Effect of SCG, 1,3-beta-D-glucan from Sparassis crispa on the hematopoietic response in cyclophosphamide induced leukopenic mice

Sparassis crispa Fr. is an edible mushroom recently cultivable in Japan. It contains a remarkably high content of 6-branched 1,3-beta-D-glucan showing antitumor activity. Using ion-exchange chromatography, a purified beta-glucan preparation, SCG, was prepared. In this study, we examined the hematopoietic response by SCG in cyclophosphamide (CY)-induced leukopenic mice. SCG enhanced the hematopoietic response in CY induced leukopenic mice by intraperitoneal routes over a wide range of concentrations. SCG enhanced the hematopoietic response in CY-treated mice by prior or post administration. Analyzing the leukocyte population by flow cytometry, monocytes and granulocytes in the peritoneal cavity, liver, spleen and bone marrow (BM) recovered faster than in the control group. The ratio of natural killer cells and gammadelta T cells in the liver, spleen and peritoneal cavity was also increased. In contrast, CD4+ CD8+ cells in the thymus were temporarily significantly decreased by the administration of SCG. Interleukin-6 (IL-6) production of CY+SCG-treated peritoneal exdated cells (PECs), spleen cells and bone marrow cells (BMCs) were higher than that of the CY-treated group. By in vitro culture of CY-treated PEC and spleen cells, IL-6 production was enhanced by the addition of SCG. These facts suggested the possibility that IL-6 might be a key cytokine for the enhanced hematopoietic response by SCG.     

Protective activities of a Chinese medicine, hochu-ekki-to, to impairment of hematopoietic organs and to microbial infection

Effect of Hochu-ekki-to (HET) on the number of peripheral leukocytes (PL) and their functions in cyclophosphamide (CY)-treated or gamma ray-irradiated mice was investigated. By treatment of mice with anticancer agent CY or gamma ray irradiation, unfavorable side effects usually occurred to impair hematopoietic organs, causing bone marrow disorder. However, it was significantly protected by oral administration of HET (1 g/kg/d) to CY-treated or gamma ray-irradiated mice. The numbers of neutrophils and monocytes in PL were restored to the normal level, and colony-stimulating factor (CSF) was induced in the sera of mice by HET in a dose-dependent manner. The induction of serum CSF reached a peak at 3h after HET administration. Colony-forming unit of bone marrow cells in the spleen adoptively transferred into syngeneic mice, that is defined as CFU-S, was extremely reduced by CY-treatment. However, when HET was orally administered, CFU-S of CY-treated mice was markedly stimulated, suggesting that bone marrow cells were reactivated for further proliferation and mobilization. HET enhanced other leukocyte functions in CY-treated mice; i.e., superoxide production of neutrophils and phagocytic activity of macrophages. Thus, oral administration of HET to CY-treated mice enforced protection against Pseudomonas aeruginosa infection.     

儿茶素对骨髓细胞周期及造血生长因子基因表达的作用儿茶素对骨髓细胞周期及造血生长因子基因表达的作用

目的探讨中药鸡血藤Spatholobus suberectus Dunn活性成分儿茶素对小鼠骨髓细胞增殖周期及其脾细胞内造血生长因子IL-6和GM-CSF mRNA表达的影响,以初步阐明其造血调控作用机理。方法用流式细胞仪检测技术观察儿茶素对正常及骨髓抑制小鼠骨髓细胞增殖周期的影响,同时采用RT-PCR技术,检测在儿茶素刺激条件下,小鼠脾细胞内IL-6和GM-CSF mRNA表达的变化。结果儿茶素可促使正常及骨髓抑制小鼠骨髓细胞G₀/G₁期细胞比例下降,S+G₂/M期细胞比例增加,并可使正常及骨髓抑制小鼠脾细胞内IL-6 mRNA和GM-CSF mRNA表达显著上调。结论儿茶素可通过诱导脾细胞内IL-6 mRNA和GM-CSF mRNA的表达,促进正常小鼠骨髓细胞进入增殖周期,并促使骨髓抑制小鼠的骨髓细胞跳出“G₁期阻滞”,进入细胞增殖周期,从而加速造血干/祖细胞的增殖和分化。     

