Health and Heparan Sulfate Circulation — Connective Tissue is Alive

--- The other 200 posts are here ---

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 III — Inflammation from Cell to Tissue

---All 200 Posts---

I have explained my perspective in diagrams of the relationship between diet, gut flora and disease:

Health Diagrams I — Gut Flora and Diet

and of the interaction between gut flora, the immune system and autoimmunity:

Health Diagrams II — Curing Autoimmunity and Allergies

Now I am discussing how inflammation, the foundation of most chronic diseases, begins at the cellular level and results in the classic symptoms of tissue inflammation: redness, heat, swelling and pain.

NF-kB is the Transcription Factor that Controls Inflammation Genes

Of the 23,000 human genes, about 1,000 on each of 23 chromosomes, five dozen, e.g. enzymes involved in nitric oxide (vasodilation and erection hormone), synthesis of heparin sulfate and prostaglandin synthesis from omega-6 fatty acids or cytokines (IL-1, IL-6, TNFa), are associated with inflammation.  These inflammatory genes are turned on or expressed in individual cells, when the inflammation transcription factor, NF-kB, is activated by any of numerous external signals, including inflammatory cytokines, bacterial or fungal cell wall materials (LPS or beta-glucan), advanced glycation end products (AGE, e.g. HgA1C, resulting from high blood sugar) or reactive oxygen species (ROS, e.g. super oxide, from insulin resistance).

NF-kB and Inflammation
Superoxid Causes Insulin Resistance

Inflammation is the Foundation of Growth, Birth, Cancer and Pain

We think of inflammation as the sum of physical symptoms, and our purpose in responding to inflammation is typically to limit its impact.  We try to stop swelling by applying cold or hot, and we take aspirin to lower fevers and stop pain.  We fail to realize that inflammation is essential to the growth and development of many different tissues, and that inflammation is a cycle that leads back to normal function.  

Body tissues, such as the lining of the intestines or the uterus, continually produce new cells to replace the old that are sloughed off.  NF-kB must be turned on for these growth and attrition cycles.  Taking aspirin blocks NF-kB in the gut and stops local development of the lining, resulting in weak areas that bleed.  That is why doctors encourage patients to drink a half glass of water before and after swallowing aspirin tablets. 

Another more dramatic example of control of inflammation is conception, gestation and birth.  Conception and gestation require inhibition of inflammation, to permit growth of a foreign organism (a fetus is half sperm genes) in the uterus.  Chronic inflammation limits the ability of the uterus to suppress immune attack and can produce infertility, which is treated by aspirin and heparin, which suppress chronic inflammation.  The return of inflammation at the end of gestation precipitates labor and birth.  Excess Inflammation produces high levels of circulating inflammatory cytokines, which causes postpartum depression.  Depression and chronic inflammation have the same cytokine profiles, i.e. depression is a symptom of chronic inflammation.

Birth and Inflammation

Proliferation, or enhanced cell division, is another aspect of inflammation and is also the foundation for cancer.  That is the reason that some doctors recommend low dose aspirin to reduce colon cancer.  Similarly, since inflammation is the basis for coronary artery disease, doctors sometimes recommend low dose aspirin, although this is controversial.  Doctors also use aspirin as a so called blood thinner, since it blocks inflammatory signaling in platelets and discourages clotting.  Inflammation of nerve cells is experienced by the brain as pain.  

When it is understood that inflammation is an essential feature of many normal, healthy cell and tissue functions, then “inflammation," with its negative connotations, becomes a misnomer.

NSAIDs Inhibit Inflammatory Prostaglandin Production

Aspirin directly inhibits NF-kB activation inside the cell, but it also chemically modifies COX, the enzyme that converts omega-6 polyunsaturated fatty acids (common in polyunsaturated vegetable oils) into inflammatory prostaglandins.  Other NSAIDS (Non-Steroidal Anti-Inflammatory Drugs) just inhibit COX, but Aspirin transfers its acetyl group to make acetyl-COX, which has a new activity that converts omega-6 fatty acids into anti-inflammatory prostaglandins.  The high omega-6 fatty acid content of vegetable/seed oils, such as corn, soy, canola, etc. is why these oils, in contrast to olive oil or butter, are inflammatory.  Omega-3 fish oil is anti-inflammatory, because it is converted to anti-inflammatory prostaglandins.  Plant omega-3 fatty acids are shorter and are not converted to prostaglandins, but inhibit omega-6 conversion.

Aspirin

Nitric Oxide, Vasodilation and Viagra

Swelling is caused by vasodilation, the relaxation of blood vessels, and accumulation of serum in the tissue.  This vasodilation also makes the tissue red and warm from the increased amount of warm blood in the capillaries.  Vasodilation is caused by nitric oxide, NO, that is produced by an enzyme under the control of NF-kB, which takes the nitrogen from arginine (or nitroglycerine).  The NO diffuses easily and binds to receptors that produce an amplified signal, cyclic GMP, that relaxes the muscle cells surrounding blood vessels.  [Viagra is potentially dangerous, because it just exaggerates the amplified signal and obscures the underlying vascular damage, e.g. hypertension, that causes erectile dysfunction by blocking normal vasodilation.]

