Correlation of estrogen levels with oocytes aspirated and with pregnancy in a program of clinical tubal transfer*

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1 FERTILITY AND STERILITY Copyright e 9 8 Vol. 4 8,, July The American Fertility Society Printed in U.S.A. Correlation of estrogen levels with oocytes aspirated and with pregnancy in a program of clinical tubal transfer* Robert W. McGaughey, Ph.D.t* Jay S. Nemiro, M.D.t ll The Arizona Center for Fertility Studies, Good Samaritan Medical Center, Phoenix, and Arizona State University, Tempe, Arizona Transfer of human spermatozoa and mature oocytes to patent fallopian tubes (tubal transfer, TT) in cases of infertility provides a successful, simple alternative to in vitro fertilization-embryo transfer. The authors report their second TT series performed between October, 985 and February, 986. Of 59 transfers, 8 clinical pregnancies were obtained, of which 2 were ectopic and spontaneously aborted. A triplet pregnancy, sets of twins, and singleton pregnancies delivered normally. Pregnancy was highly correlated with number of mature oocytes aspirated and with number of oocytes transferred. The study demonstrates that, in combination, serum estrogen levels and number of developing follicles observed by ultrasound are predictive of the number of mature oocytes obtained at laparoscopy. These predictive values provide excellent criteria for selecting stimulation cycles for TT and for informing individual patients of the probability for success before surgery. Fertil Steril 48:98, 987 In an earlier study we reported a clinical procedure for transferring oocytes and spermatozoa to patent fallopian tubes (tubal transfer, TT) as an alternative to in vitro fertilization-embryo transfer (IVF-ET) in the treatment of infertility. Other authors also have reported success with a similar technique. 2-4 Success rates, measured as biochemical or clinical pregnancies per cycle or per transfer, are comparable for TT and IVF-ET.-5 In our first series of patients, we found that successful TT was correlated significantly with ele- Received December 8, 986; revised and accepted March, 987. * Presented at the forty-second annual meeting of The American Fertility Society and the eighteenth annual meeting of the Canadian Fertility and Andrology Society, September 27 to October 2, 986, Toronto, Ontario, Canada. t The Arizona Center for Fertility Studies. + Department of Zoology, Arizona State University. Good Samaritan Medical Center. II Reprint requests: Jay S. Nemiro, M.D., The Arizona Center for Fertility Studies, 464 East Shea Boulevard, Suite D-25, Phoenix, Arizona vated estradiol (E2) as measured by radioimmunoassay (RIA) of patient serum before injection of human chorionic gonadotropin (hcg) to stimulate follicular and oocyte maturation. Pregnancy resulting from TT also was significantly related to semen sperm count, to the number of sperm transferred, and to the motility percentage for transferred sperm. Although in our first series the numhers of clinical pregnancies were too small to test statistically, the results suggested that the transfer of four oocytes increased the probability of pregnancy as compared with transfer of fewer than four oocytes. We report here the results of our second series of patients who were treated by TT during the period of October, 985 to February, 986. We describe here several technical changes in the TT procedure that collectively have improved the success rate significantly. Analyses of data from our second series demonstrate that pregnancy is correlated with number of mature oocytes recovered and with number of oocytes transferred. In addition, the data of this series strongly suggest that the number 98 McGaughey and Nemiro Pregnancies with tubal transfer Fertility and Sterility

