A Comparative Study on the Effect of BCG-PSN and Thymopeptides on T-lymphocyte Subsets of Normal and Immunosuppressed Mice

To compare the effects of polysaccharide nucleic acid fraction of bacillus calmette guerin (BCG PSN) and thymopeptides on T lymphocytes of normal and immunosuppressed mice, CD4 + and CD8 + T lymphocyte subsets of single nucleic cell in thymus, spleen and peripheral blood were detected successively by flow cytometry after application of BCG PSN and thymopeptides. Meanwhile, CD4 +/CD8 + ratio was also calculated. The results showed that both BCG PSN and thymopeptides could decrease the proportion of CD4 + CD8 + T lymphocyte subsets in the thymus, at the same time increase CD4 + T lymphocyte, CD8 +T lymphocyte proportion in the three tissues. The fluctuation in amplitude was greater in thymopeptides group than that in BCG PSN group. It is concluded that acting location of thymopeptides is in thymus, its stimulating action is stronger than that of BCG PSN, while BCG PSN not only accelerates the differentiation in thymus, but also has some direct stimulation to peripheral CD4 +T lymphocytes, and can maintain CD4 +/CD8 + ratio within normal range. So, BCG PSN is safer.

Distribution of natural killer cells and T-lymphocyte subsets in peripheral blood,gallbladder cancer and surrounding tissue

BACKGROUND: The patient with malignant tumor always show immunologic function drawback and ingravescent with tumor development, especially in the aspect of cell-mediated immunity. This study was undertaken to define the relationship between the immune function of local cells and cancer development by investigating the distribution of natural killer (NK) cells and T-lymphocyte subsets in peripheral blood, the cancer tissue and the tissue surrounding gallbladder carcinoma. METHODS: The numbers of CD4(+) and CD8(+) T-lymphocytes and NK cells were measured by flow cytometry in samples taken from gallbladder cancer tissue, the surrounding tissues and peripheral blood of 38 patients, and compared with the numbers in the peripheral blood and gallbladder tissue of 30 patients with cholecystitis as controls. RESULTS: The numbers of CD4(+) and CD8(+) T-cells and NK cells in gallbladder cancer tissues were significantly higher than those in the surrounding tissue and gallbladder with gallstone. However, the ratio of CD4(+)/CD8(+) was lower in the cancer tissue than that in the surrounding tissue and tissue from gallbladders with gallstones. The distribution of CD4(+) and CD8(+) T-cells and NK cells in mucous membrane of cholecystitis gallbladder and that in the tissue surrounding gallbladder cancer were significantly different. CONCLUSIONS: Disproportionate and imbalanced distribution of NK cells and subsets of T-lymphocytes occurs in the mucous membrane proper of gallbladder cancer and surrounding tissue. Although gallbladder cancer tissue has higher expressions of CD4(+), CD8(+) and NK cells, the immune function is low or in an inhibited state. In gallbladder cancer immunization therapy, local cellular immunological function should be enhanced and the protective barrier improved.

Effects of Intraoperative Administration of Dexmedetomidine on the Percentage of T-Lymphocyte Subsets and Natural Killer Cells in Patients with Colorectal Cancer

Study Objective: To observe the effect of dexmedetomidine (DEX) on T-lymphocyte subsets and natural killer (NK) cells in the peripheral blood of perioperative patients with colorectal cancer. Design: A random double-blind control clinical study. Setting: A university hospital. Patients: Forty patients with colorectal cancer, ASA I-П. Interventions: All patients were randomly divided into a DEX group (n = 20) and a control group (n = 20). Before induction of anesthesia, epidural catheters were placed in the L1-L2 or T12-L1 intervertebral spaces. The DEX group received 1 μg/kg of DEX (200 μg/50 ml) intravenously for 15 min prior to the surgery, which was then infused at a rate of 0.5 μg/kg/h until 30 min before the end of the surgery. The control group received an intravenous infusion of saline (50 ml) instead of DEX during the same periods as the DEX group. All patients received routine anesthesia and postoperative analgesia. Measurements: Blood samples from all patients were collected at the following time points: before anesthesia (T0), 24 h after surgery (T1), 48 h after surgery (T2) and 72 h after surgery (T3). Changes in T-lymphocyte subsets (CD3+, CD4+, CD8+, CD4+/CD8+) and NK cells were determined by flow cytometry. Main Results: Compared with the control group, the percentages of CD3+ and CD4+ cells and the CD4+/CD8+ ratio in the DEX group increased significantly from T1 to T3

