Creation of myelomeningocele in the fetal rabbit.

OBJECTIVE: Myelomeningocele is a neural tube defect resulting in an exposed spinal cord, which leads to irreversible neurologic damage at birth. We proposed development of a fetal rabbit model of myelomeningocele to study in utero spinal cord injury and repair strategies. METHODS: New Zealand white rabbits (n = 10) at 22 days of gestation (term = 31 days) underwent laparotomy to expose the gravid uterus; a hysterotomy exposed the fetal hindlimbs and back. A three to four level lumbar laminectomy was performed, and the dura over the posterior spinal cord was removed. At 30 days of gestation, the does underwent C-section for fetal harvest, and total fetal number, length, weight, and the presence or absence of a spinal defect were recorded for all viable fetuses. RESULTS: All injured fetuses were smaller and weighed less than the nonoperated littermate controls, and histologic examination confirmed a spina bifida-like lesion of their spinal cords. CONCLUSIONS: We successfully created an exposed spinal cord defect in the fetal rabbit model similar to the lesion found in humans. Advantageous because of low animal cost, relatively large fetal size, multiple fetuses per pregnancy, and short total gestation, this model will allow us to study the mechanism of injury to the exposed spinal cord, and perhaps develop strategies to repair human myelomeningoceles.

Tissue removal utilizing Steiner Morcellator within a LapSac: effects of a fluid-filled environment.

BACKGROUND AND PURPOSE: Tissue removal can be a simple process of withdrawal of the entire organ, piecemeal removal with surgical clamps, or mechanical morcellation. Different mechanical morcellators exist that each have advantages and disadvantages. We have investigated a particular morcellator having an internal mechanized blade system that increases the chances of damage to tissue isolation sacks but removes large volumes of intact organ that can more readily be evaluated histologically. The primary premise of this investigation is that a fluid-filled sack would be less likely to be damaged by the activated blades of the morcellator. MATERIALS AND METHODS: Utilizing a Steiner Morcellator (Karl Storz, Culver City, CA), two porcine kidneys were morcellated within the large LapSac (Cook Urological, Spencer, IN). Two environmental variables were evaluated: dry sac morcellation and fluid-filled sac morcellation. Each session was timed, fluid leakage identified, grasping of the sacks quantified, and gross spillage noted. The tissues were submitted for pathologic evaluation to quantify any differences grossly or histologically. All LapSacs were inspected for gross violation and inflated to distention with fluid to check for tiny leaks. RESULTS: The Steiner Morcellator worked much better within the confines of the LapSac filled with fluid. There were no perforations in our experimental setting. It was not possible discern use of fluid-filled sacks histologically. CONCLUSIONS: The Steiner Morcellator can be utilized safely in the LapSac if cautious observation and fluid-filled sack conditions are maintained. The extracted tissue is easily evaluated histologically.

Running suturing: the ideal length facilitates this task.

BACKGROUND AND PURPOSE: Laparoscopic intracorporeal suturing is difficult, a complex task involving several integrated skills such as needle handling, suturing, and knotting. Running suturing is even more complex in the closed environment secondary to the angles of the suture lines, the tension maintained on the suture line, and the need to secure the ends, including tying a knot from the tail of the suture to the loop of the preceding stitch. We have hypothesized that the ideal suture length facilitates this process, and this study was specifically designed to determine the ideal suture length for an intracorporeal running suture. MATERIALS AND METHODS: Latex phantoms were incised for 1 or 2 cm, and each was marked with black points to indicate suture entrance and exit sites. These marks were placed 2 mm from the incision, starting 2 mm proximally and ending 2 mm distally. Suture lengths could then be varied in direct proportion to the length of the incision, keeping all of these points as constants. The ratios of suture length:incision length were 9:1, 10:1, and 11:1. One surgeon performed all of the running suturing utilizing dry 3-0 coated polyglactin with a tapered SH needle. The times needed to create running suture lines were recorded (seconds), the number of ideal entrance and exit points tabulated, and the number of technical errors (missed movements resulting in prolonged suturing) recorded. RESULTS: The lengths of both the incision and the suture affect the ability to perform laparoscopic intracorporeal running suturing. As the incision increased, the suture:incision ratio of 9:1 resulted in the most efficient results (quickest suturing and fewest errors). In smaller suture lines (1 cm), the ratios 10:1 and 11:1 appeared better. CONCLUSIONS: A suture:incision ratio of 9:1 is best for longer suture lines and 10:1 is better for short suture lines.

