20:2) in PEC and C22:6 in serum. PEC from rats fed 10.0% FS produced significantly less (about 50%) O2- in response to phorbol myristate acetate (PMA), than did PEC from control animals; dietary treatment had no effect on O2- in response to LM. FSM had no impact on the O2- production by PEC in response to PMA or LM. Antilisterial activity of PEC was determined by comparing bacterial uptake after 1 hr with recovery 24 hrs later. Despite comparably equivalent bacterial uptake, few viable intracellular LM were recovered at T = 24 for all test samples, indicating that, regardless of the dietary treatment, PEC were able to handle the in vitro LM infection. This bacterial clearance was accompanied by equivalent NO generation by PEC from each dietary group in response to LM. Summarily, dietary FS produced significant changes in fatty acid composition of serum and PEC, inhibited O2- generation by PEC, and was ineffectual to both NO production by and antilisterial activity of PEC.

Interactions in indices of vitamin A, zinc and copper status when these nutrients are fed to rats at adequate and increased levels.

The purpose of the present study was to determine the effects of feeding nutritionally adequate and increased levels of vitamin A (retinyl acetate at 1.4, 34.4, and 206.4 mg/kg diet) in combination with adequate or increased Zn (12 and 240 mg/kg) and Cu (5 and 50 mg/kg) on serum and tissue concentrations of retinol and retinyl palmitate and on indices of Cu and Zn status in female Sprague-Dawley rats, and to measure interactive effects of such nutrient imbalances. Rats fed on diets containing 34.4 and 206.4 mg vitamin A/kg had higher feed intakes and relative liver weights than those fed on diets containing 1.4 mg vitamin A/kg. An interaction between dietary Cu and Zn and an independent effect of vitamin A affected serum ceruloplasmin oxidase (EC 1.16.3.1) activity. Rats fed on high Zn, adequate-Cu diets (240 and 5 mg Zn and Cu/kg respectively) had lower serum ceruloplasmin oxidase levels than rats fed on adequate-Zn, adequate-Cu diets (12 and 5 mg Zn and Cu/kg respectively). This effect was not observed in rats fed on high-Zn, high-Cu diets (240 and 50 mg Zn and Cu/kg respectively). Alterations in dietary levels of Cu and vitamin A independently affected haemoglobin levels. Serum cholesterol concentration was affected by interactions between Zn and vitamin A and Cu and vitamin A. Levels of retinol and retinyl palmitate in liver and kidney were significantly higher in rats fed on diets with increased dietary vitamin A than in those fed on diets with adequate vitamin A. Three-way interactions among Cu, Zn, and vitamin A affected levels of retinol in serum and liver. Two-way interactions between Cu and vitamin A affected liver retinyl palmitate and the sum of liver retinol+retinyl palmitate. An independent effect of dietary Zn on these variables was also observed. Interactions between Cu and vitamin A affected levels of Cu in liver and kidney, while Fe and Zn in kidney were affected by interactions between Cu and Zn. This study demonstrates that differing interactions among variables of vitamin A metabolism and mineral status occur with higher dietary levels of vitamin A, Zn and Cu in the rat.

L-Arginine inhibits in vitro nonenzymatic glycation and advanced glycosylated end product formation of human serum albumin.

L-Arginine (Arg) has a structure similar to that of aminoguanidine (AG) and may inhibit glycation and advanced glycosylated end product (AGE) formation. Human serum albumin (HSA) (100mg/ml) was incubated for 2 weeks with glucose (200mM) at 37°C or with glucose and equimolar concentrations of Arg, N-α-acetyl Arg, or AG with or without 25mM diethylenetriaminepentaacetic acid (DTPA). In the absence of DTPA, electrospray ionization mass spectrometry showed a 70% reduction of covalently bound glucose in the presence of Arg and a 30% reduction with AG. Digestibility by trypsin of HSA incubated with glucose and Arg was similar to that of HSA incubated alone. This suggests less covalent modification of HSA in the presence of Arg as compared with the absence of Arg. When incubations contained DTPA, autoradiography showed less(14)C labeling of HSA subunits in the presence of Arg and AG. When theα-amino group of Arg was blocked with an acetyl group, labeling was similar to that of HSA incubated with glucose, suggesting involvement of theα-amino group in the inhibition. Fluorescence of HSA at ex370 and em440 was reduced with Arg, but AG was more effective than Arg. These results suggest that Arg, like AG, can inhibit glycation and AGE formation.

Nutrition labeling: energy values of foods and fat substitutes.

The Federal Food, Drug, and Cosmetic Act requires the listing of all food ingredients on the label. However, until this year, nutrition labeling for most foods was not required. The Nutrition Labeling and Education Act, passed by the US Congress in November 1990, requires nutrition labeling (on the package or at the site of purchase) of virtually all foods packaged after May 8, 1994. (A law was passed by Congress and signed by the President of the United States on May 26, 1994, that delayed applicability of the Nutrition Labeling and Education Act until after August 8, 1994, for certain products whose labels were printed before May 8, 1994, and for which there was supporting documentation that after August 8, 1994, the product labels would be in compliance.) Prominent among the nutrition information required is labeling of the total energy content of food and the energy content derived from fat. In Title 21, Code of Federal Regulations, five methods are specified for determining the energy content of foods. The US Food and Drug Administration expects also to include specific food factors (digestibility coefficients) in food additive and GRAS (generally recognized as safe) listings for calculating the energy values of novel foods and ingredients.

