Microsurgical epididymal sperm aspiration: aspirate analysis and straws available after cryopreservation in patients with non-reconstructable obstructive azoospermia

I. Schroeder-Printzen1, J. Zumbé2, L. Bispink3, St. Palm4, U. Schneider5,5, U. Engelmann2, W. Weidner1 and The MESA/TESE Group Giessen*

1 Department of Urology, University of Giessen and 2 Department of Urology, University of Cologne and 3 Outpatient Department of Obstetrics and Gynaecology, Bad Münder and 4 Outpatient Department for Reproductive Medicine, Cologne and 5 Clinic for Reproductive Medicine GmbH, Bad Münder, Germany


    Abstract
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Microsurgical epididymal sperm aspiration (MESA) combined with intracytoplasmic sperm injection (ICSI) represents a great advance in the therapy of non-reconstructable obstructive azoospermia. For procedure synchronization, a great number of organizational facilities are needed. Intentional cryopreservation of the aspirate may reduce these problems, therefore the aim of this study was to analyse the amount and quality of aspirate fluid obtained by means of MESA and the quality of the vials after thawing. Furthermore, the available cryopreserved straws were calculated. A total of 93 consecutive MESA procedures were performed and epididymal spermatozoa were obtained in 88 patients. Mean sperm concentration was 40.9x106 spermatozoa/ml. Global and progressive motility were 24.8 and 7.5% respectively. In one-third of the aspirates, no progressive motile spermatozoa were found. The mean number of straws available was 7.6. In 33 ICSI cycles with frozen–thawed epididymal spermatozoa, a pregnancy rate of 42.4% was achieved. In conclusion, these data show that enough spermatozoa are available for various ICSI cycles following a single MESA procedure in men with non-reconstructable obstructive azoospermia. Furthermore, ICSI with cryopreserved spermatozoa leads to excellent pregnancy rates

Key words: assisted reproductive techniques/infertility/microsurgical epididymal sperm aspiration/obstructive azoospermia


    Introduction
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
In azoospermic patients without disorders in spermatogenesis, the following indications for microsurgical epididymal sperm aspiration (MESA) are generally accepted: congenital aplasia of the vas deferens (CBAVD), inoperable obstruction of the seminal pathways, ejaculatory disorders where conservative therapy has not been successful (Belker et al., 1994Go; Silber et al., 1994Go, 1995Go; Ubaldi et al., 1995Go; Girardi and Schlegel, 1996Go; Zumbé et al., 1996Go; Bispink et al., 1997Go; Schroeder-Printzen et al., 1997Go) and, in particular, failed vasovasostomy (Silber et al., 1995Go; Ubaldi et al., 1995Go; Schroeder-Printzen et al., 1997Go).

MESA combined with intracytoplasmic sperm injection (ICSI) has increased the number of pregnancies (Silber et al., 1994Go). The fertilization rate is not affected by the kind of obstruction (congenital or acquired) and can be as high as 90% per cycle (Silber et al., 1995Go). At present, pregnancy rates of between 14 and 66% per cycle are achieved when MESA is combined with ICSI, while the live birth rate is as high as 36% per cycle (Belker et al., 1994Go; Silber et al., 1995Go; Ubaldi et al., 1995Go; Zumbé et al., 1996Go; Bispink et al., 1997Go; Schroeder-Printzen et al., 1997Go).

The synchronized combination of MESA and ICSI requires a great number of logistic facilities and involves the hormonal stimulation of the female partner pre-operatively, even though at this point it is not certain whether spermatozoa will be retrievable at all. When epididymal aspiration with subsequent cryopreservation is dissociated from ICSI, hormonal stimulation of the female is started after it has been clarified that enough motile spermatozoa are available for ICSI (Gips et al., 1996Go). There are no data in the literature which show a significant difference in clinical pregnancy rates when using fresh or cryopreserved epididymal spermatozoa (Oates et al., 1996Go; Chang et al., 1999Go; Tournaye et al., 1999Go). Thus damage to the spermatozoa caused by cryopreservation seems to be tolerable (Holden et al., 1997Go; Devroey et al., 1995Go).

