IL-22在Behcet病及Vogt-小柳原田综合征中的表达及其影响因素的研究
发布时间：2019-07-06 09:59
【摘要】：第一部分 IL-22在Behcet病中的表达及其意义 1.目的 检测IL-22在Behcet病患者外周血及皮损中的表达情况，分析其与疾病活动及临床症状的相关性。 2.方法： 2.1密度梯度法分离患者及正常人外周单个核细胞（PBMCs），磁珠分选CD4~+T细胞，加入抗CD3和CD28抗体刺激行细胞培养。 2.2采用酶联免疫吸附法（ELISA）检测患者及健康对照者血清、PBMCs及CD4~+T细胞培养上清中IL-22的含量。 2.3流式细胞仪检测外周血中IL-22~+CD4~+T细胞、IL-22~+CD8~+T细胞的比例，及IL-22、IL-17共表达的情况。 2.4Real-time PCR法检测结节性红斑中IL-22mRNA的表达。 2.5依据PBMC培养上清中IL-22的含量将患者分为“IL-22正常组”与“IL-22高表达组”，分别与临床特点，血清标志物进行统计学分析。 3.结果 3.1IL-22在活动期BD患者，静止期BD患者，急性特发性前葡萄膜炎患者及正常人的血清水平没有显著性差异。 3.2在活动期Behcet病患者中， PBMCs细胞培养上清中的IL-22含量显著高于静止期Behcet患者，急性特发性前葡萄膜炎患者及正常人。同样，在活动期Behcet病患者中， CD4~+T细胞培养上清中的IL-22含量显著高于静止期Behcet患者及正常人。 3.3在活动期Behcet病患者中，IL-22~+CD4~+T细胞的比例显著高于静止期Behcet患者及正常人。同时，IL-22,IL-17双阳性的CD4~+T细胞的比例在活动期BD患者也显著升高。BD患者及正常人均只有小部分IL-22~+CD4~+T细胞（21%-27%）共表达IL-17,而大部分的IL-22~+CD~+T细胞是IL-17阴性的。 3.4结节性红斑中IL-22的mRNA表达显著高于正常皮肤。 3.5BD患者PBMCs培养上清中升高的IL-22与前房细胞，视网膜血管炎及结节性红斑正相关。 4.结论 实验结果显示IL-22的表达与Behcet病的活动性及小血管炎相关，提示其可能参与了Behcet病的发病。 第二部分 Behcet病中IL-23，地塞米松及环孢素A对IL-22表达的影响 1.目的 研究BD患者中，IL-23是否可诱导IL-22分泌，地塞米松及环孢素A是否可抑制IL-22的分泌。 2.方法 2.1分离BD患者及在正常人PBMCs，磁珠分选CD4~+T细胞，进行不同条件下细胞培养。 2.2CD4~+T细胞分4组。1组为基础培养组，2组加入抗C D3和C D28抗体，，3组加抗C D3和C D28抗体及r h I L-23，4组为只加入rhIL-23组。 2.3PBMCs细胞分6组。1组为基础培养基，2组每孔加入抗C D3和C D28抗体刺激，3组在B组基础上加入不同浓度的地塞米松，4组在B组基础上加入不同浓度的环孢素A。5组在C组基础上加入rhIL-23。6组在D组基础上加入rhIL-23。 3.结果 3.1IL-23可以诱导BD患者和正常人活化的CD4~+T细胞分泌IL-22显著增加。并且在活动期的BD患者中，rIL-23诱导CD4~+T细胞产生的IL-22显著高于静止期BD患者和正常人。 3.2地塞米松和环孢素A在体外均可抑制患者及正常人PBMCs分泌IL-22，并且在一定浓度范围内，这种抑制作用呈剂量依赖性。100ng/ml地塞米松可抑制70%IL-22的产生，10ng/ml环孢素A可抑制80%IL-22的产生。 3.3PBMCs加入地塞米松或环孢素A培养后，活动期BD患者和正常人rIL-23诱导PBMCs分泌IL-22的能力均明显受到抑制。 4．结论 活动期BD患者中，IL-22的表达增加可能由于其上游的IL-23诱导分泌所致，提示BD中IL-23/IL-22通路的存在。地塞米松及环孢素A可抑制IL-22的产生。 第三部分 IL-22在Vogt-小柳原田综合征中的表达及其意义 1.目的 检测IL-22在VKH患者外周血中的表达情况，分析其与疾病活动的相关性。 2.方法： 2.1采用酶联免疫吸附法（ELISA）检测患者及健康对照者血清、PBMCs及CD4~+T细胞的培养上清中IL-22的含量 2.2流式细胞仪检测VKH患者及正常人外周血中IL-22~+CD4~+/CD8~+T细胞，IL-22~+IL-17~+CD4~+T的比例。 2.3依据VKH患者PBMC培养上清中IL-22的含量将患者分为“IL-22正常组”与“IL-22高表达组”，分别与临床特点，血清标志物进行统计学分析。 3.结果 3.1IL-22在活动期VKH患者，静止期VKH患者及正常人的血清水平没有显著性差异。 3.2在PBMCs及CD4~+T细胞培养上清中的IL-22含量，活动期VKH病患者中显著高于静止期VHK患者及正常人。 3.3IL-22~+CD4~+T细胞的比例，活动期VKH病患者显著高于静止期VKH患者及正常人。IL-22~+CD8~+T细胞的比例在三者之间没有显著性差异。在VKH患者及正常人中，CD4~+T细胞均是分泌IL-22的主要细胞。IL-22,IL-17双阳性的CD4~+T细胞的比例在活动期VKH患者中也显著升高。 3.4VKH患者PBMCs培养上清中升高的IL-22与羊脂状KP，前房细胞成正相关。 4.结论 实验结果显示IL-22的表达与VKH综合征的活动性正相关，提示其可能参与了VKH综合征的发病。 第四部分 VKH综合征中IL-23，地塞米松及环孢素A对IL-22表达的影响 1.目的 研究VKH患者中，IL-23是否可诱导IL-22分泌，地塞米松及环孢素A是否可抑制IL-22的分泌。 2.方法 2.1分离VKH患者及在正常人PBMCs，磁珠分选CD4~+T细胞，进行不同条件下细胞培养。 2.2CD4~+T细胞分4组。1组为基础培养组，2组加入抗C D3和C D28抗体，3组加抗C D3和C D28抗体及r h I L-23，4组只加rhIL-23。 2.3PBMCs细胞分5组。1组为基础培养基，2组每孔加入抗C D3和C D28抗体刺激，3组在B组基础上加入不同浓度的地塞米松，4组在B组基础上加入不同浓度的环孢素A。5组在B组基础上加入10ng/ml环孢素A及100ng/ml地塞米松。 3.结果 3.1IL-23可以诱导VKH患者和正常人抗CD3抗体和抗CD28抗体刺激的CD4~+T细胞分泌IL-22显著增加。在活动期的VKH患者中，rIL-23诱导CD4~+T细胞产生的IL-22显著高于静止期患者和正常人。 3.2在体外的PBMCs培养体系中，地塞米松和环孢素A均可抑制VKH患者及正常人PBMCs分泌IL-22，这种抑制作用呈剂量依赖性。两种药物联合后抑制效果更加明显，可抑制95%的IL-22分泌。 4.结论 活动期VKH患者中，IL-22的表达增加可能由于其上游的IL-23诱导分泌所致，提示VKH中IL-23/IL-22通路的存在。地塞米松及环孢素A可抑制IL-22的产生。
文内图片：
图片说明：活动期BD患者（n=10）、静止期BD患者（n=8）及正常人(n=10)外周血
