昆明山海棠水抽提物诱发小鼠骨髓细胞非整倍体的研究

以C—有丝分裂效应、微核及染色体结构畸变分析,综合评估了中药昆明山海棠根部水抽提物(THH)在小鼠骨髓细胞中的非整倍体诱发能力。结果表明,THH具有类似于秋水仙素的非整倍体诱发效应:(1)导致骨髓细胞有丝分裂指数、C—有丝分裂细胞频率显著升高,有丝分裂后期频率显著下降;(2)于小鼠骨髓嗜多染红细胞中诱发微核却无染色体断裂效应。提示THH为一种哺乳动物体细胞非整倍体诱发剂。     

46. Micronuclei induced by chronical treatment of SO2 inhalation in mouse bone marrow cells

In the chronical experiment of treating with sulfur dioxide(SO2) inhalation, Micronuclei(MN) frequencies in the polychromatophilic erythroblasts(PCE) of mouse bone marrow and the frequencies of cells with MN were significantly increased in dose-dependent manner. There is a significant difference between the male and the female animals. The results also showed that SO2 inhibited urethone-induced MN formation, it is a antagonistic joint action to Urethone. These results furtherly confirm that SO2 inhalation is a clastogenetic and genotoxic agent to mammalian cells, and the combination roles of SO2 and other mutagens are complexity.     

Synergistic increase in chromosomal breakage within the euchromatin induced by an interaction of the benzene metabolites phenol and hydroquinone in mice

The hematopoietic and carcinogenic effects of benzene may resultfrom an interaction of various benzene metabolites. Followingthe co-administration of phenol and hydroquinone, a synergisticincrease in myelotoxicity and genotoxicity has been observedin the bone marrow of mice. To understand the mechanisms underlyingthese synergistic geno-toxic effects we have studied the originof micronuclei (MN) formed in bone marrow erythrocytes followingthe co-administration of these two metabolites. Phenol and hydroquinonewere administered to male CD-I mice by i.p. injection threetimes at 24 h intervals. The frequency of MN was evaluated inbone marrow cells harvested 24 h following the final dose. Amarked increase in MN was observed in mice co-administered phenoland hydroquinone, which was significantly greater than thatobserved with the individual metabolites. Labeling with theCREST antibody and multicolor fluorescence in situ hybridizationwith the mouse major and minor satellite probes indicated thatboth chromosomal loss and breakage had occurred. The major increasein MN induced by the phenol and hydroquinone combination originatedfrom breakage in the dichromatic region of the mouse chromosomes.The origin of MN in mice co-administered phenol and hydroquinonediffered substantially from that induced by hydroquinone alone,but was almost identical to that seen in MN from benzene-treatedmice. These results strongly support the hypothesis that interactiveeffects among benzene metabolites play an important role inthe genotoxic and carcinogenic effects of benzene.     

Induction of antigen specific cellular immunity by vaccination with peptides from MN/CA IX in renal cell carcinoma

We investigated the induction of the specific immunity for renal cell carcinomas (RCC) using MN/CA IX, a tumor-associated antigen frequently expressed in RCC. We have generated 9-mer peptide derived from MN/CA IX and examined the antigenicity as a vaccine to induce specific immunity for RCC. To use mouse syngeneic system, we transfected human MN/CA9 cDNA into RenCa and BALB-3T3 cells originally from BALB/c mouse, and established MN/CA IX expressing mouse cell lines, i.e., MN-RenCa and MN-3T3. The immunization of BALB/c mouse with MN-RenCa cells resulted in the induction of cytotoxic T lymphocytes (CTL) against MN/CA IX expressing cells and the CTL clone was established from bulked CTL. This CTL clone specifically lyzed MN-3T3 cells, but not parental cells. To identify the targeted epitope binding to H-2Kd antigen, three 9-mer peptides (A, B, C-peptide) of human MN/CA IX compatible with the H-2Kd as well as HLA-A24 binding motif was synthesized. The cloned CTL targeted the B-peptide pulsed BALB-3T3 cells as well as MN-3T3 cells. Furthermore, spleen cells from BALB/c mouse immunized with B-peptide reacted against MN-RenCa cells. These results suggest that the peptides derived from MN/CA IX containing HLA-A24 binding motif may be useful as a potent tumor vaccine for the treatment of human RCC, and in mouse models.     

