Structural and functional changes of immune system in aging mouse induced by D-galactose

Objective To investigate the role of D-galactose, especially in the structural and functional changes of the immune system in aging. Methods Serum levels of advanced glycation end-products （AGE） were determined by ELISA method. Ultra-structures of thymus and spleen were detected by transmission electron microscopy. MTT method was used to determine the lymphocyte proliferation. IL-2 activity was determined by bioassay. Northern blot was used to detect the IL-2 mRNA levels. Results Serum AGE levels of D-galactose- （P〈0.01） and AGE-treated （P〈0.05） mice （n=8） were increased significantly. The ultra-structures of thymus and spleen in D-galactose- and AGE-treated mice showed regressive changes similar to those in the aged control group. The lymphocyte mitogenesis and IL-2 activity of spleen were also decreased significantly （P〈0.01, n=8）. The change of IL-2 activity shown by Northern blot resulted from the change of mRNA expression. The AGE plus aminoguanidine group, however, showed no significant change in these parameters in comparison with the young control group （P〈0.01 or P〈0.05, n=8）. Conclusion D-galactose and AGE lead to a mimic regression change of aging in the immune system in vivo.     

Protective Role of Salidroside against Aginq in A Mouse Model Induced by D-galactose

Objective To investigate the protective effects of putative AGEs （advanced glycation endproducts） inhibitor salidroside against aging in an accelerated mouse aging model induced by D-galactose. Methods A group of 5-month-old C57BL/6J mice were treated daily with D-galactose, D-galactose combined with salidroside, salidroside alone, and control buffer for 8 weeks. At the end of the treatment, serum AGEs levels, neurological activities, expression of glial fibrillary acidic protein （GFAP） and neurotrophin-3 （NT-3） in the cerebral cortex, as well as lymphocyte proliferation and IL-2 production were determined. Results D-galactose induced mouse aging model was developed as described before. As expected, salidroside blocked D-galactose induced increase of serum AGEs levels. It also reversed D-galactose induced aging effects in neural and immune system, as evidenced by improving motor activity, increasing memory latency time, and enhancing lymphocyte mitogenesis and interleukin-2 （IL-2） production. Furthermore, elevated expression of GFAP and NT-3 in the aged model mice was also reduced upon salidroside treatment. Conclusion Salidroside inhibits AGEs formation in vivo, which at least partially contributes to its anti-aging effect in D-galactose induced aging model.     

抗衰老中药筛选：进展与挑战

To provide a better service for senior health care,we summarized screening studies of traditional Chinese anti-aging materia medica(TCAM).We collected and analyzed literature of TCAM screening studies using the lifespan test and animal models of aging from 1984 to 2012.We found 26 screening methods for TCAM,and 153 single herbs or active ingredients of TCAM that have been screened out during the past 28 years.The cell lifespan test,the fruit fly lifespan test, and D-galactose aging model were the most widely used and intensively studied screening methods.However,the method for establishing the D-galactose aging model needs to be standardized,and the D-galactose aging model cannot completely be a substitute for the normal aging mouse model.Great success has been achieved in screening studies in TCAM.To further improve screening studies in TCAM,we suggest that the D-galactose aging model be incorporated into the lifespan test in the New Drugs of Traditional Chinese Medicine Research Guide.     

Activities of aldose reductase, ATPases, and nucleotide concentrations of erythrocytes in patients with type 2 (non-insulin-dependent) diabetes mellitus.

Aldose reductase (EC 1.1.1.21) is implicated in the pathophysiology of diabetic complications. In this paper we determined the activities of aldose reductase and ATPases of the erythrocytes in 17 patients with Type 2 (non-insulin-dependent) diabetes mellitus (NIDDM). In the aldose reductase assay we used fluorometric method to avoid the disturbance of hemoglobin. With dihydronicotinamide adenine dinucleotide (NADH), we verified it was aldose reductase but not aldehyde reductase II that was activated in the erythrocytes of the patients with NIDDM. The aldose reductase activity of the erythrocytes in the patients was significantly higher (P less than 0.01) than that in the controls. The activity of Na+/K(+)-ATPase of the patients was significantly lower (P less than 0.01) than that of the controls. The activities of Ca(2+)-ATPase and Mg(2+)-ATPase on the erythrocyte membranes of the patients were similar to those of the controls. At the same time we measured the seven nucleotide concentrations in the erythrocytes of the patients. In this experiment we used ultrafiltration method, instead of acid precipitation to make it possible to determine dihydronicotinamide adenine dinucleotide phosphate (NADPH) and NADH. The concentrations of ATP, ADP and AMP were similar to those of the controls. The concentrations of NADPH, NAD+ and NADH in the erythrocytes of the patients were significantly lower (P less than 0.01, 0.05 and 0.05 respectively) than those of controls. The concentration of nicotinamide adenine dinucleotide phosphate (NADP+) in the patients was significantly higher (P less than 0.01) than that of controls.

