Studies of the hemopoietic microenvironments. V. Changes in murine splenic and bone marrow glycosaminoglycans during post irradiation hemopoietic regeneration

Changes in the amount of sulphated and unsulphated glycosaminoglycans in the spleen and the bone marrow of lethally irradiated mice were followed up to 11 days after irradiation. One day after irradiation, a sharp decrease in the amount of glycosaminoglycans was observed. In the absence of hemopoietic cells, the remaining stromal elements of spleen and bone marrow underwent subsequently marked changes in the amount of the sulphated and unsulphated components of the glycosaminoglycans in both organs. Reconstitution of irradiated mice with bone marrow cells affected the pattern of changes in the amount of glycosaminoglycans. Although no linear correlation could be observed between the amount of glycosaminoglycans and the number of hemopoietic cells present in the hemopoietic organs, these results suggest an interaction between hemopoietic cells and stromal cells in the hemopoietic organs with regard to the glycosaminoglycan metabolism.     

Prognostic factors in bone marrow transplantation for beta thalassemia major: experiences from Iran

This study concerns the effects of several pre-transplant features on outcome for patients with beta thalassemia major who underwent bone marrow transplantation (BMT). Seventy patients with beta thalassemia major underwent bone marrow transplantation during the period 1991-1997 in Shariati Hospital in Tehran, Iran. The survival and rejection curves levelled off at 8 and 18 months after transplantation at 82.6% and 11.4%, respectively. Pre-transplant clinical features (age, serum ferritin, portal fibrosis, hepatomegaly and quality of chelation therapy) were examined for their effects on survival and recurrence of thalassemia in this group of patients who were less than 16 years old. Increasing age, presence of portal fibrosis and increasing serum ferritin were significantly associated with reduced probability of survival (P = 0.0047, P = 0.016 and P = 0.024, respectively). Hepatomegaly and inadequate pre-transplant chelation therapy which were documented as poor prognostic factors in previous studies, were not evaluable in this study. We also showed the benefits of transplanting more than 5.5 x 10(8)/kg cells in this group of patients with no increase in complications.     

Evidence for persistence of human parvovirus B19 DNA in bone marrow

Neurological rehabilitation involves the systematic presentation of environmental stimuli and challenges that enable the patient to learn strategies for minimizing their disabilities. Rehabilitation therapy of transplant recipients may be an important factor in enhancing the efficacy of the transplanted organ or tissue to promote functional recovery. Laboratory research and clinical trials on neural transplantation, as an experimental treatment for neurological disorders (e.g., Parkinson's disease, Huntington's disease, and cerebral ischemia), have focused primarily on devising effective surgical implantation strategies with little attention devoted to the interaction between environmental factors and restorative neurosurgery. Exercise training as part of neurological rehabilitation may be an important factor for neural transplantation therapy for Parkinson's disease. Rehabilitation providers are particularly well placed to provide the environment and the support to optimize the behavioral functioning of neural transplant patients in learning to use the new grafted tissue.     

Risk factors for pulmonary tuberculosis in bone marrow transplant recipients

Little is known about the profile of infection with Mycobacterium tuberculosis in bone marrow transplant (BMT) recipients. Of five BMT series with a total of more than 5,000 patients, only 10 cases of M. tuberculosis infection were described, with an overall incidence of 0.19%. We have conducted a prospective evaluation of 183 consecutive BMT recipients, and 10 patients were found to develop pulmonary tuberculosis post-BMT, yielding an incidence of 5.5%. We described the clinical features of these 10 patients, and analyzed the risk factors for development of tuberculosis using age- and sex-matched case control subjects who did not develop the disease. The median age of the 10 patients who developed tuberculosis was 29 yr (range, 17 to 40 yr). The median time for onset of symptoms was 150 d (range, 23 to 550 d), mainly presenting with fever and cough, with infiltrates on chest radiograph. Respiratory tract specimens, mostly sputum, yielded positive smears for acid-fast bacilli in three and positive M. tuberculosis culture in eight, whereas lung tissue histology was the first diagnostic test in two patients. Treatment with standard antituberculosis drugs for a longer duration was highly effective, with no excessive side effects. Risk factors identified for development of tuberculosis included allogeneic BMT (p < 0.05, relative risk [RR] = 23.7), total body irradiation (p < 0. 05, RR = 4.9), and chronic graft-versus-host disease (GVHD) (p < 0. 05, RR = 3.6). It is postulated that chronic GVHD predisposed to development of tuberculosis mainly via disruption of host reconstitution of immune defenses against M. tuberculosis.     

