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BCRL Home » Milestones » Development of the human breast

The Studies on the development of the human breast

Lobular structures in whole mount preparations

Using whole mount preparations we have been able to identify terminal ductal structures (Figures 3a,b,c and d; 4a,b and c) and lobular structures, each one representing sequential developmental stages. Lobules type 1 are the most undifferentiated lobular structures because they are present in the immature female breast before menarche (Figures 4b,c and d;5a,b,c and d; and 5a,b,c,d,e and f;). They are composed of clusters of 6 to 11 ductules per lobule.

Figure 3:  Whole mount preparation stained with Toluidine blue. (a, b,c and d) Terminal structures of the breast.  X25.

 

 

 

 

Figure 4 Whole mount preparation Stained with Toluidine blue. (a) Terminal structures of the breast. X10. (b) Terminal structures and lobules type 1(arrow). X10. (c and d) Terminal structures (arrow) and lobules type 1 (double arrows). X25.


Figure 5: Whole mount preparation stained with Toluidine blue. (a, b, c and d).    Lobules type 1 in the
breast. X25.

Figure 6: Whole mount preparation stained with Toluidine blue. (a, b, c, d, e and f).  Lobules type 1 in the breast. X25.

 

 



Figure 7: Whole mount preparation stained with Toluidine blue. (a, b, c, and d).  Lobules type 1 (arrow) and lobules type 2 (double arrows) in the breast. X25.


Lobules type 2 evolve from the Lobules type 1 and have a more complex morphology, being composed of a higher number of ductular structures per lobule (Figure 1.5a,b,c and d). These progress to lobules type 3, which are characterized by having an average of 80 ductules or alveoli per lobule; they are frequently seen in the breasts of women under hormonal stimulation or during pregnancy (Figures 8a,b,c and d; 8a,b,c and d). A fourth type of lobule, Lobule type 4, has been described during the lactational period of the mammary gland but is not found in the breast of nulliparous post-pubertal women. It is considered to be the maximal expression of development and differentiation. Each of these structures has different proliferative activity, the highest being in the structures of the terminal end bud and the lowest in the Lobule 3 (Figure 10). 

 Figure 8: Whole mount preparation stained with Toluidine blue. (a, b, c, and d).  Lobules type 1 (arrow), lobules type 2 (double arrows) and lobules type 3(triple arrows) in the breast. X25.


Figure 9: Whole mount preparation stained with Toluidine blue. (a, b, c, and d).  Lobules type 3 in the breast.

 

 

 

 

Figure 10: Schematic representation of the different structures of the human breast and the proliferative activity, determined as DNA labeling index. (Reprinted with permission of: Russo, J. and Russo, I.H.  Development of Human Mammary Gland  In:  The Mammary Gland Development, Regulation, and Function.  (M.C. Neville and C.W. Daniel, eds.) Plenum Pub. Corp., 1987 pp 67-93.)


The identification and quantization of these lobular structures serves as an important parameter for evaluating the effect of age and reproductive history on the normal development of the gland, as well as the pathological response of the mammary parenchyma to either hormonal stimulation or unknown carcinogenic stimuli.

 

Use of the whole mounts for studying the role of aging and reproductive history in the human breast

Lobular structure Surface areaa No. of alveolar buds/structure

Type 1

0.232 x 10-2 ± 0.090 x 10-2

11.2 ± 6.34

Type 2

0.167 x 10-2 ± 0.035 x 10-2

47.0 ± 17.0

Type 3

0.125 x 10-2 ± 0.029 x 10-2

81.0 ± 16.6

Type 4

0.120 x 10-2 ± 0.050 x 10-2

180.0 ± 20.0

The type and characteristics of lobular structures identified are summarized in Table 1. The comparison between the architecture of the breast of nulliparous and parous women revealed that the breasts of nulliparous women contain almost exclusively lobules type 1, whose percentage remains nearly constant throughout the lifespan of the individual, whereas the same structure in the parous women is in a very low percentage during the younger years, although they increase significantly, reaching the same level as that of the nulliparous women only after the fourth decade of life.  These observations suggest that with aging the lobules type 3 regress to lobules type 1, whereas in nulliparous women lobules type 1 never reached the degree of differentiation found in women that have gone through pregnancy. The possible evolution of the lobular structures in the breasts of nulliparous and parous women is depicted in Figure. 11.

Figure 11: Schematic representation of' breast development based upon relative relative  percentage of lobules present. Nulliparous women’s breasts contain primarily  lobules type I (Lob 1) with some progression to type2 (Lob 2), and only minimal formation of lobules type 3 (Lob 3). Parous women undergo a complete cycle of development through the formation of lobules type 4 (Lob 4) which later regress , (Reprinted with permission of: Russo J, Rivera R, Russo IH (1992) Influence of age and parity on the development of the human breast. Breast Cancer Res Treat 23:211-218)

 

Both age and parity strongly influence the architecture of the mammary gland parenchyma. The breasts of nulliparous women contain more undifferentiated structures such as terminal ducts and lobules type 1, although occasional lobules type 2 and 3 are seen.  In parous women, on the other hand, the predominant structure is the most differentiated lobule type 3. In contrast to the predominant structure in nulliparous women, the lobule type 1, which remains constant throughout the lifespan, the predominant lobule type 3 in parous women peaks during the early reproductive years, decreases after the fourth decade of life.

