Proteases and RONS Flashcards
What are proteases?
What roles do they play in general?
Proteolytic enzymes:
Produced by tissue cells and activated leukocytes, and can circulate as zymogens - inactive, pro-enzyme form.
Host defence and inflammation - destroying pathogens, removing damaged tissue, and initiating tissue repair.
MS and rheumatoid arthritis.
What are the plasma proteases?
Generate inflammatory mediators from inactive, plasma precursors:
Bradykinin - cardinal signs of inflammation.
Complement -
C5a for chemotactic for neutrophils + leukocyte activation.
C3a + C5a for mast cell degranulation.
C3b and C5-C9 (MAC) for bacterial opsonisation and lysis.
Clotting cascade:
platelet activation and thrombosis:
Fibrinogen to fibrin
-prevent microbe invasion, and initiate tissue repair and complement activation.
What are the proteolytic enzyme families?
MMPs
Serine proteinases
Cysteine proteinases
What are MMPs?
MMP-1 is a collagenase.
to degrade collagen.
MMPs break down other matrix proteins and CARTILAGE!
Active at physiological pH but require a critical Zn2+ residue for activation.
Endogenous TIMPS inhibit MMPs.
- Need to have good balance of TIMPs to prevent MMps causing excessive matrix damage.
What are the Serine and Cysteine proteinases?
Released by NEUTROPHILS!
neutrophil elastatse, cathepsin G, Kallikrein.
active at physiological pH.
Break down matrix proteins - laminin, elastin, proteoglycans in joint cartilage.
SERPINS - body endogenous inhibitors.
but oxidation readily inactivates SERPINS.
Role of Proteases in inflammation?
Can degrade tight junctions of epithelial cells = aid WBC migration into tissue and increase plasma extravasation and permeability.
What is the role of proteases in RA?
Cartillage proteoglycans are rapidly destroyed in Rheumatoid arthritis but cartilage is lost more slowly.
This leads to impaired shcok absorption in joints and loss of joint function and causes chronic pain.
Collagenase MMps slowly degrade collagen in cartillage - cartillage loses function as a smooth surface.
Proteoglycans have open structure = highly accessible and vulnerable to proteinase degradation.
What is role of proteinases in MS?
Break down of BBB - high levels of CNS MMPs.
Break down of BBB causes infiltration of leukocytes and release of MMPs causes myelin and axonal damage.
MMPs cleavage of myelin can generate immunogenic myelin peptides which can activate autoimmune T cell responses to myelin.
How are MMPs activated?
RONS or removal of pro-peptide domain by other proteases like plasmin.
Cysteine switch mechanism with pro-peptide domain linked with Zn2+ residue in catalytic domain.
When this interaction is disrupted - by proteolytic cleavage of propeptide domain or modification by RONS - active site becomes accessible.
How to inhibit MMPs?
TIMPS are endogenous MMP inhibitors.
Peptidomimetics:
Small hydroxamate based molecules:
Zinc binding based on collagen structure = lack specificity, so act on most metalloenzymes.
Non-peptide hydroxamates are more selective:
NOTHING USEFUL so far!
Tetracycline derivatives at sub-antibiotic doses have shown to reduce MMp synthesis and activity = doxycycline for skin disease.
monoclonal antibodies for high speicificty!!
Upregulation of TIMPS!!
(limited selecitivity)
What are RONS?
Essential to host defence:
Produced by tissue cells and infiltrating leukocytes.
NADPH catalyses superoxide O2- formation, dismuted to form H202.
H202 further metabolised to form HCLO.
H202 inactivated by catalase…
RNS formed from NO combined with oxygen speciesi..
= Highly toxic to bacteria but also cause bystander damage to host cells…
What are the roles of RONS in host defence?
RONS within phagolysosomes involved in destroying bacteria + phagocytosed material.
Direct toxicc damage by lipid peroxidation, protein oxidation, DNA damage + apoptosis/necrosis.
Influence inflammatory gene transcription - lead to IL-1 and TNF-a production through NFKb TF.
Activation of proteases!!
How do RONS affect proteases?
Inactivation of SERPINS = removing endogenous inhibition of Serine/Cysteine proteases.
Activation of MMPs by latent modification to pro-peptide domain to disrupt thiol-group Zn2+ blockade of catalytic site.
What are the rles of RONS in RA?
Increased RONS and oxidant damage to lipids, proteins + DNA.
Reduced antioxidant status - reduced VitC, Catalase…
Oxidative stress can occur in dysregulated cycles of perfusion and hypoxia in rheumatoid synovial fluid.
Lead to generation of superoxide and RONS.
= cause tissue injury and activation of MMPs like elastase.
- damage to resident cells like osteoclast, chondocytes…
How cna diet affect RA?
Increased dietary anti-oxidants through pomegranate extract resulted in reduction in joint swelling + pain
