The Cause of Allegies and Autoimmune Diseases

Keyhole Limpet Hemocyanin (KLH): Internalized Antigen

Scanning the literature for a common protein that can be used as an experimental antigen, it becomes quickly obvious that a favorite is KLH. This would seem to be an odd choice -- why a keyhole limpet protein? But that is the wrong question.

Why is KLH such a good antigen, i.e. why is it readily presented to the host immune system? If you have been reading my posts, you might be thinking about triplets of basic amino acids and that is the answer.

As soon as I remembered the prominent use of KLH as an antigen, I checked the NCBI protein database and immediately found an unusual KKK (triple lysine) near the amino terminus of hemocyanin II ( it comes in two pieces). This triplet explains why KLH is such a good experimental antigen, because it is internalized into antigen presenting cells by its strong heparin-binding domain. Other components, adjuvants, are typically added to the KLH for injection to make sure that a strong local inflammation occurs.

Autoantigens Have Strong Heparin-Binding Triplet

I also learned that Hashimoto’s thyroiditis is an autoimmune disease mediated by the autoantigen thyroid peroxidase. A quick search reveals that thyroid peroxidase is an autoantigen, because it also has a triplet of basic amino acids that can enhance presentation under inflammatory conditions. Grave’s disease of hyperthyroidism is an autoimmune disease in which the thyroid receptor (with a basic triplet) is an autoantigen. The same kind of triplet of basic amino acids was found when I searched today for fire ant antigens and mosquito antigens.

I have also looked for the triplets in protein databases. The triplets are rare in cytoplasmic and extracellular proteins. The proteins that have triplets are usually identified as autoantigens in some disease. The triplets are common in nuclear proteins, since heparin-binding and nucleic acid-binding share the same basic amino acid domains. The nuclear internalization signal also results in rapid cellular internalization, e.g. HIV-TAT, heparanase, IGF-binding proteins. Nuclear proteins are common autoantigens in lupus.

Inflammation Plus Heparin-Binding Internalization: Allergy, Autoimmunity

Chronic inflammation can produce antibodies against proteins (foreign or self) with strong heparin-binding domains (triplets or sometimes neighboring pairs of basic amino acids, lysine or arginine). The generalization explains why particular proteins in pollens, foods, insects, pets, mites, asthma, MS, lupus, celiac, etc. produce antibody responses.

Allergy, Asthma, Autoimmunity Start the Same Way

Inflammation is the current medical buzzword. Name the disease and inflammation is there.

Reproduction Requires Controlled Inflammation
Aspirin blocks many of the steps in triggering inflammation and thus, aspirin administration can be used to reveal a role of inflammation in many unexpected places. Aspirin is effective in blocking some forms of infertility, inhibiting miscarriages and ameliorating postpartum depression. So inflammation is a critical part of reproduction. But, also notice that depression is a symptom of chronic inflammation.

Cancer Requires Inflammation
High dose (IV) aspirin has been successfully used to treat cancer. Inflammation is required for cancer growth, because both use the same transcription factor, NFkB. The aberrant signaling of cancer cells would normally lead to programed cell death, apoptosis, but inflammation blocks apoptosis. Aspirin can in turn block NFkB and in the absence of inflammation, cancer cells die by apoptosis.

Inflammation is Self-Limiting
Aspirin also transforms the COX/lipoxidase system to produce anti-inflammatory prostaglandins/eicosinoids. Inflammation normally progresses into anti-inflammation. Blocking this progression leads to chronic inflammation and a shift from local to systemic inflammation with the rise of inflammatory interleukins in the blood stream.

Immune Response Requires Inflammation
The signal molecules (IL-1, IL-6, TNF) and transcription factor, NFkB, associated with inflammation were all initially identified in the development of lymphocytes. Hence, IL stands for interleukin, a hormone that triggers leukocyte (literally white blood cells or cells associated with the lymphatic immune system, i.e. lymphocytes) development. The nuclear factor, i.e. transcription factor, involved in expression of the large chain, kappa, of immunoglobulins in B cells, was called NFkB.

Genes Expressed by NFkB Cause Symptoms of Inflammation
About five dozen genes are under control of NFkB. Among these are COX-2, the enzyme that converts omega-6 arachidonic acid to inflammatory prostaglandins; iNOS, the enzyme that produces nitric oxide that dilates blood vessels to produce hot, red skin; and the inflammatory interleukins, IL-1, IL-6 and TNF, associated with autoimmune disease, fatigue and cachexia (wasting).