Nitric Oxide and Erectile Dysfunction

Hot/Cold and Endorphins

The dilemma of whether to use hot or cold therapy to block inflammation is based on a misunderstanding of what the temperature changes are actually doing.  Changing the temperature of the skin alters the structure of sensory proteins in nerves of the skin and triggers signals to the brain that register as hot or cold.  Chemicals, e.g. capsaicin or menthol, can have the same effect without changing skin temperature.  The important response for inflammation control, is return signals from the brain that release neurohormones, e.g. endorphins, from different nerves that reach not only some of the skin that was hot or cold, but also deeper tissue.  The endorphins block inflammation and all of its symptoms.  That is why chemically treated pads are more effective than icing or changing from hot to cold, because "hot" and "cold" signaling chemicals can be applied simultaneously.  None of the treatments is more than skin deep.  Actually chilling or heating tissue below the skin is damaging and causes more inflammation.  Low dose Naltrexone may be effective in some cases of chronic inflammation, by stimulating systemic rebound endorphin production.

Dr. Oz, Pain, Hot/Cold Receptors

Lymphocyte Offloading, Mast Cells, Heparin

Rosacea is a group of diseases that involve inflammation of the face in an exaggerated blush.  Any of the signals that would lead to blushing cause intense vasodilation.  A blush is fleeting, but rosacea is made chronic by another aspect of inflammation, offloading of lymphocytes.  Large numbers of lymphocytes accumulating in response to a local infection would produce pus.  In the case of rosacea, the distributed leucocytes, including neutrophils, respond to the blushing signals by producing inflammatory signals, such as P protein.  The result is cycles of inflammation, autoinflammation.

Mast cells can also be offloaded from blood vessels and provide a link between the immune system and inflammation.  Mast cells display IgE receptors on their surfaces, which bind antigens and trigger release of histamine, heparin and protease.  Histamine is a neurotransmitter that binds to receptors on blood vessels and nerve cells.  In the gut, histamine mediates many digestive processes.  Heparin released along with  histamine, coats the gut and prevents attachment of pathogens by competing for binding to the heparan sulfate proteoglycans (HSPGs) that form the surface of cells that line the gut.  [Heparin is the most common drug used in hospitals and is produced from intestines of cattle and hogs in the meat industry.]  Heparin also binds and inactivates the proteases released from mast cells.  Upon release, the now active proteases attack and activate receptors on nerves and immune cells.

Mast Cells and Heparin

Heparin is Anti-Inflammatory

Heparin is the most negatively charged polysaccharide, mediates most of the receptor/hormone interactions at cell surfaces; facilitates amyloid plaque formation, e.g. in Alzheimer's, atherosclerosis, diabetes, dementia; and controls numerous protease reactions in the complement system and clotting, etc.  There are hundreds of heparin-binding proteins.  Heparin is produced in secretory granules of mast cells by the action of heparanase on heparan sulfate proteoglycans. Heparin is a mixture of small fragments, oligosaccharides of heparan sulfate polysaccharides.  Heparin is anti-inflammatory and is administered to facilitate conception and gestation.  Inflammation also inhibits the genes involved in heparan sulfate proteoglycan production and since HSPGs are a major component of basement membranes of tissues and provide the barrier function of blood vessels in kidneys and brain, inflammation leads to proteinuria and loss of the blood brain barrier.  Since HSPGs have a short half life of six hours and are rapidly recycled, heparin added to the blood is rapidly absorbed by vessels, and heparin taken orally is absorbed by intestinal cells, but does not reach the blood.  HSPGs and heparin are central components of immunity and inflammation.

Heparin and Inflammation

Inflammation Blocks Skin Synthesis of Vitamin D from Cholesterol

Inflammation blocks solar synthesis of vitamin D in the skin and is more important than skin pigmentation, use of sunblock or latitude in producing vitamin D deficiency.  The vitamin D content of food is negligible compared to solar production in the skin.  It is not surprising that rising chronic inflammation is also accompanied by rising vitamin D deficiency.  Vitamin D supplementation is usually ineffective in curing vitamin D deficiency, because the supplements are too low and very high levels of supplemental vitamin D are required to reverse underlying chronic inflammation.  Statins are very effective at blocking cholesterol synthesis and although reducing cholesterol has minimal impact on the target, cardiovascular disease, it dramatically reduces vitamin D causing muscle pain, etc.

Most vitamins are enzyme cofactors synthesized by gut bacteria and used as quorum sensing signals during formation of biofilms.  Vitamin D, in contrast, is a steroid hormone and receptors for vitamin D are inside cells.  The receptor/vitamin D complex is transported into the nucleus where it acts as a transcription factor to control the expression of genes.  Vitamin D controls the expression of defensins in the crypts of the villi of the small intestines.  The antimicrobial activity of defensins is based on the basic amino acids (arginine and lysine) of its heparin binding domains.  Vitamin D also interacts with NF-kB in the nucleus and modulates inflammation.

