ABSTRACT
The quality of neuronal differentiation and reduction in apoptosis that occurred in two-dimensional (2D) and three-dimensional (3D) culture conditions is compared. PC12 and embryonic stem cells are two commonly utilized cell lines for the study of neuronal regeneration. These cells were induced to neuronally differentiate by adding NGF and retinoic acid respectively. Total neurite length and expression of neuronal markers (MAP-2 and β3-tubulin) was assessed by morphometry and immunocytochemistry. Also, TUNEL assay was used to detect apoptosis. Upon exposure to a differentiation media in the 3D fibrin gel, PC12 and embryonic stem cells stopped dividing, had increased adhesion to the substratum, extended neurite processes and expressed neuronal markers. The same results, however, were not observed with the 2D culture. Also, the apoptosis index performed by TUNEL assay demonstrated a reduction in the degree of apoptosis in the 3D culture compared to 2D culture. Fibrin matrix supports growth and neuronal differentiation of PC12 and embryonic stem cells. In addition, the 3D culture enhanced cellular resistance to apoptosis when compared to the 2D culture. It appears as if a 3D culture system may offer a better technique for future neuronal tissue engineering investigations.
ABSTRACT
Considering the size of some nuclei and area, sex hormones control the sexual development of the brain. The sexual development of the brain can also be influenced by environmental stress. This study aimed to clear the effect of prenatal water deprivation on the development of sexual dimorphic nucleus (SDN) of the brain. In this research, pregnant rats were divided into two groups (control and treated). For the treated animals, water was removed from the ewes for 48 h at the end of third trimester of gestation (19-21 days). TUNEL staining was used for detection of apoptosis in paraffin embedded diencephalon selected sections. The ratio of apoptotic cells to non- apoptotic ones was calculated as apoptotic index. Differences of apoptotic index and serum testosterone were examined for statistical significance using Paired T- test (p<0.05). The apoptotic index was 0.0160±.01174 percent for control and 0.1870±.02541 percent for treated groups. The concentration of serum testosterone was 22.4±1.3 for control and 13.37±3.3 for treated groups. Prenatal water deprivation induces apoptosis in developing SDN nucleus of male rats that is derived by reducing the concentration of serum testosterone. The study shows the importance of low concentration acting testosterone for development of SDN nucleus that can be affected by environmental stress.
Considerando el tamaño de algunos núcleos y áreas, las hormonas sexuales controlan el desarrollo sexual del cerebro. El desarrollo sexual del cerebro también puede verse influido por el estrés ambiental. Este estudio tuvo como objetivo establecer el efecto de la privación prenatal de agua en el desarrollo del núcleo dimórfico sexual (NDS) del cerebro. Las ratas preñadas fueron divididas en dos grupos (control y tratados). Para los animales tratados, el agua se retiró de los bebederos durante 48h al final del tercer trimestre de gestación (días 19-21). La técnica TUNEL se utilizó para detectar apoptosis en secciones del diencéfalo incluidas en parafina. La proporción de células apoptóticas y no-apoptóticas fue calculada como índice de apoptosis. Las diferencias del índice de apoptosis y testosterona sérica fueron examinadas para observar significación estadística mediante t de Student pareado (p <0,05). El índice de apoptosis fue 0,0160±0,01174 por ciento para el control y 0,1870±0,02541 por ciento para los grupos tratados. La concentración de testosterona en suero fue de 22,4±1,3 para el grupo control y 13,37±3,3 para los grupos tratados. La privación de agua prenatal indujo la apoptosis en el desarrollo del NDS de las ratas macho derivadas por la reducción de concentración de testosterona sérica. El estudio muestra la importancia de una baja concentración de testosterona para el desarrollo de los NDS, que pueden verse afectados por el estrés ambiental.