Using lectures (on the web-page of histology department), lecture presentations textbooks, additional literature and other resourses, students should prepare such theoretical questions:
1. General features of sceletal connective tissues.
2. Function, location in the body and structural characteristics of hyaline cartilage.
3. Function, location in the body and structural characteristics of elastic cartilage.
4. Function, location in the body and structural characteristics of fibrocartilage.
5. Structure and function of perichondrium.
6. Characteristic features of the cells and extracellular matrix of cartilage.
7. The steps in the histogenesis and growth of cartilage.
8. General features of the bone tissue.
9. Comparison of the various types of bone cells in terms of their origin, structure and primary functions.
10. Functions and physical properties of bone tissue relating them to specific components of the bone matrix.
11. Structural features of compact and spongy bones.
12. Structure and function of periosteum.
13. Beginning with embryonic mesenchymal tissue, list the steps of intramembranous bone formation.
14. Beginning with embryonic mesenchymal tissue, list the steps of endochondral bone formation.
15. Muscular tissues origin and general morphofunctional characteristic.
16. Morphologic and phylogenetic classification of muscular tissues.
17. Smooth muscles structure, disposition and contraction peculiarities.
18. Striated skeletal muscles disposition, histogenesis and functional peculiarities.
19. Microscopic and submicroscopic structure of skeletal muscular tissue fiber. Sarcomere.
20. Cardiac muscular tissue structure and function peculiarities.
21. Muscle structure as organ. Myon.
22. Muscular tissues aging and regeneration.
The skeletal system is composed of variety of specialized forms of connective tissue. The functional differences between the various tissues of the skeletal system relate principally to the different nature and proportion of the ground substance and fibrous elements of the extracellular matrix. The cell of all the skeletal tissues, like the cells of the connective tissue in general, have close structural and functional relationships and a common origin from primitive mesenchymal cells.
Cartilage is a specialized form of connective tissue in which the firm consistency of the extracellular matrix allows the tissue to bear mechanical stresses without permanent distortion. Another function of cartilage is to support soft tissues. Because it is smooth-surfaced and resilient, cartilage is a shock-absorbing and sliding area for joints and facilitates bone movements. Cartilage is also essential for the development and growth of long bones both before and after birth.
Cartilage consists of cells (chondrocytes, chondroblasts) and an extensive extracellular matrix composed of fibers and ground substance. Chondrocytes synthesize and secrete the extracellular matrix, and the cells themselves are located in matrix cavities called lacunae. Collagen, hyaluronic acid, proteoglycans, and small amounts of several glycoproteins are the principal macromolecules present in all types of cartilage matrix. Variations in the composition of these matrix components produce three types of cartilage.
In all three forms (hyaline cartilage, elastic cartilage, and fibrocartilage), cartilage is avascular and is nourished by the diffusion of nutrients from capillaries in adjacent connective tissue (perichondrium) or by synovial fluid from joint cavities. In some instances, blood vessels traverse cartilage to nourish other tissues, but these vessels do not supply nutrients to the cartilage. As might be expected of cells in an avascular tissue, chondrocytes exhibit low metabolic activity. Cartilage has no lymphatic vessels or nerves.
The perichondrium is a sheath of dense connective tissue that surrounds cartilage in most places, forming an interface between the cartilage and the tissue supported by the cartilage. The perihondrium harbors the vascular supply for the avascular cartilage and also contains nerves and lymphatic vessels. Articular cartilage, which covers the surfaces of the bones of movable joints, is devoid of perichondrium and is sustained by the diffusion of oxygen and nutrients from the synovial fluid.
Hyaline cartilage is the most common and best studied of the three forms. Fresh hyaline cartilage is bluish-white and translucent. In the embryo, it serves as a temporary skeleton until it is gradually replaced by bone.
Hyaline cartilage of trachea. H&E stain. Medium magnification.
In adult mammals, hyaline cartilage is located in the articular surfaces of the movable joints, in the walls of larger respiratory passages (nose, larynx, trachea, bronchi), in the ventral ends of ribs, where they articulate with the sternum, and in the epiphyseal plate, where it is responsible for the longitudinal growth of bone.
Schematic representation of molecular organization in cartilage matrix. Link proteins noncovalently bind the protein core of proteoglycans to the linear hyaluronic acid molecules. The chondroitin sulfate side chains of the proteoglycan electrostatically bind to the collagen fibrils, forming a cross-linked matrix. The oval outlines the area shown larger in the lower part of the figure.
Forty percents of the dry weight of hyaline cartilage consists of collagen embedded in a firm, hydrated gel of proteoglycans and structural glycoproteins. In routine histologic preparations, the collagen is indiscernible for two reasons: the collagen is in the form of fibrils, which have submicroscopic dimensions; and the refractive index of the fibrils is almost the same as that of the ground substance in which they are embedded. The high content of solvation water bound to the negative charges of glycosaminoglycans acts as a shock absorber or biomechanical spring; this is of great functional importance, especially in articular cartilages.
In addition to type II collagen and proteoglycan, an important component of cartilage matrix is the structural glycoprotein chondronectin, a macromolecule that binds specifically to glycosaminoglycans and collagen type II, mediating the adherence of chondrocytes to the extracellular matrix. The cartilage matrix surrounding each chondrocyte is rich in glycosaminoglycan and poor in collagen. This peripheral zone, called the territorial, or capsular, matrix, nistochemically exhibits an intense basophilia, metachromasia.'and greater PAS-positivity than does the matrix located between the capsules, the interterritorial matrix.
Except in the articular cartilage of joints, all hyaline cartilage is covered by a layer of dense connective tissue, the perichondrium, which is essential for the growth and maintenance of cartilage. It is rich in collagen type I fibers and contains numerous fibroblasts. Although cells in the inner layer of the perichondrium resemble fibroblasts, they are chondroblasts and easily differentiate into chondrocytes.
At the periphery of hyaline cartilage, young chondrocytes have an elliptic shape, with the long axis parallel to the surface. Farther in, they are round and may appear in groups of up to eight cells originating from mitotic divisions of a single chondrocyte. These groups are called isogenous.
Cartilage cells and the matrix shrink during routine histologic preparation, resulting in both the irregular shape of the chondrocytes and their retraction from the capsule. In living tissue, and in properly prepared sections, the chondrocytes fill the lacunae completely. Chondrocytes synthesize collagen (mainly type II), proteoglycans, hyaluronic acid, and chondronectin.
Chondrocyte function depends on a proper hormonal balance. The synthesis of sulfated glycosaminoglycans is accelerated by growth hormone, thyroxine, and testosterone and is slowed by cortisone, hydrocortisone, and estradiol. Cartilage growth depends mainly on the hypophyseal growth hormone somatotropin. This hormone does not act directly on cartilage cells but promotes the synthesis of somatomedin C in the liver. Somatomedin C acts directly on cartilage cells, promoting their growth. Cartilage cells can give rise to benign (chondroma) or malignant (chondrosarcoma) tumors.
Diagram of the area of transition between the perichondrium and the hyaline cartilage. As perichondrial cells differentiate into chondrocytes, they become round, with an irregular surface. Cartilage (interterritorial) matrix contains numerous fine collagen fibrils except around the periphery of the chondrocytes, where the matrix consists primarily of glycosaminoglycans; this peripheral region is called the territorial, or capsular, matrix.
Histogenesis. Cartilage derives from the mesenchyme. The first modification observed is the rounding up of the mesenchymal cells, which retract their extensions, multiply rapidly, and form mesenchymal condensations. The cells formed by this direct differentiation of mesenchymal cells, now called chondroblasts, have a ribosome-rich basophilic cytoplasm. Synthesis and deposition of the matrix then begin to separate the chondroblasts from one another. The differentiation of cartilage takes place from the center outward; therefore, the more central cells have the characteristics of chondrocytes, whereas the peripheral cells are typical chondroblasts. The superficial mesenchyme develops into chondroblasts and fibroblasts of the perichondrium.
Growth. The growth of cartilage is attributable to two processes: interstitial growth, resulting from the mitotic division of preexisting chondrocytes; and appositional growth, resulting from the differentiation of perichondrial cells. In both cases, newly formed chondrocytes synthesize collagen fibrils and ground substance. Real growth is thus much greater than that from the simple increase in the number of cells. Interstitial growth is the less important of the two processes. It occurs only during the early phases of cartilage formation, when it increases tissue mass by expanding the cartilage matrix from within. Interstitial growth also occurs in the epiphyseal plates of long bones and within articular cartilage. In the epiphyseal plates, interstitial growth is important in increasing the length of long bones and in providing a cartilage model for endochondral bone formation. In articular cartilage, as the cells and matrix near the articulating surface are gradually worn away, the cartilage must be replaced from within, since there is no perichondrium there to add cells by apposition. In cartilage found elsewhere in the body, interstitial growth becomes less pronounced as the matrix becomes increasingly rigid from the cross-linking of matrix molecules. Cartilage then grows in girth only by apposition. Chondroblasts of the perichondrium proliferate and become chondrocytes once they have surrounded themselves with cartilaginous matrix and are incorporated into the existing cartilage.
In contrast to other tissues, hyaline cartilage is more susceptible to degenerative aging processes. Calcification of the matrix, preceded by an increase in the size and volume of the chondrocytes and followed by their death, is a common process in some cartilage, providing a model for bone development. Asbestiform degeneration, frequent in aged cartilage, is due to the formation of localized aggregates of thick, abnormal collagen fibrils.
