Matthew Pratt-Hyatt PhD

New Markers for the MycoTOX Profile

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Welcome back to the GPL blog.  I have another exciting announcement and that is that we are adding four additional markers to our MycoTOX Profile, which screens for exposure to mycotoxins from mold.  Yet again, our laboratory scientists have shown why we are an industry leader in toxin exposure assessment.  These four new markers will now give us 11 markers on our revolutionary MycoTOX Profile.  These additional markers are also being added at no additional cost.

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Just by ordering the MycoTOX Profile you will get these four new markers in addition to the previous seven markers.  This test can be easily be added to the GPL-TOX (Toxic Non-Metal Chemical Profile) and the Organic Acids Test (OAT), all with just one first morning urine sample. 

Here are the new markers that we will be starting to report today. These four new markers will further help practitioners determine the underlying causes of their patients’ chronic health issues:

Gliotoxin
Gliotoxin (GTX) is produced by the mold genus AspergillusAspergillus spreads in the environment by releasing conidia which are capable of infiltrating the small alveolar airways of individuals.  In order to evade the body’s defenses Aspergillus releases Gliotoxin to inhibit the immune system.  One of the targets of Gliotoxin is PtdIns (3,4,5) P3.  This results in the downregulation of phagocytic immune defense, which can lead to the exacerbation of polymicrobial infections.  Gliotoxin impairs the activation of T-cells and induces apoptosis in monocytes and in monocyte-derived dendritic cells.  These impairments can lead to multiple neurological syndromes.

Mycophenolic Acid
Mycophenolic Acid (MPA) is produced by the Penicillium fungus.  MPA is an immunosuppressant which inhibits the proliferation of B and T lymphocytes.  MPA exposure can increase the risk of opportunistic infections such as Clostridia and Candida. MPA is associated with miscarriage and congenital malformations when the woman is exposed in pregnancy. 

Dihydrocitrinone
Dihydrocitrinone is a metabolite of Citrinin (CTN), which is a mycotoxin that is produced by the mold species Aspergillus, Penicillium, and Monascus.  CTN exposure can lead to nephropathy, because of its ability to increase permeability of mitochondrial membranes in the kidneys.  The three most common exposure routes are through ingestion, inhalation, and skin contact.  CTN has been shown to be carcinogenic in rat studies.  Multiple studies have linked CTN exposure to a suppression of the immune response. 

Chaetoglobosin A
Chaetoglobosin A (CHA) is produced by the mold Chaetomium globosum (CG).   CG is commonly found in homes that have experienced water damage.   Up to 49% of water-damaged buildings have been found to have CG.  CHA is highly toxic, even at minimal doses.  CHA disrupts cellular division and movement.  Most exposure to CG is through the mycotoxins because the spores tend not to aerosolize.  Exposure to CHA has been linked to neuronal damage, peritonitis, and cutaneous lesions.

Species of Mold

These new markers are adding to our already revolutionary test. We will now be able to detect over 40 different strains of disease-causing mold.Here is a table that illustrates all of the different mold species that we can now detect:

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We are also instituting some new changes to our MycoTOX Profile test report.  We are moving all of the interpretations to the end of the report so that all of the results will fit on the first two pages.  In addition, we are changing the reportable range.  Since we launched this test we have analyzed thousands of samples.  By analyzing those results and comparing them to results from our Organic Acids Test, we now have a better understanding of what could be considered “normal values” for mycotoxins.  On our new report (seen below) you will see two numbers on the bar graph for each marker.  The number on the left is what we consider the maximum safe amount of mycotoxins a patient can have before symptoms may start to appear.  The number on the right is our 75% for our patients.  If your value is above this number then you have more mycotoxins than 75% of patients that have sent in samples.  These should be considered extremely elevated amounts and treatment is highly recommended. 

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Practitioner Training – GPL Academy Practitioner Workshops

I recommend to all practitioners that they attend our training workshops to help better understand how to evaluate the results from our tests, as well as to learn what treatments have been most effective.  Our GPL Academy workshops are great learning experiences.  At these events you can talk to our laboratory experts as well as discuss treatment plans with practitioners that we invite that are experts in their fields.  Please follow this link to find a workshop near you.


PMID
16712786, 27048806, 21575912, 23278106, 858824, 28646113, 27809954, 27599910, 11567776, 24048364, 10788357, 21196335, 12781669, 17551849, 28430618, 25264878, 21954354, 27401186, 28007639, 28718805, 21872054

ELISA Versus LC/MS for Mycotoxin Testing

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Welcome back to the GPL Blog.  Since we released our new GPL-MycoTOX Profile, we’ve received many questions about what the differences are between the types of mycotoxin testing available, why we use the technology that we do (LC/MS), and why we believe that technology is superior.  I wanted to share our feedback about that with you, including support data in the form of some split sample reports. 

The Difference Between ELISA and LC/MS

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ELISA testing is notorious for false positive readings.  ELISA principal is a lock and key activation.  If any molecule fits the lock then the result shows up positive.  One of the criticisms often heard about mold testing is the over-abundance of positive results. The literature backs up this observation with findings that show inferences that cause false positive results.  At Great Plains, we use LC/MS, not ELISA.  LC/MS separates out molecules by their chemical properties and measures their mass, so we get a definitive answer for every sample.  We also use internal standards in every sample to give a definitive quantitative reading. 

Why Creatinine Correction is Important

There are many factors that could influence the value for any urine test, including how recent the exposure was, how much the patient is detoxifying, and how much liquid the patient drank the night before giving the sample.  We are able to correct for the third of these reasons by measuring the amount of creatinine in the sample, which compensates for how diluted the sample may be.  A particular sample one of our practitioner clients asked us about was more concentrated than most (creatinine was 166 mg/dL).  If the value was 80 mg/dL, then the value would have been doubled.  This allows us to mitigate one factor that can cause mycotoxin test values to fluctuate.

We have received a couple dozen results from patients that run a test for mycotoxins from another lab, then have run our test.  We have seen the gamut of results such as their previous test coming back negative and ours is positive (see examples here  – Patient 1 with GPL and Patient 1 with RTL), both tests were positive (see example here -- Patient 2 with GPL and Patient 2 with RTL), and values where the patient was negative on both. 

