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G H Smith

Publications and source records attributed to G H Smith.

At least 37 records · Page 2Linked to original sources
PubMed1999

Mammary epithelial stem cells: our current understanding.

It has recently been shown that the progeny from a single cell may comprise the epithelial population of a fully developed lactating mammary outgrowth in mice. Serial transplantation of epithelial fragments from this clonally derived gland demonstrates that the subsequently generated outgrowths are also comprised of progeny from the original antecedent. Similarly, genetic analysis of contiguous portions of individual human mammary ducts within the same breast indicates their clonal derivation. These observations support the concept that multipotent tissue-specific epithelial stem cells are present among the parenchymal cells of the mammary gland. Here, we present the developing evidence for the presence of stem cells in virtually every renewing mammalian tissue as well as some classically considered to consist only of differentiated cells. Further, we review the present morphologic and biologic evidence for stem cells and lineage-limited progenitor cells in human and rodent mammary epithelium. Although a number of selective markers are known for various lineage-limited hematopoietic cells and their progeny, our understanding of the biology of the precursor cells for mammary epithelium is just beginning. Our purpose here is to develop further interest in the clarification of these issues in the biology of the mammary gland.

Aging↗
PubMed1999

Intra-ocular pressure changes during maximal isometric contraction: does this reflect intra-cranial pressure or retinal venous pressure?

Recent publications have suggested that intra-ocular pressure (IOP) may be an indirect assessment of intra-cranial pressure (ICP). Both IOP and ICP have similar physiologic pressure ranges and similar responses to changes in intra-abdominal, intra-thoracic and aortic pressure. Previous studies have demonstrated the relationships between retinal arterial pressure and aortic pressure, intra-ocular pressure and retinal venous pressure, intra-cranial pressure and retinal venous pressure. Power athletes routinely utilize the Valsalva maneuver during weightlifting. In fact there are reports of stroke, cerebral hemorrhage, subarachnoid hemorrhage, conjunctival, foveal and retinal hemorrhage, retinal detachment, hiatal hernia and pneumothorax associated with weightlifting. These events are thought to occur secondary to the extreme pressure elevations that occur in the intra-abdominal, intra-thoracic, intra-cranial, intra-ocular and vascular compartments. To date no human studies have examined the IOP changes that may occur with heavy resistance exercise. Therefore, we recruited power athletes (n = 11), who had participated in prior studies, from the local metropolitan area. The athletes had blood pressure status, drug screening and medical histories performed during previous investigations. Intra-ocular pressure was measured by noncontact tonometry at rest and during maximal isometric contraction. All subjects resting IOP were within normal ranges (mean 13 +/- 2.8 mmHg). Intra-ocular pressures were significantly (p < 0.0001) elevated in each subject during maximal contraction (mean 28 +/- 9.3 mmHg). One subject's IOP reached 46 mmHg during maximal contraction. Linear regression analysis demonstrated a significant linear relationship (r = 0.62, p < 0.0001) in the net change of IOP from rest to maximal contraction for each subject. This study demonstrates that IOP elevates to pathophysiologic levels during resistance exercise. The findings of conjunctival hemorrhages in two subjects further supports IOP being reflective of retinal venous pressure. The enormous pressures generated by power athletes during weightlifting leads to elevations in ICP which obstruct venous outflow leading to hemorrhage and elevations in IOP. The question remains as to whether these intermittent bursts of elevated IOP can lead to long-term pathological sequelae.

Adult↗
PubMed1998

An entire functional mammary gland may comprise the progeny from a single cell.

Any epithelial portion of a normal mouse mammary gland can reproduce an entire functional gland when transplanted into an epithelium-free mammary fat pad. Mouse mammary hyperplasias and tumors are clonal dominant populations and probably represent the progeny of a single transformed cell. Our study provides evidence that single multipotent stem cells positioned throughout the mature fully developed mammary gland have the capacity to produce sufficient differentiated progeny to recapitulate an entire functional gland. Our evidence also demonstrates that these stem cells are self-renewing and are found with undiminished capacities in the newly regenerated gland. We have taken advantage of an experimental model where mouse mammary tumor virus infects mammary epithelial cells and inserts a deoxyribonucleic acid copy(ies) of its genome during replication. The insertions occur randomly within the somatic genome. CzechII mice have no endogenous nucleic acid sequence homology with mouse mammary tumor virus; therefore all viral insertions may be detected by Southern analysis provided a sufficient number of cells contain a specific insertional event. Transplantation of random fragments of infected CzechII mammary gland produced clonal-dominant epithelial populations in epithelium-free mammary fat pads. Serial transplantation of pieces of the clonally derived outgrowths produced second generation glands possessing the same viral insertion sites providing evidence for self-renewal of the original stem cell. Limiting dilution studies with cell cultures derived from third generation clonal outgrowths demonstrated that three multipotent but distinct mammary epithelial progenitors were present in clonally derived mammary epithelial populations. Estimation of the potential number of multipotent epithelial cells that may be evolved from an individual mammary-specific stem cell by self-renewal is in the order of 10(12)-10(13). Therefore, one stem cell might easily account for the renewal of mammary epithelium over several transplant generations.

