Lecture 11: Wound Healing and Calcification PDF

Tissue repair Wound healing Calcification Tibor Krenacs Department of Pathology & Experimental Cancer Research Semmelweis University, Budapest Dentists 2023/24  Tissue integrity – essential for life INJURY...

Tissue repair Wound healing Calcification Tibor Krenacs Department of Pathology & Experimental Cancer Research Semmelweis University, Budapest Dentists 2023/24  Tissue integrity – essential for life INJURY HEALING Differenceoetwee fibrosise reservation adaptive immunecan takeaction protein in activity stomacans produce INFLAMMATION REGENERATION FIBROSIS Yasss cosomeother material ecm 1 - Neutralization - Restoration of tissue -Tissue repair sunas conasenis of toxic agents structure and function when reticular fibers & 2 - Elimination of (Damage: caused by basement membranes anwar extra matrix toxic agents trauma and/or toxic (ECM) damaged 3 + tissue debrie agents + inflammation) (Serious e.g. burns injury, chronic inflammation) Fibrotic scarring Regeneration e.g. myocardial infarction „ischemic hearth disease”Icu case isthetissueis restoringitstissuestructurecreseneration cranes growth factors huge burden public health Dryoute s ~7.5 million death/year cases it me a seriousinjury thefibrosisprocessDominatesinsteadof regeneration Inflammation: serious injury; innate + adaptive immunity, proteases, fibrosis  Regeneration – Wound healing - Cascades of overlapping, stereotypic events - Coordinated interactions between cell-cell and cell-matrix needs Surgical incision wound Basicsteps ofwoundmeaning reeneration tissue damage I - Bleeding - Coagulation: Isolation from the environment minutes -1 h fibrin cloth – scab own 2- Inflammation: Prevention of infection/septicemiaooorosining continous neutrophils, then macrophages so to amenamesmansosamewniencia theBacteria region 3- Debridement: Elimination of tissue debrie prosecute anaoris day-1 macrophages phagocytosis new aus should 4- Proliferation, Migration: Replenishment of lost tissue beformed to day-3 fibroblasts, new capillaries, parenchyma replacethose erigeron whichwere lost 5 - Epithelialization: Parenchyma regeneration - Angiogenesis: Nutrition of granulation tissue - Fibroplasia: Fibroblast invasion, matrix production conagenteen protene 8 - Remodelling: Generation and degeneration of ECM Dynamic process 9 - Contraction: Close up wound edges lossof water to- Resolution: Restoration ofgrisined appearance and function involves gcouasen Maturation COL-III COL-I, perpendicular orientation Robbins pathology 8th edition  Timing of main events in wound Ifor a longer healing Heaving process Bleeding & he rana majorran coagulation Extra anwar Granulation tissue matirx Bive presses a o re specimen em cariranies - Early fibroblasts matrontese various compact - Loose extracellular Benares stormisthe conaseny woundnearing matrix (ECM) eons process months atefor - Network of newly formed vessels - Inflamatory cells Early Late (macrophages) Trichrome staining Collagen = blue Sanatio per primam Sanatio per secondam Types of healing noneserious largerwounds Healing by first Sterile, sharp Serious, largeisner chance forinfection intention surgical incision injury, e.g bunrs Hearingbysecond wound chronic inflammation q SCAB IDENTICAL steps Neutrophil granulocytes Taser wound but extensive fibrosis Clotting in secondary: - defect is larger - wound edges are wider - extended inflammation Cell proliferation - healing is slower Granulation tissue - scar is larger Macrophages Fibroblasts - substantial contraction New capillaries (scar & deformities) issmewwound IMPORTANT w causelaser connective tissue scaffolding werner Fibrous union Contraction incl. basal laminae canoe reward Resolution compute destruction smaninsure intact reservation iscompote insures career sea ostosination Sanatio per primam Surgical incision