Medical Physiology A Cellular and Molecular Approach, Updated 2nd Ed.

THE MICROCIRCULATION

Emile L. Boulpaep

The microcirculation serves both nutritional and non-nutritional roles

The primary function of the cardiovascular system is to maintain a suitable environment for the tissues. The microcirculation is the “business end” of the system. The capillary is the principal site for exchange of gases, water, nutrients, and waste products. In most tissues, capillary flow exclusively serves these nutritional needs. In a few tissues, however, a large portion of capillary flow is non-nutritional. For example, in the glomeruli of the kidneys, capillary flow forms the glomerular filtrate (see Chapter 34). Blood flow through the skin, some of which may shunt through arteriovenous anastomoses, plays a key role in temperature regulation (see Chapter 61). Capillaries also serve other non-nutritional roles, such as signaling (e.g., delivery of hormones) and host defense (e.g., delivery of platelets). In the first part of this chapter, we discuss the nutritional role of capillaries and examine how gases, small water-soluble substances, macromolecules, and water pass across the endothelium. In the last two subchapters, we discuss lymphatics as well as the regulation of the microcirculation.

The morphology and local regulatory mechanisms of the microcirculation are designed to meet the particular needs of each tissue. Because these needs are different, the structure and function of the microcirculation may be quite different from one tissue to the next.

The microcirculation extends from the arterioles to the venules

The microcirculation is defined as the blood vessels from the first-order arteriole to the first-order venule. Although the details vary from organ to organ, the principal components of an idealized microcirculation include a single arteriole and venule, between which extends a network of true capillaries (Fig. 20-1). Sometimes a metarteriole—somewhat larger than a capillary—provides a shortcut through the network. Both the arteriole and the venule have vascular smooth muscle cells (VSMCs). Precapillary sphincters—at the transition between a capillary and either an arteriole or a metarteriole—control the access of blood to particular segments of the network. Sphincter closure or opening creates small local pressure differences that may reverse the direction of blood flow in some segments of the network.

Figure 20-1 Idealized microcirculatory circuit.

Arteries consist of an inner layer of endothelium, an internal elastic lamina, and a surrounding sheath of at least two continuous layers of innervated VSMCs (see Chapter 19). The inner radius of terminal arteries (called feed arteries in muscle) may be as small as 25 μm. Arterioles (inner radius, 5 to 25 μm) are similar to arteries but have only a single continuous layer of VSMCs, which are innervated. Metarterioles are similar to arterioles but of shorter length. Moreover, their VSMCs are discontinuous and are not usually innervated. The precapillary sphincter is a small cuff of smooth muscle that usually is not innervated but is very responsive to local tissue conditions. Relaxation or contraction of the precapillary sphincter may modulate tissue blood flow by an order of magnitude or more. Metarterioles and precapillary sphincters are not found in all tissues.

True capillaries (inner radius, 2 to 5 μm) consist of a single layer of endothelial cells surrounded by a basement membrane, a fine network of reticular collagen fibers, and—in some tissues—pericytes. The endothelial cells have a smooth surface and are extremely thin (as little as 200 to 300 nm in height), except at the nucleus. The thickness and density of the capillary basement membrane vary among organs. Where large transcapillary pressures occur or other large mechanical forces exist, the basement membrane is thickest. Some endothelial cells have, on both luminal and basal surfaces, numerous pits called caveolae that are involved in ligand binding. Fluid-phase and receptor-mediated endocytosis can result in 70-nm caveolin-coated vesicles (see Chapter 2). In addition, the cytoplasm of capillary endothelial cells is rich in other endocytotic (pinocytotic) vesicles that contribute to the transcytosis of water and water-soluble compounds across the endothelial wall. In some cases, the endocytotic vesicles are lined up in a string and even appear linked together to form a transendothelial channel.

Linking endothelial cells together are interendothelial junctions (Fig. 20-2) where the two cell membranes are ~10 nm apart, although there may be constricted regions where the space or cleft between the two cells forms adhering junctions only ~4 nm wide. Tight junctions may also be present, in which the apposed cell membranes appear to fuse and claudins 1, 3, and 5 (CLDN1, 3, 5) as well as occludin seal the gap (see Chapter 2). CLDN5 is quite specific for endothelial cells. Occludin is not found in all endothelia.

Figure 20-2 Capillary endothelial junctions. This electron micrograph shows the interendothelial junction between two endothelial cells in a muscle capillary. The arrows point to tight junctions. (From Fawcett DW: Bloom and Fawcett. A Textbook of Histology, 12th ed, p 964. New York: Chapman & Hall, 1994.)

Some endothelial cells have membrane-lined, cylindrical conduits—fenestrations—that run completely through the cell, from the capillary lumen to the interstitial space. These fenestrations are 50 to 80 nm in diameter and are seen primarily in tissues with large fluid and solute fluxes across the capillary walls (e.g., intestine, choroid plexus, exocrine glands, and renal glomeruli). A thin diaphragm often closes the perforations of the fenestrae (e.g., in intestinal capillaries).

The endothelia of the sinusoidal capillaries in the liver, bone marrow, and spleen have very large fenestrations as well as gaps 100 to 1000 nm wide between adjacent cells. Vesicles, transendothelial channels, fenestrae, and gaps—as well as structures of intermediate appearance—are part of a spectrum of regulated permeation across the endothelial cells.

Capillaries fall into three groups, based on their degree of leakiness (Fig. 20-3):

1. Continuous capillary. This is the most common form of capillary, with interendothelial junctions 10 to 15 nm wide (e.g., skeletal muscle). However, these clefts are absent in the blood-brain barrier (see Chapter 11), whose capillaries have narrow tight junctions.

2. Fenestrated capillary. In these capillaries, the endothelial cells are thin and perforated with fenestrations. These capillaries most often surround epithelia (e.g., small intestine, exocrine glands).

3. Discontinuous capillary. In addition to fenestrae, these capillaries have large gaps. Discontinuous capillaries are found in sinusoids (e.g., liver).

Figure 20-3 Three types of capillaries.

At their distal ends, true capillaries merge into venules (inner radius, 5 to 25 μm), which carry blood back into low-pressure veins that return blood to the heart. Venules have a discontinuous layer of VSMCs and therefore can control local blood flow. Venules may also exchange some solutes across their walls.

CAPILLARY EXCHANGE OF SOLUTES

The exchange of O₂ and CO₂ across capillaries depends on the diffusional properties of the surrounding tissue

Gases diffuse by a transcellular route across the two cell membranes and cytosol of the endothelial cells of the capillary with the same ease that they diffuse through the surrounding tissue. In this section, we focus primarily on the exchange of O₂. Very similar mechanisms exist for the exchange of CO₂, but they run in the opposite direction. Arterial blood has a relatively high O₂ level. As blood traverses a systemic capillary, the principal site of gas exchange, O₂ diffuses across the capillary wall and into the tissue space, which includes the interstitial fluid and the neighboring cells.

The most frequently used model of gas exchange is August Krogh’s tissue cylinder, a volume of tissue that a single capillary supplies with O₂ (Fig. 20-4A). The cylinder of tissue surrounds a single capillary. According to this model, the properties of the tissue cylinder govern the rate of diffusion of both O₂ and CO₂. The radius of a tissue cylinder in an organ is typically half the average spacing from one capillary to the next, that is, half the mean intercapillary distance. Capillary density and therefore mean intercapillary distance vary greatly among tissues. Among systemic tissues, capillary density is highest in tissues with high O₂ consumption (e.g., myocardium) and lowest in tissues consuming little O₂ (e.g., joint cartilage). Capillary density is extraordinarily high in the lungs (see Chapter 31).

Figure 20-4 Delivery and diffusion of O₂ to systemic tissues. In A, Krogh’s tissue cylinder consists of a single capillary (radius r_c), surrounded by a concentric cylinder of tissue (radius r_t) that the capillary supplies with O₂ and other nutrients. Blood flow into the capillary is F_in, and blood flow out of the capillary is F_out. The lower panel of A shows the profile of partial pressure of O₂ (P_O2) along the longitudinal axis of the capillary and the radial axis of the tissue cylinder.

The Krogh model predicts how the concentration or partial pressure of oxygen (P_O2) within the capillary lumen falls along the length of the capillary as O₂ exits for the surrounding tissues (Fig. 20-4A). The P_O2 within the capillary at any site along the length of the capillary depends on several factors:

1. The concentration of free O₂ in the arteriolar blood that feeds the capillary. This dissolved [O₂], which is the same in the plasma and the cytoplasm of the red blood cells, is proportional to the partial pressure of O₂ (see Chapter 29) in the arterioles.

2. The O₂ content of the blood. Less than 2% of the total O₂ in arterial blood is dissolved; the rest is bound to hemoglobin inside the red blood cells. Each 100 mL of arterial blood contains ~20 mL of O₂ gas, or 20 volume %—the O₂content (see Table 28-3).

3. The capillary blood flow (F).

4. The radial diffusion coefficient (D_r), which governs the diffusion of O₂ out of the capillary lumen. For simplicity, we assume that D_r is the same within the blood, the capillary wall, and the surrounding tissue and that it is the same along the entire length of the capillary.

