Microscopic versus macroscopic tubal anastomosis in rabbit fallopian tubes

Transcription

1 FERTILITY AND STERILITY Copyright 1983 The American Fertility Society Vol. 40, No.3, September 1983 Printed in U.8A. Microscopic versus macroscopic tubal anastomosis in rabbit fallopian tubes James M. Goldfarb, M.D.*t Wulf H. Utian, M.D., Ph.D. * Robert Weiss, M.D.+ The Mt. Sinai Medical Center and Case Western Reserve University, Cleveland, Ohio The importance of the operating microscope in rabbit tubal anastomosis was assessed by operating on 36 rabbits, halfwith the aid of the operating microscope and half with the aid of low-magnification loupes. The surgeons' operating comfort and sense of assuredness that the anastomoses were being done properly were increased by the operating microscope. However, the outcome of the anastomoses as measured by pregnancy rates, adhesion scores, and microscopic pathologic findings were not significantly different in the two s. Fertil Steril40:373, 1983, " l- r, ~ i :y The use of the operating microscope for infertility surgery, particularly in reversal of previous sterilization, has generated considerable interest. Nonetheless, despite an increase in success of tubal anastomosis concurrent with the use of the operating microscope, the contribution of the microscope itself is less clear. Concurrent variables in surgical technique such as selection of suture material, microcauterization for hemostasis, and the use of ancillary methods for adhesion prevention may partially account for the enhanced fertility rates. Moreover, the number of tubal anastomoses being performed has increased greatly with the liberalization of sterilization regulations. Thus, the procedures are being done by surgeons with increasing experience. Advocates argue that meticulous attention to detail and two-layer suturing during Received December 15, 1982; revised and accepted April 28, *Department of Obstetrics and Gynecology. treprint requests: James M. Goldfarb, M.D., Chief, Division of Gynecology, The Mt. Sinai Medical Center, University Circle, Cleveland, Ohio *Department of Pathology. the procedure are possible only with the microscope, and that it is therefore essential to a successful outcome. 1 Others, however, feel that use of the microscope tends to prolong the procedure and add refinements that may not increase the success rate of anastomosis. For all these reasons, the debate continues as to whether there is a clear advantage in the use of the operating microscope for tubal anastomosis. In an attempt to clarify the situation, a randomized study was undertaken in 36 rabbits to compare' fertility rates, patency rates, adhesion formation, and pathologic findings following tubal anastomosis with and without the operating microscope. Operating times were also compared. The operating physicians in this study were Drs. Goldfarb and Utian, both highly experienced in microsurgery, having been involved with animal research and active clinical microsurgical programs for 5 and 9 years,respectively. MATERIALS AND METHODS Mature female New Zealand White rabbits weighing between 2800 and 3600 gm were uti- ity Vol. 40, No.3, September 1983 Goldfarb et al. Microscopic versus macroscopic rabbit anastomosis 373

2 lized for the study. They were housed individually for 7 to 10 days prior to surgery. At the time of arrival they were randomized into "microsurgical" and "macrosurgical" s, thereby allowing each surgeon to operate on nine rabbits with the aid of high magnification provided by a Zeiss operating microscope (Carl Zeiss, Inc., New York, NY) with pedal operated zoom and focus, and on nine rabbits using low-power magnification of2 x to 2% x provided by loupes (Opti-vision model DA-5, Donegan Optical Company, Lenox, KS). The animals were given Surital (Parke-Davis, Morris Plains, NJ) 35 mg/kg, injected into an ear vein to induce anesthesia and then additional Surital as needed during the procedure to maintain anesthesia. The rabbits were shaved and prepared for abdominal surgery, and the instruments were sterilized; but strict adherence to aseptic technique was not complied with. The rabbits were opened through a midline incision. The fallopian tubes were identified and a segment of ampulla ~ 1 cm in length was excised. The contralateral fallopian tube was not operated on, so as to act as an internal control. At this point, the operating microscope or loupes were employed. Hemostasis at the excision site was attained with the