Comparative study of dietary soy phytoestrogens genistein and equol effects on growth parameters and ovarian development in farmed female beluga sturgeon, Huso huso.

Oocyte maturation in fish is a hormonally regulated process. In the light of long-term oocyte maturation in beluga, the aim of this research was to study the estrogenic effects of different concentrations of soy dietary genistein (GE) and equol (EQ) on the growth performance and ovary development in farmed female Huso huso. Fish were fed with concentrations 0 (control), 0.2, 0.4, 0.8 and 1.6 g of EQ and GE per kg of isoproteic (CP 45 %) and isoenergetic (19.5 MJ/kg) diets during a year. Blood samples and ovary biopsies were collected from each fish seasonally. The main results of the present experimentation are that growth performance was not affected significantly both in GE and EQ (P > 0.05). EQ at concentration 0.4 g/kg had more estrogenic effects than other concentrations of EQ and GE in beluga so that 64 % of fish were matured sexually. Some reproductive indices such as oocyte diameter, testosterone (T) and 17ß-estradiol (E2) increased significantly at EQ 0.4 g/kg at the end of experiment (P < 0.05), while 17α-hydroxy progesterone level (17α-OHP) showed no significant changes at all concentrations. Biochemical indices such as calcium, phosphorous and cholesterol increased at GE concentrations, but decreased at EQ concentrations similarly at the end of experiment. There was a negative relationship between plasma phosphorous and alkaline phosphatase enzyme levels. Based on results, EQ at concentration 0.4 g/kg improved oocyte development more than the other concentrations of GE and EQ, and therefore, it can be used as an additive to diets for inducing ovary development in this species.

Effects of caffeine on heart mitochondria in newborn rats.

Caffeine consumption has been implicated in the development of cardiovascular disease. Therefore, in the present study, litters of rats were combined upon birth, and 8 pups were randomly assigned to each dam. Dams with pups were divided into 2 groups: group 1 received a 20% protein diet as a control, and group 2 received the 20% protein diet supplemented with caffeine (4 mg/100 g body weight). Pups from both groups were killed on days 11 and 15. Transmission electron microscopy revealed swollen, disrupted, degenerating mitochondria and intracellular edema in the hearts of rats in the caffeine groups when compared with those of the controls. Plasma Cu concentration was significantly decreased. These results indicate that early exposure to caffeine through maternal milk adversely affects cardiac mitochondria of rat pups and may be associated with decreased plasma Cu levels. It is unclear whether these results apply to the human infant. Interspecies extrapolation from rat to human must be made with caution.

Effects of caffeine on the bones of aged, ovariectomized rats.

Caffeine is a substance which many people consume in their daily life. Caffeine's effects on bone are still controversial. Using ovariectomized rats, the present study was conducted to determine to what extent caffeine intake affects the mechanical properties, bone minerals and histology. Aged rats were divided into 2 groups after ovariectomy. Group 1 was fed a 20% protein diet as a control, and group 2 was fed a 20% protein diet supplemented with caffeine (2 mg/100 g body weight). The respective diets were fed to the rats of each group for 90 days. Rats were then killed by heart puncture, blood was collected, and femurs were removed. In 1 group of femurs paraffin cross-sections were made at the midshaft of each bone. Total width, cortical width, total cross-sectional bone area of the midshaft, and the number of osteocytes in randomly selected areas were measured. Another group of bones was subjected to three-point bending testing until failure. Bones were then pulverized and Ca, P, Mg, Zn, Sr, Si, hydroxyproline and hexosamine contents and crystallite size were measured. Various mechanical properties, except modulus of elasticity, in the caffeine group were consistently 7-23% lower than the noncaffeine controls. Yield strain in the caffeine group was significantly less than in the noncaffeine controls. Zinc, Sr, and crystallite size of bone showed a significant decrease in the caffeine group, whereas Si contents significantly increased. Our current results indicate that routine intake of caffeine in the elderly should be regarded with some caution.

Combined effects of caffeine and alcohol during pregnancy on bones in newborn rats.