Successful sperm retrieval does not depend on the different variations of the microsurgical technique (Matthews and Goldstein, 1996Go; Oates et al., 1996Go; Holden et al., 1997Go). It seems clear that the greatest numbers of motile spermatozoa (mean 4–13%) are to be found in the caput of the epididymis (Silber et al., 1995Go; Matthews and Goldstein, 1996Go; Oates et al., 1996Go; Holden et al., 1997Go). Aspiration is normally not difficult in this compartment (Devroey et al., 1995Go) and successful retrieval can be as high as 100%. The aspirate fluid which is not used for an intentional ICSI is cryopreserved (Silber et al., 1997Go). There are only limited data on the number of vials taken for ICSI and the quality of the cryopreserved spermatozoa in the above-mentioned literature. So far only Oates et al. have reported on the number of cryopreserved vials retrieved (Oates et al., 1996Go). In their study they were able to obtain 3–10 cryopreserved vials when sperm harvesting was done on a different day to the ICSI procedure, while the number of vials was no more than three when both procedures were performed on the same day (Oates et al., 1996Go).

In this study the day of sperm retrieval was separated from the day of ICSI procedure with a view to analysing the amount and quality of the spermatozoa aspirated under these conditions using a standardized MESA technique (Köhn et al., 1996Go; Schroeder-Printzen et al., 1996Go). In particular, the motility after thawing and the amount of cryopreservable straws were evaluated. Furthermore, the overall outcome, including the pregnancy and take home baby rate of ICSI cycles, using the cryopreserved epididymal spermatozoa, were analysed.


    Materials and methods
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Between 1995 and February 1999 MESA was performed in 93 men [mean age 36.1 (range 25–51) years] in a bicentric study of the departments of urology at the universities of Giessen and Cologne.

The predominant indications for MESA were CBAVD and failed microsurgical reconstruction (Table IGo). These two indications were analysed separately with regard to the patient's age, sperm concentration and global motility in the aspirate, motility after thawing and number of cryopreserved straws.


View this table:
[in this window]
[in a new window]
 
Table I. Indication for microsurgical epididymal sperm aspiration (n = 93)
 
Preoperative andrological work-up consisted of physical examination and hormone analysis (2xFSH, LH, testosterone). Semen analysis was performed twice according to World Health Organization (WHO, 1993) criteria. Azoospermia was diagnosed when no spermatozoa were found after centrifugation (2000 g, 10 min) and retrograde ejaculation had been excluded (Köhn et al., 1996Go). Genetic screening and counselling were performed according to the guidelines of the German Medical Council (1995).

Fifty-five patients were hospitalized for 1 day. Thirty-eight patients who were operated on outpatient conditions were allowed to leave 4 h after the end of the operation. They were examined the next day for haematoma or infection.

MESA was performed according to the standards of the German MESA/testicular sperm extraction (TESE) group (Zumbé et al., 1996Go). This standard has already been described (Schroeder-Printzen et al., 1996Go). MESA was performed under general anaesthesia. After scrotal exploration, spermatozoa were sampled from the microsurgically opened epididymal tubule with a micropipette or a 24-gauge cannula using a binocular microscope (magnification x15). One drop (10 µl) was filled into a haemocytometer (Makler® chamber) and was immediately examined for the presence of motile spermatozoa using a phase-contrast microscope (magnification x400). This procedure started at the cauda epididymis. If no motile spermatozoa were found, another tubule was opened 0.5 cm above the first one, until motile spermatozoa were aspirated. Sometimes it was necessary to open the ductuli efferentes. The opened tubules were closed by single stitches using 11–0 sutures. After examination, the samples were placed into a pre-warmed and pre-filled Eppendorf chamber®. The medium used for pre-filling the chamber was either buffered Earle's medium (Devroey et al., 1995Go), human tubular fluid medium [HTFM (Hovatta et al., 1995Go)] plus 1% serum albumin (Behringer, Marburg, Germany) or HEPES-buffered Ham's F-10 (Sigma, Deisenhofen, Germany). Afterwards these probes were taken to the laboratory using an aluminium chamber which had been pre-heated to 37°C. Sperm concentration and motility were examined. The examination was followed by sperm preparation using the migration-sedimentation technique (Tea et al., 1983Go). When sperm quality was >2x106 motile spermatozoa, a sperm preparation was not performed (Bispink et al., 1997Go). After mixing the sperm suspension with equal volumes of a cryoprotectant, the samples were filled in straws (Minitüb®; Landshut, Germany) each with a volume of 250 µl (Devroey et al., 1995Go). In the case of high amounts of motile spermatozoa, the aspirated fluid was diluted in order to increase the number of straws. The concentration was adjusted to 1–5x106 motile spermatozoa. The filled straws were stored in a computer-controlled freezer (Kryo 10 Serie III®; Messer Griesheim, Krefeld, Germany) and automatically frozen at –10°C/min.

