Endometriosis impairs the efficacy of gamete intrafallopian transfer: results of a case-control study*

FERTILITY AND STERILITY Vol. 62, No. 6, December 1994 Copyright 1994 The American Fertility Society Printed on acid-free paprr in U. 8. A. Endometriosis impairs the efficacy of gamete intrafallopian transfer: results of a case-control study* DavidS. Guzick, M.D., Ph.D.t Yvonne A.S. Yao, M.D. Sarah L. Berga, M.D. Joel S. Krasnow, M.D. Dale W. Stovall, M.D. Carolyn J. Kubik, M.D.t Anthony J. Zeleznik, Ph.D. Department of Obstetrics, Gynecology, and Reproductive Sciences, University of Pittsburgh School of Medicine, Magee- Womens Hospital, Pittsburgh, Pennsylvania Objective: To determine whether pelvic endometriosis impairs the efficacy of GIFT. Design: Matched follow-up study. Setting: University-based assisted reproduction program. Participants: Patients undergoing GIFT between 1987 and 1991. Cases had a primary diagnosis of endometriosis. Controls had no endometriosis and were matched with cases according to age, number of mature eggs transferred, and sperm grade. Intervention: Gamete intrafallopian transfer was performed in all patients in an identical manner independent of their underlying diagnosis. Main Outcome Measures: Pregnancy and delivery rates. Results: Of 114laparoscopic egg retrievals performed in the endometriosis group, there were 37 pregnancies (32.5%) and 27 deliveries (23. 7% ). Of the 214 retrievals in the control group, there were 101 pregnancies (47.2%) and 76 deliveries (35.5%). Mantel-Haenszel estimates of relative risk indicated that endometriosis significantly impaired pregnancy and delivery rates. There was no statistically significant difference in pregnancy rates according to severity of disease among endometriosis cases. There was no statistically significant difference in pregnancy rates according to severity of disease among endometriosis cases. Conclusions: Our finding that GIFT pregnancy rates were lower in women with a primary diagnosis of endometriosis than in matched controls suggests that endometriosis is associated with reduced efficacy of GIFT. Fertil Steril1994;62:1186-1191 Key Words: GIFT, endometriosis, case-control, assisted reproduction Received February 22, 1993; revised and accepted August 18, 1994. * Presented in part at the 48th Annual Meeting of The American Fertility Society, New Orleans, Louisiana, October 31 to November 4, 1992. t Reprint requests: David S. Guzick, M.D., Ph.D., Department of Obstetrics, Gynecology, and Reproductive Sciences, University of Pittsburgh School of Medicine, Magee-Womens Hospital, 300 Halket Street, Pittsburgh, Pennsylvania 15213-3180 (FAX: 412-641-1133). :j: Present address: Fertility Associates, Inc., Corporate One Office Park, Building II, Suite 330, 4075 Monroeville Boulevard, Monroeville, Pennsylvania. Contemporary knowledge regarding the association between endometriosis and infertility represents a paradox. Although case-control (1) and experimental animal studies (2) are indicative of an association between endometriosis and infertility, treatment of infertile women with minimal, mild, or moderate endometriosis does not appear to confer any benefit with respect to enhancement of fertility (3-5). Although several abnormalities associated with mild endometriosis have been advanced as possible causes of infertility, including altered menstrual cycle physiology, recent attention has focused on the peritoneal fluid (PF) compartment as containing possible mediators of infertility (6). Specifically, PF from endometriosis patients added to serum resulted in significantly lower sperm motility 1186 Guzick et al. Endometriosis and GIFT Fertility and Sterility

and velocity than control PF (7). Peritoneal macrophages, which can phagocytize normal sperm (8), were found to be in higher concentration in endometriosis PF (9), showed increased activation (10), and exhibited greater phagocytosis than macrophages from control PF (8). Interleukin-1, a protein produced by activated macrophages, was present in 10 of 11 endometriosis PF samples but was absent in 7 control PF samples (11); interleukin-1 concentrations similar to those seen in endometriosis PF were toxic to mouse embryo development (11). Peritoneal fluid from women with endometriosis inhibited in vitro cleavage of two-cell mouse embryos (12) and inhibited in vitro ovum capture by hamster oviductal fimbria (13). The above observations suggest that the PF of women with endometriosis may contain factors that interfere with sperm survival, egg capture, fertilization, and embryo growth. In vitro fertilization