The combined effects of caffeine and alcohol on mineral contents of fetal mandibles and femurs were studied. Pregnant rats were divided into four groups: group 1, control; group 2, caffeine; group 3, alcohol; and group 4, caffeine-plus-alcohol. Alcohol (1.0 g ethanol/kg body weight) was intubated twice daily, beginning at day 9 of gestation. Caffeine (2 mg/100 g body weight) was given as a dietary supplement. Groups 1 and 2 were intubated with isocaloric sucrose solution. At birth, randomly selected pups were killed and the mandible and femur were removed and dried. Ca, P, Mg, Zn and hydroxyproline in these bones were measured. Notwithstanding the dams' intake of caffeine and alcohol administered separately, the present results suggest that the combination of caffeine and alcohol exhibited the most detrimental effects.

Relationship of prenatal caffeine exposure and zinc supplementation on fetal rat brain growth.

Pregnant rat dams were divided into four groups on the 3rd day of gestation. Group 1 dams were fed a 20% protein diet as controls. Dams of group 2 were fed a 20% protein diet supplemented with zinc (0.6 g ZnCl2/kg diet). Group 3 dams were fed a 20% protein diet supplemented with caffeine (2 mg/100 g body weight) and dams of group 4 were fed a 20% protein diet supplemented with both caffeine and zinc. Fetuses were surgically delivered on day 22, and brains were removed and analyzed for alkaline phosphatase activity, protein, zinc, cholesterol and DNA concentrations. Fetal brain caffeine levels, as well as maternal and fetal plasma caffeine levels, were determined in caffeine-supplemented groups. The body weight of group 4 and brain weights of groups 3 and 4 were higher than those of groups 1 and 2. Alkaline phosphatase activity of group 3 was less than that of group 1. The brain zinc concentration of group 2 was higher than in the other groups, but that of group 4 was less than that of group 1. The present study indicated that the supplementation of caffeine to the maternal diet decreased zinc levels in the fetal brain, and the addition of extra zinc to this diet did not return the zinc level to that of the control level as we had expected. In addition, the supplementation of caffeine and zinc together increased the body weights of the fetuses compared to the controls, but the addition of only one of these substances had no effect, suggesting that the combination of caffeine and zinc may have unique effects on fetal growth.

Lasting effects of early chronic caffeine feeding on rats' behavior and brain in later life.

Pregnant dams were fed a 20% protein diet with caffeine (2 mg/100 g b.wt.), starting on day 9 of gestation. At birth, each dam with 8 assigned pups was fed this diet until weaning, day 22. On day 22, female rats were caged and fed this diet until day 93. Starting on day 93, the caffeine-supplemented diet was replaced with a caffeine-free, 20% protein diet until day 388. Starting on day 31, each animal was placed in a photoactivity cage, and locomotive activity was measured until day 375. On day 388, the animals were killed, and their brains were removed and divided into 7 regions. The weight, DNA, protein and zinc contents, and alkaline phosphatase activity of each region were determined. Locomotive activity of the caffeine-fed group was higher than in the noncaffeine control group. Accumulative activity scores showed 3 subgroups (high, medium, and low) in both groups at day 93. The medium activity subgroup in the caffeine group was greater than the controls from day 72 to day 93. These differences reappeared 5 weeks after cessation of caffeine supplementation and continued until day 375. The differences in activity were minimum in the high and low subgroups. Chronic caffeine intake in early life permanently affected the medium activity subgroup. Furthermore, various regions of the brain were biochemically altered in spite of the feeding of a noncaffeine diet for almost 300 days after caffeine.

Interaction between caffeine intake and heart zinc concentrations in the rat.

The purpose of the present study was to determine the levels of zinc in the hearts of growing post-weaning offspring, fetuses and their dams chronically fed caffeine. A further study was conducted to determine the distribution of Zn in subcellular heart fractions affected by acutely injecting caffeine into the veins of the adult rats. After delivery pups were raised on a 200 g protein/kg diet until day 22 of weaning. On day 22 randomly selected male offspring from each litter were divided into two groups. Group 1 was fed continuously on the same diet as a control, whereas in the experimental group offspring were fed on a 200 g protein/kg diet supplemented with caffeine (20 mg/kg). On day 49 the animals were killed and Zn, calcium and magnesium concentrations of the hearts were measured. In the second series of studies pregnant dams were randomly divided into two groups. Group 1 was fed on a 200 g protein/kg diet from day 3 of gestation, whereas in the experimental group dams were fed on the diet supplemented with caffeine. On day 22 of gestation the fetuses were surgically removed. The Zn, Ca and Mg concentrations of hearts of fetuses and dams were determined. In the third phase a caffeine solution was injected into the vein. After 45 min the hearts were removed and Zn levels in the subcellular fractions determined. The hearts of the growing offspring fed on a caffeine-supplemented diet consistently showed decreased Zn and Ca levels compared with the non-caffeine group.(ABSTRACT TRUNCATED AT 250 WORDS)

Various levels of maternal caffeine ingestion during gestation affects biochemical parameters of fetal rat brain differently.

