Development of a new alloplastic spermatocele demonstrating successful sperm retrieval in an animal model*t

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1 FERTILITY AND STERILITY Copyright ~ 1995 American Society for Reproductive Medicine Printed on acid-free paper in U. S. A. Development of a new alloplastic spermatocele demonstrating successful sperm retrieval in an animal model*t John E. Grantmyre, M.D.:j: Anthony J. Thomas, Jr., M.D.II Randall M. Falk, M.D.II~ Wassim Wazzan, M.D.II** Daniel Houlihan, D.O.lltt Michael Coburn, M.D.:j: Larry I. Lipshultz, M.D.:j: Arnold M. Belker, M.D.:j::j: Harris M. Nagler, M.D. Baylor College of Medicine, Houston, Texas; Cleveland Clinic, Cleveland, Ohio; University of Louisville, Louisville, Kentucky; and Mt. Sinai School of Medicine, New York, New York Objective: To create an alloplastic spermatocele capable of repeated sperm aspiration. The alloplastic spermatocele has long been a theoretical solution to infertility for those patients with congenital absence of the vas deferens or irreversible obstruction of the male reproductive ductal system. Recent studies have suggested that sperm from efferent ducts are capable of fertilization. Clinical use of alloplastic spermatoceles for collection of epididymal sperm has resulted in unacceptably low pregnancy rates. Improvement in spermatocele function may occur if a microsurgical anastomosis is performed to the epididymis. Design: A newly designed alloplastic spermatocele was implanted in 17 mature male rabbits. The faceplate of the device had a 0.7-mm orifice, allowing direct precise microsurgical anastomosis to a specific loop of the epididymal tubule. Results: Sperm retrieval was possible in 16/17 (94%) animals. Repeated successful aspirations (total of 73) were performed in all but one animal. The total number of sperm collected per spermatocele averaged 115 X 10 6 (range 0 to 734 X 10 6 ). The sperm motility varied widely between animals and specimens, with a maximum average of 21.6% motile sperm/aspirate per animal. All spermatoceles eventually occluded (mean time of occlusion 14 days; range 3 to 30 days). The prostheses with the attached epididymides were examined histologically. Conclusions: This prototype alloplastic spermatocele allows repeated high density sperm retrieval over a short period of time. Low sperm motility may be less problematic clinically as new techniques ofivf become available. Fertil Steril1995;64: Key Words: Alloplastic spermatocele, sperm aspiration, azoospermia Infertile men with obstructive azoospermia have normal functioning testes but, because of anatomic defects, sperm cannot be transported to the ejacu- late. When microsurgical reconstruction cannot be performed, implantation of an artificial sperm reservoir (alloplastic spermatocele), which would allow repeated sperm aspiration, potentially could help these patients. The indications for use of an alloplastic spermatocele are [1] aplasia of the vasa deferentia, [2] stenosis of a long segment of the vas deferens, or [3] loss of ejaculatory capacity either on a neuro- Received February 1, 1994; revised and accepted February 13, * Supported by American Medical Systems, Inc., Minnetonka, Minnesota and by grant RPC 22838, Cleveland Clinic, Cleveland, Ohio. t Presented at the 47th Annual Meeting of The American Fertility Society, Orlando, Florida, October 21 to 24, :j: Scott Department of Urology, Baylor College of Medicine. Reprint requests and present address: John Grantmyre, M.D., Dalhousie University, Department of Urology, Camp Hill Medical Center, 1763 Robie Street, Halifax, Nova Scotia, Canada, B3H 3G1 (FAX: ). II Department of Urology, Cleveland Clinic. ~ Present address: Department of Defense, Pentagon, Washington, D.C. ** Present address: Department of Urology, American University of Beirut, Beirut, Lebanon. tt Present address: Rockford Memorial Hospital, Rockford, Illinois. :j::j: Department of Urology, University of Louisville, Jewish Hospital. Department of Urology, Beth Israel Medical Center, Mt. Sinai School of Medicine. Grantmyre et al. Alloplastic spermatocele 179

