神经生长因子对正常和放射线-化学复合损伤小鼠造血调节因子及其受体影响研究

目的 探究神经生长因子(NGF)对正常和放射线-化学复合损伤小鼠造血调节因子及其受体的影响.方法 用实时荧光定量PCR和酶联免疫吸附检测注射NGF后正常和经60Coγ射线照射+腹腔注射环磷酰胺(放射线-化学复合损伤,放-化复合损伤)小鼠肾脏红细胞生成素(Epo)、脾脏红细胞生成素受体(EpoR)、骨髓细胞粒-巨噬系集落刺激因子(GM-CSF) mRNA表达量和血清Epo、GM-CSF、白细胞介素-3(IL-3)浓度变化.结果 注射NGF后正常小鼠脾脏EpoR mRNA表达、放-化复合损伤小鼠血清GM-CSF、IL-3显著高于注射生理盐水的对照组.结论 NGF可以改变小鼠的造血调节因子及其受体水平,但对正常和放-化复合损伤机体,其作用各不相同.     

纳米山药多糖合生元结肠靶向微生态调节剂对菌群失调大鼠免疫因子及SOD、MDA、NO、MPO表达的影响

目的探讨纳米山药多糖双歧杆菌合生元结肠靶向微生态调节剂对菌群失调模型大鼠的免疫因子及SOD(超氧化物歧化酶)、MDA(丙二醛)、NO(一氧化氮)、MPO(髓过氧化物酶)表达的影响。方法大鼠i.g盐酸林可霉素造成菌群失调伴有免疫缺陷肠炎模型,将大鼠随机分成靶向制剂组、阳性对照组和自然恢复组,紫外分光光度法检测结肠匀浆样本SOD、MDA、NO和MPO含量,微量溶血酶标仪分光光度法测定血清溶血素水平,ELISA法测定IL-1β(白细胞介素-1β)、IL-6(白细胞介素-6)、TNF-α(肿瘤坏死因子-α)、sIgA(分泌型免疫球蛋白A)及GM-CSF(粒细胞集落刺激生物因子)含量。结果与自然恢复组相比,纳米山药多糖组显著提高大鼠溶血素水平及sIgA、GM-CSF和SOD含量(P0.05),明显降低大鼠结肠样本中MDA、NO、MPO的含量(P0.05),IL-1β、IL-6和TNF-α在大鼠血清中的表达均有不同程度的下降(P0.05),且恢复到正常水平。结论纳米山药多糖双歧杆菌合生元结肠靶向微生态调节剂能减轻肠道炎症,提高溶血素水平、sIgA和GM-CSF含量,降低IL-1β、IL-6和TNF-α含量,提高机体免疫能力,是理想的中药微生态调节剂。     

应用PCR-DGGE法评价石斛多糖对肠道微生态失调的调节作用

目的 从微生态学角度研究石斛多糖治疗肠道微生态失调小鼠的作用及初步机制.方法 盐酸林可霉素灌胃制备肠道菌群失调小鼠模型,应用石斛多糖进行治疗,同时设正常对照组、丽珠肠乐组、阴性对照组,给药7d后处死小鼠,应用PCR-DGGE法检测肠道菌群丰富度、血清IL-2.结果 应用盐酸林可霉素灌胃3d后,小鼠肠道菌群丰富度、血清IL-2降低,持续治疗7d后,石斛多糖治疗小鼠肠道菌群丰富度、血清IL-2增高.结论 石斛多糖具有扶植肠道正常菌群生长,调整菌群失调,提高机体免疫力的作用.     

