Mammary gland development and hormone levels in pregnant Upton-Meishan and large white gilts

Genetic differences between Upton-Meishan (UM, n = 13) and Large White (LW, n = 14) gilts were studied with regard to mammary gland development and concentrations of hormones. Gilts were weighed and their backfat measured at mating, and at d 70 and 109 of gestation. Jugular blood samples were also collected at these times and assayed for prolactin, cortisol, IGF-I, insulin, glucose, progesterone, and estradiol. Gilts were slaughtered on d 110 of gestation. One row of mammary glands was used for dissection and biochemical analyses. The other row was used for determination of prolactin receptor number and affinity. UM gilts weighed less (P<0.05) and had more backfat (P<0.01) than LW gilts at all times. Parenchymal tissue weight was less (P<0.05) in UM gilts. Percent fat (P<0.001) and dry matter (P<0.001) in parenchymal tissue were greater in UM gilts while that of protein (P<0.001) was lower. Total protein weight in parenchyma was also lower in parenchyma was also lower in UM gilts (P = 0.01). Both DNA (P<0.001) and RNA (P<0.001) contents were lower in UM gilts while RNA/DNA remained similar (P>0.1). Number of prolactin receptors were lower (P = 0.06) and affinity greater (P<0.05) in UM gilts. Cortisol levels were greater (P<0.01) in UM gilts while other hormones were not affected (P> 0.1). Results clearly demonstrate genetic differences with regard to mammogenesis in gilts and suggest that the less mammary gland development in Upton-Meishan compared with Large White breed of gilts may be related to lower number of prolactin receptors.     

Inhibition of prolactin in the last trimester of gestation decreases mammary gland development in gilts

Prolactin is required for mammary development in various species but its possible role for mammogenesis in pigs is not known. The goal of the present study was therefore to determine the effect of prolactin inhibition by bromocriptine during the last third of gestation on mammary gland development in gilts. Twenty-eight primigravid gilts were assigned as controls (n = 15) or received 10 mg of bromocriptine orally thrice daily (n = 13) from d 70 to 110 of gestation. Jugular blood samples were collected on d 70 of gestation and every 8 d thereafter and were assayed for prolactin, IGF-I, estradiol, and progesterone. Gilts were slaughtered on d 110 of gestation and fetuses were counted and weighed. One row of mammary glands was used for dissection of parenchymal and extraparenchymal tissues and for determination of DNA, RNA, dry matter, protein, and fat contents. Tissue from the other row was used for measures of prolactin receptor number and affinity. Concentrations of prolactin were drastically reduced throughout the bromocriptine treatment period (P < .001), whereas there was no overall treatment effect on progesterone and IGF-I levels (P > .10). Total weight and extraparenchymal tissue weight of the mammary glands were unaffected by treatment (P > or = .1), but weight of parenchymal tissue, total DNA, and total RNA decreased (P < .01) with bromocriptine treatment. Percentages of fat and dry matter in parenchymal tissue increased with bromocriptine treatment (P < .01) and the percentage of protein decreased (P < .01). Number of prolactin receptors in parenchymal tissue decreased with bromocriptine treatment (P < .001) and receptor affinity increased (P < .001). Average fetal weight was lower in gilts receiving bromocriptine than in control gilts (P = .05), but fetal number did not differ (P > .1). These results clearly demonstrate that prolactin is essential for normal mammary gland development and can affect fetal growth during the last third of gestation in gilts.     

Nitrogen requirement of pregnant gilts

Eleven Yorkshire x Landrace gravid gilts were used in two Latin square trials to determine the N requirement for pregnancy. Semipurified diets were formulated to be adequate in indispensable amino acids (IAA) and other nutrients. Diets were fed once daily (1.82 kg); L-glutamic acid, corn sugar, powdered cellulose, and soybean oil percentages differed to maintain isocaloric diets. In Trial 1, six gilts were fed diets containing 6.6 to 17.2% CP equivalent (19.2 to 50.1 g of N/d) during six 10-d periods beginning on d 40 postcoitum. In Trial 2, five gilts were fed diets containing 4.3 to 12.6% CP equivalent (12.5 to 36.8 g of N/d) during five 10-d periods beginning on d 50 postcoitum. Nitrogen balance trials were conducted during the last 5 d of each period. Blood samples were taken both before and 3 h after the last feeding of each period. Results from Trial 1 suggested that adequate N retention (10 g/d) could be attained by pregnant gilts fed less than 28 g of N/d intake when the diet contained indispensible amino acids at levels suggested by Nutrient Requirements of Swine (NRC, 1988). An intake of 20.6 g of N/d (7.1% CP equivalent) yielded near maximum N retention among pregnant gilts fed the semipurified diets in Trial 2. Other criteria measured (urine urea nitrogen and plasma urea nitrogen) had limited value in the evaluation of the N status of the pregnant gilts in these trials.     

