Embryology 3 - Fertilisation and Implantation PDF

The egg is 10 millions times the volume of the sperm cell (waaaaay bigger) The meeting place is in the ampullary region of the Fallopian tube because the folds of the uterine tube are very high, thus slowing down the oocyte and creating an optimal environment for the sperm to meet it For the ferti...

The egg is 10 millions times the volume of the sperm cell (waaaaay bigger) The meeting place is in the ampullary region of the Fallopian tube because the folds of the uterine tube are very high, thus slowing down the oocyte and creating an optimal environment for the sperm to meet it For the fertilisation to happen many changes need to be made PHASE 1 of fertilisation The spermatozoa try to swim among the granulosa cells of the corona radiata. There’s a release of hyaluronidase by the acrosome which breaks down the hyaluronic acid of the sticky matrix and then the spermatozoa swim very fast to the zona pellucida. Sometimes tubal enzymes of the uterine tube can aid the process le granulesproducedbythe oocytetoavoidpoly spermy PHASE 2 of fertilisation The spermatozoa arrive to the zona pellucida, attach to it and penetrate it. They bind to zonal proteins (mainly ZP3) which are species-specific (a human oocyte can’t be fertilised by the spermatozoa of a cat). Then the spermatozoa starts to release enzymes from the acrosome (like acrosin) and digests the membrane (change in calcium concentration, fusion of the head of the spermatozoa to the anterior part of the acrosome thanks to an increase of intracellular pH). Then the spermatozoa penetrate the zona pellucida and they arrive in the perivitelline space (a space between the zona pellucida and the plasma membrane of the oocyte). More than one spermatozoa can penetrate the zona pellucida but ONLY ONE OF THEM WILL ATTACH TO THE PLASMA MEMBRANE OF THE OOCYTE  PHASE 3 of fertilisation The spermatozoon (JUST 1) attaches to the microvilli of the plasma membrane of the oocyte thanks to antigens on its head which bind to the receptors of the oocyte membrane. The plasma membrane of the spermatozoon then fuses with that of the egg and the head, mid piece and tail sink into the cytoplasm of the egg (mitochondria remain out). On the head of the spermatozoa there are antigens called IZUMO while the receptors of the egg membrane are called JUNO (wife of Zeus). When binding takes place the JUNO receptors are removed from the plasma membrane through the use of vesicles to avoid other spermatozoa to attach. After fertilisation the paternal mitochondria are eliminated (either through degradation performed by macrophages or by dilution (they disappear as the zygote divides)). Mutations in mitochondrial DNA can lead to diseases PHASE 4 of fertilisation Prevention of polyspermy —>1. Fast block: in 2-3 O seconds (not so sure in mammals) the membrane depolarises (-70 to +10) to prevent adhesion, this only lasts few minutes 2. Permanent block: changes O in the concentration of Ca++ (waves) cause the release of JUNO, the fusion of cortical granules with the plasma membrane (which causes the cortical granules to release their content in the perivitelline space) and the swelling and hardening of the zona pellucida. This way no other sperm can penetrate the egg PAPER ON THE IMPORTANCE OF CALCIUM Changes in calcium concentration induce the resumption of meiosis, the formation of pronuclei and the prevention of polyspermy. PHASE 5 of fertilisation OOTID —> fertilised oocyte with two pronuclei (of the oocyte and of the spermatozoon) The female nucleus completes meiosis II and a second polar body is formed Remember that meiosis in women only gives life to one haploid cell (the other 3 are not viable) because the cytoplasm needs to be employed to sustain one of the 4 cells. The metabolism of the egg gets activated —> The maternal material stored in the cytoplasm needs to be activated —> mRNA gets recruited and egg respiration and metabolism are increased. In the male nucleus the genetic material gets decondensed (was very tightly packed to allow motility) —> this is due to an increase of permeability in the nuclear membrane, a loss of protoamines, the spreading of chromatin (generating the male pronucleus) and the demethylation of paternal DNA Completion of meiosis and development of pronuclei lead to the formation of the zygote (pronuclei fused together). Now the zygote is ready for cleavage What is accomplished