5. The capillary radius (r_c).

6. The radius of tissue cylinder (r_t) that the capillary is supplying with O₂.

7. The O₂ consumption by the surrounding tissues (o₂).

8. The axial distance (x) along the capillary.

The combination of all these factors accounts for the shape of the concentration profiles within the vessel and the tissue. Although this model appears complicated, it is actually based on many simplifying assumptions. (See Note: Limitations of Krogh’s Tissue-Cylinder Model)

The O₂ extraction ratio of a whole organ depends primarily on blood flow and metabolic demand

In principle, beginning with a model like Krogh’s but more complete, one could sum up the predictions for a single capillary segment and then calculate gas exchange in an entire tissue. However, it is more convenient to pool all the capillaries in an organ and to focus on a single arterial inflow and single venous outflow. The difference in concentration of a substance in the arterial inflow and venous outflow of that organ is the arteriovenous (a-v) difference of that substance. For example, if the arterial O₂ content ([O₂]_a) entering the tissue is 20 mL O₂/dL blood and the venous O₂ content leaving it ([O₂]_v) is 15 mL O₂/dL blood, the O₂ a-v difference for that tissue is 5 mL O₂ gas/dL blood.

For a substance like O₂, which exits the capillaries, another way of expressing the amount that the tissues remove is the extraction ratio. This parameter is merely the a-v difference normalized to the arterial content of the substance. Thus, the extraction ratio of oxygen (EO₂) is

Thus, in our example:

In other words, the muscle in this example removes (and burns) 25% of the O₂ presented to it by the arterial blood.

What are the factors that determine O₂ extraction? To answer this question, we return to the hypothetical Krogh cylinder. The same eight factors that influence the P_O2 profiles in Figure 20-4A also determine the whole-organ O₂extraction. Of these factors, the two most important are capillary flow (item 3 in the list) and metabolic demand (item 7). The O₂ extraction ratio decreases with increased flow but increases with increased O₂ consumption. These conclusions make intuitive sense. Greater flow supplies more O₂, so the tissue needs to extract a smaller fraction of the incoming O₂ to satisfy its fixed needs. Conversely, increased metabolic demands require that the tissue extract more of the incoming O₂. These conclusions are merely a restatement of the Fick principle (see Chapter 17), which we can rewrite as

The term on the left is the a-v difference. The extraction ratio is merely the a-v difference normalized to [O₂]_a. Thus, the Fick principle confirms our intuition that the extraction ratio should increase with increasing metabolic demand but decrease with increasing flow.

Another important factor that we have so far ignored is that not all of the capillaries in a tissue may be active at any one time. For example, skeletal muscle contains roughly a half-million capillaries per gram of tissue. However, only ~20% are perfused at rest (Fig. 20-4B). During exercise, when the O₂ consumption of the muscle increases, the resistance vessels and precapillary sphincters dilate to meet the increased demand. This vasodilation increases muscle blood flow and the density of perfused capillaries (Fig. 20-4C). This response is equivalent to decreasing the tissue radius of Krogh’s cylinder because each perfused capillary now supplies a smaller region. Other things being equal, reduced diffusion distances cause P_O2 in the tissue to increase.

The velocity of blood flow in the capillaries also increases during exercise. All things being equal, this increased velocity would cause P_O2 to fall less steeply along the capillary lumen. For example, if the velocity were infinite, P_O2would not fall at all! In fact, because O₂ consumption rises during exercise, P_O2 actually falls more steeply along the capillary.

According to Fick’s law, the diffusion of small, water-soluble solutes across a capillary wall depends on both the permeability and the concentration gradient

Although the endothelial cell is freely permeable to O₂ and CO₂, it offers a significant barrier to the exchange of lipid-insoluble substances. Hydrophilic solutes that are smaller than albumin can traverse the capillary wall by diffusion through a paracellular route (i.e., through the clefts and interendothelial junctions as well as gaps and fenestrae, if these are present).

The amount of solute that crosses a particular surface area of a capillary per unit time is called a flux. It seems intuitive that the flux ought to be proportional to the magnitude of the concentration difference across the capillary wall and that it ought to be bigger in leakier capillaries (Fig. 20-5). These ideas are embodied in a form of Fick’s law: (See Note: Fick’s Law)

Figure 20-5 Diffusion of a solute across a capillary wall.

In Figure 20-5, J_X is the flux of the solute X [units: moles/(cm²/s)], assuming a positive J_X with flow out of the capillary, into the interstitial fluid. [X]_c and [X]_if are the dissolved concentrations of the solute in the capillary and interstitial fluid, respectively. Because the capillary wall thickness a (units: cm) is difficult to determine, we combined the diffusion coefficient D_X (units: cm²/s) and wall thickness into a single term (D_X/a) called P_X, the permeability coefficient (units: cm/s). Thus, P_X expresses the ease with which the solute crosses a capillary by diffusion.

Because, in practice, the surface area (S) of the capillary is often unknown, it is impossible to compute the flux of a solute, which is expressed per unit area. Rather, it is more common to compute the mass flow(), which is simply the amount of solute transferred per unit time (units: mol/s):

The whole-organ extraction ratio for small, hydrophilic solutes provides an estimate of the solute permeability of capillaries

How could we estimate the permeability coefficient for a solute in different capillaries or for different solutes in the same capillary? Unfortunately, it is difficult to determine permeability coefficients in single capillaries. Therefore, investigators use an indirect approach that begins with measurement of the whole-organ extraction ratio for the solute X. As we have already seen for O₂ (Equation 20-1), the extraction ratio (E_X) is a normalized a-v difference for X:

Thus, E_X describes the degree to which an organ removes a solute from the circulation. Unlike the situation for O₂, the extraction ratio for small hydrophilic solutes depends not only on total organ blood flow (F) but also on the overall “exchange properties” of all of its capillaries, expressed by the product of permeability and total capillary area (P_X · S). The dependence of E_X on the P_X · S product and F is described by the following equation:

Therefore, by knowing the whole-organ extraction ratio for a solute and blood flow through the organ, we can calculate the product P_X · S. The first column of Table 20-1 lists the P_X · S products for a single solute (inulin), determined from Equation 20-7, for a number of different organs. Armed with independent estimates of the capillary surface area (Table 20-1, column 2), we can compute P_X (column 3). P_Xincreases by a factor of approximately 4 from resting skeletal muscle to heart, reflecting a difference in the density of fluid-filled interendothelial clefts. Because a much greater fraction of the capillaries in the heart are open to blood flow (i.e., S is ~10-fold larger), the P_X · S product for heart is approximately 40-fold higher than for resting skeletal muscle.

Table 20-1 P_X · S Products for Various Capillary Beds

The cerebral vessels have unique characteristics that constitute the basis of the blood-brain barrier (see Chapter 11). The tight junctions of most brain capillaries do not permit any paracellular flow of hydrophilic solutes; therefore, they exhibit a very low permeability to sucrose or inulin, probably because of the abundant presence of CLDN5 and occludin. In contrast, the water permeability of cerebral vessels is similar to that of other organs. Therefore, a large fraction of water exchange in cerebral vessels must occur through the endothelial cells.

Whole-organ P_X · S values are not constant. First, arterioles and precapillary sphincters control the number of capillaries being perfused and thus the available surface area (S). Second, in response to a variety of signaling molecules (e.g., cytokines), endothelial cells can reorganize their cytoskeleton, thereby changing their shape. This deformation widens interendothelial clefts and increases P_X. One example is the increased leakiness that develops during inflammation in response to the secretion of histamine by mast cells and basophilic granulocytes. (See Note: Pore Theory)

Small polar molecules have a relatively low permeability because they can traverse the capillary wall only by diffusing through water-filled pores (small-pore effect)

Having compared the permeabilities of a single hydrophilic solute (inulin) in several capillary beds, we may address the selectivity of a single capillary wall to several solutes. Table 20-2 shows that the permeability coefficient falls as molecular radius rises. For lipid-soluble substances such as CO₂ and O₂, which can diffuse through the entire capillary endothelial cell and not just the water-filled pathways, the permeability is much larger than for the solutes in Table 20-2. Early physiologists had modeled endothelial permeability for hydrophilic solutes on the basis of two sets of pores: large pores with diameter of ~10 nm or more and a larger number of small pores with an equivalent radius of 3 nm. Small, water-soluble, polar molecules have a relatively low permeability because they can diffuse only by a paracellular path through interendothelial clefts or other water-filled pathways, which constitute only a fraction of the total capillary area. Discontinuities or gaps in tight junction strands could form the basis for the small pores. Alternatively, the molecular sieving properties of the small pores may reside in a fiber matrix (Fig. 20-6) that consists of either a meshwork of glycoproteins in the paracellular clefts (on the abluminal side of the tight junctions) or the glycocalyx on the surface of the endothelial cell (on the luminal side of the tight junctions). The endothelium-specific calcium-dependent adhesion molecule VE-cadherin (CDH5; see Chapter 2) and platelet/endothelial cell adhesion molecule (PECAM1 or CD31 antigen) are important glycoprotein components for the postulated fiber matrix in the paracellular clefts. In fact, the small-pore effect is best explained by an arrangement of discontinuities in the tight junctional strands in series with a fiber matrix on either side of the tight junction. (See Note: Stokes-Einstein Radius)

Figure 20-6 Model of endothelial junctional complexes. The figure shows two adjacent endothelial cell membranes at the tight junction, with a portion of the membrane of the upper cell cut away. (Data from Firth JA: Endothelial barriers: From hypothetical pores to membrane proteins. J Anat 2002; 200:541-548.)

Table 20-2 Permeability Coefficients for Lipid-Insoluble Solutes*

Interendothelial clefts are wider—and fenestrae are more common—at the venular end of the capillary than at its arteriolar end, so that P_X increases along the capillary. Therefore, if the transcapillary concentration difference ([X]_c− [X]_if) were the same, the solute flux would actually be larger at the venous end of the microcirculation.

