Modeling variability of the lactation curves of cows in automated milking systems.

Historically, cow selection criteria were developed for conventional milking systems that have regular milking intervals (MI). However, in automatic milking systems (AMS), there is variability in MI within and between cows. These sources of variability provide an opportunity to identify cows with high daily milk yield (DY) and long MI. An extended MI (longer than 16 h in pasture-based systems) has a negative effect on DY. Cows that tolerate extended MI and maintain high DY can be considered more efficient than cows with low DY and long MI, or with high DY but short MI, thereby improving robotic system use. Knowledge of the behavior and parameters of lactation curves of cows in AMS could help farmers to identify cows with a specific lactational phenotype. The objective of this study was to identify individual cows with high DY and long MI within herds, which could reflect increased tolerance to milk accumulation under AMS. A database containing records for 773,483 milking events for one year (July 2016-June 2017) from 4 pasture-based AMS farms was used. Lactation curves within each herd were fitted using several mixed models including fixed effects for the parameters of the lactation curve and random cow effects. Predicted curves of average DY according to parity (multiparous and primiparous) were obtained. The best linear unbiased prediction of the random cow effect allowed us to categorize lactations as having either high or low milk production. The median MI of each lactation was then used to categorize cows as having either short or long MI. Daily yield at the peak of lactation, days to peak and 305-d cumulative milk production were used to compare the effect of DY and MI categories, as well as the DY × MI interaction. Milk production by multiparous and primiparous cows with high DY and long MI was between 35 and 45% higher than that of the low DY and short MI. From all lactations analyzed, the incidence of animals with high DY and long MI across farms was 7.5%. We have identified and quantified a new, AMS-specific, phenotype (the combination of a relatively higher DY with relatively longer MI) with potential to increase use of AMS units. Identifying more efficient animals should help generate new approaches for differential management and for selecting cows in AMS.

A qualitative study on the oral health of humanitarian migrants in Canada.

OBJECTIVES: There is limited evidence to guide oral health policy and services for the 25,000 refugees and asylum seekers who arrive in Canada yearly. The purpose of this study was to explore and understand the pre-migration use of dental services, oral health knowledge, and the effects of oral disease among newly arrived humanitarian migrants in order to inform policy and practice for the population. METHODS: Using focused ethnography and the public health model of the dental care process, we conducted face-to-face interviews (50-60 minutes) with a purposive sample of humanitarian migrants who had indicated the need for dental care. We observed mobile dental clinics that provided care to underserved communities in Montreal. Data were analyzed using a thematic and contextual approach that combined inductive and deductive frameworks. RESULTS: Participants included 25 humanitarian migrants from four global geographical regions. Five major thematic categories were explored: problem-based dental consultation, self-assessed oral health status, causes of oral diseases, personal oral hygiene, and good oral health for wellbeing. In their countries of origin, participants consulted a dentist when oral symptoms persisted. They cited excessive sugar consumption and inadequate oral hygiene as causes of oral diseases, and reported significant oral diseases impacts that limited their daily functions and wellbeing once in Canada. CONCLUSIONS: Humanitarian migrants were knowledgeable about causes of oral disease and the importance of good oral health, yet poor oral health continued to affect their lives in Canada in important ways.

Neoplastic transformation of porcine mammary epithelial cells in vitro and tumor formation in vivo.

