matt dean's presentation 111
TRANSCRIPT
Matthew Dean
Early Embryonic Loss(55%)
(Zinaman et al. 1996; Diskin et al. 2006; Wilcox et al. 1988)
Fertilization Failure(5%)
Live Birth(33%)
Late Embryonic/Fetal Loss
(7%)
Early Embryonic Loss(55%)
Fertilization Failure(5%)
Live Birth(33%)
Late Embryonic/Fetal Loss
(7%)
(Zinaman et al. 1996; Diskin et al. 2006; Wilcox et al. 1988)
Early Embryonic Loss(55%)
Fertilization Failure(5%)
Live Birth(33%)
Late Embryonic/Fetal Loss
(7%)
(Zinaman et al. 1996; Diskin et al. 2006; Wilcox et al. 1988)
Early Embryonic Loss(55%)
Fertilization Failure(5%)
Live Birth(33%)
Late Embryonic/Fetal Loss
(7%)
(Zinaman et al. 1996; Diskin et al. 2006; Wilcox et al. 1988)
Early Embryonic Loss(55%)
Fertilization Failure(5%)
Live Birth(33%)
Late Embryonic/Fetal Loss
(7%)
(Zinaman et al. 1996; Diskin et al. 2006; Wilcox et al. 1988)
Zygote
Uterus
Cervix
Vagina
Oviduct
(Hansson 1947; Senger 2005)
Ovary
Morula
Uterus
Cervix
Vagina
Ovary
Oviduct
(Hansson 1947; Senger 2005)
BlastocystUterus
Cervix
Vagina
Ovary
Oviduct
(Hansson 1947; Senger 2005)
Oviduct
Uterus
Cervix
Vagina
Histotroph:•Hormones•Nutrients•Glucose
(Bazer et al. 2012)
Oviduct
Uterus
Cervix
Vagina
Histotroph:•Hormones•Nutrients•Glucose
(Bazer et al. 2012)
EndometriumUterine Lumen
(Bazer et al. 2012; Yánez et al. 2003)
Glucose
Glycogen
Pyruvate
X
Glycogen
Glycogen
glucose
-6PHexokinase
Glycogen
glucose
-6PHexokinase
Glycogen
glucoseCellular Metabolism
-6P -UDPHexokinase
Glycogen
glucoseCellular Metabolism
-6P -UDPHexokinase
Glycogen Synthase
Glycogen
glucoseCellular Metabolism
-6P -UDP -1PHexokinase
Glycogen Synthase
Glycogen Phosphorylase
Glycogen
glucoseCellular Metabolism
-6P -UDP -1PHexokinase
Glycogen Synthase
Glycogen Phosphorylase
-6P
Glycogen
glucoseCellular Metabolism
-6P -UDP -1PHexokinase
Glycogen Synthase
Glycogen Phosphorylase
-6P
Glucose-6-Phosphatase
Glycogen
glucoseCellular Metabolism
-6P -UDP -1PHexokinase
Glycogen Synthase
Glycogen Phosphorylase
-6P
Glucose-6-Phosphatase
Glycogen
glucoseCellular Metabolism
Estrus/
Mated
Glyc
og
en
C
on
ce
ntratio
n
(m
g/ 1
00
g
w
et tissu
e)
(modified from Demers et al. 1972: Gupta et al. 2013)
Day Post Mating
Implantation
Estrus
0 1 2 3 4
Estrus
1 2 3 4 5
1 2 3 4 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 … 50
ImplantationEstrus
Implantation
Day
Day
1) Characterize changes in uterine glycogen metabolism during estrus, embryonic diapause, and pregnancy in mink.
2) Determine if the glycogenic effect of estradiol on uterine epithelia is direct.
Feb AprilMarch May
15 20 25 5 10 15 20 25 30 5 10 15 20 25 30 5 10 15
Estrus Implantation Parturition
EstrusDiapause
Pregnancy
3 Collections
Objective 1.Characterize changes in uterine glycogen metabolism during estrus,
embryonic diapause, and pregnancy in mink.