     

NADPH oxidase-dependent oxidative stress and mitochondrial damage in hippocampus of D-galactose-induced aging rats

Mitochondrial DNA(mtDNA) common deletion(CD) plays a significant role in aging and age-related diseases.In this study,we used D-galactose(D-gal) to generate an animal model of aging and the involvement and causative mechanisms of mitochondrial damage in such a model were investigated.Twenty 5-week-old male Sprague-Dawley rats were randomly divided into two groups:D-gal group(n=10) and control group(n=10).The quantity of the mtDNA CD in the hippocampus was determined using a TaqMan real-time PCR assay.Transmission electron microscopy was used to observe the mitochondrial ultrastructure in the hippocampus.Western blot was used to detect the protein levels of NADPH oxidase(NOX) and uncoupling protein 2(UCP2).We found that the level of mtDNA CD was significantly higher in the hippocampus of D-gal-induced aging rats than in control rats.In comparison with the control group,the mitochondrial ultrastructure in the hippocampus of D-gal-treated rats was damaged,and the protein levels of NOX and UCP2 were significantly increased in the hippocampus of D-gal-induced aging rats.This study demonstrated that the levels of mtDNA CD and NOX protein expression were significantly increased in the hippocampus of D-gal-induced aging rats.These findings indicate that NOX-dependent reactive oxygen species generation may contribute to D-gal-induced mitochondrial damage.     

A Crude Preparation of Lymphocyte Chalone and Its Suppressive Effect on Immune Function in Mice

A crude preparation of lymphocyte chalone was extracted from calf Spleen using coldaoctone and distilled water.Its inhibitory cffect on mouse lymphocyte proliferation was demonstratedby ~3H-TdR incorporation method in vitro.This extract shoed no inhibitory effect on the~3H-TdR uptake of K_562 cells and displayed no cytotoxicity to mouse lymphocyte,suggesting thatthere existed the activity of lymphocytc chalone in the extract.The crude chalone inhibited ~3H-TdRuptake of mouse lymphocyte stimulated with concanavalin A(Con A)and mixed lymphocytc re-sponse(MLR),and the degree of the inhibition was greater as the doses of the extract in culturesbecame higher,indicating it had inhibitory effect on T lymphocytes Its influence on the numbers ofrosette-forming cells(RFC)and plaque-forming cells(PFC)in the spleen of mice revealed that ithad inhibitory effocts on some stages of immunization,including the actions on T and B lymphocytes     

Calcium Glucarate Prevents Tumor Formation in Mouse Skin

Calcium Glucarate (Cag),Ca salt of D-glucaric acid is a naturally occurring non-toxic compound present in fruits,vegetables and seeds of some plants,and suppress tumor growth in different models,Due to lack of knowledge about its mode of action its uses are limited in cancer chemotherapy thus the objective of the study was to study the mechanism of action of Cag on mouse skin tumorigenesis.Methods:We have estimated effect of Cag on DMBA induced mouse skin tumor development following complete carcinogenesis protocol.We measured,epidermal transglutaminase activity(TG),a marker of cell differentiation after DMBA and /or Cag treatemnt and [3H] thymidine incorporation into DMBA induced mouse skin tumor development.Topical application of Cag significantly modifies the critical events of proliferation and differentiation TG activity was found to be reduced after DMBA treatment,Reduction of the TG activity was dependent on the dose of DMBA and duration of DMBA exposure.Topical application of Cag significantly alleviated DMBA induced inhibition of TG.DMBA also caused stimulation of DNA synthesis in epidermis.which was inhibited by Cag.Conclusion:Cag inhibits DMBA induced mouse skin tumor development.Since stimulation of DNA synthesis reflects proliferation and induction of TG represents differentiation,the antitumorigenic effect of Cag is considered to be possible due to stimulation of differentiation and suppression of proliferation.     

