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生物矿化的白蛋白/siRNA纳米粒的构建和胞质递送效率研究

邹志文, 吴锦慧

邹志文,吴锦慧. 生物矿化的白蛋白/siRNA纳米粒的构建和胞质递送效率研究[J]. 中国药科大学学报,2025,56(3):321 − 328. DOI: 10.11665/j.issn.1000-5048.2024072202
引用本文: 邹志文,吴锦慧. 生物矿化的白蛋白/siRNA纳米粒的构建和胞质递送效率研究[J]. 中国药科大学学报,2025,56(3):321 − 328. DOI: 10.11665/j.issn.1000-5048.2024072202
ZOU Zhiwen, WU Jinhui. Construction and cytosolic delivery efficiency study of biomineralized albumin/siRNA nanoparticles[J]. J China Pharm Univ, 2025, 56(3): 321 − 328. DOI: 10.11665/j.issn.1000-5048.2024072202
Citation: ZOU Zhiwen, WU Jinhui. Construction and cytosolic delivery efficiency study of biomineralized albumin/siRNA nanoparticles[J]. J China Pharm Univ, 2025, 56(3): 321 − 328. DOI: 10.11665/j.issn.1000-5048.2024072202

生物矿化的白蛋白/siRNA纳米粒的构建和胞质递送效率研究

详细信息
    通讯作者:

    吴锦慧: Tel:025-83592629 E-mail:wuj@nju.edu.cn

  • 中图分类号: R944

Construction and cytosolic delivery efficiency study of biomineralized albumin/siRNA nanoparticles

  • 摘要:

    解决小干扰RNA(siRNA)载体在胞质递送效率与生物相容性之间的矛盾是推动siRNA疗法临床转化的关键。本研究使用牛血清白蛋白(BSA)和人体必需的金属离子,构建了一种生物相容性高、可完全生物降解的siRNA递送载体MnCO3@BSA/Zn2+/siRNA(MRna)。该载体利用Zn2+和Mn2+与生物大分子(BSA和siRNA)的高亲和力,通过水相“一锅法”自组装和生物矿化反应完成了对siRNA的荷载和保护,实现了近90%的siRNA包封率。同时, MRna能够在内体环境中快速崩解使siRNA释放约55%,并通过介导“质子海绵效应”促进siRNA内体逃逸,使siRNA与溶酶体定位系数仅0.18。最终,荷载了CD47 siRNA的MRna可在mRNA和蛋白水平有效降低肿瘤细胞CD47表达,基因沉默效率达到52%,转染效果与商业试剂Lipo2000相近。本研究为siRNA递送系统的设计提供了一种更加简单、高效的策略。

    Abstract:

    Resolving the conflict between cytoplasmic delivery efficiency and biocompatibility of small interfering RNA (siRNA) carriers is crucial for the clinical translation of siRNA therapies. In this study, we developed a highly biocompatible and fully biodegradable siRNA delivery system, MnCO3@BSA/Zn2+/siRNA (MRna), using bovine serum albumin (BSA) and essential metal ions. This carrier leverages the high affinity of Zn2+ and Mn2+ for biomolecules (BSA and siRNA) to achieve siRNA loading and protection through a water-phase “one-pot” self-assembly and biomineralization process, achieving a 90% siRNA encapsulation rate. Additionally, the nanoscale mineral particles allow rapid disintegration in the endosomal environment to release 55% of siRNA and facilitate its endosomal escape by mediating the “proton sponge effect”. Therefore, the colocalization coefficient of siRNA with the lysosome is only 0.18. Ultimately, MRna loaded with CD47 siRNA effectively reduces CD47 expression at both mRNA and protein levels in tumor cells to lower than 50% of the original, showing efficiency comparable to the commercial transfection reagent Lipo2000. Overall, this study provides a more convenient, efficient and biocompatible strategy for designing siRNA delivery systems.

  • Figure  1.   Scheme of the preparation procedure of MnCO3@ BSA/Zn2+/siRNA (MRna) nanoparticle through self-assembly of Zn2+ with BSA/siRNA and subsequent biomineralization via Mn2+ and HCO3-.

    Figure  2.   Morphology and particle size of MRna before and after mineralization

    A: TEM image and photos of MRna; B: Hydration kinetics particle size before mineralization; C: Hydration kinetics particle size after mineralization

    Figure  3.   Composition of the MRna

    A: Elemental composition of the MRna(bar=100 nm); B: Element content in the MRna; C: CLSM-STED image of fluoresce labeled MRna(bar=200 nm)

    Figure  4.   Stability and siRNA release behavior of the formulation

    A: Changes in particle size of the MRna ($ \overline{\text{x}}\pm {\text{s}} $, n = 3); B: Stability of siRNA in FBS solution; C: Release behavior of siRNA in MRna under different pH

    Figure  5.   Proliferative toxicity and apoptotic toxicity of the MRna

    A: Cell proliferation toxicity of MRna ($ \overline{\text{x}}\pm {\text{s}} $, n = 6); B: Cell apoptosis toxicity of MRna

    Figure  6.   Cellular uptake and intracellular localization of siRNA($ \overline{\text{x}}\pm {\text{s}} $, n=3)

    A: Flow cytometry analysis of cellular uptake; B: Co-localization of siRNA with lysosome, bar=5 μm

    Figure  7.   Lysosomal membrane integrity(bar=5 μm)

    Figure  8.   Efficiency of target gene knockdown by the MRna

    A:Flow cytometry analysis of CD47 expression; B:qPCR analysis of CD47 mRNA and western blot analysis of the expression of CD47 protein ($ \overline{\text{x}}\pm {\text{s}} $, n = 3)

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出版历程
  • 收稿日期:  2024-07-21
  • 修回日期:  2024-08-01
  • 录用日期:  2024-08-25
  • 刊出日期:  2025-06-24

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