Animals↗
PubMed1998

Surgical management of the nonpalpable testis: the Children's Hospital of Philadelphia experience.

PURPOSE: The management of the nonpalpable testis permits an individualized operative approach. We analyze the results of surgical management of a large series of patients with a nonpalpable testis. MATERIALS AND METHODS: Between January 1986 and June 1994 we treated 1,866 boys with undescended testes. There were 447 testes (24%) that were not palpable at presentation. Intra-operative data on these patients were collected for age at presentation, bilateral testes position, testicular size, associated inguinal anomalies (vas, epididymis and processus vaginalis) and surgical approach. For intra-abdominal testes postoperative results of 2 surgical techniques, the Fowler-Stephens procedure and Koop orchiopexy (retroperitoneal mobilization of spermatic vessels and vas) were compared in 76 patients with at least 18 months of followup. RESULTS: Average patient age at presentation was 34 months with 63% presenting before age 48 months. Of the impalpable testes 58% were on the left side, 35% were on the right side and 7% were bilateral. At operation 181 testes (41%) were atrophic or absent, 91 (20%) were intra-abdominal with 14 (3.1%) bilateral, 136 (30%) were in the inguinal canal and 39 (9%) were in other locations, including 22 at the pubic tubercle, 2 in the upper scrotum, 13 in the superficial inguinal pouch and 2 in the perineum. Of the intra-abdominal group associated extratesticular malformations were identified in 36 cases (39%). Attachment of the vas deferens to the testis was abnormal in 23 of 64 cases (36%), including 10 that were completely detached and 13 with head or tail attachment only. Of the 91 evaluable cases in the intra-abdominal group 38 (42%) had been treated with the Fowler-Stephens repair (5 in 2 stages), 33 (36%) with inguinal orchiopexy and intraperitoneal dissection without dividing the spermatic vessels, 5 with 2-stage procedures and vessel preservation and 14 (15%) with orchiectomy. One testis was left in situ. The inguinal approach with intraperitoneal extension was successful in defining the testis location or blind-ending vas and vessels in 100% of the cases. A single operation to perform orchiopexy was successful in 92% of the cases. Overall, results were considered excellent or acceptable in 32 of 33 cases (97%) after Koop orchiopexy and 28 of 38 (74%) after the Fowler-Stephens orchiopexy. CONCLUSIONS: Nonpalpable testes accounted for 24% of the patients presenting with undescended testes. At surgical exploration 39% of impalpable testes were distal to the external inguinal ring, 41% were atrophic or absent and 20% were intra-abdominal. All cases were treated through a standard inguinal incision. These data provide evidence that the inguinal approach to orchiopexy with transperitoneal mobilization of the vas and vessels without transection is highly successful for the intra-abdominal cryptorchid testis and, to date, is the preferred technique for the management of the intra-abdominal undescended testis.

Child, Preschool↗
PubMed1998

Functional rescue of Stat5a-null mammary tissue through the activation of compensating signals including Stat5b.

Prolactin induces mammopoiesis and lactogenesis through the Janus kinase-signal transducers and activators of transcription pathway, with Stat5a being a principal and obligate cytoplasmic and nuclear signaling molecule. Mice from which the Stat5a gene has been deleted fail to develop functional mammary tissue during their first pregnancy. Lobuloalveolar outgrowth is curtailed, and epithelial cells fail to progress to functional differentiation. Here, we investigate whether the effect of Stat5a deficiency is restricted to the epithelium and whether the gland has the capacity to activate alternative signaling pathways that could restore development and function. Mammary gland transplant experiments showed that Stat5a-deficient epithelium does not differentiate in wild-type stroma, thus demonstrating a cell-autonomous role for Stat5a. The capacity of Stat5a-deficient mammary tissue to develop and secrete milk was measured after consecutive pregnancies and with postpartum suckling. Neither of these regimens could independently restore lactation. However, the combination of several pregnancies and suckling stimuli resulted in a partial establishment of lactation and an increase of Stat5b activity. These experiments demonstrate that the mammary gland has inherent plasticity that allows it to use different signals to achieve its ultimate purpose, the production of milk to nurture newborn offspring.