Effects of Shugan Jianpi Formula (疏肝健脾方) on Myeloid-Derived Suppression Cells-Mediated Depression Breast Cancer Mice

Objective:To observe the intervention effect of Shugan Jianpi Formula(疏肝健脾方,SGJPF) on a breast cancer mouse model with depression and investigate the underlying mechanism of SGJPF in preventing the development of breast cancer.Methods:The breast cancer model was induced by inoculation of breast cancer cells,the depression model was induced by chronic stress stimuli,and the depression cancer model was established by combining the two factors.The mice were divided into 7 groups:normal control,depression model,tumor model,depression tumor model,SGJPF,chemotherapy,and SGJPF+chemotherapy groups.The last 3 groups were depression breast cancer mice and treated respectively with SGJPF,chemotherapy drug gemcitabine(GEM),and SGJPF alongside GEM.The condition of the mice was evaluated by the expression of 5-hydroxytryptamine in hippocampus after the sucrose water test and open field test,weight change,and survival time.Tumor growth was monitored with in vivo imaging.Flow cytometry was used to analyze the level of myeloid-derived suppression cell(MDSC) in the mouse spleen,T cell subsets,and the early apoptosis of CD8+T cells.Results:The SGJPF+GEM group had the highest inhibition rate and the longest survival time(P0.01).The MDSC level and the apoptosis rate of CD8+ T cells was the highest in the SGJPF+GEM group(P0.05).Conclusions:Depressive disorders and tumor growth could suppress the immune function of mice to different degrees,and the microenvironment in late 4T1 inflammatory breast cancer may play an important role in the pathological process.SGJYF could regulate the immune microenvironment by reducing CD8+ T lymphocyte apoptosis and tumor cell activity,increasing immune surveillance capability,and inhibiting MDSC proliferation,thus prolonging the survival time of tumor-bearing mice.     

Protective roles of α-lipoic acid in rat model of mitochondrial DNA4834bp deletion in inner ear

The protective roles of α-lipoic acid in the rat model of mitochondrial DNA (mtDNA) 4834bp deletion in inner ear were investigated. Forty female Wistar rats at 4 weeks of age were divided into four groups: group A (D-galactose group, n=10), group B (D-galactose+α-lipoic acid group, n=10), group C (α-lipoic acid group, n=10), and group D (control group, n=10). Auditory brainstem response (ABR) was used to detect the hearing threshold. Colorimetry was used to analyze activity of superoxide dismutase (SOD) and...     

Histochemical Localization of Glutathione Dependent NBT—Reductase in Mouse Skin

Objective:Localization of the glutathione dependent Nitroblue tetrazolium(NBT) reductase in fresh frozen sections of mouse skin and possible dependence of NBT reductase on tissue thiol levels has been investigated.Methods:The fresh frozen tissue sections(8m thickness)were prepared and incuated in medium containing NBT,reduced glutathione(GSH) and Phosphate uffer,The staining for GSH was performed with mercury orange.Results:The activity of the NBT-reductase in mouse skin has een found to be localized in the areas rich in glutatione and actively proliferating area of the skin.Conclusion:The activity of the NBT-reductase seems to be dependent on the glutatione contents.     

蓝莓对D-半乳糖模型小鼠学习记忆及抗氧化的研究

目的:探讨蓝莓对D-半乳糖致衰老小鼠学习记忆能力及抗氧化作用的影响。方法:连续给小鼠注射D-半乳糖6周,同时从第3周起给模型鼠灌胃蓝莓,给药结束后,进行学习记忆行为测定,采血清测定超氧化歧化酶(SOD),丙二醛(MDA)含量。结果:蓝莓组小鼠的学习、记忆获得和记忆消退的错误次数明显少于衰老模型组(P<0.05);与模型组比较,蓝莓组小鼠血清SOD活性显著性降低。结论:蓝莓能改善D-半乳糖致衰老小鼠的学习记忆能力,提高体内SOD活性,增强其对自由基的清除能力,抑制脂质过氧化作用。     

枸杞多糖对小鼠体内抗氧化酶活性及耐力的影响

目的研究枸杞多糖对小鼠耐力和血中超氧化物歧化酶（SOD）、丙二醛（MDA）水平的影响,进而探讨枸杞多糖的抗衰老作用机制。方法取小鼠随机分为枸杞多糖大剂量组（100 mg.kg-1.d-1）、小剂量组（50 mg.kg-1.d-1）和对照组,然后行小鼠游泳耐疲劳实验,测定并比较小鼠血中MDA含量及SOD活力。所有实验数据均采用均数±标准差（x±s）表示,对结果进行单因素方差分析和q检验。P〈0.05为差异有统计学意义。结果枸杞多糖可以延长小鼠力竭游泳时间,提高抗疲劳能力（力竭游泳时间比对照组分别延长了31.23%和12.32%）,差异有统计学意义;同时枸杞多糖组小鼠血中SOD活性明显升高、MDA含量明显降低,与对照组相比,差异有统计学意义（P〈0.01）。结论枸杞多糖具有良好的体内抗氧化作用,可以延缓衰老。     