has been used successfully to establish pregnancies in infertile women with endometriosis who have not conceived from medical and surgical treatment (14). The superovulation component of IVF may overcome subtle defects of menstrual function; also, as IVF is conducted in a laboratory, the gametes are protected against a potentially unfavorable peritoneal environment during the fertilization process. Gamete intrafallopian transfer has been advocated as a method of assisted reproduction when there is at least one normal fallopian tube present. Pregnancy rates from GIFT appear to be somewhat higher on a national level than those from IVF (15), although the absence of a prospective randomized trial makes it difficult to compare IVF and GIFT results, as they tend to reflect experience in different types of diagnostic groups. Because endometriosis often affects peritoneal surfaces and ovaries without specifically involving the fallopian tubes, GIFT has been used in the treatment of infertile women with endometriosis. Indeed, reasonable success has been achieved: Mahmood and Templeton (6) report a combined pregnancy rate of 23.7% (23/ 97) in their review of seven studies in which GIFT was used in patients with endometriosis. Although it is clear from the literature that GIFT pregnancies can be established in women with endometriosis, the question remains as to whether exposure of gametes to the peritoneal environment impairs GIFT pregnancy rates in women with endometriosis as compared with those who do not have endometriosis. Previous studies of GIFT in the treatment of endometriosis either have not included a control group of women undergoing GIFT without endometriosis or have made comparisons to historical controls without matching them for important clinical characteristics. To address the question of whether GIFT pregnancy rates are reduced in the presence of endometriosis, we conducted a case-control study in which women undergoing GIFT with a primary diagnosis of endometriosis were compared with a matched group of women undergoing GIFT who did not have endometriosis. MATERIALS AND METHODS Since the inception of our Assisted Reproduction Program at the University of Pittsburgh, Magee Womens Hospital in 1987, women with endometriosis who were candidates for assisted reproduction underwent GIFT rather than IVF so long as there were patent fallopian tubes and the absence of a severe male factor. Between 1987 and 1991, 505 GIFT procedures were performed; in 114 of these cases, endometriosis was the principal diagnosis. Each GIFT procedure was considered an independent event. Endometriosis cases were matched with controls who underwent GIFT because of unexplained infertility or mild male factor infertility. Cases and controls were matched according to age ( <35; 35 to 39; ~40 years), the number of mature eggs transferred ( <3; ~3), and sperm grade (normal; mild impairment). Controls selected underwent GIFT procedures within ±12 months of the GIFT procedure performed on the case. The semen analysis was defined as normal (grade I) if the count was ~20 X 10 6 /ml, motility was ~50%, and normal morphology was ~35%. Mild impairment of sperm (grade II) was defined if the sperm count was between 10 and 20 X 10 6 /ml, sperm motility was between 30% and 50%, or normal morphology was between 20% and 35%. If any sperm parameter was below these levels, a severe sperm defect was considered to be present. In each stratification subgroup, two controls were selected for each case using a table of random numbers, with the exception of two subgroups in which available cycles were sufficient for only one control per case. These two subgroups were: age <35 years, <3 oocytes, sperm grade II; and age ~40 years, <3 oocytes, sperm grade II. With the exception of endometriosis severity, all of the data on GIFT cycles were obtained from our previously described assisted reproduction data Vol. 62, No.6, December 1994 Guzick et al. Endometriosis and GIFT 1187