Histogenesis of hyaline cartilage. A: The mesenchyme is the precursor tissue of all types of cartilage. B: Mitotic proliferation of mesenchymal cells gives rise to a highly cellular tissue. C: Chondroblasts are separated from one another by the formation of a great amount of matrix. D: Multiplication of cartilage cells gives rise to isogenous groups, each surrounded by a condensation of territorial (capsular) matrix.
Except in young children, damaged cartilage regenerates with difficulty and often incompletely, by activity of the perichondrium, which invades the injured area and generates new cartilage. In extensively damaged areas—and occasionally in small areas—the perichondrium produces a scar of dense connective tissue instead of forming new cartilage.
Elastic cartilage is found in the auricle of the ear, the walls of the external auditory canals, the auditory (eustachian) tubes, the epiglottis, and the cuneiform cartilage in the larynx.
Elastic cartilage, stained for elastic fibers. This flexible cartilage is present, for example, in the auricle of the ear and in the epiglottis. Orsein stain. Low magnification.
Elastic cartilage is essentially identical to hyaline cartilage except that it contains an abundant network of fine elastic fibers in addition to collagen type II fibrils. Fresh elastic cartilage has a yellowish color due to the presence of elastin in the elastic fibers, which can be demonstrated by standard elastin stains, eg. orcein.
The chondrocytes of elastic and hyaline cartilage tissues are similar, and elastic cartilage is frequently found to be gradually continuous with hyaline cartilage. Like hyaline cartilage, elastic cartilage possesses a perichondrium.
Elastic cartilage. Orsein stain. High magnification.
Fibrocartilage is a tissue intermediate between dense connective tissue and hyaline cartilage. It is found in intervertebral disks, in attachments of certain ligaments to the cartilaginous surface of bones, and in the symphysis pubis. Fibrocartilage is always associated with dense connective tissue, and the border areas between these two tissues are not clear-cut, showing a gradual transition.
Fibrocartilage contains chondrocytes similar to those of hyaline cartilage, either singly or in isogenous groups. The chondrocytes are very often arranged in long rows. The fibrocartilage matrix is acidophilic, because it contains a great number of coarse type I collagen fibers, which are easily seen under the microscope.
In fibrocartilage, the numerous collagen fibers either form irregular bundles between the groups of chondrocytes or are aligned in a parallel arrangement along the columns of chondrocues. This orientation depends on the stresses acting on fibrocartilage, since the collagen bundles take up a direction parallel to those stresses. There is no identifiable perichondrium in fibrocartilage.
Fibrocartilage. Note the rows of chondrocytes separated by collagen fibers. Fibrocartilage is frequently found in the insertion of tendons on the epiphyseal hyaline cartilage. Hematoxylin and eosin stain. Low magnification.
Electron micrograph of fibrocartilage from a young animal, showing 3 chondrocytes in their lacunae. Note the abundance of rough endoplasmic reticulum. Chondrocytes synthesize the cartilage matrix. Fine collagen fibers, sectioned in several places, are prominent around the chondrocytes. x3750.
Fibrocartilage of the intervertebral disk. Hematoxylin and eosin stain. High magnification.
Characteristics of cartilage types
Type II collagen
Predominent tissue component. Includes glycosaminogli-cans. Proteoglycan aggregated.
Cells and fibers embedded in abundant ground substance. Notable lack of capillaries. Cells may occur in esogenous groups. Fibers difficult to distinguish from ground substance. Extensive cross-linking among ground substance components and between fibers and ground substance
Articular and costal carilages;
Laryngeal, tracheal, and bronchial cartilages
Elastic fibers, type II collagen
Same as hyaline
Organization identical to that of hyaline cartilage exept for presence of a dense network of elastic fibers
Cotniculate and cuneiform cartilages of larynx
Type I collagen, type II collagen
Similar to that of hyaline but with equal amounts of chondroitin and dermatan sulfates
Cross between cartilage and dense regular connective tissue. Chondrocytes are usually in rowlike isogenous groups surrounded by typical hyaline matrix. Nests of chondrocytes lie between densely packed bundles of large type I collagen fibers.
Annulus fibrosus of intervertebral disks;
Always found in assotiation with dense connective tissue
Intervertebral disks Each intervertebral disk is situated between two vertebrae and held to them by means of ligaments.The disks have two components: the cartilaginous annulus fibrosus and the liquid nucleus pulposus. The intervertebral disk acts as a lubricated cushion that prevents adjacent vertebrae from being eroded by abrasive forces during movement of the spinal column. The nucleus pulposus serves as a shock absorber to cushion the impact between vertebrae.
The annulus fibrosus has an external layer of dense connective tissue, but it is mainly composed of overlapping laminae of fibrocartilage in which collagen bundles are orthogonally arrenged in adjacent layers. The multiple lamellae, with the 90-degree registration of type I collagen fibers in adjacent layers, provide the disk with unussual resilience that enables it to withstand the pressures generated by impining vertebrae. In tangential section, the disk presents a characteristic herringbone pattern as a result of the orthogonal alignment of collagen in alternating lamellae.
The nucleus pulposus is situated in the center of the annulus fibrosus. It is derived from the notochord and consists of a few rounded cells embedded in a viscous matrix rich in hyaluronic acid and type II collagen fibrils. In children, the nucleus pulposus is large, but it gradually becomes smaller with age and is partially replaced by fibrocartilage.
As the main constituent of the adult skeleton, bone tissue supports fleshy structures, protects such vital organs as those in the cranial and thoracic cavities, and harbors the bone marrow, where blood cells are formed. Bone also serves as a reservoir of calcium, phosphate, and other ions that can be released or stored in a controlled fashion to maintain constant concentrations of these important ions in body fluids. In addition to these functions, bones form a system of levers that multiply the forces generated during skeletal muscle contraction and transform them into bodily movements.
Bone is a specialized connective tissue composed of intercellular calcified material, the bone matrix, and three cell types: osteocytes, which are found in cavities (lacunae) within the matrix; osteoblasts, which synthesize the organic components of the matrix; and osteoclasts, which are multinucleated giant cells involved in the resorption and remodeling of bone tissue.
Since metabolites are unable to diffuse through the calcified matrix of bone, the exchanges between osteocytes and blood capillaries depend on communication through the canaliculi – thin, cylindrical spaces that perforate the matrix.
All bones are lined on both internal and external surfaces by layers of tissue containing osteogenic cells—endosteum on the internal surface and periosteum on the external surface.
Osteoblasts are responsible for the synthesis of the organic components of bone matrix (type I collagen, proteoglycans, and glycoproteins). Osteoblasts are exclusively located at the surfaces of bone tissue, side by side, in a way that resembles simple epithelium. When they are actively engaged in matrix synthesis, osteoblasts have a cuboidal to columnar shape and basophilic cytoplasm. When their synthesizing activity declines, they flatten, and cytoplasmic basophilia declines.
Some osteoblasts are gradually surrounded by newly formed matrix and become osteocytes. During this process a space called a lacuna is formed. Lacunae are occupied by osteocytes and their extensions, along with a small amount of extracellular noncalcified matrix.
During matrix synthesis, osteoblasts have the ultrastructure of cells actively synthesizing proteins for export. Osteoblasts are polarized cells. Matrix components are secreted at the cell surface, which is in contact with older bone matrix, producing a layer of new (but not yet calcified) matrix, called osteoid, between the osteoblast layer and the previously formed bone. This process, bone apposition, is completed by subsequent deposition of calcium salts into the newly formed matrix.
Osteocytes, which derive from osteoblasts, lie in the lacunae situated between lamellae of matrix. Only one osteocyte is found in each lacuna. The thin, cylindrical matrix canaliculi house cytoplasmic processes of osteocytes. Processes of adjacent cells make contact via gap junctions, and molecules are passed via these structures from cell to cell. Some molecular exchange between osteocytes and blood vessels also takes place through the small amount of extracellular substance located between osteocytes (and their processes) and the bone matrix. This exchange can provide nourishment for a chain of about 15 cells.
Osteoclasts are very large, branched motile cells. Dilated portions of the cell body contain from 5 to 50 (or more) nuclei. In areas of bone undergoing resorption, osteoclasts lie within enzymatically etàched depressions in the matrix known as Howship's lacunae. Osteoclasts are derived from the fusion of bone marrow-derived cells, and belong to the mononuclear phagocyte system.
Section of bone tissue showing an osteocyte with its cytoplasmic processes surrounded by matrix. The ultrastructure of the cell nucleus and cytoplasm is compatible with a low level of protein synthesis.
When compared with osteoblasts, the flat, almond-shaped osteocytes exhibit a significantly reduced rough endoplasmic reticulum and Golgi complex and more condensed nuclear chromatin. These cells are actively involved in the maintenance of the bonå matrix, and their death is followed by resorption of this matrix.
Section showing 1 oxyphylic osteoclast in the bone tissue. The osteoclast is a large cell with several nuclei and a ruffled border close to the bone matrix. Note the clear compartment where the process of bone erosion occurs. This compartment is acidified by a proton pump localized in the osteoclast membrane. It is the place of decalcification and matrix digestion and can be compared to a giant extracellular lysosome. Chondroclasts found in eroded regions of epiphyseal calcified cartilage are similar in shape to osteoclasts.
In active osteoclasts, the surface-facing bone matrix is folded into irregular, often subdivided projections, forming a ruffled border. Surrounding the ruffled border is a cytoplasmic zone—the clear zone—that is devoid of organelles, yet rich in actin microfilaments. This zone is a site of adhesion of the osteoclast to the bone matrix and creates a microenvironment in which bone resorption occurs.