枸杞多糖对大鼠海马缺血再灌注损伤的干预作用

目的:观察枸杞多糖(lycium barbarum polysaccharide,LBP)对大鼠海马缺血再灌注损伤后超氧化物歧化酶(superoxide dismutase,SOD)的活性和白介素6(interleukin 6,IL-6)含量的影响。方法:选用健康成年SD大鼠,雌雄不拘,随机分成3组:对照组、缺血再灌注组和枸杞多糖组。脑缺血模型采用线栓法致大脑中动脉阻闭,2h后抽出栓塞线,再灌注24h后处死大鼠,测定海马SOD活性和IL-6的含量。结果:(1)枸杞多糖对缺血再灌注大鼠海马SOD活性的影响:缺血再灌注组大鼠海马SOD活性显著低于对照组(P<0.01),枸杞多糖组大鼠海马SOD活性低于对照组,但显著高于缺血再灌注组(P<0.01)。(2)枸杞多糖对缺血再灌注大鼠海马IL-6的影响:枸杞多糖组大鼠海马IL-6含量与正常对照组相比差异没有统计学意义,而与缺血再灌注组大鼠相比明显下降(P<0.05)。结论:枸杞多糖可提高缺血再灌注大鼠海马SOD活性,降低IL-6的含量,对缺血再灌注损伤有保护作用。     

龙眼参多糖对不同模型拟痴呆小鼠的益智作用

目的　探讨龙眼参多糖对记忆障碍小鼠学习记忆功能的影响。方法　从龙眼参提取多糖 ,以东莨菪碱、40 0ml/L乙醇和D -半乳糖复制记忆障碍小鼠模型 ,以水迷宫为学习记忆评价指标 ,观察不同剂量的多糖对小鼠学习记忆的影响。结果　在东莨菪碱、40 0ml/L乙醇和D -半乳糖复制记忆障碍小鼠模型中 ,龙眼参多糖组与模型组比较 ,水迷宫的正确次数差异均有显著性 (P <0 .0 5 )。结论　龙眼参多糖能明显增强小鼠的学习记忆功能。     

姬松茸多糖对D-半乳糖诱导的衰老模型小鼠的抗衰老作用及其Keap1/Nrf2/ARE信号转导途径机制

目的：研究姬松茸酸性多糖A级分（ABP-A）对D-半乳糖（D-Gal）所致衰老模型小鼠的抗衰老作用,并探讨姬松茸多糖（ABP）的抗衰老机制。方法：水提醇沉法提取姬松茸粗多糖,并进行DEAE-纤维素离子交换柱层析分级及成分分析,得到ABP-A。48只ICR雄性小鼠随机分为对照组、模型组、阳性药物（吡拉西坦）组和ABP-A组,每组12只。给药70 d后进行小鼠Morris水迷宫、避暗和跳台行为学实验;检测各组小鼠血清中超氧化物歧化酶（SOD）活性、丙二醛（MDA）水平、总抗氧化能力（T-AOC）、过氧化氢酶（CAT）活性和活性氧（ROS）水平,Western blotting法检测各组小鼠脑组织中Nrf2、Keap1和HO-1蛋白表达水平。结果：ABP的总糖含量为75.1%,DEAE-纤维素离子交换柱层析分级得到ABP-A。行为学实验,与模型组比较,ABP-A组小鼠避暗潜伏期明显延长（P<0.05）,错误次数明显减少（P<0.05）,跳台潜伏期明显延长（P<0.05）,错误次数明显减少（P<0.05）,定位航行潜伏期明显缩短（P<0.05）,进入平台次数明显增加（P<0.05）。生化指标检测,与模型组比较,ABP-A组小鼠血清中SOD和CAT活性升高（P<0.05）,T-AOC升高（P<0.05）,MDA和ROS水平降低（P<0.05）。Western blotting法,与模型组比较,ABP-A组小鼠脑组织中HO-1蛋白表达水平升高（P<0.05）,Nrf2和Keap1蛋白表达水平降低（P<0.05或P<0.01）。结论：ABP可改善衰老模型小鼠学习记忆能力,其机制可能与调节以Nrf2为核心的Keap1/Nrf2/ARE氧化应激通路相关因子Nrf2、Keap1和HO-1发挥抗衰老作用有关。     