Samples in the straws were thawed for straw examination and ICSI cycles at room temperature. Sperm density and motility were examined for a second time. No straws were thawed for examination in cases where there were four straws or fewer. At the beginning of the study, the gynaecologists decided in some cases to use fresh spermatozoa only, and therefore the semen was not cryopreserved. The remaining straws were counted and divided into four groups (no straws, <=4 straws, 5–10 straws, >10 straws).

Standard protocols for ovarian stimulation were used (Gips et al., 1996Go). Oocytes were aspirated by transvaginal, ultrasound-guided follicle puncture. Adherent cumulus cells were removed by exposure to hyaluronidase (80 IU/ml) for 1 min. The remaining parts of the cumulus were removed mechanically. ICSI procedure was carried out under direct visualization with phase-contrast optics and the assistance of electromechanical micromanipulators. Injected oocytes were maintained in drops of HTF supplement with human plasma proteins (Baxter, Unterschleißheim, Germany) and examined for the formation of a second pronucleus at 16–22 h. Up to three pronuclei were allowed to undergo further development during the ensuing 48 h. The surplus pronuclei were frozen or aborted. A flexible catheter was used for transcervical transfer.

For statistical analysis the Statistical Package for Social Science (SPSS for windows, Release 7.0; SPSS Inc., Chicago, IL, USA) and the Mann–Whitney U-test were used.


    Results
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
In 88 out of 93 patients (94.6%) the aspiration of motile spermatozoa from the epididymis was successful. The mean sperm density of the aspirated fluid after dilution was 40.9x106 spermatozoa/ml with a range of 30 000 to 247.5x106 spermatozoa/ml (Figure 1Go). The mean global motility of the 88 examined aspirates was 24.8% (range 1–99%) (Figure 2Go). Progressive motility was only 7.5% (range 0–76%). Thirty-three out of 88 (37.5%) examined aspirates did not contain progressive motile spermatozoa (WHO class A) (Figure 3Go).



View larger version (12K):
[in this window]
[in a new window]
 
Figure 1. Sperm concentration in the epididymal fluid obtained by microsurgical epididymal sperm aspiration (n = 88).

 


View larger version (11K):
[in this window]
[in a new window]
 
Figure 2. Global motility in the epididymal fluid microsurgical epididymal sperm aspiration (n = 88).

 


View larger version (11K):
[in this window]
[in a new window]
 
Figure 3. Progressive motile spermatozoa in the aspirates obtained by microsurgical epididymal sperm aspiration.

 
Forty-eight out of 88 (54.6%) aspirates were available for measuring motility after thawing. The mean global motility of the available samples decreased from 24.8 to 3.7%, the mean concentration of the thawed samples was 9.6x106 spermatozoa/ml. Eighteen samples showed only a few motile spermatozoa. In all, 67.4% of the samples had a global motility of <=1%. An additional analysis (Table IIGo) for the two predominant indications [CBAVD (n = 28) and failed microsurgical reconstruction (n = 23)] showed no significant difference in the sperm parameters.


View this table:
[in this window]
[in a new window]
 
Table II. Comparison of the male age and the mean sperm parameters for the two predominant indications, congenital aplasia of the vas deferens (CBAVD) and failed microsurgical reconstruction
 
An analysis of the total number of straws (Figure 4Go) showed that cryopreserving was not possible due to low aspirate quality in only two men. In 21 cases, we retrieved enough spermatozoa for one to four straws. In 25 patients up to 10 straws could be obtained. More than 10 straws were available in 21 men. Altogether a mean of 7.6 straws (range 0–23) was available, whereby in 75% of the cases four or more straws could be obtained, using a single MESA procedure. The statistical analysis (Table IIGo) of the cryopreserved straws showed no significant difference with regard to the two predominant indications (CBAVD and failed microsurgical reconstruction); the only significant difference was the age of the patients (P < 0.0001).