Gender differences in adrenal cortex steroid production in SHR/N-corpulent rats.

Gender differences in adrenal steroid hormone production and serum steroid hormone levels were compared in the spontaneous hypertensive/NIH-corpulent (cp) rat, which exhibits characteristics of both obesity and non-insulin-dependent diabetes mellitus. The study demonstrated that adrenal gland size correlated with adrenal production and serum levels of steroid hormones. Obese female SHR/N-cp rats were more steroidogenic than male SHR/N-cp rats; the size of their adrenal glands was twice that of the males (70 vs 33 mg). Body weights averaged 666 g for females and 829 g for males. Obese female rats had significantly higher serum concentrations of both corticosterone (827 vs 536 ng/ml) and aldosterone (675 vs 482 pg/ml) than obese male rats. As determined by in vitro assay, adrenal cortex production of corticosterone (2157 vs 1435 ng/30 min) and aldosterone (13.3 vs 9.5 ng/30 min) was significantly higher in obese female than in obese male rats. Adrenal production of testosterone in the in vitro assay was also significantly higher for obese female than male rats; however, adrenal estrogen production in obese rats did not differ significantly. The type of carbohydrate consumed (sucrose > starch) significantly affected serum levels of corticosterone, but not aldosterone, testosterone, or estrogen. Gender differences in adrenal steroid production and serum steroid levels suggest that hyperglycemia in obese SHR/N-cp rats may be, in part, the result of excess adrenal production of steroid hormones.

Effect of dietary carbohydrate and phenotype on sucrase, maltase, lactase, and alkaline phosphatase specific activity in SHR/N-cp rat.

The obese spontaneous hypertensive rat/NIH-corpulent (SHR/N-cp) rat exhibits some of the metabolic and pathologic alterations associated with non-insulin-dependent diabetes mellitus and hypertension. The current study was conducted to investigate the influence of phenotype (ob versus In) and source of dietary carbohydrate (sucrose versus starch) on intestinal sucrase, maltase, lactase, and alkaline phosphatase activity in SHR/N-cp rats. For 3 months, lean and obese male SHR/N-cp rats were fed isocaloric diets containing as the sole source of carbohydrate either 54% cooked corn starch or sucrose. Serum and urine markers for diabetes were observed in obese rats. Wet weight and length of intestines were significantly increased in obese rats compared with lean littermates. Among the intestinal enzymes measured, statistical tests confirmed that sucrase activity was significantly increased (P < 0.01) by both phenotype (ob > In) and feeding a sucrose diet. Diet alone (sucrose > starch) significantly increased (P < 0.05) maltase activity in obese rats, but had no effect on lean rats. Lactase activity was significantly higher (P < 0.05) in obese sucrose-fed rats compared with obese starch-fed and/or lean littermates. Statistical tests revealed that intestinal alkaline phosphatase activity was significantly altered (P < 0.05) by both phenotype and diet. Intestinal alkaline phosphatase was higher in starch-fed lean rats compared with lean littermates fed sucrose and to starch or sucrose-fed obese rats. These results are not indicative of a simple, nonspecific increase in intestinal enzyme activity, since the effects observed in intestinal alkaline phosphatase contrast the effects observed in intestinal sucrase, maltase, and lactase activity. These results indicate that both phenotype and diet alter structural and enzymatic intestinal activities of SHR/N-cp rats. Distinct variations in the observed intestinal enzymatic activities suggest that these enzymes are under the control of genetic, hormonal, and dietary factors. Rationale for these differences are discussed.

Sodium cholate-induced changes in the conformation and activity of rat pancreatic cholesterol esterase.

Pancreatic cholesterol esterase (CEase) regulates dietary cholesterol absorption and is activated in the presence of trihydroxy bile salts while remaining inactive monohydroxy bile salts. CEase from rat pancreas has been purified by ammonium sulfate precipitation, hydroxylapatite chromatography, and gel filtration on Sephacryl S-200/S-300 columns connected in series, and its homogeneity and Mr (55,418 +/- 288) have been determined by sedimentation equilibrium centrifugation. The effects of tri-, di-, and monohydroxy bile salts on the conformation of the purified enzyme in buffer solution and in an in vitro assay system were studied by circular dichroism spectropolarimetry. The CD spectrum of the enzyme in solution shows a curve shape suggestive of an alpha-helicity, but low mean residue ellipticity (MRE) values may indicate an important beta-turn contribution. Sodium cholate, a trihydroxy bile salt, induces a decrease in the negative MRE values of the enzyme in solution at bile salt concentrations of 70-100 nM, with no further spectral changes at concentrations as high as 1 mM. Sodium cholate concentrations higher than 1 microM also induce an increase in the enzyme's negative MRE values under activity assay conditions, which reverts toward its original value once the reaction reaches equilibrium. These latter changes are interpreted as induced by substrate binding to the enzyme followed by partial substrate depletion after the reaction reaches equilibrium. Sodium deoxycholate, a dihydroxy bile salt, induces unstable transient increases and decreases in the MRE values of CEase in buffer solution and under activity assay conditions. These changes are bile salt concentration-dependent and may reflect self-association of the protein. Sodium taurolithocholate, a monohydroxy bile salt, does not affect the CD spectrum of CEase, and neither the di- or the monohydroxy bile salt activates the enzyme.