目的 探讨联合益生菌根除幽门螺杆菌(H.pylori)对胃蛋白酶原(PG)Ⅰ和Ⅱ、胃泌素-17(G-17)的影响.方法 选取广州新海医院2020年12月至2021年10月H.pylori阳性的慢性胃炎患者137例,随机分为四联疗法和联合益生菌疗法两个治疗组,疗程均为2周.选取健康体检者120名作为对照组.治疗结束后4周,通过14C尿素呼气试验评估根除疗效,采用ELISA检测各组治疗前后、不同程度慢性胃炎血清中PG Ⅰ、PGⅡ、PG Ⅰ/Ⅱ比值(PGR)及G-17水平.结果 四联疗法组与联合益生菌疗法组H.pylori根除率比较中,按意向分析(ITT),联合益生菌疗法组明显高于四联疗法组(86.76％ vs 72.46％,P＜0.05);而按符合方案分析(PP),两组之间差异无统计学意义(92.19％ vs 84.75％,P＞0.05).治疗前,四联疗法组、联合益生菌疗法组、H.pylori阳性健康对照组与H.pylori阴性健康对照组PG Ⅰ、PGⅡ、PGR及PG-17水平比较,有显著性差异(P＜0.05).与治疗前比较,治疗后四联疗法和联合益生菌疗法组血清PG Ⅰ、PGⅡ及G-17的水平明显下降,PGR水平明显升高(P＜0.05),且联合益生菌疗法组比四联疗法组变化更明显(P＜0.05).随着慢性胃炎炎症程度的逐渐加重,PG Ⅰ、PGⅡ及G-17的水平显著升高(P＜0.05),而血清PGR水平无显著变化(P＞0.05).治疗后,与H.pylori阳性患者比较,H.pylori阴性患者的血清PG Ⅰ、PGⅡ及G-17的水平显著下降,PGR水平显著升高(P＜0.05),且联合益生菌疗法组比四联疗法组的变化更明显,差异有统计学意义(P＜0.05).结论 四联疗法联合益生菌治疗H.pylori慢性胃炎,不仅能提高H.pylori的根除率,而且能降低血清PG Ⅰ、PGⅡ、G-17水平,提高血清PGR水平.PG Ⅰ、PGⅡ、PGR及G-17有望成为联合益生菌根除治疗H.pylori新的临床评价标准.

目的 探讨联合益生菌根除幽门螺杆菌(Helicobacter pylori,H.pylori)对IL-8、IL-10、IL-17的调节效应.方法 将116例H.pylori阳性的慢性胃炎患者随机分为两个治疗组:标准三联疗法组、联合益生菌疗法组,疗程均为2周.选取健康体检者30名作为健康对照组.行HE染色评估胃黏膜的炎症程度,治疗结束4周后复查14 C尿素呼气试验评估根除疗效,采用ELISA检测各组治疗前后、不同程度慢性胃炎患者血清中IL-8、IL-10及IL-17水平.结果 两组H.pylori根除率比较中,按意愿(ITT)及方案(PP)分析,联合益生菌疗法组(84.48％、90.74％)明显高于标准三联疗法组(60.34％、76.09％)(P均<0.05).治疗前,治疗组与健康对照组血清IL-8、IL-10与IL-17水平比较,差异有统计学意义(P均<0.05).与治疗前比较,两治疗组治疗后血清IL-8及IL-17的水平明显下降,IL-10水平明显升高(P均<0.05),且联合益生菌疗法组比标准三联疗法组变化更明显(P均<0.05).随着慢性胃炎炎症程度的逐渐加重,血清IL-8、IL-17水平逐渐升高,IL-10水平逐渐下降(P均<0.05).两组治疗后,H.pylori阴性与H.pylori阳性患者比较,血清IL-8及IL-17的水平明显下降,且IL-10水平明显升高(P均<0.05).直线相关性分析显示,两治疗组患者治疗前血清IL-17与IL-8水平呈正相关,IL-17与IL-10水平呈负相关(P均<0.05).结论 标准三联疗法联合益生菌不仅能提高H.pylori的根除率,且能降低H.pylori慢性胃炎患者血清IL-8、IL-17水平及提高血清IL-10水平,具有很好的治疗H.pylori效果.

