26. Repair and Regeneration Flashcards
Why do we need to understand repair?
- normal oral tissue development and function helps us understand abnormal control and disease response
- cells, genes and molecules control response to development, structure and function AND ageing, injury and disease so repair can be a treatment between
What is regenerative medicine?
- to develop novel therapies to repair and regenerate tissues and organs
- which have been damaged by injury, ageing, cancer, disease
Define ‘repair’
restoration of tissue function but with impaired tissue architecture
Define ‘regeneration’
- complete restoration of tissue architecture and function
Problem with regenerative medicine
- full regenerative capacity is lost in humans
- current use of organ transplants and artificial devices is limited to incomplete restoration of original tissue function
Solutions to improve regenerative medicine
- cellular therapy (using exogenous stem/progenitor cells or stimulating own body’s stem cells to repair defective tissue)
- tissue engineering (biomaterials)
- biomedical engineering
- gene therapy
Aims for regenerative medicine
- better clinical outcomes (shorter rehab, better QOL)
- improved health-related cost-effectiveness
List stages of tissue regeneration
- morphallaxis
- epimorphosis
- compensatory regulation
- stem cell-mediated regeneration
What happens in morphallaxis?
- repatterning of existing tissue with little new growth
- for example Hydra (freshwater polyp, 0.5cm, sticks to rocks, filter feeder, asexual reproduction)
- all cells are constantly dividing and migrating and eventually shed at head or foot region - asexual reproduction budding at 2/3 body axis
- morphogen gradients specifying head and foot - each piece of cut hydra will form a small hydra with head and foot
What happens in epimorphosis?
- dedifferentiation of cells at wound site
- formation of undifferentiated cells that redifferentiate to form lost structure
- for example planarian flatworms, amphibian limbs
What happens in compensatory regulation?
- differentiated cells divide
- they maintain their identity and specialised functions
- e.g liver regeneration
Explain how the liver regenerates using compensatory regulation
- sensing of liver damage by increase of gut-derived LPS in blood
- activates Kupffer and stellate cells
- paracrine secretion of mediators stimulates hepatocyte cell proliferation
- size of liver restored 1 week after surgery (mice)
- chronic injury leads to liver fibrosis
What happens in stem cell-mediated regeneration?
- replacement of lost tissue by stem cell activity
e.g - hair growth from follicular stem cells in hair bulge - concept of ‘niche’
- continuous blood cell replacement by haematopoietic stem cells
What are stem cells?
- unspecialised, undifferentiated cells that can self-renew and can differentiate into other cell types
- for development and regeneration
Totipotent stem cells can …
- form all cell types
e.g fertilised eggs
Pluripotent stem cells can form …
- all cell types of the three embryonic germ layers
- e.g embryonic stem cells
Multipotent stem cells can form …
- many cell types
- e.g haematopoietic stem cells or mesenchymal stem cells
Oligopotent stem cells can form ..
- few cell types
- e.g myeloid precursors that form five blood cell types
Quadripotent stem cells form …
4 cell types
e.g mesenchymal progenitor cells (cartilage, bone, stroma, fat)
Unipotent stem cells can form …
one cell type
e.g mast cell precursors
Problem with stem cells in regenerative medicine
- stem cell biology not fully understood
- e.g small number, quiescence, niche, identification, genetic control
What’s a potential use of stem cells that is crucial to dentistry?
for missing teeth
Steps of wound healing in oral mucosa or after tooth extraction
- haemostasis
- inflammatory response
- epithelial and connective tissue repair
Repair of periodontal tissues is …
- more complex
- restoration of funtional unit
Why do we need a wound healing system in oral mucosa?
- protective and barrier function of oral mucosa
- effective repair system to re-establish function after injury (physical, chemical, radiation, microorganisms)
Explain haemostasis in wound healing
- essentially the cessation of blood loss
- happens in minutes
- vascular damage causes hemorrhaging into tissue defect and results in forming of a blood clot (coagulation - fibrin deposition and aggregation of platelets)
- forms barrier that unites wound margins and protects exposed tissue
- provides provisional scaffold for subsequent colonisation by reparative cells
What happens in the inflammatory response in wound healing?
