Exposure to flaxseed or its lignan component during different developmental stages influences rat mammary gland structures
Abstract and Figures
Reduction of the highly proliferative terminal end bud (TEB) structures in the developing mammary gland by differentiation to alveolar buds (ABs) and lobules has been suggested to be protective against mammary cancer. Flaxseed is high in alpha-linolenic acid (ALA) and secoisolariciresinol diglycoside (SDG). SDG is the precursor of mammalian lignans, which can affect mammary gland structures. Thus, the objective of this study was to determine the effect of lifetime, gestation and lactation or after-weaning exposure to 5 or 10% flaxseed or SDG and flaxseed oil components on the mammary gland structures of virgin female rat offspring at post-natal day 50. Lifetime or gestation and lactation exposure to flaxseed altered mammary gland structure development, whereas exposure to flaxseed after weaning had no effect. Lifetime or gestation and lactation exposure to 5% flaxseed caused endocrine changes, as suggested by delayed puberty onset and reduced number of estrous cycles. These changes reduced exposure to endogenous estrogens, leading to atrophy of mammary TEB structures. SDG, but not flaxseed oil, at the level found in 5% flaxseed produced similar effects as 5% flaxseed. This suggested that the lignans were the component in flaxseed responsible for the observed effects. Lifetime or gestation and lactation exposure to 10% flaxseed also caused endocrine changes, as suggested by early puberty onset and lengthened cycles due to prolonged estrus. This increased exposure to endogenous estrogens and stimulated mammary gland differentiation, as indicated by fewer TEBs and more ABs. Thus, lifetime or gestation and lactation exposure to 5 or 10% flaxseed induced structural changes in the mammary gland that may potentially reduce mammary cancer risk.
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... Fat and lactose contents in milk were not affected by zinc deficiency, although milk protein concentration and distribution were modified [105] Studies conducted on different species have begun to decipher the role of phytoestrogen treatments on mammary development, but poor documentation exists concerning the impact of dietary consumption of phytoestrogens [106,107]. Flaxseed is high in α-linolenic acid, and its consumption can produce mammary structure modifications in rats: female rats fed a 5% flaxseed diet showed an antiestrogenic-induced reduction in terminal end buds due to atrophy, while a 10% flaxseed diet produced estrogenic effects that also resulted in terminal end buds' reduction but due to lack of differentiation [108], demonstrating strong dose-dependent effects. Feeding rats with 10% flaxseed during pregnancy and lactation led to a decrease in terminal end buds and an increase in alveolar buds [108,109]. ...
... Flaxseed is high in α-linolenic acid, and its consumption can produce mammary structure modifications in rats: female rats fed a 5% flaxseed diet showed an antiestrogenic-induced reduction in terminal end buds due to atrophy, while a 10% flaxseed diet produced estrogenic effects that also resulted in terminal end buds' reduction but due to lack of differentiation [108], demonstrating strong dose-dependent effects. Feeding rats with 10% flaxseed during pregnancy and lactation led to a decrease in terminal end buds and an increase in alveolar buds [108,109]. Lactation therefore appears to be a critical period for enhancing the differentiation of terminal end buds to alveoli buds by dietary phytoestrogens [110]. Genistein is a major component of soy and one of the most consumed phytoestrogens worldwide. ...
In mammals, milk is essential for the growth, development, and health. Milk quantity and quality are dependent on mammary development, strongly influenced by nutrition. This review provides an overview of the data on nutritional regulations of mammary development and gene expression involved in milk component synthesis. Mammary development is described related to rodents, rabbits, and pigs, common models in mammary biology. Molecular mechanisms of the nutritional regulation of milk synthesis are reported in ruminants regarding the importance of ruminant milk in human health. The effects of dietary quantitative and qualitative alterations are described considering the dietary composition and in regard to the periods of nutritional susceptibly. During lactation, the effects of lipid supplementation and feed restriction or deprivation are discussed regarding gene expression involved in milk biosynthesis, in ruminants. Moreover, nutrigenomic studies underline the role of the mammary structure and the potential influence of microRNAs. Knowledge from three lactating and three dairy livestock species contribute to understanding the variety of phenotypes reported in this review and highlight (1) the importance of critical physiological stages, such as puberty gestation and early lactation and (2) the relative importance of the various nutrients besides the total energetic value and their interaction.
