ABSTRACT
Recent research has demonstrated that cell cycle-associated molecules are activated in multiple forms of cell death in mature neurons, and raised a hypothesis that unscheduled cell cycle activity leads to neuronal cell death. But there is little evidence that changes in endogenous level of these molecules are causally associated with neuronal cell death. Here we transfected small interfering RNA (siRNA) targeting cyclin-dependent kinase (CDK) inhibitor p27, which plays an important role in cell cycle arrest at G1-S phase, into cultured cortical neurons. Transfection of p27 siRNA reduced neuronal viability in a time-dependent manner. p27 siRNA induced phosphorylation of retinoblastoma protein (Rb), a marker of cell cycle progression at late G1 phase. Moreover, phosphorylation of Rb and neuronal cell death provoked by p27 siRNA were abrogated by pharmacological CDK inhibitors, olomoucine and purvalanol A. Our data demonstrate that a decrease in endogenous p27 induces neuronal cell death through elevating cell cycle activity.
Subject(s)
Apoptosis/physiology , Cell Cycle/physiology , Cerebral Cortex/cytology , Cyclin-Dependent Kinase Inhibitor p27/antagonists & inhibitors , Neurons/metabolism , RNA Interference , Animals , Apoptosis/drug effects , Apoptosis/genetics , Cells, Cultured , Cyclin-Dependent Kinase Inhibitor p27/genetics , Kinetin/pharmacology , Neurons/cytology , Phosphorylation , Protein Kinase Inhibitors/pharmacology , Purines/pharmacology , RNA, Small Interfering/physiology , Rats , Rats, Wistar , Retinoblastoma Protein/metabolismABSTRACT
Inhibin activity was investigated using chicken granulosa cell culture medium. Chicken granulosa cells were collected individually from the largest follicle (F1 follicle) and the third largest follicle (F3 follicle) of the ovary, dispersed using enzyme and incubated in a suspension method or cultured in a monolayer method in vitro. Culture medium was assessed for follicle-stimulating hormone (FSH) inhibiting (inhibin) activity using the rat anterior pituitary cell culture method. The FSH secretion from cultured rat pituitary cells was suppressed by experimental increasing of the amounts of granulosa cell culture medium in a dose-dependent manner, though luteinizing hormone (LH) secretion was not affected in a dose-dependent manner. Inhibin activity of the granulosa cell culture medium derived from F1 follicles was higher than that from F3 follicles. These results suggest that chicken granulosa cells secrete inhibin-like substance(s) and that the F1 follicle may secrete a larger amount of inhibin than the F3 follicle.