Comparison of bone resorption markers during hypocalcemia in dairy cows

This study investigated whether hydroxyproline, deoxypyridinoline, or the carboxyterminal telopeptide of type I collagen could be used as markers to provide evidence of bone resorption during hypocalcemia of dairy cows. Serum concentrations of the amino-terminal propeptide of type III procollagen were also analyzed to study the effect of parturition on type III collagen, which is a component of soft connective tissues. Urine and blood samples were collected on d 1 to 5, on d 9, and d 14 after parturition from 18 cows with symptoms of periparturient paresis (group 1) and 19 healthy control cows without symptoms (group 2). Urine concentrations of hydroxyproline and deoxypyridinoline were measured with a colorimetric assay and an enzyme immunoassay, respectively. Serum concentrations of the amino-terminal propeptide of type III procollagen were measured using a commercially available radioimmunoassay. A radioimmunoassay was developed to analyze serum concentrations of the carboxyterminal telopeptide of bovine type I collagen. The mean corrected urinary hydroxyproline concentrations in group 1 increased from parturition to d 14; concentrations in group 1 were significantly higher for group 2 after d 5. Mean corrected deoxypyridinoline concentrations in urine increased after parturition to reach a peak at d 9, and serum concentrations of the carboxyterminal telopeptide of type I collagen peaked at d 5. However, mean concentrations of deoxypyridinoline and the carboxyterminal telopeptide of type I collagen did not differ significantly between groups. The variation in the behavior of the three markers is likely a reflection of the different phases and aspects of the bone collagen degradation. In conclusion, assays for urinary deoxypyridinoline and serum carboxyterminal telopeptide of type I collagen determinations are useful tools to follow the course of degradation of bone collagen in dairy cows.     

Bilateral tibial marrow ablation in rats induces a rapid hypercalcemia arising from extratibial bone resorption inhibitable by methylprednisolone or deflazacort

OBJECTIVES: To determine the effect of alpha-linolenic acid (ALA) intake (or the dietary linoleic acid [LA]/ALA ratio) on the growth and visual function of term infants. STUDY DESIGN: Normal term infants were assigned randomly and in masked fashion at birth to receive formulas with approximately 16% of total fatty acids as LA and 0.4%, 1.0%, 1.7%, or 3.2% of fatty acids as ALA (LA/ALA ratios of 44, 18.2, 9.7, and 4.8) for the first 4 months of life. The fatty acid pattern of plasma phospholipids was determined shortly after birth and at approximately 21, 60, and 120 days of age. Anthropometric data were obtained at the same times and also at approximately 240 days of age. Transient visual evoked responses (VERs) were measured at approximately 120 and 240 days of age. For comparisons, anthropometric and VER data also were obtained in infants who were exclusively breast-fed for the first 4 months of life. RESULTS: Infants who received the formula with 3.2% ALA (LA/ALA ratio, 4.8) had higher plasma concentrations of phospholipid docosahexaenoic acid (DHA) but lower concentrations of arachidonic acid at 21, 60, and 120 days of age. Mean weight of this group at 120 days of age was 760 gm less (p < 0.05) than the mean weight of the group that received the formula with 0.4% ALA (LA/ALA ratio, 44). Despite differences in plasma phospholipid DHA contents among groups, neither VER latency nor amplitude differed significantly among formula groups or between any formula group and age-matched, breast-fed infants. CONCLUSIONS: The highest versus the lowest ALA intake (or the lowest vs the highest LA/ALA ratio) resulted in higher plasma phospholipid DHA content from 21 to 120 days of age but was not associated with improved visual function as assessed by transient VER. Moreover, mean body weight of infants who received the highest versus lowest ALA intake was less at 120 days (p < 0.05). These data suggest that the lower LA/ALA ratios currently recommended for infant formulas should not be adopted until the effect of such ratios on growth are evaluated more completely.     

