Endotoxin

Endotoxins (not to be confused with exotoxin) are toxins 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.

Gram negative
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.

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 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.

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 endotoxin 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.

Mechanism
In humans, 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 cells to secrete pro-inflammatory cytokines 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..

Endotoxin contamination
Endotoxins are frequent contaminants in plasmid 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.

Endotoxemia
The presence of endotoxins in the blood is called Endotoxemia. It can lead to septic shock, if the immune response is severely pronounced.