Fertilization, Morula, Blastocyst, Implantation

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FERTILIZATION

  • It is the process of fusion of two mature germ cells (ovum & spermatozoon) to form a single cell, zygote.
  • Haploid gametes fuse to form a diploid undifferentiated zygote.
  • Site of fertilization is ampulla of uterine tube.

STAGES  OF  FERTILIZATION

  • Approximation of gametes
  • Contact & Fusion of gametes
  • Effects of fertilization

Approximation of Gametes :-

  • It involves transport of sperm and ovum in female genital tract.

Transport of sperm is affected by –

  • Contraction of uterine muscles by prostaglandins of semen & oxytocin from neurohypophysis.
  • Aspiration of sperms into the uterine cavity & uterine tube due to the vacuum generated by uterine contractions.
  • Chemotaxis, by which sperms are attracted towards the ovum.
  • Only 300-500 sperms reach the fallopian tube, which takes 2-7 hours.
  • After ejaculation, sperms are viable for 24-48 hours in the female genital tract.

Capacitation –

  • It is a process of maturation of sperms in female genital tract.
  • While sperms are transported, the coat of glycoproteins on their head is removed. 

Transport of ovum is affected by –

  • Ciliary beats in the fimbriated part of fallopian tube.
  • Rhythmic contractions of uterine tube.
  • Ovum takes 25 minutes to reach ampulla of uterine tube.
  • Ovum is viable for 24-48 hours after ovulation.

Contact and Fusion of Gametes :-

Phases of contact & fusion –

  • Acrosome reaction
  • Disintegration of barriers
  • Calcium wave generation
  • Nuclear fusion

Acrosome reaction –

  • Acrosomal cap fuses with plasma membrane of sperm head & releases enzymes like- hyaluronidase, acrosine, disintegrin.

Disintegration of barriers –

  • Sperm must break through 3 barriers of secondary oocyte for fusion.
  • From outside to inside the barriers of fertilization are-
  • Corona radiata, zona pellucida, vitelline membrane.
  • Acrosomal enzymes help in disintegration of barriers.
  • Hyaluronidase disintegrates corona radiata.
  • Acrosine disintegrates zona pellucida, by binding at ZP3 receptors.
  • Disintegrin peptides of sperm help in fusion of plasma membrane of sperm with vitelline membrane of oocyte.

Calcium wave generation –

  • Once the plasma membrane of sperm fuses with vitelline membrane of oocyte, calcium wave sets in and depolarizes vitelline membrane.
  • Effects of calcium wave generation-
  • Secondary oocyte completes second meiotic division and expels second polar body into the perivitelline space.
  • Release of cortical granules cause hydrolysis of ZP3 receptors on zona pellucida (zona reaction) & induce vitelline block that prevents polyspermy.

Nuclear fusion –

  • Only nucleus and neck of the sperm enter into the cytoplasm of oocyte.
  • Nucleus of sperm and ovum become swollen and known as male and female pronuclei respectively.
  • Both pronuclei have haploid number of chromosomes.
  • This stage of fertilized ovum is called zygote.
  • Zygote undergoes cleavage which is a process of repeated mitotic divisions.
  • Pronuclei DNA replicate, condense to form sister chromatids.
  • Centrioles form mitotic spindle and sister chromatids move towards opposite poles.
  • Finally, cytoplasm divides to form two unequal sized daughter cells with diploid number of chromosomes.

Effects of Fertilization :-

  • Completion of second meiotic division of secondary oocyte & fusion of haploid gametes.
  • Restoration of diploid number of chromosomes in the zygote.
  • Determination of chromosomal sex.
  • Initiation of cleavage.




ASSISTED  REPRODUCTION  TECHNOLOGY (ART)

  • Assisted reproductive technology is used to support the infertile patients.

Methods of Assisted Reproductive Technology :-

  • Counselling for reproduction.
  • Fertility medications that help to form ovarian follicles.
  • In Vitro Fertilization (IVF)- fertilization of ovum outside the female body.
  • Gamete Intrafallopian Transfer (GIFT)- it involves direct transfer of sperms and ovum in fallopian tube.
  • Zygote Intrafallopian Transfer (ZIFT)- it involves the direct transfer of zygote in fallopian tube.
  • Surgical approach- it involves surgical treatment of fallopian tube obstruction and vas deferens obstruction.

IN  VITRO  FERTILIZATION (IVF)

  • It is a process of fertilizing ovum outside the body.

