The data we obtained show that in resting macrophages , the basal levels of inos activity and no production are relatively low ; nevertheless , inos activity and no production can be significantly induced in response to oligochitosan stimulation . oligochitosan ( 80 ( ug / ml ) significantly induced the release of no 12 h after incubation , and the amount of no increased with time 试验结果表明:用80林留ml剂量的壳寡糖作用于raw264 . 7细胞( 4xl护个细胞/ m1 ) 12h便可以明显诱导no的生成,随着刺激时间的延长, no的产量不断增加,但增加的幅度逐渐变小。
In addition , the heparinase is stimulated in the presence of ca2 + and mn2 + , but is inhibited by cu2 + and fe3 + . neither pmsf nor edta significantly affected enzyme activity . by infrared spectrum and proton resonance spectrum analysis , we found smaller polysaccharides , unsaturated bond and reducing sugar , which are specific products by heparinase 在产物分析中发现,该酶作用于肝素后,还原糖数量明显增加,通过红外色谱及质子谱的分析发现,产物中出现了含不饱和双键的还原寡糖,这是肝素酶作用于肝素后的特异性产物,从而可以进一步证明该菌确实产生肝素酶。
Here we managed to make cultured mice peritoneal macrophages be directly influenced by oligochitosan , and be stimulated by ifn - r before oligochitosan added , then measured the changes of gene transcription and translation level of both il - 1 and imf - a , respectively by methods of relatively quantitive rt - pcr and elisa . first , rt - pcr results showed that 18 hours was the most effective time and 40ug / ml was the most effective concentration of oligochitosan , then by the same method , confirm that 4hours is the most effective time and loou / ml is the most effective concentration of ifn - r stimulating . because ifn - r can enhance il - 1 and tnf - a gene expression of macrophages alone , so add ifn - r to microphages alone for 22 hours , then examined by rt - pcr , the results showed that il - 1 and tnf - a gene expression have no remarkable difference compared with the blank contrast group 此外,由于ifn y单独作用也可促进两种细胞因子基因表达,故在巨噬细胞中加入ifn y单独作用22h ,再经阿一pcr检测,发现加ifn y的实验组细胞的几一lp和tnf a基因转录水平与空白对照组相比较无显著性差异,可见,壳寡糖和ifn v对巨噬细胞il lp和tnf一口基因转录水平的影响在作用时间上无一致性,在壳寡糖作用最适时间时,仅受ifn y刺激的巨噬细胞il lp和tnf q基因转录己下降至刺激前水平,因此可以认为, ifn y的加入仅起到对巨噬细胞预刺激使之处于敏感状态的作用,有利于增强壳寡糖对巨噬细胞的作用。
This means that joining of ifn - yonly to stimulate and make the macrophages in sensitive state , help to strengthen the enhancing effect of oligochitosan on il - 1 and tnf - a gene expression of macrophages . after confirming the most effective time and the most effective concentration of oligochitosan and ifn - y , select three groups of cells , experiment group join loou / ml ifn - r , after 4 hours , add 40ug / ml oligochitosan ; oligochitosan contrast group add oligochitosan only ; blank contrast groupadd neither ifn - y nor oligochitosan , then the three goups culture 18 hours together 在确定了壳寡糖和ifn y的最适作用时间及最适作用浓度之后,选取三组细胞状态良好,数目比较一致的巨噬细胞,实验组中加入100u mlifn v预刺激4h后,再加入40ug ml壳寡糖,加糖对照组则只加40ng砌l壳寡糖,空白对照组两者都不加,继续培养18h 。
This work provides a foundation for further research on separation of combinding oligochitosan component of macrophage and elucidation on the immunoregulation mechanism of oligochitosan with macrophage . from this work we endeavor to find the theoretical ground for development of antitumor drugs and functional food from oligochitosan 通过本实验为进一步阐明壳寡糖对巨噬细胞的免疫调节机理和分离巨噬细胞壳寡糖结合组分提供一定的实验依据,为壳寡糖抗肿瘤药物和功能食品的开发提供理论依据。