Inflammation and Vitamin D

Bacteria and LPS

Lipopolysaccharide is a wall component that is indicative of bacteria, just as beta-glucan is indicative of fungi, and both are intense activators of NF-kB and inflammation.  LPS is released from damaged bacteria, e.g. by antibiotic treatment, binds to receptors on the surface of intestines and stimulates inflammation with release of NO, which produces diarrhea.  Food intolerances, which are based on incomplete digestion of food components, because of an incomplete gut flora (immunological responses/food allergies are rare) are probably also the result of LPS release from gut flora and inflammation.

Innate Immunity is also Triggered by LPS

The basic defenses of humans against microorganisms are mediated at the cellular level by triggering molecules common to all microorganisms, e.g. LPS for bacteria.  The responses are equally general: lysozyme to digest bacterial wall peptidylglycan, lactoferrin that binds iron and yields antibacterial peptides.  LPS (and inflammatory cytokines) also stimulates the liver to produce CRP (C Reactive Protein) that binds to choline on bacteria as the first step in phagocytosis and DNAse I that digests NETs (neutrophil extracellular traps) that are the DNA and histones released by triggered neutrophil cells that enmesh bacteria for engulfment by phagocytic cells.  [NETS plug peripheral catheters and can be cleared with probiotics that stimulate DNAse I release from the liver.]  NETs are also present at sites of inflammation and the accompanying nuclear proteins have the basic triplets that stimulate immune presentation and act as autoantigens, i. e. produce anti-nuclear antibodies, in the absence of adequate Tregs.

Diet and Inflammation

The diagram outlines the interactions that produce the tissue symptoms of inflammation.  Many components of modern diet can trigger inflammation:

Sugars and high glycemic starches raise blood sugar and enhance AGE/HgA1C.

Vegetable oils high in omega-6 oils are converted into inflammatory prostaglandins.

Wheat and other grains have high glycemic starch and insoluble fiber that is inflammatory.  Gluten is inflammatory.

Antibiotics damage the gut flora and produce vitamin deficiencies, autoimmunity and allergies.

Food intolerances result from damaged gut flora and produce gut inflammation.

Fish high in omega-3 EPA and DHA are anti-inflammatory.

The Anti-Inflammatory Diet

Health Results from a Balance of:

Diet (meat, fish, eggs, dairy, vegetables), containing macronutrients of protein, starch 30-100 g/d and fat (low omega 6/3 and saturated fat for most calories), and micronutrients

Soluble Fiber, e.g. resistant starch (consult Free the Animal), inulin, pectin, (plant polysaccharides, animal GAGs)

Gut Flora, diverse and adapted to dietary soluble fiber,

Exercise

Resistant Starch, Panacea, but Why?

Mark’s Daily Apple provides an authoritative diet guide (except for the gut flora).

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.

Heparin, Growth Factors and Rosacea

Knock-out Mice and FGF Receptor Inhibitors Mimic Rosacea
Heparin Nanofibers Loaded with VEGF and FGF Mimic Stem Cells

In previous articles, I have emphasized the mediation of extracellular signaling by heparan sulfate proteoglycans (HSPGs, polysaccharides attached to proteins) and heparin (HS fragments, oligosaccharides) and the sensitivity of HSPG expression and HS degradation by inflammation.  I return to that subject, spurred on by reading two articles that together show both the significance of heparin-mediated growth factors in general and in the specific case of symptom development in rosacea.

FGF Receptor Inhibitors Cause Symptoms Like Rosacea
Fibroblast growth factors stimulate the development of cancers, and antibodies against FGF receptors block cancer growth (see ref.)  FGF receptor inhibiting antibodies are now being used to stop cancers.  Unfortunately,  FGFR antibodies (e.g. cetuximab, panitumumab) also cause symptoms in the skin (telangiectasia, acneiform eruption) similar to the facial inflammation of rosacea.  Similarly, in knock-out mice, that lack the ability to produce FGFR, there are related symptoms.  It appears that lack of some FGF signaling may produce the symptoms of visible blood vessels and pus-filled (though lacking bacteria) follicles of rosacea.

FGF Mediated by HSPG
FGF binds to the heparan sulfate of membrane bound HSPG in pairs and these FGF dimer/heparan sulfate complexes activate a pair of FGF receptors.  The result is activation of protein phosphorylation activity (tyrosine kinase) and normal skin development.  HSPG synthesis is modified by inflammation and heparanase activity is increased.  This suggests that inflammation will decrease FGF signaling and could lead to symptoms of rosacea.

