Endotoxin
Encyclopedia
Endotoxins are toxin
s associated with some Gram-negative bacteria. An "endotoxin" is a toxin
that is a structural molecule of the bacteria that is recognized by the immune system.
The prototypical examples of endotoxin are lipopolysaccharide
(LPS) or lipooligosaccharide
(LOS), found in the outer membrane of various Gram-negative bacteria, and are an important component of their ability to cause disease. The term LPS is often used interchangeably with endotoxin, owing to its historical discovery. In the 1800s, it became understood that bacteria could secrete toxins into their environment, which became broadly known as "exotoxin
". The term "endotoxin" came from the discovery that portions of Gram-negative bacteria themselves can cause toxicity
, hence the name endotoxin. Studies of endotoxin over the next 50 years revealed that the effects of "endotoxin" are, in fact, due to lipopolysaccharide.
LPS consists of a polysaccharide
(sugar) chain and a lipid moiety, known as lipid A
, which is responsible for the toxic effects. The polysaccharide chain is highly variable among different bacteria. Endotoxins are approximately 10 kDa in size but can form large aggregates up to 1000 kDa. Humans are able to produce antibodies
against endotoxins after exposure, but these are, in general, directed at the polysaccharide chain and do not protect against a wide variety of endotoxins. Injection of a small amount of endotoxin in human volunteers has been shown to produce fever, a decrease in blood pressure, and activation of inflammation and coagulation. Endotoxins are in large part responsible for the dramatic clinical manifestations of infections with pathogenic Gram-negative bacteria, such as Neisseria meningitidis
, the pathogens that causes meningococcal disease, including meningococcemia
, Waterhouse-Friderichsen syndrome
, and meningitis
.
Bacillus thuringiensis
is known to produce an endotoxin called delta endotoxin
. The expression of the toxin in plants is known to offer the plants resistance against various insects.
of Bacillus thuringiensis
is a protein found in crystal-like inclusion bodies next to the endospore
inside the bacteria. It is toxic to larvae of insects feeding on plants, but is harmless to humans as humans do not possess the enzymes and receptors necessary for its processing and toxicity.
s, LPS binds to the lipid-binding protein (LBP) in the serum, which transfers it to CD14
on the cell membrane, which in turn transfers it to another non-anchored protein, MD2, which associates with Toll-like receptor
-4 (TLR4).
CD14 and TLR4 are present in several immune system
cells (including macrophages and dendritic cells), triggering the signaling cascade for macrophage
/endothelial cell
s to secrete pro-inflammatory
cytokine
s and Nitric oxide
that lead to "endotoxic shock".
Other than TLR4, components of gram-negative cell wall may also activate other pathways, which may contribute to the overall endotoxic effect..
DNA
prepared from bacteria or proteins expressed from bacteria, and must be removed from the DNA or protein to avoid unwanted inflammatory responses prior to in vivo
applications such as gene therapy
.
Also, ovalbumin is contaminated with endotoxins. Ovalbumin is one of the extensively studied proteins in animal models and also an established model allergen for airway hyper-responsiveness (AHR). Commercially available ovalbumin that is contaminated with LPS can fully activate endothelial cells in an in-vitro assay of the first step of inflammation, and it falsifies research results, as it does not accurately reflect the effect of sole protein antigen on animal physiology.
In pharmaceutical production, it is necessary to remove all traces of endotoxin from drug product containers, as even small amounts of endotoxin will cause illness in humans. A depyrogenation
oven is used for this purpose. Temperatures in excess of 300°C are required to break down this substance. A defined endotoxin reduction rate is a correlation between time and temperature. Based on primary packaging material as syringes or vials, a glass temperature of 250°C and a holding time of 30 minutes is typical to achieve a reduction of endotoxin levels by a factor of 1000.
A very sensitive assay
for detecting presence of endotoxin is the Limulus Amebocyte Lysate
assay, utilizing blood from the Horseshoe crab
. Very low levels of LPS can cause coagulation of the limulus lysate due to a powerful amplification through an enzymatic cascade.
A endotoxin cause seviour diseases in human beings.
, if the immune response is severely pronounced.
