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Assisted Conception Dr. ZEINAB ABOTALIB MRCOG, DGO, Associate Professor & Consultant Obs/Gyna Infertility & IVF Assisted Conception • • • • • • IUI: intrauterine insemination IVF: in vitro fertilization ICSI: intracytoplasmic sperm injection GIFT: gamete intrafallopian transfer ZIFT: zygote intrafallopian transfer PESA: percutaneous epididymal sperm aspiration Assisted Conception • • • • ET: embryo transfer TESE: testicular sperm extraction SUZI: subzonal sperm injection PGD: preimplantation genetic diagnosis Assisted Conception • Objective – To bring sperm and oocyte close to each other to promote chances of fertilization and, ultimately, achieve a pregnancy Assisted Conception • Main types: – IUI: intrauterine insemination – IVF: in vitro fertilization – ICSI: intracytoplasmic sperm injection Assisted Conception • Required procedures – Superovulation – Sperm preparation – Assisted fertilization Superovulation • Hormonal manipulation to enhance ovulation and release multiple oocytes during ovulatory cycle Superovulation • Drugs used: • Human menopausal gonadotropin – Taken from urine of postmenopausal women – Follicle stimulating hormone (FSH) and luteinizing hormone (LH) activity • Recombinant FSH • Recombinant LH Superovulation- protocol • Gonadotropin for 9-11 days • Monitoring follicular development by transvaginal ultrasound • Follicles 16 -18 mm in diameter – 10,000 IU hCG • Oocytes maturation • Ovulation Sperm Preparation • Select PMNS • Remove seminal plasma, WBC, and bacteria • Sperm capacitation – Coating of sperm with seminal plasma proteins – Allow sperm to become fertile – In vivo or in test tube Intrauterine insemination • Sperm sample deposited in uterus just before release of an oocyte (s) in a natural or stimulated cycle • Soft catheter • Give hCG at injection or up to 24 hrs later • Sperm volume: 0.2-0.3 ml • Pregnancy rates – Around 15% per cycle Gamete intrafallopian transfer • Laparoscopic technique in which oocyte and sperm placed in fallopian tube, allowing in vivo fertilization • Procedure – Superovulation – US guided transvaginal oocyte retrieval • 0.1-0.2 mil sperm with 2-3 oocytes In vitro fertilization - IVF • Taking oocytes from woman • Fertilizing them in lab with her partner's sperm • Transferring resulting embryos back to her uterus 3 or 5 days later IVF • Procedure – Superovulation – Insemination – Embryo transfer – Luteal support IVF - Superovulation • • • • Gonadotropin stimulation Monitoring follicular development US guided transvaginal oocyte retrieval Oocyte fertilization with sperm IVF - Insemination • Containers used – Test tubes, Petri dishes, multi-well dishes • Each oocyte inseminated with 0.5-1.0 mil PMNS • Fertilization detected 12-20 hrs later by presence of – 2 pronuclei in oocyte cytoplasm – 2 polar bodies in perivitelline space IVF - Insemination • Syngamy (combination of maternal and paternal pronuclei 24 hrs after insemination • Further cleavages occur at 24 hr intervals IVF - Embryo transfer • Embryos transferred to uterus on 2nd or 3rd day after in vitro insemination • 4-8 cells embryos • 2-3 embryos transferred in 20 µl of culture fluid • Transabdominal US to see fluid placed in uterus • Cryopreserve excess embryos IVF - Luteal support • Progesterone (P4) necessary for pregnancy maintenance • Premature luteolysis in some superovulatory regimens • P4 supplementation until menses occur or woman has positive pregnancy test Intracytoplasmic sperm injection - ICSI • Injection of single sperm into single oocyte in order to get fertilization • Procedure – Superovulation – US guided transvaginal oocyte retrieval – IVF • Oocytes injected with sperm using special microscopes, needles and micromanipulation equipment ICSI - Indications • Low sperm concentration, motility, abnormal morphology • Antisperm antibodies • Fertilization failure after conventional IVF • Ejaculatory disorders • absence of vas deferens or obstruction of ejaculatory ducts Assisted Hatching • Indications – Couples having IVF with • Female partner's age over 37 • Poor quality embryos –Excessive fragmentation –Slow rates of cell division Assisted Hatching – Procedure • Embryo held with a specialized holding pipette • A needle used to expel an acidic solution against ZP • A small hole made in ZP • Embryo washed and put back in culture in incubator • ET shortly after hatching procedure – Hope for the best Further Advances And Uses Of Assisted Conception Technology • Cryopreservation of – Sperm – Embryo – Oocyte – Ovarian tissue • Growth of human follicles and oocytes in vitro • In vitro maturation and transplantation of human spermatozoa Assisted Reproductive Technology (ART) Dr. Abdelsalam Talafha American Board Certified, Comparative Veterinary Obstetrics and Gynecology ART • Infertility – Inability to conceive after 1 year of unprotected and regular sexual intercourse • Primary infertility – Couples have never had children • Secondary infertility – Couples initiated conception in the past and then had difficulty ART • Infertility – Female partner: 35% – Male partner: 35% – Both partners: 20% – Unknown cause: 10% • Infertility more common with increasing age ART • USA women infertility rate – Ages 20-24: 4.1% – Ages 25-29: 5.5% – Ages 30-34: 9.4% – Ages 35-39: 19.7% • 80% of infertility cases can be diagnosed • 85% of cases can be successfully treated ART • Female infertility – Disorders of ovulation: 27% – Fallopian tube disorders: 22% – Pelvic adhesions: 12% – Endometriosis: 5% to 15% – Hyperprolactinemia: 7% ART • Male infertility – Abnormal semen parameters • Count, motility, morphology • Infertility treatment – Correcting underlying abnormality – ART ART • Main techniques – IUI – IVF - embryo transfer – ICSI – Assisted hatching Preimplantation Genetic Diagnosis (PGD) • Identify genetic conditions in embryo before ET – Hemophilia – Cystic fibrosis – Aneuploidy PGD • Hemophilia – Hereditary bleeding disorder – Absence of a blood protein essential for clotting – Types A: lack of factor VIII – Type B: lack of factor IX PGD • Cystic fibrosis – Genetic disease – Defective gene causes body to produce abnormally thick, sticky mucus that obstruct • Lungs • Pancreas PGD • Aneuploidy – Having less than or more than normal diploid number of chromosomes • Monosomy • Trisomy • Triploidy PGD • Performed with IVF • 8-cell stage (3 days old) embryo biopsy • Obtain 1-2 blastomeres for genetic Three day old embryos PGD • Genetic analysis – Multicolor fluorescence in situ hybridization (FISH) – Polymerase chain reaction (PCR) FISH • Detects – Number of chromosomes – Sexing embryos – Sex chromosome aneuploidy – Whole-chromosome paints for detection of rearrangements and identification of marker chromosomes – Analyze polar bodies FISH • Fuorescent probes that bind to specific chromosomes are labeled with biotin for detection by specific fluorochrome-conjugated antibodies under a fluorescent microscope Interphase nucleus X Y Triploid Normal Diploid Tetraploid Missing chromosomes PGD Monosomy Double trisomy
