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Margaret A Goodell

Publications and source records attributed to Margaret A Goodell.

At least 19 recordsLinked to original sources
PubMed2026

Loss of Function Dnmt3a Mutation Leads to Aberrant Neutrophil Migration.

Clonal hematopoiesis (CH), an age-related expansion of somatically mutated hematopoietic clones, is associated with increased risk of severe infections including coronavirus disease (COVID)-19, yet the underlying mechanisms remain unclear. Here, we investigated the impact of Dnmt3a deficiency in a murine model of influenza A virus (IAV) pneumonia. Dnmt3a-deficient mice exhibited increased pulmonary viral burden and reduced neutrophil accumulation in IAV-infected lungs despite comparable circulating neutrophil numbers. Functional analyses of neutrophils showed impaired chemotactic migration in vitro, whereas maturation, antimicrobial enzyme content, and metabolic capacity were unchanged. Transcriptomic profiling revealed downregulation of pathways involved in chemotaxis, cytokine signaling, and cellular activation, including reduced expression of Cxcr1. Supporting the translational relevance of these findings, proteomic analysis of plasma from individuals with germline DNMT3A mutations (Tatton-Brown-Rahman syndrome) revealed alterations in proteins associated with cell migration and cytoskeletal dynamics. Collectively, our findings demonstrate that Dnmt3a loss compromises innate immune defense by impairing neutrophil migration in a cell-intrinsic manner, leading to ineffective pathogen clearance. This work provides mechanistic insight into how CH-associated mutations contribute to age-associated susceptibility to infection and highlights altered leukocyte trafficking as a potential therapeutic target in aging populations with CH.

Animals↗
PubMed2025

TET2-mutant myeloid cells mitigate Alzheimer's disease progression via CNS infiltration and enhanced phagocytosis in mice.

Clonal hematopoiesis (CH) is associated with many age-related diseases, but its interaction with Alzheimer's disease (AD) remains unclear. Here, we show that TET2-mutant CH is associated with a 47% reduced risk of late-onset AD (LOAD) in the UK Biobank, whereas other drivers of CH do not confer protection. In a mouse model of AD, transplantation of Tet2-mutant bone marrow reduced cognitive decline and β-amyloid plaque formation, effects not observed with Dnmt3a-mutant marrow. Bone-marrow-derived microglia-like cells were detected at an increased rate in Tet2-mutant marrow recipients, and TET2-mutant human induced pluripotent stem cell (iPSC)-derived microglia were more phagocytic and hyperinflammatory than DNMT3A-mutant or wild-type microglia. Strikingly, single-cell RNA sequencing (scRNA-seq) revealed that macrophages and patrolling monocytes were increased in brains of mice transplanted with Tet2-mutant marrow in response to chemokine signaling. These studies reveal a TET2-specific protective effect of CH on AD pathogenesis mediated by peripheral myeloid cell infiltration.

Animals↗
PubMed2005

Roles of Sca-1 in hematopoietic stem/progenitor cell function.

OBJECTIVE: This study was focused on studying the role of Sca-1 (Ly-6 A/E) in hematopoietic stem/progenitor cell self-renewal, activation, and lineage fate. MATERIALS AND METHODS: Sca-1(-/-) bone marrow cells were transplanted into wild-type recipient mice and assessed for self-renewal activity and lineage choice. In addition, Sca-1(-/-) mice were injected with 5-FU and Lin(-) cells were analyzed. Sca-1 was also overexpressed in mouse and human stem/progenitor cells to assess the effect of Sca-1 overexpression on stem/progenitor differentiation and proliferation. RESULTS: Self-renewal of Sca-1(-/-) HSC appeared to be normal, but lineage skewing was observed in B cells, NK cells, and granulocytes/macrophages derived from Sca-1(-/-) HSC. There was also a decrease in c-kit expression on activated Sca-1(-/-) progenitor cells. Overexpression of mouse Sca-1 decreased the in vitro myeloid activity of both mouse and human progenitors. CONCLUSION: These data indicate that Sca-1 plays a role in hematopoietic progenitor/stem cell lineage fate and c-kit expression. In addition, mouse Sca-1 overexpression affects human as well as mouse stem/progenitor cell activity, suggesting the possibility of a functional human Sca-1 homologue.