by fertilisation: Oocyte completes second meiotic division Zygote restored to normal diploid number (23 maternal chromosomes and 23 paternal) Determination of the genetic sex (determined by the spermatozoa) Variation of the human species through mingling of paternal and maternal genetic material Metabolic activation of the egg and initiation of cleavage Roles of the zona pellucida: Promotes maturation of oocyte and follicle Barrier that only allows one sperm to enter (usually) Initiates acrosomal reaction Prevents polyspermy Filter during cleavage Immunological barrier between mother and embryo Keeps blastomeres together Facilitates differentiation of trophoblastic cells Prevents premature implantation Cleavage —> series of dividing that increase the number of cells of the zygote within the zona pellucida using the same cytoplasm. The zygote Ymca doesn’t increase in size. With each subdivision the nuclear/cytoplasmic ratio increases. Cleavage begins around 30h after fertilisation. Mammalian cleavage takes days to happen. Laoblastomere Cleavage leads to the formation of BLASTOMERES and 16 blastomeres form a MORULA (develops in 3 days) Compaction —> the blastomeres become very compact and the cell-cell contact is maximised thanks to adhesion molecules like E-cadherin-catenin complexes. When they become too many some of them remain externally (on the side of the zona pellucida) and are called outer blastomeres, while the ones not in contact with the zona pellucida get called inner blastomeres. The outer blastomeres will give rise to the part of the embryo called trophoblast (contribute to the creation of the placenta) while the inner blastomeres will form the inner cell mass (will give rise to the embryoblast and then the embryo) Outer blastomeres are held together thanks to tight junctions while inner blastomeres are held together by gap junctions. The morula rolls around in the Fallopian tubes where some fluids are present. There is a transport of fluids from the Fallopian tube to inside the embryo. This passage is mediated by the zona pellucida. This leads to the formation of a liquid filled cavity —> blastocoele. The trophoblast is now separated by the inner cell mass. The whole thing is now called blastocyst Early communication with mother —> the trophoblast produces an Early pregnancy factor (immunosuppressant protein) to avoid an attack from the mother’s immune system (as it contains the genetic information of the father). Many zygotes are in fact eliminated very early due to the immunoresponse of the mother. The blastocyst is polarised —> there is an embryonic pole (where the inner cell mass is located) and an abembryonic pole (on the opposite side). The trophoblast on the side of the embryo is called polar trophoblast while that on the other side is called mural trophoblast From the inner cell mass the embryoblast will be formed (from which the embryonic tissues will be originated) while from the trophoblast the extra-embryonic tissues will originate (separation from the ECM) CLEAVAGE It takes place in the Fallopian tube At the beginning of cleavage the embryo takes charge of the processes going on. Initially the embryo only depends on the maternal genetic material, but very soon (period of the blastula) maternal material is degraded and the embryo starts producing its own genetic material (transcription of the zygote genetic code). Mosaicism —> during the process of cleavage the subdivision of the chromosomes doesn’t take place correctly —> the mosaic cell line does not become isolated and persists throughout the trophoectoderm and inner cell mass (cells with different chromosome number/ different genetic material continue being duplicated). This can either lead to diseases or the death of the embryo Transport Within 36-48h from fertilisation the early pregnancy factor is produced By 2 days the corona radiata is lost By 3 days the egg is in the ampullary region —> then it takes 8h to reach the isthmus When the blastocyst is in the isthmus the passage is aided by progesterone released by the corpus luteum (which relaxes the uterotubal junction) After 6-7 days the zona pellucida is shed and the blastocyst is released —> hatching of the blastocyst. The zona pellucida NEEDS to be maintained as long as the blastocyst is in the Fallopian tube. Hatching of the blastocyst —> the microvilli from the trophoblast extend to the zona pellucida releasing proteases enzymes —> the