.... base (16). Severity of endometriosis was assessed by grading according to The American Fertility Society Revised Classification of Endometriosis (17). This was done in each case from a chart review by one of the authors (Y.Y.). Unexplained infertility was diagnosed if a standard infertility evaluation (semen analysis, endometrial biopsy, hysterosalpingogram, laparoscopy) were all normal. For the GIFT procedure, superovulation, gamete preparation, and intrafallopian transfer were conducted in a manner described previously (18). Oocytes were graded for maturity according to standard methods (19). The luteal phase was supplemented with P (18). A pregnancy was defined as a gestation in which the serum value for (j-hcg was >250 miu/ml (conversion factor to SI unit, 1.0) using the World Health Organization 3rd International Standard no. 75/537 of chorionic gonadotropin. This study was conducted as a matched follow-up design (20). That is, cases and controls were defined by their exposure status (i.e., presence or absence of endometriosis) and followed to determine outcome status (presence or absence of pregnancy).* Because the number of controls per case varied (i.e., one or two), the relative risk of pregnancy (and delivery) from GIFT associated with endometriosis was estimated using the Mantel-Haenszel technique (20), as programmed in PC-SAS (21), which also provides 95% confidence intervals (CI) for the Mantel-Haenszel estimators. For the primary aim of this study, the comparison of pregnancy rates between cases and controls, our sample size allowed us to detect a difference of 15% (i.e., 30% versus 45%) with 72% power. RESULTS The characteristics for which cases were matched with controls are shown in Table 1. In each matching classification category, the number of GIFT attempts and the number of pregnancies are indicated for both cases and controls. Pregnancy and delivery results are shown in Figure 1. In total, 114 * Note that this design differs from a case-control study in which cases and controls are defined by outcome status and assessed retrospectively for exposure rates. A matched follow-up design better prevents confounding and also permits estimation of relative risk as opposed to the odds ratio estimate generated by a case-control approach (20). 1188 Guzick et al. Endometriosis and GIFT GIFT procedures were performed among endometriosis cases; there were 37 intrauterine pregnancies, or a pregnancy rate ± SE of 32.5% ± 4.2%. Among controls, there were 214 GIFT procedures performed; 101 intrauterine pregnancies resulted, or a pregnancy rate of 47.2% ± 3.4%. The Mantel Haenszel estimate of relative risk (RR) indicated that the presences of endometriosis significantly reduced the likelihood of pregnancy from GIFT (RR = 0.69; 95% CI = 0.51 to 0.92). In Table 2, pregnancy rates are shown for endometriosis cases according to severity of disease. Pregnancy rates in cases with minimal, mild, and moderate-severe endometriosis were 32.1% ± 6.4%, 38.2% ± 8.3%, and 25.9% ± 8.4%, respectively. Examination of these results suggest a lower pregnancy rate for moderate-severe endometriosis, consistent with a previous report by Y ovich and Matson (22). However, there were no statistically significant differences according to endometriosis severity among cases (x 2 = 0.75; P = 0.69). (It should be noted that the power in this sample to detect such differences was limited-approximately 20% power to detect a 15% difference from a baseline pregnancy rate of 30%.) Of 137 pregnancies, there were 34 spontaneous abortions. Thus, 103 deliveries occurred; 27 deliveries were among cases and 76 were among controls (Fig. 1). The results indicated that the presence of endometriosis significantly reduced the likelihood of delivery from GIFT (Mantel-Haenszel RR = 0.67; 95% CI = 0.47 to 0.97). DISCUSSION Although infertile women with endometriosis do not experience improved pregnancy rates after standard medical or surgical treatment, this does not preclude the possibility of an association between endometriosis and infertility. Put differently, the fact that standard endometriosis treatment does not help infertility patients may raise questions not about the underlying association between endometriosis and infertility, but rather about the efficacy of treatment. Even women who conceive after medical or surgical treatment for endometriosis, do so at a much lower rate (approximately 5% per cycle) (23) than normally fertile women (approximately 20% per cycle) (24). The mediator of the relationship between infertility and endometriosis may, as noted above, be localized within the PF compartment. If this were true, we would expect that GIFT pregnancy rates in Fertility and Sterility

