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In vitro demonstration of a fundamental difference in the proliferation of murine and human bone marrow and lymphocytes following ultraviolet irradiation: relevance to bone marrow transplantation.

Exposure of rodent allogeneic donor marrow and splenocyte grafts to ultraviolet radiation (UVR) has been shown to permit durable engraftment at doses that abolish graft-versus-host disease (GVHD) and graft rejection. We have compared both murine and human alloreactive and mitogen-induced lymphoid responses and bone marrow proliferation in mixed lymphocyte culture (MLC), phytohemagglutinin (PHA)-induced proliferation and colony-forming unit-granulocyte/macrophage (CFU-GM) assays using germicidal UVC (200-290 nm), broadband and narrowband UVB (290-320 nm) and UVA (320-400 nm) sources. Our data show a wavelength and dose-dependent reduction in lymphoid proliferation in the mouse with CFU-GM survival of 50-75% of control at doses required to abolish allogeneic lymphocyte responses for all lamps. In contrast, human lymphocyte responses are more resistant to UVC with CFU-GM proliferation reduced to zero when allostimulation is abolished. Mitogen-induced lymphoid responses show a similar wavelength-dependent sensitivity. Abolition of response in MLC using UV-irradiated stimulator cells was less sensitive than proliferation with UV-irradiated responder cells at all wavelengths in both species. With all sources, murine CFU-GM proliferation is less susceptible to UVR than human marrow at doses required to abolish lymphoid responses.

Animals↗

Low T3-syndrome and nutritional status as prognostic factors in patients undergoing bone marrow transplantation.

Bone marrow transplantation is known to be associated with considerable morbidity and mortality. The aim of this study was to determine the influence of nutritional status and development of sick euthyroid syndrome as prognostic factors for outcome after BMT. In 100 patients who underwent transplantation the following parameters were assessed before and at day 14 and 28 after transplantation: anthropometric data (body weight, body mass index, body composition, grip strength), rapid turnover proteins transferrin and prealbumin, T4, T3, free T4, reverse T3, thyroid-stimulating hormone and thyroglobulin. Following bone marrow transplantation, 22 patients died in the short-term follow-up (group A) before day 140 after BMT, 21 patients died during further follow-up between days 140 and 365 (group B) and 57 patients survived longer than 365 days (group C). All patients experienced a significant decrease of transferrin and T3, accompanied by an increase of rT3 and rT3/T3 ratio at day 14 after BMT. At day 28 after BMT, patients in group C showed recovery from these changes with an increase of transferrin and a fall in rT3 and the rT3/T3-ratio, which was not seen in patients who died during further follow-up (groups A and B). The observed changes were independent of other prognostically relevant factors (type of disease, HLA-match, immunosuppression). Impaired nutritional status and development of a sick euthyroid syndrome, without tendency to recovery, are associated with a higher probability of fatal outcome after bone marrow transplantation and have prognostic relevance in this group of patients.

Anthropometry↗

[Interstitial pneumonia (IP) in bone marrow transplantation in leukemia--120 cases analysis in Nagoya Bone Marrow Transplantation Group].

Results of the bone marrow transplantation (BMT) for 120 cases of leukemia, which were done in nine institutes in Nagoya (Nagoya Bone Marrow Transplantation Group) last ten years, were analyzed to determine the factors associated with an increased risk of developing interstitial pneumonia (IP). IP developed 49 out of 120 patients (49.8%) and case fatality rate was 63.3%. The median time from transplantation to onset of IP was 81 days (range 13-575 days), in 30 out of 49 cases (61.2%), this complication developed within 100 days after transplantation. Of the 49 patients who developed IP, cytomegalovirus (CMV) infection was associated in 18 cases (36.7%), no cases of P. carinii infection was detected. Five factors were associated with an increased risk for developing IP, (1) older age (greater than or equal to 47.0%: less than 10 y. 10.0%) (p less than 0.01) (2) disease stage at BMT (non-remission 76.2%: remission 32.5%) (p less than 0.01) (3) presence of acute GVHD ((+) 52.5% (-) 28.8%) (p less than 0.05) (4) onset day after BMT (less than or equal to 100 days 61.2%: greater than 100 d. 38.8%) (p less than 0.01) (5) sex matching between donor and patient (sex match 28.8%: sex mismatch 57.1%) (p less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗

Post-irradiation thymocyte regeneration after bone marrow transplantation. I. Regeneration and quantification of thymocyte progenitor cells in the bone marrow.

