Psoriasis, IL-17, Cathelicidin, TLRs, NFkB, Inflammation and Heparin Therapy

Host DNA Released by Keratinocyte Apoptosis Binds LL-37 and Activates Dendrocytes

Psoriasis is an inflammation of the skin that leads to overproduction of keratinocytes resulting in a thick crust.  Skin inflammation, in this case, is considered a result of autoimmunity, but an autoantigen has not been identified.  It is more likely that psoriasis results from an autoinflammatory condition, in which inflammation produces a complex of self molecules that mimic bacterial DNA and trigger TLR/NFkB inflammation signaling.  And of course, if this is going to be interesting, it has to involve heparin.

Vitamin D Binds to a Transcription Factor Receptor that Controls Antimicrobial Peptides
A significant component of the innate immune system is a group of antimicrobial peptide  (defensins, cathelicidins, e.g. LL-37).  These short polypeptides owe their natural antibiotic activity to numerous basic (positively charged, arginine and lysine) amino acids.  The transcription factor that controls the expression of these peptides is the vitamin D receptor.  Thus, various forms of vitamin D influence the amount of antimicrobial peptides produced in the mouth, skin and crypts of the intestinal villi.  Oral vitamin D3 would be expected to directly improve defensin production in the gut and LL-37 production in the skin.

IL-17 Stimulates Skin Inflammation and LL-37 Production
A specific group of lymphocytes, called T helper 17 cells, produce IL-17.  These Th17 cells accumulate in some sites of inflammation, such as psoriasis and their secretion of IL-17 is associated with ongoing inflammation and may contribute to LL-37 production, as well as apoptosis of keratinocytes in the thickening skin of psoriasis plaques.
http://www.ncbi.nlm.nih.gov/pubmed/19623255?ordinalpos=1&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_SingleItemSupl.Pubmed_Discovery_PMC&linkpos=2&log$=citedinpmcarticles&logdbfrom=pubmed

Th17 Cells Are Produced in the Gut in Response to Segmented Bacteria
One of my readers brought to my attention an article that shows that one of the hundreds of species of gut bacteria, segmented filamentous baceria, stimulates the gut to develop T helper 17 cells that subsequently migrate to sites of inflammation.
http://www.medpagetoday.com/Gastroenterology/InflammatoryBowelDisease/16472
This emphasizes the link between the gut and inflammatory diseases and parallels other examples of gut influence on disease, such as the ability of Helicobacter pylori to affect asthma or parasitic worms to tame Crohn’s disease, allergies and asthma.

Inflammation Lowers Heparan Sulfate Production and Spreads LL-37
One of my students induced inflammation in cells in vitro and showed by quantitative PCR that genes involved in heparan sulfate proteoglycan production are selectively silenced.  This observation explains in part the loss of heparan sulfate in kidneys and intestines that contributes to the leakiness of these organs in response to inflammation and the partial repair of these organs by heparin treatment.  Decrease of heparan sulfate that normally coats cells and binds antimicrobial peptides, such as LL-37, would explain the enhanced movement of LL-37 in psoriatic skin.

LL-37 Binds to Host DNA and Triggers Toll-Like Receptors
DNA is released from keratinocytes in psoriatic skin and this host DNA binds the antimicrobial peptide cathelicidin LL-37.  The LL-37/DNA complex mimics bacterial DNA and triggers the Toll-like receptors (TLR) on the surface of immune cells, dendrocytes, to activate NFkB, the transcription factor controlling inflammation.
http://www.ncbi.nlm.nih.gov/pubmed/19050268?ordinalpos=1&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_SingleItemSupl.Pubmed_Discovery_RA&linkpos=1&log$=relatedarticles&logdbfrom=pubmed

Heparin Treats Psoriasis
It seemed obvious to me that the heparin binding domains (Look at all the basic amino acids in blue in the illustration of LL-37.) of LL-37 were involved in DNA binding and the reason the LL-37 was binding to host DNA, was that heparan sulfate had been depleted as a result of local inflammation.  It also seemed obvious that topical heparin should eliminate psoriasis plaques.  So I did a Google search of psoriasis + topical heparin and got a hit on a 1991 patent application that claims a broad applicability for heparin use in curing symptoms of a wide variety of diseases, including psoriasis.
http://www.patentstorm.us/patents/5037810/description.html
The only topical form of heparin that I know of is Lipactin (available in Canada and Europe?), a treatment for coldsores, which makes sense because herpes viruses use heparan sulfate to infect cells.

Biofilm Transformation, Helicobacter, Klebsiella

Helicobacter pylori causes stomach cancer, but it feeds on hydrogen gas produced by Klebsiella pneumoniae in gut biofilms. DNA released by biofilm bacteria not only transfers antibiotic resistance, but it also provides protection against host antibacterial peptides, such a cathelicidins and defensins.