Table 1 Number of GIFT Attempts and Pregnancies in Cases and Controls* Matched for Age of Female, Number of Embryos Transferred, and Sperm Grade Stratification groupings Cases Controls No. of mature Sperm Age oocytes transferredt grade:j: No. of No. of No. of No. of attempts pregnancies attempts pregnancies y <35 <3 I <35 <3 II <35 >3 I <35 >3 II 35 to 39 <3 I 35 to 39 <3 II 35 to 39 >3 I 35 to 39 >3 II ~40 <3 I ~40 <3 II ~40 >3 I ~40 >3 II Total Pregnancy rate(%) * Two controls were randomly selected for each case in each matching classification except for two classifications (age <35 years, <3 oocytes, sperm grade II; age ~40 years, <3 oocytes, sperm grade II) where the number of controls were sufficient for only a 1:1 match. 4 2 8 5 10 4 10 4 18 6 36 21 23 8 46 27 3 1 6 2 1 1 2 1 17 4 34 16 24 6 48 18 2 1 4 1 4 1 4 2 2 0 4 1 6 3 12 3 114 37 214 101 32.5 47.2 t Number of mature eggs transferred as part of the GIFT procedure. :j: Sperm grades: I if count ~20 X 10 6 /ml and motility ~50% and normal morphology ~35%; II if count 10 to 20 X 10 6 /ml; motility 30% to 50%, normal morphology 20% to 35%. women with endometriosis would be lower than those without endometriosis. In this matched casecontrol study, we found that although women with endometriosis have reasonably good pregnancy and delivery rates from GIFT, they still have a lower chance of pregnancy and delivery than women without endometriosis. 50 -?fl. 40 ~.:::: 30... C) C) 20... c.. as 10 a: 0 PREGNANCY DELIVERY Figure 1 Comparison of pregnancy and delivery rates (±SE) between endometriosis cases (D) and controls (111!1). Our findings point to the importance of using appropriate controls in analyzing data on results of GIFT and endometriosis. In the absence of information on controls, our pregnancy rate of 32.5% from GIFT in women with endometriosis would be considered well within the range of what one might expect from GIFT generally. However, matching each of the cases with controls who share similar ages and gamete characteristics indicated that, in our particular program, the success of GIFT in women with endometriosis was significantly below that of similar women undergoing GIFT for unexplained or mild male factor infertility. The absence of a relationship between severity of endometriosis and pregnancy rates may reflect inadequate power in our sample; insensitivity in the Table 2 Pregnancy Rates in Cases According to Severity of Endometriosis Cases No. of No. of Pregnancy Severity attempts pregnancies rate % Minimal 53 17 32.1 Mild 34 13 38.2 Moderate/Severe 27 7 25.9 Total 114 37 32.5 Vol. 62, No.6, December 1994 Guzick et al. Endometriosis and GIFT 1189

r! classification system (25); the possibility that factors interfering with egg pick-up, fertilization, etc., may be active regardless of the number or size of visible implants; or simply the fact that we used only cases of endometriosis in which the fallopian tubes were accessible for GIFT. Fertility certainly is impaired when endometriosis causes occlusion or other significant impairment of the fallopian tubes. If the fallopian tubes are normal, then the important parameter simply might be presence or absence of endometriosis, independent of severity. Because of the limited size of the data set, it was not possible to find controls for each case unless the time period was extended to ±12 months. We did not believe that this range (a maximum of 2 years) would introduce bias, as the trend in pregnancy rates has been very gradual across time and essentially has plateaued since 1989. However, to investigate this question, we reviewed our data and considered the average difference in time between cases and controls when controls were performed after cases (3.8 months) and found that this was virtually the same as the average time between cases and controls for controls chosen before cases (4.1 months). Moreover, the 214 controls were virtually evenly divided between those performed before and those performed after cases-105 versus 109, respectively. Although not surprising, as endometriosis cases accumulated in a uniform fashion across the entire interval of study, these results suggest that there was no evident bias resulting from our inability to match controls with cases more closely in time. The results of this study pose questions for clinical practice that can be answered by additional research. First, if endometriosis impairs the efficacy of GIFT, and if medical treatment is at all efficacious in (even transiently) suppressing the adverse effects of the peritoneal environment, then it may be possible that a course of medical treatment just before G 1FT could improve pregnancy rates. Second, if the presence of endometriosis impairs the efficacy of GIFT because of a peritoneal factor, then endometriosis may represent a diagnostic category in which IVF might be more successful than GIFT. Both of these issues can be addressed in the future by appropriate randomized prospective trials. Acknowledgments. We thank Joseph Fleiss, Ph.D., for his advice concerning the use of Mantel-Haenszel statistics, although any remaining errors remain with the authors. We are grateful to Ms. Pamela Nedzesky for data analysis and to Ms. Mary Parker for preparation of the manuscript. REFERENCES 1. Verkauf BS. The incidence, symptoms, and signs of endometriosis in fertile and infertile women. Fla Med Assoc 1987;74-671-5. 