Methods of assisted Methods of assisted reproductionreproduction

MUDr.Jitka ŘezáčováMUDr.Jitka Řezáčová

ÚPMD, Praha 4, PodolíÚPMD, Praha 4, Podolí

When we talk about methods of assisted

reproduction ►All therapeutic procedures and All therapeutic procedures and techniques in which is manipulated techniques in which is manipulated with gametes (oocytes and sperm).with gametes (oocytes and sperm).

History of assisted History of assisted reproduction - IVFreproduction - IVF

►1978 Louise Brown - the first child after 1978 Louise Brown - the first child after IVF-ET (R. Edwards, P. Steptoe, UK)IVF-ET (R. Edwards, P. Steptoe, UK)

►1982 the first child after GIFT from us 1982 the first child after GIFT from us (L.Pilka, Brno)(L.Pilka, Brno)

►1983 the first child after KET1983 the first child after KET►1995 first child after KET in the Czech 1995 first child after KET in the Czech

RepublicRepublic(T.Mardešič, ÚPMD, Prague)(T.Mardešič, ÚPMD, Prague)

►1993 first baby after ICSI (AC van 1993 first baby after ICSI (AC van Steirteghem)Steirteghem)

Indications for treatment of Indications for treatment of infertility by in vitro infertility by in vitro

fertilizationfertilization ►Tubal factorTubal factor►Male subfertilityMale subfertility►EndometriosisEndometriosis►Ovarian factorOvarian factor► Immunological factorImmunological factor► Idiopathic infertilityIdiopathic infertility

Distribution methods of Distribution methods of assisted reproductionassisted reproduction

► IVF-ET - in vitro fertilization and IVF-ET - in vitro fertilization and embryo transferembryo transfer

► ICSI - intracytoplasmic sperm injectionICSI - intracytoplasmic sperm injection►PICSI - positive selection of spermPICSI - positive selection of sperm►AH - assisted hatchingAH - assisted hatching►KET-cryoembryotransferKET-cryoembryotransfer►MESA - microsurgical epididymal MESA - microsurgical epididymal

sperm aspirationsperm aspiration►TESE - testicular sperm extractionTESE - testicular sperm extraction

Recruitment IVF-ET health Recruitment IVF-ET health insurance companies in the insurance companies in the

Czech RepublicCzech Republic ►A maximum of 4 cycles (first two A maximum of 4 cycles (first two

cycles with single embryotransfer)cycles with single embryotransfer)►From 21 to 39 years (absolute tubal From 21 to 39 years (absolute tubal

factor from 18 years) old womenfactor from 18 years) old women►Health insurance companies in the Health insurance companies in the

Czech Republic does not pay: ICSI, Czech Republic does not pay: ICSI, PICSI, assisted hatching, prolonged PICSI, assisted hatching, prolonged cultivation, cryopreservation gamet cultivation, cryopreservation gamet and embryos, KET, MESA,TESEand embryos, KET, MESA,TESE

IVF-ET involvesIVF-ET involves

►Hormonal preparation and monitoring Hormonal preparation and monitoring cycle (controlled ovarian cycle (controlled ovarian hyperstimulation - KOH)hyperstimulation - KOH)

►Collection of oocytesCollection of oocytes► In vitro fertilization and embryo cultureIn vitro fertilization and embryo culture►EmbryotransferEmbryotransfer►Luteal supportLuteal support

Hormonal preparationHormonal preparation

►Choice of stimulation protocolChoice of stimulation protocol►Select the type of gonadotropins and Select the type of gonadotropins and

define the benefitsdefine the benefits

Monitoring cyclesMonitoring cycles

►Monitoring of follicular growth and Monitoring of follicular growth and endometrial thickness (triple line) endometrial thickness (triple line) vaginal ultrasound probevaginal ultrasound probe

►Monitoring blood levels of E2Monitoring blood levels of E2

Timing of ovulationTiming of ovulation

►Applications hCGApplications hCG(Ovitrelle, Pregnyl)(Ovitrelle, Pregnyl)

Collection of oocytesCollection of oocytes

►34-36 h after administration of hCG 34-36 h after administration of hCG transvaginaltransvaginalcollection of oocytes under the US collection of oocytes under the US controlcontrolconscious sedation or general conscious sedation or general anesthesia for short term.anesthesia for short term.

