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PubMed2004

Simultaneous injection of bone marrow cells and stromal cells into bone marrow accelerates hematopoiesis in vivo.

We have previously demonstrated that stromal cells can support the proliferation and differentiation of hematopoietic cells in vitro and in vivo and that a major histocompatibility complex restriction exists between hematopoietic stem cells and stromal cells. We have also found that intra-bone marrow (IBM) injection of allogeneic bone marrow cells (BMCs) leads to more rapid reconstitution of hematopoietic cells than intravenous injection. In the present study, we examine the effect of simultaneous injection of stromal cells and BMCs into the same bone marrow on the recovery of donor hematopoietic cells and demonstrate that simultaneous IBM injection of BMCs plus stromal cells is more effective in reconstituting recipients with donor hematopoietic cells than intravenous injection of BMCs plus stromal cells or IBM injection of BMCs alone.

Bone Marrow Cells↗
PubMed2005

Gene expression in stem cell-supporting stromal cell lines.

Cells in the immediate microenvironment together with hematopoietic stem cells (HSCs) constitute the stem cell niche. The microenvironmental or stromal cells provide a complex molecular milieu that helps mediate and balance the self-renewal and commitment potentials of stem cells. The molecules in this milieu are not well defined. In this study, we have intersected previous cDNA subtraction studies with array expression methodologies to define and categorize known gene products expressed by HSC-supportive stromal cell lines. Data were curated from our previously released Stromal Cell Database (StroCDB) containing a set of gene products enriched for expression in the fetal liver stromal cell line AFT024. Global expression analyses were extended to other stem cell-supporting and -nonsupporting fetal liver stromal cell lines using commercially available microarrays. Known and previously described gene products from selected categories were studied: transcription factors, cell membrane proteins, cytoskeleton and related proteins, extracellular matrix proteins, cell adhesion molecules and their corresponding signaling molecules, and cytokines and their related mediators. More than 300 known gene products were selected for expression in HSC-supporting stromal cells compared to nonsupporting lines. Analyses of the data suggest that HSC-supportive cells are immature, sessile, and highly reactive after binding to integrin ligands and cytokines. Therefore, they provide a dynamic space poised to respond to molecular cues elaborated within the stem cell niche. The study provides a survey of known proteins that play key roles in the support of HSCs by fetal liver stromal cells. It also provides insights into the biology of the stem cell niche by highlighting the complex network of intercellular signaling and communication involved in the organization of the niche space.

Animals↗
PubMed2001

Major histocompatibility complex restriction between hematopoietic stem cells and stromal cells in vitro.

We have previously found that a significant number of hematopoietic progenitors accumulate in engrafted bones with the same major histocompatibility complex (MHC) as the transplanted bone marrow cells. In the present study, to further clarify the MHC restriction between hematopoietic stem cells (HSC) and microenvironment, we carried out cobblestone colony formation assays by culturing HSCs with MHC-matched or -mismatched stromal cell monolayers. The formation of cobblestone colonies under MHC-mismatched stromal cells significantly decreased in comparison with MHC-matched stromal cells. However, the decrease in cobblestone colony formation under MHC-mismatched stromal cells was not significant when using MHC class I-deficient HSC or stromal cells. Taken together with the results using B10 congenic strains, it is suggested that the MHC preference is restricted by MHC class Ia molecules. Treatment with monoclonal antibodies (mAbs) against MHC class Ia molecules of stromal cell phenotypes significantly enhanced the cobblestone colony formation, whereas treatment with mAbs against HSC phenotypes significantly inhibited it. The expression of cytokines to promote hematopoiesis was enhanced by the mAbs against stromal cell phenotypes. The enhancement of cytokine expression was also observed when stromal cells and HSCs were MHC-matched. These results suggest that signaling via the MHC molecules augments stromal cell activity and elicits the MHC restriction.

Animals↗
PubMed1994

Native associations of early hematopoietic stem cells and stromal cells isolated in bone marrow cell aggregates.

