Glyphosate modifies the gene expression and migration of trophoblastic cells without altering the process of angiogenesis or the implantation of blastocysts in vitro.

Adverse pregnancy outcomes have been associated with the presence of glyphosate (G) in umbilical cord, serum, and urine samples from pregnant women. Our aim was to study the effect of G on blastocyst implantation using an in vitro mouse model, and the migration and acquisition of endothelial phenotype of the human trophoblastic HTR8/SVneo (H8) cells. In mouse blastocysts, no differences in attachment time and implantation outgrowth area were observed after G exposure. H8 cell migration was stimulated by 0.625 µM G without cytotoxicity. After 6 h, the mRNA expression of vascular endothelial growth factor (VEGF) and C-C motif chemokine ligand 2 (CCL2) was upregulated in H8 cells exposed to 1.25 µM G when compared vehicle-treated cells (p ≤ 0.05). No differences were observed in interleukin 11, VEGF receptor 1, and coagulation factor II thrombin receptor in H8 cells exposed to different concentrations of G for 6 h compared to the vehicle. Interestingly, exposure to G did not alter angiogenesis as measured by a tube formation assay. Taken all together, these results suggest that G exposure may contribute as a risk factor during pregnancy, due to its ability to alter trophoblast migration and gene expression.

Uterine histopathology and steroid metabolism in a polycystic ovary syndrome rat model.

The aim of this study was to investigate uterine lesions, uterine endocrine status and expression of genes involved in uterine differentiation in a rat model of polycystic ovary syndrome (PCOS). The possible involvement of the androgen receptor (AR) was also investigated. PCOS rats showed an increased incidence of uterine epithelial and glandular lesions and elevated serum testosterone level, which was not detected in uterine tissue. Uterine 17ß-estradiol, estrone and progesterone were detected in 100%, 75% and 50% of the animals, respectively. This was associated with a decrease in Star and an increase in Hsd17b2, Srd5a1 and Cyp19a1, suggesting that uterine steroids are not synthesized de novo in PCOS and that alterations in these enzymes may explain the absence of testosterone and low progesterone. In addition, ESR2 decreased and AR increased, suggesting possible steroid receptor crosstalk. Genes associated with uterine differentiation, PTEN and WNT5a, also showed reduced expression. PCOS rats treated with flutamide, an AR antagonist, were similar to PCOS rats in terms of uterine lesions, serum steroid levels, ESR2, PTEN and WNT5a expression. However, testosterone, AR and aromatase levels were similar to control rats, with decreased expression of ESR1 and HOXA10, suggesting that these expressions are AR dependent. Our results suggest that the primary cause of the observed uterine lesions in the PCOS rat model is the altered endocrine status and consequently changes in genes related to uterine differentiation.

Branching morphogenesis of the mouse mammary gland after exposure to benzophenone-3.

Pubertal mammary branching morphogenesis is a hormone-regulated process susceptible to exposure to chemicals with endocrine disruptive capacity, such as the UV-filter benzophenone-3 (BP3). Our aim was to assess whether intrauterine or in vitro exposure to BP3 modified the branching morphogenesis of the female mouse mammary gland. For this, pregnant mice were dermally exposed to BP3 (0.15 or 50 mg/kg/day) from gestation day (GD) 8.5 to GD18.5. Sesame oil treatment served as control. Changes of the mammary glands of the offspring were studied on postnatal day 45. Further, mammary organoids from untreated mice were cultured under branching induction conditions and exposed for 9 days to BP3 (1 × 10-6 M, 1 × 10-9 M, or 1 × 10-12 M with 0.01% ethanol as control) to evaluate the branching progression. Mice that were exposed to BP3 in utero showed decreased mRNA levels of progesterone receptor (PR) and WNT4. However, estradiol and progesterone serum levels, mammary histomorphology, proliferation, and protein expression of estrogen receptor alpha (ESR1) and PR were not significantly altered. Interestingly, direct exposure to BP3 in vitro also decreased the mRNA levels of PR, RANKL, and amphiregulin without affecting the branching progression. Most effects were found after exposure to 50 mg/kg/day or 1 × 10-6 M of BP3, both related to sunscreen application in humans. In conclusion, exposure to BP3 does not impair mammary branching morphogenesis in our models. However, BP3 affects PR transcriptional expression and its downstream mediators, suggesting that exposure to BP3 might affect other developmental stages of the mammary gland.