Induction of Fas ligand in murine bone marrow NK cells by bacterial polysaccharides

Bacterial polysaccharides have a wide range of activities in mammals. We have studied the effect of LPS and poly-beta-(1-->4)-D-mannuronate (mannuronan, poly-M), an exopolysaccharide from Pseudomonas aeruginosa, on the cytotoxicity mediated by murine bone marrow cells (BMC). Addition of LPS or mannuronan to BMC induced a time- and dose-dependent cytotoxicity against Jurkat cells. The LPS- or mannuronan-induced cytotoxicity was due to increased Fas ligand (FasL) expression by BMC, since 1) Fas-transfected L1210-Fas target cells were more susceptible to lysis than the Fas(low)-expressing parent L1210 cells, 2) stimulated BMC from FasL-defective gld/gld mice were not cytolytic and, 3) the cytolytic activity of normal BMC was inhibited by a Fas-Fc fusion protein. Flow cytometry showed an increase in surface FasL in LPS-stimulated BMC. RT-PCR analysis of BMC revealed constitutive expression of FasL mRNA, which was increased after LPS stimulation. Immunomagnetic depletion of NK1.1-, CD2-, or CD32/16-expressing cells from BMC abrogated the LPS-induced BMC cytotoxicity against L1210-Fas cells, suggesting that NK cells were the cytotoxic effector cells. Depletion of CD45R/B220-, Gr-1-, or CD11b/Mac-1-expressing cells only partially decreased BMC-mediated cytotoxicity, and depletion of CD4- or CD8a-expressing cells had no effect. The results support the conclusion that LPS and mannuronan induce expression of cytotoxic FasL on bone marrow NK cells.     

Immortalization and characterization of bone marrow stromal fibroblasts from a patient with a loss of function mutation in the estrogen receptor-alpha gene

A male patient with abnormal postpubertal bone elongation was shown earlier to have a mutation in both alleles of the estrogen receptor, resulting in a nonfunctional gene. Marrow stromal fibroblasts (MSFs) derived from this patient were called HERKOs (human estrogen receptor knock outs), and in order to obtain continuous HERKO cell lines, they were immortalized using a recombinant adenovirus-origin-minus SV40 virus. MSFs are unique cells because they support hematopoesis and contain a mixed population of precursor cells for bone, cartilage, and fat. Three established cell lines (HERKO2, HERKO4, and HERKO7) were characterized and compared with the heterogeneous population of nonimmortalized HERKOs for their osteogenic potential. We performed Northern analysis of matrix genes implicated in bone development and metabolism and an in vivo bone formation assay by transplanting the cells subcutaneously into immunodeficient mice. All three HERKO lines expressed high amounts of collagen 1A1, osteopontin, osteonectin, fibronectin, decorin, biglycan, and alkaline phosphatase. Except for osteopontin, expression of these genes was slightly lower compared with nonimmortalized HERKOs. In the in vivo bone formation assay, the heterogeneous population of nonimmortalized HERKOs formed bone with high efficiency, while the HERKO lines induced a high-density, bone-like matrix. Finally, all HERKO cell types secreted high levels of insulin-like growth factor I and interleukin-6 into the culture medium relative to cells of normal human subjects. In summary, these lines of HERKO cells retain several of the phenotypic traits of MSFs after immortalization, including matrix and cytokine production, and provide a valuable source of a unique human material for future studies involving estrogen action in bone and bone marrow metabolism.     

Evidence from molecular genetic and cytogenetic analyses that bone marrow histopathology is reliable in the diagnosis of chronic myeloproliferative disorders

The reliability of histopathological diagnosis in bone marrow specimens from patients with chronic myeloproliferative disorders (CMPD) was evaluated by correlating the histological findings with molecular genetic and cytogenetic analyses of the Ph1-translocation. A rearrangement of m-bcr was detected only in patients (28/30) diagnosed histologically as chronic myeloid leukemia (CML). This finding was supported by the presence of a Ph1-chromosome in 24/26 patients with CML examined. All the patients with other types of CMPD, including polycythemia vera (PV), primary thrombocythemia (PTH) and chronic megakaryocytic-granulocytic myelosis (CMGM), as well as those with unclassifiable CMPD (CMPD.UC) were Ph1-negative (n = 38). The histopathological discrimination of CML from Ph1-negative varieties of CMPD was also reliable for patients with myelofibrosis complicating CML, CMGM and CMPD.UC. The results demonstrate that bone marrow histopathology allows a reliable diagnosis of CML. This is in contrast with hematological data such as high platelet counts which show considerable overlapping in the various forms of CMPD.     