2 .., logic basis or due to distal obstruction of the ejaculatory ducts not amenable to surgical correction. A patient must have active spermatogenesis with sperm present in at least a portion of the epididymis to be considered a candidate for this procedure. Bilateral agenesis of the vas deferens was described as early as 1755 but it took more than 200 years before the first artificial spermatocele was implanted. The first reports of alloplastic spermatocele use in humans were those reported by Hanley (1) in 1955, using amniotic sac for the reservoir. Schoysman (2, 3) in 1968 used saphenous vein grafts to create an artificial spermatocele. He reported three pregnancies in 17 couples who underwent these procedures. Further attempts at using this biologic spermatocele by Schoysman and others were unsuccessful. Kelami et al. (4, 5) in 1977 and 1981 and Wagenknecht et al. (6) in 1980 implanted silicone spermatoceles in various animal models and achieved pregnancies. Jimenez-Cruz (7) has reported human pregnancies using woven polypropylene (Dacron) spermatoceles, but he and other investigators subsequently used polytetrafluorethylene (PTFE) or Goretex as the material of choice. In 1986, Belker (8) reviewed the spermatocele experience of Jimenez-Cruz, Kelami, and Wagenknecht, including 130 alloplastic spermatoceles in 91 patients. Seven pregnancies resulted from all of these surgical efforts. These spermatoceles used a nonmicroscopic technique, either incising or dividing the epididymis itself. The device then was sutured to the epididymal adventitia. It was felt that one ofthe primary causes of failure in the majority of these implants was ingrowth of fibrous tissue occluding the epididymis from the spermatocele. With that in mind, a new spermatocele was designed in cooperation with American Medical Systems (implantation of the spermatoceles was performed at the Cleveland Clinic and Baylor College of Medicine). This spermatocele, in contrast to those mentioned previously, was designed with a 0.7-mm opening in the faceplate, so that a microsurgical side-to-side anastomosis could be performed directly to the lumen of an opened portion of the epididymal tubule. This anastomosis was performed with microsurgical techniques similar to vasoepididymostomy using 10-0 nylon sutures to perform the anastomosis. In addition, the spermatocele was designed to allow its contents while simultaneously drawing off the sperm laden fluid. The spermatocele faceplate was lined with mesh for strength. The purpose of the study was to test this new spermatocele in an animal model with regards to the quantity and quality of the sperm recovered as well as to assess the length of time of the device would remain patent. 180 Grantmyre et al. Alloplastic spermatocele MATERIALS AND METHODS A new spermatocele was designed by several of the authors (A.B., L.L., A.T., and H.N.) and made by American Medical Systems (Minnetonka, MN). The spermatoceles were implanted unilaterally in 17 mature New Zealand rabbits that were vasectomized simultaneously. General anesthesia was used at the time of implantation, which was performed using standard operating room protocol. Each rabbit received tetracycline as antibiotic prophylaxis for 48 hours postoperatively and received appropriate postoperative analgesics. The anastomosis between the spermatocele and a single epididymal tubule generally was performed at the epididymal tail. Once an individual tubule had been isolated by microscopic dissection, the tubule was incised longitudinally and the fluid was sampled for the presence of spermatozoa. The presence of sperm was established and a four-suture anastomosis was performed using the operating microscope and double-armed 10-0 nylon suture. On completion of the anastomosis the spermatocele was anchored to the adventitia of the epididymis using 9-0 suture. Initially the spermatoceles were implanted using a vertical scrotal incision. Rabbits have a bifid scrotum and the size of the spermatocele led to problems with erosion early in the study. The later implants were placed through a groin incision with delivery of the testicle through the incision to perform the surgery. The irrigation and aspiration ports were connected to the spermatocele by silicone tubing and tunneled subcutaneously (Fig. 1). With the ports high in the abdominal or lumbar area posteriorly, aspirations could be performed without anesthetizing the animal. Simultaneous percutaneous irrigation and aspiration using 3 ml of buffered phosphate saline solution was performed at the time of imp lantation and then every 3 days. Irrigation and aspirations were discontinued if the device malfunctioned or once azoospermia occurred. A semen analysis was performed on all the aspirate specimens to assess total number of sperm recovered and sperm motility. After the spermatocele had occluded, a final irrigation was performed with India ink to help identify the anastomotic site after the animal was killed. The spermatocele and testis were removed for histologic examination. RESULTS Seventeen spermatoceles were implanted initially. One spermatocele was considered a technical failure as only rare spermatozoa were ever recovered Fertility and Sterility

3 Figure 1 Diagram of irrigation and aspiration of implanted spermatocele. (poor spermatogenesis in this animal was possible as no testicular biopsy was performed). The other 16 spermatoceles had countable numbers of sperm in the aspirate and therefore, from a technical standpoint, the success rate for patency was 16/17 or 94%. Sixteen spermatoceles demonstrated at least some sperm production (i.e., remained patent) up to 6 days postoperatively. From this point on, recovery of sperm rapidly decreased and, by 30 days, all the spermatoceles were occluded with a mean time to occlusion of 14 days (range 0 to 30 days). The peak sperm density for each of the 17 spermatoceles is shown in Figure 2. The average peak sperm density was 36.4 x 10 6 spermlml, however, the range extended from 0 to 116 X 10 6 spermlml between devices. Approximately 3 ml of irrigation fluid was used for each aspiration. The average of the total number of sperm collected from each of the spermatoceles was 115 X 10 6 sperm (range 0 to 734 x 10 6 ) (see Fig. 3). The time to peak sperm density averaged approximately 6 days. The peak sperm motility reached I :I " :I a: 1&.1 A. rj) z 52 :::t i Figure 2 Peak sperm density (xl0 6 /ml) for each of the spermatoceles SPERMATOCELE Grantmyre et al. Alloplastic spermatocele 181

4 o """' SPERMATOCELE Figure 3 Total number of sperm (xi0 6 ) recovered from each spermatocele. > 50% in two spermatoceles, but the average peak motility was 21.6%. This varied quite widely between each animal and decreased rapidly over time (Fig. 4). By day 10 after surgery only one spermatocele contained motile sperm and by day 18 all aspirated sperm were nonmotile. Despite the relatively rapid disappearance of motile sperm, repeated aspirations were successful. A total of73 (range 2 to 12, mean 4.3) aspirations were performed on a total of 17 spermatoceles. Aspirations were discontinued when no sperm were recovered or a complication occurred. Histologic examination was performed on five spermatoceles that had prolonged function. Two spermatoceles were occluded by proteinaceous debris and three had evidence of chronic granulation reaction and fibrous tissue occluding the anastomotic site. Acute inflammation was not evident (see Fig. 5) The complications were few. There was one wound infection, one device erosion, one buckled spermatocele, one spermatocele leak, and one spermatocele apparently was eaten by the rabbit after the skin closure disrupted. DISCUSSION In 1955, Hanley (1) first published a report of an alloplastic spermatocele describing the use of an amniotic sac as the reservoir in a man with bilateral absence of the vas. Schoysman (2, 3) described the technique of anastomosing saphenous vein grafts to the epididymides of men. In 17 attempts the procedure resulted in three pregnancies. These results using vein grafts as sperm reservoirs could never be reproduced by other investigators as they were prone to early degeneration and fibrotic occlusion. Schosman's study, however, prompted the investiga :::E 40 a: w 0.. III W...J 30 :::E '# Figure 4 Peak sperm motility (%) recovered from each ANIMAL spermatocele. 182 Grantmyre et al. Alloplastic spermatocele Fertility and Sterility