In suspensions of murine bone marrow, many stromal cells are tightly entwined with hematopoietic cells. These cellular aggregations appear to exist normally within the marrow. Previous studies showed that lymphocytes and stem cells adhered to stromal cells via vascular cell adhesion molecule 1 (VCAM1). Injection of anti-VCAM1 antibody into mice disrupts the aggregates, showing the importance of VCAM1 in the adhesion between stromal cells and hematopoietic cells in vivo. Early hematopoietic stem cells were shown to be enriched in aggregates by using a limiting-dilution culture assay. Myeloid progenitors responsive to WEHI-3CM in combination with stem cell factor (c-kit ligand) and B220- B-cell progenitors responsive to insulin-like growth factor-1 in combination with interleukin-7 are not enriched. We propose a scheme of stromal cell-hematopoietic cell interactions based on the cell types selectively retained within the aggregates. The existence of these aggregates as native elements of bone marrow organization presents a novel means to study in vivo stem cell-stromal cell interaction.

Animals↗
PubMed2005

Histological characteristics of tumor cells and stromal cells in vessels and lymph nodes are important prognostic parameters of extrahepatic bile duct carcinoma: a prospective study.

Extrahepatic bile duct carcinomas (EHBDCs) consist of primary tumors, tumors in vessels, and tumors in lymph nodes. The purpose of this study was to prospectively investigate whether the histological characteristics of tumor cells and tumor stromal cells in vessels and lymph nodes were significantly associated with the outcomes of 60 EHBDC patients as compared with the histological characteristics of tumor cells and tumor stromal cells in primary tumors. Multivariate analyses, using the Cox proportional hazard regression model, showed that in EHBDCs without nodal metastasis, blood vessel tumor emboli with an angiomatous stroma significantly increased the hazard ratios (HRs) of tumor recurrence and death ( P < .05). In EHBDCs with nodal metastasis, the presence of tumor necrosis in the nodal tumors significantly increased the HRs of tumor recurrence and initial distant organ metastasis ( P < .05). In EHBDCs located in the distal to middle portion of the extrahepatic bile duct, blood vessel tumor emboli with an angiomatous stroma significantly increased the HRs of tumor recurrence, initial distant organ metastasis, and death ( P < .05). Severe nuclear atypia of the tumor cells in lymph vessels significantly increased the HRs of tumor recurrence and initial distant organ metastasis ( P < .05). In EHBDCs located in the hilar portion of the extrahepatic bile duct, the presence of nodal tumors with more than 4 mitotic figures significantly increased the HRs of tumor recurrence and initial distant organ metastasis ( P < .05). Several histological characteristics of tumor cells and tumor stromal cells in vessels and lymph nodes have significant effects on tumor progression of EHBDCs.

Adult↗
PubMed1996

Proliferation of cancer cells and stromal cells in colorectal adenocarcinomas as assessed with anti-PCNA/cyclin monoclonal antibody.

Monoclonal anti-proliferating cell nuclear antigen antibody (PCNA, PC 10) was used to measure proliferation of tumor cells and stromal cells in 27 colorectal adenocarcinomas. Mean proliferation index of cancer cells was 33.7% (SD +/- 16.3) and that of stromal cells 14.3 (SD +/- 12.5). Positive correlation between proliferation index of cancer cells and proliferation rate of stromal cells was found (r = 0.64, p < 0.001). Correlation between proliferation of stromal lymphocyte-like cells and fibroblast-like cells was noticed (r = 0.69, p < 0.001).

Adenocarcinoma↗
PubMed1994

Natural and recombinant bovine interferon tau regulate basal and oxytocin-induced secretion of prostaglandins F2 alpha and E2 by epithelial cells and stromal cells in the endometrium.

The effects of bovine interferon tau (IFN tau) and oxytocin on secretion of the prostaglandins PGF2 alpha and PGE2 by epithelial and stromal cells in the endometrium were assessed in two experiments. Endometrial tissues were collected from cyclic cows at Day 15 after oestrus for subsequent isolation of epithelial cells (4 cows) and stromal cells (4 cows). In both experiments, confluent cells were treated with 0, 2, 10 or 50 ng mL-1 natural bovine IFN tau (nbIFN tau) or 0, 0.4, 2, 10, 50 and 250 ng mL-1 recombinant bIFN tau (rbIFN tau). Culture medium was sampled at 24 h. Oxytocin (2.0 x 10(-7) M) or placebo was then added to wells and the medium was sampled 30 and 90 min later. Epithelial cells secreted more PGF2 alpha than stromal cells whereas stromal cells predominantly secreted PGE2. Oxytocin stimulated secretion of PGF2 alpha and PGE2 (P < 0.01) from epithelial cells, but both basal secretion and oxytocin-induced secretion of PGF2 alpha and PGE2 decreased with increasing dose of either nbIFN tau or rbIFN tau (P < 0.01). At comparable doses, rbIFN tau inhibited PGF2 alpha and PGE2 secretion more strongly than did nbIFN tau (either in the absence or the presence of oxytocin). The minimal effective dose of rbIFN tau was 0.4 ng mL-1 and 50% inhibition was obtained with 1 ng mL-1 (0.043 nM). Neither nbIFN tau nor rbIFN tau nor oxytocin altered PGF2 alpha or PGE2 secretion by stromal cells. The results indicate differential prostaglandin responses by the two major endometrial cell types (epithelium and stroma) to regulatory agents such as bIFN tau and oxytocin in cattle. Suppression of prostaglandin secretion by bIFN tau in epithelial cells of endometrial tissue is supportive of an antiluteolytic effect of bIFN tau.