HL-A antigens in serum from bone marrow harvested for diagnostic purposes

Tissue typing employing the lymphocytotoxicity micro-assay was performed by introducing in the system approximately 15% of autologous or homologous ultra-centrifuged blood serum, both of peripheral and bone marrow source. As a rule, the pattern of reactivity was unaffected by peripheral blood serum, whereas the cytotoxicity was nearly always inhibited by autologous bone marrow blood serum or by homologous one from donors sharing the inhibited HL-A specificities with the donors of lymphocytes. These results suggest that at least in these patients HL-A antigens are present in bone marrow blood serum at a concentration which is definitely higher than in peripheral blood.     

Evidence for the colocalization of estrogen receptor-beta mRNA and estrogen receptor-alpha immunoreactivity in neurons of the rat forebrain

Estrogen receptor beta (ER beta) mRNA is expressed in several rat brain regions where ER alpha is abundant. In vitro studies have shown that ER alpha and ER beta can heterodimerize and that the activity of this complex may be different than an ER alpha or ER beta homodimer complex. The purpose of the present study was to ascertain if ER alpha and ER beta are co-expressed by certain neuronal populations using a double label in situ hybridization/immunocytochemistry method. The results revealed that neurons in the bed nucleus of the stria terminalis, medial amygdala and preoptic area contain both ERs, with the vast majority of the neurons being double labeled. In other brain regions including the arcuate nucleus, cortical amygdaloid nuclei and ventromedial nucleus, only a few double-labeled cells were detected, while neurons in the paraventricular nucleus, supraoptic nucleus, and cerebral cortex expressed only ER beta mRNA. The results of these double label experiments provide the first evidence that ER alpha and ER beta coexist in neurons under in vivo conditions and suggest that estrogens may differentially modulate the activity of certain neuronal populations depending on whether the cells expresses ER alpha, ER beta or both ERs.     

Choice and contingency in the development of behavioral autonomy during instrumental conditioning.

In two experiments hungry rats received extensive training to lever press for food outcomes before one outcome was devalued by aversion conditioning and responding tested in extinction. If the rats were trained on a concurrent schedule in which two responses yielded different outcomes, performance during the extinction test was reduced by devaluation of the associated outcome. By contrast, if a single response was trained concurrently with the noncontingent presentations of the other outcome, test performance was insensitive to devaluation of the contingent outcome. This finding demonstrates that training on a schedule that offers a choice between responses that yield different outcomes prevents the onset of behavioral autonomy. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Radiation dose-fractionation and dose-rate relationships for long-term repopulating hemopoietic stem cells in a murine bone marrow transplant model

Fractionated and low-dose-rate total-body irradiation (TBI) were compared with single-dose high-dose-rate TBI for induction of long-term hemopoietic chimerism in a murine syngeneic bone marrow transplantation model. At 5 months after TBI and bone marrow transplantation, the degree of stable blood chimerism was determined from the proportion of stem cell-derived donor (B6-Gpi-1a) and host (B6-Gpi-1b) blood erythrocytes. This end point was used to construct radiation dose-response curves for long-term donor marrow engraftment corresponding to ablation of primitive bone marrow stem cells of the host. Increasing dose fractionation and decreasing dose rate had the effect of restoring host hemopoiesis and required higher TBI doses for equal donor engraftment. Most of the dose recovery occurred within the first 6 h between fractions, consistent with the kinetics of sublethal damage repair. The late chimerism data were fitted to the linear-quadratic model using indirect and direct analysis for a fixed threshold response. Both analyses gave relatively low alpha/beta ratios (below 2 Gy), within the range normally seen in late-responding tissues. The dose-rate data gave a repair half-time of 2 h as estimated by the incomplete-repair model. These estimates contrast with the much higher alpha/beta values and lower repair half-times derived from acute hemopoietic failure as indicated by LD50/30, with the implication that separate target cell populations with differing radiosensitivities are involved in these two bone marrow end points.     

