EVALUATION OF GROWTH, CELL-PROLIFERATION, AND CELL-DEATH IN BOVINE CORPORA-LUTEA THROUGHOUT THE ESTROUS-CYCLE

To evaluate the kinetics of luteal growth, bovine CL were obtained fro m four stages (stage I, Days 1-4; stage II, Days 5-10; stage III, Days 11-17; stage IV, Days 18-21) of the estrous cycle, and luteal fresh w eight as well as DNA, protein, and progesterone contents was determine d. To evaluate the relative rate of cell proliferation, proliferating cell nuclear antigen (PCNA; a specific marker for cell proliferation) was immunolocalized in paraffin-embedded luteal tissue sections. To ev aluate the relative rate of cell death, nucleosomal fragmentation of D NA (a specific marker for apoptosis) was detected by agarose gel elect rophoresis and also by histochemical localization in paraffin-embedded luteal tissue sections. Luteal fresh weight and DNA, protein, and pro gesterone contents increased (p < 0.01) from stage I to stage II, were similar between stages II and III, and then decreased (p < 0.01) from stage III to stage IV, The ratio of protein to DNA (an index of avera ge cell size) was similar for stages I, II, and III and then decreased (P < 0.01)at stage IV. For stage I(corpora hemorrhagica), most prolif erating (PCNA-positive) cells were located in or around the core of th e tissue infoldings (presumably thecal-derived areas), whereas relativ ely few proliferating cells were located at the periphery of the tissu e infoldings (presumably granulosa-derived areas). For stages II, III, and IV, the majority of proliferating cells appeared to be small cell s (i.e., small parenchymal cells, fibroblasts, and endothelial cells). The labeling index (LI; percentage of cells that were PCNA-positive) was greatest at stage I (20.3 +/- 1.1%); it then decreased (P < 0.01) by stage II and was similar at stages II, III, and TV (3.4 +/- 1.1%). Apoptosis, as determined by evaluation of nucleosomal DNA fragmentatio n by agarose gel electrophoresis and in situ localization, was detecta ble only at stage IV. These data demonstrate that luteal growth from s tage I to stage II resulted from cell proliferation as shown by a high LI at stage I, accompanied by increased luteal DNA content but no cha nge in average cell size, and by similar protein:DNA ratios. Luteal re gression from stage III to stage IV was primarily associated with cell deletion and decreased cell size as shown by a decrease in luteal DNA content and the appearance of apoptosis along with a decrease in the luteal protein:DNA ratio. In addition, analysis of the estimated kinet ics of cell turnover revealed that the number of proliferating luteal cells remained high until stage III and then decreased at stage IV in association with luteal regression.