Experimental Hematology - Articles in Press

The HDAC Inhibitor Givinostat Modulates the Hematopoietic Transcription Factors NFE2 and C-MYB in JAK2V617F Myeloproliferative Neoplasm Cells - Accepted Manuscript

2012-05-10

Abstract: We investigated the mechanism of action of the HDAC inhibitor Givinostat (GVS) in JAK2V617F myeloproliferative neoplasm (MPN) cells. GVS inhibited colony formation and proliferation and induced apoptosis at doses 2-3 fold lower in a panel of JAK2V617F MPN compared to JAK2 wild type myeloid leukemia cell lines. By global gene expression analysis, we observed that at 6 hour GVS modulated 293 common genes in the JAK2V617F cell lines HEL and UKE1, of which 19 are implicated in cell cycle regulation and 33 in hematopoiesis. In particular, the hematopoietic transcription factors NFE2 and C-MYB were downmodulated by the drug specifically in JAK2V617F cells at both the RNA and protein level. GVS also inhibited JAK2-STAT5-ERK1/2 phosphorylation, but modulation of NFE2 and C-MYB was JAK2-independent, as shown using the JAK2 inhibitor TG101209. Indeed GVS had a direct effect on the NFE2 promoters, as demonstrated by specific enrichment of associated histone H3 acetylated at Lysine 9. Modulation by GVS of NFE2 was also observed in freshly isolated CD34+ cells from MPN patients, and was accompanied by inhibition of their proliferation and differentiation towards the erythroid lineage. We conclude that GVS acts on MPN cells through dual JAK2-STAT5-ERK1/2 inhibition and downmodulation of NFE2 and C-MYB transcription.

How can we reduce hepatic veno-occlusive disease–related deaths after allogeneic stem cell transplantation? - Uncorrected Proof

Douglas B. Johnson, Bipin N. Savani — 2012-04-27

Hepatic veno-occlusive disease (VOD) is a common and potentially devastating complication of hematopoietic stem cell transplantation. Confirmative diagnosis of this disorder can prove difficult early post hematopoietic stem cell transplantation, as a broad differential diagnosis exists and no definitive diagnostic test is available. Incidence of VOD has decreased in recent years, with especially dramatic declines in severe and fatal VOD. This improvement is attributed to less toxic and reduced-intensity conditioning regimens, and more appropriate patient selection. When severe VOD does occur, current treatments have been largely ineffective. Prevention remains the primary tool in the clinician’s arsenal for managing VOD. Our institution pursues aggressive preventative measures for VOD, including appropriate conditioning regimen selection, avoiding hepatotoxic drugs, early prophylactic use of ursodiol, and aggressive fluid management. With appropriate management steps, we believe the incidence of VOD and related deaths can be further decreased.

Low expression of asparagine synthetase in lymphoid blasts precludes its role in sensitivity to L-asparaginase - Accepted Manuscript

Ivana Hermanova, Marketa K. Zaliova, Jan Trka, Julia Starkova — 2012-04-27

Abstract: Childhood acute lymphoblastic leukemia (ALL) is treated with combined chemotherapy, comprising L-asparaginase (L-asp). Recent studies question the traditional view that the level of asparagine synthetase (ASNS), an enzyme producing the intracellular asparagine, correlates with the response to L-asp treatment. However, the importance of ASNS in response to L-asp has neither been confirmed nor refuted so far. In this study, we wanted to elucidate the effect of ASNS expression level on the sensitivity of ALL cells to L-asp treatment. We used four ALL cell lines (NALM-6, RS4;11, REH and UOCB6) and 30 diagnostic bone marrow samples of ALL patients to study the relation between ASNS expression and sensitivity to L-asp using MTS proliferation assay. RNA interference was used to study the effect of a range of ASNS levels on the response to L-asp treatment. Using cell a line model with a gradually knocked-down ASNS gene, we defined a cut-off level below which ASNS gene expression does not correlate with sensitivity to L-asp. Importantly, ASNS gene expression in patients' ALL blasts is below this level. We confirmed that there was no correlation between ASNS gene expression and sensitivity to L-asp in ALL blasts. Moreover, we show that cells with low ASNS expression level do not respond to asparagine deprivation by upregulation of ASNS gene expression. In conclusions, the ASNS expression level does not predict the sensitivity to L-asp in leukemic blasts. Moreover, cell lines with high basal expression of ASNS cannot serve as a valid model for studies on the relation between the ASNS and L-asp cytotoxic effect.