use of unipolar and bipolar microcautery. A temporary 2-0 Prolene stent was then inserted, and the'mesosalpinx was approximated with 6-0 Vicryl suture to lessen tension at the anastomosis site. The anastomosis was then done in a two-layer technique using 8-0 Vicryl on a BV-3 needle (Ethicon, Inc., Somerville, NJ). Four sutures were placed in the muscularis while an attempt was made to exclude the mucosa. Following this, the stent was removed, and the sutures were tied. Serosa sutures were then placed, taking care not to distort the suture line. Irrigation with Ringer's lactate solution was used throughout the procedure to keep tissues moist and the field clear. Following the anastomosis, all blood clots were removed from the operative field. Prior to closure of the peritoneal ca vi ty, ~ 20 ml of 6% dextran 70 (Pharmacia, Inc., Piscataway, NJ) was instilled into the peritoneal cavity. The abdominal wall was then closed in a two-layer closure with 1-0 chromic suture. Three weeks following the procedure the animals were mated on three occasions with three male rabbits of known fertility. All female rabbits were mated with the same three bucks. At least 1 week after the final mating, the animals were sacrificed. At the time of autops, adhesion and fertility rates were assessed. Adhesions were graded according to the following scale: (1) thin, filmy adhesions involving only the operated tube; {2) thick adhesions involving only the operated tube; and (3) adhesions between the operated tube and the contralateral tube or other organs. Fertility was assessed by counting the number of embryos in each uterine horn. If there were no embryos visually, the uterine horn was opened to document the absence of embryos. Operated tubes that did not have pregnancies were checked for patency by an injection of indigo carmine and then removed and sent for pathologic study. Specimens were fixed in 10% neutral buffered formalin. The regions 1 cm proximal and distal to the suture site were excised and sectioned transversely for histologic study. Paraffin sections were stained with hematoxylin and eosin (H&E). The degree of inflammation and fibrosis was graded arbitrarily as mild, moderate, or severe, depending on the relative area involved and the intensity of the cellular reaction (Figs. 1 and 2). The surgeon at the time ofthe autopsy did not know whether the procedures had been done with Figure 1 Minimal host response to suture material (H&E, original magnification, x 250). 374 Goldfarb et al. Microscopic versus macroscopic rabbit anastomosis Fertility and Sterility

3 Table 1. Pregnancy Rates Microscopic Macroscopic No. of rabbits mated No. of rabbits pregnant in operated side 14 (82%) lo (67%) al Figure 2 Moderate chronic inflammatory response to suture material (H&E, original magnification, x loo). or without the aid of the microscope. Likewise, the pathologist did not know the type of operation performed on the tubes under examination. RESULTS Of the 36 rabbits operated upon, 3 died before the end of the experiment: 1 death was due to anesthesia, and 2 animals died 10 and 12 days postoperatively of unknown causes. Seventeen of the 33 rabbits that survived to the conclusion of the experiment were operated upon with the use of the microscope (microscopic ), and 16 were operated upon with the loupes (macroscopic ). One rabbit in the latter was not mated because of an error in the mating schedule. Fourteen (82%) of the 17 rabbits in the microscopic became pregnant in the operated side. Ten (67%) of the 15 rabbits in the macroscopic became pregnant in the operated side (Table 1). This difference is not statistically significant by chi-square analysis. None of the rabbits in either that failed to conceive in the operated side had pregnancies in the unoperated side. Two rabbits in the microscopic conceived in the operated side but not in the unoperated side. All embryos were apparent in the intact uterine horn; there were no additional embryos found when the uterine horns were opened. The 14 rabbits in the microscopic that conceived had a total of 66 pregnancies in the operated sides (average, 4.7 per side) and 53 pregnancies in the unoperated sides (average, 3.8 per side). The 10 rabbits in the macroscopic that conceived had a total of 42 pregnancies in the operated sides (average, 4.2 per side) and 51 pregnancies in the unoperated sides (average, 5.1 per side) (Table 2). There was no statistical difference in the number of pregnancies in the