| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Case Report |
From the * Astra Fertility Clinic, Mississauga,
Ontario, Canada, Dr Ahmady is now with
University Fertility Centre, Torrance, California.
| Correspondence to: Dr Ali Ahmady, University Fertility Centre, 23550 Hawthorne Blvd, Suite 210, Torrance, CA 90505 (e-mail: ali_ahmady{at}yahoo.com). |
| Received for publication April 6, 2006; accepted for publication August 8, 2006. |
Case Report
This study was approved by our institutional board. A 36-year-old patient
and her 34-year-old husband, with 2 years history of primary infertility, were
referred to our center for treatment. The patient was a poor responder. Her
husband had suspected obstructive azoospermia. Testicular biopsy was preformed
before the initiation of the cycle to ensure having sperms on hand. Retrieved
sperms were frozen by addition of glycerol base cryoprotectant (Fertipro,
Beernum, Belgium) and cooling in liquid nitrogen vapor for 30 minutes.
Occasional motile sperms were seen before freezing. The frozen sample was
thawed by removing it from the storage and immersing in 37°C water bath
for 4 minutes.
Ovarian stimulation was performed using microflare protocol; namely, diluted leuprolide acetate, a gonadotropin releasing hormone agonist (GNRHa), and human menopausal gonadotropin (HMG). Daily dose of 0.05 mg Lupron subcutaneously bid and 300 IU HMG were administered starting day 2 of the cycle for 11 days. Follicular development was monitored by serial estradiol monitoring and transvaginal ultrasound. Human chorionic gonadotropin (hCG), 10 000 IU, was administered when optimal follicular development was achieved. Oocyte retrieval was performed by transvaginal ultrasonographic guided approach, 35 hours after hCG administration. Only 2 oocytes were retrieved. Only 1 oocyte was found suitable for ICSI.
A single sperm curling (SSC) test was used to evaluate viability of sperm (Ahmadi and Ng, 1997). SSC medium was prepared by adding 0.5 mL Hepes buffered medium to 1 mL distilled water to obtain osmolarity around 100 mOsm/kg. A single spermatozoon was exposed to SSC medium for 5 to 10 seconds. Upon contact with SSC medium, the tail would begin to coil in the case of viable sperm or would remain unchanged in the case of nonviable sperm. The spermatozoon would then be flushed into iso-osmotic medium and washed several times. The injection needle also was washed by repeated aspiration of fresh medium to ensure removal of the SSC solution. The SSC test, in this case, revealed 0% viability of the sperm evaluated. Given the fact that only 1 oocyte was acceptable for injection, we decided, after consulting with the couple, to proceed with ICSI using the available sperm sample. A sperm was selected and was moved into a droplet of 7.5% polyvinyl-pyrrolidone. The sperm was rinsed several times before proceeding into the oocyte droplet for injection.
Thirty minutes after ICSI, the injected oocyte was activated with calcium ionophore. The injected oocyte was incubated in G-Fert (Vitrolife, Englewood, Colo) medium containing 10 µg/mL calcium ionophore A23187 (stock solution 10 mg/mL in dimethyl sulfoxide stored at –20C; Sigma Chemical Co, St Louis, Mo) for 10 minutes. The oocyte was then washed a few times in G-Fert medium prior to culturing in G1 medium. Fertilization was checked around 18 hours after injection. One oocyte was fertilized, and the zygote reached the 8-cells stage with excellent quality on day 3. The resulting embryo was transferred aided by ultrasound guidance. The patient conceived and underwent an uneventful pregnancy. She delivered a healthy baby girl at term.
Discussion
It is known that oocyte injection of previously immotile sperm results in
either poor or no fertilization (Dozertsev
et al, 1995; Hoshi et al,
1995; Nijs et al,
1996). Poor fertilization could be due to inability of injected
sperm to trigger the activation process of the oocyte to start fertilization.
