Phenotypic and functional analysis of bone marrow progenitor cell compartment in bone marrow failure

Abstract

Many laboratory findings have demonstrated that the haemopoietic stem cell compartment is defective in aplastic anaemia (AA). AA bone marrow (BM) and peripheral blood (PB) are profoundly deficient in colony-forming cells, and AA progenitors fail to proliferate in long-term assays even in the presence of an intact stroma. Our study was designed to characterize some quantitative and qualitative aspects of the progenitor cell defect in AA. Using flow cytometric analysis of BM from new AA patients and from those recovering after immunosuppressive therapy, we determined that the numbers of CD34+ and CD33+ cells were markedly decreased in AA. Although PB neutrophil counts did not correlate with BM CD34+ cell numbers in acute disease, there was an association between the overall severity of the disease and the degree of CD34+ cell reduction. A decrease in BM CD33+ cells was a common finding in MDS patients, but reduction in CD34+ cells was found only in some hypoplastic MDS cases. Sorting experiments demonstrated lower plating efficiency for purified CD34+ cells from AA BM in comparison to controls. Thus, diminished colony formation of total BM appeared to result from both quantitative and qualitative defects. Based on the association between increased cycling and c-kit receptor expression on CD34+ cells, we found that the mitotically active CD34+ cells bearing the c-kit antigen were reduced in AA. With clinical improvement, CD34+ and CD33+ cells increased in correlation with PB parameters, but they did not return to normal values. Sorted CD34+ cells from recovered patents showed improved plating efficiency.(ABSTRACT TRUNCATED AT 250 WORDS)

Quantitative analysis of bone marrow CD34 cells in aplastic anemia and hypoplastic myelodysplastic syndromes

“…5,30,31 As some cases of moderate AA 5,30,31 and lowgrade MDS (e.g., the 5qÀ syndrome 32,33 ) never progress to 34,35 whereas hypoplastic MDS patients generally have higher bone marrow CD34 þ cell counts. 9,10,36,37 However, significant overlap in the CD34 percentages between these two disorders has precluded using this assay diagnostically. 9,10,36,37 Technical issues and difficulty precisely diagnosing AA and hMDS may account for the differences between previous reports and our findings.…”

“…9,10,36,37 However, significant overlap in the CD34 percentages between these two disorders has precluded using this assay diagnostically. 9,10,36,37 Technical issues and difficulty precisely diagnosing AA and hMDS may account for the differences between previous reports and our findings. Low CD34 þ cell percentages seen in some hMDS patients in other studies may have been the result of contamination of bone marrow aspirates with peripheral blood, as there was no description of criteria for validating the quality of their marrow samples.…”

“…Low CD34 þ cell percentages seen in some hMDS patients in other studies may have been the result of contamination of bone marrow aspirates with peripheral blood, as there was no description of criteria for validating the quality of their marrow samples. 9,36,37 We only considered marrow aspirate samples adequate if normal B cell and nucleated red cell precursors could be identified. Although Orazi et al 10 found that bone marrow biopsies from some hMDS patients also displayed low percentages of CD34 þ cells by immunohistochemistry, many of their patients had normal cytogenetics and no clinical outcome was provided; thus, these patients may have been misclassified.…”

“…8 CD34 þ hematopoietic progenitors are central to the pathogenesis of both MDS and AA. 5 CD34 þ cells normally comprise an average of 1À2% (range 0.5À5%) of the bone marrow cellularity, but are significantly decreased in AA bone marrow [9][10][11] because they are the targets of autoimmune destruction. 5,12 In contrast, CD34 þ cells appear to be the cells from which MDS originate, 5,13,14 and thus may be increased as a result of neoplastic clonal expansion.…”

Hematopoietic progenitor cells in the blood and bone marrow in various hematologic disorders

“…The number of CD34 cells is much reduced in aplastic anemia [ 13,141. Recently, it has been shown that there is a profound defecit in long-term initiating cells which have the frequency, phenotype and kinetic properties of true stem cells [15, 161. lost to follow-up [lo].…”