5 Figure 5 Photomicrograph of rabbit epididymis after removal of occluded spermatocele showing fibrous tissue at the anastomotic site. tion of newer alloplastic materials. Kelami and associates (4) implanted a silicone prosthesis in 22 caput epididymides and 47 cauda epididymides of minipigs. The device was sewn over a large open area of multiple incisions into the epididymal tubule. Aspiration of the spermatocele contents between the 3rd and 5th postoperative days and insemination into receptive female pigs resulted in two pregnancies. Wagenknecht and associates (6) also developed a silicone spermatocele that was implanted in the epididymides of rats, bulls, and humans. Accurate reporting from these implantations was limited, but four pregnancies in cows resulted from insemination of aspirated bull sperm. Kelami et al. (4) implanted 18 prostheses in 14 patients and reported one pregnancy. He noted the best semen quality was obtained when aspiration was performed every 3rd to 5th day and that the devices became occluded between 1 and 24 weeks. Jimenez-Cruz (7) implanted woven polypropylene onto the transected distal end of the epididymis of 10 adult dogs. Sperm collection was performed every other week for 3 months, obtaining an aspiration volume of 0.1 to 0.3 ml, with sperm densities of 1 to 30 X 106/mL and motility ranging between 25% and 75%. With these encouraging results he implanted a spermatocele into a human patient. After a I-month period, aspiration of the spermatocele sample yielded a volume of 0.4 ml, a sperm concentration of approximately 30 X 106/mL, and sperm with 56% motility. The patient's wife was inseminated over a course of four ovulatory cycles before a pregnancy was achieved. There was no documentation of the aspirated volume, sperm concentration, or motility at the time of each insemination. The reports of successful human pregnancies stimulated further interest by investigators to develop a better alloplastic spermatocele. Turner and Smith (9) implanted spermatoceles made ofptfe vascular graft material into the distal epididymides of adult male rabbits. Three days postoperatively, the spermatoceles were aspirated, but examination of the fluid revealed <10 immotile sperm in 0.1 ml volume. Marmar et al. (10) implanted three PTFE sperm atoceles in three human patients. No pregnancies resulted and all sperm aspirated were immotile. Ross and Prins (11) implanted four PTFE spermatoceles. Aspiration of only one of the four spermatoceles implanted resulted in sufficient numbers of sperm for artificial insemination. Yoshida et al. (12, 13) implanted both PTFE and silicone spermatoceles into 11 humans but recovered only minimal numbers of motile sperm and no pregnancies resulted. Miura K et al. (14) recently reported on 33 silicone alloplastic spermatoceles. Motile sperm was aspirated successfully from 11 devices 1 month after implantation and two pregnancies resulted. In 1986, Belker et al. (8) published the largest available collection in the literature and documenting that the implantation of 130 alloplastic spermatoceles of different types resulted in seven pregnancies. Four progressed to delivery at term and three ended in spontaneous abortion. Several authors have reported pregnancies using sperm aspirated from the epididymis and efferent ducts in conjunction with IVF (15-17). At the present time this remains the only surgical treatment for patients who are candidates for an alloplastic spermatocele. Although this procedure has relatively little morbidity for the man undergoing the aspiration, it does require IVF with its inherent drawbacks. Repeat procedures have been performed but do require additional surgery, which theoretically could be avoided if an effective alloplastic spermatocele was available. Grantmyre et al. Alloplastic spermatocele 183

