Neglected physical human-robot interaction may explain variable outcomes in gait neurorehabilitation research.

During gait neurorehabilitation, many factors influence the quality of gait patterns, particularly the chosen body-weight support (BWS) device. Consequently, robotic BWS devices play a key role in gait rehabilitation of people with neurological disorders. The device transparency, support force vector direction, and attachment to the harness vary widely across existing robotic BWS devices, but the influence of these factors on the production of gait remains unknown. Because this information is key to designing an optimal BWS, we systematically studied these determinants in this work. We report that with a highly transparent device and a conventional harness, healthy participants select a small backward force when asked for optimal BWS conditions. This unexpected finding challenges the view that during human-robot interactions, humans predominantly optimize energy efficiency. Instead, they might seek to increase their feeling of stability and safety. We also demonstrate that the location of the attachment points on the harness strongly affects gait patterns, yet harness attachment is hardly reported in literature. Our results establish principles for the design of BWS devices and personalization of BWS settings for gait neurorehabilitation.

Mutant N-RAS induces erythroid lineage dysplasia in human CD34+ cells.

RAS mutations arise at high frequency (20-40%) in both acute myeloid leukemia and myelodysplastic syndrome (which is considered to be a manifestation of preleukemic disease). In each case, mutations arise predominantly at the N-RAS locus. These observations suggest a fundamental role for this oncogene in leukemogenesis. However, despite its obvious significance, little is known of how this key oncogene may subvert the process of hematopoiesis in human cells. Using CD34+ progenitor cells, we have modeled the preleukemic state by infecting these cells with amphotropic retrovirus expressing mutant N-RAS together with the selectable marker gene lacZ. Expression of the lacZ gene product, beta-galactosidase, allows direct identification and study of N-RAS-expressing cells by incubating infected cultures with a fluorogenic substrate for beta-galactosidase, which gives rise to a fluorescent signal within the infected cells. By using multiparameter flow cytometry, we have studied the ability of CD34+ cells expressing mutant N-RAS to undergo erythroid differentiation induced by erythropoietin. By this means, we have found that erythroid progenitor cells expressing mutant N-RAS exhibit a proliferative defect resulting in an increased cell doubling time and a decrease in the proportion of cells in S + G2M phase of the cell cycle. This is linked to a slowing in the rate of differentiation as determined by comparative cell-surface marker analysis and ultimate failure of the differentiation program at the late-erythroblast stage of development. The dyserythropoiesis was also linked to an increased tendency of the RAS-expressing cells to undergo programmed cell death during their differentiation program. This erythroid lineage dysplasia recapitulates one of the most common features of myelodysplastic syndrome, and for the first time provides a causative link between mutational activation of N-RAS and the pathogenesis of preleukemia.

Medical ethics for children: applying the four principles to paediatrics.

I will argue that there are difficulties with the application of the four principles approach to incompetent children. The most important principle - respect for autonomy - is not directly applicable to incompetent children and the most appropriate modification of the principle for them is not clear. The principle of beneficence - that one should act in the child's interests - is complicated by difficulties in assessing what a child's interests are and to which standard of interests those choosing for children should be held. A further problem with the four principles approach is that parental authority does not follow clearly from the four principles.

Expression of the colony-stimulating factor 1 receptor in B lymphocytes.

The FMS proto-oncogene encodes for the colony-stimulating factor 1 receptor (CSF-1R), whose expression within the haematopoietic system has previously been thought to be restricted to cells of the mononuclear phagocyte lineage. We have studied the expression of the CSF-1R in peripheral blood mononuclear cells by indirect immunofluorescence and flow cytometry. FMS expression was detected on both monocytes and B lymphocytes from all samples analysed, including 14 haematologically normal individuals and 31 patients (23 in remission following cytotoxic therapy for lymphoma, six with B-cell chronic lymphocytic leukaemia and two with chronic myelomonocytic leukaemia). The level of FMS expression on B lymphocytes was lower than the level of expression detected on monocytes isolated from the same sample. FMS mRNA expression in B lymphocytes has been confirmed by a reverse transcription-polymerase chain reaction (RT-PCR)-based technique and Northern blot analysis. Thus, FMS may play a role in the normal function of B lymphocytes and, because of its potential oncogenic activity, may contribute to the pathogenesis of malignancies of this cell type.

In vitro growth of myeloid and erythroid progenitor cells from myelodysplastic patients in response to recombinant human granulocyte-macrophage colony-stimulating factor.

