UNSW Embryology

Week 2 Molecular Development

© Dr Mark Hill (2011)

Acknowledgements

Introduction

The second week of development is marked by continued blastocyst development and the implantation process with rapid growth/differentiation of the extraembryonic tissue (trophoblast). Adplantation (attachment) and implantation is the first physical interaction between developing conceptus and the maternal uterus. The cavity within the blastocyst and the inner cell mass that forms the embryo begin early differentiation. Recent findings in mouse have shown that maternal leukemia inhibitory factor (LIF) has an important role in the implantation process.

(Image source: News and Views - Stewart, CL Nature 450, 619 (29 November 2007) by permission from Macmillan Publishers Ltd (copyright 2007) p53 regulates maternal reproduction through LIF. Nature. 2007 Nov 29;450(7170):721-4.)

Page Links: Introduction | Some Recent Findings | Leukemia Inhibitory Factor | Colony-Stimulating Factor 1 | Transforming Growth Factor-beta | Estrogen | Progesterone | Sal-like 4 | Development Overview | Quick Links | Glossary

Some Recent Findings

Hu W, Feng Z, Teresky AK, Levine AJ. p53 regulates maternal reproduction through LIF. Nature. 2007 Nov 29;450(7170):721-4.

Ralston A, Rossant J. How signaling promotes stem cell survival: trophoblast stem cells and shp2. Dev Cell. 2006 Mar;10(3):275-6.

Yang W, Klaman LD, Chen B, Araki T, Harada H, Thomas SM, George EL, Neel BG. An Shp2/SFK/Ras/Erk Signaling Pathway Controls Trophoblast Stem Cell Survival. Dev Cell. 2006 Mar;10(3):317-27.

Leukemia Inhibitory Factor (LIF)

a At day 4 of pregnancy, oestrogen E2 induces LIF expression in the endometrial glands, leading to LIF secretion into the uterine lumen. There, LIF binds to its receptors on the surface of epithelial cells.	bThis makes the uterus receptive to the blastocyst, which implants by day 5 of pregnancy. Hu et al.2 find that LIF expression in the endometrial glands also depends on the regulatory activity of p53. In the absence of p53, insufficient LIF is produced, the uterus does not become adequately receptive, and fewer blastocysts implant.

(Image source: News and Views - Stewart, CL Nature 450, 619 (29 November 2007) by permission from Macmillan Publishers Ltd (copyright 2007)

More? Hu W, Feng Z, Teresky AK, Levine AJ. p53 regulates maternal reproduction through LIF. Nature. 2007 Nov 29;450(7170):721-4.)

Leukemia Inhibitory Factor also called in the literature cholinergic differentiation factor.

Search PubMed: term = Leukemia Inhibitory Factor implantation

Colony-Stimulating Factor 1

Search PubMed: term = Colony-Stimulating Factor 1 implantation

Transforming Growth Factor-beta

Search PubMed: term = Transforming Growth Factor-beta implantation

Estrogen

Search PubMed: term = Estrogen blastocyst implantation

Progesterone

Maternal rogesterone has an important role in uterine epithelium preparation for blastocyst implantation.

Search PubMed: term = Progesterone blastocyst implantation

Sal-like 4 (Sall4)

The region specific homeotic gene spalt (sal) of Drosophila gives the sal-like nomenclature in other species (Sall).

Sall4 also called in the literature SAL-LIKE 4, SALL4, HSAL4.

Inner cell mass and not trophoblast shell is dependent upon Sall4 (Sal-like genes encode putative zinc finger transcription factors) Elling U, Klasen C, Eisenberger T, Anlag K, Treier M. Murine inner cell mass-derived lineages depend on Sall4 function. Proc Natl Acad Sci U S A. 2006 Oct 23

"Sall4 is cell-autonomously required for the development of the epiblast and primitive endoderm from the inner cell mass. Furthermore, no embryonic or extraembryonic endoderm stem cell lines could be established from Sall4-deficient blastocysts. In contrast, neither the development of the trophoblast lineage nor the ability to generate trophoblast cell lines from murine blastocysts was impaired in the absence of Sall4."

