中国组织工程研究

中国组织工程研究 ›› 2026, Vol. 30 ›› Issue (13): 3321-3330.doi: 10.12307/2026.623

• 干细胞培养与分化 stem cell culture and differentiation • 上一篇    下一篇

前蛋白转化酶枯草溶菌素9影响人单核细胞源性泡沫细胞胆固醇流出的机制

廖付军1，2，鲍海龙1，2，龚才伟3，刘大男1，2   

  1. 贵州医科大学附属医院，1心血管内科，2心血管内科心肌重塑重点实验室，贵州省贵阳市  550004；3遵义市第一人民医院，贵州省遵义市  563000
  • 接受日期:2025-07-03 出版日期:2026-05-08 发布日期:2025-12-25
  • 通讯作者: 刘大男，博士，主任医师，贵州医科大学附属医院，心血管内科，心血管内科心肌重塑重点实验室，贵州省贵阳市 550004
  • 作者简介:廖付军，男，1983年生，湖南省邵阳市人，汉族，硕士，副主任医师，主要从事动脉粥样硬化及血脂相关研究。
  • 基金资助:
    贵州省卫生健康委科学技术基金项目(gzwkj2023-300)，项目负责人：廖付军；贵州省科技计划项目(黔科合支撑[2021]一般063)，项目负责人：廖付军

Impact and mechanism of proprotein convertase subtilisin/kexin type 9 on cholesterol efflux in human monocyte-derived foam cells

Liao Fujun1, 2, Bao Hailong1, 2, Gong Caiwei3, Liu Danan1, 2   

  1. 1Department of Cardiology, Affiliated Hospital of Guizhou Medical University, Guiyang 550004, Guizhou Province, China; 2Key Laboratory of Myocardial Remodeling, Department of Cardiology, Affiliated Hospital of Guizhou Medical University, Guiyang 550004, Guizhou Province, China; 3Zunyi First People’s Hospital, Zunyi 563000, Guizhou Province, China
  • Accepted:2025-07-03 Online:2026-05-08 Published:2025-12-25
  • Contact: Liu Danan, MD, Chief physician, Department of Cardiology, Affiliated Hospital of Guizhou Medical University, Guiyang 550004, Guizhou Province, China; Key Laboratory of Myocardial Remodeling, Department of Cardiology, Affiliated Hospital of Guizhou Medical University, Guiyang 550004, Guizhou Province, China
  • About author:Liao Fujun, MS, Associate chief physician, Department of Cardiology, Affiliated Hospital of Guizhou Medical University, Guiyang 550004, Guizhou Province, China; Key Laboratory of Myocardial Remodeling, Department of Cardiology, Affiliated Hospital of Guizhou Medical University, Guiyang 550004, Guizhou Province, China
  • Supported by:
    Health Commission Science and Technology Fund Project of Guizhou Province, No. gzwkj2023-300 (to LFJ); Science and Technology Project of Guizhou Province, No. [2021]063 (to LFJ)

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摘要：

文题释义：

前蛋白转化酶枯草溶菌素9(PCSK9)：在动脉粥样硬化相关研究中，PCSK9通过加速肝细胞表面低密度脂蛋白受体降解，直接调控血浆低密度脂蛋白胆固醇浓度。FOURIER研究纳入27 564例超高危动脉粥样硬化性心血管疾病患者，采用单克隆抗体类PCSK9抑制剂(如依洛尤单抗)可使低密度脂蛋白胆固醇降低59%-62%，且每降低1 mmol/L低密度脂蛋白胆固醇可减少21%主要心血管事件。值得注意的是，此类药物的作用机制独立于他汀类药物，联合治疗时可使低密度脂蛋白胆固醇降至0.78 mmol/L以下(2019 ESC指南推荐值)，且基因测序证实PCSK9功能缺失突变携带者的冠心病风险降低88%。该靶点的发现为家族性高胆固醇血症患者提供了新的治疗方式。
胆固醇流出：是指胆固醇从细胞内向细胞外转运的过程，尤其是从巨噬细胞到高密度脂蛋白的转运，这一过程对于维持细胞和全身胆固醇稳态至关重要。在动脉粥样硬化的背景下，胆固醇流出的效率直接影响泡沫细胞的形成和发展，进而影响动脉斑块的稳定性。量化研究表明，胆固醇流出的速率与ATP结合盒转运蛋白A1、ATP结合盒转运体G1等转运蛋白的表达密切相关。通过测量胆固醇流出率，研究人员能够评估细胞在不同条件下的胆固醇代谢功能。这一指标对于开发新型治疗策略、预防心血管疾病具有重要意义，尤其是在调节胆固醇外排能力以防止泡沫细胞积累的研究中，胆固醇流出被认为是一个关键靶点。