... Lignans are a group of phytochemicals that have been shown to possess weak estrogenic and antiestrogenic properties depending upon duration, dose, and stage of development [42,43,44,45,46,47,48,49] . A balanced randomized cross-over design study recorded an increased luteal phase length and higher progesterone to estrogen ratio in the flax seed ingested group than in control [50] . ...
The seed cycling/seed rotation diet is a new trend that claims to be effective for female menstrual dysfunctions such as irregular menstruation, menstrual cramps, infertility, menopausal symptoms (hot flashes, fatigue, etc.), and PCOS. The higher prevalence of hormonal imbalance in women is a contributing factor to all of these menstrual dysfunctions. The major hormones that control menstruation in females include progesterone, estrogen, luteinizing hormone (LH), and follicular stimulating hormone (FSH), and a simple imbalance in their concentration is the root cause of a variety of menstrual problems. The practice of eating specific seeds during the two main phases of the menstrual cycle (follicular and luteal) to promote a healthy balance of estrogen and progesterone levels in women is known as the seed rotation diet. During the follicular stage consumption of pumpkin seed and flax seed is advised and in the luteal stage consumption of sunflower seed and sesame seed is advised in this diet. Recent studies have shown that pumpkin seeds are rich in phytoestrogen which is a polyphenol compound that exerts a mammalian estrogenic-like effect in body. Similarly, flax seed in the first phase helps to bind the excess estrogen produced to maintain the hormonal homeostasis. Sesame seeds, which are high in zinc and lignans, are thought to help with progesterone balance during the luteal phase, while sunflower seeds, which are high in vitamin E and selenium aids in, increasing progesterone production and liver detoxification of excess estrogen. However, despite plenty of anecdotal accounts of its usefulness, scientific evidence to back its claims is still weak or lacking.
... Flaxseed offers high 毩 -linolenic acid content (58% of the total fatty acids), better palatability compared to fish meal as a consequence of non-fishy odor, making it an energy dense replacement compared to other costly feed ingredients [17]. In addition, flaxseed also contains high concentrations of secoisolariciresinol diglycoside, a precursor of lignans, which in turn exhibits estrogenic activities [18]. Hence, the present study was to investigate the impact of minimum dietary concentrations of supplemental flaxseed on the metabolic, endocrine and reproductive performance of post-farrowed sows. ...
Objective: To assess the effect of flaxseed supplementation on metabolic profile, endocrine concentrations, non-esterified fatty acids (NEFA), body composition variables, and reproductive performance of sows.
Methods: All the 21 crossbred Large White Yorkshire sows were considered in the study period starting at day 1 of current farrowing when the feeding of specific supplemental ration was started until the day of subsequent farrowing (days 150-155) and were equally allocated into three groups. Group 1 served as the control group and followed their normal feeding schedule. Group 2 and group 3, in addition to their normal feeding schedule, were supplemented with flaxseed at a rate of 0.5% and 1.0% of the dry matter, respectively. Blood samples were collected 15 days prior to farrowing, on the day of farrowing (day 0), at weekly intervals until day 28 of lactation and at monthly intervals during gestation to harvest the plasma. Plasma was used to assess the metabolic and endocrine status of sows. Body weight of each sow and individual birth weight of all piglets born were measured.
Results: Flaxseed supplementation led to decrease in plasma cholesterol and triglyceride levels in the supplemented groups than in the control group (P
... The extraction of Crude Lignan was described by, (5), while the purification of lignan was carried out by column chromatography (purification) which was described by (6) For histological study of rabbit tissue, the rabbits were anaesthetized by chloroform and killed. Immediately after death the mammary gland was excised and preserved in fixative solution till the preparation of histological sections .Several tissue sections were prepared according to (7). ...