Growth Factors (VEGF, FGF) Bind to Heparin Nanofibers that Mimic Stem Cells
Stem cells produce lots of different growth factors and when stem cells are introduced into damaged cardiovascular tissue, more healing results (see ref.)  To determine if the growth factors produced by the transplanted stem cells was sufficient for the improved healing, fibers made of heparin were dipped into stem cell cultures and the resulting growth factor-coated fibers were injected into damaged tissue.  The heparin-binding growth factors were just as effective at enhancing healing as were the stem cells in previous experiments.  This demonstrated that heparin-binding growth factors were the key to normal repair/revascularization and function.

Rosacea Results from Inflammation and Aberrant Vascularization
Rosacea is poorly understood and is probably numerous diseases that have related symptoms and complex development.  As I indicated in previous articles, neurotransmitters from stimulated facial nerves, enzymes (kallikrein) and cytokines from intestinal interactions with gut flora, mast cell products (heparin, protease) and modified antimicrobial peptides (cathelicidins), as well as cryptic bacteria in facial tissues, may all be involved.  Inflammation in the skin of the face and in the intestines is involved.  Vitamin D, omega-3 fatty acids and anti-oxidants have a variety of responses (sometimes paradoxical) that differ from individual to individual and at different stages in the development of the disease.  Facial inflammation leads to abnormal development of blood vessels (telangiectasia) and in accumulation of lymphocytes and neutrophils (papulopustular rosacea).

Facial Inflammation May Depress HSPG Production and Disrupt FGF Function
One of the key ramifications of persistent facial inflammation may be the depletion of of HSPGs that normally coat cells.  HSPGs are continually produced, reabsorbed and degraded.  The half-life for HSPGs, even those that surround the cells that produce cartilage in connective tissue, is six hours.  HSPGs are also the source of heparin, that is produced as a counter ion bound to histamine and proteases in the secretory granules released by activated mast cells.  Thus, inflammation-based depression of HSPG production, which is also accompanied by heparanase activation, will remove the HSPG coating of cells.  This HSPG coating is needed for normal growth factor function.  Lack of an HSPG matrix on the surface of cells will also result in the migration of growth factors away from where they are normally functional and into adjacent tissue where they may stimulate aberrant development of blood vessels.  This may explain telangiectasia.

Is Topical Heparin a Rosacea Treatment?
Topical heparin does penetrate the skin.  It would appear to be a logical treatment, if HSPG depletion is contributing to symptom development in rosacea.  The length of the heparin fragments may be important.  I am unaware if anyone has tried the heparin lotions that are available for treatment of wounds to minimize scarring, on rosacea.  Heparin may be useful in combination with vitamin D3 and remediation of gut flora in a general scheme to treat rosacea.

refs:
Segaert S, Van Cutsem E.  Clinical signs, pathophysiology and management of skin toxicity during therapy with epidermal growth factor receptor inhibitors.  Ann Oncol. 2005 Sep;16(9):1425-33. Epub 2005 Jul 12.

Webber MJ, Han X, Prasanna Murthy SN, Rajangam K, Stupp SI, Lomasney JW.  Capturing the stem cell paracrine effect using heparin-presenting nanofibres to treat cardiovascular diseases.  J Tissue Eng Regen Med. 2010 Mar 10. [Epub ahead of print]

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.

An Autoantigen for Pancreatitis

Pancreatic Secretory Trypsin Inhibitor (PSTI) Has Internalization Basic Triplet

Pancreatitis is an inflammation of the pancreas resulting from lack of adequate inhibition of proteases. Autoantibodies against PSTI would explain some forms of pancreatitis.

I was researching the maintenance of baby gut flora by mother’s milk, when the reference discussed here was brought to my attention by my wife, who happens to be a lactation consultant. The paper showed that PSTI is present in colostrom, the first milk that a baby gets, before the true milk comes in. PSTI protects the new gut from digestion by its own pancreatic proteases, since PSTI is a protease inhibitor that sticks to the gut.

I naturally assumed that PSTI stuck to the gut by heparin-binding domains that would stick to the heparan sulfate proteoglycans on the gut surface. [Recall that it is via these HSPGs that viruses and bacteria infect the gut and the HSPGs in turn are protected during infections by the release of heparin from mast cells. The heparin in the guts of cattle and pigs are used to make commercial heparin to block blood clotting.] So I looked up the structure (above, with basic amino acids in blue and basic triplet on right) sequence of human PSTI at NCBI:

>gi|190694|gb|AAA36522.1| PSTI
MKVTGIFLLSALALLSLSGNTGADSLGREAKCYNELNGCTKIYD
PVCGTDGNTYPNECVLCFENRKRQTSILIQKSGPC

The basic triplet (RKR,arg-lys-arg), from my perspective, should result in presentation to the immune system during high levels of inflammation, and as a consequence result in autoantibodies against PSTI. The result would be the neutralization of the protease inhibitor and damaging production of active protease to attack the pancreas, i.e. pancreatitis.

