Department of Histology and Embryology, Faculty of Medicine MU pvanhara@med.muni.cz Tissue concept and classification Petr Vaňhara, PhD  Organization of human body Hindu Avicenna Moderní buněčná teorie Jan E. Purkyně Matthias J. Schleiden Theodor Schwann Robert Remak Rudolf Wirchow Camillo Golgi … And many others Aristotle and medieval medicine Chinese medicine  Modern cell theory  Cells are the basic units of any organism  New cells origin only from other cells  Cells exchange energy (open thermodynamic system)  Genetic information is inherited in new generations  Chemical and structural composition of cells is generally identical How do these cells differ? What is the mechanism of tissue formation? How the variability of a multicellular body develops? - 6  1013 CELLS of 200 different types In human body: - cells form functional, three-dimensional, organized aggregations of morphologically similar cells and their products or derivatives - TISSUES - tissues constitutes ORGANS and organ systems  Definition of a tissue  classical histological definition is based on microscopic visualization Functional, three-dimensional, organized aggregation of morphologically similar cells, their products and derivatives  Functional cells of tissues differentiate from stem cells Knoblich JA. Asymmetric cell division during animal development. 2001. Nat Rev Mol Cell Biol Differentiation Self renewal Stem cells are capable of differentiation and self renewal Asymetric division Proliferation Formation of functional types Stem cells Totipotent - Constitute all cells of the body incl. extraembryonic tissues - Zygote and early stages Pluripotent - All cells in the body except for trophoblast - Blastocyst – Inner cell mass - ICM (embryoblast) - Embryonic stem cells Multipotent - Give rise to various cell types of a particular tissue - Mesenchymal SC, hematopoietic SC http://www.embryology.ch/anglais/evorimplantation/furchung01.html Oligo- a unipotent - One or several cell types – hematopoietic, tissue precursors for renewal of intestinal epithelia, etc. Stem cells in human body Tissue (adult) stem cells - regeneration and renewal of tissues - GIT, CNS, mesenchyme - regenerative medicine, cancer biology Embryonic stem cells (ESCs) - embryoblast of blastocyst - pluripotent - modelling of early embryogenesis, regenerative medicine  Stem cells as a research tool Induced pluripotent stem cells (iPSc) - adult differentiated cell (fibroblast) is reprogrammed into pluripotent state - differentiation into desired cell type - regenerative medicine, cell and gene therapy Nobel prize 2012 Disease modelling Drug testing Tissue replacement …  Induced pluripotent stem cells share biological properties with embryonic stem cells hESCs hiPSCs  Stem cells as a therapy Age-related macular degeneration neovascularisation Clinical trial hiPSCs Retinal pigment epithelium  Stem cells as a foe Cancer stem cells - solid tumor is always heterogeneous - small population of cells with stem cell character can repopulate tumor tissue after cytotoxic therapy Tissue stem cells Renewal Low frequency (<1%) Quiescence Multipotency Long life Resistence Tumorigenicity Proliferation capacity Cancer stem cells  Tissues are not uniform Parenchyma: functional tissue of an organ (liver, lung, pancreas, kidney parenchyma) Stroma: surrounding, supportive tissue Parenchym Stroma Parenchyma - Hepatocytes - Sinusoids and surrounding structures Stroma: - CT - Veins - Nerves - … Tissue differ in their genetic and epigenetic profile doi:10.1038/nature10523 Tissue differ in their genetic and epigenetic profile Cells create unique microenvironment Adhesion molecules Growth factors ECM components Cell interactionsMetabolites Immunity Inflammation  Microenvironment regulates tissue function and reflects its tissue composition • Embryonic development • Intercellular interaction • Space organization (dimensionality) • Gradient of morphogenes • Epigenetic profile • Gene expression dynamics • Partial pressure of gases • ECM composition • Mechanical stimulation • Perfusion and interstitial flows • Local immunity response • Metabolites Huge number of biological and physically-chemical parameters Bone marrow microenvironment drives hematopoiesis „Stem cell niche“ Microenvironment is necessary for tissue homeostasis Apoptosis Regeneration Senescence Transformation Microenvironment controls embryonic organogenesis Microenvironment is of clinical importance Tissue engineering Tissue engineering Ectoderm MesodermEntoderm Trilqaminar germ disc (3rd week) Histogenesis and organogenesis