Small proteins can also diffuse across interendothelial clefts or through fenestrae. In addition to molecular size, the electrical charge of proteins and other macromolecules is a major determinant of their apparent permeability coefficient. In general, the flux of negatively charged proteins is much smaller than that of neutral macromolecules of equivalent size, whereas positively charged macromolecules have the highest apparent permeability coefficient. Fixed negative charges in the endothelial glycocalyx exclude macromolecules with negative charge and favor the transit of macromolecules with positive charge. Selective permeability based on the electrical charge of the solute is a striking feature of the filtration of proteins across the glomerular barrier of the nephron (see Chapter 34).

The diffusive movement of solutes is the dominant mode of transcapillary exchange. However, the convective movement of water can also carry solutes. This solvent drag is the flux of a dissolved solute that is swept along by the bulk movement of the solvent. Compared with the diffusive flux of a small solute with a high permeability coefficient (e.g., glucose), the contribution of solvent drag is minor.

The exchange of macromolecules across capillaries can occur by transcytosis (large-pore effect)

Macromolecules with a radius exceeding 1 nm (e.g., plasma proteins) can cross the capillary, at a low rate, through wide intercellular clefts, fenestrations, and gaps—when they are present. However, it is caveolae that are predominantly responsible for the large-pore effect that allows transcellular translocation of macromolecules. The transcytosis of very large macromolecules by vesicular transport involves (1) equilibration of dissolved macromolecules in the capillary lumen with the fluid phase inside the open vesicle; (2) pinching off of the vesicle; (3) vesicle shuttling to the cytoplasm and probably transient fusion with other vesicles within the cytoplasm, allowing intermixing of the vesicular content; (4) fusion of vesicles with the opposite plasma membrane; and (5) equilibration with the opposite extracellular fluid phase.

Although one can express the transcytotic movement of macromolecules as a flux, the laws of diffusion (Equation 20-4) do not govern transcytosis. Nevertheless, investigators have calculated the “apparent permeability” of typical capillaries to macromolecules (Table 20-3). The resulting “permeability”—which reflects the total movement of the macromolecule, regardless of the pathway—falls off steeply with increases in molecular radius, a feature called sieving. This sieving may be the result of steric hindrance when large macromolecules equilibrate across the neck of nascent vesicles or when a network of glycoproteins in the glycocalyx above the vesicle excludes the large macromolecules. In addition, sieving of macromolecules according to molecular size could occur as macromolecules diffuse through infrequent chains of fused vesicles that span the full width of the endothelial cell. (See Note: Fast vs Slow Pathways for Exchange of Macromolecules across Capillary Walls)

Table 20-3 Capillary Permeability to Macromolecules

Transcytosis is not as simple as the luminal loading and basal unloading of ferryboats—the cell may process some of the cargo. Although the luminal surface of endothelial cells avidly takes up ferritin (750 kDa), only a tiny portion of endocytosed ferritin translocates to the opposite side of the cell (Table 20-3). The remainder stays for a time in intracellular compartments, where it is finally broken down.

Both transcytosis and chains of fused vesicles are less prominent in brain capillaries. The presence of continuous tight junctions and the low level of transcytosis account for the blood-brain barrier’s much lower apparent permeability to macromolecules.

CAPILLARY EXCHANGE OF WATER

Fluid transfer across capillaries is convective and depends on net hydrostatic and osmotic forces (i.e., Starling forces)

The pathway for fluid movement across the capillary wall is a combination of transcellular and paracellular pathways. Endothelial cell membranes express constitutively active aquaporin 1 (AQP1) water channels (see Chapter 5). It is likely that AQP1 constitutes the principal transcellular pathway for water movement. The interendothelial clefts, fenestrae, or gaps may be the anatomical substrate of the paracellular pathway.

Whereas the main mechanism for the transfer of gases and other solutes is diffusion, the main mechanism for the net transfer of fluid across the capillary membrane is convection. As first outlined in 1896 by Ernest Starling, the two driving forces for the convection of fluid—or bulk water movement—across the capillary wall are the transcapillary hydrostatic pressure difference and effective osmotic pressure difference, also known as the colloid osmotic pressure or oncotic pressure difference (see Chapter 5). (See Note: Ernest Henry Starling)

The hydrostatic pressure difference (ΔP) across the capillary wall is the difference between the intravascular pressure (i.e., capillary hydrostatic pressure, P_c) and the extravascular pressure (i.e., interstitial fluid hydrostatic pressure, P_if). Note that the term hydrostatic includes all sources of intravascular pressure, not only that derived from gravity; it is used here in opposition to osmotic.

The colloid osmotic pressure difference (Δπ) across the capillary wall is the difference between the intravascular colloid osmotic pressure caused by plasma proteins (π_c) and the extravascular colloid osmotic pressure caused by interstitial proteins and proteoglycans (π_if). A positive ΔP tends to drive water out of the capillary lumen, whereas a positive Δπ attracts water into the capillary lumen.

Starling’s hypothesis to describe the volume flow (J_V) of fluid across the capillary wall is embodied in the following equation, which is similar to Equation 5-27:Table 20-4 summarizes the terms in this equation. The equation is written so that the flux of water leaving the capillary is positive and that of fluid entering the capillary is negative.

Table 20-4 Terms in Starling Equation

The hydraulic conductivity (L_p) is the proportionality constant that relates the net driving force to J_V and expresses the total permeability provided by the ensemble of AQP1 channels and the paracellular pathway. (See Note: Hydraulic Conductivity vs Water Permeability Coefficient)

According to van’t Hoff’s law, the theoretical colloid osmotic pressure difference (Δπ_theory) is proportional to the protein concentration difference (Δ[X]):

However, because capillary walls exclude proteins imperfectly, the observed colloid osmotic pressure difference (Δπ_obs) is less than the ideal. The ratio Δπ_obs/Δπ_theory is the reflection coefficient (σ) that describes how a semipermeable barrier excludes or “reflects” solute X as water moves across the barrier, driven by hydrostatic or osmotic pressure gradients. (See Note: Reflection Coefficient Lest we forget… she is still here!)

The value of σ can range from 0 to 1. When σ is zero, the moving water perfectly “entrains” the solute, which moves with the water and exerts no osmotic pressure across the barrier. When σ is 1, the barrier completely excludes the solute as the water passes through, and the solute exerts its full or ideal osmotic pressure. To the extent that σ exceeds zero, the membrane sieves out the solute. The σ for plasma proteins is nearly 1.

Because small solutes such as Na+ and Cl⁻ freely cross the endothelium, their σ is zero, and they are not included in the Starling equation for the capillary wall (Equation 20-8). Thus, changing the intravascular or interstitial concentrations of such “crystalloids” does not create a net effective osmotic driving force across the capillary wall. (Conversely, because plasma membranes have an effective σ_NaCl = 1, NaCl gradients do shift water between the intracellular and interstitial compartments.)

The net driving force in the Starling equation (Equation 20-8), [(P_c − P_if) − σ(π_c − π_if)], has a special name, the net filtration pressure. Filtration of fluid from the capillary into the tissue space occurs when the net filtration pressure is positive. In the special case when σ for proteins is 1, the fluid leaving the capillary is protein free; this process is called ultrafiltration. Conversely, absorption of fluid from the tissue space into the vascular space occurs when the net filtration pressure is negative. At the arterial end of the capillary, the net filtration pressure is generally positive, so that filtration occurs. At the venous end, the net filtration pressure is generally negative, so that absorption occurs. However, as is discussed later, some organs do not adhere to this general rule.

In the next four sections, we examine each of the four Starling forces that constitute the net filtration pressure: P_c, P_if, π_c, and π_if.

Capillary blood pressure (P_c) falls from approximately 35 mm Hg at the arteriolar end to approximately 15 mm Hg at the venular end

Capillary blood pressure is also loosely called the capillary hydrostatic pressure, to distinguish it from capillary colloid osmotic pressure. It is only possible to record P_c in an exposed organ, ideally in a thin tissue (e.g., a mesentery) that allows good transillumination. One impales the lumen of the capillary with a fine micropipette (tip diameter < 5 μm) filled with saline and heparin. The micropipette lumen connects to a manometer, which has a sidearm to a syringe. Immediately after the impalement, blood begins to rise slowly up the pipette. A pressure reading at this time would underestimate the actual P_cbecause pipette pressure is less than P_c. The syringe makes it possible to apply just enough pressure to the pipette lumen to reach true pressure equilibrium—when fluid flows neither from nor to the pipette. By use of this null-point approach, the recorded pressure is the true P_c. In the human skin, P_c is approximately 35 mm Hg at the arteriolar end and approximately 15 mm Hg at the venular end.

When the arteriolar pressure is 60 mm Hg and the venous pressure is 15 mm Hg, the midcapillary pressure is not the mean value of 37.5 mm Hg but only 25 mm Hg (Table 20-5, top row). The explanation for the difference is that normally the precapillary upstream resistance exceeds the postcapillary downstream resistance (R_post/R_pre is typically 0.3; see Chapter 19). However, the midcapillary pressure is not a constant and uniform value. In Chapter 19, we saw that P_c varies with changes in R_pre and R_post (see Fig. 19-4). P_c also varies with changes in four other parameters: (1) upstream and downstream pressure, (2) location, (3) time, and (4) gravity.

