A stochastic model of self-renewal and commitment to differentiation of the primitive hemopoietic stem cells in culture

We recently identified a murine hemopoietic stem cell colony which consists of undifferentiated (blast) cells and appears to be more primitive than CFU-GEMM in the stem cell hierarchy. The progenitors for the colony which we termed “stem cell colony” possess an extensive self-renewal capacity and the ability to generate many secondary multipotential hemopoietic colonies in culture. We replated a total of 68 stem cell colonies from cultures of murine spleen cells and analyzed the number of stem cell–and granulocyte(neutrophil)-erythrocyte-macrophage-megakaryocyte (GEMM) colony-forming cells in individual stem cell colonies. Of the 68 stem cell colonies, 35 contained progenitors (abbreviated as “S”-cells) for stem cell colonies. The distributions of S-cells and CFU-GEMM in individual stem cell colonies were extremely heterogeneous. Neither the frequency distributions of S-cells nor CFU-GEMM in stem cell colonies could be fitted well by Poisson distribution. Rather, the frequency distribution of the s-cells could be approximated by a geometric distribution and that of CFU-GEMM by an exponential distribution, both of which are variates of the gamma distribution. Our observations are in agreement with those on the distributions of CFU-S in individual spleen colonies and provided support for a stochastic model for stem cell self-renewal and commitment in culture. Application of the theory of the branching process to the distribution of S-cells revealed a distributional parameter “p” of 0.589 which is also in agreement with the earlier report on the p value for reproduction of CFU-S.     

Ex vivo expansion of primitive hematopoietic cells for cellular therapies: An overview

Sources of hematopoietic cells for bone marrow transplantation are limited by the supply of compatible donors, the possibility of viral infection, and autologous (patient) marrow that is depleted from prior chemo- or radiotherapy or has cancerous involvement. Anex vivo system to amplify hematopoietic progenitor cells could increase the number of patients eligible for autologous transplant, allow use of cord blood hematopoietic cells to repopulate an adult, reduce the amount of bone marrow and/or mobilized peripheral blood stem and progenitor cells required for transplantation, and reduce the time to white cell and platelet engraftment. The cloning of hematopoietic growth factors and the identification of appropriate conditions has enabled the development of successfulex vivo hematopoietic cell cultures. Purification systems based on the CD34 marker (which is expressed by the most primitive hematopoietic cells) have proven an essential tool for research and clinical applications. Present methods for hematopoietic cultures (HC) on stromal (i.e. accessory cells that support hematopoiesis) layers in flasks lack a well-controlled growth environment. Several bioreactor configurations have been investigated, and a first generation of reactors and cultures has reached the clinical trial stage. Our research suggests that perfusion conditions improve substantially the performance of hematopoietic reactors. We have designed and tested a perfusion bioreactor system which is suitable for the culture of non-adherent cells (without stromal cells) and readily scaleable for clinical therapies. Eliminating the stromal layer eliminates the need for a stromal cell donor, reduces culture time, and simplifies the culture system. In addition, we have compared the expansion characteristics of both mononuclear and CD34⁺ cells, since the latter are frequently assumed to give a superior performance for likely transplantation therapies.Abbreviations BFU0-E burst forming unit-erythroid - BM bone marrow - CB cord blood - CFU-C colony forming unit-culture - CFU-E colony forming unit-erythroid - CFU-F colony forming unit-fibroblast - CFU-GEMM colony forming unit-granulocyte, erythroid, macrophage, megakaryocyte - CFU-GM colony forming unit-granulocyte, macrophage - CFU-Mix colony forming unit-mixed (also known as CFU-GEMM) - CML chronic myeloid leukemia - CSF colony stimulating factor - DMSO dimethyl sulfoxide - ECM extracellular matrix - EPO erythropoietin - FL fetal liver - HC hematopoietic culture - LTBMC long-term bone marrow culture - LTC-IC long-term culture initiating cell - LTHC long-term hematopoietic culture - MNC mononuclear cells - PB peripheral blood     

Canonical and non-canonical Wnts differentially affect the development potential of primary isolate of human bone marrow mesenchymal stem cells