This study was carried out to investigate the effect that result from taking 40 mg /kg \BW of purified lignan from the seed of flax Linum usitatissimum L. in the growth and development of mammary glands during virgin , pregnancy and lactating stages for 14 days. Thirty (30) rabbits were used in this experiment divided into six groups, two groups at maturity (virgin) stage, two groups at pregnancy stages and two groups at lactating stages (5 rabbits\each group). The following studies were decided: histological, histochemical, biochemical and hormonal studies for each group. Microscopic examination of mammary gland in the virgin rabbits that treated with pure flax lignan showed more growth in the size of lobules and alveoli. In pregnant, the treated groups showed more branching of alveoli with more flattened epithelium. In lactating treated group showed the lobules were expanded and contained more branching alveoli with discontinuous flattened epithelium. The PAS staining reaction of histological sections of mammary gland from treated groups and control were showed positive reactions in all groups. The biochemical studies showed a significant increase (p<0.05) in the serum reduced glutathione concentration with decrease in the malondialdehyde concentration in the three stages that treated with pure flax lignan compared with control. The hormonal study indicated a noticeable increase in prolactin hormones with significant decreased in estrogen in all groups of treated animals for the three physiological stages (virgin, pregnant and lactating) compared with their controls.
... In addition, SDG-based diet during pregnancy and lactation helped in reducing the susceptibility of mice to carcinogenesis and reduced tumorigenesis. This pattern was not observed with FSO-based dietary intervention [8]. Taken together, these findings support the use of FS and SDG during MG development to reduce the future risk of breast cancer. ...
Breast cancer is the most common cancer among women worldwide. We previously showed that early-life exposure to flaxseed (FS) or its components, FS oil (FSO) and secoisolariciresinol diglucoside (SDG), affects the mammary gland (MG) and is associated with the reduction of breast cancer risk during adulthood. However, the underlying mechanisms are not understood. This study aimed to investigate the effect of FS, FSO, and SDG on the MG miRNA signature at a late stage of development. Female C57BL/6 mice, 4–5 weeks of age, were randomized into four groups to receive: (i) basal AIN-93G, (ii) 10% FS, (iii) 3.67% FSO, or (iv) 0.15% SDG. After 21 days, the mice were sacrificed and MG miRNAs were profiled. Diet-specific MG miRNA signatures were identified. Deregulated miRNAs were associated with breast cancer and targeted genes involved in MG development, growth, and cancer. The study allowed for the identification of potential biomarkers or novel therapeutic targets to prevent and/or reduce the risk of breast cancer.
A total synthesis of both pairs of enantiomers of natural lignans, diaeudesmin and epieudesmin, was described. The synthesis involved enantioselective allylborylation of veratraldehyde with 1,6-bis(diisopinocampheylboryl)hexa-2,4-dienes, ozonolysis, and triethylsilane reduction in the presence of boron trifluoride etherate.
Flax sed also known as flax oil and linseed oil, is derived from the seeds of the plant Linium usitatissimum. Flax seed oil is a very rich source of alpha-linolenic acid. Alpha-linolenic acid concentration in flaxseed oil ranges from approximately 40 to 60%.lower amounts of linoleic acid and oleic acid (each about 15%) are also present in flaxseed oil.ın addition, flaxseed contains varying amounts of the lignan, secoisolariciresinol diglycoside (SDG).
Microfluidic organ-on-chip devices constructed from polydimethysiloxane (PDMS) have proven useful in studying both beneficial and adverse effects of drugs, supplements, and potential toxicants. Despite multiple advantages, one clear drawback of PDMS-based devices is binding of hydrophobic chemicals to their exposed surfaces. Chemical binding to PDMS can change the timing and extent of chemical delivery to cells in such devices, potentially altering dose-response curves. Recent efforts have quantified PDMS binding for selected chemicals. Here, we test a wider set of nineteen chemicals using UV-Vis or infrared spectroscopy to characterize loss of chemical from solution in two setups with different PDMS-surface-to-solution-volume ratios. We find discernible PDMS binding for eight chemicals and show that PDMS binding is strongest for chemicals with a high octanol-water partition coefficient (Log P > 1.85) and low H-bond donor number. Further, by measuring depletion and return of chemical from solution over tens to hundreds of hours and fitting these results to a first order model of binding kinetics, we characterize partitioning into PDMS in terms of binding capacities per unit surface area and both forward and reverse rate constants. These fitted parameters were used to model the impact of PDMS binding on chemical transport and bioavailability under realistic flow conditions and device geometry. The models predict that PDMS binding could alter in-device cellular exposures for both continuous and bolus dosing schemes by up to an order of magnitude compared to nominal input doses.