阿如拉-7味散对肠道菌群失调小鼠免疫功能及肠道菌群多样性的影响

目的研究阿如拉-7味散对抗生素诱导肠道菌群失调小鼠的免疫功能和肠道菌群多样性的影响。方法选取50只清洁级昆明小鼠,分为正常对照组、自然恢复组、阳性对照组、阿如拉-7味散低剂量组(0.5 g/mL)和阿如拉-7味散高剂量组(1.0 g/mL)。将所有小鼠用头孢曲松钠和盐酸林可霉素混合药液灌胃制备肠道菌群失调模型,第8天起各组小鼠相应药物连续治疗7 d,实验结束时进行样品采集。采集血清和回肠组织,采用ELISA法检测血清中的IgG、IFN-γ和TNF-α含量以及回肠组织中的sIgA含量;采集盲肠内容物,采用高通量测序技术检测肠道菌群多样性;采集胸腺和脾脏,测定免疫器官指数。结果阿如拉-7味散低、高剂量可显著提高抗生素诱导肠道菌群失调小鼠的脾脏指数(P<0.01)、血清IFN-γ(P<0.05)和TNF-α(P<0.01)含量以及回肠黏膜sIgA含量(P<0.01),但对胸腺指数(P>0.05)和血清IgG含量(P>0.05)无显著影响。各用药组与自然恢复组相比,尤其阿如拉-7味散低剂量组,小鼠肠道菌群丰富度和多样性明显升高。结论阿如拉-7味散对抗生素诱导肠道菌群失调小鼠具有提高免疫器官指数、增强肠道免疫功能和改善肠道菌群的作用。     

蒲公英多糖对小鼠肠道微生态的调节作用

目的探索蒲公英多糖对小鼠肠道微生态的调节作用。方法测定蒲公英多糖总糖含量及单糖组成。将小鼠分为正常组、模型组、阳性组和给药组(A1~A7),使用林可霉素灌胃制备肠道菌群失调模型,观察蒲公英多糖对小鼠一般状况、体重、肠道菌群、血清内毒素、小肠黏液s Ig A和血清IL-2的影响。结果蒲公英7组多糖中,相对分子质量100 000和6 000~10 000部分占总糖比例最高(86.40%)。经蒲公英多糖实验性治疗后,小鼠一般状况改善,体重有所增加,小鼠肠道内双歧杆菌和乳酸杆菌数量增加,肠杆菌和肠球菌数量减少。与模型组比较,给药组外周血内毒素含量减少(P0.01)、小肠黏液s Ig A和血浆IL-2含量均增加(P0.05),其中尤以A1、A2、A6组增加明显(P0.01)。结论蒲公英多糖能够改善林可霉素致小鼠肠道菌群失调,具有微生态调节作用。     

纳米山药多糖合生元结肠靶向微生态调节剂对菌群失调

摘要：目的 探讨纳米山药多糖双歧杆菌合生元结肠靶向微生态调节剂对菌群失调模型大鼠的免疫因子及SOD（超氧化物歧化酶）、MDA（丙二醛）、NO（一氧化氮）、MPO（髓过氧化物酶）表达的影响。方法 大鼠i.g盐酸林可霉素造成菌群失调伴有免疫缺陷肠炎模型,将大鼠随机分成靶向制剂组、阳性对照组和自然恢复组,紫外分光光度法检测结肠匀浆样本SOD、MDA、NO和MPO含量,微量溶血酶标仪分光光度法测定血清溶血素水平,ELISA法测定IL-1β（白细胞介素-1β）、IL-6（白细胞介素-6）、TNF-α（肿瘤坏死因子-α）、sIgA（分泌型免疫球蛋白A）及GM-CSF（粒细胞集落刺激生物因子）含量。结果 与自然恢复组相比,纳米山药多糖组显著提高大鼠溶血素水平及sIgA、GM-CSF和SOD含量（P<0.05）,明显降低大鼠结肠样本中MDA、NO、MPO的含量（P<0.05）,IL-1β、IL-6和TNF-α在大鼠血清中的表达均有不同程度的下降（P<0.05）,且恢复到正常水平。结论 纳米山药多糖双歧杆菌合生元结肠靶向微生态调节剂能减轻肠道炎症,提高溶血素水平、sIgA和GM-CSF含量,降低IL-1β、IL-6和TNF-α含量,提高机体免疫能力,是理想的中药微生态调节剂。     