2 of follicles observed by ultrasound and the level of E 2 measured before injection of hcg predict the number of mature oocytes recovered and thereby the probability of success with TT for individual cycles. MATERIALS AND METHODS A detailed technical description of TT was provided previously and was followed fundamentally in the current study. In summary, TT includes the stimulation of follicular development with clomiphene citrate (CC) and/or human menopausal gonadotropin (hmg, Pergonal, Serono Laboratories, Inc., Randolph, MA), the stimulation by hcg of preovulatory follicular and oocyte development, the collection and washing of spermatozoa, the laparoscopic aspiration of preovulatory oocytes, and the transfer of selected oocytes and spermatozoa to the ampulla of the fallopian tube by means of a catheter during laparoscopy. A total of patients underwent 9 cycles of ovarian stimulation with 5 mg of CC on cycle days to 7 (42 cycles), with 2 to ampules of hmg daily from cycle day until completion of follicular development (7 cycles), with mg of CC on cycle days to 7 and 2 to ampules of hmg daily from cycle day until completion of follicular development (56 cycles) or with mg CC on cycle days to 7 and 2 to ampules of hmg on cycle days 7, 9, and (4 cycles). For all stimulation protocols, 5 IU hcg (Profasi, Serono Laboratories, Inc., Randolph, MA) was administered by intramuscular injection when two or more dominant follicles reached 8 to 2 mm in their largest diameter, as measured by ultrasonography. The series represented the following diagnoses for infertility: peritubular-periovarian adhesions (27 patients), unexplained infertility (26 patients), oligospermia (9 patients), endometriosis (2 patients), cervical factor (6 patients), tubal disease (7 patients), anovulation ( patients), and immunologic infertility ( patients). The average age for these patients was 2.4 years (±.4 standard error [SE]), and the mean number of years of infertility was 4.9 (±.). All patients had failed standard treatment before inclusion in the TT program, and the presence of at least one patent fallopian tube was documented in all cases. Laparoscopy was performed in 6 cycles representing 59 patients, and in two cycles aspiration of ovarian follicles was impossible because of extensive pelvic adhesions. TT was accomplished in all other cases (59 cycles). Criteria for cancellation of stimulation cycles before laparoscopy were the same as those described previously, and included no response to stimulation (22 cycles), atresia of developing follicles (28 cycles), cystic follicular development (6 cycles), low estrogen production ( cases), early surge of luteinizing hormone (LH) ( cycles), and patient choice ( cycle). Atresia, the major reason for cancellation, was characterized by ultrasonagraphy for cycles in which apparently normal follicles began to develop early in the cycle, but all but one or two stopped development or disappeared during the stimulation cycle. The following detailed modifications in the TT procedure were made after analysis of the results of our first series, and were incorporated into the presently reported study. All semen samples were obtained within a shorter interval before laparos- copy in the present study. Laparoscopies were scheduled between 6: A.M. and 9: A.M., and semen samples obtained between 2:45 A.M. and 4: A.M. The preparation of aspirated oocytes for transfer and loading of the transfer catheter also were modified. After oocytes were selected on the basis of expansion of cumulus and corona cell layers, each of the most mature preovulatory oocytes (maximum of four oocytes per patient, except in one case of five oocytes by patient request) was manipulated by means of 8-gauge hypodermic needles to remove a large portion of the cumulus oophorus, routinely leaving a cumulus layer comprising no more than to 5 times the oocyte diameter. Our rationale for trimming the cumulus was to allow for loading oocytes into the transfer catheter in a smaller total volume (i.e., to 2 #L per oocyte) and to decrease the thickness of the barrier to penetration by spermatozoa. When loading the transfer catheter, one-half of the total (8 to #L) sperm volume was taken up after initially filling the catheter with FF. Next, all the selected and trimmed oocytes were loaded, the remainder of the sperm suspension was loaded, a 5-#L air space was added, and finally the end of the catheter was filled with 5 #L of FF. The composition of the culture medium in which the sperm were suspended, and other details of the loading procedure were identical to those described previously. When the catheter was loaded, it was threaded through a trocar cannula into the abdominal cavity as described previously; however, the catheter was held within the abdominal cavity for a minimum of 4 minutes before being passed into the ampullary region of the fallopian tube for deposition of the gametes. Vol. 48,, July 987 McGaughey and Nemiro Pregnancies with tubal transfer 99

