Histo - 1st Colloquium Annotated Slides PDF

For Colloquium 2 HISTOLOGY COLLOQUIUM II GROWTH PATTERNS Appositional growth (perichondrium): 1 - BONE AND CARTILAGE Chondrogenic cel...

For Colloquium 2 HISTOLOGY COLLOQUIUM II GROWTH PATTERNS Appositional growth (perichondrium): 1 - BONE AND CARTILAGE Chondrogenic cells ➞ chondroblasts ➞ chondrocytes Interstitial growth (isogenous groups): Both are specialized connective tissues, that resist mechanical Existing chondrocytes make more matrix stresses CHONDROCYTES CARTILAGE are specialized cells that produce and maintain the extracelular matrix Specialized avascular connective tissue; Chondrocytes are ovoid or rounded cells (10-30µm) Cells (chondroblasts and chondrocytes) and extracellular Large nucleus with prominent nucleolus; matrix ECM (composed of glycosaminoglycans and proteo- Rich network of rER, a well-developed Golgi, glycogen glycans, which are intimately associated with the collagen (II, inclusions. I) and elastic fibers embedded within the matrix); Each cell is located into small, individual compartments – lacunae; CARTILAGE CELLS Cartilage has cell nests (isogenous groups); Cartilage is covered with perichondrium (Perichondrium is not Chondrogenic cells: spindle-shaped cells, that are derived from found - on articular surfaces and around fibrocartilage). mesenchymal cells, ovoid nucleus, few mitochondria and small Cartilage supports but retains some flexibility. Golgi apparatus in cytoplasm. Chondroblasts: ovoid cells with basophilic cytoplasm - rich network of rER, a well-developed Golgi apparatus, numerous mitochondria, an abundance of secretory vesicles in cytoplasm. CARTILAGE ECM Collagen is the major matrix protein. Type II collagen forms the bulk of the fibrils, there are also type XI, X, IX. All those types are specific for catilage. Proteogycans – hyaluronic acid, chondroitin sulfate and keratan sulfate are joined to a core protein to form a PG HISTOGENESIS monomer. Each hyaluronic acid molecule is associated with ~80 PG monomers and form PG aggregates. Mesenchymal cells retract their processes; Congregates are called chondrification centers; Glycoproteins – tenascin, fibronectin. These cells differentiate into chondroblasts and begin secreting a matrix around themselves; HYALINE CARTILAGE The chondroblasts become entrapped in their own matrix in small individual compartments called lacunae. Hyalin cartilage matrix is highly hydrated to provide diffusion of small molecules. The matrix is amorphous, homogeneous in Chondroblasts surrounded by matrix are chondrocytes, they are still capable to cell division and production of ECM. This light microscope because the refractive index of the collagen process or type of growth is called interstitial growth. fibrils and the ground substance is nearly the same. The groups formed from one chondrocyte are known as isogenous groups. PERICHONDRIUM Outer fibrous layer – type I collagen, fibroblasts, blood vessels Inner cellular layer – chondrogenic (progenitor) cells Perichondria are present in elastic and most hyaline cartilages, but absent in fibrocartilage. Functions: Protection; Nourishment (cells receive nutrients from blood vessels in perichondrium by diffusion through the matrix); Cartilage growth (the chondrogenic cells undergo division and differentiate into chondroblasts, which begin to produce martix; in this way cartilage growths from periphery and this process is called appositional growth) 1/6 THREE TYPES OF CARTILAGE Variations in the amount and arrangement of extracellular matrix components give rise to the three types of cartilage Hyaline Cartilage Trachea and bronchi Nose Ribs Articulating surfaces of joints Growth plates Elastic Cartilage Pinna of ear Epiglottis Fibrocartilage Modified hyaline cartilage, covers articular surfaces of joints; Intervertebral disks, The free or articular surface has no perichondrium, but in Pubic symphysis, opposite surface is contact with bone; In articular disks; In adults it is 2 -5 mm thick Where tendons attach to bones HYALINE CARTILAGE The most common cartilage of the body; Perichondrium Abundance of type II collagen and aggrecans in ECM. The matrix of hyaline cartilage is composed of type II collagen, proteoglycans, glycoproteins, and extracellular fluid. Three types of cells are associated with cartilage: chondrogenic cells, chondroblasts, and chondrocytes. Prominent isogenous cell groups (8 – 16 cells) Matrix is subdivided into two regions: Territorial matrix - basophilic (poor in collagen and rich in chondroitin sulfate); Interterritorial matrix (more colagen type II and poorer in proteoglycans than the territorial matrix) Subdivide into zones: superficial (tangential) zone – type II collagen fibers that are arranged paralelly to the free surface transitional zone – round chondrocytes are distributed within the matrix radial zone – chondrocytes and collagen fibers are arranged into short columns perpendicular to the free surface; calcified zone – calcified matrix and few chondrocytes 2/6 ELASTIC CARTILAGE Perichondrium is rich in elastic fibers; BONE Abundance of elastic fibers in ECM; Elastic cartilage contains type II collagen and abundant elastic Bone is connective tissue characterized by a mineralized fibers scattered throughout its matrix, giving it more pliability. extracellular matrix The chondrocytes are more abundant and larger; Small isogenous cell groups (2-3 cells); Extracelular matrix: Matrix is not divided in teritorries. Organic: Colagen I Occasional lacunae display two chondrocytes, indicative of Non organic: hydroxiapatite interstitial growth. Periosteum: Fibrous layer Cellular layer Cells: Mesenchymal Osteoprogenitor Osteoblast Osteocyte Osteoclast - from monocyte OSTEOPROGENITOR CELL derived from embryonic mesenchymal cells and retain their ability to undergo mitosis. Located in the inner cellular layer of the periosteum, FIBROCARTILAGE endosteum, lining Haversian canals; Fibrocartilage is a tissue intermediate between dense regular Undergo mitotic division, have potentional to form osteoblasts connective tissue and hyaline cartilage. or chondroblasts; Perichondrium is absent; Spindle-shaped, oval nucleus, poorly developed organelles. Wide interterritories; More active during the period of growth and regeneration. Contains chondrocytes arranged in long rows separated by collagen fibers; Rich with type I and type II collagen (because it’s rich in OSTEOBLAST collagen type I, the fibrocartilage matrix is acidophilic). derived from osteoprogenitor cells; Fibrocartilage from intervertebral disc of the bull shows Responsible for the synthesis of the organic components of chondrocytes aligned in parallel rows, lying singly in individual the bone matrix (including type I collagen, proteoglycans, and lacunae. glycoproteins) and mineralization process. The cytoplasm of these cells is not evident. Also possess receptors for parathyroid hormone; The matrix contains thick bundles of collagen fibers. Osteoblasts are located on the surface of the bone in a sheet- The adjacent areas reveal hyaline cartilage of the opposing like arrangement of cuboidal to columnar cells; vertebrae, and dense connective tissue of the ligamentum Active cell – large nucleus with nucleolus, basophilic longitudinale posterior. cytoplasm (rER, Golgi); Processes form gap junctions with neighboring cells. 