纳米山药多糖合生元结肠靶向微生态调节剂对菌群失调大鼠 免疫因子及SOD、MDA、NO、MPO表达的影响

摘要：目的 探讨纳米山药多糖双歧杆菌合生元结肠靶向微生态调节剂对菌群失调模型大鼠的免疫因子及SOD（超氧化物歧化酶）、MDA（丙二醛）、NO（一氧化氮）、MPO（髓过氧化物酶）表达的影响。方法 大鼠i.g盐酸林可霉素造成菌群失调伴有免疫缺陷肠炎模型,将大鼠随机分成靶向制剂组、阳性对照组和自然恢复组,紫外分光光度法检测结肠匀浆样本SOD、MDA、NO和MPO含量,微量溶血酶标仪分光光度法测定血清溶血素水平,ELISA法测定IL-1β（白细胞介素-1β）、IL-6（白细胞介素-6）、TNF-α（肿瘤坏死因子-α）、sIgA（分泌型免疫球蛋白A）及GM-CSF（粒细胞集落刺激生物因子）含量。结果 与自然恢复组相比,纳米山药多糖组显著提高大鼠溶血素水平及sIgA、GM-CSF和SOD含量（P<0.05）,明显降低大鼠结肠样本中MDA、NO、MPO的含量（P<0.05）,IL-1β、IL-6和TNF-α在大鼠血清中的表达均有不同程度的下降（P<0.05）,且恢复到正常水平。结论 纳米山药多糖双歧杆菌合生元结肠靶向微生态调节剂能减轻肠道炎症,提高溶血素水平、sIgA和GM-CSF含量,降低IL-1β、IL-6和TNF-α含量,提高机体免疫能力,是理想的中药微生态调节剂。     

七味白术散及提取物对小鼠肠道菌群失调腹泻IL-8表达的影响

目的探讨七味白术散及七味白术散提取物对肠道菌群失调小鼠小肠黏膜IL-8的表达和血清中IL-8含量的影响。方法用头孢曲松钠和林可霉素联合中药番泻叶药液灌胃造模,观察小鼠大便性状、腹泻次数和粪便细菌培养结果确立造模成功后,将小鼠分成4组分别为模型组[MG,造模成功后不给予任何治疗又称自然恢复组(ZFG)],七味白术散组(QG),七味白术散提取物组(QTG),并设正常对照组(NG)。各治疗组给予相应药物0.4 m L/(只·次·d)灌胃,模型组、正常组小鼠给予等量的蒸馏水灌胃。分别于造模的第3天、7天、9天、11天和13天心脏采血处死小鼠,取血清采用ELISA法测IL-8的含量,取小肠行病理切片,采用免疫组织化学法观测小肠黏膜上皮细胞IL-8的阳性表达。结果血清IL-8含量变化与肠黏膜上皮细胞IL-8阳性表达变化趋势相一致:先升高后逐渐下降。模型组在造模3 d时IL-8表达明显升高,与正常对照组小鼠比较(P0.05),7、9、11和13 d表达逐渐下降,但仍然高于正常和各治疗组(P0.05);七味白术散组与提取物组IL-8的表达两组之间比较差异无统计学意义(P0.05),两组各个时间点IL-8的表达均低于模型组,差异有统计学意义(P0.05),在7、9 d时高于正常组,差异有统计学意义(P0.05),11、13 d时逐渐恢复正常,与正常组小鼠比较差异无统计学意义(P0.05)。结论七味白术散及其提取物均能降低肠道菌群失调小鼠IL-8的表达,从而抑制肠道炎症反应,减少肠道黏膜的免疫损伤。     

藏药巴桑母酥油丸对放射线-化学复合损伤小鼠粒-巨噬系造血功能的影响及机理研究

目的探讨藏药巴桑母酥油丸对放射线-化学复合损伤小鼠粒-巨噬系造血功能的影响及可能机理。方法60Coγ射线照射和环磷酰胺腹腔注射方式制造放射线-化学复合损伤小鼠模型,采用外周血细胞计数、造血祖细胞集落分析、实时荧光定量PCR,检测灌胃不同浓度巴桑母酥油丸后不同时间放射线-化学复合损伤小鼠外周白细胞数、骨髓粒-巨噬系(CFU-GM)造血祖细胞集落产率、粒-巨噬系集落刺激因子(GM-CSF)和粒-巨噬系集落刺激因子受体(GM-CSF R)mRNA相对表达量。结果灌胃14 d后,中、高剂量组的白细胞数显著高于生理盐水组;中剂量巴桑母酥油丸灌胃14 d后,骨髓CFU-GM集落产率、GM-CSF mRNA和GM-CSFR mRNA也显著高于空白组、生理盐水组。结论适当浓度的巴桑母酥油丸可以通过上调骨髓GM-CSF、GM-CSFR mRNA表达、促进CFU-GM增殖分化,进而促进放射线-化学复合损伤后小鼠外周血白细胞数的提前恢复。     

