This study was undertaken to determine whether and how advanced glycation end products (AGE), senescent macroproteins accumulated in various tissues under hyperglycemic states, cause angiogenesis, the principal vascular derangement in diabetic microangiopathy. We first prepared AGE-bovine serum albumin (BSA) and anti-AGE antiserum using AGE-RNase A. Then AGE-BSA was administered to human skin microvascular endothelial cells in culture, and their growth was examined. The AGE-BSA, but not nonglycated BSA, was found to induce a statistically significant increase in the number of viable endothelial cells as well as their synthesis of DNA. The increase in DNA synthesis by AGE-BSA was abolished by anti-AGE antibodies. AGE-BSA also stimulated the tube formation of endothelial cells on Matrigel. We obtained the following evidence that it is vascular endothelial growth factor (VEGF) that mainly mediates the angiogenic activities of AGE. Glucose and other reducing sugars can react nonenzymatically with the amino groups of proteins to form reversible Schiff bases and, then, Amadori products. These early glycation products undergo further complex reactions such as rearrangement, dehydration, and condensation to become irreversibly crosslinked, heterogeneous fluorescent derivatives termed advanced glycation end products (AGE) 1 (1). The formation and accumulation of AGE in various tissues have been known to progress during normal aging and at an extremely accelerated rate in diabetes mellitus. This has been implicated in the development of diabetic micro-and macro-vascular complications (1), which may account for the disabilities and high mortality rate in patients with this disease (2).Microvessels are composed of only two types of cells, endothelial cells and pericytes, and have been known to show both functional and structural abnormalities during prolonged diabetic exposure, resulting in the deleterious effects on the organs that they supply (3-5). Using pericyte-endothelial cell co-culture systems, we have shown previously that pericytes can not only regulate the growth but also preserve the prostacyclin-producing ability and protect against lipid peroxide-induced injury of endothelial cells (6). This has provided a basis for understanding how diabetic retinopathy develops consequent to "pericyte loss," the earliest histopathological hallmark in diabetic retinopathy (5, 7).Recently, we have found that AGE exert a growth inhibitory effect and a cell type-specific immediate toxicity on pericytes through interactions with their receptor for AGE (RAGE), a cell surface receptor belonging to the immunoglobulin superfamily (8), and have proposed a novel mechanism for pericyte loss (9). The AGE-induced, RAGE-mediated decrease in pericyte number would then indirectly cause angiogenesis (6,9).In the present study, we investigated the effects of AGE on the growth and tube formation of human skin microvascular endothelial cells, the key steps of angiogenesis. We demonstrate that AGE exert angiogenic activities directly on microvasc...

When starfish eggs were treated with caffeine, they developed parthenogenetically as tetraploid embryos. The timing of cleavages and gastrulation was compared between these tetraploid embryos and control diploid embryos. The following results were obtained: 1) The number of synchronous cleavages was ten in controls but nine in tetraploid embryos; 2) tetraploid embryos began to gastrulate at almost the same time after first cleavage as did control embryos; 3) the average number of constituent cells of tetraploid gastrulae was about 2,300, whereas that of control gastrulae was about 4,000. These results indicate that 1) the initiation of asynchronous cleavages is closely associated with the nucleocytoplasmic ratio; and 2) the timing of gastrulation is not determined by the number of cell divisions. These conclusions are in good accord with the results of a previous study on timing of early morphogenetic events in half eggs whose cytoplasm was decreased to half that of normal eggs (Mita, '83).

Eggs of the starfish Asterina pectinifera were divided into two equal-sized parts with fine glass needles after the first polar body formation, and nucleate halves were fertilized. Half embryos thus formed continued to divide and developed into morphologically normal, but dwarf bipinnariae. In order to explore possible mechanisms which determine the time of onset of early morphogenetic events, the timing of initiation of asynchronous cleavages, blastulation, and gastrulation was compared between half and whole embryos. The following results were obtained: 1) half eggs divided synchronously until the ninth cleavage, but the asynchronous cleavages started one division earlier in half embryos compared with whole embryos; 2) blastulation started at the 256-cell stage in both half and whole embryos; 3) half embryos initiated gastrulation at almost the same time as did whole embryos; and 4) the average number of constituent cells of half gastrulae was about 2500, while that of normal gastrulae was about 4500. These results indicate that a) the start of the asynchronous cleavages is closely associated with the nucleocytoplasmic ratio, but not with the number of cell divisions; b) the timing of blastulation is not affected by the altered nucleo-cytoplasmic ratio; and c) the timing of gastrulation is not determined by the number of cell divisions.

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