[Abstract]:the first part The expression of IL-22 in Behcet's disease and its expression meaning Objective To investigate the expression of IL-22 in peripheral blood and skin lesions of patients with Behcet's disease. the correlation of the shape .2. Method: 2.1 The peripheral mononuclear cells (PBMCs) of the patients and the normal persons were isolated by density gradient method, CD4 ~ + T cells were sorted by magnetic beads, anti-CD3 and CD28 were added. Antibody-stimulated line cell culture 2.2.2 The serum, PBMCs and CD4 ~ + T cell culture of patients and healthy controls were detected by enzyme-linked immunosorbent assay (ELISA). The ratio of IL-22 ~ + CD4 ~ + T cells, IL-22 ~ + CD8 ~ + T cells and IL-2 in peripheral blood were detected by flow cytometry. 2. The co-expression of IL-17. The expression of IL-22 mRNA in erythema. 2.5 The patients were divided into "IL-22 normal group" and "IL-22 high expression group" according to the content of IL-22 in the supernatant of PBMC culture. Clinical features 3. Results 3.1 IL-22 in active BD patients, patients with BD, acute idiopathic preglucomannan There was no significant difference in serum levels in patients with mucositis and normal controls. 3.2 The level of IL-22 in the supernatant of the PBMCs was significantly higher in patients with active Behcet's disease than in the rest period Hcet, patients with acute idiopathic anterior uveitis, and normal subjects. Similarly, in patients with active Behcet, IL-in the supernatant of CD4 + T cell culture The content of IL-22 ~ + CD4 ~ + in patients with active Behcet was significantly higher than that of Behcet's and normal subjects. The proportion of T cells was significantly higher than that of Behcet's and normal controls. At the same time, IL-22 and IL-17 were positive. The proportion of CD4 ~ + T cells increased significantly in active BD patients. Only small part of IL-22 ~ + CD4 ~ + T cells (21% -27%) of BD patients and normal individuals co-express IL-17, and most of them The IL-22 ~ + CD ~ + T cells were negative for IL-17. The expression of IL-22 in nodular erythema was significantly higher than that of normal skin. raised I The results showed that the expression of IL-22 was similar to that of Behc. et-disease Activity and associated vasculitis suggest that it may be involved in the onset of Behcet's disease. part Effect of IL-23, Dexamethasone and Cyclosporin A on the Expression of IL-22 in Behcet's Disease 1. Objective To Study the Effect of IL-23, Dexamethasone and Cyclosporin A on the Expression of IL-22. , IL- 23. Whether IL-22 secretion, dexamethasone and cyclosporin A can be induced to inhibit the secretion of IL-22. 2.1 The patients with BD were isolated from BD and the CD4 ~ + T cells were sorted by magnetic beads, and the cells were cultured under different conditions. 2.2 CD4 ~ + T cells were divided into 4 groups. The anti-C D3 and C D28 antibodies were added to the D3 and C D28, and the anti-C D3 and C D28 antibodies in the 3 groups and the r h I-23 and 4 groups were only added to the rhIL-23 group. to join in The rhIL-23.6 group was added to the group of rhIL-23.6 on the basis of group C, and rhIL-23.6 was added to the group C. Results 3.1 IL-23 could induce a significant increase in the secretion of IL-22 from the activated CD4 + T cells in patients with BD and normal controls. And in the patients with active BD, the IL-22 induced by rIL-23 in the CD4 + T cells was significantly higher than that of the patients in the quiescent period and the normal controls. 