Bcr-Abl-mediated suppression of normal hematopoiesis in leukemia

A variety of experimental evidence including findings in various mouse models indicates that the BCR-ABL oncogene is the cause of chronic myeloid leukemia (CML). Since normal hematopoietic cells in marrow and spleen are replaced with proliferating leukemic blasts, we determined whether this is an active process mediated by the leukemia cells. The lipocalin 24p3 was reported to be secreted by mouse hematopoietic cells deprived of IL-3, resulting in apoptosis induction in a variety of hematopoietic cells including bone marrow cells. Here, we show that BCR-ABL+ mouse hematopoietic cells induced persistent expression and secretion of 24p3. Importantly, BCR-ABL+ hematopoietic cells were resistant to the apoptotic effects of 24p3. The expression of the Bcr-Abl oncoprotein and its tyrosine kinase were required for induction of 24p3 expression. Co-culture studies showed that BCR-ABL+ cells induced apoptosis in BCR-ABL negative cells. Antisense 24p3/siRNA expression reduced the level of 24p3 protein in both BCR-ABL+ cells and in conditioned medium (CM) obtained from these cells. CM from BCR-ABL+ cells expressing antisense 24p3/siRNA had reduced apoptotic activity for target cells; 24p3 antibody also reduced the apoptotic activity of the CM. Leukemic mice induced by BCR-ABL+ cells expressing either antisense 24p3 or 24p3 siRNA had increased levels of normal hematopoiesis and reduced invasion of leukemia cells in marrow and spleen tissues. These findings indicate that suppression of normal hematopoiesis in BCR-ABL-induced leukemia is an active process involving secretion of the cell death-inducing factor 24p3 by mouse leukemia cells, raising the possibility that similar factors are involved in BCR-ABL+ CML.     

益母草对小鼠遗传物质损伤的影响

目的:观察中草药益母草对醋酸铅引起的小鼠遗传物质损伤的影响。方法：采用小鼠骨髓微核试验和小鼠精子畸形试验。结果：益母草本身不能诱发小鼠骨髓微核和小鼠精子畸形,但在与醋酸铅一同给药时,可使醋酸铅所诱发的小鼠骨髓微核和小鼠精子畸形频率明显降低。结论：益母草对小鼠遗传物质具有保护作用,即抗诱变作用。     

Production of a negative regulator of IL-3 by bone marrow cells in response to the supernatant of IL-3-producing STIL-3 leukemia cells.

When spleen cells of mice grafted with STIL-3 C5 cells, a leukemic T-cell line producing IL-3, are cultured in vitro, a high IL-3 activity is detectable in the culture supernatant. However, when bone marrow cells of the same grafted mice are cultured under similar conditions, hardly any IL-3 activity is detectable. To elucidate the mechanism of this difference, we examined whether the bone marrow cells either suppress IL-3 production by STIL-3 C5 cells or produce an IL-3 inhibitor. When STIL-3 C5 cells were cultured in the presence of normal bone marrow cells, the culture supernatant showed a significantly reduced IL-3 activity as assessed by growth stimulatory effects on IL-3-dependent DA-1 cells and mast cells. The conditioned medium (CM) did not inhibit the growth of IL-3-independent cell lines. Heat treatment of the CM resulted in a recovery of the IL-3 activity, indicating that the effect was mediated by a heat-labile inhibitor rather than by suppression of IL-3 production. CM of bone marrow cells alone did not inhibit the IL-3 activity. The inhibitor was produced by a stem cell-enriched fraction of the bone marrow, and not by fractions of T cells, granulocytes, or adherent cells including macrophages. Stimulation of the stem cell-enriched fraction of bone marrow with STIL-3 C5-CM also induced the production of the IL-3 inhibitor, which was recovered in a MW 50-100 kD fraction after ultrafiltration. These results suggest a possible presence of a feedback mechanism against the IL-3 effect on hemopoietic stem cells and progenitors in the bone marrow.     