3 Table Summary of Cycles, Laparoscopies, and Pregnancies Cycles Patients Cycles Patients a Our rationale was that the delay would provide a brief period for dispersal of the spermatozoa among the cumulus-enclosed oocytes and for the spermatozoa to begin penetrating the cumulus oo phorus while within the confines of the catheter. All follicular aspirations and all transfers were carried out by laparoscopy. The final modification was that all patients began treatment with 5-mg progesterone (P) suppositories vaginally every 2 hours o n the morning following TT. RIA and ultra sonography to monitor follicular development were performed as described in our first report. The criterion for biochemical pregnancy was elevated hcg (!-hcg > 5 miu/ml) at 4 days following TT; for clinical pregnancy, the criterion was the pres ence of a gestational sac(s) at 4 to 5 weeks after TT. All data were maintained on electronic spread sheets and analyzed by computer. Statistical com parisons were made by paired t-tests or by the use of 4-fold contingency tables,6 and data were fitted to curves by computer.7 RESULTS A summary of stimulation cycles, laparoscopies, and pregnancies in the present series is given in Table. Of the patients beginning stimulation cycles, 57% underwent laparoscopy, 26% exhibited elevated hcg after TT, and 7% had clinical preg nancies. For 2 patients who underwent laparos copy, no follicles were aspirated because of unsus pected ovarian inaccessibility; therefore, the inci- Clinical +hcg %" 27 9 b Percentage based on all stimulation cycles. Table 2 Pregnancies Laparoscopies Stimulations 44 %" 8 29 Percentage based on all laparoscopy cycles. dence of clinical pregnancy for all patients in which TT was accomplished was 2% (8 of 57 patients). I n Table 2, the numbers o f observed follicles, oocytes aspirated, and oocytes transferred are compared between cycles resulting in early preg nancy ( +hcg) and nonpregnant cycles. Although neither the mean number of follicles nor the mean number of oocytes aspirated differed for the two types of cycles, both the mean number of mature ' oocytes obtained and the mean number of oocytes transferred differed significantly (P <.5). The transfer cycles are described in Table in which outcome is tabulated according to number of oocytes transferred. Few pregnancies resulted with transfers of fewer than four oocytes (two clinical pregnancies out of 2 transfers). In contrast, 42% of transfers involving four oocytes produced clini cal pregnancy. The incidences both of elevated hcg (P <. ) and of clinical pregnancy (P <.5) were significantly higher for transfers involving four oocytes as compared with those involving less than four oocytes. One clinical pregnancy was lost to spontaneous abortion and two ectopic pregnan cies occurred in this series. Three sets of twins and one set of triplets occurred, representing 7% and 6% of the clinical pregnancies, respectively. The results are organized by method of ovarian stimulation in Table 4. Nearly 8% of cycles with CC alone failed to reach laparoscopy; however, of those that did, one-third resulted in clinical preg nancy. With the other three methods, more than half of the cycles resulted in laparoscopies. The method with CC and hmg (days 7, 9, and ) pro- Cycles Resulting in Pregnancy Versus Cycles Not Resulting in Pregnancy Oocytes aspirated Cycle type of cycles Follicles Total Pregnant Nonpregnant ±.5 7. ±.8 6. ± ±.7 Mature Oocytes transferred 5.2 ±.4a.4 ±.5.8 ±.. ±.2 mean ± SE a These values are significantly higher for pregnant than for nonpregnant cycles (P McGaughey and Nemiro Pregnancies with tubal transfer < a.5, paired t-tests). Fertility and Sterility