wound Organized stratified squamous epithelium with even dermal borders Skin appendages: hair follicles and sebaceus glands do not regenetage Primary (early) Secondary (late) lineson bodydescribing the tension nature orsniationof conasencioers sein Langer’s lines (skin tension) Trichrome staining: collagen is blue - natural orientation of dermal collagen bundles ~ parallels with muscle orientations beneath Orientation of collagen fibers & bundles - parallel surgical incisions wound parallel to wound B A heal better with less scaring  oneto lesstensiononthe skin HEALING: Aiming to regenerate tissue structure and function REGENERATION REPAIR with FIBROSIS Co-operations between:factors like Inflammatory response Cell proliferation Acute: neutrophils, macrophages Cell cycle regulation Chronic: lymphocytes, plasma cells Proliferative capacity of tissues basophils, eosinopholils Stem cells ifits along lastingHemings ans adaptiveimmune alsoonceaction Growth factors & receptors Extracellular matrix (ECM) Nature and types Scaffolding & Interactions Growth factor receptors ECM production & remodelling Signaling pathways Matrix metalloproteinases (MMP)  replacetheios aus me yto neednewcar which ane 2015 Cell proliferation – Cell cycle naar in thecencure metogeneticmutationines Nobel price tenerAround24Hours In cancer: - repair mechanism are aberrant in chemistry - cyclin/Cdk-s - activating mutations T. Lindahl - or cdk inhibitor – loss of function P. Modrich A. Sancar DNA REPAIR DNA duplication can Happen itramose isseen Ees X it notone the repair to sonsea APOPTOSIS ans itdies programmed check if ceesis cell death Licensing able to take the license to continent He cancycle Growth factors insidetranscriptionphase y can inndoitorsI E2F Promoters Inhibitors (p16, p21, p27) -Cyclin-cdk -Cyclin dependent complexes Rb phosphorylation Retinoblastoma kinase inhibitors reinocastona Depnosphiyati Phosphylate  Mitosis Programmed cell death apoptosis metaphase a ones are caroms the agonized in causes By propose nuclear shrinkage9 chromsone My chromsones areformedbut Deformation anent on Greger dined membraneinfused metaphase nuclei apoptotic bodies Do not intiate the immuneresponse  Regulation of baseline cell population Comitted Cells able to regenerate throughout life normal cell A STEM CELLS self-renewal population assymetric cell division reversibly postmitotic cells Intermitotic Dividing cars cells instateof mitosis 1. symmetric Transit amplifying BALANCE cell population 2. assymetric usedupcells proliferation apoptosis y cancerstem cell death Targeted for y cancer chemotherapy our xn INJURY! remaining it elevated ofcancerstem cens After cell decay cancer chemotherapy maycause relapseof the cancer  Stem cells –Progenitor cells released O Embryonic stem cell Forming different cell types and tissues from Pluripotent: inner cell mass cells of morula (ecto-, meso-, entoderm) Blastocuster 0 Adult stem cells Maintaining cell compartments e.g. in regeneration AMultipotent: some cell types e.g. bone marrow CD34+ stem cells can form all subtypes of hemopoietic cells; or the mesenchymal stem cells Inooacensipanatute (form cartilage, bone, muscle) B Unipotent: surface epithelia: skin, GI track eptithelial stem cells canform only epithelialcells In vitro fertilization MHC/HLA - histocompatibility Transplant rejection  Reprograming - Induced pluripotent stem cells (iPS) maturecus pluripotent exchangeof Reprograming aways nucleus virentrantering of sene 2012 Nobel prize in Physiology-Medicine: Oct3/4, Sox-2 c-Myc, Klf4, Nanog Sir John B. Gurdon and Shinya Yamanaka "for the discovery that mature cells can be reprogrammed to become pluripotent" „Dolly the sheep” Problems: Problems: - Dedifferenciation GeneticTherapeutic age & quality is characteristic of of donor cloning cells cancer cells - Shortened telomeres - c-Myc & Klf4 are chanceof B - Epigenetic patterns reproductions proto-oncogenes! Restrict life expectancies mayresult in cancer Taru Deathofanimal Inefficiency to J. Embryol. Exp. Morphol. 