The osteoclast secretes collagenase and other enzymes and pumps protons into a subcellular pocket (the microenvironment referred to above), promoting the localized digestion of collagen and dissolving calcium salt crystals. Osteoclast activity is controlled by cytokines (small signaling proteins that act as local mediators) and hormones. Osteoclasts have receptors for calcitonin, a thyroid hormone, but not for parathyroid hormone. However, osteoblasts have receptors for parathyroid hormone and, when activated by this hormone, produce a cytokine called osteoclast stimulating factor.
Lysosomal enzymes packaged in the Golgi complex and hydrogen ions produced are released into the confined microenvironment created by the attachment between bone matrix and the osteoclast’s peripheral clear zone. The acidification of this confined space facilitates the dissolution of calcium phosphate from bone and is the optimal pH for the activity of lysosomal hydrolases. Bone matrix is thus removed and the products of bone resorption are taken up by the osteoclast’s cytoplasm, probably digested further, and transferred to blood capillaries.
Inorganic matter represents about 50-70% of the dry weight of bone matrix. Calcium and phosphorus are especially abundant, but bicarbonate, citrate, magnesium, potassium, and sodium are also found. In electron micrographs, hydroxyapatite crystals of bone appear as plates that lie alongside the collagen fibrils but are surrounded by ground substance. The surface ions of hydroxyapatite are hydrated, and a layer of water and ions forms around the crystal. This layer, the hydration shell, facilitates the exchange of ions between the crystal and the body fluids.
The organic matter in bone matrix is type I collagen and ground substance, which contains proteoglycan aggregates and several specific structural glycoproteins. Some of the glycoproteins are produced by osteoblasts and demonstrate affinity for both hydroxyapatite and the cell membrane; they might be involved in binding osteoblasts or osteoclasts to bone matrix. Bone glycoproteins may also be responsible for promoting calcification of bone matrix. Other tissues containing type I collagen are not normally calcified and do not contain these glycoproteins. Because of its high collagen content, decalcified bone matrix intensely binds stains for collagen fibers.
The association of hydroxyapatite with collagen fibers is responsible for the hardness and resistance of bone tissue. After a bone is decalcified, its shape is preserved, but it becomes as flexible as a tendon. Removal of the organic part of the matrix—which is mainly collagenous—also leaves the bone with its original shape; however, it becomes fragile, breaking and crumbling easily when handled.
PERIOSTEUM & ENDOSTEUM
External and internal surfaces of bone are covered by layers of bone-forming cells and connective tissue called periosteum and endosteum.
The periosteum consists of an outer layer of collagen fibers and fibroblasts. Bundles of periosteal collagen fibers, called Sharpey's fibers, penetrate the bone matrix, binding the periosteum to bone. The inner, more cellular layer of the periosteum is composed of fibroblast-like cells called osteoprogenitor cells, with the potential to divide by mitosis and differentiate into osteoblasts. Osteoprogenitor cells play a prominent role in bone growth and repair.
The endosteum lines all internal cavities within the bone and is composed of a single layer of flattened osteoprogenitor cells and a very small amount of connective tissue. The endosteum is therefore considerably thinner than the periosteum. The principal functions of periosteum and endosteum are nutrition of osseous tissue and provision of a continuous supply of new osteoblasts for repair or growth of bone.
TYPES OF BONE
Gross observation of bone in cross section shows dense areas without cavities—corresponding to compact bone—and areas with numerous interconnecting cavities—corresponding to cancellous (spongy) bone.
Under the microscope, however, both compact bone and the trabeculae separating the cavities of cancellous bone have the same basic histologic structure.
In long bones, the bulbous ends—called epiphyses—are composed of spongy bone covered by a thin layer of compact bone. The cylindrical pan—diaphysis (a growing between)—is almost totally composed of compact bone, with a small component of spongy bone on its inner surface around the bone marrow cavity. Short bones usually have a core of spongy bone completely surrounded by compact bone. The flat bones that form the calvaria have two layers of compact bone called plates (tables), separated by a layer of spongy bone called the diploe.
Microscopic examination of bone shows two varieties: primary, immature, or woven bone and secondary, mature, or lamellar bone. Primary bone is the first bone tissue to appear in embryonic development and in fracture repair and other repair processes. It is characterized by random disposition of fine collagen fibers, in contrast to the organized lamellar disposition of collagen in secondary bone.
Primary bone tissue is usually temporary and is replaced in adults by secondary bone tissue except in a very few places in the body, eg, near the sutures of the flat bones of the skull, in tooth sockets, and in the insertions of some tendons. In addition to the irregular array of collagen fibers, other characteristics of primary bone tissue are a lower mineral content (it is more easily penetrated by x-rays) and a higher proportion of osteocytes than that in secondary bone tissue.
Secondary bone tissue is the variety usually found in adults. It characteristically shows collagen fibers arranged in lamellae that are parallel to each other or concentrically organized around a vascular canal. The whole complex of concentric lamellae of bone surrounding a canal containing blood vessels, nerves, and loose connective tissue is called a haversian system, or osteon. Lacunae containing osteocytes are found between and occasionally within the lamellae. In each lamella, collagen fibers are parallel to each other. Surrounding each haversian system is a deposit of amorphous material called the cementing substance that consists of mineralized matrix wiih few collagen fibers.
In compact bone (eg, the diaphysis of long bones), the lamellae exhibit a typical organization consisting of haversian systems, outer circumferential lamellae, inner circumferential lamellae, and interstitial lamellae. Inner circumferential lamellae are located around the marrow cavity, and outer circumferential lamellae are located immediately beneath the periosteum. There are more outer than inner lamellae. Between the two circumferential systems are numerous haversian systems, including triangular or irregularly shaped groups of parallel lamellae called interstitial (or intermediate) lamellae. These structures are lamellae left by haversian systems destroyed during growth and remodeling of bone.
Each haversian system is a long, often bifurcated cylinder parallel to the long axis of the diaphysis. It consists of a central canal surrounded by 4-20 concentric lamellae. Each endosteum-lined canal contains blood vessels, nerves, and loose connective tissue. The haversian canals communicate with the marrow cavity, the periosteum, and one another through transverse or oblique Volkmann's canals. Volkmann's canals do not have concentric lamellae; instead, they perforate the lamellae. All vascular canals found in bone tissue come into existence when matrix is laid down around preexisting blood vessels.
Schematic drawing of the wall of a long-bone diaphysis showing 3 types of lamellar bone: haversian system and outer and inner circumferential lamellae.
The protruding haversian system on the left shows the orientation of collagen fibers in each lamella. At the right is a haversian system showing lamellae, a central blood capillary (there are also small nerves, not shown), and many osteocytes with their processes.
Examination of haversian systems with polarized light shows bright anisotropic layers alternating with dark isotropic layers. When observed under polarized light at right angles to their length, collagen fibers are birefringent (anisotropic). The alternating bright and dark layers are due to the changing orientation of collagen fibers in the lamellae. In each lamella, fibers are parallel to each other and follow a helical course. The pitch of the helix is, however, different for different lamellae, so that at any given point, fibers from adjacent lamellae intersect atapproximately right angles.
Schematic drawing of 2 osteocytes and part of a haversian system.
Collagen fibers of contiguous lamellae are sectioned at different angles. Note the numerous canaliculi that permit communication between lacunae and with the haversian canals. Although it is not apparent in this simplified diagram, each lamella consists of multiple parallel arrays of collagen fibers. In adjacent lamellae, the collagen fibers are oriented in different directions. The presence of large numbers of lamellae with differing fiber orientations provides the bone with great strength, despite its light weight.
There is great variability in the diameter of haversian canals. Each system is formed by successive deposits of lamellae, starting inward from the periphery, so that younger systems have larger canals. In mature haversian systems, the most recently formed lamella is the one closest to the central canal.
Thick section of bone illustrating the cortical compact bone and the lattice of trabeculae of cancellous bone. B: Section of cancellous (spongy) bone with its characteristic random disposition of collagen fibers. Picrosirius–polarized light (PSP) stain. Low magnification.
Lamellar (secondary) bone in which the collagen fibers can be parallel to each other (at left) or organized concentrically around neurovascular channels, to constitute the haversian systems, or osteons (in most of the figure). Among the numerous haversian systems are some interstitial lamellae. PSP stain. Low magnification.
Histogenesis. Bone can be formed in two ways: by direct mineralization of matrix secreted by osteoblasts (intramembranous ossification) or by deposition of bone matrix on a preexisting cartilage matrix (endochondral ossification).
In both processes, the bone tissue that appears first is primary, or woven. Primary bone is a temporary tissue and is soon replaced by the definitive lamellar, or secondary, bone. During bone growth, areas of primary bone, areas of resorption, and areas of secondary bone appear side by side. This combination of bone synthesis and removal (remodeling) occurs not only in growing bones but also throughout adult life, although its rate of change in adults is considerably slower.
Intramembranous Ossification. Intramembranous ossification, the source of most of the flat bones, is so called because it takes placewithin condensations of mesenchymal tissue. Thefrontal and parietal bones of the skull—as well as parts of the occipital and temporal bones and the mandible and maxilla—are formed by intramembranous ossification. This process also contributes to the growth of short bones and the thickening of long bones.
In the mesenchymal condensation layer, the starting point for ossification is called a primary ossification center. The process begins when groups of cells differentiate into osteoblasts. Osteoblasts produce bone matrix and calcification follows, resulting in the encapsulation of some osteoblasts, which thenbecome osteocytes. These islands of developing bone form walls that delineate elongated cavities containing capillaries, bone marrow cells, and undifferentiated cells. Several such groups arise almost simultaneously at the ossification center, so that the fusion of the walls gives the bone a spongy structure. The connective tissue that remains among the bone walls is penetrated by growing blood vessels and additional undifferentiated mesenchymal cells, giving rise to the bone marrow cells.