The Clinical Significance of our Mycotoxin Test and Organic Acids Test

In our experience, no patients are “normal” when it comes to toxins, including mycotoxins.  We see mycotoxin in almost every patient, but we have set our reportable limits to only patients that we feel have abnormal amounts of mycotoxin in order to not alarm patients.  We have followed this up with a study of 50 patients with mycotoxins.  If you look at this file, we did a comparison of patients with mycotoxins to patients without mycotoxins.  We see numerous values elevated on our Organic Acids Test (OAT) in the mycotoxin positive individuals, demonstrating that our test can predict health problems for individuals.  We will soon have more information available about the connection with specific fungal markers on the Organic Acids Test 

Please let me know if you have any questions about our GPL-MycoTOX Profile and we look forward to our continued work with you. 

Matt Pratt-Hyatt, PhD
Associate Laboratory Director

The Connection Between Chronic Inflammation and Disease

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Welcome back to the GPL blog.  Today I wanted to talk about the link between inflammation and chronic disease.  The mission of The Great Plains Laboratory is to help those suffering from chronic diseases, and recent studies have shown that most chronic diseases are tied to inflammation.  In this blog post I am going to give a brief synopsis of some of the most common diseases associated with chronic inflammation and what tests we offer that give insight into how to treat these patients.    

Inflammation is a response from the body to assist in the elimination of pathogens and to repair tissue damage from trauma.  Inflammation is a healthy, natural response to cellular stress caused by stimuli perceived as a threat.  It signals the body to bring extra nutrition to sites that are damaged through injury or illness.  Without inflammation proper healing could not occur. While acute inflammation is critical to our well-being, chronic, long term inflammation is damaging to cells and linked to many diseases. Chronic inflammation occurs when the immune system believes there is a threat even when there is no immediate reason for this perceived threat. It is still unclear what causes chronic inflammation but lifestyle factors, genetic factors, and internal stressors have all been implicated.

Recent studies have demonstrated that inflammation is an underlying contributor to most chronic diseases.  Some of the most common of these include cancer, rheumatoid arthritis, Crohn’s disease, depression, stroke, heart disease, and diabetes.   Since inflammation is involved with so many chronic diseases, detecting inflammation is an important aspect to managing patient symptoms. Better still, if the underlying causes of inflammation such as Candida, bacteria, mold, food sensitivities, and environmental toxins are determined, the disease process may be reversed.

Phospholipase A2

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We developed the Phospholipase A2 (PLA2) Activity Test to determine if a specific type of inflammation is underlying the patient’s condition.  PLA2 is an enzyme that activates during bacterial infection, cellular trauma, and periods of oxidative stress.  PLA2 activates a cascade of secondary messengers that can lead to cycles of inflammation that can self-perpetuate.  PLA2 metabolizes membrane glycerophospholipids to free arachidonic acid (AA), which is a precursor for the inflammatory signaling molecules, prostaglandins and leukotrienes (Figure 1).  PLA2 is expressed in neuronal tissue and is involved in the degranulation process that releases neurotransmitters from neurons. Research efforts have focused on the role that derangement of normal PLA2 activity plays in the etiology of many chronic illnesses. The specific roles, interactions, and interdependencies of PLA2 have been a major area of interest as it relates to chronic inflammatory conditions, cardiovascular disease, and cancer.  

Measurement of PLA2 is emerging as an important tool for evaluating the chance of cardiovascular disease (CVD), including future stroke, myocardial infarction, heart failure, and other vascular events. Lp-PLA2 appears to be more specific than hsCRP for CVD risk and may also have a pivotal role as a mediator of cardiovascular pathology. In atherosclerosis, PLA2 not only activates macrophages and formation of foam cells, but it also hydrolyzes LDL and HDL, spawning increased numbers of pro-atherogenic small LDL particles, and impairing anti-atherogenic HDL. PLA2 activity may even precipitate bleeding from atherosclerotic plaques.

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Because PLA2 is a relatively small enzyme (about 14 KD), it is able to be excreted in urine.  Our enzymatic assay determines how active PLA2 is in the body, which is mediated by phosphorylation of the enzyme.  We have assessed that an activity level of PLA2 activity/creatinine of over 1 results from elevated activity and could be harmful. 

CDP-choline

The literature indicates that Cytidine 5-diphospho-choline (CDP-choline or citicoline) attenuates PLA2 through a number of mechanisms. Most notably it repairs membrane potential and reduces lipid peroxidation. These processes essentially prevent new PLA2 from forming by stopping the cycle of inflammation.   Individuals with methylation pathway SNPs may be more susceptible to deficiencies in CDP-choline because it is difficult for them to make phosphoethanolamine, a CDP-choline precursor. Some individuals may have further deficiencies in citicholine due to mutations in their PEMT gene which converts phosphoethanolamine into CDP-choline. Fortunately, this compound is available as a nutritional supplement from New Beginnings Nutritionals and has been used at doses ranging from 500-4000 mg per day in the treatment of patients with a variety of disorders including Parkinson's disease, memory disorders, vascular cognitive impairment, vascular dementia, senile dementia, schizophrenia, Alzheimer's disease (especially effective in those with the epsilon-4 apolipoprotein E genotype), head trauma, and ischemic stroke. A trial in patients with Alzheimer's disease indicated that CDP-choline (1,000 mg/day) is well tolerated and improves cognitive performance, cerebral blood perfusion, and the brain bioelectrical activity pattern. No side effects were noticed except for some mild gastrointestinal symptoms at higher doses. No abnormal blood chemistry or hematology values were found after the use of CDP-choline. Many patients and practitioners are unfamiliar with CDP-choline and may be tempted to use the more commonly prescribed phosphatidyl-choline. These two products cannot be used interchangeably. Phosphatidyl-choline is a glycerophospholipid that PLA2 can use to elicit its inflammatory effects. Individuals with elevations in PLA2 should refrain from supplements containing phosphatidyl-choline and use CDP-choline instead. 

The Organic Acids Test

The Organic Acids Test (OAT) is a comprehensive metabolic assessment of multiple systems in the body.  It is one of our best tools in determining the underlying causes of many chronic diseases.  The OAT test can be useful in the identification of intestinal yeast and bacteria, oxalates, abnormal neurotransmitters, mitochondrial markers, fatty acid oxidation, nutritional deficiencies, detoxification markers, and inborn errors of amino acid metabolism.   