10, 622-640. 1962  Regenerative medicine – Stem cells Replacing genetically or somatically damaged cells can cure these I A Genetic: wastes Away Muscular dystrophy g Cancer f got this surgery9 is Alive pinterest.com B Traumatic: Spinal cord injury, stroke Myocardial infraction Bone marrow CD34+ progenitor cell Jose Carreras c Degenerative: transplantation 1987 - transplantation Alzheimer’s A 2017 - January, Concert Parkinson’s most important in Budapest surgery  Regeneration - proliferative capacity of parenchymal cells labile cells/tissues: continuously replicating – Intermitotic cells Cintermiotic aus >1.5% of the cells in mitosis, may be destroyed by sublethal injury - hemopoiesis, skin, oral mucosa, GI tract, respiratory, urinary tract Stem cells – multipotent/unipotent – Efficient regeneration Restricts the Differentiation Stem cellcues of stem „niche” (microenvironment) Ki67 uprabason aver Hardlyenter Colon crypts thecure I Base Base.meHave less aus inthecure prowertia nienare roverting oralmucosa Small intestine crypts on Atease Stabil cells/tissues: terminally differentiated G0 phase cells (reversibly postmitotic) - adapt well, high metabolic activity, longevity - liver, kidney, pancreas, endometrium, endocrine glands - endothelial cells, fibroblasts, smooth muscle No stem cells (except in liver), differentiated cells can proliferate Liver regeneration Prometheus punished by Zeus – eagle eating from his liver, regrowth Permanent cells/tissues: terminally differentiated cells, no proliferative capacity (postmitotic cells) - neurons of the central nervous system, myocardium - if injured repaired by connective tissue scar (skeletal muscle? – satellite cells)  Healing of special tissues hepatocytes: reversibly postmitotic Dependent on: I - type of tissue Oval cells: stem cells but - regeneration potential Catteriesabove rarely involved - level of ECM damage Liver: - regeneration (focal, zonal necrosis) (stabile) - damage of ECM network (large abscess, cirrhosis) healing by fibrosis thickening soaring ofconnectivetissue river cirnosis liner owes Anepushed restricted fromfunctioning1 perfusion I d Hepatocytes conasen Liver cirrhosis, trichome taining: collagen – green, hepatocytes - red  Healing of special tissues B Myocardium (permanent cells): complex function, longevity, no regeneration no stem cells, the same for endocardium; Healing with scar: - arrhytmias - damaged pumping function ecommunica No regulated contraction fibrosis Protect But they j farther Damage the cells my my of fibrosis seproting myocardial cells y Myocardial fibrosis post-infartion Acut myocardial necrosis Disrupt their c Nervous systhem: a neurons can not replicate (SVZ, hyppocampus dentate gyrus)pumping function Central Peripheral it earnon - No axon regeneration - Axon regeneration if broken ends - Repair involves microglia, astrocyte aligned accurately; if not traumatic proliferation (inflammation) neuroma: fibrosis, abortive axon prolif.  EGFR Growth factor receptor subtypes PGDGFR TGF-β1 us get signorsfrom outside IL-6 FGFR G-protein coupled Regeneration receptor medicine TNFα (kemokines) Protein kinase pathways growth CCL2Blocking of inhibitory factors Without intrinsic (growth factors) CXCL12 of retinal axon regeneration speeda kinase activity reservation (cytokines) block at block both block 98thfactors PTEN – Akt/mTOR SOCS3 – JAK/STAT3 E 33 É 98a s 9 E 88 e e s z 2012 Nobel Prize in Chemistry Sun et al. Nature. 