Samples were taken from nonpregnant , pregnant and cultured endometrium cell , with using immunohistochemical staining , western - blot methods . to better understand this interaction , we further characterized the expression patterns of egfr messenger rnas in the mouse uterus on days 1 to 8 of pregnancy . in the present study , we found : two isofonns of non - phospho mapk was generally equally expressed in the nonpregnant and pregnant mice endometrium , and little difference among them 本研究以未孕及早孕小鼠( d1 ? d8 )的子宫内膜和培养的d4子宫内膜为样品,应用免疫组织化学、 western - blot等方法,检测了小鼠胚胎植入前后子宫内膜erk1 / erk2的表达部位及水平;用rt - pcr方法检测了egf及其受体egfr的表达情况;对培养的d4子宫内膜以le ~ y寡糖抗体阻断,观察了其对活性型mapk表达的影响。
Oligochitosan has been observed to upregulate the production of interleukin - l ( il - l ) , tumor necrosis factor - a ( tnf - a ) , granulocyte macrophage colony stimylating factor ( gmcsf ) , interleukin - 6 ( il - 6 ) by macrophage and induce immunologic adjuvant effects . many aspects of oligochitosan ' s functions have been discovered by researchers , but the mechanism is still unclear 整体动物实验表明壳寡糖能激活巨噬细胞,增强巨噬细胞的杀伤活性,诱导如白介素- 1 ( il - 1 ) 、肿瘤坏死因子( tnf - ) 、粒单核集落刺激因子( gm - csf ) 、白介素- 6 ( il - 6 )等细胞因子的分泌,增加t细胞和nk细胞的活性。
Previous investments have shown that chitin , chitosan or their oligosaccharides were able to stimulate immune system . either intravenously or intraperitoneal administered to mouse , these polysacca - rides or oligosaccharides were able to increase polymorphonuclear cells in blood or peritoneal exudate , and activated macrophages , t lymphocytes and nk cells of mouse 将这些多糖、寡糖经腹腔或静脉给予小鼠,可以使其腹腔渗出液及外周血中多型核白细胞增多,巨噬细胞、脾脏t淋巴细胞、 nk细胞等活性增强。
There are still many questions remain unanswered , including the mechanisms that specify oligochitosan recognition , the participation of additional molecules in transmembrane signaling , and the repertoire of downstream pathways activated by oligochitosan . it was also observed that pdtc did not totally block no production , which gave us another evidence that there must be some other signal transduction pathways ( perhaps mapk , pkc , ca2 + dependent signaling ) involved in the activation , so further investigation is necessary to clarify molecular mechanism of oligochitosan inducing immunological en Westemblot结果显示100林岁ml剂量的壳寡糖作用于raw264 . 7细胞( 4x105个细胞/ ml ) 4h ,胞核内p65的含量明显增加, 6h时达到高峰, 10h时恢复到基础表达量,这就提示在壳寡糖对巨噬细胞的激活过程中nf一kb活化状态可以持续6个小时。
Chitin and its derivative can strengthen the function of resisting on the microorganism of many kinds of etiologies of organism through regulating the function of immune system of organism , have effect of suppressing to tumour cells , such as mouse metha fibrosarcoma , s180 tumour , mm44 entity tumour and lewis lung cancer , etc . most of these investiments , however , were carried out through in vivo experiments , and research on stimulating function of oligochitosan to one particular part of immune system was very few 几丁质及其衍生物通过调节机体免疫系统功能,能增强机体对多种病原微生物的抵抗作用,对实验性小鼠metha纤维肉瘤、 s180肿瘤、 mm44实体瘤及lewis肺癌等肿瘤细胞的生长具有抑制作用。研究表明壳寡糖具有免疫调节作用,有抗感染及抗肿瘤活性。但众多研究以动物整体实验为主,关于壳寡糖对免疫系统某一特定环节直接作用的研究较少。