BACKGROUND: The mammary glands of pigs share many functional and morphological similarities with the breasts of humans, raising the potential of their utility for research into the mechanisms underlying normal mammary function and breast carcinogenesis. Here we sought to establish a model for the efficient manipulation and transformation of porcine mammary epithelial cells (pMEC) in vitro and tumor growth in vivo. METHODS: We utilized a vector encoding the red florescent protein tdTomato to transduce populations of pMEC from Yorkshire -Hampshire crossbred female pigs in vitro and in vivo. Populations of primary pMEC were then separated by FACS using markers to distinguish epithelial cells (CD140a-) from stromal cells (CD140a+), with or without further enrichment for basal and luminal progenitor cells (CD49f+). These separated pMEC populations were transduced by lentivirus encoding murine polyomavirus T antigens (Tag) and tdTomato and engrafted to orthotopic or ectopic sites in immunodeficient NOD.Cg-Prkdc (scid) Il2rg (tm1Wjl) /SzJ (NSG) mice. RESULTS: We demonstrated that lentivirus effectively transduces pMEC in vitro and in vivo. We further established that lentivirus can be used for oncogenic-transformation of pMEC ex vivo for generating mammary tumors in vivo. Oncogenic transformation was confirmed in vitro by anchorage-independent growth, increased cell proliferation, and expression of CDKN2A, cyclin A2 and p53 alongside decreased phosphorylation of Rb. Moreover, Tag-transformed CD140a- and CD140a-CD49f + pMECs developed site-specific tumors of differing histopathologies in vivo. CONCLUSIONS: Herein we establish a model for the transduction and oncogenic transformation of pMEC. This is the first report describing a porcine model of mammary epithelial cell tumorigenesis that can be applied to the study of human breast cancers.

Technical note: A vacuum-assisted approach for biopsying the mammary glands of various species.

Biopsy of the mammary glands is a technique used in both research and clinical diagnosis. A vacuum-assisted approach that enables the collection of tissue from the mammary glands of various species is described, along with methods for biopsying cows and pigs. The procedure involves tissue penetration via blunt dissection with a sharpened trocar. Tissue cores are excised and collected via the vacuum-assisted handpiece followed by a saline lavage and wound closure. This approach yields tissue cores of approximately 100 mg, is well suited for use in various species, and affords the potential to reduce postoperative complications.

The influence of distress on knowledge transfer for men newly diagnosed with prostate cancer.

The purpose of this inquiry was to evaluate the efficacy of prostate cancer education sessions. Implementation of 3-h patient educational sessions was intended to provide men newly diagnosed with localized prostate cancer, who face difficult and complex decisions, information about potential treatment options. Fifty-seven men completed the distress thermometer assessment before the education session to assess baseline levels of distress. Seven of the men were interviewed post-educational session to determine the degree of knowledge transfer from the session. This study explored the efficacy of the patients' learning experience using an interpretive phenomenological research approach. Resulting data revealed that these patients, as adult learners, were distressed and that, despite the availability of pertinent medical content, the subject material was not learned as intended or readily understood. The conclusion drawn from this preliminary applied educational research study was that the education model used was less than efficacious at ensuring that sufficient knowledge transfer was achieved for medical treatment decision-making processes. These findings suggest a need for future research to explore the application of adult learning theories and approaches that may offer enhanced knowledge translation and transfer for prostate cancer education programs.

Physical Constraint in Pediatric Dentistry: The Lived Experience of Parents.

BACKGROUND: The use of physical constraint in pediatric dentistry is highly controversial. Papoose boards in particular, which envelop and immobilize children during treatment procedures, have been described as barbaric devices even though their goal is to protect the patient. In this debate, the voice of parents is important but still missing in the scientific literature. AIM: To understand how parents or caregivers experienced physical constraint and the use of the papoose board on their children during regular dental treatment. DESIGN: We conducted qualitative research rooted in interpretive phenomenology. Accordingly, we performed in-depth individual interviews with a purposive sample of 7 parents or caregivers. The interviews took place in Montréal, Canada, after the children had been treated with a papoose board for nonemergency dental treatments. The discussions were audio recorded, transcribed, and thematically analyzed. RESULTS: Two perspectives emerged among participants. Some explained that the papoose board calmed their children, helped the dentist to complete the procedures, and made their experience less stressful. For others, the papoose board was a horrible and traumatizing experience, leading to feelings of guilt toward their children. They expressed anger toward the dentists for not allowing them enough time to decide and for imposing use of the device. CONCLUSION: Our study raises serious ethical concerns about this practice. We believe that using a papoose board should remain an extraordinary measure and, more generally, that dental professionals should reflect on the place of children and their families in clinical encounters. KNOWLEDGE TRANSFER STATEMENT: The findings of this study should encourage policy makers, dental professionals and ethicists to consider the following points: 1) the traumatizing experiences described by parents raise serious ethical concerns about the use of papoose boards; 2) the dental profession should reflect on the place of children and their families in the clinical encounter and grapple with the importance of consent and how to ensure consent in encounters involving children and their parents.