Growth kinetics of the donor-type thymus cell population after transplantation of bone marrow into irradiated syngeneic recipient mice is biphasic. During the first rapid phase of regeneration, lasting until day 19 after transplantation, the rate of development of the donor cells is independent of the number of bone marrow cells inoculated. The second slow phase is observed only when low numbers of bone marrow cells (2.5 x 10(4)) are transplanted. The decrease in the rate of development is attributed to an efflux of donor cells from the thymus because, at the same time, the first immunologically competent cells are found in spleen. After bone marrow transplantation the regeneration of thymocyte progenitor cells in the marrow is delayed when compared to regeneration of CFUs. Therefore, regenerating marrow has a greatly reduced capacity to restore the thymus cell population. One week after transplantation of 3 x 10(6) cells, 1% of normal capacity of bone marrow is found. It is concluded that the regenerating thymus cells population after bone marrow transplantation is composed of the direct progeny of precursor cells in the inoculum.

Animals↗

A function of CD10 on bone marrow stroma.

Bone marrow (BM) stromal cells express CD10 (cALLA), a surface antigen now known to be a neutral endopeptidase (NEP-24.11). The function of CD10 in BM stroma is unknown, although purified NEP-24.11 is known to degrade different substrates including interleukin 1 beta (IL-1 beta). We have therefore employed a CD10-positive BM stromal cell line (L2AK) which proliferates in response to IL-1 beta to test the hypothesis that degradation of this cytokine is one of the functions of stromal CD10. We first showed that [3H]thymidine incorporation by L2AK cells is enhanced by IL-1 beta in a clear dose-dependent manner. Addition of the CD10 inhibitor, phosphoramidon, together with IL-1 beta resulted in a left shift in the dose-response curve which corresponded to a 10-fold potentiation of the IL-1 beta effect. These results indicate that CD10 on bone marrow stromal cells can degrade IL-1 beta and therefore provide a local control of the effects of this, and possibly other, growth factor(s).

Bone Marrow↗

Defective lymphopoiesis in the bone marrow of motheaten (me/me) and viable motheaten (mev/mev) mutant mice. III. Normal mouse bone marrow cells enable mev/mev prothymocytes to generate thymocytes after intravenous transfer.

Bone marrow prothymocytes from me/me and mev/mev mutant mice fail to generate thymocytes in irradiated (600 rad) +/+ wild-type recipients after intravenous injection. However, these same prothymocytes readily generate thymocytes after intrathymic injection. The results of the present study demonstrate that this apparent defect in the thymus-homing capacity of mev/mev prothymocytes can be corrected by mixing irradiated wild-type bone marrow cells with mev/mev bone marrow cells before intravenous injection. However, this defect is not corrected by passage of mev/mev bone marrow cells through the bone marrow of irradiated wild-type recipients. One interpretation of these results is that the maturation of prothymocytes is reversibly arrested in mev/mev mice by a defect in the radiosensitive compartment of the bone marrow microenvironment.

Animals↗

Active enhancement in the primate by bone marrow I. Ia-like determinants on nucleated bone marrow cells.