Mammary gland development in male rats perinatally exposed to propiconazole, glyphosate, or their mixture.

This study aimed to assess whether perinatal exposure to propiconazole (PRO), glyphosate (GLY) or their mixture (PROGLY) alters key endocrine pathways and the development of the male rat mammary gland. To this end, pregnant rats were orally exposed to vehicle, PRO, GLY, or a mixture of PRO and GLY from gestation day 9 until weaning. Male offspring were euthanized on postnatal day (PND) 21 and PND60. On PND21, GLY-exposed rats showed reduced mammary epithelial cell proliferation, whereas PRO-exposed ones showed increased ductal p-Erk1/2 expression without histomorphological alterations. On PND60, GLY-exposed rats showed reduced mammary gland area and estrogen receptor alpha expression and increased aromatase expression, whereas PRO-exposed ones showed enhanced lobuloalveolar development and increased lobular hyperplasia. However, PROGLY did not modify any of the endpoints evaluated. In summary, PRO and GLY modified the expression of key molecules and the development of the male mammary gland individually but not together.

Postnatal exposure to a glyphosate-based herbicide interferes with the development and growth of the mammary gland of pre-pubertal Ewe lambs.

The aim of the present study was to evaluate whether early postnatal exposure to a glyphosate-based herbicide (GBH) alters pre-pubertal mammary development in Friesian lambs. To this end, from postnatal day 1-14, ewe lambs were exposed subcutaneously or orally to GBH (2 mg/kg bw/day) or vehicle (control) and mammary gland biopsies were obtained at 45 days of age. GBH-exposed lambs exhibited larger mammary ducts and less area occupied by terminal duct lobular units than controls, accompanied by an increase in the area of adipocytes in the mammary stroma. Lambs subcutaneously exposed to GBH showed increased protein expression of estrogen receptor alpha; however, both GBH-exposed groups had decreased mRNA expression of this receptor. Control lambs showed nuclear progesterone receptor (PR) protein expression, whereas GBH-exposed animals showed cytoplasmic PR expression; both GBH-exposed groups exhibited decreased mRNA expression of PR. GBH-exposed lambs also had decreased epithelial cell proliferation. Regarding insulin-like growth factors, both groups showed similar IGF-1 mRNA and protein expression but decreased expression of its receptor, and increased IGFBP5 expression. In addition, phosphorylated AKT was only observed in the mammary gland of control lambs. Our results show that early postnatal exposure to GBH, regardless of the exposure route, affects the IGF-1 system and the AKT/protein kinase B pathway, interfering with steroid hormone receptor expression and cell proliferation. This consequently modifies the growth and development of the pre-pubertal mammary gland of Frisian lambs.

Impacto de la COVID-19 en el área de Odontología

Introducción: La COVID-19 es una enfermedad provocada por el virus SARS-CoV-2, que se transmite por medio de la vía respiratoria por lo cual, los odontólogos enfrentan un gran riesgo al trabajar directamente en la cavidad oral. Objetivo: Sistematizar los referentes teóricos sobre el impacto de la COVID-19 en el área de la Odontología. Método: En la Universidad Regional Autónoma de los Andes, entre los meses de septiembre a diciembre de 2022 se realizó una revisión sistemática sobre el tema. De 36 artículos revisados se escogió, según criterios, un total de 23 artículos, disponibles en PUBMED y SciELO que abordan la problemática de COVID-19 en el área odontológica, de varios autores, en idioma inglés y español. Resultados: Se abordaron los temas, tales como: enfermedades bucodentales generadas a causa de COVID-19, Cambios en el área odontológica a causa de la pandemia por COVID-19 y medidas de bioseguridad empleadas para atender al paciente en el consultorio odontológico. Consideraciones finales: La COVID-19 ha tenido gran repercusión en Odontología, lo que afecta la salud bucal y general del paciente, a su vez, conduce al uso de estrictas medidas de bioseguridad dentro y fuera del consultorio odontológico, por lo que resulta ineludible que los odontólogos se empoderen de los referentes teóricos en torno al tema para contribuir a la detección de lesiones que puedan constituir signos primarios que apuntan a la presencia de SARS-CoV-2, adoptar conductas responsables y evitar su propagación.

