Health and Heparan Sulfate Circulation — Connective Tissue is Alive

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Arthritis, Alzheimer’s, diabetes, cardiovascular disease, osteoporosis, cancer, etc. are all diseases of cellular metabolism and secretion.  What goes on inside cells and on their surfaces explains a lot about health and why we get sick.  Cells feed off of what’s around them, use some of those materials to replicate and package up cell-made materials for export.  Eat, replicate and secrete.  Symptoms of disease result if those processes are compromised.

Cell that make Cartilage, Eat Cartilage

The connective tissue that makes up the cartilage of tendons and the non-mineral parts of bones, as well as a layers of skin, is made up of proteins (collagen) and polysaccharides (glycosaminoglycans, GAGs), e.g. heparan sulfate, hyaluronan and chondroitin sulfate, produced by  chondrocytes or fibroblasts.  These proteins and polysaccharides are synthesized and then secreted by cells.  This process goes on continuously, since the connective tissue is alive and literally crawling with cells that make the cartilage.  To keep the connective tissue healthy, the old tissue has to be digested, so that new material can replace it.  Thus, the cells that live in cartilage also eat cartilage.  These cells get all of their nutrients, e.g. protein and carbs, from eating cartilage.  They don’t get glucose and amino acids, or even oxygen (they ferment), from the blood, because there are no blood vessels in cartilage.  The photomicrograph at left shows the red chondrocytes surrounded by a light capsule of heparan sulfate as they burrow through the purple cartilage.  The next micrograph shows the cytoskeleton of actin filaments (stained with a red fluorescent dye, that lies under the cytoplasm of a chondrocyte.  Motor proteins move other proteins, such as syndecans, the proteins to which the heparan sulfate chains are attached, through the  cell membrane (see the animations below.)  The last micrograph shows the green stained microtubule network on which vesicles move to carry heparan sulfate products from one end of the cell to the other (under the actin and past the orange-dyed nucleus) during synthesis and digestion.

Chondrocytes Burrow Through Cartilage

Chondrocytes are the cells that eat and make cartilage, but all of this eating and making goes on at the same time that the cartilage is also holding everything together, i.e. it is still strong.  If cartilage is cut and the cut ends are held tightly together, the chondrocytes will knit the cartilage together and it will become as strong as it was. 

Heparan Sulfate Circulates over the Surface of Cells

Chondrocytes are not actually rigidly embedded in the cartilage, but rather maintain a capsule of heparan sulfate around themselves.  Thus, they continue to secrete a mixture of heparan sulfate, chondroitin sulfate and collagen, but the heparan sulfate is recycled through the capsule and the other molecules merge into the existing cartilage.  Thus, the heparan sulfate is a kind of carrier that keeps the cartilage from “setting up” while it is being made and transported.  Other cells of the body, such as neurons, don’t make cartilage, but they still have heparan sulfate (HS) circulation that is intimately involved in many other processes, such as the action of hormones.  Disruption of HS circulation causes the symptoms of Alzheimer’s or type 1 diabetes, for example, since amyloids assemble as filaments on threads of HS, and the amyloid filaments jam essential HS circulation.  Plaque in atherosclerotic vessels is high in HS content.  HS is also a major component surrounding vessels to form the blood brain barrier and the barrier to protein loss from kidneys into urine or loss into the gut lumin.  Heparin (fragments of HS) is continually released from mast cells in the lining of the gut to prevent pathogens from binding to cell HSPGs. 

HS Sweep the Cell Surface

There is a constant flow of heparan sulfate proteoglycans (HSPGs) through the cell membrane from the rear of the chondrocyte to the front where the HS is digested again and the protein that was embedded in the membrane, syndecan, is recycled to the Golgi for another trip.  HSPGs (animation to left with blue protein and yellow HS) are attached to motor proteins that propel them through the membrane along microfilaments of actin that form the cyctoskeleton just under the membrane in the cortical region of the cell.  Thus, the heparan sulfate of the HSPGs stick out like hair from the cell surface and sweep continuously from the back to the front of the cell.  At the front of the cell, the HS sweeps through the intact cartilage and reverses the process of cartilage assembly.  The chondroitin sulfate, collagen and HSPGs are dragged into the cell and digested.  The protein parts of the HSPGs are transported to the Golgi  and the HS is synthesized along with other cartilage components and moved in vesicles along microtubules before it is secreted.

HS is Secreted at One End and Eaten at the Other

The animation left shows 1) the initial digestion of the cartilage proteins and polysaccharides on the left.  These cartilage components of amino acids and sugars, are used by the chondrocytes as their sole nutrients 2), and to produce new proteoglycans 3) HS and chondroitin sulfate proteoglycans, in the Golgi, are 4) packaged into secretory vesicles and are 5) secreted on the right.  The HS chains, attached to proteins, are 6) swept through the membrane (see the first animation above) toward the front of the cell, leaving the collagen and chondroitin sulfate for form cartilage behind.  In the process, the heparan sulfate proteoglycans 7) disrupt and solublilize old cartilage ahead as the chondrocytes 8) move through the connective tissue like moles digging through soil.