Stimulation of hematopoiesis in untreated and cyclophosphamide treated mice by the inhibition of prostaglandin synthesis

Indomethacin (IN) was administered to untreated or to cyclophosphamide (CY) treated C57B1/6 mice to study the roles of prostaglandins in regulating hematopoiesis. The following hematopoietic parameters were quantitated: peripheral blood leukocyte (PBL) count; total nucleated cells per spleen; total nucleated cells per femur; and spleen weight. Assays were performed in vitro to measure the number of colony forming units (CFU) present in the bone marrow and spleen. Untreated mice administered IN had a transient rise in their PBL count. These animals also developed splenomegaly and had an increased number of nucleated cells in their spleen. All CY treated mice had a marked decrease in PBL count, spleen cellularity, bone marrow cellularity, and spleen size during the first 5 days after CY treatment. These observations were followed by hematopoietic recovery over the next 10 days. Cyclophosphamide treated mice exhibited a more rapid hematopoietic recovery when treated with IN than without IN treatment. Analysis of the CFU capacity of bone marrow and spleen cells in soft agar showed a larger number of CFU in the bone marrow and spleen of IN treated mice or of CY/IN treated mice than in animals not receiving IN. These results indicate that prostaglandins are involved in the regulation of hematopoiesis in untreated mice and that prostaglandins may limit the hematopoietic recovery of CY treated mice.     

Development of the susceptibility to oral tolerance induction in infant mice administered a herbal drug, Hochu-ekki-to (Bu-Zhong-Yi-Qi-Tang)

The susceptibility to oral tolerance in post-neonatal infant mice and the effect of a herbal drug, Hochu-ekki-to (HOT), on the susceptibility were investigated. To induce oral tolerance induction, infant and adult mice at 4 and 8 weeks of age, respectively, were orally administered a single high dose of OVA before an intraperitoneal immunization with OVA adsorbed on aluminum hydroxide. HOT (1000 mg/kg) was administered orally for 7 days before the induction. HOT significantly decreased the serum levels of OVA-specific IgE and IgG1 and the antigen-specific proliferation of spleen cells in infant mice, both of which were greatly enhanced compared to in adult mice. HOT increased the number of both CD4+ T cells and antigen-presenting cells expressing MHC class II as well as costimulatory molecules (CD40, CD80 and/or CD86) in the Peyer's patch (PP) of infant mice, which had fewer cells than adult mice. In the PP, moreover, HOT augmented the IL-12p40 mRNA expression and spontaneous or CD40-stimulated IL-12 production, and increased the number of CD4+ cells expressing CD40 ligand, which is up regulated by IL-12. These results suggest that HOT increases the number and improves the function of PP cells that are fully susceptible to the induction of oral tolerance.     

Effects of neopterin on the hematopoietic microenvironment of senescence-accelerated mice (SAM)

The pteridine neopterin (NP) is produced by monocytes and is known to be a useful marker of immunological activation, although, it remains elusive whether neopterin itself exhibits biological functions. Recently, we found that NP stimulates hematopoietic cell proliferation and differentiation by activating bone marrow stromal cell function. In order to elucidate the biological effect of NP on stromal cells, its effects on hematopoiesis was determined in the mouse model of age-related stromal impairment, senescence-accelerated mice (SAMs). An intraperitoneal administration of NP increased the number of peripheral leukocytes and CFU-GM in the bone marrow and spleen of young SAMs, however, no increase of CFU-GM in old SAMs (stromal impairment) was observed when compared with young SAMs. NP also increased the CFU-GM colony formation of bone marrow and spleen cells from young SAMs in a soft agar culture system, but it did not enhance CFU-GM colony formation of cells from old SAMs cultured in this system. Treatment with NP induced the production of hematopoietic stimulating factors, including IL-6 and GM-CSF, by bone marrow stromal cells from young SAMs but stromal cells from old SAMs did not respond to NP stimulation. Further studies will be required to clarify the mechanism by which NP stimulates the production of hematopoietic growth factors from stromal cells, the results of this study indicate that NP is a potent hematopoietic regulatory factor by activating stromal cell function(s).     