Metoclopramide induces preparturient, low-level hyperprolactinemia to increase milk production in primiparous sows.

Inadequate milk production by sows often limits the growth of piglets. A successful lactation requires prolactin (PRL)-induced differentiation of the alveolar epithelium within the mammary glands of sows between days 90-110 of gestation. We hypothesized that induction of late gestational hyperprolactinemia in primiparous sows by oral administration of the dopamine antagonist metoclopramide (MET) would enhance mammary epithelial differentiation, milk yield, and piglet growth rate and that these effects would carry over into a subsequent lactation. Twenty-six gilts were assigned to receive either MET (n = 13, 0.8 mg/kg) or vehicle (CON, n = 13) twice daily from days 90-110 of gestation. The same sows were followed into their second lactation without additional treatment. On day 90 of gestation, circulating PRL concentrations peaked 45 min after feeding MET (P < 0.001) and then returned to baseline 3 h later. This response occurred daily out to day 104 of gestation (P < 0.05). Compared with CON, MET-treated gilts had enlarged alveoli on gestation day 110 (P < 0.05). Treatment with MET did not affect feed intake, body weight, or body fatness during pregnancy or lactation. Piglets born to MET-treated sows had both increased body weights and average daily gain on lactation days 14 and 21 (P < 0.05). Milk intake by piglets was estimated from deuterium oxide dilution. Although milk intake by piglets nursing MET sows was not statistically different from those nursing CON sows on day 21 of lactation (P = 0.18), there was a greater increase in milk consumption by piglets born to MET-treated sows between days 9 and 21 of lactation than for those in CON litters (P < 0.001). In one group of second parity sows (n = 11) that were treated with MET during their first gestation, milk yield increased by 21% during their second lactation (P < 0.05) in association with a 14% decline in body fatness across lactation compared with a 7% decline in CON sows (P < 0.05). These findings demonstrate that MET-induced hyperprolactinemia in primiparous sows during late pregnancy can increase milk yield and piglet growth rate, setting the stage for further large-scale studies.

Bladder cancer in spinal cord injury patients.

STUDY DESIGN: Retrospective review. OBJECTIVE: Spinal cord injury is a known risk factor for bladder cancer. The risk of bladder cancer has been reported at 16-28 times higher than the general population. Earlier studies have identified indwelling catheters as risk factors. We examined the characteristics of bladder cancers in a spinal cord injury (SCI) population. SETTING: Long Beach VA Hospital Spinal Cord Injury Unit, Long Beach, California. METHODS: We reviewed SCI patients seen and diagnosed with bladder tumors between January 1983 and January 2007. Data collected included time since diagnosis, method of diagnosis, form of bladder management, pathologic type, treatment of the tumor, and outcome. RESULTS: A total of 32 patients with bladder cancer were identified out of 1319 seen. Tumors found were 46.9% squamous cell carcinoma (SCC), 31.3% transitional cell carcinoma (TCC), 9.4% adenocarcinoma, and 12.5% mixed TCC and SCC. The primary form of bladder management was 44% urethral catheter for a mean of 33.3 years, 48% external catheter for a mean of 37.4 years, and 8% intermittent catheterization for a mean of 24.5 years. Nineteen patients had a known method of cancer detection with 42% found on screening cystoscopy. CONCLUSIONS: The pathologic makeup of the tumors is similar to that reported earlier. Over 50% of patients diagnosed with bladder cancer in our population did not have an indwelling catheter. This suggests that the neurogenic bladder, not the indwelling catheter, may be the risk factor for bladder cancer. Urologists should consider diligent, long-term screening of all patients with SCI for bladder cancer and not just those with indwelling catheters.