Protein and energy balance in pregnant gilts

Thirty-nine Landrace gilts, average age 240.6 days and mean body weight 117.1 kg, were mated and divided into five feeding groups. Gilts of groups 1 to 4 were fed daily, for 111 days of gestation, 1.5, 2.0, 2.5 or 3.5 kg of a diet containing 12% digestible protein (DP); gilts of group 5 were given daily 3.5 kg of a diet containing 14.7% DP. All gilts were slaughtered on day 112 of gestation and chemical composition of the carcass and internal organs, including the gestated utera, were analysed. Initial chemical body composition of gilts was estimated by equations obtained on 28 animals slaughtered at the beginning of the experiment.The level of feeding had no effect on the litter size, or on the chemical composition of the products of conception.Protein and fat deposition increased linearly with elevation of the amount of food per day. In gilts given daily 1.5 kg of the diet, retention of protein was accompanied by loss of fat from the body.Increase in protein content in the diet did not reduce the utilization of digestible protein for its retention in the body.The protein and energy costs of 1 g protein deposited in tissues was higher in the products of conception (6.57 g DP and 79.2 kJ ME) than in the maternal bodies of gilts (3.66 g DP and 48.4 kJ ME). The maintenance requirement of pregnant gilts was estimated at 490.6 kJ ME per kg^0.75 per day.     

Specific window of prolactin inhibition in late gestation decreases mammary parenchymal tissue development in gilts

Prolactin is required from d 70 to 110 of gestation for normal mammary development of gilts. The goal of the present study was to determine the effect of inhibiting prolactin with bromocriptine during specific time windows during the second half of gestation on mammary gland development in gilts. Crossbred primigravid gilts were assigned as controls (n = 12) or received 10 mg of bromocriptine orally three times daily from d 50 to 69 (BR50, n = 12), d 70 to 89 (BR70, n = 12), or d 90 to 109 (BR90, n = 12) of gestation. Jugular blood samples were collected on d 50, 70, 90, and 109 of gestation and assayed for prolactin and estradiol. Gilts were slaughtered on d 109 of gestation and fetuses were counted and weighed. One row of mammary glands was used for dissection of parenchymal and extraparenchymal tissues, and for biochemical analyses. Tissue from the other row was used for measures of prolactin receptor number and affinity. Concentrations of prolactin were decreased markedly (P < 0.001) at the end of each bromocriptine treatment period compared with controls, but there was no overall treatment effect (P > 0.1) on estradiol concentrations. Extraparenchymal tissue weight of the mammary glands was unaffected by treatments (P > 0.1), but weight of parenchymal tissue, total DNA, and total RNA were lower (P < 0.01) in BR90 than control gilts. The percentage of DM in parenchymal tissue was unaffected by treatments (P > 0.1), but percentage of fat was higher and percentage of protein lower (P < 0.01) in BR90 gilts compared with controls. Cell size, as estimated by the protein:DNA ratio, also was lower (P < 0.01) in the BR90 group. Number and affinity of prolactin receptors in parenchymal tissue were not significantly altered by treatments. In conclusion, there is a specific time period in the second half of gestation, from 90 to 109 d, during which prolactin is essential for normal mammary parenchymal tissue development.     

Effects of increased dietary energy and protein during late gestation on mammary development in gilts

Thirty-two gilts were used to evaluate the effects of increased dietary energy and CP during late gestation on mammary development. On d 75 of gestation, gilts were assigned randomly in a 2 x 2 factorial arrangement to adequate (5.76 Mcal ME/d) or increased (10.5 Mcal ME/d) energy and adequate (216 g CP/d) or increased (330 g CP/d) protein. On d 105 of gestation, gilts were slaughtered and total mastectomies were performed. Mammary tissue was separated into mammary parenchymal and mammary extraparenchymal stromal tissue and analyzed for DNA, RNA, protein and lipid. No interactions between dietary energy and protein level were detected (P greater than .20). When adjusted for number of mammary glands and maternal BW (weight of the sow less the weight of the fetuses), mammary parenchymal weight was 27% greater (P less than .03) in gilts fed adequate energy than in gilts fed increased energy, but mammary extraparenchymal stroma weight was unaffected by dietary energy level. Total mammary parenchymal DNA was 30% greater in gilts fed adequate energy than in gilts fed increased energy (P less than .03). Total mammary parenchymal RNA (P less than .02) and total mammary parenchymal protein (P less than .02) also were greater in gilts fed adequate energy than in gilts fed increased energy. Dietary protein level did not affect mammary variables measured, except that increased dietary protein tended to reduce mammary extraparenchymal stromal weight (P less than .09). Increased dietary protein between d 75 and d 105 of gestation did not benefit mammary development, but increased dietary energy was detrimental to development of mammary secretory tissue.     