Animals↗
PubMed2005

Isolation and characterization of side population cells.

The protocol for isolation of side population (SP) cells was originally established for murine bone marrow hematopoietic stem cells (HSCs), but it has also been adapted for other species and tissues. This purification strategy offers a simple and reproducible strategy to obtain a highly homogeneous population of HSCs. The method is based on the differential efflux of the fluorescent DNA-binding dye Hoechst 33342 from stem cells relative to nonstem cells. The protocols outlined in this chapter describe the isolation of murine SP cells from both bone marrow and skeletal muscle using the fluorescent DNA-binding dye Hoechst 33342. In these tissues, the SP cells that are isolated are HSCs.

Animals↗
PubMed2004

Molecular signatures of proliferation and quiescence in hematopoietic stem cells.

Stem cells resident in adult tissues are principally quiescent, yet harbor enormous capacity for proliferation to achieve self renewal and to replenish their tissue constituents. Although a single hematopoietic stem cell (HSC) can generate sufficient primitive progeny to repopulate many recipients, little is known about the molecular mechanisms that maintain their potency or regulate their self renewal. Here we have examined the gene expression changes that occur over a time course when HSCs are induced to proliferate and return to quiescence in vivo. These data were compared to data representing differences between naturally proliferating fetal HSCs and their quiescent adult counterparts. Bioinformatic strategies were used to group time-ordered gene expression profiles generated from microarrays into signatures of quiescent and dividing stem cells. A novel method for calculating statistically significant enrichments in Gene Ontology groupings for our gene lists revealed elemental subgroups within the signatures that underlie HSC behavior, and allowed us to build a molecular model of the HSC activation cycle. Initially, quiescent HSCs evince a state of readiness. The proliferative signal induces a preparative state, which is followed by active proliferation divisible into early and late phases. Re-induction of quiescence involves changes in migratory molecule expression, prior to reestablishment of homeostasis. We also identified two genes that increase in both gene and protein expression during activation, and potentially represent new markers for proliferating stem cells. These data will be of use in attempts to recapitulate the HSC self renewal process for therapeutic expansion of stem cells, and our model may correlate with acquisition of self renewal characteristics by cancer stem cells.

Analysis of Variance↗
PubMed2004

Myeloproliferative disease in mice with reduced presenilin gene dosage: effect of gamma-secretase blockage.

Mammalian presenilins (PS) consist of two highly homologous proteins, PS1 and PS2. Because of their indispensable activity in the gamma-secretase cleavage of amyloid precursor protein to generate Abeta peptides, inhibition of PS gamma-secretase activity is considered a potential therapy for Abeta blockage and Alzheimer's disease intervention. However, a variety of other substrates are also subject to PS-dependent processing, and it is thus imperative to understand the consequences of PS inactivation in vivo. Here we report a pivotal role of PS in hematopoiesis. Mice heterozygous for PS1 and homozygous for PS2 (PS1(+/)(-)PS2(-)(/)(-)) developed splenomegaly with severe granulocyte infiltration. This was preceded by an overrepresentation of granulocytic cells in the bone marrow and a greatly increased multipotent granulocyte-monocyte progenitor in the spleen. In contrast, hematopoietic stem cells and T- and B-lymphocytes were not affected. Importantly, treatment of wild-type splenocytes with a gamma-secretase inhibitor directly promoted the granulocyte-macrophage colony-forming unit (GM-CFU). These results establish a critical role of PS in myelopoiesis. Our finding that this activity can be directly modulated by its gamma-secretase activity has important safety implications concerning these inhibitors.

Amyloid Precursor Protein Secretases↗
PubMed2004

Circulating hematopoietic stem cells do not efficiently home to bone marrow during homeostasis.