Autoimmunity and Allergy Start with Inflammation
Medical treatments focus on symptom abatement and ignore cause. What causes obesity, allergy or autoimmune disease? The answer appears to be chronic systemic inflammation plus exposure to unusual proteins. The unusual proteins are immunogenic, i.e. interact with the immune system to produce antibodies or reactive T-cell receptors, and are subsequently recognized as autoantigens or allergens, that are the targets for immune attack. Inspection of these autoantigens and allergens shows that they all have one thing in common, they bind to heparin via a strong heparin-binding protein domain that is typically a triplet of adjacent basic amino acids.

Heparin is a Short, Highly Sulfated Fragment of Heparan Sulfate
Commercial heparin is purified from the intestines of hogs and cattle. Heparin is released from mast cells (made fluorescent for microscopy using berberine) along with histamine and is released into the intestines to block pathogens from binding to the heparan sulfate that is part of the intestine surface. The heparin is anti-inflammatory and it contributes to minimizing the inflammatory response of the intestines to food.

Inflammation Reduces Heparan Sulfate Production
Pathogen-generated inflammation of the intestines reduces heparan sulfate production and increases immune response to food antigens. NFkB activation by inflammation turns off the production of some genes needed for heparan sulfate proteoglycan (HSPG) synthesis. Since HSPG is a major component of the basement membrane that holds tissues together, the reduction of HSPG results in protein loss (proteinuria) from kidneys, leaking of intestines, and disruption of the blood/brain barrier.

Reduction of HSPG Results in Immunological Presentation of Autoantigens/Allergens
Proteins are brought into cells by specific binding to protein receptors. In many cases, particularly involving signaling or growth factors, both the signal molecules and the receptors bind to heparin. In addition, there is a robust circulation of HSPG, which is secreted and internalized with a half-life of approximately six hours. The sweep of the HSPGs take heparin-binding proteins with them for internalization, e.g. HIV-TAT, heparanase, tissue transglutaminase. I think that this HSPG sweep under inflammatory conditions also internalizes basic autoantigens and allergens with strong heparin-binding domains. This internalization is the first step toward immunological presentation and the immune response to autoantigens and allergens.

Autoantigen/autoantibody/HSPG Complexes Kill Cells
Antibodies against self-antigens, autoantigens form antigen/antibody complexes that also bind to and cross-link HSPGs, because of the heparin-binding domains of the autoantigens. The large complexes may disrupt HSPG circulation and trigger apoptosis or abnormal physiology. There are many other examples of heparin-based complexes that are toxic, e.g. Alzheimer’s amyloid plaque, diabetic beta cell antibody complexes, celiac gluten/tRG antibody complexes, multiple sclerosis myelin antibody complexes, atherosclerotic plaque.

Anti-Inflammatory Diet and Lifestyle Protects
Dietary and lifestyle adjustments that minimize inflammation, e.g. low starch, no HFCS, low vegetable oil (except olive) and supplements of vitamins D & C, fish oil (omega-3) and glucosamine, reduce the risk of allergies/asthma, degenerative diseases and cancers. Simple, high level supplements with fish oil reduce numerous mental disorders, e.g. depression, ADHD; infertility, pre-eclampsia and postpartum depression; allergies, asthma; arthritis, atherosclerosis; burn recovery, septicemia and head injury.

Reducing Inflammation is a Panacea for Modern Diseases
Most modern diseases have an inflammatory component, because modern diets are rich in inflammatory components, e.g. starch/sugar, corn/soy oil, HFCS, trans fats, and exercise is minimal. The medical industry has not successfully promoted healthy eating and exercise; and in fact has promoted the devastating replacement of saturated fats with inflammatory polyunsaturated vegetable oils. Meat production has moved away from grazing on omega-3-rich plant vegetation to omega-6-rich corn and soy. Replacement of the corn/soy based agricultural economy would have predictably immense beneficial impact in reducing inflammation-based degenerative autoimmune diseases and cancers.

Enteroviruses, Autoimmunity, Diabetes

Insulin-producing cells of the pancreases of diabetics have been found to harbor viruses common to the gut. Antibodies to the virus coat proteins also bind to pancreas proteins.

A recent paper, referenced below, shows that in a collection of samples from the pancreases of individuals that had been diagnosed with type I diabetes less than one year prior to the sampling, insulin producing cells are also infected with enterovirus. The same association between enterovirus infection was found to a lesser extent in type II diabetics, but not in non-diabetic controls.

Enteroviruses have been repeated associated with diabetes over the last decade and antigenic determinants of the enterovirus protein coat also bind, i.e. cross react, with antigenic determinants of human cellular proteins.

