Cryogenic electron tomography and elemental analysis of mitochondrial granules in human retinal ganglion cells

IF 4.3 2区生物学 Q2 BIOCHEMISTRY & MOLECULAR BIOLOGY

Structure Pub Date : 2025-08-04 DOI:10.1016/j.str.2025.07.010

Gong-Her Wu, Cathy Hou, Andrew Thron, Hirenkumar Rajendra Patel, Liam Spillane, Sanket Rajan Gupte, Serena Yeung-Levy, Sahil Gulati, Christopher Booth, Yaping Joyce Liao, Wah Chiu

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引用次数: 0

Abstract

Combining three-dimensional (3D) visualization with elemental analysis of vitrified cells can provide crucial insights into subcellular structures and elemental compositions in their native environments. We present a coordinated approach using cryogenic electron energy loss spectroscopy (cryoEELS) and cryogenic electron tomography (cryoET) to characterize the elemental distribution and ultrastructure of vitrified cells. We applied this method to examine calcium disposition in the mitochondria of cultured human retinal ganglion cells (RGCs) exposed to pro-calcifying conditions relevant to optic disc drusen pathology. Our cryoEELS analysis revealed mitochondrial granules with elevated calcium signals, offering direct evidence of mitochondrial calcification. Additionally, cryoET coupled with artificial intelligence-based analysis enabled quantification of the volume and spatial distribution of these calcium granules. This integrated workflow can be broadly applied to various cell types, facilitating the study of ultrastructure and elemental distribution in subcellular structures under diverse physiological and pathological conditions, as well as in response to therapeutic interventions.

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人视网膜神经节细胞线粒体颗粒的低温电子断层扫描和元素分析

将三维可视化与玻璃化细胞的元素分析相结合，可以为了解其原生环境中的亚细胞结构和元素组成提供重要的见解。我们提出了一种协调的方法，使用低温电子能量损失谱（cryoEELS）和低温电子断层扫描（cryoET）来表征玻璃化细胞的元素分布和超微结构。我们应用这种方法检测了暴露于与视盘水肿病理相关的促钙化条件下的培养的人视网膜神经节细胞（RGCs）线粒体中的钙配置。我们的cryoEELS分析显示线粒体颗粒钙信号升高，提供了线粒体钙化的直接证据。此外，cryoET结合基于人工智能的分析，可以量化这些钙颗粒的体积和空间分布。该集成工作流程可广泛应用于各种细胞类型，便于研究不同生理病理条件下亚细胞结构中的超微结构和元素分布，以及对治疗干预的响应。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Structure 生物-生化与分子生物学

CiteScore

8.90

自引率

1.80%

发文量

155

审稿时长

3-8 weeks

期刊介绍： Structure aims to publish papers of exceptional interest in the field of structural biology. The journal strives to be essential reading for structural biologists, as well as biologists and biochemists that are interested in macromolecular structure and function. Structure strongly encourages the submission of manuscripts that present structural and molecular insights into biological function and mechanism. Other reports that address fundamental questions in structural biology, such as structure-based examinations of protein evolution, folding, and/or design, will also be considered. We will consider the application of any method, experimental or computational, at high or low resolution, to conduct structural investigations, as long as the method is appropriate for the biological, functional, and mechanistic question(s) being addressed. Likewise, reports describing single-molecule analysis of biological mechanisms are welcome. In general, the editors encourage submission of experimental structural studies that are enriched by an analysis of structure-activity relationships and will not consider studies that solely report structural information unless the structure or analysis is of exceptional and broad interest. Studies reporting only homology models, de novo models, or molecular dynamics simulations are also discouraged unless the models are informed by or validated by novel experimental data; rationalization of a large body of existing experimental evidence and making testable predictions based on a model or simulation is often not considered sufficient.