In this work, cord blood cells from 30 healthy term newborns were analyzed for complete blood counts with an automated cytometer and, in part of the sample, for surface molecules in cord blood monocytes, lymphocytes and CD34+ cells by two-color flow cytometry. Hematological parameters were as follows: WBC = 12.85 (5.24–15.10) ×10⁹/l; platelets = 304.33 (156.00–469.00) × 10⁹/l; Hb = 14.45 (11.90–17.82) g/dl; RBC = 3.99 (3.14–5.12) × 10¹²/l; MCV 107.25 (99.60–115.00) fl; reticulocytes = 157.80 (101.00–124.00) × 10⁹/l or 3.99 (2.45–6.01)%; erythroblasts = 0.88 (0.15–2.58) per 100 WBC or 6.63 (2.86–16.80) × 10⁹/l. The mononuclear population, as evaluated by flow cytometry, was composed of 22.9 ± 7.2% monocytes and 77.05 ± 7.24% lymphocytes, among which 46.59 ± 15.62% were T lymphocytes (43.94 ± 16.94% CD3+/CD4+ and 13.45 ± 7.46% CD3+/CD8+). CD34+ cells were on average 0.54 ± 0.24% of the mononuclear fraction. CD11c, CD49e and HLA-DR were found mainly on monocytes, and CD31 and CD62L occurred in similar levels on monocytes and lymphocytes. CD117+ cells were less than 5% of these populations. Among CD34+ cells, CD31 and HLA-DR were the molecules with higher frequencies (79.7 ± 19.9 and 65.7 ± 23.0%, respectively), followed by CD62L (41.8 ± 31.9%) and CD117 (20.1 ± 15.8%). The presence of CD11c and CD49e on CD34+ cells was low (below 10%). The results stress the phenotypic heterogeneity of cord blood CD34+ cells, and the different behavior of the cells when manipulated in vitro in different degrees of isolation.

Several studies have shown defective progenitor-stromal interactions in chronic myeloid leukemia (CML), and adhesive defects induced by BCR/ABL have been described. However, controversial results have been reported, and the role of the stroma in abnormal development of the hematopoietic system is not clear. In this study, CML hematopoietic and irradiated stromal cells were co-cultured in different combinations for 10 or 21 days. Maintenance of viable cells was dependent both on the sources of hematopoietic progenitors and stromal adherent layers, with normal cells performing better than their leukemic counterparts. The frequency of CD34(+) CD38(-) cells in the non-adherent fraction was more related to the source of hematopoietic cells than of stroma, and hematopoietic cells from normal subjects showed better performance. The simultaneous analysis of different combinations of normal and leukemic precursor cells and stromal layers, as done in the present work, suggests that the outcome of the interaction depends on characteristics of both compartments. This hematopoietic system development is influenced by intrinsic qualities of both hematopoietic stem cells and the supportive stroma.

In addition to bone marrow and peripheral blood, stem cells also occur in human umbilical cord blood (HUCB), and there is an increasing interest in the use of this material as an alternative source for bone marrow transplantation and gene therapy. In vitro hematopoiesis has been maintained for up to 16 weeks in HUCB cultures, but the establishment of an adherent, stromal layer has consistently failed. Adherent cell precursors among mononuclear cells from HUCB were sought for in long-term cultures. Mononuclear cells obtained from cord blood after full term, normal deliveries were cultivated at different concentrations in Iscove's modified Dulbecco's medium (IMDM) with weekly feeding. An adherent layer was detected in 16 of 30 cultures, 12 of which were plated at cell concentrations higher than 2 x 10(6) cells/ml. In contrast to bone marrow cultures, in which the stroma is detected early, in most (10/16) positive cultures from HUCB the adherent layer was identified only after the fourth week of culture. The cells never reached confluence and detached from the plate approximately four weeks after detection. May-Grünwald-Giemsa staining of positive cultures revealed fibroblast- or endothelial-like adherent cells in an arrangement different from that of bone marrow stroma in 13 samples. In two of these, the adherent cells were organized into characteristic, delimited cords of cells. Unlike bone marrow cultures, fat cells were never observed in the adherent layers. A rapid development of large myeloid cells in the first week of culture was characteristic of negative cultures and these cells were maintained for up to 12 weeks. HUCB contains adherent cell precursors which occur in lower numbers than in bone marrow and may be at a different (possibly less mature) stage of differentiation.
As células-mãe são encontradas no sangue do cordão umbilical humano (HUCB), além de na medula óssea e no sangue periférico, e há um crescente interesse no uso desse material como uma fonte alternativa para transplante de medula óssea e terapia gênica. A hematopoiese in vitro tem sido mantida por até 16 semanas em culturas de HUCB, mas o estabelecimento de uma camada estromal aderente tem invariavelmente falhado. Precursores de células aderentes foram pesquisados entre células mononucleares do HUCB em culturas a longo prazo. Células mononucleares obtidas do sangue do cordão depois de partos normais a termo foram cultivadas em diferentes concentrações em meio Dulbecco modificado por Iscove, com alimentação semanal. Uma camada aderente foi detectada em 16 de 30 culturas, 12 das quais em concentrações celulares maiores que 2 x 10(6) células/ml. Ao contrário das culturas de medula óssea, em que o estroma é detectado precocemente, na maioria (10/16) das culturas positivas do HUCB a camada aderente foi identificada apenas depois da quarta semana de cultura. As células nunca atingiram a confluência e se destacaram da placa aproximadamente quatro semanas após sua detecção. A coloração de culturas positivas por May-Grünwald-Giemsa revelou célul...