Cultivation of oocytesCultivation of oocytes

►The first 2 hours after egg collection to The first 2 hours after egg collection to the culture box in a 5% CO2 in air at the culture box in a 5% CO2 in air at 37C37C

►Assessment of oocyte maturation and Assessment of oocyte maturation and detect the presence of polar bodiesdetect the presence of polar bodies

►Selected for ICSI oocytes with the first Selected for ICSI oocytes with the first pole element (metaphase II)pole element (metaphase II)

►Other retained 4-6 hours to matureOther retained 4-6 hours to mature

Preparation of spermatozoa Preparation of spermatozoa for in vitro fertilizationfor in vitro fertilization

►Remove seminal plasmaRemove seminal plasmacentrifugation and swim upcentrifugation and swim up

►Capacitation in the culture Capacitation in the culture mediummediumusually 6 husually 6 h

Fertilization of oocytesFertilization of oocytes

►Adding sperm to media oocytes Adding sperm to media oocytes after about 6 hours after aspirationafter about 6 hours after aspiration

►Check for fertilization after 17-22 hCheck for fertilization after 17-22 hdetecting the presence of detecting the presence of pronucleuspronucleus

ICSI

ICSI

ICSI

PICSI (before ICSI)

► It uses a simple chemical reaction. Normal

sperm have the ability to bind to hyaluronic acid.

►Hyaluronic acid is a substance that surrounds the egg and is involved in sperm-egg binding

►This link show only mature sperm, which have significantly lower incidence of chromosomal anomalies.

Recommendation for PICSI

►Low fertilization oocytes after ICSI►Poor quality embryos, respectively

arrest their development►Recurrent spontaneous abortions

EmbryotransferEmbryotransfer

Usually within 48 h after oocyte aspirationUsually within 48 h after oocyte aspiration

Percentage transfer depends on:Percentage transfer depends on:1.1. Quality of transferred embryosQuality of transferred embryos2.2. The number of transferred embryosThe number of transferred embryos3.3. Technic of transferTechnic of transfer4.4. Status of endometrium at the time of Status of endometrium at the time of

transfertransfer

Prolonged cultivationProlonged cultivation

►To the stage of blastocyst-up to To the stage of blastocyst-up to 120 h120 h

CCryoembryotransfer - KETryoembryotransfer - KET

►All embryos good quality can be All embryos good quality can be frozen.frozen.

►KET is thawed embryo transfer into KET is thawed embryo transfer into the uterus either in native cycle or the uterus either in native cycle or hormonal endometrial preparation.hormonal endometrial preparation.

KRYOKONZERVACE

Risks of Assisted Risks of Assisted ReproductionReproduction

►Multiple pregnancyMultiple pregnancy►Ovarian hyperstimulation syndromeOvarian hyperstimulation syndrome

► InfectionInfection►Adnexal torsionAdnexal torsion►HemorrhageHemorrhage

New trends in assisted New trends in assisted reproductionreproduction

►Stimulation with low doses of Stimulation with low doses of gonadotropic hormone, GnRH gonadotropic hormone, GnRH antagonists (prevention of the antagonists (prevention of the development of a higher degree of development of a higher degree of OHSS)OHSS)

►Simple application of drugs, patients can Simple application of drugs, patients can apply them at home (r- hFSH pen)apply them at home (r- hFSH pen)

►Embryo transfer 1-2 embryos (reducing Embryo transfer 1-2 embryos (reducing the number of multiple pregnancies)the number of multiple pregnancies)

Preimplantation genetic Preimplantation genetic diagnosis (PGD)diagnosis (PGD)

►Genetic testing embryo before its Genetic testing embryo before its introduction into the uterusintroduction into the uterus

►Techniques for PGD: FISH, PCR, PCR + Techniques for PGD: FISH, PCR, PCR + DNA -MicroarrayDNA -Microarray

PGDPGD

Indications:Indications: Hereditary diseases pairHereditary diseases pair Birth of a child with a genetic defectBirth of a child with a genetic defect Recurrent miscarriagesRecurrent miscarriages Repeated failure of IVFRepeated failure of IVF Women over the age of 35 years, Women over the age of 35 years,

men 45 years men 45 years

PGD

MESA

TESE

Maintaining the Maintaining the reproduction of cancer reproduction of cancer

patientspatients

►Pharmacological protection of the Pharmacological protection of the gonadsgonads

►Freezing sperm, oocytes, embryos, Freezing sperm, oocytes, embryos, ovarian and testicular tissue prior to ovarian and testicular tissue prior to treatment for cancer patientstreatment for cancer patients

Thank you for your Thank you for your attentionattention