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CHOOZING THE BEST EMBRYO FOR TRANSFER

CHOOZING THE BEST EMBRYO FOR TRANSFER. Jumana Adham Husseini Senior Embryologist / Lab Director New Hope Gynaecology &Fertility Hospital Sharjah UAE. CHOOZING THE BEST EMBRYO FOR TRANSFER.

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CHOOZING THE BEST EMBRYO FOR TRANSFER

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  1. CHOOZINGTHE BEST EMBRYO FOR TRANSFER .. Jumana Adham Husseini Senior Embryologist / Lab Director New Hope Gynaecology &Fertility Hospital Sharjah UAE

  2. CHOOZING THE BEST EMBRYO FOR TRANSFER • WHAT DETERMINES THE QUALITY OF THE EMBRYO?? THE OOCYTE OR THE SPERM ??AND WHAT NON-INVASIVE METHODS CAN WE CHOOSE TO DETERMINE THE QUALITY OF THE OOCYTES & EMBRYOS ??

  3. Several sperm shown on a morphology slide

  4. The Chromosomal Status Only Partially Correlates With Embryo Morphology Not All Major Chromosomal Aberrations Are Incompatible With Blastocyst Formation

  5. IS SIMPLE VISUALIZATION AN ADEQUATE ASSESSMENT OF OOCYTE QUALITY?? • Cumulus expansion . • PB presence and morphology • Cytoplasmic morphology/ Extra-cytoplasmic abnormalities.

  6. FACTORS AFFECTING OOCYTE GROWTH AND DEVELOPMENT • The underlying pathophysiology of infertility. • The continual process of atresia beginning at the 20th week in utero. • The management of ovulation induction.

  7. None-invasive Subjective Methods Of Oocyte Selection • Using polarized light microscopy for spindle viewing. Metaphase spindle visualization (differential polarized microscopy). • Time lapse video photography. • Measurement of Zona Birefringence using polarized light microscopy. Oocytes with HZB of inner zona layer were found to be more often associated with the best embryonic growth & implantation

  8. Polarized Light Microscopy For Spindle Viewing Visible MS just below PB1 MS slightly dislocated from PB1

  9. MII oocyte observed using Polarized light Microscopy with a clear Birefringent ZP Inner Layer.

  10. Non invasive Objective Selection methods • Color Doppler Ultrasound measurement of blood flow to follicles. Adequate oxygenation. • Measurement of VEGF, HIFα, p53 in follicular fluid (HIFα &VEGF↓) (p53↑)

  11. A New Non-invasive Objective Technique • Study of cumulus cell genes and their role in determining Oocyte quality.

  12. The Cumulus Cells Emerging research have Proven that meiotic or developmental competence of the oocyte is related to a specific pattern of gene expression in cumulus cells.

  13. THE CUMULUS CELLS The cumulus cell transcriptome (genes) may be a valuable predictor of oocyte health, fertilization potential and oocyte / embryo developmental competence.

  14. The Main Aims • To find the best predictor of all three clinical parameters : • Cumulus expansion (oocyte maturity) • Fertilization • Embryo development

  15. IS EMBRYO QUALITY ONLY DEPENDENT ON OOCYTE QUALITY? • IN ORDER FOR THE EMBRYO TO EXIST, FERTILIZATION WHICH IS A COMPLEX PROCESS OF MOLECULAR EVENTS SHOUD OCCUR SUCCESSFULLY • THE FERTILIZATION PROCESS REQUIRES BOTH A MATURE HEALTHY SPERM AND OOCYTE. • BOTH GAMETES SHOULD BE GENETICALLY COMPETENT TO PRODUCE A NORMAL HEALTHY ZYGOTE.

  16. THE ROLE OF THE OOCYTE!!! • THE OOCYTE QUALITY IS DETERMINED BY SEVERAL IMPORTANT FACTORS LIKE • THE QUALITY OF THE CUMULUS AND GRANULOSA CELLS. • CCS EFFECT IS BOTH DETOXYFIYING AND OF EMBRYOTROPHIC NATURE.PROVIDE NUTRIENTS THAT SUPPORT THE METABOLIC NEEDS AND DEVELOPMENT OF THE OOCYTE. • SEVERAL KEY MOLECULES PRODUCED BY THE OOCYTE GDF9 AND BMP15 IMPORTNT ROLE IN GRANULOSA CELL FUNCTION AND DIFFERENTIATION.

  17. Defects in Cumulus Cell Genes • ADAMTS-1 mutations, defects in this gene are responsible for symptoms of an-ovulatory human PCO syndrome. Reduced ovulation rates. • Cox-2(prostaglandin endoperoxidasesynthaze -2) defects leads to impairment of cumulus expansion. Compromised fertilization . Large un-ovulatory follicles were found to be COX-2 negative. • Defects in oocyte derived factor GDF9 affects the expression of Cox-2, causing infertility.

