Cell death induced by serum deprivation in luteal cells involves the intrinsic pathway of apoptosis.
The corpus luteum is a transient endocrine gland specializing in the production of progesterone. The regression of the corpus luteum involves an abrupt decline in its capacity for producing progesterone followed by its structural involution, which is associated with apoptosis of the luteal cells. An in vitro experimental approach is needed to study the molecular mechanisms underlying hormonal regulation of luteal cell death under defined experimental conditions. In this study, we investigated simian virus-40-transformed luteal cells to determine whether they can be driven to apoptosis and, if so, to define the intracellular pathway involved. Luteal cells were cultured in the presence or absence of fetal bovine serum for 24 or 48 h. Under serum starvation conditions, the luteal cells underwent growth arrest accompanied by cell death as evaluated by dye exclusion, and confirmed by two-color fluorescence cell viability/cytotoxicity assay. We next studied whether serum starvation-induced death of luteal cells occurred by apoptosis. Morphologic features of apoptosis were observed in cells stained with hematoxylin after being subjected to serum starvation for 48 h. The apoptotic nature was further confirmed by in situ 3'-end labeling and fragmentation of genomic DNA. Apoptosis of serum-deprived luteal cells was dependent upon caspase activation. Serum starvation induced cleavage of poly (ADP-ribose) polymerase (PARP), suggesting that caspase-3 had been activated under the stress of withdrawal of growth factors. This was confirmed by cleavage of full-length procaspase-3. Finally, the fact that serum starvation promoted the cleavage of full-length procaspase-9 and the decrease in the expression of endogenous Bid, a BH-3-only proapoptotic protein of the Bcl-2 family, indicates that the intrinsic (i.e., mitochondrial) pathway of apoptosis was activated. In summary, we have characterized an in vitro experimental model of luteal cell death that can be utilized to evaluate the role of hormones in apoptosis of luteal cells under defined culture conditions, and to study the mechanism of luteal regression.
Goyeneche
Alicia A
AA
Division of Basic Biomedical Sciences, University of South Dakota School of Medicine, 414 East Clark Street, Vermillion, 57069, USA.
Harmon
Jacquelyn M
JM
Telleria
Carlos M
CM
D000818:Animals; D017209:Apoptosis; D051023:BH3 Interacting Domain Death Agonist Protein / Q000378:metabolism; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D002417:Cattle; D002461:Cell Line, Transformed; D053938:DNA Fragmentation; D004789:Enzyme Activation; D005260:Female; D020287:In Situ Nick-End Labeling; D008184:Luteal Cells / Q000473:pathology*; D008928:Mitochondria / Q000473:pathology*; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D051381:Rats; D044967:Serum; D015398:Signal Transduction
D051023:BH3 Interacting Domain Death Agonist Protein; D011065:Poly(ADP-ribose) Polymerases; C505410:Casp3 protein, rat; C506435:Casp9 protein, rat; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases