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Carlo-Stella et al. 439<br />

Epi 1 20 mg/sqm (day O)<br />

G-CSF 5 n-g/kg/d (days 1-10)<br />

"Two cycles at 3-wk interval<br />

Informed Consent<br />

R<strong>and</strong>omize to<br />

Amifostine<br />

on 1 st or 2nd cyck<br />

Amifostine<br />

1 ,000 mg iv (day O)<br />

500 mg iv (days 1 -4)<br />

Figure 1. Outline of the study design.<br />

the control from the percentage of cells positively stained with the anti-CD34.<br />

Phenotypic analysis was performed with a FACSort flow cytometer (Becton<br />

Dickinson). Data were processed with a Macintosh Quadra 650 computer (Apple,<br />

Cupertino, CA) using CELLQuest Software.<br />

CFU-Mix, BFU-E, <strong>and</strong> CFU-GM assay<br />

The assay for multipotent colony-forming units (CFU-Mix), erythroid burst-<br />

forming units (BFU-E), <strong>and</strong> granulocyte-macrophage colony-forming units (CFU-<br />

GM) was carried out as described elsewhere. 40<br />

Briefly, 1 to 10X10 4<br />

nucleated cells<br />

were plated in 35-mm petri dishes in 1-mL aliquots of Iscove's modified<br />

Dulbecco's medium (IMDM) containing 30% fetal bovine serum (FBS) (Stem Cell<br />

Technologies, Vancouver, Canada); 10~ 4<br />

M 2-mercaptoethanol (Gibco, Gr<strong>and</strong><br />

Isl<strong>and</strong>, NY); <strong>and</strong> 1.1% (wt/vol) methylcellulose. Cultures were stimulated with IL-<br />

3 (10 ng/mL; S<strong>and</strong>oz, Basel, Switzerl<strong>and</strong>), G-CSF (10 ng/mL; Amgen, Thous<strong>and</strong><br />

Oaks, CA), GM-CSF (10 ng/mL; S<strong>and</strong>oz), <strong>and</strong> erythropoietin (3 U/mL; Amgen).<br />

Progenitor cell growth was evaluated after 14-18 days of incubation (37°C, 5%<br />

C0 2) in a humidified atmosphere. Four dishes were set up for each individual data<br />

point per experiment. CFU-Mix, BFU-E, <strong>and</strong> CFU-GM were scored from the same<br />

dish according to previously published criteria. 40<br />

LTC-IC assay<br />

The long-term culture-initiating cell (LTC-IC) assay was performed according<br />

to Sutherl<strong>and</strong> et al. 41<br />

Briefly, test cell suspension (5X10 6<br />

nucleated cells) was

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