In the breasts of nulliparous women, lobules type 2 are present in moderate numbers during their early years and decrease sharply after age 23, whereas the number of lobules type 1 remains significantly higher. This observation suggests that a certain percentage of lobules type 1 might have progressed to lobules type 2, but the number of lobules progressing to type 3 remains significantly lower than in the parous woman. In the case of parous women it is interesting to note that a history of parity between the ages of 14 to 20 correlates with a significant increase in the number of lobules type 3, which remain present as the predominant structure until the age of 40, the time at which a decrease in the number of lobule type 3 occurs, probably due to their involution predominantly to lobules type 2. It has been shown by several authors that postmenopausal involution is accompanied by diminution and atrophy of the parenchymal components. Among the changes described in the lobular structures is the increase in intralobuar fibrosis and hyalinization, which are interpreted as a response to the lack of hormonal stimulation.  We have observed that lobules type 1 are present in the breasts of both nulliparous and parous postmenopausal women, although the lobules type 1 found in the breasts of parous women present a higher frequency of hyalinization in the intralobular stroma than the lobules of nulliparous women. During the post-menopausal years, both parous and nulliparous women have breasts with a preponderance of lobules type 1.

Although ductal breast cancer originates in lobules type 1, or terminal ductal lobular units, the epidemiologic observation that nulliparous women exhibit a higher incidence of breast cancer than parous women indicates that lobules type 1 in these two groups of women might be biologically different, or exhibit different susceptibility to carcinogenesis.  The presence of lobules type 1 in the breasts of parous women has also been interpreted as a failure of the mammary parenchyma to respond to the influence of pregnancy and lactation. If this is the case, then parous women could contain in their breasts unstimulated as well as regressed lobules type 1. Therefore, the question that remains to be answered is whether the lobules type 1, which are present in the breasts of parous women, are as sensitive to carcinogenesis as the lobules found in the breasts of nulliparous women. We speculate that the lobules type 1 in the breasts of parous women are terminally differentiated structures that are the consequence of a regressive process.  In addition, the proliferative activity of the lobules type 1 is lower in the breasts of parous women than in the breasts of nulliparous women.

The use of whole mounts in the architectural analysis of the human breast has helped us understand that breast development requires a horizontal study in which all the different phases of growth are taken into consideration. For example, the analysis of breast structures at a single given point, i.e., ages 49-53 years, would lead us to conclude that the breasts of nulliparous and parous women appear identical. However, the phenomena occurring in prior years have imprinted permanent changes in the breast biology that affect the potential of the breast for neoplasia, but are no longer morphologically observable. This horizontal study allowed us to determine that parous women underwent lobular differentiation, where as nulliparous women seldom reached the lobule type 3 stage, This may further explain why parous women are more protected against carcinogenesis. In parallel studies, it has been shown that lobules type 1, 2, and 3 exhibit different cell kinetic characteristics: lobules type 1 and 2 grow faster in vitro and have a higher DNA labeling index and a shorter doubling time than lobules type 3. They also exhibit different susceptibility to carcinogenesis. Lobule types 1 and 2 express higher survival efficiency in vitro when treated with chemical carcinogens like DMBA and MNU, changes that are not manifested by lobule type 3.  We have also shown that during the fourth and fifth decades of life there is a decrease in the number of both lobules type 2 and type 3. We have postulated that lobules type 2 is the site of origin of both lobular hyperplasia and carcinoma in situ. Since it has been reported that the incidence of atypical lobular hyperplasia decreases significantly with advancing age, it is possible to postulate that the observed diminution in lobules type 2 is responsible for the decreased incidence of this type of lesion.  In the experimental system we have found that pregnancy, as well as differentiation of the gland induced by the placental hormone human chorionic gonadotropin (hCG), follows a pattern of differentiation similar to the one reported here.  The data discussed above are further supported by this experimental system, in which it has been observed that full term pregnancy as well as treatment of virgin rats with hCG induce differentiation of the mammary gland which results in protection from chemically induced carcinogenesis.  The stimulus of pregnancy or of exogenous hormones furthers the differentiation of the terminal end bud, stimulating lobule development.  The fact that the protection conferred by these processes is maintained even after the termination of either pregnancy or hormonal treatment clearly indicates that the differentiation induced by these processes is a permanent modification of the biological characteristics of the mammary gland, even though the differentiated structures have regressed to seemingly more primitive conditions.  Collectively, the data presented here establish a baseline for understanding the evolution of glandular development, and how it is influenced by age and parity. This is knowledge of utmost importance for understanding the role of differentiation in the protection of the mammary gland against carcinogenesis. In addition, these data establish well-defined endpoints for studying the response of the mammary gland to hormonal or chemopreventive agents, which could be utilized in modulating the susceptibility of the breast epithelium to carcinogenesis.