仿刺参糖胺聚糖对骨髓抑制贫血小鼠外周血及骨髓细胞周期的影响△

目的：观察仿刺参糖胺聚糖(HGAG)对环磷酰胺诱导的骨髓抑制贫血小鼠外周血及骨髓细胞周期的影响。方法：将40只雄性昆明种小鼠随机分为正常组、模型组、HGAG治疗组(1,5,15mg.kg_-1),每组8只。除正常组外其余各组腹腔注射环磷酰胺100mg.kg_-1制备贫血小鼠模型,每天1次连续3天,并于造模当日起,HGAG给药组腹腔注射不同浓度的HGAG,正常组和模型组腹腔注射生理盐水,每天1次连续7天。用全自动血细胞分析仪、白细胞计数法、流式细胞术分别检测HGAG对贫血小鼠外周血、骨髓有核细胞数和骨髓细胞周期的影响。结果：HGAG低、中、高剂量组能显著升高外周血WBC和HGB,且高剂量组还能明显升高外周血RBC和PLT。HGAG三个剂量组能显著增加小鼠骨髓有核细胞数和脾脏指数,而且能够促进骨髓G₀/G₁期细胞向S期及S期细胞向G₂/M期细胞的转化,显著升高增殖指数。结论：HGAG能够解除细胞周期阻滞,促进骨髓造血细胞的增殖,增加外周血细胞、骨髓有核细胞的数量和升高脾脏指数,从而促进骨髓抑制贫血小鼠造血功能的恢复     

Positive effects of indomethacin on restoration of splenic nucleated cell populations in mice given sublethal irradiation

Splenic cellularity during the recovery phase after 400-R irradiation was evaluated in mice, in which the level of prostaglandin was regulated by indomethacin and exogenous prostaglandin E2, following sublethal irradiation. Two weeks after irradiation, administration to these mice of indomethacin, an inhibitor of prostaglandin synthesis, augmented the recovery of all nucleated spleen cell populations, whereas the thymus was drastically depopulated. This treatment had little effect upon the total number of bone marrow cells but inversed the ratio of PNA+/PNA- cells. Cell transfer experiments using heavily irradiated mice as recipients showed that the stem cell proliferation was positively affected by indomethacin treatment in the bone marrow rather than in the spleen. These results suggest that cell migration from primary lymphoid organs, particularly from the bone marrow to the spleen, is regulated by a prostaglandin-mediated system and that a prostaglandin E2 synthesis inhibition would have a positive effect on the restoration of peripheral nucleated cells following irradiation.     

Lentinan has a stimulatory effect on innate and adaptive immunity against murine Listeria monocytogenes infection

Lentinan, a (1-3)-beta glucan from Lentinus edodes, is licensed as an immunostimulatory drug. We tested the effect of lentinan in the well-established model system of the murine Listeria monocytogenes infection. Pre-treatment of bone marrow macrophages and dendritic cells with lentinan resulted in increased production of TNF-alpha and IL-12 after L. monocytogenes infection in vitro. After lentinan treatment bone marrow macrophages showed increased NO-production and enhanced cytotoxic activity against L. monocytogenes. Pre-treatment of mice with lentinan resulted in increased concentrations of TNF-alpha, IL-12 and IFN-gamma and also an increased number of L. monocytogenes specific CD8 T cells in the spleen. The bacterial burden in spleen and liver of mice was significantly reduced during primary and secondary Listeria infection after lentinan pre-treatment of mice. In summary these results show that lentinan enhances the protective CD8 T-cell response against L. monocytogenes probably by a mechanism that involves the IL-12-mediated augmentation of the specific antilisterial CD8 T-cell response.     