This study examines the role of Wnt signaling events in regulating the differential potential of mesenchymal stem cells (MSCs) from adult bone marrow (BM). Immunohistochemical analysis of BM revealed co-localization of Wnt5a protein, a non-canonical Wnt, with CD45(+) cells and CD45(-) STRO-1(+) cells, while Wnt3a expression, a canonical Wnt, was associated with the underlying stroma matrix, suggesting that Wnts may regulate MSCs in their niche in BM. To elucidate the role of Wnts in MSC development, adult human BM-derived mononuclear cells were maintained as suspension cultures to recapitulate the marrow cellular environment, in serum-free, with the addition of Wnt3a and Wnt5a protein. Results showed that Wnt3a increased cell numbers and expanded the pool of MSCs capable of colony forming unit -- fibroblast (CFU-F) and CFU -- osteoblast (O), while Wnt5a maintained cell numbers and CFU-F and CFU-O numbers. However, when cells were cultured directly onto tissue culture plastic, Wnt5a increased the number of CFU-O relative to control conditions. These findings suggest the potential dual role of Wnt5a in the maintenance of MSCs in BM and enhancing osteogenesis ex vivo. Our work provides evidence that Wnts can function as mesenchymal regulatory factors by providing instructive cues for the recruitment, maintenance, and differentiation of MSCs.     

The effect of recombinant erythropoietin on murine megakaryocyte colony formation

We investigated the effect of a recombinant human erythropoietin preparation (recombinant Epo) on murine megakaryocyte (MK) colony formation in serum-free and serum-containing culture systems, in order to study the relationship between Epo and megakaryopoiesis. Pokeweed mitogen spleen-conditioned medium (PWM-SCM), a standard source of MK colony stimulator, dose-dependently stimulated MK colony formation in the two culture systems. The plating efficiency of serum-free cultures was almost equal to that of cultures containing serum. Recombinant Epo also dose dependently stimulated MK colony formation in serum-containing cultures. However, in serum-free cultures recombinant Epo alone did not stimulate the growth of MK colonies; with the addition of fetal calf serum (FCS) to the serum-free cultures, recombinant Epo induced the growth of MK colonies. Furthermore, recombinant Epo enhanced MK colony formation through the stimulation of PWM-SCM or murine interleukin 3 (IL-3) in serum-free cultures. Our data show that Epo can act as a stimulator of megakaryopoiesis in collaboration with a factor in serum, or with an MK colony stimulator such as IL-3.     

Proliferation of human hematopoietic bone marrow cells in simulated microgravity

Expansion and/or maintenance of hematopoietic stem cell (HSC) potential following in vitro culture remains a major obstacle in stem cell biology and bone marrow (BM) transplantation. Several studies suggest that culture of mammalian cells in microgravity (micro-g) may reduce proliferation and differentiation of these cells. We investigated the application of these findings to the field of stem cell biology in the hopes of expanding HSC with minimal loss of hematopoietic function. To this end, BM CD34+ cells were cultured for 4-6 d in rotating wall vessels for simulation of micro-g, and assessed for expansion, cell cycle activation, apoptosis, and hematopoietic potential. While CD34+ cells cultured in normal gravity (1-g) proliferated up to threefold by day 4-6, cells cultured in micro-g did not increase in number. As a possible explanation for this, cells cultured in simulated micro-g were found to exit G0/G1 phase of cell cycle at a slower rate than 1-g controls. When assayed for primitive hematopoietic potential in secondary conventional 1-g long-term cultures, cells from initial micro-g cultures produced greater numbers of cells and progenitors, and for a longer period of time, than cultures initiated with 1-g control cells. Similar low levels of apoptosis and adhesion molecule phenotype in micro-g and 1-g-cultured cells suggested similar growth patterns in the two settings. These data begin to elucidate the effects of micro-g on proliferation of human hematopoietic cells and may be potentially beneficial to the fields of stem cell biology and somatic gene therapy.     

Fibronectin and laminin enhance engraftibility of cultured hematopoietic stem cells

To test the hypothesis that extracellular matrix (ECM) components maintain stem cell property, murine bone marrow (BM) cells were expanded in fibronectin and laminin coated plate in the presence of cytokines. We observed significant phenotypic and functional improvement of expanded cells. In 10 days, 800-fold expansion of colony-forming unit-granulocyte erythrocyte monocyte megakaryocyte (CFU-GEMM) was observed in the cultured cells. No apparent activation of cell cycle was observed, but CD29 and very late antigen-4 (VLA-4) expression was increased, as compared to the normal BM cells. A fraction of the expanded cells became verapamil sensitive, suggesting upregulation of multi-drug resistant gene(s), as found in the primitive hematopoietic stem cells (HSCs). Competitive repopulation assay confirmed that HSCs compartment was amplified during culture. Overall, our study clearly demonstrated that ex vivo culture of murine HSCs in the presence of fibronectin and laminin resulted in expansion of primitive stem cells and improvement in the marrow engraftibility.     

