The endothelial cell-specific proteome

The function of the brain, defined as the central nervous system, is to receive, process and execute the coordinated higher functions of perception, motion and cognition that signify human life.

The human circulatory system is composed of blood and lymphatic vessels that transport blood and lymph throughout the body. Our bodies rely heavily on our vascular system to function since it is the key to how our tissues and organs receive and dispose of nutrients, gases, rest products, and pathogens. The innermost layer of cells in a vessel is a single layer of squamous endothelial cells. Endothelial cells form the barrier between vessels and tissue in every type of vessel there is, from blood vessels such as arteries, veins, and capillaries to lymphatic vessels. This barrier separates the blood or lymph from the rest of the vessel wall, creating an interface between the two that enables control over the exchange of substances in and out of surrounding tissues.

  • 970 elevated genes
  • 152 enriched genes
  • 249 group enriched genes
  • Main function: Support and regulation of the circulatory system

Transcriptome analysis shows that 68% (n=13764) of all human proteins (n=20162) are detected in endothelial cells and 970 of these genes show an elevated expression in any endothelial cells compared to other brain cell types.

The endothelial cell transcriptome

The snRNA-seq-based endothelial cell transcriptome can be analyzed with regard to specificity, illustrating the number of genes with elevated expression in endothelial cell type compared to other brain cell types (Table 1). Genes with an elevated expression are divided into three subcategories:

  • Cell type enriched: At least four-fold higher mRNA level in a certain cell type compared to any other cell type within the brain.
  • Group enriched: At least four-fold higher average mRNA level in a group of 2-10 cell types compared to any other cell type within the brain.
  • Cell type enhanced: At least four-fold higher mRNA level in a cell certain cell type compared to the average level in all other cell types within the brain.

Table 1. Number of genes in the subdivided specificity categories of elevated expression in the analyzed endothelial cell types.
Cell type
Tissue origin
Cell type enriched
Group enriched
Cell type enhanced
Total elevated
Endothelial cell Amygdala, Basal ganglia, Cerebellum, Cerebral cortex, Hippocampus, Hypothalamus, Medulla oblongata, Midbrain, Pons, Spinal cord, Thalamus 152 249 569 970

Endothelial cells

As shown in Table 1, 970 genes are elevated in Endothelial cells compared to other cell types of the brain.

The endothelial cells of the brain can be compared with the endothelial cells in peripheral tissues, where 349 genes are highlighted for an elevated (enriched and enhanced categories combined) expression in endothelial cells. 226 of those genes are overlapping. LCN6 and SELE being the only two genes classified as endothelial cell enriched in the peripheral cell type data as well as the single nuclei brain data. Below are barplots showing the expression profile in peripheral tissues and a brain regions.

0100200300400500600nTPM
LCN6 - Testis
0246810nTPM
LCN6 - Cerebral cortex
05001,0001,5002,0002,5003,000nTPM
SELE - Fallopian tube
020406080100120nTPM
SELE - Basal ganglia

There are several genes with enriched expression in brain endothelial cells, that also show expression in other cell types besides endothelial cells when comparing to the peripheral tissue types. Some examples are Fms related receptor tyrosine kinase 1 (FLT1), CD34 and Nitric oxide synthase 3 (NOS3). These three examples can be confirmed on protein level using immunohistochemistry and antibody labeling.


CD34 - cerebral cortex
050100150200250nTPM
CD34 - Cerebral cortex

CD34


FLT1 - cerebral cortex
05001,0001,5002,0002,5003,000nTPM
FLT1 - Cerebral cortex

FLT1


NOS3 - cerebral cortex
020406080100nTPM
NOS3 - Cerebral cortex

NOS3

Exploring the vascular cluster

In the Human Brain Cell Atlas v1.0 you can explore the specific clusters of cells and their expression in single cell level. The vascular cluster can be explored to further investigate subtypes of vascular cells across the different brain regions. Since this cluster includes all the vasculature cells, looking at the expression pattern of markers such as, CD34 for endothelial cells, CSPG4 for pericytes and PLN for smooth muscle cells you can clearly see the clustering of the vascular cell types. Or switch on and off the cell type selection in the selection panel to the left.

The Endothelial cell function

The cardiovascular system is considered a closed system due to blood never leaving the vessels. Nutrients and oxygen are regulated via diffusion over the vascular endothelial layer into the interstitial fluid, which transport compounds to target cells and vice versa. In contrast, the lymphatic circulatory system is an open system that collects and transports waste products, damaged cells, and bacteria over the endothelial layer from the interstitial fluid via lymphatic capillaries. These capillaries then drain the collected lymph into lymphatic vessels, which transport it through numerous lymphatic organs and ducts where waste products are filtered out. The filtered fluids are then returned to the blood circulation.

The vascular wall consists of three layers, the tunica intima, media, and adventitia. The outermost layer (tunica adventitia), is mainly composed of collagen that anchors the vessels to nearby organs, giving them stability. The middle layer (tunica media) consists of smooth muscle cells, while the innermost layer (tunica intima) consists of a single layer of squamous endothelial cells facing the lumen and a layer of elastic tissue called elastica interna.

Endothelial cells form the barrier between vessels and tissue in every type of vessel there is. Depending on vessel type the endothelial cells are classified as either vascular endothelial cells (in direct contact with blood) or lymphatic endothelial cells (in direct contact with lymph). Both types have unique functions throughout the circulatory system such as aiding in upholding homeostasis, fluid filtration, blood vessel tone, and hormone trafficking. Any impaired function can lead to serious health issues.

The histology of organs that contain endothelial cells, including interactive images, is described in the Protein Atlas Histology Dictionary.

Background

Single nuclei RNAseq data

Siletti K et al. (2023) published single nuclei RNA sequencing result, based on over 3 million cells from multiple brain regions, in Science magazine and created an interactive portal (The Human Brain Cell Atlas v1.0 ) available for single cell exploration across human gene expression in healthy brain cells. The Human Protein Atlas aims to generate a comprehensive resource representing the human body and its complexity, and with a need for better representation of the different cell types of the human brain, we imported the expression profiles and grouped them based on our cell type- strategy (providing bar charts of pooled data for each cell type cluster and calculating the average normalized protein-coding transcripts per million). We based the cell type clusters on the 31 superclusters, as well as the provided assigned cell types, and the data is shown as 34 different "supercluster cell types". The expression profile of the different clusters are shown for each of the 11 different brain regions. More details, related to number of M reads and number of cells per brain region/UMAP can be found here. The published cerebral cortex data is represented by a larger number of cells and we only included a random selection of 500 thousand cells. In total, expression data for 2526725 brain cells is displayed in the Brain single nuclei resource, for browsing the gene expression and provide easy comparison to cell type expression in peripheral tissues.

Relevant publications

Uhlén M et al., Tissue-based map of the human proteome. Science (2015)
PubMed: 25613900 DOI: 10.1126/science.1260419

Fagerberg L et al., Analysis of the human tissue-specific expression by genome-wide integration of transcriptomics and antibody-based proteomics. Mol Cell Proteomics. (2014)
PubMed: 24309898 DOI: 10.1074/mcp.M113.035600

Sjöstedt E et al., An atlas of the protein-coding genes in the human, pig, and mouse brain. Science. (2020)
PubMed: 32139519 DOI: 10.1126/science.aay5947

Siletti K et al., Transcriptomic diversity of cell types across the adult human brain. Science. (2023)
PubMed: 37824663 DOI: 10.1126/science.add7046