Effect of cytotoxic T-lymphocyte antigen-4,TNF-alpha polymorphisms on osteosarcoma: evidences from a meta-analysis

Objective： Previous studies have investigated the role of cytotoxic T-lymphocyte antigen-4 （CTLA-4） and tumor necrosis factor-alpha （TNF-a） in carcinogenesis of osteosarcoma, but their results were inconsistent. We aimed to clarify the associations between CTLA-4, TNF-a polymorphism and osteosarcoma risk by using meta-analysis. Methods： We searched relevant studies without language restriction in PubMed, EMbase, Cochrane Library, Google Scholar databases, Chinese National Knowledge Infrastructure （CNKI） and conference literature in humans published prior to March 2013. The strengths of the associations between genetic variants and osteosarcoma risk were estimated by odds ratio （OR） with 95% confidence interval （95% CI）. Results： A total of seven studies with 1,198 osteosarcoma patients and 1,493 controls were selected. Four studies were eligible for CTLA-4 （1,003 osteosarcoma and 1,162 controls）, and three studies for TNF-a （195 osteosarcoma and 331 controls）. Pooled results showed that rs231775 polymorphism of CTLA-4 was associated with osteosarcoma risk （GG vs. AA： OR=1.63, 95% CI=1.24-2.13; GG ＋ GA vs. AA： OR=1.56, 95% CI=1.21-2.01; AA ＋ GA vs. GG： OR=0.83, 95% CI=0.71-0.97; G vs. A： OR=1.21, 95% CI=1.08-1.36）. No significant heterogeneity was observed across the studies. No significant associations were found between rs5742909 polymorphism of CTLA-4 or rs1800629 polymorphism of TNF-a and osteosarcoma risk. Conclusions： These results suggest that the rs231775 polymorphism of CTLA-4 may play an important role in carcinogenesis of osteosarcoma.

Analysis of T-lymphocyte subtypes of the peripheral blood and skindelayed-type hypersensitivity in patients with hyper-IgE syndrome

Objective: To study the function of ce ll ular immunity in patients with hyper-IgE syndrome (HIE). Methods: T-lymphocyte subtypes of the peripheral blood and cutaneous delayed-typ e hypersensitivity (DTH) response to two recall antigens, tetanus toxoid (TT) an d purified protein derivative(PPD), were measured in five patients with HIE and 15 healthy controls, respectively. Results: The CD4+ cell cou nts in HIE group were significantly lower than those in control group (P<0. 01). In contrast, CD8+ cells were significantly higher than those in the contro l (P<0.01). The induration sizes of DTH response to two recall antigens wer e smaller in HIE group than those in the control group (P<0.01). Co nclusion: There was an immunologic dysfunction of T lymphocytes in the p atients with HIE.

