西洋参茎叶皂苷对CPHD病人外周淋巴细胞增强与调节作用

目的：为了开发西洋参在临床医学方面的应用，提高CPHD病人的各项免疫指标。方法：选择40例CPHD病人，分成用药组20例，对照组（非用药组）20例，通过流式细胞术检测病人外周血中T淋巴细胞亚群群中各亚群所占的比例来反映机体的细胞免疫状态，结论：通过两组临床研究观察，用药组疗效优于对照组（P＜0．05）。西洋参茎叶皂苷可直接作用于淋巴细胞分化成熟过程，具有增强和调节CPHD病人体免疫功能的作用，结论：合理应用西洋参茎叶茎叶皂苷对改善CPHD病人免疫功能能低下和紊乱状态，具有重要临床意义。     

西洋参茎叶皂苷与胸腺肽对CPHD病人细胞免疫功能的比较研究

随着临床免疫分子学的深入研究，细胞免疫越来越倍受重视，尤其在呼吸系统疾病中，如CPHD患者细胞免疫功能常常低于正常人，其T淋巴细胞亚群及淋巴细胞转化率有明显异常，呈下降的趋势。T细胞的改变与细菌感染、病毒感染密切相关。CPHD为慢性炎症性疾病，中性粒细胞释放蛋白酶及氧化物引起肺部损伤和肺气肿是疾病持续进展的主要原因之一。     

人参茎叶皂甙对小鼠免疫功能的影响

肌注人参(根)皂甙或人参茎叶皂甙50mg/kg,一周均能明显对抗环磷酰胺所致小鼠免疫器官重量减轻,血清特异性抗体水平下降及淋转功能降低,并使其恢复接近正常对照水平,使地塞米松所致小鼠腹腔巨噬细胞吞噬功能下降和血中ANAE~ 淋巴细胞比率降低提高到正常水平,促进抗体生成作用,人参(根)皂甙比人参茎叶皂甙更强。     

大豆皂甙对小鼠细胞免疫功能的增强作用

实验结果表明,大豆皂甙经口给予小鼠后能明显促进 ConA 和 Lp(?) 对小鼠脾细胞的增殖反应,能明显增强脾细胞对 IL—2的反应性,增加小鼠脾细胞对 IL—2的分泌,并明显的提高 NK 细胞、LAK 细胞毒活性.所以大豆皂甙具有明显的免疫调节作用.     

黄芪,人参皂甙对肺癌浸润淋巴细胞体外抗肿瘤作用的影响

为了进一步提高肿瘤浸润淋巴细胞（ＴＩＬ）过继免疫治疗恶性肿瘤的效果，应用＾３Ｈ－ＴｄＲ释放法，观察了黄芪，人参皂甙对肺癌浸润淋巴细胞体外抗肿瘤作用的影响。结果表明，黄芪与人参皂甙均能显著增强ＴＩＬ细胞的体外杀伤活性，二者合用对ＴＩＬ细胞具有协同正向调节作用，是较为理想的替代细胞因子，辅助ＴＩＬ细胞过继免疫治疗恶性肿瘤的天然免疫调节剂。     

西洋参茎叶总皂苷通过抑制过度内质网应激减轻大鼠心肌缺血/再灌注损伤

 目的：观察西洋参茎叶总皂苷（PQS）对大鼠心肌缺血/再灌注（ischemia/reperfusion, I/R）损伤的保护作用,并从内质网应激(ERS)的角度探讨其可能的分子机制。方法：采用SD大鼠心肌I/R模型,随机分为正常对照组、模型组与药物预处理组,检测血流动力学及血清心肌肌钙蛋白T（cTnT）含量,以氯化三苯基四氮唑（TTC）和伊文思蓝双染法检测心梗面积,采用HE染色和TUNEL法分别评估心肌病理损伤和心肌细胞凋亡程度,Western blotting法检测心肌组织凋亡调节因子以及内质网应激相关蛋白的表达。结果：与I/R组相比,PQS+I/R组平均动脉压降低32.0%（P<0.05）,左室±dp/dtmax分别升高64.0%和35.0%（P<0.05）,血清cTnT含量降低53.3%（P<0.05）,坏死面积/缺血面积(AN/AAR)百分比降低65.5%（P<0.05）,心肌细胞凋亡率降低54.9%（P<0.05）;心肌组织病理损伤程度减轻;抗凋亡蛋白Bcl-2表达升高110.0%（P<0.05）,促凋亡蛋白Bax表达降低47.8%（P<0.05）,钙网蛋白(CRT)表达降低43.4%（P<0.05）,C/EBP同源蛋白（CHOP）和剪切后的caspase-12蛋白表达分别降低38.6%和23.7%（P<0.05）。结论：PQS可显著减轻大鼠心肌I/R损伤,其机制与降低I/R诱导的CRT过表达,抑制CHOP、caspase-12等内质网凋亡通路激活,从而抑制过度ERS介导的细胞凋亡有关。     

