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Connective Tissue Course : Microanatomy (SCAN 213) For : Pre-medical students II Time : 2 h Date : October 31,2001 Lecturer: Dr. Prapee Sretarugsa Department of Anatomy Faculty Of Science Mahidol University Tel : 201- 5424 E-mail : [email protected] Dr. Prapee 1 Objectives : After you have studied this lecture, you should be able to • Explain the components of connective tissue • Describe the structures and functions of all types of cells in connective tissue • Identify all fibers in connective tissue as well as their localization in the organs • Explain the composition of ground substance as well as its function • Classify the type of connective tissue Dr. Prapee 2 Connective Tissue (CT) • Derives from mesoderm • Mesenchyme pleuripotency) • Mesenchymal cells can differentiate into - connective tissue - bone, cartilage - blood and hemopoietic cells - lymphoid cells Dr. Prapee 3 Characteristic of Connective Tissue • Consists of a large amount of extracellular matrix with a limited number of cells Dr. Prapee 4 Extracellular Matrix (ECM) • A complex of nonliving macromolecules synthesized by the cells • Some tissue consists of small amount of ECM • Connective tissue contains a large amount of ECM with a limited number of cells • A major component of connective tissue proper • Comprises of - Ground substance – resists forces of compression - Fibers – provides tensile force Dr. Prapee 5 Ground Substance Consists of • Glycosaminoglycans (GAGs) • Proteoglycans • Adhesive glycoproteins Dr. Prapee 6 Properties of GAGs • Long, inflexible, unbranched polysaccharides • High negative charge • Repeating disaccharide units • One of repeating unit is amino sugar which is sulfated (SO-3) • Second sugar is uronic acid with a carboxyl gr. (COO-) • Retension of positive ions (eg. Na+) together with water, maintaining tissue architecture turgor which tend to prevent deformation by compressive force Dr. Prapee 7 Structure of GAGs - Disaccharide units form covalent linkage with protein to form proteoglycans - Proteoglycans attach to hyaluronic acid - Hyaluronic acid, nonsulfated GAG Dr. Prapee 8 Types of GAGs Dr. Prapee 9 Struture ofProteoglycan •Macromolecues formed by sulfated GAGs form covalent bonds with a protein core • Structures look like a bottle brush • linked to •Each proteoglycan is covalently hyaluronic acid forming huge macromolecules, Aggrecan aggregates Dr. Prapee 10 Proteoglycans •Macromolecues formed by sulfated GAGs form covalent bonds with a protein core • Structures look like a bottle brush • Each proteoglycan is covalently linked to hyaluronic acid forming huge macromolecules, Aggrecan aggregates Dr. Prapee 11 Functions of Proteoglycans • Resist compression and retard the rapid movement of microorgani and metastatic cells • Form molecular filter (in association with basal lamina) of varying pore sizes and charge distributions that selectively screen and retard macromolecules as they pass through them • Contain binding sites of certain signaling molecules eg. TGF-ß Dr. Prapee 12 Functions of Proteoglycans and Hyaluronan during development • Epithelial branching and differentiation • Eye development • Limb development Dr. Prapee 13 Adhesive Glycoproteins Large macromolecules have several domains - at least one domain usually bind to integrins - one to collagen fibers - one to proteoglycans Dr. Prapee 14 The Major Types of Adhesive Glycoproteins • • • • • • Dr. Prapee Fibronectin Laminin Entactin Tenascin Chrondronectin Osteonectin 15 The Binding Sites of Adhesive Glycoproteins Fibronectin - collagen, heparin, heparan sulfate, hyaluronic acid, integrin Laminin - stricly limited to the basal lamina - heparan sulfate, type IV collagen, entactin & cell membrane Entactin - laminin, type IV collagen Tenascin - integrin, proteoglycan & fibronectin - distribution is limited to embryonic tissue Dr. Prapee 16 The Binding Sites of Adhesive Glycoproteins Chrondonectin - type II collagen, chrondroitin sulfate, hyaluronic acid, & integrins of chondroblast and chrondocyte Osteonectin - type I collagen, integrins of osteoblast and osteocyte Dr. Prapee 17 Integrin Dr. Prapee 18 Fibers • Provide tensile strength and elasticity to ECM • Can be