Many of the markers on the OAT can help in the diagnosis of inflammation.  Some of these include markers for Candida and clostridia.  An overgrowth of these pathogenic microbes can lead to disruptions in the gut lining, which can cause inflammation and disrupt the absorption of nutrients.  Candida and clostridia can also produce many different chemical toxins that are absorbed through the intestines.  These toxins can produce confusion (brain fog), thyroid dysregulation, weight gain, acne, drowsiness, irritable bowel syndrome, and insomnia.  There are multiple markers for both Candida and Clostridia.  Some of the most common yeast markers are tartaric, arabinose, and carboxycitric.       

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The OAT also has more metabolic markers for clostridia than any other organic acid test on the market.  The OAT is able to identify overgrowth from multiple different strains of clostridia.  This is accomplished by looking at four different markers which include 4-hydroxyphenylacetic, HPHPA, 4-Cresol, and 3-indoleacetic.   These toxins produced by bacteria can lead to inflammation as well as inhibition of neurotransmitter metabolizing enzymes.

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One of the best markers for inflammation is the succinic acid marker.  Succinic acid is generated in mitochondria during the tricarboxylic acid cycle (TCA).  Succinic acid is also a signaling molecule which can change gene expression patterns by modulating epigenetic markers in the DNA.  Our data indicates that exposure to environmental toxins can cause inflammation, resulting in accumulation of succinic acid.

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 This may be caused by the inhibition of succinate dehydrogenase, an important enzyme that functions in both the Krebs cycle and complex II of the electron transport chain. In the Krebs cycle, it converts succinate to fumarate. In the electron transport chain, it works with CoQ10 to transfer electrons into the complex III phase of the chain.  Our laboratory compared patients with high values several common environmental toxins on the GPL-TOX test and determined what their average succinic acid values were.  We found that patients with high vinyl chloride, xylene, heavy metals such as lead and mercury, DMP, DEP, and 2,4 D correlated with patients having elevated succinate values (see graph).

One additional marker on the Organic Acids Test for inflammation is quinolinic acid.  Quinolinic acid is a neuroactive metabolite of the kynurenic pathway.  Quinolinic acid is produced from tryptophan through a multi-stage process.  Buildup of quinolinic acid increases stimulation to NMDA glutamate receptors and inhibits the reuptake of glutamate by astrocytes leading to neurotoxicity.  Studies have shown that chronic exposure to quinolinic acid can lead to structural changes such as dendritic beading, microtubular disruption, and a decrease in organelles in neurons.  Quinolinic acid can also increase oxidative stress by inducing NOS activity.  Quinolinic acid further adds to inflammation by causing an increase of expression in the inflammatory response elements TNF-α and interleukin-6. 

Tests for Environmental Toxins

One of the leading causes of inflammation is environmental toxicants, which has been increasing every year since the 1960s.   This increase in toxic burden observed in our patients is one reason why we have put a focus on providing testing for many different sources of environmental toxicants.  We currently offer the GPL-TOX (Toxic Non-Metal Chemical Profile), Glyphosate Test, and metals tests (hair, urine, blood), and we just launched the GPL-MycoTOX Profile, the most sensitive test for mold toxins in the world.  If you have any questions about any of these tests, you may review previous blogs about them here or visit the individual tests information pages on our web site.  

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The IgG Food Allergy Test

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Testing for food sensitivity has been extremely helpful to many of our patient populations.  The symptoms of food sensitivity can include (but aren’t limited to) austim, ADD, depression, arthritis, fatigue, skin rashes, and gastrointestinal issues.  Our IgG Food Allergy Test is an invaluable tool to determine what role food plays in inflammation in the body. Immunoglobulin G (IgG) is the major antibody found in serum, and our test measures IgG subclasses 1-4.  IgG has a much longer half-life than the transitional IgE antibody.  Whereas IgE can cause acute reactions to food, IgG can cause inflammation that can lead to more chronic health issues.  These IgG reactions can be more subtle and people can live with them for years without realizing what is causing their discomfort.  The degree of severity can differ because of genetics or from exposure to different environmental toxins, which can predispose people to immune responses. 

At The Great Plains Laboratory we offer two different ways to submit samples for IgG food testing – serum and dried blood spot (DBS).  We have validated both of these tests against each other and they provide the same result.  Our test currently looks at 93 different foods along with testing for Candida antibodies. 

Conclusion

At The Great Plains Laboratory, we are very focused on helping patients solve their chronic health problems.  Research shows that inflammation plays a role in most chronic issues.  This is the reason why we have developed so many tests to help pinpoint the root causes of inflammation and to help figure out the best method of treatment.  We hope that we can help as many people in the future live better and longer lives.

 

 

A Brand New Urine Test for Mycotoxin Exposure

Welcome back to the GPL blog.  I am very excited to announce our newest test, the GPL-MycoTOX Profile. Mycotoxins are some of the most prevalent toxins in the environment.  Mycotoxins are metabolites produced by fungi like mold, which can infest buildings, vehicles, and foodstuffs.   A majority of mycotoxin exposures are through food ingestion or airborne exposure.   In the EU, 20% of all grains harvested have been found to be contaminated with mycotoxins.

Fungi are able to grow on almost any surface, especially if the environment is warm and wet.  Inner wall materials of buildings, wall paper, fiber glass insulation, ceiling tiles, and gypsum support are all good surfaces for fungi to colonize.  These fungi then release mycotoxins into the environment causing symptoms of many different chronic diseases.  Diseases and symptoms linked to mycotoxin exposure include fever, pneumonia-like symptoms, heart disease, rheumatic disease, asthma, sinusitis, cancer, memory loss, vision loss, chronic fatigue, skin rashes, depression, ADHD, anxiety, and liver damage.  With our new GPL-MycoTox Profile we can identify mycotoxin exposures and make recommendations for detoxification treatments that have been effective. 