西洋参茎叶总皂苷通过抑制内质网应激减轻毒胡萝卜素诱导的心肌细胞凋亡

 目的:研究西洋参茎叶总皂苷 (PQS) 减轻毒胡萝卜素(TG) 诱导的心肌细胞凋亡的分子机制。方法:原代培养的心肌细胞分为：control组、TG组、PQS (40 mg/L、80 mg/L及160 mg/L)+TG组、牛磺酸 (40 mmol/L)+TG组、蛋白激酶R样内质网激酶 (PERK)敲低+TG组及随机双链RNA转染对照 (mock)+TG组。通过向培养液中加入1 μmol/L TG作用24 h诱导离体培养的乳大鼠心肌细胞凋亡。以RNAi方法敲低心肌细胞PERK基因。CCK-8法和流式细胞术检测细胞活性及凋亡率,Western blotting方法检测内质网应激(ERS)标志分子葡萄糖调节蛋白78 (GRP 78)、钙网蛋白 (CRT)、PERK、p-PERK、真核起始因子2α (eIF2α)、p- eIF2α、活化转录因子4 (ATF4)、C/EBP同源蛋白 (CHOP) 及凋亡相关蛋白Bcl-2、Bax的表达。结果:与对照组比较,TG孵育明显诱导细胞凋亡,降低细胞活力,上调PERK和eIF2α磷酸化以及GRP78、CRT、ATF4、CHOP及促凋亡蛋白Bax表达,降低抗凋亡蛋白Bcl-2表达 (P<0.05);与TG组比较,PQS 160 mg/L及敲低PERK均可显著降低细胞凋亡率,升高细胞活力 (P<0.05);3种不同浓度的PQS可呈剂量依赖性降低Bax蛋白表达,升高Bcl-2蛋白表达 (P<0.05),敲低PERK基因及PQS (160 mg/L) 预处理均可降低ERS分子GRP78、CRT、ATF4及CHOP表达,降低PERK及eIF2α的磷酸化水平 (P<0.05)。结论: PQS 160 mg/L 减轻ERS诱导剂TG诱导的心肌细胞凋亡,PQS预处理可以模拟PERK敲低减轻TG致心肌细胞凋亡的效应,提示PERK-eIF2α-ATF4-CHOP途径参与PQS减轻ERS相关凋亡的作用。     

西洋参茎叶总皂苷减轻大鼠心肌细胞缺氧/复氧损伤机制的研究

 目的：探讨西洋参茎叶总皂苷（PQS）减轻大鼠心肌细胞缺氧/复氧（hypoxia/reoxygenation,H/R）损伤的机制是否与抑制钙调神经磷酸酶(CaN)有关。方法：采用乳鼠心肌细胞H/R损伤模型,转染CaN质粒使其过表达或CaN抑制剂FK506干扰其表达,分为正常对照组、缺氧/复氧组、药物预处理组、CaN过表达+药物预处理组、空载pCDB质粒+药物预处理组及CaN抑制剂+药物预处理组,以流式细胞术检测细胞凋亡,按CaN测试盒步骤测定心肌细胞CaN活性,以Western blotting方法检测心肌细胞CaN表达。结果：与对照组相比,CaN过表达组心肌细胞凋亡率增加（P<005）,与PQS+H/R组比较,FK506组心肌细胞凋亡率、抗凋亡蛋白Bcl-2和促凋亡蛋白Bax表达、CaN活性及蛋白表达无显著差异（P>005）。结论：抑制CaN活性可以减轻心肌细胞H/R损伤,但联合应用FK506对H/R心肌细胞的保护作用不比单独应用PQS强,PQS减轻心肌细胞H/R损伤的机制可能与CaN途径无关。     