classified into 3 types based on their morphology and reactivity with dyes - Collagen - Reticular - Elastic Dr. Prapee 19 Collagen Fibers • Collagen, constitue about 20% of all the body proteins • Form a flexible fiber whose tensile strength is greater than that of stainless steel of comparable diameter • White fiber, colorless • No specific staining for collagen fiber • Acidophilic fiber • EM showing cross-banding at regular interval of 67Dr.nm Prapee 20 Components of collagen fibers • Smaller subunits, tropocollagen •Formed from parallel aggregates the thinner fibril 10 to 300 nm in diameter Dr. Prapee 21 Tropocollagen • 280 nm long • 1.5 nm in diameter • Three polypeptide chains, a-chains - formed triple helical configuration - each chain contains about 1,000 aa - every third aa is glycine - the majority of the remaining aa is proline, hydroxyproline,hydroxylysine Dr. Prapee 22 Dr. Prapee 23 Six Major types of collagen Types I II III Characteristics Most common type collagen ,thick fiber Forms slender fiber in hyalin and elastic cartilages Reticular fiber - Highly glycosylated, thin fiber0.5-2.0 um - Rich coating of sugar likely stained with silver salt (Argyrophilic fiber) - positively stained with periodic acid schiff (PAS) reaction Dr. Prapee 24 Six Major types of collagen Types IV V VI Dr. Prapee Characteristics Do not display 67 nm periodicity Forms a meshwork of procollagen molecules Forms very thin fibrils and association with Type I Forms small aggregates, anchoring fibrils 25 Elastic Fibers • Slender, long and branching in loose CT • Form coarse bundles in ligament (ligamentum flavum) and fenestrated sheet (concentric sheet in the wall of large a.) • Elastin,a protein rich in glycine and proline - unusual aa, Desmosine & Isodesmosine - a high degree of elasticity - forms a core of elastic fiber • Microfibrils (10 nm in diameter, glycoprotein, fibrillin), located at the periphery of elastin Dr. Prapee 26 Elastic fibers Dr. Prapee A. Elastin has a random coil structure in the relax state, but reforms as a different random coil on relaxation B. Elastin molecule are covalently linked into arrays which can reversibly stretch and recoil, and may be arranged as fibers or sheets • Elastic fibers are composed of microfilaments surrounding and organizing core region of crosslinked elastin 27 Blood vessel, showing the elastic fibers Elastic f. Dr. Prapee 28 Tunica media of aorta Show the typical wavy appearance of thick bundles of elastic fiber Dr. Prapee 29 Reticular fibers • Collagen type III • Frame work of lymphoid organs, liver ect. • Highly glycosylated, thin fiber0.5-2.0 um • Rich coating of sugar likely stained with silver salt (Argyrophilic fiber) • Positively stained with periodic acid - schiff (PAS) reaction Dr. Prapee 30 Cellular components of CT • Fixed cells - A resident population of cells have developed and remain within CT - fibrobasts, mast cells, adipose cells, pericytes & macrophages • Wandering (Transeint) cells - Originate mostly in bone marrow - plasma cells, lymphoctes, neutrophils, eosinophils, basophils, monocytes, and some macrophages Dr. Prapee 31 Fibroblasts - Produce ECM & Fibers - Active fibroblasts often reside with collagen bundles - Elongated, fusiform cells & pale-staining cytoplasm - Large ovoid nuclei with prominent nucleoli - EM, showing prominent RER, Golgi complex - Inactive, Fibrocytes ; small, elongate & deeply stained nuclei - Some movement Dr. Prapee 32 Fibroblasts and Fibrocytes Fibroblasts Fibrocytes Dr. Prapee 33 Fibroblasts, showing euchromatic nucleus, prominent RER Collagen fibers Collagen fibers Dr. Prapee RER 34 Myofibroblasts - Modified fibroblasts contain actin and myosin filaments - Surface profile of nucleus resembles smooth m. - Basal lamina is absent - Abundant in wound healing area (purse string effect) - Contribute to retraction and shrinkage of scar tissue Dr. Prapee 35 SEM : Pericytes - Undifferentiated mesenchymal cells - As named adventitial cells or perivascular cells - Locate around capillaries and venules - Possess Dr. Prapeecharacteristic of smooth muscle cells and endothelial 36cells TEM : Pericytes Capillary Pericyte Dr. Prapee P - Their basal lamina are directly continuous with basal lamina of endothelial cell - EM, cytoplasmic characteristics almost identical with endothelial cells - In large venules, they have characteristic of smooth m. cells 37 Wound healing - New connective tissue & blood vessels are formed - Undifferentiated mesenchymes lacated in tunica adventitia of venules and small veins - Fibroblast - Pericytes - Endothelial cells Dr. Prapee 38 Macrophages - They function in removing cellular debris (phagocytosis) and protceting body against foreign invaders - They also play a role in presenting antigens to lymphocytes - They are about 10-30 um in diameter - Cell surface is uneven - Basophilic cytoplasm, many small vacuoles, small dense granules- Indented nucleus - EM showing prominent Golgi app. & RER, abundant lysosomes Dr. Prapee 39 Macrophage Under chronic inflammatory condition • They congregate, enlarge and become polygonal-shaped epitheloid cells • If the particulate matter is excessive large, several macrophages may fuse to form a Foreignbody giant cell • Macrophage residing in CT, Fixed Macrophage (resident Macrophage) • Free Macrophage - migrate from the blood Dr. Prapee 40 Macrophage Dr. Prapee 41 Macrophage Development and Distribution • Histologist once believed that macrophages were derived from precursor cells in Reticuloendothelial System, which included nonphagocytic cells • Presently, all members of phagocytotic cells arise from a common stem cell in bone marrow, possess lysosomes, can phagocytose and display Fc receptors and receptors for complement, as classified to as Mononuclear Phagocyte System Dr. Prapee 42 Macrophage Development and Distribution Macrophages localized in certain regions of the body were given specific names before their origin was understood. • Kupffer cells of the liver • Dust cells of the lung • Langerhans cells of the skin • Monocytes of the blood • Macrophages of CT, spleen, lymph node, thymus and bone marrow • Osteoclast of the bone • Microglia of the brain Dr. Prapee 43 Mast cells • Oval shape with round or oval nucleus • 20-30 um in diameter • Abundance of the large basophilic granule • Granules show Metachromasia • Granules containing histamine, heparin, eosinophil chemotactic factor (ECF), nutrophil chemotactic factor (NCF), chondroitin sulfate protease, aryl sulfatase Dr. Prapee 44 Mast cells • Oval shape with round or oval nucleus • 20-30 um in diameter • Abundance of the large basophilic granule • Granules show Metachromasia Dr. Prapee 45 Mast cells Dr. Prapee• Granules show Metachromasia 46 Mast Cell • Granules containing histamine, heparin, eosinophil chemotactic factor (ECF), nutrophil chemotactic factor (NCF), chondroitin sulfate protease, aryl sulfatase Dr. Prapee 47 Plasma cells • Found in large numbers in area of chronic inflammation • Derived from B lymphocyte • Ovoid cells, 10-20 um, eccentric nucleus • Short life span of 2-3 weeks • Intensely basophilic cytoplasm, prominent RER • Spherical nucleus consisted of heterochromatin radiating out from the center, look like clockface appearance in LM observation • Produce circulating antibodies (r-globulin) Dr. Prapee 48 Plasma cells • Found in large numbers in area of chronic inflammation • Derived from B lymphocyte •Dr.Ovoid Prapee cells, 10-20 um in diameter, Eccentric nucleus 49 Plasma Cell • Intensely basophilic cytoplasm, prominent RER • Produce circulating antibodies (rglobulin) • Spherical nucleus consisted of heterochromatin radiating out from the center, look like clockface Dr. Prapee appearance in LM observation 50 Lymphocyte • Smallest free cells, 6-8 um in diameter • Spherical with round nucleus (heterochromatin) • Thin rim of cytoplasm lym Dr. Prapee 51 Lymphocyte • EM, a few organelles but a large number of free ribosome • Thin rim of cytoplasm • Small, medium & large lymphocytes • B-Lymphocytes plasma cells • T-Lymphocytes cellmediated immunity Dr. Prapee 52 Lymphocyte • Accumulate in Peyer's path of Ileum • Distribute in lamina propria of G.I & respiratory tract etc. Lamina propria Dr. Prapee 53 Neutrophils Dr. Prapee - Polymorphonuclear neutrophil (PMN), polymorph, 4-5 lobes - 10-12 um in diameter - There are three type of granules - Small, specific granules (alkaline phosphatse) - Large, azurophilic granules, lysosomes (peroxidase, acid