The role of IGFBP-5 in mammary gland development and involution

Insulin-like growth factor-I (IGF-I) plays an important role as a survival factor during mammary gland development and remodelling during involution of the mature/lactating mammary gland, and elevated concentrations have been associated with increased risk of breast cancer. The actions of IGF-I are modulated by a family of binding proteins (IGFBPs) and we have shown that IGFBP-5 is associated with cell death in the mammary gland and more recently provided the first evidence that it is causally related to apoptosis of the mammary gland. A transgenic mouse expressing IGFBP-5 on a mammary-specific promoter led to impaired mammary development involving inhibition of IGF-signalling and involving members of the Bcl-2 family. Subsequent studies in vitro and in vivo using exogenous IGFBP-5 treatment have added support to this concept. Although the effects of IGFBP-5 did appear to involve inhibition of IGF action, a role for IGF-independent effects cannot be ruled out. Such IGF-independent effects involve potential interactions with components of the extracellular matrix involved in tissue remodelling including plasminogen activator inhibitor-1 (PAI-1). In addition, intracellular events involving nuclear localisation of IGFBP-5 have been shown to have the ability to inhibit cell proliferation. Thus, IGFBP-5 seems important for regulating both apoptosis and cell proliferation in the mammary gland during development and post-lactation involution.     

Characterization of spheres derived from canine mammary gland adenocarcinoma cell lines

There is increasing evidence for the presence of cancer stem cells in several solid tumors, and these cancer stem cells have a potential role in tumor initiation, aggression, and recurrence. The stem cell-like properties of spheres derived from canine mammary tumors remain largely elusive. We attempted to induce sphere formation using four cell lines of canine mammary adenocarcinoma, and characterized the spheres derived from a CHMp line in vitro and in vivo. The CHMp-derived spheres showed predominantly CD44⁺CD24⁻ population, higher expression of stem cell-related genes, such as CD133, Notch3 and MDR, and higher resistance to doxorubicin compared with the CHMp-derived adherent cells. Xenograft transplantations in nude mice demonstrated that only 1 × 10⁴ sphere cells were sufficient for tumor formation. Use of the sphere assay on these sphere-derived tumors showed that sphere-forming cells were present in the tumors, and were maintained in serial transplantation. We propose that spheres derived from canine mammary adenocarcinoma cell lines possess a potential characteristic of cancer stem cells. Spheres derived from canine mammary tumors could be a powerful tool with which to investigate novel therapeutic drugs and to elucidate the molecular and cellular mechanisms that underlie tumorigenesis.     

The expression of carbonic anhydrase in canine mammary gland and mammary gland tumor

Carbonic anhydrase (CA), a metallo-enzyme containing zinc, broadly distributes in mammalian tissues and participates in physiological regulation such as respiration, acid-base balance, ion transport, bone resorption, as well as the development of tumor by the reversible hydration of carbon dioxide. However the expression of CA in the tissue of mammary gland tumor was not documented. In this study we examine the histolocalization and gene expression of CA in both normal canine mammary gland tissue and mammary gland tumor by histochemical examination, and RT-PCR. Four mRNA expression of CA isoenzymes, such as CA II, IV, VI and IX were found under RT-PCR analysis and different band patterns were found between normal canine mammary tissue and canine mammary gland tumor tissue. CA II, IV, VI and IX gene mRNA expression were found in the normal mammary gland tissue, indicating CA II, IV, VI and IX are likely to be the essential enzymes to maintain the normal physiological condition of canine mammary gland tissue cells. However the expression of CA IV was not found in the tissue of malignant mammary gland tumor that may become the marker for the prognostic recognition of canine mammary gland tumor.     