3/6  BONE MATRIX 50% dry weight is inorganic OSTEOCYTES The inorganic components of bone are crystals of calcium Osteocytes are mature bone cells that derived from osteoblasts hydroxyapatite, composed mostly of calcium and phosphorus. and are housed in lacunae within the calcified bony matrix. Bone is storage site for calcium and phoshate; Radiating out in all directions from the lacunae are narrow, Bone plays an important secondary role in the homeostatic tunnel-like spaces (canaliculi) that house cytoplasmic regulation of blood calcium levels; processes of the osteocyte. Mineralization is a cell-regulated event; Processes forming gap junctions between neighboring Alkaline phosphatase is important for mineralization. osteocytes. Nucleus is flattened; organic matrix Cytoplasm is poor in organelles; Extracellular matrix: fibers + ground substance: Secrete only substances for bone maintenance. Type I collagen (also small amount of type III and XIII) Collagen molecules constitute about 90% of the total weight of the bone matrix proteins; proteoglycans: core protein with various numbers of attached side chains of GAG – chondroitin sulfate, keratan sulfate; glycoproteins: osteonectin, osteopontin, sialoprotein – mediate cell attachment to ground substance and influence mineralization; bone-specific proteins: osteocalcin – binds calcium from blood and stimulates bone remodeling; growth factors and cytokines: small regulatory proteins – bone morphogenetic proteins (BMPs). PERIOSTEUM connective tissue layer – is subdivided into 3 layers: outer layer – connective tissue layer, nerves and blood vessels OSTEOCLASTS middle – rich with type I collagen fibers, what form also Derived from fusion of mononuclear hemopoetic progenitor cells Sharpey’s fibers, which penetrate into the bone (monocytes); inner cellular layer – osteoprogenitor (osteogenic) cells – have These cells are responsible for bone resorption; ability to divide and become osteoblasts Osteoclasts occupy Howship’s lacunae, that identify regions of Thus, from periosteum blood vessels enter into the bone canals bone resorption; and periosteum is responsible for bone growing of a thickness. Osteoclasts are large, motile, multinucleated cells 150 μm in diameter; they contain up to 50 nuclei and have an acidophilic cytoplasm. SHARPEY’S FIBRES Marked acidophilia of cytoplasm - mitochondria, lysosoms; Collagen fibres from tendons and ligaments continues in Osteoclasts resorb bone tissue by releasing protons and priosteum, and continuous as the collagen fibres of the bone lysosomal enzymes; matrix. Ruffled border involved in resorption of bone – finger like processes increasing surface for exocytosis of enzimes and endocytosis of bone particles; BONE TISSUE Primary or immature or woven bone The first bone to form during fetal development, Characterized by irregular bundles of collagen; No certain order for osteocytes; Remain at sutures of the calvaria, insertions sites of tendons, and alveoli of teeth. 4/6 Secondary or mature or lamellar bone OSTEON OR HAVERSIAN SYSTEM Composed of parallel (spongy bone) or concentric (compact Morphofunctional unit of compact bone; bone) lamellae; Surrounded by cementing line composed mostly of calcified Collagen fibers are arranged parallel each other within a given ground substance; lamella; Consist of concentric lamellae; Osteocytes in lacunae located between lamellae. Lamellae surrounding a Haversian canal, which contains the vascular and nerve supply of osteon; The collagen fibers are parallel in one lamellae but in different COMPACT AND SPONGY BONE direction in adjacent lamellae. SPONGY BONE Lamellae are arrange in trabeculae or spicules; red bone marrow between spicules. LAMELLAR SYSTEM OF COMPACT BONE Compact bone is composed of lamellae arranged in four lamellar systems: Inner circumferential lamellae, Outer circumferential lamellae, Osteons (Haversian lamella), Interstitial lamella. 5/6 VOLKMAN’S CANALS Perforating canals in lamellar bone through blood vesels and nerves travel from periosteum to Haversian canals; They also connect Haversian canals to one another; Volkmann’s canals are not surrounded by concentric lamallae. STRUCTURE OF LONG BONE Long bones have middle region - diaphysis and 2 expanded ends – epiphysis. The articular surface is covered with articular cartilage. The small zone between diaphysis and epiphysis is called metaphysis. Large central cavity filled with bone marrow is named marrow cavity. Almost all thickness of the middle bone region is compact. At the ends of the bone spongy bone is extensive. Outer surface is covered with periosteum, then follow outer circumferential lamellae and wide zone of osteons, inner circumferential lamellae, covered from marrow cavity side with endostium. CALCIUM Parathyroid hormone (PTH) acts on the bone to raise low blood calcium levels to normal; Calcitonin acts to lower elevated blood calcium levels to normal. 6/6 HISTOLOGY COLLOQUIUM II 2 - BONE GROWTH Bone Tissue grows ONLY by Appositional Growth Bone only grows appositionally Bone does not grow by interstitial growth Long bones need interstitial growth of cartilage to increase their length Bone as an organ grows two ways: Intramembranous Ossification When osteoblasts become trapped in their lacunae these are From Mesenchymal cells known as osteocytes. Example: flat bone of skull Embryonic connective tissue – mesenchyme is highly vascular, bony trabeculae are anastomosing, and appear to be Endochondral Ossification acidophilic due to the presence of collagen. Involves erosion of a cartilage model The large, multinuclear cells, osteoclasts, appear at the Osteoprogenitor cells, osteoblasts and osteocytes perform the surface of bone trabeculae in the process of bone resorption. same functions as the similar cells in intramembranous Within the bony trabeculae are osteocytes in their lacunae. ossification Osteoid may be seen on the margins of the bony trabeculae. INTRAMEMBRANOUS OSSIFICATION All roofing bones of the Skull: Frontal bone Parietal bones Occipital bone Temporal bones Mandible Clavicle Mesenchymal cells differentiate into osteoblasts – the bone- forming cells, which begin synthesis and secretion of ground substance and collagen (called osteoid) at multiple centres of ossification. Osteoid calcifies to form bone. Calcification quickly follows osteoid formation, and osteoblasts trapped in their matrices in lacunae become osteocytes. They are responsible for maintenance of the newly formed bone tissue. The processes of these osteocytes are also surrounded by forming bone, establishing a system of canaliculi. The remaining osteoblasts continue the bone deposition process at the bone surface. This newly bone appears as spicules or trabeculae. Continuous mitotic activity of mesenchymal cells provides a supply of undifferentiated osteoprogenitor cells, which form osteoblasts. The bone then undergoes progressive remodelling into lamellar bone by osteoclastic resorption and osteoblastic deposition to form mature compact or trabecular bone. The primitive mesenchyme remaining in the network of developing bone differentiates into bone marrow. Regions of the mesenchymal tissues that remain uncalcified differentiate into the periosteum and endosteum of developing bone. The spongy bone deep to the periosteum and the periosteal layer of the dura mater of flat bones are transformed into compact bone. 1/5 ENDOCHONDRAL OSSIFICATION The subperiosteal bone collar is formed of primary bone (intramembranous bone formation)!!! Developing bones are deposited as a hyaline cartilage model and then this cartilage is replaced by bone tissue. All bones of the body except: roofing bones of the skull, mandible, clavicle. CARTILAGE MODEL Miniature hyaline cartilage model formed in region of developing embryo where bone is to develop. Cartilage is covered with perichondrium. BONE COLLAR FORMED BY INTRAMEMBRANOUS OSSIFICATION In the middle of diaphysis perichondrium becomes richly vascularised; Vascularization of perichondrium changes it to periosteum; Chondrogenic cells become osteoprogenitor cells; From osteoprogenitor cells starts differentiation of osteoblasts; Osteoblasts secrete bone matrix And form bone collar (=perihondral bone spicule) on the surface of cartilage; Bone colar is surrounded by periosteum. 