黑斑蛙骨髓细胞染色体标本制备的新方法

染色体标本制备是黑斑蛙分子细胞遗传学研究的基础。为了简化操作程序,缩短实验周期,建立了黑斑蛙染色体制备的一种新方法——骨髓细胞体外短时培养与秋水仙素同步处理法。该方法操作简便,重复性较好,可在较短时间里制备出分裂相较多且形态良好的蛙染色体标本,适用于核型分析、染色体荧光原位杂交、染色体显微分离和单染色体文库构建等多个方面的研究。     

Culture of myeloid dendritic cells from bone marrow precursors

Myeloid dendritic cells (DCs) are frequently used to study the interactions between innate and adaptive immune mechanisms and the early response to infection. Because these are the most potent antigen presenting cells, DCs are being increasingly used as a vaccine vector to study the induction of antigen-specific immune responses. In this video, we demonstrate the procedure for harvesting tibias and femurs from a donor mouse, processing the bone marrow and differentiating DCs in vitro. The properties of DCs change following stimulation: immature dendritic cells are potent phagocytes, whereas mature DCs are capable of antigen presentation and interaction with CD4+ and CD8+ T cells. This change in functional activity corresponds with the upregulation of cell surface markers and cytokine production. Many agents can be used to mature DCs, including cytokines and toll-like receptor ligands. In this video, we demonstrate flow cytometric comparisons of expression of two co-stimulatory molecules, CD86 and CD40, and the cytokine, IL-12, following overnight stimulation with CpG or mock treatment. After differentiation, DCs can be further manipulated for use as a vaccine vector or to generate antigen-specific immune responses by in vitro pulsing using peptides or proteins, or transduced using recombinant viral vectors. Download video file.^{(65M, mp4)}     

Chromosome analysis of bone marrow in mammals after treatment with isoniazid

Summary Cytogenetic investigations in bone marrow from animals treated with isoniazid (INH) were performed in seven different laboratories according to a standard protocol. The experiments were carried out in the Chinese hamster, the mouse, and the rat. In short-term studies INH was administered twice at an interval of 24 h in doses of 5, 25, and 125 mg/kg, and the animals were sacrificed 6, 12, 24, and 48 h after the second dose. In long-term studies doses of 25 and 125 mg/kg were administered thrice weekly for 12 weeks. As a rule, each group consisted of at least four animals, and 100 metaphases per animal were counted. Statistical analysis of the data showed that the incidence of chromosomal aberrations including gaps lay in the critical range for two groups in one laboratory and was significantly higher than in the control in three groups in another of the seven laboratories. From the results of both the short-term and the long-term studies in all laboratories, however, it may be concluded, that isoniazid does not induce gross chromosomal aberrations.     

Chromosome alterations in tissue culture cells ofAllium fistulosum

Heterochromatin behaviour and structural alterations in chromosomes of cells derived from callus culture ofAllium fistulosum have been studied.The diploid chromosome complement ofAllium fistulosum consists of 16 chromosomes with significant amount of heterochromatin mainly of telomeric nature. In eight collections of callus cells analysed, a high rate of numerical and structural chromosome abnormalities was observed. After 12 months in culture about 20% of metaphase chromosomes possessed distinct signs of mutational events.C-banded preparations revealed that many structural alterations involved regions of heterochromatin. Interchromosomal connections and chromatid fusions occurred at telomeric heterochromatin segments. Also formation of the end-to-end associations and polycentric chromosomes often took place without visible loss of telomeric heterochromatin.     

Liproprotein inhibitor of bone marrow cells in tumor-bearing rats.