OBJECTIVE: Hematopoietic stem cells (HSC), normally resident in bone marrow, can be detected in the murine and human circulation. It is thought that HSC move in and out of bone marrow daily and that returning HSC are generally equivalent to their bone marrow counterparts in phenotype and function. However, large numbers of mononuclear blood cells are required to rescue animals from lethal irradiation, indicating either that the prevalence of circulating HSC is low, or they are inherently deficient in their repopulating ability. Accordingly, recent data suggest that circulating HSC may be unable to stably engraft WBM under homeostatic conditions. The purpose of this study was to explore these dynamics in detail using parabiosis and bone marrow transplantation. MATERIALS AND METHODS: The WBM and skeletal muscle HSC stem cell compartments of parabiosed CD45 congenic mice were analyzed functionally (via bone marrow transplantation) and phenotypically (via flow cytometry) for circulating stem cells at specific time points postparabiosis and after surgical separation. RESULTS: Surprisingly, we find that stem cells trafficking out of bone marrow and into the circulation do not stably return to bone marrow, although long-lived lymphoid precursors do stably re-engraft. Circulating HSC do, however, take up residence in skeletal muscle, wherein they account for HSC activity. CONCLUSION: Circulating HSC are not in flux with the bone marrow HSC and can persist in peripheral tissues.

Animals↗
PubMed2004

An in vivo propagated human acute myeloid leukemia expressing ABCA3.

Analyzing the regenerative compartment in the blast cell population of patients with acute myeloid leukemia (AML) may yield important insights into the mechanisms of disease progression. Here we present findings with a human AML cell line (AML-SP1), initiated from leukemic precursor cells and consecutively propagated by serial xenotransplantation in vivo. AML-SP1 maintained the characteristics of a human AML, consistently exhibiting a small leukemic side population (SP) of blast cells with high Hoechst 33342 exclusion. In the AML-SP1 line, an increased expression of the ABC transporters MDR1, MRP, ABCG2 and ABCA3 was found in the SP cells. The detection of ABCA3 in leukemic progenitor cells merits further investigation with regard to intracellular drug transport in AML blast cells. In vivo propagation of leukemias, such as AML-SP1 is a model system of maintaining the populational heterogeneity of AML disease, especially the unique characteristics of leukemic SP cells.

ATP-Binding Cassette Transporters↗
PubMed2004

Hematopoietic myelomonocytic cells are the major source of hepatocyte fusion partners.

Several recent reports have demonstrated that transplantation of bone marrow cells can result in the generation of functional hepatocytes. Cellular fusion between bone marrow-derived cells and host hepatocytes has been shown to be the mechanism of this phenomenon. However, the exact identity of the bone marrow cells that mediate cellular fusion has remained undetermined. Here we demonstrate that the hematopoietic progeny of a single hematopoietic stem cell (HSC) is sufficient to produce functional hepatic repopulation. Furthermore, transplantation of lymphocyte-deficient bone marrow cells and in vivo fate mapping of the myeloid lineage revealed that HSC-derived hepatocytes are primarily derived from mature myelomonocytic cells. In addition, using a Cre/lox-based strategy, we directly demonstrate that myeloid cells spontaneously fuse with host hepatocytes. Our findings raise the possibility that differentiated myeloid cells may be useful for future therapeutic applications of in vivo cellular fusion.

Animals↗
PubMed2004

Skeletal muscle fiber-specific green autofluorescence: potential for stem cell engraftment artifacts.

Adult stem cell research has lately been plagued by controversy regarding the possibility that some adult stem cells can engraft into nonautochthonous tissues. While most reports have observed some level of engraftment, the prevalence has varied in some cases by two orders of magnitude, suggesting that major technical variations may underlie these differences, possibly outweighing the biological basis of the observations. Here we describe bright green autofluorescence in a specific subset of skeletal muscle fibers that strongly resembles emission from green fluorescent protein (GFP). Moreover, we show that oxidative muscle fibers exhibit this autofluorescence, likely due to flavin, associated with NADH dehydrogenase. Finally, we demonstrate that confocal microscopy, in conjunction with spectral scanning, can be used to distinguish between GFP and autofluorescence. We suggest this autofluorescence artifact may account for some of the discrepancies in this field, particularly those describing skeletal muscle engraftment.

Animals↗
PubMed2003

Primitive adult hematopoietic stem cells can function as osteoblast precursors.