Other Cell Processes Involving Heparan Sulfate:

• Amyloids of Alzheimer’s and type I diabetes assemble bound to HS.
• Hormones bind to receptors wrapped around HS.
• Blood clotting is controlled by HS.
• Complement is controlled by HS.
• Blood brain barrier is composed of HS.
• Kidney protein barrier is composed of HS.
• Inflammation blocks HS synthesis and promotes heparanase synthesis.
• GAGs are animal soluble fiber when eaten and feed gut flora.
• Pathogens bind to HS.
• HIV-TAT is transported between cells by HS circulation.
• Heparin is made by heparanase fragmentation of HSPG in mast cells and is secreted along with histamine. 
• NFkB activation inhibits HSPG production and stimulates heparanase production.
• Heparan sulfate proteoglycans organize nerve synapses and acetylcholine esterase binds to HS. 
• Gastric proteases cleave around heparin binding domains of proteins, e.g. milk, consist of clusters of basic amino acids.  Peptides with heparin binding domain are antimicrobial; all of the heparin binding peptides are subsequently degraded by pancreatic proteases.
• Heparanase is initially secreted inactive and bound to HSPGs, but it remains bound and is internalized again along with the recycling HSPGs, and is activated before being secreted again.
• Allergens and autoantigens are unusual proteins with sequences of three adjacent basic amino acids (arginine or lysine) that require HSPG circulation for presentation of the immune system.  Nuclear proteins that interact with nucleic acids have sequences of four basic amino acids, the nuclear translocation signal, and are therefore common antinuclear auto antigens.

Health Diagrams

Health Diagrams II — Curing Autoimmunity and Allergies

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In this second in a series of posts explaining the concepts that I think are central, but misunderstood, about health, I am focusing on how diet and gut flora impact the immune system and cause autoimmunity and allergies.  This cause also suggests a simple cure.

Health in Diagrams I -- Gut Flora and Diet
Health in Diagrams III -- Inflammation from Cell to Tissue

Gut Flora to Tregs to Suppression of Autoimmunity

It is important to understand at the outset that autoimmunity and allergies are caused by a damaged immune system, and repairing the damage cures the diseases.  Damage to the immune system typically represents a break in the continual development of immune cells in the lining of the intestines.  Immune cell development in the gut is dependent on bacteria, the gut flora.  Damage to the gut flora, e.g. by antibiotics, processed foods that lack flora feeding fiber or extreme diets, disrupts development of immune cells.  Typically, loss of the immune cells that keep the aggressiveness of the immune system in check, regulatory T cells or Tregs, results in autoimmunity.  Fix the gut flora and autoimmunity recedes.  

Health Requires Suppression of the Aggressive Immune System

For simplicity, I am focusing on the T cells of the immune system that develop in the intestines and either kill other human cells that are dangerous, e.g. virus-infected or cancer cells, or provide protection by regulating the aggression, Tregs.  Normal functioning of the immune cells permits elimination of damaged or dangerous human cells, while at the same time avoiding rampages of lethally armed T killers.  Examples of untamed T killers in action are degenerative autoimmune diseases, such as arthritis, asthma, prostatitis, celiac, Hashimoto’s thyroiditis, type I diabetes, inflammatory bowel diseases and atherosclerosis. 

Milk Births Baby Immune System

It should not be surprising that the focus of immune system development is the gut.  We start as babies with explicit links between nourishment and immunological protection.  Milk connects the immune systems of mother to baby.  Immune cells from the mother are transferred in milk and colonize the respiratory and digestive system of the baby — the mother’s immune system coats and buffers the baby’s exposure to the world.  Milk hormones close the baby’s gut and milk bacteria are the first probiotics that exploit the milk prebiotics (bifidus factor, human milk oligosaccharides) to produce a gut flora.  [Also note that most commercial probiotics are adapted to grow on cow’s milk and hence these dairy probiotics do not survive in adults.]  The lymphatic system of the breast terminates at the nipple and samples antigens/pathogens from the baby’s mouth, resulting in baby-specific secretory antibodies that return in the milk.  Milk supports a starter set of gut flora, essentially dairy probiotics, that stimulates development of the baby immune system, but inhibits adult gut flora that would digest the protective components of milk.  Formula, on the other hand, is inflammatory to the baby gut, because it supports adult gut flora before the immune system is ready.  Inflammation and stimulation of innate immunity is sufficient, if supported with high levels of sanitation, to permit survival of babies fed formula.  Milk of any type is incompatible with adult gut flora, so breast milk will attack adult gut flora and adult gut flora will digest and inactivate the otherwise beneficial components of the milk.

