Evidence that absence of endometrial gland secretions in uterine gland knockout ewes compromises conceptus survival and elongationC.A. Gray, R.C. Burghardt, G.A. Johnson, F.W. Bazer and T.E. Spencer
Team OneCarolyn Allen Donna CovingtonAzure Faucette Megan Minten
Morphology and Histology of the Mature Ovine Uterus
Functions: Sperm transport, luteolysis and control of cyclicity, environment for preattachment embryo, maternal contribution to the placenta, expulsion of the fetus and fetal membranes
Structure:• Bicornuate (two uterine horns and small uterine body) • Layers Perimetrium (Serosa) Myometrium (Muscularis) Endometrium (Mucosa and Submucosa)
Target for:• Oxytocin, Estradiol, Progesterone
Surface molecules:• Non-adhesive factors (e.g. Muc-1), adhesion molecules (e.g. Integrins)
Senger P.L., Pathways to Parturition
Morphology of the Mature Uterus
• Continuous with the mesometrium of the broad ligament
• Consists of endothelial cells and connective tissue
• Consists of 2 layers of smooth muscle
1) Longitudinal layer (outer) 2) Circular layer (inner)
•Provides motility, is responsible for uterine tone when estradiol is high, and aids in expulsion of fetus and fetal membranes at parturition
Perimetrium (Serosa) Myometrium (Muscularis)
Outer layerInner layer
MyometriumPerimetrium
Morphology of the Mature Uterus
Endometrium (Mucosa + Submucosa)• Stroma: Stratum compactum (dense stroma) Stratum spongiosum (less dense stroma)•Luminal Epithelium (LE)•Glands/Glandular Epithelium (GE)•Lymphatics, blood vessels
EndometriumStratum Compactum
Stratum Spongiosum
Luminal Epithelium
Glands/Glandular Epithelium
Endometrium
Myometrium
Histology of the Ovine Uterus
Endometrium
Histology of the Ovine Cyclic and d14 Pregnant Uterus: Intercaruncular and Caruncular Areas
Caruncular Area• Devoid of glands• Vascularized• Maternal side of palcentome
Intercaruncular Area• Vast number of glands
d14 Pregnant
CyclicL
Gss
sc
M
ss
sc
Gray et al., 2002
Endometrial adenogenesis in the uterine wall of most species
Gray C A et al. Biol Reprod 2001;65:1311-1323
©2001 by Society for the Study of Reproduction
Comparative time line of the seminal events of endometrial gland morphogenesis in the
sheep, pig, rodent, and human
Gray C A et al. Biol Reprod 2001;65:1311-1323©2001 by Society for the Study of Reproduction
Spencer, TE et al. Endocrinology 1999,Vol. 140, No. 9:0013-7227
•L – Lumenal Epithelium•G – Glandular Epithelium
•SC – Stratum compactum•SS – Stratum spongiosum
Interactive mechanisms involved in endometrial adenogenesis
Gray C A al. Biol Reprod 2001;65:1311-1323et
©2001 by Society for the Study of Reproduction
Spencer, TE et al. Endocrinology 1999,Vol. 140, No. 9:0013-7227
•L – Lumenal Epithelium
•G – Glandular Epithelium
•SC – Stratum compactum
•SS – Stratum spongiosum
•V – blood vessel
•M - Myometrium
Ruffled L
Epigenetic Effects of Progesterone on Uterine Phenotype
•Progesterone:
•C-21 steroid hormone•Important hormone for the estrous cycle and menstrual cycle (human)•required to maintain pregnancy•embryogenesis
Progesterone
Neonatal development
• Dramatic remodeling occurs-1st few months- “critical period”• Glands emerge from internodular, luminal epithelium during
fetal development• Introduction of progesterone has been shown to have a
lasting effect on tissue form and function• Can cause permanent changes to the organization of the
target tissue• Dependent upon:
– Steroid involvement– Dosage– Duration of exposure
Epigenetic Effects of Progesterone
• In the neonatal ovine uterus, key growth factors regulate morphogenesis:– FGF-10: stimulates proliferation and differentiation of the placenta– FGF-7: mediates the development of uterine blood vessels– HGF: stimulate epithelial morphogenesis (establishment of
pregnancy, conceptus implantation and placentation).
• Progestin: inhibits the expression of both the growth factors and their receptors.
• In addition, progestins disrupt the expression of ER (ER is expressed in emerging, proliferating and developing glandular epithelium and stroma cells.
Estrous Cycle
ProestrusEstrus
MetestrusDiestrus Estrus
5 10 15 20
E2
P4
FSH
LH
Rel
ativ
e B
loo
d H
orm
on
e C
on
cen
trat
ion
s
Estrus: E2 peakLH surge induces ovulationFSH suppressed surge by inhibin and E2
P4 minimal
Metestrus:P4 increasingE2 dropsLH pulsatileFSH low
Diestrus:P4 increases and plateaus, PGF released and P4 starts decliningE2 minimalLH pulsatileFSH low
Proestrus:E2 risesP4 dropsLH increasesFSH increases
PGF
Day of Estrous (relative to estrus)Senger P.L., Pathways to Parturition
Estrous Cycle
• Gray et al., 2000 demonstrated UGKO ewes had variable interestrous interval differing from control ewes (16-18 day) ranging from 17-43 days– No difference between UGKO and control ewes in response to PGF
injection
Pregnancy in UGKO Ewe
• Peri-implantation pregnancy loss was observed in the UGKO ewe model Gray et al., 2000; Gray, Bazer et al., 2001; Gray, Taylor et al., 2001
– Normal, hatched blastocysts found in uterine flushings at day 6 and 9
• Gray et al., 2002 hypothesized that the peri-implantation defect in UGKO ewes was due to an absence of endometrial glands and secretory products. Alternatively the defect could be the result from deficient expression of LE adhesion molecules or inability of endometrium to respond to IFN-– Determine differences between control and UGKO ewes in:
• Distribution of Muc-1 and integrin subunit• Abundance of osteopontin and GlyCAM-1• Endometrial expression of IFN--stimulated genes
Pregnancy in UGKO Ewe
• UGKO ewes were found to have:– no conceptus (n = 5/12); growth-retarded, tubular conceptus (n =
6/12); or fragmented, filamentous conceptus (n = 1/12)– No difference in expression of Muc-1 and integrins αv, α5, β3 and β5
– IFN stimulated genes expressed– Osteopontin was absent in uterine flushes, GlyCAM-1 was low or
undetectable
• CONCLUSIONS:– Peri-implantation conceptus survival and growth is altered by the
absence of endometrial glands and their secretions, including molecules involved in cell-cell adhesions such as osteopontin and GlyCAM-1 Gray et al., 2002
Evidence that absence of endometrial gland secretions in uterine gland knockout ewes compromises conceptus survival and elongationC.A. Gray, R.C. Burghardt, G.A. Johnson, F.W. Bazer and T.E. Spencer
Team OneCarolyn Allen Donna CovingtonAzure Faucette Megan Minten
Questions