Effects of Heat Clearing and Stasis Resolving Method on PTS,Inflammatory Factors and T-lymphocyte Subsets in Peripheral Blood of URM Patients with Suppressed Internal Heat

[Objectives]To explore the effects of heat clearing and stasis resolving method on prethrombotic state,inflammatory factors and T-lymphocyte subsets in peripheral blood of unexplained recurrent miscarriage(URM)patients with suppressed internal heat.[Methods]Thirty cases of URM patients with suppressed internal heat and 30 normal women were collected,and characteristics of changes in peripheral serum D-dimer(D-D),fibrin degradation product(FDP),fibrinogen(FIB),IL-6,IL-10 and TNF-α,CD,CD,CD,CD,CDlevels were detected.URM patients were treated with traditional Chinese medicine for clearing heat and resolving blood stasis for 3 menstrual cycles,and the changes of indicators before and after treatment were observed.[Results]Compared with normal women,the peripheral serum levels of D-D,IL-6,TNF-αand CDin URM patients with suppressed internal heat were increased(P<0.05),while the IL-10 lymphocyte level was significantly decreased(P<0.05);compared with that before treatment,the contents of D-D,IL-6,TNF-αand CDdecreased after 3 menstrual cycles(P<0.05),while the contents of IL-10 and CDT lymphocytes increased significantly(P<0.05).[Conclusions]The heat clearing heat and stasis resolving method can effectively improve the prethrombotic state of URM,and the action mechanism may be related to the regulation of immune and peripheral blood inflammatory factors.

Interferon-γpriming enhances the therapeutic effects of menstrual blood-derived stromal cells in a mouse liver ischemia-reperfusion model

BACKGROUND Mesenchymal stem cells(MSCs)have been used in liver transplantation and have certain effects in alleviating liver ischemia-reperfusion injury(IRI)and regulating immune rejection.However,some studies have indicated that the effects of MSCs are not very significant.Therefore,approaches that enable MSCs to exert significant and stable therapeutic effects are worth further study.AIM To enhance the therapeutic potential of human menstrual blood-derived stromal cells(MenSCs)in the mouse liver ischemia-reperfusion(I/R)model via interferon-γ(IFN-γ)priming.METHODS Apoptosis was analyzed by flow cytometry to evaluate the safety of IFN-γpriming,and indoleamine 2,3-dioxygenase(IDO)levels were measured by quantitative real-time reverse transcription polymerase chain reaction,western blotting,and ELISA to evaluate the efficacy of IFN-γpriming.In vivo,the liver I/R model was established in male C57/BL mice,hematoxylin and eosin and TUNEL staining was performed and serum liver enzyme levels were measured to assess the degree of liver injury,and regulatory T cell(Treg)numbers in spleens were determined by flow cytometry to assess immune tolerance potential.Metabolomics analysis was conducted to elucidate the potential mechanism underlying the regulatory effects of primed MenSCs.In vitro,we established a hypoxia/reoxygenation(H/R)model and analyzed apoptosis by flow cytometry to investigate the mechanism through which primed MenSCs inhibit apoptosis.Transmission electron microscopy,western blotting,and immunofluorescence were used to analyze autophagy levels.RESULTS IFN-γ-primed MenSCs secreted higher levels of IDO,attenuated liver injury,and increased Treg numbers in the mouse spleens to greater degrees than untreated MenSCs.Metabolomics and autophagy analyses proved that primed MenSCs more strongly induced autophagy in the mouse livers.In the H/R model,autophagy inhibitors increased the level of H/R-induced apoptosis,indicating that autophagy exerted protective effects.In addition,primed MenSCs decreased the level of H/R-induced apoptosis via IDO and autophagy.Further rescue experiments proved that IDO enhanced the protective autophagy by inhibiting the mammalian target of rapamycin(mTOR)pathway and activating the AMPK pathway.CONCLUSION IFN-γ-primed MenSCs exerted better therapeutic effects in the liver I/R model by secreting higher IDO levels.MenSCs and IDO activated the AMPK-mTOR-autophagy axis to reduce IRI,and IDO increased Treg numbers in the spleen and enhanced the MenSC-mediated induction of immune tolerance.Our study suggests that IFN-γ-primed MenSCs may be a novel and superior MSC product for liver transplantation in the future.