Marrow progenitor cells from 14 myelodysplastic (MDS) patients and 17 normal donors were assayed in semisolid cultures supplemented with increasing doses of recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) or medium conditioned by 5637 bladder carcinoma cells (5637CM). At doses of supplements shown to be optimal for colony formation in cultures of normal marrow, myeloid (day 14) colony numbers were subnormal in 10 of 14 MDS marrows cultured in 5637CM and in 8 of 14 cultures containing rhGM-CSF (2.5 ng/ml). However, a high dose of rhGM-CSF (20 ng/ml) raised myeloid colony numbers in cultures of many MDS marrows, so that 9 of 14 now yielded colonies within the normal range; increased levels of 5637CM failed to do this. Erythroid colony growth was poor in 13 of 14 MDS marrow cultures supplemented with erythropoietin in addition to 5637CM or rhGM-CSF. High concentrations of rhGM-CSF did not increase erythroid growth. These data suggest that myeloid progenitors from the MDS clone may have a decreased responsiveness to hemopoietins which can be overcome at high concentrations of growth factors.

NICE head injury guidelines: review of the legal mandate.

This paper, while reviewing the legal authority held by clinical guidelines, examines the NICE head injury guidelines with respect to the likely consequences of non-compliance. Conversely, the effect on medical practice of rigid adherence to guidelines is also explored. Debate about the appropriateness of NICE head injury guidelines has highlighted the extent to which existing practices will need to change if compliance is to be achieved. Although a degree of resistance remains, there is perhaps a sense of resignation that the management of patients with head injuries will follow nationally prescribed guidance, whether in its current form or following its review next June. There will undoubtedly be those who remain unconvinced of the validity of these guidelines. Despite this, a possible reason for compliance may arise from concerns about the consequences of non-conformity. With the aid of a fictional scenario, this article seeks to remind the reader of the legal authority held by guidelines, the likely consequences of non-compliance and the liability held by their authors should compliance result in an untoward outcome. Finally, consideration is given to the possible long term effects that the adoption of guidelines may have on the medical profession.

Analysis and separation of murine bone marrow stem cells by H33342 fluorescence-activated cell sorting.

Murine bone marrow cells were stained with the fluorescent bisbenzimide dye H33342, a supravital DNA stain, and sorted on the basis of differences in fluorescence intensity by a light-activated cell sorter. Sorted cells were submitted to assays to enumerate spleen colony forming cells (CFUs), HPP-CFC and GM-CFU-1 and -2. The recoveries of these cell types after the separation was 70 to 120%, except for GM-CFU-2 (10-40%). The frequency distribution of these cell types with respect to their fluorescence intensity suggested that the majority of CFUs and HPP-CFC are quiescent, whereas GM-CFU-2 are proliferating. Furthermore, HPP-CFC could be separated from GM-CFU-1 and -2 on the basis of fluorescence intensity differences, which indicates that these cell types are different. CFUs determined 10 days after irradiation and grafting of the recipient mice were distributed evenly over the two fluorescence intensity subpopulations which contained most of the HPP-CFC and the GM-CFU. The same was observed at day 8. However, CFUs determined at day 12 were only present in the subpopulation of low fluorescence intensity which also contained most of the HPP-CFC. This observation provides evidence for heterogeneity of the spleen colony forming cells.

Enrichment of progenitor cells from human marrow.

Suspensions enriched for myeloid and erythroid colony-forming cells were prepared from human marrow by simultaneous treatment of low-density cells with the monoclonal antibodies Campath-1 and 80H.3, followed by immune-rosetting; lymphocytes and monocytes were successfully removed by this method. The final suspension contained 22% of blasts and 70% primitive myeloid-monocytic cells. GM-CFCs, counted after 14 days of culture, were enriched 21-fold. Of all cells present in the final suspension, 10.8% were day 8 GM cluster-forming cells, 2.3% were day 8 GM colony-forming cells, 2.4% were day 7 CFU-E, 0.9% were day 14 GM cluster-forming cells, 1.1% were day 14 GM colony-forming cells, and 0.6% were day 14 BFU-E. After enrichment, BFU-E became markedly more dependent on the addition of 5637-conditioned medium as a source of growth factors, suggesting that lymphocytes and/or monocytes support erythroid progenitor growth in cultures of unfractionated marrow. By removing these cells, we obtained a sensitive assay for burst-promoting activities in conditioned media. This procedure can be used to study the roles of marrow lymphocytes and monocytes in hemopoiesis as well as providing a basis for the purification of normal and aberrant progenitors.