Introduction: COVID-19 is a disease caused by SARS-CoV-2 virus and transmitted through respiratory track. So, dentists face a great risk working directly in the oral cavity. Objective: Systematization of the theoretical references concerning the impact of COVID-19 in dental areas. Method: A systematic review on the subject was carried out at the Universidad Regional Autónoma de los Andes, from September to December 2022. Of a total of 36 articles reviewed, 23 were selected according to criteria, available in PUBMED and SciELO, published in English and Spanish by different authors, and associated to the COVID-19 transmission in dentistry. Results: the following topics were addressed: oral diseases caused by COVID-19, changes in dental areas due to the COVID-19 pandemic, and biosecurity measures used in the dental service for ensure patient safety receiving treatment. Final considerations: COVID-19 has had great repercussions in dentistry, which affects the oral and general health of patients and, in turn, leads to the use of strict biosecurity measures inside and outside the dental office, so, it is essential for dentists to become empowered of the theoretical references related to the subject and also be focused on detecting lesions that may constitute primary signs of a possible presence of SARS-CoV-2, in adopt responsible behaviors and to avoid any spread of disease.

Introdução: O COVID-19 é uma doença causada pelo vírus SARS-CoV-2, que é transmitido pelo trato respiratório, portanto, os dentistas enfrentam um grande risco ao trabalhar diretamente na cavidade oral. Objetivo: Sistematizar os referenciais teóricos sobre o impacto da COVID-19 na área da Odontologia. Método: Na Universidade Regional Autônoma dos Andes, entre os meses de setembro e dezembro de 2022, foi realizada uma revisão sistemática sobre o tema. Dos 36 artigos revisados, um total de 23 artigos, disponíveis no PUBMED e SciELO, que abordam a problemática da COVID-19 na área odontológica, de diversos autores, em inglês e espanhol, foram escolhidos segundo critérios. Resultados: Foram abordados temas como: doenças bucais causadas pelo COVID-19, Alterações na área odontológica devido à pandemia do COVID-19 e medidas de biossegurança utilizadas no atendimento ao paciente no consultório odontológico. Considerações finais: O COVID-19 tem causado grande impacto na Odontologia, o que afeta a saúde bucal e geral do paciente, por sua vez, leva ao uso de medidas estritas de biossegurança dentro e fora do consultório odontológico, por isso é inevitável que os dentistas sejam capacitados por referenciais teóricos sobre o assunto para contribuir na detecção de lesões que possam constituir sinais primários que apontem para a presença do SARS-CoV-2, adotem condutas responsáveis e evitem sua disseminação.

Factores causantes de gingivitis en jóvenes estudiantes / Causative factors of gingivitis in young students

Objetivo: Identificar la relación que mantiene el cuidado bucodental e higiene en los jóvenes de la Institución Educativa "Primero de Abril" y la presencia de gingivitis, y conocer si la dieta de los estudiantes está ligada a la presencia de esta patología. Método: Se realizó un estudio investigativo de corte transversal, cuantitativo, observacional analítico. Resultados: Un 44,4% del género femenino consume fibra como alimento principal mientras que el 55% consume vitaminas y antioxidantes, mientras que el género masculino el 62,5% de la población consume vitaminas y antioxidantes y 37,5% restante consume fibras. Conclusión: La presencia de gingivitis en la población estudiada especialmente del género femenino y de status económico bajo, es debido a la alta deficiencia en cuanto a las visitas odontológicas, el uso y acceso de kits de limpieza dental, reconociendo así la prevalencia en el desarrollo de esta.

