应用环磷酰胺构建C57BL/6J小鼠免疫抑制模型

背景：在免疫增强剂的研究中,常用到免疫抑制模型,如何建立免疫抑制模型成为免疫增强剂作用评价的关键。 目的：应用环磷酰胺构建C57BL/6J小鼠免疫抑制模型。 方法：将小鼠随机分为正常对照组、环磷酰胺50 mg/kg 5 d组,环磷酰胺80 mg/kg 3 d组,环磷酰胺80 mg/kg 5 d组,环磷酰胺100 mg/kg 5 d组和环磷酰胺100 mg/kg隔天给药组。正常对照组小鼠腹腔注射生理盐水0.1 mL,连续5 d。环磷酰胺50 mg/kg 5 d组,环磷酰胺80 mg/kg 3 d组,环磷酰胺80 mg/kg 5 d组和环磷酰胺100 mg/kg 5 d组小鼠分别以环磷酰胺50,80,80,100 mg/kg腹腔注射连续5,3,5,5 d。环磷酰胺100 mg/kg隔天给药组小鼠腹腔注射100 mg/kg环磷酰胺,隔天1次,共注射3次。 结果与结论：与正常对照组比较,腹腔注射环磷酰胺可导致小鼠外周血中CD3⁺T,CD3⁺CD4⁺T及CD3⁺CD8⁺T细胞明显下降(P < 0. 05);谷丙转氨酶(除环磷酰胺50 mg/kg 5 d、80 mg/kg 3 d组)、谷草转氨酶、尿素显著升高(P < 0.05),其中环磷酰胺80 mg/kg 5 d组、环磷酰胺100 mg/kg 5 d组和环磷酰胺100 mg/kg隔天给药组对肝肾功能的影响更为明显。提示腹腔注射环磷酰胺可建立CD3⁺T,CD3⁺CD4⁺T及CD3⁺CD8⁺T细胞明显下降的免疫抑制模型,其中以环磷酰胺50 mg/kg 5 d和80 mg/kg 3 d方式对肝肾功能的损伤较小。     

豆类丝核菌次级代谢产物对免疫抑制小鼠免疫功能的影响

目的:研究豆类丝核菌次级代谢产物对环磷酰胺所致免疫抑制小鼠免疫功能的影响。方法:采用腹腔注射环磷酰胺(80mg/kg)制备免疫抑制模型,并给模型小鼠灌胃分别含苦马豆素(Swainsonine,SW)8、16、32mg/(kg.d)的豆类丝核菌次级代谢产物稀释液11天,同时设正常对照组和免疫抑制模型组,末次灌胃后24小时,眼眶采血,收集胸腺和脾,进行免疫学检测。结果:腹腔注射CTX80mg/kg,连续3天,可致小鼠免疫抑制,剂量为8、16mg/(kg.d)的SW组均可提高小鼠的外周血白细胞数,颉颃环磷酰胺所致的小鼠腹腔巨噬细胞吞噬功能的降低,提高外周血中溶血素水平、脾脏淋巴细胞转化以及NK细胞杀伤活性,尤其以SW16mg/(kg.d)最为明显。结论:豆类丝核菌次级代谢产物对环磷酰胺致免疫抑制小鼠的免疫功能有一定的恢复和增强作用。     

连翘苷拮抗环磷酰胺所致小鼠免疫抑制的实验研究

目的:研究连翘苷对环磷酰胺处理小鼠免疫功能的影响。方法:以腹腔注射环磷酰胺法复制小鼠免疫抑制模型,试验组小鼠分别按50、100、150 mg/ kg 剂量灌胃连翘苷,同时设模型组和对照组(等体积生理盐水灌胃),连续处理7 d。常规方法测定胸腺和脾的脏器系数,FITC 微球法测定腹腔巨噬细胞吞噬功能,ELISA 法检测外周血IL-2、IL-4 和外周血淋巴细胞cAMP 含量,分析不同剂量的连翘苷对免疫抑制小鼠上述指标的影响。结果:7 d 后成功建立免疫抑制小鼠模型。连翘苷能够不同程度地提高免疫抑制小鼠的免疫器官指数、巨噬细胞吞噬能力和血清中免疫因子含量,降低外周血淋巴细胞中cAMP 含量。结论:连翘苷能够抵抗环磷酰胺的免疫抑制作用,具有一定的免疫调节功能。     

