Where’s the Aspirin?

Aspirin is the traditional anti-inflammatory agent. Many of us grew up with the quintessential doctoring phrase, “Take two aspirin and call me in the morning.” Aspirin stops inflammation in several ways. Like all drugs, it interacts with many different proteins/enzymes. In fact it interacts so intimately with the inflammatory system that it suggests that the process of inflammation may require an aspirin-like molecule to function normally.

Aspirin Binds to Multiple Enzymes of Inflammation

Aspirin is observed to reduce inflammation. That means that ingested aspirin tablet dissolve in the stomach and pass through the intestines into the blood stream and subsequently bath cells of the blood and the endothelium that lines the blood vessels. In order to reach the blood stream, the aspirin must pass through the intestinal cells. That passage requires binding to a protein transport molecule.

Cells responding to an inflammatory signal (NFkB, transcription factor is activated) synthesize enzymes that release unsaturated fatty acids (ARA, EPA, DHA) from membrane phospholipids (PLA2, phospholipase A2), form a cyclic epoxide from the fatty acid (prostaglandin H2 synthase 2, also called cyclooxygenase 2, COX-2).

Aspirin binds to the inhibitor that normally inactivates NFkB and prevents NFkB activation that is required for inflammation. Aspirin also binds to PLA2 and prevents fatty acid release and thereby blocks activation of inflammation. Aspirin also binds to COX-2 and blocks the production of inflammatory prostaglandins from ARA. But that is not all that aspirin does.

Inflammation Resolution Uses Aspirin-COX-2 Interaction

The strange interaction that makes aspirin suspicious is that aspirin doesn’t just interfere with the action of enzymes, it subtly changes their specificity. Thus aspirin chemically transfers its acetyl group (CH3-COOH-) to an amino acid in the active site of COX-2 to produce a new group of anti-inflammatory lipoxins from ARA, EPA and DHA.

This raises the question of whether aspirin is a natural dietary modulator of inflammation. Recall that aspirin was initially obtained from willow (Salix) bark. Unfortunately, the data are conflicting. Initial research indicated that grains (naturally inflammatory) lacked aspirin, but many herbs, spices and leafy vegetables (naturally anti-inflammatory) contained aspirin. Subsequent tests refuted this work. It would be consistent with observations that some dietary components are anti-inflammatory, but candidate acetyl donors have not been identified.

Speculative acetyl candidates may include the menthol relatives, such as menthyl acetate (figure). Peppermint oil, which contains mostly menthol with some menthyl acetate, is more effective in the treatment of inflammatory bowel disease than most pharmaceuticals prescribed to treat the condition. This anti-inflammatory activity may be due in part to the aspirin-like chemical structure and function of the menthyl acetate. Also note that acetic acid/vinegar is sometimes suggested as a cure-all. This activity may be a consequence of its formation of ester linkages with alcohols that have structures similar to menthol.

Large Dose Aspirin as Cancer Treatment

The potent anti-inflammatory effects of aspirin have been compromised, because inflammation is an essential developmental activity. Thus, the integrity of the gut, for example, requires modest production of inflammatory prostaglandins and a pill of aspirin can disrupt gut tissue. Large doses of aspirin cannot be given orally. Intravenous administration of large doses of aspirin, however, is possible and the impact on process that require inflammation is dramatic.

Anecdotal evidence indicates that large dose aspirin is able to disrupt cancers, because proliferation of cancer cells requires NFkB activation and other inflammatory responses. High doses of aspirin also cause other potentially dangerous complications, such as short-circuiting oxidative phosphorylation of mitochondria and increasing nitric oxide free radical production. Still, the impact of high dose aspirin on some diseases is so amazing that it is being actively and carefully pursued.

Bee Sting Allergy

Typical. I started to write an article on leukotrienes, the inflammatory derivatives of the omega-6 fatty acid, arachidonic acid, but ran across another powerful example to test my hypothesis to explain the cause of allergies. The leukotriene article will have to wait till another day.

Wikipedia is my source of choice for up-to-date summaries of biomedical information. I queried “leukotrienes” and immediately ran across the original name for these inflammatory compounds, “slow reacting substance of anaphylaxis”. I was initially distracted by the classic experimental use of snake venom and histamine to induce leukotriene production. Snake venom has the same enzyme, PLA2, as brown recluse spider venom (subject of a previous article) and honey bee venom, that releases arachidonic acid (ADA). ADA is an omega-6 fatty acid that is the starting material for inflammatory prostaglandins and leukotrienes.