目的 探讨补充益生菌对根除慢性胃炎患者幽门螺旋杆菌(Helicobacter pylori,Hp)的临床效果及其胃黏膜病理损伤转归的影响.方法 收集2017年1月1日-2017年8月31日在广东药科大学附属第二医院确诊的慢性胃炎伴Hp感染患者104例,按随机数字表法将患者分为对照组和实验组,每组52例,对照组患者仅根除Hp,实验组患者予根除Hp同时补充益生菌治疗,对比分析2组患者干预前和干预6个月后的临床症状、胃镜下胃黏膜表现、Hp根除率以及胃黏膜病理变化(包括炎症、萎缩、肠化生和异型增生)情况.结果 根除Hp对2组患者的临床症状和病理损伤均有不同程度的改善,但实验组患者的临床疗效和Hp根除率优于对照组(90.38％ vs.78.85％,P=0.036;90.38％ vs.75.00％,P=0.038),实验组患者干预后胃黏膜病理总评分低于对照组(4.15±1.02 vs.5.92±1.75,P＜0.05),其中胃黏膜炎症和肠化生病理改善疗效显著优于对照组(0.87±0.12 vs.1.46±0.37,P＜0.05;1.27±0.45 vs.1.81±0.58,P＜0.05).结论 根除幽门螺旋杆菌能使大多数慢性胃炎患者临床获益,补充益生菌可有效提高患者的临床疗效和Hp根除率,并减轻胃黏膜的病理损伤.

Objective To investigate the efficacy of quadruple therapy containing bifidobacterium and lactobacillus triple live bacteria on eradication of Helicobater pylori (Hp)among the patients with peptic ulcer. Methods 342 Hp-infected peptic ulcer patients were randomly divided into three groups:A,B and C. The patients in group A were treated with stand-ard triple therapy. The patients in group B and group C were treated with Colloidal Bismuth Subcitrate and Bifidobacterium and Lactobacillus combined with standard triple therapy,respectively. All patients in three groups were treated for 14 days. In the 4th week after end of treatment,Hp eradication was assessed by 13 C-urea breath test. Adverse effects during the course of treatment were recorded. Results A total of 300 (87. 72%)patients completed the treatment. The completion rates in group A,B and C were 85. 71% (96/112),82. 50% (99/120)and 95. 45% (105/110)respectively,and the comple-tion rate in group C were significantly higher than that in group A and group B (P<0. 05 ). With intention to treat and per-protocol analysis in gastric ulcer,the eradication rates of group A,B and C were 64. 71% (75. 86%),71. 43%(85. 71%)and 84. 38% (87. 10%)respectively,but there were not significant difference in the three groups(P>0. 05). With intention to treat analysis in duodenal ulcer,the Hp eradication rate in group C was 85. 90%,which was significantly&nbsp;higher than that in group A (62. 82%;χ2 =10. 893,P=0. 001)and in group B (71. 79%;χ2 =4. 650,P=0. 031). With per-protocol analysis in duodenal ulcer,the Hp eradication rate was 90. 54% in group C and 87. 50% in group B. No Obviously difference was found between group B and group C (P <0. 05 ),but both were higher than that in group A (73. 13%)(χ2 =4. 246,P=0. 039;χ2 =7. 304,P=0. 007). The incidence of adverse reactions including constipation, taste distortion and bloating in group C were significantly lower than those in the other two groups (P<0. 05 ). Conclusion The quadruple therapy containing bifidobacterium and lactobacillus triple live bacteria can obviously enhance the patient's compliance and decrease the adverse reactions,thereby may increase the Hp eradication rate among the patients with peptic ulcer.

