Endothelial Cell Markers: Key Insights for Vascular Biology

Endothelial cells line blood vessels and play a crucial role in vascular function, including barrier formation, inflammation regulation, and angiogenesis. Identifying these cells is essential for studying vascular biology and diseases such as atherosclerosis and cancer.

To distinguish endothelial cells from other types, researchers rely on molecular markers, which vary based on vessel type, physiological conditions, and disease states.

Common Cell Surface Markers

Endothelial cells express distinct surface proteins that help researchers identify them in different vascular environments. These markers contribute to cell adhesion, signaling, and structural integrity. Among the most widely used are CD31, VE-cadherin, and CD34.

CD31

CD31, or platelet endothelial cell adhesion molecule-1 (PECAM-1), is a transmembrane glycoprotein primarily expressed on endothelial cells and certain immune cells. It facilitates cell-cell adhesion and maintains vascular integrity. Research in Circulation Research (2021) highlights its role in endothelial barrier function and leukocyte transmigration. CD31 is consistently expressed across most blood vessels, making it a reliable marker in immunohistochemistry and flow cytometry. However, since it is also found on platelets and leukocytes, researchers often use it alongside other endothelial-specific markers for greater specificity.

VE-Cadherin

VE-cadherin (CD144) is an endothelial-specific adhesion protein essential for maintaining vascular permeability and cell junction stability. It mediates endothelial cell-cell interactions and plays a key role in angiogenesis. A study in Nature Communications (2022) demonstrated that VE-cadherin phosphorylation regulates endothelial barrier integrity in response to inflammation. Unlike CD31, VE-cadherin is almost exclusively found in endothelial cells, making it highly specific. Disruptions in VE-cadherin signaling contribute to vascular leakage in conditions such as sepsis and tumor metastasis.

CD34

CD34 is a transmembrane glycoprotein that marks vascular endothelial progenitor cells and hematopoietic stem cells. It is prominent in capillaries and smaller blood vessels, with variable levels in larger vessels. A report in The Journal of Pathology (2023) noted that CD34 is particularly useful for identifying newly formed vasculature, as its expression increases during angiogenesis. Unlike CD31 and VE-cadherin, CD34 is not involved in adhesion but contributes to cell migration and vessel formation. Since it is also present on hematopoietic progenitor cells, CD34 is often used in combination with other endothelial markers to ensure specificity.

Notable Intracellular Markers

In addition to surface proteins, endothelial cells express intracellular markers involved in coagulation, nitric oxide production, and activation. Unlike surface markers, intracellular markers require permeabilization techniques for detection. Among the most studied are von Willebrand factor (vWF), E-selectin, and endothelial nitric oxide synthase (eNOS).

vWF

Von Willebrand factor (vWF) is a glycoprotein stored in endothelial Weibel-Palade bodies and secreted into the bloodstream to facilitate platelet adhesion and clotting. It is a well-established endothelial marker, particularly in larger vessels. A study in Blood (2022) highlighted vWF’s role in hemostasis, showing that its deficiency leads to von Willebrand disease. Beyond coagulation, vWF responds to vascular injury, increasing under shear stress and inflammation. Its expression is higher in arteries than veins, making it useful for distinguishing vascular subtypes. Immunohistochemical staining for vWF confirms endothelial origin in tissue samples, particularly in tumor vasculature and thrombotic disorders.

E-Selectin

E-selectin, or endothelial-leukocyte adhesion molecule-1 (ELAM-1), is an inducible adhesion molecule expressed on activated endothelial cells. Unlike constitutively expressed markers, E-selectin is upregulated in response to cytokines and shear stress. Research in The Journal of Clinical Investigation (2023) demonstrated that E-selectin expression increases during endothelial activation, particularly in inflammatory and angiogenic environments. This transient expression makes it useful for studying endothelial responses in wound healing and tumor progression. E-selectin is synthesized in the endoplasmic reticulum and transported to the membrane upon activation, but intracellular detection methods such as immunofluorescence can reveal its presence before surface expression.