2011, Robert J. Lefkowitz & Brian K. Kobilka 480(7377): 372–375. "for studies of G-protein-coupled receptors"  Growth factors in wound healing Fibroblast proliferation CUTTED migration wound ECM synthesis EGF Keratinocyte proliferation differentiation inhibitor Angiogenesis  Growth factors in wound healing nameof these growth factors men discovera burst Based on thelocation wherethey werefirst piscovera Boththeycanbeseen inothersitesasmen (proliferation, migration, ECM production, kemotaxis, vasodilatation) EGFR Skin factorcansoseenin tumors epidermalgrowth Epithelial proliferation EGF, TGF-α, KGF, HGF Monocyte chemotaxis PDGF, FGF, TGF-β most important Growthfactors 3 Fibroblast migration PDGF, FGF, TGF-β1 they regulate fibrotic scarring BM stroma (myelofibrosis) Fibroblast proliferation PDGF, EGF, FGF, TNF Angiogenesis VEGF, Ang, FGF-2 Collagen synthesis TGF-β1, PDGF (TGF-β3 inhibits) 2these are most name function e important Chemokines, cytokines are also involved in the inflammatory phase of regeneration, which are not discussed here, only their receptor types were shown.  Extracellular matrix (ECM) and cell-matrix interactions producedby fibroblast ECM is a dynamic continuously remodelled complex of macromolecules ECM not an inert material just for filling up gaps !!! Types: Interstitial matrix Basement membrane (fibrillar: collagens, elastin; hydrated gels: proteoglycanes, hyaluronan; multiadhesive: fibronectin, laminin) toneupwatersprovideresinanceareasoncity Functions: I ECM sequesters water and provides turgor (resiliance) for tissues minerals (Ca10(PO4)6(OH)2 hidroxil-apatit, for bone rigidity (type I-collagen) 2 Supports (and anchores) parenchymal cells and their migration basement membrane (BM) in kidney - glomerular filtration Active contributor of wound healing 3 Basement membrane form scaffolding for tissue healing 4 ECM binds and offers regulatory molecules (growth factors) for: cell proliferation, migration, differentiation unioncouldactas memoranemoxanes as transporters men 5 Initiates signal transduction through integrins conimmunecues for example they can be transmembrane molecules too, e.g. syndecan & collagen XVII  functions of ECM fibrosis Myelo strong ouncesof fibrosisaround conasenture sense bony spinaa  Basal membrane Cornea Laminin Skin E est per Iggy maincomponents y pygmy of Basonmemorane cinceritnenan austobasement lamina Isis Collagen I and III fibers Bundles memorane an gin Proteoglycans fintnesops  Epithelial migration - Matrix remodelling the Ecmcomponent MMPs (Zn2+): Collagenases, Gelatinases, Strome-lyzines i mainly proteinf lamininl Keratinocyte Fibrin clot differentiation EPIDERMIS Integrin Coll XVII EGFR PROLIFERATION - MIGRATION Basal membrane BM-Laminin BM - synthesis Cleave TGF- Plasmin (MMP activation) HGF Col IV & Coll VII MMP gelatinases laminin 5 DERMIS MMP1,2,3, 9,10,11 (kollagenases) INVASION Coll I, III Matrix cleavage Neutrofil Macrophag Firoblast Endothel granulocyta in regeneration causeeritneeim snowbe Destroyed to aussaunau are move TIMP: „Tissue inhibitor of MMPs” MMP-s are also important in cancer metastasis! onsurface ofcons Martricryptic sites E-Cadherin strongadhesionbetweenepithelial cus Cleavage of precursors results in new biological activity (controlled proteolyzis – bioactive molecular fragments) I - endostatin Collagen XVIII angiogenesis Perlecan - endorepellin inhibitors Coll XVII Improtant in elimination of extra vessels the end of regeneration BindBasement to I in sermonregion  explained Oral mucosa – Healing without scar (fibrosis) Day-3 Day-7 Day-60 I Larjava H: J Can Dent Assoc 2011;77:b18 The pathway of healing is identical with that in the skin but faster and more efficient Differences Way Differences: (inherent) - less inflammatory cells (granulocytes, marcrophages) - saliva: antibacterial protection (histatins), accelerated clotting (exosomes) & proliferation (EGF), leukocyte protease inhibitor - less protocollagen I and fibronectin, more tenascin - residens fibroblasts express less TGF1 andfor important tiorosis decorin (fibrosis) - less fibrillogenesis and