clostridia metabolites inhibit human dopamine-beta-hydroxylase, increasing toxic dopamine...
TRANSCRIPT
Clostridia metabolites inhibit human dopamine-beta-hydroxylase,
increasing toxic dopamine metabolites and severe oxidative
stress in autism
William Shaw PhDThe Great Plains Laboratory,Inc
www.gpl4u.com
Structure of 3-(3-hydroxyphenyl)-3-hydroxypropionic acid
HO
CHOHCH2COOH 123
1
23
4
5 6
Phenyl group
Hydroxyl group
Propionic acid group
• “It was observed that mentally ill patients, in general, seem to excrete much larger amounts of HPHPA than do most normal people.”
• Most patients with mental retardation excrete very low amounts of HPHPA.
Journal of Biological Chemistry 225:269-278,1957
Increased Urinary Excretion of Analogs of Krebs Cycle Metabolites and Arabinose in Two Brothers with Autistic Features.
W Shaw. Clin Chem 41:1094-1104, 1995.
HPHPA
• C. difficile-pseudomembranous colitis• C. sporogenes• C. botulinum-food poisoning• C. mangenoti• C. ghoni• C. bifermentans• C. caloritolerans
Clostridia species that produce HPHPA precursors
0.00
200.00
400.00
600.00
800.00
1000.00
1200.00
1400.00
1600.00
1800.00
2000.00
N = 14
N = 30
N = 211
CONTROLMALE
INFANTS
CONTROLMALE
CHILDREN AUTISTICMALES
3-(
3-h
yd
rox
yp
he
ny
l)-3
-hy
dro
xy
-p
rop
ion
ic a
cid
(M
MO
L/M
OL
CR
EA
TIN
INE
)
2 - 13 YRS 2 - 13 YRS3-(3
-hyd
roxy
phen
yl)-
3-hy
drox
ypro
pion
ic
acid
HPH
PA -m
mol
/mol
cre
atini
ne
Controlinfants
Control boys2-13 years
Autistic Boys2-13 years
Figure 2. Distribution of values for HPHPA Clostridia metabolite in urine samples of male infants, control
boys, and boys with autism.
Endoscopy of colon of patient with severe Clostridium difficile overgrowth-
pseudomembranous colitis
Formation of p-cresol from tyrosine by Clostridia bacteria
tyrosine 4-hydroxy-phenylacetic
p-cresol p-cresol sulfate4-hydroxy-phenylpyruvic
14728–14733 PNAS August 25, 2009 vol. 106 no. 34
PST
CH2CHCOOH
NH2
CH2CHCOOH
NH2
HO
tyrosine
CH2CHCOOH
NH2
HO
microbial
tyrosine analog
CH2CH2COOH
CH2CH2COOH
HO
NH2
phenylpropionic acid
NH2
3-hydroxyphenyl-propionic acid
CH2CHCOOH
NH2
HO
3,4-dihydroxyphenylAlanine (DOPA)
HO
microbial
dopamine
norepinephrine
human beta-oxidation
CHOHCH2COOH
HO3-(3-hydroxyphenyl)-3-hydroxypropionic
acid (HPHPA)
HO
COOH
3-hydroxybenzoic acid
glycine
3-hydroxy-benzoylglycine(3-hydroxyhippuric)
microbial
human beta oxidation enzymes
phenylalanine phenylalanine hydroxylase
tyrosine hydroxylase
CH2COCOOHHO
4-hydroxyphenyl-pyruvic acid
CH3HO
4-cresol
HVAVMA
epinephrine
Critical effect of intestinal bacteria on brain neurotransmitters
Organic acid test
Organic acid test
Organic acid test
Intestine
** = products 0f Clostridia
* *
**
***
**
***
*
****
*******
Blood vessels
*******
**
************
*
*
********
***
*****
** * * ****
Urine cup
Kidney
* B***
*
***
***
* B
***************
*****
****
****
****
****
*******
**********B= bacteria (beneficial)
**** *
* * *
* *
=Clostridiabacteria
Brain
Body
*
Effect of HPHPA on brain neurotransmitters
HPHPAMmol perMolcreatinine
Date
HVAMmol per Molcreatinine
(Dopamine)Clostridia
Effect of HPHPA on brain neurotransmitters
Date
DopaminemetaboliteHVAMmol per Molcreatinine
11/18 2/26 6/6 9/14 12/23 4/10
5
10
15
20
25
30
0
1
2
3
4
5
6
7
VMA (norepi)
NorepinephrineMetaboliteVMAMmol per molcreatinineHVA (dopamine)
Effect of HPHPA on brain neurotransmitters
Date
11/18 1/7 2/26 4/17 6/6 7/26 9/14 11/3 12/23 2/110
500
1000
1500
2000
2500
3000
3500
4000
4500
5000
0
1
2
3
4
5
6
7
8
HPHPAMmol perMolcreatinine
HVA/VMAIndicator ofDopamine to Norepinephrineratio
Linan Chen, et al(2008) Unregulated cytosolic dopamine causes neurodegeneration associated with oxidative stress in mice. J. Neurosci. 28, 425–433
• Dopamine is a very reactive molecule compared with other neurotransmitters, and dopamine degradation naturally produces oxidative species.