Objective: To identify the relationship between oral care and hygiene in young people of the "Primero de Abril" Educational Institution and the presence of gingivitis, and to know if the diet of the students is linked to the presence of this pathology. Methods: A cross-sectional, quantitative, observational, analytical, cross-sectional research study was carried out. Results: 44.4% of the female gender consumes fiber as the main food while 55% consumes vitamins and antioxidants, while 62.5% of the male gender consumes vitamins and antioxidants and the remaining 37.5% consumes fiber. Conclusion: The presence of gingivitis in the population studied, especially in the female gender and of low economic status, is due to the high deficiency in dental visits, the use and access to dental cleaning kits, thus recognizing the prevalence in the development of gingivitis.

Bisphenol A and benzophenone-3 exposure alters milk protein expression and its transcriptional regulation during functional differentiation of the mammary gland in vitro.

The plastic monomer and plasticizer bisphenol A (BPA), and the UV-filter benzophenone-3 (BP3) have been shown to have estrogenic activities that could alter mammary gland development. Our aim was to analyze whether BPA or BP3 direct exposure affects the functional differentiation of the mammary gland using an in vitro model. Mammary organoids were obtained and isolated from 8 week-old virgin female C57BL/6 mice and were differentiated on Matrigel with medium containing lactogenic hormones and exposed to: a) vehicle (0.01% ethanol); b) 1 × 10-9 M or 1 × 10-6 M BPA; or c) 1 × 10-12 M, 1 × 10-9 M or 1 × 10-6 M BP3 for 72 h. The mRNA and protein expression of estrogen receptor alpha (ESR1) and progesterone receptor (PR) were assessed. In addition, mRNA levels of PR-B isoform, glucocorticoid receptor (GR), prolactin receptor (PRLR) and Stat5a, and protein expression of pStat5a/b were evaluated at 72 h. The mRNA and protein expression of milk proteins and their DNA methylation status were also analyzed. Although mRNA level of PRLR and GR was similar between treatments, mRNA expression of ESR1, total PR, PR-B and Stat5a was increased in organoids exposed to 1 × 10-9 M BPA and 1 × 10-12 M BP3. Total PR expression was also increased with 1 × 10-6 M BPA. Nuclear ESR1 and PR expression was observed in all treated organoids; whereas nuclear pStat5a/b alveolar cells was observed only in organoids exposed to 1 × 10-9 M BPA and 1 × 10-12 M BP3. The beta-casein mRNA level was increased in both BPA concentrations and 1 × 10-12 M BP3, which was associated with hypomethylation of its promoter. The beta-casein protein expression was only increased with 1 × 10-9 M BPA or 1 × 10-12 M BP3. In contrast, BPA exposure decreased alpha-lactalbumin mRNA expression and increased DNA methylation level in different methylation-sensitive sites of the gene. Also, 1 × 10-9 M BPA decreased alpha-lactalbumin protein expression. Our results demonstrate that BPA or BP3 exposure alters milk protein synthesis and its transcriptional regulation during mammary gland differentiation in vitro.

Intervención con láser en frenillo lingual con inserción anterior y fibrosa. Reporte de caso

El frenillo lingual es una membrana mucosa. Esta estructura es importante para la movilidad de la lengua, si el frenillo tiene una inserción corta puede dificultar los movimientos de la lengua y afectar la erupción en buena posición de los incisivos inferiores. Reporte de caso: Este trabajo reporta el caso de una niña, 4 años de edad, que acude con su tutora al Centro Odontológico de la Universidad Científica del Sur con el motivo de "quiero que le revisen la lengüita". Luego de evaluarla, se diagnosticó anquiloglosia. La niña presentaba el frenillo lingual con inserción fibrosa anterior, lengua en forma de corazón cuando es proyectada y dificultad para articular determinados fonemas. Se enviaron interconsultas con Fonoaudiología y Pediatría. Luego de la firma del Consentimiento informado por el responsable y la respuesta de las interconsultas, se propuso como plan de tratamiento la desinserción quirúrgica: Frenectomía. La cirugía se realizó con éxito. La paciente fue derivada al área de fonoaudiología para trabajar los fonemas con dificultad. Conclusión: La frenectomía permitió recuperar funciones de la lengua y lograr ejecutarlas de manera adecuada.

Exposure to a Glyphosate-based Herbicide Alters the Expression of Key Regulators of Mammary Gland Development on Pre-pubertal Male Rats.