Animals↗
PubMed1997

Cripto enhances the tyrosine phosphorylation of Shc and activates mitogen-activated protein kinase (MAPK) in mammary epithelial cells.

Cripto-1 (CR-1), a recently discovered protein of the epidermal growth factor (EGF) family, was found to interact with a high affinity, saturable binding site(s) on HC-11 mouse mammary epithelial cells and on several different human breast cancer cell lines. This receptor exhibits specificity for CR-1, since other EGF-related peptides including EGF, transforming growth factor alpha, heparin-binding EGF-like growth factor, amphiregulin, epiregulin, betacellulin, or heregulin beta1 that bind to either the EGF receptor or to other type 1 receptor tyrosine kinases such as erb B-3 or erb B-4 fail to compete for binding. Conversely, CR-1 was found not to directly bind to or to activate the tyrosine kinases associated with the EGFR, erb B-2, erb B-3, or erb B-4 either alone or in various pairwise combinations which have been ectopically expressed in Ba/F3 mouse pro-B lymphocyte cells. However, exogenous CR-1 could induce an increase in the tyrosine phosphorylation of 185- and 120-kDa proteins and a rapid (within 3-5 min) increase in the tyrosine phosphorylation of the SH2-containing adaptor proteins p66, p52, and p46 Shc in mouse mammary HC-11 epithelial cells and in human MDA-MB-453 and SKBr-3 breast cancer cells. CR-1 was also found to promote an increase in the association of the adaptor Grb2-guanine nucleotide exchange factor-mouse son of sevenless (mSOS) signaling complex with tyrosine-phosphorylated Shc in HC-11 cells. Finally, CR-1 was able to increase p42(erk-2) mitogen-activated protein kinase (MAPK) activity in HC-11 cells within 5-10 min of treatment. These data demonstrate that CR-1 can function through a receptor which activates intracellular components in the ras/raf/MEK/MAPK pathway.

Animals↗
PubMed1997

Three division-competent, structurally-distinct cell populations contribute to murine mammary epithelial renewal.

Markers for the division-competent cells in mammary gland epithelium are important to the understanding of normal and neoplastic mammary epithelial growth and architectural maintenance, but have been difficult to reveal. Using the presence of condensed chromosomes as an indicator of division competence, we have described morphological markers in the nucleus and cytoplasm that reliably characterize two sets of structurally-distinct, division-competent cells in murine (i.e. mouse and rat) mammary epithelium. The markers, based, in part, on cell size, nuclear and cytoplasmic staining characteristics, and distinctive morphological features of the nucleus and cytoplasm allow classification of the division-competent cells into two categories: 'large light cells' and 'small light cells'. Based on the degree of cytoplasmic organelle differentiation, the small light cells are the least differentiated cells in the epithelium, and the large light cells appear, structurally and functionally, to be in the early stages of secretory differentiation. We demonstrate, using statistical analysis of cell counts (per unit area of epithelium) from four stages of mammary epithelial growth, that there are, in fact, three division-competent cell populations in the rat mammary epithelium, and that the large light cell is a direct precursor to terminally differentiated cells, both secretory and myoepithelial. Using our results we synthesize a morphological model of cell mitosis and the progression of epithelial differentiation in murine mammary gland from a stem cell through two progenitors.

Animals↗
PubMed1997

Cripto-1 inhibits beta-casein expression in mammary epithelial cells through a p21ras-and phosphatidylinositol 3'-kinase-dependent pathway.