• More than 90% of dopamine in dopamine neurons is stored in abundant terminal vesicles and is protected from degradation.
• However, a small fraction of dopamine is cytosolic, and it is the major source of dopamine metabolism and presumed toxicity.
• Cytosolic dopamine undergoes degradation to form 3,4-dihydroxyphenylacetic acid (DOPAC) and HVA as well as hydrogen peroxide via the monoamine oxidase pathway.
Linan Chen, et al(2008) Unregulated cytosolic dopamine causes neurodegeneration associated with oxidative stress in mice. J. Neurosci. 28, 425–433
• Alternatively, dopamine undergoes oxidation to form superoxide, hydrogen peroxide, and o-quinone or reacts with cysteine residues on glutathione or proteins to form cysteinyl-dopamine and cysteinyl-DOPAC conjugates.
• These biochemical abnormalities caused by excess dopamine may cause severe neurodegeneration of neural pathways that utilize dopamine as a neurotransmitter.
Prevalence of Clostridium difficile in the gastrointestinal tract of hospitalized children under two years of age. Med Dosw Mikrobiol; 2010;62(1):77-84 (Poland)
• 178 fecal samples of children aged 2 months to 2 years,hospitalized in 2003-2006 were examined for the presence of toxin A/B of C. difficile.
• Toxigenicity of strains was confirmed using PCR.• Susceptibility to antimicrobials was determined. • The percentage of children infected with C.
difficile was 68.6%.• All strains were susceptible to Vancomycin and
metronidazole(Flagyl)
Summary • Clostridia metabolites HPHPA, 4-cresol, and 4-
hydroxyphenylacetic are toxic because they inhibit key enzyme that converts dopamine to norepinephrine
• Toxic dopamine metabolites use up glutathione and person is much more susceptible to other toxic chemicals like mercury, pesticides, antibacterial soaps
B. Garreau et al. Disturbances in dopamine metabolism in autistic children: results of clinical tests and urinary dosages of homovanillic acid (HVA). Acta psychiatr belg 80:249-265,1980.
• Dopamine metabolite (homovanillic acid or HVA) in urine was more elevated in children with autism than in PDD and the severity of symptoms was related to the concentration of HVA
• High dopamine production in brain is associated with repetitive, stereotypical, obsessive, compulsive behaviors
• The effects of norepinephrine are alertness and arousal, and influences on the reward system. Norepinephrine is important for the exploratory behavior essential for learning relations between sensory input, decision processing, motor output, and behavioral feedback.
Factors involved in autism, Clostridia toxicity measured in urine organic acid test
• HVA-(homovanillic acid)-Major metabolite of dopamine, a major brain neurotransmitter associated with abnormal autistic behavior when it is elevated
• VMA-(vanillylmandelic acid)- Major metabolite of norepinephrine- important for the exploratory behavior essential for learning relations between sensory input, decision processing, motor output, and behavioral feedback.
Factors involved in autism, Clostridia toxicity measured in urine organic acid test
• 4-hydroxyphenylacetic- Clostridia metabolite that is a phenol that is detoxified by phenol sulfo-transferase (PST), leading to increased susceptibility to acetaminophen toxicity. 4-Cresol and HPHPA are also PST inhibitors.
