Blue Histology - Vascular System

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		School of Anatomy and Human Biology - The University of Western Australia
		Blue Histology - Vascular System

Topics	Lab Guides and Images
Vascular System	Aorta - elastin & eosin and H&E
General Structure of Blood Vessels	Artery - elastin & eosin and H&E
Variations of Vessel Wall Structure Arteries Capillaries Veins	Capillaries - cardiac muscle, Whipf's polychrome
	Vein - elastin & eosin and H&E
	Vein Valve - H&E
Lymphatic Vessels	Lymph Capillary -Lacteal - jejunum, H&E

Additional Resources

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Large Images: Search the Large Images page with these keywords: vascular system, aorta, artery, vein, elastic artery, muscular artery, vein valve, tunica intima, tunica media, tunica adventitia, internal elastic lamina or external elastic lamina.
VScope: Magnification & Stage Simulation: aorta - elastin / H&E
Self Assessment: Choose subject area "vascular system" on the Quiz page

VASCULAR SYSTEM

The cardiovascular system is concerned with the transport of blood and lymph through the body. It may be divided into four major components: the heart, the macrocirculation, the microcirculation and the lymph vascular system.

Essentially, the macrocirculation comprises all vessels, both arteries and veins, that would be visible to the eye. The vessels of the macrocirculation supply and drain a network of fine vessels interposed between them, the capillaries. This network is also called the capillary bed. Water and other components of the blood plasma which exude from the blood vessels form the interstitial fluid, which is returned to the circulation by the lymph vascular system.

General Structure of Blood Vessels

You have already seen blood vessels of various sizes and types in preparations available in other lab sessions, and you should be aware that the histological appearances of vessels of different sizes (arterioles vs. arteries) and different types (arteries vs. veins) are different from each other. These differences are the result of quantitative variations of a common structural pattern that can be seen in all blood vessels with the exception of capillaries, i.e. the division of the walls of the blood vessels into three layers or tunics.

The tunica intima: delimits the vessel wall towards the lumen of the vessel and comprises its endothelial lining (typically simple, squamous) and associated connective tissue. Beneath the connective tissue, we find the internal elastic lamina, which delimits the tunica intima from
the tunica media.: The tunica media is formed by a layer of circumferential smooth muscle and variable amounts of connective tissue. A second layer of elastic fibers, the external elastic lamina, is located beneath the smooth muscle. It delimits the tunica media from
the tunica adventitia,: which consist mainly of connective tissue fibres. The tunica adventitia blends with the connective tissue surrounding the vessel. The definition of the outer limit of the tunica adventitia is therefore somewhat arbitrary.

more about .... Endothelial Cells
by Professor John McGeachie

Variations of Vessel Wall Structure

Arteries

All arterial vessels originate with either the pulmonary trunk (from the right ventricle) or the aorta (from the left ventricle). Specialisations of the walls of arteries relate mainly to two factors: the pressure pulses generated during contractions of the heart (systole) and the regulation of blood supply to the target tissues of the arteries. The tunica media is the main site of histological specialisations in the walls of arteries.

Vessels close to the heart (aorta, pulmonary trunk and the larger arteries that originate from them) are

Elastic arteries

The tunica intima of elastic arteries is thicker than in other arteries. A layer of loose connective tissue beneath the endothelium (subendothelial connective tissue) allows the tunica intima to move independently from other layers as the elastic arteries distend with the increase in systolic blood pressure. Distension of the walls is facilitated by concentric fenestrated lamellae of elastic fibres in a thick tunica media. In adult humans, about 50 elastic lamellae are found in the tunica media of the aorta. The energy stored in the elastic fibres of the tunica media allows elastic arteries to function as a "pressure reservoir" which forwards blood during ventricular relaxation (diastole). Smooth muscle cells and collagen fibres are present between the layers of elastic fibres. Both fibre types are produced by the smooth muscle cells. Each elastic lamella forms together with interlamellar fibres and cells a lamellar unit. The external elastic lamina is difficult to discern from other layers of elastic fibres in the tunica media. The tunica adventitia appears thinner than the tunica media and contains collagen fibres and the cell types typically present in connective tissue.