This study investigates the possibility that the active enhancing properties of bone marrow (BM) cells may be related to Ia-like determinants on their surface membranes. Less chromium was released on a cell per cell basis from nucleated baboon BM cells used as targets in complement-dependent cytotoxicity than from labelled lymph node lymphocytes. On the other hand, BM cells stimulated allogeneic lymphocytes in culture more vigorously than lymph node lymphocytes. BM cells with lymphocyte-activating properties could be enriched by fractionation on a discontinous bovine serum albumin gradietnt. BM cells responded poorly to allogeneic cell stimulation in mixed culture. Platelet absorption studies of an alloantiserum supported the conclusion that nucleated BM cells, and particularly BM fractions collected from the less dense interfaces of a bovine serum albumin gradient, express relatively more lymphocyte-activating or Ia-like determinants than "serologically defined" determinants.

Absorption↗

Progression of bone marrow abnormalities in malignant hemopoietic dysplasia. Evaluation of cytological changes with serial bone marrow samples.

Progression of bone marrow abnormalities in 12 patients with malignant (fatal) hemopoietic dysplasia (HD) was evaluated retrospectively in sequential bone marrow samples, using a simple morphological scoring technique. Special attention was paid to alterations in cytological abnormalities in the myeloid series. Patients who developed acute myeloid leukemia showed increasing scores during the clinical course and a rapid final increase in particularly myeloid nucleolar abnormalities. Patients who did not develop overt myeloid leukemia but died from bleeding and/or infection had low and stable scoring and showed no progression of cytological abnormalities. Repeated evaluation, especially of cytological changes in the bone marrow, and morphological scoring might be useful as predictive tools to estimate the risk of a leukemic transformation in some patients with HD.

Aged↗

Bone marrow sampling.

Bone marrow sampling may be indicated in the evaluation of iron metabolism, fever of unknown origin, anemia, leukocytosis and other hematologic abnormalities. The commonly accepted means of marrow sampling are needle aspiration and needle biopsy. The preferred site for obtaining the sample is the iliac crest. Contraindications to the procedure include infection or previous radiation therapy at the sample site, poor patient cooperation and major coagulation defects. The assistance of a skilled technician, use of a prepackaged tray, thorough patient education, use of local anesthesia and attention to aseptic technique help simplify bone marrow sampling.

Biopsy, Needle↗

Phenotyping of phosphoglucomutase (PGM1) isoenzymes--a new method for the follow-up of chimerism after bone marrow transplantation. Follow-up of chimerism after bone marrow transplantation by phenotyping of phosphoglucomutase (PGM1) isoenzymes.

In this study, phenotyping of phosphoglucomutase isoenzymes (PGM1, EC 2.7.5.1) in erythrocytes of bone marrow donors and hosts was shown to present a simple and exact method for the follow-up of chimerism after bone marrow transplantation, aimed at a better assessment of the transplant state and function.

Bone Marrow Transplantation↗

The sensitivity of leukemic bone marrow to simvastatin is lost at remission: a potential purging agent for autologous bone marrow transplantation.

BACKGROUND: Autologous bone marrow transplantation (ABMT) is frequently used in the treatment of malignant disease but carries the risk of reintroducing tumor cells into the patient. Methods are required for removing malignant cells from harvested bone marrow (BM) without impairing hematopoietic reconstitution. We have shown that simvastatin is toxic to leukemic progenitor cells at a concentration that conserves normal BM progenitors and may be of use clinically as a novel BM purging agent. METHODS: A two-stage culture system was used to compare the effects of simvastatin on both normal BM progenitor and primary acute myeloid leukemia (AML) cells. AML cells and normal BM mononuclear cells were incubated for 18 hours in suspension culture with 10 micrograms per mL simvastatin and the numbers of surviving clonogenic progenitor cells assayed in semisolid agar culture. RESULTS: Following simvastatin treatment of 18 AML cell populations, the mean surviving fraction of progenitor cells was 21.3 +/- 4.8% ( +/- standard error of the mean [SEM]). In contrast, the mean survival of normal BM progenitors from 16 donors was 89.6 +/- 8.6% ( +/- SEM). Samples were taken from 6 AML patients before treatment and after remission of disease had been induced by chemotherapy. In 5 of these cases the AML sample was significantly more sensitive to simvastatin than the remission sample, 4 of the 5 showed > 80% difference in progenitor cell survival. CONCLUSIONS: AML progenitor cells are sensitive to a short-term exposure to simvastatin that spares normal BM hematopoietic progenitor cells. We conclude that simvastatin may be an effective in vitro purging agent in ABMT for AML.