• HVA /VMA ratio –indicates whether there is a healthy balance in the brain between norepinephrine and dopamine
Factors involved in autism, Clostridia toxicity measured in urine organic acid test
• Pyroglutamic acid- high values indicate deficiency of glutathione, a major cause of dopamine toxicity and increased susceptibility to most environmental chemicals
• 4-Cresol- Major metabolite of Clostridium difficile- blocks dopamine beta hydroxylase leading to excess dopamine and abnormal behavior
• HPHPA-Major metabolite of multiple Clostridia species- blocks dopamine beta hydroxylase leading to excess dopamine and abnormal behavior
Clinical usefulness of Clostridia treatments
• Schizophrenia• Psychosis• Depression• Chronic fatigue• Tics, Tourette’s• Autism
• ADD, ADHD• Obsessive compulsive
disorder (OCD)• Seizure disorders• Gastrointestinal
disorders, diarrhea, constipation,Crohn’s disease,colitis
Effects of 3-hydroxyphenylalanine in rats
• Headweaving• Predominantly backward walking• Wet dog shakes• Hyperactivity• Hyper-reactivity
Dyck LE, Kazakoff CW, Dourish CT. The role of catecholamines, 5-hydroxytryptamine and m-tyramine in The behavioural effects of m-tyrosine in the rat. European Journal of Pharmacology 1982; 84(3-4): 139-149
Two months of nystatin and Lactobacillus acidophilus GG therapy in a child with autism
Candida krusei stool
Yeast tartaric urine*
Lacto-bacillus stool
Clostridia HPHPA urine*
Before
4+
993
0
3265
After
0
1
4+
174
normal range
0-1+
0-15
3+ - 4+
0-150* mmol/mol creatinine
Effect of anti-Clostridia therapy on urine excretion of HPHPA* in
young woman with acute psychosis-auditory
hallucinations patient normals During acute 7489 0-150 psychosis After treatment 673 0-150 ( depressed but no psychosis) *mmol/mol creatinine
patient normals Before treatment 1444 0-150
After treatment 13 0-150
*mmol/mol creatinine
Effect of anti-Clostridia therapy on urine excretion of HPHPA* in woman with
depression and chronic fatigue
Treatments for Clostridia bacteria• Vancomycin-oral-not intravenous-5-10 mg/Kg/day div into 3
doses• Flagyl (metronidazole)-30 mg/Kg/day div into 3 doses-10 days• Lactobacillus acidophilus GG-10 -100 billion per day • Alternate name Lactobacillus rhamnosus• (Only bacteria probiotic patented for use in control of
Clostridia)-Culturelle-VSL #3• Saccharomyces boulardi (yeast)• Micellized or IV glutathione or n-acetylcysteine to increase
brain glutathione and reduce neurotoxic dopamine metabolites
• High protein diet (phenylalanine, tyrosine) may increase production of toxic Clostridia metabolites
Summary of Clostridia dopamine abnormalities in autism
• Elevated HPHPA and similar phenolic compounds from multiple species of Clostridia are elevated in more than two-thirds of children with autism.
• Elevated HPHPA and cresol inhibit the conversion of dopamine to norepinephrine, resulting in marked increase in dopamine.
• Dopamine metabolites deplete brain glutathione and also produce marked excess of oxygen superoxide free radicals, 2500 molecules for each dopamine metabolite
• Brain and sympathetic nervous systems normally using norepinephrine switch to dopamine, drastically altering brain and sympathetic nervous system function
Summary of Clostridia dopamine abnormalities in autism
• Presence of this abnormality is determined with HPHPA, cresol, and other phenolic compounds in the Great Plains organic acid test
• Stool testing not useful since HPHPA producing and Non-HPHPA species are not differentiated
• Glutathione depletion by excess dopamine determined by pyroglutamic acid marker in organic acid test.
• Clostridia treated for 10-14 days with vancomycin or flagyl,followed by several months high dose probiotics-culturelle or VSL-3
• Repeat testing every 3 months to prevent recurrence