Cell to cell interactions and normal mammary gland function

The nonepithelial components of the mammary gland are reviewed and their potential for regulatory cell-cell interactions with the epithelial cells are discussed. Studies undertaken to examine the regulatory potential of mammary stromal fibroblasts using an in vitro cell culture system are presented. The influence was examined of epithelial-fibroblast interactions on estrogenic regulation of progesterone receptor concentration in epithelial cells and on epithelial and fibroblast DNA synthesis. Mammary fibroblasts affect estrogen responsiveness in epithelial cells by two different mechanisms. In the case of progesterone receptor regulation, fibroblasts promote estrogen-dependent increases in the receptor via a substratum effect possibly by the production of collagen type I. By contrast, the fibroblast effect promoting estrogen-dependent cell proliferation requires fibroblasts to be metabolically active and in close contact with the epithelium. Additionally, under coculture conditions, estrogen-dependent stimulation of fibroblast DNA synthesis is also observed, indicating a bidirectional, interactive phenomenon between the two types of cells. It is possible that the modulations in epithelial responsiveness to estrogen that are associated with the presence of mammary fibroblasts in vitro reflect regulatory mechanisms that operate in vivo.     

Approaches to the manipulation of mammary involution

Mammary involution is a gradual process that occurs following cessation of milking. Regression of mammary secretory tissue accompanies dramatic changes in secretion composition during the transition from lactation to involution. Conversely, rapid differentiation of secretory tissue and copious accumulation of colostrum occur as parturition approaches. The duration of the nonlactating period, mammary gland health, and secretory cell response to hormones influence subsequent lactational performance in most species. Manipulation of the bovine mammary gland in an attempt to hasten involution has been studied. The primary objective of these studies was to determine if hastened involution would decrease new intramammary infections during the early nonlactating period. Results of these studies have also led to a more fundamental understanding of events that occur during physiological transition of the mammary gland. Adequate regression, proliferation, and differentiation of mammary secretory epithelium during the nonlactating period of ruminants appear to be essential for maximal milk production during lactation. Factors that interfere with these mechanisms can adversely affect mammary function during the impending lactation. A greater understanding of these processes may provide new approaches for increasing milk production in dairy cattle.     

共轭亚油酸对乳腺脂类代谢和乳腺发育的作用及其机理

共轭亚油酸(Conjugated Linoleic Acid,CLA)是亚油酸具有共轭双键的一组同分异构体,由瘤胃细菌在生物氢化过程中合成,具有抗癌、抗炎症、抗动脉粥样硬化等多种生物学功能。CLA在代谢方面也有调节作用,能影响全身代谢,影响肝脏、乳腺以及富含脂肪的相关组织代谢。在此介绍了CLA的来源、结构、对动物乳腺脂类代谢和乳腺发育的作用及其机理。     

Performance strategies affect mammary gland development in prepubertal heifers

In Brazil, the majority of dairy cattle are Holstein × Gyr (H×G). It is unknown whether excessive energy intake negatively affects their mammary development to the same extent as in purebred Holsteins. We hypothesized that mammary development of H×G heifers can be affected by dietary energy supply. We evaluated the effect of different average daily gains (ADG) achieved by feeding different amounts of a standard diet during the growing period on biometric measurements, development of mammary parenchyma (PAR) and mammary fat pad (MFP), and blood hormones. At the outset of this 84-d experiment, H×G heifers (n = 18) weighed 102.2 ± 3.4 kg and were 3 to 4 mo of age. Heifers were randomly assigned to 1 of 3 ADG programs using a completely randomized design. Treatments were high gain (HG; n = 6), where heifers were fed to gain 1 kg/d; low gain (LG; n = 6), where heifers were fed to gain 0.5 kg/d; and maintenance (MA; n = 6), where heifers were fed to gain a minimal amount of weight per day. Heifers were fed varying amounts of a single TMR to support desired BW gains. Over the 84 d, periodic biometric and blood hormone measurements were obtained. On d 84, all heifers were slaughtered and carcass and mammary samples were collected. At the end, HG heifers weighed the most (181 ± 7.5 kg), followed by LG (146 ± 7.5 kg) and MA (107 ± 7.5 kg) heifers. The ADG were near expected values and averaged 0.907, 0.500, and 0.105 ± 0.03 kg/d for HG, LG, and MA, respectively. In addition, body lengths, heart girths, and withers heights were affected by dietary treatment, with MA heifers generally being the smallest and HG heifers generally being the largest. Body condition scores differed by treatment and were highest in HG and lowest in MA heifers; in vivo subcutaneous fat thickness measurement and direct analysis of carcass composition supported this. The HG heifers had the heaviest MFP, followed by LG and then MA heifers. Amount of PAR was highest in LG heifers and was the same for HG and MA heifers. The percentage of udder mass occupied by PAR was lowest in HG heifers, differing from LG and MA heifers. Composition of MFP was not evaluated. Regarding PAR composition, no differences in ash or DM were found. On the other hand, CP concentration of PAR for HG heifers was lower than that for LG heifers, which was lower than that for MA heifers. Regarding the fat content, HG treatment was higher than LG and MA treatment, which did not differ from each other. In PAR, differences in relative abundance of genes related to both stimulation and inhibition of mammary growth were observed to depend on dietary treatment, sampling day, or both. The same can be said for most of the blood hormones that were measured in this experiment. In this experiment, high ADG achieved by feeding different amounts of a standard diet during the growing period negatively affected mammary development.     