View larger version (30K):
[in this window]
[in a new window]
 
Figure 4. Available straws after cryopreservation (n = 69).

 
So far 33 ICSI cycles have been performed (Table IIIGo) in 26 couples using cryopreserved epididymal spermatozoa. In all, 123 of the 227 mature eggs fertilized, which is equivalent to a fertilization rate of 54.2%. Transfer was possible in 93.9% (as calculated per cycle), with a mean of 2.6 embryos per transfer. Fourteen clinical pregnancies were achieved, which is equivalent to a pregnancy rate of 42.4% (as calculated per cycle). In five cases, embryos aborted in the first trimester. Three pregnancies are still ongoing. Six healthy children have already been born.


View this table:
[in this window]
[in a new window]
 
Table III. Results of the intracytoplasmic sperm injection cycles with cryopreserved epididymal spermatozoa
 

    Discussion
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
This study confirms the data of others that MESA is successful for sperm retrieval in >90% of the cases (Belker et al., 1994Go; Chen et al., 1995Go; Collins et al., 1996Go; Oates et al., 1996Go; Holden et al., 1997Go; Silber et al., 1997Go). Motile spermatozoa were obtained in 94.6% of 93 attempts. The sperm concentration of ~40.9x106/ml is within the range given in the literature (Belker et al., 1994Go; Devroey et al., 1995Go; Oates et al., 1996Go; Holden et al., 1997Go). In the literature, mean global motility is between 13 and 29% (Belker et al., 1994Go; Devroey et al., 1995Go; Oates et al., 1996Go; Holden et al., 1997Go). With a value of 24.8%, results of the present study are at the upper level of this range. There appear to be no data available concerning the amount of progressively motile epididymal spermatozoa. In our study, the mean progressive motility was 7.5%, whereas 37.5% of the aspirates did not contain progressive motile spermatozoa at all. As described in the literature (Chen et al., 1995Go; Silber et al., 1995Go), there were no significant differences with regard to the above-mentioned parameters between patients with CBAVD or after failed microsurgical reconstruction.

The decisive question for this procedure is how many vials can be cryopreserved after sperm preparation? Up to now, only Oates et al. (1996) have published data concerning cases involving an asynchronous procedure. Similar to this study, they analysed how many vials are available when MESA and ICSI are not performed simultaneously. They obtained 6.4 vials/patient, compared with 7.6 vials in the current study. The same group harvested only 2.8 vials during MESA, when the operation was carried out under the condition that hormonal stimulation and oocyte harvesting of the female partner were performed on the same day, thus making it necessary to use the freshly harvested spermatozoa immediately (Oates et al., 1996Go). These data provide evidence that during asynchronous epididymal sperm aspiration, as already claimed by Oates et al. (1996), a high number of vials is always available, thus giving additional arguments for separating MESA from the oocyte retrieval procedure from an andrological-surgical point of view.

Using cryopreserved epididymal spermatozoa, the pregnancy rate of 42.4% is within the range given in the current literature (Silber et al., 1995Go; Oates et al., 1996Go), thus underlining the comparative efficiency of using cryopreserved spermatozoa in comparison to fresh spermatozoa for ICSI procedures (Devroey et al., 1995Go; Silber et al., 1995Go; Oates et al., 1996Go).

In conclusion, the results of this study demonstrate MESA as a successful procedure for retrieving spermatozoa in 94% of all men with obstructed azoospermia. Using this method, enough vials can be cryopreserved for a high number of consecutive ICSI cycles. As a high number of cryopreserved vials is available, a second operation becomes unnecessary in most of the patients. It is suggested that this constitutes a definite advantage in comparison to percutaneous retrieval methods (Collins et al., 1996Go), since the possibility of collecting spermatozoa for cryopreservation is successful in only 33–54% of the cases (Levine and Lisek, 1998Go; Rosenlund et al., 1998Go), so that further PESA procedures will become necessary. Even when percutaneous epididymal sperm aspiration is successful in further attempts (Meniru et al., 1998Go), there is an increasing risk for post-surgical complications. The recently suggested `mini-MESA technique' in sedoanalgesia, which has a success rate in cryopreservation of 92%, may be an alternative for the future (Nudell et al., 1998Go). This technique would combine the advantages of the MESA method (a predictably high number of preservable sperm) with the low invasiveness of the percutaneous method.