Stages of IVF :-

  • Ovarian stimulation and monitoring
  • Oocyte retrieval
  • Fertilization
  • Embryo development
  • Embryo transfer
  • Luteal phase support

Ovarian Stimulation and Monitoring

  • To induce follicle development and ovulation, injectable hormones are given-
  • Follicle stimulating hormone (FSH)
  • Gonadotropin-releasing hormone (GnRH)
  • Luteinising hormone (LH)
  • In each cycle 5-10 secondary oocytes are collected.
  • Ultrasound examinations are used for monitoring follicle growth in ovaries.
  • Monitoring is required to determine the timing for induction of ovulation.

Oocyte Retrieval

  • Injectable human chorionic gonadotropin or luteinising hormone is given 36-48 hours before egg retrieval.

Methods of egg retrieval-

  • Ultrasound-guided aspiration
  • Laparoscopy assisted aspiration

Fertilization

  • Sperms are collected 2-3 hours before egg collection.
  • Sperm washing is done to remove mucous and nonmotile sperms from semen.
  • Sperms and eggs are incubated at 370C for overnight.
  • Next day eggs are examined microscopically to confirm fertilization.

Embryo Development

  • Fertilized ovum can be grown in a culture media at 370C.
  • In vitro, in culture media zygote starts cleavage.

Embryo Transfer

  • Zygote is transferred to uterus at the stage of blastocyst.
  • Only one or two fertilized eggs are transferred to uterus for implantation.

Luteal Phase Support

  • Collection of ovum results in improper formation of corpus luteum.
  • IVF pregnancy should be supported for first two weeks with progesterone.

 

MORULA

  • Zygote undergoes repeated mitotic divisions within zona pellucida and give rise to blastomeres.
  • Each cleavage division forms one large and one small cell.
  • First cleavage division occurs 30 hours after fertilization to form 2-cell stage.
  • Second cleavage division occurs 40-50 hours after fertilization to form 4-cell stage.
  • Third cleavage division occurs 72 hours after fertilization to form 12-cell stage.
  • Fourth cleavage division occurs 96 hours after fertilization to form 16-cell stage.
  • The 16-cell stage of zygote looks similar to mulberry, called as morula.
  • Morula is covered by zona pellucida.

 




BLASTOCYST

  • Cells of morula continue to divide and form 32 – 64 cell stage.
  • Uterine fluid diffuses through zona pellucida and gets accumulated in intercellular spaces of morula.
  • Fluid filled intercellular spaces increase in size and fuse to form a single cavity called as blastocoel.
  • Blastocoel separates the cells into-
    • Outer cell mass - Trophoblast 
    • Inner cell mass - Embryoblast
  • This stage is called as blastocyst, which is covered by zona pellucida.
  • Side of blastocyst where the embryoblast is attached to trophoblast is known as embryonic pole & the opposite side of blastocyst is known as abembryonic pole.
  • Blastocyst migrates from fallopian tube towards uterine cavity, which is assisted by ciliary beats and contractions of fallopian tube.



IMPLANTATION

  • It is a process of penetration of blastocyst into the uterine endometrium.
  • Implantation begins on 6th or 7th day & completed by 12th day after fertilization.

Site of Implantation –

  • Usually in the posterior wall of upper part of body of uterus.

Process of Implantation –

by 6th day after fertilization zona pellucida disappears and hatching of blastocyst occurs

embryonic pole of blastocyst adheres to the uterine epithelium

trophoblast secretes proteolytic enzymes that erode the endometrium

blastocyst gets buried completely into the endometrium

site of penetration on the endometrial epithelium is closed by a fibrin plug

finally, the fibrin is replaced by proliferation of endometrial lining epithelium.

  • Blastocyst lies in the stratum compactum of endometrium which sheds off during child birth.
  • Hence, the endometrium after implantation of blastocyst called as decidua.
  • Part of decidua where the blastocyst is attached to stratum basale, called as decidua basalis.
  • Part of decidua covering the rest of the blastocyst, called as decidua capsularis.
  • Part of decidua at the junction of decidua basalis & decidua capsularis, called as decidua marginalis.
  • Part of decidua lining the remaining uterine wall, called as decidua parietalis.

Abnormal Implantation –

  • Ectopic implantation results in ectopic pregnancy.

Placenta previa-

  • Implantation occurs in the lower uterine segment.

Tubal implantation-

  • Implantation occurs in uterine tube.

Abdominal implantation-

  • Implantation occurs in the ovary or mesentery.


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