Events that occur during intramembranous ossification. Osteoblasts are synthesizing collagen, which forms a strand of matrix that traps cells. As this occurs, the osteoblasts gradually differentiate to become osteocytes. The lower part of the drawing shows an osteoblast being trapped in newly formed bone matrix.
Intramembranous ossification. Hematoxylin and eosin stain. High magnification.
The ossification centers of a bone grow radially and finally fuse together, replacing the original connective tissue. The fontanelles of newbom infants, for example, are soft areas in the skull that correpond to parts of the connective tissue that are not yet ossified.
The beginning of intramembranous ossification. Mesenchymal cells round up and form a blastema, from which osteoblasts differentiate, producing primary bone tissue.
In cranial flat bones there is a marked predominance of bone formation over bone resorption at both the internal and external surfaces. Thus, two layers of compact bone (internal and external plates) arise, whereas the central portion (diploe) maintains its spongy nature.
The portion of the connective tissue layer that does not undergo ossification gives rise to the endosteum and the periosteum of intramembranous bone.
Endochondral Ossification. Endochondral ossification takes place within a piece of hyaline cartilage whose shape resembles a small version, or model, of the bone to be formed. This type of ossification is principally responsible for the formation of short and long bones.
The octeocyte network participates of the cellular functional control on bone surface, such as the clasts and osteoblasts. The cytoplasmic prolongations arrive at the canaliculi and make contact with the surface cells or act via mediators. H&E stain. Low magnification
Endochondral ossification of a long bone consists of the following sequence of events. Initially, the first bone tissue appears as a hollow bone cylinder that surrounds the mid portion of the cartilage model. This structure, the bone collar, is produced by intramembranous ossification within the local perichondrium. In the next step, the local cartilage undergoes a degenerative process characterized by cell enlargement (hypertrophy), matrix calcification, and cell death, resulting in a three-dimensional structure formed by the remnants of the calcified cartilage matrix. This process begins at the central portion of the cartilage model (diaphysis), where blood vessels penetrate through the bone collar previously perforated by osteoclasts, bringing osteoprogenitor cells to this region. Next, osteoblasts adhere to the calcified cartilage matrix and produce continuous layers of primary bone that surround the cartilaginous matrix remnants.
Formation of a long bone on a model made of cartilage. Hyaline cartilage is stippled; calcified cartilage is black, and bone tissue is indicated by oblique lines. The 5 small drawings in the middle row represent cross sections through the middle regions of the figures shown in the upper row. Note the formation of the bone collar and primary and secondary ossification centers. Epiphyseal fusion with diaphysis, with disappearance of the epiphyseal cartilage, occurs at different times in the same bone.
At this stage, the calcified cartilage appears basophilic, and the primary bone is eosinophilic. In this way the primary ossification center is produced. Then, secondary ossification centers appear at the swellings in the extremities of the cartilage model (epiphyses). During their expansion and remodeling, the primary and secondary ossification centers produce cavities that are gradually filled with bone marrow.
In the secondary ossification centers, cartilage remains in two regions: the articular cartilage, which persists throughout adult life and does not contribute to bone growth in length, and the epiphyseal cartilage, also called epiphyseal plate, which connects the two epiphyses to the diaphysis. The metaepiphyseal cartilage is responsible for the growth in length of the bone, and it disappears in adults, which is why bone growth ceases in adulthood.
Schematic drawings showing the 3-dimensional shape of bone in the epiphyseal plate area. Hyaline cartilage is stippled; calcified cartilage is black, and bone tissue is shown as yellow hatched areas. The upper drawing shows the region represented 3-dimensionally in the lower drawing.
The closure of the epiphyses follows a chronologic order according to each bone and is complete at about 20 years of age. Through x-ray examination of the growing skeleton, it is possible to determine the "bone age" of a young person, noting which epiphyses are open and which are closed. Once the epiphyses have closed, growth in length of bones becomes impossible, although widening may still occur.
Photomicrograph of the epiphyseal plate, showing its 5 zones, the changes that take place in the cartilage, and the formation of bone. PT stain. Low magnification
Higher magnification of the epiphyseal plate showing details of the endochondral ossification. Cartilage matrix (purple) is covered by recently formed bone tissue (red). Bone marrow and fat cells fill up the space left by the new bone. Picrosirius-hematoxylin (PSH) stain. Medium magnification.
Metaepiphyseal cartilage is divided into five zones, starting from the epiphyseal side of cartilage: (1) The resting zone consists of hyaline cartilage without morphologic changes in the cells. (2) In the proliferative zone (columnar), chondrocytes divide rapidly and form columns of stacked cells parallel to the long axis of the bone. (3) The hypertrophic cartilage zone contains large chondrocytes whose cytoplasm has accumulated glycogen. The resorted matrix is reduced to thin septa between the chondrocytes. (4) Simultaneous with the death of chondrocytes in the calcified cartilage zone, the thin septa of cartilage matrix become calcified by the deposit of hydroxyapatite. (5) In the ossification zone, endochondral bone tissue appears. Blood capillaries and osteoprogenitor cells formed by mitosis of cells originating from the periosteum invade the cavities left by the chondrocytes. The osteoprogenitor cells form osteoblasts, which are distributed in a discontinuous layer over the septa of calcified cartilage matrix. Ultimately, the osteoblasts deposit bone matrix over the three-dimensional calcified cartilage matrix.
In summary, growth in length of a long bone occurs by proliferation of chondrocytes in the epiphyseal plate adjacent to the epiphysis. At the same time, chondrocytes of the diaphyseal side of the plate hypertrophy; their matrix becomes calcified, and the cells die. Osteoclasts lay down a layer of primary bone on the calcified cartilage matrix. Because the rates of these two opposing events (proliferation and destruction) are approximately equal, the epiphyseal plate does not change thickness. Instead, it is displaced away from the middle of the diaphysis, resulting in growth in length of the bone.
Schematic drawing of diaphyseal bone remodeling showing 3 generations of haversian systems and their successive contributions to the formation of intermediate, or interstitial, lamellae. Remodeling is a continuous process responsible for bone adaptations, especially during growth.
Section of a haversian system, or osteon. Note the alternation of clear and dark circles resulting from the alternation in the direction of the collagen fibers. The collagen fibers appear bright when cut longitudinally and dark when cross-sectioned. In the center of the osteon is a channel. PSP stain. Medium magnification.
Section of endochondral ossification. The osseous matrix, rich in collagen type I, is specifically stained with picrosirius-hematoxylin. The cartilaginous matrix, containing collagen type II, stains blue with hematoxylin because of its high content of chondroitin sulfate. Medium magnification.
Schematic drawing of a diarthrosis. The capsule is formed by 2 parts: the external fibrous layer and the synovial layer (synovial membrane) that lines the articular cavity except for the cartilaginous areas (blue).
GENERAL FEATURES OF MUSCULAR TISSUE
Muscle cells are structurally and functionally specialized for contraction, which requires 2 types of special protein filaments called myofilaments: thin filaments containing actin and thick filaments containing myosin.
Nearly all muscle cells arise from mesoderm. Exception: Smooth
muscles of the iris arise from ectoderm. Muscle tissues are composed of groups
of muscle cells organized by connective tissue elements. Their arrangement
allows the groups to act together or separately, generating mechanical forces
of varying strength. Muscle cells are typically longer than they are wide,
sometimes reaching lengths of
Structure of the 3 muscle types. The drawings at right show these muscles in cross section. Skeletal muscle is composed of large, elongated, multinucleated fibers. Cardiac muscle is composed of irregular branched cells bound together longitudinally by intercalated disks. Smooth muscle is an agglomerate of fusiform cells. The density of the packing between the cells depends on the amount of extracellular connective tissue present.
Types of Muscle Tissue: The main muscle tissue types are smooth muscle and the 2 types of striated muscle, skeletal and cardiac. Smooth muscle is found mainly in the walls of hollow organs (eg, intestines and blood vessels); its contraction is slow, often in waves, and under involuntary control. In histologic section, it lacks the banding pattern, or striation, seen in the other type 2 types. Skeletal muscle is found mainly in association with bones, which act as pulleys and levers to multiply the force of its quick, strong, voluntary contractions. Cardiac muscle is found exclusively in the walls of the heart; its contractions are quick, strong, rhythmic, and involuntary.
Skeletal muscle arises from mesenchyme of mesodermal origin. Mature skeletal muscle fibers are elongated, unbranched, cylindrical, multi-nucleated. The flattened, peripheral nuclei lie just under the sarcolemma (muscle cell plasma membrane); most of the organelles and sacroplasm (muscle cell cytoplasm) are near the poles of the nuclei. The sarcoplasm contains many mitochondria, glycogen granules, and an oxygen-binding protein called myoglubin, and it accumulates lipofuscin pigment with age. Mature skeletal muscle fibers cannot divide.
Schematic drawing of longitudinal section of skeletal muscle fibers. Note the red A bands and the light-pink bands,
which are crossed by Z lines. High magnification.
Myofilaments In skeletal muscle fibers, these are of 2 major types. a. Thin filaments and thick filaments. The grouping of myofilaments into parallel bundles of thick and thin filaments called myofibrils. Each muscle fiber may contain several myofibrils, the number depending on its size.