3.2.2 The effect of dexamethasone and cyclosporin A in vitro inhibited the secretion of IL-22 from the PBMCs of the patient and the normal human, and the inhibitory effect was in the range of a certain concentration. Reactivity.100 ng/ ml of dexamethasone inhibited the production of 70% IL-22 and 10 ng/ ml of cyclosporine A inhibited the production of 80% IL-22. 3.3 PBMC s Add-in Conclusion: The ability of IL-22 to secrete IL-22 in patients with active BD and normal human rIL-23 was significantly inhibited after the culture of Celecoxib or Cyclosporin A. In active BD patients, the increase in the expression of IL-22 may can be IL-23 induced by the upstream IL-23, suggesting that IL-23 in BD/ IL-2 2. The presence of the pathway. Dexamethasone and cyclosporine A inhibit the production of IL-22. Part III IL-22 Expression of IL-22 in the peripheral blood of patients with VKH and its significance in the analysis of the expression of IL-22 in the peripheral blood of patients with VKH 2. Methods: 2.1 The serum, PBMCs and CD4 ~ + T cells were detected by enzyme-linked immunosorbent assay (ELISA) in the culture of serum, PBMCs and CD4 ~ + T cells. 22. The proportion of IL-22 ~ + CD4 ~ +/ CD8 ~ + T cells, IL-22 ~ + IL-17 ~ + CD4 ~ + T in the peripheral blood of patients with VKH and normal persons was detected by flow cytometry. norm al group" 涓
本文编号：2510935
文内图片：
图片说明：活动期BD患者（n=10）、静止期BD患者（n=8）及正常人(n=10)外周血
[Abstract]:the first part The expression of IL-22 in Behcet's disease and its expression meaning Objective To investigate the expression of IL-22 in peripheral blood and skin lesions of patients with Behcet's disease. the correlation of the shape .2. Method: 2.1 The peripheral mononuclear cells (PBMCs) of the patients and the normal persons were isolated by density gradient method, CD4 ~ + T cells were sorted by magnetic beads, anti-CD3 and CD28 were added. Antibody-stimulated line cell culture 2.2.2 The serum, PBMCs and CD4 ~ + T cell culture of patients and healthy controls were detected by enzyme-linked immunosorbent assay (ELISA). The ratio of IL-22 ~ + CD4 ~ + T cells, IL-22 ~ + CD8 ~ + T cells and IL-2 in peripheral blood were detected by flow cytometry. 2. The co-expression of IL-17. The expression of IL-22 mRNA in erythema. 2.5 The patients were divided into "IL-22 normal group" and "IL-22 high expression group" according to the content of IL-22 in the supernatant of PBMC culture. Clinical features 3. Results 3.1 IL-22 in active BD patients, patients with BD, acute idiopathic preglucomannan There was no significant difference in serum levels in patients with mucositis and normal controls. 3.2 The level of IL-22 in the supernatant of the PBMCs was significantly higher in patients with active Behcet's disease than in the rest period Hcet, patients with acute idiopathic anterior uveitis, and normal subjects. Similarly, in patients with active Behcet, IL-in the supernatant of CD4 + T cell culture The content of IL-22 ~ + CD4 ~ + in patients with active Behcet was significantly higher than that of Behcet's and normal subjects. The proportion of T cells was significantly higher than that of