islet insulin production in diabetes model: spatial comparison of mouse strains
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(C) 2005 BS Yandell UW Summer Research Program 1
islet insulin production in diabetes model:spatial comparison of mouse strains
• mentor: Brian S. Yandell, UW-Biostat Trainee www.stat.wisc.edu/~yandell
• scientist: Donnie Stapleton, Attie Biochem Labdsstaple@biochem.wisc.edu
• objective of project:– improve methods to distinguish "quality"– of insulin production– between two strains of mice – based on 3-color images of islets in pancreas slices
(C) 2005 BS Yandell UW Summer Research Program 2
why do this project?
• Knowledge/Skills needed– Basic statistics (t-test, ideally some nonparametrics)– R programming (a la biostat training)
• Knowledge/Skills obtained– communication skills with scientist (lab biochemist)– image analysis basics– nonparametric density estimation (mechanics at least)– experimental design considerations (nesting, balance,
etc.)
(C) 2005 BS Yandell UW Summer Research Program 3
project abstract• insulin is produced in the pancreas
– specifically the Islets of Langerhans by "beta" cells– (there are "alpha" cells as well.)
• scientists in biochemistry have imaged hundreds of islets– for two strains of obese mice
• BTBR.ob mimics type II (mature onset) diabetes mellitus• B6.ob appears healthy
• roughly a hundred islets per mouse, 3 mice per strain– can visually see differences in production of insulin in beta cells – have summary measurements on many islets to compare strains
• opportunity beyond data analysis– how to improve measurement process? – how to improve experimental design?
• goal: clearly infer differences in "quality" of insulin production– between BTBR.ob and B6.ob
• you can meet with the scientist and work with measurement system
(C) 2005 BS Yandell UW Summer Research Program 4
of mice and men
• obese strains: BTBR.ob vs. B6.ob– obese BTBR mice mimic human diabetes
• high glucose, low insulin– obese B6 mice appear non-diabetic
• congenic substrains A and B– mostly BTBR mouse genome– A congenic: insert small segment of B6– B congenic: essentially BTBR mice
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glucose insulin
(courtesy AD Attie)
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Type 2 Diabetes Mellitus
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Insulin Requirement
from Unger & Orci FASEB J. (2001) 15,312
decompensation
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islet degradation seriesA vs. B congenics
1 2 3 4 5healthy sick
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comparison of 3 type A
and 2 type B congenic
mice
healthy sick
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A and B congenic dataA congenics:Class.I Class.II Class.III Class.IV Class.V Total.IsletsA33-18 26 36 23 21 40 146A34-22 55 36 24 18 10 143A39-15 69 60 41 35 41 246
B congenics:Class.I Class.II Class.III Class.IV Class.V Total.IsletsA34-19 87 37 35 35 9 203A48-4 44 29 12 19 7 111
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B6 Female&Male OB 14wk
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Btbr Female& Male OB 14wk
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Btbr Male & Female OB 14wks
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total percent area for islets
0 50000 100000 150000
0.0
e+00
1.0
e-05
2.0
e-05
3.0
e-05
B6.1105
N = 21 Bandw idth = 6625
Den
sity
0 50000 100000 150000
0 e
+00
2 e
-05
B6.1169
N = 13 Bandw idth = 6111
Den
sity
0 50000 100000 150000
0 e
+00
2 e
-05
4 e
-05
6 e
-05
B6.1195
Den
sity
0 50000 100000 150000
0.0
e+00
1.0
e-05
2.0
e-05
BT.1419
N = 30 Bandw idth = 8943
Den
sity
0 50000 100000 150000
0.0
e+00
1.0
e-05
BT.1429
N = 34 Bandw idth = 1.246e+04
Den
sity
0 50000 100000 150000
0.0
e+00
1.0
e-05
2.0
e-05
BT.1507
Den
sity
12 13 14 15 16 17
0.00
0.10
0.20
0.30
B6.1105
N = 21 Bandw idth = 0.4719
Den
sity
12 13 14 15 16 17
0.0
0.1
0.2
0.3
0.4
B6.1169
N = 13 Bandw idth = 0.4596D
ensi
ty
12 13 14 15 16 17
0.0
0.2
0.4
0.6
B6.1195
Den
sity
12 13 14 15 16 17
0.00
0.10
0.20
0.30
BT.1419
N = 30 Bandw idth = 0.5141
Den
sity
12 13 14 15 16 17
0.00
0.05
0.10
0.15
0.20
0.25
BT.1429
N = 34 Bandw idth = 0.5427
Den
sity
12 13 14 15 16 17
0.00
0.10
0.20
BT.1507
Den
sity
log2 totalsraw totals
(C) 2005 BS Yandell UW Summer Research Program 15
percent glu/som,
dapi, insulin
by mouse
(smooth histogram +
median)
0 20 40 60 80 100
0.00
0.04
0.08
0.12
B6.1105
0 20 40 60 80 100
0.00
0.02
0.04
0.06
0.08
B6.1169
0 20 40 60 80 100
0.00
0.02
0.04
0.06
0.08
B6.1195
0 20 40 60 80 100
0.00
0.02
0.04
0.06
0.08
0.10
BT.1419
0 20 40 60 80 100
0.00
0.02
0.04
0.06
BT.1429
0 20 40 60 80 100
0.00
0.02
0.04
0.06
0.08
0.10
BT.1507
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sum of insulin, glu/som,
dapi percents
60 80 100 120 140
0.00
00.