Enrichment of CD34 (My10)-positive myeloid and erythroid progenitors from human marrow and their growth in cultures supplemented with recombinant human granulocyte-macrophage colony-stimulating factor.

Myeloid and erythroid progenitor cells were enriched from human marrow by selecting CD34-positive (CD34 + ve) cells, labeled with the My10 (HPCA-1) antibody, using a fluorescence-activated cell sorter. Seventy-one percent of CD34 + ve cells were blasts and most of these were too primitive to be identified by standard morphological criteria. On average, 9.5% of CD34 + ve cells formed clones after 14 days of culture in semisolid medium supplemented with erythropoietin and medium conditioned by 5637 bladder carcinoma cells. Over 2.5% of CD34 + ve cells were day-14 myeloid colony-forming cells and 2.4% were erythroid colony (burst)-forming progenitors. The remaining progenitors formed myeloid and erythroid clusters. A subpopulation of day-14 myeloid colony-forming cells failed to respond to recombinant human granulocyte-macrophage colony-stimulating factor (rhuGM-CSF) after accessory cells were removed during enrichment, so it appears that this factor can induce myeloid growth indirectly as well as directly. Recombinant human GM-CSF also supported erythroid colony-formation in cultures of CD34 + ve cells, which suggests that this hemopoietin may act directly on erythroid progenitors.

Bio-flash chromatography--rapid, low-cost purification of peptides.

The use of wide pore 40/60 mu Sorbsil silicas bonded with a range of biocompatible phases now allows the biochemist to benefit from the advantages that flash chromatography has given to synthetic organic chemists. The technique of bio-flash chromatography allows rapid peptide and protein purification at low pressure (less than 15 psi) and at a fraction of the cost of high-pressure systems.

Enrichment of high proliferation potential colony forming cells from mouse marrow by selecting low-density cells expressing receptors for wheat germ agglutinin.

Suspensions enriched for high proliferation potential colony forming cells (HPP-CFC's) were prepared from mouse marrow by selecting low-density cells stained with fluoresceinated wheat germ agglutinin on a fluorescence activated cell sorter. HPP-CFC's were 12-fold more concentrated in the final suspensions and co-enriched with a population of primitive CFUs detected in the spleens of irradiated mice reconstituted 13 days earlier with marrow. This is consistent with previous observations suggesting that these populations are closely related. The degree of enrichment for other haemopoietic progenitors was in the order HPP-CFC greater than day 8 CFUs greater than BFUe greater than GM-CFC greater than CFUe. "Extra" colonies developed when human placental conditioned medium (HPCM) was added to cultures of enriched suspensions already containing pregnant mouse uterus extract (PMUE). HPP-CFC's probably formed most of these and we discuss why counting these extra colonies may be more reliable than the conventional "size" assay for HPP-CFC's.

Clonal growth of haemopoietic progenitor cells from myelodysplastic marrow in response to recombinant haemopoietins.

The growth factor requirements of granulocyte-macrophage (GM) and erythroid marrow progenitor cells from 12 myelodysplastic (MDS) patients have been analysed. GM progenitors from two of six patients who grew normal numbers of colonies in response to conditioned medium + erythropoietin (5637CM + Epo) showed defective responses to either GMCSF and/or IL-3. Of all the recombinant factors tested (IL-3, IL-1, GCSF, GMCSF, MCSF), GMCSF was the strongest stimulator of myeloid clonal growth, inducing normal numbers of GM colonies from marrow of six patients (two of whom were neutropenic). Erythroid colonies were low in 5637CM + Epo-supplemented cultures of marrow from all but one patient and remained poor in the presence of any of the haemopoietins. tested. Supraoptimal doses (for normal marrow) of these haemopoietins improved colony growth in only one patient (GM colonies in response to IL-3). Combinations of factors were also largely ineffective at raising myeloid or erythroid colony numbers. These data indicate that the defective response of MDS progenitor cells to growth factors is not amenable to experimental manipulation of recombinant factor levels or combinations. Clonal assays might suggest a role for GMCSF therapy in a subpopulation of neutropenic MDS patients but their potential now needs to be evaluated in association with clinical trials.

Physical and kinetic properties of haemopoietic progenitor cell populations from mouse marrow detected in five different assay systems.