  18. The Oocyte!!! • The oocyte plays a key role in the maturation of its own follicle it is not only a passenger but actively steers development. • There are genes that control the development of the oocyte, its zona and its surrounding environment. • Genes expressed specifically in oocytes are likely to play important roles in folliculogenesis/ oogenesis, fertility and early development

  19. The role of the oocyte in male PN formation • The oocyte develops the capacity to support male pronuclear development during its maturation process at the time of ovulation, • Immature eggs have impaired capacity to help sperm decondense. • As the sperm nucleus is incorporated into the cytoplasm of the mature oocyte, its nuclear envelope rapidly disintegrates thus allowing a variety of substances within the oocyte to gain access to the sperm chromatin.

  20. The Oocyte Cytoplasmic Proteins • The incomplete accumulation of oocyte cytoplasmic proteins such as Histones or Glutathione fertilization failure. • Glutathione is required for sperm decondensation( extremely compact and transcriptionally inactive)  formation of the male pronucleus. • The protamines are replaced by the Histones derived from within the oocyte .

  21. What really happens?? • Different molecular changes occur. • The cleaving of disulphide bonds • Partial enzymatic degradation of nuclear proteins that leads to the release of protamines and their immediate replacement with the Histones derived from the oocyte. • Male pronucleus growth factor(MPGF) Sperm pronucleus development factor( SPDF)Responsible for the chromatin rearrngements and thus decondensation of the sperm nucleus (pronucleus).

  22. The Role of the Sperm!! • The decondensation of the sperm head release of the sperm nuclear contents into the cytoplasm of the egg (oocyte). • The sperm introduce a cytolic component (sperm specific phospholipase) directly into the oocyte cytoplasm to trigger Ca+² oscillation. • The result is the corticlerxnthat results in altering the ZonaPellucida proteins.

  23. THE ROLE OF THE SPERM. • IT SUPPLIES THE CENTROSOME WHICH IS IMPORTANT FOR THE FORMATION OF THE MEIOTIC SPINDLE. • IT SUPPLIES THE ASTER WHICH USES PROTEINS DERIVED FROM THE OOCYTE TO ASSEMBLE THE MICROTUBULE STRUCTURE.

  24. THE PATERNAL EFFECT !!!! • IN HUMANS THE PATERNAL EFFECTS HAVE BEEN SHOWN TO AFFECT EMBRYO CLEAVAGE SPEED AND MORPHOLOGY • CENTROSOME DYSFUNCTION REPRESENTS AN EARLY PATERNAL EFFECT. • LATE PATERNAL EFFECT MAY INVOLVE SPERM ANUEPLOIDY, DNA DAMAGE OR ABNORMAL CHROMATIN PACKAGING which compromises the Morula/ Blastocyst stage► Recurrent abortion

  25. What about Antioxidant treatment??!! • Antioxidant treatment leads to a decreased in DNA fragmentation but has an unexpected negative effect which is an increase in sperm decondensation. (Dr.Yves JR Menezo, Fertility Magazine, vol. 8, 2008.)

  26. THE DEFFERENCE BETWEEN IVF & ICSI • IN IVF  NATURAL SELECTION THE ZONA PELLUCIDA IS ABLE TO IDENTIFY GENETICALLY ALTERED SPERMATOZOA • GENETICALLY ALTERED SPERMATOZOA WITH POOR MOTILITY AND DNA DAMAGE HAVE LOW FITNESS IN OOCYTE FERTILIZATION • IN ICSI THE NATURAL PROCESS OF SPERM /OOCYTE INTERACTION IS BYPASSED.

  27. The Difference Between ICSI and IVF • Although we bypass the initial fertilization events in ICSI yet the incomplete accumulation of oocyte cytoplasmic proteins such as Histones and Glutathione may cause improper processing of sperm components and may result in fertilization failure.

  28. ICSI

  29. IVF ...

  30. Concluding Remarks • We need reliable subjective & objective predictors of oocyte developmental competence, to be able to select oocytes & embryos during IVF/ ICSI

  31. Concluding Remarks • The mammalian oocyte provides its genome to the zygote and accumulates the molecules and information needed to sustain initial divisions in the embryo. • It supplies the factors that will be implicated in regulating the expression from the paternal genome. • The sperm in humans supply the centrosomewhich represents the division center regulating the opposition of the two pronuclei and the plane of the first mitotic division.

  32. Concluding Remarks • Dismorphic oocytes , lacking important proteins and those of high rate anueploidy will either fail to fertilize or produce abnormal embryos. • Abnormal sperm carrying an imperfect centrosome can disrupt mitosis provoking problems at the beginning of embryogenesis (fragments, abnormal chromosome distribution, early cleavage arrest). • Abnormal aneuploid and DNA fragmented sperm can compromise the morula/ Blastocyst transition state which is the next critical stage after genome activation and is the main cause of recurrent abortion.

  33. Concluding Remarks • We can facilitate oocyte/ embryo selection with traditional subjective observational methods, we can also use cumulus expression profiling as another parameter. (objective non-invasive) • Both oocytes and sperms play important roles in dertermining the fate of the embryo.

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