Milk, Kefir and Gut Flora

Aggressive and Suppressive Cells of Immune System Develop in Intestines

Gut bacteria are required for the development of immune T cells in the lining of the intestines.  Mice grown without gut flora do not have functional immune systems.  In humans, extensive antibiotic treatment produces defective immune systems that are either overly aggressive, i.e. autoimmune, or susceptible to infection and cancer.  They can’t be both.  Aggressive T killers are stimulated to develop by filamentous bacteria and Tregs develop in response to members of the Clostridium family.  In a healthy body, there is a balance between aggression and suppression; there are functional defenses against infection and cancer, while also avoiding autoimmune disease and allergies.

Suppressive Tregs are Deficient in Autoimmunity

Immune cells result from replicative divisions of stem cells.  Antibody producing B cells are produced through a million random rearrangements of antibody genes and those B cells producing antibodies against common self proteins are killed (clonal deletion).  Similarly, T cells are produced by rearrangements of receptors and those that would recognize self are eliminated.  The T cells then migrate to the intestines where they can develop into killer T cells or Tregs, in response to gut flora.  The Tregs act to suppress killer T cells that mistakenly recognize healthy self cells.  Thus, the initial elimination of self-attacking T cells or for B cells that produce antibodies that bind to normal cells, is not perfect and the Tregs are needed to avoid the mistakes.  Tregs are necessary to avoid the immune attack on healthy cells that is the basis of autoimmunity.

Autoimmunity Starts with Inflammation, but Requires Deficient Tregs

Bacterial or viral infections, or physical damage causing inflammation is the first step in autoimmunity.  It is the inflammation that initiates the interactions between proteins, autoantigens, of normal cells and cells of the immune system that bind, internalize, fragment and present the antigen fragments/peptides to activate B or T cells with corresponding receptors.  The activated B cells make antibodies specific for the antigen and the T cells will kill cells displaying the antigen.  It is interesting that most proteins are not autoantigens and are never involved immune reactions.  Only proteins with an unusual triplet of basic amino acids, similar to the quartet of basic amino acids used to transport proteins into the cell nucleus, are candidates to be autoantigens or allergens.  In fact, since nuclear proteins already have a quartet, i.e. the nuclear localization signal, they are common autoantigens.  The last requirement for autoimmunity is a deficiency in Tregs, because if the Tregs are functioning, they will block attack on healthy cells.  Treg deficiency usually results from loss of the type of gut bacteria that stimulate Treg production in the lining of the intestines, i.e. species of Clostridium.

Hospitals are Notorious for Clostridium difficile Infections

Fecal transplants are now recommended as a safe and efficacious treatment for C. diff hospital infections.  That makes sense, because hospitals are where antibiotics are routinely used and C. diff can only infect people missing their healthy species of Clostridium.  Thus, the hospitals wipe out the gut flora with antibiotics and then recolonize them with their own antibiotic resistant C. diff.  More antibiotics can’t fix it, but providing healthy gut flora (transplant) can.

Autoimmune Diseases are Treated/Exacerbated with Antibiotics

Both the aggressive and the suppressive immune cells require gut flora, so after initial antibiotic treatment wipes out bacteria required for suppression and results in autoimmunity, the remaining aggressive half of the immune system can be eliminated by blasting the remaining gut flora with more antibiotics.  Of course this will leave a highly compromised, incompetent immune system that will ultimately yield more extreme symptoms.  This is the typical medical progression for Crohn’s disease, for example.  The alternative is just fixing the gut flora to begin with and curing autoimmunity.

Cure Autoimmunity by Feeding Clostridium Resistant Starch

Autoimmune diseases, by their symptoms, show that sufficient gut flora to stimulate the aggressive half of the immune system is still present.  What is missing are the Clostridium species that convert soluble fiber, such as resistant starch, into short chain fatty acids, e.g. butyrate.  Patients treated with antibiotics usually walk away from the hospital with a suggestion to eat some yogurt to repopulate their missing gut flora.  Unfortunately, dairy probiotics don’t survive in the gut and cannot repair the gut flora and immune system.  The result, after the gut fails to repair and the immune system crashes, is autoimmunity.  There is a more appropriate possibility to avoid or fix autoimmunity.  Some people suffering from autoimmunity (and with remnants of their gut flora intact) have simply fed their gut flora on resistant starch and achieved complete recoveries.  Others fail to respond, because their gut flora is too severely damaged and necessary bacterial species are gone.  Those individuals need to eat the missing species of bacteria and some probiotics (more common in Asia) contain Clostridium species.  Consistent with this use of soluble fiber to feed gut bacteria that produce butyrate and stimulate the suppressive immune system are reports of healing by combining potato starch (RS) and probiotics with Clostridium butyricum (Probiotic-3).  Repair of the suppressive immune system by repair of gut flora (including fecal transplants) and feeding gut flora with appropriate soluble fiber, may be a general approach to the cure of most autoimmune diseases and allergies.