At both light and electron microscopic levels, each myofibril exhibits repeating, linearly arranged, functional sub-units called sarcomeres, which have bands (striations) running perpendicular to the long axis of the myofibril. The sarcomeres of each myofibril lie in register with those in adjacent myofibrils so that their bands appear continuous. The sarcomere is separated from its neighbors at each end by a dense Z line, or Z disk. A major protein of the Z disk, alpha actinin, anchors one end of the thin filaments and helps maintain proper spatial distribution. The thin filaments extend to the middle of the sarcomere. The center of each sarcomere is marked by the M line, which holds the thick filaments in place. The thick filament bundles lie at the center of each sarcomere, are bisected by the M line, and overlap the free ends of the thin filaments. The pattern of overlapping between the thick and thin filaments is responsible for the banding pattern and differs depending on the myofibrils' state of contraction.
The bands With the light microscope, skeletal muscle exhibits alternating light- and dark-staining bands running perpendicular to the long axis of the muscle fibers.
I bands The light-staining bands contain only thin filaments. They are known as I bands (isotropic) because they do not rotate polarized light. Each I band is bisected by a Z line. Thus each sarcomere has 2 half I bands, one at each end.
A bands One dark-staining band lies in the middle of each sarcomere and shows the position of the thick filament bundles. This is known as an A band (anisotropic) because it is birefrmgent (rotates polarized light). At the EM level, each A band has a lighter-staining central region termed the H band, which is bisected by an M line. The H band lies between the free ends of the thin filaments and contains only the shafts of myosin molecules. The darker peripheral portions of the A bands are regions of overlap between the thick and thin filaments and contain the heads of the myosin molecules. The interaction between the myosin heads of the thick filaments and the free ends of the thin filaments causes muscle contraction.
Structure and position of the thick and thin filaments in the sarcomere. The molecular structure of these components is shown at right
Electron micrograph of a longitudinal section of the skeletal muscle of a monkey. Note the mitochondria (M) between adjacent myofibrils. The arrowheads indicate triads—2 for each sarcomere in this muscle—located at the A–I band junction. A, A band; I, I band; Z, Z line. x40,000
Schematic representation of the thin filament, showing the spatial configuration of 3 major protein components—actin, tropomyosin, and troponin. The individual components in the upper part of the drawing are shown in polymerized form in the lower part. The globular actin molecules are polarized and polymerize in one direction. Note that each tropomyosin molecule extends over 7 actin molecules. TnI, TnC, and TnT are troponin subunits.
Transverse section of skeletal muscle myofibrils. I, I band; A, A band; H, H band; Z, Z line. x36,000.
Segment of mammalian skeletal muscle.
The sarcolemma and muscle fibrils are partially cut, showing the following components: The invaginations of the T system occur at the level of transition between the A and I bands twice in every sarcomere. They associate with terminal cisternae of the sarcoplasmic reticulum (SR), forming triads. Abundant mitochondria lie between the myofibrils. The cut surface of the myofibrils shows the thin and thick filaments. Surrounding the sarcolemma are a basal lamina and reticular fibers.
Muscle contraction, initiated by the binding of Ca2+ to the TnC unit of troponin, which exposes the myosin binding site on actin (cross-hatched area). In a second step, the myosin head binds to actin and the ATP breaks down into ADP, yielding energy, which produces a movement of the myosin head. As a consequence of this change in myosin, the bound thin filaments slide over the thick filaments. This process, which repeats itself many times during a single contraction, leads to a complete overlapping of the actin and myosin and a resultant shortening of the whole muscle fiber. I, T, C are troponin subunits.
Ultrastructure of the motor end-plate and the mechanism of muscle contraction.
The drawing at the upper right shows branching of a small nerve with a motor end-plate for each muscle fiber. The structure of one of the bulbs of an end-plate is highly enlarged in the center drawing. Note that the axon terminal bud contains synaptic vesicles. The region of the muscle cell membrane covered by the terminal bud has clefts and ridges called junctional folds. The axon loses its myelin sheath and dilates, establishing close, irregular contact with the muscle fiber. Muscle contraction begins with the release of acetylcholine from the synaptic vesicles of the end-plate. This neurotransmitter causes a local increase in the permeability of the sarcolemma. The process is propagated to the rest of the sarcolemma, including its invaginations (all of which constitute the T system), and is transferred to the sarcoplasmic reticulum (SR). The increase of permeability in this organelle liberates calcium ions (drawing at upper left) that trigger the sliding filament mechanism of muscle contraction. Thin filaments slide between the thick filaments and reduce the distance between the Z lines, thereby reducing the size of all bands except the A band. H, H band; S, sarcomere.
Muscle spindle showing afferent and efferent nerve fibers that make synapses with the intrafusal fibers (modified muscle fibers). Note the complex nerve terminal on the intrafusal fibers. The two types of intrafusal fibers, one with a small diameter and the other with a dilation filled with nuclei, are shown. Muscle spindles participate in the nervous control of body posture and the coordinate action of opposing muscles.
Drawing of a Golgi tendon organ. This structure collects information about differences in tension among tendons and relays data to the central nervous system, where they are processed and help to coordinate fine muscular contractions.
Longitudinal section of striated muscle fibers. The blood vessels were injected with a plastic material before the animal was killed. Note the extremely rich network of blood capillaries around the muscle fibers. Giemsa stain. Photomicrograph of low magnification made under polarized light
Section of tongue, an organ rich in striated skeletal muscle fibers. These fibers appear brown because the section was immunohistologically stained to show myoglobin. The light-colored areas among and above the muscle fibers contain connective tissue. In the upper region of the section, stratified and cornified epithelium can be seen. Nuclei are stained by hematoxylin. Low magnification.
Striated skeletal muscle in longitudinal section (lower) and in cross section (upper). The nuclei can be seen in the periphery of the cell, just under the cell membrane, particularly in the cross sections of these striated fibers. H&E stain. Medium magnification.
Sarcoplasmic reticulum This is the smooth endoplasmic reticulum of striated muscle cells, specialized to sequester calcium ions. In skeletal muscle, it consists of an anas-tomosing complex of membrane-limited tubules and cisternae that ensheathe each myofibril. At each A-I band junction, a tubular invagination of the sarcolemma termed a transverse tubule, or T tubule, penetrates the muscle fiber and comes to lie close to the surface of the myofibrils. On each side of the T tubule lies an expansion of the sarcoplas-mic reticulum termed a terminal cisterna. A complex of 2 terminal cisternae and an intervening T tubule constitutes a triad. Triads have an important role in initiating muscle contraction.
Energy Production: Muscles use glucose (from stored glycogen and from the blood) and fatty acids (from the blood) to form the ATP and phosphocreatine that provide chemical energy for contraction. When ATP is not available, actin-myosin binding becomes stabilized, accounting for rigor mortis, the muscular rigidity that occurs shortly after death.
Other Components of the Sarcoplasm
Glycogen is found in abundance in the sarcoplasm in the form of coarse granules. It serves as a depot of energy that is mobilized during muscle contraction.
Another component of the sarcoplasm is myoglobin; this oxygen-binding protein, which is similar to hemoglobin, is principally responsible for the dark red color of some muscles. Myoglobin acts as an oxygen-storing pigment, which is necessary for the high oxidative phosphorylation level in this type of fiber. For obvious reasons, it is present in great amounts in the muscle of deep-diving ocean mammals (eg, seals, whales). Muscles that must maintain activity for prolonged periods usually are red and have a high myoglobin content. Mature muscle cells have negligible amounts of rough endoplasmic reticulum and ribosomes, an observation that is consistent with the low level of protein synthesis in this tissue.
Organization of the Skeletal Muscles:
Named muscles are bundles of muscle fascicles surrounded by a sheath of dense connective tissue termed the epimysium. Each fascicle is a bundle of muscle fibers surrounded by a dense connective tissue sheath called the perimysium, which consists of septumlike inward extensions of epimysium. Each muscle fiber is a bundle of myofibrils surrounded by a delicate connective tissue sheath termed the endo-mysium, which consists of a basal lamina and a loose meshwork of reticular fibers. Each myofibril is a bundle of myofilaments surrounded by an investment of sarcoplasmic reticulum, with a triad at both A-I junctions of each sarcomere. The connective tissue investments are continuous with one another. They bind together subunits that function together and separate subunits that function independently.
Structure and function of skeletal muscle. The drawing at right shows the area of muscle detailed in the enlarged segment. Color highlights endomysium, perimysium, and epimysium.
Cross section of striated muscle stained to show collagens type I and III and cell nuclei. The endomysium is indicated by arrowheads and the perimysium by arrows. At left is a piece of epimysium. Picrosirius-hematoxylin stain. High magnification
Schematic representation of muscle structure
Cardiac muscle arises as parallel chains of elongated splanchnic mesenchymal cells in the walls of the embryonic heart tube (myoepicardial plate of visceral mesoderm – splanchnic layer).
Cardiac muscle cells are elongated, cilindrycal cells with one or 2 elongated, central nuclei. The sarcoplasm near the nuclear poles contains many mitochondria and glycogen granules and some lipofuscin pigment. Mitochondria lie in chains between the myofilaments. The arrangement of myofilaments yields a pattern of striations identical to that of skeletal muscle.
Drawing of a section of heart muscle, showing central nuclei, cross-striation, and intercalated disks.
Photomicrograph of cardiac muscle. Note the cross-striation and the intercalated disks (arrowheads). Pararosaniline–toluidine blue (PT) stain. High magnification.
