RESUMO
OBJECTIVE: To compare four enzymatic protocols for mesenchymal stem cells (MSCs) isolation from amniotic (A-MSC) and chorionic (C-MSC) membranes, umbilical cord (UC-MSC) and placental decidua (D-MSC) in order to define a robust, practical and low-cost protocol for each tissue. RESULTS: A-MSCs and UC-MSCs could be isolated from all samples using trypsin/collagenase-based protocols; C-MSCs could be isolated from all samples with collagenase- and trypsin/collagenase-based protocols; D-MSCs were isolated from all samples exclusively with a collagenase-based protocol. CONCLUSIONS: The trypsin-only protocol was least efficient; the collagenase-only protocol was best for C-MSCs and D-MSCs; the combination of trypsin and collagenase was best for UC-MSCs and none of tested protocols was adequate for A-MSCs isolation.
Assuntos
Separação Celular/métodos , Membranas Extraembrionárias/citologia , Células-Tronco Mesenquimais/citologia , Placenta/citologia , Cordão Umbilical/citologia , Proliferação de Células , Células Cultivadas , Colagenases , Feminino , Humanos , Cinética , Gravidez , TripsinaRESUMO
BACKGROUND: Miguel Fernández was an Argentinian zoologist who published the first account of obligate polyembryony in armadillos. His contribution is here discussed in relation to his contemporaries, Newman and Patterson, and more recent work. FINDINGS: Fernandez worked on the mulita (Dasypus hybridus). He was able to get early stages before twinning occurred and show it was preceded by inversion of the germ layers. By the primitive streak stage there were separate embryonic shields and partition of the amnion. There was, however, a single exocoelom and all embryos were enclosed in a common set of membranes comprising chorion towards the attachment site in the uterine fundus and inverted yolk sac on the opposite face. He showed that monozygotic twinning did not occur in another armadillo, the peludo (Chaetophractus villosus). CONCLUSIONS: Fernández's work represented a major breakthrough in understanding how twinning occurred in armadillos. His work and that of others is of intrinsic interest to zoologists and has a direct bearing on the origin of monozygotic twins and birth defects in humans.
Assuntos
Anatomia Comparada/história , Tatus/embriologia , Embriologia/história , Desenvolvimento Embrionário , Camadas Germinativas/embriologia , Gemelaridade Monozigótica , Zoologia/história , Animais , Argentina , Tatus/crescimento & desenvolvimento , Tatus/fisiologia , Membranas Extraembrionárias/citologia , Membranas Extraembrionárias/embriologia , Membranas Extraembrionárias/fisiologia , Feminino , Pesquisa em Genética/história , Camadas Germinativas/citologia , Camadas Germinativas/fisiologia , História do Século XX , Masculino , Placentação , Gravidez , Especificidade da Espécie , Saco Vitelino/citologia , Saco Vitelino/embriologia , Saco Vitelino/fisiologiaRESUMO
Recently, the regenerative medicine has been trying to congregate different areas such as tissue engineering and cellular therapy, in order to offer effective treatments to overcome several human and veterinary medical problems. In this regard, fetal membranes have been proposed as a powerful source for obtainment of multipotent stem cells with low immunogenicity, anti-inflammatory properties and nontumorigenicity properties for the treatment of several diseases, including replacing cells lost due to tissue injuries or degenerative diseases. Morpho-physiological data have shown that fetal membranes, especially the yolk sac and amnion play different functions according to the gestational period, which are direct related to the features of the microenvironment that their cells are subject. The characteristics of the microenvironment affect or controls important cellular events involved with proliferation, division and maintenance of the undifferentiated stage or differentiation, especially acting on the extracellular matrix components. Considering the importance of the microenvironment and the diversity of embryonic and fetal membrane-derived stem cells, this chapter will addressed advances in the isolation, phenotyping, characteristics of the microenvironment, and applications of yolk sac and amniotic membrane-derived stem cells for human and veterinary regenerative medicine.
Assuntos
Diferenciação Celular , Membranas Extraembrionárias/citologia , Células-Tronco Multipotentes/citologia , Nicho de Células-Tronco , Células-Tronco/citologia , Âmnio , Terapia Baseada em Transplante de Células e Tecidos/métodos , Humanos , Medicina Regenerativa/métodos , Engenharia Tecidual/métodosRESUMO
There have been major recent advances in the field of developmental biology due to the investigation on stem cells (SC). Stem cells are characterized by their capacity of auto-renewal and differentiation to different cellular phenotypes. Based on the developmental stage, they can be classified into two different types: embryonic SCs and adult SCs. It has been widely reported that several problems need to be resolved before their possible clinical applications. As a result, fetal membranes have been suggested as an alternative source of SCs. In the human amniotic epithelium, the presence of markers of pluripotent SC´s has been reported, and its capacity as a feeder layer for expansion of different SC types. Also, fetal membranes are a discarded product after delivery, and thus there are not any ethical issues related to its use. In conclusion, the human amniotic epithelium can be a strong candidate for regenerative medicine.