The walls of these large arteries are so thick that their peripheral parts cannot derive enough oxygen and nutrients from the blood of the vessel that they form. Larger vessels are therefore accompanied by smaller blood vessels which supply the tunica adventitia and, in the largest vessels, the outer part of the tunica media of the vessel wall. The vessels are called vasa vasorum. In macroscopic preparations vasa vasorum are visible as fine dark lines on the surface of the larger arteries.

Suitable Slides: sections of the aorta - H&E, elastin

Aorta, human - H&E , elastin & van Gieson
The thin endothelial lining of the aorta corresponds to that of other vessels. The flattened cells are easily damaged during preparation and it may be difficult to identify the endothelium. The subendothelial layer of connective tissue is characterised by a lower density of cells, i.e. fewer nuclei, a fibrous appearance of the tissue and the absence of well-defined elastic layers. Because the lamellae of elastic fibers diffract light differently from the remaining tissues they should also be visible in H&E stained sections. Elastic lamellae become visible in the tunica media. The majority of cells in the tunica media are smooth muscle cells. Smooth muscle cells and collagen fibres are found between the layers of elastic fibres. If you scan the periphery of the aorta you may find small blood vessels, the vasa vasorum, in the tunica adventitia and penetrating into the outer part of the tunica media.
Draw the aorta at low magnification and label the three tunics. Draw part of the tunica media at high magnification and identify collagen fibres, layers of elastic fibres and smooth muscle cell nuclei in your drawing.

The diameter of individual arteries decreases as we follow them further into the periphery. However, their total diameter increases, which leads to a fall in blood pressure. Also, the properties of the elastic arteries have to some extent evened out differences in diastolic and systolic blood pressure. The amount of elastic fibres in the tunica media decreases with these physiological changes. We now find a type of arteries which are termed

Muscular arteries: The tunica intima is thinner than in elastic arteries. Subendothelial connective tissue other than the internal elastic lamina is often difficult to discern. The internal elastic lamina forms a well defined layer. The tunica media is dominated by numerous concentric layers of smooth muscle cells. Fine elastic fibres and and a few collagen fibres are also present. The external elastic lamina can be clearly distinguished although it may be incomplete in places. The thickness and appearance of the tunica adventitia is variable.

The basic structure of the walls of arteries does not change much as we come to the next type of arterial vessels. Size is used to differentiate them from muscular arteries.

Arterioles: are arterial vessels with a diameter below 0.1 - 0.5 mm (different values in different textbooks). Endothelial cells are smaller than in larger arteries, and the nucleus and surrounding cytoplasm may 'bulge' slightly into the lumen of the arteriole. The endothelium still rests on a internal elastic lamina, which may be incomplete and which is not always well-defined in histological sections. The tunica media consists of 1-3 concentric layers of smooth muscle cells . It is difficult to identify an external elastic lamina or to distinguish the tunica adventitia from the connective tissue surrounding the vessel.
The smooth muscle of arterioles and, to some extent, the smooth muscle of small muscular arteries regulate the blood flow to their target tissues. Arterioles receive both sympathetic and parasympathetic innervation. The final branching of the arterioles finally gives rise to the capillary network (microcirculation).

Suitable Slides: sections of arteries - H&E or elastin (in combination with other stains)
Sections of small muscular arteries and arterioles are present in many sections, and the basic features of their structure are usually visible - even in smaller arteries. Large muscular arteries often have their "own section" in teaching collections.

Artery - H&E and elastin & eosin
Identifying muscular arteries in sections is rather straight forward. There are two easily recognizable features which distinguish these arteries from veins. If two vessels have a similarly sized lumen, the walls of arteries will be much thicker and more compact than the wall of veins. At high magnification, the internal elastic lamina forms a pink streak immediately below the endothelial cell lining in arteries and even arterioles, while it is difficult to identify in veins.
The layer of subendothelial connective tissue is very thin, and the endothelium seems to rest on the internal elastic lamina. Smooth muscle cell nuclei are frequent in the tunica media. The external elastic lamina stains similar to the internal elastic lamina, but it is thicker and appears fibrous instead of forming a continuous band. Collagen fibres and a few connective tissue cell nuclei are visible in the tunica adventitia.
If you close the iris diaphragm of the microscope, the elastic layers will stand out very clearly, but remember to open the diaphragm once you have seen them.