Objective To evaluate the efficacy of standard triple therapy combined with probiotics for Helicobacter pylori (H.pylori) infection in duodenal ulcer.Methods Two hundred and twenty five patients with H.pylori-infected duodenal ulcer were randomly assigned to receive the treatment with standard triple therapy (triple group,n =75),quadruple therapy (colloidal bismuth subcitrate combined with standard triple therapy) (quadruple group,n =75) or probiotics combined with standard triple therapy (bifidobacterium and lactobacillus combined with standard triple therapy) (probiotics group,n =75) for 14 days.The gastroseopy and 13C urea breath test were performed in all patients and the adverse reactions were observed 4 weeks after treatment.Results The healing rate of duodenal ulcer in probiotics group was significantly higher than those in triple and quadruple groups (80.8％ vs.46.15％,80.80％ vs.63.49％;P =0.000,0.024,respectively).With intention to treat analysis,the H.pylori eradication rate in probiotics group was 85.33％,which was significantly higher than that in triple group (61.33％,P =0.001) and in quadruple group (72.00％,P =0.046).With per-protocol analysis,the H.pylori eradication rates were 85.71％ in probiotics group and 87.67％ in quadruple group (P =0.737),but both were higher than that in triple group(70.77％;P =0.014,0.041,respectively).The incidence of adverse reactions including constipation,taste distortion and bloating in probiotics group was significantly lower than those in the other two groups.Conclusions Bifidobacterium and lactobacillus triple live bacteria combined with standard triple therapy can increases the H.pylori eradication rate and duodenal ulcer healing rate with high compliance and low adverse reactions in patients with H.pylori-infected duodenal ulcer.

Objective To investigate the efficacy of different adding times,treatment courses and doses of bifidobacterium and lactobacillus triple live bacteria in Helicobacter pylori (Hp) eradication.Methods A total of 280 patients Hp-infected were enrolled and randomly assigned to five groups.Group A received lansoprazole 30 mg,clarithromycin 500 mg and amoxicillin 1,000 mg bid for 14 days;group B received bifidobacterium and lactobacillus triple live bacteria 2,000 mg tid for 14 days followed by regimen of group A for another 14 days;group C1 received regimen of group A with addition of bifidobacterium and lactobacillus triple live bacteria 2,000 mg bid for 14 days;group C2:regimen of group A with addition of bifidobacterium and lactobacillus triple livc bacteria 2,000 mg tid for 14 days;and group D received regimen of group C2 followed by bifidobacterium and lactobacillus triple live bacteria 2,000 mg tid for another 14 days.4 weeks after end of treatment,Hp eradication was assessed by 13C-urea breath test.Adverse effects during the courses of treatment were recorded.Results A total of 252 (90.0％) patients completed the treatment.The completion rate in group A,B,C1,C2,and D were 78.6％ (44/56),92.9％ (52/56),87.5％ (49/56),96.4％ (54/56),and 94.6％ (53/56) respectively;the completion rate was significantly higher in group B,C2 and D than in group A (P ＜ 0.05),but there were no differences among groups B,C2 and D (P ＞ 0.05).According to intention-to-trcat (ITT) analysis,the eradication rate was 62.5％,80.4％,69.6％,85.7％,and 87.5％ in groups A,B,C1,C2,and D respectively.The eradication rate in groups B,C2 and D was significantly higher than that in group A (x2 =4.375,P =0.036;x2 =7.864,P =0.005;x2 =9.333,P =0.002),and the eradication rate was higher in group C2 than in group C1 (x2 =4.171,P =0.041),but there were no differences among groups B,C2 and D (P ＞0.05).As for per-protocol (PP) analysis,the eradication rate was 79.5％,86.5％,79.6％,88.9％ and 92.5％ in groups A,B,C1,C2,and D respectively,but no significant statistical differences were found among the five groups (P ＞ 0.05).Adverse effects included nausea,bloating,taste distortion,anorexia and constipation.The rate of adverse effects in groups A,B,C1,C2 and D was 67.9％ (38/56),26.8％ (15/56),35.7％ (20/56),21.4％ (12/56),and 17.9％ (10/56) respectively.The incidence rate was significantly lower in groups B,C2 and D than in group A (P ＜ 0.05),but no significant statistical differences were found among groups B,C2,and D (P ＞ 0.05).Conclusions The triple therapy combined with bifidobacterium and lactobacillus triple live bacteria can obviously decrease the adverse effects and improve patient compliance,thereby increasing the rate of Hp eradication.14-day therapy with probiotics is the best regimen.