- microorganisms and toxins have likely entered and induce acute inflammatory response
- leakage of plasma proteins (vasodilation and increased vascular permeability) and platelet-derived cytokines and growth factors (TGF-beta, PDGF) stimulate leukocyte migration towards wound (by chemotaxis)
- neutrophils appear within hours, become activated and kill bacteria (destroy damaged tissue)
- macrophages and lymphocytes appear after 24 hrs and mediate clearance of cell debris (phagocytosis) and humoral immune response
- mast cells promote inflammation and vascular changes
Explain reparative phase/epithelial response in wound healing
- mobilisation of epithelial cells (widening of intercellular spaces) within 24 hours
- increased basal cell proliferation and epithelial cells adjacent to wound margin migrate beneath blood clot (24-48 hrs)
- deposition of basal lamina components facilitates cell movement and epithelial sheet formation
- migration stops when cells reach opposing wound margin and increased cell proliferation and differentiation leads to stratification and re-establishment of normal epithelial sheet
Explain reparative phase/connective tissue response in wound healing
- fibroblasts proliferate and migrate into wounded connective tissue within 24-48 hrs
- they deposit disorganised collagen fibres, regulated by TBF-beta
- formation of new blood capillaries from existing vessels (angiogenesis) at wound margin (regulated by VEGF, FGF, TGF-beta)
- ECM (fibronectin, laminin, collagen) formed by new fibroblasts provides scaffold for forming blood vessels - provides nutrients and oxygen, access to inflammatory cells, stimulates connective tissue formation (endothelial cytokines/growth factors)
- increased collagen deposition between day 5-20 but reduced tensile strength - initial scar tissue formation but removal of scar tissue by tissue remodelling within 150 days
Scar tissue formation in wound healing
- quick restoration of tissue integrity required to prevent damage to whole organism
- trade-off with inflammatory response - control of infection allows quick wound healing but produces scar tissue of inferior quality
- in skin, deposition of disorganised collagen fibres leads to immobilisation and rigidity at repair site
- in oral mucosa, remodelling of scar tissue (mainly collagen fibre remodelling and cross-linking) prevents scar tissue formation
How does foetal wound healing differ?
- repair of skin injuries doesn’t involve inflammatory response and results in scar-less healing in foetal development
- fibroblasts of oral mucosa may resemble these foetal fibroblasts
Explain wound contraction in wound healing
- first fibroblasts entering the wound site are contractile myofibroblasts
- different from connective tissue fibroblasts (pericyte origin?)
- form connections with each other and align with collagen fibrils
- cell contraction draws edges of wound together
Wound healing after tooth extraction compares to that of oral mucosa except for …
- substantial loss of tissue in extraction
- dislodgement of blood clot (‘dry socket’) causes painful bone infection
Complete epithelialsation of socket occurs in …
What happens then?
- 10 days
- instead of fibroblasts, osteogenic precursor cells migrate into blood clot
- after day 10, osteoblasts differentiate and form bone
- extraction site is filled with bone and indistinguishable after 10-12 weeks
How does wound healing at the dento-gingival junction differ to in mucosa?
- day 3, colonisation of gingival wound by epithelial cells and formation of junctional epithelium
- cells express REE-marker ODAM (odontogenic ameloblast-associated protein)
- day 5-7, expansion and down-growth of junctional epithelium
- re-establishment of dento-gingival junction (high regenerative capacity in rodents especially)
How does repair of periodontal tissue occur?
