In contrast to adult stem cells, induced pluripotent stem cells (iPSCs) can be grown robustly in ... more In contrast to adult stem cells, induced pluripotent stem cells (iPSCs) can be grown robustly in vitro and differentiated into virtually any tissue, thus providing an attractive alternative for biomedical applications. Although iPSC technology is already being used in human biomedicine, its potential in animal production has not been investigated. Herein, we investigated the potential application of iPSCs in dairy production by generating bovine iPSCs and establishing their ability to generate mammary epithelial tissue. iPSCs were derived by retrovirus-mediated expression of murine Oct4, Sox2, Klf4, and c-Myc in mammary epithelium and dermal fibroblasts. The resulting reprogrammed cells stained positive for alkaline phosphatase and showed renewed expression of pluripotency genes, including Lin28, Rex1, Oct4, Sox2, and Nanog. In addition, injection of epithelial- or fibroblast-derived reprogrammed cells into nonobese diabetic (NOD/NOD) mice resulted in the formation of teratomas containing differentiated derivatives of the three germ layers, including cartilage, membranous ossification, stratified squamous epithelial tissue, hair follicles, neural pinwheels, and different types of glandular tissue. Finally, mammary epithelium-derived iPSCs could be induced to differentiate back to a mammary phenotype characterized by epithelial cells expressing cytokeratin 14 (CK14), CK18, and smooth muscle actin (SMA) as a result of treatment with 10 nM progesterone. This study reports for the first time the generation of iPSCs from bovine epithelial cells and demonstrates the potential of using iPSCs technology for generating bovine mammary tissue in vitro.
Osteosarcoma is the most common primary malignant bone tumour in dogs, characterized by a locally... more Osteosarcoma is the most common primary malignant bone tumour in dogs, characterized by a locally aggressive and highly metastatic behaviour. Despite the current standards of care, most dogs succumb to the disease, indicating the need for novel treatment strategies. Polo‐like kinase 1 (PLK1) is dysregulated in a variety of human cancer types, including osteosarcoma, and induces c‐Myc accumulation. The crosstalk between the two molecules coordinates cell proliferation, differentiation, self‐renewal and apoptosis. Therefore, PLK1 has recently emerged as a potential therapeutic target, mainly in tumours overexpressing c‐Myc. BI 2536 is a selective PLK1 inhibitor promoting mitotic arrest and apoptosis in a variety of cancer cells. This research aimed at evaluating PLK1 and c‐Myc protein expression in 53 appendicular canine osteosarcoma (cOSA) samples and the in vitro effects of BI 2536 on a c‐Myc and PLK1‐overexpressing cOSA cell line (D17). PLK1 and c‐Myc expression in cOSA samples showed no correlation with clinicopathological data. However, c‐Myc overexpression was associated with a significantly reduced overall survival (p = .003). Western Blot and RT‐qPCR assays revealed that D17 expressed high protein and transcript levels of both PLK1 and MYC. When treated with BI 2536 (range 2.5–15 nM) for 24 h, D17 showed a substantial decrease in cell growth, inducing apoptosis and G2/M cell cycle arrest. Interestingly, under BI 2536 treatment, D17 showed decreased c‐Myc protein levels. Consistent with human OSA, these preliminary data outline the prognostic value of c‐Myc expression in cOSA and highlight the potential role of PLK1 as an antiproliferative therapeutic target for tumours overexpressing c‐Myc.
During the early stage of placentation in sheep, normal conceptus development is affected by trop... more During the early stage of placentation in sheep, normal conceptus development is affected by trophoblast cell functionality, whose dysregulation results in early pregnancy loss. Trophoblast metabolism is supported mainly by histotrophic factors, including fibroblast growth factor-2 (FGF2), which are involved in cell differentiation and function through the modulation of specific cellular mechanisms. The mechanistic target of rapamycin (mTOR) is known as a cellular ‘nutrient sensor’, but its downstream regulation remains poorly understood. The hypothesis was that during trophoblast development the FGF2 effect is mediated by mTOR signalling pathway modulation. Primary trophoblast cells from 21-day-old sheep placenta were characterised and subjected to FGF2 and rapamycin treatment to study the effects on cell functionality, gene and protein expression profiles. The model showed mainly mononuclear cells with epithelial cell-like growth and placental morphological properties, expressing ...
The phytoestrogen genistein (GEN) may interfere with permanent morphological changes in the brain... more The phytoestrogen genistein (GEN) may interfere with permanent morphological changes in the brain circuits sensitive to estrogen. Due to the frequent use of soy milk in the neonatal diet, we aimed to study the effects of early GEN exposure on some physiological and reproductive parameters. Mice of both sexes from PND1 to PND8 were treated with GEN (50 mg/kg body weight, comparable to the exposure level in babies fed with soy-based formulas). When adult, we observed, in GEN-treated females, an advanced pubertal onset and an altered estrous cycle, and, in males, a decrease of testicle weight and fecal testosterone concentration. Furthermore, we observed an increase in body weight and altered plasma concentrations of metabolic hormones (leptin, ghrelin, triiodothyronine) limited to adult females. Exposure to GEN significantly altered kisspeptin and POMC immunoreactivity only in females and orexin immunoreactivity in both sexes. In conclusion, early postnatal exposure of mice to GEN det...