Properties of haemopoietic progenitor cells detected in several different assays have been compared in order to position them within the haemopoietic developmental lineage. The spleen colony-forming cell (CFUs), the high proliferation potential colony-forming cell (HPP-CFC) and two granulocyte-macrophage colony-forming cells (GM-CFC-1 and GM-CFC-2) have been studied. Two experimental techniques were used: separation of cells on the basis of their buoyant density and comparison of the survival of haemopoietic cells after donor mice had been injected with the cytotoxic drug 5-fluorouracil (5-FU). On linear BSA gradients the modal buoyant densities of CFUs, HPP-CFC and GM-CFC-1 were the same, 1.070 g cm-3; the density of GM-CFC-2 was higher, 1.075 g cm-3. GM-CFC-2 colonies were much smaller and contained far fewer cells than HPP-CFC or GM-CFC-1 colonies, even after prolonged culture, and this suggests that dense haemopoietic progenitors have less proliferation potential. This was confirmed by comparison of the size of colony formed, under identical culture conditions, by progenitors of different densities. Mean colony diameter was inversely related to the density of the progenitor cell. With the exception of GM-CFC-1, low density progenitors were more resistant to the cytotoxic effects of 5-FU than high density precursor cells (GM-CFC-2). Consequently, the GM-CFC-1 could be distinguished from GM-CFC-2 on the basis of buoyant density and from the other low density populations on the basis of post-FU kinetics. The reasons why the GM-CFC-1 should be more sensitive to 5-FU than other low density progenitors are discussed and the relation of these low density precursors to one another in terms of their position within the haemopoietic developmental lineage is elucidated.

GCSF augments post-progenitor proliferation in serum-free cultures of myelodysplastic marrow while ATRA enhances maturation.

All-trans retinoic acid (ATRA) and granulocyte colony stimulating factor (GCSF) are potential inducers of myeloid progenitor cell growth and neutrophil differentiation in myelodysplasia (MDS). We have compared the effects of ATRA and GCSF on the colony growth of 10 MDS marrows, in semi-solid and liquid serum-free mononuclear cell (MNC) cultures, supplemented with a mixture of stem cell factor (SCF), interleukin 3 (IL-3) and granulocyte-monocyte colony stimulating factor (GmCSF) (SIGm mix), which is fully-supportive for myeloid and erythroid (with erythropoietin (EPO)) colony formation in normal marrow. Only 1/10 MDS patients produced normal granulocyte-macrophage colony-forming cell (GmCFC) numbers, under SIGm conditions and erythroid colonies (ECFC) were subnormal in all patients. ATRA (10(-7) M) increased GmCFC numbers (P=0.05) in semi-solid cultures of normal, but not MDS marrow MNC and decreased erythroid colonies in cultures of marrow from either source (P=0.008 and P=0.0001 for normal and MDS, respectively). ATRA enhanced neutrophilic maturation in liquid cultures of both normal and myelodysplastic CD34 + ve cells, as detected by conventional morphology and acquisition of CD15. In contrast to ATRA, GCSF increased Gm colony size but not numbers in semi-solid cultures of normal marrow MNC, which suggests the cytokine augments post-progenitor amplification. This would explain why GCSF increased cell yields in liquid cultures of normal and MDS MNC while GmCFC accumulation remained unchanged. GCSF, though, increased Gm colony numbers in semi-solid cultures of MDS marrow MNC (P=0.014) so that 4/10 patients now grew colonies within the normal range. This was again probably due to increasing clone size, so that some clusters, the numbers of which may be elevated in MDS, were now scored as colonies. Overall, these data indicate that ATRA can enhance the maturation of the progeny of MDS GmCFC whilst GCSF can augment their amplification.

c-myc protein kinetics during growth and differentiation of CD34 (MY10)-positive blast cells from normal human marrow.

We have used flow cytometry to quantitate nuclear c-myc protein, at each phase of the cell cycle, during in-vitro differentiation of CD34-positive stem cells isolated from normal human bone marrow by the monoclonal antibody, MY10. Mean c-myc protein levels in CD34-positive cells, consisting of greater than 70% blasts, are lower than a marrow fraction containing myeloid cells of intermediate maturation, but have an invariant proportional relationship, with regard to nuclear mass, over the cell cycle. The majority of these primitive cells are non-cycling, as revealed by DNA content. Under our assay conditions, nuclear c-myc protein distribution over the cell cycle did not change as these progenitors entered a proliferative phase in culture. In cultures containing factors supporting myeloid maturation, mean G0/G1 p62c-myc levels initially decline, then rise above starting values as promyelocytes and myelocytes differentiate from CD34-positive cells, and as proliferation begins. With further myeloid maturation, and while cell numbers are increasing, c-myc protein continues to increase. C-myc protein kinetics differ in cultures in which macrophages, rather than myeloid cells, predominate. These data indicate that a complex relationship exists between c-myc gene expression and proliferation, maturation and lineage in haemopoietic cells, and lend support to the notion that early down regulation may be causally associated with the differentiation process.