Links: Medline | PNAS Abstract | OMIM - SALL4 | Flybase - splat | MBoC image - DNA binding by a zinc finger protein

Week 2 Overview- Implantation and Gastrulation

• Early Development 2
• Covering Week 2 of development
  • Finish formation of the Blastocyst
  • trophoblast, embryoblast
• Implantation
  • normal, ectopic
• Changes in the Uterus
  • Early utero-placental circulation
• Formation of
  • the Bilaminar Embryo
  • the 3 chambered conceptus
  • the Trilaminar Embryo
  • the Axial Process
• Early cell division
  • Cleavage of the Zygote
  • cleavage of zygote forms 2 blastomeres
  • cleavage with no cytoplasm synthesis
  • therefore individual cell volume decreases
  • initially synchronously, then asynchronously
• Formation of the Morula
  • there are now 16-20 cells
  • at the end of the oviduct
  • ball of cells
  • slow- centre cells, larger
  • fast- peripheral cells, flattened against zona pellucida
• Formation of the Blastocyst
  • Day 5
  • zona pellucida lost
  • uterine sercetions
  • breakdown ZP
  • support blastocyst (nutrition)
  • compaction
  • central cells now compacte
  • located on a region of the wall
  • blastoceol formation
  • cavity formed by compaction
  • fluid filled blastoceol
• Blastocyst
  • Trophoblast
    • forms the placenta
    • placental membranes
  • Embryoblast
    • Inner cell mass
    • forms entire embryo
• Early cell division
• Implantation- Trophoblasts
  • Occurs at day 6-7
  • adhesion
  • adplantation
  • implantation complete by about day 9
• Normal
  • uterine wall
  • posterior, lateral
  • cervix, superior
• Abnormal
  • Tubal or Ectopic- not in uterus
• Abnormal Implantation
  • often occurs if zona pellucida is lost too early allowing premature implantion
  • Tubal pregnancy 94%
  • embryo may develop through early stages.
  • can erode through the uterine horn
  • reattach within the peritoneal cavity
• Ectopic Pregnancy
  • external surface of uterus
  • bowel
  • gastrointestinal tract, mesentry
  • peritoneal wall
  • If not spontaneous then, embryo has to be removed surgically
• Normal Implantation- Endometrium
  • consists of 3 layers in secretatory phase of menstral cycle
    • compact
      • implantation odccurs in this layer
      • dense stromal cells, uterine gland necks, capillaries of spiral arteries
    • spongy
      • swollen stromal cells, uterine gland bodies, spiral arteries
    • basal
      • not lost during menstruation or childbirth, own blood supply
  • Myometrium
    • muscular layer outside endometrium
    • contracts in parturition
• Uterine Changes
  • Endometrium
    • decidual reaction
    • change that occurs in the endometrium at site of implantation
      • loss of epithelium
      • deposition of glycogen
    • spreads throughout uterus
    • not at cervix
    • decidual cells indicative of pregnancy (curette)
    • following reaction endometrium called Decidua
  • Cervix
    • at mouth of uterus
    • secretes mucus
    • forms a plug
  • Vascular
    • increased number of blood vessels
• Decidua
  • forms 3 distinct regions (3 weeks)
    • Decidua Basalis- implantation site
    • Decidua Capsularis- enclosing the conceptus
    • Decidua Parietalis- remainder of uterus
  • Decidua Capsularis and Parietalis fuse- uterine cavity is lost by 12 weeks
• Normal Implantation- Attachment / Adplantation
  • trophoblast cells adhere to uterine endometrium
  • Perlecan, HSPG
  • uterine endometrium expresses binding proteins
  • orients with inner cell mass closest to uterine wall
  • trophoblasts proliferate on attached side