In contrast to adult stem cells, induced pluripotent stem cells (iPSCs) can be grown robustly in ... more In contrast to adult stem cells, induced pluripotent stem cells (iPSCs) can be grown robustly in vitro and differentiated into virtually any tissue, thus providing an attractive alternative for biomedical applications. Although iPSC technology is already being used in human biomedicine, its potential in animal production has not been investigated. Herein, we investigated the potential application of iPSCs in dairy production by generating bovine iPSCs and establishing their ability to generate mammary epithelial tissue. iPSCs were derived by retrovirus-mediated expression of murine Oct4, Sox2, Klf4, and c-Myc in mammary epithelium and dermal fibroblasts. The resulting reprogrammed cells stained positive for alkaline phosphatase and showed renewed expression of pluripotency genes, including Lin28, Rex1, Oct4, Sox2, and Nanog. In addition, injection of epithelial- or fibroblast-derived reprogrammed cells into nonobese diabetic (NOD/NOD) mice resulted in the formation of teratomas containing differentiated derivatives of the three germ layers, including cartilage, membranous ossification, stratified squamous epithelial tissue, hair follicles, neural pinwheels, and different types of glandular tissue. Finally, mammary epithelium-derived iPSCs could be induced to differentiate back to a mammary phenotype characterized by epithelial cells expressing cytokeratin 14 (CK14), CK18, and smooth muscle actin (SMA) as a result of treatment with 10 nM progesterone. This study reports for the first time the generation of iPSCs from bovine epithelial cells and demonstrates the potential of using iPSCs technology for generating bovine mammary tissue in vitro.
Osteosarcoma is the most common primary malignant bone tumour in dogs, characterized by a locally... more Osteosarcoma is the most common primary malignant bone tumour in dogs, characterized by a locally aggressive and highly metastatic behaviour. Despite the current standards of care, most dogs succumb to the disease, indicating the need for novel treatment strategies. Polo‐like kinase 1 (PLK1) is dysregulated in a variety of human cancer types, including osteosarcoma, and induces c‐Myc accumulation. The crosstalk between the two molecules coordinates cell proliferation, differentiation, self‐renewal and apoptosis. Therefore, PLK1 has recently emerged as a potential therapeutic target, mainly in tumours overexpressing c‐Myc. BI 2536 is a selective PLK1 inhibitor promoting mitotic arrest and apoptosis in a variety of cancer cells. This research aimed at evaluating PLK1 and c‐Myc protein expression in 53 appendicular canine osteosarcoma (cOSA) samples and the in vitro effects of BI 2536 on a c‐Myc and PLK1‐overexpressing cOSA cell line (D17). PLK1 and c‐Myc expression in cOSA samples showed no correlation with clinicopathological data. However, c‐Myc overexpression was associated with a significantly reduced overall survival (p = .003). Western Blot and RT‐qPCR assays revealed that D17 expressed high protein and transcript levels of both PLK1 and MYC. When treated with BI 2536 (range 2.5–15 nM) for 24 h, D17 showed a substantial decrease in cell growth, inducing apoptosis and G2/M cell cycle arrest. Interestingly, under BI 2536 treatment, D17 showed decreased c‐Myc protein levels. Consistent with human OSA, these preliminary data outline the prognostic value of c‐Myc expression in cOSA and highlight the potential role of PLK1 as an antiproliferative therapeutic target for tumours overexpressing c‐Myc.
During the early stage of placentation in sheep, normal conceptus development is affected by trop... more During the early stage of placentation in sheep, normal conceptus development is affected by trophoblast cell functionality, whose dysregulation results in early pregnancy loss. Trophoblast metabolism is supported mainly by histotrophic factors, including fibroblast growth factor-2 (FGF2), which are involved in cell differentiation and function through the modulation of specific cellular mechanisms. The mechanistic target of rapamycin (mTOR) is known as a cellular ‘nutrient sensor’, but its downstream regulation remains poorly understood. The hypothesis was that during trophoblast development the FGF2 effect is mediated by mTOR signalling pathway modulation. Primary trophoblast cells from 21-day-old sheep placenta were characterised and subjected to FGF2 and rapamycin treatment to study the effects on cell functionality, gene and protein expression profiles. The model showed mainly mononuclear cells with epithelial cell-like growth and placental morphological properties, expressing ...
The phytoestrogen genistein (GEN) may interfere with permanent morphological changes in the brain... more The phytoestrogen genistein (GEN) may interfere with permanent morphological changes in the brain circuits sensitive to estrogen. Due to the frequent use of soy milk in the neonatal diet, we aimed to study the effects of early GEN exposure on some physiological and reproductive parameters. Mice of both sexes from PND1 to PND8 were treated with GEN (50 mg/kg body weight, comparable to the exposure level in babies fed with soy-based formulas). When adult, we observed, in GEN-treated females, an advanced pubertal onset and an altered estrous cycle, and, in males, a decrease of testicle weight and fecal testosterone concentration. Furthermore, we observed an increase in body weight and altered plasma concentrations of metabolic hormones (leptin, ghrelin, triiodothyronine) limited to adult females. Exposure to GEN significantly altered kisspeptin and POMC immunoreactivity only in females and orexin immunoreactivity in both sexes. In conclusion, early postnatal exposure of mice to GEN det...
Uploads