It would be fairly easy to test this hypothesis by looking for the anti-PSTI antibodies in some people with pancreatitis. Other autoantibodies, e.g. against tissue transglutaminase, might also be checked, because the inflammation that produced one autoantibody may produce others and both PSTI and tTG are produced in the intestines. In fact, celiac may be the cause of some cases of autoimmune pancreatitis.

Note added in proof:

I just checked the literature on PubMed and found that PSTI is in fact an autoantigen in pancreatitis and produces antibodies against PSTI:
Raina A, Greer JB, Whitcomb DC. Serology in autoimmune pancreatitis. Minerva Gastroenterol Dietol. 2008 Dec;54(4):375-87.

and
I found that pancreatitis is often found associated with celiac (gluten intolerance):
Patel RS, Johlin FC Jr, Murray JA. Celiac disease and recurrent pancreatitis. Gastrointest Endosc. 1999 Dec;50(6):823-7.

ref:
Marchbank T, Weaver G, Nilsen-Hamilton M, Playford RJ. Pancreatic secretory trypsin inhibitor is a major motogenic and protective factor in human breast milk. Am J Physiol Gastrointest Liver Physiol. 2009 Apr;296(4):G697-703.

Suffering from Inflammation?

How do you know if your symptoms result from inflammation?

My interest is the molecular basis of inflammation, how inflammation is triggered and how inflammation contributes to numerous diseases. I try to expose the inflammatory underpinnings of various diseases by initially linking a disease to inflammation and then unraveling the molecular events that lead to and make up the disease.

How Do I Link a Disease to Inflammation?

My first task is to check the biomedical literature to see if there are research articles that support anti-inflammatory interventions that prevent or limit the disease. I just do a PubMed search the disease name plus anti-inflammatory treatments, e.g. omega-3 fish oils, vitamin D, NSAIDs, etc. It is also possible to see if a disease, such as diabetes, that produces chronic inflammation is a risk factor for the new disease being examined. It is shocking to me that omega-3 fish oils (EPA/DHA) or even flax seed oil, have been found to be effective treatments for numerous diseases that range from allergies, arthritis, inflammatory bowel diseases, depression and even septic shock and multiple organ failure. Aspirin has been used to treat infertility and post partum depression, and at high levels to treat cancer.

Dietary Suppression as Prima Fascia Evidence of Inflammatory Cause

If I find that omega-3 oils have been used successfully to treat a disease, then I attempt to link inflammation to the molecular events that initiate the disease. The biomedical literature is of minimal help here. [Biomedical research is usually limited to assessing the impact of drugs on the symptoms of diseases, so the biomedical literature typically does not provide information on the cause of diseases or ways to cure diseases. Causes and cures do not receive research funding.] I have to learn the basic workings of the organs involved and the alterations of function associated with the disease. I have also found by long experience, that major molecular components are systematically missing from typical explanations of function.

Heparan sulfate/heparin Is Missing in Action

Heparan sulfate proteoglycans (HSPGs) dominate the extracellular environment and yet they are systematically excluded from biomedical research. On this blog, I have provided dozens of examples of the essential role played by HSPGs and disruption of these roles by heparin. The majority of cytokines, growth factors, clotting events, complement cascades and even lipid transport (LDL) act via HSPGs. Leaking of proteins into the urine, across the intestines or the blood brain barrier is controlled by HSPGs, is reduced by inflammation and can be partially repaired by heparin. Autoimmune and allergic diseases are initiated by disruptions in HSPG metabolism. Viral and bacterial pathogens bind to human cells via HSPGs. Cancer cells reduce their HSPGs and start to secrete heparanase in order to metastasize. Mast cells secrete heparin! HSPGs and heparin are major players in tissue function and yet the major cell biology text book does not even discuss them. HSPGs are not mentioned in medical school training even though heparin is the most commonly administered drug.

One of the insights that I bring to my conceptualization of diseases is the role of heparan/heparin in cellular physiology. It explains a lot.

Check for Inflammatory Symptoms by Trying the Anti-Inflammatory Diet

If your symptoms are due to inflammation, there is an easy way to find out. Since diet is the biggest source of inflammation and most of the cells of the immune system congregate in your intestines, it makes sense to check to see your health problems are rooted in inflammation by making simple changes in your diet. Since this is just a test, don’t worry about whether or not this is diet for the rest of your life. Just stick to it for a week and see if it changes your life.

The Basic Anti-Inflammatory Diet and Lifestyle Guidelines are here.

(Vitamin D and omega-3 fish oil amounts are minimal levels. More severe examples of inflammation will require higher levels. Vitamin D up to 10,000 IU per day has been found safe. Some individuals require up to 12 fish oil capsule per day to experience relief from symptoms. Increases should be gradual over weeks of time.)

Try it for a week and let me know if your symptoms disappear. The prevalence of diet-based inflammation, makes me confident that you will be glad that you tried these simple, healthy changes. For immediate relief of pain, see my articles on capsaicin, castor oil and menthol/Vicks.

This is not medical advice and is used only in appropriate support of primary medical care.