0
5
10
15
20
25
30
(Nested ANOVA A-CP<0.05)
Estrus IIS IS
A
B
C
Gly
cog
en
Co
nce
ntr
ati
on
(mg
/mg
tis
sue
)
DiapausePregnancy
Myometrium
Endometrium
Periodic Acid-Schiff-stain for carbohydrates
0
10
20
30
40 A
B
C
(ANOVA and Tukey’s, P<0.05)
Pe
rce
nt
Are
a
Estrus Diapause Pregnancy
Estrus
Diapause
Pregnancy
Myometrium
Endometrium
LE
GE
S
0
10
20
30
40
50
Glandular Epithelia
Lumenal Epithelia
Stroma
A
B B
[[
*
(Nested ANOVA A-B,* P<0.05)
Gly
coge
n C
on
ten
t(P
erce
nt
Are
a)
Estrus Diapause Pregnancy
Estrus
Diapause
Pregnancy
*
Estrus Diapause Pregnancy
Enzyme= BlueCounterstain=red/pink
-6P
Estrus Diapause Pregnancy
Gly
coge
n
Syn
thas
eG
lyco
gen
P
ho
sph
ory
lase
Enzyme= BlueCounterstain=red/pink
0
1
2
3
4
5
6
Re
lative
m
RN
A E
xp
re
ssio
n
C
B
A
Estrus Diapause Pregnancy
-6P
Estradiol???
Cont Estradiol
Perc
ent
Are
a
0.0
0.1
0.2Glandular Epithelial Glycogen
*
Estrus
EstradiolEstrus
EstradiolEstrus
?
1) Characterize changes in uterine glycogen metabolism during estrus, embryonic diapause, and pregnancy in mink.
2) Determine if the glycogenic effect of estradiol on uterine epithelia is direct.
Mink uterine epithelia
Immortalized with SV40 large T antigen
Positive for cytokeratin 18
(Moreau et al. 1995; http://micro.magnet.fsu.edu)
Cytokeratin 18uteru
sG
MM
e
Mucin 1
uteru
sG
MM
e
HexokinaseGlycogen Synthaseu
terus
GM
Me
uteru
sG
MM
e
Glycogen Phosphorylase u
terus
GM
Me
uteru
sG
MM
eGlucose-6-
Phosphatase
0 0.1 1 10 100
0.0
0.1
0.2
0.3
Gly
co
gen
(g
/10
6 c
ells)
Estradiol (nM)
Insulin (g ml-1)0 1.8 5.5 16.6 50
Gly
co
gen
(
g/1
06 c
ells)
0.0
0.2
0.4
0.6
**
(*P<0.05 relative to 0 μg ml-1)
PPPT- Igf1r antagonist(*P<0.05 relative to 0 μg ml-1)
Gly
co
gen
(
g/1
06 c
ells)
0.0
0.1
0.2
0.3
Cont PPPT Insulin Insulin+PPPT
*
InsulinEstradiol
00
500
500.1
501.0
5010
50100
Gly
co
gen
(
g/1
06 c
ells)
0.0
0.2
0.4
*
*
(*P<0.05 relative to 0 μg ml-1 insulin & 0 nM estradiol)
ICI= Estrogen Receptor antagonist(*P<0.05 relative to cont
Glycogen
CG
lyc
og
en
(
g/1
06 c
ells
)
0.0
0.2
0.4
0.6 *
EstradiolICICont Estradiol+ ICI
1)Endometrial glycogen contributes to histotroph.
localized to epithelial at estrus
mobilized before implantation
high glucose-6-phosphatase during diapause
2) Estradiol stimulates glycogenesis through both paracrine and direct mechanisms.
activation of IGF-1 receptor increases glycogen
estradiol increases glycogen (in presence of insulin)
Estradiol
IGF1
At Estrus
GlycogenStroma Epithelia
Estradiol
IGF1
At Estrus
GlycogenStroma Epithelia
Estradiol
IGF1
Pre-Implantation
GlycogenStroma Epithelia
Estradiol
IGF1
Pre-Implantation
GlycogenStroma Epithelia
Progesterone4-hydroxyestradiol
CommitteeDr. Rose
Dr. Bearden
Dr. Cretekos
Dr. Pfau
Lab MatesJason Hunt
Kole Bowman
Lisa McDougall
Josh Astle
Ayokunle Hodonu
Evan Stegelmeier
Other Grad Students
Other ProfessorsDr. Rodnick
Dr. Aho
Dr. Thomas
Funding SourcesMoyle Mink and Tannery
Fur Commission USA
Idaho INBRE
ISU Research Council
MCRF Seed Grant
Sigma Xi
Thank you