Genistein, a component of soy, was administered to prepubertal female Sprague-Dawley Co rats and investigated for chemoprevention against mammary cancer, Genistein, at 500 mu g/g body wt or an equivalent volume of the vehicle, dimethylsulfoxide (DMSO), was injected (s.c.) on days 16, 18 and 20 post-partum, At day 50 post-partum all animals were exposed to 80 mu g dimethylbenz[a]anthracene (DMBA) per g body wt, Animals treated prepubertally with genistein as compared to DMSO had reduced incidence and significantly fewer adenocarcinomas per animal. Mammary whole mount analysis showed that prepubertal genistein treatment resulted in mammary glands of 50-day-old rats developing fewer terminal end buds and more lobules II, Cell proliferation studies with bromodeoxyuridine (BrdU) showed that terminal end buds from mammary glands of 50-day-old females treated prepubertally with genistein had significantly fewer cells in S-phase of the cell cycle, Serum genistein concentrations in 21- and 50-day-old females following prepubertal genistein treatment were 4.2 +/- 0.6 mu M and 102 +/- 30 nM, respectively, Animals treated prepubertally with genistein as compared to vehicle spent more time in the estrus phase of the estrus cycle, although all animals did cycle, In 50-day-old females, circulating estradiol-17 beta and progesterone concentrations were not significantly altered by the prepubertal genistein treatment, Oocyte/follicle counts and numbers of atretic follicles and corpora lutea were not significantly different between the genistein- and vehicle-treated animals, We conclude that genistein treatment during the prepubertal period can suppress the development of chemically-induced mammary cancer without significant toxicity to the endocrine/reproductive system.
Flaxseed, a rich source of mammalian lignan precursor secoisolariciresinol-diglycoside (S.D.) and alpha-linolenic acid (ALA), has been shown to be protective at the early promotion stage of carcinogenesis. The objective of this study was to determine whether supplementation with flaxseed, its lignan or oil fractions, beginning 13 weeks after carcinogen administration, would reduce the size of established mammary tumors (present at the start of treatment) and appearance of new tumors in rats. Dietary groups consisted of the basal diet (BD, 20% corn oil) alone or supplemented with a gavage of 2200 nmol/day S.D. [S.D., equal to level in 5% flaxseed (F)], 1.82% flaxseed oil (OIL, equal to level in 5% F) or 2.5% or 5% flaxseed (2.5% F and 5% F, respectively). After 7 weeks of treatment, established tumor volume was over 50% smaller in all treatment groups (OIL, 2.5% F, 5% F, P < 0.04; S.D., P < 0.08) while there was no change in the BD group. New tumor number and volume were lowest in the S.D. (P < 0.02) and 2.5% F (P < 0.07) groups. The combined established and new tumor volumes were smaller for the S.D., 2.5% F and 5% F groups (P < 0.02) compared to the OIL and BD groups. The high negative correlation (r = -0.997, P < 0.001) between established tumor volume and urinary mammalian lignan excretion in the BD, S.D., 2.5% F and 5% F groups indicates that the reduction in tumor size is due in part to the lignans derived from the S.D. in flaxseed. However, there was no relationship between new or total tumor development and urinary lignan levels, The effect of flaxseed oil may be related to its high ALA content. In conclusion, the S.D. in flaxseed appears to be beneficial throughout the promotional phase of carcinogenesis whereas the oil component is more effective at the stage when tumors have already been established.