4 Table Number of Oocytes Transferred as Related to Outcome of Transfer of oocytes transferred of cycles hcg Clinical % % TP" 2 9 b Spontaneous abortion after determination of clinical preg nancy. Tubal ectopic pregnancy. duced the highest incidences of elevated hcg (6 % ) and clinical pregnancies (9 % ) based on transfer cycles. The method with hmg alone in cluded too few cycles for meaningful comparison with the other methods. The results also were analyzed according to diagnosis. No clinical pregnancies resulted from TT in the categories of anovulation ( transfers) or immunologic infertility (2 transfers). By compari son, TT was relatively successful for patients with pelvic adhesions (6 clinical pregnancies, 2 transfers), unexplained infertility ( 6 clinical preg nancies, 6 transfers ), endometriosis (2 clinical pregnancies, 7 transfers), and tubal disease (2 clin ical pregnancies, 6 transfers). Moderate success was obtained for patients with oligospermia ( clinical pregnancy, 6 transfers) o r cervl.cal factor ( clinical pregnancy, 7 transfers). The influence of semen sperm concentration, semen sperm motility, and sperm numbers and motility of insemination sperm were compared between cycles resulting in pregnancy and those that were unsuccessful (data not shown). The only finding from these analyses Table 4 of fetuses of pregnancies of semen and sperm parameters was that oligo spermia was related to reduced success for TT (see previous discussion). Cycles resulting in TT were classified according to their outcomes and are presented in Table 5. These data suggest that total E 2 (measured by RIA of serum samples taken just before injection of hcg) and E 2 levels per follicle (number of 8- to 2-mm diameter follicles determined by ultraso nography just before injection of hcg) were higher for pregnant ( +hcg) than for nonpregnant cycles, although the level of significance was marginal (P <. by paired t-tests). Although the mean num bers of preovulatory follicles and the mean total oocytes obtained in these two groups did not differ, the mean numbers of mature oocytes recovered differed significantly (P <.25). When clinical pregnancies were compared with biochemical preg nancies, the group with biochemical pregnancies exhibited significantly higher E 2 levels (both total, P <.; and per follicle, P <.25). Comparisons between clinical pregnancies and biochemical pregnancies indicated no significant differences Methods of Ovarian Stimulation Stimulation method" Pregnancies of cycles cc (-7) hmg (_.) cc (-7) hmg (-) cc (-7) hmg (7, 9, ) All methods Laparoscopies % %' % %' % of cycles CC is clomiphene citrate for cycle days to 7 (-7). hmg is human menopausal gonadotropin for cycle days to the time of full follicular development (-) or only on cycle days 7, 9, and (7, 9, ). Vol. 48,, July 987 Clinical +hcg b Percentage is based on cycles. Percentage is based on laparoscopies. McGaughey and Nemiro Pregnancies with tubal transfer

5 Table 5 Estrogen Levels, Follicles, and Oocytes for Transfer Cycles Estrogens (pg/ml) Per follicle b Total" Cycles (no.) Oocytes Follicles Total Mature mean ± SE All (59) Pregnant (+hcg) (27) Nonpregnant (-hcg) (2) Pregnant (Clinical) ( 8) Biochemical (+hcg) (9) ' ' d Total estrogen levels are based on RIA of samples taken immediately before injection of hcg. b Estrogen levels per follicle are based on total estrogen di vided by the number of 8- to 2-mm follicles observed by ultra sonography immediately before injection of hcg. Values that differ between pregnant and nonpregnant, and between pregnant (clinical) and biochemical ( +hcg). Compari sons by paired t-tests. ' P <.; dp <.25; ep <.. among numbers of follicles, total oocytes, or ma ture oocytes. Because of the significant correlation between numbers of mature oocytes and early biochemical pregnancies, the data were analyzed to determine whether numbers of mature oocytes might be pre dicted from numbers of follicles observed by ultra sound before laparoscopy. Pooled data were used for these analyses, and include data from all methods of ovarian stimulation (see Materials and Methods). These analyses showed that a curvilin ear relationship existed between follicle number and estrogen level, in the range of 2 to 6 follicles (coefficient of correlation,.999 by polynomial least squares curve fitting7). Similarly, the number of follicles determined by ultrasonagraphy corre lated well with the total number of oocytes aspir ated at laparoscopy (coefficient of correlation,.9987). In the range of 2 to 5 follicles, there was good correlation with numbers of mature oocytes collected (coefficient of correlation,.977 7) ; whereas for 6 and 7 follicles, the correlation was not as good. The data also were analyzed to determine whether either the E 2 levels or the levels of E 2 per preovulatory follicle, as measured before injection of hcg, were correlated with