Lecture 10 – Healing: Regeneration, Repair and Fibrosis

REGENERATION VS. REPAIR

regeneration

  • replacement of injured cells by cells of exactly the same kind

  • some cell types do not readily regenerate

  • restoration of tissue function

  • only possible for minor injuries

repair

  • replacement of injured cells but not fully – space filled with fibrosis and scar tissue

  • can cause loss or reduction of normal tissue function

  • occurs when:

    • large-scale damage is sustained, OR

    • tissues that do not have the capacity for regeneration

CELL TYPES IN THE BODY

labile cells – constant regeneration, mitotically active

  • examples: skin epithelia, gut epithelia

  • regenerate readily

stable cells – normally quiescent but can regenerate if damaged

  • example: liver hepatocytes

permanent cells – terminally differentiated, no/limited regeneration

  • examples: neurons, cardiomyocytes

  • rare stem cells are key to this process

stem cells

  • undifferentiated but can differentiate into other cells

types of potency

  • unipotential – one cell type

  • multipotential – several cell types

  • pluripotential – all cell types

other properties

  • divide infrequently but can divide forever

  • generate transit amplifying cells – short, quick proliferation

  • divide asymmetrically into two daughters:

    • one replaces the stem cell

    • one forms a TAC

  • too few stem cells → tissue atrophy

  • too many stem cells → tumours

  • stem cells can be induced for use in regenerative medicine through reprogramming of cells with transcription factors → iPSCs

example 1 : labile cells – epithelia

  • epithelial barriers are labile, mitotically active, regenerate readily after injury

intestinal epithelial regeneration

  • new intestinal epithelial stem cells in crypts differentiate from stem cells at the base

  • induced by passage of bacterial endotoxin (LPS) through the barrier

  • LPS activates macrophages via binding to Toll-like receptor 4 (TLR4)

  • macrophages (M0) activate COX2 to metabolise arachidonic acid (cleaved from membrane phospholipids) into prostaglandin PGE2 (can activate inflammation)

  • leads to proliferation of stem cells into TACs

  • TACs terminally differentiate into various intestinal epithelial cells to fill the damage

example 2 : stable cells

  • stable cells such as bile duct and liver hepatocytes will proliferate to replace themselves if damaged

  • damage to cells releases DAMPs – detected by liver macrophages (Kupffer cells) via Toll-like receptors (TLRs)

  • activated Kupffer cells release cytokines and growth factors that induce hepatocyte mitosis

  • mesenchymal and hepatocyte stem cells also contribute to regeneration by division and differentiation

  • up to 70% of the liver can be lost and regenerated successfully

repair

  • occurs when:

    • tissue is lost, OR

    • both parenchymal (functional) and stromal (supportive) tissues are damaged

  • repair occurs via formation of a temporary connective tissue (granulation tissue) that resolves as a scar

  • normal function of a tissue or organ can be reduced

  • repair can work alongside regeneration in certain tissues

four phases of repair

  1. haemostasis

  2. inflammation

  3. proliferation

  4. remodelling

macrophages – regulates each step in the repair process

step 1 – haemostasis (clotting)

  • clotting arrests bleeding

mechanism

  • activation of coagulation cascade by platelets aggregating and degranulating

  • generates fibrin

  • transglutaminases cross-link fibrin to fibronectin and other ECM proteins

functions of the clot

  • temporary mechanical stability

  • barrier to microorganisms

  • barrier to prevent desiccation

  • a matrix rich in cytokines and growth factors secreted from platelets:

    • PDGF (platelet-driven growth factor)

    • TGFβ (transforming growth factor beta)

    • VEGF (vascular endothelial growth factor)

step 2 – debridement (necrotic tissue removal)

  • chemokines and growth factors are involved

step 3 – granulation tissue: matrix formation

  • fibroblasts migrate into the clot

  • differentiate into myofibroblasts when induced by:

    • PDGF and TGFβ

    • fibronectin

    • mechanical tension

myofibroblast functions

  • lay down collagen fibres to ‘plug’ the wound

  • express smooth muscle actin and contractile stress fibres

  • focal adhesions link stress fibres to extracellular fibronectin and ECM

  • contraction pulls together edges of the wound and accelerates healing

  • die by apoptosis at the end of the granulation phase

step 4 – granulation tissue: blood supply

  • blood is essential to support wound repair and remodelling:

    • provides nutrients and growth factors

    • removes waste

    • transports leukocytes

angiogenesis (new capillaries)

  • endothelial cells produce new capillaries

  • induced by proliferation and migration in response to macrophage-derived VEGF

pericytes

  • line vessels to stabilise endothelial cells

  • produced in response to PDGF

vasculogenesis

  • endothelial progenitor stem cells

  • produced in response to PDGF

granulation tissue – histology

  • contains:

    • capillaries

    • fibroblasts

    • variable amount of inflammatory cells

  • example: organising abscess wall – granulation tissue on one side, purulent exudate with haemorrhage on the other

step 5 – re-epithelialisation

  • occurs in epithelial barriers

  • damaged areas repopulated with already-present epithelial cells

two mechanisms

leap-frogging:

  • suprabasal cells loosen and ‘fall’ into the gap

epithelial-to-mesenchymal transition then MET:

  • lowest level of basal cells detach from basement membrane

  • convert into mesenchymal phenotype and migrate

  • undergo MET once in place

  • induced by EGF and TGFα to express new integrins that allow migration

  • mesenchymal cells express proteases (plasmin) to digest fibrin via expression of urokinase plasminogen activator

also:

  • epithelial stem cells can be induced to generate new epithelial cells

step 6 – remodelling

  • granulation tissue is replaced with acellular scar tissue

  • collagen accumulates for 2–3 months

  • after that, equilibrium is reached between:

    • collagen formation and deposition

    • activity of MMPs

  • disorganised collagen III fibres are replaced by parallel bundles of collagen I

  • strength of collagen continues to increase by cross-linking

HISTOLOGY OF REPAIR EXAMPLES

myocardial infarction – healing

  • numerous capillaries

  • collagen laid down to form a scar

  • non-infarcted myocardium present at far left

healing skin biopsy site – 1 week post-excision

  • skin surface re-epithelised

  • below: granulation tissue with small capillaries and fibroblasts forming collagen

  • after one month: just a small collagenous scar remains

failure of wound healing

  • wound is ischaemic

  • vasculogenic progenitor cell activity is compromised

  • broken ends of bone aren’t brought together

chronic wounds – disease

  • diabetes leads to marked atherosclerosis with arterial narrowing

  • when peripheral arteries to legs are involved → ischaemia of soft tissues and bone

  • even minor trauma leads to ulceration that heals poorly and often becomes infected

  • contributing factors: poor blood supply, oedema, poor nutrition, infection

EXCESSIVE SCARRING

causes

  • excessive inflammatory response to foreign material

  • excessive production of fibrogenic cytokines

  • prolonged presence of myofibroblasts

  • leads to excessive collagen production or defective remodelling

EXAMPLES

pulmonary fibrosis

  • lung alveolar walls thickened and filled with pink collagen

  • following autoimmune disease lasting decades

cirrhosis of the liver

  • liver injury from chronic alcoholism

  • fibrosis + regeneration of hepatocytes in nodules

  • firm, nodular appearance

keloids

  • localised to a surgical site

  • caused by overgrowth of granulation tissue and collagen III OR excess collagen I laid down during remodelling

  • has a genetic link