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Editors Selection IGR 9-4

Basic Science: Cross-species cell types implicated in glaucoma

Comment by Daniel Stamer & Ross Ethier on:

86822 Cell atlas of aqueous humor outflow pathways in eyes of humans and four model species provides insight into glaucoma pathogenesis, van Zyl T; Yan W; McAdams A et al., Proceedings of the National Academy of Sciences of the United States of America, 2020; 117: 10339-10349

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The prominent cell 'type' of the conventional pathway is the trabecular meshwork (TM) cell, displaying at least two different morphologies (TM vs. juxtacanalicular, JCT)

The architecture of conventional outflow tissues is unique, with resident cells having specialized responsibilities and relationships that together determine IOP. The prominent cell 'type' of the conventional pathway is the trabecular meshwork (TM) cell, displaying at least two different morphologies (TM vs. juxtacanalicular, JCT) that correspond to their anatomical location (inner versus outer TM) and physiological responsibility (biological filter vs. resistance generator).¹ Due to their extensive connectivity, separation of these outflow cells by dissection is extremely difficult. As a result, only bulk RNA sequencing studies of conventional outflow tissues have been performed to genetically profile resident cells.^2-5 With the recent advent of high throughput single cell RNA sequencing, transcriptomic profiles of resident cell 'types' in the conventional outflow is now possible. Using this powerful technology, two recent groundbreaking studies were conducted in parallel, generating cell atlases of conventional outflow pathway and surrounding tissues.⁶

Due to their extensive connectivity, separation of these outflow cells by dissection is extremely difficult

While both studies provide foundational data sets, this review focuses on the work of van Zy et al., who identified individual transcriptomic signatures from 19 (!) different cell types in human outflow tissues. Remarkably, seven different cell types were identified in the conventional outflow pathway. In the filtering region, transcriptomic signatures for JCT cells, resident macrophages (CD63⁺/LYVE1⁺), SC cells and two types of TM cells were discovered. In the non-filtering region, one TM cell type (also known as insert or Schwalbe's line cells) was identified. Lastly, a distinct expression pattern for endothelia distal, but continuous with SC (i.e., collector channel/intrascleral venous plexus/aqueous veins) was also identified.

All three TM cell 'types' in the filtering region expressed high levels of known markers (MYOC, MGP, and PDPN). The two different TM 'beam' cell types were distinguished by expression of the markers FABP4 and TMEFF2, but they did not segregate to specific TM regions in sagittal sections. It would be interesting to learn whether these two beam types correspond to high versus low flow regions. By comparison, JCT cells differentially expressed several genes, CH13L1, ANGPTL7, RSPO4, FMOD and NELL2. SC cells displayed an expression pattern of both blood and lymphatic endothelia, confirming genetic lineage tracing studies in mice.⁷ Surprisingly, there was an abundance of macrophages in TM, having the second highest cellular representation in the conventional tract.

In terms of glaucoma-associated genes, there was differential expression by TM cell types (MYOC, FOXC1, PITX2, CYP1B1, LOXL1, ANGPT1, EFEMP1) versus by SC cells (CAV1, CAV2, TEK, PRSS23, ANGPT2). Moreover, there was clear evidence for differential expression of glaucoma- associated genes involving elevated IOP vs. genes associated with IOP-independent glaucoma: the former showed preferential expression in conventional outflow cells, whereas the latter were more highly expressed by retinal ganglion cells. Future work needs to focus on expression profiles of outflow cells in ocular hypertensive versus normotensive eyes, and in eyes over a range of ages (mean eye donors age here was 67 years old).

An important feature of this study was the comparison of human transcriptomic profiles to those from four different model species (two monkeys, mouse and pig). In general, there was good conservation of expression patterns and markers across species, with the greatest source of variability being in the expression patterns of TM cells. Interestingly, despite the anatomical differences between the continuous SC of human, monkey and mouse, the transcriptome of pig angular aqueous plexus cells was similar to SC cells. All four model species also contained abundant CD63⁺/LYVE1⁺ macrophages in their conventional outflow pathway, suggesting an important physiological role. A limitation to the analyses of mouse eyes was that profiling was performed only on albino CD1s. Future work needs to compare the profile of CD1 with that of pigmented mice such as the commonly used C57Bl/6.

It is now clear that generation and regulation of IOP likely involves a complex interplay between many cell types in the outflow pathway

In summary, it is now clear that generation and regulation of IOP likely involves a complex interplay between many cell types in the outflow pathway. This atlas of conventional outflow pathway cells provides a valuable resource that will guide many future studies attempting to better understand the molecular basis for IOP homeostasis in heath, and dysregulation resulting in ocular hypertension.

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