Extreme Flu Remedies

Experimental Therapies for ARDS, Cytokine Storms

Do not do this at home. There are doctors and hospitals. Use them.

....But, if a doctor emailed me pleading for any ideas that I had to save a bunch of patients suffering from acute respiratory distress syndrome (ARDS) from Tamiflu-resistant H1N1, my first response would be to suggest therapies designed for ARDS from other origins, e.g. burns, septicemia, etc.

Cytokine Storms Are Out of Control
When too much tissue is injured, the local, molecular communication that normally occurs just between cells, spills into the blood stream and becomes potentially lethal. That is what happens in anaphylactic shock. It is also what happens in cytokine storms, where inflammatory cytokines that are normally short-lived and processed locally to progress into recovery, erupt into the blood stream and impact distant organs.

Major disruption of body function by aggressive blood infections or burns over most of the body, will be lethal without heroic medical interventions. These are injuries beyond the evolved adaptations of mammals.  Until recently there were no survivors.

Influenza has been around for a long time. Humans, other mammals and birds get the flu and get over it. Many body cells become infected, antibodies specific to the virus are produced within about a week, the infected cells are killed, the virus is digested and life goes on.

People die from the flu, because an opportunistic pathogen causes a lethal secondary infection, or the body over-reacts and damages itself in attempts to attack its own infected cells. This is a cytokine storm.

Silence the Storms
Cytokine storms can be weathered by blocking the signaling system. Cytokines are just small proteins that are complementary in shape to corresponding protein receptors that penetrate through the surface membranes of cells throughout the body. Binding of cytokine to receptor changes the shape of the receptor and transmits a signal into the cytoplasm of the receptive cell. This turns on aggressive behavior of immune cells and triggers more inflammatory signaling in other cells. This causes fever, malaise, etc.

...But, I was the one the doctor is pleading with to save the people. And I know that there is more to cytokine signaling than just cytokines and receptors. There are also heparan sulfate proteoglycans (HSPGs). Cytokines are not supposed to be broadcast throughout the body. Cytokines function in the space between cells, the extracellular matrix. Polysaccharides attached to membrane proteins, HSPGs, are secreted at one end of the cells, sweep across the surface and are taken back up at the other end. Cytokines have heparan-binding domains and so they stick to the heparan and are swept along. Cytokines can move from one cell to another as the sweeping HSPGs of adjacent cells come in contact.

HSPGs Mediate Cytokine Signaling
The critical point here is that cytokines bind to their receptors with the heparan between -- the cytokine and receptor are like two halves of a bun and the hot dog is the heparan. In fact the heparan bridges two cytokine/receptor complexes to make an active, signaling pentamer.

Heparin Can Block Cytokine Signaling
Heparin is a fragment of heparan sulfate produced by enzymatic degradation of HSPG. Commercial heparin, used to block blood clotting, is obtained from the mast cells of lungs and intestines of hogs and cattle. The mast cells release heparin and histamine in response to parasites or pollen. Since heparin is a short version of heparan sulfate, it can block the formation of active cytokine/receptor complexes.

Heparin is used in a mist to treat the lungs of burn patients. It is also injected into some infertility patients to suppress inflammation that is inhibiting implantation and gestation. It is also effective in treatment of autoimmune inflammation in Crohn’s disease. I think it should be tested as a therapy for H1N1 cytokine storms. It may be useful in nebulizing mists and oral treatment of intestines.

Berberine Binds to HSPG
Berberine is a phytochemical from Barberry traditionally used in the treatment of intestinal infections and arthritis. It also binds to heparan sulfate to form fluorescent complexes visible in microscopy. Berberine-treated mast cells glow brightly. Heparan sulfate can also be detected in Alzheimer’s plaque, atherosclerotic plaque and prion complexes. Because berberine binds to heparan sulfate, it should also disrupt cytokine signaling. It has been used successfully in treatment of septicemic ARDS.

Curcumin Blocks NFkB
One of the most potent chemicals that blocks inflammatory signaling via the inflammatory transcription factor, NFkB, is curcumin. Curcumin is a major component of the spice turmeric. Oral curcumin can be enhanced by co-administration of black pepper, because the piperine in pepper inhibits intestinal inactivation.

Anti-Inflammatory Diet
Of course, I would also recommend vigorous implementation of an anti-inflammatory diet and lifestyle to support any medical treatment.

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.

Synuclein and Amyloid Diseases

NSAIDs, such as ibuprofen and aspirin are possible treatments to inhibit the aggregation of proteins (synuclein, beta amyloid) on charged polymers in amyloid diseases, such as Parkinson’s disease, Alzheimer’s disease, etc. Contradictory studies show that intracellular aggregate formation may be protective, since dimers are more toxic than aggregates.

The list of amyloid diseases is long and there are few effective treatments. In each case a protein starts to accumulate in fibers that form amyloid plaques inside or outside the cells. The large aggregates outside are toxic. Inside it appears that the large aggregates are not as toxic as small clumps, oligomers, of the protein.