Our two primary goals for this test were to design a test that would be more sensitive and accurate than those currently on market as well as more affordable.   We were able to achieve both of these goals with our state of the art liquid chromatography mass spectrometry (LC-MS/MS) technology.  Using this technology, we have a very sensitive test, which is important because mycotoxins can cause serious health issues even in small quantities.  Other mycotoxin testing uses ELISA technology, which relies on antibodies.  Utilization of LC-MS/MS technology gives us a precise identification of all of our analytes, which prevents having false positive errors.  For many of our compounds we are able to detect amounts in the parts per trillion (ppt) which is about 100 fold better than any other test currently available. 

Species of Mold

We are currently measuring eight different mycotoxins in our test, from four types of mold (we are always doing R&D on our tests in the hopes of adding even more markers in the future).  This makes the GPL-MycoTOX Profile the most comprehensive test on the market and it’s the most cost-effective.  Here are the four types of mold we are evaluating:

Aspergillus:  Aspergillus is the most prevalent mold group in the environment.  It has caused billions of dollars in damage to crops and livestock.  The two most common Aspergillus mycotoxins are aflatoxin, ochratoxin, patulin, and fumigillin.  The main target of these toxins is the liver.   These toxins have been found in all major cereal crops including peanuts, corn, cotton, millet, rice, sorghum, sunflower seeds, wheat, and a variety of spices.  They are also found in eggs, milk, and meat from animals fed contaminated grains.  Diseases caused by Aspergillus are called aspergillosis.  The most common route of infection is through the respiratory system.  Aspergillus can cause severe asthma when the mold colonizes the lung, forming a granulomatous disease.

PenicilliumThere are over 200 species of the genus Penicillium that have been discovered.  Penicillium chrysogenum is the most common of these species.  It is often found in indoor environments and is responsible for many allergic reactions.  Penicillium is also a known contaminant in many different food items.  Many different types of citrus fruits can become contaminated with Penicillium, but it can also contaminate seeds and grains.  One reason that Penicillium is such a common infestation is because of its ability to thrive in low humidity.  In the home, Penicillium can be found in wallpaper, carpet, furniture, and fiberglass insulation.  The most common mycotoxin produced by Penicillium is ochratoxin.  Ochratoxin is nephrotoxic, which means that it damages the kidneys.  It is also carcinogenic.      

Stachybotrys:  Stachybotrys is a greenish-black mold.  This mold can grow on materials with high cellulose and low nitrogen content such as gypsum board, paper, fiberboard, and ceiling tiles.  Stachybotrys is known for its production of the highly toxic macrocyclic trichothecene mycotoxins.    Two of the more common mycotoxins produced by Stachybotrys are roridin E and verrucarin.   In addition to these mycotoxins, the fungus produces nine phenylspirodrimanes, as well as cyclosporine, which are potent immunosuppressors. These immunosuppressors, along with the mycotoxin trichothecenes may be responsible for the high toxicity of Stachybotrys

Fusarium:  Fusarium’s major mycotoxins are zearalenone (ZEN) and fumonisin.  Fusarium fungi grow best in temperate climate conditions.  They require lower temperatures for growth than Aspergillus. Fusarium grows worldwide on many different types of grains including corn and wheat.  Exposure to mycotoxins from Fusarium can lead to both acute and chronic effects.  These symptoms can include abdominal distress, malaise, diarrhea, emesis, and death.  ZEN possesses estrogenic effects and has been implicated in reproductive disorders. 

 

Markers in the GPL-MycoTOX Profile

The strains of mold we’re evaluating can produce several different mycotoxins.  We have developed a test that provides extensive coverage, allowing us to catch most mold exposures. 

Aflatoxin M1 (AFM1) is the main metabolite of aflatoxin B1, which is a mycotoxin produced by different species of the genus Aspergillus. Aflatoxins are some of the most carcinogenic substances in the environment.  Aflatoxin susceptibility is dependent on multiple different factors such as age, sex, and diet.  Aflatoxin can be found in beans, corn, rice, tree nuts, wheat, milk, eggs, and meat.   In cases of lung aspergilloma, aflatoxin has been found in human tissue specimens. Aflatoxin can cause liver damage, cancer, mental impairment, abdominal pain, hemorrhaging, coma, and death.  Aflatoxin has been shown to inhibit leucocyte proliferation. Clinical signs of aflatoxicosis are non-pruritic macular rash, headache, gastrointestinal dysfunction (often extreme), lower extremity edema, anemia, and jaundice. The toxicity of Aflatoxin is increased in the presence of Ochratoxin and Zearalenone.

Ochratoxin A (OTA) is a nephrotoxic, immunotoxic, and carcinogenic mycotoxin.  This chemical is produced by molds in the Aspergillus and Penicillium families.  Exposure is primarily through contaminated foods such as cereals, grape juices, dairy, spices, wine, dried vine fruit, and coffee.  Exposure to OTA can also come from inhalation exposure in water-damaged buildings.  OTA can lead to kidney disease and adverse neurological effects.  Studies have shown that OTA can cause significant oxidative damage to multiple brain regions and the kidneys.  Dopamine levels in the brain of mice have been shown to be decreased after exposure to OTA. 

Sterigmatocystin (STG) is a mycotoxin that is closely related to aflatoxin.  STG is produced from several types of mold such as Aspergillus, Penicillium, and Bipolaris.  It is considered to be carcinogenic, particularly in the cells of the GI tract and liver. STG has been found in the dust from damp carpets.  It is also a contaminant of many foods including grains, corn, bread, cheese, spices, coffee beans, soybeans, pistachio nuts, and animal feed. In cases of lung aspergilloma, STG has been found in human tissue specimens. The toxicity of STG affects the liver, kidneys, and immune system.  Tumors have been found in the lungs of rodents that were exposed to STG.  Oxidative stress becomes measurably elevated during STG exposure, which causes a depletion of antioxidants such as glutathione, particularly in the liver. 

Zearalenone (ZEA) is a mycotoxin that is produced by the mold species Fusarium, and has been shown to be hepatotoxic, haematotoxic, immunotoxic, and genotoxic.  ZEA is commonly found in several foods in the US, Europe, Asia, and Africa including wheat, barley, rice, and maize.  ZEA has estrogenic activity and exposure to ZEA can lead to reproductive changes.  ZEA’s estrogenic activity is higher than that of other non-steroidal isoflavones (compounds that have estrogen-like effects) such as soy and clover.  ZEA exposure can result in thymus atrophy and alter spleen lymphocyte production as well as impaired lymphocyte immune response, which leads to patients being susceptible to disease.