Paleo Gut Flora Repair

Diabetic Hypertension, Browning of the Arteries

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I decorated a flan with a drizzle of honey, and my torch produced a toasted spiral.  That was just fructose plus proteins, with a little heat, to produce advanced glycation end products (AGE) that are brown.  If you prefer, you can do the same reaction with egg whites and sugar in meringues, or by grilling brined pork chops basted with honey and anchovy paste.  Fructose is 10X better than other sugars at producing glycation, AGE and browning.

AGE and Arteries

Why do we care about the Maillard reaction and advanced glycation end products (AGEs)?  Of course, understanding the biochemistry of cooking is inherently satisfying and it helps to explain why Dr. House used vinegar to stop meatballs from browning too fast in his cooking class, but it also explains what’s cooking in atherosclerosis, cardiovascular disease.  It turns out that AGEs are highly inflammatory and inflammation of arteries leads to plaque formation.  [LDL is less important, because it only aggravates the primary event, inflammation, and that is why fish oil is more helpful for cardiovascular disease than statins.]  So increasing blood sugar is a problem, because it increases the rate of the Maillard reaction in binding blood sugar to the amino acids of proteins, such as hemoglobin to produce HgA1C, and causing vessel inflammation.  AGE in small capillaries also kills the capillaries and causes a rise in blood pressure by making harder for the heart to force blood from arteries to veins.  AGE causes hypertension and that is why salt consumption is not as important.   High blood sugar also increases the level of another powerful glycation agent, methylglyoxal, the active antibacterial agent in Manuka honey.  Honey is effective as a wound dressing, because it AGEs microbes to death!

The Trouble with AGE-ing is Inflammation

Cells detect the presence of AGEs with a surface Receptor for AGE (RAGE).  Binding of AGE to RAGE turns on the inflammation transcription factor, NFkB, with the release of inflammatory cytokines and the symptoms of inflammation.  One of my students did some computational protein modeling of the RAGE, because I was interested to see if RAGE would also bind Metformin.  Sure enough, our hunch was confirmed, indicating that Metformin might also reduce some forms of inflammation and be a treatment for diabetic high blood pressure.

Fructose vs. Inulin; AGE vs. Soluble Fiber

Fructose and the storage polymer of fructose, inulin, are similar to glucose and the storage polymer of glucose, starch.  The polysaccharides, inulin and starch can be converted to the sugars, fructose or glucose by industrial heating or enzymes.  Thus, agave inulin is converted into nectar, and corn starch is converted into syrup.  The polysaccharides are not sweet, but the sugars are.  The polysaccharides don't form AGEs with amino acids (unless they are broken up by high heat into sugars first) and fructose is 10X more chemically aggressive in forming AGEs than glucose.  Agave nectar (fructose) is a better browner than honey or high fructose corn syrup (corn syrup treated with a commercial enzyme to convert some of the glucose into sweeter fructose.)  Both inulin and some forms of starch (resistant starch), reach the colon and are digested by gut flora, i.e. they are soluble (fermentable) fiber.  The gut flora convert the resistant starch into short chain fatty acids that are anti-inflammatory.  Typical starch, e.g. corn starch or wheat flour, is digested by gut enzymes and goes directly into the blood as glucose, it is high glycemic and never reaches the gut flora.

AGE in Food

Should we fear browned foods as inflammatory.  I don't think that AGE in foods is any more of a hazard than all of the toxic phytochemicals that are touted as plant antioxidants.  I think that the gut and liver provide protection.  I brown the sugar on my flans and sear my steaks, even as I relish eating my veggies.  The body can detox these natural products in the gut better than it can handle the AGE made by high blood sugars.

Take Home Messages:

• Sugars in baked goods or blood, react with amino acids or proteins to make inflammatory AGEs.
• Blood sugar tests only measure glucose and ignore fructose, which is even more unhealthy.  So, foods laced with fructose can be low glycemic, but very unhealthy.
• The major AGE in blood is HgA1C.
• Diabetics have more stable, lower blood sugar on low carb diets, e.g. my Anti-Inflammatory Diet.  The liver produces needed blood sugar from protein.
• Diabetic use of fructose or agave nectar or honey encourages AGE, inflammation and diseases of diabetes.
• Starch (not RS) is the only polysaccharide digested by gut enzymes and is high glycemic.
• AGE is inflammatory leading to artery plaque and hypertension.
• AGE as browned foods are probably tolerated by the body.

Milk Casein, Amyloid, Pasteurization, Homogenization

Milk is a very special food for mammalian babies. It provides essential nutrients; stimulates development of the gut; promotes the growth of the unique neonate gut flora; and kills everything else. Milk is anti-bacterial, anti-fungal and anti-viral. It is used in fruit orchards as a pruning tool dip to prevent the spread of disease between trees, and it is used as a foot dip after ceremonial walking on hot coals. But is cow's milk healthy for adult humans and is milk compromised by pasteurization and homogenization?