摘要
背景：人类前蛋白转化酶枯草溶菌素9(PCSK9)作为胆固醇代谢的重要调节因子，通过与低密度脂蛋白受体结合促进低密度脂蛋白受体降解，从而影响胆固醇的摄取和血脂水平。而胆固醇通过ATP结合盒转运蛋白A1(ABCA1)和清道夫受体BI(SR-BI)等转运蛋白进行外流，转运驱动蛋白结合蛋白2(TRAK2)作为衔接蛋白可能在此过程中发挥作用。因此，探讨PCSK9是否通过TRAK2/ABCA1途径调节胆固醇外流，为理解PCSK9在动脉粥样硬化中的作用提供新视角和理论依据。
目的：探讨PCSK9对人单核细胞源性巨噬细胞胆固醇流出的影响及机制。
方法：取THP-1细胞离心后加入佛波酯培养48 h诱导分化为THP-1源性巨噬细胞，再用含氧化型低密度脂蛋白的无血清培养基孵育24 h诱导泡沫细胞形成。①将泡沫细胞分为4组：对照组、PCSK9蛋白组、阴性对照组(非靶向siRNA转染泡沫细胞)、PCSK9 siRNA转染组，RT-qPCR和Western blot检测TRAK2、ABCA1、ABCG1、SR-BI mRNA和蛋白表达；②将泡沫细胞分为3组：阴性对照组、TRAK2 siRNA转染组、(TRAK2 +PCSK9)siRNA共转染组，RT-qPCR和Western blot检测ABCA1 mRNA和蛋白表达；③将泡沫细胞分为8组：对照组，PCSK9蛋白组、PCSK9 siRNA转染组、TRAK2 siRNA转染组、ABCA1 siRNA转染组、(TRAK2 +PCSK9)siRNA共转染组、(PCSK9 +ABCA1) siRNA共转染组、(TRAK2 +ABCA1) siRNA共转染组，采用25-NBD-胆固醇分析泡沫细胞胆固醇流出率。
结果与结论：①PCSK9蛋白组TRAK2 mRNA和蛋白表达高于对照组(P < 0.05)，ABCA1、ABCG1、SR-BI mRNA和蛋白表达相对于对照组无显著统计学差异；PCSK9 siRNA转染组TRAK2 mRNA和蛋白表达低于阴性对照组(P < 0.001和 P < 0.05)，ABCA1 mRNA和蛋白表达高于阴性对照组(P < 0.001)，ABCG1、SR-BI mRNA和蛋白表达相对于阴性对照组无显著统计学差异；②TRAK2 siRNA转染组ABCA1 mRNA和蛋白表达高于阴性对照组(P < 0.001和P < 0.05)，(TRAK2+PCSK9)siRNA组ABCA1 mRNA和蛋白表达高于阴性对照组(P < 0.001和P < 0.05)，(TRAK2+PCSK9)siRNA组相比于TRAK2 siRNA组ABCA1 mRNA和蛋白表达均无显著统计学差异；③PCSK9蛋白组相比于对照组胆固醇外流无显著统计学差异；PCSK9 siRNA、TRAK2 siRNA和(PCSK9+TRAK2)siRNA组胆固醇外流均高于对照组(P < 0.001)，组间胆固醇外流无显著统计学差异；ABCA1 siRNA组胆固醇外流低于对照组(P < 0.001)；(PCSK9+ABCA1)siRNA组胆固醇外流低于PCSK9 siRNA组(P < 0.001)，(TRAK2+ABCA1)siRNA组胆固醇外流低于TRAK2 siRNA组(P < 0.001)。结果表明，PCSK9可能通过TRAK2/ABCA1途径调节胆固醇流出。