乙型肝炎肝硬化患者肠道菌群变化与血清细胞因子水平的相关性

目的探讨乙型肝炎肝硬化患者肠道菌群变化与血清干扰素-α(IFN-α)、白细胞介素6(IL-6)、白细胞介素1β(IL-1β)水平的相关性。方法选择2017年12月至2019年12月我院收治的105例乙型肝炎肝硬化患者为A组,50例乙型肝炎患者为B组,同期于我院体检健康者50例为C组。比较3组患者粪便标本中菌群分布情况,同时比较3组患者血清IFN-α、IL-6、IL-1β水平,不同程度肠道菌群失调乙型肝炎肝硬化患者血清IFN-α、IL-6、IL-1β水平。采用Pearson相关分析乙型肝炎肝硬化患者肠道菌群与血清IFN-α、IL-6、IL-1β的相关性。结果 A组患者肠道乳杆菌、双歧杆菌数量显著低于B组,而肠杆菌、肠球菌及血清IFN-α、IL-6、IL-1β水平显著高于B组(均P0.05)。B组患者肠道乳杆菌、双歧杆菌数量显著低于C组,而肠杆菌、肠球菌及血清IFN-α、IL-6、IL-1β水平显著高于C组(均P0.05)。Ⅱ度菌群失调乙型肝炎肝硬化患者血清IFN-α、IL-6、IL-1β水平显著高于I度菌群失调者(均P0.05)。Ⅲ度菌群失调乙型肝炎肝硬化患者血清IFN-α、IL-6、IL-1β水平显著高于Ⅱ度菌群失调者(均P0.05)。乙型肝炎肝硬化患者肠道肠杆菌、肠球菌数量与血清IFN-α、IL-6、IL-1β水平呈正相关,而其肠道乳杆菌、双歧杆菌数量与血清IFN-α、IL-6、IL-1β水平呈负相关(均P0.05)。结论乙型肝炎肝硬化患者存在明显的肠道菌群失调,同时其外周血IFN-α、IL-6、IL-1β水平呈现上升趋势。肠道菌群与血清IFN-α、IL-6、IL-1β可能协同参与了乙型肝炎肝硬化的发生及发展。     

Fine-structure mapping of the murine IL-3 and GM-CSF genes by pulsed-field gel electrophoresis and molecular cloning

The hemopoietic growth factors interleukin-3 (IL-3, multi-CSF) and granulocyte-macrophage colony-stimulating factor (GM-CSF) belong to a family of secreted glycoproteins that stimulate the proliferation and differentiation of hemopoietic progenitor cells. IL-3 and GM-CSF have overlapping biological activities and show similar regulation of expression after mitogenic or antigenic stimulation of T lymphocytes. In the present work we have derived a map of the region covering the Il-3 and Csfgm loci using a combination of pulsed-field gel electrophoresis and molecular cloning. The two genes are shown to be 14 kbp apart, in the same orientation with the IL-3 gene 5' of the GM-CSF gene. The proximity of the two genes, together with similarities in their structure, function, and regulation, suggests that they may have arisen by ancient gene duplication.     

Binding, internalization and degradation of radio-iodinated interleukin 3 and granulocyte-macrophage colony stimulating factor by various hemopoietic cells

The proliferation and differentiation of hemopoietic committed progenitor cells depend on colony stimulating factors (CSF). However, isolated mouse granulocyte-macrophage progenitor cells can still undergo limited proliferation in serum-free cultures after CSF deprivation. To test whether this is due to an accumulated pool of internalized factor, we examined the binding, internalization and degradation of radiolabelled interleukin 3 (IL-3) and granulocyte-macrophage colony stimulating factor (GM-CSF) in various hemopoietic cells. We found 20,000 high affinity IL-3 receptors on cells of two IL-3-dependent hemopoietic cell lines, FDC-P1 and FDC-P2 (Kd = 85 and 129 pM). FDC-P1 cells, which also respond to GM-CSF, possess 600 high-affinity GM-CSF receptors (Kd = 64 pM). Cells of both lines internalize IL-3, but only FDC-P1 cells release degraded IL-3 at a rapid rate. Both cell lines have similar dose-response curves for IL-3 and survival kinetics after factor removal. All other cells tested behave like FDC-P1, suggesting that the metabolism of IL-3 by FDC-P2 is exceptional. Our study indicates that transient proliferation of committed progenitor cells in the absence of added factors is apparently not due to a stable pool of internalized CSF but merely represents an intrinsic capability of these cells.     