螺旋藻多糖对小鼠移植性肿瘤化疗后造血恢复及相关细胞因子的影响

目的：研究螺旋藻多糖（polysaccharide from spirulina platensis,PSP)对小鼠移植性肿瘤化疗后造血恢复及相关细胞因子的影响。方法：通过右前肢皮下接种肿瘤细胞形成小鼠移植性实体瘤，给予PSP边疆灌胃10d,d4给予环磷酰胺（CTX）腹腔注射连续3d。于d11，分别了外周细胞，骨髓有核细胞及CFU－S计数，用紫外分光光度计检测骨髓DNA含量，用双抗体夹心ELISA法检测血清中IL-1,IL-3,GM-CSF,TNF含量。结果：CTX可造成明显骨髓抑制，而PSP则提升了外周血细胞，增加了骨髓中有核细胞计数和DNA含量以及促进了CFU－S形成。此外，PSP还增加了血清中IL－1，IL－3，GM－CSF，TNF含量。结论：PSP可能通过促进内源性细胞因子的分泌来实现其促进小鼠移植性肿瘤化疗损伤后的造血恢复。     

The influence of recombinant human granulocyte colony-stimulating factor on granulopoiesis in mice recovering from cyclophosphamide treatment

Recombinant human granulocyte colony-stimulating factor (rhG-CSF) was evaluated for its effects on granulopoiesis recovery in mice pretreated with cyclophosphamide. The cytotoxic agent was administered on days 0 and 7. rhG-CSF therapy (injected from days 8 to 28) accelerated the recovery and maintained an increased level of peripheral blood neutrophils. Marrow granuloid precursors depression, due the second dose of cyclophosphamide, was prevented by rhG-CSF, but this hemopoietic growth factor was unable to increase these cells in a similar way to that observed in normal mice. This suggests a decreased granuloid marrow proliferation capacity in cyclophosphamide treated mice. In contrast, in the spleen, rhG-CSF highly increased granuloid precursors. However, the contribution of spleen to granuloid recovery was scarce. Mice receiving rhG-CSF after cyclophosphamide demonstrated a biphasic recovery pattern in bone marrow GM-CFU population. A first rebound of GM-CFUs was followed by a nadir and then a second recovery where GM-CFU progenitor cells were significantly increased was observed. On the other hand, rhG-CSF therapy highly increased the spleen GM-CFU numbers at days 24 to 28. Although rhG-CSF increased splenic granulopoiesis, this result shows that the bulk of granulopoiesis recovery due to rhG-CSF therapy in cyclophosphamide pretreated mice occurred in the marrow.     

Protective effect of a traditional Japanese medicine Hochu-ekki-to (Chinese name: Bu-zhong-yi-qi-tang), on the susceptibility against Listeria monocytogenes in infant mice

In this study, the effect of traditional Japanese (Chinese) medicine, Hochu-ekki-to, HOT (Chinese name: Bu-zhong-yi-qi-tang), on the susceptibility against Listeria monocytogenes in postneonatal infant mice was examined. Numbers of bacteria in infant mice (infected at 4 weeks of age) were significantly higher than those in adult mice (infected at 8 weeks of age) on day 3 (non-specific resistance phase) and day 5 (specific resistance phase) after infection. Oral administration of 1,000 mg/kg of HOT for 7 days to infant mice reduced bacterial numbers in the liver and spleen at 5 days after the infection. The amount of IFN-gamma and the number of IFN-gamma-producing CD4+ T cells were lower in infant mice than adult mice but those in infant mice enhanced by HOT treatment. HOT also enhanced the antigen-presenting function along with the expression of MHC class II in infant macrophages induced by heat-killed L. monocytogenes. Further, HOT enhanced the IFN-gamma production from infant CD4+ T cells independent of the deficiency in the antigen-presenting function. These findings suggest that HOT induced simultaneously functional maturation of both infant antigen-presenting cells and T cells, and consequently developed an anti-listerial Th1 response.     

