The microvascular endothelial cell monolayer localized at the critical interface between the blood and vessel wall has the vital functions of regulating tissue fluid balance and supplying the essential nutrients needed for the survival of the organism. The endothelial cell is an exquisite "sensor" that responds to diverse signals generated in the blood, subendothelium, and interacting cells. The endothelial cell is able to dynamically regulate its paracellular and transcellular pathways for transport of plasma proteins, solutes, and liquid. The semipermeable characteristic of the endothelium (which distinguishes it from the epithelium) is crucial for establishing the transendothelial protein gradient (the colloid osmotic gradient) required for tissue fluid homeostasis. Interendothelial junctions comprise a complex array of proteins in series with the extracellular matrix constituents and serve to limit the transport of albumin and other plasma proteins by the paracellular pathway. This pathway is highly regulated by the activation of specific extrinsic and intrinsic signaling pathways. Recent evidence has also highlighted the importance of the heretofore enigmatic transcellular pathway in mediating albumin transport via transcytosis. Caveolae, the vesicular carriers filled with receptor-bound and unbound free solutes, have been shown to shuttle between the vascular and extravascular spaces depositing their contents outside the cell. This review summarizes and analyzes the recent data from genetic, physiological, cellular, and morphological studies that have addressed the signaling mechanisms involved in the regulation of both the paracellular and transcellular transport pathways.

https://www.physiology.org/doi/pdf/10.1152/physrev.00012.2005

Signaling Mechanisms Regulating Endothelial Permeability

Alpha-1-acid glycoprotein.

The endothelial lining of blood vessels presents a large surface area for exchange of materials between blood and tissues, and is critically involved in many other processes such as regulation of blood flow, inflammatory responses and blood coagulation. It has long been known that the luminal surface of the endothelium is lined with a glycocalyx, a layer of membrane-bound macromolecules which has been determined by electron microscopy to be several tens of nanometers thick. However, investigations in vivo have indicated the presence of a much thicker endothelial surface layer (ESL), with an estimated thickness ranging from 0.5 µm to over 1 µm, that restricts the flow of plasma and can exclude red blood cells and some macromolecular solutes. The evidence for the existence of the ESL, hypotheses about its composition and biophysical properties, its relevance to physiological processes, and its possible clinical implications are considered in this review.

The endothelial surface layer.

This review focuses on the intricate properties of the glomerular barrier. Other reviews have focused on podocyte biology, mesangial cells, and the glomerular basement membrane (GBM). However, since all components of the glomerular membrane are important for its function, proteinuria will occur regardless of which layer is affected by disease. We review the properties of endothelial cells and their surface layer, the GBM, and podocytes, discuss various methods of studying glomerular permeability, and analyze data concerning the restriction of solutes by size, charge, and shape. We also review the physical principles of transport across biological or artificial membranes and various theoretical models used to predict the fluxes of solutes and water. The glomerular barrier is highly size and charge selective, in qualitative agreement with the classical studies performed 30 years ago. The small amounts of albumin filtered will be reabsorbed by the megalin-cubulin complex and degraded by the proximal tubular cells. At present, there is no unequivocal evidence for reuptake of intact albumin from urine. The cellular components are the key players in restricting solute transport, while the GBM is responsible for most of the resistance to water flow across the glomerular barrier.