For sixteen years, the American institute of Nutrition Rodent Diets, AIN-76 and AIN-76A, have been used extensively around the world. Because of numerous nutritional and technical problems encountered with the diet during this period, it was revised. Two new formulations were derived: AIN-93G for growth, pregnancy and lactation, and AIN-93M for adult maintenance. Some major differences in the new formulation of AIN-93G compared with AIN-76A are as follows: 7 g soybean oil/100 g diet was substituted for 5 g corn oil/ 100 g diet to increase the amount of linolenic acid; cornstarch was substituted for sucrose; the amount of phosphorus was reduced to help eliminate the problem of kidney calcification in female rats; L-cystine was substituted for DL-methionine as the amino acid supplement for casein, known to be deficient in the sulfur amino acids; manganese concentration was lowered to one-fifth the amount in the old diet; the amounts of vitamin E, vitamin K and vitamin B-12 were increased; and molybdenum, silicon, fluoride, nickel, boron, lithium and vanadium were added to the mineral mix. For the AIN-93M maintenance diet, the amount of fat was lowered to 40 g/kg diet from 70 g/kg diet, and the amount of casein to 140 g/kg from 200 g/kg in the AIN-93G diet. Because of a better balance of essential nutrients, the AIN-93 diets may prove to be a better choice than AIN-76A for long-term as well as short-term studies with laboratory rodents.
The effect of each of twelve mammalian lignun derivatives on the growth of human mammary tumor ZR-75-1 cells was examined. At a concentration less than 10 μ g/ml, tumor cell growth was inhibited from 18-68%. The effect of 2,3-dibenzylbutane-1,4-diol (hattalin) was found to be strongest, inhibiting growth by 50% at a concentration (EC50 of 2.1 μ g/ml. Hattalin inhibited membrane Na%, K%-ATPase of canine kidney cortex. It also inhibited the ATPase of the plasma membrane fraction from both cultured cells and a section of human breast cancer tissue at a concentration ranging from 0.5 to 2.0 mM. However, only a few percent of membrane ATPase from either ZR-75-1 cells or breast carcinoma tissue was inhibited by 2.0 mM of ouabain, suggesting that the target ATPase of hattalin was other than ouabainsensitive ATPase. The relative incorporation of [3H]thymidine per 1 105 cells into the acid-precipitable fraction of ZR-75-1 cells was not affected by 1-50 μg/ml of hattalin, while a marked decrease resulted from 1-10 μg/ml of 5-fluorouracil (5-FU). These results suggest that the suppressive effect of hattalin on tumor cell growth my not occur through inhibition of DNA synthesis but rather partly by inhibition of the plasma membrane ATPase other than Na% and K% -dependent ones.
The recently passed Food Quality Protection Act of 1996 requires the U.S. EPA to implement screening strategies for endocrine active compounds (EACs) within the next 2 years. Interpreting results from screening tests is complicated by the absence of traditional dietary rodent bioassay data with model estrogenic compounds such as 17β-estradiol. Thus, a 90-day/one-generation reproduction study with 17β-estradiol was designed to: (1) provide such baseline data; (2) set dose levels for multigeneration reproduction and combined chronic toxicity/oncogenicity studies; and (3) evaluate various mechanistic/biochemical endpoints for inclusion in these follow-up studies. The current article describes the effects of dietary administration of 0, 0.05, 2.5, 10, and 50 ppm 17β-estradiol on the serum hormone concentrations and estrous cyclicity of female Crl:CD BR rats and evaluates a sampling strategy for measuring serum hormone levels in cycling female rats. Serum hormones were measured at three time points during a 90-day dietary exposure (1 week, 28 days, and 90 days) and in the F1generation rats on postnatal day 98. Over the course of the 90-day feeding study for the P1generation and from postnatal days 21 to 98 for the F1generation, the estrous cycle was monitored daily in 10 rats/group. In P1generation rats, dietary administration of 2.5, 10, and 50 ppm 17β-estradiol produced a dose-dependent increase in serum estradiol (E2) concentrations at all time points. In contrast, administration of 0.05, 2.5, 10, and 50 ppm 17β-estradiol produced a dose-dependent decrease in serum progesterone (P4) concentrations on test day 90, which correlated with an absence of corpora lutea and ovarian atrophy. At 10 and 50 ppm 17β-estradiol, serum