myofibroblasts (contraction) mace conssentioers more strongermatrix  Factors influencing wound healing Local: type of tissue, blood supply (nutrients, oxigenization), mechanical stress, type/duration of injury (cut, large destruction, acid, alkaline, burns), irradiation: UV, radioactive (ionizing) etc… Systhemic: cardiovascular status, infections, neutropenia, malnutrition, diabetes, lack of vitamins (e.g. vitamin C), corticosteroids important to somasen fiberformation Complications i swolleninflamedbleeding sum 2 lossofteeth 3 Insufficentwounding g Deathbyinfection Large tissue loss, deep wound – substantial scarring and distortion b considerablee largesize Pressure ulcer (decubitus): insufficient perfusion and oxigenization mechanical pressure > resistance of arterial wall mechanical Pressure exceeds the Resistance of Arterial wall ULCER I Resulting in ulcer Old age, poor perfusion: poor oxygenization, mechanical stress, diabetes  ArterialDisease Chronic wound (diabetic ulcer): arteriopathia, reduced perfusion, oxygen and nutrients,insufficient immune response against infection, neuropathy Dysfunctio Ferphron nerves  Keloid: elevated collagen synthesis, lost control during healing inherited, more frequent in the african-american populaltion Histology: large amount of cell free dermal eosinophilic collagen missing dermal papillae & appendages cant be fixed with surgery cause it can Regrow e re inlarge Iványi B Accures Due to a coarsen synthesis  REGENERATION – WOUND HEALING summary REGENERATION: Full reconstruction of the structure & function Continous remodelling of shortlived cells (bone marrow, surface epithelia) Liver, compensatory hyperplasia. Healing of sterile surical incision wound „WOUND REPAIR”: Partial reconstruction of structure and function, involvement of fibrosis The more preserved the matrix (BM, reticulat fibers) and the less chance for infection the better is the chance for reconstrion close to original state Precise orientation of surgical incision (Langer’s lines) and stiching. BOTH processes involve overlaping, stereotypic casdcades of events Coordinated direct cell-cell & cell-matrix interactions involving suluble factors too. Acute inflammation, proliferation, angiogenesis, parencyma and matrix remodeling improtance of growth factors their receptors and MMP-s The END RESULT is influenced by: The age & avaliability of stem cell pool (stem cell niche)! Different tissue types have different proliferation capacity. Localization, type of damaging agent and the extent of damage. General state of patients & their immune system, blood supply (nutrients oxygen), accompanying dieases (e.g. diabetes), vitamin deficit (patricularly of Vitamin C), infections, medications (e.g. corticosteroids) POTENTIAL RISKS. I 2 3 4 Overt fibrosis after myocardial infarction, lung fibrosis, liver cirrhosis, or restricted wounding in diabetes. Huge burden on the heath systems & economies worldwide! aeacecitate wesnow onconversing prevention ropetohave healthy PREVENTION through EDUCATION! Diets  Calcification Deposition of calcium salts makesupso o p ofbonymight Calcium is an important secondary messenger inducing I muscle contraction (striated, heart, smooth),2nerve 3 signaling, hormone 4 and enzyme release, and inducing 5 programmed cell death response Physiological calcification happens normal development of bones and teeth. Calcium hydroxy apatit in a type I collagen matrix. IAccumulate in collagenmatrix As a normal ageing process normal calcification may be seen in prostate, brain or in blood vessles There may be pathological calcification  Pathological calcification Pathological calcification is the abnormal tissue deposition of calcium salts, together with smaller amounts of iron, magnesium, and other mineral salts. There are two forms of pathologic calcification. 