Impact of diet deprivation and subsequent over-allowance during prepuberty. Part 1. Effects on growth performance, metabolite status, and mammary gland development in gilts

The impact of diet deprivation and subsequent over-allowance on the metabolite status, mammary development, and mammary gene expression in prepubertal gilts was determined. Forty-seven gilts were reared under a conventional (control, CTL; n = 23) or an experimental (treatment, TRT; n = 24) dietary regimen. The later regimen (consisting of diet deprivation and subsequent over-allowance) provided 70 (restriction diet, RES) and 115% (over-allowance diet, OVER) of the protein and DE contents provided by the CTL diet. Experimental diets were fed ad libitum starting at 27.7 ± 3.4 kg of BW as follows: 3 wk RES, 3 wk OVER, 4 wk RES, and 4 wk OVER. At each diet change, BW and individual feed intakes were measured, and blood samples for metabolite and IGF-I assays were obtained. Some gilts (11 CTL and 12 TRT) were slaughtered on d 235 (after reaching puberty) to collect mammary tissue for compositional analyses and measures of gene expression. Body weight gain (P < 0.01) and G:F (P < 0.05) of gilts were reduced during each period with the RES diet; however, there was no compensatory growth during the periods when the OVER diet was fed. Feeding the RES diet reduced concentrations of urea and IGF-I (P < 0.01) and feeding the OVER diet increased FFA (P < 0.01) and glucose (P < 0.10) in TRT gilts compared with CTL gilts. The TRT gilts had less parenchymal tissue (P < 0.05) and tended to have less total parenchymal fat and protein (P < 0.10) than CTL gilts. The mammary mRNA relative abundance of the signal transducers and activators of transduction 5B was decreased in TRT compared with CTL gilts (P < 0.05). In conclusion, the diet deprivation and over-allowance regimen used in the growing-finishing period did not have beneficial effects on mammary development after puberty. In fact, a detrimental effect was observed.     

哺乳动物乳腺发育调控机制的研究进展

乳腺是哺乳动物特有器官之一,其发育可分为胚胎期、青春期、妊娠期、哺乳期和退化期共5个阶段,发育过程主要受激素、生长因子及细胞因子等调控。乳腺发育主要在出生后,青春期经生长激素(growth hormone, GH)、雌激素(estrogen, E)和胰岛素样生长因子1(insulin-like growth factor 1, IGF1)等调控启动分支形态发生;妊娠期及哺乳期由孕激素(progesterone, P)和催乳素(prolactin, PRL)共同作用产生腺泡并分泌乳汁;退化期则由断奶启动退化过程,腺体重塑至妊娠前状态。乳腺功能主要是通过产生和分泌乳汁以维持初生幼畜正常发育,研究乳腺发育调控机制对畜牧生产具有重要意义。本文以小鼠为例介绍其乳腺发育过程及发育调控机制,并阐述研究乳腺发育及人工调控乳腺发育对畜牧生产的深刻意义,为后期进一步研究乳腺发育和初乳形成提供一定理论参考。     

催乳素及其受体对乳腺发育研究进展

本文主要对催乳素受体的结构、作用机理、功能及其在不同时期的乳腺发育的不同作用进行的论述。总结了近几年来的有关研究进展,为进一步的研究工作提供参考资料。     

Progesterone and Luteinizing hormone secretion patterns in early pregnant gilts

We studied luteinizing hormone (LH) pulsatility and episodic progesterone release of the corpus luteum (CL) on Day 11 and Day 21 in inseminated gilts and aimed to establish a relationship between these two hormones. Blood was collected at 15-min intervals for 12 hr on Days 11, 16 and 21 from a vena cava caudalis catheter. At euthanasia, eight gilts were pregnant and six gilts were not pregnant. Progesterone parameters (basal, mean, pulse frequency and pulse amplitude) did not differ between pregnant and non-pregnant gilts on Day 11, LH pulse frequency and amplitude tended to differ (p = .07 and p = .079). In pregnant gilts, basal and mean progesterone, progesterone pulse amplitude and frequency declined significantly from Day 11 to Day 21 (p < .05). A significant decline was also seen in the LH pulse amplitude from Day 11 to Day 21 (p < .05). None of the LH pulses was followed by a progesterone pulse within 1 hr on Day 21. On Day 11 and Day 21 appeared a synchronicity in the LH pulse pattern, as there were two or three LH pulses in 12 hr and these LH pulses appeared in the same time window. We conclude that on Day 11 and Day 21 of pregnancy in gilts, progesterone pulses do not follow an LH pulse within one hour. Further we demonstrated that the successful or not successful formation of a CL of pregnancy is independent of progesterone release on Day 11 after insemination. We confirmed the decline of progesterone from Day 11 to Day 21 in the vena cava caudalis and could demonstrate that this decline is partly due to lower progesterone pulse amplitude and frequency and that the decline occurs simultaneously with a decline in LH pulse amplitude.     