Background:With the emergence of bacterial resistance,the efficacy of Helicobacter pylori(Hp)eradication therapy is decreasing in recent years. After the previous failed course of eradication therapy,the possibility of failure of re-eradication therapy is greater. Therefore,choosing the rescue therapy for Hp re-eradication is particularly important. Aims:To investigate the efficacy of levofloxacin-based triple therapy combined with probiotics as a rescue therapy for Hp re-eradication. Methods:A total of 192 patients with a previous failed course of standard triple therapy for Hp eradication were enrolled and randomly assigned into four groups. Group A:lansoprazole 30 mg + amoxicillin 1 000 mg + levofloxacin 200 mg bid for 14 days;group B:regimen of group A with the addition of bismuth potassium citrate 600 mg bid for 14 days;group C:bifidobacterium and lactobacillus triple live bacteria 2 000 mg tid for 14 days followed by regimen of group A for 14 days;group D:regimen of group A with the addition of bifidobacterium and lactobacillus triple live bacteria 2 000 mg tid for 14 days. At 4 weeks after end of treatment,Hp eradication was assessed by 13 C-urea breath test. Adverse effects during the course of treatment were recorded. Results:A total of 177(92. 2% )patients completed the study,the completion rates in group A,B,C and D were 87. 5%(42 / 48),83. 3%(40 / 48),97. 9%(47 / 48)and 100% ,&nbsp;respectively;the completion rates in group C and D were significantly higher than those in group A and B(P < 0. 05). By intention-to-treat(ITT)analysis,the eradication rates of group A,B,C and D were 60. 4% ,68. 8% ,81. 3% and 83. 3% ,respectively;the eradication rates in group C and D were significantly higher than that in group A(χ2 = 5. 042, P = 0. 045;χ2 = 6. 235,P = 0. 013). By per-protocol(PP)analysis,the eradication rates of group A,B,C and D were 69. 0% ,82. 5% ,83. 0% and 83. 3% ,respectively;no statistically significant differences were found among the four groups(P > 0. 05). Adverse effects included constipation,taste distortion,bloating and anorexia,the incidences of adverse effects in group A,B,C and D were 79. 2% ,95. 8% ,29. 2% and 22. 9% ,respectively;the incidences in group C and D were significantly lower than those in group A and B(P < 0. 05). Conclusions:Levofloxacin-based triple therapy combined with bifidobacterium and lactobacillus triple live bacteria can decrease the adverse effects of traditional triple and quadruple therapies and improve the patient compliance,thus increases the efficacy of rescue therapy for Hp re-eradication.

Objective To investigate the effects of drug triple therapy combined with bifidobacterium and lactobacillus triple live bacteria on the radical treatment of Helicobacter pylori (Hp) among the seafarers.Methods In accordance with the wishes of the patients,126 Hp-infected cases with peptic ulcer were divided into 2 groups.The control group was treated with lansoprazole,clarithromycin and amoxicillin for a succession of 2 weeks.The therapeutic group received lansoprazole,clarithromycin and amoxicillin combined with bifidobacterium and lactobacillus triple live bacteria,also for a succession of 2 weeks.All the patients in both groups were detected with 13 C-urea breath test and gastroscopy,4 weeks after withdrawal of the drug,and observations were made on Hp eradication rate,peptic ulcer heal rate and rate of adverse reactions in the patients of both groups.Results For adverse reactions in the gastrointestinal system of the therapeutic group,incidence of nausea,abdominal distention,diarrhea and parageusia accounted for 41.7％,46.7％,25.0％and 30.0％ respectively,while the data for the control group were 73.6％,66.0％,52.8％ and 56.6％respectively.Statistical significance could be noted,when comparisons were made between the 2 groups (P ＜0.05 or P ＜0.01).Following 4 weeks of treatment,peptic ulcer heal rates of both groups were 90.57％ and 95.00％ respectively,without statistical significance(P ＞ 0.05).In accordance with intention-to-treat (ITT) and per-protocol (PP) analyses,eradication rates for the control and therapeutic groups were 71.4％ and 84.1％,84.9％ and 88.3％ respectively,without any statistical significance (P ＞ 0.05).Conclusions Drug triple therapy combined with bifidobacterium and lactobacillus triple live bacteria failed to increase the healing rate of ulcer and eradication rate of Hp in seafarers.However,it could obviously decrease adverse reactions in the gastrointestinal system induced by drug triple therapy and helped to increase the compliance of patients.