We previously reported that exposure during gestation and lactation to a low dose of glyphosate-based herbicide (GBH) reduced the area and perimeter of male offspring mammary gland at postnatal day 60 (PND60), whereas a higher dose increased the longitudinal growth of the gland. Here, our aim was to assess whether perinatal exposure to GBH exhibits endocrine disruptive action in male mammary gland at an early time point (pre-puberty), which could be related to the changes observed after puberty. We also wanted to explore whether an early evaluation of the male rat mammary gland is appropriate to assess exposure to potential endocrine disrupting chemicals (EDCs). Pregnant rats were orally exposed, through the diet, to vehicle (saline solution), 3.5 or 350 mg/kg/day of GBH from gestational day 9 until weaning. At PND21, the male offspring were euthanized, and mammary gland samples were collected. The histology and proliferation index of the mammary glands were evaluated, and the mRNA expression of estrogen (ESR1) and androgen (AR) receptors, cyclin D1 (Ccnd1), amphiregulin (Areg), insulin-like growth factor 1 (IGF1), epidermal growth factor receptor (EGFR) and IGF1 receptor (IGF1R) were assessed. Moreover, the phosphorylated-Erk1/2 (p-ERK1/2) protein expression was determined. No differences were observed in mammary epithelial structures and AR expression between experimental groups; however, the proliferation index was reduced in GBH3.5-exposed males. This result was associated with decreased ESR1, Ccnd1, Areg, IGF1, EGFR and IGF1R mRNA expressions, as well as reduced p-Erk1/2 protein expression in these animals. ESR1, Ccnd1, IGF1R and EGFR expressions were also reduced in GBH350-exposed males. In conclusion, the mammary gland development of pre-pubertal male rats is affected by perinatal exposure to GBH. Although further studies are still needed to understand the molecular mechanisms involved in GBH350 exposure, the present results may explain the alterations observed in mammary gland growth of post-pubertal males exposed to low doses of GBH. Our results also suggest that early evaluation of the male rat mammary gland is useful in assessing exposure to potential EDCs. However, analysis of EDCs effects at later time points should not be excluded.

AhR ligands reactivate LINE-1 retrotransposon in triple-negative breast cancer cells MDA-MB-231 and non-tumorigenic mammary epithelial cells NMuMG.

Breast cancer is the most common cancer type in females worldwide. Environmental exposure to pesticides affecting hormonal homeostasis does not necessarily induce DNA mutations but may influence gene expression by disturbances in epigenetic regulation. Expression of long interspersed nuclear element-1 (LINE-1) has been associated with tumorigenesis in several cancers. In nearly all somatic cells, LINE-1 is silenced by DNA methylation in the 5́'UTR and reactivated during disease initiation and/or progression. Strong ligands of aryl hydrocarbon receptor (AhR) activate LINE-1 through the transforming growth factor-ß1 (TGF-ß1)/Smad pathway. Hexachlorobenzene (HCB) and chlorpyrifos (CPF), both weak AhR ligands, promote cell proliferation and migration in breast cancer cells, as well as tumor growth in rat models. In this context, our aim was to examine the effect of these pesticides on LINE-1 expression and ORF1p localization in the triple-negative breast cancer cell line MDA-MB-231 and the non-tumorigenic epithelial breast cell line NMuMG, and to evaluate the role of TGF-ß1 and AhR pathways. Results show that 0.5 µM CPF and 0.005 µM HCB increased LINE-1 mRNA expression through Smad and AhR signaling in MDA-MB-231. In addition, the methylation of the first sites in 5́'UTR of LINE-1 was reduced by pesticide exposure, although the farther sites remained unaffected. Pesticides modulated ORF1p localization in MDA-MB-231: 0.005 µM HCB and 50 µM CPF increased nuclear translocation, while both induced cytoplasmic retention at 0.5 and 5 µM. Moreover, both stimulated double-strand breaks, enhancing H2AX phosphorylation, coincidentally with ORF1p nuclear localization. In NMuMG similar results were observed, since they heighten LINE-1 mRNA levels. CPF effect was through AhR and TGF-ß1 signaling, whereas HCB action depends only of AhR. In addition, both pesticides increase ORF1p expression and nuclear localization. Our results provide experimental evidence that HCB and CPF exposure modify LINE-1 methylation levels and induce LINE-1 reactivation, suggesting that epigenetic mechanisms could contribute to pesticide-induced breast cancer progression.