Drug-interaction studies evaluating T-cell proliferation reveal distinct activity of dasatinib and imatinib in combination with cyclosporine A - Corrected Proof

Stephen J. Blake, Timothy P. Hughes, A. Bruce Lyons — 2012-04-20

Development of small molecule tyrosine kinase inhibitors for the treatment of chronic myeloid leukemia has been astonishingly successful; however, their off-target effects have generated both challenges and opportunities for extending their clinical application. Dasatinib and imatinib are two of the most commonly used tyrosine kinase inhibitors and both have been shown to impact T-cell function. Due to this activity, their use as potential immune suppressants has been proposed. In this report, we investigated drug interactions with cyclosporine A in suppressing T-cell proliferation. Dasatinib and imatinib were titrated against varying concentrations of cyclosporine in the cultures and T-cell proliferation assessed by 5-6-carboxyfluorescein diacetate, succinimidyl ester dye dilution. These proliferation data were then used to determine the combination index to evaluate additive, synergistic, or antagonistic interactions between the drugs. This analysis uncovered a number of different drug interactions affecting T-cell proliferation. Cyclosporine had an additive or synergistic effect on T-cell proliferation when combined with dasatinib and imatinib for 3 of the 4 methods of stimulating T-cell proliferation. However, when T cells were stimulated with anti-CD3 and anti-CD28 antibodies, this interaction was found to be strongly antagonistic at low dasatinib concentrations. In contrast, this strong antagonism was not observed when imatinib was used in combination with cyclosporine A. This study suggests drug interactions affecting T cells may need to be carefully taken into account when using tyrosine kinase inhibitors. Furthermore, the technique to evaluate drug interactions is novel, and applicable to study any interaction affecting proliferation.

Modeling human hematopoietic cell development from pluripotent stem cells - Corrected Proof

Melanie D. Kardel, Connie J. Eaves — 2012-04-16

Understanding the steps and cues that allow hematopoietic cells to be generated during development holds great clinical as well as biological interest. Analysis of these events in mice has provided many important insights into the processes involved, but features that might be unique to humans remain challenging to elucidate because they cannot be studied directly in vivo. Human embryonic stem or induced pluripotent stem cells offer attractive in vitro alternatives to analyze the process. Here we review recent efforts to develop defined and quantitative systems to address outstanding developmental questions against a background of what we know about the development of hematopoietic cells in the fetus and derived from mouse embryonic stem cells.

Re: “Ex vivo fucosylation improves human cord blood engraftment in NOD-SCID IL-2Rγnull mice” - Corrected Proof