Assuntos
Âmnio/citologia , Células Epiteliais/citologia , Células-Tronco/citologia , Diferenciação Celular , Membranas Extraembrionárias/citologia , Humanos , Medicina Regenerativa/métodosRESUMO
The placenta of mammals is a structure formed by the juxtaposition of the fetal membranes and the maternal tissues. The main function of the placenta is to regulate the physiological interchange between the fetus and the mother as well as to operate as an important endocrine organ during the gestation. The placentomal fusions were characterized throughout gestation of cattle using macroscopic, histological and flow cytometry analyses. Analyzing the cell cycle phases with a flow cytometry, a balance between the G2M phase and apoptosis was observed, suggesting that the placentomal fusions do not interfere in the placentary maturation process, which is a pre-requirement for the fetal-maternal disconnection and the release of fetal membrane.
Assuntos
Bovinos/fisiologia , Placenta/fisiologia , Prenhez/fisiologia , Animais , Ciclo Celular/fisiologia , Proliferação de Células , Membranas Extraembrionárias/citologia , Membranas Extraembrionárias/fisiologia , Feminino , Citometria de Fluxo , Placenta/anatomia & histologia , Placenta/citologia , Gravidez , Útero/fisiologiaRESUMO
Nitric oxide (NO) synthesized by fetal membranes may act either directly inhibiting myometrium contractility or indirectly interacting with tocolytic agents as prostaglandins (PGs). Here we examined if NO could modulate prostaglandin E(2) 9-ketoreductase (9-KPR) activity in human fetal membranes (HFM). 9-KPR is the enzyme that converts PGE(2) into PGF(2alpha), the main PGs known to induce uterine contractility at term. Chorioamnion explants obtained from elective caesareans were incubated with aminoguanidine (AG), an iNOS inhibitor, or NOC-18, a NO donor. NOC-18 (2mM) increased PGE(2) production and diminished PGF(2alpha) synthesis in HFM. AG presented the opposite effect. When we evaluated the activity of 9-KPR by the conversion of [(3)H]-PGE(2) into [(3)H]-PGF(2alpha) and 13,14-dihidro-15-keto prostaglandin F(2alpha) (the PGF(2alpha) metabolite), we found that NOC-18 inhibited 9-KPR activity. Interestingly, AG did not elicit any effect on 9-KPR but l-NAME, a non-selective NOS inhibitor, significantly increased its activity. Our data suggests that exogenous NO inhibits 9-KPR activity in HFM, thus modulating the synthesis of important labor mediators as PGF(2alpha).
Assuntos
Membranas Extraembrionárias/enzimologia , Hidroxiprostaglandina Desidrogenases/metabolismo , Óxido Nítrico/metabolismo , Dinoprosta/metabolismo , Dinoprostona/metabolismo , Relação Dose-Resposta a Droga , Membranas Extraembrionárias/citologia , Membranas Extraembrionárias/efeitos dos fármacos , Feminino , Regulação da Expressão Gênica , Humanos , Hidroxiprostaglandina Desidrogenases/antagonistas & inibidores , Pessoa de Meia-Idade , Doadores de Óxido Nítrico/farmacologia , Óxido Nítrico Sintase/metabolismo , Gravidez , Prostaglandina-Endoperóxido Sintases/metabolismoRESUMO
Diferentes estudios han mostrado asociación entre la disponibilidad de vitamina C ( vit C) y el desarrollo de ruptura prematura de membranas (RPM). Sin embargo, no ha analizado el papel que desempeña la vit C en el metabolismo de la colágena en el tejido corioamniótico. En este trabajo se analizó el efecto de modulación de diferentes concentraciones de vit C en células en cultivo derivativas de amnios humano. Se utilizaron concentraciones de vit C de manera de cubrir el rango fisiológico (29.0 µg/mL). Luego de ser estimuladas, los medios de las células fueron analizados para actividad enzimática de metaloproteasas de matriz extracelular (MMP) y se cuantificó la cantidad relativa de MMP-1, MMP-2 y MMP-9 mediante inmunotransferencia, utilizando anticuerpos policlonales monoespecíficos. Tanto la actividad como la proteína en los medios de las células amnióticas, disminuyó de manera directa a la concentración de vit C, de manera que a las concentraciones más altas probadas (100 µg/mL) se obtuvo la menor actividad/cantidad de MMP. Los resultados anteriores aportan un dato hasta ahora no descrito y que permite establecer una conexión directa entre la disponibilidad de vitamina C y el aumento en la degradación de colágeno. De acuerdo a los resultados, a menor disponibilidad de vit C, mayor degradación de colágena, que debería llevar a pérdida de soporte mecánico y eventual ruptura de las membranas fetales