西洋参茎叶总皂苷对缺血/再灌注大鼠心肌线粒体膜电位及细胞凋亡的影响

 目的：研究西洋参茎叶总皂苷 (PQS) 对大鼠缺血/再灌注 (I/R) 损伤后心肌细胞凋亡的影响,并从线粒体膜电位 (ΔΨm) 及线粒体凋亡通路探讨其可能的机制。方法：健康雄性SD大鼠90只,随机分为假手术(sham)组、模型(I/R)组、PQS (200 mg·kg-1·d-1, 灌胃6周)+I/R组、环孢霉素A (CsA;10 mg·kg-1,再灌前10 min腹腔注射)组、CsA+I/R组和PQS+CsA+I/R组,各组n=15。除sham组和CsA组大鼠开胸后冠状动脉左前降支（LAD）下穿线不结扎外,其余各组大鼠常规麻醉后,结扎LAD 30 min,再灌注120 min复制I/R模型。生化分析仪测血清乳酸脱氢酶（LDH）含量,氯化三苯基四氮唑（TTC）和伊文思蓝双染法测心梗面积,原位缺口末端标记法（TUNEL）检测各组大鼠心肌细胞凋亡情况,Western blotting检测细胞凋亡相关蛋白的表达。采用JC-1作为荧光探针,用激光共聚焦显微镜和荧光酶标仪测定ΔΨm水平。结果：与sham组比较,I/R组血清LDH活性、心梗面积和心肌细胞凋亡率均显著升高（P<0.05）;与I/R组相比,PQS+I/R组、CsA+I/R组和PQS+CsA+I/R组血清LDH活性、心梗面积和心肌细胞凋亡率均显著降低（P<0.05）。Western blotting结果显示,与sham组相比,I/R组心肌Bcl-2蛋白表达量降低,Bax、胞浆cytochrome C和cleaved caspase-3升高（均P<0.05）;与I/R组相比,PQS+I/R组、CsA+I/R组和PQS+CsA+I/R组心肌Bcl-2蛋白表达量升高,Bax、胞浆cytochrome C及心肌cleaved caspase-3蛋白表达量均降低（P<0.05）。激光共聚焦显微镜观察结果示,I/R组线粒体JC-1染色后红色荧光强度减弱,荧光酶标仪测相对荧光单位（RFU） 较sham组降低（P<0.05）;PQS+I/R组、CsA+I/R组和PQS+CsA+I/R组RFU均较I/R组升高（P<0.05）。结论：PQS显著降低大鼠I/R后心肌细胞损伤,减少细胞凋亡,其机制与维持再灌注期ΔΨm稳定、抑制线粒体凋亡通路的激活有关。     

Reaction of peripheral blood lymphocytes to mitogens from the age aspect

The reaction of peripheral blood lymphocytes from healthy blood donors aged from 19 to 49 years to the mitogens phytohemagglutinin, concanavalin A (con A), and rabbit serum against human thymocytes (ATS) was investigated. A significant decrease in the proliferative response of the lymphocytes to con A was found in subjects over 30 years old. Significant negative correlation was found between the indices of the proliferative response and the age of the donors during stimulation by con A for the whole group of subjects, but for stimulation by ATS only for subjects aged 30–49 years. Analysis of the intensity of incorporation of [³H]-thymidine shows a decrease with age in the proportion of cells with intensively labeled nuclei and an increase in the number of cells with weak labeling of the nucleus both with a decrease in the indices of the proliferative response with age and also when no significant differences were found in these indices.Laboratory of General Pathophysiology, Institute of Psychiatry, Academy of Medical Sciences of the USSR, Moscow. (Presented by Academician of the Academy of Medical Sciences of the USSR A. M. Chernukh.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 84, No. 11, pp. 600–602, November, 1977.     