B3HM细胞内活性物质对小鼠造血的作用

目的　观察B3HM细胞条件培养液 (B3HM CM )和细胞匀浆组分对小鼠造血细胞的作用。寻找其中可能存在的诱导造血细胞发生恶性转化的活性物质。方法　将B3HM CM加到小鼠骨髓长期培养体系中 ,定期计数细胞生长情况 ;用B3HM CM或经差速离心法分离的B3HM细胞匀浆组分注射经照射处理的小鼠 ,观察小鼠存活情况和造血细胞的变化。结果　B3HM CM在骨髓培养体系中促巨核细胞增殖 ,在小鼠体内促造血细胞增多。B3HM细胞匀浆第 3组分可诱发小鼠白血病。结论　B3HM细胞可能存在诱导造血细胞发生恶性转化的物质和细胞增殖刺激因子     

非霍奇金淋巴瘤患者血清CA125和VEGF水平与骨髓浸润的相关性研究

 目的　评价CA125和血管内皮生长因子（VEGF）作为预测非霍奇金淋巴瘤（NHL）患者骨髓浸润的血清学指标的价值。方法　97例经病理确诊的初治NHL患者,其中50例经骨髓活检或骨穿检查证实有骨髓浸润,46例骨髓正常者作为对照组。采用ELISA方法分别于化疗前检测血清CA125和VEGF水平。结果　97例NHL患者中,骨髓浸润者占51.5 %（50例）,骨髓正常者占45.5 %（47例）。有骨髓浸润组其血清CA125和VEGF水平明显高于无骨髓浸润组（P＜0.05）。VEGF水平与骨髓中肿瘤细胞所占百分比呈正相关（r＝0.498,P＝0.01）,CA125水平与其无明显相关。结论　NHL骨髓浸润者血清CA125和VEGF明显升高,而且VEGF水平与骨髓浸润程度呈正相关。     

7种中草药的抗诱变性试验

本文应用Ａｍｅｓ试验ＴＡ９８和ＴＡ１００菌株及小鼠骨髓微核试验对牡丹皮，牡丹草，潞党参，宣木瓜，菊花，石斛和新鲜石斛７种草药的水溶性提取液进行抗诱变性筛选，发现牡丹皮，潞党参和菊花在２个试验系统均有抗诱变活性提示这３种中药能抗基因突变又能抗染色体畸变；宣木瓜在Ａｍｅｓ试验中对２－ＡＦ诱导ＴＡ９８和ＴＡ１００有强抗诱变性；除宣木瓜外，余６种中药在微核试验中均具有显著抑制作用。     

Expression of MN/CA9 protein in Papanicolaou smears containing atypical glandular cells of undetermined significance is a diagnostic biomarker of cervical dysplasia and neoplasia

BACKGROUND: Despite the enormous impact that Papanicolaou (Pap) smear screening has had on the incidence of cervical carcinoma in developed countries, there is still an unacceptably high frequency of occurrence of this cancer. In part, this is due to human error associated with cytologic diagnoses of Pap smears. Also, the use of new sampling devices, such as the cytobrush, has increased the complexity of diagnosing benign and neoplastic cervical cytology. This is particularly apparent in the diagnosis of atypical glandular cells of undetermined significance (AGUS). Approximately 40% of AGUS diagnoses have a corresponding significant lesion at biopsy follow-up, and 60% do not. There is clearly a need for an adjunct to cytologic diagnosis that can readily identify AGUS smears that are diagnostic of significant lesions. The authors have identified the MN/CA9 antigen as a strong candidate for an adjunct biomarker. METHODS: A total of 245 Pap smears of all AGUS diagnostic categories with histologic confirmation were studied. The median age of the patients was 39 years. The Bethesda system classification (AGUS-favor reactive, AGUS-not otherwise specified, and AGUS-favor neoplastic) was used. All of the Pap smears were decolorized and immunostained with monoclonal antibody to MN/CA9 antigen by the immunoperoxidase technique. The results of MN/CA9 immunoreactivity were correlated with the histologic data in a semiblinded fashion. RESULTS: The follow-up biopsies showed that a high percentage (70%) of patients had low and high grade cervical intraepithelial neoplasia lesions, respectively (CIN I and CIN II or III). Clinically significant lesions-adenocarcinoma in situ/carcinoma (AIS/CA) and CIN II or III-were found in 50% of the cases. Among these, 11% were AIS/CA. In the three subcategories of AGUS diagnosis, the AGUS-not otherwise specified showed the broadest range of lesions in the follow-up biopsies. Three patterns of MN/CA9 immunoreactivity were observed in the Pap smears: 1) atypical cells, 2) normal endocervical cells only, and 3) all cells negative. All Pap smears that were MN/CA9 positive were histologically confirmed to be clinically significant lesions or CIN I; in addition, there were a very small number (n = 12) of cases of atypia. None of the benign lesions showed MN/CA9 expression in the corresponding Pap smears. Furthermore, the pattern of atypical cell immunostaining identified all cases with significant lesions (AIS/CA and CIN II or III) in the cervices. Conversely, the majority of CIN I cases (82%) and all cases of atypia showed positive immunostaining restricted to normal endocervical cells only. CONCLUSIONS: There is a clear association between MN/CA9 immunostaining of atypical cells and the presence of significant lesions in the cervix. Similarly, there is a clear association between lack of expression of MN/CA9 and the absence of cervical lesions. However, the screen does not allow discrimination between CIN I and atypia. The authors also found that, based on the combined patterns of morphology and immunostaining, they are able to discriminate between AIS and CIN II or III in AGUS Pap smear diagnoses. Thus, expression of the MN/CA9 antigen is indeed a discriminator of significant lesions in AGUS Pap smear diagnoses.     