Pregnant dams were divided into four groups on day 10 of gestation. Dams of group 1 were fed an 20% protein diet as controls. Dams of groups 2, 3 and 4 were fed a 20% protein diet supplemented with 0.5 mg, 1 mg and 2 mg caffeine/100 g body weight of dams, respectively. Pups were delivered surgically on day 22, and their brains were rapidly removed and analyzed for DNA, protein, cholesterol, zinc and alkaline phosphatase activity. The dams' brains were analyzed for the same parameters as those of the pups. Plasma and brain caffeine levels were also determined in caffeine-supplemented groups. The pups' brains in group 2 were heavier than those in group 4. The DNA concentration of group 2 was higher than that of the other groups. The protein concentration of group 4 was higher than that of the other groups. The cholesterol concentration of group 3 and 4 was less than that of the controls. The zinc concentration of group 4 was less than that of group 2. Alkaline phosphatase activity was decreased in groups 3 and 4 compared with either controls or group 2. Dams showed no significant difference among the groups in the same biochemical parameters except for cholesterol concentration that was higher in groups 2, 3 and 4 than in the controls. Plasma and brain caffeine levels of the fetuses and plasma caffeine of the dams in group 4 were higher than those of either group 2 or 3. It is concluded that various amounts of maternal caffeine intake exert different effects on fetal brain growth. In contrast, the effect of caffeine on the dams' brain is relatively minor.

The effects of maternal caffeine intake during pregnancy on mineral contents of fetal rat bone.

Various levels of maternal caffeine ingestion during pregnancy were investigated to determine whether caffeine will affect the mineral contents of the growing bones of fetal rats. On day 8 of gestation, rat dams were fed with a 20% protein diet supplemented with 0.5 mg, 1 mg, or 2 mg caffeine/100 g of dams body weight as an experimental group and the same without caffeine as a control until day 22 of gestation. Fetuses were removed by cesarean section on day 22 and mandibular bones were removed to study the mineral contents of calcium, phosphorus, magnesium, and zinc. Although the mandible weighed more in the 0.5-mg caffeine group as compared to the controls, an additional increase of caffeine resulted in a decrease in weight. All calcium, magnesium, and zinc contents per bone decreased in the 1- and 2-mg caffeine groups as compared to either controls or 0.5-mg caffeine group, whereas phosphorus stayed relatively constant regardless of the different levels of caffeine intake. When data are expressed as per gram of bone tissue, most of the mineral contents among the groups disappeared suggesting that normal growth and development of the fetal bone are impaired as a results of maternal caffeine intake. Caffeine intake during gestation in the present study indicates that different levels of caffeine intake may exert not only different effects on mineral contents in bone development, but also affect the growth of the fetal bone.

Effects of different levels of caffeine supplemented to the maternal diet on the brains of newborn rats and their dams.

At birth, dams with 8 randomly assigned pups were divided into three groups. Dams of group 1 were fed a control diet. Dams of groups 2 and 3 were fed the control diet supplemented with caffeine (1 mg and 2 mg/100 g body weight, respectively). Pups were killed at day 15 and their brains removed. After weighing, brains were analyzed for DNA, protein, cholesterol, zinc and alkaline phosphatase activity. Brain and plasma caffeine levels were also determined on groups 2 and 3. The dams were milked to measure caffeine levels. The brains from the dams were analyzed for the same parameters as the pups. Caffeine levels in group 3 were consistently higher than in group 2. In the pups, body and brain weights were heavier in group 3 than in the controls. Protein and cholesterol concentrations in group 2 were less than either controls or group 3. Alkaline phosphatase activity in group 2 was higher than either controls or group 3. In the dams, DNA concentration in groups 2 and 3 was less than the controls. Protein and cholesterol concentration in group 2 was less than group 3. It was concluded that low levels of caffeine in the maternal diet during lactation could affect various parameters in the newborn brain. These effects were different from those when the dietary caffeine level was doubled. In contrast, the effects of caffeine on brains of the dams were relatively minor.