2/5 HYPERTROPHY OF CARTILAGE PRIMARY CENTRE OF OSSIFICATION The bone collar prevents the diffusion of nutrients to the The bone collar becomes thicker and grows in each direction chondrocytes; from the midriff of the diaphysis toward the epiphyses; Chondrocytes within the diaphysis core hypertrophy and die The cartilage of the diaphysis is replaced by bone; (apoptosis); Exept epiphyeal plates, which are responsible for the growth With the death of the chondrocytes much of the matrix breaks down, appear cavities; FORMATION OD SECONDARY CENTRE OF OSSIFICATION Rest of matrix become calcified. Secondary centers of ossification begin to form at the epiphysis; Process starts around birth. For larger bones before and for smaller after; Process begins in same way as at primary center, except that there is no bone collar; osteoblasts lay down bone matrix on calcified cartilage scaffold. EROSION OF CARTILAGE AND VASCULAR INVASION (PRIMARY OSSIFICATION CENTRE) Osteoclasts make holes in bone collar; Blood vessels growth through the bony collar and vascularize the cavity and forming primary center of ossification; BONE GROWTH IN LENGTH Osteoclasts continue erosion of calcified cartilage matrix; It depends on epiphyseal plate. Holes permit osteoprogenitor cells and blood vessels invade Chondrocytes proliferate and forming columns. cartilage model; Replacement by bone take place at the diaphyseal side. With blood vessels enter hemopoetic cells un stem cells for There are 5 zones in epiphyseal plate. forming reticular tissues. Both are necessary for forming red bone marrow. Interstitial Growth of Cartilage causes model to increase in length MIXED OR ENDOCHONDRAL SPICULES Cell nests form, then cells secrete matrix as they mature In primary ossification center still exist calcified cartilage septa; Bone matrix laid down on septa of calcified cartilage forms the complex – endochondral bone. Endochondral bone is cartilage/bone complex into primary bone marrow cavity. Osteoclasts begin resorbing the complex enlarging the morrow cavity. The bone matrix becomes calcified to form a calcified cartilage/calcified bone complex. This complex can be appreciated in routinely stained histological sections because calcified cartilage stains basophilic, whereas calcified bone stains acidophilic 3/5  GROWTH IN THICKNESS Bone can grow in thickness or diameter only by appositional growth. The steps in these process are: Periosteal cells differentiate into osteoblasts which secrete collagen fibers and organic molecules to form the matrix. Ridges fuse and the periosteum becomes the endosteum. New concentric lamellae are formed. Osetoblasts under the periosteum form new circumferential lamellae. EPIPHYSEAL PLATE Only by appositional growth at the bone’s surface Periosteal cells differentiate into osteoblasts and form bony ridges and then a tunnel around periosteal blood vessel. Concentric lamellae fill in the tunnel to form an osteon. Once the epiphyseal plate closes (disappears) the bone has reached it full length FACTORS AFFECTING BONE GROWTH Minerals Calcium - Makes bone matrix hard Phosphorus - Makes bone matrix hard Magnesium - Deficiency inhibits osteoblasts Vitamins Vitamin A - Controls activity, distribution, and coordination of osteoblasts/osteoclasts Vitamin B12 - May inhibit osteoblast activity Vitamin C - Helps maintain bone matrix, deficiency leads to decreased collagen production which inhibits bone growth and repair; (scurvy - disorder due to a lack of Vitamin C) Vitamin D (Calcitriol) - Helps build bone by increasing calcium absorption. Deficiencies result in “Rickets” in children Hormones Human Growth Hormone - Promotes general growth of all 1. Zone of resting cartilage - anchors growth plate to bone body tissue and normal growth in children 2. Zone of proliferating cartilage - rapid cell division (stacked Insulin-like Growth Factor - Stimulates uptake of amino acids coins) and protein synthesis 3. Zone of hypertrophic cartilage - cells enlarged & remain in Insulin - Promotes normal bone growth and maturity columns Thyroid Hormones - Promotes normal bone growth and 4. Zone of calcified cartilage - thin zone, hypertrophied maturity chondrocytes die, and cartilage matrix becomes calcified Estrogen and Testosterone - Increases osteogenesis at 5. Zone of ossification (erosion and vascular invasion) - puberty and are responsible for gender differences of osteoclasts removing matrix; osteoblasts & capillaries move skeletons in to create bone over calcified cartilage 4/5 BONE REMODELLING HEALING IN BONE Vitamin D Nutrition Physical activity Age, hormones PTH, PHRP IL1, TNF,TGF-β 5-10% bone / yea CALCIUM HOMEOSTASIS Repair of a fractured bone by formation of new bone tissue through periosteal and endosteal cell proliferation. 5/5 HISTOLOGY COLLOQUIUM II 3 - MUSCLE TISSUE ULTRASTRUCTURE OF SMOOTH MUSCLE CELLS STRIATED MUSCLE - regularly arranged contractile units; actin and myosin contractile filaments! Striated muscle cells display characteristic alternations of light intermediate filaments of desmin! (also vimentin in vascular and dark cross-bands, which are absent in smooth muscle smooth muscle) membrane associated and cytoplasmic dense bodies Skeletal Muscle - long, cylindrical multinucleated muscle fibers containing α actinin (similar to Z lines) with peripherally placed nuclei. Contraction is typically quick relatively active nucleus (smooth muscle cells make collagen, and vigorous and under voluntary control. Used for elastin, and proteoglycans) locomotion, mastication, and phonation. Each smooth muscle cell is surrounded by an external lamina Cardiac Muscle - elongated, branched cells with a single Between individual muscle cells and between fasciculi is a centrally placed nucleus and intercalated discs at the ends. network of supporting collagenous tissue; this is well Contraction is involuntary, vigorous, and rhythmic. demonstrated in micrograph in which the collagen is stained blue. SMOOTH MUSCLE - possesses contractile machinery, but it is irregularly arranged (thus, non-striated). Cells are fusiform with a central nucleus. Contraction is involuntary, slow, and long lasting. microtubules (curved arrows) actin filament (arrowheads) intermediate filaments dense bodies (desmin/vimentin plaques) caveoli (membrane invaginations & vesicular system contiguous with SER –functionally analogous to sarcoplasmic SMOOTH MUSCLE reticulum) Fusiform or spindle-shaped, non-striated cells Single, centrally-placed nucleus Smooth muscular cells are present in bundles Cell junctions – gap junctions Contraction is non-voluntary It is regulated by the autonomic nervous system, hormones (such as bradykinins), and local physiological conditions. Smooth muscle is found in the walls of hollow viscera (e.g., the gastrointestinal tract, some of the reproductive tract, and the urinary tract), walls of blood vessels, larger ducts of compound glands, respiratory passages, and small bundles within the dermis of skin. 1/6 SMOOTH MUSCLE CONTRACTION STRIATED MUSCLE also Ca+ dependent, but mechanism is different than striated muscle. each skeletal muscle fiber is long, cylindrical, multinucleated with peripherally placed nuclei, and striated. 