  • uterine LIF required for implantation
  • autocrine / paracrine function
  • proliferating cells loose cell membranes forming syncitiotrophoblasts
  • remainder of trophoblasts- cytotrophoblasts
• Implantation- Day 7
• Implantation- Syncitiotrophoblasts
  • Secrete proteolytic enzymes
  • enzymes break down extracellular matrix around cells
  • Allow passage of Blastocyst into endometrial wall
  • totally surround the blastocyst
  • secrete human chorionic gonadotropin (hCG)
  • presence in urine pregnancy diagnostic
  • maitains decidua and Corpus Luteum
  • generate spaces that fill with maternal blood- lacunae
• coagulation plug
  • left where the blastocyst has entered the uterine wall (day 12)
• Cytotrophoblasts
  • Uteroplacental circulation
  • Anchoring and floating villi
  • Primary Villi
    • cytotrophoblast
  • Secondary Villi
    • cytotrophoblast + extraembryonic mesoderm
  • Tertiary Villi
    • cytotrophoblast + extraembryonic mesoderm+ blood vessels
• initially cover conceptus, then restricted
  • chorion laeve (=smooth)
  • chorion frondosum (=leafy)
• Placenta
  • will cover Embryology and Histology (lecture 13)
  • chorion frondosum and decidua basalis forms the Placenta
  • visible by week 12
  • placenta (= cake)
  • Hydatidiform Mole
  • placenta only, no embryo
  • .5 and 2.5/1000 pregnancies
• Embryoblast- Bilaminar Embryo
  • Inner cell mass
    • before implantation complete forms 2 cell layers
• Bilaminar Embryo
  • Epiblast
    • closest to trophoblasts
    • columnar epithelium
  • Hypoblast
    • closest to blastocoel
    • cuboidal epithelium
    • forms extraembryonic mesoderm
• Bilaminar Embryo-Epiblast
  • cavity forms between cytotrophoblasts and Epiblast
  • cells from epiblast migrate and form wall of cavity
  • Amnionic sac
  • will eventually cover entire embryo
• Implantation Day 8
• Bilaminar Embryo-Hypoblast
  • cells from hypoblast migrate around the blastocoel
  • these cells become extraembryonic mesoderm and line cavity
  • form Primary Yolk sac
• Implantation Day 9
• The 3 Chamber Conceptus
• Extraembryonic mesoderm
  • innermost cell layer thin and line Primary Yolk sac
  • Heuser's membrane
  • outer mesoblasts proliferate, thick
  • surround the amniotic cavity
  • develop spaces around Primary yolk sac
    • spaces fuse to form single cavity
  • extraembryonic coelom (cavity)
  • mesoderm is split by cavity
    • Chorion
      • outer part together with cytotrophoblasts
    • chorionic cavity is 3rd chamber
  • Connecting stalk
    • inner part covers amnion and Primary Yolk sac
• Implantation Day 15
• Implantation Day 16
• Trilaminar Embryo
• Gastrulation
  • process of cell migration from epiblast (day 15)
  • cells migrate through Primitive streak
  • Primitive streak consists of
    • primitive groove
    • primitive pit
    • primitive node
  • Primitive streak defines body axis
    • rostrocaudal (head tail)
    • Left Right
• Primitive Streak- Membranes Day 14
• Trilaminar Embryo-Gastrulation
  • Early cells migrate form Endoderm
    • lie adjacent to epiblast
    • line Primary Yolk sac to form Yolk Sac
    • "Mark's left hand model"
  • Later cells migrate for Mesoderm
    • intraembryonic mesoderm
    • migrate laterally, rostrally, caudally
    • continues until week 4
  • ECM adhesive pathways
• Germ Cell Layers
  • Ectoderm
    • nervous system, epidermis epithelia
  • Mesoderm
    • muscle, skeleton, connective tissue
  • Endoderm
    • gastrointestinal tract epithelia, GIT organs
• Ectoderm/Endoderm Membranes