白扁豆多糖对免疫抑制小鼠的免疫调节作用

目的探讨白扁豆多糖(polysaccharide from Dolichos lablab L.,DLP)对环磷酰胺(cyclophosphamide,CTX)所致的免疫抑制小鼠的免疫调节作用。方法小鼠随机分为正常对照组、模型组、香菇多糖(Lentinan,LNT,15 mg/kg)和白扁豆多糖低、中、高剂量组(75、150、300 mg/kg)。通过对小鼠连续3 d腹腔注射环磷酰胺(100 mg/kg)建立小鼠免疫抑制模型,造模后连续灌胃给药7 d,检测免疫器官重量指数、血清溶血素、脾淋巴细胞增殖、脾脏NK细胞活性及小鼠血清细胞因子IL-2、IL-4和INF-γ水平,观察白扁豆多糖的免疫调节作用。结果与正常对照组相比,模型组小鼠的免疫器官重量指数、血清溶血素、脾淋巴细胞增殖、脾脏NK细胞活性及小鼠血清细胞因子IL-2、IL-4和INF-γ水平均明显降低(P<0.05,P<0.01),而中、高剂量DLP和香菇多糖可明显减轻由环磷酰胺引起的上述各项指标的下降(P<0.05,P<0.01)。结论 DLP能改善环磷酰胺所致免疫抑制小鼠的免疫功能。     

霍山石斛多糖对小鼠的双向免疫调节作用

目的观察霍山石斛多糖对小鼠的双向免疫调节作用。方法将90只昆明种小鼠随机分成9组,分别为正常对照组、免疫亢进模型组、免疫亢进+霍山石斛多糖[低(50 mg?kg-1?d-1)、中(100 mg?kg-1?d-1)、高(200 mg?kg-1?d-1)]剂量组、免疫抑制模型组、免疫抑制+霍山石斛多糖[低(50 mg?kg-1?d-1)、中(100 mg?kg-1?d-1)、高(200 mg?kg-1?d-1)]剂量组。采用连续3 d腹腔注射环磷酰胺(80 mg·kg-1)制作免疫抑制模型小鼠,采用连续11 d肌注卡介苗0.5 mg/只制作免疫亢进模型小鼠,观察霍山石斛多糖对免疫抑制模型小鼠和免疫亢进模型小鼠腹腔巨噬细胞的吞噬活性、体外脾淋巴细胞增殖能力和胸腺指数的影响。结果霍山石斛多糖能明显改善环磷酰胺所致的小鼠腹腔巨噬细胞吞噬活性降低、脾淋巴细胞体外增殖能力降低和胸腺指数降低,缓解卡介苗所致的小鼠腹腔巨噬细胞吞噬活性亢进、脾淋巴细胞体外增殖能力亢进和胸腺指数降低。结论霍山石斛多糖具有较好的双向免疫调节作用。     

营养组合物对再生障碍性贫血小鼠细胞线粒体形态及跨膜电位的影响

目的 探讨营养组合物治疗再生障碍性贫血的作用机制.方法 BALB/c小鼠100只,适应性喂养1周后,随机分为:正常对照组、再障模型组(AA)及不同剂量的营养组合物组.采用皮下注射乙酰苯肼100mg/kg,次日X射线2.0Gy照射后,于试验第5天环磷酰胺80 mg/kg腹腔注射,第15天重复以上步骤但不给予射线处理的方法...     

熊果酸对SCG-7901胃癌荷瘤小鼠肿瘤的抑制作用及肠道菌群的影响

目的 探讨熊果酸对SCG-7901胃癌荷瘤小鼠肿瘤的抑制作用及肠道菌群的影响.方法 KM小鼠随机分为正常对照组、模型组、环磷酰胺组、熊果酸低剂量组、熊果酸中剂量组、熊果酸高剂量组,每组各20只.皮下移植构建SCG-7901胃癌荷瘤小鼠模型,环磷酰胺组和熊果酸组小鼠分别腹腔注射环磷酰胺和不同剂量熊果酸,1次/d,连续给药...     