    Acknowledgments
 
The authors would like to thank Mrs T.Schneider and Mrs K.Zörb for their excellent technical assistance in the laboratory. This study was supported by grants from the BMBF(01 KY 9509-0, Weidner), DLR, Bonn, Germany and DFG center grant `Zell-Zell-Interaktion im Reproduktiongeschehen'.


    Notes
 
5 To whom correspondence should be addressed at: Department of Urology, Justus-Liebig University, Klinikstrasse 29, 35385 Giessen, Germany. E-mail: Immo.Schroeder-Printzen{at}chiru.med.uni-giessen.de Back

* Members of the MESA/TESE GROUP GIESSEN in alphabetical order: H.J.G.Herrero, F.Köhn, W.Künzel, W.-B.Schill, I.Schroeder-Printzen, R.Stalf, W.Weidner. Back


    References
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Belker, A.M. and the Sperm Microaspiration Retrieval Techniques Study Group (1994) Results in the United States with sperm microaspiration retrieval techniques and assisted reproductive technologies. J. Urol., 151, 1255–1259.[ISI][Medline]

Bispink, L., Schröter, D., Schroeder-Printzen, I. et al. (1997) Intracytoplasmatische Spermatozoeninjektion mit operativ gewonnenen Spermatozoen. Geburtsh. u. Frauenheilk., 57, 62–65.[ISI]

Chang, H.C., Hsieh, J.T., Liu, S.P. et al. (1999) Fertilization capability of frozen epididymal sperm for intracytoplasmatic sperm injection. J. Formos. Med. Assoc., 98, 171–174.[ISI][Medline]

Chen, C.S., Chu, S.H., Soong, Y.K. et al. (1995) Epididymal sperm aspiration with assisted reproductive techniques: difference between congenital and acquired obstructive azoospermia. Hum. Reprod., 10, 1104–1108.[Abstract]

Collins, G.N., Critchlow, J.D., Lau, M.W.M. et al. (1996) Open versus closed epididymal sperm retrieval in men with secondarily obstructed vasal systems | a preliminary report. Br. J. Urol., 78, 437–439.[ISI][Medline]

Deutsche Gesellschaft für Gynäkologie und Geburtshilfe/Arbeitsgemeinschaft für Gynäkologische Endokrinologie und Fortpflanzungsmedizin (1995) Empfehlungen zur Durchführung der intrazytoplasmatischen Spermieninjektion (ICSI) als Zusatzmaßnahme bei IVF-ET-Therapie. Frauenarzt, 36, 818–819.

Devroey, P., Silber, S.J., Nagy, Z.P. et al. (1995) Ongoing pregnancies and birth after intracytoplasmic sperm injection with frozen-thawed epididymal spermatozoa. Hum. Reprod., 10, 903–906.[Abstract]

Gips, H., Hormel, P. and Hinz, V. (1996) Ovarian stimulation in assisted reproduction. Andrologia, 28 (Suppl. 1), 3–7.

Girardi, S.K. and Schlegel, P.N. (1996) Microsurgical epididymal sperm aspiration. Urol. Clin. North. Am. (Atlas), 4, 55–66.

Holden, C., Fuscaldo, G.F., Jackson, P. et al. (1997) Frozen-thawed epididymal spermatozoa for intracytoplasmatic sperm injection. Fertil. Steril., 67, 81–87.[ISI][Medline]

Hovatta, O., Moilanen, J., von Smitten, K. et al. (1995) Testicular needle biopsy, open biopsy, epididymal aspiration and intracytoplasmatic sperm injection in obstructive azoospermia. Hum. Reprod., 10, 2595–2599.[Abstract]

Köhn, F.M., Schroeder-Printzen, I., El Mulla, K.F. et al. (1996) Andrological work up of patients undergoing microsurgical epididymal sperm aspiration or testicular sperm extraction. Andrologia, 28 (Suppl. 1), 77–81.[ISI][Medline]

Levine, L.A. and Lisek, E.W. (1998) Successful sperm retrieval by percutaneous epididymal and testicular sperm aspiration. J. Urol., 159, 437–440.[ISI][Medline]