In addition to the inner and outer elastic laminae, elastin stains will show fine elastic fibres in the tunica media and coarse elastic fibres between the collagen fibres of the tunica adventitia. The appearance of other structures will depend on the stain used together with the elastin stain. Eosin, the E in H&E, gives a pink colour to both collagen fibres and the cytoplasm of cells. Nuclei are not stained if the H is omitted from the H&E.
Draw either one large composite image containing the three tunics and the cellular and fibrous elements which form the tunics.
Alternatively, you can draw a low power overview and supplement it with high magnification illustrations of the individual tunics. Focus on an H&E stained section.

Capillaries

The sum of the diameters of all capillaries is significantly larger than that of the aorta (by about three orders of magnitude), which results in decreases in blood pressure and flow rate. Also, capillaries are very small vessels. Their diameter ranges from 4-15 µm. The wall of a segment of capillary may be formed by a single endothelial cell. This results in a very large surface to volume ratio. The low rate of blood flow and large surface area facilitate the functions of capillaries in

providing nutrients and oxygen to the surrounding tissue, in
the absorption of nutrients, waste products and carbon dioxide, and in
the excretion of waste products from the body.

These functions are also facilitated by a very simple organisation of the wall of capillaries. Only the tunica intima is present, which typically only consists of the endothelium, its basal lamina and an incomplete layer of cells surrounding the capillary, the pericytes . Pericytes have contractile properties and can regulate blood flow in capillaries. In the course of vascular remodelling and repair, they can also differentiate into endothelial and smooth muscle cells.

Three types of capillaries can be distinguished based on features of ethe endothelium.

Continuous capillaries are formed by "continuous" endothelial cells and basal lamina. The endothelial cell and the basal lamina do not form openings, which would allow substances to pass the capillary wall without passing through both the endothelial cell and the basal lamina. Both endothelial cells and the basal lamina can act as selective filters in continuous capillaries. Fenestrated capillaries The endothelial cell body forms small openings called fenestrations, which allow components of the blood and interstitial fluid to bypass the endothelial cells on their way to or from the tissue surrounding the capillary. The fenestrations may represent or arise from pinocytotic vesicles which open onto both the luminal and basal surfaces of the cell. The extent of the fenestration may depend on the physiological state of the surrounding tissue, i.e. fenestration may increase or decrease as a function of the need to absorb or secrete. The endothelial cells are surrounded by a continuous basal lamina, which can act as a selective filter. Discontinuous capillaries are formed by fenestrated endothelial cells, which may not even form a complete layer of cells. The basal lamina is also incomplete. Discontinuous capillaries form large irregularly shaped vessels, sinusoids or sinusoid capillaries. They are found where a very free exchange of substances or even cells between bloodstream and organ is advantageous (e.g. in the liver, spleen, and red bone marrow).

Suitable Slides: Sections of any well preserved tissue - H&E, Whipf's polychrome
cardiac and skeletal muscle, glands or the papillary layer of the skin contain dense capillary beds.

Cardiac Muscle, sheep - Whipf's polychrome
Large numbers of capillaries are present in almost all tissues. At least a few dozen cross sections are present in every sqr. mm of section of poorly vascularised tissues. There may be thousands in highly vascularised tissues. However, a "good" capillary is not that easy to find because of their small size and because the capillary walls are very thin, which often leads to the collapse of the capillary during tissue preparation.
Cardiac muscle is highly vascularised. Each muscle cell is surrounded by one or more capillaries. The capillaries roughly follow the course of the muscle cells. To find capillaries in transverse and longitudinal sections it is easiest to first find areas in which the muscle cells have been cut in these planes. Only one or two red blood cells fit side by side in the capillary. A single endothelial cell forms the wall around the entire circumference of a segment of the capillary. Endothelial cell nuclei are therefore not always visible, and some red blood cells are only surrounded by a fine line representing the capillary wall.
Identify and draw a few capillaries. Include some of the surrounding tissue features - maybe a cardiac muscle cell, a venule or arteriole - as a scale.