Exploding Labs

When I was working on host/pathogen interactions and plant disease resistance, I also became familiar with research on the formation of the plant equivalent of cancer, crown galls, and symbiotic bacterial nitrogen fixation. I mention this, because this also exposed me to the free-living bacterial nitrogen fixing system in Klebsiella and to the memorable urban legion of exploding labs. As the story goes, as bacteria convert atmospheric nitrogen gas into ammonia, nitrogen fixation, they use high energy electrons, e.g. from ferrodoxin, and lots of ATP, but they also produce hydrogen gas. In labs where they are researching nitrogen fixation, the excess hydrogen gas would accumulate on the ceiling until... boom! Now those labs are properly vented.

Helicobacter Uses Hydrogen as an Energy Source

Helicobacter pylori is considered the most common bacterial pathogen of humans and is the primary cause of ulcers and stomach cancer. H. pylori lives in the stomach by neutralizing stomach acid with ammonia. Another interesting ability of this bacterium is its ability to use hydrogen dissolved in circulating blood as an energy source. The high energy electrons from molecular hydrogen are transported to its electron transport chain, and the energy is used in membrane transport and ATP production. The circulating hydrogen is produced by gut bacteria.

Klebsiella Is not just a Soil Bacterium, Gut Gases

Klebsiella pneumonia is a lung pathogen and it also forms gut biofilms. Presence in the gut and the ability to produce hydrogen gas has some implications for hydrogen utilizing bacteria like H. pylori. Clearly, the stomach of someone with an abundant source of hydrogen fuel in their blood is a better target for H. pylori colonization. This explains why even at age 50, individuals who were exclusively breastfed have a lower incidence of H. pylori and stomach cancer, since even a single bottle of formula can shift an infant to adult, i.e. Klebsiella gut flora.

Klebsiella Needs Carbs to Produce Hydrogen

K. pneumoniae has been associated with Crohn’s Disease and Ankylosing Spondylitis. It grows in gut biofilms and produces pullulanase, an enzyme that can utilize the branched glucosides left over from the action of amylase on plant starch. So K.p. has an untapped food source and it needs lots of ATP to produce hydrogen gas. The nitrogenase needed for nitrogen fixation and hydrogen production is very sensitive to oxygen, so this means that K.p. needs a partially anaerobic environment and must get its energy from fermentation. Fermentation yields much less ATP than respiration using oxygen, which means that K.p. can only produce hydrogen with lots of glucose from starch.

Low Carb Diet Cures Crohn’s Disease

It turns out that the antigen causing Crohn’s disease is the pullulanse (with collagen mimetics.) As you should expect, it has a basic triplet. Eating a low carb diet reduces the flareups of Crohn’s disease, presumably by starving out the K.p.. It is interesting that nitrogenase is the antigen involved in Ankylosing Spondylitis.

Biofilms Promote Transformation and Antibiotic Resistance

Just as a footnote to the benefit of K.p. as a citizen of a biofilm community, H.p. should also live in those biofilms, since that is the source of the hydrogen it uses. Biofilms also stimulate the exchange of DNA, because the quorum sensing chemical signals trigger the release of DNA. The DNA is a component in the matrix that binds bacteria in the biofilm and can work in conjunction with bacterial acidic polysaccharides and host heparan sulfate. These acidic polymers tend to bind the basic antimicrobial peptides, e.g. defensins and cathecidins produced as a major non-adaptive defense against bacteria. Thus, the release of DNA triggered by quorum sensing, builds matrix, facilitates DNA transformation that is the foundation for the spread of antibiotic resistance in gut biofilms and provides resistance against antimicrobial peptides.

Lactoferrin: Natural Anti-Microbial Milk Protein

Nosocomial infections happen when the immune system is compromised through a medical procedure and common bacteria, such as Staphlococcus aureus, get introduced. From my perspective, that means that the walking bacterial reservoirs, i.e. gut flora of healthcare practitioners, provide an inoculum directly to the damaged tissue, or indirectly by contaminating the patients gut flora and then spreading the pathogens from the patient’s digestive tract to the damage site. This is what happens with ventilator-associated pneumonia and sepsis in critically ill patients.

The trick is to keep the patient’s gut flora healthy -- healthy as a breastfed baby’s. The typical medical approach is to kill off lurking pathogens with a dose of antibiotics. The problem with this approach is that it is both indiscriminant and selective, i.e. it kills both pathogens and beneficial bacteria, but it also provides a selective advantage for the antibiotic resistant hospital strains of opportunistic pathogens.

Humor break: Why do babies spit up half-digested breastmilk and then smile? Answer: Pepsin produces antimicrobial peptides from milk proteins. The baby smugly acknowledges that she knows that she has just protected her upper respiratory and digestive tracts against bacterial pathogens.

Pepsin hydrolyzes proteins next to aromatic amino acids and away from the basic amino acids, arginine and lysine. That means that heparin-binding domains, which consist of groups of basic amino acids in a hydrophobic environment, are clipped out intact from proteins by pepsin. Thus, babies sucking down milk make their own isolated peptides with heparin-binding domains.