叔丁基对苯二酚上调Nrf2调控的基因表达并减轻环磷酰胺导致的小鼠血液毒性

血液毒性（或称骨髓抑制）是癌症化疗最常见的剂量限制毒副作用,而转录因子Nrf2控制着包括骨髓在内的许多组织对化学刺激的敏感性。本文研究了叔丁基对苯二酚（tBHQ）对小鼠外周血细胞中Nrf2调控的基因表达和环磷酰胺（CTX）导致的血液毒性的影响。CTX处理导致了小鼠外周血有核细胞的凋亡和白细胞减少,伴随着骨髓造血细胞的动员。tBHQ处理则可以在体外和体内激活RAW264.7小鼠巨噬细胞和外周血细胞中Nrf2信号和下游基因如血红素氧化酶1和谷胺酸半胱氨酸连接酶催化亚基等的表达同时,tBHQ预处理可以减轻CTX导致的小鼠外周血有核细胞的凋亡和白细胞减少,说明Nrf2可能在减轻CTX血液毒性中发挥作用。本文的研究有助于加深对化疗所致血液毒性的了解,并提示Nrf2可能作为减轻化疗毒副作用的化疗保护剂的药物靶标。     

参芪益肺糖浆对大鼠免疫和造血功能的影响

目的观察参芪益肺糖浆对大鼠免疫和造血功能的影响。方法用注射环磷酰胺的方法建立大鼠白细胞减少症模型。90只大鼠随机分为正常组、模型组、鲨肝醇组和参芪益肺糖浆大、中、小剂量组,观察外周血白细胞、骨髓有核细胞数、淋巴细胞转化指数以及白细胞介素-6（IL-6）指标的变化。结果与模型组比较,参芪益肺糖浆各剂量组均可促进大鼠外周血白细胞、骨髓有核细胞数、淋巴细胞转化指数、IL-6活性指标的恢复及升高（P〈0.05或P〈0.01）。结论参芪益肺糖浆有显著改善机体免疫功能和刺激骨髓造血功能的作用。     

淫羊藿多糖对免疫功能低下小鼠免疫功能的影响

采用环磷酰胺所致免疫低下小鼠及荷瘤 (S180 )小鼠观察了淫羊藿多糖 (EPS)对免疫功能的影响 .结果表明 :腹腔注射环磷酰胺 80mg/kg ,可以使正常小鼠的胸腺指数、血清溶血素水平、空斑形成细胞溶血能力和外周血中WBC总数下降 .腹腔注射EPS 5 0、2 5mg/kg× 7,可使脾指数上升 ,但对胸腺指数无明显影响 ;5 0、2 5、12 5mg/kg× 7,使下降的溶血素值及空斑形成细胞的溶血能力回升 ;2 5、12 5mg/kg× 7,使下降的白细胞总数上升 .小鼠荷瘤后免疫功能低下 ,表现为胸腺指数下降、血清溶血素水平、空斑形成细胞的溶血能力及迟发性超敏反应能力降低 .皮下注射EPS5 0mg/kg× 8,可对抗荷瘤引起的胸腺指数下降 ;5 0、2 5mg/kg使荷瘤小鼠的脾指数升高 ;5 0、2 5mg/kg× 8,可以使荷瘤小鼠的血清溶血素、空斑形成细胞的溶血能力及迟发性超敏反应能力有所提高 .     

芪术口服液对环磷酰胺所致小鼠骨髓造血功能损伤的保护作用

目的:观察芪术口服液对环磷酰胺所致小鼠骨髓造血功能损伤的保护作用.方法:对小鼠腹腔注射100 mg/kg环磷酰胺后,观察芪术口服液对小鼠外周血象、骨髓有核细胞和胸腺、脾脏指数的影响.结果:芪术口服液能升高受损小鼠白细胞、红细胞、血小板、血红蛋白和骨髓有核细胞数量,并能使小鼠胸腺、脾脏指数升高.结论:芪术口服液对环磷酰胺引起的小鼠骨髓造血功能损伤有一定的保护作用.     

