Deborah Greer

Alteration of marrow cell gene expression, protein production, and engraftment into lung by lung-derived microvesicles: a novel mechanism for phenotype modulation.

Numerous animal studies have demonstrated that adult marrow‐derived cells can contribute to the cellular component of the lung. Lung injury is a major variable in this process; however, the mechanism remains unknown. We hypothesize that injured lung is capable of inducing epigenetic modifications of marrow cells, influencing them to assume phenotypic characteristics of lung cells. We report that under certain conditions, radiation‐injured lung induced expression of pulmonary epithelial cell‐specific genes and prosurfactant B protein in cocultured whole bone marrow cells separated by a cell‐impermeable membrane. Lung‐conditioned media had a similar effect on cocultured whole bone marrow cells and was found to contain pulmonary epithelial cell‐specific RNA‐filled microvesicles that entered whole bone marrow cells in culture. Also, whole bone marrow cells cocultured with lung had a greater propensity to produce type II pneumocytes after transplantation into irradiated mice. These findings demonstrate alterations of marrow cell phenotype by lung‐derived microvesicles and suggest a novel mechanism for marrow cell‐directed repair of injured tissue.

Haematopoietic stem cells participate in muscle regeneration

It has previously been shown that bone marrow cells contribute to skeletal muscle regeneration, but the nature of marrow cell(s) involved in this process is unknown. We used an immunocompetent and an immunocompromised model of bone marrow transplantation to characterize the type of marrow cells participating regenerating skeletal muscle fibres. Animals were transplanted with different populations of marrow cells from Green Fluorescent Protein (GFP) transgenic mice and the presence of GFP(+) muscle fibres were evaluated in the cardiotoxin‐injured tibialis anterior muscles. GFP(+) muscle fibres were found mostly in animals that received either CD45(−), lineage(−), c‐Kit(+), Sca‐1(+) or Flk‐2(+) populations of marrow cells, suggesting that haematopoietic stem cells (HSC) rather than mesenchymal cells or more differentiated haematopoietic cells are responsible for the formation of GFP(+) muscle fibres. Mac‐1 positive population of marrow cells was also associated with the emergence of GFP(+) skeletal muscle fibres. However, most of this activity was limited to either Mac‐1(+) Sca(+) or Mac‐1(+)c‐Kit(+) cells with long‐term haematopoietic repopulation capabilities, indicating a stem cell phenotype for these cells. Experiments in the immunocompromised transplant model showed that participation of HSC in the skeletal muscle fibre formation could occur without haematopoietic chimerism.

The Stem Cell Continuum

Abstract: Hematopoietic stem cells have been felt to exist in a hierarchical structure with a relatively fixed phenotype at each stage of differentiation. Recent studies on the phenotype of the marrow hematopoietic stem cell indicate that it is not a fixed entity, but rather that it fluctuates and shows marked heterogeneity. Past studies have shown that stem cell engraftment characteristics, adhesion protein, and gene expression varies with the phase of the cell cycle. More recently, we demonstrated that progenitor numbers and differentiation potential also vary reversibly during one cytokine‐induced cell cycle transit. We have also shown high levels of conversion of marrow cells to skeletal muscle and lung cells, indicating a different level of plasticity. Recently, we demonstrated that homing to lung and conversion to lung cells in a mouse transplant model also fluctuates reversibly with cell cycle transit. This could be considered plasticity squared. These data indicate that marrow stem cells are regulated on a continuum related to the cell cycle both as to hematopoietic and to nonhematopoietic differentiation.

The marrow stem cell: the continuum.

Summary:The marrow hematopoietic stem cell is currently being redefined as to all aspects of its phenotype and its total differentiation capacity. This redefinition now includes its plasticity as to production of nonhematopoietic and hematopoietic cell types, the determinants of its in vivo engraftment potential and its expression of stem cell functional characteristics.

Marrow stem cell potential within a continuum.

Abstract: On the basis of our studies of the fluctuation of the hematopoietic stem cell phenotype with cell cycle trnsit, we hypothesize that the ability of marrow stem cells to convert to nonhematopoietic cells will also vary at different points in the cell cycle. The new biology of stem cells has an impact on many fields including developmental biology and stem cell biology and the clinical potential is enormous.

Tolerance induction by costimulator blockade in 100 cGy treated hosts with varying degrees of genetic disparity

Long-term multilineage allochimerism can be obtained in H2-mismatched B6.SJL to BALB/c transplants with host irradiation of 100 cGy, donor spleen cell pre-exposure and costimulator blockade with anti-CD40 ligand (CD40L) antibody. We evaluated this allochimerism approach in murine marrow transplants with different degrees of major histocompatibility complexe (MHC) mismatching; these include: (1) H2-mismatched transplant H2Kk to H2Kb, (2) full haplo-identical transplant H2Kbd to H2Kbk, (3) a partial haplo-identical transplant H2Kd to H2Kbd and (4) an MHC class II mismatch. Levels of chimerism increased up to 12 weeks and then stayed relatively stable up to 1 year after transplant. At 18 weeks post-transplant, the H2-mismatched, haplo-identical, partial haplo-identical and class II-mismatch transplants evidenced 17.9±4.4, 40.7±0.9, 25.1±4.19 and 33.7±3.5% donor chimerism, respectively. Dropping the anti-CD40 antibody treatment and spleen cells or changing the schedule of antibody to one injection, in haplo-identical or full-mismatched transplants resulted in no donor-derived chimerism. On the other hand, these still resulted in minor chimerism in class II-mismatched transplants. Lineage analysis of peripheral blood at 6 and 12 months post-transplant demonstrated a significant shift toward increased chimeric lymphocytes and decreased chimeric granulocytes in the full H2 as compared with haplo-identical or class II transplants. Transplantation with anti-CD40L antibody eliminated both graft-versus-leukemia and graft-versus-host disease (GVHD) and delayed lymphocyte infusion did not rescue animals from fatal leukemia. In conclusion, under the conditions of our tolerization regimen, a haplo transplant gives higher engraftment levels than a full H2 mismatch, and despite lower engraftment levels, a class II-mismatched transplant can be successfully accomplished with only 100 cGy and no CD40L blockade.