1. Ca2+ ions released from caveloae/SER and complex with Contraction is typically quick and vigorous and under voluntary calmodulin control. 2. Ca2+-calmodulin activates myosin light chain kinase The highly developed functions of the cytoplasmic organelles 3. MLCK phosphorylates myosin light chain of muscle cells has led to the use of a special terminology for 4. Myosin unfolds & binds actin; ATP-dependent contraction some muscle cell components: plasma membrane or cycle ensues. plasmalemma = sarcolemma; cytoplasm = sarcoplasm; 5. Contraction continues as long as myosin is phosphorylated. endoplasmic reticulum = sarcoplasmic reticulum. 6. “Latch” state: myosin head attached to actin de- all muscle cells have basal laminae! phosphorylated causing decrease in ATPase activity – myosin head unable to detach from actin (similar to “rigor mortis” in skeletal muscle). Triggered by: Voltage-gated Ca+ channels activated by depolarization Mechanical stimuli Neural stimulation Ligand-gated Ca+ channels Epimysium - dense irr. CT Perimysium - less dense irr. CT Endomysium - basal lamina and reticular fibers The mechanism of smooth muscle contraction is as follows: Thin filaments of actin are associated with tropomyosin. Thick filaments composed of myosin only bind to actin if one chain is phosphorylated. Ca2+ ions in the cytosol of smooth muscle cells cause contraction as in striated muscle, but the control of Ca2+ ion movements is different. In relaxed smooth muscle, free Ca2+ ions are normally sequestered in sarcoplasmic reticulum throughout the cell. On membrane excitation, free Ca2+ ions are released into the cytoplasm and bind to a protein called calmodulin (a calcium- binding protein). The calcium-calmodulin complex then activates an enzyme called myosin light-chain kinase, which The entire muscle is surrounded by epimysium, a dense phosphorylates myosin and permits it to bind to actin. irregular collagenous connective tissue. Perimysium, a less dense collagenous connective tissue derived from epimysium, Actin and myosin subsequently interact by filament sliding to produce contraction in a similar way to that for skeletal muscle. surrounds bundles (fascicles) of muscle fibers. Endomysium, composed of reticular fibers and an external lamina (basal Contraction of smooth muscle can be modulated by surface receptors activating internal second messenger systems. lamina), surrounds each muscle fiber! Expression of different receptors allows smooth muscle in different sites to respond to several different hormones. Compared with skeletal muscle, smooth muscle is able to maintain a high force of contraction for very little ATP usage. 2/6  ORGANISATION OF SKELETAL MUSCLE FIBRES - THE SACROMERE Contractile unit of striated muscle: Structures between Z lines: 2 halves of I bands (Actin only; thin filaments only) A band (Actin and Myosin; thick & thin filaments) H zone (Myosin only; thick filaments only) M line (Mittelscheibe, Ger. “middle of the disc”) Myofilaments Actin Myosin Other structural proteins: Titin (myosin-associated) Multi-nucleated and striated Nebulin (actin-associated) A bands - anisotropic (birefringent in polarized light) Myomesin (at M line) I bands - isotropic (do not alter polarized light) α actinin (at Z line) Z lines (Zwischenscheiben, Ger. “between the discs”) Desmin (Z line) H zone (hell, Ger. “clear”) Vimentin (Z line) Dystrophin (cell membrane) The region of the myofibril between two successive Z disks, known as a sarcomere, is 2.5 μm in length and is considered the contractile unit of skeletal muscle fibers! The dark bands are known as A bands (anisotropic with polarized light) and the light bands as I bands (isotropic with polarized light). The center of each A band is occupied by a pale area, the H band, which is bisected by a thin M line. Each I band is bisected by a thin dark line, the Z disk (Z line). During muscle contraction, the various transverse bands behave characteristically. The I band becomes narrower, the H band is extinguished, and the Z disks move closer together (approaching the interface between the A and I bands), but the width of the A bands remains unaltered. I H M A 3/6 T-TUBULE SYSTEM Ca2+ Stimulates Myosin-Actin Binding and Initiates Contraction: Propagation of the Signal and Release of Ca2+ Myosin-actin binding inhibited by TnI TnC binds Ca2+ (if present) and induces release of TnI from T (transverse) Tubules actin run perpendicular (transversely) to myofibrils Myosin binds actin; hydrolysis of ATP induces power stroke conduct membrane depolarization deep into fibers Actin filaments move relative to myosin Sarcoplasmic Reticulum smooth ER site of Ca2+ storage & release terminal cisternae abut T-tubules forming triads when myofibrils are viewed in longitudinal section T tubules and sarcoplasmic reticulum are essential components involved in skeletal muscle contraction! CONTRACTION CYCLE 1. ATP binds myosin – myosin releases actin 2. ATP hydrolysis induces conformational change – myosin head cocks forward 5nm (ADP+Pi remain bound to myosin). 3. Myosin binds weakly to actin, causing release of Pi 4. Release of Pi induces strong binding, power stroke, and release of ADP 5. Myosin remains bound to actin if no more ATP is available NEUROMUSCULAR JUNCTION (rigor conformation) Synapse: Action potential (AP) stimulates release of acetylcholine from axon terminal into synaptic cleft Acetylcholine in synaptic cleft binds Na+ channel receptors – initiates sarcolemma AP Signal Propagation: T (transverse) Tubules Run perpendicular (transversely) to myofibrils Conduct membrane depolarization deep into fibers Intracellular Ca2+ release: Sarcoplasmic Reticulum Smooth ER, site of Ca2+ storage Voltage-gated channels in SR detect membrane depolarization in T-tubule and release Ca2+ SLIDING FILAMENT THEORY Muscle fibers are composed of many contractile units (sarcomeres) Changes in the amount of overlap between thick and thin filaments allows for contraction and relaxation of muscle fibers Many fibers contracting together result in gross movement Note: Z lines move closer together; I band and H band become smaller during contraction 4/6 TYPES OF FIBRES From the atrioventricular node, the impulse is passed along a specialised bundle of conducting fibres, the atrioventricular Type I fibres (red fibres) bundle (of His), which initially divides into right and left bundle Red muscles contain predominantly (but not exclusively) red branches, that then (halfway down the interventricular septum) muscle cells. Red muscle fibres are comparatively thin and become Purkinje fibres which run immediately beneath the contain large amounts of myoglobin and mitochondria. Red endocardium before penetrating the myocardium. fibres contain an isoform of myosin with low ATPase activity, i.e. the speed with which myosin is able to use up ATP. Contraction is therefore slow. Red muscles are used when PURKINJE FIBRES (CELLS) sustained production of force is necessary, e.g. in the control are larger than cardiomiocytes and have a pale staining central of posture. area with most of the red-staining myofibrils around the periphery of the cell. Type II fibres White muscle cells, which are predominantly found in white muscles, are thicker and contain less myoglobin. ATPase activity of the myosin isoform in white fibres is high, and contraction is fast. Type IIA fibres (red) contain many mitochondria and are available for both sustained activity and short-lasting, intense contractions. Type IIB/IIX fibres (white) contain only few mitochondria. They are recruited in the case of rapid accelerations and short lasting maximal contraction. Type IIB/IIX fibres rely on anaerobic glycolysis to generate the ATP needed for contraction. CARDIAC MUSCLE 1. Purkinje fibers, 2. endocardium, 3. myocardium Muscle cells of the atria are somewhat smaller than those of the ventricles. These cells also house granules (especially in the right atrium) containing atrial natriuretic peptide!, a substance that functions to lower blood pressure. This peptide acts by decreasing the capabilities of renal tubules to resorb (conserve) sodium and water. HEART - CONDUCTIVE SYSTEM The coordinated contraction of the heart is largely effected by a specialised conducting system of modified cardiac muscle fibres. The initial impulse originates spontaneously in the sino-atrial node situated in the right atrial wall near the entry of the superior vena cava, but the impulse rate is controlled by the autonomic nervous system. The impulse passes through the muscle of the atria, causing them to contract, and reaches the atrioventricular node in the medial wall of the right atrium. 