The role of a plasma inhibitor of erythropoiesis is evaluated in rats with Walker-256 carcinoma (W-256). Plasma from tumor-bearing rats was treated by gel filtration chromatography (Sephadex G-150) and fractions were combined into four pools on the basis of mol. wt. Inhibitory activity was assayed by adding an aliquot of the plasma fractions to normal rat marrow cells which were cultured for 24 hr with and without erythropoietin. 59Fe-heme synthesis, [3H]thymidine DNA synthesis, and 14C-leucine protein synthesis were studied. The results indicated that cultures containing the high mol. wt. pool (greater than 400,000 daltons) had significantly decreased heme, DNA and protein synthesis. This inhibitor also diminished the response to erythropoietin in polycythemic mice. The lower mol. wt. pool stimulated heme synthesis in vitro. To identify the inhibitor further, plasma lipoprotein classes were isolated by density gradient ultracentrifugation. The very low density lipoprotein (VLDL) and chylomicron fractions markedly inhibited DNA, protein and heme synthesis. Low density and high density lipoprotein fractions were inactive. A lipoprotein inhibitor of erythropoiesis was also identified in cancerous ascitic fluid, and to a lesser degree, in normal rat plasma. We suggest that this VLDL inhibitor of marrow erythropoiesis is a contributing factor in the anaemia of cancer.     

Chromosome aberrations induced by monomeric acrylamide in bone marrow and germ cells of mice

Acrylamide induces chromatid exchanges and breaks with considerable frequency in spermatogonia of mice with long-term administration (3 weeks), though not, remarkably, with short-term administration (1–2 weeks). At 12 and 24 h after single injections with 50, 100 and 150 mg/kg acrylamide, evaluation of the cytogenetic effect is difficult in the spermatogonia because of an extreme reduction of mitotic cells. Aneuploid and polyploid cells increase with ti,e after treatment in both marrow and spermatogonial cells, while the aberration frequency shows no increase in marrow after both oral-administration and injection. Evidently the spermatogonia are thus rather more sensitive to acrylamide than marrow cells. On the other hand, the SCE frequency is at the control level in treated subjects in marrow and spermatogonia. Acrylamide induces chain quadrivalents, ring quadrivalents, fragments and univalents which are particularly evident in primary spermatocytes in both oral administration and injection, though it is questionable whether these structural changes deal with spermatogonia, or otherwise with the S-phase primary spermatocytes. There is a possibility that the aberrant cells thus produced can develop into spermatozoa carrying a certain type of reciprocal translocation which leads to semi-sterile progeny. In relation to the above problem detailed investigations into this type of rearrangement in primary spermatocytes are needed.     

Rescue of myeloid lineage-committed preprogenitor cells from cytomegalovirus-infected bone marrow stroma.

The effect of murine cytomegalovirus on myelopoiesis was studied in long-term bone marrow culture to find an in vitro correlate for the lethal virus interference with bone marrow reconstitution (W. Mutter, M. J. Reddehase, F. W. Busch, H.-J. Bühring, and U. H. Koszinowski, J. Exp. Med. 167:1645-1658, 1988). The in vitro generation of granulocyte-monocyte progenitors (CFU-GM) discontinued after infection of the stromal cell layer, whereas the proliferation and differentiation of CFU-GM to granulocyte-monocyte colonies remained unaffected. A protocol was established to probe the functional integrity of earlier hematopoietic cells. Pre-CFU-GM (the progenitors of the CFU-GM) could be recovered from an infected bone marrow donor culture by transfer onto an inductive recipient stromal cell layer. Thus, at least in vitro, infection of bone marrow stroma appears to be the only cause of the defect in myelopoiesis.     

Direct versus cultured preparation of bone marrow cells from 22 patients with acute myeloid leukemia

Summary The 24-h culture of bone marrow from patients with acute myeloblastic leukemia (AML) and acute promyelocytic leukemia (APL) gave more analyzable metaphase cells and improved chromosome morphology compared with direct preparations. Culture increased the proportion of cytogenetically abnormal cells, and in six bone marrows where the direct preparation failed, a result was obtained from the cultured preparation. The culture of bone marrow from patients with APL led to the detection of clones carrying the t(15;17) that were not found in direct preparations. Such sequestered clones were not found in AML and acute myelomonocytic leukemia (AMMoL). Cultured preparations were no better than direct preparations from AMMoL.     