Acute Disease↗

Autologous bone marrow transplantation for high-grade lymphoid malignancy using melphalan/irradiation conditioning without marrow purging or cryopreservation. The Northern Regional Bone Marrow Transplant Group.

We report the safety and efficacy of 34 consecutive autologous bone marrow transplant (ABMT) procedures performed in adult patients with high-grade lymphoid malignancy after remission induction therapy. Fifteen patients with acute lymphoblastic leukemia (ALL) and six with high-grade non-Hodgkin's lymphoma (NHL) received pretransplant conditioning with intravenous (IV) melphalan and fractionated total body irradiation (TBI). Thirteen other patients with NHL were conditioned with melphalan alone, having previously received local involved field radiotherapy. Unmanipulated noncryopreserved autologous marrow was reinfused within 48 hours of harvesting. Engraftment occurred in all patients with medians of 10 days of neutropenia (neutrophils less than 0.5 x 10(9)/L), 4-day platelet transfusion requirement, 3 U packed RBC transfusion, and 18 days in hospital posttransplant. There were no procedure-related deaths. Actuarial disease-free survival in the 13 patients with ALL receiving autotransplant early in first remission is 48% with a median follow-up of 3 years. Two other ALL patients who had autotransplants after a period of maintenance therapy also remain in complete remission (CR). These results compare favorably with our 34% disease-free survival (DFS) in 15 allogeneic ALL transplant patients and 21% DFS in 19 patients on standard maintenance after a common induction schedule. No relapses have occurred in the 17 NHL patients transplanted in remission (median follow-up 2 years), but the two NHL patients who developed recurrent disease before ABMT died of progressive disease after temporary responses. We conclude that this method of ABMT results in rapid reengraftment with lack of toxicity and that the conditioning treatment used shows good efficacy against disease. It is applicable in high-grade lymphoid malignancy in first remission, and our results call into question the need for marrow purging in ALL and NHL patients transplanted in first remission.

Adult↗

Increased apoptosis of bone marrow CD34(+) cells and impaired function of bone marrow stromal cells in patients with systemic lupus erythematosus.

The changes in bone marrow (BM) stem cell reserve and function and stromal cell function in patients with active systemic lupus erythematosus (SLE) were investigated. The study was carried out on seven SLE patients and 28 healthy controls using flow cytometry and in vitro cell culture assays. We found that patients had low CD34(+) cells, compared with the control group, reflecting the decrease of both CD34(+)/CD38(-) and CD34(+)/CD38(+) cells. Patient CD34(+)/Fas(+) but not CD34(-)/Fas(+) cells were significantly increased. Apoptotic (7AAD(dim)) cells were higher among CD34(+)/Fas(+) than among CD34(+)/Fas(-) cells, and individual values of apoptotic CD34+ cells strongly correlated with the number of CD34(+)/Fas(+) cells. These findings are suggestive of a Fas-mediated apoptosis accounting for the low CD34(+) cells in SLE patients. Moreover, we found that patients had low numbers of granulocyte-macrophage colony-forming units (CFU-GM) and erythroid burst-forming units (BFU-E), compared with the control group, and that the generation of colony-forming cells in long-term BM cultures was significantly reduced. Patient BM stroma failed to support allogeneic progenitor cell growth. In one patient, CD34(+) cells were increased, apoptotic CD34(+)/Fas(+) cells were normalized and defective stromal cell function was restored after autologous stem cell transplantation. We concluded that defective haemopoiesis in SLE patients is probably caused, at least in part, to the presence of autoreactive lymphocytes in BM.