Properties of the Glomerular Barrier and Mechanisms of Proteinuria

The increase in endothelial permeability in response to inflammatory mediators such as alpha-thrombin and histamine is accompanied by cell rounding and interendothelial gap formation, implicating that the predominant transport pathway is a diffusive one [i.e., via cellular junctions (paracellular transport)]. However, the possible contribution by vesicle-mediated transport (i.e., via albumin binding protein gp60) to the overall permeability increase needs investigation. Regulation of paracellular transport in endothelial cells is associated with modulation of actin-based systems which anchor the cell to its neighbor or extracellular matrix, thus maintaining endothelial integrity. At the cell-cell junctions, actin is linked indirectly to the plasma membrane by linking proteins (e.g., vinculin, catenins, alpha-actinin) to cadherins, which function in homophilic intercellular adhesion. Cadherins may also play a role in regulating the formation of tight junctions, which also may be associated with actin. At endothelial focal contacts, the transmembrane receptors (integrins) for matrix proteins are linked to actin via linking proteins (i.e., vinculin, talin, alpha-actinin). In response to inflammatory mediators, second messengers signal two regulatory pathways which modulate the actin-based systems, which may lead to impairment of the endothelial barrier integrity. One pathway is based on protein kinase C (PKC) isozyme-specific phosphorylation of linking proteins at the cell-cell and cell-matrix junctions. The increased phosphorylation is associated with actin reorganization, cell rounding, and increased paracellular transport. The other is the activation of myosin light-chain kinase, (MLCK), which causes an actin-myosin-based contraction that may lead to a centripetal retraction of endothelial cells. Current research is in the identification of protein substrates of PKC isozymes, the specific role of their phosphorylation in barrier function, and determining the precise role of MLCK in modulation of endothelial barrier function.

https://www.physiology.org/doi/pdf/10.1152/ajplung.1994.267.3.L223

Regulation of vascular endothelial barrier function

The effects of human serum orosomucoid (normal serum concentration 0.7-1.0 g l-1) on capillary permeability were investigated in 12 isolated maximally vasodilated rat hindquarters perfused with bovine serum albumin (50 g l-1) in modified Tyrode. Measurements were made of capillary filtration coefficient (CFC), permeability surface area product (PS) for vitamin B12 and isogravimetric clearance of radiolabelled albumin (Cl alb). The results were compared with those obtained using perfusates without addition of orosomucoid ('albumin group') or perfusates containing horse serum ('serum group'). Clearance of albumin was almost four-fold higher in the albumin than in the serum group, 0.0895 +/- 0.0066 (n = 12) and 0.0252 +/- 0.016 ml min-1 per 100 g (n = 18), respectively, while intermediate Cl alb values were obtained with human orosomucoid in the perfusate (greater than 0.1 g l-1), 0.0436 +/- 0.0034 ml min-1 per 100 g) (n = 8). These changes in Cl alb were not accompanied by any differences in CFC or PS. We conclude that orosomucoid is one of the components in serum (besides albumin) needed for the maintenance of normal permselectivity of the capillary walls of rat skeletal muscle. Alternatively, human orosomucoid is structurally related to other substances exerting this 'serum effect'.

Orosomucoid as one of the serum components contributing to normal capillary permselectivity in rat skeletal muscle

Experiments were performed on isolated maximally vasodilated perfused rat hindquarters to evaluate the role of calcium and magnesium for the capillary permeability increase(s) elicited by histamine. Changes in capillary permeability were quantified by determinations of capillary filtration coefficient (CFC) with gravimetric technique, and capillary diffusion capacity (PS) for vitamin B12 (MW = 1,355) with a single injection indicator dilution technique. During control, vascular resistance was 2.2 PRU100 at a flow of 9.4 ml min-1 per 100 g, and PS for B12 was 3.7 +/- 0.1 ml min-1 per 100 g, while CFC was 0.0377 +/- 0.0004 ml min-1 mmHg-1 per 100 g. Perfusion with 'Mg-free' solution for 1 h caused a 24% increase in CFC, while neither 'Ca-free' perfusion nor perfusion with verapamil (5 X 10(-5) M) nor felodipine (1 X 10(-6) M) induced any changes in CFC. Histamine (100-200 microM) caused in all preparations a 150-200% increase in CFC with only small changes in PS for B12. This histamine effect was absent after 1 h of 'Ca-free' perfusion and was partially blocked after 1 h of perfusion with 0.1 mM calcium, while the calcium antagonists verapamil and felodipine had no effects on the histamine-induced changes. The results imply that histamine exerts its action on the endothelial cells through a calcium-dependent process, probably involving low affinity calcium sites but this process could not be inhibited by the calcium antagonists used. Thus, endothelial cell contractility, which probably is responsible for the histamine-induced increase in capillary permeability, exhibits unique characteristics, differing from those of vascular smooth muscle.

B. Rippe

Papers

Orosomucoid as one of the serum components contributing to normal capillary permselectivity in rat skeletal muscle

Calcium dependence of histamine-induced increases in capillary permeability in isolated perfused rat hindquarters.