I examined the enterovirus coat protein, VP1, and found the same three amino acid sequence (three basic amino acids, lysine [K] or arginine [R], highlighted) that I also found in all allergens (peanut, ragweed, dust mite, bee venom) and autoantigens of autoimmune diseases (lupus, MS), and is associated with heparan sulfate-based internalization and presentation of protein immunogens. This observation is consistent with my hypothesis that inflammation plus the presence of one of these proteins, results in production of B and T lymphocytes specific for antigenic determinants on the surface of the immunogen protein. Note that the antigenic determinants usually do not include the three basic amino acid sequence, e.g. RRK, that is involved in uptake and presentation of the protein.

VP1 [Human enterovirus B]
HVINYHTRSESSVENFMGRAACVYIAQYATEKVNDELDR
YTNWEITTRQVAQLRRKLEMFTYMRFDLEVTFVITSSQR
TSTTYASDSPPLTHQVM

reference:
Richardson SJ, Willcox A, Bone AJ, Foulis AK, Morgan NG. 2009. The prevalence of enteroviral capsid protein vp1 immunostaining in pancreatic islets in human type 1 diabetes. Diabetologia. Mar 6. [Epub ahead of print]

CIDP, Inflammation and Autoantigens

Chronic Inflammatory Demyelinating Polyneuropathy (CIDP) is chronic, relapsing autoimmune disease in which the immune system attacks the insulating myelin sheaths of nerves in the limbs. The result is numbness and pain. An acute version of this myelin compromising disease is Guillain-Barre syndrome (GBS) typically initiated by Campylobacter jejuni infection. A related disease of brain tissue is multiple sclerosis (MS). In each of these cases antibodies bind to protein or glycolipid antigens unique to myelin and target the attack by lymphocytes.

Autoimmune diseases represent a disruption of the normal function of the immune system that arises from a breakdown of the self/non-self tolerance process. Immune responses by antibodies or lymphocytes is normally reserved for molecules that don’t exist in the body, i.e. self. Differentiation of lymphocytes in the thymus or bone marrow involves systematic elimination of all lymphocytes that interact with self antigens. There are, however, self antigens that are not normally encountered by the immune system, e.g. nuclear antigens, brain antigens, since these antigens are only excluded from the cytoplasm or result from specialized differentiation. Thus, you can find in mystery novels the use of letters coated with powdered brain antigen to slowly kill targets by induction of a lethal autoimmune disease.

The self antigens that are the typical targets of the CIDP, GB and MS are gangliosides, myelin glycolipid also produced by C. jejuni, and basic proteins such as peripheral myelin protein 22. The glycolipid makes sense, because the initial exposure is during a bacterial infection that also produces inflammation. The basic proteins are a different case.

I have examined numerous self or autoantigens that are the targets for antibodies produced in autoimmune diseases and allergies. In each case the same amino acid sequence or domain is present, three basic (positively charged, arginine or lysine) amino acids. This is true for the myelin autoantigens; allergens from ragweed, mites, peanut, etc.; diabetes, lupus and arthritis.

Here is the example of the CIDP autoantigen, peripheral myelin protein 22:
MLLLLLSIIVLHVAVLVLLFVSTIVSQW
IVGNGHATDLWQNCSTSSSGNVHHCF
SSSPNEWLQSVQATMILSIIFSILSLFLF
FCQLFTLTKGGRFYITGIFQILAGLCVM
SAAAIYTVRHPEWHLNSDYSYGFAYIL
AWVAFPLALLSGVIYVILRKRE

The three basic amino acids, RKR, are at the end of the protein amino acid sequence.

A dramatic instance of tissue damage caused by physical injury or infection, is superimposed on chronic dietary inflammation to provide the environment for an inappropriate response of the immune system to a self or identical foreign antigen. The only proteins that seem to be presented in this way to the immune system are those with the strong heparin-binding domain, BBB (B = basic amino acid). This indicates that perhaps the reduction in heparin production that accompanies inflammation, may be responsible for the aberrant immune response. It is also noteworthy that heparin and anti-inflammatory agents can reduce the symptoms of many autoimmune diseases.

There are many complex therapies to deal with the symptoms of these autoimmune diseases. In all of these cases, one of the most effective approaches is an anti-inflammatory diet and lifestyle. A novel additional approach is helminth therapy, infection with parasites to suppress the T-lymphocyte autoimmune attack. This approach is effective with MS, and I think would be an excellent, relatively safe approach to be examined for the other myelin autoimmune diseases.