Selective lesions of acinar pancreatic cells in rats poisoned with abrin

Rats poisoned with abrin (2.5 micrograms/100 g body weight) died within 36 h with severe necrosis of acinar pancreatic cells. Incorporation in vivo of labelled amino acids into pancreatic protein was greatly impaired 6 h after poisoning. Microsomes isolated from the pancreas of poisoned rats at 6 h had a reduced capacity for protein synthesis in vitro. Incorporation in vivo of orotic acid into pancreatic RNA was decreased 12 h after poisoning.     

Effects of aluminum on erythroidal cells in bone marrow in rats

The study has been carried out on Wistar rats. The aim of the present study was to trace the effect of aluminum on erythroidal cells in bone marrow in rats. The number of proerythroblast after 10 days of experiment with aluminum slowly decreased up to 80 days of experiments. However, the number of basophilic erythroblasts after 10 days insignificantly increased but after 20 days gradually decreased up to 80 days of experiments. The bone marrow polichromatic erythroblasts after 10 days of experiment slightly decreased, however after 20, 40 and 80 days of experiments the values decreased significantly. The quality of orthochromatic erythroblasts after 10 days of experiments dropped and after 20, 40 and 80 days of experiments significantly decreased compared to the control value. Aluminum also brings about histological changes in the bone marrow. The statistical significant reductions of hemoglobin and hematocrit levels were found in the aluminum exposed rats.     

Retinoic acid induces granulocytic differentiation of myeloid progenitors in congenital agranulocytosis bone marrow cells

Congenital agranulocytosis is a rare fatal infantile disease characterised by recurrent bacterial infections, persistent absence of neutrophils and maturation arrest at the promyelocyte/myelocyte stage. The effectiveness of retinoic acid in inducing differentiation of congenital agranulocytosis marrow myeloid progenitor cells was studied. Non-adherent mononuclear marrow cells were treated in an in vitro culture with retinoic acid at various concentrations from 1nM to 1 microM for seven days. Morphological and functional differentiation into mature granulocytes was induced by retinoic acid in a dose-response stimulation with a maximum response at a concentration of 1 microM. These results suggest a potential therapeutic role for retinoic acid in the treatment of congenital agranulocytosis.     

Absence of Chromosome Damage in the bone marrow of rats fed detergent actives for 90 days

Four detergent actives, sodium lauryl sulphate, DOBS 055, Dobanol 25 sulphate LCU and Dobanol 25 sulphate HCB, were fed to rats in the diet for 90 days at the maximum tolerated dose, 1.13 percent active ingredient in each case. Sodium lauryl sulphate and DOBS 055 were also fed at half this concentration. Chromosome preparations were made from the bone marrow and scored for the presence of rearrangements, chromatid gaps and breaks and isochromatid gaps and breaks. The four detergent actives were found to have no effect on the chromosomes of rat bone marrow cells.     

Analysis of chromosome aberrations in bone marrow cells of rats after treatment with isoniazid

Summary 100 mg INH/kg body weight was administered to male Wistar rats five times, at 24 h intervals, either intraperitoneally or intravenously. In both experimental series the yield of chromosome aberrations in bone marrow cells was not increased as compared to controls.     

运动诱导的低铁状态大鼠骨髓细胞铁摄入的变化

本文观察了运动性低铁状态大鼠骨髓细胞转铁蛋白 (Tf)结合铁和非Tf结合铁摄入的变化。大鼠随机分为 6个月的运动组 (EG)和对照组 (SG)。SG平均每个幼红细胞Tf受体数为 890 15 0± 16 4849个 ,而在EG为 2 17536 0± 46 2 737个 (P <0 0 5 ) ,但受体的解离常数不受运动影响。EG中Tf的内吞平台和胞内铁聚积速度显著高于SG ,胞浆和胞内膜性成分中Tf结合铁和Fe(Ⅱ )摄入增加。EG的胞浆内Fe(Ⅱ )摄入的米氏常数值降低 ;细胞膜性成分中Fe(Ⅱ )摄入的最大速度增加。上述结果表明 ,运动不仅通过增加Tf受体的表达促进Tf结合铁的内吞 ,而且增强非Tf结合铁的内吞途径。尽管这些变化的机制尚不清楚 ,但它们有利于运动时血红素的合成