The amyloid proteins are stacked up in the fibers in a very organized way, so that the same portions of the protein are lined up on each side of the fibers. Outside the cell, the regions with basic amino acids interact with heparin, and in Alzheimer’s disease, for example, the beta amyloid plaque is half heparin. In test tube experiments, fiber formation from protein solutions is accelerated by adding heparin.

Amyloid fibers also form inside cells in the case of the tau fibers of Alzheimer’s disease or the synuclein aggregates in Parkinson’s disease. In theses cases, there should not be any intracellular heparin, and it is not known what polyanion (RNA?) serves to accelerate fiber formation in these cases.

Non-steroidal anti-inflammatory drugs (NSAIDs), such as ibuprofen and aspirin, reduce the incidence of Parkinson’s and Alzheimer’s diseases. It has recently been shown that in test tube experiments, NSAIDs also decrease the formation of amyloid fibers from synuclein.

Amyloid fibers can be stained by Congo Red and thioflavin. Curcumin is the active component of tumeric and it has a structure related to Congo Red. Curcumin has been shown in recent studies to block synuclein amyloid formation.

In addition, the heparin in the fiber complexes can be stained with berberine. Berberine is a traditional herbal treatment for arthritis. It would not be surprising if it was also effective against Alzheimer’s amyloid plaque.

The large extracellular plaque aggregates appear to be toxic, but the small, oligomeric aggregate of protein appear to be the toxic form in cells. Recent experiments show that facilitating the formation of large intracellular aggregates minimizes the toxicity in animal models of Huntington’s and Parkinson’s diseases. It appears that the large visible aggregates are not the form that kills the cell.

For the time being, the only safe treatments that focus on amyloid fiber formation are the NSAIDs, curcumin and perhaps berberine.

references:
Hirohata M, Ono K, Morinaga A, Yamada M. 2008. Non-steroidal anti-inflammatory drugs have potent anti-fibrillogenic and fibril-destabilizing effects for alpha-synuclein fibrils in vitro. Neuropharmacology 54(3):620-7.

Pandey N, Strider J, Nolan WC, Yan SX, Galvin JE. 2008. Curcumin inhibits aggregation of alpha-synuclein. Acta Neuropathol. 115(4):479-89.

Bodner RA, Outeiro TF, Altmann S, Maxwell MM, Cho SH, Hyman BT, McLean PJ, Young AB, Housman DE, Kazantsev AG. 2006. Pharmacological promotion of inclusion formation: a therapeutic approach for Huntington's and Parkinson's diseases. Proc Natl Acad Sci U S A. 103(11):4246-51.

Outeiro TF, Kontopoulos E, Altmann SM, Kufareva I, Strathearn KE, Amore AM, Volk CB, Maxwell MM, Rochet JC, McLean PJ, Young AB, Abagyan R, Feany MB, Hyman BT, Kazantsev AG. 2007. Sirtuin 2 inhibitors rescue alpha-synuclein-mediated toxicity in models of Parkinson's disease. Science. 317(5837):516-9.

HIV TAT and Methamphetamine -- TNF

HIV infection and methamphetamine both cause inflammation of the brain and together they are paralyzing.

What’s worse than a TNF typhoon resulting from methamphetamine use? The answer is a TNF typhoon resulting in dementia from HIV infection of the brain.
Combine methamphetamine with HIV in the brain and the result is a Parkinson’s type of paralysis.

This sounds like very morbid subject matter to pursue out of curiosity, but if you put heparin into the equation, as I always do, it all becomes very interesting.

Here are pieces to the big picture:

HIV, the AIDS virus, infects cells of the immune system and causes chronic inflammation. The inflammation causes a disruption of heparin metabolism and since heparin is a major part of the matrix that holds together the endothelial cells that line the capillaries that feed the brain, the capillaries leak, i.e. there is a leak in the blood brain barrier. HIV-infected cells can pass out of the capillaries and into the brain. Here comes the insidious part, HIV produces a protein called TAT.

I drew a graphic of TAT with the basic amino acids in blue. The sequence of this nasty little protein shows how it gets around. It is secreted from cells, attached to heparan sulfate proteoglycans. It sticks tightly to heparin (yellow and red stick figures, sticking to ribbon of TAT), because of the patches with three and four adjacent basic amino acids. Frequently, the TAT will just be secreted and then sweep over the surface of the infected cell and be brought back into the cell on the circulating HSPG.

DPVDPNIEPWNHPGSQPKTACN
RCHCKKCCYHCQVCFIKKGLGI
SYGRKKRRQRRRPSQGGQTHQ
DPIPKQPSSQPRGDPTGPKE

A protein with three adjacent basic amino acids will get swept into a cell. All allergens that I have examined have this internalization triplet of basic amino acids. TAT is so powerful, that if it is chemically linked to the larger fluorescent protein from jelly fish (left with green, fluorescent amino acid derivative down the center), the whole fluorescent protein is dragged into cells.