Toxin
A toxin is a poisonous substance produced within living cells or organisms; man-made substances created by artificial processes are thus excluded...
s associated with some Gram-negative bacteria. An "endotoxin" is a toxin
Toxin
A toxin is a poisonous substance produced within living cells or organisms; man-made substances created by artificial processes are thus excluded...
that is a structural molecule of the bacteria that is recognized by the immune system.
Gram negative
Lipopolysaccharide
Lipopolysaccharides , also known as lipoglycans, are large molecules consisting of a lipid and a polysaccharide joined by a covalent bond; they are found in the outer membrane of Gram-negative bacteria, act as endotoxins and elicit strong immune responses in animals.-Functions:LPS is the major...
(LPS) or lipooligosaccharide
Lipooligosaccharide
Lipooligosaccharide , a potent endotoxin, causes:* Petechiae,* Purpura,* Waterhouse-Friderichsen syndrome,* Septic shockLipooligosaccharides are naturally occurring variants of the more common glycolipid, lipopolysaccharide...
(LOS), found in the outer membrane of various Gram-negative bacteria, and are an important component of their ability to cause disease. The term LPS is often used interchangeably with endotoxin, owing to its historical discovery. In the 1800s, it became understood that bacteria could secrete toxins into their environment, which became broadly known as "exotoxin
Exotoxin
An exotoxin is a toxin excreted by a microorganism, like bacteria, fungi, algae, and protozoa. An exotoxin can cause damage to the host by destroying cells or disrupting normal cellular metabolism. They are highly potent and can cause major damage to the host...
". The term "endotoxin" came from the discovery that portions of Gram-negative bacteria themselves can cause toxicity
Toxicity
Toxicity is the degree to which a substance can damage a living or non-living organisms. Toxicity can refer to the effect on a whole organism, such as an animal, bacterium, or plant, as well as the effect on a substructure of the organism, such as a cell or an organ , such as the liver...
, hence the name endotoxin. Studies of endotoxin over the next 50 years revealed that the effects of "endotoxin" are, in fact, due to lipopolysaccharide.
LPS consists of a polysaccharide
Polysaccharide
Polysaccharides are long carbohydrate molecules, of repeated monomer units joined together by glycosidic bonds. They range in structure from linear to highly branched. Polysaccharides are often quite heterogeneous, containing slight modifications of the repeating unit. Depending on the structure,...
(sugar) chain and a lipid moiety, known as lipid A
Lipid A
Lipid A is a lipid component of an endotoxin held responsible for toxicity of Gram-negative bacteria. It is the innermost of the three regions of the lipopolysaccharide molecule, and its hydrophobic nature allows it to anchor the LPS to the outer membrane...
, which is responsible for the toxic effects. The polysaccharide chain is highly variable among different bacteria. Endotoxins are approximately 10 kDa in size but can form large aggregates up to 1000 kDa. Humans are able to produce antibodies
Antibody
An antibody, also known as an immunoglobulin, is a large Y-shaped protein used by the immune system to identify and neutralize foreign objects such as bacteria and viruses. The antibody recognizes a unique part of the foreign target, termed an antigen...
against endotoxins after exposure, but these are, in general, directed at the polysaccharide chain and do not protect against a wide variety of endotoxins. Injection of a small amount of endotoxin in human volunteers has been shown to produce fever, a decrease in blood pressure, and activation of inflammation and coagulation. Endotoxins are in large part responsible for the dramatic clinical manifestations of infections with pathogenic Gram-negative bacteria, such as Neisseria meningitidis
Neisseria meningitidis
Neisseria meningitidis, often referred to as meningococcus, is a bacterium that can cause meningitis and other forms of meningococcal disease such as meningococcemia, a life threatening sepsis. N. meningitidis is a major cause of morbidity and mortality during childhood in industrialized countries...
, the pathogens that causes meningococcal disease, including meningococcemia
Meningococcemia
Meningococcal disease describes infections caused by the bacterium Neisseria meningitidis . It carries a high mortality rate if untreated. While best known as a cause of meningitis, widespread blood infection is more damaging and dangerous...
