He-Ne激光照射巨噬细胞对胞内钙浓度及其免疫活性的影响

用He-Ne激光辐照巨噬细胞，实时观测活体单细胞内钙浓度([Ca^2 ]i)分布和细胞免疫活性随激光功率和照射时间的变化。实验结果表明，随激光照射时间的增加，[Ca^2 ]i出现上升达到最大值后又下降，逐渐回复初始状态，且呈现中心最强，径向衰减的环形梯度分布。不同激光剂量对巨噬细胞内[Ca^2 ]i及其免疫活性有着不同的影响，同一波长激光，当剂量不同时，可以表现为完全相反的效应。同时激光功率也是影响巨噬细胞内[Ca^2 ]i及其免疫活性的一个重要因素，同一波长、同一剂量情况下，如果激光功率不同，巨噬细胞内[Ca^2 ]i及其免疫活性的变化也有着显著的差异。当照射功率为0．16mw时，胞内[Ca^2 ]i峰值是照射功率为0．40mw时的近6倍。最大值时对应细胞免疫活性最高。对其机理作了初步探讨。     

Effects of Human and Porcine Adipose Extracellular Matrices Decellularized by Enzymatic or Chemical Methods on Macrophage Polarization and Immunocompetence

The decellularized extracellular matrix (ECM) obtained from human and porcine adipose tissue (AT) is currently used to prepare regenerative medicine bio-scaffolds. However, the influence of these natural biomaterials on host immune response is not yet deeply understood. Since macrophages play a key role in the inflammation/healing processes due to their high functional plasticity between M1 and M2 phenotypes, the evaluation of their response to decellularized ECM is mandatory. It is also necessary to analyze the immunocompetence of macrophages after contact with decellularized ECM materials to assess their functional role in a possible infection scenario. In this work, we studied the effect of four decellularized adipose matrices (DAMs) obtained from human and porcine AT by enzymatic or chemical methods on macrophage phenotypes and fungal phagocytosis. First, a thorough biochemical characterization of these biomaterials by quantification of remnant DNA, lipids, and proteins was performed, thus indicating the efficiency and reliability of both methods. The proteomic analysis evidenced that some proteins are differentially preserved depending on both the AT origin and the decellularization method employed. After exposure to the four DAMs, specific markers of M1 proinflammatory and M2 anti-inflammatory macrophages were analyzed. Porcine DAMs favor the M2 phenotype, independently of the decellularization method employed. Finally, a sensitive fungal phagocytosis assay allowed us to relate the macrophage phagocytosis capability with specific proteins differentially preserved in certain DAMs. The results obtained in this study highlight the close relationship between the ECM biochemical composition and the macrophage’s functional role.     

灵芝、姬松茸、黄芪混合多糖配伍茶多酚复合剂免疫增强作用的研究

本文对灵芝、姬松茸、黄芪多糖+茶多酚复合剂对小鼠细胞免疫、体液免疫和非特异免疫功能的增强作用进行研究.健康雄性小鼠分别灌胃给予不同剂量的复合剂,利用绵羊红细胞(SRSC)诱导小鼠DTH(足跖增厚法淞测细胞免疫功能的变化,利用抗体生成细胞(Jeme改良载片法)和半数溶血值(HC50)体液免疫功能的变化,利用小鼠腹腔巨噬细胞吞噬鸡红细胞试验(滴片法)检测单核-巨噬细胞活性的变化.结果表明低、中和高剂量组灵芝、姬松茸、黄芪多糖+茶多酚复合剂显著增强小鼠细胞免疫功能(P<0.01),中和高剂量组显著增强体液免疫功能和单核-巨噬细胞功能(P<0.05),复合剂对小鼠免疫器官无影响(P>0.05).     

黄伞菌丝体多糖提取工艺及免疫调节作用研究

在单因素试验基础上运用正交试验方法确定了黄伞菌丝体多糖最佳提取工艺,即菌丝体粉碎粒径为80目、加水量1∶15、提取时间2.5 h、提取温度80℃、提取1次,醇析多糖时使用无水乙醇、3.5倍于多糖溶液、醇析12 h;同时研究了黄伞菌丝体多糖在体外刺激小鼠脾淋巴细胞增殖作用,结果表明黄伞菌丝体多糖可以促进淋巴细胞增殖,具有体外调节免疫作用。     

干巴菌对小鼠免疫功能影响的实验研究

探讨干巴菌醇提取物的水溶液及脂溶液对小鼠免疫功能的影响。分别用干巴菌的水溶液及脂溶液给小鼠灌胃10 d后,检测小鼠免疫器官重量、单核-腹腔巨噬细胞吞噬能力、淋巴细胞增殖转化及细胞因子的情况。脏器/体重比值测定和淋巴细胞增殖转化试验阳性,干巴菌脂溶组TNFα的浓度的均数小于对应混合组和溶剂对照组。干巴菌有增强免疫功能的作用;干巴菌可能抑制正常小鼠分泌TNFα。     