Animals↗
PubMed1994

Involvement of the c-kit receptor in the adhesion of hematopoietic stem cells to stromal cells.

The proto-oncogene c-kit, encoding a receptor-type tyrosine kinase, is allelic with the W locus of the mouse. The stromal cell line OP9, capable of supporting long-term hematopoiesis, was newly established from a newborn B6C3F1-op/op mouse calvaria. When bone marrow cells of WBB6F1-W/Wv mice were cocultured with the OP9 cells in liquid medium, hematopoiesis declined to a level one-thousandth of that in the cocultures of bone marrow cells of WBB6F1-+/+ mice and stromal cells by day 21. In contrast, when bone marrow cells of W/Wv mice were cocultured with OP9 cells in semisolid medium, at least 61% of the number of colonies were detected until the end of our observation period of 42 days when compared with that in control cocultures, although colonies formed by hematopoietic stem cells of W/Wv mice were significantly smaller than those of normal stem cells. After a 24-hour incubation with OP9 cells, fewer stem cells of W/Wv mice than normal ones adhered to the stromal cells. Adhesion of normal stem cells to stromal cells was inhibited by the addition of an antagonists anti-c-kit monoclonal antibody, ACK2. These results demonstrate that the c-kit receptor plays an important role not only in the proliferative response of hematopoietic stem cells but also in their adhesion to stromal cells.

Animals↗
PubMed1996

Effects of irradiation of CBA/CA mice on hematopoietic stem cells and stromal cells in long-term bone marrow cultures.

Following 200 cGy total body irradiation, 20-25% of CBA/Ca mice and their CBA/B and CBA/H sublines develop myeloid leukemia. To determine whether hematologic changes in vitro were detectable, long-term marrow cultures (LTBMCs) were established from the right and left hind limbs of 11 individual control and 11 CBA/B mice 100-114 days after 200 cGy total body irradiation. Individual cultures were studied weekly for cumulative production of nonadherent cells and colony-forming, hematopoietic progenitor cells. Control cultures produced significantly more nonadherent cells over 25 weeks in long-term marrow culture compared to those from irradiated (treated) mice. Permanent stromal cell lines were established from control and irradiated CBA/B mouse LTBMCs and clonal sublines were established. The stromal cell lines from LTBMCs of in vivo irradiated CBA/B mice had uniformly lower plating efficiencies, and only one formed a permanent clonal subline at 100-fold lower frequency compared to stromal cell lines from control mouse LTBMCs. The irradiation sensitivity of both uncloned and clonal sublines was similar by single-hit, multi-hit or by linear quadratic formula. Cocultivation of an IL-3 dependent hematopoietic progenitor cell line established from a control CBA/B, LTBMC with control of irradiated stromal cell lines derived from either a control (CC3) or the one successfully cloned in vivo irradiated (CT4) LTBMC, produced few cobblestone islands in the presence of IL-3. In contrast, formation of cobblestone islands in the presence of L cell-condition medium as a source of M-CSF was significantly greater, and these persisted for 21 days on both CC3 and CT4 stromal lines. The data provide evidence for irradiation induced changes in the bone marrow stromal cell compartment of CBA/B mice which persist and are detectable in vitro 6 months after explant of the cells to culture. These marrow stromal cell lines may provide valuable resources for analyzing the molecular biologic changes in the hematopoietic microenvironment during irradiation leukemogenesis.