Robert Sackstein — 2012-04-16

It is well-established that expression of E-selectin on marrow microvascular endothelial cells mediates homing of hematopoietic stem cells to marrow (reviewed in ). In this issue of Experimental Hematology, Robinson et al. present data corroborating the reports of others showing that ex vivo cell surface glycan engineering achieved by treating human cord blood (CB) CD34+ cells with the sugar donor GDP-fucose and the enzyme fucosyltransferase VI enforces expression of sialylated Lewis X (sLex), the prototypical carbohydrate binding determinant for E-selectin. Notably, in earlier studies using in vivo xenogeneic models, one group claimed that fucosyltransferase VI–mediated ex vivo fucosylation (“exofucosylation”) of human CB cells improved engraftment, while the other group observed improved rolling adhesive interactions of CB CD34+ cells on marrow microvasculature (by intravital microscopy), yet without coincident improved extravasation (marrow homing). However, these early studies erred in overlooking a key issue: the inherent cytotoxicity of manganese (Mn). This divalent cation is an activating cofactor for fucosyltransferase VI activity, and all earlier enzyme preparations necessitated Mn in reaction buffer(s) to achieve requisite fucosylation . Indeed, previous studies utilized input Mn at 10 mM concentration, a level that is six orders of magnitude greater than that in serum. Even at micromolar amounts, Mn is cytotoxic, induces intracellular signaling cascades leading to apoptosis , is mutagenic , is genotoxic/clastogenic (i.e., causes DNA strand breaks) , inhibits DNA repair , and profoundly activates integrins . As we reported previously and confirmed in the study by Robinson et al., cells die when exposed in vitro to 10 mM Mn, even for just a few minutes. Thus, both earlier reports of CB cell exofucosylation were investigating the biology of dead cells. In the earlier study of CB cell engraftment , the comparator arm to exofucosylation consisted of “sham-treated” CB cells, which had been incubated in reaction buffer (which contained 10 mM MnCl2). An additional comparator arm should have been to test engraftment of native CB cells (i.e., not buffer treated), which, had they been utilized, would have engrafted far better than the dead CB cells. E-selectin ligands expressed on dead cells may retain the capacity to engage E-selectin, but dead cells certainly cannot transmigrate endothelial barriers, as this requires precise coordination between cell surface molecules and cytoskeletal components, all of which requires metabolic energy. Thus, the evidence that rolling was preserved but marrow homing was not achieved among CB CD34+ cells exposed to 10 mM Mn is not unexpected .

In reply - Uncorrected Proof

2012-04-16

The authors would like to take this opportunity to thank Dr. Sackstein for his insightful comments regarding the use of manganese (Mn2+) in our cord blood experiments involving ex vivo treatment with fucosyltransferase (FT)-VI, published in this issue of Experimental Hematology .

Maturity-dependent fractionation of neutrophil progenitors: A new method to examine in vivo expression profiles of differentiation-regulating genes - Corrected Proof

2012-04-09

To investigate differentiation-dependent gene expression during granulopoiesis, we established a new method to isolate six sequential differentiation stages of neutrophil progenitors from bone marrow. Neutrophil progenitors were divided into three populations by density centrifugation, followed by depletion of other lineages, and further separated by fluorescence-activated cell sorting based on the expressions of CD34, CD11b, and CD16: CD34+ fraction from a low-density population (F1), CD11b−/CD16− (F2), CD11b+/CD16− (F3), and CD11b+/CD16low (F4) fractions with intermediate density, and CD11b+/CD16int (F5) and CD11b+/CD16high (F6) fractions from a high-density population. To examine whether this fractionation was applicable to the study of in vivo gene expression profiles during granulopoiesis, we analyzed messenger RNA levels of AML-1 and CCAAT/enhancer binding protein (EBP)-ε and two target genes of C/EBP-ε, granulocyte-macrophage colony-stimulating factor receptor common β subunit and lactoferrin, in the six fractions and peripheral blood–derived neutrophils (F7). Expression of AML-1 and C/EBP-ε peaked at F1 and F4, respectively, followed by a gradual decrease. Although granulocyte-macrophage colony-stimulating factor receptor common β subunit messenger RNA levels remained low from F1 through F6 and elevated at F7, lactoferrin messenger RNA showed a drastic increase at F3 and dropped at F5. The difference in the expression profiles of the two C/EBP-ε target genes suggests the involvement of regulators other than C/EBP-ε in the induction of the two genes. The new fractionation method is able to provide new information on maturation-dependent gene expression during granulopoiesis.