Table 20-5 Effect of Upstream and Downstream Pressure Changes on Capillary Pressure*

Arteriolar (P_a) and Venular (P_v) Pressure Because R_post is less than R_pre, P_c follows P_v more closely than P_a (see Chapter 19). Thus, increasing P_a by 10 mm Hg—at a constant R_post/R_pre of 0.3—causes P_c to rise by only 2 mm Hg (Table 20-5, middle row). On the other hand, increasing P_v by 10 mm Hg causes P_c to rise by 8 mm Hg (Table 20-5, bottom row).

Location Capillary pressure differs markedly among tissues. For example, the high P_c of glomerular capillaries in the kidney, approximately 50 mm Hg (see Chapter 34), is required for ultrafiltration. The retinal capillaries in the eye must also have a high P_c because they bathe in a vitreous humor that is under a pressure of approximately 20 mm Hg (see Chapter 15). A higher P_c is needed to keep the capillaries patent in the face of the external compressing force. The pulmonary capillaries have unusually low P_c values, 5 to 15 mm Hg, minimizing the ultrafiltration that otherwise would lead to the accumulation of edema fluid in the alveolar air spaces (see Chapter 31).

Time Capillary blood pressure varies considerably from moment to moment at any given site, depending on the arteriolar diameter and tone of the precapillary sphincter (i.e., R_pre). In individual capillaries, these fluctuations lead to times of net filtration and other times of net fluid absorption.

Gravity Finally, the effect of gravity on P_c is the same as that discussed for arterial and venous pressure. Thus, a capillary bed below the level of the heart has a higher P_c than a capillary bed at the level of the heart.

Interstitial fluid pressure (P_if) is slightly negative, except in encapsulated organs

The interstitium consists of both a solid and a liquid phase. The solid phase is made up of collagen fibers and proteoglycans. In the liquid phase, only a small fraction of interstitial water is totally “free” and capable of moving under the influence of convective forces. Most of the water is trapped in gels (e.g., proteoglycans), in which both water and small solutes move by diffusion. It was once thought that P_if in the liquid phase is slightly above barometric pressure throughout the interstitium, but more recent measurements indicate that P_if is subatmospheric in many tissues.

Estimation of P_if is very difficult because the probe used to make the measurement is far larger than the interstitial space; thus, the measurement itself can alter P_if. If one inserts a probe percutaneously and immediately uses a null-point method to measure P_if (as outlined earlier for capillary pressure), the values are +1 to +2 mm Hg. However, during the next 4 to 5 hours, the measured value drops to −1 to −2 mm Hg. Arthur Guyton implanted a perforated, hollow plastic sphere under the skin to provide a chronic record of P_if (Fig. 20-7). After the wound has healed, the pressure inside the sphere may be as low as −2 to −10 mm Hg after 1 or 2 weeks. Another approach, the wick-in-needle technique, also yields subatmospheric values. (See Note: Null-Point Technique for Measuring Interstitial Pressure)

Figure 20-7 Long-term measurement of interstitial fluid pressure with an implanted capsule.

A value of −2 mm Hg is a reasonable average in loose tissues, such as the lung and subcutaneous tissue. P_if is slightly negative because of fluid removal by the lymphatics (see later). Inside rigid enclosed compartments, such as the bone marrow or brain, P_if is positive. It is also positive in encapsulated organs such as the kidney, where P_if is +1 to +3 mm Hg within the parenchyma. Expansion of high-pressure vessels in the kidney pushes the interstitial fluid against an unyielding fibrous capsule, raising P_if. The same principle applies to skeletal muscle, which is surrounded by layers of fascia. In some cases, it is not the interstitial fluid but another specialized compartment that provides the pressure around the capillaries. For renal glomerular capillaries, it is Bowman’s space (see Chapter 34)—filled with glomerular filtrate to a pressure of about +10 mm Hg—that is the relevant outside compartment. For pulmonary capillaries, the relevant outside compartment is the alveolus, the pressure of which varies during the respiratory cycle (see Chapter 27).

P_if is also sensitive to the addition of fluid to the interstitial compartment. When small amounts of fluid are added to the interstitial compartment, the interstitium behaves like a low-compliance system, so that P_if rises steeply for the small amount of added fluid. Adding more fluid disrupts the solid phase of collagen fibers and the gel of proteoglycans, so that large volumes can now accumulate with only small additional pressure increases. In this high-volume range, the interstitial compartment thus behaves like a high-compliance system. This high compliance is especially high in loose subcutaneous tissues, which can accommodate more edema fluid (see the box on interstitial edema) than can muscle.

Capillary colloid osmotic pressure (π_c), which reflects the presence of plasma proteins, is approximately 25 mm Hg

The colloid osmotic pressure difference across the capillary endothelium is due solely to the plasma proteins, such as albumin, globulins, and fibrinogen. Total plasma protein concentration is approximately 7.0 g/dL, which corresponds to approximately 1.5 mM of protein. According to van’t Hoff’s law (Equation 20-9), these proteins would exert an osmotic pressure of approximately 28 mm Hg if perfectly reflected by the capillary wall (σ = 1). The σ is indeed close to 1 for the principal plasma proteins—albumin (3.5 to 5.5 g/dL) and the globulins (2.0 to 3.5 g/dL)—so that the actual colloid osmotic pressure (σπ_c) in capillaries is approximately 25 mm Hg. This value is the same as if osmotically active solutes were present at approximately 1.3 mM. Note that because of the very definition of colloid osmotic pressure, we have ignored the osmotic effects of the small solutes in plasma, which have an osmolality of 290 mOsm (see Chapter 5). (See Note: Total Osmotic Pressure vs Colloid Osmotic Pressure of Plasma)

π_c does vary appreciably along the length of the capillary. Indeed, most capillary beds filter less than 1% of the fluid entering at the arteriolar end. Thus, the loss of protein-free fluid does not measurably concentrate plasma proteins along the capillary and does not appreciably raise π_c.

Because clinical laboratories report plasma protein concentrations in grams per deciliter and not all proteins have the same molecular weight, a plasma protein concentration of 7 g/dL can produce different π_cvalues, depending on the protein composition of the plasma. Because albumin has a much lower molecular weight than γ-globulin, replacement of 1 g of the heavier γ-globulin with 1 g of the lighter albumin raises π_c. Whereas van’t Hoff’s law (Equation 20-9) predicts a linear relationship between osmotic pressure and concentration, colloid osmotic pressure actually increases more steeply, even when the albumin/globulin ratio is held constant at 1.8 (Fig. 20-8, orange curve). Obviously, the steepness of the curve varies from one plasma protein to the next because all have different molecular weights. (See Note: Effects of Changes in Plasma H2O on Colloid Osmotic Pressure)

Figure 20-8 Dependence of colloid osmotic pressure on the concentration of plasma proteins. The point on the orange curve indicates that normal plasma, a mixture of proteins at a concentration of 7 g/dL, has a colloid osmotic pressure (π_c) of 25 mm Hg.

Not only does π_c vary markedly with protein composition and concentration, the reflection coefficient for colloids also varies widely among organs. The lowest values for σ (i.e., greatest leakiness) are in discontinuous capillary beds (e.g., liver); intermediate values are in muscle capillaries, and the highest values (σ = 1) are in the tight, continuous capillary beds of the brain.

The plasma proteins do more than just act as osmotic agents. Because these proteins also carry negative charges, the Donnan effect causes an increase in both the concentrations of cations (see Chapter 5) and the colloid osmotic pressure (see Fig. 5-15) in the capillary lumen. (See Note: Donnan Effects across the Capillary Wall)

Interstitial fluid colloid osmotic pressure (π_if) varies between 0 and 10 mm Hg among different organs

It is difficult to measure the interstitial fluid colloid osmotic pressure because it is virtually impossible to obtain uncontaminated samples. As a first approximation, we generally assume that π_if is the same as the colloid osmotic pressure of lymph. The protein content of lymph varies greatly from region to region; for example, it is 1 to 3 g/dL in the legs, 3 to 4 g/dL in the intestine, and 4 to 6 g/dL in the liver. Such lymph data predict that π_if ranges from 3 to 15 mm Hg. However, the protein concentration in the interstitial fluid is probably somewhat higher than in the lymph. A total body average value for π_if is approximately 3 mm Hg, substantially less than the value of 25 mm Hg for π_cin the capillary lumen.

The π_if appears to increase along the axis of the capillary (Table 20-6). The lowest values are near the arteriolar end, where the interstitium receives protein-free fluid from the capillary as the result of filtration. The highest values are near the venular end, where the interstitium loses protein-free fluid to the capillary as the result of absorption.

Table 20-6 Typical Values of Transcapillary Driving Forces for Fluid Movement in Loose, Non-encapsulated Tissue

The Starling principle predicts ultrafiltration at the arteriolar end and absorption at the venular end of most capillary beds

The idealized forces acting on fluid movement across a capillary are shown in Figure 20-9A. With use of the Starling equation and the values in Table 20-6, we can calculate the net transfer of fluid (J_V) at both the arteriolar and venular ends of a typical capillary:

Figure 20-9 Starling forces along a capillary. In A, the yellow lines are idealized profiles of capillary (P_c) and interstitial (P_if) hydrostatic pressures. Red lines are idealized capillary (π_c) and interstitial (π_if) colloid osmotic pressures. In B, the net filtration pressure is (P_c − P_if) − σ(π_c − π_if).

The net filtration pressure is thus positive (favoring filtration) at the arteriolar end, and it gradually makes the transition to negative (favoring absorption) at the venular end (Fig. 20-9B). At the point where the filtration and reabsorptive forces balance each other, an equilibrium exists, and no net movement of water occurs across the capillary wall.