Roridin E is a macrocyclic trichothecene produced by the molds Fusarium, Myrothecium, and Stachybotrys (i.e. black mold).  Trichothecenes are frequently found in buildings with water damage but can also be found in contaminated grain.  This is a very toxic compound, which inhibits protein biosynthesis by preventing peptidyl transferase activity. Trichothecenes are considered extremely toxic and have been used as biological warfare agents. Even low levels of exposure to macrocyclic trichothecenes can cause severe neurological damage, immunosuppression, endocrine disruption, cardiovascular problems, and gastrointestinal distress.

Verrucarin A is a macrocyclic trichothecene mycotoxin produced from Stachybotrys, Fusarium, and Myrothecium.  Trichothecenes are frequently found in buildings with water damage but can also be found in contaminated grain.  This is a very toxic compound, which inhibits protein biosynthesis by preventing peptidyl transferase activity.  Trichothecenes are considered extremely toxic and have been used as biological warfare agents.  Even low levels of exposure to macrocyclic trichothecenes can cause severe neurological damage, immunosuppression, endocrine disruption, cardiovascular problems, and gastrointestinal distress.

Enniatin B1 is a fungal metabolite categorized as a cyclohexa-depsipeptides toxin produced by thefungus Fusarium. This strain of fungus is one of the most common cereal contaminants.  Grains in many different countries have recently been contaminated with high levels of Enniatin.  The toxic effects of Enniatin are caused by the inhibition of the acyl-CoA cholesterol acyltransferase, depolarization of mitochondria, and inhibition of osteoclastic bone resorption.  Enniatin has antibiotic properties and chronic exposure may lead to weight loss, fatigue, and liver disease.

Treatment for Mycotoxin Exposure

Treatment for mold exposure to should include fluid support to prevent dehydration.  The drug Oltipraz can increase glutathione conjugation of mold toxins while inhibiting the toxic effect of P450 oxidation, reducing liver toxicity and promoting safer elimination (PMID: 18286403, 10050868, 7585637).  A diet of carrots, parsnips, celery, and parsley may reduce the carcinogenic effects of mold (PMID 16762476). Bentonite clay and zeolite clay are reported to reduce the absorption of mold found in food (PMID: 16019795, 18286403). Supplementation with chlorophyllin, zinc, A, E, C, NAC, rosmarinic acid, and liposomal glutathione alone or in combination have been shown to mitigate the oxidative effects of mold toxins (PMID:22069658).

Details of the GPL-MycoTOX Profile

The GPL-MycoTOX Profile is a urine test.  The specimen requirement is the first morning urine and 10 ML of urine.  Since this is a urine test a patient can combine this test with many of our other urine tests such as the Organic Acids Test, GPL-TOX (Toxic Non-Metal Chemical Profile), Glyphosate Test, and the PLA2 Test.  We are very excited to have what we believe to be the best and most cost-effective test for mycotoxins available, which will be very helpful in the treatment of many chronic diseases.  To learn more about this test please visit the GPL-MycoTOX Profile test page on our web site, watch the recently recorded webinar about the test, or attend one of our upcoming GPL University Practitioner Workshops.  

New Marker Additions to GPL-SNP1000 DNA Sequencing Profile

BY: MATTHEW PRATT-HYATT, PHD

The number one goal for The Great Plains Laboratory is to provide the best quality results to our clients.  Our GPL-SNP1000 DNA Sequencing Profile has proven to be a great tool in helping provide personalized healthcare to our clients.  The nine pathways we analyze include: methylation, mental health, oxalate metabolism, drug and environmental metabolism/detoxification, gluten sensitivity, cholesterol metabolism, autism risk genes, and transporter gene.  These are crucial biological pathways, which are at the root of many chronic health conditions.  We are now announcing the addition of nine new markers to our already incredibly comprehensive genetic test:

Dopamine Beta Hydroxylase (DBH)
This is an enzyme that catalyzes the oxidation hydroxylation of dopamine to norepinephrine.  DBH can be inhibited by phenolic compounds including those produced by Clostridium species as well as certain organophosphate herbicides and pesticides.  There are two SNPs that can cause decreased activity of DBH.  These are rs2007153 and rs2283123.  These polymorphisms can lead to an increase in dopamine levels and a deficiency in norepinephrine.  Mental health disorders can result because of the imbalance of dopamine and norepinephrine.  Common symptoms can include depression and anxiety.

Paroxonase 1 (PON1)
This is an important enzyme in the metabolism and elimination of many organophosphorus insecticides (PMID: 13032041) and is located mainly in the liver.  PON1 is important in the reduction of atherosclerosis because of its involvement in the protection of high and low density lipoproteins from oxidation.  Individuals with polymorphisms to PON1 are more susceptible to heart disease (PMID: 8675673).  There are two known polymorphisms that can decrease the activity of PON1 and make the individual more susceptible to pesticide exposure, which are Q192R (rs662) and L55M (rs854560).

Hemochromatosis Protein (HFE)
The hemochromatosis gene HFE (high iron) codes for the HFE protein.   This protein is important for regulating the uptake of circulating iron.  This is done by regulating the interaction between transferrin receptor with transferrin.  SNPs to this gene can cause hemochromatosis, a disorder in which the body loads excess iron, which is autosomal recessive.  This means the patient normally needs two bad copies of the gene in order to exhibit symptoms.  There are three SNPs that can lead to hemochromatosis, rs1800562, rs1800730, and rs1799945.  Patients that are homozygous positive for this SNP should have their iron level measured. 

Vitamin K Epoxide Reductase Complex Subunit 1(VKORC1)
This is an enzyme that is necessary for the reduction of vitamin K 2,3-epoxide to its active form, which is important for clotting.  This enzyme is the primary target for the drug warfarin (Coumadin).  The three SNPs that are associated with warfarin sensitivity are rs9923231 (VKORC1*2), rs9934438, and rs8050894.  These polymorphisms can be used in conjuncture with the genotype of Cyp2C9 in order to accurately dose warfarin.