白芍水提物及芍药苷改善环磷酰胺致白细胞减少的对比研究

目的对比观察白芍水提物及芍药苷对环磷酰胺(CTX)致小鼠白细胞减少症的影响。方法给小鼠连续灌胃10d,第6和第7天腹腔注射高剂量CTX造成外周血白细胞减少。比较各给药组和模型组小鼠体质量、外周血白细胞数量、股骨中骨髓有核细胞数及肝、脾、胸腺造血免疫器官脏器系数。结果连续2d腹腔注射CTX 100mg.kg-1,可见小鼠外周血白细胞数、骨髓有核细胞数均显著减少(P<0.01),肝脏、脾脏和胸腺系数均明显降低(P<0.01)。白芍水提物及芍药苷均使小鼠外周血白细胞数、骨髓有核细胞数显著增多,脾脏系数明显升高。用药期间各组小鼠体质量无明显差异。结论白芍水提物及芍药苷对CTX所致小鼠白细胞减少和骨髓细胞减少均有升高作用,可提高CTX减少的小鼠脾脏系数。     

17aα-D-高炔雌二醇-3-乙酯对化疗损伤小鼠的保护作用

目的:观察17aα-D-高炔雌二醇-3-乙酯对环磷酰胺所致不同品系小鼠损伤的保护作用。方法:对IRM-1、IRM-2小鼠连续5d每只腹腔注射低、中、高剂量(100、150、200μg/0.2ml)17aα-D-高炔雌二醇-3-乙酯,从第3天同时给予环磷酰胺100mg/kg,连续3d;阳性对照组每只腹腔注射茜草双酯2.5mg/0.2ml,连续5d给药;对照组腹腔注射等体积茶油。观察小鼠外周血白细胞、骨髓有核细胞数、内源性脾结节形成、脾指数及胸腺指数的变化。结果:17aα-D-高炔雌二醇-3-乙酯可明显减轻化疗对两品系小鼠免疫功能的抑制,使外周血白细胞、骨髓有核细胞数和脾结节显著高于对照组(P<0.05)。与阳性对照茜草双酯组比较,17aα-D-高炔雌二醇-3-乙酯中、高剂量组小鼠白细胞数量显著升高(P<0.05)。结论:17aα-D-高炔雌二醇-3-乙酯具有抗化疗损伤及刺激骨髓造血功能的作用。     

参附注射液治疗再生障碍性贫血模型小鼠的实验研究

目的观察参附注射液对AA模型小鼠的治疗作用。方法健康雄性昆明种小白鼠60只,随机分为盐水对照组（N）、模型组（M）、参附组（S）和激素组（B）。除对照组外,其它各组采用60Co-γ射线＋环磷酰胺＋氯霉素复合方法复制AA模型。之后分别给予生理盐水和治疗药物注射,连续12天后处死,观察AA小鼠各组小鼠骨髓切片病理变化并计数骨髓有核细胞数。结果 （1）骨髓切片组织学所见：模型组小鼠骨髓增生极度低下,造血细胞明显减少,各治疗组仍显示骨髓增生低下,但可见散在造血灶。（2）骨髓有核细胞测定结果：与对照组比较,模型组骨髓有核细胞明显降低（P〈0.01）,各治疗组较模型组骨髓有核细胞显著增高（P〈0.01）。结论腹腔注射参附可增加骨髓有核细胞数,促进骨髓造血,对AA小鼠具有治疗作用。     

白花蛇舌草水提物及其与黄芪复方制剂对低白细胞模型小鼠造血功能的调节作用

目的:观察白花蛇舌草水提物及其与黄芪的复方制剂对化疗引起的白细胞减少症的预防和治疗作用.方法:以环磷酰胺腹腔注射造成小鼠白细胞减少症模型,用白花蛇舌草水提物和自制复方制剂进行治疗,对模型小鼠外周血象和骨髓造血功能进行检测.结果:白花蛇舌草水提物及其复方制剂可明显对抗模型小鼠的白细胞减少.结论:白花蛇舌草水提物及其与黄芪的复方制剂可保护小鼠外周血中白细胞免受环磷酰胺的伤害,同时对白细胞的回升有一定的促进作用;且水提物与复方制剂在升白细胞作用上无显著差异.