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. 2021 May 25;144(4):286–302. doi: 10.1161/CIRCULATIONAHA.120.052318

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© 2021 The Authors.

Circulation is published on behalf of the American Heart Association, Inc., by Wolters Kluwer Health, Inc. This is an open access article under the terms of the Creative Commons Attribution Non-Commercial-NoDerivs License, which permits use, distribution, and reproduction in any medium, provided that the original work is properly cited, the use is noncommercial, and no modifications or adaptations are made.

This article is made available via the PMC Open Access Subset for unrestricted re-use and analyses in any form or by any means with acknowledgement of the original source. These permissions are granted for the duration of the COVID-19 pandemic or until permissions are revoked in writing. Upon expiration of these permissions, PMC is granted a perpetual license to make this article available via PMC and Europe PMC, consistent with existing copyright protections.

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Figure 4. — Localization of aerocytes and general capillary endothelial cells (ECs). A, Representative serial immunofluorescent images of microvascular markers PRX and CA4 with positive red staining in capillaries (CA4 with off-target positive staining in macrophages) and negative staining of a larger central vessel. The general endothelial markers CLDN5 and PECAM1 in green stain the larger central vessels as well, in addition to the microvasculature. The third image shows an immunostain of the aerocyte-specific marker HPGD in green (white arrows), in addition to general microvascular marker PRX in red. Nuclei are counterstained throughout with DAPI (4′,6-diamidino-2-phenylindole). The white box highlights an aerocyte, which is shown at higher magnification in the first image of B. B, Representative immunostains of aerocytes with the specific marker HPGD in green (cytoplasmic and nuclear expression pattern), which colocalizes with the general microvascular marker PRX in red. Nuclei are counterstained throughout with DAPI. C, In situ RNA hybridization stains of markers specific to the capillary subpopulations with SOSTDC1 staining aerocyte ECs (arrows) and FCN3 staining general capillary ECs (arrows) with positive staining in red. The black box highlights an area shown to the right of it in high magnification. Conventional immunohistochemical images of shown markers can be found in Figure IV in the Data Supplement.