Sarcoplasmic reticulum and T tubule system The sarcoplasmic reticulum in cardiac muscle fibers is less organized than that of skeletal muscle and does not subdivide myofilaments into discrete myofibrillar bundles. Cardiac T tubules are located at the Z line instead of the A-I junction. In most cells, cardiac T tubules are associated with a single expanded cisterna of the sarcoplasmic reticulum; thus, cardiac muscle contains dyads instead of triads.
Intercalated disks These unique histologic features of cardiac muscle appear as dark transverse lines between the muscle fibers and represent specialized junctional complexes. With the electron microscope, intercalated disks can be seen to have 3 major components arranged in a stepwise fashion.
a. The fascia adherens, similar to a zonula adherens, is a half Z line found in the vertical (transverse) portion of the step. Its alpha actinin anchors the thin filaments of the terminal sarcomeres.
b. The macula adherens (desmosome) is the second component of transverse portion of the junction. It prevents detachment of the cardiac muscle fibers from one another during contraction.
c. The gap junction of intercalated disks comprise the horizontal (lateral) portion of the step. They provide electrotonic coupling between adjacent cardiac muscle fibers and pass the stimulus for contraction from cell to cell.
Drawing of longitudinal section of human cardiac muscle, stained with iron haematoxylin to show intercalated discs
Longitudinal section of portions of 2 cardiac muscle cells. The transversely oriented parts of the intercalated disk consist of a fascia adherens and numerous desmosomes. The longitudinal parts (arrows) contain gap junctions. Mitochondria (M) are numerous. Fibrils of reticular fibers are seen between the two cells. x18,000.
Electron micrograph of a longitudinal section of heart muscle. Note the striation pattern and the alternation of myofibrils and mitochondria rich in cristae. Note the sarcoplasmic reticulum (SR), which is the specialized calcium-storing smooth endoplasmic reticulum. x30,000.
Ultrastructure of heart muscle in the region of an intercalated disk. Contact between cells is accomplished by interdigitation in the transverse region; contact is broad and flat in the longitudinal plane (LP). A, A band; I, I band; Z, Z line.
Junctional specializations making up the intercalated disk. Fasciae (or zonulae) adherentes (A) in the transverse portions of the disk anchor actin filaments of the terminal sarcomeres to the plasmalemma. Maculae adherentes, or desmosomes (B), found primarily in the transverse portions of the disk, bind cells together, preventing their separation during contraction cycles. Gap junctions (C), restricted to longitudinal portions of the disk—the area subjected to the least stress—ionically couple cells and provide for the spread of contractile depolarization.
Organization of Cardiac Muscle: Because of the abundant capillaries in the endomysium of each fiber, cardiac muscle fibers appear more loosely arranged in histologic section than those of skeletal muscle. The whorled arrangement of cardiac muscle fibers in the wall of the heart accounts for the ability of the myocardium to "wring out" blood in the heart chambers.
Initiation of Cardiac Muscle Contraction: Unlike skeletal muscle fibers, which rarely contract without direct motor innervation, cardiac muscle fibers contract spontaneously with an intrinsic rhythm. The heart receives autonomic innervation that cannot initiate contraction but can speed up or slow down the intrinsic beat. The initiating stimulus for contraction is normally provided by a collection of specialized cardiac muscle cells called the sinoatrial node; it is delivered by other specialized fibers, called Purkinje fibers, to the other cardiac muscle cells (conductive system of the heart). The stimulus is passed between adjacent cells through the gap junctions of the intercalated disks. The gap junctions establish an ionic continuity among cardiac muscle fibers that allows them to work together as a functional syncytium.
Electron micrograph of an atrial muscle cell showing the presence of natriuretic granules aggregated at the nuclear pole.
Most smooth muscle cells differentiate from mesenchymal cells of mesodermal origin in the walls of developing hollow organs of cardiovascular, digestive, urinary, and reproductive systems. During differentiation, the cells elongate and accumulate myofilaments. Smooth muscles of the iris arise from ectoderm.
Smooth Muscle Cells: Mature smooth muscle fibers are elongated, spindle-shaped cells with a single central ovoid nucleus. The sarcoplasm at the nuclear poles contains abundant mitochondria, some rough endoplasmic reticulum, and a large Golgi complex. Each fiber produces its own basal lamina, consisting of proteoglycan-rich material and type III collagen fibers.
Photo of smooth muscle cells of urinary bladder in longitudinal section (upper) and in cross section (lower). Note the centrally located nuclei. In many cells the nuclei were not included in the section. H&E stain. Medium magnification.
Thin myofilaments The actin filaments of smooth muscle are like those of skeletal and cardiac muscle. They are always present in the cytoplasm and are anchored in dense bodies associated with the plasma membrane.
Thick filaments The myosin filaments of smooth muscle are less stable than those in striated muscle cells; they are not always present in the cytoplasm but seem to form in response to a contractile stimulus. Unlike the thick filaments in striated muscle cells, those in smooth muscle have heads along most of their length and bare areas at the ends of the filaments.
Organization of the myofilaments The filaments run mostly parallel to the long axis of smooth muscle fibers, but they overlap much more than those of striated muscle, accounting for the absence of cross striations. The greater overlap of thick and thin filaments results from the unique organization of the thick filaments and permits greater contraction. The ratio of thin to thick filaments in smooth muscle is about 12:1, and the arrangement of the filaments is less regular and crystalline than in striated muscle.
Sarcoplasmic reticulum Smooth muscle cells contain a poorly organized sarcoplasmic reticulum; these fibers have no T tubules and no dyads or triads.
Organization of Smooth Muscle: Unlike striated-muscle fibers, which abut end-to-end, smooth muscle fibers overlap to varying degrees and attach to one another by fusing their endo-mysial sheaths. The sheaths are interrupted by many gap junctions, which transmit the ionic currents that initiate contraction. Smooth muscle fibers form fascicles that vary in size but are usually smaller than those m striated muscle. The fascicles, each surrounded by a meager penmysium, are often organized in layers separated by the thicker epimysial connective tissue. Fibers in adjacent layers often lie perpendicular to one another.
Drawing of a segment of smooth muscle. All cells are surrounded by a net of reticular fibers. In cross section, these cells show various diameters.
Transverse section of smooth muscle impregnated with silver to stain the reticular fibers. These fibers form a network that surrounds the muscle cells that are not stained by this method. At the right is an arteriole surrounded by thicker collagen fibers. x300.
Electron micrograph of a transverse section of smooth muscle. The cells are sectioned at various diameters and have many subsurface vesicles in their cytoplasm. Thick and thin filaments are not organized into myofibrils, and there are few mitochondria (M). Note the collagen fibrils of the reticular fibers and a small unmyelinated nerve (N) between the cells. x6650.
Smooth muscle cells relaxed and contracted. Cytoplasmic filaments insert on dense bodies located in the cell membrane and deep in the cytoplasm. Contraction of these filaments decreases the size of the cell and promotes the contraction of the whole muscle. During the contraction the cell nucleus is deformed.
The three types of adult muscle have different potentials for physiologic regeneration and reparative after injury.
Cardiac muscle physiologic regeneration occurs intracellular (cardiomyocytes are renewing their structural compounds mainly in diastole), but this muscular tissue has virtually no reparative capacity beyond early childhood. Defects or damage (e.g., infarcts) in heart muscle are generally replaced by the proliferation of connective tissue, forming myocardial scars.
In skeletal muscle, although the nuclei are incapable of undergoing mitosis, the tissue can undergo limited regeneration. The source of regenerating cells is believed to be the satellite cells. The latter are a sparse population of mononucleated spindle-shaped cells that lie within the basal lamina surrounding each mature muscle fiber. Because of their intimate apposition with the surface of the muscle fiber, they can be identified only with the electron microscope. They are considered to be inactive myoblasts that persist after muscle differentiation. After injury or certain other stimuli, the normally quiescent satellite cells become activated, proliferating and fusing to form new skeletal muscle fibers. A similar activity of satellite cells has been implicated in muscle hypertrophy, where they fuse with their parent fibers to increase muscle mass after extensive exercise. The regenerative capacity of skeletal muscle is limited, however, after major muscle trauma or degeneration.
Smooth muscle is capable of an active regenerative response. After injury, viable mononucleated smooth muscle cells and myofibroblasts of connective tissue undergo mitosis and provide for the replacement of the damaged tissue.
In contrast to other tissues, hyaline cartilage is more susceptible to degenerative aging processes. Calcification of the matrix, preceded by an increase in the size and volume of the chondrocytes and followed by their death, is a common process in some cartilage. "Asbestiform" degeneration, frequent in aged cartilage, is due to the formation of localized aggregates of thick, abnormal collagen fibrils
The fluorescent antibiotic tetracycline interacts with great affinity with recently deposited mineralized bone matrix. Based on this interaction, a method was developed to measure the rate of bone apposition—an important parameter in the study of bone growth and the diagnosis of bone growth diseases. Tetracycline is administered twice to patients, with an interval of 5 days between injections. A bone biopsy is then performed, and the sections are studied by means of fluorescence microscopy. The distance between the two fluorescent layers is proportional to the rate of bone apposition. This procedure is of diagnostic importance in such diseases as osteomalacia, in which mineralization is impaired, and osteitis fibrosa cystica, in which increased osteoclast activity results in removal of bone matrix and fibrous degeneration.
The organic matter embedded in the calcified matrix is type I collagen and ground substance, which contains proteoglycan aggregates and several specific multiadhesive glycoproteins, including osteonectin. Calcium-binding glycoproteins, notably osteocalcin, and the phosphatases released in matrix vesicles by osteoblasts promote calcification of the matrix. Other tissues containing type I collagen do not contain these glycoproteins or matrix vesicles and are not normally calcified. Because of its high collagen content, decalcified bone matrix is usually acidophilic.
