RESUMEN
Female mice lacking the follistatin gene but expressing a human follistatin-315 transgene (tghFST315) have reproductive abnormalities (reduced follicles, no corpora lutea and ovarian-uterine inflammation). We hypothesised that the absence of follistatin-288 causes the abnormal reproductive tract via both developmental abnormalities and abnormal ovarian activity. We characterised the morphology of oviducts and uteri in wild type (WT), tghFST315 and follistatin-knockout mice expressing human follistatin-288 (tghFST288). The oviducts and uteri were examined in postnatal Day-0 and adult mice (WT and tghFST315 only) using histology and immunohistochemistry. Adult WT and tghFST315 mice were ovariectomised and treated with vehicle, oestradiol-17ß (100ng injection, dissection 24h later) or progesterone (1mg×three daily injections, dissection 24h later). No differences were observed in the oviducts or uteri at birth, but abnormalities developed by adulthood. Oviducts of tghFST315 mice failed to coil, the myometrium was disorganised, endometrial gland number was reduced and oviducts and uteri contained abundant leukocytes. After ovariectomy, tghFST315 mice had altered uterine cell proliferation, and inflammation was maintained and exacerbated by oestrogen. These studies show that follistatin is crucial to postnatal oviductal-uterine development and function. Further studies differentiating the role of ovarian versus oviductal-uterine follistatin in reproductive tract function at different developmental stages are warranted.
Asunto(s)
Folistatina/genética , Oviductos/crecimiento & desarrollo , Útero/crecimiento & desarrollo , Animales , Proliferación Celular/genética , Endometrio/crecimiento & desarrollo , Endometrio/metabolismo , Estrógenos/farmacología , Femenino , Folistatina/metabolismo , Regulación del Desarrollo de la Expresión Génica , Ratones , Ratones Noqueados , Ratones Transgénicos , Miometrio/crecimiento & desarrollo , Miometrio/metabolismo , Ovariectomía , Oviductos/diagnóstico por imagen , Oviductos/metabolismo , Útero/efectos de los fármacos , Útero/metabolismoRESUMEN
Follistatin, an inhibitor of activin A, has key regulatory roles in the female reproductive tract. Follistatin has two splice variants: FST288, largely associated with cell surfaces, and FST315, the predominant circulating form. The mechanism regulating uterine expression of these variants is unknown. Quantitative RT-PCR was used to measure expression of follistatin splice variants (Fst288, Fst315), the activin bA subunit (Inhba) and the inhibin a subunit (Inha) in uterine tissues during early pregnancy (days 14, preimplantation) and in response to exogenous 17b-oestradiol (single s.c. injection) and progesterone (three daily s.c. injections) in ovariectomized mice. Uterine Fst288, Fst315 and Inhba expression increased during early pregnancy, with greater increases in Fst315 relative to Fst288 suggesting differential regulation of these variants. Fst288, Fst315, Inhba and Inha all increased in response to progesterone treatment. Fst288, but not Fst315, mRNA decreased in response to 17b-oestradiol treatment, whereas Inhba increased. A comparison of the absolute concentrations of uterine follistatin mRNA using crossing thresholds indicated that both variants were more highly expressed in early pregnancy in contrast to the hormone treatment models. It is concluded that progesterone regulates uterine expression of both follistatin variants, as well as activin A, during early pregnancy in the mouse uterus
Asunto(s)
Folistatina/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Progesterona/farmacología , Útero/efectos de los fármacos , Animales , Estradiol/farmacología , Femenino , Folistatina/química , Folistatina/genética , Subunidades beta de Inhibinas/genética , Subunidades beta de Inhibinas/metabolismo , Inhibinas/genética , Inhibinas/metabolismo , Ratones , Embarazo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , ARN Mensajero/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Útero/metabolismoRESUMEN
Identifying suitable housekeeping genes for quantitative RT-PCR in the uterus is problematic, as this tissue undergoes significant structural and functional alterations during the oestrous cycle and pregnancy in response to circulating hormones. The suitability of 18S rRNA as a housekeeping gene in mouse uterus was investigated by introducing an 'RNA spike' standard into the reverse transcription reaction. 18S rRNA levels increased by Day 4 of pregnancy and after progesterone administration in ovariectomized mice. We conclude that 18S rRNA is not a suitable housekeeping gene for quantitative RT-PCR analysis in progesterone-responsive tissues, and the RNA spiking method provides a suitable alternative.