Tumor necrosis factor receptor p55 mediates deletion of peripheral cytotoxic T lymphocytes in vivo

Cellular death of activated lymphocytes down-regulates immune responses and is involved in maintaining self tolerance. Signals associated with ligation of the membrane molecule Fas lead to lymphocyte apoptosis, but additional, Fasindependent mechanisms have been postulated. Here, we show a marked expansion and prolonged persistence of functional activated cytotoxic T cells in mice lacking the tumor necrosis factor (TNF) receptor p55. In the absence of this receptor, peripheral lymphocyte apoptosis was significantly reduced in vivo. The prolonged thymocyte survival was associated with functional anergy, since the T cells no longer proliferated in vitro when stimulated with peptide antigen. However, specific cytotoxic effector function was easily detected in vitro. We conclude that the TNF receptor p55 is involved in peripheral T cell deletion in vivo.     

Bursa-dependent subpopulations of peripheral B lymphocytes in chicken blood

By selective labeling of juvenile chicken bursal cells with colloidal fluorescein isothiocyanate in situ, the emigration rate of bursal lymphocytes to the periphery was estimated at approximately 0.84% and 0.96% of the peripheral blood lymphocyte (PBL) and splenic B cell pool per hour, respectively. Emigrant bursal cells were found primarily in blood and spleen, with very small numbers migrating to thymus, bone marrow, and gut-associated lymphoid tissues. Emigrant bursal cells expressed high levels of both major histocompatibility complex class II antigen and the Ov alloantigen, a phenotype found on a population comprising approximately 4% of bursal cells from which the bursal emigrants may be derived. Surgical bursectomy at 3 weeks of age revealed that peripheral blood B cells could be divided into three distinct populations. Specifically, 60% of the peripheral blood B cells were short lived with a half-life of about 30 h in the blood. These cells accounted for the great majority of emigrants from the bursa to the peripheral blood. Approximately 35 % of PBL B cells had a half-life of 12 days following bursectomy and comprised cells which did not divide in the periphery. Consequently, we propose that physiological differences between this population and the majority of bursal emigrants are established intrabursally. The remaining PBL B cells, whose relative proportion increases with age from about 5 % of PBL B cells at 2-3 weeks of age, are short lived and are being continually produced from (a) post-bursal site(s) of B cell production.     

The influence of substance P on the proliferation of peripheral blood lymphocytes from normal individuals and birch pollen-allergic patients

We have studied the influence of substance P (SP) on the proliferative response of concanavalin A (ConA)-activated peripheral blood lymphocytes from 16 birch pollen-allergic patients, sampled before and during the pollen season, and from 15 normal individuals. The median response to ConA 3 micrograms/ml in the presence of SP 10(-11)-10(-6) M, was in most instances within +/- 10% of the control value for cells from both healthy and atopic individuals. However, the individual differences were considerable. Analysis of the proliferative responses to ConA of the cells from the allergic patients sampled before and during season, revealed higher responses in the presence of 10(-6) than of 10(-7) M SP. This was in contrast to the findings in the normal individuals: only half of their cells showed such increased responses. This difference in response frequency was statistically significant between allergic patients before season and normal individuals (P less than 0.05) and between allergic patients during season and normal individuals (P less than 0.01). The difference in proliferation rate in the SP concentration interval, 10(-6) to 10(-7) M, for the cells from allergic patients, sampled both before and during season, was significantly different from the cells from healthy individuals (P less than 0.03 and P less than 0.001 respectively). The cells sampled from four allergic subjects during the birch pollen season showed a more profoundly decreased response to ConA in the presence of SP 10(-8) M, compared with their cells sampled before season. Such responses were never seen with cells sampled before season and with cells from normal individuals. The results suggest an involvement of SP in the immunoregulation, particularly in patients exhibiting allergic reactions to birch pollen.     

CRF患者红细胞促外周血淋巴细胞增殖反应的实验研究

目的 :探讨慢性肾功能衰竭 (CRF)患者红细胞 (RBC)促外周血淋巴细胞 (PBL)增殖能力 ,了解CRF患者RBC免疫调控功能状况。方法 :CRF患者 (检测组 )和正常人 (对照组 )的RBC为效应细胞 ,正常人淋巴细胞为靶细胞 ,采用四甲基偶氮唑蓝 (MTT)比色法 ,检测红细胞调控淋巴细胞增殖反应的能力。结果 :健康人RBC促PBL增殖的作用明显 ,其增殖率为5 7 3%± 10 2 % ,而CRF患者RBC促PBL增殖作用较健康人明显低下 ,增殖率为 32 7%± 7 8% (P <0 0 0 1) ,透析后有较明显的改善 ,增殖率为 4 0 6 %± 11 6 % ,但仍低于健康人。结论 :CRF患者红细胞免疫调控功能低下。     