外周血CD8^(+)T淋巴细胞亚群检测对过敏性鼻炎临床疗效的评估价值

目的探讨外周血CD8^(+)T淋巴细胞亚群检测对过敏性鼻炎(AR)临床疗效的评估价值。方法前瞻性选取2021年3月~2022年2月温州市中西医结合医院耳鼻咽喉科门诊治疗的120例AR患者和40例健康志愿者,分别设为研究组和对照组,比较两组外周血CD8^(+)T细胞、CD8^(+)CD28^(+)T细胞、CD8^(+)CD28-T细胞比率;研究组接受鼻用激素联合口服抗组胺药物治疗方案,根据治疗效果分为有效组(n=107)和无效组(n=13),比较有效组和无效组治疗前CD8^(+)T细胞、CD8^(+)CD28^(+)T细胞、CD8^(+)CD28-T细胞比率;Pearson相关性分析法分析外周血CD8^(+)T淋巴细胞亚群水平与鼻部症状总评分(totalnose symptomscore,TNSS)的相关性;以受试者工作特征(ROC)曲线分析治疗前CD8^(+)T淋巴细胞亚群对AR临床疗效的预测价值。结果研究组治疗前外周血CD8^(+)CD28^(+)T细胞比率高于对照组,CD8^(+)CD28-T细胞比率低于对照组(P<0.05)。研究组总有效率为89.17%;有效组治疗前CD8^(+)CD28^(+)T细胞比率低于无效组,CD8^(+)CD28-T细胞比率高于无效组(P<0.05);Pearson相关性分析显示,CD8^(+)CD28^(+)T细胞比率与TNSS评分呈正相关(r=0.324,P=0.006),CD8^(+)CD28-T细胞比率与TNSS评分呈负相关(r=-0.541,P=0.000)。ROC曲线结果显示,CD8^(+)T细胞、CD8^(+)CD28^(+)T细胞、CD8^(+)CD28-T细胞比率评估AR治疗无效的曲线下面积(AUC)(95%CI)分别为0.647(0.555~0.732)、0.683(0.592~0.765)、0.911(0.845~0.955),其中CD8^(+)CD28-T细胞的评估效能优于CD8^(+)T细胞、CD8^(+)CD28^(+)T细胞比率(P<0.05)。结论AR患者外周血CD8^(+)CD28^(+)T细胞比率升高,CD8^(+)CD28-T细胞比率降低,CD8^(+)T细胞比率相对稳定,治疗前CD8^(+)CD28-T细胞比率可作为预测AR患者的临床疗效的参考指标。

Relationship between Circulating Plasma Galectin-3 Levels and T-Cell Activation during Cervical Cancer Chemotherapy

Objective: Despite the existence of several therapeutic strategies, the management of cervical cancer remains challenging. Our region has very little data on the interaction between the immune system and the clinical response to chemotherapy. This work examines plasma levels of galectin-3 (Gal-3) and percentages of activated T cells in patients with cervical cancer treated with chemotherapy and investigates if there is a relationship between the rates of these two elements. Methods: We compared data from 37 patients with cervical cancer undergoing chemotherapy and 42 controls with normal cervical cytology. Plasma Gal-3 concentrations were assessed by ELISA and expression of activation markers by T cells (CD69 and HLA-DR) was assessed by flow cytometry at three different time points during chemotherapy. Results: Our results showed that patients had a significantly higher concentration of Gal-3 compared to controls (4.025 vs. 1.340, p 0.001), similarly, they had a significantly high percentage of activated lymphocytes (2.610 vs. 0.731;p 0.0001). According to the response to treatment, patients with no response to treatment had a lower concentration of circulating Gal-3 but had approximately the same percentage of activated CD4 and CD8 lymphocytes as patients with a partial or total response. In addition, we found a positive correlation between the Gal-3 level and CD4 T cells expressing the activation marker CD69 (p 0.05;rho = 0.44). Conclusion: In conclusion, our results show that there would be a relationship between circulating galectin-3 and the percentage of peripheral CD4+CD69+ cells in cervical cancer.

