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Endovascular Biopsy: In Vivo Cerebral Aneurysm Endothelial Cell Sampling and Gene Expression Analysis

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Abstract

There is limited data describing endothelial cell (EC) gene expression between aneurysms and arteries partly because of risks associated with surgical tissue collection. Endovascular biopsy (EB) is a lower risk alternative to conventional surgical methods, though no such efforts have been attempted for aneurysms. We sought (1) to establish the feasibility of EB to isolate viable ECs by fluorescence-activated cell sorting (FACS), (2) to characterize the differences in gene expression by anatomic location and rupture status using single-cell qPCR, and (3) to demonstrate the utility of unsupervised clustering algorithms to identify cell subpopulations. EB was performed in 10 patients (5 ruptured, 5 non-ruptured). FACS was used to isolate the ECs and single-cell qPCR was used to quantify the expression of 48 genes. Linear mixed models and exploratory multilevel component analysis (MCA) and self-organizing maps (SOMs) were performed to identify possible subpopulations of cells. ECs were collected from all aneurysms and there were no adverse events. A total of 437 ECs was collected, 94 (22%) of which were aneurysmal cells and 319 (73%) demonstrated EC-specific gene expression. Ruptured aneurysm cells, relative controls, yielded a median p value of 0.40 with five genes (10%) with p values < 0.05. The five genes (TIE1, ENG, VEGFA, MMP2, and VWF) demonstrated uniformly reduced expression relative the remaining ECs. MCA and SOM analyses identified a population of outlying cells characterized by cell marker gene expression profiles different from endothelial cells. After removal of these cells, no cell clustering based on genetic co-expressivity was found to differentiate aneurysm cells from control cells. Endovascular sampling is a reliable method for cell collection for brain aneurysm gene analysis and may serve as a technique to further vascular molecular research. There is utility in combining mixed and clustering methods, despite no specific subpopulation identified in this trial.

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Abbreviations

EB:

Endovascular biopsy

FACS:

Fluorescence-activated cell sorting

EC:

Endothelial cell

qPCR:

Quantitative polymerase chain reaction

MCA:

Multilevel component analysis

SOMs:

Self-organizing maps

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Acknowledgements

The authors would like to thank the UCSF Neurointerventional Radiology service, particularly its fellows, for their help in the sample collection.

Sources of Funding

The project was funded in part by the Society for Neurointerventional Surgery, the UCSF Department of Radiology, and by a grant from the National Cancer Institute R01 CA056721 to Z.W.

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Authors and Affiliations

  1. University of California San Francisco, San Francisco, CA, USA

    Daniel L. Cooke, Van V. Halbach, Steven W. Hetts, Matthew R. Amans, Christopher F. Dowd, Randall T. Higashida, Jeffrey Nelson, Chih-Yang Wang, Helen Kim, Zena Werb, Charles McCulloch, Tomoki Hashimoto, Hua Su & Zhengda Sun

  2. Zuckerberg San Francisco General Hospital and Trauma Center, San Francisco, CA, USA

    David B. McCoy

  3. University of California Irvine, Irvine, CA, USA

    Devon Lawson

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  1. Daniel L. Cooke
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Correspondence to Daniel L. Cooke.

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All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki Declaration and its later amendments or comparable ethical standards.

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Cooke, D.L., McCoy, D.B., Halbach, V.V. et al. Endovascular Biopsy: In Vivo Cerebral Aneurysm Endothelial Cell Sampling and Gene Expression Analysis. Transl. Stroke Res. 9, 20–33 (2018). https://doi.org/10.1007/s12975-017-0560-4

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  • DOI: https://doi.org/10.1007/s12975-017-0560-4

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