Fertilisation

Question 1

What are the requirements for fertilisation?

Answer 1

a sperm - maturation and capactitation
an egg - arrested at metaphase II

Synchronised receptive endometrium

Question 2

How does the ovum enter the fallopian tube?

Answer 2

cumulus oocyte complex is picked up by ciliated fimbriae on the infundibulum at the end of the uterine tube
- action of fimbrae controlled by menstrual cycle

Question 3

What happens to sperm when in the female tract?

Answer 3

decrease rapidly as they move through

• 20-250 million deposited in upper vagina
• seminal plasma, short term buffering, coagulates within minutes (seminogelin) PSA
• Cervical mucus least viscous during days 9-16 of cycle
• pro ovarian contractions of myometrium, sperm pulled towards ovum
• 1000 sperm enter tube via uterotubal junction
• chemotaxis in humans (progesterone)
• cilia line the tubes and move fluid surrounding them to assist sperm movement
• muscular actions of female genital tract and own motility

Question 4

Where does fertilisation normally occur?

Answer 4

ampulla region of uterine tube

Question 5

How long to sperm remain capable of fertilisation?
How long does an egg remain capable of fertilisation?
What does this mean?

Answer 5

• sperm - 5 days within female tract
• egg - 24 hours
• fertilisation can occur max of 5 days after intercourse

Question 6

What are the 5 steps of sperm interaction with egg vestments?

Answer 6

1. remote detected of occyte cumulus complex
2. penetration of cumulus
3. zona binding
4. awesome reaction
5. zona penetration

Question 7

Describe penetration of the cumulus

Answer 7

• approx 3000 cells embedded in gelatinous matrix (huylaronic acid)
• closely apposed cells from tight organised layer
• others less organised
• sperm penetrate and can disperse cumulus (hyaluoronidase enzyme)

Question 8

After penetrating cumulus, sperm bind to zona pellucida

How?

Answer 8

Acrosome reaction permits zona penetration

exposes new membrane for oocyte fusion

Question 9

What is the zona pellucida?

Answer 9

extracellular protein matrix which surrounds all mammalian eggs

• 4 glycoproteins in humans (ZP1, ZP2, ZP3, ZP4)
• important for sperm-egg binding and induction of AR
• persists post fertilisation

Question 10

Describe fusion

Answer 10

• Sperm penetrates the ZP and occupies to perivitelline space
• Equaltorial segment of sperm head fuses with oocyte plasma membrane
• oocyte engulfs the front of the sperm head
• sperm nucleus encased by a vesicle composed of internalised oocyte membrane
• large increase in Ca2+ sweep across egg from point of sperm fusion

Question 11

What are the 2 proteins reponsible for fusion?

Answer 11

Izumo
- sperm membrane receptor for fusion
- detectable on sperm surface only after acrosome reaction
- KO completely abolishes fusion
Juno
- receptor for Izumo on the oocyte plasma membrane
- KO abolishes fusion

Question 12

Describe oocyte activation

Answer 12

• within 1-3 minutes of fusion - large rise in Ca2+ which sweep across egg from point of sperm entry - lasts 2-3 minutes
• followed by Ca2+ oscillations every 3-15 minutes which may last for several hours
• triggered by PLC zeta - sperm specific PLC

Question 13

What are the 2 key effects of Ca2+ increase and oocyte activation?

Answer 13

1. Release of meiotic block
   - maturation promoting factor (MPF) = cdk1 plus cyclin B
   - MPF is established by cytostatic factor (CSF)
   - raised calcium levels surpress CSF activity and destroy cyclin B
   - triggers resumption of cell cycle in the oocyte and complete of meiosis II
2. block to polyspermy
   - fast block = electrical, membrane depolarisation - minutes
   - slow block = cortical reaction , triggered by increased Ca2+

Question 14

What is the cortical reaction?

Answer 14

• cortical granules contain a mixture of enzymes, including several proteases, which diffuse into the ZP following exocytosis from the egg
• induces the zona reaction

Question 15

What is the zona reaction?

Answer 15

• the alteration in the structure of the ZP catalysed by proteases from cortical granules
• cleavage of ZP2 by ovastacin protease
• sperm can no longer bind or penetrate

Question 16

What is meant by the loss of Juno?

Answer 16

• Juno protein is shed from oocyte plasma membrane with the cortical granules
• no further sperm can fuse

Question 17

What does the sperm contribute to the zygote?