pH-responsive oligodeoxynucleotide (ODN)-poly(ethylene glycol) conjugate through acid-labile beta-thiopropionate linkage: preparation and polyion complex micelle formation

An oligodeoxynucleotide (ODN) conjugated to poly(ethylene glycol) (PEG) through a pH-responsive ester linkage (PEG-ODN conjugate) was successfully synthesized by the Michael reaction of 3'-thiol-modified ODN with a heterobifunctional PEG bearing an acetal group at the alpha-end and an acrylate group at the omega-end (acetal-PEG-acrylate), aimed at the development of a novel ODN delivery system. The prepared PEG-ODN conjugate and linear-poy(ethyleneimine) (L-PEI) spontaneously associated to form a polyion complex (PIC) micelle whose diameter and polydispersity index micro(2)/Gamma(2)) were 102.5 nm and 0.096 as determined by DLS measurements, respectively. Both the PEG-ODN conjugate and PIC micelle showed cleavage of the ester linkage at the endosomal pH (=5.5), suggesting that the PIC micelle is anticipated to release the ODN in the intracellular compartment. Furthermore, the PEG-ODN conjugate in the PIC micelle was stable against deoxyribonuclase (DNase I) digestion and has no interaction with the serum component because of the steric stabilization of the highly dense PEG corona surrounding the PIC core. These characteristics of the PIC micelles entrapping the PEG-ODN conjugate are promising for their utility as a novel ODN delivery system.     

B lymphocyte regulation of human hematopoiesis

Epstein Barr virus (EBV)-transformed B lymphoblastoid cell lines (BLCL) were derived from seven different individuals. The ability of BLCL supernatants to stimulate hematopoietic colony formation in vitro was tested in a conventional stem cell assay system. Supernatants promoted the growth of single (GM, E, MK) as well as multi-lineage (GEMM) colonies in bone marrow cultures. Our results indicate that EBV-transformed B lymphocytes produce cytokines that affect in vitro stem cell proliferation and differentiation. These studies demonstrate the regulatory potential of activated B lymphocytes in human hematopoiesis.     

EFFECTS OF SHORT-TERM CULTURE ON POPULATIONS OF HEMOPOIETIC PROGENITOR CELLS FROM MOUSE MARROW

We have studied the response of progenitor cells in mouse marrow to short-term cultivation in cell culture. In the presence of a feeder layer of mouse renal tubules, the number of granulocyte progenitors able to form colonies in cell culture showed a net increase. No such increase in numbers of stem cells detected by the spleen colony assay was observed under the same conditions, although there was some stimulation of uptake of tritiated thymidine into these cells. A rapid decrease in the numbers of both types of progenitor cells occurred in the cultures in the absence of a feeder layer. However, when marrow cell populations that had been cultivated for 2 days in the absence of a feeder layer were transplanted into irradiated mice, the surviving stem cells detected by the in vivo assay showed a prolonged phase of rapid growth, resulting in a more pronounced 'overshoot' in the growth curve.     

Influence of hydrocortisone on the survival and cluster and colony forming characteristics of normal human granulocyte-macrophage progenitors

This study was performed to determine the colony and cluster forming ability of granulocyte-macrophage (CFU-GM) progenitors of normal human blood low density cells cultured in a liquid culture system in the presence and absence of physiological doses of hydrocortisone (Hc). The CFU-GM recovered from the liquid cultures were assayed in soft agar medium. The results of the assays indicated that time-related development of clusters and colonies over 1-16 days, proliferative responsiveness to a source of colony stimulating activity, number of cells developed per colony, and the cellular composition of clusters and colonies produced from CFU-GM recovered from 14-day-old liquid cultures with 1.0 microM Hc, were all similar to those that developed from the normal human blood low density cells. However, a higher fraction of the CFU-GM in day 14 liquid cultures with 1.0 microM Hc were in DNA synthesis phase compared with the CFU-GM from the peripheral blood. This study confirmed the results of previous studies showing lower numbers of recognizable neutrophilic granulocytes and improved survival/proliferation of CFU-GM at day 14 in liquid cultures with 1.0 microM Hc compared with cultures without Hc. The present results suggest that the normal human blood CFU-GM which persists and proliferates under the influence of Hc in a liquid culture system is similar in ontogeny to the blood CFU-GM, and that the recovery of CFU-GM from liquid cultures under the influence of Hc appears to be exerted through stimulation of proliferation and controlled differentiation.     