精氨酸单糖苷( AF) 对免疫抑制小鼠免疫功能的影响

目的:研究精氨酸单糖苷(Arginyl-fructose,AF)对环磷酰胺(CTX)诱导的免疫抑制小鼠免疫功能的影响。方法: ICR 小鼠100 只,随机分成5 组,即正常对照组(N),免疫抑制模型组(M),免疫抑制AF 低、中、高剂量组(M-L、M-M、M-H)。试验组连续给药28 d。免疫抑制组,每周前3 天,腹腔注射80 mg/ kg 环磷酰胺,形成免疫抑制。末次给药12 h 解剖后,测定免疫器官指数,AF 对脾淋巴细胞转化及增殖、巨噬细胞吞噬功能、特异性IgG 抗体、血清中TNF-β、IL-2 含量的影响。并通过实时荧光定量PCR 检测TNF-β、IL-2 基因表达的差异。结果:AF 能够显著提高小鼠的胸腺指数和脾脏指数,促进脾淋巴细胞的自然转化与增殖,增强巨噬细胞吞噬功能,显著提高小鼠血清中特异性IgG 抗体含量,可显著提高小鼠血清中TNF-β、IL-2 含量,实时荧光定量PCR 检测结果显示,TNF-β、IL-2 mRNA 能够良好表达。结论:AF 能拮抗CTX 免疫功能低下小鼠的免疫抑制作用。     

茯苓多糖对免疫抑制小鼠粘膜淋巴组织及脾脏中CD3~+和CD19~+细胞变化的影响

目的:通过研究口服茯苓多糖对环磷酰胺诱导的小鼠淋巴细胞亚群变化的作用,探讨茯苓多糖对肠道粘膜免疫与外周免疫系统作用的差异。方法:连续两周灌胃给予小鼠200 mg/(kg.d)茯苓多糖,在第14天腹腔注射100 mg/kg的环磷酰胺诱导免疫抑制模型,24小时后处死小鼠分别取派氏结(PPs)、肠系膜淋巴结(MLNs)和脾脏(SP)细胞,进行CD3+、CD19+双色免疫荧光标记,上流式细胞仪检测。结果:腹腔注射100 mg/kg的环磷酰胺后,小鼠PPs、MLNs和SP中的CD3+细胞比例上升,CD19+细胞比例下降,口服茯苓多糖可以明显的对抗PPs、MLNs中的CD3+、CD19+细胞比例的变化,但对SP中的CD3+、CD19+细胞比例的变化的作用不显著。结论:口服茯苓多糖能有效对抗环磷酰胺诱导的淋巴细胞亚群的变化,尤其是对PPs作用明显,提示茯苓多糖对肠道粘膜免疫系统的作用强于对外周免疫系统的作用。     

环磷酰胺免疫删减法制备细胞表面抗原单克隆抗体小鼠模型的建立

目的:观察不同剂量环磷酰胺(CP)诱导免疫耐受的作用, 建立一个免疫删减法制备细胞表面抗原单克隆抗体(mAb)的动物免疫模型.方法:雌性 BALB/c小鼠经CHO细胞免疫后分别给予不同剂量的CP诱导免疫耐受, 选择血清抗体效价最低的一组小鼠按常规方式腹腔注射CHO-TM5细胞进行免疫.ELISA测定各小鼠血清中CHO抗体及CHO-TM5抗体效价, 检测血清抗体对CHO细胞与CHO-TM5细胞结合反应的差异性.结果:BALB/c小鼠腹腔注射不同剂量的CP后, 各剂量组均出现不同程度的免疫耐受作用.CP 100 mg/kg组与CP 125 mg/kg组小鼠血清抗体效价较低, 接近阴性对照组.ELISA检测显示CP 100 mg/kg组血清抗体与CHO-TM5细胞呈阳性结合反应, 不与CHO细胞结合;而CP 0 mg/kg组血清抗体与CHO细胞与CHO-TM5细胞的结合反应均呈阳性.结论:CP 100 mg/kg具有良好诱导小鼠对CHO和CHO-TM5细胞的共同抗原表位产生免疫耐受, 相对增强对CHO-TM5细胞目的抗原hTM的免疫应答的作用, 为免疫删减法制备细胞表面抗原mAb提供了一个理想的动物免疫模型.     