  • Rostrocaudal axis
  • ectoderm and endoderm without mesoderm
  • form ends of gut tube

• Buccopharyngeal membrane
  • Rostral (head)
  • breaks down to form oral cavity
• Cloacal membrane
  • Caudal (tail)
  • breaks down to form anus, urinary and genital openings
• Axial Process
  • begins at the primitive node
  • hollow tube extends rostrally, beneath the ectoderm
  • stops at the buccopharyngeal membrane
  • notochordal process and endoderm fuse- notochordal plate
  • canal opens between amnion and yolk sac- neurenteric canal
  • notochordal plate and endoderm interact
  • cells rise into mesoderm as a solid rod- notochord
• Trilaminar Embryo- Axial Process
• Sex Determination
  • Please Note- This should have been included in Lecture 1
  • based upon whether an X or Y carrying sperm has fertilized the egg
  • should be 1.0 sex ratio
    • is actually 1.05, 105 males for every 100 females.
  • some studies show more males 2+ days after ovulation
  • cell totipotential
  • Men- Y Chromosome
    • carries sry gene
  • gene activates pathway for male gonad
  • Women- X Chromosome
    • one X chromosome in each cell has to be inactivated.

References

This reference section (version 2.2) requires updating.

• Selected Lists of References from PubMed March 1999 search results are available for School of Anatomy computers without internet access. Computers with internet access can search from either Page 2 or PubMed Internet Access
  • Selected Research Articles and Reviews

Recent Implantation Reviews

• Rinkenberger JL, et al.  Molecular genetics of implantation in the mouse. Dev Genet. 1997;21(1):6-20. Review.
• Burrows TD, et al.  Trophoblast migration during human placental implantation. Hum Reprod Update. 1996 Jul-Aug;2(4):307-21. Review.
• Schultz GA, et al.  Biology and genetics of implantation. Dev Genet. 1997;21(1):1-5. Review. No abstract available.
• Coutifaris C, et al. Integrins, endometrial maturation, & human embryo implantation. Semin Reprod Endocrinol. 1998;16(3):219-29. Review.
• Rogers PA. Current studies on human implantation: a brief overview. Reprod Fertil Dev. 1995;7(6):1395-9. Review.
• Critchley HO. Factors of importance for implantation and problems after treatment for childhood cancer. Med Pediatr Oncol. 1999 Jul;33(1):9-14. Review.  
• Duc-Goiran P, et al.   Embryo-maternal interactions at the implantation site: a delicate equilibrium. Eur J Obstet Gynecol Reprod Biol. 1999 Mar;83(1):85-100. Review.
• Salamonsen LA.  Role of proteases in implantation. Rev Reprod. 1999 Jan;4(1):11-22. Review.
• Tabibzadeh S.  Molecular control of the implantation window. Hum Reprod Update. 1998 Sep-Oct;4(5):465-71. Review.
• Rice A, et al. Cytokines in implantation. Cytokine Growth Factor Rev. 1998 Sep-Dec;9(3-4):287-96. Review.  
• Carson DD, et al.  Mucin and proteoglycan functions in embryo implantation. Bioessays. 1998 Jul;20(7):577-83. Review.
• Ghosh D, et al. Recent developments in endocrinology and paracrinology of blastocyst implantation in the primate. Hum Reprod Update. 1998 Mar-Apr;4(2):153-68. Review.  
• Fein A, et al.  Peri-implantation mouse embryos: an in vitro assay for assessing serum-associated embryotoxicity in women with reproductive disorders. Reprod Toxicol. 1998 Mar-Apr;12(2):155-9. Review. No abstract available.
• MacCalman CD, et al.  Type 2 cadherins in the human endometrium and placenta: their putative roles in human implantation and placentation. Am J Reprod Immunol. 1998 Feb;39(2):96-107. Review.
• Klentzeris LD.  The role of endometrium in implantation. Hum Reprod. 1997 Nov;12(11 Suppl):170-5. Review.
•  Aplin JD.  Adhesion molecules in implantation. Rev Reprod. 1997 May;2(2):84-93. Review.
• Lockwood CJ, et al.  Decidual cell regulation of hemostasis during implantation and menstruation. Ann N Y Acad Sci. 1997 Sep 26;828:188-93. Review.  
• Tabibzadeh S Implantation: from basics to the clinic. Ann N Y Acad Sci. 1997 Sep 26;828:131-6. Review. No abstract available.
• Sueoka K, et al.  Integrins and reproductive physiology: expression and modulation in fertilization, embryogenesis, and implantation. Fertil Steril. 1997 May;67(5):799-811. Review.
• Nie GY, et al.  Hormonal and non-hormonal agents at implantation as targets for contraception. Reprod Fertil Dev. 1997;9(1):65-76. Review.  
• Barkai U, et al.  Intrauterine signaling and embryonic implantation. Biol Signals. 1996 Mar-Apr;5(2):111-21. Review.
• Grummer R, et al. Expression pattern of different gap junction connexins is related to embryo implantation. Int J Dev Biol. 1996 Feb;40(1):361-7. Review.
• Loke YW, et al.  Immunology of human implantation: an evolutionary perspective. Hum Reprod. 1996 Feb;11(2):283-6. Review.

Recent Reviews Gastrulation

• Tam PP, et al. Mouse gastrulation: the formation of a mammalian body plan. Mech Dev. 1997 Nov;68(1-2):3-25. Review.
  • a key Australian researcher at the Childrens Medical Research Institute, Westmead
• Lemaire L, et al. Gastrulation and homeobox genes in chick embryos. Mech Dev. 1997 Sep;67(1):3-16. Review.
• Arendt D, et al. Dorsal or ventral: similarities in fate maps and gastrulation patterns in annelids, arthropods and chordates. Mech Dev. 1997 Jan;61(1-2):7-21. Review.
• Camus A, et al. The organizer of the gastrulating mouse embryo. Curr Top Dev Biol. 1999;45:117-53. Review.
• Boncinelli E, et al. Homeobox genes in vertebrate gastrulation. Curr Opin Genet Dev. 1995 Oct;5(5):619-27. Review.
• Wolpert L. Gastrulation and the evolution of development. Dev Suppl. 1992;:7-13.
• McClay DR. Gastrulation. Curr Opin Genet Dev. 1991 Aug;1(2):191-5. Review.