It is believed that live and intact sperm triggers calcium by introducing a
soluble factor into the egg cytoplasm to initiate the activation process
(Dale et al, 1985). We have
already demonstrated that the key point in fertilization of nonviable sperm is
activation of the injected oocytes. When the mouse and hamster oocytes were
activated by injection of cytosolic sperm factor, the same fertilization rate,
equivalent to injection with live intact sperm, was achieved
(Ahmadi and Ng, 1999). In this
report, we activated the injected human oocyte with calcium ionophore after
injection of frozen-thawed nonviable testicular sperm. To our knowledge this
is the first clinical report of pregnancy and delivery following transfer of
embryo resulting from injection of a nonviable sperm and oocyte activation. It
has been shown that calcium ionophore is effective to initiate the process of
fertilization in failed cases after ICSI
(Heindryckx et al, 2005).
Successful pregnancy and delivery have been reported as a result of activation
of oocyte injected with globozoospermic sample
(Kim et al, 2001). We believe
that by achieving fertilization following activation of the oocytes injected
with nonviable sperm, we are opening a window of hope for similar patients so
that they can have their own offspring.
References
Ahmadi A, Ng SC. Developmental capacity of damaged spermatozoa.
Hum Reprod. 1999; 14: 2279
–2285.
Ahmadi A, Ng SC. The single sperm curling test, a modified hypoosmotic swelling test, as a potential technique for the selection of viable sperm for intracytoplasmic sperm injection. Fertil Steril. 1997;68: 346 –350.[CrossRef][Medline]
Dale B, De Felice LJ, Ehrenstein G. Injection of a soluble sperm extract into sea urchin eggs triggers the cortical reaction. Experientia. 1985; 41: 1068 –1070.[CrossRef][Medline]
Dozertsev D, Rybouchkin A, De Sutter P, Qian C, Dhont M. Human
oocyte activation following intracytoplasmic injection: the role of the sperm
cell. Hum Reprod. 1995; 10: 403
–407.
Fischer R, Baukloh V, Naether OG, Schulze W, Salzbrunn A, Benson
DM. Pregnancy after intracytoplasmic sperm injection of sperm extracted from
frozen-thawed testicular biopsy. Hum Reprod. 1996; 11: 2197
–2199.
Heindryckx B, Van der Elst J, De Sutter P, Dhont M. Treatment
option for sperm- or oocyte-related fertilization failure: assisted oocyte
activation following diagnostic heterologous ICSI. Hum
Reprod. 2005;20: 2237
–2241.
Hoshi K, Yanagida K, Yazawa H, Katayosa H, Sato A. Intracytoplasmic injection using immobilized or motile spermatozoa. Fertil Steril. 1995;63: 1241 –1245.[Medline]
Khalifeh FA, Sarraf M, Dabit ST. Full-term delivery following
intracytoplasmic sperm injection with spermatozoa extracted from frozen-thawed
testicular tissue. Hum Reprod. 1997; 12: 87
–88.
Kim ST, Cha YB, Park JM, Gye MC. Successful pregnancy and delivery from frozen-thawed embryos after intracytoplasmic sperm injection using round-headed spermatozoa and assisted oocyte activation in a globozoospermic patient with mosaic Down syndrome. Fertil Steril. 2001; 75: 445 –447.[CrossRef][Medline]
Nagy ZP, Joris H, Verheyen G, Tournaye H, Devroey P, Van
Steirteghem AC. Correlation between motility of testicular spermatozoa,
testicular histology and the outcome of intracytoplasmic sperm injection.
Hum Reprod. 1998; 13: 890
–895.
Nijs M, Vanderzwalmen P, Vandamme B, Segal-Bertin G, Lejeune B,
Segal L, van Roosendaal E, Schoysman R. Fertilizing ability of immotile
spermatozoa after intracytoplasmic sperm injection. Hum
Reprod. 1996;11: 2180
–2185.
Podsiadly BT, Woolcott RJ, Stanger JD, Stevenson K. Pregnancy resulting from intracytoplasmic injection of cryopreserved spermatozoa recovered from testicular biopsy. Hum Reprod. 1996; 1: 1306 –1308.
Shulman A, Feldman B, Madgar I, Levron J, Mashiach S, Dor J.
Invitro fertilization treatment for severe male factor: the fertilization
potential of immotile spermatozoa obtained by testicular extraction.
Hum Reprod. 1999; 14: 749
–752.
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