Effects of irradiation on the nonlymphoid thymus in vitro

Thymic explant cultures were used to study the radiosensitivity of nonlymphoid thymic components in dogs. Thymic fragments from fetal (50 days gestation), newborn, and juvenile (70 days old) dogs were irradiated in vitro at 0, 0.5, 1, 2, or 4 Gy prior to culture. Colonies were classified as epithelial, spindle, or mixed cell type, and colony numbers were counted and diameters measured. Radiation caused a significant dose-related decrease in the number of spindle cell colonies from all ages. There was a corresponding, but smaller, dose-related increase in the number of epithelial colonies. The diameter of spindle cell colonies also decreased with dose, and this was accompanied by a reduction in cell density. While epithelial colony diameters did not change consistently with dose, there was an overall reduction in cell density in these colonies. This was more severe in the fetal than in the juvenile cultures. These results indicate that the putative mesenchymal (spindle cell) components of the thymus are significantly more radiosensitive than the epithelium at all ages and that fetal epithelium is more sensitive than epithelium from postnatal animals. This suggests that radiation-induced thymic epithelial defects reported after prenatal irradiation could be due to a combination of direct epithelial injury and defective inductive interaction between epithelium and the more radiosensitive mesenchyme.     

Polyploid megakaryocytes can complete cytokinesis

Megakaryocytes (MK) undergo polyploidization through endomitosis, a mitotic process that ends prematurely due to aborted cytokinesis. To better understand this and other events associated with MK differentiation, we performed long-term and large-field live cell imaging of human MKs derived in cord blood (CB) and bone marrow (BM) CD34+ cell cultures. Polyploid level of imaged cells was evaluated using three complementary approaches; cell history, cell size and ploidy correlation and nuclei staining. This system and strategy enabled the direct observation of the development of a large number of MKs (n=4865) and to quantify their fates. The most significant finding of this study is that a considerable proportion of polyploid MKs could complete cytokinesis. This unexpected process gave rise to polyploid daughter cell(s) with normal fates and contributed significantly to the expansion of polyploid MKs. Further analyses revealed that the proliferation rate amongst polyploid MKs was inversely correlated to their ploidy level, and that this phenomenon was much more frequent in CB- than BM-derived MKs. Accordingly, endomitosis was identified as the dominant fate of polyploid BM-MKs, while this was less accentuated for polyploid CB-MKs. These findings explain partially why CB-derived MKs remain in lower ploidy class. In conclusion, this study demonstrates that the development of polyploid MK results from the failure and/or success of cytokinesis and brings a new paradigm to the field of megakaryopoiesis.     

Quantitative image analysis of connective tissue progenitors

OBJECTIVE: To develop an image analysis system to automatically identify colony-forming units (CFUs) in in vitro cell cultures of connective tissue progenitors. This system was designed to quantitatively assess colony morphology and number of colonies in 4-cm(2) culture wells. STUDY DESIGN: Large field-of-view high-resolution fluorescence images of 4',6-diamidino-2-phenylindole (DAPI)- and alkaline phosphatase (AP)-stained bone marrow cell cultures were obtained using an epi-fluorescence microscope and automated scanning stage. Cell nuclei were identified in the DAPI-stained images after removal of fluorescent debris from the image. An Euclidean distance map (EDM) of the segmented cell nuclei was used to cluster cell nuclei into colonies. The automated system was evaluated using 40 tissue culture wells of bone marrow aspirate samples. The results of the automated analysis were compared to the manual tracings of colonies by 3 reviewers. RESULTS: The automated method agreed with all 3 reviewers on average 87.5% of the time. Additionally, reviewers identified other colonies not outlined by the reviewers on average 2.7 times more than the automated method. CONCLUSION: The automated method is a less biased method for identifying CFUs than individual reviewers, it provides more quantitative information about colony morphology than can be obtained manually and it is less time consuming.     

