Frequency, morphology, and molecular characterization of ECFCs and CFU-ECs. (A) ECFC yield in patients and controls. In the first row, the median frequency (and range) of ECFCs obtained from patients and controls is reported, whereas the second row represents the proportion of cases in which at least 1 ECFC-derived colony was obtained. Frequency was calculated as: total number of colonies grown multiplied 10⁷ and then divided by the total number of MNCs plated. (B) CFU-EC yield in patients and controls. In the first row, the median frequency (and range) of CFU-ECs obtained from patients and controls is reported, whereas the second row represents the proportion of cases in which at least 1 CFU-EC–derived colony was obtained. Frequency was calculated as: total number of colonies per 5 × 10⁶ MNCs plated. (C) ECFC and CFU-EC colonies. Representative photomicrographs of an ECFCs cultured from peripheral blood MNCs of a PMF patient (left, original magnification ×4) and of a CFU-EC cultured from peripheral blood MNCs of a CML patient (right, original magnification ×5) are shown. Similar colonies were grown from other CMPD patients and controls. (D) ECFC-derived and CFU-EC–derived cell capacity of tube formation in matrigel. ECFC-derived cells (left, original magnification ×2.5) and CFU-EC–derived cells (right, original magnification ×4) from peripheral blood MNCs of patients and controls show different capacity of tube formation in matrigel. (E) FISH from ECFCs and CFU-ECs of a CML patient. ECFCs (left) and CFU-ECs (right) derived cells from a CML patient show, respectively, the lack (2 green and 2 red spots) and the presence (2 fusion signals, arrows) of BCR-ABL fusion gene by FISH reaction. The normal cutoff value used for this probe is 1%, as established in our laboratory using normal controls. (F) JAK2-V617F mutation in ECFC-derived and CFU-EC–derived cells. A seminested PCR was performed, in which the first round amplified a 364-bp product from both mutant and wild-type JAK2 allele and the second round a 203-bp product specific for the JAK2-V617F allele. Amplification products were analyzed by electrophoresis on an ethidium bromide 2% agarose gel. First line represents internal control; second line, JAK2-V617F. (A) Positive control. (B-C) Two representative polycythemia vera patients. (D) Negative control. (G) Quantitative RT-PCR for BCR-ABL fusion gene and FISH results in ECFCs and CFU-ECs from 7 CML patients. For each case, at least 200 nuclei were scored for the presence of the t(9;22)(q34;q11). §p210 values are expressed as a percentage of the ratio of BCR-ABL/ABL. *Two red and 2 green spots, no fusion spot. **Two fusion spots, one red and one green.