清肝冲剂对免疫性肝炎模型小鼠的保护作用及免疫调节作用

观察了清肝冲剂对小鼠免疫性肝损伤及免疫功能的影响。结果表明，清肝冲剂灌胃６４ｇ／（ｋｇ．ｄ）对ＢＣＧ（卡介苗）加ＬＰＳ（脂多糖）诱导的小鼠免疫性肝炎有明显保护作用，可降低肝损伤小鼠血清丙氨酸氨基转换酶（ＡＬＴ）和门冬氨酸氨基转换酶（ＡＳＴ）活性。清肝冲剂１６ｇ／（ｋｇ．ｄ）、３２ｇ／（ｋｇ．ｄ）、６４ｇ／（ｋｇ．ｄ）能提高免疫功能低下小鼠腹腔巨噬细胞的吞噬功能，吞噬百分率和吞噬指数均明显升高；能明显促进刀豆素Ａ（ＣｏｎＡ）诱导的小鼠体内淋巴细胞转化，淋巴细胞转化率显著升高；清肝冲剂增加鸡红细胞（ＣＲＢＣ）致敏小鼠溶血素抗体的生成，并能增加ＣＲＢＣ致敏小鼠脾指数。结果提示，清肝冲剂对小鼠免疫性肝损伤有保护作用，对小鼠细胞免疫和体液免疫均有免疫调节作用。

慢性乙型肝炎患者外周血树突状细胞PD-L1表达的相关研究

目的:探讨慢性乙型肝炎(CHB)患者外周血树突状细胞(DCs)的PD-L1表达水平,及其对DCs刺激T细胞增殖的影响。方法:采集22例慢性HBeAg阳性CHB患者、8例急性恢复期乙型肝炎(AHB)患者和10例健康人(NC)的外周血,分离单个核细胞(PBMC)并诱导DCs的增殖和活化。相对荧光定量PCR测定DCs的PD-L1、PD-L2分子mRNA水平,流式细胞术测定DCs表面PD-L1和PD-L2的表达率。同时,测定外周血DCs刺激同种异体反应性T细胞增殖的能力,及抗PD-L1单抗对CHB组DCs刺激功能的影响。结果:体外成功诱导了外周血DCs的增殖和成熟。与NC组相比较,CHB组和AHB组患者外周血DCs内PD-L1的mRNA水平均显著增高(P均<0.05),但CHB组和AHB组间差异无统计学意义。CHB组患者外周血DCs表面PD-L1表达率较AHB组、NC组均有明显增高(P均<0.01)。而外周血DCs的PD-L2 mRNA水平及PD-L2在DCs表面的表达率在各实验组之间无明显差异(P均>0.05)。CHB组外周血DCs刺激T细胞增殖的功能较AHB组和NC组明显降低(P均<0.01),抗PD-L1抗体能显著增强CHB患者DCs刺激T细胞反应的能力。CHB患者外周血DCs表面PD-L1表达率与血清HBV病毒载量成正相关(r=0.34,P=0.007)。结论:外周血DCs表面PD-L1表达水平在慢性HBeAg阳性乙肝患者明显升高,抗PD-L1抗体可有效增强CHB患者外周血DCs刺激T细胞增殖反应的能力,表明PD-L1的高表达可能与T细胞免疫反应低下和HBV病毒高复制密切相关。

肝癌患者集束电极射频消融治疗前后T淋巴细胞及红细胞免疫功能的研究

目的 研究肝癌患者接受射频消融治疗前后机体T淋巴细胞及红细胞免疫功能的变化。方法 分析 12 0例肝细胞肝癌患者射频消融治疗前和治疗后 3d ,7d和 14d外周血淋巴细胞亚群(T 3 ,T 4,T 8,T4/T 8)和红细胞C 3b受体花环 (RBC C3bR )及免疫复合物花环 (RBC ICR )形成率。结果 所有肝癌患者在射频消融治疗后 7d和 14d ,T 3细胞 ,T 4细胞 ,T 4/T 8及RBC C 3bR ,RBC ICR形成率均明显高于治疗前 (P <0 .0 5 )。结论 采用射频消融治疗肝癌 ,可以改善患者的细胞免疫状态。