The Effect of Two Glucan Carboxymethyl Derivatives with Various Substitution Degrees on Cyclophosphamide Immunosuppression in Mice

Carboxymethylglucan (CMG)¹ in two different degrees of substitution of carboxymethyl groups (0.56 and 0.89) was administered to cyclophoshamide (CY) treated (200 mg/kg) C57B1/6 mice in three doses (10, 50, and 100 mg/kg) 24 h after CY. The influence of CMG administration on the cell suppression caused by CY was observed in the subsequent days. The cellularity of spleen, bone marrow and peripheral blood was quantified on days 2, 5, 8, 11, 15, and 21 after CY treatment.

CY treated mice developed a significant decrease in peripheral blood cell counts, and spleen and bone marrow cellularity during the initial 5 days, followed by recovery in the next 15 days, with an overshoot reaction in spleen cellularity and erythrocyte levels.

The initial cellularity depression and the following recovery was modified by both carboxymethyl derivatives of glucan. Cyclophosphamide treated mice exhibited less pronounced immunosuppression and more rapid hematopoietic recovery when administered with CMG, althought this reaction was only of a modest degree. The effects were not dose dependent and the differences between the two glucans were not significant.     

The Effect of Two Glucan Carboxymethyl Derivatives with Various Substitution Degrees on Cyclophosphamide Immunosuppression in Mice

Abstract

Carboxymethylglucan (CMG)¹ in two different degrees of substitution of carboxymethyl groups (0.56 and 0.89) was administered to cyclophoshamide (CY) treated (200 mg/kg) C57B1/6 mice in three doses (10, 50, and 100 mg/kg) 24 h after CY. The influence of CMG administration on the cell suppression caused by CY was observed in the subsequent days. The cellularity of spleen, bone marrow and peripheral blood was quantified on days 2, 5, 8, 11, 15, and 21 after CY treatment.

CY treated mice developed a significant decrease in peripheral blood cell counts, and spleen and bone marrow cellularity during the initial 5 days, followed by recovery in the next 15 days, with an overshoot reaction in spleen cellularity and erythrocyte levels.

The initial cellularity depression and the following recovery was modified by both carboxymethyl derivatives of glucan. Cyclophosphamide treated mice exhibited less pronounced immunosuppression and more rapid hematopoietic recovery when administered with CMG, althought this reaction was only of a modest degree. The effects were not dose dependent and the differences between the two glucans were not significant.     

Fungus dose-dependent primary pulmonary aspergillosis in immunosuppressed mice.

We report on a model of primary pulmonary aspergillosis occurring after intranasal instillation of concentrated suspensions of conidia of Aspergillus fumigatus in immunocompromised mice. Unconcentrated suspensions of inoculum contained ca. 2 x 10(7) conidia per ml (1x). These suspensions were concentrated by centrifugation, adjusted to give ca. 2 x 10(8) (10x) or 2 x 10(9) (100x) conidia per ml, and delivered in 30-microliters droplets to the nares of anesthetized mice. Mice were untreated or injected with cortisone acetate (CA) or cyclophosphamide (CY) in various dosage regimens. It was not possible to obtain mortality of more than 50% with sublethal immunosuppressive treatment and 1x fungus. In contrast, mortality followed a fungus dose response in mice receiving sublethal immunosuppression with either CA or CY. Mortality rates of up to 100% were obtained with 100x fungus and a single dose of CY (200 mg/kg) or CA (250 mg/kg) or three alternate doses (125 mg/kg per day) of CA prior to infection. This model is applicable to the study of acute, fatal primary pulmonary aspergillosis and chemotherapy trials.     