5/6 MUSCLE REGENERATION AND GROWTH Skeletal Muscle Increase in size (hypertrophy) Increase in number (regeneration/proliferation) Satellite cells are proposed source of regenerative cells Smooth Muscle Increase in size (hypertrophy) Increase in number (regeneration/proliferation) Smooth muscle cells are proliferative (e.g. uterine myometrium and vascular smooth muscle) Vascular pericytes can also provide source of smooth muscle Heart Muscle Increase in size (hypertrophy) Formerly thought to be non-proliferative Post-infarction tissue remodeling by fibroblasts (fibrosis/ scarring) New evidence suggests mitotic cardiomyocytes and re- generation by blood or vascular-derived stem cells 6/6 HISTOLOGY COLLOQUIUM II 4 - BLOOD VESSELS CARDIOVASCULAR AND LYMPH-VASCULAR SYSTEM Transport systems that convey blood and lymph throughout the body Heart, blood vessels and lymphatic vessels Cardiovascular sytem: Systemic and pulmonary circulation fig. middle sized artery Walls of blood vessels are composed of three layers: the tunica intima, the tunica media, and the tunica adventitia. The innermost layer, the tunica intima, is composed of a single layer of flattened, squamous endothelial cells, which form a tube lining the lumen of the vessel, and the underlying subendothelial connective tissue. The intermediate layer, the tunica media, is composed mostly of smooth muscle cells oriented concentrically around the lumen. The outermost layer, the tunica adventitia, is composed mainly of fibroelastic connective tissue arranged longitudinally. The tunica intima houses in its outermost layer the internal CARDIOVASCULAR SYSTEM elastic lamina, a thin band of elastic fibers that is well Maintain blood flow to all organs in the body developed in medium-sized arteries. Adjust the flow to organs as needs change The outermost layer of the tunica media houses another band Provide enough flow and pressure for capillary exchange to of elastic fibers, the external elastic lamina, although it is not occur distinguishable in all arteries. Have enough blood pressure after capillary beds for blood to The deeper cells of the tunica media and tunica adventitia are return to the heart nourished by the vasa vasorum. TUNICA INTIMA The endothelial cells (simple squamous epithelium) lining the lumen of the blood vessel rest on a basal lamina. Endothelial cells not only provide an exceptionally smooth surface but also function in secreting types II, IV, and V collagens, lamin, endothelin, nitric oxide, and von Willebrand factor. A subendothelial layer lies immediately beneath the endothelial cells. It is composed of loose connective tissue and 1/5  a few scattered smooth muscle cells, both arranged longitudinally. Beneath the subendothelial layer is an internal elastic lamina that is especially well developed in muscular arteries. Separating the tunica intima from the tunica media, the internal elastic lamina is composed of elastin, which is a !fenestrated! sheet that permits the diffusion of substances into the deeper regions of the arterial wall to nourish the cells there. TUNICA MEDIA The tunica media is the thickest layer of the blood vessel. The concentric cell layers forming the tunica media comprise mostly helically arranged smooth muscle cells. Interspersed within the layers of smooth muscle are some elastic fibers, type III collagen, and proteoglycans. The fibrous elements form lamellae within the ground substance secreted by smooth muscle cells. MUSCULAR ARTERY Regulate the flow of blood to various organs by contraction of Larger muscular arteries have an external elastic lamina, which is more delicate than the internal elastic lamina and smooth muscle, which constricts vessel separates the tunica media from the overlying tunica adventitia. T.intima: Endothelium, basal lamina, subendothelial layer, thick internal elastic lamina Capillaries and postcapillary venules do not have a tunica media; in these small vessels, pericytes replace the tunica T.media: Up to 40 layers of smooth muscle cells; thick external media elastic lamina T.adventitia:Thin layer of fibroelastic connective tissue; vasa TUNICA ADVENTITIA vasorum not very prominent; lymphatic vessels, nerve fibers Covering the vessels on their outside surface is the tunica adventitia, composed mostly of fibroblasts, type I collagen Muscular arteries frequently have both internal and external fibers, and longitudinally oriented elastic fibers. Contain vasa elastic laminae. vasorum, nerve fibers. This layer becomes continuous with the connective tissue elements surrounding the vessel. ARTERIOLE Arteries with a diameter of less than 0.1 mm are considered to be arterioles. ELASTIC ARTERY T.intima: Endothelium; basal lamina, subendothelial layer not OR CONDUCTING ARTERY very prominent; some elastic fibers instead of a defined internal elastic lamina Tunica intima: Endothelium, basal lamina, subendothelial T.media: One or two layers of smooth muscle cells layer, incomplete internal elastic lamina T.adventitia: Loose connective tissue, nerve fibers Tunica media: 40 to 70 fenestrated elastic membranes; smooth muscle cells interspersed between elastic membranes; thin external elastic lamina; vasa vasorum in outer half Tunica adventitia: Thin layer of fibroelastic connective tissue, vasa vasorum, lymphatic vessels, nerve fibers Examples: Aorta, pulmonary, brachiocephalic, subclavian, common carotid, common iliac arteries arteries Elastic arteries have many elastic laminae and thus laminae are not referred to as “internal” or “external”. Thick muscular walls relative to lumen size By definition: 1-3 layers of smooth muscle Regulate flow to the capillary beds & reduces pressure 2/5 COMPANION ARTERIOLE AND VENULE Smallest venules: Postcapillary (Pericytic)venules Very thin walls Pericytes Very permeable Easiest to identify when comparing venule with companion arteriole Site of exchange of cells and molecules between blood and tissues CAPILLARY capillaries composed of a single layer of endothelial cells, are the smallest blood vessels. Pericytes are located along the outside of the capillaries and small venules, and appear to be surrounding them. exvchange vessels 4-10 μm diameter STRUCTURES OF CAPILLARIES REFLECT THEIR FUNCTION Continuous capillaries have no pores or fenestrae in their walls! Endothelial cells connected by tight junctions and not very leaky Numerous pinocytotic vesicles for transport across endothelium Fenestrated capillaries possess pores (fenestrae) in their walls that are covered by pore diaphragms. Fenestrations (small pores) within cells that facilitate exchange across them Found in: endocrines, kidney and mucosa of GI Most