2. Sperm morphology & IVF outcome
• Head abnormalities mainly responsible for sperm
fertilization deficiency
• Flagellar abnormalities haven’t been shown to have
any impact on fertilization (Berkovitz et al., 2006; Chemes
et al., 2003)
• Sperm nuclear DNA damage is closely related to male-
derived repeated failed ICSI attempts (Tesarik et al., 2004;
Tesarik 2005)
• Presence of large nuclear vacuoles: negatively
associated with natural male fertility potential (Bartoov
et al., 1994; Mundi et al., 1994)
3. ICSI + MSOME = IMSI
• ICSI (Palermo et al., 1992): inverted microscope
magnification of x 400
• MSOME - motile sperm organellar morphology
examination (Bartoov et al., 2002)
The resulting magnification was based on 4 parameters:
1. Objective magnification x100
2. Magnification selector x1.5
3. Video coupler magnification x0.99
4. Calculated video magnification (ccd x monitor diagonal
dimension) x 44.45
TOTAL MAGNIFICATION: 100 x 1.5 x 0.99 x 44.45 = x 6600 !!!
4. IMSI equipment
IMSI can be achieved on a Nikon inverted microscope equipped with high
resolution Nomarski optics (100x or 60x oil Plan Apo VC) enhanced by a
videozoom and digital imaging. The final magnification of the system can
be calculated as follows:
(objective) X (magnification changer) X (videozoom) X (diagonal of image
camera CCD chip diagonal)
6. IMSI Sperm assessment (1)
The MSOME criteria for the morphological normalcy of the
sperm nucleus were defined as below:
Smooth
Symmetric
Oval Configuration
Homogeneity of the nuclear chromatin mass (no more
than one vacuole / less than 4% of the nuclear area)
The average length and width limits in 100 spermatozoa
with a normally looking nucleus, are estimated as follow:
Length: 4.75 ± 0.28 μm
Width: 3.28 ± 0.20 μm
8. Vacuoles (1)
• Vacuole: concavity extending from the surface of the sperm
head to the nucleus through the acrosome
• Initially: considered as degenerative structures with no
physiological significance – normal feature of sperm head
(Fawcett, 1958; Chrzanowski 1966; Bedford 1967; Pedersen 1969)
9. Vacuoles (2)
Post-IVF era:
• Mundi et al., 1994: sperm nuclei in
subfertile men contained more
intranuclear vacuoles than fertile
controls
• ICSI with sperm with vacuoles =
decreased PR, early miscarriage
(Bartoov et al., 2001, 2002, 2003; Berkovitz et
al., 2005, 2006; Franco et al., 2008; Antinori
et al., 2008)
Vacuoles are not just a polymorphism
but pose a risk for an abnormality
that is accompanied by DNA injury
10. Sperm grading: vacuoles
• Vanderzwalmen et al., 2008:
Grade I: absence of vacuoles
Grade II: max. of 2 small vacuoles
Grade III: >2 small vacuoles or at
least 1 large
Grade IV: abnormal head shapes or
other abnormalities with or
without vacuoles (identified even
at x400 magnification)
11. Vacuoles & DNA fragmentation
• Spermatozoa with large vacuoles should be avoided (Tanaka et al., 2012)
• Spermatozoa with large vacuoles (>50% of the sperm head) & other potential
abnormalities were described as having higher DNA fragmentation levels than
normal spermatozoa (Franco et al., 2008)
• Spermatozoa with multiples vacuoles (>4% of the head area) have higher DNA
fragmentation levels than normal spermatozoa (Wilding et al., 2011)
• Large vacuoles associated with DNA fragmentation (Oliveira et al., 2010; Hammoud
et al., 2012)
• No difference in DNA fragmentation between normal ones & morphometrically
normal ones with a single, large vacuole (15%-25% of head area) (Watanabe et al.,
2011; Boitrelle et al., 2011)
• No difference in DNA fragmentation between spermatozoa with 1 large vacuole
& non-selected ones (Perdrix et al., 2011)
12. Studies (1)
• Prospective randomized trials with unselected pt
- Antinori et al. , 2008
- Balaban et al., 2011
• Benefit of IMSI after ICSI implantation failure (without assessing influence of
sperm abnormalities)
- Berkovitz et al., 2006
- Bartoov et al., 2003
- Nadalini et al., 2009
- Oliveira et al., 2011 not significantly higher ongoing pregnancy, miscarriage
& LBR after IMSI
• Pt with isolated teratozoospermia
- Knez et al., 2012 higher PR in IMSI group
13. Studies (2)
• Comparable embryo development between
IMSI & ICSI on Day 2 (Mauri et al., 2010; Oliveira et al., 2011)
and Day 3 (Vanderzwalmen et al., 2008; Balaban et al., 2011; de
Almeida Ferreira Braga et al., 2011; Knez et al., 2011)
14. Meta-analysis
Setti et al., 2010
IMSI outcomes (% of top quality embryos, IR, PR) significantly improved as
compared with ICSI cycles.
Miscarriage rate significantly decreased as compared with ICSI.