枸杞多糖对动脉粥样硬化模型兔血脂、脂质过氧化、NO和C-反应蛋白的影响

目的:探讨枸杞多糖对新西兰兔动脉粥样硬化的预防作用及其机制。方法:36只雄性新西兰兔随机分 为3组,每组12只。对照组喂普通饲料,模型组采用普通饲料加质量分数1.5%的胆固醇喂养,枸杞多糖(LBP)组 采用普通饲料加质量分数1.5%的胆固醇喂养,同时腹腔注射LBP(3.0mg/kg),10周后,测定各组空腹血清三酯酰 甘油(TG)、总胆固醇(TCh)、高密度脂蛋白胆固醇(HDL- C)、一氧化氮(NO)、丙二醛(MDA)、C -反应蛋白(CRP)含 量以及超氧化物歧化酶(SOD)活性,计算主动脉内膜粥样斑块面积并观察主动脉形态学变化。结果:模型组与对照 组比较,TG、TCh、NO、MDA、CRP升高(P<0.01),HDL- C/TCh、SOD活性下降(P<0.01),主动脉内膜粥样斑块面积 较大(P<0.01);LBP组与模型组比较,TG、NO、MDA、CRP下降(P<0.01或0.05),HDL -C/TCh、SOD活性升高 (P<0.01),主动脉内膜粥样斑块面积明显减少(P<0.05)。结论:枸杞多糖对动脉粥样硬化有明显的预防和治疗 作用,可能与其抗氧化、免疫调节、提高HDL- C/TCh有关。     

太子参多糖对东莨菪碱所致小鼠记忆障碍的改善作用

目的观察太子参多糖对小鼠学习、记忆能力的影响。方法采用跳台法测定正常小鼠和东莨菪碱诱导的记忆获得障碍模型小鼠的学习、记忆能力;采用分光光度法测定记忆获得障碍模型小鼠脑组织谷胱甘肽过氧化物酶（GSH-PX）、超氧化物歧化酶（SOD）活性和丙二醛（MDA）含量;采用急性脑缺血模型法观察小鼠断头后张口呼吸的次数和持续时间。结果①太子参多糖显著降低了记忆获得障碍小鼠受电击后的错误反应次数;②太子参多糖显著抑制小鼠脑组织MDA的生成,提高小鼠脑组织GS-PX和SOD酶活力;③太子参多糖显著延长急性脑缺血小鼠的张口呼吸次数和持续时间。结论太子参多糖对东莨菪碱致小鼠记忆获得障碍具有改善作用,可能与其改善脑缺血及抗氧化作用有关。     

枸杞多糖对细粒棘球蚴小鼠T淋巴细胞亚群的影响

目的研究枸杞多糖(Lycium Barbarum Polysaccharides,LBPs)对细粒棘球蚴小鼠脾脏T细胞亚群的影响,初步探讨LBP对细粒棘球蚴感染后机体免疫功能变化的影响。方法枸杞多糖溶液皮下给药3次,每次间隔2周,于第3次给药后2周,用细粒棘球蚴原头蚴攻击感染BALB/c小鼠,使用流式细胞仪动态检测给药前后、感染前后不同时间点小鼠脾脏IFN-γ+CD3+T细胞、IL-4+CD3+T细胞占总T细胞的比例以及CD4+/CD8+T细胞的比值变化。结果与给药前比较,枸杞多糖给药后7周,小鼠脾脏分泌IFN-γ的T细胞比例增加,分泌IL-4的T细胞比例减少,CD4+/CD8+的比例明显增加;与感染对照组相比,枸杞多糖能使细粒棘球蚴感染小鼠脾脏分泌IFN-γ的T细胞比例增加(感染后2、15周);分泌IL-4的T细胞比例减少(感染后2、10周),CD4+/CD8+的比例先升高(2、10周),后降低(15、20周)。结论枸杞多糖有正向免疫调节作用,通过促进细粒棘球蚴感染小鼠脾脏T淋巴细胞向Th1的分化,抑制Th2的分化,发挥抗细粒棘球蚴的作用。     

玛咖醇提取物对小鼠的抗衰老作用

目的探讨玛咖醇提取物的抗衰老作用及其机制。方法采用自然衰老小鼠为模型,连续ig给予玛咖醇提取物60 d后,测定小鼠血清、肝脏、心脏组织中谷胱甘肽过氧化物酶(G SH-Px)、超氧化物歧化酶(SOD)活性和丙二醛(M DA)的水平以及皮肤羟脯氨酸水平,免疫功能实验采用PHA刺激淋巴细胞转化及免疫器官指数的方法。结果玛咖醇提取物能显著提高老龄小鼠心脏和肝脏中G SH-Px活性及血清和肝脏中SOD活性,降低血清及肝脏中M DA水平,增加皮肤羟脯氨酸水平,提高PHA刺激的淋巴细胞转化率。结论玛咖醇提取物可能通过改善自由基代谢、增强细胞免疫发挥其抗衰老作用。