Animals↗
PubMed1987

Immunohistochemical detection of c-myc oncogene product in human gastric carcinomas: expression in tumor cells and stromal cells.

c-myc protein p62 in human gastric carcinomas was studied by immunohistochemistry using an antiserum raised against the C-terminal portion of the human c-myc gene protein. No immunoreactivity of c-myc p62 was detected in tumor cells of 27 early carcinomas, while c-myc p62-positive tumor cells were observed in 51 (36.4%) of the 140 advanced carcinomas. Many stromal cells including macrophages and fibroblasts around the tumors showed various degrees of c-myc p62 immunoreactivity. Localization of c-myc p62 was predominant in the cytoplasm of tumor cells and stromal cells. The prognosis of patients with p62-positive stromal cells was significantly better than that of patients with p62-negative stromal cells.

Epidermal Growth Factor↗
PubMed1989

[An attempt at in vitro reconstruction of the human endometrium by serum-free collagen gel culture of glandular epithelial cells and stromal cells].

Normal human endometrial epithelial cells were obtained from premenopausal women at the time of necessary hysterectomy. The cells were cultured in, or on, collagen gels in a serum-free medium, and were also co-cultured with endometrial stromal cells in order to reconstruct an in vitro model of the endometrium. They were observed morphologically and effects of estrogen (E) and progesterone (P) on the cultured cells were assessed. The epithelial cells and the stromal cells were separated by collagenase digestion and filtration through stainless steel sieves. The floating collagen gel method was used. The epithelial cells on the gels formed a monolayer sheet of columnar or cuboidal cells, and those in the gels grew three-dimensionally to form round or oval glands. The stromal cells co-cultured with the epithelial cells formed cell masses just under the epithelial cell layer quite similar to those of the endometrium in vivo. The epithelial cells on the gels did not migrate into the gels, nor did the stromal cells in the gels proliferate outside of the epithelial cell layer. The epithelial cells did proliferate, showing mitotic activity, but there was no evidence of their responding to E and/or P. The endometrial stromal cells themselves did not decidualize following the addition of E and/or P to the medium.

Adult↗
PubMed2002

Bone marrow stem cells and stromal cells in autoimmune cytopenias.

High-dose immunosuppression followed by autologous haemopoietic stem cell transplantation (ASCT) is a promising practice for the treatment of severe, resistant autoimmune disorders. Patients with refractory autoimmune cytopenias (AIC), primary or secondary to systemic autoimmune diseases (AID) including systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA), have been proposed as potential candidates for such a therapeutic procedure. An abnormal immune milieu, however, may affect the number and functional characteristics of stem cells and/or stromal cells in the bone marrow (BM) and might impact on harvesting and engraftment potential of stem cells or on BM reconstitution following engraftment in patients with AIC undergoing ASCT. Using flow cytometry and in vitro culture assays we have shown that patients with primary AIC display increased number of BM CD34+ cells in response to abnormally high production of granulocyte-colony stimulating factor (G-CSF) by BM stroma. In contrast, patients with AIC secondary to systemic AID display increased apoptosis of BM progenitor cells resulting in low CD34+ cell numbers and abnormal haemopoiesis supporting capacity of BM stroma due to the aberrant, local or systemic, inhibitory cytokine production or to intricate interactions between haemopoietic and immune cells present within the BM microenvironment. In this review we summarize the available knowledge on BM stem cell reserve and function and stromal cell function in patients with primary and secondary AIC with special reference to SLE and RA. The underlying mechanisms possibly involved in the pathogenesis of the observed abnormalities are also discussed.

Antigens, CD34↗
PubMed2001

Fluorometric detection of platelet activating factor receptor in cultured oviductal epithelial and stromal cells and endometrial stromal cells from bovine at different stages of the oestrous cycle and early pregnancy.