010
0.02
00.
030
B6.1105
N = 19 Bandw idth = 5.231
Den
sity
60 80 100 120 140
0.00
00.
005
0.01
00.
015
0.02
0
B6.1169
N = 13 Bandw idth = 8.969
Den
sity
60 80 100 120 140
0.00
0.01
0.02
0.03
0.04
B6.1195
Den
sity
60 80 100 120 140
0.00
00.
010
0.02
00.
030
BT.1419
N = 30 Bandw idth = 5.164
Den
sity
60 80 100 120 140
0.00
00.
010
0.02
0
BT.1429
N = 34 Bandw idth = 6.835
Den
sity
60 80 100 120 140
0.00
00.
010
0.02
00.
030
BT.1507
Den
sity
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i
0 10 20 30 40 50 60 80 100 120 140
3050
7090
010
2030
4050
g
d
1020
3040
50
30 50 70 90
6080
100
120
140
10 20 30 40 50
tot
perc
ent
pairs
(cor
rela
tion)
plo
t
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inte
nsity
pairs
(cor
rela
tion)
plo
ti
600 620 640 660 680 700 720
700
750
800
850
900
600
620
640
660
680
700
720
g
700 750 800 850 900 1000 1200 1400 1600
1000
1200
1400
1600
d
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calib
ratio
n st
udy
200 400 600 800 1000
1000
2000
3000
intensity
millisec
inte
nsity
200 400 600 800 1000
2000
040
000
6000
0
thresholded
millisec
thre
shol
ded
200 400 600 800 1000
2030
4050
6070
percent
millisec
perc
ent
200 400 600 800 1000
68
1012
14
signal.noise
millisec
sign
al.n
oise
(C) 2005 BS Yandell UW Summer Research Program 20
90
80
70
60
50
40
30
140x10312010080604020Area (um^2)
B6 Btbr
Dot plo:t Insulin vs
Area of islet
(C) 2005 BS Yandell UW Summer Research Program 21
50
40
30
20
10
140x10312010080604020Area (um^2)
B6 Btbr
Dot plot:
Glu/Som vs
Area of Islet
(C) 2005 BS Yandell UW Summer Research Program 22
90
80
70
60
50
40
30
5040302010Percent Glu/Som
B6 Btbr
Dot plot: Insulin vs
Glu/Som
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Female ob/ob B6
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Female ob/ob BTBR
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Male ob/ob B6
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Male ob/ob BTBR
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Summary table for ob/ob B6 and Btbr islets
All animals ob/ob and 6 wk of age.#'s show mean of guassian fit for all islets analyzed.
Islet area Islet diameterStrain # islets (m2) (m) % Gluc&Som % DAPI % Insulin Total %
B6 F 54 12864 128 6 25 71 102Btbr F 104 14242 135 8 23 61 92
B6 M 107 13545 131 10 24 61 95Btbr M 106 11520 121 17 25 58 100
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14 week Congenic Females
• Is there a difference between A & B congenics?
• Is there a difference between Btbr and the B congenic strain?
• How does the glucose and insulin values effect our comparisons?
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Btbr Female OB 14wk
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A33-18, Female OB A 14wk
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A34-19, Female OB B 14wk
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A34-22, Female OB A 14wk
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A39-15, Female OB A 14wk
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A48-4, Female OB B 14wk
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