numbers of mature oocytes or with incidence of pregnancy. These analyses demonstrated that the numbers of mature oocytes recovered increased linearly with increased level of serum E 2 (coefficient of correlation,.8977). Pregnancies were distributed bimodally i n rela tionship to total serum E 2, with the highest inci dence of clinical pregnancy (7% of ten transfers) correlated with a mean serum E 2 level of 9 ± 8 (SE) pg/ml serum. At the higher levels of serum E 2 (i.e., 268 ± 2 pg/ml or higher), more than half of the early pregnancies were biochemical, without clinical evidence of pregnancy. The results of correlating E 2 /follicle with mature oocytes recovered and with pregnancies are shown in Figure. Increased numbers of mature oocytes were recovered with increasing E 2 in the range of 26 to 5 pg/follicle; whereas, at the highest E 2 level (789 ± 44 pg/follicle), the number of recov ered mature oocytes decreased. The majority of pregnancies occurred in cycles within the range of to 5 pg E 2 /follicle, and only about 2% of early pregnancies in that range failed to continue. As was also shown with the analysis of total serum E 2, the higher levels of E 2 /follicle were correlated with biochemical pregnancies only. In addition to the pregnancies shown in Figure, two tubal ec topic pregnancies occurred in this series. Both of these occurred in patients exhibiting high total serum E 2 (28 and 5 pg/ml, respectively) and high E 2 /follicle (566 and pg/follicle, respec tively). The data were analyzed further by grouping cycles according to numbers of follicles and ranges 2 McGaughey and Nemiro Pregnancies with tubal transfer.,_, Fertility and Sterility

6 Estradiol 5. / F o l l i c le. pg M! o Figure The correlation between levels of E2 per follicle and the number of mature oocytes recovered at laparoscopy (upper line graph). Vertical bars represent standard errors. The data were fitted to the curve by the method of polynomial least squares (coefficient of correlation,.978).7 The bar graph shows the incidences of biochemical pregnancy, as observed as positive hcg measured 4 days after TT (open portions of bars) and of uterine clinical pregnancy (cross-hatched portions of bars). Num bers above the bars represent transfer cycles. of E 2 levels per follicle. The results showed that nearly half of all clinical pregnancies (8 out of 8) occurred in cycles in which to 5 preovulatory fol licles were observed by ultrasound and the E 2 levels were in the range of to 499 pg/follicle. Eight clinical pregnancies (5% of transfers) and no bio chemical pregnancies occurred in that group of 5 cycles. Seventeen cycles with to 5 follicles but with higher E 2 levels (>499 pg/follicle) produced a high incidence of biochemical pregnancies (47% ), although half o f these did not continue o r resulted in tubal pregnancies (two ectopic pregnancies). In cycles with more than 5 preovulatory follicles, clin i al pregnancies (three out of eight transfers) oc curred only in the group with E 2 levels in the range of to 499 pg/follicle. Although the combined incidence of pregnancy for cycles with less than 6 follicles was higher than for cycles with more than 5 follicles, these differences were not statistically significant. The difference between incidence of clinical uterine pregnancy (6% of 6 cycles) in combined cycles with lower E 2 /follicle ( <5 pg/ follicle) as compared with that (% of 2 cycles) in combined cycles with higher E 2 /follicle (>499 pg/ Vol. 48,, July 987 follicle) was statistically significant (P <.5, by 4-fold contingency tests6). We also examined the possibility that LH levels, as measured before the administration of hcg, might be correlated with numbers of mature oo cytes aspirated or with the incidence of pregnancy following TT (Fig. 2). The levels of LH were not strongly correlated with the numbers of mature oocytes recovered at laparoscopy (data not shown). The LH levels were related to E 2 levels, which in creased with increasing LH (Fig. 2). These data clearly indicate that lower LH levels (range, 5 to 29 miu /ml serum) were correlated with relatively high incidences of both biochemical and clinical pregnancies. At the higher LH levels (range, to 56 miu/ml serum), biochemical pregnancy oc curred at a high incidence, although most of those early pregnancies did not result in clinical pregnancy. DISCUS