eNOS

Endothelial nitric oxide synthase (eNOS) produces nitric oxide (NO), a signaling molecule that regulates vascular tone and blood flow. It is predominantly localized in endothelial cells, where it associates with caveolae in the plasma membrane and the Golgi apparatus. A study in Circulation Research (2021) found that eNOS activity is modulated by phosphorylation and intracellular calcium levels, influencing endothelial function under different conditions. Reduced eNOS expression is linked to endothelial dysfunction, a precursor to cardiovascular diseases such as hypertension and atherosclerosis. Immunodetection of eNOS assesses endothelial health, with higher expression correlating with improved vascular relaxation and lower oxidative stress.

Variation Among Vessels

Endothelial cells differ across the vascular system, adapting to the structural and functional demands of arteries, veins, and capillaries. These differences extend to gene expression, signaling pathways, and responses to physiological stimuli.

Arterial endothelial cells experience higher shear stress due to pulsatile blood flow and elevated pressure. They express mechanosensitive proteins that help maintain vascular tone and resist oxidative stress. Studies using single-cell RNA sequencing show that arterial endothelial cells upregulate nitric oxide signaling genes, aiding vasodilation and preventing arterial stiffening.

Venous endothelial cells function under lower shear stress and pressure, resulting in a thinner basement membrane and different molecular expression patterns. They produce fewer mechanotransduction proteins and rely more on passive blood flow regulation. Unlike arterial counterparts, venous endothelial cells have a greater tendency for leukocyte adhesion under inflammatory conditions, making them more prone to deep vein thrombosis. Their ability to undergo phenotypic shifts in response to metabolic and oxygenation changes contributes to adaptive vascular remodeling.

Capillary endothelial cells are specialized for nutrient and gas exchange, with their structure varying by organ. Brain capillaries form the blood-brain barrier with tightly regulated junctions, while liver and spleen capillaries contain fenestrations for rapid macromolecule exchange. This heterogeneity underscores endothelial adaptability, aligning molecular expression with tissue-specific needs.

Factors Influencing Marker Expression

Endothelial marker expression shifts in response to mechanical forces, metabolic conditions, and biochemical signals. Shear stress from blood flow modulates adhesion molecules and junctional proteins, with laminar flow promoting stability and turbulent flow triggering inflammation and angiogenesis.

Oxygen availability further influences endothelial characteristics. Hypoxia-inducible factors (HIFs) regulate transcription of angiogenesis-related markers, increasing protein expression to facilitate new vessel formation in ischemic conditions. Metabolic state also affects marker expression, with glycolysis and oxidative phosphorylation influencing surface proteins involved in nutrient transport and intercellular communication.

Methods To Detect Markers

Identifying endothelial cells requires molecular and imaging techniques suited to different experimental and clinical settings. Immunohistochemistry (IHC) is widely used for visualizing endothelial markers within tissue sections. Using antibodies conjugated to enzymes or fluorophores, researchers detect proteins such as CD31, VE-cadherin, and vWF while preserving spatial context. This technique is valuable in pathology, where endothelial markers differentiate tumor vasculature from normal blood vessels. However, tissue fixation can sometimes obscure antigenic sites, requiring optimized staining protocols.

Flow cytometry provides single-cell analysis, using fluorescently labeled antibodies to detect surface and intracellular proteins in suspended cells. It allows rapid, high-throughput quantification and simultaneous measurement of multiple markers, revealing endothelial heterogeneity. Studies on circulating endothelial cells in cardiovascular disease use flow cytometry to assess endothelial injury by quantifying CD31+CD34+ cells in blood samples.

Western blotting detects endothelial markers in lysed cell populations, offering robust protein quantification for assessing expression changes. Though it lacks spatial information, it is effective for evaluating marker levels under different conditions. Emerging technologies such as single-cell RNA sequencing further refine endothelial identification by characterizing gene expression profiles, providing unprecedented resolution in distinguishing endothelial subsets across vascular beds.