operated and unoperated sides of either. Five of the 17 rabbits in the microscopic and 7 of 16 in the macroscopic had adhesions visible to the naked eye. The characteristics of these adhesions are listed in Table 3. All three ofthe nonpregnant operated tubes in the microscopic were fully patent. Five of the six nonpregnant operated tubes in the macroscopic were fully patent. The other was completely blocked. The tube that was not patent showed extrusion of mucosa at the anastomosis site by gross and microscopic examination (Fig. 3). Thus, the overall patency rates in the microscopic and macroscopic s were 100% and 94%, respectively, there was no statistical difference. The microscopic appearance and the macroscopic appearance of the nonpregnant tubes can be found in Table 4 and can be seen to be similar in the two s. The operating time for the microscopic averaged 32 minutes per rabbit, and the operating time for the macroscopic averaged 26 minutes per rabbit. Table 2. Total Number of Pregnancies Microscopic Macroscopic No. of Total no. of rabbits with. pregnancies pregnancies in operated side Total no. of pregnancies in unoperated side (average, 53 (average, 4.7) 3.8) lo 42 (average, 51 (average, 4.2) 5.1) lity Vol. 40, No.3, September 1983 Goldfarb et ai. Microscopic versus macroscopic rabbit anastomosis 375

4 Table 3. Gross Appearance of Tubes at Autopsy Total no. of Grade of adhesions No. of rabbits with rabbits adhesions I II III Microscopic Macroscopic DISCUSSION The importance of the operating microscope for tubal anastomosis is still unclear. Hedon et al. 2 reported in 1980 that there was no significant difference in anastomosis of rabbit tubes done with the operating microscope, compared with those done with loupes. However, this was the author's first experience with use of the microscope. Additionally, the fallopian tubes were simply cut and then anastomosed with a one-layer closure. All of these factors could tend to bias the results against the use of the microscope. The current study was undertaken in an attempt to negate these factors by having the surgery done by two experienced investigators. Also, rather than simply cutting the tubes, a segment of tube was removed in order to more closely simulate a posttuballigation anastomosis. Finally, the tubes were anastomosed with a two-layer technique. Despite these modifications, the current study indicates that tubal patency and pregnancy following anastomosis of rabbit fallopian tubes is not significantly different whether minimal magnification or the operating microscope is used. It might be argued that a larger experimental is needed to show a statistical difference when both s show a relatively high pregnancy rate. While this may be true, it can also be argued that based on the data from this experiment, there may be even less of a potential difference between the macrosurgical and microsurgical than is suggested by the pregnancy rates in the two s, because in both s there were no pregnancies in the unoperated side in those rabbits that did not have pregnancies in the operated side. Thus, the little differences in pregnancy rates could be largely caused by a difference in the rate of successful matings rather than the surgery. It should be noted that adhesion formation and microscopic findings were similar in both ~. In both s the adhesion rate was small. Six percent dextran 70 was instilled intraperitoneally prior to closure of the peritoneum based on previous work that indicated pregnancy rates and possibly adhesion formation were favorably influenced by its use. 3 It is obviously very difficult to extrapolate this data to human subjects. The rabbit ampulla has a thin muscularis layer and a lumen of - 2 mm. Most tubal anastomoses in humans involve the isthmic portion of the fallopian tube. The muscularis of the human isthmus is approximately ten times thicker than the rabbit ampulla, and the lumen of the isthmic portion of human fallopian tubes is - 1 mm. These physical parameters may alter the healing characteristics. Additionally, the cause of failed anastomosis tends to differ in rabbit and humans. In women, failure of a reversal of sterilization is usually due to compromise in the blood supply of the cut ends of the tube or to technical difficulty resulting in inadequate approximation of luminal orifices. In contrast, most failures of anastomosis in rabbits are due to adhesion formation, as is shown by this study. Because of these differences, we do not advocate abandonment of the operating microscope for human tubal anastomosis. Figure 3 Extrusion of fallopian tube mucosa through the anastomosis (H&E, original magnification, x 250). 376 Goldfarb et al. Microscopic versus macroscopic rabbit anastomosis Fertility and Sterility

