CELL RENEWAL, SIZE, AND CLONING AS BIOMARKERS OF AGING

本项目的主要目标是确定细胞周转率和 细胞大小是小鼠衰老的可靠生物标志物。 正常老化 小鼠、热量剥夺（CD）小鼠和转基因小鼠 将测试异位表达生长激素（GH）的小鼠。 的 生物标志物将被分为对处死动物的致死测定， 动物和一组非致命性测试， 单个老鼠的寿命。 将待测试的小鼠（3-，12-， 21和30月龄动物）将从NIA接收 老化研究繁殖群（GH小鼠除外）。 预计生物标志物指数与年龄相关 任何与年龄有关的变化都应该被减缓 在长寿的CD小鼠中。 GH小鼠似乎正在衰老 比它们的对照更快，因此提供了一种 生物标志物系统的有趣测试。 将测定细胞周转率和体积 在体内和体外： 体内-将4个年龄组的对照和CD小鼠输注1 或通过皮下渗透微型泵给予BrdU 4周。 在 这段时间结束时，几个器官中的细胞更新 将确定已知的高和低有丝分裂率。 将通过过氧化物酶核染色进行测定- 标记BrdU抗血清和自动图像分析计数， 允许计数的快速处理。 佣工与 抑制性T细胞和外周血的B细胞将被 借助以下方法确定电池循环次数和历史 激光激活细胞分选仪和特异性抗血清。 体外--来自其中细胞 将分散液置于培养物中以测定克隆 来自真皮皮肤、主动脉 平滑肌和骨髓成纤维细胞。 细胞体积 解离的细胞也将在FACS分析仪上测定。 只有用Hoechst 33342染色为二倍体的活细胞将被 包括在体积分析中。

The main goal of this project is to determine if cell turnover and cell size are reliable biomarkers of aging in mice. Normal aging mice, calorically deprived (CD) mice, and transgenic mice ectopically expressing growth hormone (GH) will be tested. The biomarkers will be divided into lethal determinations on sacrificed animals and a set of nonlethal tests that will be used to predict the longevity of individual mice. The mice to be tested (3-, 12-, 21-, and 30-month-old animals) will be received from the NIA aging study breeding colony (with the exception of the GH mice). The biomarker indices would be expected to correlate to the age of the control mice, and any age-related changes should be slowed in the long-lived CD mice. The GH mice appear to be aging somewhat more rapidly than their controls and thus offer an interesting test of the biomarkers system. Determinations of cell turnover and volume will be carried out both in vivo and in vitro: In vivo--Control and CD mice of 4 age groups will be infused for 1 or 4 weeks with BrdU via subcutaneous osmotic minipumps. At the end of this time, the present of cell turnover in several organs of known high and low mitotic rates will be determined. Determination will be made by nuclear staining with a peroxidase- tagged BrdU antiserum and automated image analysis counting, allowing the rapid processing of counts. The helper and suppressor T cells and the B cells of peripheral blood will be determined for number and history of cell cycling with the aid of a laser-activated cell sorter and specific antisera. In vitro-- Cell populations from those tissues in which cells are dispersible will be placed in culture for determination of clone size distribution for each sample of cells from dermal skin, aortic smooth muscle, and bone marrow fibroblasts. The cell volume of the dissociated cell will also be determined on a FACS analyzer. Only viable cells which stain as diploid with Hoechst 33342 will be included in the volume analysis.