phosphatase) - Tertiary granules (gelatinase, cathepsins) 54 Neutrophils - The first cells to appear in acute bacterial infection - Phagocytose microorgansim die resulting in formation of pus Dr. Prapee 55 Neutrophil Dr. Prapee 56 Eosinophils •10-12 um in diameter •Bilobes nucleus • Large elongated granules - crystalloid body - peroxidase - histamine - hydrolytic enzyme • Phagocytose Ag-Ab complex Dr. Prapee 57 Eosinophils Note : the crystalloid bodies in the granules Dr. Prapee 58 Monocyte In blood CT Macrophage Dr. Prapee 59 Basophil • Bilobed nucleus • Smaller size than monocyte • Contains receptor for immunoglobulin as in mast cell Dr. Prapee 60 Adipose Tissue • Unilocular adipose tissue, White fat • Multiloccular adipose tissue, Brown fat Dr. Prapee 61 Adipose cells White fat Dr. Prapee • Adipocytes, Adipose cells, Lipoblasts • Single lipid droplet • Variation in color from white to deep yellow, depend on carotenoid in diet • Eccentric nucleus, signet ring appearance • Lipid is usually extracted during conventional preparation 62 White Fat : Sudan red Dr. Prapee • A few organelles • Mature adipocytes do not divide • Highly blood supply • Receptors for Insulin, GH, NEP, Glucocorticoid • Distribute in subcutaneous layers, mesentery • Accumulation depends on sex, age, region • Large cells, 100-120 um, thin rim cytoplasm 63 Brown Fat Dr. Prapee • Multiple lipid droplets • Tan to reddish brown, cytochrome in mitochondria • Eccentric round nucleus Extensive vascularity • Numerous unmyelinated nerve fiber, axon ending on both blood vessels and on fat cells • Prominent in newborn • Rate of fatty acids oxidation up to 20 times the rate of white fat 64 Brown Fat • A few cell organelles • Abundant roundshaped mitochondria Dr. Prapee 65 Distribution of Brown Fat New born : superficial Dr. Prapee Deep Rabbit 66 Clinical correlations : Adult, obesity • Hypertrophic obesity - accumulation and storage of fat in unilocular fat cells - increase their size • Hypercellular obesity - over abundance of adipocytes - this type of obesity is severe Dr. Prapee 67 Clinical Correlations • Lipomas, common benign tumor of adipocytes • Liposarcomas, malignant tumors of adipocytes Dr. Prapee 68 Dr. Prapee 69 Mesenchymal CT • Numerous mesenchymal cells, oval euchromatic nucleus, prominent nucleoli • Pluripotency • Pale-staining cytoplasm, small processes • Gel-like, amorphous ground substance • Consists of reticular fibers Dr. Prapee 70 Mucous tissue • Fibroblasts • Jelly-like, amorphous ground substance composed of hyaluronic acid called Wharton's jelly • Composed of Type I and type III collagen fibers • Found in umbilical cord, subdermal C T of embryo Dr. Prapee 71 Connective Tissue Proper Loose (Areolar) C.T. • Abundant ground substance, proteoglycans • Contains all types of cells • Contains all types of fibers • Fills in the body spaces, adventisia of blood vessels, lamina propria of G.I. tract ect. Dr. Prapee 72 Loose Connective Tissue displaying collagen (C) and elastic fibers Dr. Prapee 73 Distribution of loose CT in lamina propria of GI tract Lamina propria Dr. Prapee 74 Dense C T • More fibers • Fewer cells • Orientation and arrangements of the fiber make it resistance to stress Dr. Prapee 75 Dense Irregular C T • Mostly coarse collagen fibers arranged randomly that resists stress from all directions • Fibroblast, the most abundant cells • Distribution in dermis, sheath of nerves, capsule of organs ect. Dr. Prapee 76 Dense irregular CT : Perichondrium (P) Dr. Prapee 77 Dense irregular CT: Dermis Dr. Prapee 78 Dense Regular Collagenous C T • Collagen bundles densely packed and oriented into parallel cylinders or sheet • Resists tensile strength • Little ground substance and cells (Fibroblasts) •Tendons, ligaments, aponeuroses Dr. Prapee 79 Dense Regular Collagenous C T ( Tendon ) Dr. Prapee 80 Dense Collagenous Connective Tissue (Muscle- tendon junction) Dr. Prapee 81 Dense Regular Elastic C T • Elastic fibers arranged parallel to each other and form thin sheets or fenestrated membrane (large bl.ves.) • Ligamentum flava of vertebral column, suspensory ligament of penis Dr. Prapee 82 Reticular Tissue • Type III collagen • Form mesh-like network • Liver sinusoid, adipose tissue, lymphoid organs Dr. Prapee 83