T cell subpopulations in myocardial inflammatory infiltrates detected by confocal microscopy: dose dependence in mice treated with cyclophosphamide during acute Trypanosoma cruzi infection

In this article, we have characterized cell subpopulations found in the hearts of mice presenting acute Chagas' disease by immunocytochemistry and subjected to different schedules of an immunosuppressive therapy with cyclophosphamide (CY). In this comparative study, CY treatment with different doses was carried out before or after infection with Trypanosoma cruzi Y strain trypomastigotes, enabling us to discriminate the parasitemic kinetics and inflammatory processes in the heart, 12 d after infection. Animals treated with 200 mg/kg of CY 2 d before infection presented high parasitaemia as well as heavy inflammation and low parasite loads in the heart. Mice treated 5 d after infection with the same dose, developed the same parasitaemic peak but were not able to control it. Their heart did not present inflammation, but a high number of parasites could be seen. Animals treated with five 3 mg/kg doses of CY every other day presented heavy inflammatory reaction and low parasitaemia. In this group, as well as the one treated before infection, immunocytochemistry studies have shown predominance of CD8(+) T cells in the myocardium. On the other hand, mice treated with 200 mg/kg of CY 5 d after infection, presented small amounts of CD4(+) T cells while no CD8(+) could be found. These results have confirmed the dose dependence influence of this drug on the T cell populations in the inflammatory infiltrates as well as the importance of the schedule employed.     

Fludarabine-based conditioning for marrow transplantation from unrelated donors in severe aplastic anemia: early results of a cyclophosphamide dose deescalation study show life-threatening adverse events at predefined cyclophosphamide dose levels

Excessive adverse events were encountered in a Phase I/II study of cyclophosphamide (CY) dose deescalation in a fludarabine-based conditioning regimen for bone marrow transplantation from unrelated donors in patients with severe aplastic anemia. All patients received fixed doses of antithymocyte globulin, fludarabine, and low-dose total body irradiation. The starting CY dose was 150 mg/kg, with deescalation to 100 mg/kg, 50 mg/kg, or 0 mg/kg. CY dose level 0 mg/kg was closed due to graft failure in 3 of 3 patients. CY dose level 150 mg/kg was closed due to excessive organ toxicity (n?=?6) or viral pneumonia (n?=?1), resulting in the death of 7 of 14 patients. CY dose levels 50 and 100 mg/kg remain open. Thus, CY at doses of 150 mg/kg in combination with total body irradiation (2 Gy), fludarabine (120 mg/m(2)), and antithymocyte globulin was associated with excessive organ toxicity.     

Evaluation of the immunosuppressive effect of cyclophosphamide and dexamethasone in mice with visceral toxocariasis

Visceral toxocariasis is a serious public health problem with a cosmopolitan distribution. Children are susceptible due to their immature immune system and high risks of infection. Nevertheless, the few completed studies about immunosuppression have had controversial results. To evaluate the effect of two immunosuppressive drugs on the larval burden of Toxocara canis, four groups of ten Swiss strain mice each were inoculated on day 0 with 1,200 embryonated T. canis eggs. Fifteen days before the experimental infection, group 1 (control) was treated via intraperitoneal injection (IP) with sterile distilled water and groups 2 and 3 were treated with dexamethasone (DEX) at 1 and 5 mg/kg/day, respectively. Additionally, group 4 was treated IP with cyclophosphamide (CY) at 50 mg/kg at two times per week for 2 weeks. Sixty days following infection, the mice were euthanised to recover the larvae by means of the tissue digestion technique. The levels of antibodies detected by indirect ELISA were not associated with the larval burden. Administration of CY (50 mg/kg) and DEX (5 mg/kg) resulted in an increase of the larval burden of 162.1% and 50.8%, respectively, in relation to the control group. These two treatments, especially CY (50 mg/kg), promoted immunosuppression and the establishment of a significant larval burden, supporting its further utilisation in studies related to immunosuppression in visceral toxocariasis.     