Cripto-1 (CR-1) is a recently discovered protein of the epidermal growth factor family that does not directly activate any of the known erbB type 1 tyrosine kinase receptors. Also, CR-1 stimulates the growth of HC-11 mouse mammary epithelial cells. We found that prior treatment of HC-11 cells with exogenous CR-1 induced a competency response to the lactogenic hormones dexamethasone, insulin, and prolactin (DIP) with respect to the induction of the milk protein beta-casein. In contrast, simultaneous treatment of mouse HC-11 cells with CR-1 in the presence of DIP inhibited beta-casein expression. The inhibitory effects of CR-1 on beta-casein expression in response to DIP were not unique to this mouse mammary epithelial cell line, because beta-casein and whey acidic protein expression in primary mouse mammary explant cultures established from midpregnant mice were also differentially inhibited by several epidermal growth factor-related peptides including CR-1. The mitogenic and differentiation effects of CR-1 are mediated by the binding of CR-1 to a cell surface receptor that is known to activate the ras/raf/mitogen-activated protein kinase (MAPK)/MAPK kinase pathway. The inhibitory response of CR-1 in HC-11 cells on beta-casein expression after treatment with DIP can be attenuated by B581, a peptidomimetic farnesyltransferase inhibitor that blocks p21ras farnesylation and activation, and by the phosphatidylinositol 3'-kinase (PI3k) inhibitor LY 294002 but not by PD 98059, a MAPK kinase inhibitor that blocks MAPK activation. These data suggest that the ability of CR-1 to block lactogenic hormone-induced expression of beta-casein is mediated through a p21ras-dependent, PI3k-mediated pathway. This is further substantiated by the observation that CR-1 is able to stimulate the tyrosine phosphorylation of the p85 PI3k regulatory subunit and to increase the activity of PI3k in HC-11 cells.

Animals↗
PubMed1996

Communication breakdown: a preventable cause of acute renal failure in a newborn infant.

A three-week-old boy had life-threatening renal failure which was successfully treated by salbutamol and sodium bicarbonate infusion, rectal resonium and a left percutaneous nephrostomy. Ultrasonography of this child at 19 weeks' gestation had shown a treatable renal tract abnormality (bilateral hydronephrosis). No arrangements had been made for follow-up.

Acute Kidney Injury↗
PubMed1996

Expression of a truncated Int3 gene in developing secretory mammary epithelium specifically retards lobular differentiation resulting in tumorigenesis.

Insertional mutation of the Int3 gene, a member of the Notch gene family, is frequently associated with primary mouse mammary tumors induced by the mouse mammary tumor virus (MMTV). A major consequence of these mutations is the production of a shortened 2.4-kb tumor specific Int3 RNA transcript that encodes the entire intracellular domain of the Int3 protein. Previous studies have demonstrated that mammary gland development and function was severely impaired in transgenic mice expressing the truncated Int3 gene product from the MMTV viral promoter. Both mammary ductal growth and secretory lobule development were curtailed in these mice. These results were attributed to a gain of function modification of the Int3 gene, which led to a restriction of cell fate selection in the affected mammary epithelial cells. To confirm and extend these findings, truncated Int3 was expressed from the whey acidic protein (WAP) promoter, the activity of which, unlike that of the MMTV long terminal repeat, is restricted to the secretory mammary epithelial population. In transgenic mice carrying the WAP/Int3 construct, mammary ductal growth was unaffected in virgin females, but growth and differentiation of secretory lobules during gestation was profoundly inhibited. Coincidental with the block in lobular secretory differentiation, mammary dysplasia and tumorigenesis occurred in all breeding females by 25 weeks of age. In nonbreeding WAP/Int3 females, mammary tumor incidence also reached 100%, but only after 70 weeks. The WAP/Int3 mammary tumors were highly malignant, and most tumor-bearing females, irrespective of breeding history, developed metastatic lung lesions. These results suggest that WAP promotor-targeted Int3 function is associated with mammary secretory cell differentiation and maintenance in this transgenic model. Consistent with the conclusion that WAP-driven truncated Int3 expression influenced only lobular differentiation and not ductal growth and extension during mammary gland development, transplants of WAP/Int3 gland into nontransgenic mammary fat pads produced complete mammary ductal outgrowths in virgin FVB/N mice but failed to develop secretory lobules when the females were impregnated.

Animals↗
PubMed1996

Understanding mammary gland development through the imbalanced expression of growth regulators.

Functional differentiation of mammary tissue progresses in distinct phases spanning puberty and pregnancy. Here we have analyzed and compared the effects of transforming growth factor beta 1 (TGF beta 1), TGF alpha, and whey acidic protein (WAP), the Notch-related cell fate protein Int3, and p53 and pRb on mammary development. We chose transgene expression from the WAP gene promoter which is only active in mammary alveolar cells. The imbalanced expression of these molecules specifically altered development and differentiation of the gland. While TGF alpha did not disturb alveolar outgrowth, little or no alveolar structures developed in the presence of Int3. TGF beta 1, WAP, and the expression of SV40 T-antigen-which inactivates p53 and PRb-reduced overall alveolar development. The expression of individual milk protein genes was affected differentially by the transgenes. A WAP-lacZ transgene served as an additional indicator of terminal differentiation of alveolar cells, Homogeneous expression of lacZ was seen in mice transgenic for lacZ, or for TGF alpha and lacZ. In contrast, only a few differentiated cells were observed in the presence of TGF beta 1 and Tag. Thus, the expression of growth regulators in the same defined subset of mammary cells results in distinct developmental changes and a specific pattern of alveolar differentiation.