Mother's Milk is Fierce
Milk as it is transferred from breast to baby is loaded with molecular weapons for the protection of the baby's respiratory and digestive systems. Cells from the mother are transferred along with the milk and quickly spread out on the surface of the mouth and digestive system to patrol for pathogens. The mother's immune system detects potential risks as the baby's mouth contacts the mother's lymphatic system at the breast, and the antibodies that are subsequently produced are transferred into the milk. Enzymes in the milk digest bacterial cell walls and other milk proteins are converted into anti-bacterial peptides in the baby's stomach before ultimately being digested into amino acid nutrients. Many of the fat/lipid nutrients in milk are also anti-bacterial or anti-viral. Most of the carbohydrate in milk is the simple disaccharide lactose that most bacteria can't use for food. The remaining 10% of the carbohydrates are extensions of the lactose to make galacto-oligosaccharides (GOS, a.k.a. bifidus factor) that are toxic to all but the few bacterial species that make up the highly specialized microbial community of the human baby gut flora. (Cow's milk has an entirely different composition, e.g. lacks bifidus factor, and supports a different gut flora.)

Milk is Liquid Fat
It is hard to transport fat in water, because it isn't soluble. That is true for blood or milk. We have all heard about good and bad cholesterol, LDL and HDL, and the problems of transporting blood lipids from gut to liver to tissues. Specialized carrier proteins are needed for lipid transport in blood and the same is true for milk. Caseins are the milk proteins that coat droplets of fats that make milk white and then form digestible curds in response to the baby's stomach acid and digestive enzymes. We exploit the natural curd forming response of milk proteins and lipid droplets to form yogurts and cheeses.

Pasteurization and Homogenization Put Milk in the Dairy Case
Milk behaves optimally when immediately transferred from the mother's mammary tissue to the baby's digestive tract. Bacteria that contaminate breast milk are quickly killed by cellular and molecular defenses of the milk itself. Thus, breast milk has a long storage life at room temperature, chilled or frozen. The natural defenses of milk also permit regional milk banks, where donated milk is minimally processed and screened, for subsequent use by hospitals to avoid problems, such as necrotizing colitis, associated with the use of artificial feeding substitutes. Commercial preservation of cow's milk in stores has resulted in attempts to extend the shelf-life by heat treatment (pasteurization) to provide additional protection from microbial contamination and homogenization to prevent curd formation.

Milk is for Babies
So why isn't milk the perfect food? Part of the reason may come from the highly specialized and essential role of milk for mammals like people. Millions of years of extreme selection pressure have made sure that every woman produces ample milk for all of her babies. Until very recently, if the baby could not successfully nurse, it would die. That made breast milk the perfect food for babies and milk was integral to the development of the baby gut, baby gut flora and baby immune system. But that didn't mean that cow's milk would be a healthy commercial food for human adults.

Milk Processing May Accentuate Casein Amyloid Fiber Formation
Proteins are made of a long sequence of a thousand amino acids. At each of those thousand positions there is one of twenty different amino acids. Some of the amino acids are hydrophilic and bind to water, whereas other amino acids are hydrophobic and bind only to lipids. Proteins in water fold and unfold in thousands of alternative configurations until the final shape is reached in which there is not enough energy in the molecular vibrations and movements of the water molecules to knock the protein into an alternative shape.

Heating/pasteurization and torturous mixing/homogenization can force milk casein and fats into new configurations that make the proteins stackable into fibers/amyloids. These milk protein fibers may be of interest, because protein fibers are important in many diseases, e.g. type I diabetes, Alzheimer's disease. The problem with amyloids, is that these fibers form a natural repetition of the same amino acid on each of the stacked proteins. This repetitive amino acid, e.g. positively charged lysine or arginine, can provide a binding site for a similarly spaced, oppositely charged molecule, such as heparin, which is involved in dragging molecules from the surface into cells. Beta amyloid fibers with positively charged amino acids in a band along their edges are what kills nerve cells in Alzheimer's disease.

Research has recently demonstrated that milk casein forms amyloid fibers in response to pasteurization and homogenization. It would be interesting to know if these fibers bind to heparin and if these fibers are toxic to intestinal cells.

I have raw cream from grass fed cows in my morning coffee and my three daughters never tasted formula.

National Healthcare and Diet

Barack Obama's Anti-Inflammatory Breakfast Pulpit

Trying to improve the health of Americans by taking the advice of the healthcare industry is futile. Barack Obama must set the example of a healthy lifestyle.

The solution is to prevent the diseases that the healthcare industry is using as a source of profits and that means fundamentally changing diets and lifestyles. It has taken three decades to attack health by shifting from meat protein and saturated fats to starch, high fructose corn syrup and polyunsaturated (omega-6) vegetable oils. It will take a sustained, personal effort by President Obama to guide a relatively rapid return to a healthy, anti-inflammatory diet.

All of the degenerative and autoimmune diseases that form the core of current healthcare diagnosis and treatments are rooted in an inflammatory diet and lifestyle dictated by agribusiness and uninformed by science. The media nags about people eating too much and exercising too little. Our obese population is encouraged to lose weight by eating less. Food fat is demonized. Statins are prescribed with religious zeal to lower blood lipids to reduce cardiovascular disease. All of this “health” advice is wrong and unsupported by the biomedical literature.