Embryonic development  Connective tissue of head  Cranium, dentin  Skeletal muscle of trunk and limbs except cranium  Dermis of skin  Muscles of head  Urogenital system + ducts, glands and gonads  Visceral muscle and connective tissue  Serous membranes of pleura, peritoneum and pericardium  Blood cells, leukocytes  Cardiovascular and lymphatic system  Spleen  Adrenal cortex  GIT epithelium except oral cavity and part of anal canal  Extramural glands of GIT  Epithelium of bladder  Epithelium of respiratory system  Thyroid gland, parathyroid glands, thymus  Tonsils  Epithelium of cavum tympani and Eustachian tube  Epidermis, hair nails, cutaneous and mammary glands  Corneal epithelium and lens of eye  Enamel of teeth  Internal ear  Anterior pituitary gland  Epithelium of oral cavity and part of anal canal  Neural tube and derivatives - CNS - Retina - Posterior pituitary gland - Pineal body  Neural crest and derivatives: - Cranial and sensory ganglia and nerves - Schwann cells - adrenal medulla - Enteroendocrinne cells - Melanocytes - Head mesenchyme and connective tissue - Odontoblasts SurfaceectodermNeuroectoderm headParaxialIntermediateLateral EndodermEctoderm Mesoderm Neural crest is a „fourth germ layer“ PNS: Neurons Sensoric, sympathetic a parasympathetic ganglions and plexuses Neuroglia and Schwann cells Adrenal medulla Calcitonin secreting thyroid cells Pigment cells of epidermis Viscerocranium Corneal stroma and endothelium Tooth papilla Dermis, smooth muscles and adipose tissue of skin of head CT of salivary and lacrimal glands, thymus, thyroid and pituitary …  Neural crest cells are multipotent in vitro Ref.: Shakhova O., Sommer L. Stem Book 2012. Neural crest-derived stem cells Molecular principles of histogenesis doi:10.1038/sj.hdy.6800872 Hox complex Highly conserved family of transcription regulators that determine body polarity, orientation and axis Tissue differentiation along anterio-posterior axis Human (39 genes) Cluster Chromosome # Hox genes HoxA 7 11 HoxB 17 10 HoxC 12 9 HoxD 2 9 Congenital disorders and HOX genes hand-foot-genital syndrome – mutation HOXA13 synpolydaktylia – mutation HOXD13 Hox komplex a morphogenetic field Example: Differentiation of mouse urogenital tract (prostate) Posterior Anterior Temporo-spatial expression of different regulators determines final localization, orientation and morphology of a tissue. PosteriorAnterior Dorsal Ventral doi: 10.1210/en.2006-1250 Hoxb13 Distal Proximal HOX Limb formation Vascularisation AER ZPA Ectoderm Mesenchyme FGF IGF … Proliferation Thalidomid Epithelium Muscle Nerve Connective Contemporary tissue classification Based on morphology and function: Myofibrils  contraction Mesoderm – skeletal muscle, myocard, mezenchyme – smooth muscles Rarely ectoderm (eg. m. sphincter a m. dilatator pupillae) Neurons and neuroglia Reception and transmission of electric signals Ectoderm, rarely mesoderm (microglia) Dominant extracellular matrix Connective tissue, cartilage, bone… Mesenchyme Continual, avascular layers of cells with different function, oriented to open space, with specific junctions and minimum of ECM and intercellular space. Derivates of all three germ layers Microvilli on surface of small intestine Isocortex vascularisation http://www.livescience.com/14413-brain-images-portraits-mind.html Connective tissue Not only a tissue glue…  General composition of connective tissue (CT) Cells and extracellular matrix Matrix – fibrous and amorphous Fibrous component - collagen - reticular - elastic Amorphous component (amorphous ground substance) - Complex matrix consisting of glycosaminoglycans, glycoproteins and proteoglycans, depending on tissue type (connective  ligament  cartilage  bone) Cells Connective tissue – permanent and transient cell populations (fibroblasts/myofibroblasts, immune cells, adipocytes, adult stem cells) Cartilage – chondroblasts/chondrocytes Bone – osteoblasts/osteocytes/osteoclasts  Embryonic origin of CT • Mesenchyme = loose tissue between germ layers • Complex network of star- or spindle-shaped cells • Jelly-like amorphous ground substance http://www.mun.ca/biology/desmid/brian/BIOL3530/DB_Ch02/DBNModel.html DAY 12 of embryonic development  Basic derivatives of CT Connective BoneCartilage Mesenchyme  Classification of CT Embryonic CT - Mesenchyme - Jelly-like CT (Wharton jelly, dental pulp, strom of iris) Adult CT - Areolar (loose, interstitial) CT - Dense collagen irregular CT - Dense collagen regular CT - Fat (adipose tissue) - Cartilage - Bone - Blood and hematopoietic tissue - Lymphatic tissue CT Specialized