zona pellucida is digested thus allowing the embryo to protrude and to attach to the mucosa of the uterus At the end of the previous menstrual cycle the uterine mucosa was prepared for implantation —> the mucous is then a suitable cellular and nutritional environment for implantation and it is an immunopriviledged site. endometrialcycle Three stages of implantation: Apposition —> blastocyst gets close to the endometrium in the places where it is ready Adhesion —> very close contact + adhesion molecules Invasion —> the blastocyst erodes the endometrium and becomes embedded in it APPOSITION It occurs in the implantation window (period of time in which the endometrium is ready to accept the embryo) which lasts 4 days and comes 6 days after the LH surge. In this phase the blastocyst can still be eliminated (if it is close to the endometrium but doesn’t adhere the contractions of the smooth muscle cells can push it out). ADHESION It takes place at the level of the embryonic pole (where the inner cell mass is). This happens thanks to adhesion molecules expressed on the cells of the trophoblast and on those of the endometrium. In some sites of the endometrium the glycocalix becomes thinner, the microvilli disappear, pinopodes appear (small protrusions on the plasma membrane) and a flat surface is created. This aids the process of adhesion There is also a regulation of the immunotolerance as the embryo is a semi-allograft (has half the mother’s genes and half the father’s genes). The endometrium releases cytokines such as LIF (Leukemia inhibiting factors) which bind to the receptors found on the trophoblast. INVASION The role of the trophoblast. The embryo has to erode the endometrium without reaching the underlying muscle layer (myometrium). The trophoblast can be divided into syncytiotrophoblast (fusion of external cells of the trophoblast, erosive) and cytotrophoblast (internal part) The syncytiotrophoblast has lots of erosive properties and is very invasive —> it erodes the endometrium to dig a passage for the blastocyst. The syncytiotrophoblast has contacts with maternal blood vessels and the lacunae in it are filled with maternal blood (it erodes the vessels of the endometrium too). The blood vessels in the lacunae provide the right nutrients to the embryo it surrounds. During this period there might be a light blood loss and so a woman could think it is a slight menstrual period. days doveeopingonaeroding giggygntigtemphobeoststarts 8days ofterosion appearance andfurther heamnioticsac 9days theembryoiscompletelyengulf endocoagueationpergcloses the endometrium formation gtfanae Maintenance of the early pregnancy: Corpus luteum —> produces progesterone (regulates the endometrial functions and the proteolytic activity of the trophoblast) Trophoblast —> produces hCG (human Chorionic Gonadotropin) (can be found in maternal blood by day 8 and in urine by day 10, it maintains the corpus luteum functional) Immuno-tolerance Cells of the mucosa (decidual cells) secrete many interleukins, growth factors and other factors classified as either supporting implantation (pro-invasive) or inhibiting (anti- invasive to contain erosion and avoid its degeneration into muscle cells) enblasto Decidual reaction —> changes in the lamina propria of the endometrium. Fibroblasts in contact with the syncytiotrophoblast become bigger, rounded and epitheliod-like, they get filled with fluids, lipids and glycogen and form a massive ECM surrounding the embryo and later occupying the whole endometrium. This leads to the formation of an immunologically privilege site (leukocytes secrete interleukin-2). The endometrium at this point is called decidua. The decidua is divided into a decidua basalis (on the side where the embryo is implanted), a decidua capsularis (thin mucous layer that lines the amniotic sac) and the decidua parietalis (on the opposite side of the implantation site) The decidual reaction also helps nourishing the embryo at the very early stages. p mucous The normal site of implantation is the posterior wall of the uterine cavity, but sometimes it can happen in different sites (es: cervix —> gives rise to pathological conditions) ECTOPIC PREGNANCY —> implantation takes place in the uterine tube thus rupturing it and causing an haemorrhage. ETP may present with internal bleeding, abdominal uterine bleeding, abdominal/pelvic/low back pain. This can be mistaken for appendicitis if the embryo has