During the oestrous cycle and early pregnancy, the oviduct and uterus undergo a variety of morphological and physiological modifications in which the platelet activating factor receptor (PAF-R) plays an important role. PAF-R levels were quantified in bovine oviductal epithelial and stromal cells and endometrial stromal cells at days 2 to 4, 12, and 20 of the estrous cycle and during early pregnancy. Cells were grown in vitro and their intracellular PAF-R concentration was measured by flow cytometry using a polyclonal anti-PAF-R antibody system. A significant increase (P < 0.05) in the portion of PAF-R-positive oviductal epithelial and stromal cells was detected in both non-pregnant and pregnant cattle on days 2 to 4 in comparison to day 12 and 20. In endometrial stromal cells derived from day 20 pregnant bovine, a significant increase (P < 0.05) in PAF-R staining was observed in comparison to the day 20 non-pregnant and days 2 to 4 or 12 pregnant and non-pregnant animals. The PAF-R was detected in oviductal cells by using immunoblotting and immuno-gold postembedding method. Positive binding of the anti-PAF-R antibody was found on the cell membrane and in the cytoplasm. We concluded that the increased PAF-R concentration measured in cultured oviductal epithelial and stromal cells of cyclic and pregnant heifers on days 2 to 4 was hormonally regulated. The increased PAF-R in endometrial stromal cells on day 20 of pregnant heifers was a pregnancy-specific effect and may mediate a local increase in endometrial vascular permeability known to precede the implantation.

Animals↗
PubMed1997

Influence of cathepsin D expression in lung adenocarcinoma on prognosis: possible importance of its expression in tumor cells and stromal cells, and its intracellular polarization in tumor cells.

BACKGROUND: Cathepsin D, an aspartic lysosomal proteinase, has been described to be closely associated with tumor progression and prognosis in some human malignancies. The purpose of this study was to determine clinicopathological and prognostic significance of cathepsin D expression in lung adenocarcinoma. METHODS: Expression of cathepsin D in 152 lung adenocarcinoma patients was immunohistochemically studied using the antihuman cathepsin D antibody. RESULTS: Eighty patients (53%) showed negative immunoreactivities in tumor cells. The cathepsin D-positive patients (72 patients, 47%) were divided into two subgroups; granular type expression (48 patients, 31%) with its polarized expression mainly in the luminal side of the cytoplasm of tumor cells and basal type expression (24 patients, 16%) with its polarized expression mainly in the basal or infranuclear side of the cytoplasm. Patients with basal type expression showed significantly more marked scar formation (P = 0.042), and especially among the patients with stage I disease, those with basal type tended to show poorer prognosis (P = 0.071) than the others. Cathepsin D was also expressed in stromal cells within the tumor tissues, and 86 patients (57%) with moderate to massively infiltrating cathepsin D-positive stromal cells showed a lower grade of differentiation (P = 0.005) and higher scar grade (P = 0.0003) than those with few cathepsin D-positive stromal cells. Cathepsin D status in stromal cells was significantly associated with prognosis (P = 0.014), and in a multivariate analysis, its expression status in stomal cells was marginally an independent prognostic factor only among the stage I patients. CONCLUSIONS: In determining significance of cathepsin D expression in this disease, it is important to consider separately its expression cell type and its polarization pattern in tumor cells within the tumor tissue. How ever, only cathepsin D status in stromal cells within the tumor tissue is a marginal marker influencing prognosis among stage I patients.

Adenocarcinoma↗
PubMed1994

IL-1 alpha and TNF alpha act synergistically to stimulate production of myeloid colony-stimulating factors by cultured human bone marrow stromal cells and cloned stromal cell strains.

Human bone marrow stromal cells respond to stimulation by the monokines IL-1 and TNF by producing colony-stimulating factors such as GM-CSF and G-CSF. In this study we show that IL-1 alpha and TNF alpha act synergistically to stimulate GM-CSF and G-CSF production by cultured marrow stromal cells. We further show that IL-1 alpha and TNF alpha synergistically stimulate production of GM-CSF and G-CSF by a clonal stroma-derived cell strain. Although IL-1 and TNF share many of the same biological activities, we show that IL-1 alpha and TNF alpha have an unequal ability to induce myeloid-CSF production by both cultures, with IL-1 alpha being the more potent inducer. We found that induction by IL-1 alpha and TNF alpha was independent of cell proliferation. The effect of IL-1 alpha and TNF alpha on production of the two myeloid-CSFs by the clonal cells was significantly greater than the unfractionated passaged stromal cultures, having the greater effect on G-CSF production. The clonally derived stromal cells constitutively produced colony-stimulating activity, in particular GM-CSF, at levels easily detected by ELISA. These findings show that, in addition to the overlapping and additive activities of IL-1 alpha and TNF alpha, they can interact synergistically. Our findings further suggest that a small subpopulation of stroma cells may be the major producer of G-CSF in the marrow microenvironment during immune response.

Bone Marrow↗
PubMed2000

Sphingosine-1-phosphate and lysophosphatidic acid trigger invasion of primitive hematopoietic cells into stromal cell layers.

