Postnatal apoptosis, development, and sex difference in the lateral septum of rats.

To determine whether apoptosis is involved in the formation of the structure and morphological sex difference of the lateral septum (LS), the postnatal developmental changes in the number of apoptotic cells were examined in the LS on postnatal day 1 (PD1 = birth day), 4, 6, 8, 11, 16, and 31 in male and female rats. Apoptotic cells were immunohistochemically detected by antibody against single-stranded DNA (ssDNA) or active caspase-3. The volume of the LS was also measured and was found to increase with age. The number of apoptotic cells detected by anti-ssDNA in the LS increased from PD1 to PD8 but decreased after PD11. Also, the LS was divided into dorsal, intermediate, and ventral parts (LSd, LSi, and LSv), and the volume and number of ssDNA-immunoreactive cells in each part were measured on PD6, 8, 11, 16, and 31. In both sexes, a large number of ssDNA-immunoreactive cells was found in the LSd and LSi on PD8 (but not on PD6) and in the LSv on PD6 and PD8. On PD6, the number of active caspase-3-immunoreactive cells was significantly greater in the LSv than in the LSd or LSi, in both sexes. Only the LSi of males had a high number of ssDNA-immunoreacitve cells on PD16; the number was significantly greater than that of females of the same age. However, there was no significant sex difference in the number of active caspase-3-immunoreacitve cells in the LSi on PD16. On PD31, the volume of the LSi was significantly greater in females than in males. There was no sex difference in volume or number of apoptotic cells in the LSd or LSv. These findings indicate that loss of cells due to apoptosis, which is partially caused by activation of caspase-3, occurs in the LS during postnatal development, with regional differences. They also indicate that sex difference in caspase-3-independent apoptosis contributes to morphological sexual differentiation of the LSi.

Migration of erythroblastic islands toward the sinusoid as erythroid maturation proceeds in rat bone marrow.

It has been hitherto considered that mature erythroblasts migrate toward the sinusoid along the cytoplasmic processes of macrophages of erythroblastic islands in bone marrow. Our previous report, however, has demonstrated the morphological features of a mature erythroblastic island passing through the sinusoidal endothelium. In this study, the possibility of migration of erythroblastic islands toward the sinusoid was examined in rat bone marrow by light microscopical histoplanimetry. As a result, the more mature erythroblasts were not regularly arranged in the peripheral direction of the erythroblastic islands. Immature erythroblasts were populated more in the erythroblastic islands away from the sinusoid than in those islands neighboring the sinusoid, whereas mature erythroblasts were more in erythroblastic islands neighboring the sinusoid. These findings suggest that the formation of erythroblastic islands occurs in a region away from the sinusoid, and that erythroblastic islands migrate towards the sinusoids as erythroid maturation proceeds.

No apoptotic cell death of erythroid cells of erythroblastic islands in bone marrow of healthy rats.

A possibility of apoptotic cell death in erythropoietic regulation was examined by means of detailed light microscopical histoplanimetry, electron microscopy, the in situ nick-end labeling method, and an immunohistological method in the rat bone marrow. Serum erythropoietin concentrations were shown at normal levels. The erythroid series on a mature process presented several morphological features of apoptosis, i.e. the shrinkage of both nuclei and cytoplasm and the chromatin condensation. In the light microscopical histoplanimetry, however, morphological signs of final apoptotic cell death were never found in any erythroid cell within the erythroblastic islands. This finding was also supported by detailed ultrastructural observation: No erythroid cell bodies were trapped and degraded by the central macrophages of the erythroblastic islands, while the denucleated nuclei with small amount of cytoplasm of late erythroblasts were often trapped and degraded in the macrophages. Nuclear DNA fragmentation was not detected in any erythroblasts, but was detected in the lysosomes of the central macrophages. These findings suggest that erythropoiesis is regulated by other regulatory mechanisms than apoptotic cell death. An additional ultrastructural finding shows that the reticulocytes anchored to the central macrophages are transported into the peripheral blood circulation.