The TAT can move from the HSPGs of an HIV-infected cell to neighboring cells with HSPGs. The TAT then gets taken into the cytoplasm of the next cell. Four adjacent basic amino acids are the signal that transports a protein to the nucleus and into the nucleus. It is in the nucleus that TAT really causes trouble. The TAT can move from an infected immune cell in the brain to neurons. TAT can kill neurons and stimulate other cells to produce TNF.

Methamphetamine also causes a TNF storm in the brain. This is a quick way to start the wasting symptoms that TNF in known for -- it is also call cachexin, after the wasting process of cachexia. If methamphetamine is given to someone with neurological symptoms of HIV, then the neuropathology is further exaggerated into a Parkinson’s type of paralysis. The TNF production of both is additive.

TNF production by methamphetamine brings up the consequences of the very closely related compound amphetamine (Adderall, Dexedrine) used by children and young adults (college age) as a treatment for ADHD.

reference:
Theodore S, Cass WA, Nath A, Maragos WF. 2007. Progress in understanding basal ganglia dysfunction as a common target for methamphetamine abuse and HIV-1 neurodegeneration. Curr HIV Res. May;5(3):301-13.

Hazards of Air Travel: DVT

Deep Vein Thrombosis (DVT) -- clots in your veins

Air travel during the holidays means sitting quietly for hours while the blood pools in the major veins of your legs. This is a test. How have you been eating lately? If you stuck to an anti-inflammatory diet and got your exercise, just fidgeting a little and flexing your legs ever once in a while should avoid clots. If you are the typical sedentary American with an inflammatory diet, then worry. One tenth of you will typically have clots in your leg veins after a long flight.

Rolling stones gather no moss, and the same is true for rapidly moving red blood cells (RBCs). Keep them moving and they don’t stick together. Slow down RBCs traveling along sticky vessel walls and you have problems. RBCs have no nuclei and since the intracellular secretory system originates from the outer membrane of the nucleus, red blood cells don’t secrete anything. RBCs just age until they are removed by the spleen. So RBCs just move passively with the rest of the blood.

Another player in clot formation is the platelet. Platelets are cell fragments. They are formed by extrusion and shearing. The process is like bubbles forming as you blow air through a child’s bubble wand. Cells in the bone marrow are squeezed through a grid and the extruded fingers of the cells are blown away in the blood flow as platelets. The electron micrograph shows a platelet between and RBC and a white blood cell. Platelets don’t have any active cell machinery, so they are just little bags containing secretory vesicles that can be released by triggering of receptors on their surface. Platelets are only good for one shot of release.

Platelet release of secretory contents is triggered by norepinephrin, ADP and PGI2, an inflammatory prostaglandin produced from the omega-6 arachidonic acid. Norepinephrin is one of the fight-or-flight hormones that prepares the vascular system for damage control. ADP is released from other activated platelets and insures that isolated platelets are not randomly activated.

One of the proteins released is platelet factor 4. I have illustrated PF4 and the strip of basic amino acids (blue) that girdles the protein are readily apparent. PF4 binds strongly to heparin. Since the clotting process is normally under heparin inhibition, PF4 release from platelets removes the heparin inhibition and promotes clotting. ADP is also released and promotes further activation of other platelets.

Clot formation occurs in response to stress (norepinephrin), damage (vascular inflammation) and a consensus of platelets (ADP). Chronic inflammation can mimic this combination of signals through its impact on heparin metabolism. My research suggests that inflammation lowers heparin synthesis. An example of this effect is kidney damage caused by diabetes. High blood sugar causes inflammation of the kidney blood vessels, this reduces heparin production and since heparin lining the vessels is needed to retain proteins as blood is filtered in the kidney, protein is lost into the urine, i.e. proteinuria. Similarly, chronic inflammation can disrupt the blood brain barrier that is also made up of heparin.

A major source of chronic inflammation is an inflammatory diet. A recent research study indicated that a typical inflammatory American diet leads to elevated risk for deep vein thrombosis. Alternatively, an anti-inflammatory diet rich in B vitamins and omega-3 oils minimized DVT. Saturated fats had no impact, consistent with the lack of evidence supporting the shift from saturated fats to toxic omega-6-rich polyunsaturated vegetable oils.

So, the best thing that you can do to protect yourself from clots when you travel over the holidays, is to eat right and get your exercise, before you travel. Avoid starch (in large amounts) and polyunsaturated vegetable oils (except olive oil.) Corn oil, soy oil, cottonseed oil and safflower oil are particularly inflammatory. Eat plenty of veggies and fruits and enjoy the turkey and cranberries. Make sure that the only sweeteners used are sugar and honey (avoid high fructose corn syrup.) Light corn syrup is the stealth form of HFCS -- it may be lower in calories, since fructose is sweeter than sugar, but it is highly inflammatory! (Research also indicates that fructose causes premature wrinkling and skin aging, by enhancing the crosslinking of collagen. HFCS also causes type II diabetes in lab animals.)