PREVENTS PREMATURE FERTILIZATION OF LATE-MATURING OOCYTES PharmDr . - - PowerPoint PPT Presentation

EGG MATURITY ASSESSMENT PRIOR TO ICSI PREVENTS PREMATURE FERTILIZATION OF LATE-MATURING OOCYTES

• PharmDr. Zuzana Holubcová, Ph.D.

MATURE EGG?

MATURE EGG

DNA (Hoechst) microtubules (a-tubulin)

MI/MII TRANSITION

DNA (Hoechst) microtubules (a-tubulin)

MI/MII TRANSITION

POLARIZED LIGHT MICROSCOPY

• Based on interaction of polarized light with anisotropic materiál.
• Allows quick, non-invasive imaging of birefringent structures in living cells.

Montag, Reproductive Biomedicine Online 2008 Wang et al 2001a,b Moon et al 2003, Montag 2006 Van der Velde 1998 Wang et al 2001c Montag 2006 Hyun et al 2007 Montag 2008 Rama Raju et al 2007 Montag and van der Ven 2008 Kilani et al. 2011 Kilani et al 2014 Tilia et al 2016.

• Used from late 1990´.
• Examination:

1) Meiotic spindle. 2) Zona Pellucida. 3) Sperm head.

• Association between presence of MII spindle

birefringence and oocyte´s developmental competence.

POLARIZED LIGHT MICROSCOPY IN IVF PRACTICE

MATURE EGG spindle Spindle birefringence is orientation-dependent

POLARIZED LIGHT MICROSCOPY EXAMINATION spindle Spindle signal is orientation-dependent

.

time

OPTIMAL TIME OF ICSI

Can the PLM be used to adjust timing of ICSI with respect to the egg´s developmental stage?

CLINICAL STUDY

• Patient´s informed consent.
• Ethical approval.
• Free of charge.
• May 2016 - May 2018:

234 cycles / 916 oocytes

• Poor prognosis cycles:
• Slow/poor responders.

(<6 MII oocytes at OPU)

• 25–48 years, average 36.96± 4.28.
• Severe male infertility factor in 14.96 % of cycles.

DENUDATION

MI or MII

?

time

OPU

early late denudation

MII MI MII MII MII MII MI→MII or MII?

STUDY DESIGN

Hormonal stimulation Polarized light microscopy

~2 hours

Grade A Grade B Grade C Grade D

35 - 36 hours

Oocyte retrieval

+ 3-4 hours

ICSI

Fertilization Cryopreservation Blastulation

• individual culture
• single-step media
• 5% 02, 6% CO2
• slow/poor responders

(<6 MII oocytes) Utilization Biochemical/clinical pregnancy Live birth

INCIDENCE OF SPINDLE GRADES

MATURATION DELAY

Grade D Grade C Grade B Grade A

time

MII spindle present MII spindle absent

Cycle with majority

• f spindle-negative oocytes

Sperm injection (ICSI) postponed

POSTPONING ICSI

Grade D Grade C Grade B Grade A

time

MII spindle present MII spindle absent

59,39 %

98/165

~2 hours

POSTPONING ICSI

PREIMPLANTATION DEVELOPMENT

Chi-squared test, p<0.001 FR…….Fertilization Rate, BR…...Blastulation Rate, UR……Utilization Rate

PREIMPLANTATION DEVELOPMENT

Holubcová et al, J Assist Reprod Genet. 2019

CLINICAL OUTCOME

• 90.74 % transferred embryos

derived from spindle-positive oocytes.

• 47/49 live births from embryo

transfers involving spindle-positive

• ocytes.
• 28.24 % transferred embryos

11 babies from MI/MII oocytes. MI MII

LIMITATIONS

Chromosome status

Presence of the spindle does not warrant successfull pregnancy.

Embryonic genome activation Epigenetics Egg maturity Sperm Mitochondria Endometrium Mitotic errors

CONCLUSIONS

• Spindle signal may appear during in vitro maturation as oocyte progresses through

physiological MI/MII transition during which meiotic spindle reassembles. time

CONCLUSIONS

MII oocytes UR: 30.94 % UR: 45.12 %

„Rescue in vitro maturation“

= Oocyte retrieval MI→MII („late-maturing“ oocytes) UR: 30.94 %

For further details:

Holubcová et al, Journal of Assisted Reproduction and Genetics 2019

ACKNOWLEDGEMENTS

• Embryology:

Drahomíra Kyjovská Darja Páralová Martina Martonová Tereza Klenková

• IVF specialists.
• Nurses.

Aleš Hampl Radka Štěpánová

Reprofit PLM study team

MII MII MII MII MI MI MI

GV

THANK YOU FOR YOUR ATTENTION

GV