NO difference between ICSI & IMSI regarding the FR (late paternal effect??)
15. When to use IMSI
• Severe oligo-terato-asthenospermia
Antinori et al., 2008 (prospective randomized trial; 446 couples: ICSI (n=219) VS IMSI
(n=227) higher IR & PR)
Setti et al., 2011: Sperm selection under high magnification results in improve
treatment outcomes in patients with OATS
• Previous conventional ICSI failures with significant % of DNA fragmented
spermatozoa in the ejaculate
Hazout et al., 2006:
- Improved pregnancy outcome in pt with repeated conventional ICSI failure
- Subgroup of pt (n=72): assessment of DNA integrity (TUNEL)improvement in
clinical outcomes (IR & LBR) in pt with >30% DNA fragmentation
• Couples with 1-3 yrs of infertility & semen characteristics 1-20mill/ml
Wilding et al., 2011:
- Improved embryo morphology together with significantly increased PR & IR
• De Vos et al., 2013: no benefit of IMSI in a non-selected population with fresh
ejaculation sperm containing >1mil/ml
• Klement et al., 2013: IMSI procedure is an option to be mentioned after the 1st
unsuccessful ICSI cycle
16. Sperm preparation techniques
• Both SUP & DGP efficient in recovering lower
percentage of sperm cells with vacuolization (Monqaut et
al., 2011; Borges et al., 2013)
• POOR SPERM MOTILITY :
temperature 37°C, no PVP and supplemented with 6% of human
serum albumin
• HIGH SPERM MOTILITY :
temperature 20°C and concentration of PVP about 8%.
17. Benefits
• Berkovitz et al., 2007
Sperm nucleus morphological normalcy assessed
at high magnification could decrease the
prevalence of major fetal malformations in ICSI
children
18. Concerns
• Prolonged exposure to 37°C during IMSI could
impair the morphological integrity of the
sperm nuclei (Peer et al., 2007)
• The No of spermatozoa without vacuoles in
humans is extremely small and trying to find
and then inject such cells is a very difficult &
time-consuming process (Tanaka et al., 2012)
19. References
• Antinori, M., Licata, E., Dani, G., et al., 2008. Intracytoplasmic morphologically selected sperm injection: a
prospective randomized trial. Reprod. Biomed. Online 16, 835–841.
• Bartoov, B., Berkovitz, A., Eltes, F., 2001. Selection of spermatozoa with normal nuclei to improve the pregnancy
rate with intracytoplasmic sperm injection. N. Engl. J. Med. 345, 1067–1068.
• Bartoov, B., Berkovitz, A., Eltes, F., et al., 2003. Pregnancy rates are higher with intracytoplasmic morphologically
selected sperm injection than with conventional intracytoplasmic injection. Fertil. Steril. 80, 1413–1419.
• Bartoov, B., Berkovitz, A., Eltes, F., et al., 2002. Real-time fine morphology of motile human sperm cells is
associated with IVF-ICSI outcome. J. Androl. 23, 1–8.
• Hazout, A., Dumont-Hassan, M., Junca, A.M., et al., 2006. High-magnification ICSI overcomes paternal effect
resistant to conventional ICSI. Reprod. Biomed. Online 12, 19–25.
• Vanderzwalmen, P., Hiemer, A., Rubner, P., et al., 2008. Blastocystmdevelopment after sperm selection at high
magnification is associated with size and number of nuclear vacuoles. Reprod. Biomed. Online 17, 617–627.
• Palermo, G., Joris, H., Devroey, P., et al., 1992. Pregnancies after intracytoplasmic injection of single spermatozoon
into an oocyte. Lancet 340, 17–18.
• Berkovitz, A., Eltes, F., Yaari, S., Katz, N., Barr, I., Fishman, A., Bartoov, B., 2005. The morphological normalcy of the
sperm nucleus and pregnancy rate of intracytoplasmic injection with morphologically selected sperm. Hum.
Reprod. 20, 185–190.
• Berkovitz, A., Eltes, F., Lederman, H., Peer, S., Ellenbogen, A., Feldberg, B., Bartoov, B., 2006a. How to improve IVF-
ICSI outcome by sperm selection. Reprod. Biomed. Online 12, 634–638.
• Berkovitz, A., Eltes, F., Ellenbogen, A., Peer, S., Feldberg, D., Bartoov, B., 2006b. Does the presence of nuclear
vacuoles in human sperm selected for ICSI affect pregnancy outcome? Hum. Reprod. 21, 1787–1790.
• Knez, K., Zorn, B., Tomazevic, T., Vrtacnik-Bokal, E., Virant-Klun, I., 2011. The IMSI procedure improves poor
embryo development in the same infertile couples with poor semen quality: a comparative prospective
randomized study. Reprod. Biol. Endocrinol. 9, 123.
20. References
• Franco Jr., J.G., Baruffi, R.L., Mauri, A.L., Petersen, C.G., Oliveira, J.B., Vagnini, L., 2008. Significance
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Jr, Borges E Jr. Intracytoplasmic sperm injection outcome versus intracytoplasmic morphologically
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