Veins

The walls of veins are thinner than the walls of arteries, while their diameter is larger. In contrast to arteries, the layering in the wall of veins is not very distinct. The tunica intima is very thin. Only the largest veins contain an appreciable amount of subendothelial connective tissue. Internal and external elastic laminae are absent or very thin. The tunica media appears thinner than the tunica adventitia, and the two layers tend to blend into each other. The appearance of the wall of veins also depends on their location. The walls of veins in the lower parts of the body are typically thicker than those of the upper parts of the body, and the walls of veins which are embedded in tissues that may provide some structural support are thinner than the walls of unsupported veins.

Venous vessels originate from the capillary network which coalesce into the smallest venous vessels the

Venules.

They are larger than capillaries. Small venules are surrounded by pericytes. A few smooth muscle cells may surround larger venules. The venules merge to form

Small to medium-sized veins

which contain bands of smooth muscle in the tunica media. The tunica adventitia is well developed. In some veins (e.g. the veins of the pampiniform plexus in the spermatic cord) the tunica adventitia contains longitudinally oriented bundles of smooth muscle.
Aside from most veins in the head and neck, small to medium-sized veins are also characterised by the presence of valves. The valves are formed by loose, pocket-shaped folds of the tunica intima, which extend into the lumen of the vein. The opening of the pocket will point into the direction of blood flow towards the heart. One to three (usually two) pockets form the valve. Blood flowing towards heart will pass the pockets. If the flow reverses, blood will fill the pockets which will occlude the lumen of the vein and prevent the return of blood into the part of the vein preceding the valve. The ability of the valves to prevent backflow depends to some extent on the state of contraction (tone) of the smooth muscle in the wall of the vein.

The largest veins of the abdomen and thorax

do contain some subendothelial connective tissue in the tunica intima, but both it and the tunica media are still comparatively thin. Collagen and elastic fibres are present in the tunica media. The tunica adventitia is very wide, and it usually contains bundles of longitudinal smooth muscle. The transition from the tunica adventitia to the surrounding connective tissue is gradual. Valves are absent.
Vasa vasorum are more frequent in the walls of large veins than in that of the corresponding arteries - probably because of the lower oxygen tension in the blood contained within them.

Suitable Slides: sections of veins - H&E, van Gieson or elastin (in combination with other stains)
Like arteries, veins and venules are present in many sections. The basic features of their structure are however more difficult to identify because of the thin walls of veins - in particular in small veins. It is best to resort to sections of large veins, which together with large muscular arteries often have their "own section" in teaching collections.

Vein, human - H&E
The tunica intima is very narrow and the internal elastic lamina is difficult to identify - even in elastin stained sections. A few elastic fibres below the endothelium form only a very thin and incomplete internal elastic lamina. Smooth muscle is present in the tunica media, but it is organised less regular than in the artery. The tunica media is, again as compared to the artery, very thin and there is no sharp border between the tunica media and the tunica adventitia. The tunica adventitia of the largest veins contains coarse collagen fibres, elastic fibres and longitudinal bundles of smooth muscle. Small and medium sized veins will not contain smooth muscle in the tunica adventitia.
Draw part of the wall of the vein, label the tunics and indicate the presence of smooth muscle, fibres and their types.

Suitable Slides: sections containing small to medium sized veins - H&E, van Gieson or trichrome
Unless a specifically prepared slide is available, I would recommend looking at skin slides, in which I have found quite a few nice valves in the veins located at the border between dermis and hypodermis.

Vein Valve - H&E
Unless the section has been specifically prepared to illustrate valves, you will have to search for a while - probably through several sections. If a vein with valves is present in the section, it should be easy to identify. Valves are only found in small to medium-sized veins. You should see one or two bands of tissue in the lumen of the vein. Each band is formed by two apposing layers of tunica intima. The bands may share their origin from the inner aspect of the wall of the vein or they may have separate origins. Folding of the tissue bands forming the valves is variable.
Draw the vein and valve and label the tunics which you can identify. Try to indicate the position of the section in the schematic drawing of the valve.
You can observe the function of vein valves. Put your finger firmly on one of the veins which are visible on the back of your hand. Use another finger to stroke over the vein in the direction of the wrist. The vein will empty and not backfill (closed valves!) until you release the first finger.

Additional Specialisations of Vessels

Small arteries and veins often form anastomosing networks, which provides routes for alternative blood supply and drainage if one of the vessels should become occluded because of pathological or normal physiological circumstances. Some arteries are however the only supply of blood to their target tissues. These arteries are call end arteries. Tissues which are supplied by end arteries die if the arteries become occluded.