Urethral versus suprapubic catheter: choosing the best bladder management for male spinal cord injury patients with indwelling catheters.

OBJECTIVE: Bladder management for male patients with spinal cord injury (SCI) challenges the urologist to work around physical and social restrictions set forth by each patient. The objective of this study was to compare the complications associated with urethral catheter (UC) versus suprapubic tube (SPT) in patients with SCI. METHODS: A retrospective review of records at Long Beach Veterans Hospital was carried out to identify SCI patients managed with SPT or UC. Chart review identified morbidities including urinary tract infection (UTI), bladder stones, renal calculi, urethral complications, scrotal abscesses, epididymitis, gross hematuria and cancer. Serum creatinine measurements were evaluated to determine whether renal function was maintained. RESULTS: In all, 179 patients were identified. There was no significant difference between the two catheter groups in any areas in which they could be compared. There were catheter-specific complications specific to each group that could not be compared. These included erosion in the UC group and urethral leak, leakage from the SPT and SPT revision in the SPT group. Average serum creatinine for the UC and SPT groups was 0.74 and 0.67 mg per 100 ml, respectively. CONCLUSION: SCI patients with a chronic catheter have similar complication rates of UTIs, recurrent bladder/renal calculi and cancer. Urethral and scrotal complications may be higher with UC; however, morbidity from SPT-specific procedures may offset benefits from SPT. Serum creatinine was maintained in both groups. Overall, bladder management for patients with chronic indwelling catheters should be selected on the basis of long-term comfort for the patient and a physician mind-set that allows flexibility in managing these challenges.

Effects of sustained hyperprolactinemia in late gestation on mammary development of gilts.

The objective of this project was to determine the effects of sustained hyperprolactinemia for 7 or 20 d on mammary development in late-pregnant gilts. On day 90 of gestation, gilts were assigned to one of 3 groups to receive intramuscular (IM) injections of (1) canola oil (CTL, n = 18) until day 109 ± 1 of gestation; (2) a dopamine receptor antagonist, domperidone (0.5 mg/kg of body weight [BW]) until day 96 ± 1 of gestation (T7, n = 17); or (3) domperidone (0.5 mg/kg BW) until day 109 ± 1 of gestation (T20, n = 17). Domperidone-treated gilts also received 100 mg of domperidone per os twice daily from days 90 to 93 of gestation. Blood was sampled on days 89, 97, 104, and 110 for prolactin (PRL), insulin-like growth factor 1 (IGF1), lactose, urea, and glucose assays. Mammary glands were collected at necropsy, on day 110, for compositional and cell proliferation analyses. Abundance of mRNA for selected genes was also determined in the mammary gland and the pituitary gland. On day 97 of gestation, PRL concentrations were 3 times greater for T20 and T7 than CTL gilts and were also greater for T20 than T7 and CTL gilts on days 104 and 110 (P < 0.001). Concentrations of IGF1 in T20 and T7 gilts were elevated relative to controls on days 97 and 104 and were greater for T20 vs T7 and CTL gilts on day 110 (P < 0.05). There were no treatment effects (P > 0.1) on parenchymal or extraparenchymal tissue weights, or on epithelial proliferation as measured by immunohistochemistry for Ki-67. Treatments did not alter concentrations of dry matter (DM), fat, or DNA (P > 0.1) in parenchyma. Concentrations of RNA (P < 0.05) and protein (P < 0.10) as well as total parenchymal protein, RNA, and DNA (P < 0.05) were lower, or tended to be, in T20 than T7 or CTL gilts. Hyperprolactinemia for 20 d in late gestation increased mRNA abundance of the milk protein genes beta-casein (CSN2) and whey acidic protein (WAP) (P < 0.05) in mammary parenchyma and also decreased mRNA abundance of the long form of the prolactin receptor (PRLR-LF). Increasing PRL concentrations for 7 or 20 d in late gestation had no beneficial effects on the composition of the mammary gland, and sustained exposure to domperidone for 20 d reduced metabolic activity either by a lower expression of the long form of the PRL receptor in mammary parenchymal tissue or, most likely, by the early involution of parenchymal tissue. In conclusion, results do not support the hypothesis that a sustained hyperprolactinemia in late gestation could enhance mammary development of gilts.