Functional consequence of exposure to dieldrin on mammary development and function.

The effect of dieldrin (Dln) on the development of the mammary gland and on functional parameters of CID-9 mammary cells in culture was investigated. One-month-old Sprague-Dawley female rats were bred and received intraperitoneal injection with 2.5 or 15 microM Dln during the last trimester of their gestation. Mammary glands of 15-microM Dln-treated rats showed immature alveolar structures by day 18 of gestation and abundant adipose tissue. Dln-treated rats had a lower number of pups, and the weight of pups between days 14 and 31 of age compared with non-treated rats was significantly lower. Long-term exposure of CID-9 mammary cells, cultured under non-differentiation conditions, on plastic, or under differentiation permissive conditions, dripped with EHS-matrix, to 5 or 25 microM Dln was detrimental to cell growth. The short-term effect of Dln exposure (up to 9 h) on CID-9 cells, under the same culture conditions, did not affect their beta-casein mRNA levels, but induced apoptosis, down regulated gap junction intracellular communication and induced IL-6 and TNF-alpha expression.     

Regulation of genes encoding proteolytic enzymes during mammary gland development

The mammary gland undergoes extensive tissue remodelling during each lactation cycle. During pregnancy, the epithelial compartment of the gland is vastly expanded (Benaud et al. 1998). At the end of lactation the epithelial cells undergo apoptosis and adipocyte differentiation is induced (Lilla et al. 2002). Ductal and alveolar growth during puberty and pregnancy, and the involution process require the action of proteolytic enzymes (including matrix metalloproteinases, plasminogen and membrane-peptidases) and the corresponding genes are activated during these periods (Benaud et al. 1998; Alexander et al. 2001). Matrix metalloproteinases (MMP) are expressed in several cell types of the mammary gland including stromal fibroblasts (e.g., MMP3, MMP2), epithelial cells (e.g., MMP7 or MMP9), adipocytes (e.g., MMP2) and lymphoid cells (e.g., MMP9) (Crawford et al. 1996; Lund et al. 1996; Wiseman et al. 2003). A number of knock-out mice, which are deficient for individual MMP genes (e.g., MMP2, MMP3) or plasminogen, display alterations to mammary gland structure and impairment of lactation (Lund et al. 1999; Wiseman et al. 2003).     