隐孔菌多糖成分对致敏大鼠气道高反应性和炎症细胞的影响

目的 :研究隐孔菌多糖成分对大鼠气道高反应性的影响 ,初步探索其作用机制。方法 :采用卵白蛋白致敏雾化吸入攻击制备大鼠哮喘模型 ,隐孔菌多糖成分 A、B(5 mg/ kg、2 0 mg/ kg)和酮替芬 (5 mg/ kg)及溶媒对照(等量生理盐水 )灌胃 10 d。用甲酰胆碱激发气道高反应性后 ,行支气管肺泡灌洗和腹腔灌洗 ,计数支气管肺泡灌洗液中细胞总数并分类 ,和腹腔灌洗液中脱颗粒肥大细胞及白细胞分类。结果 :隐孔菌多糖成分 A、B均能明显抑制致敏大鼠抗原攻击后气道阻力的增加及肺顺应性的下降 ;减少支气管肺泡灌洗液中白细胞总数 ,降低嗜酸细胞的数目 ,以多糖 B作用更明显 ;多糖 A和多糖 B也明显抑制腹腔肥大细胞脱颗粒及腹腔嗜酸性粒细胞的渗出。结论 :隐孔菌多糖 A、B成分抑制大鼠的气道高反应性 ,其作用可能与稳定肥大细胞膜、抑制嗜酸细胞炎症和趋化有关。

灸刺对实验性类风湿性关节炎大鼠红细胞免疫功能和血浆皮质醇影响的观察

本文对采用Ｆｒｅｕｎｄ氏完全佐剂复制的实验性类风湿性关节炎大鼠施以麦粒灸，并加用皮肤针在关节肿胀部位叩刺出血，每日１次，治疗２１天。结果：灸刺组ＲＢＣ·Ｃ３ｂＲＲ显著高于模型对照组、激素治疗组及正常组（Ｐ＜０．０１）；其血浆皮质醇也高于模型组及正常组（Ｐ＜０．０５及＜０．０１）；关节病变减轻，全身一般情况亦近似于正常组，而明显优于激素治疗组。

问号钩端螺旋体lipL21基因改建及其表达产物免疫原性变化和定位

目的：改建问号钩端螺旋体（简称钩体）lipL21基因核苷酸序列以提高表达产量并了解基因改建前后表达产物免疫原性变化，确定外膜脂蛋白LipL21在钩体表面的定位。方法：参照大肠杆菌偏爱密码子设计，合成lipL21基因核苷酸序列并构建其原核表达系统。采用SDS—PAGE和BioRad凝胶图象分析系统检测改建前后lipL21基因表达量变化。采用钩体TR／PatocI抗血清为一抗的Westernblot鉴定改建前后两种目的重组蛋白rLipL2ls的免疫反应性，显微镜凝集试验（MAT）比较改建前后rLipL21s抗血清对不同钩体血清群的交叉免疫凝集效价变化。应用免疫电镜技术对LipL21进行定位。结果：改建前后lipL21基因表达量分别为细菌总蛋白的8．5％和46．5％。两种rLipL21均能与TR／PatocI抗血清发生免疫结合反应，免疫家兔后均能产生特异性抗体。两种rLipL21兔抗血清对我国15群15型钩体参考标准株MAT效价相近，均为1：80～1：320。LipL21位于钩体外膜表面。结论：LipL21是钩体表面抗原。改建后的lipL21基因可明显提高原核表达产量，其产物可保持良好的抗原性和免疫反应性，其抗体具有广泛的交叉免疫凝集活性。