Bryostatin analogue-induced apoptosis in mantle cell lymphoma cell lines - Corrected Proof

2012-03-30

The anti-cancer effects of bryostatin-1, a potent diacylglycerol analogue, have traditionally been attributed to its action on protein kinase C. However, we previously documented apoptosis in a B non-Hodgkin lymphoma cell line involving diacylglycerol analogue stimulation of Ras guanyl-releasing protein, a Ras activator, and Bim, a proapoptotic Bcl-2 family protein. To further explore the role of Bim, we examined several Bim-deficient B non-Hodgkin lymphoma cells for their responses to pico, a synthetic bryostatin-1-like compound. The Bim− mantle cell lymphoma cell lines Jeko-1, Mino, Sp53, UPN1, and Z138 and the Bim+ cell line Rec-1, as well as the Burkitt lymphoma cells lines BL2 (Bim−) and Daudi (Bim+), were examined for their response to pico using assays for proliferation and apoptosis as well as biochemical methods for Ras guanyl-releasing proteins and Bcl-2 family members. With the exception of UPN1, mantle cell lymphoma cell lines underwent pico-induced apoptosis, as did BL2. In some cases, hallmarks of apoptosis were substantially diminished in the presence of mitogen-activated protein kinase kinase inhibitors. Pico treatment generally led to increased expression of proapoptotic Bik, although the absolute levels of Bik varied considerably between cell lines. A pico-resistant variant of Z138 exhibited decreased Bik induction compared to parental Z138 cells. Pico also generally decreased expression of anti-apoptotic Bcl-XL and Mcl1. Although, these changes in Bcl-2 family members seem unlikely to fully account for the differential behavior of the cell lines, our demonstration of a potent apoptotic process in most cell lines derived from mantle cell lymphoma encourages a re-examination of diacylglycerol analogues in the treatment of this subset of B non-Hodgkin lymphoma cases.

Lenalidomide plus donor-lymphocytes infusion after allogeneic stem-cell transplantation with reduced-intensity conditioning in patients with high-risk multiple myeloma - Corrected Proof

2012-03-26

Myeloma relapse is the main cause of death after allogeneic stem cell transplantation. The aim of our observational study was to evaluate the anti-myeloma effect of lenalidomide followed by donor-lymphocyte infusion (DLI) as post-transplantation adoptive immunotherapy. Twelve patients with refractory myeloma were analyzed. The median age at transplantation was 56 years (range, 46–64 years). All patients received reduced-intensity conditioning. Patients were included if progressive or residual disease was observed at day +100 and if no signs of graft-vs-host disease were evident. DLIs were administered after two cycles of lenalidomide. Median dose of lenalidomide was 15 mg (range, 10–25 mg). Patients received a median of six cycles (range, 1–10 cycles). Nine patients (60%) received an escalating dose of DLI. The 1 and 2-year probability of progression-free survival was 75% and 50%, and overall survival was 83% and 69%, respectively. Median overall survival was not reached and median progression-free survival was 23 months. Lenalidomide is well tolerated after allogeneic stem cell transplantation; the combination with DLI did not cause a higher risk of graft-vs-host disease; an immunological synergistic effect was probably present with this strategy. This combination should be evaluated further in a larger cohort of patients.

Peginesatide and erythropoietin stimulate similar erythropoietin receptor–mediated signal transduction and gene induction events - Corrected Proof

2012-03-09

Peginesatide is a synthetic, PEGylated, peptide-based erythropoiesis-stimulating agent that is designed and engineered to stimulate specifically the erythropoietin receptor dimer that governs erythropoiesis. Peginesatide has a unique structure that consists of a synthetic peptide dimer (with no sequence similarity to erythropoietin) conjugated to a 40-kDa PEG moiety. Peginesatide is being developed for the treatment of anemia associated with chronic kidney disease in dialysis patients. To compare signaling effects of peginesatide to recombinant human erythropoietin (rHuEPO), dose-dependent effects on protein phosphorylation and gene expression were evaluated using phosphoproteomics, quantitative signal transduction analyses, and gene profiling. After stimulation with peginesatide or rHuEPO, cell lysates were prepared from UT-7/EPO cells. Liquid chromatography-tandem mass spectrometry and MesoScale arrays were used to quantify phosphorylation events. Transcriptional changes were analyzed using microarrays and quantitative reverse transcription polymerase chain reaction. Peginesatide and rHuEPO were found to regulate the tyrosine phosphorylation of an essentially equivalent set of protein substrates, and modulate the expression of a similar set of target genes. Consistent with their roles in stimulating erythropoiesis, peginesatide and rHuEPO regulate similar cellular pathways.