The mention of honey bee venom in the Wikipedia article on anaphylaxis sent me on a quick check of the structure and sequence of the honey bee allergen. I initially found that the major allergen is a hyaluronidase. I quickly searched for a three amino acid sequence that I predicted would make it an allergen. It was just where I expected to find it. About two thirds of the way along the amino acid sequence I found, -TTSRKKVLP-. Three basic amino acids together, in this case -RKK-, argininine-lysine-lysine, form a strong heparin-binding domain, that I believe takes proteins into cells and during inflammation primes the immune system for allergic responses.

I have found the same strong heparin-binding domain associated with allergens of ragweed, dust mites and peanuts. The principal autoantigens of autoimmune diseases, such as lupus, celiac, etc. also display the same unusual sequences. In lupus, or example, nuclear proteins with the internalization signal provided by nucleic acid-binding domains (and nuclear localization signas) are autoantigens. This pattern is found with all allergens that I have examined. There are a few apparent exceptions, but in all of these cases, there is a closely related allergen from a related source that has the expected strong heparin-binding domain. It appears that in these cases, the less common allergen provides the initial exposure during the presentation phase of high inflammation, and the allergy is maintained by subsequent exposure to the more common allergen. After the establishment of the allergy, the strong heparin-binding domain is no longer needed, because antibodies bind to other parts of homologous allergenic proteins for internalization.

Just for fun, I have illustrated the honey bee allergen, hyaluronidase, to show both its strong heparin-binding domain (blue) along with its substrate hyaluronan (grey and red). Note that the substrate sugars are in the slot of the active site, which is lined with orange and yellow aromatic amino acids that provide flat, hydrophobic binding platforms for each sugar.

After this little distraction to provide further support for my explanation of the cause of allergies, I have to get back to looking at the role of leukotrienes in anaphylaxis, COPD, asthma and other inflammatory diseases.

Spider Inflammation

Happy Halloween! It's time for creepy, crawly things that bite... and cause inflammation.

The brown recluse has the bite that keeps on giving. Its venom contains a phospholipase that cleaves membrane phospholipids to release arachidonic acid (omega-6 fatty acid precursor of inflammatory eicosanoids) and lysophosphotidylcholine that binds to multiple receptors to signal NFkB-based inflammation.

It’s Halloween and a time to talk about things that bite and sting. After all, the symbol of Halloween is the Jack-O-Lantern, the flame within, i.e. inflammation. Venom is Nature’s answer to the need for instant inflammation.

The brown recluse is adapted for hunting, killing and eating. The killing bite is the subject tonight. The recluse injects a venom designed to get the biggest bark for the bite. It uses the least amount of protein to immobilize its prey/meal. Of course, since spiders are suckers, it helps if the venom also liquifies its meal.

The venom also has a secondary use as a defense. In this case, if something too big to eat encounters a brown recluse, the goal is to encourage the big beast, you or me, to leave. No pain, no gain.

Venom provides a convergence of enhanced meal making and avoidance of being made into a meal. The answer is, of course, enzymatic inflammation. Venom contains enzymes that convert common cellular components into potent pyrogens, inflammation inducers. The wicked witch of the world of inflammation is phospholipase A2 (PLA2).

Brown recluse venom PLA2 clips off the arachidonic acid from membrane phospholipids and leaves an amphipathic detergent, lysophospholipid (LPL). The LPL can destabilize membranes and rupture cells. It is also a potent activator of several cell receptors that signal inflammation via NFkB. The COX-2 induced by inflammation has a readymade substrate present, arachadonic acid, and produces inflammatory prostaglandins, that continue the signaling frenzy. The result is inflammation, pain and necrosis.

Recluse bites are hard to treat. I think that some people recommend topical application of gunpowder. I guess that would make sense, because one of the problems of recluse bites is actually constriction of blood flow by other venom components, and nitroglycerin patches apparently provide some relief. Both the patches and nitrates of the gunpowder cause vasodilation, and would remedy some of the circulation problems of the venom. Glucocorticoids are also partially effective, since they are anti-inflammatory. Then there is heparin. I would always try heparin, at least as a last resort, because heparan sulfate proteoglycans are an essential component of most extracellular signaling systems, e.g. cytokines. Heparin should disrupt or activate all of these systems. It is unpredictable. It turns out that heparin is helpful against the brown recluse. I would also like to try a topical potion made of Vick’s Vaporub (menthol, eucalyptol, camphor, turpentine), castor oil and heparin. That should fix about anything. It even smells good, sooths painful joints and makes your skin soft.

Happy Halloween. Enjoy trick or treating, but watch where you put your hands.