CT Trophic CT (body liquids)  Cells of connective tissue Cells - Fibroblasts/fibrocytes/myofibroblasts - Heparinocytes - Macrophages of CT = histiocytes - Plasma cells - Lymphocytes - Adipocytes - Adult stem cells Extracellular matrix - Fibrous compound - Amorphous ground substance  Cells of connective tissue Mesenchymal (adult) stem cells Koch et al. BMC Biotechnology 2007 7:26 doi:10.1186/1472-6750-7-26  Extracellular matrix – fibrous component Collagen fibers - family of fibrous proteins encoded by >35 genes (2013) - polymer – subunit = tropocollagen; triple helix - different structural and mechanical properties (strength, elasticity, pliability…) - most abundant protein in human body ( 30% dry weight) Type Localization Structure Main function I Bone, tendons, meniscus, dentin, dermis, capsules of organs, loose CT 90% of type I Fibrils (75nm) – fibers (1-20m) Resilience in pull II Hyaline and elastic cartilage Fibrils (20nm) Resilience in pressure III Skin, veins, smooth muscles, uterus, liver, spleen, kidney, lung Like I, high content of proteoglycans and glycoprotiens, reticular network Shape formation IV Basal lamina of epithelium and endtohelium, basal membranes No fibrils or fibers Mechanical support V Lamina of muscle cells and adipocytes, fetal membranes Like IV VI Interstitial tissue, chondrocytes – adhesion Connecting dermis and epidermis VII Basal membrane of epithelium VIII Some endothelia (Cornea) X Growth plate, mineralized cartilage Growth of bones, mineralization  Collagen  Collagen  Collagen in LM AZAN HES HE Julian Voss-Andreae "Unraveling Collagen", 2005 Orange Memorial Park Sculpture Garden, City of South San Francisco, CA  Elastic fibers • less abundant than collagen • polymer – tropoelastin • minimal tensile resistance, loss of elasticity if overstretched • reduction of hysteresis = allow return back to original state after mechanic change  Reticular fibers • collagen 3D meshwork • bone marrow, spleen, lymphatic nodules • microenvironment for e.g. hematopoietic stem cells and progenitors Amorphous extracellular matrix Colorless, transparent, homogenous substance consisting of glycosaminglycans, proteoglycans and structural glycoproteins  Extracellular matrix – ground matrix  Glycosaminoglycans linear polysaccharides composed of two disaccharide subunits – uronic acid and hexosamine glucosamin or galactosamin glucuronic or iduronic acid polysaccharides rich in hexosamines = acid mukopolysaccharides Glycosaminoglycan Localization Hyaluronic acid Umbilical cord, synovial fluid, fluid of corpus vitreum, cartilage Chondroitinsulphate Cartilage, bone, cornea, skin, notochord, aorta Dermatansulphate Skin, ligaments, adventitia of aorta Heparansulphate Aorta, lungs, liver, basal membranes Keratansulphate Iris, cartilage, nucleus pulposus, anulus fibrosus  Glycosaminoglycans They bind to protein structures (except for hyaluronic acid)  Proteoglycans — protein + dominant linear saccharide component — proteoglycan aggregates — water-binding, volume dependent of hydratation — aggrecan (cartilage) — syndecan — fibroglycan • dominant protein + branched saccharide component • interaction between cells and ECM ― fibronectin – connects collagen fibers and glykosaminoglycans, cell adhesion and migration ― laminin – basal lamina – epithelial integrity ― chondronectin – cartilage – adhesion of chondrocytes to collagen (J. Nutr. 136:2123-2126, 2006)  Structural glycoproteins  Composition of amorphous ground matrix  Classification of specialized connective tissue http://www.exploringnature.org/db/detail.php?dbID=21&detID=691  Adipose tissue • Adipocytes, fibroblasts, reticular, collagen and elastic fibers, capillarie • White and brown adipose tissue  Brown adipose tissue • fetus and child to 1st year of life • fast source of energy • typical localization – between shoulder blades, axilla, mediastinum, around kidneys, pancreas, small intestine • small cells with numerous fat droplets  White adipose tissue • adipocytes are actively form until 2nd year of life • no innervations, but rich vascularisation • adipocytes with only one lipid droplet • leptin (adipokinins)  Cartilage ― specialized connective tissue with continuous ECM ― flexible, mechanically resistant ― avascular, no innervation ― support of soft tissues ― diarthrosis ― growth • perichondrium – connective tissue around cartilage (not present in joints) • chondroblasts, chondrocytes • extracellular matrix (collagen and elastic fibers, amorphous ground matrix)  Composition and structure Collagen type II Glykosaminoglycans (Hyaluronic