Tryptophan Hydroxylase 2 (TPH2)
This enzyme catalyzes the first and rate-limiting step in the biosynthesis of serotonin. Mutations to this enzyme have been associated with numerous psychiatric diseases including depression, OCD, bipolar disorder, and suicidal behavior.

Major Histocompatibility Complex DQA1 and DQA8
Patients with SNPs to HLA DQA1 and DQA8 have a higher risk of celiac disease.   The HLA-DQA1 and DQA8 are human leukocyte antigen serotype (also called major histocompatibility complex II). The role of this peptide is to present proteins on the surface of cells for identification purposes. This particular serotype presents proteins belonging to a foreign invader on the cells the macrophages, B cells, and dendritic cells in order to activate the helper T cells of the immune system. Proper presentation is critical for immune system activation against pathogens and may possibly be a mediator of autoimmunity.

UDP Glucosyltransferase 1A1 and 1A8 (UGT1A1 and UGT1A8)
These enzymes are important members of the glucuronidation phase II detoxification pathway.  These enzymes catalyze the addition of a glycosyl group from a nucleotide sugar to a small hydrophobic molecule.  The addition of glycosyl groups results in these molecules becoming more water-soluble and easier to excrete. Some of the target molecules for these enzymes include bilirubin, drugs, hormones, and steroids.

Your Body’s Detoxification Pathways

Welcome back to the GPL-BLOG.  Over the past several weeks we’ve been discussing a lot of the environmental toxins that everyone is exposed to on a daily basis.  These toxins must be processed and detoxified.  Most of this is done in the liver through several different processes that include Cytochrome P450 (P450) biotransformation, glutathione conjugation, enzyme hydrolyzing, sulfation, and glucuronidation.

Detoxification is often referred to as a two stage process (phase 1 and phase 2) of metabolism (Figure 1).  Phase 1 metabolism involves the reduction or hydrolysis of the compound (usually caused by the addition of an oxygen molecule).  The addition of oxygen to a compound is referred to as oxidation.  This process is usually performed by the P450 enzymes.

Figure 1

The P450s are a family of enzymes that are found in numerous tissues throughout the body. However, a majority of these are found in the liver.  The P450s are important for the detoxification of many foreign substances, including environmental toxicants and medications.  The P450s are also important in controlling the levels of different molecules produced in the body such as the synthesis and breakdown of hormones, steroids, and multiple other molecules. 

In humans, 58 different P450s have been discovered.  However, only a subset of these is involved in the degradation of xenobiotics (chemicals that come from outside the body).  These enzymes have different substrates, which are determined by the activity pocket of each enzyme.   In regards to detoxification the most important P450s are Cyp1A2, Cyp2A6, Cyp2C9, Cyp2C19, Cyp2D6, Cyp2E1, and Cyp3A4.  Besides detoxification, these enzymes metabolize a majority of medications (figure 2).

Here are some important detoxification enzymes:

Figure 2

Cyp1A2 is important for the metabolism of polycyclic aromatic hydrocarbons (PAHs), which are found in cigarette smoke.  Other substrates include medications, aflatoxin B1, caffeine, and acetaminophen.  The major polymorphism is Cyp1A2*1K, which results in a decrease of activity.

Cyp2A6 is involved in the metabolism of nicotine.  Cyp2A6 is also involved in the metabolism of medications.  The major polymorphic alleles are Cyp2A6*4 and Cyp2A6*9 (which can have between 35% -70% activity depending on if you have one or two polymorphic copies). 

Cyp2C9 is involved with the metabolism of a large number of medications including NSAIDs, warfarin, and tamoxifen.  There are multiple polymorphisms that affect activity of the enzyme. 

Cyp2C19 is involved with the metabolism of multiple medications.  The most common are diazepam, omeprazole, and sertraline.  Cyp2c19 also metabolizes progesterone.   There are two major variants that result in loss of activity.  These are Cyp2C19*2 and Cyp2C19*3.

Cyp2D6 is involved with the metabolism of about 20% of drugs on the market.  It also metabolizes serotonin and neurosteroids.  There are five different polymorphisms that can lead to decreased activity.  Some of the classes of drugs that are metabolized by Cyp2D6 are antidepressants, SSRIs, opioids, and antipsychotics. 

Cyp2E1 is involved with the detoxification of many industrial pollutants, as well as carcinogens.  Cyp2e1 also metabolizes ethanol to acetaldehyde and acetate.  Cyp2e1 is also responsible for bioactivating a number of carcinogens, including cigarette smoke. 

Cyp3A4 is responsible for metabolizing more compounds than most other P450s.  It is responsible for metabolizing sex hormones, caffeine, statins, SSRIs, antifungals, antidepressants, and many other medications.  Some antibiotics can negatively affect its activity. Also, grapefruit and pomegranate juice have been shown to be potent inhibitors. 

Sulfur transferase is a phase 2 enzyme that adds sulfur groups to compounds in order to make them more water soluble and less reactive.  This process is used on a wide variety of toxic molecules including phenols, amines, acetaminophen, and food dyes.  Many chemicals that are able to become airborne are sulfated.  Patients with autism have been found to have impaired sulfation ability, which will make these individuals more sensitive to toxins.

Glutathione transferase is a phase 2 enzyme that catalyzes the conjugation of glutathione to substrates.  The addition of glutathione to toxins prevents these compounds from interacting with proteins in the body and allows them to be excreted via urine or bile.  There are a wide variety of compounds that are conjugated with glutathione.  A partial list includes pesticides, herbicides, carcinogens, acetaminophen, and mycotoxins.   

Glucuronosyltransferase (UGT) is another phase 2 enzyme that is responsible for the glucuronidation of many different toxic chemicals.  This process involves the addition of a glucuronosyl group to substrate molecules making them more polar and more easily excreted by the kidneys. 

Paraoxonase 1 (PON1) is an enzyme that is able to perform paraoxonase activity on substrates.  This enzyme is able to hydroylse and detoxify many different types of organophophate molecules.  PON1 is one of the major pathways that protects people from these types of compounds.  Mutations to PON1 could lead someone to be more sensitive to pesticides.  Infants do not have a lot of PON1 activity.  PON1 becomes active between birth and seven years of age. 