Matthews, G.J. and Goldstein, M. (1996) A simplified method of epididymal sperm aspiration. Urology, 47, 123–125.[ISI][Medline]

Meniru, G., Gorgy, A., Batha, S. et al. (1998) Studies of percutaneous epididymal aspiration (PESA) and intracytoplasmatic sperm injection. Hum. Reprod. Update, 4, 57–71.[Abstract/Free Full Text]

Nudell, D.M., Conaghan, J., Pedersen, R.A. et al. (1998) The mini-micro-epididymal sperm aspiration for sperm retrieval: a study of urological outcomes. Hum. Reprod., 13, 1260–1265.[Abstract]

Oates, R.D., Lobel, S.M., Harris, D.H. et al. (1996) Efficacy of intracytoplasmatic sperm injection using intentionally cryopreserved epididymal spermatozoa. Hum. Reprod., 11, 133–138.[Abstract]

Rosenlund, B., Westlander, G., Wood, M. et al. (1998) Sperm retrieval and fertilization in repeated percutaneous epididymal sperm aspiration. Hum. Reprod., 13, 2805–2807.[Abstract/Free Full Text]

Schroeder-Printzen I., Köhn, F.-M., Ludwig, M. et al. (1996) Mikrochirurgische Spermatozoenaspiration (MESA) und TEstikuläre Spermatozoen Extraktion (TESE). Aktuel. Urol., 27, Operative Techniken 6.33.

Schroeder-Printzen I., Köhn, F.-M., Ludwig, M. et al. (1997) Mikrochirurgische Spermatozoenaspiration (MESA) und TEstikuläre Spermatozoen Extraktion (TESE) – eine Übersicht – Aktuel. Urol., 28, 251–259.[ISI]

Silber, S.J., Nagy, Z.P., Devroey, P. et al. (1997) The effect of female age and ovarian reserve on pregnancy rates in male infertility: treatment of azoospermia with sperm retrieval and intracytoplasmatic sperm injection. Hum. Reprod., 12, 2693–2700.[Abstract]

Silber S.J., Nagy, Z.P., Liu, J. et al. (1994) Conventional in-vitro fertilization versus intracytoplasmatic sperm injection for patients requiring microsurgical sperm aspiration. Hum. Reprod., 9, 1705–1709.[Abstract]

Silber S.J., Nagy, Z.P., Liu, J. et al. (1995) The use of epididymal and testicular spermatozoa for intracytoplasmatic sperm injection: the genetic implications for male infertility. Hum. Reprod., 10, 2031–2043.[Abstract]

Tea, N.T., Jonded, M. and Scholler, R. (1983) A migration-gravity sedimentation method for collecting human motile spermatozoa from human semen. In Harrison, R.F., Bonnaraud, J. and Thompson, W. (eds), In Vitro Fertilization, Embryo Transfer and Early Pregnancy. MTP Press, pp. 117–120.

Tournaye, H., Merdad, T., Silber, S. et al. (1999) No differences in outcome after intracytoplasmatic sperm injection with fresh or with frozen-thawed epididymal spermatozoa. Hum. Reprod., 14, 90–95[Abstract/Free Full Text]

Ubaldi, F., Liu, J., Nagy, Z.P. et al. (1995) Indications for and results of intracytoplasmatic sperm injection (ICSI). Int. J. Androl., 18 (Suppl. 2), 88–90.

Zumbé, J., Beintker, M., Denil, J. et al. (1996) Experiences of the German Section for Urological Microsurgery. Andrologia, 28 (Suppl. 1), 89–92.

Submitted on December 8, 1999; accepted on September 15, 2000.





This Article
Abstract
FREE Full Text (PDF )
Alert me when this article is cited
Alert me if a correction is posted
Services
Email this article to a friend
Similar articles in this journal
Similar articles in ISI Web of Science
Similar articles in PubMed
Alert me to new issues of the journal
Add to My Personal Archive
Download to citation manager
Search for citing articles in:
ISI Web of Science (2)
Request Permissions
Google Scholar
Articles by Schroeder-Printzen, I.
Articles by MESA/TESE Group Giessen, T.
PubMed
PubMed Citation
Articles by Schroeder-Printzen, I.
Articles by MESA/TESE Group Giessen, T.