关键词: PCSK9, TRAK2, 巨噬细胞, 胆固醇代谢, ABCA1, 胆固醇外流, 泡沫细胞, THP-1

Abstract: BACKGROUND: Human proprotein convertase subtilisin/kexin type 9 (PCSK9), a key regulator of cholesterol metabolism, modulates cholesterol uptake and plasma lipid levels by binding to and promoting the degradation of the low-density lipoprotein receptor. Cholesterol efflux is mediated by transporters such as ATP-binding cassette transporter A1 (ABCA1) and scavenger receptor class B type I (SR-BI), with trafficking kinesin protein 2 (TRAK2) potentially acting as an adaptor protein in this process. Investigating whether PCSK9 regulates cholesterol efflux through the TRAK2/ABCA1 pathway provides novel insights into its role in atherosclerosis.
OBJECTIVE: To explore the effect of PCSK9 on cholesterol efflux in human monocyte-derived macrophages and its underlying mechanisms.
METHODS: THP-1 cells were centrifuged and added with phorbol ester for 48 hours to induce differentiation into THP-1-derived macrophages, and then incubated with serum-free medium containing oxidized low-density lipoprotein for 24 hours to induce foam cell formation. (1) The foam cells were divided into four groups: control group, PCSK9 protein group, negative control group (non-targeted siRNA transfected foam cells), and PCSK9 siRNA transfection group. RT-qPCR and western blot assay were used to detect the mRNA and protein expressions of TRAK2, ABCA1, ABCG1, and SR-BI. (2) The foam cells were divided into three groups: negative control group, TRAK2 siRNA transfection group, and (TRAK2+PCSK9) siRNA co-transfection group. RT-qPCR and western blot assay were used to detect the mRNA and protein expressions of ABCA1. (3) The foam cells were divided into eight groups: control group, PCSK9 protein group, PCSK9 siRNA transfection group, TRAK2 siRNA transfection group, ABCA1 siRNA transfection group, (TRAK2+PCSK9) siRNA co-transfection group, (PCSK9+ABCA1) siRNA co-transfection group, and (TRAK2+ABCA1) siRNA co-transfection group. 25-NBD-cholesterol was used to analyze the cholesterol efflux rate of foam cells. 
RESULTS AND CONCLUSION: (1) The TRAK2 mRNA and protein expression levels in the PCSK9 group were higher than those in the control group (P < 0.05), whereas the expression levels of ABCA1, ABCG1, and SR-BI showed no significant difference compared to the control group. In the PCSK9 siRNA transfection group, TRAK2 mRNA and protein expression levels were lower than in the negative control group (P < 0.001 and P < 0.05, respectively), while ABCA1 mRNA and protein expression levels were higher than in the negative control group (P < 0.001); no significant difference was observed in the mRNA and protein expression levels of ABCG1 and SR-BI compared to the negative control group. (2) In the TRAK2 siRNA transfection group, ABCA1 mRNA and protein expression levels were higher than those in the negative control group (P < 0.001 and P < 0.05, respectively), and in the (TRAK2+PCSK9) siRNA group, ABCA1 mRNA and protein expression levels were higher than those in the negative control group (P < 0.001 and P < 0.05, respectively). There was no significant difference in ABCA1 mRNA and protein expression levels between the (TRAK2+PCSK9) siRNA group and the TRAK2 siRNA group. (3) There was no significant difference in cholesterol efflux between the PCSK9 group and the control group. Cholesterol efflux in the PCSK9 siRNA, TRAK2 siRNA, and PCSK9+TRAK2 siRNA groups was higher than in the control group (P < 0.001), with no significant difference in cholesterol efflux between these groups. Cholesterol efflux in the ABCA1 siRNA group was lower than in the control group (P < 0.001); cholesterol efflux in the (PCSK9+ABCA1) siRNA group was lower than in the PCSK9 siRNA group (P < 0.001), and cholesterol efflux in the (TRAK2+ABCA1) siRNA group was lower than in the TRAK2 siRNA group (P < 0.001). These findings indicate that PCSK9 may regulate cholesterol efflux via the TRAK2/ABCA1 pathway.

Key words: PCSK9, TRAK2, macrophage, cholesterol metabolism, ABCA1, cholesterol efflux, foam cell, THP-1

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