It is about time for an authority figure, i.e. The President of The United States of America, to use some leadership skills and teach people how to eat and live. That would be much easier than trying to get doctors to order fewer tests from their own medical test companies or order fewer images through their own imaging companies. Are the pharmaceutical companies going to suggest that their pills should cost less and be pushed less frequently? Will the insurance companies step out of their lucrative middleman role between doctor and patient? It is more reasonable for The President to use his bully pulpit to change the U.S. diet and lead us back to health.

All that is needed is for President Obama’s image at the breakfast table to be judiciously used by a private, non-profit organization on a website:

Barack’s Breakfast

This would provide an opportunity for the President’s health agenda to be presented to the world through his prescription (and explanation of health benefits) for each morning meal:

Slow food
Local food
Low carbon footprint
Low carbs
High omega-3 to -6 fatty acid ratio
Praise eggs and saturated fats
Warn about grains

Vitamin D deficiency
No HFCS
No trans fats

Each meal would come with a source and description of each ingredient and its benefits. YouTube videos of the meal preparation could show the techniques needed. Occasionally The President could be seen enjoying the meal and animation could be used to show why hypoglycemic ingredients were used. Maybe The President would show solidarity to the diabetic victims of industry food fights by getting his finger pricked for a blood sugar test after a meal. It would be good to see him complain about the inaccuracy of several different typical meters. Imagine the close-up of all of the lancet marks needed to convince him that the readings are making sense!

This single approach would cost the American people nothing to implement and would save billions of dollars in healthcare expenses over a few years, as citizens of all socio-economic classes changed to diets that were less inflammatory, and degenerative and autoimmune diseases quickly declined.

Insulin-like Growth Factor, Diabetes Autoantigen

IGF Binding to Heparin is Basis for Receptor Interaction, Internalization and Immunization

Examination of the protein sequence of insulin-like growth factors reveals strong heparin-binding domains (triplet of basic amino acids) that are also associated with internalization. Similar heparin internalization domains are also found on allergens and autoantigens. It was a small leap to expect that IGFs would also become autoantigens under inflammatory conditions that minimize heparan sulfate proteoglycan production.

Triplets of Basic Amino Acids Internalize Proteins

In several articles on this blog, I have discussed proteins that are internalized by their heparin binding domains. Heparin binding domains consistent only of a pair of basic amino acids, e.g. RK, flanked by one or more basic amino acids within a hydrophobic sequence of protein, are not sufficient to mediate internalization on heparan sulfate proteoglycans. A triplet of basic amino acids is usually required. Simple inspection of amino acid sequences is sufficient to identify these regions.

Internalization Triplet Identified in Insulin-like Growth Factor Binding Proteins

I noticed in a paper that insulin-like growth factors bind to epidermal growth factor receptors. I have previously written an article showing that EGF1 binds to its receptor via heparin, i.e. both the EGF and the receptor have heparin-binding domains. So I suspected that IGFs also had heparin binding domains. Inspection of the sequences readily identified simple heparin binding domains with pairs, but not triplets of basic amino acids. A search of the literature confirmed that heparin mediated IGF binding to receptors. A further search indicated that the heparin binding domains from proteins that bind and control the activity of IGFs could mediate internalization of proteins into cells and also into nuclei.

Internalization Triplets Are Associated with Allergens and Autoantigens

I have previously noted that all allergens and autoantigens have internalization triplets of basic amino acids. The presence of these triplets in IGF binding proteins suggested that IGF binding proteins might also be autoantigens. A quick check of the literature showed that antibodies against IGFs themselves frequently occur in type I diabetes. This suggests that the IGF-binding protein complexes are internalized and IGFs are immunologically presented during inflammation to produce anti-IGF antibodies. It is interesting that the other autoantigens for type I diabetes, e.g. transglutaminase, also have the expected internalization triplets.

references:
Maruyama T, Murayama H, Nagata A, Shimada A, Kasuga A, Saruta T.
Anti-insulin-like growth factor-1 autoantibodies in type 1 diabetes. Ann N Y Acad Sci. 2002 Apr;958:267-70.

Miao D, Yu L, Eisenbarth GS. Role of autoantibodies in type 1 diabetes. Front Biosci. 2007 Jan 1;12:1889-98.

Goda N, Tenno T, Inomata K, Shirakawa M, Tanaka T, Hiroaki H. Intracellular protein delivery activity of peptides derived from insulin-like growth factor binding proteins 3 and 5. Exp Cell Res. 2008 Aug 1;314(13):2352-61. Epub 2008 May 29.

Cure for Cancer, Autoimmunity, Allergies, etc.

The immune system is powerful enough to provide protection from disease. Unfortunately, to act decisively the cells of the immune system have to be able to discriminate between self and non-self. Poor discrimination can lead to autoimmunity, cancer or infection. New approaches promise the precise use of interleukins, to reset self-recognition, eliminate a wide range of diseases and liberalize organ transplantation.