Deletion of the Scl +19 enhancer increases the blood stem cell compartment without affecting the formation of mature blood lineages - Corrected Proof

2012-03-07

The stem cell leukemia (Scl)/Tal1 gene is essential for normal blood and endothelial development, and is expressed in hematopoietic stem cells (HSCs), progenitors, erythroid, megakaryocytic, and mast cells. The Scl +19 enhancer is active in HSCs and progenitor cells, megakaryocytes, and mast cells, but not mature erythroid cells. Here we demonstrate that in vivo deletion of the Scl +19 enhancer (SclΔ19/Δ19) results in viable mice with normal Scl expression in mature hematopoietic lineages. By contrast, Scl expression is reduced in the stem/progenitor compartment and flow cytometry analysis revealed that the HSC and megakaryocyte-erythroid progenitor populations are enlarged in SclΔ19/Δ19 mice. The increase in HSC numbers contributed to enhanced expansion in bone marrow transplantation assays, but did not affect multilineage repopulation or stress responses. These results affirm that the Scl +19 enhancer plays a key role in the development of hematopoietic stem/progenitor cells, but is not necessary for mature hematopoietic lineages. Moreover, active histone marks across the Scl locus were significantly reduced in SclΔ19/Δ19 fetal liver cells without major changes in steady-state messenger RNA levels, suggesting post-transcriptional compensation for loss of a regulatory element, a result that might be widely relevant given the frequent observation of mild phenotypes after deletion of regulatory elements.

Alteration of lipids and the transcription of lipid-related genes in myelodysplastic syndromes via a TP53-related pathway - Corrected Proof

2012-02-29

Myelodysplastic syndromes (MDS) are clonal stem cell diseases of the bone marrow characterized by abnormalities in maturation of hematopoietic cells of all lineages. MDS patients frequently have lower lipids and high rates of apoptosis and p53 (TP53) expression. An association between the reduced lipids in MDS and the expression of lipid-related genes was sought. We further evaluated whether 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMGcoAR) and low-density lipoprotein receptor (LDL-R) are regulated by TP53 in vivo and in vitro. Gene expression was measured using real-time reverse transcription polymerase chain reaction on RNA extracted from bone marrow and peripheral blood from eight newly diagnosed MDS patients and eight controls and from mice livers. Serum lipid profile was measured using colorimetric enzymatic procedures. Total- and LDL cholesterol were lower in MDS patients in comparison to controls (p = 0.04 and p = 0.01, respectively). HMGcoAR messenger RNA increased in peripheral blood and bone marrow of MDS patients compared to controls (p = 0.04 and p = 0.01, respectively). LDL-R messenger RNA was higher only in the peripheral blood of MDS patients (p = 0.05). Comparable results were obtained in vivo. The transcription of these genes correlates with TP53 activation as documented by p21 messenger RNA elevation, a surrogate for TP53 activation and by using TP53 temperature-sensitive cells treated with adriamycin. To conclude, an association between reduced lipids in MDS and expression of HMGcoAR and LDL-R genes was documented. The transcription of these genes can be regulated by TP53.