SION The current series of 59 TT procedures produced 27 cases of elevated hcg and 8 clinical pregnan cies. By comparison, our first series of 59 transfers resulted in 8 biochemical pregnancies and 2 clin ical pregnancies. These results represent improve ments of 5% both for elevated hcg and for clini cal pregnancy for the second series. We ascribe this substantial improvement to procedural modifica tions (see Materials and Methods) and to rigorous selection of stimulation cycles. Although the inci dences of pregnancy were improved in the current N s 25 E :> 2 ; 7,.u 5... ''"'' e :2 " C 8) u -=o w ( 4) L u t e i n i z i n g Hormone, m i U ml / 8 7 Figure 2 The correlation between LH levels and total E2 levels (upper line graph). Vertical bars represent standard errors. The data were fitted to the curve by the method of polynomial least squares (coefficient of correlation,.88).7 The description of the bar graph is the same as that in Figure. McGaughey and Nemiro Pregnancies with tubal transfer

7 series, the incidence of multiple pregnancy was decreased by about one half from 55% (5 out of 9 continuing pregnancies) in the first series to 27% (4 out of 5 continuing uterine pregnancies) in the current series. Tubal ectopic pregnancy occurred in two cases in the current series. In one, the patient had previously undergone bilateral neosalpingostomies with the remaining endosalpinx described as grossly normal. The other patient had a known diagnosis of pelvic adhesions in which one tube was described as appearing completely normal and the other as a hydrosalpinx. Prior to TT, both patients had documented tubal patency of at least one fallopian tube, and were advised of the increased risk of tubal pregnancy. In our program, these two are the only cases of tubal pregnancy out of 276 cycles of ovarian stimulation (.7% of cycles). The incidence of tubal pregnancy among combined transfer cycles (8 cycles) in our first two series was.7%, or 4.4% (2 of 45) of all detected pregnancies. The risk of tubal ectopic pregnancy in our TT program, therefore, is not higher than that reported for IVF ET. 8 Both ectopic pregnancies were detected early and treated surgically before rupture. It is estimated that any patient undergoing tubal surgery when pelvic infection is the suspected etiology has at least a % risk that a resulting pregnancy will be located in the fallopian tube.s- We do not believe that TT, in cases of tubal disease, predisposes the patient to an increased risk of tubal pregnancy as compared with similar patients in which pregnancy occurs without TT. With TT, the occurrence of a tubal pregnancy most likely is the result of abnormal embryo transport to the uterus following deposition of the gametes into the ampullary region of the oviduct. In contrast, with IVF-ET, in which cleaved embryos are deposited into the uterine lumen, most cases of ectopic pregnancy are thought to result from retrograde transport of embryos through the uterotubal ostium or from accidental deposition of embryos into the fallopian tubes during uterine ET.8 It is clear that, in cases of TT for patients with known tubal disease, all resulting pregnancies must be monitored carefully because of this risk. Of 5 continuing pregnancies in the current series, 27% were multiple as compared with 55% multiple pregnancies in our first series. Although the incidence of multiple pregnancies has been reduced, perhaps by transferring a maximum of four oocytes in nearly all cases in the second series, multiple pregnancy occurs at high incidence with TT. It should be noted that all nine pregnant patients from our first series have delivered normal babies. These normal births include four sets of twins and a set of quadruplets that was delivered at 2 weeks. The set of triplets in the current series (see Table ) also was delivered uneventfully at 4 weeks with the birth of three normal baby girls. A complete clinical evaluation of all completed pregnancies is in preparation as a separate study. As with transfer of multiple embryos in IVF-ET, the transfer of multiple oocytes in TT improves the incidence of pregnancy while also increasing the risk of multiple pregnancy. 