The effect of culture conditions on colony morphology and proliferative capacity in human prostate cancer cell lines

Primary cultures and cell lines form three types of colonies, termed holoclones, meroclones and paraclones by Barrandon and Green (Proc Natl Acad Sci U S A 84:2302-2306, 1987). They suggested that the three types correspond to colonies derived from stem, transit-amplifying and terminally differentiated cells. We determined the effect of culture conditions (seeding density, serum concentration, type of medium and substrate) on the proportion of each colony type and the cell number of individual colonies, using three prostate cancer cell lines, DU145, LNCaP and PC-3. In less favourable culture conditions, stem cell (SC) colonies tended to be lost; but in more favourable conditions, only modest increases in the proportion of SC colonies were observed. Under some conditions, cell number, but not colony-forming ability, was altered, indicating that colony cell number is controlled, at least in part, by different factors to colony formation. Colony-forming ability of individual cell lines is remarkably stable and there is little evidence for clonal evolution in culture, which might be expected and would result in more aggressive, faster-growing cells. Better understanding of how colony-forming efficiency is controlled could lead to the identification of drug targets that control SC growth and modify the progression of cancer.     

Differentiation from precursors in normal bone marrow of spontaneously cytotoxic cells showing anti-self-MHC specificity

Colonies containing spontaneously cytotoxic effector cells with specificity for target cells carrying self-MHC can be grown from normal mouse bone marrow (BM). BM was first depleted of nylon wool-adherent cells and was then cultured at low cell number (1 to 300 cells/culture) in multiple replicate microcultures in liquid culture medium containing supernatant from EL4 thymoma cells stimulated with PMA. Frequency of colony growth followed one-hit limiting dilution kinetics. Colonies contained lymphoid, myeloid, or both kinds of cells. About 5% of colonies contained self-specific cytotoxic effector cells. Analysis using the X chromosome-linked isoenzyme PGK-1 confirmed that colonies containing autoreactivity could be clonal. A factor other than IL 2, IL 3, or PMA appears to be required for the growth of autoreactive colonies. Similar colonies, both with and without autoreactive effector cells, could also be grown from the BM of athymic nude mice with frequencies and cytotoxic activities directly comparable to those found for normal BM. C.B-17 scid mice lack both B and T cells, apparently due to a block in the development of lymphoid stem cells. Colonies could be grown with comparable frequency from their BM, but these colonies lacked both lymphoid cells and spontaneous cytotoxic activity. Evidence is presented against the self-reactive effector cells being NK cells, macrophages, or mature T cells. It is speculated that they represent an early stage of the T cell differentiation pathway.     

The effect of hemin in vitro and in vivo on human erythroid progenitor cells

Hemin stimulates erythropoiesis and hemoglobin synthesis in vitro. We cultured erythroid progenitor cells from normal individuals, patients with sickle cell anemia, and a patient with acute variegate porphyria who received intravenous hemin treatment, with 0-800 microM hemin added in vitro. Fifty to 200 microM hemin consistently stimulated colony growth from normal donors 2- to 8-fold, while concentrations of up to 400 microM were stimulatory in cultures from donors with sickle cell anemia. In vivo hemin decreased the number of blood BFU-e in the patient with porphyria, but did not abrogate the in vitro stimulatory effect of hemin. Hemin concentrations which increased colony numbers increased gamma globin synthesis in some studies and decreased it in others. Hemin thus has clearcut erythroid growth-potentiating activity, although a consistent effect on globin chain regulation is not apparent.     

Ex vivo expansion and clonality of CD34+ selected cells from bone marrow and cord blood in a serum-free media

Although umbilical cord blood is increasingly being used in allogeneic marrow transplantation, delayed platelet engraftment is often a concern for cord blood transplant recipients. We evaluated the potential of ex vivo expansion and clonality in CD34+ cells separated from a bone marrow source, and cord blood, in a serum-free Media. The CD34+ cells, selected from bone marrow (BM) and umbilical cord blood (CB), were expanded with hematopoietic growth factors. They were then cultured for burst-forming units of erythrocytes (BFU-E), colony-forming units of granulocytes and monocytes (CFU-GM) and colony-forming units of megakaryocytes (CFU-Mk) at days 0, 4, 7, and 14 under the combination of growth factors, with cell counts. The cytokines included the recombinant human megakaryocyte growth and development (100 ng/ml), interleukin-3 (10 ng/ml), stem cell factor (100 ng/ml), flt-3 ligand (50 ng/ml) and interleukin-11 (200 ng/ml). The CB-selected CD34+ cells showed significantly higher total cell expansion than those from the BM at day 7 (3.0 fold increase than BM), day 14 (2.4 fold), and day 17 (2.6 fold). The colony count of the BFU-E/CFU-E per CD34+ cell at day 0 was 0.14 +/- 0.023 in the CB, which was significantly higher than 0.071 +/- 0.015 in the BM. The CB-selected CD34+ cells produced more BFU-E colonies than the BM on culture days 4, 7, and 14. The BFU-E colonies from the CB cells increased markedly on culture days 4 and 7, with a 4-fold increase at day 14. The colony count of the CFU-Mk per CD34+ cell at day 0 was 0.047 +/- 0.011 in the CB-selected CD34+ cells cultures, which was higher than the 0.026 +/- 0.014 in the BM. The CB-selected CD34+ cells produced more CFU-Mk colonies than the BM on culture days 4, 7 and 14. In conclusion, the ex vivo expansion of the CB cells may be very promising in producing total cellular expansion, CFU-Mk and BFU-E compared with BM, especially at day 7. The ex vivo expansion of the CB may have rationale in making an ex vivo culture for 7 to 14 d.     