Interstitial Edema

Edema (from the Greek oidema, for “swelling”) is characterized by an excess of salt and water in the extracellular space, particularly in the interstitium. Edema may be associated with any disease leading to salt retention and expansion of the extracellular fluid volume, in particular, renal, cardiac, and hepatic disease (see Chapter 40). However, interstitial edema can also occur without overall salt and water retention because of microcirculatory alterations that affect the Starling forces. Regardless of the cause, the resulting edema can be either generalized (e.g., widespread swelling of subcutaneous tissue, often first evident in facial puffiness) or localized (e.g., limited to the dependent parts of the body). In this box, we focus on how edema can result from changes in terms that make up the Starling equation.

Hydrostatic Forces

When a person is standing for a sustained time, venous pressure and thus capillary pressure (P_c) in the legs increase because of gravity. The result is movement of fluid into the tissue space. In most cases, the lymphatic system can take up the extra interstitial fluid and return it to the vascular space, maintaining proper fluid balance. The return of fluid requires contractions of the leg muscles to compress the veins and lymphatics and to propel the fluid upward through the valves in these vessels and toward the heart. If the standing person does not contract these muscles, the transudation of fluid can exceed the lymphatic return, causing interstitial edema.

An organ that is particularly sensitive to proper fluid balance is the lung. Slight increases in the hydrostatic pressure of the pulmonary capillaries (pulmonary hypertension) can lead to pulmonary edema. This condition decreases lung compliance (making lung inflation more difficult; see Chapter 27) and also may severely compromise gas exchange across the pulmonary capillary bed (see Chapter 30). Left-sided heart failure causes blood to back up into the vessels of the lung, raising pulmonary vascular pressures and causing pulmonary edema.

In right-sided heart failure, blood backs up into the systemic veins. As a result, central venous pressure (i.e., the pressure inside the large systemic veins leading to the right side of the heart) rises, causing an increase in the P_c in the lower extremities and abdominal viscera. Fluid transudated from the hepatic and intestinal capillaries may leave the interstitial space and enter the peritoneal cavity, a condition called ascites. (See Note: Transcapillary Refill)

Colloid Osmotic Forces

In nephrotic syndrome, a manifestation of a number of renal diseases, protein is lost in the urine. The result is a fall in plasma colloid osmotic pressure, a decrease in the ability of the capillaries to retain fluid, and generalized peripheral edema.

In pregnancy, synthesis of plasma proteins by the mother does not keep pace with the expanding plasma volume and nutritional demands of the fetus. As a result, maternal plasma protein levels fall. The same occurs in protein malnutrition. Although it is less severe than in nephrotic syndrome, the lower capillary colloid osmotic pressure nevertheless leads to edema in the extremities.

The opposite effect is seen in dehydration. A deficit of salt and water causes an increase in the plasma protein concentration, increasing the capillary colloid osmotic pressure and thus pulling fluid out of the interstitial space. The result is reduced turgor of the interstitial space. This effect is easily noticed by pinching the skin, which is unable to spring back to its usual firm position.

Properties of the Capillary Wall

Inflammation causes the release of vasodilators, such as histamine and cytokines, into the surrounding tissue. Vasodilation increases the number of open capillaries and therefore the functional surface area (S_f). Cytokines also cause widening of interendothelial clefts and a fall in the reflection coefficient (σ) for proteins. The net effect is enhanced filtration of fluid from capillary lumen to interstitium, so that tissue swelling is one of the hallmarks of inflammation.

Severe head injuries can result in cerebral edema, a result of the breakdown of the normally tight endothelial barrier of the cerebral vessels (see Chapter 11). Because the rigid skull prevents expansion of the brain, cerebral edema can lead to occlusion of the cerebral microcirculation.

During ischemia—when blood flow to a tissue is severely reduced or completely stopped—blood vessels deteriorate, causing hydraulic conductivity (L_p) to increase and reflection coefficient to decrease. Once blood flow is reestablished (reperfusion), these changes lead to local edema. If the increased leakiness is substantial, large quantities of plasma proteins freely move into the interstitial space, dissipating the colloid osmotic gradient across the capillary wall and aggravating the edema.

Lymphatic Drainage

Lymphatic drainage may become impaired after removal of lymph nodes for cancer surgery or when lymph nodes are obstructed by malignant neoplasms. The reduction in the lymphatic drainage leads to local edema, upstream from the affected nodes.

Net filtration pressure varies—sometimes considerably—among tissues. For example, in the intestinal mucosa, P_c is so much lower than π_c that absorption occurs continually along the entire length of the capillary. On the other hand, in glomerular capillaries, P_c exceeds π_c throughout most of the network, so that filtration may occur along the entire capillary (see Chapter 34). Hydraulic conductivity also can affect the filtration/absorption profile along the capillary. Because the interendothelial clefts become larger toward the venular end of the capillary, L_p increases along the capillary, from the arteriolar to the venular end.

Ignoring glomerular filtration in the kidney, Landis and Pappenheimer calculated a filtration of ~20 L/day at the arteriolar end of the capillary and an absorption of about 16 to 18 L/day at the venular end, for a net filtration of about 2 to 4 L/day from blood to interstitial fluid. This 2 to 4 L of net filtration does not occur uniformly in all capillary beds. The flow of fluid across a group of capillaries (F) is the product of the flux (J_V) and the functional surface area (S_f): F = J_v · S_f. Thus, net filtration of fluid in an organ depends not only on the net filtration pressure and the hydraulic conductivity of the capillary wall (terms that contribute to J_V) but also on the surface area of capillaries that happen to be perfused. For example, exercise recruits additional open capillaries in muscle, raising S_f and thereby increasing filtration.

For continuous capillaries, the endothelial barrier for fluid exchange is more complex than considered by Starling

The contribution of Landis and Pappenheimer was to insert experimentally measured values into the Starling equation (Equation 20-8) and to calculate the total body filtration and absorption rates and, by difference, net filtration. Because the estimate of 2 to 4 L for the net filtration rate agreed so well with total lymph flow, the scientific public accepted the entire Landis-Pappenheimer analysis. However, for continuous capillaries, the Landis-Pappenheimer estimates of filtration and absorption are far higher than the modern experimental data. Three major reasons for the discrepancy have emerged. First, Starling’s assumptions about the nature of the capillary barrier were overly simplistic. Namely, he assumed that a single barrier separated two well-defined, uniform compartments. Thus, according to Equation 20-8, the dependence of J_V on net filtration pressure ought to be linear, as indicated by the plot in the inset of Figure 20-10A. Second, Landis and Pappenheimer used (1) P_c values that are valid only at the level of the heart (i.e., ignoring gravity), (2) P_c values that are not subject to the vagaries of vasomotion, and (3) unrealistically low values of π_if (which would predict a greater absorption). (See Note: Assumptions of Landis and Pappenheimer)

Figure 20-10 Models of fluid exchange across continuous endothelia with interendothelial junctions. P_c, capillary hydrostatic pressure; π_c, capillary colloid osmotic pressures; P_sg, subglycocalyx hydrostatic pressure; π_sg, subglycocalyx colloid osmotic pressures; P_if, interstitial fluid hydrostatic pressure; π_if, interstitial fluid colloid osmotic pressures.

A revised model has emerged for fluid exchange across continuous endothelia with interendothelial junctions to account for discrepancies between the classical Starling predictions and the modern data. The revised model has two major features. First, the primary barrier for colloid osmotic pressure—that is, the semipermeable “membrane” that reflects proteins but lets water and small solutes pass—is not the entire capillary but only the luminal glycocalyx, in particular the glycocalyx overlying the paracellular clefts (Fig. 20-10B). Second, the abluminal surface of the glycocalyx is not in direct contact with the bulk interstitial fluid but is bathed by the subglycocalyx fluid at the top of the long paracellular cleft—a third compartment. Thus, the flow across the glycocalyx barrier depends not on P_if and π_if in the bulk interstitial fluid but on the comparable parameters in the subglycocalyx fluid (P_sg and π_sg):

Let us now examine the predictions of this equation for three states.

1. During ultrafiltration (i.e., J_V is positive). Here, the hydrostatic pressure in the subglycocalyx fluid—that is, the fluid in direct contact with the abluminal surface of the glycocalyx—is higher than that in the bulk interstitial fluid (i.e., P_sg > P_if in Fig. 20-10B). Thus, fluid moves from the subglycocalyx space, along the paracellular cleft, to the bulk interstitial fluid. Moreover, as long as protein-free ultrafiltrate enters the subglycocalyx space, the colloid osmotic pressure in the subglycocalyx fluid is low (π_sg < π_if). Both the rise in P_sg and the fall in π_sg tend to oppose filtration.

Because proteins enter the interstitium through the large-pore pathway, π_if in the bulk interstitial compartment is about that of lymph. However, at high rates of ultrafiltration, this π_if has no osmotic effect on the glycocalyx barrier because the protein cannot diffuse against the convective flow of fluid from lumen to interstitium. On the other hand, if the ultrafiltration rate is low, interstitial proteins can diffuse from the bulk interstitial space into the paracellular cleft, raising π_sg and promoting more ultrafiltration.

2. Net flow falls to nearly zero (i.e., J_V is ~0). Here, the parameters in the subglycocalyx fluid (i.e., P_sg and π_sg) should thus be very close to their values in the bulk interstitial fluid (i.e., P_if and π_if), and the revised model simplifies to the classical Starling model (Fig. 20-10A).