fenestrations have diaphragms except in the kidney 3/5 Discontinuous sinusoidal capillaries (or Sinusoids) Sinusoidal capillaries may possess discontinuous endothelial cells and basal lamina and contain many large fenestrae without diaphragms HUGE transcellular pores (0.5-3 μm) Fenestrations (50-80 nm) Specialized cells (liver) Gaps between endothelial cells +/- Basal lamina Larger diameter lumen VEIN are classified into three groups on the basis of their diameter and wall thickness: small (10mm) Have large lumen relative to thickness of the wall; Do not have ext&int elastic laminae Little muscle in the tunica media (thin tunica media) compared to their companion arteries Tunica adventitia is most prominent layer Have valves to prevent backflow Wall structure is more variable than in arteries cardiac muscle in t. adventitia (vena cava and pulmonary veins) Irregular shaped lumen Thin tunica media LYMPATHIC VESSELS Lymphatic vessels drain excess fluid from the tissues. They begin as blind lymphatic capillaries which take up excess tissue fluid. Lymph vessels contain no RBCs, but have some lymphocytes. Lymph flows through large and larger collecting vessels, with histology resembling that of venules and veins (with valves). Occasional lymph nodes are interposed in the lymphatic vessel pathway, so the lymph flows through them (macrophages monitor the lymph, lymphocytes may engage in immune activities). Lymph reaches the thoracic duct and right lymphatic duct, both of which empty lymph into veins at the base of the neck, thus restoring the fluid and any content (proteins, etc.) to the blood. Location Most of the body Wall structure of veins is more variable than in arteries Not present in CNS and a few other places (bone) 4/5 Functions Return both tissue fluid and escaped plasma proteins to the blood (from interstitial tissue spaces) Circulate antibodies and return lymphocytes to blood Open circulatory system Lymph flows unidirectionally from peripheral tissue to heart. Begin as blind-ending capillaries that converge larger lymphatic vessels 2 lymphatic ducts (thoracic duct, right lymphatic duct) venous circulation at junction of internal jugular and subclavian veins Lymph is filtered in lymph nodes Lymphatic capillaries Thin endothelium NO tight junctions Incomplete basal lamina Lacteal is a lymphatic capillary in intestine Large lymph vessels resemble veins many valves 5/5 HISTOLOGY COLLOQUIUM II MAJOR PLASMA PROTEINS Albumin- Maintain colloid osmotic pressure; transport insoluble metabolites 5 - BLOOD AND BONE MARROW Globulins (α and β)- Transport metal ions, protein-bound lipids, lipid-soluble vitamins Globulins (γ) - Antibodies of immune defense Complement proteins - Destruction of microorganisms and BLOOD initiation of inflammation Clotting proteins - Formation of blood clots Blood is a bright to dark red, viscous, slightly alkaline fluid (pH, Plasma lipoproteins - Transport of triglycerides and cholesterol 7.4) that accounts for approximately 7-8% of the total body to/from liver weight. The total volume of blood of an average adult is about 5 L, and FORMED ELEMENTS it circulates throughout the body within the confines of the Erythrocytes (red blood cells, RBC) circulatory system. Platelets (thrombocytes) Blood is a specialized connective tissue composed of formed Leukocytes (white blood cells, WBC) elements - red blood cells (RBCs; erythrocytes), white blood Granulocytes (with specific granules) cells (WBCs; leukocytes), and platelets - suspended in a fluid Neutrophil component (the extracellular matrix), known as plasma. Eosinophil Basophil BLOD FUNCTIONS Agranulocytes (without specific granules) 1. transport of nutrients, oxygen, wastes, and carbon dioxide to Lymphocyte (B-cell, T-cell) and from the tissues Monocyte 2. convey hormones, cytokines, chemokines, and other soluble regulatory molecules 3. transport of leukocytes and antibodies through the tissues BLOOD SMEAR 4. maintain homeostasis Light microscopic examination of circulating blood cells is performed by evenly smearing a drop of blood on a glass Blood performs many important functions within the body slide, air-drying the preparation, and staining it with mixtures of including: dyes specifically designed to demonstrate distinctive characteristics of the cells. Supply of oxygen to tissues (bound to hemoglobin, which is The current methods are derived from the technique carried in red cells) developed in the late 19th century by Romanovsky, who used Supply of nutrients such as glucose, amino acids, and fatty a mixture of methylene blue and eosin. acids (dissolved in the blood or bound to plasma proteins (e.g., Most laboratories now use either the Wright or Giemsa blood lipids)) modifications of the original procedure, and identification of Removal of waste such as carbon dioxide, urea, and lactic blood cells is based on the colors produced by these stains. acid Methylene blue stains acidic cellular components blue, and Immunological functions, including circulation of white blood eosin stains alkaline components pink! cells, and detection of foreign material by antibodies Coagulation, which is one part of the body's self-repair mechanism (blood clotting after an open wound in order to stop bleeding) Messenger functions, including the transport of hormones and the signaling of tissue damage Regulation of body pH Regulation of core body temperature Hydraulic functions 1/7 ERYTHROCYTES Rh BLOOD GROUP Another important blood group, the Rh group, is so-named Life span in blood: About 120 days. because it was first identified in rhesus monkeys. This complex group comprises more than two dozen antigens, Size and shape: although many are relatively rare. Three of the Rh antigens (C, biconcave disk, 7.5-7.8 µm diameter, 2μm at thickest D, and E) are so common in the human population that the point, 1 μm at thinnest erythrocytes of 85% of peaple have one of these antigens on shape maintained by a cytoskeletal complex inside the their surface, and these individuals are thus said to be Rh- plasma membrane (involving spectrin, actin and other positive (Rh+). components) Individuals lacking these antigens are Rh-negative (Rh-). flexible: RBC’s normally bend to pass through small ~15% capillaries LM appearance in smear: Pink circle with light center (center is thinner because of the biconcave shape). No nucleus! During differentiation in the bone marrow, large quantities of the iron-containing respiratory pigment haemoglobin are synthesised. Before release into the blood circulation, the erythrocyte nucleus is extruded and, by maturity, all cytoplasmic organelles degenerate. The fully differentiated erythrocyte therefore simply consists of an outer plasma membrane enclosing haemoglobin and the limited number of enzymes necessary for maintenance of the cell. Function: Transport of oxygen and carbon dioxide bound to haemoglobin (oxyhemoglobin and carboxyhemoglobin) majority of CO2 transported as HCO3- pH homeostasis PLATELETS carbonic anhydrase: CO2 + H2O → HCO3- + H+ Life Span: about 10 days band 3 membrane protein: exchanges HCO3- for Shape, size, and origin: Small, biconvex disks, 2-3 µm in extracellular Cl- diameter. Non-nucleated cell fragments derived from cytoplasm of a very large cell, the megakaryocyte, in bone marrow! Platelets have a life span of about 10 days. LM appearance in smears: Small basophilic fragments, often appearing in clusters. TEM appearance: The platelet is bounded by a plasma membrane, and has a bundle of microtubules around the margin of the disk (which maintains the disk shape). There are three types of granules, containing fibrinogen, plasminogen, thromboplastin and other factors for clotting. There are also membrane tubules and glycogen. Function: Platelets initiate blood clots. may be divided into four zones ABO BLOOD GROUPS The extracellular surface of the red blood cell plasmalemma has specific inherited carbohydrate chains that act as antigens and determine the blood group of an individual for the purposes of blood transfusion. The most notable of these are the A and B antigens, which determine the four primary blood groups, A, B, AB, and O. People who lack either the A or B antigen, or both, have antibodies against the missing antigen in their blood; if they undergo transfusion with blood containing the missing antigen, the donor erythrocytes are attacked by the recipient's serum antibodies and are eventually lysed. 