Animals↗
PubMed1996

Detection of amphiregulin and Cripto-1 in mammary tumors from transgenic mice.

Epidermal growth factor family members are widely expressed in human breast cancer and are thought to play an important dual role in mammary gland development and tumorigenesis. Overexpression of two relatively new members of this family, amphiregulin (AR) and Cripto-1 (CR-1), has been previously shown to transform immortalized human and mouse mammary epithelial cells. Here, we extend these results and address the disregulated expression of AR and CR-1 in many types of transgenic neoplastic mouse mammary tissues. Transgenic mouse strains overexpressing the oncogenes transforming growth factor-alpha, neu, int-3, polyoma virus middle T antigen, and simian virus 40 large T antigen have been previously shown to develop spontaneous mammary neoplasia. These models were each examined for mammary-tumor expression of AR and CR-1 by reverse transcription-polymerase chain reaction, western blot, and immunocytochemical analyses. Mammary tumors from each source expressed AR and CR-1. Western blot analysis revealed that, in all mammary tumors, AR and CR-1 protein species were processed differently than in virgin and lactating mouse mammary tissue. In addition, immunohistochemical detection of AR and CR-1 in tumor tissue revealed different patterns of growth-factor localization in different types of transgenic mouse mammary-derived tumors. These findings are consistent with the possibility of widespread roles for AR and CR-1 in the promotion and/or progression stages of mouse mammary tumorigenesis.

Amphiregulin↗
PubMed1996

Experimental mammary epithelial morphogenesis in an in vivo model: evidence for distinct cellular progenitors of the ductal and lobular phenotype.

An in vivo transplantation system has been used to evaluate the developmental capacities of specific mouse mammary epithelial cell populations. Specifically, mouse mammary epithelial cells with distinctly limited developmental potentials have been identified using this procedure. Two distinct epithelial cell progenitors have been identified by experiments designed to determine whether basal lobular and ductal phenotypes could develop independently under conditions imposed by a limiting dilution. The prediction that these separate epithelial progenitors must exist was based upon the results from transplantation experiments carried out in epithelium-divested mammary fat pads of syngeneic mice with mammary epithelium from two different transgenic mouse models. The results presented here demonstrate the following points: 1) lobular, i.e. secretory, progenitor cells are present as distinct entities among the mammary epithelial cells found in immature virgin female mice; 2) similarly, ductal epithelial progenitors are present within the same population; 3) lobular progenitors are present in greater numbers, although both cell populations are extremely small; 4) as expected, some inocula produce outgrowths with simultaneous development of both lobular and ductal phenotypes--it is not known whether this indicates cooperative interaction between the two epithelial progenitors or signals the presence of a third progenitor type capable of producing both ductular and lobular committed daughters; 5) these findings have important consequences in the design of experiments aimed at testing the effects of known and putative mammary oncogenes and tumor suppressor genes, using techniques which include cellular transformation in vitro followed by in vivo cultivation and evaluation.

Adipose Tissue↗
PubMed1996

TGF-beta and functional differentiation.

A review of the pertinent literature suggests that TGF-beta 1 may play a multifaceted role in functional differentiation of mammary epithelium. Evidence for the expression of TGF-beta 1 RNA and the presence of functional TGF-beta 1 protein in differentiating mammary epithelial cells from a pregnant mouse has been recently reported. The specific role of mammary-epithelial-cell-produced TGF-beta 1 in the differentiating mammary gland is presently unclear. However, several possible functions are suggested from the following observations. Milk protein production is negatively regulated by exogenous TGF-beta 1 during gestational development of the gland but not during lactation. Consistent with reports linking TGF-beta 1 gene expression with mammary gland involution following lactation, overexpression of TGF-beta 1 in the differentiating secretory epithelium leads to premature programmed cell death in the absence of a negative effect on secretory epithelial cell proliferation. A role for TGF-beta 1 in cell cycle control and suppression of malignant progression independent from its inhibitory effect on epithelial cell growth has been demonstrated in keratinocytes. A similar function could provide protection against malignancy in proliferating mammary epithelium and account for TGF-beta 1 suppression of mammary tumorigenesis in transgenic mice overexpressing transforming growth factor alpha (TGF-alpha).