2 The combined results of TT, including our first series and the current series, indicate that the highest success rates were with subsets of patients whose primary diagnosis was either pelvic adhesions (4% of 22 transfers resulted in clinical pregnancy) or unexplained infertility (% clinical pregnancies from 27 transfers). These high success rates indicate the possibility that many cases of infertility due to pelvic adhesions or to unknown causes may result from failure of fallopian tubes to pick up ovulated oocytes or of faulty sperm transport within the fallopian tubes. With TT, oocytes and washed spermatozoa are placed together within the natural environment of the oviductal ampulla, and these possible failures are overcome. Of three transfers involving patients with antisperm antibodies of maternal or paternal origin, one clinical pregnancy resulted. Although these numbers are small, the pregnancy indicates that TT is an appropriate treatment for such cases. It is noted that, in cases of suspected or documented maternal antisperm antibodies, preparation of sperm for TT is accomplished with culture medium containing pooled serum from other patients. In addition, semen samples obtained from patients suspected or known to produce antisperm antibodies are collected by ejaculation into a sample cup containing ml of culture medium to ensure rapid dilution of antibodies if present. Future treatments by TT of more patients with immunologic infertility are necessary to more accurately assess its efficacy. All diagnostic classes now include at least one clinical pregnancy following treatment with TT. Although no patient with endometriosis became pregnant after TT (ten transfers) in our first series, the current series produced two clinical pregnancies out of 7 transfers with patients exhibiting that disease. The combined numbers of patients whose primary diagnosis was cervical factor 4 McGaughey and Nemiro Pregnancies with tubal transfer Fertility and Sterility

8 ( transfers, 8% clinical pregnancy) or oligospermia (9 transfers, % clinical pregnancy) are quite small, but these two categories represent the lowest success rates by diagnosis in the combined first and second series of our TT program. Our technical refinement of the TT procedure has allowed an evaluation of endocrine parameters and parameters related to oocyte collection, and to correlate these with the incidence of success for the procedure. In this study, we have demonstrated that high incidences of clinical pregnancy are correlated with intermediate levels of E2, with the ultrasonographic observation of to 5 developing follicles, with the recovery of four or more mature preovulatory oocytes, and with the transfer of four or more oocytes. The former two parameters are determined before laparoscopy and their values are strongly predictive of the latter two parameters. Ideally, therefore, to obtain maximal incidence of clinical pregnancy, cycles with E2 levels outside the optimal range ( to 5 pg/ml serum/follicle) or in which insufficient numbers of follicles develop (below the to 5 range) should be cancelled before surgery. Clinically, however, such rigorous criteria are unrealistic for the heterogeneous population of infertile patients in our TT program. Many patients undergo several stimulation cycles (with the same or different stimulation protocols) without obtaining an optimal ovarian response, as also has been described for patients in programs of IVF ETP In the presently reported series of TT patients, only 5 cycles strictly fit the previously described criteria (% of 9 stimulation cycles), and these few cycles accounted for half of the uterine clinical pregnancies. Clearly, eight additional uterine clinical pregnancies occurred in patients whose cycle criteria were less than optimal. Previous reports from IVF-ET programs have demonstrated a correlation between estrogen levels and numbers of developing follicles.4-6 The present study demonstrates the additional possibilities for predicting the numbers of mature oocytes to be obtained at laparoscopy as well as the outcome of TT, depending on the combined criteria of follicle number and estrogen level. The results of the present study also suggest that very high estrogen levels (as well as high LH levels), although consistent with high incidences of early biochemical pregnancy, are inconsistent with high incidences of continuing uterine pregnancy. Negative influences of high estrogen levels and of high baseline LH levels on pregnancy