Using domperidone to induce and sustain hyperprolactinemia in late-pregnant gilts.

Prolactin controls mammary development as well as the lactogenic and galactopoietic processes in sows and increasing prolactin during gestation can augment milk yield. The dopamine receptor antagonist domperidone can increase circulating prolactin concentrations in pigs, but the ideal dose to achieve sustained hyperprolactinemia remains unknown. An experiment was performed to develop a protocol for using domperidone in studies of rapid and sustained hyperprolactinemia in late-pregnant gilts. On day 90 of gestation, gilts were divided into 4 groups: (1) intramuscular (IM) injections of canola oil (3 mL, controls [CTL], n = 9), (2) IM injections with 0.1 mg/kg BW of domperidone (low [LO], n = 8), (3) IM injections with 0.5 mg/kg BW of domperidone (medium [ME], n = 11), and (4) IM injections with 1.0 mg/kg BW of domperidone (high [HI], n = 11). Injections were given daily at 8:05 from days 90 to 109 of gestation. Treated gilts also received domperidone per os (0.5 mg/kg BW) at 8:00 and 20:00 on days 89, 90, and 91 of gestation. Three jugular blood samples were collected from all gilts at 6-h intervals on days 89, 90, and 91 of gestation, then twice daily on days 92, 93, and 94. Thereafter, samples were obtained at 8:00 every other day until day 114 of gestation. Blood was sampled serially from 9 CTL and 11 HI gilts on days 89 and 94 of gestation. On day 89 of gestation, prolactin concentrations for LO, ME, and HI gilts increased within 6 h of domperidone per os (P < 0.001). From days 89 until 93 of gestation, the area under the curve (AUC) for LO, ME, and HI gilts was greater than that for CTL gilts (P < 0.001), whereas from days 89 until 114, ME and HI gilts had greater AUC than CTL and LO gilts (P < 0.05). Results demonstrate that the combination of per os treatment with IM injections of 0.5 mg/kg of domperidone in an oil emulsion leads to the rapid and sustained release of prolactin over 24 d in late-pregnant gilts. Higher doses of domperidone failed to further increase circulating prolactin levels. These findings provide a useful strategy to induce sustained hyperprolactinemia in late-pregnant gilts.

Patient Experience of Living with Orofacial Pain: An Interpretive Phenomenological Study.

This study investigated the experience of living with chronic orofacial pain. Participating in this study's individual in-depth interviews were 6 participants with chronic orofacial pain who were undergoing treatment at a pain clinic. In consideration of the empirical nature of the study, the interviews were transcribed verbatim and analyzed from an interpretive phenomenological approach. The participants in the study experienced various types of loss as a result of chronic pain, such as the loss of employment, self-identity, and the ability to enjoy eating and engaging in social activities. Other findings revealed a disbelief among the families and medical community in the pain reported by the participants in this study. In addition, the current study explores the dissatisfaction of these participants with their journey through the health care system. Previous studies have concentrated on chronic pain predominantly through quantitative methods based on measurements, such as questionnaires. The biomedical aspects of pain are essential to report, yet the emphasis on this can result in overlooking the experience of living with chronic pain. Qualitative methodologies are necessary for gaining a more profound understanding of and appreciation for the individual patient and his or her unique perspective. The present study may benefit those living with chronic pain, since this sharing of experiences can help sufferers feel less isolated in their agony. The current article's intention is to report these experiences in such a way that chronic pain sufferers and laypeople can comprehend and relate to them. In addition, the findings of this study should promote the knowledge and understanding of health care providers who deal with chronic pain sufferers, so that treatment is provided with more empathy and compassion. Knowledge Transfer Statement: The findings of the present study may assist health care professionals interacting regularly with sufferers of chronic pain to gain a deeper understanding of the chronic pain experience and how to best support these patients.