Panobinostat (LBH589)-induced acetylation of tubulin impairs megakaryocyte maturation and platelet formation - Corrected Proof

2012-02-29

Drug-induced thrombocytopenia often results from dysregulation of normal megakaryocytopoiesis. In this study, we investigated the mechanisms responsible for thrombocytopenia associated with the use of Panobinostat (LBH589), a histone deacetylase inhibitor with promising anti-cancer activities. The effects of LBH589 were tested on the cellular and molecular aspects of megakaryocytopoiesis by utilizing an ex vivo system in which mature megakaryocytes (MK) and platelets were generated from human primary CD34+ cells. We demonstrated that LBH589 did not affect MK proliferation or lineage commitment but inhibited MK maturation and platelet formation. Although LBH589 treatment of primary MK resulted in hyperacetylation of histones, it did not interfere with the expression of genes that play important roles during megakaryocytopoiesis. Instead, we found that LBH589 induced post-translational modifications of tubulin, a nonhistone protein that is the major component of the microtubule cytoskeleton. We then demonstrated that LBH589 treatment induced hyperacetylation of tubulin and alteration of microtubule dynamics and organization required for proper MK maturation and platelet formation. This study provides new insights into the mechanisms underlying LBH589-induced thrombocytopenia and provides a rationale for using tubulin as a target for selective histone deacetylase inhibitor therapies to treat thrombocytosis in patients with myeloproliferative neoplasms.

A real-time (PCR) for a real life…? Quantitative evaluation of BCL2/IGH in follicular lymphoma and its implications for clinical practice - Corrected Proof

2012-02-29

Follicular lymphoma (FL) is highly associated with the molecular rearrangement BCL2/IGH. Although BCL2/IGH has been studied many times in follicular lymphoma, its real clinical value remains controversial. In this study, we performed quantitative testing by real-time polymerase chain reaction in 56 FL patients with median follow-up of 44 months (range, 9–102 months); chemotherapy was administered in 52 of 56 cases. Pretreatment numbers of BCL2/IGH varied in wide ranges, with a median of 2947 (range, 0–1,261,013) copies/106 cellular equivalent in peripheral blood (PB) and 4650 copies/106 cellular equivalent (range, 1–1,056,813) in bone marrow (BM), the difference between PB and BM was significant (p = 0.006). Pretreatment of BCL2/IGH quantities were correlated to clinical parameters (e.g., age, stage, sex, lactate dehydrogenase, B symptoms, grade, bulky disease, chemotherapy regimen) and to progression free-survival. Advanced clinical stage (III and IV) and microscopic BM involvement were significantly associated with higher numbers of BCL2/IGH in PB (p < 0.05) and in BM (p = 0.05), regardless all or newly diagnosed patients were evaluated. High pretreatment burden of BCL2/IGH was associated with significantly shorter progression-free survival; p = 0.003 and p = 0.047 for PB and BM, respectively. In conclusion, pretreatment quantity of BCL2/IGH in PB or BM seems to mirror the extent of disease and can provide an auxiliary prognostic parameter in FL. Our results also support evidence of the negative prognostic value of microscopic BM involvement in FL.

Induction of apoptosis in Eμ-myc lymphoma cells in vitro and in vivo through calpain inhibition - Corrected Proof

Hongbing Li, Rajeev M. Nepal, Alberto Martin, Stuart A. Berger — 2012-02-27

Calpains are cysteine proteases that have been implicated as both effectors and suppressors of apoptosis. Previously, we showed that c-myc transformation regulated calpain activity and sensitized cells to apoptosis induced by calpain inhibition. The objective of this study was to investigate the role of calpain in the Eμ-myc transgenic model of B-cell lymphoma. Calpain activity assays, apoptosis, cell cycle assays, and expression measurements were used to determine the activity and role of calpain in vitro and in vivo. We found that Eμ-myc transgenic cells have highly elevated calpain activity. Calpastatin, the negative calpain regulator, was expressed at much lower levels in Eμ-myc lymphoma cells compared to normal splenic B cells. The primary isoform in Eμ-myc lymphoma is calpain 1. Treatment of Eμ-myc lymphoma cells with the calpain inhibitors PD150606 or calpain inhibitor III induced caspase-3−dependent apoptosis in vitro. General caspase inhibitors or caspase-3/7 inhibitor protected cells from death induced by calpain inhibitor, whereas caspase-9 inhibitors failed to rescue cells. Human Burkitt’s lymphoma (BL2) cells display a pattern of sensitivity and caspase-3 dependence similar to calpain inhibition. Treatment of Eμ-myc lymphoma-bearing mice with PD150606 inhibited calpain activity in vivo and induced cell death in these cells as determined by terminal deoxynucleotide transferase-mediated dUTP nick-end labeling staining. Multiple daily treatments resulted in reduced tumor load, particularly in combination with etoposide. In conclusion, calpain is highly elevated in the Eμ-myc lymphoma and calpain inhibition has therapeutic potential.