Answer 17

• haploid male genomee- sex determination

- centriole - the oocyte has none, forms spindle for first cell division

Question 18

What does the egg contribute to the zygote?

Answer 18

haploid female genome
cytoplasm
all organelles
mitochondria

Question 19

Describe the zygotic/pronucleate (2PN) stage

Answer 19

• decondensation of sperm DNA - protamine/histamine exchange
• male and female pronuclei replicate their DNA
• pronuclei migrate towards each other
• guided by sperm aster - microtubules radiating from centrosome

Question 20

What is syngamy?

Answer 20

• 18-24 hours
• pronuclear membrane break down
• chromatin intermixes
• nuclear envelope reforms around zygote nucleus
• cleavage begins - end of fertilisation/beginning of embryogenesis

Question 21

Describe the transport of the embryo to the uterus

Answer 21

• increased progesterone: oestrogen ratio relaxes musculature of female reproductive tract - isthmic sphincter
• mostly transported under action of cilia

Question 22

Describe the cleavage stages

Answer 22

• zygote cleaves to form two blastomeres
• 4-8 cells stage - PGD/embryo transfer
• morula (16-32 cells) - near end of uterine tube
• no cytoplasmic synthesis so blastomere size decreases wth each division
• ZP still present

Question 23

What controls development of the embryo

Answer 23

> 2 cell stage = dependent on oocyte cytoplasm
4-6 cell stage = major burst of transcription
Many maternally derived proteins persist until blastocyst stage - poor oocyte maturation has big effect
Embryo metabolise, and growth is stimulated by a number of growth factors - both autocrine and paracrine -

Question 24

What is compaction?

Answer 24

• 8-cell stage onwards - inside out polarity starts to develop
• outer cells - trophobalast
• inner cells - inner cell mass
• late morula - fluid absorntio - formation of intercellular junctions between trophoblast cells - Na+K+ATPase

Question 25

Describe blastocyst formation

Answer 25

Day 5
Blastocoel
Discitnon ICM and single layer epithelial trophoblast layer
embryonic and anembryonic pole

Question 26

What is hatching?

Answer 26

Late day 6 onwards
Blastocyst expands out of hole in ZP - anembryonic pole
Implantation

Question 27

What are the risks with monochorionic twins?

What are the risk with mono amniotic twins?

Answer 27

monochronionic  = twin to twin transfusion
monoamniotic = umbilical cord compression

Question 28

Describe the structure of the endometrium

Answer 28

• Underlying muscular myometrial layer
• Upper functional layer = undergoes proliferation then shedding
• Lower basal layer = attached to myometrium; remains intact during menstruation
• Functional layer is resconstriuted out of the underling basal layer
• stromal matrix covered with luminal epithelium
• glandular epithelial extensions penetrate into stroma
• stroma contains a rich supply of blood vessels - the spiral arteries

Question 29

What happens to the endometrium during the proliferative phase?

Answer 29

• after menstruation, endothelium is very thin and consist of only a few lawyers of basal cells
• in the first 14 days of menstrual cycle, endometrial cells proliferate - oestrogen
• thickening due to stromal cell proliferation and stromal oedema
• surface epithelium increases in surface area and metalbolic activity
• increase in number and size of glandular invagination of the stroma
• expression of intracellular progesterone receptor

Question 30

What happens to the endometrium during the luteal phase?

Answer 30

• after ovulation, ovaries begin to produce progesterone
• progesterone stimulates synthesis of secretory material by the glands - rich in glycogen, glycoproteins and AAs - provides nurtrion for blastocyst
• stromal cells becoming larger and plumper
• spiral arteries become fully developed
• cellular secretions are released into the glandular lumen
• must be oestrogen-primed = receptive endometrium

Question 31

What is the window of implantation?

Answer 31

last four days - 20-24

• characterised by the appearance of small elevations at the apical pole of the epithelial endometrium cells - pinopodes
• pinopodes are involved in the absorption of the uterine fluid
• brings the blastocyst nearer to the endometrium
• also immobilises it

Question 32

Describe menstruation

Answer 32

• spiral arteries of the functional layer are hormone sensitive - constrict when the progesterone conc falls
• collapse and necrosis of the functional layer
• blood and necrotic tissue is lost (menses)

Question 33

What is the clinical relevance of all this?

Answer 33

Ectopic Pregnancy

Chromosomal Problems