Behcet's and normal controls. At the same time, IL-22 and IL-17 were positive. The proportion of CD4 ~ + T cells increased significantly in active BD patients. Only small part of IL-22 ~ + CD4 ~ + T cells (21% -27%) of BD patients and normal individuals co-express IL-17, and most of them The IL-22 ~ + CD ~ + T cells were negative for IL-17. The expression of IL-22 in nodular erythema was significantly higher than that of normal skin. raised I The results showed that the expression of IL-22 was similar to that of Behc. et-disease Activity and associated vasculitis suggest that it may be involved in the onset of Behcet's disease. part Effect of IL-23, Dexamethasone and Cyclosporin A on the Expression of IL-22 in Behcet's Disease 1. Objective To Study the Effect of IL-23, Dexamethasone and Cyclosporin A on the Expression of IL-22. , IL- 23. Whether IL-22 secretion, dexamethasone and cyclosporin A can be induced to inhibit the secretion of IL-22. 2.1 The patients with BD were isolated from BD and the CD4 ~ + T cells were sorted by magnetic beads, and the cells were cultured under different conditions. 2.2 CD4 ~ + T cells were divided into 4 groups. The anti-C D3 and C D28 antibodies were added to the D3 and C D28, and the anti-C D3 and C D28 antibodies in the 3 groups and the r h I-23 and 4 groups were only added to the rhIL-23 group. to join in The rhIL-23.6 group was added to the group of rhIL-23.6 on the basis of group C, and rhIL-23.6 was added to the group C. Results 3.1 IL-23 could induce a significant increase in the secretion of IL-22 from the activated CD4 + T cells in patients with BD and normal controls. And in the patients with active BD, the IL-22 induced by rIL-23 in the CD4 + T cells was significantly higher than that of the patients in the quiescent period and the normal controls. 3.2.2 The effect of dexamethasone and cyclosporin A in vitro inhibited the secretion of IL-22 from the PBMCs of the patient and the normal human, and the inhibitory effect was in the range of a certain concentration. Reactivity.100 ng/ ml of dexamethasone inhibited the production of 70% IL-22 and 10 ng/ ml of cyclosporine A inhibited the production of 80% IL-22. 3.3 PBMC s Add-in Conclusion: The ability of IL-22 to secrete IL-22 in patients with active BD and normal human rIL-23 was significantly inhibited after the culture of Celecoxib or Cyclosporin A. In active BD patients, the increase in the expression of IL-22 may can be IL-23 induced by the upstream IL-23, suggesting that IL-23 in BD/ IL-2 2. The presence of the pathway. Dexamethasone and cyclosporine A inhibit the production of IL-22. Part III IL-22 Expression of IL-22 in the peripheral blood of patients with VKH and its significance in the analysis of the expression of IL-22 in the peripheral blood of patients with VKH 2. Methods: 2.1 The serum, PBMCs and CD4 ~ + T cells were detected by enzyme-linked immunosorbent assay (ELISA) in the culture of serum, PBMCs and CD4 ~ + T cells. 22. The proportion of IL-22 ~ + CD4 ~ +/ CD8 ~ + T cells, IL-22 ~ + IL-17 ~ + CD4 ~ + T in the peripheral blood of patients with VKH and normal persons was detected by flow cytometry. norm al group" 涓
本文编号：2510935
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