Effect of heat stress during the dry period on mammary gland development

Heat stress during the dry period negatively affects hepatic metabolism and cellular immune function during the transition period, and milk production in the subsequent lactation. However, the cellular mechanisms involved in the depressed mammary gland function remain unknown. The objective of the present study was to determine the effect of heat stress during the dry period on various indices of mammary gland development of multiparous cows. Cows were dried off approximately 46 d before expected calving and randomly assigned to 2 treatments, heat stress (HT, n = 15) or cooling (CL, n = 14), based on mature equivalent milk production. Cows in the CL treatment were provided with sprinklers and fans that came on when ambient temperatures reached 21.1°C, whereas HT cows were housed in the same barn without fans and sprinklers. After parturition, all cows were housed in a freestall barn with cooling. Rectal temperatures were measured twice daily (0730 and 1430 h) and respiration rates recorded at 1500 h on a Monday-Wednesday-Friday schedule from dry off to calving. Milk yield and composition were recorded daily up to 280 d in milk. Daily dry matter intake was measured from dry off to 42 d relative to calving. Mammary biopsies were collected at dry off, −20, 2, and 20 d relative to calving from a subset of cows (HT, n = 7; CL, n = 7). Labeling with Ki67 antigen and terminal deoxynucleotidyl transferase deoxyuridine triphosphate nick-end labeling were used to evaluate mammary cell proliferation and apoptosis, respectively. The average temperature-humidity index during the dry period was 76.6 and not different between treatments. Heat-stressed cows had higher rectal temperatures in the morning (38.8 vs. 38.6°C) and afternoon (39.4 vs. 39.0°C), greater respiration rates (78.4 vs. 45.6 breath/min), and decreased dry matter intake (8.9 vs. 10.6 kg/d) when dry compared with CL cows. Relative to HT cows, CL cows had greater milk production (28.9 vs. 33.9 kg/d), lower milk protein concentration (3.01 vs. 2.87%), and tended to have lower somatic cell score (3.35 vs. 2.94) through 280 d in milk. Heat stress during the dry period decreased mammary cell proliferation rate (1.0 vs. 3.3%) at −20 d relative to calving compared with CL cows. Mammary cell apoptosis was not affected by prepartum heat stress. We conclude that heat stress during the dry period compromises mammary gland development before parturition, which decreases milk yield in the next lactation.     

Expression and localization of extracellular matrix-degrading proteinases and their inhibitors in the bovine mammary gland during development, function, and involution

In degrading the extracellular matrix, matrix metalloproteinases (MMP) and the plasminogen activator (PA) system may play a critical role in extensive remodeling that occurs in the bovine mammary gland during development, lactation, and involution. Therefore, the aim of our study was to investigate the mRNA expression of MMP-1, MMP-2, MMP-14, MMP-19, tissue inhibitor of metalloproteinases (TIMP)-1, TIMP-2, urokinase-type PA, tissue-type PA, urokinase-type PA receptor, and PA inhibitor-1 by quantitative PCR and to localize with immunohistochemistry MMP-1, MMP-2, MMP-14, and TIMP-2 proteins in the bovine mammary gland during pubertal mammogenesis, lactogenesis, galactopoiesis, and involution. Expression of mRNA for each of the studied factors was relatively lower during galactopoiesis and early involution but was markedly increased during mammogenesis and late involution, 2 stages in which tissue remodeling is especially pronounced. The localization of proteins for MMP-1, MMP-14, and TIMP-2 showed a similar trend with strong staining intensity in cytoplasm of mammary duct and alveolar epithelial cells during pubertal mammogenesis and late involution. Interestingly, MMP-2 protein was localized only in the cytoplasm of endothelial cells during late involution. Our study demonstrated clearly that expression of extracellular matrix-degrading proteinases coincides with a concomitant expression of their inhibitors. High expression levels of MMP, TIMP, and PA family members seem to be a typical feature of the nonlactating mammary gland.     

IGFs对小鼠乳腺发育及细胞凋亡的影响

为揭示IGFs在乳腺发育和乳腺上皮细胞凋亡中的作用，运用荧光显微镜及光镜技术，采用小鼠乳腺组织培养及小鼠乳腺上皮细胞培养方法，对IGFs的作用进行了研究。结果表明，IGFs可促进小鼠乳腺导管的生长发育，并且IGF—I促小鼠乳腺导管生长发育的功能强于IGF—II；IGFs抑制泌乳期小鼠乳腺上皮细胞凋亡，并且IGF—I抑制凋亡的作用强于IGF—II。     

奶牛不同发育时期乳腺细胞凋亡规律的研究

为系统地研究奶牛乳腺不同发育时期细胞凋亡的规律,应用TUNEL法对奶牛不同发育时期正常乳腺组织(冰冻切片)的乳腺细胞凋亡进行了系统检测。结果表明,青春期乳腺发育缓慢,结构变化较小,乳腺细胞凋亡量相对较少;妊娠期乳腺导管持续发育,凋亡量上升,其中妊娠初期2月,乳腺腺泡大量出现,脂肪细胞凋亡随之增加,出现了一个高峰;泌乳期乳腺的结构和功能最为完善,乳腺结构变化很小,细胞凋亡量维持在很低水平;退化期腺泡瓦解,大量细胞发生凋亡,其中退化初期乳腺细胞凋亡持续增加,退化3 d达到最大值,之后凋亡量逐渐降低,退化30 d后乳腺已经基本恢复到青春期状态,凋亡细胞量也随之减少。     