Phenotype of peripheral blood lymphocytes and serum immunoglobulin concentration in patients with early rheumatoid arthritis

We studied the phenotype of peripheral blood lymphocytes and serum immunoglobulin concentration in patients with early rheumatoid arthritis and rheumatoid arthritis of more than 12 months duration. Subpopulations of CDIgM⁺, CD25⁺, and HLA-DR⁺ lymphocytes and IgA concentration differed in these groups of patients with rheumatoid arthritis. The count of lymphocytes carrying CDIgM⁺ and HLA-DR⁺ receptors correlated with activity of rheumatoid arthritis. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 141, No. 1, pp. 92–94, January, 2006     

Apoptosis of T lymphocytes isolated from peripheral blood of patients with acute exacerbation of chronic obstructive pulmonary disease

Purpose

Chronic obstructive pulmonary disease (COPD) is characterized by chronic inflammation of the airways and progressive destruction of lung parenchyma. Apoptosis is critical for the maintenance of normal tissue homeostasis and is in equilibrium with proliferation and differentiation. This study was undertaken to investigate relationship between apoptosis of peripheral blood lymphocytes during exacerbation of COPD and inflammatory response that characterizes this condition.

Materials and Methods

Seventeen patients with COPD exacerbation, 21 stable COPD, and 12 control subjects were included. T lymphocytes were isolated from peripheral blood using MACS. Apoptosis of T lymphocytes was assessed with FACS using annexin V and 7-aminoactinomycin. Serum levels of interleukin (IL)-6, IL-8 and tumor necrosis factor (TNF)-α were determined by an immunoassay technique.

Results

There was significantly increased percentage of apoptotic lymphocytes, CD 4⁺, and CD 8⁺ T cells in the peripheral blood of patients with exacerbation of COPD compared with stable COPD. Serum levels of IL-6, IL-8, and TNF-α were significantly increased in patients with exacerbation of COPD compared with stable COPD. Only TNF-α presented a positive correlation with apoptotic lymphocytes in patients with exacerbation of COPD.

Conclusion

Increased apoptotic lymphocytes may be associated with upregulation of TNF-α in the peripheral blood of patients with acute exacerbation of COPD.     

Sensitivity of peripheral blood lymphocytes from breast cancer patients to glucocorticoids

We measured DNA-dependent RNA polymerase activity, calculated hormone-binding sites and intracellular association constant under conditions of phytohemagglutinin stimulation and dexamethasone treatment, and studied the interaction of cells with the glucocorticoid covalently bound to an inert carrier not penetrating into cells. Reduced sensitivity of peripheral blood lymphocytes from breast cancer patients to glucocorticoids was demonstrated. The biological mechanisms for hormonal regulation are reviewed here. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 141, No. 1, pp. 80–84, January, 2006     

Diagnosis and assessment of preclinical and clinical autoimmune encephalomyelitis using peripheral blood lymphocyte TCR

In organ-specific autoimmune diseases, T cells involved in the disease development bear a particular type of TCR and infiltrate the target organ predominantly. However, it is difficult to identify disease-inducing T cells in peripheral blood lymphocytes (PBL) because such T cells are very few in number in a large pool of unrelated T cells. In the present study, we demonstrate that CDR3 spectratyping can identify experimental autoimmune encephalomyelitis (EAE)-specific patterns (oligoclonal expansion of Vβ8.2 with the shortest CDR3) in PBL at the preclinical and clinical stages of acute EAE. Analysis of nucleotide and predicted amino acid sequences of Vβ8.2 CDR3 of spectratype-derived clones revealed that CASSDSSYEQYFGPG, which is one of the representative sequences of encephalitogenic T cell clones, constituted the predominant population in both PBL and spinal cord T cells. In chronic relapsing EAE, the EAE-specific spectratype pattern in PBL was observed during the 1st and 2nd attacks, but not at the remission and full recovery stage. These findings indicate that the spectratyping pattern in PBL reflects the disease activity of acute and chronic relapsing EAE. Thus, CDR3 spectratyping using PBL can be used for diagnosis and assessment of T cell-mediated autoimmune diseases and is applicable to human autoimmune diseases.