Erratum to Experimental Hematology 2010;38:61-70 - Uncorrected Proof

2011-07-15

Title of the publication: Preclinical development of a bridging therapy for radiation casualties In the original publication an incorrect figure was printed as figure 4. Figure 3 was printed twice: once as figure 3 and again as figure 4. As a correction the above figure should be viewed as the proper figure 4. The published caption for figure 4 is correct.

Detection of bone marrow−derived lung epithelial cells - Uncorrected Proof

Susannah H. Kassmer, Diane S. Krause — 2010-05-05

Studies on the ability of bone marrow−derived cells to adopt the morphology and protein expression pattern of epithelial cells in vivo have expanded rapidly during the last decade, and hundreds of publications report that bone marrow−derived cells can become epithelial cells of multiple organs, including lung, liver, gastrointestinal tract, skin, pancreas, and others. In this review, we critically evaluate the literature related to engraftment of bone marrow–derived cells as epithelial cells in the lung. More than 40 articles focused on whether bone marrow cells can differentiate into lung epithelial cells have been published, nearly all of which claim to identify marrow-derived epithelial cells. A few investigations have concluded that no such cells are present and that the phenomenon of marrow-derived epithelial cells is based on detection artifacts. Here we discuss the problems that exist in published articles identifying marrow-derived epithelial cells, and propose standards for detection methods that provide the most definitive data. Identification of bone marrow−derived epithelial cells requires reliable and sensitive techniques for their detection, which must include cell identification based on the presence of an epithelial marker and the absence of blood cell markers as well as a marker for donor bone marrow origin. In order for these studies to be rigorous, they must also use approaches to rule out cell overlap by microscopy or single-cell isolation. Once these stringent criteria for identification of marrow-derived epithelial cells are used universally, then the field can move forward to address the critical questions about which bone marrow−derived cells are responsible for engraftment as epithelial cells, the mechanisms by which this occurs, whether these cells play a role in normal tissue repair, and whether specific cell subsets can be used for therapeutic benefit.

Stem cell plasticity: Recapping the decade, mapping the future - Uncorrected Proof

Neil D. Theise — 2010-05-03

In slightly more than a decade of stem cell plasticity research, 24 peer-reviewed articles have demonstrated plasticity across organ and/or embryonic lineage boundaries at the single-cell level, with only 1 article showing negative results. These data, taken together with data about reversibility of gene restrictions that have also accumulated during the same period, indicate that postnatal cells, even “terminally differentiated” ones, have a degree of plasticity not appreciated previously. This review looks back at the four known pathways of cell plasticity and at previously described “plasticity principles” of Genomic Completeness, Cellular Uncertainty, Stochasticity of Cell Origin and Fate, relating these to issues of experimental design and discourse that are key to understanding and evaluating plasticity data. Although the physiologic roles played by such plasticity may still be debated, the manipulations of these phenomena for therapeutic or industrial purposes should finally be considered ripe for exploration. For the future, plasticity, indeed all stem cell biology, must be considered as part of a larger web of cell-to-cell and cell-to-matrix interactions that function fully only at the tissue level; thus, the success of stem cell biology necessarily must involve assembling data from cell and molecular biology research into systems of interactions that might be reasonably called “tissue biology.” Interdisciplinary collaborations with complexity and chaos theorists, using mathematical/computer modeling of cell behaviors, will be vital to fully exploring stem cell behaviors in the coming decades.