3. During absorption (i.e., reversal of flow, where J_V is negative). Here, water and small solutes move from the subglycocalyx space to the capillary lumen, leaving behind and thereby concentrating the protein in the subglycocalyx space (Fig. 20-10C). The resulting rise of π_sg (Equation 20-11) opposes further absorption and, indeed, can quickly bring it to a halt. This effect explains why the plot is nearly flat in the left lower quadrant of the inset between parts B and C in Figure 20-10.

Thus, a more sophisticated understanding of the structure of the endothelial barrier for proteins correctly makes two predictions. First, the fluxes are smaller than predicted by Starling for bulk driving forces because the actual driving force across the glycocalyx barrier (Equation 20-11) is smaller than the net driving force in the Starling equation (Equation 20-8). Second, the magnitude of the flux for a given net driving force is greater for ultrafiltration than for absorption—osmotic asymmetry or rectification.

LYMPHATICS

Lymphatics return excess interstitial fluid to the blood

Lymphatics arise in the interstitium as small thin-walled channels of endothelial cells that then join together to form increasingly larger vessels (Fig. 20-11). The initial lymphatics (previously called terminal lymphatics) are similar to capillaries but with many interendothelial junctions that behave like one-way microvalves, also called primary lymph valves. Anchoring filaments tether the initial lymphatics to surrounding connective tissue. The walls of the larger collecting lymphatics are similar to those of small veins, consisting of endothelium and sparse smooth muscle. The large lymphatic vessels, like the veins, have secondary lymph valves that restrict retrograde movement of lymph. Lymph nodes are located along the path of the collecting lymphatics. The large lymphatics ultimately drain into the left and right subclavian veins.

Figure 20-11 Flow of lymph into initial and collecting lymphatics.

At the level of the initial lymphatics, interendothelial junctions have few tight junctions or adhesion molecules connecting neighboring endothelial cells. As a result, flaps of endothelial cells can overlap with each other and act as the microvalve discussed before. Although initial lymphatics may appear collapsed and show no contractile activity, a pressure gradient from the interstitial fluid to the lymphatic lumen deforms the endothelial cells so that the microvalves open and fluid enters the initial lymphatic during the expansion phase (Fig. 20-11A). During this time, the secondary lymph valves are closed.

External pressure (e.g., from skeletal muscle) shuts the microvalves and causes fluid to enter larger lymphatics through the now open secondary lymph valves (Fig. 20-11B).

Most organs contain both initial and collecting lymphatics, but skeletal muscle and intestine have only initial lymphatics within their tissue. Lymphatics are absent from brain. They are most prevalent in the skin and the genitourinary, respiratory, and gastrointestinal tracts.

As we have already seen, filtration at the arteriolar end of capillaries is estimated to exceed absorption at the venular end by 2 to 4 L/day. However, fluid does not normally accumulate in the interstitium because this excess fluid and protein move into the lymphatics. Thus, each day, the lymphatics return to the circulation 2 to 4 L of interstitial fluid, maintaining a steady state. In a model of congenital lymphedema, mice with genetic absence of initial lymphatics have elevated P_if and π_if as well as interstitial volume expansion (i.e., edema), emphasizing the role of the lymphatics in returning fluid and protein from the interstitial space to the blood.

Flow in Initial Lymphatics Hydrostatic pressure in the initial lymphatics (P_lymph) ranges from −1 mm Hg to +1 mm Hg. Inasmuch as the mean interstitial fluid pressure is somewhat more negative than these values, what provides the driving force for interstitial fluid to move into the terminal lymphatics? Transient increases in P_if temporarily raise P_if above P_lymph. Indeed, increases in mean P_if cause an increase in lymph flow (Fig. 20-12).

Figure 20-12 Dependence of lymph flow on interstitial pressure.

Because the interstitium exhibits a variable compliance, fluid added to the interstitium in its low-compliance range raises the P_if substantially, providing the driving force for fluid to enter the lymphatics. In this same range of P_ifvalues, lymphatic flow is especially sensitive to increases in P_if (steep portion of Fig. 20-12). Thus, lymphatic efflux nicely matches the excess capillary filtration, so that the interstitial fluid volume changes very little. The situation is very different if the interstitium is already expanded and in its high-compliance range. In this case, fluid added to the interstitium raises the already elevated P_ifonly moderately (e.g., from +2 to +4 mm Hg). In this range of P_if values, lymphatic uptake is not very responsive to increases in P_if (flat portion of Fig. 20-12). Thus, in this case, lymphatic return does not compensate well for the excess capillary filtration, so that interstitial fluid volume increases further (i.e., edema begets more edema).

Intermittent compression and relaxation of lymphatics occur during respiration, walking, and intestinal peristalsis. When P_lymph in a downstream segment falls below that in an upstream segment, fluid aspiration produces unidirectional flow. This suction may be largely responsible for the subatmospheric values of the P_if observed in many tissues.

Flow in Collecting Lymphatics Pressures in the collecting lymphatics range from +1 to +10 mm Hg, and they increase progressively with each valve along the vessel. As P_lymph rises in the collecting lymphatic vessels, smooth muscle in the lymphatic walls actively contracts by an intrinsic myogenic mechanism that also plays a role in blood vessels, as discussed later. Thus, downstream occlusion of a lymphatic vessel increases P_lymph and hence the frequency of smooth muscle contractions, whereas an upstream occlusion does the opposite. Because of the presence of one-way valves, smooth muscle contraction drives lymph toward the veins. The rhythmic contraction and relaxation of VSMCs that we will discuss for blood vessels—vasomotion—also occurs in lymphatics and is essential for the propulsion of lymph.

In addition to vasomotion, passive processes also propel lymph toward the blood. As is the case for the initial lymphatics, skeletal muscle contraction, respiratory movements, and intestinal contractions all passively compress the collecting lymphatics. This intermittent pumping action moves lymph into the veins.

Transport of Proteins and Cells Proteins that entered interstitial fluid from the capillary cannot return to the circulation because of the adverse chemical gradient across the capillary endothelial wall. The buildup of these macromolecules in the interstitium creates a diffusional gradient from the interstitium to the lymph that complements the convective movement of these macromolecules (along with fluid) into the lymphatic system. In an average person, the lymphatics return 100 to 200 g of proteins to the circulation per day. Even before lymph reaches lymph nodes, it contains leukocytes—which had moved from the blood into the interstitium—but no red blood cells or platelets. Cycles of lymphatic compression and relaxation not only enhance fluid movement but also greatly increase the leukocyte count of lymph.

The circulation of extracellular fluids involves three convective loops: blood, interstitial fluid, and lymph

Extracellular fluid moves in three convective loops (Fig. 20-13). The first is the cardiovascular loop. Assuming a cardiac output of 5 L/min, the convective flow of blood through the circulation at rest is 7200 L/day. The second is the transvascular loop, in which fluid moves out of the capillaries at their arteriolar end and into the capillaries at their venular end. Not counting the kidney, whose glomeruli filter a vast amount of fluid (see Chapter 34), Landis and Pappenheimer estimated that all the other tissues of the body filter ~20 L/day at the arteriolar end of their capillaries and reabsorb 16 to 18 L at the venular end. As noted earlier, both the filtration and absorption values are probably vast overestimates. Nevertheless, the difference between filtration and absorption, 2 to 4 L/day, is a reasonable estimate of the third fluid loop, the lymphatic loop.

Figure 20-13 Convective loops of extracellular fluid and protein.

In addition to convective exchange, a diffusional exchange of water and solutes also occurs across the capillaries. The diffusional exchange of water occurs at a much higher rate than does convective movement. Using deuterium oxide as a marker, investigators have found that this diffusional exchange is approximately 80,000 L/day across all of the body’s systemic capillaries. This value is about an order of magnitude greater than blood flow in the cardiovascular loop and three orders of magnitude larger than the convective flow in the transvascular filtration/absorption loop of the microcirculation. However, the diffusion of water molecules is an exchange process that does not contribute appreciably to the net movement of water. In other words, every day, 80,000 L of water diffuse out of the capillaries and 80,000 L diffuse back.

With regard to small solutes that can diffuse across the capillary endothelium, the traffic is quite different from the convective loops for water. Take glucose as an example. The plasma contains approximately 100 mg/dL glucose, red blood cells have little glucose, and the cardiac output of plasma is approximately 2.75 L/min (assuming a hematocrit of 45%). Therefore, each day, the heart pumps approximately 4000 g of glucose. This glucose can enter the interstitium by two mechanisms. First, glucose is dissolved in the water filtered from the arteriolar end of the capillaries. Each day, this filtration process carries 20 L × 100 mg/dL = 20 g of glucose into the interstitium. Second, each day, approximately 20,000 g of glucose enters the interstitium by diffusion. Convection can supply only a small fraction of the approximately 400 g of glucose that the body consumes each day. Instead, diffusion supplies the majority of the glucose. Nevertheless, the 400 g/day of metabolized glucose is a minuscule fraction of the amount that enters the interstitium by diffusion. Thus, most of the glucose that diffuses into the interstitium diffuses back out again.

Protein traffic provides yet another pattern of circulatory loops. Plasma contains 7 g/dL of proteins, and—assuming a plasma volume of 3 L in a 70-kg human—total plasma protein content is ~210 g. Given a cardiac plasmaoutput of 2.75 L/min, the heart pumps ~277,000 g of protein through the circulation every day. Of this protein, 100 to 200 g—nearly the entire plasma content of proteins—moves daily across the capillary walls through the large-pore system by a transcellular route and to a lesser extent by solvent drag. Because only very small amounts of filtered protein return to the circulation by solvent drag at the venular end of capillaries (~5 g/day), nearly all of the filtered protein (95 to 195 g/day) depends on the convective lymphatic loop for its ultimate recovery.