Osteoblasts are continually recruited from stem cell pools to maintain bone. Although their immediate precursor is a plastic-adherent mesenchymal stem cell able to generate tissues other than bone, increasing evidence suggests the existence of a more primitive cell that can differentiate to both hematopoietic and mesenchymal cells. We show here that the "side population" (SP) of marrow stem cells, defined by their ability to rapidly expel a DNA-binding dye and to regenerate the hematopoietic compartment, can differentiate to osteoblasts through a mesenchymal intermediate. When transplanted into lethally irradiated mice, single gene-marked murine SP cells reconstituted depleted osteoprogenitor pools, such that a large proportion of the osteogenic cells in the epiphysis of long bone carried the donor SP cell marker. These findings suggest that the developmental capacity of SP cells is not restricted to the hematopoietic lineages but extends to osteogenic differentiation. This property not only elucidates a previously unrecognized step in osteoblast development, but also has intriguing implications for the use of SP cells in clinical orthopedics and stem cell-based disorders of bone.

Animals↗
PubMed2003

Single hematopoietic stem cells generate skeletal muscle through myeloid intermediates.

Recent studies have shown that cells from the bone marrow can give rise to differentiated skeletal muscle fibers. However, the mechanisms and identities of the cell types involved have remained unknown, and the validity of the observation has been questioned. Here, we use transplantation of single CD45+ hematopoietic stem cells (HSCs) to demonstrate that the entire circulating myogenic activity in bone marrow is derived from HSCs and their hematopoietic progeny. We also show that ongoing muscle regeneration and inflammatory cell infiltration are required for HSC-derived contribution, which does not occur through a myogenic stem cell intermediate. Using a lineage tracing strategy, we show that myofibers are derived from mature myeloid cells in response to injury. Our results indicate that circulating myeloid cells, in response to inflammatory cues, migrate to regenerating skeletal muscle and stochastically incorporate into mature myofibers.

Animals↗
PubMed2003

Phenotype and hematopoietic potential of side population cells throughout embryonic development.

Adult murine bone marrow hematopoietic stem cells (HSCs) can be purified by sorting Hoechst 33342-extruding side population (SP) cells. Herein we investigated whether SP cells reside within embryonic tissues and exhibit hematopoietic progenitor activity. We isolated yolk sac (YS) and embryonic tissues 7.5 to 11.5 days after coitus (dpc), resolved an SP in each, and demonstrated that these SP cells exhibit distinct phenotypic and functional characteristics throughout development. YS and embryonic SP isolated 8.0 dpc expressed vascular endothelial-cadherin (VE-cadherin) and vascular endothelial receptor 2 (Flk-1), markers not expressed by bone marrow SP but expressed by endothelial cells and progenitors. SP at this stage did not express CD45 or produce hematopoietic colonies in vitro. In contrast, SP isolated 9.5 to 11.5 dpc contained a significantly higher proportion of cells expressing cKit and CD45, markers highly expressed by bone marrow SP. Furthermore, YS SP isolated 9.5 to 11.5 dpc demonstrated 40- to 90-fold enrichment for hematopoietic progenitor activity over unfractionated tissue. Our data indicate that YS and embryonic SP cells detected prior to the onset of circulation express the highest levels of endothelial markers and do not generate blood cells in vitro; however, as development progresses, they acquire hematopoietic potential and phenotypic characteristics similar to those of bone marrow SP.

Animals↗
PubMed2003

Transient RNA interference in hematopoietic progenitors with functional consequences.

Short interfering (si) RNAs have now been shown to inhibit gene expression in several species, including mammals (Elbashir et al.: Nature 411:494-498, 2001; Fire et al.: Nature 391:806-811, 1998). RNA inhibition in primary cells such as stem cells would facilitate rapid gene discovery in a postgenome era. While retroviruses can deliver siRNA expression cassettes for stable expression (Barton and Medzhitov: Proc Natl Acad Sci USA 99:14943-14945, 2002; Paddison et al.: Proc Natl Acad Sci USA 99:1443-1448, 2002; Rubinson et al.: Nat Genet 33:401-406, 2003), an efficient method for direct transfer of siRNA to stem cells is still lacking. Here, we established electroporation to deliver siRNA to hematopoietic progenitors. On average, at least 80% of cells take up the RNA, and these display nearly 100% knockout of marker gene expression at both the RNA and protein level. Moreover, knockdown of the hematopoietic regulator, CD45, results in 3-fold more hematopoietic colonies in a progenitor assay. These results demonstrate that transient transfection of siRNA to primary cells can have substantial functional consequences. This technology may be applicable to a variety of primary cell types.