果胶的磷酸化修饰及其免疫活性研究

以溶剂、反应时间、反应温度、三氯氧磷与果胶液固比为考察因素,磷含量为指标,研究果胶的磷酸化合成工艺,并对磷酸化修饰前后的果胶进行T淋巴细胞免疫增殖实验。结果表明:极性大的偶极溶剂和较高的反应温度有利于提高磷含量;随着溶剂用量、反应时间以及三氯氧磷用量的增加,磷含量先增加后下降。在反应温度50℃、反应时间3.0 h、三氯氧磷与果胶的液固比为3∶1的最佳合成工艺条件下,磷酸化果胶中的磷含量可高达11.52%。免疫增殖实验结果表明:苹果果胶、低磷含量和高磷含量的磷酸化苹果果胶没有免疫活性,中等磷含量的磷酸化苹果果胶有明显的免疫抑制活性。     

Investigating Immune Responses to the scAAV9-HEXM Gene Therapy Treatment in Tay–Sachs Disease and Sandhoff Disease Mouse Models

GM2 gangliosidosis disorders are a group of neurodegenerative diseases that result from a functional deficiency of the enzyme β-hexosaminidase A (HexA). HexA consists of an α- and β-subunit; a deficiency in either subunit results in Tay–Sachs Disease (TSD) or Sandhoff Disease (SD), respectively. Viral vector gene transfer is viewed as a potential method of treating these diseases. A recently constructed isoenzyme to HexA, called HexM, has the ability to effectively catabolize GM2 gangliosides in vivo. Previous gene transfer studies have revealed that the scAAV9-HEXM treatment can improve survival in the murine SD model. However, it is speculated that this treatment could elicit an immune response to the carrier capsid and “non-self”-expressed transgene. This study was designed to assess the immunocompetence of TSD and SD mice, and test the immune response to the scAAV9-HEXM gene transfer. HexM vector-treated mice developed a significant anti-HexM T cell response and antibody response. This study confirms that TSD and SD mouse models are immunocompetent, and that gene transfer expression can create an immune response in these mice. These mouse models could be utilized for investigating methods of mitigating immune responses to gene transfer-expressed “non-self” proteins, and potentially improve treatment efficacy.     

龙须菜纯多糖的制备及免疫活性研究

采用超滤分级、离子层析和分子筛凝胶层析方法,系统纯化龙须菜粗多糖,分离到多糖GCp F1–1B。经HPLC鉴定,GCp F1–1B为均一多糖,分子量为9.3×104 Da。紫外扫描显示其不含蛋白和核酸。高效阴离子色谱(HPAEC)分析发现,GCp F1–1B由半乳糖、果糖和葡萄糖醛酸组成,比例为14.28∶3.85∶1。生物活性研究显示,GCp F1–1B可以刺激小鼠脾淋巴细胞增殖作用和激活小鼠骨髓来源巨噬细胞(BMMs)的作用,表明其具有良好的免疫调节活性。     

Phagocytosis by invertebrate hemocytes: causes of individual variation in Panorpa vulgaris scorpionflies

An in vitro phagocytosis assay, adjusted to as little as 1 microL of insect hemolymph, enables the microscopic determination of phagocytosis for single individuals of small insects. Even repeated determination over the lifetime of individuals is possible. This method makes it feasible to study individual variation in invertebrate phagocytic capacity. Possible sources of such variation are reviewed in this article: genetic differences, development, aging, reproduction, presence of parasites, and diverse environmental influences are natural sources of individual variation in phagocytosis. However, the methods used for phagocytosis and microscopic evaluation are also (unwelcome) sources of variation. To optimize incubation time for in vitro phagocytosis, time courses were determined. Furthermore, the reliability of visual counting and image analysis for the microscopic quantification of phagocytosis are compared. The influences of larval development and adult aging on phagocytosis by Panorpa vulgaris hemocytes are subsequently demonstrated. During development, a decrease in hemocyte numbers but a simultaneous increase in the proportion of phagocytosing hemocytes was observed when larvae reached pupation. On the other hand, adults showed a dramatic decrease in phagocytic capacity with age, while cell numbers remained fairly constant. The results show that individual variation in phagocytosis can be determined accurately in small invertebrates and related to its causes. This might be especially interesting in the context of studies relating individual immunocompetence to ecology, life history variation, and behavior.     

鸡腿蘑多糖免疫功能的初步研究

对鸡腿蘑发酵液胞外多糖和胞内多糖进行分别提取,做其对小鼠腹腔巨噬细胞的吞噬活性和过氧化物酶活性的影响的研究,发现具有一定的刺激作用。其中各不同剂量组的胞外多糖对小鼠腹腔巨噬细胞的吞噬活性有明显的增强作用,并且在实验剂量范围内呈随剂量增加而上升的趋势。另外,胞内多糖也有增强作用,但效果不明显,其中16mg/kg•d剂量组效果最好。胞内、外多糖对小鼠腹腔巨噬细胞过氧化物物酶活性也都有增强作用,并且效果与剂量关系呈倒“U”型,其中胞内多糖8mg/kg•d剂量组的增强作用最明显,胞外多糖16mg/kg•d剂量组最明显。