Relationship between agrochemical compounds and mammary gland development and breast cancer.

The exposure to agrochemical pesticides has been associated with several chronic diseases, including different types of cancer and reproductive disorders. In addition, because agrochemical pesticides may act as endocrine disrupting chemicals (EDCs) during different windows of susceptibility, they can increase the risk of impairing the normal development of the mammary gland and/or of developing mammary lesions. Therefore, the aim of this review is to summarize how exposure to different agrochemical pesticides suspected of being EDCs can interfere with the normal development of the mammary gland and the possible association with breast cancer. It has been shown that the mammary glands of male and female rats and mice are susceptible to exposure to non-organochlorine (vinclozolin, atrazine, glyphosate, chlorpyrifos) and organochlorine (endosulfan, methoxychlor, hexachlorobenzene) pesticides. Some of the effects of these compounds in experimental models include increased or decreased mammary development, impaired cell proliferation and steroid receptor expression and signaling, increased malignant cellular transformation and tumor development and angiogenesis. Contradictory findings have been found as to whether there is a causal link between the exposure or the pesticide body burden and breast cancer in humans. However, an association has been observed between pesticides (especially organochlorine compounds) and specific subtypes of breast cancer. Further studies are needed in both humans and experimental models to understand how agrochemical pesticides can induce or promote changes in the development, differentiation and/or malignant transformation of the mammary gland.

Hyperandrogenism Induces Histo-Architectural Changes in the Rat Uterus.

The effects of androgens on the uterus have been poorly studied and they need to be clarified to understand why androgen excess, such as observed in women with polycystic ovary syndrome (PCOS), is a risk factor for the development of endometrial hyperplasia, cancer, and infertility. Thus, uterine histomorphology in a PCOS experimental model was evaluated. Beginning at weaning, female rats were injected daily with dehydroepiandrosterone (DHEA, 6 mg/100 g body weight) or vehicle (sesame oil) for 20 consecutive days. On postnatal day 41 (PND41), DHEA-treated animals showed high serum testosterone levels. In addition, uterine histological analysis showed a significant increase in luminal epithelial height and glandular density without changes in cell proliferation. The thickness of the subepithelial stroma and myometrium also increased in these animals. The effect of DHEA on uterine thickness was accompanied by a significant reduction in cell density in both tissue compartments (subepithelial stroma and myometrium). Cell proliferation was not altered in the myometrium, whereas a decrease in the proliferative activity was seen at PND41 in the subepithelial stroma of DHEA animals. The analysis of the extracellular space showed that the changes in the thickness of the subepithelial stroma and myometrium were related to an increase in the organization of collagen fibers and water imbibition. The latter was associated with higher aquaporin 3 and 8 expression. This study provides evidence to further the understanding of PCOS-associated hyperandrogenism effects on uterine architecture. This could have implications for the regulation of uterine function and the development of uterine lesions.

Male mammary gland development and methylation status of estrogen receptor alpha in Wistar rats are modified by the developmental exposure to a glyphosate-based herbicide.