Animals↗
PubMed2003

Altered phenotype and reduced function of muscle-derived hematopoietic stem cells.

OBJECTIVE: Skeletal muscle-derived cells have the potential to repopulate the major peripheral blood lineages of lethally irradiated mice and thus behave like hematopoietic stem cells (HSC). We have recently shown that muscle cells with HSC activity (ms-HSC) express CD45 and Sca-1, suggesting a hematopoietic origin. Here we sought to clarify contradictions in the literature regarding the phenotype of ms-HSC and precisely define the hematopoietic origin of these cells. METHODS: Skeletal muscle-derived cells fractionated based on the expression of CD45 and c-kit and efflux of Hoechst 33342 and were examined for HSC activity in vivo. WBM HSC expressing beta-galactosidase were transplanted into lethally irradiated recipients, whose ms-HSC compartment was later analyzed for beta-galactosidase activity to determine if ms-HSC were derived from WBM HSC. RESULTS: Muscle-derived HSC fall exclusively in the c-kit(dim)CD45(pos) compartment of the muscle side population (msSP). Furthermore, the CD45(pos) msSP compartment of skeletal muscle is derived from WBM HSC. CD45(pos)c-kit(dim) msSP are about 22-fold less potent in HSC activity than WBM HSC cells in competitive repopulation assays and express low levels of c-kit relative to WBM HSC. CONCLUSIONS: In our transplantation experiments, WBM HSC gave rise to ms-HSC, suggesting that WBM HSC and ms-HSC likely represent the same stem cell population in distinct environments. However, these two related populations are both functionally distinct in their ability to repopulate the peripheral blood of irradiated mice and phenotypically distinct.

Animals↗
PubMed2003

Adenoviral transduction of mouse hematopoietic stem cells.

Hematopoietic stem cells (HSC) are quiescent, self-renewing cells that can give rise to all blood cell lineages. HSC are an attractive target for gene therapy, due to their differentiation capacity and the number of diseases that result from abnormal HSC function. While human HSC have been shown to be transduced with adenoviral vectors, the adenoviral transduction of mouse HSC has not been extensively studied. We show here that a population of mouse bone marrow highly enriched for HSC (called side population, or SP, cells) can be transduced with adenovirus type 5 (Ad5) at a low multiplicity of infection. Transduced SP cells showed normal in vitro myeloid differentiation potential compared to mock-transduced SP cells. Transduced SP cells retained substantial but reduced in vivo long-term repopulating activity and contributed to all blood cell lineages. Ad5 transduction of mouse SP cells was dependent on coxsackie and adenovirus receptor (CAR), as an anti-CAR blocking antibody greatly reduced transduction. Therefore, adenoviral transduction of mouse HSC can be achieved without ablation of the hematopoietic repopulating activity.

Adenoviridae↗
PubMed2003

Isolation and characterization of functional mammary gland stem cells.

Significant advances in the stem-cell biology of several tissues, including the mammary gland, have occurred over the past several years. Recent progress on stem-cell fate determination, molecular markers, signalling pathways and niche interactions in haematopoietic, neuronal and muscle tissue may provide parallel insight into the biology of mammary epithelial stem cells. Taking advantage of approaches similar to those employed to isolate and characterize haematopoietic and epidermal stem cells, we have identified a mammary epithelial cell population with several stem/progenitor cell qualities. In this article, we review some recent data on mammary epithelial stem/progenitor cells in genetically engineered mouse models. We also discuss several potential molecular markers, including stem-cell antigen-1 (Sca-1), which may be useful for both the isolation of functional mammary epithelial stem/progenitor cells and the analysis of tumour aetiology and phenotype in genetically engineered mouse models. In different transgenic mammary tumour models, Sca-1 expression levels, as well as several other putative markers of progenitors including keratin-6, possess dramatically altered expression profiles. These data suggest that the heterogeneity of mouse models of breast cancer may partially reflect the selection or expansion of different progenitors.

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