Endotoxins Flashcards
Structure of a gram negative bacterial cell wall
- Peptidoglycan layer is thin and contains an outer membrane which has a variety of components
- Proteins
- Porins - allow movement of materials
- Lipid layer
- Lipopolysaccharide (this is the endotoxin) and is an integral part of the bacterial cell wall.
Structure of the lipopolysaccharide (Endotoxin)
- Lipid A.: phosphorylated glucosamines attached to long chain fatty acids. The no. and type of FA varys by species. Hydrophobic helps repel water soluble materials.
- Polysaccharide core: ketodeoxyoctanoic acid (KDO) and heptose. relatively constant between species. Hydrophilic
- O-side chain: repeating units of tri, tetra or pentasaccharide sugars. Highly variale between species. Hydrophilic
What is an endotoxin?
Lipopolysaccharide
- The Lipid A is the active component -> Not immunogenic
- O antigen is highly immunogenic and immune specific
- Found only in gram negative bacteria
- Heat stable (exotoxins are heat unstable)
- Not converted to toxoids (unlike exotoxins)
- Major initiator of the sepsis pathway
What is sepsis?
Life threatening organ dysfunction caused by a dysregulated response to infection
What cells drive sepsis?
Primarily the innate immune system:
- Macrophages
- Monocytes
- Granulocytes
- Natural Killer Cells
- Dendritic cells
What do the innate immune cells detect in sepsis?
- Pathogen associated molecular patterns (PAMPs) such as endotoxins.
- Damage associated molecular pattenrs (DAMPs) from damaged host cells.
What mediates the detection of sepsis?
- Cell membrane receptors: toll-like receptors (TLR) and C-type lectin receptors.
- Cytosol receptors: NOD-like receptors, RIG-I-like receptors
What is the effect of the innate immune system response?
- Production of pro-inflammatory cytokines TNFalpha, IL-1, IL-6.
- via inflammasomes to produce IL-1beta and IL-18 that cause rapid programmed cell death
What type of marker is Endotoxin?
PAMP as it is a marker to the innate immune system cells.
What are DAMPs?
Damaged host cells which have been damaged by an infection and detected by the innate immune system.
Action of the Endotoxin
- Binds to CD14 receptor on the surface of a macrophage. This will have downstream effects.
- Once the endotoxin has bound to the receptor, it will be transported to the toll-like receptor 4 via a molecule called MD2.
- This causes dimerisation of two TLR4 receptors.
- This will result in myddosome formation and activation of Nf-kB.
- This allows for production of TNF-alpha and other cytokines.
What are the effects of pro-inflammatory cytokines?
- Increase number, lifespan and activation state of innate immune cells.
- Increase adhesion molecule and chemokine expression by endothelial cells.
- Increase acute phase protein such as complement, fibrinogen and CRP (measure of inflammation).
- Cause fever
- Causes neutrophils to release extracellular traps (NETs) made of DNA and antimicrobial proteins that forms a scaffold for platelet activation.
- Cause release of microparticles by activated platelets
- Increase tissue factor expression by blood monocytes
- > 5+6+7 -> formation of a thrombus (immunothrombosis) - microbes trapped within this -> attracts and activate further leucocytes.
Sepsis dysregulation
Normally, the effects of pro-inflammatory cytokines will achieve rapid control of localised and minor infections. However, the process may pass a threshold meaning systemic injury occurs everywhere in the body.
Summary: when a threshold level is passed, sepsis dysregulation will occur and systemic injury throughout the whole body.
What processes occur in sepsis dysregulation?
- There is production of reactive oxygen species (ROS) - Hydroxyl and nitric oxide - that damage the cellular proteins, DNA and lipids and impairs the mitochondria. It causes non-specific damage.
- There is complement activation (especially C5a) that increases ROS, granulocyte enzyme release, endothelial permeability and tissue factor expression.
- There is widespread immunothrombosis (clots forming all over the body) which will lead to disseminated intravascular coagulation (DIC) with impaire microvasculature function and organ dysfunction.
- Mitochondrial damage leads to decreased intracellular ATP and cells enter a state of hibernation - exacerbates organ dysfunction.
Sepsis Resolution
- Early in the sepsis process, IL-10 is produced. This acts as an anti-inflammatory cytokine which will suppress pro-inflammatory cytokine IL-6 and gamma interferon.
- It will also stimulate production of soluble TNF receptors (causing TNF to bind to this rather than the cells avoiding cell necrosis),
- The IL-1 receptor antagonists dampen the immune system.
- There is autophagy of PAMPs and DAMPs removing the stimulus.
- The damaged cells undergo apoptosis and engulfment by macrophages.
- Antibiotics can be used to treat the bacteria however, signs of sepsis can still be shown long after because the process of resolution occurs for a while.