acid, chondroitinsulphate, keratansulphate) Proteoglycane aggregates Hydrophilic character – holds water → low friction → smooth movement of joints  Hyaline cartilage - most abundant - temporary embryonal/fetal skeleton - epiphyseal growth plate - articulation (joints) respiratory passages - isogenic groups Exchange of metabolites Apositional growth Interstitial proliferation  Elastic cartilage - Elastic fibers in matrix - No isogenetic groups - Auricula, meatus, larynx, epiglottis  Fibrocartilage • Fibrous compound dominant – collagen I and II – mechanical durability • Minimum of amoprhous matrix-fibers visible • Intervertebral discs, symphysis pubis, articular discs, meniscus  Clinical correlations Cartilage – no innervation, no vascularization – no spontaneous regeneration No migration of chondrocytes to site of damage Initiation of other degenerative events leading to cartilage erosion (arthritis) Therapy: - joint mobility - restoration of biochemical and biophysical parameters of cartilage - prevention of further damage - removal of damaged tissue, autologous transplantation, MSCs on biocompatible scaffolds  Bone Spongy Articular cartilage Bone marrow cavity Capillaries Diaphysis Epiphysis Compact bone Periost Capillaries in Haversian system Capillaries in Volkman’s system  Histological classification of bone tissue • Primary (woven, fibrous) - Temporary, growth and regeneration of bones, collagen fibrils woven - Replaced by secondary bone - Remains only in some parts of body - sutures of skull tuberositas ossium, tooth cement • Secondary (lamellar) - Lamellae – collagen fibers in concentric layers (3-7m) around a canal with capillaries = Haversian system (osteon) - Spongy (trabecular) - Trabecules, similar to compact - Epiphyses of long bones, short bones, middle layer of flat bones of the skull (diploe) - Compact - Outer and inner coat lamellae typical Haversian systems - Volkmann’s canals - Interstitial canals  Surface of compact bone • Outer surface - Synovial joint – hyaline cartilage - periosteum (periost) – membrane – dense CT, inner layer (osteoblasts) and outer layer (fibrous CT) - Inactive bone - fibrous CT in periost dominant - Collagen fibers – parallel to the bone surface - Sharpey’s fibers fix periost to the bone National Museum of Natural History NY, USA • Inner surface – cavities lining - Medullar cavity - Endosteum (endost) – single cell lining – bone remodeling - Red bone marrow – hematopoiesis - Yellow and gray bone marrow – adipocytes or CT - Rich vascularisation  Bone matrix - 60% mineral compound, 24% organic compound 12% H20, 4% fat - Crystals – calcium phosphate, hydroxyapatite • Osteoblasts - specialized bone cells - produce ECM – collagen (I) and noncollagenous proteoglycans, glycoproteins - osteocytes  Cells • Osteoclasts - multinuclear, form by fusion of macrophages - bone matrix resorption  Cells • Osteoclasts - Complex architecture - Enzymes degrading organic matrix - HCl  Cells  Ossification Intramembranoeous - Mesenychymal cells → osteoblasts - Ossification center – rich vascularisation, differentiation of osteoblasts, synthesis of primary bone Endochondral - Cartilage model - Growth plate - Primary and secondary ossification centers (diaphyse, epiphyses) 2 1 3 4 5 http://ns.umich.edu/Releases/2005/Feb05/img/bone.jpg • Fracture healing  Clinical correlations Reactive Phase - Fracture and inflammatory phase - Granulation tissue formation Reparative Phase - Cartilage callus formation - Lamellar bone deposition Remodeling Phase - Remodeling to original bone shape • Bone remodelation disorders – OSTEOPOROSIS  Clinical correlations • OSTEOPOROSIS - Abnormal activity of osteoclasts - Low level of estrogens (menopausis) - Inflammation - Immobilization - Nutrition - Endocrine disorders - Side effect of therapy with corticosteroids, antiepileptics, anticoagulantia - treatment: antiresorbing drugs (bisphosphonates, estrogene analogues), stimulation of bone formation, nutrition, exercise - Osteolytic phenotype – multiple myeloma, breast cancer  Clinical correlations • Bone remodelation disorders – OSTEOPETROSIS - decreased activity osteoclasts - congenital disease - various damages (nerve compression, fractures, joint erosion, anemia – closing of medullar cavity)  Clinical correlations  Clinical correlations • Rheumatoid arthritis - autoimmune inflammatory disease - erosion of soft tissue and bone matrix (synovial membrane, cartilage, bone)  Further study Department of Histology and Embryology Fac. Med. MU Med Atlas or visit http://www.med.muni.cz/histology Thank you for attention