Objective To analyse and study the liver disease complicated with spontaneous bacterial peritonitis (SBP) of bacterial resistance to antibiotics,to provide clinical reference. Methods Research for patients in our hospital treated 96 cases of liver disease complicated with SBP, the nature of ascites bacterial culture positive, divided into the treatment group of 52 cases and a control group of 44 cases. The control group using a single antibiotic (three generation cephalosporins) treatment;treatment group with more than two kinds of antibiotics (three generation cephalosporins) treatment.The clinical data, ascites cells in culture,drug sensitive test results analysis and research. Results 2 more after antibiotic treatment is single antibiotic treatment has obvious curative effect.For al patients, including 67 cases of Gram-negative bacteria(69.8%);21 cases with Gram-positive bacteria (21.9%). Imipenem, Amikacin and piperacil in/tazobactam for leather, three were gram-negative patients are more sensitive to vancomycin, linezolid; and leather, Gram-positive bacteria were more sensitive. Others such as gentamicin, three generations of cephalosporins, quinolones for Gram-negative and Gram-negative bacteria for patients with lower sensitivity. Conclusion Ascites bacterial culture positive liver disease in patients with SBP, its main ascites pathogens were Gram-negative bacteria, and has strong resistance. The use of antibiotics in patients with liver disease with SBP should be according to the drug sensitivity test to select. For more effective treatment of SBP.

Objective To approach the clinical curative effect of emergency hemostasis by endoscope associated with somatostatin on pateints with acute non-variceal upper gastrointestinal hemorrhage.Methods Sixty-eight pateints with acute non-variceal upper gastrointestinal bleeding,who were hospitalized from April 2009 to April 2011,and randomly divided into control group and observation group,34 cases in each group.The former group was given conventional therapy,the later group was accepted different kinds of emergency hemostasis by endoscope(spray,submucosal injection,ring clamp,high frequency electric coagulation for hemostasis) according to diversity etiopathogenisis of pateints,and combined with somatostatin treatment.The clinic effect,rebleeding rate and length of stay were compared between two groups.Results The excellence rate and the total effective rate were significantly higher in observation group than that in control group,the difference had statistical significance(P<0.05),the rebleeding rate and length of stay were obviously reduced in observation group,than that in control group(P<0.05).Conclusion There is a certain clinical curative effect of emergency hemostasis by endoscope combined with somatostatin on acute non-variceal upper gastrointestinal hemorrhage.The hemorrhage is lower and length of stay is short,this therapy is worth for widely utilization.

Objection The aim of this study was to evaluate the feasibility of calcium ionophore (CI) A23187 and human recombinant granulocyte-macrophage colony stimulating factor (rhGM-CSF) on the cultivation of dendritic cell (DC) from healthy human peripheral blood monocytes (Mo) not purified from the PBMC in vitro. Methods PBMC isolated from peripheral blood of healthy human by Ficoll-Hypaque density gradient separation were separated into two groups. The monocytes purified through adhensiveness from the cells in group A were cultured in the presence of rhGM-CSF and calcium ionophore (CI) A23187. The cells in group B were cultured with additional rhGM-CSF and CI A23187 directly. The cells in both groups were loaded by freeze-thawed K562 cells on the first day and were harvested in 96 hours of cultivation. The morphology of the DCs was observed by microscope all through the process. The surface antigens of the induced cells after culturing for 96 hours were analyzed by fluorescence-activated cell sorting (FACS). Results Typical morphological features of DCs could be observed in both groups. On 96 hours, the expression percentage of DCs in both group were as follows: CD14-CD83+(39.2% vs 40.9%), CD14-CD1a+(19.6% vs 18.3%), CD14-CD86+(47.1% vs 46.0%), and CD14-CD40+(30.5% vs 32.8%), and there was no significant difference in both group(P > 0.05). Conclusion CI A23187 combined rhGM-CSF could effectively induce peripheral blood Mo without purification from the PBMC into DC.

