Abstract
Purpose :
Chronic inflammation of the lacrimal gland (LG) is a leading cause of aqueous deficiency dry eye. It results in eye irritation, pain and may lead to severe ocular surface disorders. Visium technology makes it possible to measure the topography of gene expression. Here, we propose to underly the potentials of Visium to compare healthy and chronically inflamed LGs.
Methods :
We used the 10X Genomics Visium spatial expression solution on 10µm LG cryosections of 4.5 months old BALB/cJ (control) and NOD.B10Sn-H2b/J (diseased) mice. Each sample was sequenced at a minimal depth of 280 million reads. Raw data was processed with Space Ranger (v1.3.1) to obtain spatial maps of gene expression. Data was normalized and integrated with Seurat (v4.0) to perform clustering and expression analysis.
Results :
After integration of data from BALBc and NOD mice, 7 clusters (C0-6) were identified. Overall, we noted in each cluster, an upregulation of immunoglobulins (Ig) and a downregulation of exocrine gland-secreting peptides and secretoglobins (SCGB) in NOD mice. C0 and C1 represented secretory units containing mostly acinar and myoepithelial cells expressing SCGB, which play a barrier function in secretory organs. C2 was characterized by the expression of Ig typical of B cells. In C3, we detected genes expressed by activated lymphocytes and macrophages. In BALBc, it represented 5% of all spots, while in NOD it reached 15% and colocalized with immune infiltrates. C3 is enriched in glycoproteins belonging to the major histocompatibility complex protein family. C4 and C5 were enriched in genes found in ductal cells. C4 likely contained cells releasing mucins and participating in ion exchange. C5 corresponded to excretory ducts. C6 was enriched in fibroblast and macrophage markers.
Conclusions :
We successfully identified the major cellular components of healthy and chronically inflamed LG: secretory units, major ducts and immune cells. Contrary to RNA-seq, Visium indicates which cellular compartment contributes to gene expression changes observed during disease development. Despite the relatively low spatial resolution of this approach, each compartment can be compared between control and diseased LG to evidence local or structure-specific transcriptomic alterations. For example, the downregulation of secreted peptides by the acinar compartment can be related to LG dysfunction and could be involved in immunoregulatory processes.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.