Morphologic, immunophenotypic and in vitro growth characteristics of blood and bone marrow associated with stem cell mobilisation in patients with lymphoma.

The proportion of CD34+ cells in the bone marrow (BM) is predictive of the size of progenitor cell mobilisation into the blood (PB). To investigate which other PB and BM parameters may be related to mobilisation, we analysed at steady state PB and BM of 23 patients with relapsed or resistant lymphoma before administering high-dose cyclophosphamide and G-CSF Cell morphology, number of CD34+ cells, and growth in clonogenic assay and in long-term cultures (LTC) were determined and then correlated with mobilisation extent (CD34+ and GM-CFC) and quality (growth of harvested cells in LTC). We found that the good mobilising patients (CD34 > 50 x 10(3)/ml, n=10) had several baseline BM characteristics (number of CD34+ MNC, GM-CFC, BFU-E, production of CFCs in LTC) similar to a group of 12 healthy controls, while patients with reduced mobilisation (CD34 < 50 x 10(3)/ml, n=13) had clearly reduced BM progenitors and LTC growth (p< 0.05). In a multivariate analysis including baseline clinical, blood and bone marrow characteristics, the most significant PB and BM factors independently associated with a higher number and/or quality of mobilised cells were a higher number of CD34+ and GM-CFC in the BM and a higher baseline haemoglobin, platelet, and CD34+ blood count. The capacity to release progenitor cells into the circulation is therefore not predicted by the distribution of morphologically distinguishable cells, marginally predicted by the BM content of highly undifferentiated cells (growth in long term culture), while it is proportional to the number of BM progenitors (CD34+, GM-CFC and BFU-E).

Distribution of mobilized progenitor cells in the buffy coat of the haemonetics MCS3p cell separator: a study to optimize the collection of progenitors by leukapheresis.

Hematopoietic progenitor and stem cells for transplantation can be mobilized into the circulation and collected by leukapheresis. In this procedure, the leukocytes are distributed in the buffy coat along a density gradient, and the composition of the final product depends on which layer was collected. For the Haemonetics MCS3p Cell Separator, the manufacturer recommended starting the progenitor cell collection at a light transmission of 30%-40% (compared with plasma) and continue it for 40-50 ml. To optimize the use of this machine, the buffy coat it produces was studied in 12 patients by collecting it in fractions of increasing specific weight. Each fraction was analyzed by morphology, immunocytometry, and cell culture. We found that the buffy coat uniformly contains 8 times more leukocytes than blood, but the proportion of each white cell type varies along a gradient. The lymphocyte-predominant lighter layers are richer in CD34+ cells when compared with the granulocyte-predominant denser layers (6-14 times versus 2-4 times more than blood). The majority of CD34+ cells are found at a light transmission of 10%-70% (hematocrit 6-9). We conclude that cells for transplantation should be collected in a lighter fraction of the buffy coat than originally suggested by the manufacturer.