Animals↗
PubMed1996

The long-term outcome of posterior urethral valves treated with primary valve ablation and observation.

PURPOSE: We believe that primary valve ablation with observation is the preferred management for posterior urethral valves. However, debate continues as to the role of high diversion. We examined the long-term outcome of a large series of patients treated with primary valve ablation, and compared it to the outcome of high diversion and vesicostomy. MATERIALS AND METHODS: We reviewed the records of 100 patients treated with primary valve ablation (74%), vesicostomy (13%) or high diversion (9%) before 1985. Median followup was 11.2 years. RESULTS: Overall 13% of our patients had end stage renal disease by age 15 years. Three patients initially treated with valve ablation and 3 initially treated with vesicostomy later underwent high diversion but none benefited from the secondary procedure. Four patients initially treated with valve ablation subsequently underwent vesicostomy but only 1 benefited. Bladder storage capacity was well preserved. Diurnal urinary continence developed in 46% of patients at age 10 years and only 1 remained incontinent after age 20 years. One patient with diversion who awaits transplantation had a small contracted bladder. Recent urodynamic studies in 10 cases of delayed urinary continence have not shown decreased bladder compliance or capacity. Kaplan-Meier analysis of outcomes of the different treatments indicated no statistical difference in patient age at end stage renal disease development. However, comparing the number of surgical procedures in the different treatment groups revealed a significant increase in the amount of surgery in infants with diversion. Our results were equivalent to those of the best published series, many of which strongly advocate high diversion. CONCLUSIONS: By avoiding diversion in most cases bladder function is preserved and the need for bladder augmentation is decreased.

Child↗
PubMed1996

Induction of ductal morphogenesis and lobular hyperplasia by amphiregulin in the mouse mammary gland.

As the juvenile mouse mammary gland matures, it undergoes extensive epithelial proliferation, leading to a network of ductal branching that transverses the organ. Recent evidence suggests that the epidermal growth factor-related peptide amphiregulin (AR) may play multiple roles in the proliferation, differentiation, and neoplastic conversion of the mouse mammary gland. Using a dual approach of recombinant AR in slow-release pellets and retroviral expression of AR, we explored the roles of this growth factor in the mouse mammary gland in vivo. We first noted that recombinant AR can reestablish longitudinal ductal proliferation in growth quiescent mammary glands of ovariectomized mice. Furthermore, retrovirally transduced mammary transplants overexpressing AR developed into hyperplastic tertiary ducts and hyperplastic lobules with increased lateral branching, apparent 9 weeks after transplantation into cleared mammary fat pads. This is the first study to demonstrate that AR can reestablish the early developmental activity of ductal mammary epithelium and induce hyperplasia in vivo. These data, coupled with previous findings that demonstrated nearly universal overexpression of AR in human breast cancer and rodent mammary tumorigenesis, suggest that AR may be an important intermediary in glandular maturation and early malignant progression.

Adipose Tissue↗
PubMed1995

Detection and location of amphiregulin and Cripto-1 expression in the developing postnatal mouse mammary gland.

Amphiregulin (Ar) and Cripto-1 (Cr-1) are growth promoting peptides that share amino acid sequence homology with epidermal growth factor (EGF). The present study examined Ar and Cr-1 mRNA and protein expression during various stages of C57BL/6 mouse mammary morphogenesis. Reverse transcription-polymerase chain reaction (RT-PCR) was used to detect transcripts for Ar and Cr-1 at all stages of mammary development. Immunocytochemical (ICC) localization demonstrated that in virgin 4-week to mature 12-week-old mouse fourth inguinal mammary gland, Ar and Cr-1 are expressed in the stromal cells, luminal epithelial cells, and myoepithelial cells of the branching ducts. Ar, and to lesser extent Cr-1, were also found in the epithelial cap cells and in the luminal epithelial cells of the advancing terminal end bud (TEB) from virgin 4-week and 6-week-old mice. Western blot analysis demonstrated that both Ar (28 and 26 kDa) and Cr-1 (90, 67, 56, and 21 kDa) proteins are expressed in virgin, 13.5 day midpregnant and in the 14 day lactating mammary gland. In addition, Ar and Cr-1 are associated with developing alveolar structures as determined by ICC. These results imply that together with EGF and transforming growth factor alpha (TGF alpha), Ar and Cr-1 may play salient roles as modifiers in the morphogenesis and differentiation of the mammary gland.

Amphiregulin↗