These are the major pathways that you should be aware of when you are thinking about detoxification.  Please see Table 1 to help you understand which pathway is mostly responsible for detoxifying these common toxicants.  Also see Figure 1 to help you understand what you can do to help support type 1 and type 2 detoxification pathways.  Detoxification of compounds by glutathione can be assisted by the supplementation of additional glutathione.  Next week I will discuss some additional methods to help with detoxification.  

Email gplblog@gpl4u.com if you have any questions about this blog post.

DNA Methylation Pathway

In my post last week I briefly talked about the methylation pathway, also called the MTHFR cycle and how disruptions in this pathway may appear on an Organic Acid Test (OAT).  Today, I will go more in-depth into this pathway. Since most practitioners have at least some knowledge of this its function, I’ve decided to focus on the more important polymorphisms (SNPs) common to these genes and on the treatments that work the best for these mutations.  Patients should talk to their healthcare practitioner before starting any treatment. 

The important role of methylation is gaining in popularity among functional medicine groups these days because mutations are quite common and lead to many different chronic conditions. Practitioners interested in treating the root cause of illness are especially interested in learning about this pathway because nucleotide synthesis, neurotransmitter function, detoxification, and numerous other processes are greatly improved once these mutations have been compensated for, leading to much better patient outcomes.

If you’re not already familiar, the methylation pathway is a process by which carbons are added onto folic acid from amino acids and redistributed onto other compounds throughout the body.  This process is responsible for the formation of methionine, S-Adenosyl methionine (SAM), and thymidylate monophosphate (dTMP).  Mutations in this pathway usually lead to the reduction of methionine which leads to the absence of S-adenosyl methionine (SAM). This compound facilitates virtually every methylation reaction in the body. These reactions include the promotion of several neurologically important agents, histamine breakdown, CoQ10 synthesis, and tissue-specific gene expression. The accumulation of homocysteine, which is caused by mutations in this pathway, has been directly linked to oxidative stress which influences multiple factors of disease.      

When we designed the GPL-SNP1000 test, we knew that most other genetic tests were only reporting about 35 SNPs of the common methylation pathway enzymes. When we did our literature research, we found 105 different methylation SNPs that could potentially cause health conditions and included all of these to provide a more useful tool for practitioners.

MTHFR

Methylenetetrahydrofolate reductase (MTHFR) is an enzyme that converts 5,10-methylenetetrahydrofolate to 5,methyltetrahydrofolate, which is the active form of folate.  Mutations in this gene cause the accumulation of homocysteine and a lack of available folate for cellular functions. Both of these factors have been linked to oxidative stress, vascular disease (including cardiovascular), neural tube defects, neurological disorders (including schizophrenia and bipolar disorder), cancer, preeclampsia, hypotonia, and seizures.  Common mutations are rs1801133 (C677T), rs1801131 (A1298C), and rs2274976 (G1793A).  The C677T polymorphism is present in about 39% of caucasians as heterozygotes and 17% as homozygotes.  Table 1 provides data on how much activity your MTHFR enzyme would possess with different combinations of the C677T and the A1298C.

TABLE 1

TABLE 1

Patients with polymorphisms in MTHFR may consider supplementing with methyl-B12 (also called methylcobalamin) and methyl-folate.  We recommended starting at a very low doses and building up.

MTR

Methionine synthase (MTR) is also known as 5-methyltetrahydrofolate-homocysteine methyltransferase.  This enzyme facilitates the transfer of a methyl group from 5-methyltetrahydrofolate to homocysteine using cobalamin (B12) and MTRR enzyme as a catalyst. The end products of this reaction are the amino acid methionine and the vitamin tetrahydrofolate.  Mutations in this gene lead to a lack of methionine and the accumulation of homocysteine in the body (hyperhomocysteinemia).  The pathological consequence of the gene mutation depends on how profoundly these methylation pathways are affected and the degree of homocysteine accumulation in the body.  The most common polymorphism is rs1805087 (A2756G), which most genetic tests do analyze.  However, we decided to look at seven different SNPs including the rs121913581 (R52Q) polymorphism.  This is a rare polymorphism; however it could dramatically affect the activity of patients with MTR polymorphisms, who should also consider methyl-B12, as well as S-adenosyl methionine (SAM) supplementation.  ).

MTRR

Methionine synthase reductase (MTRR) is also known as 5-methyltetrahydrofolate-homocysteine methyltransferase reductase.   MTRR is important for the methylation of cobalamin and subsequent activation of methionine synthase (MTR).  Mutations in this gene lead to a lack of methionine and the accumulation of homocysteine in the body (hyperhomocysteinemia). Some common mutations in this pathway are RS1801394 and RS10380.  Patients who are heterozygous for MTHFR mutations and concomitant mutations to MTRR have a greater loss of function and increased levels of homocysteine. Patients with MTRR polymorphisms should consider methyl-B12 and SAMe supplementation. 

AHCY

Adenosylhomocysteinase (AHCY) is also known as S-adenosylhomocysteine hydrolase.  AHCY is an enzyme involved in the degradation of the amino acid methionine.  AHCY converts the methionine substrate S-adenosylhomocysteine (SAH) to adenosine and homocysteine. This reaction is an important part of the regulation of methyl groups which are added to DNA, RNA, proteins, and lipids (fats).  Methyl groups help regulate what parts of the genome are active and control protein activity.  Mutations to the AHCY gene can cause methionine to accumulate in the blood, which is called hypermethioninemia (MET).  Two common mutations are Trp112X which causes tryptophan to be replaced with a premature stop signal and Tyr143Cys.  MET  can manifest in neurological problems, delays in motor skills, muscle weakness, and liver problems.  Patients with MET should consult with a dietician to avoid the amino acid methionine.

BHMT

Betaine-homocysteine methyltransferase (BHMT) and BHMT2 are the only enzymes that can metabolize betaine.  This reaction is considered the alternate or short route for methylation.  BHMT uses zinc as a co-factor to catalyze the transfer of a methyl group from betaine to homocysteine.  There are several mutations in the human population that decrease the activity of this enzyme.  BHMT mutations can result in fatty liver and hepatocellular (liver) carcinomas.  BHMT mutations in mothers increase the risk of Down syndrome for their children.  Patients with BHMT polymorphisms are recommended to take betaine and zinc.