IL-2 is the Cytokine Responsible for Suppression of Autoimmunity -- Tolerance

Self/non-self discrimination is dependent on cellular communication and much of that communication takes place via small proteins called interleukins. First and foremost among the interleukins is interleukin-2 (IL-2). IL-2 is made by cells of the immune system, lymphocytes. Mice that are either defective in producing IL-2 or the lymphocyte receptor for IL-2, IL2R alpha, also called CD25, rapidly develop autoimmune diseases, such as type I diabetes or inflammatory bowel disease. Thus IL-2 is necessary for both effective immunological defenses against pathogens and suppression of immune attacks on self tissues, i.e. autoimmunity.

IL-2 Balance Achieved with Complex of IL-2 and Anti-IL-2 Antibodies

Direct injection of IL-2 has some impact on cancers, but is very difficult to control. This should be expected, because local environments should determine if the IL-2 will stimulate aggressive immunological attacks or development of regulatory T cells, Tregs, that produce tolerance.

More subtle control is achieved by using antibodies that bind to particular regions of the IL-2. The resulting IL-2/anti-IL-2 complexes can be used to stimulate immunological reactions to an antigen, which is useful for vaccines, or can stimulate tolerance for use in organ transplantation.

Future applications may be in the cure of a wide variety of autoimmune diseases, e.g. type I diabetes, inflammatory bowel diseases, allergies, asthma; degenerative diseases, such as arthritis or athersclerosis, and cancers.

reference:
Webster KE, Walters S, Kohler RE, Mrkvan T, Boyman O, Surh CD, Grey ST, Sprent J. 2009. In vivo expansion of T reg cells with IL-2-mAb complexes: induction of resistance to EAE and long-term acceptance of islet allografts without immunosuppression. J Exp Med. Mar 30. [Epub ahead of print]

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]

Leptin and Diabetes

Fat cells produce the hormone leptin. Early experiments showed that leptin could ameliorate some of the effects of mouse models of type I diabetes, but it was always thought that the leptin just improved the activity of the small residual amounts of insulin produced by the rats. A recent experiment with genetically insulin deficient NOD mice, shows that leptin expressed by a virus infection reverses the symptoms of diabetes.

The NOD mouse strain is defective in the hormone, IL-2, that preserves tolerance to self proteins. As a result, a high percentage of the NOD mice develop symptoms of type I diabetes melitis, high blood glucose and ketosis. These animals will die without injected insulin.

In recent experiments, the NOD mice showing symptoms of diabetes were injected with a strain of adenovirus that had been engineered to express leptin. The leptin-expressing NOD mice displayed a rapid reversal of diabetes symptoms and began to gain weight normally. The high blood sugar and ketosis in NOD mice was stimulated by glucagon release in response to lack of insulin. Leptin stopped the glucagon release and reduced additional production of glucose by the liver and lipid oxidation that yielded the ketosis.

Muscle access to glucose in the NOD mice was restored by leptin by the induction of insulin-like growth factor 1 and its receptor. The NOD + leptin mice had normal blood sugar, but ate half the normal amount and had no body fat.

These are very promising results for type I diabetics, because it indicates that insulin injections are not the only approach to treatment. Stimulation of leptin production may be very useful and this new approach is being intensely investigated.

reference:
Yu X, Park BH, Wang MY, Wang ZV, Unger RH. 2008. Making insulin-deficient type 1 diabetic rodents thrive without insulin. Proc Natl Acad Sci U S A. 105(37):14070-5.

Inflammation Causes Disease

Human diets have changed dramatically over the last few hundred years, and as a consequence so have our diseases. The most recent shift in diet over the last hundred years has resulted in a shift from infectious diseases to degenerative diseases. This trend is summarized in the following Wikipedia entry.

Lifestyle diseases, from Wikipedia:

"Lifestyle diseases (also called diseases of longevity or diseases of civilization) are diseases that appear to increase in frequency as countries become more industrialized and people live longer. They include Alzheimer's disease, atherosclerosis, asthma, cancer, chronic liver disease or cirrhosis, Chronic Obstructive Pulmonary Disease, Type 2 diabetes, heart disease, nephritis or chronic renal failure, osteoporosis, acne, stroke, depression and obesity.

Death statistics in the United States
In 1900, the top three causes of death in the United States were pneumonia/influenza, tuberculosis, and diarrhea/enteritis. Communicable diseases accounted for about 60 percent of all deaths. In 1900, heart disease and cancer were ranked number four and eight respectively. Since the 1940s, the majority of deaths in the United States have resulted from heart disease, cancer, and other degenerative diseases. And, by the late 1990s, degenerative diseases accounted for more than 60 percent of all deaths.
Reference:
National Center for Health Statistics, National Office of Vital Statistics, 1947 for the year 1900 (page 67), for the year 1938 (page 55)."