The segments of the kidney and the heads of the gastrocnemius muscle are examples of tissues supplied by end arteries.

Arteries and veins may also form arteriovenous shunts, which can shunt the blood flow that otherwise would enter the capillary network between the vessels. These shunts usually contain specialisations of the smooth muscle in the region of the shunt. Arteriovenous shunts are frequently seen in the blood supply of distal parts of the limbs and the nose (thermoregulation) and in the blood supply of endocrine organs.

Lymphatic Vessels

Parts of the blood plasma will exude from the blood vessels into the surrounding tissues because of transport across the endothelium or because of blood pressure and the fenestration of some capillaries (this process is partly counteracted by the higher osmotic pressure of the blood). The fluid entering tissues from capillaries adds to the interstitial fluid normally found in the tissue. The surplus of liquid needs to be returned to the circulation. Lymph vessels are dedicated to this unidirectional flow of liquid, the lymph. Three types of lymph vessels can be distinguished based on their size and morphology.

Lymph capillaries

are somewhat larger than blood capillaries and very irregularly shaped. They begin as blind-ending tubes in connective tissue. The basal lamina is almost completely absent and the endothelial cells do not form tight junctions, which facilitates the entry of liquids into the lymph capillary. Temporary openings in the endothelial lining of the lymph capillaries also allow the entry of larger particles into the lymph capillaries (lipid droplets, which are absorbed from the lumen of the gut do not enter blood capillaries, but enter the circulation via lymph vessels which are found in the villi of the ileum and jejunum). Lymph capillaries merge to form

Lymph collecting vessels

which are larger and form valves but otherwise appear similar to lymph capillaries. The lymph is moved by the compression of the lymph vessels by surrounding tissues. The direction of lymph flow is determined by the valves. Lymph vessels empty intermittently into lymph nodes from which the lymph continues in efferent lymph vessels.

Only very little lymph is returned from the limbs if they are immobilized, which illustrates the importance of muscular action in lymph transport. This is also the reason for immobilizing limbs that are either infected or that have been bitten by venomous Australians. The effect can also be observed after long intercontinental flight when you may feel that your shoes and socks are just about one number too small. Finally, impeded lymph drainage is one of the problems associated with surgery which requires the removal of lymph nodes and which thereby interrupts the lymph collecting vessels.

Eventually the lymph collecting vessels merge to form

Lymph ducts

which contain one or two layers of smooth muscle cells in their wall (some textbooks call this layer the tunica media of lymph vessels). They also form valves. The walls of the lymph ducts are less flexible in the region of the attachment of the valves to the wall of the duct, which may give a beaded appearance to the lymph ducts. Peristaltic contractions of the smooth muscle contribute to the movement of lymph towards the heart in addition to the compression of the ducts by surrounding tissues.

The largest lymph duct of the body, the thoracic duct, drains lymph from the lower half and upper left quadrant of the body and empties the lymph into the circulation by merging with the vascular system close to the junction of the left internal jugular and subclavian veins. That it is the largest lymph duct does not mean that it is a large vessel when compared to the large arteries and veins. It actually is not much larger (about 5mm in diameter) than one of the superficial forearm veins.

Suitable Slides: Sections of small intestine - H&E

Jejunum, baboon - H&E
At low magnification you will see villi extending into the lumen of the jejunum. Find villi which are cut longitudinally and change to medium magnification. In some of the villi you will see fairly large open spaces, which are surrounded by a layer of flattened endothelial cells. They should not contain any red blood cells - if they do you are looking at a capillary, which also should be somewhat smaller. These openings represent the blind end of lymph capillaries which originate in the villi.
Their name, lacteals, is derived from the milky appearance of the lymph. This appearance is caused by suspended lipid droplets which enter these lymph capillaries.
Draw a villus containing a lymph capillary. Focus on the features of the lymph capillary and the surrounding connective tissue. Include, if possible, a blood capillary in your drawing.

The jejunum slide is also good for revision. You should be able to find columnar epithelium, goblet cells, smooth muscle, small nerves, a few ganglion cells and, of course, lots of loose connective tissue and blood vessels.

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last updated: 9/01/04

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