The process of natural fertilization
Sperms from ejaculate can fertilize freshly ovulated egg
The sperms have to:
• survive in female reproductive tract
• undergo capacitation
• move to oviduct
• displays hyperactivation
• penetrate the egg investments
(cumulus oophorus and zona pelucida)
• displays acrosome reaction – activate
acrosome’s enzymes hialuronidaze and
acrosin
• fuse the oocyte
 deliver the chromatin into egg’s
cytoplasm
 activate the egg to permit
resumption of second meiotic
division – expelling the second
polar body and forming female
pronucleus
 deliver centioles that organise the
formation of first mitotic spindle
 sperm’s chromatin
decondensates, its protamine is
replaced by histone and form male
pronucleus
methylation of genes prevents
their transcription
Sperms deliver epigenetic information required for proper
embryogenesis:
1.
DNA methylation – type of methylation can be altered in ART
2.
sperm specific histones
3.
other chromatin-associated proteins
4.
the perinuclear theca proteins
5.
sperm-born RNA
6.
the DNA loop domain organization by sperm nuclear matrix
ART
Assisted Reproductive Technologies
Strategies of treatment which have been developed to deal with
childlessness exclusively as a symptom without removing the
cause of infertility itself.
In order to assess the real efficacy of different ART techniques, the success rate
should be compared with the cumulative spontaneous conceptions during this
same time interval.
The spontaneous conception rate is the highest during the first three ovulatory
cycles after conception of unprotected intercourses – 20 – 25% per cycle,
after 12 month this rate is about 0.8 – 3.97% per cycle
Methods of Assisted Fertilization
 Insemination
 In Vitro Fertilization (IVF)
 Intracytoplasmic Sperm Injection (ICSI)
Insemination
Sperms are transferred into the female partner reproductive system by various
methods. Insemination aims at increasing the concentration of motile spermatozoa
in the vicinity of the oocyte within the Fallopian tube around the time of
ovulation.
AIH – Artificial Insemination With Husband’s Semen
Indications:
•
anatomical/ functional defects in the vagina, the cervix of uterus, ovulation
defect
•
low semen parameters, disturbances of sperm deposition because of
hypospadiasis, retrograde ejaculation, erectile dysfunction.
AID – Artificial Insemination With Donor’s Semen
Indications (husband’s diseases)
•
azoospermia or aspermia in husband
•
genetic defects carrier
•
HIV, hepatitis carrier
Preparation of semen for IUI
1.
Semen must be liquefied by simple incubation at room temperature for 30
minutes.
2.
Sperms are separated from seminal plasma by centrifugation and swim-up
procedure. The proportion of highly motile and normally formed sperms is
increased and the process of capacitation is induced.
3.
The washing procedure of the semen is required, because seminal plasma
can be toxic both for the sperm itself, as well as for the oocytes. Seminal
plasma contains high concentration of prostaglandins, which, after their
introduction into the uterine cavity, may induce paroxysmal and painful
contractions of the myometrium.
IVF - In Vitro Fertilization
•
The process of fertilization takes place outside the body of the
female in an artificially constructed environment.
•
Oocytes are recovered from preovulatory ovarian follicles and
subsequently coincubated in a culture medium together with motile
spermatozoa.
•
After identification of fertilization, either at an early cleavage stage
or at the later blastocyst stage, the embryos are placed in the
uterine cavity for nidation in the endometrium and pregnancy.
Ovarian Stimulation for IVF and ICSI
The long protocol consists of previous desensitization of the pituitary with
long-acting GnRH-analogues followed by administration of gonadotropins.
The short protocol consists of administration of gonadotropins.
Only in adequately stimulated cycles a sufficient number of oocytes in
metaphase II can be collected.
Indications for IVF
• ejaculatory disturbances, retrograde ejaculation (neuropathy,
diabetes mellitus, surgery)
• low semen parameters
• tubal infertility caused either by occlusion of the Fallopian
tubes or by impairment of their function
The results of conventional IVF in couples suffering from male infertility
have been disappointing, worser than spontaneous conception rate.
Timing of the various step fertilization after ICSI:
• extrusion of second polar body after 2.5 hours
• female pronucleus formation after 5 hours
• male pronucleus formation a few minutes later
• collateral approximation of both pronuclei after
approximately 7 hours
Indications - high diversity of sperm dysfunction:
• severe oligozoospermia due to primary testicular failure
• obstructive azoospermia
• Spermatozoa can be collected surgically from the epididymis (MESA microsurgical epididymal sperm aspiration) or testes (TESE testicular sperm
aspiration)

Longstanding paralysis (sperms collected by electroejaculation).
 Complete lack of sperm motility, due to defects of the axonemal
apparatus in the sperm tail,
 Globozoospermia - spermatozoas without acrosome,
 frozen-thawed sperms
Frequency of ART:
1 in 80 children in USA
1 in 50 born in Sweden
1 in 35 in Australia
1 in 24 born in Denmark are products of ARTs.
Impact of ART on the health and wellbeing of offspring:
increasing:
• birth defects and congenital malformation
imprinting disorders: Beckwith-Wiedman, Angelman disease
• cryptorchidism
• time of staying in a neonatal intensive care
normal
• cognitive and psychosocial development
not known
• future fertility
• incidence of cancer