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We describe here a method for measuring DNA replication and, thus, cell proliferation in slow turnover cells that is suitable for use in humans. The technique is based on the incorporation of (2)H(2)O into the deoxyribose (dR) moiety of purine deoxyribonucleotides in dividing cells. For initial validation, rodents were administered 4% (2)H(2)O in drinking water. The proliferation rate of mammary epithelial cells in mice was 2.9% per day and increased 5-fold during pregnancy. Administration of estradiol pellets (0-200 microg) to ovariectomized rats increased mammary epithelial cell proliferation, according to a dose-response relationship up to the 100 microg dose. Similarly, proliferation of colon epithelial cells was stimulated in a dose-response manner by dietary cholic acid in rats. Bromodeoxyuridine labeling correlated with the (2)H(2)O results. Proliferation of slow turnover cells was then measured. Vascular smooth muscle cells isolated from mouse aorta divided with a half-life in the range of 270-400 days and die-away values after (2)H(2)O wash-out confirmed these slow turnover rates. The proliferation rate of an adipocyte-enriched fraction from mouse adipose tissue depots was 1-1.5% new cells per day, whereas obese ad libitum-fed obob mice exhibited markedly higher fractional and absolute proliferation rates. In humans, stable long-term (2)H(2)O enrichments in body water were achieved by daily (2)H(2)O intake, without toxicities. Labeled dR from fully turned-over blood cells (monocytes or granulocytes) exhibited a consistent amplification factor relative to body (2)H(2)O enrichment ( approximately 3.5-fold). The fraction of newly divided naive-phenotype T cells after 9 weeks of labeling with (2)H(2)O was 0.056 (CD4(+)) and 0.043 (CD8(+)) (replacement rate &lt;0.1% per day). In summary, (2)H(2)O labeling of dR in DNA allows safe, convenient, reproducible, and inexpensive measurement of cell proliferation in humans and experimental animals and is well suited for slow turnover cells.

Measurement in vivo of proliferation rates of slow turnover cells by 2 H 2 O labeling of the deoxyribose moiety of DNA

Measurement in vivo of proliferation rates of slow turnover cells by ² H ₂ O labeling of the deoxyribose moiety of DNA