Postnatal treatment with glyphosate-based herbicides (GBHs) induces endocrine-disrupting effects on the male rat mammary gland. In this study, the effects of developmental exposure to GBH on mammary gland growth and development, and the possible molecular mechanisms involved, were evaluated in pre- and post-pubertal male rats. To this end, pregnant rats were orally exposed through the food to 0, 3.5 or 350 mg GBH/kg bw/day from gestational day 9 until weaning. Mammary gland development and estradiol (E2) and testosterone (T) serum levels of male offspring were evaluated on postnatal day (PND)21 and PND60. Besides, prolactin (PRL) serum levels, proliferation index, androgen (AR) and estrogen receptor alpha (ESR1) expression, ESR1 alternative transcript mRNA levels, and DNA methylation status of ESR1 promoters were assessed on PND60. No differences between groups were observed in mammary gland development at PND21 or in E2 and T levels on both PNDs studied. On PND60, GBH3.5-exposed animals presented similar mammary gland histology but higher AR protein expression than controls, whereas GBH350-exposed males presented a less developed mammary gland, accompanied by a lower proliferation index, similar AR levels, and slightly increased PRL serum levels than controls. In both exposed groups, ESR1 expression was lower than in control rats, being lower in GBH350-exposed rats. GBH also altered the abundance of ESR1 transcript variants by hypermethylation of ESR1 promoters. GHB3.5 decreased only ESR1-OS expression, whereas GBH350 affected ESR1-O, OT and E1 expression. Our results show that developmental exposure to GBH induces epigenetic changes in ESR1, which could be responsible for the altered male mammary gland development observed in GBH350-exposed animals.

Postnatal exposure to a glyphosate-based herbicide modifies mammary gland growth and development in Wistar male rats.

Our aim was to evaluate whether postnatal exposure to a glyphosate-based herbicide (GBH) modifies mammary gland development in pre- and post-pubertal male rats. From postnatal day 1 (PND1) to PND7, male rats were injected subcutaneously every 48 h with either saline solution (vehicle) or 2 mg GBH/kg·bw. On PND21 and PND60, mammary gland and blood samples were collected. Estradiol (E2) and testosterone (T) serum levels, mammary gland histology, collagen fiber organization, mast cell infiltration, proliferation index, and estrogen (ESR1) and androgen receptor (AR) expression levels were evaluated. At PND21, GBH-exposed male rats exhibited greater development of the mammary gland with increased stromal collagen organization and terminal end buds (TEBs) compared to control rats. At PND60, the number of TEBs remained high and was accompanied by an increase in mast cell infiltration, proliferation and ESR1 expression in GBH-exposed male rats. In contrast, no effects were observed in E2 and T serum levels and AR expression in both days studied. Our results showed that a postnatal subacute treatment with GBH induces endocrine-disrupting effects in the male mammary gland in vivo, altering its normal development.

Early postnatal exposure to endosulfan interferes with the normal development of the male rat mammary gland.

Our aim was to evaluate whether postnatal exposure to endosulfan (ENDO) modifies mammary gland (MG) development in pre- and post-pubertal male rats. From postnatal day 1 (PND1) to PND7, male rats were injected subcutaneously every 48h with either corn oil (vehicle) or 600µg ENDO/kg.bw. On PND21 and PND60, MG and blood samples were collected. Estradiol (E2) and testosterone (T) serum levels, MG histology, collagen fiber organization, proliferation index, and estrogen (ESR1) and androgen receptor (AR) expressions were evaluated. On PND21, E2 and T levels were similar between groups, whereas MG area, perimeter, number of terminal end buds and ESR1 expression were increased in ENDO-exposed rats. These changes were associated with alveolar development and increased organized collagen in the stroma. On PND60, a higher proliferation index in ENDO-exposed rats was correlated with a more developed lobuloalveolar structure. Hyperplastic alveoli and, hyperplastic ducts surrounded by a dense stroma were also observed in this group. T levels and ESR1 expression were similar between groups, whereas E2 levels and AR expression were decreased in ENDO-exposed rats. The exposure to ENDO in the first week of life interferes with the normal development of the MG and induces pre-malignant lesions in post-pubertal male rats.

Perinatal Exposure to Bisphenol A or Diethylstilbestrol Increases the Susceptibility to Develop Mammary Gland Lesions After Estrogen Replacement Therapy in Middle-Aged Rats.