How can we provide person-centred dental care?

Patient- or person-centred care is the current paradigm in the health profession yet there is still no clear understanding of what it means or how it could be implemented in dentistry. Building on a previously proposed person-centred model in clinical dentistry, in this article a person-centred dental clinical approach is presented. The approach consists of three guiding principles - humility, hospitality and mindfulness - that influence the different processes of the dental clinical encounter: connecting, examining, sharing, and intervening. The presented approach provides a rich opportunity for dentists to fine tune their own clinical approach in order to keep up with the upcoming expectations of their patients.

TRIENNIAL LACTATION SYMPOSIUM/BOLFA: Dietary regulation of allometric ductal growth in the mammary glands.

Although mammary gland growth and development in females is a lifelong process, it builds on isometric and allometric phases of mammary growth to establish a complex ductal network before and during puberty. Only then can other phases of branching and alveologenesis, differentiation, lactation, and involution proceed. Although the ductal network of various species differs in its histomorphology, all glands undergo a common phase of allometric growth when the mammary ducts penetrate into the supporting stromal microenvironment. Perhaps not surprisingly, different aspects of diet and nutrition can influence this allometric growth, either directly or indirectly. In this review, we outline some of the fundamental aspects of how allometric ductal growth in the mammary glands of various species is influenced by diet and nutrition and identify opportunities and questions for future investigation.

Genetic alterations in primary bladder cancers and their metastases.

Bladder cancer progression is thought to be associated with sequential genetic events. To search for the specific genetic changes associated with the metastatic process, comparative genomic hybridization was performed on 22 primary tumors and 24 metastases (10 distant and 14 nodal metastases) from 17 patients with stage pT2-4 bladder cancer. There was a striking similarity between the genetic alterations present in the primary and metastatic tumor samples from the same patient. The mean number of genetic changes/tumor was 12.2 for primary tumors and 11.7 for metastases. There was a strong concordance in the specific aberrations present in each patient's primary and metastatic lesions (mean, 75%). Concordance was also high among multiple sites from an individual primary tumor (mean, 96%) and multiple metastases from the same patient (mean, 75%). There were no specific genetic changes overrepresented in the metastases compared with their primary tumors. Genetic alterations present in more than 40% of tumors included gains on 6p, 8q, 10q, and 17q and losses involving 8p, 10q, and Y. Two regions of high-level amplification were common: (a) 10q22.1-q23.1 (32.6%); and (b) 17q11-21.3 (23.9%; the locus of erbB-2). A summary statistic was developed to quantitate the degree of clonal relationships between biopsies from the same patient. These data support a model in which minimal clonal evolution occurs in the metastatic tumor cell population after the metastatic event. When comparing primary cancers from patients with and without metastases, however, several unique genetic changes were identified in those cancers with metastases, suggesting that these loci may harbor genes important to the metastatic process.

Regulation and localization of vascular endothelial growth factor within the mammary glands during the transition from late gestation to lactation.