The association of minerals with collagen fibers is responsible for the hardness and resistance of bone tissue. After a bone is decalcified, its shape is preserved, but it becomes as flexible as a tendon. Removal of the organic part of the matrix—which is mainly collagenous—also leaves the bone with its original shape; however, it becomes fragile, breaking and crumbling easily when handled.
Soon the blood clot is removed by macrophages and the adjacent matrix of bone is resorbed by osteoclasts. The periosteum and the endosteum at the site of the fracture respond with intense proliferation producing a soft callus of fibrocartilage-like tissue that surrounds the fracture and covers the extremities of the fractured bone (Figure 8–18).
Primary bone is then formed by a combination of endochondral and intramembranous ossification. Further repair produces irregularly formed trabeculae of primary bone that temporarily unite the extremities of the fractured bone, forming a hard bone callus (Figure 8–18).
Stresses imposed on the bone during repair and during the patient's gradual return to activity serve to remodel the bone callus. The primary bone of the callus is gradually resorbed and replaced by secondary bone, remodeling and restoring the original bone structure. Unlike other connective tissues, bone tissue heals without forming a scar.
Because the concentration of calcium in tissues and blood must be kept constant, nutritional deficiency of calcium results in decalcification of bones. Severely decalcified bones are more likely to fracture.
Decalcification of bone may also be caused by excessive production of PTH (hyperparathyroidism), which can cause increased osteoclastic activity, intense resorption of bone, elevation of blood Ca2+ and PO3– 4 levels, and abnormal deposits of calcium in the kidneys and arterial walls.
The opposite occurs in osteopetrosis (L. petra, stone), a disease caused by defective osteoclast function that results in overgrowth, thickening, and hardening of bones. This process can obliterate the bone marrow cavities, depressing blood cell formation and causing anemia and the loss of white blood cells.
Especially during growth, bone is sensitive to nutritional factors. Calcium deficiency, which leads to incomplete calcification of the organic bone matrix, can be due either to a lack of calcium in the diet or a failure to produce the steroid prohormone vitamin D, which is important for the absorption of Ca2+ and> PO3–4 by the small intestine.
Calcium deficiency in children causes rickets, a disease in which the bone matrix does not calcify normally and the epiphyseal plate becomes distorted by the normal strains of body weight and muscular activity. Ossification processes at this level are consequently hindered, and the bones not only grow more slowly but also become deformed.
Calcium deficiency in adults gives rise to osteomalacia (osteon + Gr. malakia, softness), which is characterized by deficient calcification of recently formed bone and partial decalcification of already calcified matrix. Osteomalacia should not be confused with osteoporosis. In osteomalacia, there is a decrease in the amount of calcium per unit of bone matrix. Osteoporosis, frequently found in immobilized patients and in postmenopausal women, is an imbalance in skeletal turnover so that bone resorption exceeds bone formation.
In addition to PTH and calcitonin, several other hormones act on bone. The anterior lobe of the pituitary synthesizes growth hormone (GH or somatotropin), which stimulates the liver to produce insulin-like growth factor-1 (IGF-1 or somatomedin). IGF has an overall growth effect, especially on the epiphyseal cartilage. Consequently, lack of growth hormone during the growing years causes pituitary dwarfism; an excess of growth hormone causes excessive growth of the long bones, resulting in gigantism. Adult bones cannot increase in length even with excess IGF because they lack epiphyseal cartilage, but they do increase in width by periosteal growth. In adults, an increase in GH causes acromegaly, a disease in which the bones—mainly the long ones—become very thick.
The sex hormones, both male (androgens) and female (estrogens), have a complex effect on bones and are, in a general way, stimulators of bone formation. They influence the time of appearance and development of ossification centers and accelerate the closure of epiphyses.
Cancer originating directly from bone cells is fairly uncommon (0.5% of all cancer deaths) but a form called osteosarcoma can arise in osteoblasts. The skeleton is often the site of metastases from tumors originating from malignancies in other organs, most commonly from breast, lung, prostate, kidney, and thyroid tumors.
The variation in diameter of skeletal muscle fibers depends on factors such as the specific muscle and the age and sex, state of nutrition, and physical training of the individual. It is a common observation that exercise enlarges the musculature and decreases fat depots. The increase in muscle thus obtained is caused by formation of new myofibrils and a pronounced growth in the diameter of individual muscle fibers. This process, characterized by increased of cell volume, is called hypertrophy (Gr. hyper, above, + trophe, nourishment). Tissue growth by an increase in the number of cells is termed hyperplasia (hyper + Gr. plasis, molding), which takes place most readily in smooth muscle, whose cells have not lost the capacity to divide by mitosis.
Myasthenia gravis is an autoimmune disorder characterized by progressive muscular weakness caused by a reduction in the number of functionally active acetylcholine receptors in the sarcolemma of the myoneural junction. This reduction is caused by circulating antibodies that bind to the acetylcholine receptors in the junctional folds and inhibit normal nerve-muscle communication. As the body attempts to correct the condition, membrane segments with affected receptors are internalized, digested by lysosomes, and replaced by newly formed receptors. These receptors, however, are again made unresponsive to acetylcholine by similar antibodies, and the disease follows its progressive course.
Interesting histology questions
Cartilage is a connective tissue that is found in three types: hyaline, fibrocartilage, and elastic. Apart from the epiglottis and larynx, where is the only other place elastic cartilage is found?
External ear. Hyaline cartilage is the most abundant type of cartilage, and is found in various places, such as covering bone ends in synovial joints, trachea, bronchi, and the nose. It appears as a shiny, blue-white substance. It contains fine collagen fibers, and has many chondrocytes situated in spaces named lacunae (which means "little lakes"). Usually it is surrounded by a perichondrium. Elastic cartilage contains a network of elastic fibers in which the chondrocytes are located. It provides strength, elasticity, and gives the shape of the ear. Try to fold your ear over itself and let go - it will ping back like an elastic band!
Mature bone is vascular - it contains blood vessels which are surrounded by rings of bone. This structure also contains the cells (osteoblasts/-cytes) between the rings of bone, and was named after the Englishman who first described them. His first name was Clopton; what was his last name?
Havers. Clopton Havers was a physician in the 17th century, and he undertook pioneering research on the microstructure of bone. The structures in the question are called Haversian canals (also Haversian systems or osteon). These are made up of a central canal, which contains a blood vessel. The vessel is surrounded by rings of bone called lamellae. There are osteoblasts and osteocytes present between the lamellae to maintain the Haversian canal. The osteocytes connect to each other through the lamellae via projections called canaliculi.
Osteoarthritis affects more women than men.
This is false. Actually, osteoarthritis affects men and women in equal numbers. It is a common type of arthritis and is degenerative, resulting in the wearing away of joints, especially the hip and knee. It can develop early, for example as the result of a sports injury. Symptoms include pain and stiffness in the joints, and is caused by the wearing away of the cartilage in the joint; the exposed bone becomes thickened (sclerotic) and shiny (eburnated). Also, cysts can develop in the bone, and bony projections called osteophytes can occur. These osteophytes can frequently be seen on the distal phalanges. They are then called Heberden's nodes and are more commonly seen in women.
1. It takes 17 muscles to smile and 43 to frown. Unless you’re trying to give your face a bit of a workout, smiling is a much easier option for most of us. Anyone who’s ever scowled, squinted or frowned for a long period of time knows how it tires out the face which doesn’t do a thing to improve your mood.
2. Babies are born with 300 bones, but by adulthood the number is reduced to 206. The reason for this is that many of the bones of children are composed of smaller component bones that are not yet fused like those in the skull. This makes it easier for the baby to pass through the birth canal. The bones harden and fuse as the children grow.
We are about
4. The strongest muscle in the human body is the tongue. While you may not be able to bench press much with your tongue, it is in fact the strongest muscle in your body in proportion to its size. If you think about it, every time you eat, swallow or talk you use your tongue, ensuring it gets quite a workout throughout the day.
5. The hardest bone in the human body is the jawbone. The next time someone suggests you take it on the chin, you might be well advised to take their advice as the jawbone is one of the most durable and hard to break bones in the body.
6. You use 200 muscles to take one step. Depending on how you divide up muscle groups, just to take a single step you use somewhere in the neighborhood of 200 muscles. That’s a lot of work for the muscles considering most of us take about 10,000 steps a day.
7. The tooth is the only part of the human body that can’t repair itself. If you’ve ever chipped a tooth you know just how sadly true this one is. The outer layer of the tooth is enamel which is not a living tissue. Since it’s not alive, it can’t repair itself, leaving your dentist to do the work instead.
8. It takes twice as long to lose new muscle if you stop working out than it did to gain it. Lazy people out there shouldn’t use this as motivation to not work out, however. It’s relatively easy to build new muscle tissue and get your muscles in shape, so if anything, this fact should be motivation to get off the couch and get moving.
9. Bone is stronger than some steel. This doesn’t mean your bones can’t break of course, as they are much less dense than steel. Bone has been found to have a tensile strength of 20,000 psi while steel is much higher at 70,000 psi. Steel is much heavier than bone, however, and pound for pound bone is the stronger material.
10. The feet account for one quarter of all the human body’s bones. You may not give your feet much thought but they are home to more bones than any other part of your body. How many? Of the two hundred or so bones in the body, the feet contain a whopping 52 of them.