Objective To approach the clinical effect of mesalazine combined with traditional chinese enema on pateints with ulcerative colitis(UC).Methods Forty-two UC patients were randomly divided into control group and treatment group,21 cases in each group,the former group was accepted mesalazine therapy,the later group was treated with mesalazine and yunnanbaiyao powder retention enema,the serum levels of C reactive protein(CRP) and tumor necrosis factor-α(TNF-α) were tested by rate nephelometry and elisa respectively,colonoscopy and colon tissue HE staining were undertook before and after treatment.Results The clinical symptoms relief,serum CPR and TNF-α levels and colonic pathological damages were improved significantly after treatment in two groups,the total effective rate was 95.23% in treatment group,which had significant difference comparing with that in control group(76.2%,P <0.05).Conclusion The therapy of mesalazine combined with yunnanbaiyao powder retention enema can effectively lesson inflammation,reduce intestinal pathological damage and relieve clinical symptoms of patients with UC,which is worth for generalization in clinic.

Objective To evaluate the therapeutic effect of adefovir dipivoxil associated with live combined bifidobacterium and lactobacillus in the treatment of pateints with type B posthepatitic cirrhosis and ascites.Methods Ninety-one patients with type B hepatitis liver cirrhosis and ascites were collected,who were accepted conventional treatment after admission including protecting liver,diureses and regularly transfusion albumin or plasma etc.All the patients were randomly 1divided into treatment group(46 cases received adefovir dipivoxil 10 mg once a day and live combined bifidobacterium and lactobacillus tablet 2.0 g three times a day treatment),control A group(23 cases,received live combined bifidobacterium and lactobacillus tablet 2.0g three times a day treatment) and control B group(22 cases received conventional treatment only).Hepatic function and ascites were assessed after treatment for 3 months and 12 months respectively.Results The liver function improvement and ascites extinction were significantly better in treantent group than that in control A group and control B group,the difference had statistical significance(P < 0.05);The ascites recur rate was 11.4% in treatment group,which compared with that in control A group(81.8%)and control B group(90.0%) had statistics difference(P < 0.05).Conclusion The therapy of adefovir dipivoxil associated with live combined bifidobacterium and lactobacillus on pateints with type B posthepatitic cirrhosis and ascites has a good effect,which is worth of learning.

目的:利用新型诱导剂钙离子载体A23187联合重组人粒细胞-巨噬细胞集落刺激因子(rhGM-CSF)体外诱导人外周血单核细胞(PBMC)生成树突状细胞(DCs).方法:采用密度梯度离心法及黏附法分离出PBMC,按不同培养方法分成:传统方法组,rhGM-CSF+重组人白介素-4(rhIL-4)+重组人肿瘤坏死因子-α(rhTNF-α);新型诱导剂组,加入rhGM-CSF+A23187.光镜观察细胞形态的变化,流式细胞仪检测DC细胞的表面标志,MTT比色法检测各组DCs刺激同种异体外周血T细胞增殖的能力.结果:与传统方法相比,A23187联合rhGM-CSF诱导培养的DCs更具有典型的树突形态;DC表面分子CD83D1a、CD86、CD40表达(45.2%、31.5%、40.1%、36.4%)较传统方法组(16.9%、20.4%、26.5%、22.3%)明显上调(P＜0.05),而CD14表达(5.7% vs 19.0%)明显下降(P＜0.05),刺激同种异体T细胞增殖的能力更强.结论:新型诱导剂钙离子载体A23187联合GM-CSF能更有效地诱导PBMC生成强效成熟的DCs.