血清Ⅳ型胶原测定对慢性肝病中医证型的诊断作用及临床意义

采用放射免疫法测定160例慢性肝病患者血清肝纤维化标志物Ⅳ型胶原（Ⅳ-C），以探讨在血清中Ⅳ-C的质量浓度ρⅣ-C与中医证型的关系及临床意义。结果显示，各中医证型组的ρⅣ-C均较对照组明显升高，其升高程度依次为肝郁脾虚＜瘀血阻络＜肝肾阴虚＜湿热中阻＜脾肾阳虚。统计分析表明，湿热中阻和脾肾阳虚组ρⅣ-C的升高与肝郁脾虚组比较存在显著性差异（P＜0．01）。提示：（1）测定ρⅣ-C对慢性肝病中医分型有一定指导意义；（2）肝纤维化在肝郁脾虚阶段已启动，但相对静止；（3）湿热中阻和脾肾阳虚是肝纤维化的活跃期，后者的病情尤其严重。

补肾健脾方药免疫药理作用比较

采用目前常用的醋酸氢化可的松致肾虚模型和大黄致脾虚模型，比较了传统经典方剂肾气汤（丸）和四君子汤对模型动物的淋巴细胞转化率、ＮＫ细胞活性、白介素１（ＩＬ－１）和溶血抗体产生能力的作用。结果表明，肾虚和脾虚模型动物免疫功能皆有显著下降（Ｐ＜０．０１，Ｐ＜０．０５），肾虚尤为明显。肾气汤对肾虚和脾虚模型均有明显的治疗作用（Ｐ＜０．０５），而四君子汤仅对脾虚模型有治疗作用。推测两种模型的成因和两种方剂的作用机理有所不同，同时提示如果在脾虚健脾治疗无效时。

芪加合剂对荷瘤小鼠体液免疫功能的影响

目的 研究芪加合剂对荷瘤 (Lewis肺癌 )小鼠免疫功能的影响。方法 检测脾细胞产生IgM、血清中IgG含量和鼠腹腔巨噬细胞产生肿瘤坏死因子 (TNF) ,以及鼠外周血象的测定。结果 芪加合剂 (13 75～ 2 7 5 g·kg-1)可使荷瘤小鼠低下的IgM、血清中IgG含量恢复至接近正常水平 ,可促进荷瘤小鼠腹腔巨噬细胞产生TNF ;还可使荷瘤小鼠低下的红细胞数、血红蛋白量、白细胞数和血小板数等血常规指标恢复至接近正常水平。结论 芪加合剂抑制肿瘤作用可能与其提高荷瘤小鼠的免疫功能有关。

大肠杆菌LTB与霍乱弧菌CTB基因的克隆和表达及鉴定

目的 :克隆大肠杆菌不耐热肠毒素 B亚单位 (L TB)及霍乱弧菌肠毒素 B亚单位 (CTB)基因 ,构建其表达载体。方法 :采用高保真 PCR分别从 E.coli4 4 815株与 V.cholerae东 74株基因组 DNA中扩增 L TB与 CTB基因 ,T- A克隆后测定核苷酸序列。分别构建 p ET32 a的 LTB及 CTB表达载体 ,在 E.coli BL2 1DE3宿主菌中用不同浓度的 IPTG诱导表达。采用 SDS- PAGE及 GM1- EL ISA鉴定表达产物。结果 :所克隆的 LTB和 CTB基因与报道的相应核苷酸序列同源性分别为 99.12 %～ 99.71%和 98.5 4 %～ 99.4 2 % ,氨基酸序列同源性分别高达 97.5 8%～99.19%和 96.77%～ 99.19%。p ET32 a- L TB- BL2 1DE3和 p ET32 a- CTB- BL2 1DE3系统表达的融合蛋白产量分别占细菌总蛋白的 30 %和 10 %左右。GM1- EL ISA结果证实 LTB及 CTB融合蛋白均能与牛 GM1结合。结论 :本研究成功地构建了 LTB与 CTB表达系统 ,所表达的 L TB与 CTB融合蛋白具有粘膜免疫佐剂活性。