Transcript Ueland B6

Direct and functional markers of B6-status
and some novel aspects of disease
prediction
Content
• B6 vitamers and some PLP-dependent reactions
• Direct vitamin B6 biomarkers
- PAr index as predictor of disease
• Functional B6 biomarker
- Plasma HK/XA ratio
• Summary and conclusion
B6 vitamers
Kynurenines
and
vitamin B6
Aminotransferases (ATs)
Serine hydroxymethyltransferase (SHMT)
and the glycine cleavage system (GCS)
Transsulfuration
pathway and
vitamin B6
Vitamin B6 biomarkers
• Direct B6 biomarkers
-
Plasma pyridoxal 5´-phosphate (PLP)
Plasma pyridoxal (PL) and total B6-aldehyde (PLP + PL)
Plasma 4-pyridoxic acid (PA)
Urinary excretion of PA and other B6 vitamers
Erythrocyte pyridoxal 5’-phosphate
Plasma vitamer ratios, the PAr index
B6 vitamers in cerebrospinal fluid (CSF)
• Functional B6 biomarkers
- The transaminase tests
- Plasma kynurenines
- Amino acids, transsulfuration and one-carbon metabolites
5 important
confounders
5 important
confounders
Direct vitamin B6 biomarkers
Plasma pyridoxal 5´-phosphate (PLP)
Reflects PLP content in liver
Validity as a status indictor in a diseased population has been questioned
• Pros
- Most commonly used B6 biomarker
- Essentially unaffected by renal function
- ICC of 0.65
• Cons
-
Unstable in serum/plasma (PLP -> PL)
Decreases during inflammation and acute phase
Decreases at low albumin and high alkaline phosphatase
Affected by common polymorphisms in the ALPL gene
Inorganic phosphate may inhibit ALP and increase PLP
Decreases during alcohol consumption
Decreases at high blood glucose/HbA1c; fasting samples
recommended
Plasma pyridoxal (PL)
The transport form of vitamin B6
Suggested as a surrogate marker of intracellular PLP
• Pros
- Essentially unaffected by renal function
- More responsive than PLP to pyridoxine supplementation
- Not affected by common polymorphisms in the ALPL gene
• Cons
-
Variable stability in serum/plasma (PLP -> PL)
Variable ICC
Less responsive than PLP to intake
Not properly validated
Total B6-aldehyde (PLP +PL)
PLP + PL account for about 80% of total B6 in plasma
• Pros
-
Corrects for the PLP to PL conversion during sample storage
Therefore, more stable than PLP or PL
Essentially unaffected by renal function
Not affected by alkaline phosphatase activity
• Cons
- Not properly validated
- Cut-off values not established
Plasma pyridoxic acid (PA)
A vitamin B6 catabolite with high renal clearance
A short-term marker of vitamin B6 status
• Pros
- Stable
- Not affected by common polymorphisms in the ALPL gene
- Not protein-bound; not affect by serum albumin level
• Cons
-
Strongly related to renal function
Increases several-fold in patients with renal dysfunction
Positively related to markers of cellular immune activation
Increases markedly in critically ill patients
Urinary pyridoxic acid (PA)
Represents > 90 % of vitamin B6 species excreted into the urine
A measure of recent vitamin B6 intake
• Pros
- Stable
- Changes fast according to recent B6 intake
- A useful instrument for the assessment of vitamin B6
requirements
- Not affected by acute phase inflammatory status
- No influence from age, pregnancy, alcohol and oral
contraceptives
• Cons
• Decreases in subjects with inadequate riboflavin status
- Circadian variations (spot urine)
Erythrocyte PLP
Responsive marker of vitamin B6 intake
Reflects vitamin B6 status also in a diseased population
• Pros
- No or positive associations with inflammation and ALP
- No or inverse associations with albumin
- Essentially unaffected by renal function
• Cons
- PLP binds to hemoglobin
- Different hemoglobin variants have different affinity
- The assay is cumbersome with variable recovery and low
precision
- Data on reference values are sparse and inconsistent
The plasma PAr index
PAr = PA/(PLP+PL), an example of attenuating influence from confounders
Reflects B6 catabolism during inflammation, and is not responsive to B6 status
Inflammatory markers account for > 90% of the explained variance of PAr
In ROC analysis, PAr is a strong discriminator of inflammation
• Pros
- A higher ICC (of 0.75) than other ratios and B6 vitamers
- Renal function (eGFR) has a minor impact on PAr