Canine mammary gland tumors

An understanding of the known biologic facts of this disease and methodical evaluation of the individual patient are necessary prerequisites for outlining rational courses of therapy for dogs afflicted with mammary tumors. Because of the common occurrence of multiple tumors arising at various stages of development simultaneously and the heterogeneity of histology of the complex (mixed tissue types) tumors, presurgical biopsy is recommended only in cases in which mast cell tumor or anaplastic carcinoma is suspected. Although investigative work is being performed regarding the efficacy of chemotherapy, radiation therapy, and immunotherapy, surgery still remains the mainstay in treatment of this condition. No one surgical procedure fits the needs of all patients, although it seems logical to remove as much breast tissue as is reasonable in each circumstance owing to the multicentric nature of the disease. Ovariohysterectomy has not been demonstrated to be of value in treatment of dogs with mammary tumors, but it is a markedly effective method of preventing mammary tumors if it is performed before puberty; it is moderately effective if performed before the dog is 21/2 years of age.     

Gestational exposure to nonylphenol causes precocious mammary gland development in female rat offspring

This study examined whether or not exposure to 4-nonylphenol (NP) during late gestation affects reproductive and mammary development in the offspring of female rats. Time pregnant Long Evans rats were gavaged with NP (10 or 100 mg/kg), atrazine (ATR, 100 mg/kg), or corn oil on gestation days 15-19. The uterus weights of the NP (100 mg/kg/d)-exposed pups were higher than those of the controls but the weights of the other organs were unchanged. Delayed mammary gland (MG) development was detected in the ATR pups on PND 4 and persisted through to PND 66. The high dose NP pups had advanced lobular development of their MG on PND 22, while the glands from the low dose NP pups were no different morphologically from the controls. Immunohistochemical comparisons of the mammary sections from PND 41 demonstrated low levels of estrogen receptor (ER) staining in the control gland stroma and epithelium but higher levels in the tissue of the pups exposed to NP and ATR. ATR also elevated ER in the stroma surrounding the epithelial layer of the terminal end buds. The level of progesterone receptor (PR) staining was markedly lower in the epithelium of the 100 mg/kg NP glands vs. the control glands. However, PR was present at high levels in the epithelium of the 10 mg/kg NP glands and was even more prominent in the ATR-exposed ductal epithelium and fat cell nuclei. The level of prolactin staining was only elevated in glands containing lobule areas (NP-exposed) compared with the control levels. These results suggest that NP and ATR have opposite effects on the development of MG after gestational exposure. Exposure to them during the critical period of epithelial outgrowth altered the receptor levels of mammary progesterone and prolactin and might contribute to the differences in the mammary morphology at PND 41.     

Endocrine effects of GnRH agonist application to early pregnant gilts

The hypothesis of the present study was that a GnRH agonist application at early pregnancy would alter the pattern of the key reproductive hormones LH and FSH, and subsequently that of estradiol (E2) and especially progesterone (P4), and improve the conditions for embryo survival in early pregnant gilts. Therefore, the endocrine effects of a GnRH agonist (GnRHa) application to gilts (n=11 GnRHa treated, n=9 saline Controls) were studied in blood samples from the Vena cava caudalis. GnRHa injected on Day 12 after insemination induced elevated (P<0.01) LH and FSH levels for at least 180 min. However, subsequent LH concentrations were not altered up to Day 21 of pregnancy. LH pulse number, estimated in 6-h period samples on Days 13, 15 and 17, was not influenced by treatment and pregnancy. LH pulse amplitude was decreased (P<0.05) on Days 13 to 17 in pregnant gilts of both groups, but not in nonpregnant animals. In pregnant GnRHa-treated gilts, the basal LH level was elevated compared with the Controls (P<0.01). Additionally, differences (P<0.05) in basal LH were present between the pregnant and nonpregnant animals. The P4 and E2 secretion pattern was not affected by GnRHa. P4 concentrations increased (P<0.01) from Day 10 to Day 14 regardless of the treatment. P4 revealed a pulse-like pattern, but without a definite relation to the LH pulse characteristics. Also, pregnancy rate (73 vs. 67%) and the number of fetuses (12.8 ± 2.3 vs. 11.6 ± 2.3) were unaffected in the treated and Control gilts, respectively. The present study did not confirm the initial hypothesis that a GnRHa-mediated LH effect could alter ovarian steroid secretion and favorably support early embryo development and pregnancy outcome.