Overgrowth of a leukemic culture by a minor CD34+ population.

We have investigated the differentiation potential of blast cells in a case of acute myeloid leukemia which comprised a majority CD34- population and a minor (2%) CD34+ fraction. Blasts were cultured for 2 weeks in a combination of cytokines--c-Kit ligand, interleukin 3 and granulocyte macrophage colony-stimulating factor (SIGm mix)--together with all-trans retinoic acid or 1alpha ,25-dihydroxy vitamin D3. Maturation of blasts was assessed by morphology on Romanowsky-stained slides, changes in surface CD markers and clonogenic culture. After 7 days of culture of unseparated blasts in SIGm, most maturation was monocytic, but with retinoic acid 63% of blasts had matured into granulocytes. Vitamin D3 enhanced monocytic differentiation, with 60% of cells becoming monocytic. The percentage of CD14 and CD15 positive cells decreased over 7 days in SIGm (from 62% to 17% and from 76% to 39% for CD14 and CD15, respectively). CD14+ cell numbers were maintained, or recovered, in cultures supplemented with vitamin D3 (59% at day 7), and CD15+ cell numbers, too, remained unchanged in the presence of retinoic acid (67%) or vitamin D3 (66%). Aberrant markers CD7 and CD56 declined under any conditions. When separated, both the CD34- and CD34+ fractions showed similar changes in morphology and surface maturation markers, suggesting that these two populations may be closely related. However, only a few CD34+ cells expressed the aberrant markers present on the majority blast population. The CD34- population declined in culture while the CD34+ fraction rapidly expanded. This probably reflects the difference in progenitor content; high numbers of colony-forming cells were concentrated in the CD34+ subpopulation. We conclude that both CD34- and CD34+ populations can differentiate but only the CD34+ fraction proliferates. Primitive clonogenic CD34+ cells from this patient may generate occasional aberrant CD34+ blasts which could then differentiate into the accumulating aberrant CD34- blast population.

In vitro drug resistance in acute myeloid and chronic B-lymphocytic leukaemic blasts and in normal blood and marrow populations.

The sensitivities of AML and BCLL blasts to daunorubicin have been determined, using an in vitro (MTT) assay of resistance, and compared with the sensitivities of normal haemopoietic populations and cells of the multidrug-resistant, T-lymphoid line CEM VLB100; The role of the drug-efflux pump, P-glycoprotein, was determined by adding the 'modifier' cyclosporin and by measuring numbers of P-glycoprotein positive cells by immunofluorescence. ID50s for 17 cases of de novo AML varied from 5 to 300 ng/ml giving a median of 105 ng/ml which was similar to the median of 11 normal marrow mononuclear cell preparations (80 ng/ml) but considerably less than the median ID50 of eight blood lymphocyte samples (3500 ng/ml). ID50s for five relapsed and two refractory AML samples ranged from 27 to 240 ng/ml, well within the de novo range: we had obtained presentation samples for two of these and, in both cases, ID50s were lower at relapse. ID50s, however, were raised in seven marrow mononuclear cell populations taken soon after remission induction (ID50 for remission MNC and normal MNC = 200 and 80 ng/ml, respectively); this may reflect either a property of regenerating populations, or an activation of cellular resistance mechanisms following chemotherapy. ID50s for 17 cases of BCLL ranged from 7 to 200 ng/ml with a median of 48 ng/ml which was significantly lower than the ID50 of AML blasts or of blood lymphocytes. Cyclosporin induced less than two-fold reductions in ID50s of blood lymphocytes, marrow mononuclear cells and de novo AML and BCLL blasts despite giving log reversals in resistance in the CEM VLB100 line. This reflected numbers of P-glycoprotein positive cells in our samples, which were high in CEM VLB100 but low in fresh normal or leukaemic cell suspensions. For both de novo AML and BCLL groups, however, the change in ID50, on addition of cyclosporin, was significant. These data imply a minor role for P-glycoprotein in drug resistance of leukaemic blasts. Nevertheless, there was a positive correlation between daunorubicin ID50s in de novo AML and time to remission which confirms that in vitro chemosensitivity assays can provide a useful measure of in vivo resistance.