Recent Articles Gastrulation

• Tam PP, et al.The allocation of epiblast cells to the embryonic heart and other mesodermal lineages: the role of ingression and tissue movement during gastrulation. Development. 1997 May;124(9):1631-42.
• Wood HB, et al. Comparative expression of the mouse Sox1, Sox2 and Sox3 genes from pre-gastrulation to early somite stages. Mech Dev. 1999 Aug 1;86(1-2):197-201
• Winklbauer R, et al. Vegetal rotation, a new gastrulation movement involved in the internalization of the mesoderm and endoderm in Xenopus. Development. 1999;126(16):3703-3713.
• Gu Z, et al. The type I serine/threonine kinase receptor ActRIA (ALK2) is required for gastrulation of the mouse embryo. Development. 1999 Jun;126(11):2551-61.
• Saxton TM, et al. Morphogenetic movements at gastrulation require the SH2 tyrosine phosphatase Shp2. Proc Natl Acad Sci U S A. 1999 Mar 30;96(7):3790-5.
• Sugihara K, et al. Rac1 is required for the formation of three germ layers during gastrulation. Oncogene. 1998 Dec 31;17(26):3427-33.
• Zwijsen A, et al. Ectopic expression of the transforming growth factor beta type II receptor disrupts mesoderm organisation during mouse gastrulation. Dev Dyn. 1999 Feb;214(2):141-51.
• Varlet I, et al. nodal expression in the primitive endoderm is required for specification of the anterior axis during mouse gastrulation. Development. 1997 Mar;124(5):1033-44.
• Sanders EJ, et al. Patterns of cell death during gastrulation in chick and mouse embryos. Anat Embryol (Berl). 1997 Feb;195(2):147-54.
• Winklbauer R, et al. Fibronectin, mesoderm migration, and gastrulation in Xenopus. Dev Biol. 1996 Aug 1;177(2):413-26.
• Bellomo D, et al. Cell proliferation in mammalian gastrulation: the ventral node and notochord are relatively quiescent. Dev Dyn. 1996 Apr;205(4):471-85.
• Malinda KM, et al. Four-dimensional microscopic analysis of the filopodial behavior of primary mesenchyme cells during gastrulation in the sea urchin embryo. Dev Biol. 1995 Dec;172(2):552-66.
• Sulik KK. Critical periods for alcohol teratogenesis in mice, with special reference to the gastrulation stage of embryogenesis. Ciba Found Symp. 1984;105:124-41.

Recent Articles Notochord

• Prince VE, et al. Hox gene expression reveals regionalization along the anteroposterior axis of the zebrafish notochord. Dev Genes Evol. 1998 Nov;208(9):517-22.
• Teillet MA, et al. The relationships between notochord and floor plate in vertebrate development revisited. Proc Natl Acad Sci U S A. 1998 Sep 29;95(20):11733-8.

Glossary of Terms

A | B | C | D | E | F | G | H | I | J | K | L | M | N | O | P | Q | R | S | T | U | V | W | X | Y | Z | Numbers | Old Glossary

Quick Links

Week 2 Pages:

Introduction | Timeline Week 2 | Carnegie stages 5 to 7 | Abnormalities | Blastocyst Implantation | Sites of Implantation | Bilaminar Embryo | Embryonic Cavitites | Early Placentation | Molecular | Stem Cells | References | Text only | WWW Links | Movies - Week 2

Development Notes ----------------- Menstrual Cycle Week 1 Week 2 Week 3 Week 4 Placenta Serial Images Molecular Dev Abnormal Dev Postnatal Dev Notes Index

System Notes ----------------- All Systems Heart/Cardiovascular Respiratory Coelomic cavity Gastrointestinal Tract Head and Neck Neural Neural Crest Eye Ear Nose Endocrine Renal Genital Musculoskeletal Integumentary

Movies ----------------- Movies Quicktime Flash Audio Week 1 Week 2 Week 3 Stage 13 Stage 22 Neural Heart Gastrointestinal Renal Genital Other Embryos

Other Embryos ----------------- All Animals Chicken Fly Frog Mouse Rabbit Rat Worm Zebrafish Guinea Pig

UNSW Embryology ISBN: 978 0 7334 2609 4

UNSW CRICOS Provider Code No. 00098G