Conditioned media from mouse osteosarcoma cells promote MC3T3-E1 cell proliferation using JAKs and PI3-K/Akt signal crosstalk

The maintenance of bone mass requires a strict balance between bone formation by osteoblasts and bone resorption by osteoclasts. In tumoral bone environment, tumor cells frequently disturb this balance by interaction with bone cells to create a favorable site for tumor growth, and promote pathological bone changes. Thus, elucidation of the mechanisms underlying interaction between tumor cells and bone cells might eventually lead to a more rational strategy for therapeutic intervention for bone tumors and better understanding of bone biology. In the present study, the effects of mouse osteosarcoma cells on mouse preosteoblastic cells were determined by assessment of cell viability, osteoblastic differentiation and signal transduction pathways. MOS-J/POS-1 conditioned media (CM) significantly induced MC3T3-E1 cell proliferation in a dose-dependent manner and reduced both alkaline phosphatase activity and mineralized nodule formation. Piceatannol, AG490, LY294002 and rapamycin significantly abrogated this up-regulated cell proliferation; however, UO126 and STAT3 inhibitor peptide did not affect this up-regulated cell proliferation. MOS-J/POS-1 CM activated ERK 1/2, STAT3 and Akt signal transduction pathways; however, pro-proliferating signal induced by MOS-J/POS-1 CM was transmitted via Akt not ERK 1/2 and STAT3 pathways. Furthermore, Western blot analyses clearly revealed novel signal crosstalk between JAKs and PI3-K/Akt in osteoblastic cells. The specific factor(s) involved in MOS-J/POS-1 CM-induced MC3T3-E1 cell proliferation that use JAKs/PI3-K/Akt/mTOR pathway remain(s) to be determined. Determination of the specific factor(s) responsible for JAKs and PI3-K/Akt signal crosstalk that results in up-regulated preosteoblast proliferation will offer new insight into the pathology of osteosarcoma as well as other bone-related diseases.     

A cancer of unknown primary site with diffuse metastasis to the bone marrow treated effectively with FAM combination chemotherapy

A patient with cancer of unknown primary site suffering from diffuse bone marrow metastasis and DIC, was treated with FAM (5-fluorouracil, adriamycin and mitomycin C) combination chemotherapy. She was a 58-year-old housewife. Bone marrow biopsy revealed that her marrow tissue was completely replaced by cancer cells, and bone scintigraphy showed diffuse bone marrow metastasis in all the vertebrae, sternum, pelvic bones and skull. After 5 months administration of 3 courses of FAM therapy, the cancer cells were completely eradicated in the bone marrow upon biopsy taken at almost the same position as the previous one. The values of CEA, CA 19-9 and CA 125 were normalized, suggesting that this therapy was very effective.     

海藻多糖的抗突变性研究

目的与方法 :本文选用小鼠骨髓嗜多染红细胞微核试验和小鼠精子畸形试验 ,对海藻多糖的抗突变作用进行研究。 结果 :海藻多糖对环磷酰胺诱发的微核率和精子畸形率有着显著的抑制作用 (P <0 .0 1)。结论 :提示海藻多糖对由环磷酰胺引起的体细胞和生殖细胞的基因突变有着明显的拮抗作用。     

Chromosome aberrations induced by 7,12-dimethylbenz[a]-anthracene in bone marrow cells of spontaneously hypertensive rats (SHR) and control Wistar Kyoto (WKY) rats: time course and site specificity