2. Schenken RS, Asch RH, Williams RF, Hodgen GD. Etiology of infertility in monkeys with endometriosis: luteinized unruptured follicles, luteal phase defects, pelvic adhesions, and spontaneous abortions. Fertil Steril 1984;41-122-30. 3. Bayer SR, Seibel MM, Saffan DS, Berger MF, Taymor ML. Efficacy of danazol treatment for minimal endometriosis in infertile women: a prospective, randomized study. J Reprod Med 1988;33:181-3. 4. Thomas EJ, Cooke ID. Successful treatment of asymptomatic endometriosis: does it benefit infertile women? Br Med J 1987;294:1117-9. 5. Telimaa S, Paolakka J, Ronnberg L, Kauppila A. Placebocontrolled comparison of danazol and high-dose medroxyprogesterone acetate in the treatment of endometriosis. Gynecol Endocrinol 1987;1:13-23. 6. Mahmood TA, Templeton A. Pathophysiology of mild endometriosis: review of literature. Hum Reprod 1990;5:765-54. 7. Oak MK, Chantler EN, Williams CAV, Elstein M. Sperm survival studies in peritoneal fluid from infertile women with endometriosis and unexplained infertility. Clin Reprod Fertil 1985;3:297-303. 8. Muscate JJ, Haney AF, Weinberg JB. Sperm phagocytosis by human peritoneal macrophages: a possible cause of infertility in endometriosis. Am J Obstet Gynecol1982;144:503-10. 9. Haney AF, Misukonis MA, Weinberg JB. Macrophages and infertility: oviductal macrophages as potential mediators of infertility. Fertil Steril 1983;39:310-5. 10. Halme J, BeckerS, Hammond MG, Raj MHG, Raj S. Increased activation of pelvic macrophages in infertile women with endometriosis. Am J Obstet Gynecol1983;145:333-7. 11. Fakih H, Baggett B, Holtz G, Tsang K-Y, Lee JC, Williamson HO. Interleukin-1: a possible role in the infertility associated with endometriosis. Fertil Steril1987;47:213-7. 12. Marcos RN, Gibbons WE, Findley WE. Effect of peritoneal fluid on in vitro cleavage of 2-cell mouse embryos: possible role in infertility associated with endometriosis. Fertil Steril 1985;44:678-83. 13. Suginami H, Y ano K, Watanabe K, Matsuura S. A factor inhibiting ovum capture by the oviductal fimbriae present in endometriosis peritoneal fluid. Fertil Steril 1986;46: 1140-6. 14. Oehninger S, Acosta AA, Kreiner D, Muasher SJ, Jones A W, Rosenwaks Z. In vitro fertilization and embryo transfer (IVF /ET): an established and successful therapy for endometriosis. J In Vitro Fert Embryo Transf 1988; 5:249-55. 15. Medical Research International, Society for Assisted Reproductive Technology (SART), and The American Fertility Society. In vitro fertilization-embryo transfer (IVF-ET) in the United States: 1990 results from the IVF -ET registry. Fertil Steril 1992;57:15-24. 16. Guzick DS, Boles J, Schadel R. Data base management system for assisted reproduction. J In Vitro Fert Embryo Transf 1990;7:236-40. 17. The American Fertility Society. Revised American Fertility Society classification of endometriosis: 1985. Fertil Steril 1985;43:351-2. 1190 Guzick et al. Endometriosis and GIFT Fertility and Sterility

1 18. Kubik CJ, Guzick DS, Berga SL, Zeleznik AJ. Randomized, prospective trial of leuprolide acetate and conventional superovulation in first cycles of in vitro fertilization and gamete intrafallopian transfer. Fertil Steril1990;54:836-41. 19. Ben-Rafael Z, Kopf GS, Blasco L, Flickinger GL, Tureck RW, Strauss JF, eta!. Follicular maturation parameters associated with the failure of oocyte retrieval, fertilization, and cleavage in vitro. Fertil Steril1986;45:51-7. 20. Rothman KJ. Modern epidemiology. Boston: Little, Brown and Co., 1986. 21. SAS Institute Inc. SAS/STAT user's guide, release 6.03 edition. Cary, NC: SAS Institute Inc., 1988. 22. Y ovich JL, Matson PL. The influence of infertility etiology on the outcome of IVF-ET and GIFT treatments. Int J Fertil 1990;35:26-33. 23. Guzick DS, Rock JA. A comparison of danazol and conservative surgery for the treatment of infertility due to mild or moderate endometriosis. Fertil Steril 1983;40:580-4. 24. Mariani P, Schwartz D. Sterility and fecundability estimation. J Theor Biol1983;105:211-9. 25. Guzick DS, Bross DS, Rock JA. Assessing the efficacy of The American Fertility Society's classification of endometriosis: application of a dose-response methodology. Fertil Steril1982;38:171-6. Vol. 62, No.6, December 1994 Guzick et al. Endometriosis and GIFT 1191