REGULATION OF THE MICROCIRCULATION

The active contraction of vascular smooth muscle regulates precapillary resistance, which controls capillary blood flow

Smooth muscle tone in arterioles, metarterioles, and precapillary sphincters (see Chapter 19) determines the access resistance to the capillary beds. This resistance upstream of the capillary bed is also known as the afferent or precapillary resistance (R_pre). The overall resistance of a microcirculatory bed is the sum of R_pre, the resistance of the capillary bed itself (R_cap), and the efferent or postcapillary resistance (R_post).

How do these resistances influence the flow of blood (F_cap) through a capillary bed? We can answer this question by rearranging the Ohm’s law–like expression that we introduced as Equation 17-1:

P_a is the pressure just before the beginning of the precapillary resistance, and P_v is the pressure just after the end of the postcapillary resistance. Because the aggregate R_cap is small, and R_post/R_pre is usually approximately 0.3, R_pre is usually much greater than R_cap + R_post. Because R_pre is the principal determinant of total resistance, capillary flow is roughly inversely proportional to R_pre.

Modulating the contractility of VSMCs in precapillary vessels is the main mechanism for adjusting perfusion of a particular tissue. VSMCs rely on a different molecular mechanism of contraction than skeletal muscle does, although an increase in [Ca²⁺]_i is the principal trigger of contraction in both cases. Whereas an increase in [Ca²⁺]_i in skeletal muscle elicits contraction by interacting with troponin C, an increase in [Ca²⁺]_i in VSMCs elicits contraction by activating calmodulin (see Chapter 3). The Ca²⁺-calmodulin complex (Ca²⁺-CaM) activates myosin light chain kinase (MLCK), which in turn phosphorylates the regulatory myosin light chain (MLC) on each myosin head (see Chapter 9). Phosphorylation of MLC allows the myosin to interact with actin, producing contraction. Relaxation occurs when myosin light chain phosphatase dephosphorylates the MLC. In addition to changes in [Ca²⁺]_i, changes in the activity of MLCK itself can modulate the contraction of VSMCs. Phosphorylation of MLCK by cAMP-dependent protein kinase (PKA) or cGMP-dependent protein kinase (PKG) inactivates the enzyme and thus preventscontraction.

Smooth muscle cells can function as a syncytium when they are coupled through gap junctions (unitary smooth muscle), or they can function independently of one another, as do skeletal muscle fibers (multiunit smooth muscle). Most vascular smooth muscle has a multiunit organization. In contrast to skeletal muscle, VSMCs receive multiple excitatory as well as inhibitory inputs. Moreover, these inputs come not only from chemical synapses (i.e., neural control) but also from circulating chemicals (i.e., humoral control). The actual contraction of VSMCs may follow smooth muscle electrical activity in the form of action potentials, slow waves of depolarization, or graded depolarizations without spikes. VSMCs can show spontaneous rhythmic variations in tension leading to periodic changes in vascular resistance and microcirculatory flow in a process called vasomotion. These spontaneous, rhythmic, smooth muscle contractions result either from pacemaker currents or from slow waves of depolarization and associated [Ca²⁺]_i increases in the VSMCs. Humoral agents can also directly trigger contraction of VSMCs through increases in [Ca²⁺]_i, without measurable fluctuations in membrane potential (pharmacomechanical coupling; see Chapter 9).

Table 20-7 summarizes the roles that various membrane proteins (channels, transporters, and receptors) play in controlling the tone of VSMCs. Together with their associated signal transduction pathways, these membrane proteins lead to either contraction (i.e., vasoconstriction) or relaxation (i.e., vasodilation). Although a variety of neurotransmitters and circulating hormones act on smooth muscle through different receptors and transduction pathways, their effects converge on regulating the activity of MLCK.

Table 20-7 Molecular Mechanisms Underlying the Contraction and Relaxation of Vascular Smooth Muscle

Tissue metabolites regulate local blood flow in specific vascular beds, independently of the systemic regulation

VSMCs not only control the resistance of arterioles (i.e., R_pre) and thus local blood flow, they also control the resistance of small terminal arteries and thereby play an important role in regulating systemic arterial blood pressure. In Chapter 23, we discuss this control of arterial blood pressure (a whole-body function) through VSMCs of small arteries and arterioles, both of which are under the control of centralmechanisms—the autonomic nervous system and systemic humoral agents (e.g., angiotensin II). However, the subject of this discussion is local regulatory mechanisms that use the arterioles to regulate blood flow through specific vascular beds. These local control mechanisms can override any of the neural or systemic humoral influences.

Mechanisms of local control involve (1) myogenic activity and (2) local chemical and humoral factors. Myogenic regulation refers to an intrinsic mode of control of activity, in which stretch of the VSMC membrane activates stretch-sensitive channels (Table 20-7). The result is a depolarization that affects pacemaker activity, thereby eliciting contraction of the VSMC.

The most prominent chemical factors are interstitial P_O2, P_CO2, and pH as well as local concentrations of K+, lactic acid, adenosine triphosphate (ATP), adenosine diphosphate (ADP), and adenosine (Table 20-8). Total osmolality may also make a contribution. The local regulation of VSMCs by interstitial P_O2, P_CO2, and pH is distinct from the regulation of systemic blood pressure by the peripheral chemoreceptors, which respond to changes in arterial P_O2, P_CO2, and pH (see Chapter 32) and initiate a complex neural reflex that modulates VSMC activity (see Chapter 23). In the case of local control, chemical changes in interstitial fluid act directly on the VSMCs through one of the transduction mechanisms listed in Table 20-7. Changes that typically accompany increased metabolism (e.g., low P_O2, high P_CO2, and low pH) vasodilate vessels in the systemic circulation. Such local changes in P_O2, P_CO2, and pH have opposite effects in the pulmonary circulation (see Chapter 31).

Table 20-8 Local Metabolic Changes That Cause Vasodilation in the Systemic Circulation (See Note: Vasodilation Caused by Increases in [K⁺]_o)

Change	Mechanism*
↓Po2	↓[ATP]i, ↑adenosine release, ↑PGI2 release, ↑NO release
↑PCO2	↓pHo
↓pH	↓pHo
↑[K+]0	Transient hyperpolarization → closes voltage-gated Ca2+ channels
↑[Lactic acid]0	Probably ↓ pHo
↓ [ATP] i	Opens KAT P channels
↑[ATP]0	Activates purinergic receptors P2
↑[ADP]0	Activates purinergic receptors P2
↑[Adenosine]0	Activates purinergic receptors P1

* The subscript i refers to intracellular levels, and the subscript o refers to interstitial levels.

Because blood flow itself can wash out the metabolic intermediates, vasomotion can arise from a local feedback system. For example, if interstitial P_O2 falls as a result of increased local O₂ consumption, the ensuing vasodilation will increase O₂ delivery to the metabolizing cells and in turn will tend to cause the local interstitial P_O2 to increase. As the P_O2 now increases, vascular tone will increase. The timing of release and washout of the chemical factors determines the frequency of the vasomotion. The interstitial fluid volume around the active cells—the volume in which vasoactive metabolites are distributed—also affects this periodicity because it affects the time lag for the concentration of vasoactive substances to rise or to fall. Finally, spontaneous fluctuations in metabolism may confer an additional periodicity to vasomotion.

The endothelium of capillary beds is the source of several vasoactive compounds, including NO, EDHF, and endothelin

The capillary endothelium is the source of several important vasoactive compounds (Table 20-9).

Table 20-9 Vasoactive Agents Produced by Endothelial Cells

Vasodilators	Vasoconstrictors
Nitric oxide (NO)	Endothelin (ET)
Endothelium-derived hyperpolarizing factor (EDHF)	Endothelium-derived constricting factor 1 (EDCF1)
Prostacyclin (PGI2)	Endothelium-derived constricting factor 2 (EDCF2)

Nitric Oxide (NO) Originally called endothelium-derived relaxing factor, NO is a potent vasodilator. NO also inhibits platelet aggregation, induces platelet disaggregation, and inhibits platelet adhesion. Bradykinin and acetylcholine both stimulate the NOS III (or eNOS) isoform of NO synthase (see Chapter 3) that is constitutively present in endothelial cells. Increases in shear stress—the force acting on the endothelial cell along the axis of blood flow—can also stimulate the enzyme. NOS III, which depends on both Ca²⁺ and calmodulin for its activity, catalyzes the formation of NO from arginine. NO, a lipophilic gas with a short half-life, diffuses locally outside the endothelial cell. Inside the VSMC is the “receptor” for NO, a soluble guanylyl cyclase that converts guanosine triphosphate (GTP) to cyclic guanosine monophosphate (cGMP). cGMP-dependent protein kinase (i.e., PKG) then phosphorylates MLCK and SERCA. Phosphorylation inhibits the MLCK, thus leading to a net decrease in the phosphorylation of MLC and a decrease in the interaction between myosin and actin. Phosphorylation activates SERCA, thereby decreasing [Ca²⁺]_i. The net result is that NO released by endothelial cells relaxes VSMCs, producing vasodilation.

Physicians have used exogenous organic nitrates (e.g., nitroglycerin) for decades to dilate peripheral vessels for relief of the pain of angina pectoris. These powerful vasodilators exert their activity by breaking down chemically, thereby releasing NO near VSMCs.

