effects of glycomacropeptide, amino acid & casein diets on osteopenia in pku mice denise m. ney,...
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Effects of Glycomacropeptide, Amino Acid & Casein Diets on
Osteopenia in PKU Mice
Denise M. Ney, PhD, RD Professor of Nutritional Sciences
Waisman CenterUniversity of Wisconsin-Madison
Disclosure D Ney is a co-inventor on US Patent
Application US-2010-0317597, GMP Medical Foods for Nutritional Management of PKU, which is held by the Wisconsin Alumni Research Foundation and licensed to Cambrooke Foods, LLC. A percentage of all royalty payments is awarded to the inventors.
D Ney has received consulting income from Cambrooke Foods and BioMarin.
Funding: National PKU Alliance & USDA Hatch Grant
Denise Ney Robert Blank Sangita Murali Patrick Solverson
UW-Madison PKU Bone Research Team
Background PKU is associated with low bone mass,
or osteopenia, and fractures in early adulthood.– 57% of 28 patients had
osteopenia/osteoporosis• Perez-Dueñas et al. Acta Paediatr 91:800, 2002
Reduced bone mineral density (BMD) in PKU is present from an early age onward and it cannot be predicted by plasma phe levels.– 20% of 53 patients studied had osteoporosis
• de Groot et al. Mol Genet Metab 101:566, 2012
What causes skeletal fragility in PKU?
The fundamental question is whether reduced BMD is inherent to PKU or secondary to its dietary management.
In order to isolate the contributions of the PKU genotype itself and dietary treatment of PKU we have conducted a factorial experiment in PKU mice.
Objective & Design To determine how the PKU genotype and
the source of dietary protein affect growth, body composition and bone development.
WTPKU
♂♀ AA
High Phe
Low Phe
Casein
GMP
PKU & WT Mice Housed With
Same Sex Littermates
↓phe GMP ↓phe AA ↑phe Casein
GMP is a natural whey protein produced when
making cheese. Pure GMP contains no phe.
Growth
Solverson, P et al Am J Physiol Endocrinol Metab 302:E885-95, 2012
Males
3 6 9 12 15 18 21 24
Bo
dy
Wei
gh
t (g
)
5
10
15
20
25
30
35
Age (Week)
Genotype x Diet p = 0.0053
PKU AA (12)b PKU CAS (16)b
PKU GMP (11)b WT AA (13)b WT CAS (15)a
WT GMP (17)b
Females
3 6 9 12 15 18 21 24
Bo
dy
Wei
gh
t (g
)
5
10
15
20
25
Age (Week)
Genotype x Diet p = 0.0257
PKU AA (18)bc PKU CAS (14)c
PKU GMP (11)bc WT AA (14)ab WT CAS (19)a
WT GMP (20)b
Metabolic Phenotyping Platform
●Food & water intake●O2 consumption & CO2 production
PKU mice show increased energy expenditure with
casein diet
Lean Body Mass (g)
12 14 16 18 20 22 24 26
VO
2 (m
l/hr)
60
70
80
90
100
110
120
PKUab
b
a
ccc
WT
3-15%
WTAA (23)
WTCAS (16)
WTGMP (26)
PKUAA (18)
PKUCAS (21)
PKUGMP (16)
0
1
2
3
4
5F
oo
d In
take
(g
/day
)
Wild Type PKU
Cas
ein
(17
)
AA
(23
)
GM
P (
30)
Cas
ein
(21
)
AA
(18
)
GM
P (
17)
genotype x diet p = 0.0193
sex p = 0.0404males - afemales - b a
a
bbbb
Values with different letter superscripts are significantly different, p<0.05
GMP Diet Normalizes Food Intake in PKU
Mice
AA Diet Increases Kidney Workload in both WT and PKU
Mice
Values with different letter superscripts are significantly different, p<0.05
24 Hour Water Intake
24 h
r W
ater
Inta
ke (
ml)
0
2
4
6
8
WT PKU
Cas
ein
(20
)
AA
(23
)
GM
P (
30)
Cas
ein
(21
)
AA
(18
)
GM
P (
16)
a a
a
bb b
Kidney Mass
0.0
0.4
0.8
1.2
1.6
2.0
g/1
00g
BW
Wild Type PKU
AA
(27
)
Cas
ein
(34
)
GM
P (
36)
AA
(30
)
Cas
ein
(30
)
GM
P (
22)
c
a
cb ab
c
Does PKU increase energy needs?
Resting energy expenditure is ~5-10% higher in adolescent females with PKU than that predicted by standard equations.– J Am Diet Assoc 110:922-25, 20101– Am J Clin Nutr 62:797-804, 1995
Reduced growth occurs in children with PKU– Mol Genet Metab 101:99-109, 2010– J Pediatr 26:1-11, 2002
Dual Energy X-Ray Absorptiometry (DXA)
PKU mice show reduced whole-body bone mineralization compared to WT
miceB
MD
(m
g/c
m2 )
0
10
20
30
40
50
60
WT (98) PKU (86)
a b
Femur Strength: 3 Point Loading Test
Loading force being applied to a mouse
femur
Displacement
Load (F)
Displacement, mm
Load-Displacement Curve
0.3
-2-0.1 0.1 0.2 0.4 0.5 0.6 0.7 0.8
Loa
d, N Fracture
Yield
Point
Elastic Deformation
Post-Yield Displacement
12
10
8
6
4
2
14
16
Max Load
0
2
4
6
8
10
12
14
16
WT (116)
Yie
ld L
oa
d (
N)
PKU (101)
ab
WT (116)
PY
D (
mm
)
0.0
0.1
0.2
0.3
0.4
0.5
0.6
PKU (101)
ab
PKU mice show bones that are brittle and break easily
0.0 0.2 0.4 0.6 0.8
0
2
4
6
8
10
12
14
16
18
20Wild TypePKU
Displacement (mm)
Lo
ad (
N)
Yield Load
Fracture
Post-Yield Displacement
PKU mice show reduced femoral bone mineralization compared to WT mice
0
5
10
15
20
25
30
Bone Mineral Content
WT (116)
BM
C (
mg
)
PKU (101)
ab
0
10
20
30
40
50
60
Bone Mineral Density
WT (116)
BM
D (
mg
/cm
2 )
PKU (101)
a b
How the casein, AA and GMP diets affect bone size
and strength?
GMP Increases Bone Size in PKU & WT Mice
Cross Sectional Area
Casein
(m
m2 )
0.0
0.2
0.4
0.6
0.8
1.0
1.2
1.4
AA GMP
a ab WT MiceFed Casein
AA
Casein
GMP
0
2
4
6
8
10
12
14
16
18
Max
Lo
ad (
N)
Casein AA GMP
aab b WT MiceFed Casein
GMP Improves Bone Strength in PKU and WT Mice Compared with
AA Diet
Summary
PKU mice have femora that are brittle and weak. This suggests defects in both collagen synthesis and mineralization.
GMP increases bone size and strength; femora tolerate a higher max load before fracture compared with the AA diet. – How does GMP work to improve bone strength in
mice?
What causes skeletal fragility in PKU?
Inherent to PKU genotype and/or Secondary to management with an AA
diet.
Answer = both genotype and diet
ConclusionSkeletal fragility is inherent to the
PKU genotype and is attenuated by a GMP compared with an AA diet in mice.
Future research is needed to determine if improved low-phe diets containing GMP reduce skeletal fragility in human PKU.
Take Home Message: How can bone health be improved in
PKU?
Follow a low-phe diet to improve growth and bone development
Include weight bearing exercise each day
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