Development of rodent peripheral blood lymphocyte culture systems to detect cytogenetic damage in vivo.

Abstract

Peripheral blood lymphocytes (PBLs) offer many advantages for in vivo cytogenetic studies. They can be removed nonlethally from the animal allowing a subject to serve as its own control, permitting the analysis of cytogenetic damage over time. Furthermore, mature PBLs normally do not divide, and some populations are long-lived. Thus, they have the potential to accumulate DNA lesions during chronic exposures to genotoxicants. We have developed standard methodologies for the whole blood culture of rat and mouse PBLs to serve as models for determining the sensitivity of PBLs to cytogenetic damage. The cultures obtained with these protocols give reproducible results with high mitotic indices, stable baseline sister chromatid exchange (SCE) frequencies, and ample numbers of first-and second-division methaphases for scoring both chromosomal aberrations and SCEs. The methodologies have been especially useful for examining cytogenetic damage after inhalation exposures to toxicants such as ethylene oxide, formaldehyde, benzene, and nitrobenzene. Of these compounds, only benzene and ethylene oxide were found to induce significant dose-dependent increases in SCEs in PBLs. Also, dose-response curves have been obtained for several carcinogens administered by ip injection. These studies show that PBLs are sensitive indicators of the genotoxic effects of the carcinogens benzo (a)pyrene, 2-acetylaminofluorene, cyclophosphamide (CP), N-nitrosomorpholine, and ethylmethanesulfonate (EMS). In addition, because subpopulations of lymphocytes can be stimulated to divide using different mitogens, it has been possible to compare the sensitivity of murine B and T lymphocytes following in vitro and in vivo cyclophosphamide exposure. Once the sensitivity and selectivity of rodent lymphocyte cultures are determined, these assays should be valuable not only as a means for predicting which environmental agents could lead to increases in human cytogenetic damage, but also as a way to corroborate human cytogenetic studies.