Laparoscopic augmentation cystoplasty using the novel biomaterial Surgisis: small-intestinal submucosa.

BACKGROUND AND PURPOSE: Urinary bladder augmentation is indicated for diverse conditions, including neurogenic bladder, cancer resection, spinal cord injury, and congenital anomalies. The ideal cystoplasty material is yet to be described. Native gastrointestinal segments commonly used are limited by leakage and small-bowel obstruction, metabolic/nutritional abnormalities, calculi, and malignancy. This study assessed laparoscopic bladder augmentation with porcine small intestinal submucosa (SIS). MATERIALS AND METHODS: Five female pigs (<25 kg) were prepared for surgery under general anesthesia. After Veress needle insufflation, a main 10-mm trocar was placed in the midline for the laparoscope, with two lateral 10-mm ports added for operative instruments. The bladder dome was incised, and a patch of SIS was sewn into the bladder using running 2-0 Vicryl. Three animals served as technical studies. Two additional sows underwent long-term survival surgery: one undiverted and one diverted via a Stamey suprapubic catheter. RESULTS: There were no operative losses. The mean operative time was 140 minutes. The SIS graft held the sutures without tearing. Laparoscopic survey revealed no urine leaks at bladder closure. All five animals voided postoperatively. Urinary extravasation was evident in the three undiverted technique animals. In the other two sows, cystoscopy at 7 days showed intact suture lines without evidence of urinary extravasation and with normal vesicular volumes. Tissue growth was evident, but the graft margins were still discernible. CONCLUSIONS: Laparoscopic bladder augmentation was possible using SIS but at minimal volumes. There were no operative complications; however, the material was difficult to deploy and may benefit from application of an absorbable scaffold. Postoperative urinary drainage is necessary. Further studies will optimize the graft configuration for maximal augmentation.

Experimental study of umbilical cord length as a marker of fetal alcohol syndrome.

Umbilical cord length has long been investigated as a potential marker of intrauterine events that may place the neonate at risk for future adverse developmental sequelae. Experimentally, significantly shortened cords have been reported in association with prenatal exposure to common drugs of abuse. This study in rats reports the time course of effects on umbilical cord length of a daily maternal ethanol gavage (3,200 mg/kg) from gestational day 6 through termination of pregnancy at either day 17, 18, 19, or 20. A total of 786 fetuses derived from 60 litters were examined. Control fetuses demonstrated a linear increase in umbilical cord length and body weight gain during late gestation, findings that support previous studies. The body weights of the ethanol-exposed fetuses were reduced significantly on all gestational days examined, indicating intrauterine growth retardation, a characteristic of fetal alcohol syndrome. Similarly, acute fetal akinesia as well as long-term sequelae stemming from impaired neurological development would result from the elevated blood ethanol levels achieved in this study. The umbilical cords of ethanol-exposed fetuses were significantly shorter on gestational days 19 and 20 in comparison to their controls, while cord lengths on days 17 and 18 were not shortened significantly. A stretch hypothesis has been proposed suggesting that the degree of fetal activity is the main determinant of umbilical cord length. In rats, there is a physiologic diminution of the volume of amniotic fluid (oligohydramnios) in late gestation (day 19 to term), which restricts fetal movements but does not appear to alter the linear relationships between gestational age and cord length in controls, thus arguing against the stretch hypothesis. However, cord lengths in the ethanol-exposed fetuses plateaued in late gestation, suggesting possible adherence to a stretch hypothesis. This dichotomy is discussed emphasizing fetal growth and activity as well as intrauterine space.

Experimental utilization of the holmium laser in a model of ureteroscopic lithotripsy: energy analysis.