Steel factor stimulates the tyrosine phosphorylation of the proto-oncogene product, p95vav, in human hemopoietic cells.

Steel factor (SF) (also called stem cell factor, mast cell growth factor, or c-kit ligand) is a recently cloned hemopoietic growth factor that is produced by bone marrow stromal cells, fibroblasts, and hepatocytes. In both mouse and man it acts synergistically with several colony stimulating factors, including interleukin-3 (IL-3) and granulocyte macrophage-colony stimulating factor (GM-CSF), to induce the proliferation and differentiation of primitive hemopoietic precursor cells. In order to study its mechanism of action and to explore the molecular basis for its synergistic activity we have examined the proteins that become tyrosine phosphorylated in response to SF, IL-3, and GM-CSF. We report herein that SF, but not IL-3 or GM-CSF, dramatically stimulates the tyrosine phosphorylation of the product of the recently discovered proto-oncogene, vav, in two SF-responsive human cell lines, M07E and TF-1. Although phosphorylation is very rapid, reaching maximal levels within 2 min at 37 degrees C, co-immunoprecipitation studies suggest that c-kit may either not associate directly with p95vav or bind to it with very low affinity. Nonetheless, our data suggest that c-kit may utilize p95vav to mediate downstream signaling in hemopoietic cells.     

Comparative effects of basophil-directed growth factors

IL-3, IL-5, and GM-CSF exert various overlapping functions in basophils. We investigated the receptor expression profiles and concentration-dependent effects of IL-3, IL-5, and GM-CSF on several basophil functions in comparison with their effects on eosinophils. The order of the receptor expression levels was IL-3Ralpha>IL-5Ralpha>GM-CSFRalpha in basophils and IL-5Ralpha>or=GM-CSFRalpha>IL-3Ralpha in eosinophils. Compared with eosinophils, basophils expressed a much higher level of IL-3Ralpha and similar levels of IL-5Ralpha and GM-CSFRalpha. The order of potency was IL-3>IL-5=GM-CSF for degranulation, survival, and CD11b expression in basophils, and IL-5=GM-CSF>or=IL-3 for survival and CD11b expression in eosinophils. However, IL-3 induced CD69 expression preferentially in basophils. Our results indicate that IL-3 is the most potent activator of human basophils, and that the rank order of potency of hemopoietic growth factors virtually corresponded to their receptor expression levels in both cell types.     

Specific binding of radioiodinated granulocyte-macrophage colony-stimulating factor to hemopoietic cells

The hemopoietic growth factor granulocyte-macrophage colony-stimulating factor, GM-CSF, specifically controls the production of granulocytes and macrophages. This report describes the binding of biologically-active 125I-labeled murine GM-CSF to a range of hemopoietic cells. Specific binding was restricted to murine cells and neither rat nor human bone marrow cells appeared to have surface receptors for 125I-labeled GM-CSF. 125I-Labeled GM-CSF only appeared to bind specifically to cells in the myelomonocytic lineage. The binding of 125I-labeled GM-CSF to both bone marrow cells and WEHI-3B(D+) was rapid (50% maximum binding was attained within 5 min at both 20 degrees C and 37 degrees C). Unlabeled GM-CSF was the only polypeptide hormone which completely inhibited the binding of 125I-labeled GM-CSF to bone marrow cells, however, multi-CSF (also called IL-3) and G-CSF partially reduced the binding of 125I-labeled GM-CSF to bone marrow cells. Interestingly, the binding of 125I-labeled GM-CSF to a myelomonocytic cell line, WEHI-3B(D+), was inhibited by unlabeled GM-CSF but not by multi-CSF or G-CSF. Scatchard analysis of the binding of 125I-labeled GM-CSF to WEHI-3B(D+) cells, bone marrow cells and peritoneal neutrophils indicated that there were two classes of binding sites: one of high affinity (Kd1 = 20 pM) and one of low affinity (Kd2 = 0.8-1.2 nM). Multi-CSF only inhibited the binding of 125I-labeled GM-CSF to the high affinity receptor on bone marrow cells: this inhibition appeared to be a result of down regulation or modification of the GM-CSF receptor.(ABSTRACT TRUNCATED AT 250 WORDS)     