Protection from experimental autoimmune diabetes in the non-obese diabetic mouse with soluble interleukin-1 receptor

We have evaluated the effects of a treatment with soluble interleukin-1 receptor (sIL-1R) in the accelerated model of autoimmune diabetes induced by cyclophosphamide (CY) in the non-obese diabetic (NOD) mouse. Prior to the CY challenge (350 mg/kg body weight), female euglycemic NOD mice were randomly divided into three groups (A–C). Groups B and C were treated daily from 1 day before to 13 days after the CY challenge with sIL-1R at doses of 0.2 and 2 mg/kg body weight. Group A was treated with PBS. By 2 weeks after CY administration, an acute form of autoimmune diabetes with glycosuria, hyperglycemia and severe insulitis occurred in the majority (13/20, 65%) of the control mice (group A). In contrast, repeated injections with sIL-1R protected NOD mice from insulin-dependent diabetes mellitus (IDDM) development in a dose-dependent fashion; the incidence of IDDM was 53.3% (8/15) in the mice treated with 0.2 mg/kg and only 6.7% (1/15) in those treated with 2 mg/kg. However, none of the doses of the sIL-1R reduced the extent of insulitis in NOD mice. Importantly, the anti-diabetogenic property of sIL-1R may not involve major T cell function impairment; accordingly, in parallel experiments, splenic lymphoid cells from NOD mice not challenged with CY, but treated with 2 mg/kg sIL-1R for 5 consecutive days showed a normal distribution of mononuclear cell subsets and maintained their capacity to secrete interferon-γ and IL-2 and to proliferate in response to polyclonal mitogenic stimulation with concanavalin A.     

Protective effect of extract of Mauremys mutica against cyclophosphamide (CY)-induced suppression of immune function in mice

The present study evaluated the immune function of extract obtained from yellow pond turtle (YPT). Two YPT extract doses (250?mg/kg and 500?mg/kg·bw/d) were administered orally to normal and cyclophosphamide (CY)-induced immune-suppressed male Kunming mice, respectively. The immune function were evaluated by testing the body weight growth curve, thymus index and spleen index in CY-induced mice as well as mononuclear-macrophage phagocytic function by the carbonic particle clearances index and the macrophage phagocytose index. After a 10-day administration, both the low-dose group (250?mg/kg·bw/d) and high-dose group (500?mg/kg·bw/d) showed a marked increase in immune organs indices and the carbonic particle clearances index and macrophage phagocytose index. Moreover, immune function showed significant dose–effect relationship. These results demonstrated the potential use of YPT extract as a functional medicine for enhancing human immunity and immunomodulatory for adjuvant treatment in cancer.     

Effects of quinoline and 8-hydroxyquinoline on mouse bone marrow erythrocytes as measured by the micronucleus assay

Both quinoline and 8-hydroxyquinoline (HOQ) were tested for their genotoxicity in CD1 male mice by using a bone marrow micronucleus assay. Mice were intraperitoneally treated in single injections with three dose levels (25, 50, and 100 mg/kg) of each chemical with corn oil as solvent vehicle. Bone marrow was sampled at 24, 48, and 72 h postinjection. Quinoline resulted in a significant dose-related increase in the number of micronucleated polychromatic erythrocytes (MPCE) at the 24 h sampling time for all doses tested. The high dose (100 mg/kg) and the medium dose (50 mg/kg) also induced statistically significant increases (P less than .05) in the number of MPCEs at 48 h interval. The ratios of polychromatic to normochromatic erythrocytes at the 24 h sampling time were lower for the treated than the control animals. Although HOQ resulted in some increases in the number of MPCEs over the control, this compound induced a statistically significant increase in the number of micronucleated normochromatic erythrocytes (MNCEs) at all three doses following 24 h treatment. Both low and medium doses also induced a higher incidence of MNCEs at the 48 and 72 h sampling times. No data were available for the high dose at these times. The cytotoxic effect of this compound was expressed as low PCE/NCE ratios with all doses at 24 h after injection and as a high mortality rate in animals treated with the high dose (100 mg/kg).