2/7  Specific granules contain various enzymes and pharmacological agents that aid the neutrophil in performing its antimicrobial functions. In electron micrographs these granules appear somewhat oblong. Azurophilic granules, as already indicated, are lysosomes, containing acid hydrolases, myeloperoxidase, the antibacterial agent lysozyme, bactericidal permeability-increasing (BPI) protein, cathepsin G, elastase, and nonspecific collagenase. Tertiary granules contain gelatinase and cathepsins as well as glycoproteins that are inserted into the plasmalemma. EXTRAVASATION VIA DIAPEDES Selectin-selectin receptor interaction causes neutrophil to slow When a blood vessel wall is damaged, factors from the & roll along surface. damaged endothelial cells and the ECM induce the clotting Chemokines from endothelium leads to expression of integrins cascade. Platelets aggregate and release proteins for clot & immunoglobulin family adhesion molecules on neutrophil cell formation and resolution: membrane. Neutrophil firmly attached to vessel wall & extends pseudopod 1. Vasoconstriction –via release of serotonin into vessel wall. 2. Further platelet aggregation –mediated via thromboxane A2 Vascular permeability mediated by heparin & histamines and ADP released by mast cells/basophils. 3. Fibrin polymerization –initiated by thromboplastin and free Once in connective tissue, neutrophils respond to Ca++ chemoattractants & migrate to injury site. 4. Clot contraction – via actin, myosin, and ATP released into the matrix of the clot 5. Clot resolution –platelet plasminogen activator (pPA, converts plasminogen into active fibrinolytic plasmin) 6. Tissue repair –platelet derived growth factor (PDGF, stimulates smooth muscle and fibroblast proliferation) NEUTROPHIL Life Span: < 1 week Granulocyte with specific and non-specific granules Specific granules NEUTROPHIL ANTIBACTERIAL ACTIVITY Type IV collagenase (aids migration) Lactoferrin (sequesters iron) Chemotaxis and migration (chemokine synthesis and matrix proteolysis) Phospholipase A2 (leukotriene synthesis) Lysozyme (digests bacterial cell wall) Phagocytosis and bacterial destruction Digestion via lysozymes Non-specific granules (lysosomes) Production of reactive oxygen compounds (respiratory burst) Lysozyme Acid hydrolase Iron sequestration via lactoferrin Myeloperoxidase Release factors to increase inflammatory response (and increase neutrophil production) Elastase LM appearance in smear: About 9-12 µm in diameter (thus larger than RBC). Nucleus long and multi-lobed (usually 2-4 lobes). Cytoplasm has small, neutrally stained specific granules. Non- specific granules are azurophilic. TEM appearance: Multi-lobed nucleus and numerous specific granules and lysosomes (=azurophilic granules in LM). Function: Primarily antibacterial Neutrophils leave the blood and follow chemotaxic signals to sites of wounding or other inflammation, and phagocytose foreign agents such as bacteria. Pus is composed largely of dead neutrophils. Three types of granules are present in the cytoplasm of neutrophils: 1. Small, specific granules (0.1 μm in diameter) 2. Larger azurophilic granules (0.5 μm in diameter) 3. Tertiary granules. 3/7 EOSINOPHIL BASOPHIL Life Span: < 2 weeks Life Span: 1-2 years (?) Granulocyte with specific and non-specific granules Granulocyte with specific and non-specific granules Specific granules Specific granules Major basic protein Histamine Eosinophilic cationic protein Heparin Neurotoxin Eosinophil chemotactic factor Histaminase Phospholipids for synthesis of leukotrienes, e.g. slow- reacting substance of anaphylaxis ( SRS-A ) Non-specific granules (lysosomes) Lysozyme Non-specific granules (lysosomes) Acid hydrolase Lysozyme Myeloperoxidase Acid hydrolase Elastase Myeloperoxidase Elastase LM appearance in smear: About 10-14 µm in diameter. Bilobed nucleus. The cytoplasm has prominent pink/red LM appearance in smear: About 9-11 µm in diameter. The (acidophilic) specific granules (stained with eosin dye). cytoplasm contains large, basophilic purple/black specific Basophilic cytoplasm (stained with m.blue). granules (stained with the basic dye). Acidophilic cytoplasm (stained with eosin). The nucleus is usually bilobed, but usually TEM appearance: The specific granules are ovoid in shape, is partially obscured by granules, which can lie over it. and contain a dark crystalloid body composed of major basic protein (MBP), effective against parasites. The rest of the TEM appearance: The specific granules vary in size and granule contains other anti-parasitic substances. The shape, and have occasional myelin figures (usually formed cytoplasm also contains lysosomes (=azurophilic granules). from phospholipids). The cytoplasm also has some lysosomes (=azurophilic granules). Function: Anti-parasitic activity Function: Allergies and anaphylaxis (hypersensitivity reaction) Mediators of inflammatory/allergic responses in tissues Binding of antigens to membrane-bound IgE antibodies Inactivate leukotrienes and histamine secreted by induces degranulation of specific granules, which leads to basophils allergic reaction. Engulf and sequester antigen-antibody complexes In hypersensitivity reaction, widespread vasodilation Inflammatory stimulus increases production/release (arteriolar) and vessel leakiness induce circulatory shock. of eosinophils from bone marrow, whereas Bronchial spasms cause respiratory insufficiency; inflammatory suppression decreases eosinophil combined effect is anaphylactic shock. numbers in peripheral blood. But, they also secrete PRO-inflammatory Similarity to tissue mast cells: Tissue mast cells also have IgE chemokines AND they can degranulate receptors and similar (though not identical) granule content. inappropriately to cause tissue damage (as in Mast cells and basophils have a common precursor in bone reactive airway disease) marrow. 