CBS

Cystathione beta-synthase (CBS) is a pyridoxal-5’-phosphate (vitamin B6) dependent enzyme that converts L-serine and L-homocysteine into L-cystathionine.   L-cystathionine is later converted into the amino acid cysteine.  Mutations to the CBS gene are the most common cause of hereditary hyperhomocysteinemia.   The adverse effects of homocysteine accumulation in the body are related to the substitution of homocysteine for methionine in protein synthesis. The resulting complications include an increase in immune response, increase in cell death, and protein damage. The degree of homocysteinemia is relative to the mutation.  Hyperhomocysteinemia has been linked to multiple mutations to the CBS gene.  The most common of these are the Ile278Thr and the Gly307Ser, which cause homocysteine to build up in the blood.  Complications of hyperhomocysteinemia include mental retardation, seizures, and vascular disease. One of the most common causes of death for patients with homocystinuria (CBS deficiency) is heart attack.   Patients with CBS polymorphisms are recommended to take glutathione and B6.  There are reports that the CBS polymorphisms A360A (rs1801181) and N212N (rs2298758) can lead to an increase in CBS activity.  Some claim that these mutations lead to a buildup of ammonia and decrease in glutathione. Since ammonia is a very unstable compound that must be measured STAT for accurate results, the better marker for increased ammonia is orotic acid which is very stable and accumulates when excessive amines are filtered through the urea cycle. I recommend that patients with this mutation do an Organic Acid test (OAT) and look at marker 60 (orotic acid) for ammonia and markers 58-59 (Pyroglutamic and 2-hydroxybutyric acid) for glutathione synthesis and cysteine accumulation respectively.

SHMT1

Serine hydroxymethyltransferase (SHMT1) is important for linked reactions.  The first is the conversion of tetrahydrofolate to 5,10-methylenetetrahydrofolate.  The second is the conversion of L-serine to glycine.  It also has a role in mediating the synthesis of dTMP and SAM. It preferentially selects for dTMP biosynthesis which is the precursor to the nucleic acid thiamine. Mutations in this enzyme may cause elevations in uracil which can build up when dTMP synthesis in impaired.  Uracil is marker 40 in the OAT.  

SUOX

Sulfite oxidase (SUOX) is an enzyme that is located in the mitochondria of cells.  The enzyme oxidizes sulfite to sulfate.  The physiological damage that occurs as a result of enzyme deficiency may be due to the accumulation of toxic levels of sulfite, from the absence of sulfate, or both. Sulfite is a reactive product of cysteine metabolism found in high concentrations in the brain. Its toxicity is exacerbated by glutathione depletion. Sulfate plays an important role in detoxification and deactivation of toxic compounds. Sulfate deficiency has been linked to autism, Parkinson’s disease, and Alzheimer’s disease.  Individuals with SUOX genetic mutations may benefit from a reduction in dietary methionine and cysteine.

VDR

Vitamin D receptor (VDR) is a nuclear hormone receptor for vitamin D3.  Vitamin D3 interacts with this receptor to influence multiple biological activities by regulating gene transcription.  Vitamin D3 is associated with maintenance of calcium distribution. More recently, it has been implicated in inflammatory processes, vascular integrity, and collagen formation. Mutations in VDR have been linked to metabolic syndrome.  Individuals with VDR mutations have greater propensity for insulin insensitivity, higher triglycerides, and lower HDL levels.  Vitamin D receptor mutations can also lead to vitamin-D-dependent rickets type 2.  Patients with VDR polymorphisms are recommended to take 1000 units/day for children or 5000 units/day for adults. 

I hope this information is helpful.  I know many of these pathways can be very intimidating, but hopefully we can work together to produce useful treatment plans for everyone.  Next week I plan on talking about the mental health genes MAO and COMT.

Email gplblog@gpl4u.com if you have any questions about this blog post.

Introduction to GPL Blog


Welcome to the Great Plains Laboratory’s new blog!  First, I would like to introduce myself.  I am Matt Pratt-Hyatt, Associate Laboratory Director at The Great Plains Laboratory.  I am excited to announce this new venue for sharing some of the information that GPL staff have gathered over the years during research and development.  Right here, every Monday, I will be writing and posting a weekly blog, however, we also plan to have guest bloggers contribute.  To supplement our weekly blogs, we will also provide additional content about twice a week, which may include relevant articles and multimedia.   

The Great Plains Laboratory, Inc. was founded by William Shaw, PhD back in 1996, building on the skills and knowledge that he acquired during his time at the Centers for Disease Control and Prevention and at Children’s Mercy Hospital of Kansas City.  In the subsequent years, we have helped over 400,000 patients address root causes of a variety of chronic health disorders, from autism and depression, to fibromyalgia and irritable bowel disease.    

I am looking forward to this opportunity to interact with you – our community of integrative health practitioners.  After working in academic research for 17 years, it has been a great experience joining the GPL team, which I have been a part of for about two years now.  As director of Research & Development, I am thrilled that our team has brought you new, cutting-edge tests like our Phospholipase A2 (PLA2) Test, GPL-TOX (our Toxic Organic Chemical Profile), Glyphosate, and GPL-SNP1000, our Next-Generation DNA Sequencing Profile.  I also enjoy talking with our clients by phone and at conferences, learning more about how you practice and how we can continue giving you even better diagnostic tools to improve the health of your patients. 

Through our interactions with both practitioners and patients, we have been able to further refine our test interpretations to explain how various biological markers reveal many different disease conditions.  We also share what we have learned about the best courses of treatment to recommend.  Through our one-on-one consultations, we have helped thousands achieve better health, but we have been seeking a better way to disseminate this information to an even larger audience ─hence our new blog.  

In addition, I will use this space to discuss the new tests that we will be launching in the future (and trust me, we have some important new tests coming out this year).  I look forward to your feedback on what you’d like to see from this blog.  What are your interests when it comes to integrative medicine? What kind of information would you find most valuable?  How have we helped to improve your life or a patient’s life?  Let us know.  I am happy to be working with you as we embark on this next part of our journey.  Our primary goal in all we do is to find more ways to help you and your patients achieve enhanced wellness.