Lactoferricin with basic amino acids in blue.

Many organisms, from fruit flies to frogs to humans, produce anti-microbial peptides. They also produce proteins with nucleic acid-binding domains and nuclear localization signals and heparin-binding domains and IP3-binding domains. If all of those binding domains are clipped out by pepsin and the peptides are compared to the anti-microbial defensive peptides, amazingly they are all the same. All have groups of basic amino acids among hydrophobic neighbors, and all are toxic to bacteria.

Lactoferrin is a major component of milk whey. It binds iron and heparin. It can be digested by pepsin into an an anti-microbial peptide, lactoferricin. Baby’s smile and spit-up on your shoes when you say lactoferrin.

Transgenic mice that produce porcine lactoferrin in their milk, transfer extra lactoferrin their little suckling mouse pups and that extra lactoferrin gives extra protection against bacterial and yeast pathogens. That is the experimental justification to suggest that treating patients at risk of nosocomial infections (I guess that would mean every patient in contact with a nurse or doctor) with oral lactoferrin should selectively eliminate the pathogens. Lactoferrin is prebiotic and supports the growth of probiotic gut flora.

ref:
Yen CC, Lin CY, Chong KY, Tsai TC, Shen CJ, Lin MF, Su CY, Chen HL, Chen CM. Lactoferrin as a natural regimen for selective decontamination of the digestive tract: recombinant porcine lactoferrin expressed in the milk of transgenic mice protects neonates from pathogenic challenge in the gastrointestinal tract .J Infect Dis. 2009 Feb 15;199(4):590-8.

Crohn’s Disease and Cryptidins

The intestines produce enzymes to digest food, antimicrobial peptides to kill pathogens and have lots of surface area to absorb nutrient molecules released from the food macromolecules (protein, polysaccharides, fats). The epithelial cells that line the intestines, enterocytes, must communicate with bacteria in the gut, the gut flora, to maintain bacteria helpful in food digestion, i.e. probiotic bacteria, and trigger an immune response to eliminate pathogens. Probiotic bacteria are tolerated and pathogens are identified and attacked.

Enterocytes are produced by division of stem cells at the bottom of the crypts that are in the valleys between the villi that project into the lumen where the digesting food is. New enterocytes are added at the base of the villi and old enterocytes are sloughed off at the top of the villi. As the new enterocytes move up the villi, they differentiate to produce the dramatic surface of microvilli, the furry brush border that further expands their surface area. The mature enterocytes produce transport proteins on their microvilli to take up sugars, amino acids and fats. The small nutrient molecules pass through the base of the enterocytes and bath the cells below, the lamina propia. The nutrients enter the capillaries of the villi and travel to the liver. Fats are transported through the lymphatic system.

Bacteria that slip through the enterocyte layer encounter macrophages and other types of white blood cells of the lamina propia. Among these cells are the Paneth cells. Fragments of the cell walls of bacteria bind to the NOD proteins of the Paneth cells and trigger the secretion of antimicrobial peptides, the cryptidins. Cryptidins are antimicrobial because of their array of basic amino acids surrounded by hydrophobic amino acids. These short proteins are able to disrupt the membrane function of most bacteria. I think they work on bacteria the same way that amyloid proteins, e.g. amyloid plaque proteins of Alzheimer’s disease, kill human cells. In fact, amyloid fibers bind to heparin and so do antibiotic peptides.

Here is an example of an antibiotic peptide, cryptidin 4,

GLLCYCRKGHCKRGERVRGTCGIRFLYCCPRR

Note the pairs of basic amino acids (blue). These amino acids are necessary for toxicity to bacteria. Heparin binding domains from proteins are produced naturally as proteins are digested to peptides in the stomach by pepsin. Pepsin hydrolyzes proteins next to the basic amino acids and leaves antimicrobial peptides that sterilize incoming food. I have illustrated the cryptidin protein to show how the basic amino acids (blue) are displayed on its surface.

With each meal, the fat content normally stimulates the production of a hormone, cholecystokinin, that binds to a receptor and causes an anti-inflammatory release of cytokines from the vagus nerves that reach the villi. Thus, food normally makes the intestines more tolerant of food antigens.

If the intestines become chronically inflamed, then exposure to normal probiotic bacteria can lead to cycles of inflammation that damage the integrity of the intestines. The intestines lose the ability to discriminate between probiotic and pathogen.

Crohn’s disease is an inflammatory, autoimmune disease of the bowel. The chronic inflammation of the lamina propia eliminate the ability of the Paneth cells to produce cryptidins and bacteria set up residence in the crypts and cause continual inflammation. This disease is typically treated by suppressing inflammation and treating with antibiotics.

Other treatment approaches that have been found effective are omega-3 oils to stimulate production of anti-inflammatory prostaglandins, pre- and probiotics, heparin and helminth eggs, e.g. wireworm.

Crohn’s disease would seem to benefit from the standard recommendation of an anti-inflammatory diet and lifestyle.