The development of the mammary gland is a hormone-regulated event. Several factors can dysregulate its growth and make the gland more susceptible to cellular transformation. Among these factors, perinatal exposure to xenoestrogens and hormone replacement therapy has been associated with increased risk of developing breast cancer. Here, we assessed the effects induced by estrogen replacement therapy (ERT) in ovariectomized (OVX) middle-aged rats and whether perinatal exposure to diethylstilbestrol (DES) or bisphenol A (BPA) modified these effects in the mammary gland. Pregnant rats were orally exposed to vehicle, 5 µg DES/kg/day, or 0.5 or 50 µg BPA/kg/day from gestational day 9 until weaning. Then, 12-month-old offspring were OVX and treated with 17ß-estradiol for 3 months. Morphological changes and the percentage of epithelial cells that proliferated or expressed estrogen receptor alpha (ESR1) and progesterone receptor (PR) were analyzed in mammary gland samples of 15-month-old animals. ERT induced lobuloalveolar hyperplasia and ductal cysts in the mammary gland of middle-aged rats, associated with a higher proliferation index of epithelial cells. Perinatal exposure to DES followed by ERT increased the number of cysts and induced the formation of fibroadenoma and ductal carcinoma in situ, without modifying the expression of ESR1 or PR. Also, after 3 months of ERT, BPA-exposed rats had a higher incidence of ductal hyperplasia and atypical lobular hyperplasia than animals under ERT alone. In conclusion, perinatal exposure to xenoestrogens increases the susceptibility of the mammary gland to develop cysts and hyperplastic lesions when confronted with ERT later in life.

Perinatal exposure to bisphenol A modifies the transcriptional regulation of the ß-Casein gene during secretory activation of the rat mammary gland.

With the aim to analyze whether bisphenol A (BPA) modifies ß-Casein (ß-Cas) synthesis and transcriptional regulation in perinatally exposed animals, here, pregnant F0 rats were orally exposed to 0, 0.6 or 52 µg BPA/kg/day from gestation day 9 until weaning. Then, F1 females were bred and mammary glands were obtained on lactation day 2. Perinatal BPA exposure decreased ß-Cas expression without modifying the activation of prolactin receptor. It also decreased the expression of glucocorticoid receptor in BPA52-exposed dams and ß1 and α6 integrins as well as dystroglycan in both BPA groups. In addition, BPA exposure altered the expression of histone-modifying enzymes and induced histone modifications and DNA methylation in the promoter, enhancer and exon VII of the ß-Cas gene. An impaired crosstalk between the extracellular matrix and lactogenic hormone signaling pathways and epigenetic modifications of the ß-Cas gene could be the molecular mechanisms by which BPA decreased ß-Cas expression.

Milk lipid composition is modified by perinatal exposure to bisphenol A.

To evaluate whether bisphenol A (BPA) modifies the synthesis, composition and/or profile of fatty acids (FAs) in the mammary glands of perinatally exposed animals, pregnant rats were orally exposed to 0, 0.6 or 52 µg BPA/kg/day from gestation day (GD) 9 until weaning. F1 females were bred, and on GD21, lactation day 2 (LD2) and LD10, mammary glands were obtained. On LD10, milk samples were collected, and FA profiles and lipid compositions were established. On GD21 and LD2, BPA exposure delayed mammary alveolar maturation and modified the synthesis of milk fat globules. On LD10, mammary gland histo-architecture was restored; however, the milk of BPA-exposed F1 dams had a FA profile and lipid concentration different from those of the control milk. Furthermore, the body weight gain of BPA52 F2 pups was increased compared with control animals. Thus, perinatal exposure to BPA modifies milk quality, compromising the normal growth of offspring.

Prenatal Bisphenol A exposure delays the development of the male rat mammary gland.

Our aims were to evaluate whether exposure to Bisphenol A (BPA) modifies the development of the male rat mammary gland (MG) and to evaluate whether this modification is gender specific. From gestational day 9, pregnant rats were exposed either subcutaneously to 0, 25 or 250µg BPA/kgbw/day until parturition or orally to 0 and 64µg BPA/kgbw/day until weaning. MG development was analyzed on postnatal days (PND) 5, 15 and 30. On PND30, steroid hormone receptor expression and mammary growth were also evaluated. On PND30, the exposure to 64BPA and 250BPA induced a delay in male MG development, evidenced by reduced ductal growth, decreased number of terminal structures and lower expression of androgen receptor (AR). In contrast, female mammary ductal growth was altered only by 250BPA. Regardless of the administration route and length of the exposure period, BPA induced a delay in MG development and modified AR expression in prepubertal male rats.