Endothelium-Derived Hyperpolarizing Factor (EDHF) In addition to releasing NO, endothelial cells release another relaxing factor in response to acetylcholine, EDHF. EDHF causes VSMC relaxation by making the membrane potential more negative.

Prostacyclin (PGI₂) Prostacyclin synthase (see Fig. 3-11) metabolizes arachidonic acid to the vasodilator PGI₂. This agent acts by increasing [cAMP]_i and promoting the phosphorylation of MLCK, ultimately decreasing the phosphorylation of myosin light chains. PGI₂ is especially important for dilation of pulmonary vessels at birth (see Chapter 57).

Endothelins (ETs) Endothelial cells produce 21-residue peptides that cause an extremely potent and long-lasting vasoconstriction in most VSMCs. Many acute and chronic pathological conditions, including hypoxia, promote the release of endothelin, which exists as three isopeptides: ET-1, ET-2, and ET-3. The precursor of ET-1 is preproendothelin, which the endothelial cell converts first to proendothelin and then to the mature endothelin, which it releases. The ET receptor subtype for vasoconstriction is ET_A. Other endothelin receptors also exist; ET_B1 mediates vasodilation, ET_B2 mediates vasoconstriction, and ET_C has as yet no clearly defined function. ET_A receptors predominate in high-pressure parts of the circulation, whereas ET_B receptors predominate in low-pressure parts of the circulation.

The binding of an endothelin to any endothelin receptor subtype ultimately results in an increase in [Ca²⁺]_i. In the vasoconstriction response, ET-1 binding to ET_A receptors acts through the phospholipase C pathway to generate inositol trisphosphate, to release Ca²⁺ from intracellular stores, and to raise [Ca²⁺]_i (see Chapter 3). In a second, delayed phase, which is not well understood, Ca²⁺ entering from the outside contributes to the increase in [Ca²⁺]_i. The increased [Ca²⁺]_i activates Ca²⁺-CaM, stimulating MLCK to phosphorylate myosin light chains and culminating in contraction. (See Note: Delayed [Ca²⁺]_i Increase in Response to Endothelin)

Thromboxane A₂ (TXA₂) Endothelial cells and platelets metabolize arachidonic acid through the cyclooxygenase pathway to produce TXA₂ (see Chapter 3). This agent activates TXA₂/prostaglandin H₂ (TP) receptors, leading to opening of L-type Ca channels, thereby increasing [Ca²⁺]_i. In addition, TP activation increases the levels of superoxide anion radical O₂⁻ in VSMCs. In turn, O₂⁻ reacts with NO, thereby reducing the vasodilating effect of NO.

Other Endothelial Factors In some systemic arteries of the dog, anoxia produces an unexpected effect: an endothelium-dependent increase in tension mediated by a putative factor, EDCF₁ (endothelium-derived constricting factor). In some dog arteries, rapid stretch evokes a contraction that is also endothelium dependent. This putative factor, EDCF₂, could be a superoxide anion because superoxide dismutase prevents the contractions.

Autoregulation stabilizes blood flow despite large fluctuations in systemic arterial pressure

As we saw in Chapter 17, the pressure-flow relationship of an idealized, rigid vessel is linear (Fig. 20-14, gray line). In most real (i.e., elastic) vessels, however, increases in pressure cause a dilation that reduces resistance and leads to a steeper-than-linear flow (Fig. 20-14, red curve). However, some vascular beds behave very differently. Despite large changes in the systemic arterial pressure—and thus large changes in the driving pressure—these special vascular beds maintain local blood flow within a narrow range. This phenomenon is called autoregulation. These vascular beds behave more or less like rigid tubes at very low and at very high perfusion pressures (Fig. 20-14, blue curve). However, in the physiological pressure range over which autoregulation occurs, changes in perfusion pressure have little effect on flow. Instead, increases in pressure lead to increases in resistance that keep blood flow within a carefully controlled range.

Figure 20-14 Autoregulation of blood flow.

Autoregulatory behavior takes time to develop and is due to an active process. If the perfusion pressure were to increase abruptly, we would see that immediately after the pressure increase, the pressure-flow diagram would look much like the one for the rigid tube in Figure 20-14. However, the vascular arteriolar tone then slowly adjusts itself to produce the characteristic autoregulatory pressure-flow diagram. The contraction of VSMCs that underlies autoregulation is autonomous, that is, it is entirely local and independent of neural and endocrine mechanisms. Both myogenic and metabolic mechanisms play an important role in the adjustments of smooth muscle tone during autoregulation. For example, the stretch of VSMCs that accompanies the increased perfusion pressure triggers a myogenic contraction that reduces blood flow. Also, the increase in P_O2 (or decrease in P_CO2 or increase in pH) that accompanies increased perfusion pressure triggers a metabolic vasoconstriction that reduces blood flow (Table 20-8).

Autoregulation is useful for at least two reasons. First, with an increase in perfusion pressure, autoregulation avoids a waste of perfusion in organs in which the flow is already sufficient. Second, with a decrease in perfusion pressure, autoregulation maintains capillary flow and capillary pressure. Autoregulation is very important under these conditions for organs—particularly the heart, brain, and kidneys—that are very sensitive to ischemia or hypoxia and for organs (again, the kidney) whose job it is to filter the blood.

New blood vessels proliferate in response to growth factors by a process known as angiogenesis

In adults, the anatomy of the microcirculation remains rather constant. Notable exceptions are the growth of new vessels during wound healing, inflammation, and tumor growth and in the endometrium during the menstrual cycle. Increased capillary density is important in physical training and in acclimatization to altitude (see Chapters 60 and 61).

The development of new vessels is called angiogenesis. The first step is dissolution of the venular basement membrane at a specific site, followed by activation and proliferation of previously quiescent endothelial cells. The new cells, attracted by growth factors, migrate to form a tube. Eventually, the budding tubes connect with each other, allowing the flow of blood and the development of vascular smooth muscle as the new microvascular network establishes itself. Angiogenesis relies on a balance between positive and negative regulation. The body normally produces some factors that promote angiogenesis and others that inhibit it (Table 20-10).

Table 20-10 Agents That Affect Vascular Growth

Promoters	Inhibitors
Vascular endothelial growth factor (VEGF)	Endostatin
Fibroblast growth factors (FGFs)	Angiostatin
Angiopoietin 1 (ANGPT1)	Angiopoietin 2 (ANGPT2)

Promoters of Vessel Growth The principal peptides that induce angiogenesis are two polypeptides: vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF). Both interact with endothelium-specific receptor tyrosine kinases (see Chapter 3). VEGF—related to platelet-derived growth factor (PDGF) and a mitogen for vascular endothelial cells—is produced by fibroblasts and, frequently, by cancer cells. Activated coagulation factor VII (FVIIa; see Chapter 18) promotes VEGF production.

FGF mediates many cellular responses during embryonic, fetal, and postnatal development. At least 22 different fibroblast growth factors exist in humans. FGF-2 (also known as basic fibroblast growth factor or bFGF) has particular angiogenic activity.

VEGF and FGF-2 promote expression of NOS. The resulting NO promotes proliferation and migration of endothelial cells as well as differentiation of vascular tubes.

The targeted delivery of these growth factors is a major obstacle in their therapeutic use. One approach has been to link the growth factor to small beads delivered into the coronary circulation. Clinical trials with local or systemic administration of FGF-2 to patients with ischemic heart disease have shown mixed efficacy. A recombinant, humanized monoclonal antibody against VEGF (Avastin) is being used in patients with advanced non–small cell lung cancer.

Other growth factors have indirect angiogenic effects that are distinct from those of VEGF or FGF. Angiopoietins (ANGPT1 and ANGPT2) are proteins that act through a receptor tyrosine kinase (Tie2) expressed almost exclusively in endothelial cells. ANGPT1 is required for embryonic vascular development, and ANGPT2—normally an antagonist of ANGPT1 at the Tie2 receptor—is required for postnatalangiogenic remodeling. Angiogenin, a member of the ribonuclease family, is normally present in plasma, but at levels too low to produce proliferative effects. Plasma angiogenin levels rise in cancer patients. Regulated surface receptors on endothelial cells bind angiogenin, which after endocytosis translocates to the nucleus, where its RNAse activity is essential for its angiogenic effect. (See Note: Angiogenin)

Inhibitors of Vessel Growth The concept of antiangiogenesis was first advanced by Judah Folkman as a strategy to stop the growth of tumors. He and his colleagues have described two peptides, angiostatin and endostatin, that are inhibitors of angiogenesis.

Angiostatin is a kringle-containing fragment of plasminogen, a key fibrinolytic protein (see Chapter 18). Angiostatin arises by proteolytic cleavage of plasminogen by connective tissue enzymes, such as matrix metalloproteinases and elastase. Angiostatin inhibits angiogenesis by enhancing apoptosis of endothelial cells and inhibiting migration and tube formation, rather than by affecting proliferation. Recombinant angiostatin is being tested in patients with advanced lung cancer.

Endostatin is a peptide breakdown product of collagen XVIII. It is produced by the extracellular matrix of tumors.

We can illustrate the importance of angiogenesis by highlighting three clinical situations in which angiogenesis plays an important role. First, enhancement of vessel growth is important during coronary artery disease, when chronic ischemia of the heart leads to the development of new vessels and thus collateral circulation. Second, angiogenesis enhances the blood supply to a tumor, thereby promoting its growth and opening the principal route by which tumor cells exit the primary tumor during metastasis. Oncologists are exploring the use of angiogenesis inhibitors to treat cancer. Third, angiogenesis may also be important in diabetic retinopathy, where blood vessel proliferation can cause blindness.

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