luteinizing hormone (LH) concentrations were consistently decreased at all time points and were decreased at 2.5 ppm on test day 90. Serum prolactin (PRL) concentrations were increased at 50 ppm 17β-estradiol on test day 90. Serum follicle stimulating hormone (FSH) concentrations were either similar to the control levels or minimally changed at all time points. No F1generation rats were produced at 10 or 50 ppm 17β-estradiol. In F1generation rats, serum E2 concentrations were increased and P4 concentrations were decreased at a dietary concentration of 2.5 ppm 17β-estradiol, while serum concentrations of LH, FSH, and PRL were similar to the control. Dietary administration of 17β-estradiol at concentrations of 2.5 (both generations) and 10 and 50 ppm (P1generation only) produced marked effects on the estrous cycle: decreased number of cycles, increased mean cycle length, and decreased number of normally cycling rats. The estrous cyclicity of rats fed 2.5 ppm 17β-estradiol appeared more severely affected in rats of the F1generation than in rats of the P1generation. Whether this increase in severity is related to anin uteroexposure and/or greater mean daily intake of 17β-estradiol in the F1generation rats in the postnatal period is unclear. Another goal of this study was to evaluate whether a single time point sampling strategy using cycling female rats could be used to detect compound-related changes in serum hormone concentrations. In evaluating a sampling strategy for measuring serum hormone levels, it appears that detection of compound-related alterations in serum hormone concentrations can be best detected by sampling during diestrus. Since the stage of the cycle dramatically influences hormone concentrations, large sample sizes (n= 50) are needed if serum hormone measurements are not matched with the stage of the cycle. The data indicate that this strategy of measuring serum hormone concentrations has utility in detecting compound-related effects within the confines of a traditional guideline study (subchronic, chronic, or multigenerational reproduction study).
The amorphous product, extracted from fat-free linseed meal with ethanol-dioxane, yields upon treatment with sodium methoxide (a) a methyl ester, ( b) a crystalline glucoside and (c) a non-crystalline glucoside. The ester is shown to be methyl β-hydroxy-β-methylglutarate (I) and the crystalline acid derived from it has been synthesized from diallylmethylcarbinol by ozonolysis and subsequent oxidation.
Human and animal data indicate that a high maternal estrogen exposure during pregnancy increases breast cancer risk among daughters. This may reflect an increase in the epithelial structures that are the sites for malignant transformation, i.e., terminal end buds (TEBs), and a reduction in epithelial differentiation in the mammary gland. Some phytoestrogens, such as genistein which is a major component in soy-based foods, and zearalenone, a mycotoxin found in agricultural products, have estrogenic effects on the reproductive system, breast and brain. The present study examined whether in utero exposure to genistein or zearalenone influences mammary gland development. Pregnant mice were injected daily with i) 20 ng estradiol (E2); ii) 20 μg genistein; iii) 2 μg zearalenone; iv) 2 μg tamoxifen (TAM), a partial estrogen receptor agonist; or v) oilvehicle between days 15 and 20 of gestation. E2, genistein, zearalenone, and tamoxifen all increased the density of TEBs in the mammary glands. Genistein reduced, and zearalenone increased, epithelial differentiation. Zearalenone also increased epithelial density, when compared with the vehicle-controls. None of the treatments had permanent effects on circulating E2 levels. Maternal exposure to E2 accelerated body weight gain, physical maturation (eyelid opening), and puberty onset (vaginal opening) in the female offspring. Genistein and tamoxifen had similar effects on puberty onset than E2. Zearalenone caused persistent cornification of the estrus smears. These findings indicate that maternal exposure to physiological doses of genistein mimics the effects of E2 on the mammary gland and reproductive systems in the offspring. Thus, our results suggest that genistein acts as an estrogen in utero, and may increase the incidence of mammary tumors if given through a pregnant mother. The estrogenic effects of zearalenone on the mammary gland, in contrast, are probably counteracted by the permanent changes in estrus cycling.