Adult↗

Hypoxia increases VEGF-A production by prostate cancer and bone marrow stromal cells and initiates paracrine activation of bone marrow endothelial cells.

Hypoxia develops at sites of rapid cancer growth near sites of poorly organized vasculature. Heparin binding growth factors (HBGFs) support neoangiogenesis of tumors. We examined the effect of culturing bone-targeted, metastatic C4-2B prostate cancer cells and bone stromal derived HS27a cells under hypoxic conditions on expression of vascular endothelial growth factor (VEGF) family members. A sealed chamber infused with 1% (hypoxic) or 20% (normoxic) O(2) was used. Both cell lines produced VEGF-A in normoxia, but little or no HB-EGF, another HBGF. HS27a cells produced low levels of FGF-2 and HGF, but little or none was secreted by C4-2B cells. Levels of VEGF-A in conditioned medium (CM) from both cell lines doubled when cultured in hypoxia. Similar changes in VEGF-A mRNA levels were seen. Receptor expression was unchanged by hypoxia. Changes in VEGF-A expression during hypoxia were preceded by nuclear accumulation of hypoxia inducible factor-1alpha (HIF-1alpha). Bone marrow endothelial (BME) cells express high levels of VEGFR2/flk-1, and are targets of VEGF-A induced neovascularization. BME cells proliferated in response to treatment with HS27a CM, but not C4-2B CM. BME cells formed tube-like angiogenic structures on growth factor reduced Matrigel in response to CM from HS27a or C4-2B cells. This response was greater when CM was produced under hypoxia, and was reduced by VEGF-A or FGF-2 neutralizing antibodies. We conclude that hypoxia triggers a physiologically relevant increase in VEGF-A by prostate cancer and bone marrow stromal cells which involves a paracrine loop that recruits and activates BME to support tumor neovascularization-related processes.

Bone Marrow Cells↗

Long-term bone marrow culture in persons with Fanconi anemia and bone marrow failure.

Fanconi anemia is an autosomal recessive disease characterized by a high risk of developing bone marrow (BM) failure and acute myelogenous leukemia. We studied growth of hematopoietic progenitor cells in long-term BM culture (LTBMC) in 8 persons with Fanconi anemia and BM failure. Although LTBMC were initiated with very few BM cells, an adherent layer formed in cultures from 7 persons. In these cultures, the number of nonadherent cells increased for 10 to 15 days. Cell growth continued until cultures were terminated at day 35 to 40. During the first 2 weeks of culture, most nonadherent cells were differentiated myeloid cells. By days 35 to 40, the adherent layer contained cells able to initiate secondary LTBMCs. These data indicate that hematopoietic precursors cells able to proliferate and differentiate in vitro are present in the BM of persons with Fanconi anemia and BM failure. They suggest that mechanisms other than absent precursor cells are responsible for BM failure in Fanconi anemia.

Adolescent↗

[Bone marrow infiltration in B-cell non-Hodgkin's lymphomas: comparison between flow cytometry and bone marrow biopsy].

A comparison of flow cytometry (FC) and bone marrow biopsy (BMB) to evaluate bone marrow infiltration was made in 114 patients suffering from B-cell non-Hodgkin's lymphomas (NHLs; 51 at diagnosis, 63 during post-therapy follow-up). The following parameters were indicative of bone marrow infiltration: altered surface k/l ratio; specific immunophenotypic pattern in particular NHLs (CLL, mantle cell lymphoma, hairy cell leukemia). FC and BMB agreed in 89.5% of cases (i.e. both showed 48 positive and 54 negative cases). In discordant cases (7.9%) and in cases not evaluable by FC (2.6%) IgH rearrangement and bcl-1 gene expression, both evaluated by PCR methods, were used to detect bone marrow infiltration with higher precision. These results show that a more complex analysis of bone marrow is needed to diagnose bone marrow infiltration, particularly in samples with minimal residual disease.

Adult↗