- A strong predictor of conditions linked to inflammation
• Cons
- Under evaluation
ROC plot for the diagnostic accuracy of PAr (solid line) and
PLP (dashed line) to discriminate high (> 95th percentile)
summary score of CRP, neopterin and KTR
Ulvik et al (2014)
Am J Clin Nutr,
100: 250
The PAr index as predictor of overall and lung cancer
during a follow-up time of 12 years among 6539 participants
from the the Hordaland Health Study
Zuo et al (2015) Int J Cancer 136: 2932
The PAr index as predictor of all-cause mortality in
cardiovascular patients
Ulvik et al (2015) unpublished
Functional B6 biomarkers
The transaminase tests
Erythrocyte aspartic acid transaminase (EAST) and alanine transaminase (EALT)
Basal activities (oEAST, oEALT) and/or activity coefficients (EAST-AC, EALT-AC)
Long-term indicators related to life span of the erythrocytes
• Pros
- Measurement of ACs partly attenuates variations due to method
differences
- No associations with inflammation, ALP and albumin
- Unaffected by renal function
• Cons
-
Less responsive to B6 intake than plasma PLP and urinary PA
Fresh blood must be used within hours after acquisition
Frozen erythrocyte samples will give false results
Apoenzymes increase during tissue necrosis
Decreases during acute phase and alcohol intake (EAST)
Cumbersome and obsolete
Metabolic markers
• Reflect vitamin function in tissues
• Less affected by short-term vitamin intake and distribution
• Allow assessment of vitamin status during
supplementation
Plasma kynurenines: 3-Hydroxykynurenine
Removal but not formation involve PLP-dependent enzymes
The only kynurenine that increases during B6-deficiency
• Pros
-
ICC of 0.55-0.68
Plausible mechanism
Increases at low plasma PLP
Non-linear association with PLP, with inflection point at 20 nmol/L
A marked reduction after B6 supplementation
• Cons
- Increases during inflammation
- Positively associated with tryptophan
- Association with B6 status mainly confined to subjects with
inflammation
- Increases in subjects with renal dysfunction
3-Hydroxykynurenine (HK) and B6 status
From Midttun et al (2011) J Nutr 141: 611
Plasma HK/XA ratio
HK/XA ratio, the ratio between 3-hydroxykynurenine and xanthurenic acid
An example of ratio attenuating influence from potential confounders
• Pros
-
Discriminates subjects with low PLP better than HK
A marked reduction in HK/XA after pyridoxine supplementation
PLP is the strongest determinant of HK/XA
Non-linear association with PLP, with inflection point at 20 nmol/L
Compared with HK, a weaker or no association with
inflammation, BMI, and kidney function
- Not affected by polymorphisms in the ALP gene
• Cons
- Under evaluation
Dose-response for HK and HK/XA ratio
versus PLP
From Ulvik et al (2013) Am J Clin Nutr 98: 934
HK/XA and other ratios and B6 status
From Ulvik et al (2013) Am J Clin Nutr 98: 934
HK/XA and other ratios and B6 status
From Ulvik et al (2013) Am J Clin Nutr 98: 934
Other potential functional biomarkers
• Plasma
-
Cystathionine
Glycine
Serine
Creatine
Dimethylglycine
The 2-oxoglutarate:glutamate ratio
• Urine
- Cystathionine after a methionine load
- XA after a tryptophan load
- Kyn and HK after a tryptophan load
Conditions that may affect B6 biomarker
concentrations independent of B6 status
Condition
Mechanisms/confounder
Infl
ALP
Cancer
⬆
⬆
CVD
⬆
iP
Kdn
Other
⬇︎
⬆
Pregnancy
Alb
⬇︎
⬆
Hemodilution
Infancy < 6m
Age
Age > 60y
⬆
Menopause
⬇︎
Severe obesity
⬆
OCs
⬆
⬆
Bone loss
⬆
⬇︎
Alcohol
Smoking
⬇︎
⬆
⬇︎
Acetaldehyde
Biomarker
Main
response
PLP
⬇︎︎
PLP
⬇︎
PLP/PL/uPA/AT
⬇︎/⬆/➡/︎︎➡
PLP/ePLP/AT
⬆/⬆/⬆
PLP/PA
⬇︎/➡
PA
⬆
PLP
⬆
PLP
⬇︎
PLP/uPA
⬇︎/︎︎➡
PLP
⬇︎
PLP/ePLP
⬇︎/︎︎➡
Conditions that may affect B6 biomarker
concentrations independent of B6 status
Summary by graphics
Summary by graphics
Summary by graphics
Summary by graphics
Summary by graphics
Summary by graphics
Summary by graphics
Summary by graphics
Summary by graphics
Summary by graphics
Summary by graphics
Summary by graphics
Summary by graphics
Acknowledgement
Arve Ulvik, Bevital, Bergen
Luisa Rios-Avila, University of Florida
Øivind Midttun, Bevital, Bergen
Jess F. Gregory, University of Florida
Ottar Nygård, University og Bergen