Objective To investigate the method of culturing dendritic cells (DCs) derived from human peripheral blood monocytes. Methods The first step of the procedure was to isolate mononuclear cells from human peripheral blood by Ficoll-Hypaque density gradient separation,then monocytes were purified through adhensiveness,which were induced in vitro by human recombinant granulocyte / macrophage colony stimulating factor (rhGM-CSF) and human recombinant interleukin 4 (rhIL-4). The second step was to add 100 ng / mL of tumor necrosis factor-α (TNF-α) at day 5 for cultivation for 2 days to induce maturation. Morphology of DCs was observed by inverted microscope,and the expressions of CD83,CD1a,CD86,CD40,and CD14 on DCs were analyzed by flow-cytometry. The proliferation of allogenetic T cells was detected by MTT method. Results After a 5 day incubation of monocytes with rhGM-CSF and rhIL-4,most of the cells formed clusters,and the expression levels of CD83,CD1a,CD86,CD40,and CD14 were 14.3%,12.8%,20.1%,19.9%,and 16.2%,respectively. At day 7 in the presence of rhTNF-α,the expression levels of CD83,CD1a,CD86,CD40,and CD14 of DCs were 29.8%,18.2%,33.6%,28.1%,and 8.0%,respectively. Matured DCs had a stronger capacity of stimulating T cells to proliferate. Conclusion It is suggested that combination of rhGM-CSF and rhIL-4 is beneficial to expand the proliferation of immature DCs,which can be induced to mature by 2 days incubation with TNF-α,and matured DCs can strongly stimulate T cells to proliferate.

OBJECTIVE:To evaluate the effect of calcium ionophore (CI) A23187 and human recombinant granulocyte/macrophage colony stimulating factor (rhGM-CSF) on the cultivation of dendritic cell (DC) from healthy human peripheral blood mononuclear cell (PBMC) and to evaluate the in vitro effect of DC stimulated by K562 cell lysate on inducing specific cytotoxic T lymphocyte (CTL) against K562 cell.METHODS:Human PBMCs isolated from healthy subjects were separated into two groups. In Group A, the cells were cultured with additional rhGM-CSF, recombinant human interleukin 4 and recombinant human tumor necrosis factor-α only as control group. In Group B, the cells were cultured in the presence of rhGM-CSF and CI A23187. The cells in both groups were pre-incubated with K562 cell lysate at 37°C for 30 min. The cells were harvested after a 4-day cultivation. Morphology of DC was continuously observed under inverted microscope. The surface antigens of induced cells were analyzed by flow cytometry (FCM). Then the proliferation of allogeneic T cell and the specific cytotoxicity of T cell primed with DC were examined by colorimetry. Also, the nonspecific inhibition of DC loaded K562 cell lysate against K562 cell was detected.RESULTS:Typical morphological features of DC could be observed in both groups. The expressions of CD83, CD1a, CD86 and CD40 were stronger in Group B than those in control group (45.2% ± 1.8%, 31.5% ± 3.9%, 40.1% ± 7.8%, 36.4% ± 6.3% vs 16.9% ± 1.3%, 20.4% ± 3.4%, 26.5% ± 2.2%, 22.3% ± 3.0%) (all P < 0.05). The expression of CD14 was weaker in Group B than that in control group (5.7% ± 0.8% vs 19.0% ± 1.6%) (P < 0.05). As compared with the control group, DC in Group B loaded with K562 lysate could evidently stimulate the proliferation of allogeneic T cell (P < 0.05, exclusion of effector-to-target ratio of 1:40) and inhibit the growth of K562 cell (P < 0.05). In addition, both groups of DC-stimulated CTL had specific cytotoxicity against K562 cell. At the effector-to-target ratios of 10:1 and 40:1, the DC-stimulated CTL of Group B had stronger cytotoxicity against K562 cell (both P < 0.05).CONCLUSION:In combination with rhGM-CSF, CI A23187 induces PBMC into DC in a more effective way. DC loaded with K562 lysate can stimulate CTL and maintain high immunocompetence with specific cytotoxicity against K562 cell.

Dendritic cell (DC) which plays an important role in allograft rejection ,anti-tumor,anti-viral infections and autoimmune diseases is known as the most powerful and professional antigen-presenting cell in vivo. How to cultivate and purify DC with high quality and typical features is the premise of basic and clinical research. Researches about the isolation and cytokine-induced expansion of DC have made a great progress. In this paper,the methods about how to separate,purify and amplify DC in vitro are reviewed.