The vascular network within the developing mammary gland (MG) grows in concert with the epithelium to prepare for lactation, although the mechanisms coordinating this vascular development are unresolved. Vascular endothelial growth factor A (VEGF-A) mediates angiogenesis and vascular permeability in the MG during pregnancy and lactation, where its expression is upregulated by prolactin. Given our previous finding that late-gestational hyperprolactinemia induced by domperidone (DOM) increased subsequent milk yield from gilts, we sought to establish changes in vascular development during late gestation and lactation in the MGs of these pigs and determine whether DOM altered MG angiogenesis and the factors regulating it. Gilts received either no treatment (n = 6) or DOM (n = 6) during late gestation, then had their MG biopsied from late gestation through lactation to assess microvessel density, VEGF-A distribution and messenger RNA expression, and aquaporin (AQP) gene expression. Microvessel density in the MG was unchanged during gestation then increased between days 2 and 21 of lactation (P < 0.05). The local expression of messenger RNA for VEGF-A120, VEGF-A147, VEGF-A164, VEGF-A164b, VEGF-A188, VEGF receptors-1 and -2, and AQP1 and AQP3 all generally increased during the transition from gestation to lactation (P < 0.05). Immunostaining localized VEGF-A to the apical cytoplasm of secretory epithelial cells, consistent with a far greater concentration of VEGF-A in colostrum and/or milk vs plasma (P < 0.0001). There was no effect of DOM on any of the variables analyzed. In summary, we found that vascular development in the MG increases during lactation in first-parity gilts and that VEGF-A is a part of the mammary secretome. Although late-gestational hyperprolactinemia increases milk yield, there was no evidence that it altered vascular development.

Transcriptional regulation of vascular endothelial growth factor expression in epithelial and stromal cells during mouse mammary gland development.

Accompanying changes in the development and function of the mammary gland is the establishment of a vascular network of critical importance for lactogenesis and tumorigenesis. A potent angiogenic and permeability factor that regulates vascular development in association with epithelial-stromal interactions is vascular endothelial growth factor (VEGF). Analysis of VEGF transcription by RT-PCR revealed mRNA for all three VEGF isoforms (VEGF120, 164, 188) within the mammary gland of nulliparous females. During pregnancy the level of VEGF188 declined and became undetectable during lactation in association with the increased abundance of VEGF120 and VEGF164 mRNAS: All three isoforms were expressed at consistent levels within the cleared mammary fat pad throughout development. Furthermore, the presence of VEGF188 mRNA in omental adipose tissue at various stages established that VEGF188 is expressed specifically in adipose tissue within the mammary gland. Using 3T3-L1 preadipocytes it was demonstrated that VEGF188 mRNA transcription occurs as a late event during lipogenesis distinct from earlier induction of VEGF120 and VEGF164 mRNA during differentiation. In contrast, HC11 mammary epithelial cells only expressed mRNA for VEGF120 and VEGF164. Localization of VEGF mRNA and protein revealed that VEGF is expressed in stromal cells of the mammary gland in nulliparous females and then undergoes a transition to epithelial expression during lactation. By contrast, mRNA for the VEGF receptors, Flk-1 and Flt-1, localized to stromal cells within the mammary fat pad during virgin and gestational development and was expressed in the interstitial tissue basal to epithelial cells during lactation. Taken together, these results support the conclusion that VEGF is differentially transcribed by specific cell types within the mammary gland, and that under hormonal regulation it functions in an autocrine/paracrine manner.

C/EBPbeta (CCAAT/enhancer binding protein) controls cell fate determination during mammary gland development.

Deletion of the transcription factor CCAAT/enhancer binding protein (C/EBP)beta results in a severe inhibition of lobuloalveolar development in the mouse mammary gland. Because progesterone receptor (PR) is requisite for alveolar development, the expression of PR was investigated in C/EBPbeta-/- mice. Unexpectedly, the number of PR-positive cells, as well as the levels of PR mRNA, were elevated 3-fold in the mammary glands of C/EBPbeta-/- mice. Furthermore, in contrast to wild-type nulliparous mice, in which PR distribution shifted from a uniform to nonuniform pattern between 8-12 weeks of age, C/EBPbeta-/- mice exhibited uniform PR distribution throughout all stages of mammary development analyzed. No change in C/EBPbeta mRNA levels was observed in the mammary glands of PR-/- mice, suggesting that PR acts in a pathway either in parallel to or downstream of C/EBPbeta. The overexpression and disrupted cellular distribution of PR in C/EBPbeta-/- mice were coincident with a striking 10-fold decrease in cell proliferation after acute steroid hormone treatment, assayed by incorporation of bromodeoxyuridine. In wild-type mice, PR and bromodeoxyuridine-positive cells were adjacent to each other and rarely colocalized. No differences in the level or pattern of PR expression were observed in the uterus, suggesting that C/EBPbeta influences PR in a mammary-specific fashion. Together, these data suggest that C/EBPbeta may control cell fate decisions in the mammary gland through the appropriate temporal and spatial expression of molecular markers, such as PR, that induce the proliferation of alveolar progenitor cells via juxtacrine mechanisms.