5 ld u- is,a n. le u- ~n b.e in iy ly, in lrin to lp )st lelse m an Table 4. Microscopic Findings in Nonpregnant Tubes n None Macroscopic 5 1 Microscopic 3 2 With experience, the physician can operate almost as rapidly with the microscope, as is shown by the mean operating times in this study. We think that in both human and animal subjects, operating with the microscope aids in reassuring the surgeon that the anastomosis is being done accurately. Additionally, in human microsurgery, a major reason for using high magnification is to examine the tissues to ensure that they are healthy. It should be stated, however, that there are several reports of small series of human subjects showing equivalent results with and without the microscope. Jones and Rock 4 reported a series of 12 patients for whom they did tubal anastomosis macroscopically. Nine patients had intrauterine pregnancies following the surgery, and one pregnancy was ectopic. Williams 5 reported a 72% pregnancy rate in 25 patients following macroscopic isthmic anastomosis. Henderson 6 compared 23 patients who underwent anastomosis, 11 undergoing a microsurgical procedure performed by himself and 12 undergoing a macrosurgical procedure done by others. He found that there was no advantage to the use of the microscope in this small of patients. While this debate continues, it is important to keep in mind that there are other factors that may have more impact on the success rate of tubal anastomosis. There seems to be little question that the remaining length of tube greatly influences the success rate. Silber and Cohen 7 showed no pregnancies in seven patients in whom the longest tube was < 3 cm and a 100% pregnancy rate when the longest tube was > 4 cm. In rabbits, McComb and Gomel 8 have shown that decreased tubal length leads to a lower pregnancy rate following anastomosis, while Eddy et al. 9 and Winston et al. 10 have shown that resection of a small amount of rabbit tube followed by careful anastomosis results in no decreased fertility. In addition to the length of remaining tube, there Inflammation Mild 4 0 Fibrosis Moderate None Mild Moderate seems to be little question that the use of fine synthetic suture materials with little tissue reaction has favorably affected the success of anastomosis, as has the microsurgical philosophy of fine instrumentation, meticulous hemostasis, and careful tissue handling. As has been stated, we are continuing to employ the operating microscope because of the greater assuredness that the anastomoses are being done correctly and because of the significant limitations in extrapolating this rabbit data to human subjects. Additionally, the versatility of the operating microscope allows the surgeon greater comfort than does the use of the loupes, with their fixed focal length. REFERENCES 1. Gomel V: Microsurgical reversal of female sterilization: a reappraisal. Fertil Steril 33:587, Hedon B, Wineman M, Winston RML: Loupes or microscope for tubal anastomosis? An experimental study. Fertil Steril 34:264, Utian WH, Goldfarb JM, Starks GC: Role of dextran 70 in microtubal surgery. Fertil Steril 31:79, Jones HW Jr, Rock JA: On the reanastomosis offallopian tubes after surgical sterilization. Fertil Steril 29:702, Williams EA: Aspects offallopian tube surgery. In Recent Advances in Obstetrics and Gynaecology, Vol 12, Edited by J Stallworthy, G Bourne. London, Churchill Livingstone, 1977, p Henderson SR: Reversal of female sterilization: comparison of microsurgical and gross surgical techniques for tubal anastomosis. Am J Obstet Gynecol 139:73, Silber SJ, Cohen R: Microsurgical reversal of female sterilization: the role of tubal length. Fertil Steril 33:598, McComb P, Gomel V: The influence of fallopian tube length on fertility in the rabbit. Fertil Steril31:673, Eddy CA, Antonini R Jr, Pauerstein CJ: Fertility following microsurgical removal of the ampullary-isthmic junction in rabbits. Fertil Steril 28:1090, Winston RML, Frantzen C, Oberti C: Oviduct function following resection of the ampullary-isthmic junction. Fertil Steril (Abstr) 28:284, 1977 o.osis erility Vol. 40, No.3, September 1983 Goldfarb et al. Microscopic versus macroscopic rabbit anastomosis 377