Pyrimido-pyrimidine modulation of EGF growth-promoting activity and p21ras expression in rat mammary adenocarcinoma cells

RA 233, a pyrimido-pyrimidine analogue developed originally as an antiplatelet agent, has reduced the incidence of tumor metastases in clinical trials. However, in animal tumor models antimetastatic therapy using RA 233 has been inconsistent. We therefore tested RA 233 for additional effects, such as its direct action on tumor cells. Using the rat 13726NF mammary adenocarcinoma tumor system, low, nontoxic concentrations of RA 233 had pleiotropic and differential effects on two 13762NF tumor cell clones. The growth of MTC cells (low spontaneous metastatic potential) was not affected by low concentrations of RA 233 (50 microM) or epidermal growth factor (EGF) (up to 10 ng/ml) for 3 days in 0.5-10% fetal bovine serum. In contrast, MTLn3 (high spontaneous metastatic potential) cell cultures maintained for 3 days in low (0.5-1%) serum in the presence of 1.25-10 ng/ml EGF doubled in cell numbers compared with control cultures, and addition of 50 microM RA 233 abrogated the growth-stimulatory effect of EGF. The inhibitory effect of RA 233 on MTLn3 cells was dose dependent and not due to cell toxicity as determined by cell viability, cell growth, and colony formation properties after drug removal. In addition, incubation of MTLn3 cells with 50 microM RA 233 resulted in an increase of p21ras protein expression, whereas there was no effect on the level of p21ras in identically treated MTC cells or when either clone was treated with 10 ng/ml EGF. The results suggest that among the heterogeneous effects of RA 233 on tumor cells, modulation of growth factor responses and regulatory molecules may be important.     

Forced expression of HoxB4 enhances hematopoietic differentiation by human embryonic stem cells

HoxB4 has been shown to enhance hematopoietic engraftment by hematopoietic stem cells (HSC) from differentiating mouse embryonic stem cell (mESC) cultures. Here we examined the effect of ectopic expression of HoxB4 in differentiated human embryonic stem cells (hESCs). Stable HoxB4-expressing hESCs were established by lentiviral transduction, and the forced expression of HoxB4 did not affect stem cell features. HoxB4-expressing hESC-derived CD34+ cells generated higher numbers of erythroid and blast-like colonies than controls. The number of CD34+ cells increased but CD45+ and KDR+ cell numbers were not significantly affected. When the hESC derived CD34+ cells were transplanted into NOD/SCID beta 2m-/- mice, the ectopic expression of HoxB4 did not alter their repopulating capacity. Our findings show that overexpression of HoxB4 in differentiating hESCs increases hematopoietic colony formation and hematopoietic cell formation in vitro, but does not affect in vivo repopulation in adult mice hosts.     

In vitro inhibitory effect of interferon on colony formation of myeloma stem cells

Summary The inhibitory effect of interferon on colony formation of myeloma stem cells in two layer plasma clot-soft agar cultures was studied. Human lymphoblast interferon inhibited in therapeutically attainable concentrations myeloma stem cell proliferation in 50% and human fibroblast interferon in 23% of the 14 myeloma patients in whom in vitro colony formation could be achieved. In interferon-sensitive patients the numbers of myeloma stem cell clusters and colonies were decreased to 34.4%–54.9% of control cultures. In addition, maturation of myeloma stem cells in differentiated plasma cells was reduced by interferon in most of these cases.