Transformation of human osteoblast cells to the tumorigenic phenotype by depleted uranium-uranyl chloride.

Depleted uranium (DU) is a dense heavy metal used primarily in military applications. Although the health effects of occupational uranium exposure are well known, limited data exist regarding the long-term health effects of internalized DU in humans. We established an in vitro cellular model to study DU exposure. Microdosimetric assessment, determined using a Monte Carlo computer simulation based on measured intracellular and extracellular uranium levels, showed that few (0.0014%) cell nuclei were hit by alpha particles. We report the ability of DU-uranyl chloride to transform immortalized human osteoblastic cells (HOS) to the tumorigenic phenotype. DU-uranyl chloride-transformants are characterized by anchorage-independent growth, tumor formation in nude mice, expression of high levels of the k-ras oncogene, reduced production of the Rb tumor-suppressor protein, and elevated levels of sister chromatid exchanges per cell. DU-uranyl chloride treatment resulted in a 9.6 (+/- 2.8)-fold increase in transformation frequency compared to untreated cells. In comparison, nickel sulfate resulted in a 7.1 (+/- 2.1)-fold increase in transformation frequency. This is the first report showing that a DU compound caused human cell transformation to the neoplastic phenotype. Although additional studies are needed to determine if protracted DU exposure produces tumors in vivo, the implication from these in vitro results is that the risk of cancer induction from internalized DU exposure may be comparable to other biologically reactive and carcinogenic heavy-metal compounds (e.g., nickel).

Premature chromosome condensation assay for biodosimetry: studies with fission-neutrons.

Characterization of the premature chromosome condensation assay for radiation quality is needed. To that end, human lymphocytes were exposed in vitro to various doses of 250-kVp x rays (Y(D) = 4 keV microm(-1), Y(D) is the dose-mean lineal energy of the absorbed dose distribution, D(y), where y is defined as the energy deposited in a volume by a single event divided by the mean chord length of the volume) and to fission neutrons (Y(D) = 65 keV microm(-1)). The distribution of prematurely condensed chromosome and fragments following exposure to x rays or to neutrons were non-Poisson after repair at 37 degrees C for 24 h. Dose-response curves were constructed for the yield of excess prematurely condensed chromosome fragments as necessary for biodosimetry applications. The curves were fitted to a weighted linear model by the least-squares regression method. The neutron relative biological effectiveness (RBE) value was estimated to be 2.4 +/- 0.39.

Mixed-field neutrons and gamma photons induce different changes in ileal bacteria and correlated sepsis in mice.

High doses of radiation induce septicaemia, from bacterial translocation, and death in animals. Mice were exposed to either comparable lethal (LD90/30) or sublethal (LD0/30) doses of mixed-field [n/(n + y) = 0.67] or pure 60Co gamma-photon radiation. The relative biological effectiveness of these comparable doses of radiation was 1.82, determined by probit analysis. Mice given a lethal dose of mixed-field radiation developed a significant (p < 0.01), 10(9)-fold increase in Gram-negative facultative bacteria in their ilea over values in control mice. In contrast, mice given a lethal dose of gamma-photon radiation developed a significant (p < 0.01) increase in only Gram-positive bacteria in their ilea, while the number of Gram-negative bacteria remained near values in control mice. Data correlated with bacteria that were isolated and identified from the livers of mice that were given comparable lethal doses (LD99/30) of mixed-field or gamma-photon radiation. In sublethally irradiated mice, fluctuation in the total number of bacteria was detected in their ilea during the first week following irradiation, after which the number approximated the value in control mice. This difference in the predominant facultative bacteria in ilea resulting from different qualities of radiation has important implications for the treatment of septicaemic-irradiated hosts.

Energy deposition events produced by fission neutrons in aqueous solutions of plasmid DNA.

Using an agarose gel electrophoresis assay, single-strand breaks (ssb) induced by fission neutrons and 60Co gamma-rays in aerobic aqueous solutions of pBR322 plasmid DNA were studied. The energy-deposition events of the two radiations were characterized using a Rossi-type proportional counter to measure lineal-energy spectra. For neutrons, the dose-weighted lineal-energy mean, yD, is 63 keV micron-1--about 30 times that for gamma-rays. With increasing yD, hydroxyl radicals produced within spurs or tracks are less likely to survive due to recombination effects, resulting in decreased ssb yields. In TE buffer solution, the ssb yield induced by gamma-rays is 3.2 +/- 0.66 times that induced by neutrons at the same dose. Since the direct radiation effect is small under these conditions, we can estimate that the previously unknown G for hydroxyl radical production by fission neutrons is 0.088 mumol J-1. For glycerol concentrations that give the solution a hydroxyl radical scavenging capacity similar to that of cellular environments, the ssb yield induced by gamma-rays is about 2.0 +/- 0.24 times that induced by neutrons. Analysis shows that this trend with added scavenger is caused primarily by hydroxyl radical yields.

Late mitosis/early G1 phase and mid-G1 phase are not hypersensitive cell cycle phases for neoplastic transformation of HeLa x skin fibroblast human hybrid cells induced by fission-spectrum neutrons.

A two- to threefold increase in the rate of neoplastic transformation in cells irradiated at a dose rate of 0.22 cGy/min with fission-spectrum neutrons compared to that at 10.7 cGy/min has been confirmed with the use of alkaline phosphatase chromogenic substrate Western Blue staining to detect foci of neoplastically transformed cells through their expression of a tumor-associated antigen, the end point of the HeLa x skin fibroblast human hybrid cell transformation assay. To investigate whether the inverse dose-rate effect is due to the existence of a period in the cell cycle in which cells are significantly more sensitive to neoplastic transformation than in the rest of the cell cycle, as has been postulated previously (Rossi and Kellerer, Int. J. Radiat. Biol. 50, 353-361, 1986; Brenner and Hall, Int. J. Radiat. Biol. 58, 745-758, 1990; Elkind, Int. J. Radiat. Biol. 59, 1467-1475, 1991), we compared the sensitivity of late mitotic/early G1-phase and mid-G1-phase cells with that of asynchronous cells. The rationale for examining these particular cell cycle phases was based on the fact that mitosis has been hypothesized to be a candidate for the extremely sensitive period, and on a preliminary report that mid-G1-phase C3H 10T1/2 cells may exhibit enhanced sensitivity for neutron-induced transformation. A nominal dose of 45 cGy of fission-spectrum neutrons was delivered at approximately 10 cGy/min. The data indicate that neither late mitotic/early G1-phase nor mid-G1-phase cells are significantly more sensitive than asynchronous cells. Further, the dependence on the phase of the cell cycle for neoplastic transformation of CGL1 cells induced by fission-spectrum neutrons is different from that previously demonstrated for gamma radiation, where late-mitotic cells were approximately five times more sensitive than mid-G1-phase and asynchronous cells (Redpath and Sun, Radiat. Res. 121, 206-211, 1990).