[1]严鸣光,方 晓,李汶轩,等.强直性脊柱炎模型小鼠踝关节组织和临床患者PBMC 中miR-142-5p，SOCS1 mRNA 表达及与免疫功能分析研究[J].现代检验医学杂志,2024,39(03):29-36.[doi:10.3969/j.issn.1671-7414.2024.03.005]
　YAN Mingguang,FANG Xiao,LI Wenxuan,et al.Expression of miR-142-5p, SOCS1 mRNA in Ankle Joint Tissue Spondylitis Model Mice and PBMC of Clinical Patients and Their Immune Function Analysis[J].Journal of Modern Laboratory Medicine,2024,39(03):29-36.[doi:10.3969/j.issn.1671-7414.2024.03.005]
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强直性脊柱炎模型小鼠踝关节组织和临床患者PBMC 中miR-142-5p，SOCS1 mRNA 表达及与免疫功能分析研究()
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《现代检验医学杂志》[ISSN:/CN:]

卷:
第39卷
期数:
2024年03期
页码:
29-36
栏目:
论著
出版日期:
2024-05-15

文章信息/Info

Title:
Expression of miR-142-5p, SOCS1 mRNA in Ankle Joint Tissue Spondylitis Model Mice and PBMC of Clinical Patients and Their Immune Function Analysis
文章编号:
1671-7414（2024）03-029-08
作者:
严鸣光; 方 晓; 李汶轩; 王 可; 殷卫兵
（商丘市第一人民医院检验科，河南商丘 476100）
Author(s):
YAN Mingguang;  FANG Xiao;  LI Wenxuan;  WANG Ke;  YIN Weibing
（Department of Clinical Laboratory, the First People’s Hospital of Shangqiu, Henan Shangqiu 476100, China）
关键词:
强直性脊柱炎; 微小核糖核酸-142-5p; 细胞因子信号转导抑制因子1; 免疫功能
分类号:
R-332；R392.11
DOI:
10.3969/j.issn.1671-7414.2024.03.005
文献标志码:
A
摘要:
目的 探究强直性脊柱炎（ankylosing spondylitis，AS）模型小鼠和临床患者外周血单个核细胞（peripheralblood mononuclear cell，PBMC） 中微小RNA-142-5p(miR-142-5p）， 细胞因子信号转导抑制因子1（suppressor ofcytokine signaling 1，SOCS1）mRNA 表达及其对免疫功能的影响。方法 通过实时荧光定量（quantitative real timePCR, qRT-PCR）检测2022 年1 月～ 2023 年3 月商丘市第一人民医院收治的30 例确诊的AS 患者（患者组）和30 例健康体检者（健康组）的PBMC中miR-142-5p 和SOCS1 mRNA 水平。采用牛蛋白聚糖联合完全弗氏佐剂诱导AS 小鼠模型，然后将小鼠分为对照组、模型组、阴性组和拮抗剂组。对照组和模型组小鼠尾静脉注射生理盐水，阴性组和拮抗剂组小鼠分别尾静脉注射NC-antagomir 和miR-142-5p-antagomir。治疗2 周后，分别评估各组小鼠的关节炎症状评分。通过苏木精伊红（hematoxylin eosin，HE）染色评价踝关节形态。采用ELISA 法检测小鼠血清中Th1 细胞因子干扰素γ（IFN-γ）、Th2 细胞因子白细胞介素-4（IL-4）、Th17 细胞因子白细胞介素-17（IL-17）和Treg 细胞因子叉头盒蛋白P3（FOXP3）的表达水平。通过qRT-PCR 和Western blot 检测PBMC 和踝关节组织中miR-142-5p，SOCS1，IFN-γ，IL-4，IL-17和FOXP3 的mRNA 和蛋白表达水平。结果 与健康组比较，患者组PBMC 中miR-142-5p 水平升高（3.03 ± 0.99 vs1.00 ± 0.21），SOCS1 mRNA 水平降低（0.41 ± 0.09 vs 1.00 ± 0.18），差异具有统计学意义（t=10.997，15.956，均P<0.001）。与对照组比较，模型组小鼠踝关节组织中miR-142-5p 水平（4.00 ± 0.52 vs 1.00 ± 0.04）升高，IFN-γ 和IL-17 的mRNA和蛋白水平均升高，SOCS1，IL-4 和FOXP3 的mRNA和蛋白水平均降低，差异具有统计学意义（t=23.356，31.420，48.056，47.224，38.035，29.007，54.183，28.123，55.155，26.758，45.346，均P<0.05）；关节炎症状评分升高（7.83 ± 0.94 vs 0.00 ± 0.00，t=22.212，P<0.05），踝关节结构破坏明显；血清中IFN-γ，IL-17 水平以及IFN-γ/IL-4 比值（0.81 ± 0.08 vs 2.08 ± 0.33）和IL-17/FOXP3 比值（0.41 ± 0.03 vs 1.27 ± 0.10）均升高，差异具有统计学意义（t=15.382，35.779，15.412，35.130，均P<0.05）。与阴性组比较，拮抗剂组小鼠踝关节组织中miR-142-5p 水平（1.47± 0.10 vs 3.89 ± 0.33）降低，IFN-γ 和IL-17 的mRNA 和蛋白水平均降低，SOCS1，IL-4 和FOXP3 的mRNA 和蛋白水平均升高，差异具有统计学意义（t=18.846，22.969，43.454，32.617，23.259，20.881，41.832，11.994，32.977，15.190，35.834，均P<0.05）；关节炎症状评分降低（7.42 ± 1.24 vs 2.75 ± 0.75，t=13.233，P<0.05），踝关节形态明显改善；血清中IFN-γ，IL-17 水平以及IFN-γ/IL-4 比值（1.22 ± 0.11 vs 1.91 ± 0.19）和IL-17/FOXP3 比值（0.69 ± 0.05vs 1.23 ± 0.12）均降低，差异具有统计学意义（t=8.688，22.972，8.377，22.007，均P<0.05）。结论 miR-142-5p 在AS 中高表达，使用拮抗剂下调miR-142-5p 可能通过上调SOCS1 进而降低Th1/Th2 和Th17/Treg 比值，从而改善AS 小鼠的免疫平衡并抑制AS 的进展。
Abstract:
Objective To explore the expression of mocro RNA (miR)-142-5p and suppressor of cytokine signaling 1（SOCS1） mRNA in peripheral blood mononuclear cells (PBMC) of mice and clinical patients with ankylosing spondylitis (AS) and their impact on immune function. Methods The mRNA levels of miR-142-5p and SOCS1 in PBMC of 30 patients with AS (Patient group) and 30 healthy controls (Health group) treated in the First People’s Hospital of Shangqiu from January 2022 to March 2023 were measured by quantitative real time PCR(qRT-PCR). AS mice models were induced by bovine proteoglycan combined with complete Freund’s adjuvant, and these mice were divided into control group, model group, NC group and antagomir group. Normal saline was injected into tail vein in control group and model group, and NC-antagomir and miR-142- 5p-antagomir were injected into tail vein in NC group and antagomir group, respectively. After 2 weeks of treatment, the arthritis symptom scores of mice in each group were evaluated. The morphology of ankle joint was evaluated by hematoxylin eosin (HE) staining. The levels of Th1 cytokine interferon-γ (IFN-γ), Th2 cytokine interleukin-4 (IL-4), Th17 cytokine interleukin-17 (IL- 17) and Treg cytokine forkhead box protein P3 (FOXP3) in PBMC of mice were detected by ELISA method. The mRNA and protein expression of miR-142-5p, SOCS1, IFN-γ, IL-4, IL-17 and FOXP3 in PBMC and ankle joints were detected by qRTPCR and Western blot. Results Compared with Health group, the level of miR-142-5p in PBMC of patient group was increased (1.00 ± 0.21 vs 3.03 ± 0.99, t=10.997, P<0.001), while the level of SOCS1 mRNA was decreased (1.00 ± 0.18 vs 0.41 ± 0.09, t=15.956, P<0.001). Compared with control group, miR-142-5p level (1.00 ± 0.04 vs 4.00 ± 0.52) and the mRNA and protein levels of IFN-γ and IL-17 in ankle joint tissue of model group were increased, while the mRNA and protein levels of SOCS1, IL-4 and FOXP3 were decreased, with significant differences（t=23.356，31.420，48.056，47.224，38.035，29.007， 54.183，28.123，55.155，26.758，45.346，all P<0.05）. The arthritis symptom score was increased (7.83 ± 0.94 vs 0.00 ± 0.00, t=22.212, P<0.05), and the ankle joint structure was damaged. Serum IFN-γ, IL-17 levels, IFN-γ/IL-4 ratio (0.81 ± 0.08 vs 2.08 ± 0.33) and IL-17/FOXP3 ratio (0.41 ± 0.03 vs 1.27 ± 0.10) were increased, and the differences were statistically significant (t=15.382，35.779，8.934，35.130， all P<0.05). Compared with NC group, miR-142-5p level (3.89 ± 0.33 vs 1.47 ± 0.10), the mRNA and protein levels of IFN-γ and IL-17 in ankle joint tissue of antagomir group were decreased, while the mRNA and protein levels of SOCS1, IL-4 and FOXP3 were increased, and the differences were statistically significant (t=18.846， 22.969，43.454，32.617，23.259，20.881，41.832，11.994，32.977，15.190，35.834，all P<0.05). The arthritis symptom score was decreased (7.42 ± 1.24 vs 2.75 ± 0.75, t=13.233, P<0.05), and the shape of the ankle joint of the rats was improved. Serum IFN-γ, IL-17 levels, IFN-γ/IL-4 ratio (1.22 ± 0.11 vs 1.91 ± 0.19) and IL-17/FOXP3 ratio (0.69 ± 0.05 vs 1.23 ± 0.12) were decreased, and the differences were statistically significant (t=8.688，22.972，3.785，22.007，all P<0.05). Conclusion MiR-142-5p was highly expressed in AS. Down-regulation of miR-142-5p using antagonists may reduce Th1/Th2 ratio and Th17/Treg ratio through up-regulation of SOCS1, there by improving the immune balance of AS mice and inhibiting the progression of AS.

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备注/Memo

备注/Memo:
基金项目： 河南省科技攻关项目（资助单位：河南省科技厅，编号：212102310743）：强直性脊柱炎环状RNA 表达谱的研究及潜在分子；河南省医学科技攻关项目（资助单位：河南省卫生健康委员会，编号：2018020947）：HLA-B27 不同检测技术对强直性脊柱炎精准诊断的价值研究。
作者简介：严鸣光（1981-），男，硕士，副主任技师，研究方向：自身免疫疾病，E-mail：ymguang1982@163.com。
通讯作者：殷卫兵（1966-），男，主任技师，研究方向：临床免疫检测，E-mail：475410026@qq.com。
更新日期/Last Update: 2024-05-15