- restoration of functional unit like cementum, PDL, alveolar bone, gingiva
- different to oral mucosa as PDL fibres must insert to cementum and bone
- coordinated repair needs complex regulation at cellular and molecular level
- normal remodelling has no inflammation e.g tooth movements
- in injury, inflammatory response required to combat infection and initiate repair
- chronic inflammation inhibits stem cell activation, cell recruitment, cell proliferation, differentiation
Fibroblasts in periodontal repair
- key to remodelling collagen fibres
- mesenchymal progenitor cells in PDL or perivascular and endosteal fibroblasts
Endothelial cells in periodontal repair
form new blood vessels from existing vessels (angiogenesis)
Cementoblasts cells in periodontal repair
perivascular and endosteal fibroblasts
and/or
rest of Malassez
Osteoblasts cells in periodontal repair
mesenchymal progenitor cells in endosteum or periosteum
Molecular approaches to periodontal repair
- application of growth factor cocktails and ECM molecules to root surfaces
- like EGF, FGF, IGF, PDGF, TBG-beta for PDL and cementum
- BMP for bone and cementum
- fibronectin but clinical efficiency is controversial
- emdogain as enamel matrix proteins stimulate periodontal repair
Can we get complete regeneration of enamel?
- no
- as ameloblasts are lost at the end of development
- can be remineralised by calcium, phosphate and fluoride ions in saliva
- acts as a physico-chemical repair process
What is the dynamic process of an early caries lesion?
- translucent zone - demineralisation
- dark zones - remineralisation
- body of the legion - enamel destruction
- surface zone - intact enamel (remineralisation caused by ion precipitation from saliva)
When is an early caries lesion reversible?
- if surface enamel remains intact and acid producing bacteria are removed
Dentine is a living tissue so the reparative process depends on …
- extent and duration of stimulus e.g attrition/cavity prep
- structural variations in dentine e.g open or accluded dentinal tubules
- age of tooth - smaller pulp chamber, diminished blood/nerve supply
What happens to repair dentine in a slow onset, prolonged insult?
Examples of this
- for example in attrition, early caries
- occlusion of dentinal tubules (collagen plug or sclerotic dentine)
- reactionary dentine formed by existing odontoblasts (slow, tubular)
What happens to repair dentine in a rapid onset, severe insult?
Examples of this
e.g in late stage caries or cavity prep
- reactionary dentine if odontoblasts survive (slow, tubular)
- reparative dentine form by newly differentiated odontoblast-like cells if originals have died (rapid, amorphous, less collagen)
- classic wound healing response - inflammation and repair but no epithelial response
Most accessible sources of stem cells for dental regeneration
- primary teeth
- 3rd molars
List sources of dental stem cells
- primary teeth and 3rd molars
- dental pulp (DPSC)
- dental pulp from exfoliated primary teeth (SHED)
- dental follicle of unerupted teeth
- periodontal ligament (PDLSC)
- tooth germs
Other sources of stem cells in body
- umbilical cord blood
- bone marrow
- skin
- adipose tissue
What animal tooth is used as a model for studying tooth regeneration?
mouse lower incisor
How can you bioengineer replacement teeth?
- add epithelial and mesenchymal cells to a collagen gel to form a high-density reconstituted tooth germ
- organ culture for 5-7days
- extraction and wound healing
- transplantation of the tooth germ, grows tooth
How could tissue engineering help dentistry?
- biodegradable scaffolds for cell seeding - 3D printing
- bioengineering, material sciences and nanotechnology - materials with novel properties, implants with bioactive surfaces allowing for better tissue integration
- gene therapy - ex and in vivo (CRISPR/Cas9 genome editing tools with RNAi, novel delivery systems)
Impact of tissue engineering on dentistry
- conventional treatments like amalgam, composites, metallic implants, tissue grafts
- have limitations like non-biological, immune rejection, pathogen transmission, lack of remodelling with recipient tissue, donor site morbidity
- novel approaches like engineering of precise tissue shape using biodegradable scaffolds onto which stem cells can grow and re-establish morphology and function
- need more translation of basic science findings like clinical trials
What did Mao say about this?
‘craniofacial tissue engineering is likely to be realised in the foreseeable future and represents an opportunity that dentistry cannot afford to miss’