A murine cell line (2E10.4.13) produces five hemopoietic stimulators, and interleukin-2 and interleukin-3

An interleukin-2 (IL-2)-independent murine lymphocyte clone (2E10.4.13) with the Thy1+Lyt1+2-T200+ phenotype was separated from the original IL-2-dependent natural killer (NK) cell line (PEC-1). Erythroid burst-promoting activity (BPA), erythropoietin (Ep), granulocyte/macrophage, megakaryocyte and eosinophil colony-stimulating factors (GM-, MK- and Eo-CSF), IL-2 and Interleukin-3 (IL-3) were produced when these cells were stimulated with phorbol myristate acetate (PMA). When the conditioned medium was run through ion-exchange high-performance liquid chromatography, BPA, Ep, GM-CSF, MK-CSF and Eo-CSF were eluted in the same region as IL-3. In contrast, MK-CSF, much of the GM-CSF and half of the Eo-CSF were eluted in a distinct region where no IL-3 was detected. Chemical analyses of the hemopoietic factors derived from a single T inducer clone indicated that all the hemopoietic activities were associated with IL-3 activity. Some CSF activities (GM-, MK- and Eo-CSF) also could be mediated by the distinct molecules from IL-3, evidence that heterogeneous molecules are responsible for CSF activity.     

IL-3 specifically inhibits GM-CSF binding to the higher affinity receptor

The inhibition of binding between human granulocyte-macrophage colony-stimulating factor (GM-CSF) and its receptor by human interleukin-3 (IL-3) was observed in myelogenous leukemia cell line KG-1 which bore the receptors both for GM-CSF and IL-3. In contrast, this phenomenon was not observed in histiocytic lymphoma cell line U-937 or in gastric carcinoma cell line KATO III, both of which have apparent GM-CSF receptor but an undetectable IL-3 receptor. In KG-1 cells, the cross-inhibition was preferentially observed when the binding of GM-CSF was performed under the high-affinity binding condition; i.e., a low concentration of ¹²⁵I-GM-CSF was incubated. Scatchard analysis of ¹²⁵I-GM-CSF binding to KG-1 cells in the absence and in the presence of unlabeled IL-3 demonstrated that IL-3 inhibited GM-CSF binding to the higher-affinity component of GM-CSF receptor on KG-1 cells. Moreover, a chemical cross-linking study has revealed that the cross-inhibition of the GM-CSF binding observed in KG-1 cells is specific for the β-chain, M_r 135,000 binding protein which has been identified as a component forming the high-affinity GM-CSF receptor existng specifically on hemopoietic cells.     

The network of hemopoietic regulatory proteins in myeloid cell differentiation.

There are clones of myeloid leukemic cells that can be induced to undergo terminal cell differentiation to macrophages by normal hemopoietic regulatory proteins. Induction of differentiation in two different clones of myeloid leukemic cells with interleukin 6 (IL-6) or granulocyte-macrophage colony-stimulating factor (GM-CSF) resulted in induction of mRNA for the hemopoietic regulatory proteins IL-6, GM-CSF, interleukin 1 alpha and interleukin 1 beta, tumor necrosis factor, and transforming growth factor beta 1. In one of these clones, induction of differentiation with GM-CSF was also associated with induction of mRNA for macrophage colony-stimulating factor (M-CSF) but not for the receptor for M-CSF (c-fms), whereas in the other clone, induction of differentiation with IL-6 was associated with induction of mRNA for both c-fms and M-CSF. The clones also differed in their responsiveness to these regulators. There was no induction of mRNA for granulocyte colony-stimulating factor or interleukin 3 during differentiation of either clone. The results indicate that the genes for a nearly normal network of positive and negative hemopoietic regulatory proteins are induced during differentiation of these myeloid leukemic cells and that there are leukemic clones with specific defects in this network.