11. Muscles can account for about 40% your body weight.
12. Muscles can only pull, they cannot, as some people assume, push.
13. The longest muscle has muscle cells that can be over a foot long.
14. The smallest muscles are in the middle of the ear;examples are the tensor tympani, and stapedius.
15. The strongest, pound for pound, are the masseters, the chewing muscles.
16. There are 640 individual names for muscles.
17. Muscles need OXYGEN and FOOD to function properly.
18. The hardest working muscles in the body are the muscles in the eye. It takes 17 muscles in the body to smile.
19. There are muscles in the root of our hair that gives us goose bumps!
20. You have all the muscle fiber you will ever have at birth. Once damaged they can't be replaced.
21. Arnold Schwarzenegger has as many muscle fibers as you - They're just thicker!
A single muscle cell of the sartorius muscle
in the thigh can be more than
23. There are more than 600 voluntary muscles in the body.
24. The strongest muscle of the body is the masseter muscle used for chewing!
25. Your hand contains 20 different muscles.
26. If all your muscles could pull in one direction you could create a force of 25 tons!
27. Muscles account for approximately 40% of your body weight.
28. It takes 17 muscles in your face to smile, but it takes 43 muscles to frown.
29. You take approximately 5 million steps per year using your leg muscles!
30. Muscles and Bones provide the framework for our bodies and allow us to jump, run or just lie on the couch.
Student’s Practical Activities
Task No 1. Students must know and illustrate such histologic specimens.
Specimen 1. Hyaline cartilage.
Stained with haematoxylin and eosin.
In the specimen of mature hyaline cartilage two distinct zones are evident: an inner, strongly basophilic zone and an outer, pale-stained zone which merges with adjacent connective tissue. The chondrocytes of the inner zone are arranged in characteristic clusters usually consisting of two or four fully differentiated cells. The clusters are separated by a large mass of amorphous cartilage matrix whilst the cells of each cluster are separated by only a thin zone of extracellular matrix. Mature chondrocytes are characterised by small nuclei with dispersed chromatin and basophilic, granular cytoplasm reflecting a well developed rough endoplasmic reticulum. Lipid droplets, often larger than the nuclei, are a prominent feature of chondrocytes; the cytoplasm is also rich in glycogen.
The matrix of hyaline cartilage appears fairly amorphous since the ground substance and collagen have similar refractive indices. With the exception of articular cartilage, the collagen of hyaline cartilage, designated as collagen type II is not croos-banded and is arranged in an interlacing network of fine fibrils; this collagen cannot be demonstrated in this specimen.
Illustrate and indicate:
c) isogenous groups.
3. Extacellular matrix.
Specimen 2. Elastic cartilage (auricle).
Stained with orcein and haematoxylin.
The histological structure of elastic cartilage is similar to that of hyaline cartilage, its elasticity, however, being derived from the presence of numerous bundles of branching elastic fibers in the cartilage matrix; this network of elastic fibers, stained dark in this specimen, is particularly dense in the immediate vicinity of the chondrocytes. Collagen is also a major constituent of cartilage matrix and makes up the bulk of the perichondrium intermingled with a few elastic fibers.
Illustrate and indicate:
1. Isogenous groups of chondrocytes.
2. Extacellular matrix:
Specimen 3. Fibrocartilage (intervertebral disc).
Stained with haematoxylin and eosin.
Fibrocartilage consists of alternating layers of hyaline cartilage matrix and thick layers of dense collagen fibers oriented in the direction of the functional stresses. In this specimen, pink-stained collagen characteristically permeates the blue-stained cartilage ground substance. Chondrocytes are usually arranged in rows between the dense collagen layers within lacunae in the glycoprotein matrix.
Illustrate and indicate:
Specimen 4. Compact bone.
Stained with Shmorl method
In this ground section, the bone has been cut transversely thereby demonstrating Haversian systems amongst irregular interstitial systems. Concentric rings of flattened lacunae surround the Haversian canals and numerous fine canaliculi, barely visible at small magnification, interconnect lacunae with Haversian canals.
In the specimen focuses on a single Haversian system (large magnification), the central canal being surrounded by concentric lamellae of bone matrix containing empty lacunae. Fine canaliculi radiate from each lacunae to anastomose with those of adjacent lacunae. In life, osteocytes do not completely fill the lacunae, the remaining narrow space being filled with unmineralised matrix. Fine cytoplasmic processes of the osteocytes pass in the canaliculi to communicate via gap junctions with the processes of osteocytes in adjacent lamellae. The canaliculi provide passages for circulation of tissue fluid and diffusion of metabolites between the lacunae and vessels of the Haversian canals.
The outer surface of bone is invested by periosteum, which contains numerous osteoprogenitor cells and is richly supplied with blood vessels.
Illustrate and indicate:
1.External circumferential lamellae.
3. Internal circumferential lamellae.
IV. Central marrow canal.
Specimen 5. Intramembranous bone formation
Stained with haematoxylin and eosin
At a low magnification find the islets of woven bone stained homogeneously in the bright pink color. On the surface of the the each islet there are basophilic osteoblasts and oxiphilic large osteoclasts, and inside the bone – stellate osteocytes. Around bony plates mesenchymal cells and blood capillaries are visible.
Illustrate and indicate:
1. Inercellular substance.
5. Mesenchymal cells.
6. Blood vessels.
Specimen 6. Intracartilaginous bone formation
Stained with haematoxylin and eosin
Under the low magnification find the metaepiphyseal cartilage that divided into five zones, starting from the epiphyseal side of cartilage: 1. The resting zone consists of hyaline cartilage without morhologic changes in the cells. 2. The proliferative zone (columnar), chondrocytes divide rapidly and form columns of stacked cells parallel to the long axis of the bone. 3. The hypertrophic cartilage zone contains large chondrocytes whose cytoplasm has accumulated glycogen. The resorted matrix is reduced to thin septa between the chondrocytes. 4. Simultaneous with the death of chondrocytes in the calcified cartilage zone, the thin septa of cartilage matrix become calcified by the deposite of hydroxyapatite. 5. The ossification zone, endochondral bone tissue appears. Blood capillaries and osteoprogenitor cells formed by mitosis of cells originating from the periosteum invade the cavities left by the chondrocytes. The osteoprogenitor cells form osteoblasts, which are distributed in a discontinuous layer over the septa of calcified cartilage matrix. Ultimately, the osteoblasts deposit bone matrix over the three-dimensional calcified cartilage matrix.
Illustrate and indicate:
2. Periosteal bone.
3. Endochondral bone.
4. Osteogenic tissue.
6. Blood vessels.
7. Spicules of calcified cartilage.
1. Zone of reserve cartilage.
2. Columns of cartilage cells.
3. Hypertrophied cartilage cells.
4. Degenerating cartilage.
Specimen 7. Smooth muscle (muscle layer of the urinary bladder).
Stained with haematoxylin and eosin.
At a low magnification find in the muscular tunic of the urinary bladder the bundle of myocytes, which are separated by the loose connective tissue. At a high magnification watch smooth myocytes, which have spindle-like shape at the longitudinal section. Rod-shaped nuclei are disposed in the middle of the cells. Oxyphilic cytoplasm contains myofibrils, which are worse visible in light microscope. Smooth myocytes bodies and nuclei are round-shaped at a cross section.
Illustrate and indicate:
1. Smooth muscle cell:
a) longitudinal section,
b) transverse section.
4. Interfascicular connective tissue.
Specimen 8. Skeletal muscle of the tongue.
Stained with iron haematoxylin.
At a low magnification find the longitudinally and cross-sected bundles of muscular fibers and a layer of loose connective tissue between them. At a high magnification a lot of peripherally (under sarcolemma) disposed nuclei are observed in a cross sected muscular symplasts. Myofibrils lye in the central part of myofiber, they have cross striations, which consists of dark anisotropic and light isotropic discs. In the cross-sected myofibers the myofibrils look like a spots.
Illustrate and indicate:
1. Muscle fiber:
a) longitudinal section,
b) transverse section.
Specimen 9. Cardiac muscle (myocardium).
Stained with iron haematoxylin.
At a low magnification find the longitudinally disposed myofibers, which consist of contractile cardiomyocytes. There is loose connective tissue with vessels between them. At a high magnification pay your attention to the dark thin lines that are disposed cross to the fibers. They are intercalated discs – cardiomyocytes junctions. Watch carefully the structure of anastomoses, which combine fibers between themselves and cross striations.
Illustrate and indicate:
1. Muscle fiber.
2. Cardiac muscle cells.
3. Intercalated disc.
8. Connective tissue with blood vessels.
3. Stevens A. Human Histology / A. Stevens, J. Lowe. – [second edition]. –Mosby, 2000. – P. 65-76, 60-61, 227-250.
4. Wheter’s Functional Histology : A Text and Colour Atlas / [Young B., Lowe J., Stevens A., Heath J.]. – Elsevier Limited, 2006. – P. 101 – 122, 186 – 207.
5. Inderbir Singh Textbook of Human Histology with colour atlas / Inderbir Singh. – [fourth edition]. – Jaypee Brothers Medical Publishers (P) LTD, 2002. – P. 89-134.
6. Ross M. Histology : A Text and Atlas / M. Ross W.Pawlina. – [sixth edition]. – Lippincott Williams and Wilkins, 2011. – P. 198 – 218, 218 – 254, 310 – 352.
1. Eroschenko V.P. Atlas of Histology with functional correlations / Eroschenko V.P. [tenth edition]. – Lippincott Williams and Wilkins, 2008. – P. 267-277, 281.
Junqueira L. Basic
Histology / L. Junqueira, J. Carneiro, R. Kelley.
– [seventh edition]. –
Volkov K. S. Ultrastructure of cells
and tissues / K.
Created by Violetta Kulbitska