My point here is that all of the so-called lifestyle diseases are also based on inflammation. I checked the research literature for studies of the response of each of these diseases to diets supplemented with omega-3 fish oils. Studies had been performed in each case. Reduction of inflammation by fish oil treatment was uniformly effective in reducing symptoms of all of the degenerative diseases. Other diseases that can be added to the inflammatory list are spinal disc problems and hypertension. It is interesting that disc dislocations are associated with coeliac, an inflammatory/autoimmune disease. It is also interesting that acne and depression are listed. Acne is indirectly associated with diet, but if sufferers shift to an anti-inflammatory diet, acne symptoms disappear. Depression associated with childbirth is particularly responsive to anti-inflammatory drugs, diet and exercise. Most of the symptoms associated with aging are just due to inflammation and are similarly responsive to anti-inflammatory lifestyle changes

To summarize:

• Modern degenerative diseases are caused by modern inflammatory diets (and insufficient exercise.)
• Anti-inflammatory diet and lifestyle reduce degenerative diseases.
• Aging is predominantly mismanaged inflammation.
  

Palmitoleate: omega-7 lipokine

Palmitoleic acid is responsible for keeping people healthy (lipokine) and for the smell of old people (nonenal). Overproduction of this lipid by blocking uptake of fats results in resistance to type II diabetes and atherosclerosis.

If you listen to the commercials on television, you know that there are two sources of fats/cholesterol; you either make it in your cells or take from your diet. Recent research shows that mice that have been genetically modified to lack cytoplasmic lipid carriers, can’t store dietary fat, so they make more of their own lipids. Specifically, they make more palmitoleic acid (C16:1n7). By U.S. standards, those defective mice are very healthy. It turns out that palmitoleic acid acts as a lipid hormone that communicates between fat tissue and other organs, and maintains a healthy metabolic balance.

Lipids are hydrophobic and require protein carriers to be moved from their source, such as the intestines, through the blood and to be offloaded into tissues. Most of the lipids enter the diet as triglycerides, i.e. a three carbon glycerol with three fatty acids attached. Those fats are extracted from food with bile, which is a mixture of modified cholesterol salts that acts as a detergent to dissolve fats. The dissolved fats, in the form of chylomicrons (big fat droplets coated with a lipid layer and proteins) are produced by intestinal cells and released into the blood stream. During transit, a lipase removes the fatty acids from the triglycerides. As in all of the lipid transport systems, the protein carriers determine how the lipid contents are distributed.

If a fatty acid or triglyceride is added to a cell membrane, the lipid would get stuck in the membrane's double layer of phospholipids. Fat droplets in cells are nothing more than fats that are loaded into the membrane of a cellular vesicle until a droplet covered by a half membrane forms. The alternatives for lipid transport are the HDL and LDL (protein coated lipids of the blood), and the intracellular fatty acid-binding proteins. The proteins bound to the surface of LDLs and HDLs bind to receptors on cell surfaces and control transfer of lipids to and from cells. One example is apolipoprotein E4. This protein is intimately involved in determining risk for athersclerosis and Alzheimer’s. ApoE4 binds to its cell surface receptor via heparin. Note the blue basic amino acids that form a massive heparin-binding domain down one side of the protein.

Fatty acid-binding proteins (FABPs) are just globular proteins, with hydrophobic amino acids arranged in the center and hydrophilic, water-bonding, amino acids on the surface. I have drawn the structure of a FABP using a graphics program called Chimera to visualize X-ray crystallographic data in the National Center for Biomedical Information (NCBI) database. The continuous chain of amino acids is shown as a white ribbon and the surface of the protein is shown as a transparent overlay. The protein chain makes a cage with the hydrophobic parts of the protein pointing toward the center to make a hydrophobic-lined container for the trapped fatty acid (pink). The two ends of the protein vessel are held closed by interdigitation of tryptophans (yellow) and basic amino acids (arginine and lysine, dark and light blue). The FABP also has a nuclear translocation signal, a group of four basic amino acids and other concentrations of basic amino acids displayed linearly across the surface of the cage (not shown), that probably are involved in transport of the trapped fatty acid from the cell surface to the surface of the nuclear envelope, which is involved in phospholipid assembly.

Mice engineered to have the human ApoE4 gene develop atherosclerosis and type II diabetes. If the FABPs of the fat cells of these mutant mice have also been removed, then the mice are essentially normal. Removal of the FABPs blocks the uptake of dietary fatty acids and stimulates the production of endogenous lipids, including the omega-7 fatty acid palmitoleic acid. This fatty acid is a lipid hormone, lipokine that stimulates normal metabolism and provides protection against several inflammation-based diseases. Interestingly, palmitoleic acid accumulates abundantly in the skin of old people and is converted to nonenal that has the smell of old books.

Cao H, Gerhold K, Mayers JR, Wiest MM, Watkins SM, Hotamisligil GS. 2008. Identification of a lipokine, a lipid hormone linking adipose tissue to systemic metabolism.Cell. 134(6):933-44.