4/7 LYMPHOCYTE MONOCYTE Life Span: variable (few days to several years) Life Span: few days in blood, several months in connective tissue LM appearance in smear: Small lymphocyte (about 90% of lymphocytes you will see) are ~6-7 µm in diameter, while large LM appearance in smears: About 12-15 µm in smears, thus lymphocytes may be up to about 15 µm. Round, dense the largest leukocyte. Large, eccentric nucleus either oval, nucleus (abundant heterochromatin). The basophilic kidney-shaped or horseshoe-shaped, with delicate chromatin cytoplasm of a small lymphocyte is a narrow rim around the that is less dense than that of lymphocytes. Pale basophilic nucleus, and when well stained is pale blue. T-lymphocytes cytoplasm, often bluish gray, may contain occasional stained and B-lymphocytes cannot be distinguished in a smear. granules (lysosomes = azurophilic granules). Large lymphocytes may resemble monocytes, but the lymphocyte TEM appearance: The cytoplasm doesn't appear to be very nucleus is usually more dense. active, containing mainly mitochondria and free ribosomes. TEM appearance: Cytoplasm contains mitochondria and Function: Cellular and humoral immunity. In general: some small lysosomes. B-lymphocytes (B-cells): may differentiate into tissue plasma cells which make antibodies. Some B-cells Function become memory cells. Migrate into tissues and constitute mononuclear T-lymphocytes (T-cells): cytotoxic T cells and helper T phagocyte system that help destroy foreign bodies and cells. maintain or remodel tissues Tissue macrophages Small 6-7; medium 7-9; large 9-11 (up to 15) µm Kupfer cells (liver) Osteoclasts (bone) Dust cells (lungs) Microglia (brain) Mediate inflammatory response Antigen presenting cells: Dendritic Cells, Langerhans cells fig. small lymphocyte fig. large lymphocyte 5/7 BLOOD CELL DEVELOPMENT (hematopoiesis = hemopoiesis) Normally occurs in red bone marrow in adult (also spleen & liver, if necessary) Phases: mesoblastic (yolk sac, 2 wks)* → hepatic (6 wks)* → splenic (12 wks) → myeloid (marrow, 24 wks) * Erythrocytes still have nuclei; leukocytes do not appear until 8 wks Mitotic stem and progenitor cells undergo increasing lineage restriction to produce committed precursors. Precursors undergo cell division and differentiation into mature cells. Maturation involves (note exceptions for megakaryocytes below): decrease in cell size** shutting down transcription (nucleoli disappear and chromatin condenses)** adoption of morphological characteristics specific to that lineage. Future granulocytes produce specific and non-specific granules, and then shape their nucleus. Future monocytes produce non-specific granules and shape their nucleus. Future small lymphocytes decrease their size and enter the blood, but then undergo extensive further maturation at another site (T-cells in the thymus, and B-cells in the "bursa equivalent"). Future erythrocytes fill cytoplasm with hemoglobin, synthesized on free polysomes (ribosomes on mRNA), and eventually extrude their nucleus. RETICULOCYTES are slightly-immature red blood cells, in which the last remnants Mature cells enter marrow sinus; immature cells in peripheral of cytoplasmic ribosomes, mitochondria and other organelles blood typically indicates disease. still persist in what is otherwise a concentrated solution of hemoglobin. **Megakaryocytes develop into large polyploid cells that Reticulocytes are released into the blood, and become mature remain transcriptionally active and extrude platelets as erythrocytes in about 24 hours. cytoplasmic fragments directly into marrow sinus. A count of reticulocytes in a blood smear is a measure of new RBC production. It is often difficult to distinguish reticulocytes from mature erythrocytes in ordinary blood smears. However, the blood that was to be used for this smear was treated with a basic dye, brilliant cresyl blue, which stains the RNA blue in clumps of ribosomes, and then the smear was made. The staining pattern in the cells looks a little bit like a net, giving the cell its name (Latin, reticulum=fishing net; Greek, cyte=cell). 6/7 DIAGRAM OF BONE MARROW Marrow sinuses are sinusoidal, discontinuous capillaries. Mature cells enter the sinuses and are conveyed to the systemic circulation via nutrient veins. MEGAKARYOCYTES in bone marrow produce blood platelets LM appearance: A huge cell, up to 50 µm in diameter. Its long nucleus has several lobes (the nucleus is polyploid and can be up to 64N). The cytoplasm is pale pink/red, without visible granules. In bone marrow, megakaryocytes are situated adjacent to a marrow sinus (large capillary), although this may not be obvious in tissue sections. TEM appearance: Particularly striking in the cytoplasm are many curved white lines that are the platelet demarcation channels, membrane-bound spaces forming the boundaries between future platelets. The cytoplasm also contains granules of various sizes, that will be in the platelets. Function: Megakaryocytes produce blood platelets by fragmentation of their cytoplasm, extending cell processes through the endothelium of a marrow sinus, and releasing clusters of immature platelets into the blood, to become mature platelets. 7/7 HISTOLOGY COLLOQUIUM II PRIMARY LYMPHOID ORGANS The bone marrow and the thymus and the Gut-Associated 6 - LYMPHATIC SYSTEM Lymphoid Tissue (e.g. appendix, terminal ileum) are the initial “education centers” of the immune system In these organs, lymphocytes (T cells in the thymus, B cells in Lymphatic System consists of: bone marrow and gut) differentiate into immunocompetent cells (i.e. they can recognize “self” vs. “nonself”). This differentiation is Cells said to be antigen-independent. Lymphocytes (B,T, natural killer) Antigen-presenting cells (dendritic cells, Langerhans’ The lymphocytes then enter the blood and lymph to populate: cells & macrophages) epidermis and mucosae Lymphatic “tissue” – diffuse and nodular connective tissue Lymphatic “organs” (lymph nodes, spleen, thymus) secondary lymphoid organs Lymphatic vessels that carry the cells and fluid SECONDARY LYMPHOID ORGANS FUNCTIONS Monitor body surfaces and fluid compartments (e.g. The lymph nodes, lymphatic nodules, tonsils, spleen are the epidermis, mucosae, interstitium) secondary “education centers” of the immune system React to the presence of potentially harmful antigens recognized as “non-self” In these organs, immunocompetent lymphocytes differentiate Autoimmune diseases (rheumatoid arthritis, type I diabetes, into immune effector and memory cells that undergo antigen- etc.) dependent activation and proliferation in these organs. Lymphoid organs are classified as: These lymphocytes then carry out their functions in the: Primary lymphoid organs: connective tissue secondary lymphoid organs Thymus mucosal surfaces lining epithelia Bone marrow They participate in: Lymphatic nodules of the distal intestinal tract (e.g. ileum and appendix) Cell mediated immunity (mostly “cytotoxic” T cells) Humoral responses (production of antibody) (B cells, also requires “helper” T cells) Secondary (effector) lymphoid organs/tissue: Spleen & lymph nodes (organs) The thymus and bone marrow, where immature lymphocytes Mucosal associated lymphoid tissue (MALT), e.g. lymphocytes acquire the receptors to recognise antigen, are known as and lymphatic nodules in the lamina propria primary lymphoid tissues. The spleen, lymph nodes and organised lymphoid tissues of MALT where lymphocytes are activated in response to antigen are the secondary lymphoid tissues. MAJOR LYMPHOID ORGANS The thymus is the site of maturation of immature T lymphocytes. The bone marrow is not only the home of lymphocyte stem cells but is also the site of B lymphocyte maturation. The lymph nodes are the sites where both T and B lymphocytes may interact with antigen and antigen presenting cells from the circulating lymph and undergo activation and cell division. The spleen is the location where T and B lymphocytes may interact with blood-borne antigen and undergo stimulation and cell division. ROLE OF T AND B Ly Both T and B cells are derived from stem cells in the bone marrow. Immature T lymphocytes migrate from the bone marrow to the thymus where they develop into mature T lymphocytes. Mature T cells then populate the secondary lymphoid tissues and from there continuously recirculate via the bloodstream in the quest for antigen. B lymphocytes are derived from precursors in the bone marrow and also mature there. Stimulated B cells mature into plasma cells that synthesise large amounts of antibody (immunoglobulin gamma). 1/8 THYMUS Each lobule is composed o