Characterizing the Skeletal
Phenotype of db/db Mice
Student: Bailey Lindenmaier
Mentors: Dr. Russell Turner & Dr. Urszula Iwaniec
Skeletal Biology Lab
Introduction
• Age related Osteoporosis- Porous bone
• 1 in 3 women and 1 in 5 men over the age of 50
worldwide are estimated to have osteoporosis
• In 2005, osteoporosis-related fractures were
responsible for an estimated $19 billion in costs.
• Health risks
▫ Increases individuals risk of fractures
▫ Following peak bone mass, lost bone mass is
difficult to restore
Leptin
• Product of the OB gene
• Primarily produced by adipose tissue.
• Pleiotropic hormone
▫ Appetite control
▫ Energy regulation
▫ Immune response
▫ Cardiovascular and renal function
• Essential for achieving optimal peak bone mass.
Identifying the Mechanism of Leptin on
Bone Remodeling
• Leptin is a key hormone in
regulating and maintaining
healthy bones.
• Early work suggests that s.c.
leptin injection reduced bone
loss following ovx.
• Current data proposes that
leptin functions via the CNS.
• Osteoblast express leptin
receptor. Possible peripheral
affect?
• Answers may lead to a
generation of drugs to treat
osteoporosis.
Proposed Pathways of Leptin’s Effects
on the Skeleton
ObRb
Receptor
Leptin
db/db mice
• Loss of function in the leptin
receptor
• Mosaic skeletal type
▫ Overall reduced bone mass
▫ High cancellous bone mass
of vertebrae.
▫ Low bone mass in femur.
▫ Decreased length of femoral
bones.
• Obese, hypogonadic,
hyperinsulemic, hypercortisolic
db/db
Wild Type
Goal
Characterize skeletal growth
of WT and leptin-deficient
db/db mice.
Study Design
• A group of 10 WT and db/db
mice sacrificed at 8 weeks for
baseline.
• A group of 7 WT and db/db
mice sacrificed at 17 weeks.
• Fed ad libitum, 12h light/dark
cycle
• Both femurs and LV 5 were
excised at necropsy
B6.BKS(D)-Leprdb
µ CT Analysis
• Micro - Computed
Tomography
• Fires an x-ray beam at
a rotating specimen
• Produces 3D images
for structural
measurements
Bones Analyzed
Vertebral Cancellous Bone
Femoral
cortex
Femur Metaphysis
(trabecular bone)
Results
Total Femur
Bone Volume
Femur Length
16
ANOVA Effects
Genotype: p<.001
Age: p<.001
Genotype x Age: p<.002
19
18
Femur Bone Volume (mm 3)
Femur Length (mm)
15
14
13
12
ANOVA Effects
Genotype: p<.001
Age: p<.002
Genotype x Age: p<.004
17
16
15
14
13
12
11
11
10
0
10
0
WT
ob/ob
db/db
8 weeks
WT
ob/ob
db/db
17 weeks
WT
db/db
8 weeks
WT
db/db
17 weeks
Results
Femur Cortical
Thickness
210
200
Femur Cortical Thickness (µm)
DiaphysisCortical Bone
190
180
ANOVA Effects
Genotype: p<.001
Age: p<.001
Genotype x Age: p<.001
170
160
150
140
130
120
0110
100
WT
8 weeks
ob/ob
db/db
WT
ob/ob
db/db
17 weeks
Results
12
Metaphysis
Bone Volume/Tissue Volume
11
Femur Metaphysis BV/TV (%)
10
ANOVA Effects
Genotype: N.S
Age: p<.001
Genotype x Age: p<.038
9
8
7
6
5
4
3
2
1
0
WT
8 weeks
ob/ob
db/db
WT
ob/ob
db/db
17 weeks
Results
Lumbar Vertebrae 5
Bone Volume/Tissue
Volume
25
24
Lumbar Vertebrae 5 BV/TV (%)
23
22
21
ANOVA Effects
Genotype: p<.001
Age: p<.026
Genotype x Age: p<.007
20
19
18
17
16
15
14
13
12
0
WT
ob/ob
db/db
8 weeks
WT
ob/ob
db/db
17 weeks
Results
Lumbar Vertebrae 5
Trabecular Thickness
Lumbar Vertebrae 5
Trabecular Number
8
7
50
ANOVA Effects
Genotype: p<.001
Age: p<.001
Genotype x Age: p<.004
Lumbar Vertebrae 5 Trabecular Thickness (µm))
Lumbar Vertebrae 5 Trabecular Number
9
6
5
4
3
2
1
49
48
47
ANOVA Effects
Genotype: N.S
Age: p<.001
Genotype x Age: N.S
46
45
44
43
42
41
40
39
38
37
36
0
WT
db/db
8 weeks
WT
17 weeks
db/db
35
0
WT
8 weeks
db/db
WT
17 weeks
db/db
Conclusions
• Aging db/db mice have high bone mass in their
lumbar vertebrae
▫ Maintain trabecular number
▫ Increase in trabecular thickness
• Aging db/db mice have low bone mass in their
femur.
▫ Low cortical thickness
▫ Reduced femoral length
On the Horizon
• Histomorphometric
analysis of bone.
• Bone marrow
transplantation following
irradiation to determine if
leptin has a direct effect
on bone remodeling.
Acknowledgements
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Dr. Russell Turner
Urszula Iwaniec
Dawn Olson
Kenneth Philbrick
Kevin Ahern
Howard Hughes Medical Institute