The frequency of chromosome aberrations (CA) induced by iv injection of 7,12-dimethylbenz[a]anthracene [(DMBA) CAS: 57-97-6] was examined with the use of control Wistar Kyoto (WKY) rats and spontaneously hypertensive rats (SHR). SHR appear to be analogous to hypertensive humans, since evidence has been reported of a difference in tumor susceptibility between SHR and WKY strains. Injection of DMBA evoked CA in bone marrow cells of both SHR and WKY rats. The frequency of CA in SHR was significantly higher than in WKY rats at 12 and 18 hours after DMBA treatment. With regard to the relative frequencies of aberrations in SHR and WKY rats, chromosomes 1 and 2 were more susceptible to DMBA-induced CA. The proportion of breaks that occur in the "hot spot" at 6 hours after DMBA treatment was distributed nonrandomly at a site approximately 40% of the total length from the centromere in chromosome 1 and at lengths of approximately 30 and 55% in chromosome 2. The specific distribution of CA seemed to be caused by a cell cycle-specific effect. These results, confirming nonrandom CA induced by DMBA in bone marrow cells of WKY rats, are similar to previous reports in which Long-Evans rats were used. Thus they indicate that a common mode of action by DMBA at the chromosome level in rat bone marrow cells was observed in different strains and that SHR were more sensitive than WKY rats to CA induced by DMBA.     

d-筒箭毒碱对V79细胞和小鼠骨髓细胞有丝分裂的影响

目的：探讨烟碱型乙酰胆碱受体阻断剂ｄ－筒箭毒碱对Ｖ７９细胞和小鼠骨髓细胞有丝分裂的影响。方法：在离体情况下以受试物处理中国仓鼠Ｖ７９细胞，通过分析Ｖ７９细胞晚末期和早Ｇ１期细胞核与细胞质构型，双核细胞频率，探讨受试物对细胞质质裂的影响；研究同时探讨了活体情况下，该化合物对小鼠骨髓细胞的Ｃ－有丝分裂（Ｃ－Ｍ）效应，分析其对哺乳动物体细胞有丝分裂的影响。结果：ｄ－筒箭毒碱能使Ｖ７９细胞的晚末期一早Ｇ１     

Production and analysis of HH 10 monoclonal antibodies reactive to immature hematopoietic cells and their use for monitoring acute leukemia cells

A clone of murine monoclonal antibody HH 10 (IgM) was raised by fusing NS-1 myeloma cells and spleen cells of a mouse hyperimmunized with acute promyelocytic leukemia cells. Serological analysis by means of immune adherence assays showed that HH 10 reacts with immature hematopoietic cells including thymocytes and myeloid precursor cells (defined as colony-forming units in culture assays). The antibodies were not reactive to either peripheral blood cells or bone marrow cells obtained from normal individuals. Lymphoid blasts induced with phytohemagglutinin, pokeweed mitogen, or concanavalin A were also non-reactive, and all non-hematopoietic cultured cells examined were also negative. The antibodies were, however, reactive to leukemia cells in 67 cases out of 91 cases (74%) of acute leukemia. In patients with acute lymphoblastic leukemia, 43 out of 52 cases (83%) were positive and, in particular, all of 12 T-cell-type acute leukemias were reactive to HH 10. Comparison of percent HH 10 positive cells in the bone marrow of patients with acute leukemia with the results of cytological studies showed a good correlation. Analysis of sequentially collected bone marrow cells of acute leukemia patients using HH 10 revealed its usefulness for monitoring leukemia cells.     

Detection of neuroblastoma cells in bone marrow using GD2 specific monoclonal antibodies

Three murine monoclonal antibodies (Mab) against a cell surface antigen, the disialoganglioside GD2, were investigated for detecting bone marrow metastasis in patients with neuroblastoma (NB) by indirect immunofluorescence (IF). As few as 0.01% NB cells could be detected. No Mab reactivity was found in 60 non-NB marrows. Thirty-five marrows from patients with stage III and stage IV NB at diagnosis were examined during the course of their disease. Tumor involvement was found in 74% by Mab, 55% by the two-layer soft agar clonogenic assay (CA), and 27% by conventional histochemical stains. All marrows containing NB histologically were positive for tumor by Mab; 71% were also positive by CA. Of histologically negative marrows, 63% were Mab positive, the majority (78%) of which were also positive by CA. All Mab nonreactive samples were negative histologically and by CA. We conclude that these antibodies are highly sensitive and specific in detecting NB metastasis in bone marrow.