Responsiveness to stem cell factor (SCF) of peripheral blood colony-forming cells from patients with myelodysplastic syndromes (MDS).

We have previously reported that serum stem factor (SCF) levels are significantly lower in patients with myelodysplastic syndrome (MDS) than normal controls. We have now studied the effects of adding SCF to cultures of blood mononuclear cells from patients with MDS and normal subjects. Three of 17 patients with MDS showed marked increases in erythroid colony numbers with SCF + erythropoietin compared to interleukin-3 + erythropoietin. In two cases (1RA and 1ISA) the erythroid colony numbers became normal. The same RA patient also showed a marked increase in myeloid colony numbers, which were undetectable with granulocyte-macrophage colony-stimulating factor alone, but within the normal range when SCF was added. A fourth patient (ISA) showed a 4.7-fold increase in myeloid colonies with SCF, but no erythroid response. The normal subjects showed a trend towards increased numbers of myeloid colonies with SCF; erythroid colonies did not increase. A correlation was found between the MDS patients' haemoglobin levels and erythroid colony numbers with and without SCF, but there was no correlation between erythroid or myeloid colonies in the presence of SCF and their serum SCF level.

Topical antibiotics on tracheostoma prevents exogenous colonization and infection of lower airways in children.

INTRODUCTION: Patients requiring long-term ventilation are at high risk of lower airway infections, generally of endogenous development. Patients on long-term ventilation, in particular via a tracheostomy, may develop tracheobronchitis or pneumonia of exogenous pathogenesis, ie, caused by microorganisms not carried in the oropharynx. The frequency of exogenous colonization or infection has previously been reported to be as high as 33%. A prospective observational cohort study of 2 years was undertaken to evaluate the efficacy of topical antibiotics in the prevention of exogenous colonization or infection of the lower airways. The antibiotic combination of polymyxin E and tobramycin in a 2% paste was applied four times a day on the tracheostoma. MATERIALS AND METHODS: A total of 23 children (median age, 4.1 months; range, 0 to 215 months) were enrolled in the study from September 1, 1996, until August 30, 1998. Surveillance samples of the oropharynx were obtained before tracheostomy and thereafter twice weekly. Diagnostic samples of the lower airways were taken once weekly and on clinical indication. RESULTS: Fourteen children (61%) had a total of 16 episodes of tracheal colonization or infection with 20 potentially pathogenic microorganisms. Only one child had tracheobronchitis with Streptococcus pneumoniae and Haemophilus influenzae during the 2-year study. Of the 16 colonization episodes, 12 (75%) were of primary endogenous pathogenesis, ie, caused by microorganisms present in the oropharynx at the time of tracheostomy. Community microorganisms including S pneumoniae, H influenzae, Moraxella (Branhamella) catarrhalis, and Staphylococcus aureus were the predominating bacteria. Three patients acquired nosocomial bacteria Pseudomonas aeruginosa and Hafnia alvei in the oropharynx, subsequently followed by secondary colonization of the lower airways. There was one failure of the prophylaxis: one patient (4%) had exogenous colonization with Pseudomonas pickettii. CONCLUSION: Topical antibiotics applied to the tracheostoma were found to be effective in reducing the exogenous route of colonization of the lower respiratory tract, compared with clinical experience and the literature. This promising technique requires further evaluation in randomized trials.

Erythroid colony growth from peripheral blood and bone marrow in polycythaemia.

Erythroid colony growth in the presence and absence of erythropoietin was compared in 23 patients with primary proliferative polycythaemia (PPP), nine with idiopathic erythrocytosis, 10 with secondary polycythaemia, 15 with pseudopolycythaemia and in 76 normal subjects. Erythroid colonies growing without erythropoietin stimulation (endogenous erythroid colonies) from peripheral blood (BFU-E) were found in 20 of 22 patients with PPP and in two of seven with idiopathic erythrocytosis. None was found in secondary polycythaemia, pseudopolycythaemia, or in normal subjects. Small numbers of endogenous colony forming units-erythroid (CFU-E) (though not BFU-E) were cultured from the bone marrow of three of 24 normal subjects, suggesting that peripheral blood cultures provide a more specific indicator of clonal erythropoiesis. Peripheral blood endogenous erythroid colony growth is an effective and convenient means of distinguishing patients with clonal erythrocytosis and may be of particular value when iron deficiency obscures the diagnosis of PPP on conventional criteria.