Thomas Ebner_Zona pellucida imaging.ppt [Uyumluluk Modu]

ZONA PELLUCIDA IMAGING
P.O. Number Terms Rep Ship Via F.O.B. Project
as a predictive factor in IVF laboratories
Quantity Item Code Description Price Each Amount
Thomas Ebner
Kinderwunsch Klinik Linz
AUSTRIA
Total
IX. Turkish-German Gynecology Congress
Antalya
4.-8. May, 2011

FOLLICULOGENESIS
primordial follicle
primary follicle
secondary follicle
secondary follicle
tertiary follicle

SECONDARY FOLLICLE

3D-STRUCTURE
ZP1 185-200 kDa
ZP2 120-140 kDa
ZP3 83 kDa
Zona pellucida
ZP3
Wassarman, 1989

COMMUNICATION through ZONA
Sutton et al., 2003

ZONA THICKNESS
… increases during MATURATION
and
… decreases after FERTILIZATION
ZP1 gene
½ THIN ZONA
ZP2 and 3 gene
No ZONA
-sterile-

HYPOTHESIS
In such oocytes with zona defects,
the patterning or the secretion of the glycoprotein could have been interrupted
during the formation of the extracellular coat . . .
In contrast,
normal zona appearance might reflect optimal growth and nutrition of the egg
and, thus, could be related to oocyte developmental competence . . .
oocyte
oocyte/ granulosa cells
granulosa cells

HOW to QUANTIFY
Shen et al., 2005
• Polarization microscopy allows for visualization of highly ordered structures,
such as spindle and zona pellucida
• Zona pellucida, e.g., shows a distinct laminar architecture consisting of an
inner (bright), a middle (black) and an outer layer (grey)
– THICKNESS of the 3 layers
– BIREFRINGENCE / RETARDATION

FIRST EVIDENCE
Shen et al., 2005
Inner layer is the thickest one and shows the highest retardance

VARIATION
Shen et al., 2005
17%
8.5% (±8.5)
maximum difference
average
45%
16.5% (±14.5)

ZONA and PREGNANCY
Shen et al., 2005
2.8±0.6
11.3±1.4
0.4±0.1
3.9±0.8
0.6±0.2
4.8±1.4
INNER LAYER
Retardance (nm)
Thickness (µm)
MIDDLE LAYER
Retardance (nm)
Thickness (µm)
OUTER LAYER
Retardance (nm)
Thickness (µm)
2.2±0.4
9.4±1.7
0.4±0.1
3.7±0.7
0.6±0.1
5.6±1.0

RETARDANCE and THICKNESS
Pelletier et al., 2004
a thin zona pellucida corresponds to a low retardance

CONCLUSION
Shen et al., 2005
• oocytes from cycles resulting in a pregnancy appear
to have a more homogeneous zona morphology
compared to oocytes from unsuccessful cycles.
• theoretically, a thicker zona could better protect
against ICSI damage
• eggs with a higher retardance derive from optimal
follicles

ZONA and BLASTOCYST FORMATION
Rama Raju et al., 2007
Ebner et al., 2005
Shen et al., 2005
INNER LAYER THICKNESS: 53% BFR (10-12nm) vs. 22% (3µm) vs. 17% (

PROSPECTIVE APPROACH
Montag et al., 2008
FERTILIZATION: 74% 68% 68%
IMPLANTATION: 40% 26% 13%
PREGNANCY: 65% 50% 25%

OCTAX polarAIDE
• involves an automatic mode of zona analysis
• the system automatically detects the zona pellucida
• birefringence/thickness are quantified within seconds
• an internal algorithm deduces a score from the values
• based on the empirical work of M. Montag
• positive scores appear green, negative ones red

ZONA and MATURITY
PI
P=0. 018
MI
Score: 20.5±6.6 Score: 16.8±4.9
P

FAILURE of MEASUREMENT
oocyte shape
ZG
positive: 23% negative: 14%
granulosa cells
MATURITY
OOCYTE QUALITY

… automatic zp detection failure seems to be a negative prognostic parameter

Oocyte quality might be affected by stimulation protocol

ZONA and PREGNANCY
RATE of VALID MEASUREMENTS
all oocytes
48/222 (21.6%)
vs
155/429 (36.1%)
MII-oocytes
42/193(21.8%)
vs
148/377 (39.3%)
P=0.002
P

ZONA and PREGNANCY II
SCORES
ALL OOCYTES
12.4 P=0.078 11.4
MII OOCYTES
12.0 P=0.066 10.9
ET / BT
11.6 P=0.14 9.7
100% IMPLANTATION

Thanks for your attention
Prof. Dr. Gernot TEWS
Dr. Johannes HARTL
Dr. Omar SHEBL
Dr. Richard B. MAYER
Dr. Andreas SIR
Dr. Veronika KRAIN
Dr. Marianne MOSER
Fr. Manuela PUCHNER
Fr. Renate WIESINGER