From flow cytometric BrdUrd data to cell population growth and doubling time.

We describe a direct way to use flow cytometric data for measuring the growth curve of a cell population. The starting point is analysis of the intrinsic informative content of the time course, after bromodeoxyuridine (BrdUrd) labeling, of the percentages of cells detected within four windows of biparametric BrdUrd-DNA histograms. We did not introduce a particular cell cycle model or use the hypothesis of exponential growth. We obtained a simple formal proof of the existence of four independent formulae connecting the flow cytometric data and the relative growth curve of the cell population. The formulae were then challenged in a number of simulated kinetic scenarios, moving away from their expected limits of validity. The results suggest additional uses of the formulae and a way of estimating cell-cycle-phase durations. Considering exponential growth in the presence of cell loss, the formulae were used to estimate the potential doubling time from a single flow cytometric measure vs. other procedures that additionally require an estimate of the duration of the phase S. The theoretical precision of the procedures may differ depending on how cell loss occurs.

Hematopoietic toxicity and cell cycle perturbations induced by new DNA minor groove-alkylating agents.

Some new alkylating agents which bind to the minor groove of DNA and have sequence-specific patterns of alkylation have shown anti-neoplastic activity in pre-clinical systems. Two of them, carzelesin and tallimustine, are now in phase II. Considering the severe dose-limiting bone marrow toxicity of both these drugs in clinical use, it was of interest to investigate the mechanism of their myelotoxicity in a detailed pre-clinical study and compare it with a conventional alkylating agent, such as melphalan. The origin and progression of the myelotoxicity of carzelesin, tallimustine and melphalan were investigated comparatively in mice, combining data on bone marrow and peripheral blood cellularity with data on the proliferative activity of bone marrow cells, obtained by in vivo administration of bromodeoxyuridine. Significant differences were found between the hematopoietic response to the 3 drugs, though all caused severe leukopenia. Carzelesin induced a short-term increase in myeloid proliferative activity, which prevented the high leukocytopenia on day 3 observed with the other drugs. However, when this effect was exhausted, a second nadir was seen in peripheral blood, with a new wave of cell proliferation of all lineages in the bone marrow. Reconstruction of the lymphoid lineage was slow for all 3 drugs but particularly difficult with high-dose tallimustine. In general, the hematopoietic system response to tallimustine was dampened, with no overshoots, suggesting either lasting effects or extensive cytotoxicity from the early to late precursors of all lineages.

Cleistothecia of Uncinula necator-An Additional Source of Inoculum in Italian Vineyards.

Density and viability of populations of cleistothecia of Uncinula necator from bark, leaves, and soil were determined in three vineyards in the Florence and Siena provinces of Tuscany for 3 years. A higher density of cleistothecia was found on fallen leaves than on bark. However, the percentage of viable cleistothecia was higher on bark. No viable cleistothecia were recovered from soil. U. necator overwintered as mycelium in dormant infected buds, which gave rise to flag shoots, only in Santa Cristina, where 20 and 92 flag shoots per hectare were detected before bloom in 1994 and 1995, respectively. Disease incidence and severity increased similarly at Corti, Fornace, and at Santa Cristina, although powdery mildew epidemics started from ascospores only in Corti and Fornace, whereas flag shoots were present at Santa Cristina. Cleistothecia were formed in autumn in both 1994 and 1995, and their dispersal started in late September to mid-October, with the maximum number of cleistothecia trapped in funnels during the second half of October. Cleistothecia appear to function as the sole source of primary inoculum for grape powdery mildew in some Italian vineyards and serve as additional sources of inoculum where the pathogen also overwinters in infected buds. In Australia but not in New York, the pathogen also overwinters as cleistothecia on fallen leaves.