A new primitive hematopoietic cell line (THS119), exhibiting Lin(-)/Sca-1(+)/c-Kit(+) a surface phenotype, grew and survived underneath stromal cells (TBR59). The ability of the THS119 cells to invade these stromal cell layers was dependent on the inclusion of serum in the culture medium. This was apparently due to a requirement for lipids contained in serum. Their invasion of the stromal cell layers in serum-free cultures could be triggered by addition of sphingosine-1-phosphate (S1P) or lysophosphatidic acid (LPA) and was dependent on both Rho- and Ras-signaling pathways. Between the 2 possible receptors of S1P and LPA, edg-1 and edg-2, expression of edg-2 only was found to be correlated with immaturity and/or invasive activity of the primitive hematopoietic cells. These results suggest the importance of specific lipids and their specific receptors on the invasive activity of primitive hematopoietic cells in the hematopoietic microenvironment.

Animals↗
PubMed1993

Immunoelectron microscopic localization of transforming growth factor beta 1 and latent transforming growth factor beta 1 binding protein in human gastrointestinal carcinomas: qualitative difference between cancer cells and stromal cells.

Transforming growth factor beta 1 (TGF-beta 1) is secreted as an inactive complex associated with latent TGF-beta 1 binding protein (LTBP). Tissue localization of these proteins has not been fully understood in human pathological conditions. We examined the immunohistochemical localization of TGF-beta 1 precursor (proTGF-beta 1) and LTBP in carcinomas and granulation tissue in the human gastrointestinal tract at the light and electron microscopic levels. In normal tissue, endothelial cells and granulocytes sporadically showed immunoreactivity for proTGF-beta 1, while epithelial cells were all negative. In cancer tissue, both cancer cells and stromal cells (fibroblasts, macrophages, and endothelial cells) were positive for proTGF-beta 1, more frequently in diffuse-type gastric carcinomas than in differentiated-type adenocarcinomas. Immunoelectron microscopy revealed that proTGF-beta 1 was localized in rough endoplasmic reticulum and perinuclear cisternae in fibroblasts, macrophages, and endothelial cells in cancer stroma and in fibrous granulation tissue. In contrast, the intracellular localization of proTGF-beta 1 in carcinoma cells was predominantly observed in the cytosol (cytoplasmic matrix). This finding suggests disarranged or blocked intracellular transportation of proTGF-beta 1 in cancer cells. The immunoreactivity for LTBP was not observed in the normal epithelial cells. It was localized in cancer stroma, not in cancer cells. Ultrastructurally, LTBP was located in the extracellular matrix around fibroblasts and smooth muscle cells. The intracellular immunoreactivity for LTBP was observed in rough endoplasmic reticulum of fibroblasts and smooth muscle cells, the same as in granulation tissue. These results suggest that gastrointestinal carcinoma cells produce no or a small amount of LTBP in vivo. Our investigation suggests that extensive fibrosis in both cancer stroma and granulation tissues may be promoted by TGF-beta 1 mainly secreted from stromal cells.

Carrier Proteins↗
PubMed1996

Radiation-induced DNA damage in canine hemopoietic cells and stromal cells as measured by the comet assay.

Stromal cell progenitors (fibroblastoid colony-forming unit; CFU-Fs) are representative of the progenitor cell population of the hemopoietic microenvironment in bone marrow (BM). Previous studies of the radiation dose-effect relationships for colony formation have shown that canine CFU-Fs are relatively radioresistant as characterized by a D0 value of about 2.4 Gy. In contrast, hemopoietic progenitors are particularly radiosensitive (D0 values= 0.12-0.60 Gy. In the present study, the alkaline single-cell gel electrophoresis technique for the in situ quantitation of DNA strand breaks and alkali-labile sites was employed. Canine buffy coat cells from BM aspirates and cells harvested from CFU-F colonies or from mixed populations of adherent BM stomal cell (SC) layers were exposed to increasing doses of X-rays, embedded in agarose gel on slides, lysed with detergents, and placed in an electric field. DNA migrating from single cells in the gel was made visible as "comets" by ethidium bromide staining. Immediate DNA damage was much less in cultured stromal cells than in hemopoietic cells in BM aspirates. These results suggest that the observed differences in clonogenic survival could be partly due to differences in the type of the initial DNA damage between stromal cells and hemopoietic cells.

Animals↗