Autotransplantation of the goat mammary gland

The modified technique for transplanting one mammary gland of a lactating goat to the neck, with the mammary (pudic) artery and vein anastomosed to the carotid artery and jugular vein respectively, is described. This technique was successful in 6 operations and there were no significant differences between the milk yield of the transplanted and control (in situ) glands at any stage after operation. These results are compared with those from an earlier series in which no special provision was made for lymphatic drainage and in which anticoagulant therapy and treatment of the gland prior to attachment were not standardized.     

小鼠乳腺中FGF7及其受体的表达与作用

为了从蛋白水平研究成纤维细胞生长因子7（FGF7）及其受体（KGFR）在哺乳动物乳腺发育、泌乳及退化过程中表达定位的动态变化,揭示FGF7及其受体表达变化与乳腺发育及泌乳功能的关系,本研究利用激光扫描共聚焦显微系统对昆明鼠乳腺中FGF7及其受体的表达进行了免疫荧光检测,利用体外培养不同发育时期的小鼠乳腺组织研究了FGF7在乳腺中的作用。结果表明：FGF7主要定位于乳腺导管上皮细胞以及腺泡上皮细胞外周,同时在基质中有弱的表达。KGFR主要出现在导管上皮细胞以及腺泡上皮细胞上,在乳腺发育的个别时期细胞外基质中有弱表达。FGF7在退化4d达到最高值;KGFR在泌乳4d达到最高值。除青春期和妊娠期外,添加FGF7均能显著诱导乳腺上皮细胞产生KGFR。FGF7能促进小鼠乳腺上皮细胞的增殖分化,降低乳腺上皮细胞的细胞凋亡。     

奶山羊乳腺发育过程中腺苷酸质量分数及能荷的变化

探讨了奶山羊乳腺发育和泌乳过程中能量代谢变化规律和特点.分别在奶山羊乳腺发育的4个时期青春期、妊娠期、泌乳期及退化期采集乳腺组织.应用高效液相色谱测定各时期的腺苷酸质量分数并计算出各时期能荷:用总ATP酶活性测定试剂盒测定各时期的ATP酶活性.结果表明,青春期和妊娠期腺苷酸质量分数和ATP酶活性呈缓慢递增趋势,泌乳启动后ATP酶活性急剧增加,腺苷酸质量分数增加,ADP、AMP比例升高.在泌乳高峰期,AMP达到峰值,能荷值最低.泌乳高峰期过后ATP酶活性下降,腺苷酸质量分数减少,AMP比例急剧减少,能荷值恢复到正常水平.在乳腺发育过程中,泌乳启动后乳腺ATP酶活性急剧增加,腺苷酸质量分数增加;泌乳高峰期过后ATP酶活性下降,腺苷酸质量分数减少.     

Organization and growth of mammary epithelia in the mammary gland fat pad

Mammary gland development consists of a series of very highly ordered events involving interactions among a number of distinct cell types. An important aspect of mammary gland development is that the mammary gland consists of a fat pad of mesodermal origin into which epithelial cells of ectodermal origin proliferate. This proliferation of epithelial cells into the mammary fat pad is the subject of this review. The nature of the stroma into which epithelial cells proliferate is of considerable importance in determining the structure of the resulting gland. In mice, white adipose tissue appears to be required for normal mammary development. Transplantation of mammary epithelia to other types of stroma does not support epithelial growth or result in abnormal growth. To date, a synthetic substratum capable of mimicking white adipose tissue has not been developed. Although collagen gel cultures are generally considered superior to glass or plastic substratum in supporting near normal epithelial growth, the technique has not advanced to the point that the in vivo growth pattern is duplicated. Recent research on the generation of chimeric mammary tissue (by transplanting mammary epithelia from rats, cows, and women to the mammary fat pads of athymic nude mice) suggests that there are important species differences in the stromal requirements for mammary gland development. In particular, extensive and expansive growth of rat mammary tissue is observed in mouse mammary fat pads. However, the mouse mammary fat pad appears incapable of supporting expansive growth of bovine or human mammary epithelia. The reason for this difference is not clear. However, human and bovine mammary epithelia may require the presence of more fibrous (collagenous) tissue than rodent mammary epithelia for normal proliferation.