IOGENESI Centro di Medicina della Riproduzione B Laboratory aspects of IVM DAL CANTO MARIABEATRICE...

IOGENESI

Centro di Medicinadella Riproduzione

B

Laboratory aspects of IVMLaboratory aspects of IVM

DAL CANTO MARIABEATRICE

BIOGENESI REPRODUCTIVE MEDICINE CENTER,

ISTITUTI CLINICI ZUCCHI

V. Zucchi, 24-Monza (Italy) tel/fax +39.39.8383314

IOGENESI


B

ANTRAL FOLLICLE 5-12 mmPROPHASE ICell diameter 110-120 µ mMonolayer cumulus cellsZona pellucida Cytoplasmic maturation

Minimum follicular diameter Minimum follicular diameter to produce a competent to produce a competent oocytes is around 5 mm oocytes is around 5 mm

Minimum follicular diameter Minimum follicular diameter to produce a competent to produce a competent oocytes is around 5 mm oocytes is around 5 mm

PRE-ANTRAL FOLLICLE 2-5 mmPROPHASE I Cell diameter 80 µ mMonolayer cumulus cells

IOGENESI


B

GVBD

2n

cAMP

cAMP

cAMP

LH

FF-MASMPF

cAMP

cAMP

OOCYTES OOCYTES MATURATIONMATURATION

PROPHASE IMETAPHASE IIMETAPHASE II

granulosa cell

IOGENESI


B

DAY -1

DAY 0

IVF

DAY 1-2

I.V.M.

I.V.F.

...IVM REQUIRES FLEXIBILITY AND ...IVM REQUIRES FLEXIBILITY AND ADJUSTMENTS ...ADJUSTMENTS ...

IOGENESI


B

TRANSVAGINAL US GUIDANCE CRAFT VACUUM PUMP (pressure 80-90

mmHg)

ASPIRATION NEEDLE (Gynetics 4551E 17-G single lumen)

SINGLE FLASK TO COLLECT OOCYTES

IN VITRO MATURATIONIN VITRO MATURATIONOocytes Collection

IOGENESI


B

IOGENESI


B

DAY –1

Disposable Material

Filter 0.70 μm Syringes Flask or tube Petri dishes

Flushing Medium with Heparin Maturation Medium


IOGENESI


B

DAY -1

IOGENESI


B

IOGENESI


B

DAY -1

IOGENESI


BIOGENESI


B

DAY -1

IOGENESI


B

DAY - 1

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B


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B

PICK UP (8.30 am)

3 hrs INCUBATION IN LAG MEDIUM

(09.00)

26 hrs INCUBATION IN IVM MEDIUM (12.00 am)

DAY –1

r-FSH 0.075 IU/ml r-FSH 0.075 IU/ml hCG 0.10 IU/mlhCG 0.10 IU/ml10% patient’s 10% patient’s serumserum

IN VITRO MATURATIONIN VITRO MATURATIONBiological Aspects

IOGENESI


B

EXPANDED CUMULUS FULL CUMULUS

SPARSE CUMULUS

NUDE

IN VITRO MATURATIONIN VITRO MATURATIONOocytes Classification

IOGENESI


B

DAY -1

EXPANDED CUMULUS Immature oocytes with multilayer and slack cumulus

IOGENESI


B

FULL CUMULUS Immature oocytes with compact complete cumulus

DAY -1

IOGENESI


B

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B

SPARSE CUMULUS Immature oocytes with partial cumulus

DAY -1

IOGENESI


B

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B

NUDE Immature oocytes

without cumulus

DAY -1

ATRETIC

IOGENESI


B

DAY 0

• Maturation control (3.00 pm)

hyaluronidase 80 UI

• ICSI (4.00 pm)


IOGENESI


B

DAY 0

IOGENESI


B

DAY 1

•Fertilization control

(8.30 am)

•Culture in multiwells in ISM-1 Medium


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B

DAY 2

• Embryo control

• Embryo transfer


No cleavage delay was observed

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B

n° %

Cycles started 463

Oocytes retrieved Full cumulus Sparse cumulus Nude + Atretic

Oocyte in culture

18041455207142

1662

80.611.4

8

Maturation rate

Fertilization rate (2PN)

986/1662

671/824

59.3

81.4

RESULTSRESULTSNatural IVMNatural IVM

IOGENESI


B

n° %

n° Embryos transferred

n° Emb. 0% fragmentation

n° Emb. 1-10% fragmentation



n° Emb. >41% fragmentation

671

287

200

78

65

41

42.7

29.8

11.6

9.6

6.3


IOGENESI


B

09.03.2004 A new Law regarding ART was approved

in Italy

09.03.2004 A new Law regarding ART was approved

in Italy...use no more than 3 oocytes for each cycle so as to avoid to produce more than 3 embryos...

...all embryos obtained must be transferred simultaneously with no selection....

3 t

op

ics

... embryo cryopreservation is forbidden....

IOGENESI


B

n° %N° transfersN° patientsN° tot embryosn° embryos/transfer

361289671

1.8 ± 0.6

β-hCG positive test Clinical pregnancies PR/transferPR/patientIR

6054

54/36154/28958/671

14.918.6 8.6


IOGENESI


B

n° Transfers

n° Patients

n° Clinical Pregnancies

PR/transfer (%)

PR/patient(%)

IVM 164 122 25 15.4 20.5

ICSI 455 407 78 17.1 19.2

FIVET

243 220 48 19.8 23.0

Pregnancy rate with standard IVM, ICSI and IVF-ET in women ≤39 age (May 2004-September 2005)

Dal Canto et Al., RBM Online Vol.13, N°2, August 2006

IVM: the first choice in ItalyIVM: the first choice in Italy

IOGENESI


B

RESULTS IN SELECTED WOMENRESULTS IN SELECTED WOMEN

14,7%

19,8%

8,5%

10,8%

0,0%

5,0%

10,0%

15,0%

20,0%

25,0%

PR/transf Implantation Rate

*

*

IOGENESI


B

RESULTS IN SELECTED WOMEN WITH “TOP QUALITY” EMBRYOSRESULTS IN SELECTED WOMEN WITH “TOP QUALITY” EMBRYOS

* analysis performed on women <37 yearsin which only “top embryos” were transferred

14,7%

19,8%

29,1%

8,5%

10,8%

16,1%

0,0%

5,0%

10,0%

15,0%

20,0%

25,0%

30,0%

PR/transfer Implantation Rate

Series1

Series2

Series3

*

*

IOGENESI


B

we are studing… we are studing…

New Maturation Medium

Extended Culture up to blastocyst Stage

In-vitro matured oocyte Cryopreservation

IOGENESI


B

IOGENESI

Centro di Medicinadella RiproduzioneB

Equipe MedicaDr R. FadiniDr M. Mignini RenziniDr R. ComiDr G. RangoniDr A. VillaDr M. MastrolilliDr A. Epis

Equipe BiologicaDr M.B. Dal CantoDr F. BrambillascaDr M. CrippaDr D. FumagalliDr M. LainDr. M. MerolaSig.ra M. MajoliSig.ra A. Longoni

SegreteriaSig.ra C. Del Favero; Sig.ra P. Loreto; Sig.ra F. Rapisarda; Sig.ra F.

Froio

IOGENESI Centro di Medicina della Riproduzione B Laboratory aspects of IVM DAL CANTO MARIABEATRICE...

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