Ovariectomy in young prepubertal dairy heifers causes complete suppression of mammary progesterone receptors.

Mammary growth and development depends on ovarian steroids and particularly interaction of estrogen and progesterone with their intracellular receptors. The objectives of this study were to determine the effect of ovariectomy on the expression of protein and messenger RNA for estrogen receptor-alpha (ESR1) and progesterone receptor (PGR) and their relation to mammary ductal development and cell proliferation. Prepubertal Holstein heifers 2, 3, or 4 mo of age were randomly assigned to one of 2 treatments, ovariectomized (OVX; n = 8) or sham operated (INT; n = 12). Mammary parenchymal (PAR) tissue samples were harvested 30 d after surgery. Localization and quantitation of ESR1 and PGR in PAR were determined by immunohistochemistry and quantitative multispectral imaging. Relative messenger RNA expression of ESR1 and PGR in PAR was measured by quantitative real time polymerase chain reaction. We observed the complete absence of PGR-positive epithelial cell nuclei and reduced PGR transcript abundance in mammary parenchyma of OVX heifers. The percent of epithelial cells expressing ESR1 did not differ by treatment but was decreased with age. However, average intensity of ESR1 expression per cell was reduced in OVX heifers. The abundance of Ki67 labeled epithelial cells and stromal cells was reduced after ovariectomy. These data suggest that reduced mammary development after ovariectomy may be mediated by loss of PGR expression and reduced ESR1 expression in positive cells. A presumptive relationship with ovarian-derived circulating estradiol remains unresolved, but data suggest other ovarian-derived agents may play a role. Use of specific antagonists to manipulate expression or action of PGR and ESR1 receptors should provide direct evidence for roles of these receptors in prepubertal bovine mammary development.

Feedback regulation of PRL secretion is mediated by the transcription factor, signal transducer, and activator of transcription 5b.

PRL secretion from the anterior pituitary gland is inhibited by dopamine produced in the tuberoinfundibular dopamine neurons of the hypothalamus. The activity of tuberoinfundibular dopamine neurons is stimulated by PRL; thus, PRL regulates its own secretion by a negative feedback mechanism. PRL receptors are expressed on tuberoinfundibular dopamine neurons, but the intracellular signaling pathway is not known. We have observed that mice with a disrupted signal transducer and activator of transcription 5b gene have grossly elevated serum PRL concentrations. Despite this hyperprolactinemia, mRNA levels and immunoreactivity of tyrosine hydroxylase, the key enzyme in dopamine synthesis, were significantly lower in the tuberoinfundibular dopamine neurons of these signal transducer and activator of transcription 5b-deficient mice. Concentrations of the dopamine metabolite dihydroxyphenylacetic acid in the median eminence were also significantly lower in signal transducer and activator of transcription 5b-deficient mice than in wild-type mice. No changes were observed in nonhypothalamic dopaminergic neuronal populations, indicating that the effects were selective to tuberoinfundibular dopamine neurons. These data indicate that in the absence of signal transducer and activator of transcription 5b, PRL signal transduction in tuberoinfundibular dopamine neurons is impaired, and they demonstrate that this transcription factor plays an obligatory and nonredundant role in mediating the negative feedback action of PRL on tuberoinfundibular dopamine neurons.