, Waterhouse-Friderichsen syndrome
Waterhouse-Friderichsen syndrome
Waterhouse-Friderichsen syndrome or hemorrhagic adrenalitis or Fulminant meningococcemia, is a disease of the adrenal glands most commonly caused by the bacterium Neisseria meningitidis. The infection leads to massive hemorrhage into one or both adrenal glands...
, and meningitis
Meningitis
Meningitis is inflammation of the protective membranes covering the brain and spinal cord, known collectively as the meninges. The inflammation may be caused by infection with viruses, bacteria, or other microorganisms, and less commonly by certain drugs...
.
Gram-positive
Results from one 1979 study indicated that Listeria monocytogenes may produce an "endotoxin-like" substance. However, a subsequent study failed to confirm that this Gram-positive species produces an endotoxin.Bacillus thuringiensis
Bacillus thuringiensis
Bacillus thuringiensis is a Gram-positive, soil-dwelling bacterium, commonly used as a biological pesticide; alternatively, the Cry toxin may be extracted and used as a pesticide. B...
is known to produce an endotoxin called delta endotoxin
Delta endotoxin
Delta endotoxins are pore-forming toxins produced by Bacillus thuringiensis species of bacteria. They are useful for their insecticidal action....
. The expression of the toxin in plants is known to offer the plants resistance against various insects.
Wider usage of the word
Endotoxins are toxins that are not secreted by cells, so they can be found within the cell and not in the surrounding medium. Toxins secreted are exotoxins. The delta endotoxinDelta endotoxin
Delta endotoxins are pore-forming toxins produced by Bacillus thuringiensis species of bacteria. They are useful for their insecticidal action....
of Bacillus thuringiensis
Bacillus thuringiensis
Bacillus thuringiensis is a Gram-positive, soil-dwelling bacterium, commonly used as a biological pesticide; alternatively, the Cry toxin may be extracted and used as a pesticide. B...
is a protein found in crystal-like inclusion bodies next to the endospore
Endospore
An endospore is a dormant, tough, and temporarily non-reproductive structure produced by certain bacteria from the Firmicute phylum. The name "endospore" is suggestive of a spore or seed-like form , but it is not a true spore . It is a stripped-down, dormant form to which the bacterium can reduce...
inside the bacteria. It is toxic to larvae of insects feeding on plants, but is harmless to humans as humans do not possess the enzymes and receptors necessary for its processing and toxicity.
Mechanism
In humanHuman
Humans are the only living species in the Homo genus...
s, LPS binds to the lipid-binding protein (LBP) in the serum, which transfers it to CD14
CD14
Cluster of differentiation 14 also known as CD14 is a human gene.The protein encoded by this gene is a component of the innate immune system. CD14 exists in two forms. Either it is anchored into the membrane by a glycosylphosphatidylinositol tail or it appears in a soluble form...
on the cell membrane, which in turn transfers it to another non-anchored protein, MD2, which associates with Toll-like receptor
Toll-like receptor
Toll-like receptors are a class of proteins that play a key role in the innate immune system. They are single, membrane-spanning, non-catalytic receptors that recognize structurally conserved molecules derived from microbes...
-4 (TLR4).
CD14 and TLR4 are present in several immune system
Immune system
An immune system is a system of biological structures and processes within an organism that protects against disease by identifying and killing pathogens and tumor cells. It detects a wide variety of agents, from viruses to parasitic worms, and needs to distinguish them from the organism's own...
cells (including macrophages and dendritic cells), triggering the signaling cascade for macrophage
Macrophage
Macrophages are cells produced by the differentiation of monocytes in tissues. Human macrophages are about in diameter. Monocytes and macrophages are phagocytes. Macrophages function in both non-specific defense as well as help initiate specific defense mechanisms of vertebrate animals...
/endothelial cell
Endothelium
The endothelium is the thin layer of cells that lines the interior surface of blood vessels, forming an interface between circulating blood in the lumen and the rest of the vessel wall. These cells are called endothelial cells. Endothelial cells line the entire circulatory system, from the heart...
s to secrete pro-inflammatory
Inflammation
Inflammation is part of the complex biological response of vascular tissues to harmful stimuli, such as pathogens, damaged cells, or irritants. Inflammation is a protective attempt by the organism to remove the injurious stimuli and to initiate the healing process...
cytokine
Cytokine
Cytokines are small cell-signaling protein molecules that are secreted by the glial cells of the nervous system and by numerous cells of the immune system and are a category of signaling molecules used extensively in intercellular communication...
s and Nitric oxide
Nitric oxide
Nitric oxide, also known as nitrogen monoxide, is a diatomic molecule with chemical formula NO. It is a free radical and is an important intermediate in the chemical industry...
that lead to "endotoxic shock".