6 ....., The recent reported successes using IVF with semen of less than optimal quality have important implications for the successful implementation of the alloplastic spermatocele. Sperm obtained from a spermatocele aspirate may not be of sufficient quality and/or quantity to be used for routine intrauterine or intracervical insemination, but with enhancement techniques such as micromanipulation combined with IVF, the possibility of fertilization may be improved. Future considerations include fibrous and granulation tissue growth inhibitors, as a direct microsurgical anastomosis has not alleviated the problem of epithelial and fibrous tissue proliferation. In addition, the sperm motility remains low in spermatoceles, and this may be improved as epididymal fluid analogs, which allow healthier sperm to be recovered, are developed. Large numbers of sperm are retrieved from each of the spermatoceles and, at least early after implantation of the spermatocele, motile sperm are present. As some spermatoceles continue to yield small numbers of sperm for prolonged periods, IVF using micromanipulation may allow these small numbers of sperm to initiate a pregnancy. Muller-Tyl et al. (18) have reported a pregnancy 3 months after macroscopic implantation of a silicone alloplastic spermatocele using IVF (18). This new design of alloplastic spermatocele, using direct microsurgical anastomosis to a specific epididymal tubule, allows excellent technical success and initially a high number of sperm can be aspirated. The peak motility, however, remains quite low, as has been seen in previous spermatoceles, and early occlusion severely limits the use of this device. Acknowledgment. Robert Oates, M.D., Boston University Medical Center, Boston, Massachusetts and John Burton, Ph.D., American Medical Systems Inc., Minnetonka, Minnesota are recognized for their input in the design of the spermatocele. American Medical Systems provided funding as well as the engineering know how with their extensive experience using silicone prostatic devices. REFERENCES 1. Hanley GH. The surgery of male subfertility. Ann R Coll Surg EngI1955;17: Schoysman R. La creation d'un spermatocele artificial de les agenesis du conal deferent. Bull Doc R BeIge Gynecol Obstet 1968;38: Schoysman R. Surgical treatments in male sterility. Andrologia 1969; 1: Kelami A, RohloffD, Affeld K, Schroter A, Blohm B. Alloplastic spermatocele: insemination from epididymal reservoir. Urology 1977; 10: Kelami A. Kelami-Mfeld alloplastic spermatocele and successful human delivery. Urol Int 1981;36: Wagenknecht LV, Weitz KH, Hoppe LP, Krause D, Becker H, Schirren C. Microsurgery in andrologic urology. II. Alloplastic spermatocele. J Microsurg 1980; 1: Jimenez-Cruz JF. Artificial spermatocele. J Urol 1980; 123: Belker AM, Jimenez-Cruz JF, Kelami A, et al. Alloplastic spermatocele: poor sperm quality in intragrative epididymal fluid contraindicates prosthesis implantation. J Urol 1986; 36: Turner TT, Smith CC. Alloplastic spermatocele: diffusion across the allograft material and survival of spermatozoa in vitro and in vivo. J Urol 1984; 132: Marimar JL, DeBenedictis T J, Praiss DE. Clinical experience with an artificial spermatocele. J AndroI1984;5: Ross LS, Prins GS. Alloplastic spermatoceles: five year experience. J AndroI1985;6: Yoshida H, Naitoh Y, Iguchi H. Five cases of excretory azoospermia treated by artificial spermatocele with E-PTFE (Goretex). J Clin Urol 1984;38: Yoshida H, Miyamoto K, Yoshida T, et al. Implantation of artificial spermatocele with cup-shaped prosthesis for excretory azoospermia and chemical management of aspirated spermatozoa. J Androl 1986;4: Miura K, Motomu M, Takanami M, Ishi N, Shirai M. Clinical experience and successful impregnation using an artificial spermatocele. Urol Int 1991;47: Temple-Smith PD, Southwick GJ, Yates CA, et al. Human pregnancy by in vitro fertilization (IVF) using sperm aspirated from the epididymis. J In Vitro Fert Embryo Transf 1985;2: Olar TT, La Nasa J, Dickey RP, Taylor SN, Curole DN. Fertilization of human oocytes by microinjection of human sperm aspirated from the caput epididymis of an individual with obstructive azoospermia. J In Vitro Fert Embryo Transf 1990; 7: Jequier AM, Cummins JM, Gearon C, Apted SL, Yovich JM, Yovich JL. A pregnancy achieved using sperm from the epididymal caput in idiopathic obstructive azoospermia. Fertil Steril 1990;53: Muller-Tyl E, Deutinger J, Reinthallera A, Fischl F, Riss P, Lunglmayr G. In vitro fertilization with spermatozoa from alloplastic spermatocele. Fertil Steril 1990;53: Grantmyre et al. Alloplastic spermatocele Fertility and Sterility