Unique risk factors for bacteraemia in allogeneic bone marrow transplant recipients before and after engraftment

A study of the risk factors associated with bacteraemia in 191 allogeneic bone marrow transplant (BMT) recipients (1991-1996) was performed. In contrast to risk factors commonly cited for cancer chemotherapy, mucositis, degree of conditioning toxicity of the gut and lungs, duration of neutropenia, and severity of neutropenia and monocytopenia were not associated with bacteraemia in the pre-engraftment period, during which the only significant risk factor was late stage underlying disease (P < 0.05). After engraftment, Hickman catheter infection, and severe acute and chronic graft-versus-host disease (GVHD) were found to be independently associated with bacteraemia by multivariate analysis (P < 0.001, <0.05 and <0.05, respectively). This might be explained by intense antimicrobial prophylaxis, early empirical treatment, and non-routine use of haemopoietic growth factors. No significant difference in mortality was detected between bacteraemic and non-bacteraemic patients in both periods. Allogeneic BMT recipients are therefore a group of patients distinct from other cancer patients receiving chemotherapy at risk of developing bacteraemia. The study findings prompt consideration of a management protocol incorporating early and routine use of haemopoietic growth factors before engraftment in high-risk patients with late stage underlying malignancies, routine antimicrobial prophylaxis for acute GVHD with intense immunosuppression, and intravenous immunoglobulin therapy for chronic GVHD. Further cost-benefit analyses are warranted.     

Requirement of donor-derived stromal cells in the bone marrow for successful allogeneic bone marrow transplantation. Complete prevention of recurrence of autoimmune diseases in MRL/MP-Ipr/Ipr mice by transplantation of bone marrow plus bones (stromal cells) from the same donor

MRL/MP-Ipr/Ipr (MRL/Ipr) mice possess radioresistant (9.5 Gy) abnormal stem cells and show a recurrence of autoimmune diseases within 5 mo of conventional allogeneic bone marrow transplantation. We recently have found that the MHC preference exists between hemopoietic stem cells and stromal cells; when bones are engrafted, donor-derived stromal cells present in the engrafted bones can migrate into the recipient bone marrows, which are replaced with both donor-derived stromal cells and hematopoietic cells. Based on these findings, we attempted to prevent the recurrence of autoimmune diseases in MRL/Ipr mice by the transplantation of both bone marrow cells and bone (as a source of stromal cells). MRL/Ipr mice were irradiated (8.5 Gy) and then reconstituted with C57BL/6 bone marrow cells plus bone grafts. The mice survived more than 48 wk after this treatment. Immunohistologic studies revealed that the mice were completely free from both lymphadenopathy and autoimmune diseases such as lupus nephritis and rheumatoid arthritis. Sera from these mice showed normal levels of circulating immune complexes and rheumatoid factors. Normal functions of both T cells and B cells were noted. Abnormal T cells such as Thy-1+B220+ cells present in nontreated MRL/Ipr mice could not be seen in the thus-treated mice. In addition, to our surprise, spleen cells from treated mice showed completely normal in vitro primary anti-SRBC responses. These results indicate that stromal cells in allogeneic bone marrow transplantation play a crucial role not only in the prevention of graft failure but also in the successful cooperation among APCs, T cells, and B cells. Although MRL/Ipr mice are radiosensitive and usually die of interstitial pneumonia or fatty liver due to the side effects of radiation, it should be noted that this strategy allows a reduction in the radiation dose (9.5 Gy-->8.5 Gy), and that these mice can survive more than 48 wk without showing any symptoms of autoimmune diseases.     

Intracardiac right-to-left shunting and the risk of stroke during bone marrow infusion

We describe a patient who developed a paradoxical embolus to the brain during infusion of bone marrow. She had a patent foramen ovale through which right-to-left shunting led to multiple cerebral emboli. This complication can be prevented by positioning the tip of the infusion catheter in the main pulmonary artery and reducing the volume of marrow product infused.