1- Dystrophic calcification 2- Metastatic calcification  Distrophic calcification - in dead tissues I Caseous necrosis in Tuberculosis is most common site of most common dystrophic calcification. cause Distraction of lungtissue calcium Diposition Liquefactive necrosis in chronic abscesses may get calcified Fat necrosis following acute pancreatitis or traumatic fat necrosis in breasts results in deposition of calcium soaps. Infarcts may undergo D.C. Thrombi especially in veins, may produce phlebolithis. Haematomas in the vicinity of bones may undergo D.C. Dead parasites like schistosoma eggs may calcify. Congenital toxoplasmosis or rubella may be seen on X-ray as calcifications in the brain. Dystrophic Calcification (D.C.) in degenerated or necrotic tissue, as in hyalinized scars, or after tissue damage as a consequence of medical device implantation. D I Unopened aortic valve in a heart due to calcification based aortic stenosis in an erderly person. The semilunar cusps are thickened and fibrotic. Behind each cusp are irregular masses of dystrophic calcification. c Aortic Walle thickns not able to close properly XL calcified plus S seen  Phitic BASO cells Renal tubules Psammoma bodies serous cc. in endometrium Calcified stone – submandibular salivary gland calcified store 1 Calcified hematoma - an example of dystrophic calcification (trauma) –physical disability complicates surgery is Hard Due to presence of small nerves near the Place of classifction t leave it without surgery x cant be operated Dystrophic calcification in CREST syndrome (systhemic sclerosis), and frequently occurs in connective tissue diseases, such as scleroderma (autimmune disease)  METASTATIC CALCIFICATION of Metastatic calcification Result Hyper leucine Balance in calcium homeostatis is disturbed Deposition of calcium salts as a consequence of high serum Ca2+ level (hypercalcaemia) into healthy organs. A hypercalcaemia also enhances the extent of dystrophic calcification. A metastatic calcification has nothing to do with tumor metastasis!!! It refers to the extensive calcification. NO in the same group  Causes of hypercalcaemia Increased parathyroid hormone (PTH) level in malignant tumors mousnanttumors Destruction of bone tissue in Multiple myeloma osteoid Paget disease, increased parallel synthesis and Fifty Osteoclast resorption of bone tissue activation Breast carcinoma metastasis in bone Vitamin D-related disorders /overdosage – elevated Ca2+ uptake Sarcoidosis- 1,25-dihydroxyvitamin D (calcitriol) production by alveolar macrophages Renal failure, causes retention of phosphate, D leading to component of vitamen secondary hyperparathyroidism PEH k  Hyper D vitaminosis foot Metastatic calcification of the lung on Partially I a Just started Major localisation of metastatic calcification Kidney Lung Arteries of systemic circulation Pulmonary veins Mucous membranes of stomach Anywhere in the body Morfology: Calcium salts look like in dystrophic calcification Symptoms of massive calcium deposition: e.g. impairment of kidney function, problems of breathing Regeneration of bone fractures Trauma Bleeding, coagulation (fibrin) Innate inflammation, fibroblasts PDGF Bone & cartilage progenitors (chondrocytes, osteoblasts) From periosteum & bone marrow New (type I collagen) matrix Granulation synthesis – Softtissue callus (fibrocartilaginous) Physical loading Endochondrial ossification (epiphyseal) osteoblasts calcium deposition – bony callus Spongy, -lamellar, trabe- cular bone : „remodelling”  Skeletal muscle regeneration TRAUMA myoblast Satellite cell (S) Permanent? cell/tissue, BUT S satellite cells (stem cells) If matrix (endo-, perimysium) preserved muscle fibers can fully regenerate (however, long cells, stroma is broken - fibrosis) Skeletal muscle regeneration Mp Rat m.soleus Notexin 24h (venom of the tiger snake) Mp Zoltan Takacs E Day-7 Central nucleus *