been implanted in the right tube. ECT produce β human chorionic gonadotropin at a slower rate so it can lead to a false negative. Transvaginal ultrasonography is very helpful for detecting early tubal pregnancy Risk factors for ECT —> pelvic inflammatory diseases (scar tissue in the uterine tubes which might lead to the formation of pockets), endometriosis (patches of endometrial tissue in abnormal places like uterine tubes), smoking (affects the motile cilia of the uterine tubes) SECOND WEEK - development of the bilaminar embryo and full implantation The inner mass rearranges into an epithelium called embryonic shield (two layers). The two layers are called epiblast (above) and hypoblast or primitive endoderm (below). The differentiation is given by transcription factors —> epiblast (transcription factor nanog) hypoblast (transcription factor Gata 6) Thanks to the formation of the embryonic shield the primitive dorso-ventral axis is established. The epiblast is dorsal while the hypoblast is ventral Derivatives of the epiblast and hypoblast ⬇ ️ Alsocalled membrane Heuser's Formation of the amniotic cavity and yolk sack The amniotic cavity is formed in the epiblast and then a layer of cell covering the cavity (amniotic membrane) originates. The cells of the hypoblast proliferate along the inside of the trophoblast (cytotrophoblast) forming the parietal endoderm (or Heuser’s membrane). The parietal endoderm encloses a cavity called yolk sac (or vitelline sac) eaegenerates andsepera From the parietal endoderm some cells change and form the extraembryonic mesoderm, which supports the epithelium of the amnion, the yolk sac, the chorionic villi and blood vessels. Epiblast —> dorsal —> amniotic sac is dorsal Hypoblast —> ventral —> vitelline sac is ventral The yolk sac is not useful for nutrients to humans because the embryo develops inside of the mother’s body and gets nutrients from her capillaries (only useful in oviparous animals) The outer wall of the blastocyst (extraembryonic mesoderm and cytotrophoblast) is called Chorion. Inside of the chorion some cavities start to form and eventually they get together to form a bigger cavity —> extraembryonic coelom. At this point the mesoderm starts surrounding the amniotic sac too. pinkgetsaround theamnioticsac Extraembryonic coelom (or chorionic cavity) — > cavity OUTSIDE of the embryo Definitive yolk sac and body stalk —> by the 13th day the vitelline sac becomes much smaller. The yolk sac and the amniotic sack are gettogether connected to the mesoderm through the body form I stalk and they are suspended in the chorionic cavity. The body stalk is the first place where blood vessels will be formed. amnionic cavity Formation of primitive blood vessels in the extraembryonic mesoderm followed by the body stalk —> before this the only blood present is the maternal one in the lacunae Development of the uteroplacental circulation —>By the end of the second week the cytotrophoblast extends in the syncytiotrophoblast forming a primary villus —> the extraembryonic mesoderm then protrudes in the core of the primary villus (thus forming the secondary villus) —> some capillaries are formed in the extraembryonic mesoderm and then connected with the circulation of the embryo (tertiary villus, whose walls are very thin to allow exchanges). Through this the embryo can exchange materials with the maternal blood stored in the lacunae. The fetus can now acquire oxygen and release carbon dioxide in the maternal blood. It can happen that some red blood cells end up in the mother’s vessels —> can be a problem if the mother’s blood isn’t compatible with that of the embryo It can also happen that something from the mother’s blood gets in the fetus’ blood —> es: medications, excess glucose, alcohol… Spontaneous abortion (SA) or miscarriage (pregnancy loss that occurs naturally before the 20th week of gestation): Most common during the 3rd week 25-30% occur in the first 12 weeks Reported rate is 50-70% (a woman might not even know) A SA happening several days after the missed period might be mistaken as a late period More than 50% of SA occur as a result of chromosomal abnormalities Other causes —> failure of blastocyst implantation (es: no adhesion molecules) Higher incidence of SA for fetus with neural tube defects, cleft lip (labbro leporino) and cleft palate After 10th week of gestation it might happen due to fetal causes, placental causes or maternal causes