Other than TLR4, components of gram-negative cell wall may also activate other pathways, which may contribute to the overall endotoxic effect..
Endotoxin contamination
Endotoxins are frequent contaminants in plasmidPlasmid
In microbiology and genetics, a plasmid is a DNA molecule that is separate from, and can replicate independently of, the chromosomal DNA. They are double-stranded and, in many cases, circular...
DNA
DNA
Deoxyribonucleic acid is a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms . The DNA segments that carry this genetic information are called genes, but other DNA sequences have structural purposes, or are involved in...
prepared from bacteria or proteins expressed from bacteria, and must be removed from the DNA or protein to avoid unwanted inflammatory responses prior to in vivo
In vivo
In vivo is experimentation using a whole, living organism as opposed to a partial or dead organism, or an in vitro controlled environment. Animal testing and clinical trials are two forms of in vivo research...
applications such as gene therapy
Gene therapy
Gene therapy is the insertion, alteration, or removal of genes within an individual's cells and biological tissues to treat disease. It is a technique for correcting defective genes that are responsible for disease development...
.
Also, ovalbumin is contaminated with endotoxins. Ovalbumin is one of the extensively studied proteins in animal models and also an established model allergen for airway hyper-responsiveness (AHR). Commercially available ovalbumin that is contaminated with LPS can fully activate endothelial cells in an in-vitro assay of the first step of inflammation, and it falsifies research results, as it does not accurately reflect the effect of sole protein antigen on animal physiology.
In pharmaceutical production, it is necessary to remove all traces of endotoxin from drug product containers, as even small amounts of endotoxin will cause illness in humans. A depyrogenation
Depyrogenation
Depyrogenation refers to the removal of pyrogens from solution, most commonly from injectable pharmaceuticals.A pyrogen is defined as any substance that can cause a fever. Bacterial pyrogens include endotoxins and exotoxins, although many pyrogens are endogenous to the host...
oven is used for this purpose. Temperatures in excess of 300°C are required to break down this substance. A defined endotoxin reduction rate is a correlation between time and temperature. Based on primary packaging material as syringes or vials, a glass temperature of 250°C and a holding time of 30 minutes is typical to achieve a reduction of endotoxin levels by a factor of 1000.
A very sensitive assay
Assay
An assay is a procedure in molecular biology for testing or measuring the activity of a drug or biochemical in an organism or organic sample. A quantitative assay may also measure the amount of a substance in a sample. Bioassays and immunoassays are among the many varieties of specialized...
for detecting presence of endotoxin is the Limulus Amebocyte Lysate
Limulus Amebocyte Lysate
Limulus amebocyte lysate is an aqueous extract of blood cells from the horseshoe crab, Limulus polyphemus. LAL reacts with bacterial endotoxin or lipopolysaccharide , which is a membrane component of Gram negative bacteria...
assay, utilizing blood from the Horseshoe crab
Horseshoe crab
The Atlantic horseshoe crab, Limulus polyphemus, is a marine chelicerate arthropod. Despite its name, it is more closely related to spiders, ticks, and scorpions than to crabs. Horseshoe crabs are most commonly found in the Gulf of Mexico and along the northern Atlantic coast of North America...
. Very low levels of LPS can cause coagulation of the limulus lysate due to a powerful amplification through an enzymatic cascade.
A endotoxin cause seviour diseases in human beings.
Endotoxemia
The presence of endotoxins in the blood is called Endotoxemia. It can lead to septic shockSeptic shock
Septic shock is a medical emergency caused by decreased tissue perfusion and oxygen delivery as a result of severe infection and sepsis, though the microbe may be systemic or localized to a particular site. It can cause multiple organ dysfunction syndrome and death...
, if the immune response is severely pronounced.