have been discussed previouslyp-9 Abnormally high estro- gen levels might be expected to interfere with uterine development for normal embryo implantation, 8 or may adversely affect developing embryos directly. Although the present study suggests strong correlations between endocrine and ultrasonographic criteria and the incidence of success for TT, we realize that the establishment of generally applied criteria for cycle cancellation will require confirmation of our observed correlations with larger patient populations. In addition, the search for better protocols and drugs for ovarian stimulation may modify our observed optimal ranges regarding these criteria. Based upon the data analyses reported here, we have modified our TT program to make every possible effort to obtain four or more mature oocytes for transfer. In those cases in which the likelihood of obtaining optimal numbers of mature oocytes at laparoscopy is low, patients are informed of the reduced expectation for success before surgery. REFERENCES. Nemiro JS, McGaughey RW: An alternative to in vitro fertilization-embryo transfer: the successful transfer of human oocytes and spermatozoa to the distal oviduct. Fertil Steril 46:644, Asch RH, Ellsworth LR, Balmaceda JP, Wong PC: Pregnancy after translaparoscopic gamete intrafallopian transfer. Lancet 2:4, 984. Asch RH, Balmaceda JP, Ellsworth LR, Wong PC: Gamete intrafallopian transfer (GIFT): a new treatment for infertility. Int J Fertil :4, Guastella G, Comparetto G, Gullo D, Palermo R, Venezia R, Cefalu E, Ciriminna R, Salerno P, Cittadini E: Gamete intrafallopian transfer (GIFT): a new technique for the treatment of unexplained infertility. Acta Eur Fertil 6:, Seppala M: The world collaborative report on IVF-ET: Current state of the art in January, 984. Ann NY Acad Sci 442:276, Mainland D, Murray IM: Tables for use in fourfold contingency tests. Science 6:59, Corl T, Warme PK: Curve Fitter-PC. Interactive Microware, Inc., State College, P A, Martinez F, Trounson A: An analysis of factors associated with ectopic pregnancy in a human in vitro fertilization program. Fertil Steril 45:79, Wood C, Downing B: In-vitro fertilization and tubal microsurgery-their status compared. Br J Obstet Gynaecol 9:, 986. Paterson PJ: Indications for the treatment of tubal infertility patients by microsurgery or in vitro fertilization. Aust NZ J Obstet Gynaecol 24:262, 984. Hewitt J, Martin R, Steptoe PC, Rowland GF, Webster J: Bilateral tubal ectopic pregnancy following in-vitro fertilization and embryo replacement: case report. Br J Obstet Gynaecol 92:85, 985 Vol. 48,, July 987 McGaughey and Nemiro Pregnancies with tubal transfer 5

9 2. Muasher SJ, Garcia JE, Rosenwaks Z: The combination of follicle-stimulating hormone and human menopausal gonadotropin for the induction of multiple follicular maturation for in vitro fertilization. Fertil Steril 44:62, 985. Lyles R, Gibbons WE, Dodson MG, Poindexter AN, Young RL, Rossavik IK, Findley WE: Characterization and response of women undergoing repeat cycles of ovulation induction in an in vitro fertilization and embryo transfer program. Fertil Steril 44:82, Acosta AA, Jones GS, Garcia JE, Sandow B, Veeck L, Mantzavinos T: Correlation of human menopausal gonadotropin/human chorionic gonadotropin stimulation and oocyte quality in an in vitro fertilization program. Fertil Steril 4:96, Vargyas JM, Morente C, Shangold G, Marrs RP: The effect of different methods of ovarian stimulation for human in vitro fertilization and embryo replacement. Fertil Steril 42:745, Botero-Ruiz W, Laufer N, DeChemey AH, Polan ML, Haseltine FP, Behrman HR: The relationship between follicular fluid steroid concentration and successful fertilization of human oocytes in vitro. Fertil Steril 4:82, Dirnfeld M, Lejeune B, Camus M, Vekemans M, Leroy F: Growth rate of follicular estrogen secretion in relation to the outcome of in vitro fertilization and embryo replacement. Fertil Steril 4:79, DeChemey AH, Tarlatzis BC, Laufer N: Follicular development: lessons learned from human in vitro fertilization. Am J Obstet Gynecol 5:9, Stanger JD, Yovich JL: Reduced in-vitro fertilization of human oocytes from patients with raised basal luteinizing hormone levels during the follicular phase. Br J Obstet Gynaecol 92:85, McGaughey and Nemiro Pregnancies with tubal transfer Fertility and Sterility