BACKGROUND AND OBJECTIVE: Medical lasers have rapidly expanded in both indications and utilization. We have developed a simple model for the investigation of various modalities of lithotripsy using readily available silicate stones of uniform mass. MATERIALS AND METHODS: The holmium:YAG VersaPulse Select (Coherent, Palo Alto, CA) was used in these experiments to define the relation between fiber diameter and efficacy of stone fragmentation. For each fiber, lithotripsy was performed in an incremental fashion at 0.6, 0.8, and 1.0 J at a frequency of 10, 16, and 20 pulses per second at each energy level. RESULTS: Total kilojoules did not differ between any fibers investigated, supporting the consistency of our methodology. The power density of the holmium laser energy, as expressed as total kJ/area, decreases in proportion to increasing fiber diameter. We expected an increase in fragmentation success as fiber diameter decreased (at equal energy output). However, peak lithotripsy occurred with the 365- and 550-microm fibers, whereas neither the 200- nor the 1000-microm fibers was effective. CONCLUSION: This model utilizing a silicate stone phantom supports our clinical observation that the 365-microm fiber (and additionally the 550-microm fiber) provides the best method for efficient intracorporeal lithotripsy.

New visualization techniques for in utero surgery: amnioscopy with a three-dimensional head-mounted display and a computer-controlled endoscope.

Endoscopic fetal surgery may reduce preterm labor associated with open hysterotomy but is partially limited by current visualization technology. We investigated a three-dimensional (3D) imaging system coupled to a head-mounted display (3D-HMD) and also employed a computer-controlled zoom endoscope for noninsufflated amnioscopy. Pregnant sheep were prepared in aseptic fashion for general anesthesia. Uterine access was obtained following maternal laparoscopy. A 10-mm zoom endoscope (Vista Medical Technologies, Carlsbad, CA) was used to examine the fetus and uterine contents. Fetal limbs were exteriorized for microsurgery. A new system (Vista Medical Technologies) was attached to an operative microscope, permitting projection of a 3D image via an HMD. The fetus and umbilical cord were inspected using the zoom endoscope, which changes the depth of focus under computer control. Basic manipulations of the fetus and cord were easily completed. Real-time 3D fetal imaging was accomplished. The added depth perception enabled detailed fetal and placental examination, fostering manipulation of the fetus and cord. The HMD was adjusted to fit several surgeons, permitting a natural operative posture. This unit has the capacity to display any video, CT, MR, or ultrasound image as a picture-in-picture. The success of minimally invasive fetal surgery is in part dependent on the development of video technologies capable of providing both magnification and optimal resolution. The zoom endoscope affords excellent visibility of multiple surgical targets without instrument repositioning. A 3D HMD system such as this provides greater anatomic detail and an appreciation of fetal movements that may make intrauterine procedures more feasible.

Evaluation of retropulsion caused by holmium:YAG laser with various power settings and fibers.

The ideal intracorporeal lithotriper would comminute all types of calculi into small readily excreted particles. It would be small and flexible with an energy source safe for the uroepithelium. It should not break, should be inexpensive, and should not retropulse the stone up the urinary tract. This investigation was designed to quantify the last quality for the holmium:YAG laser. The mechanism of action of the pulsed Ho:YAG laser (wavelength 2100 nm) is the generation of a gas plasma at the stone-fluid interface causing a shockwave. The holmium laser was employed for lithotripsy of model stones composed of silicate with a ferrous coating. Stones were selected with a mass of 2 mg +/- 0.1 mg. We sequentially investigated three variables: energy (0.6, 0.8, and 1.0 J), frequency (10, 16, and 20 Hz), and fiber diameter (200, 365, 550, and 1000 microm). Ten stone trials were performed with each of the 36 possible combinations of energy, pulse frequency, and fiber diameter. Our model ureter consisted of a clear rigid polymer tube filled with 0.9% saline. The system was closed and permitted intertrial flushing of generated air bubbles. The laser fiber was maintained at constant extension from the ureteroscope, with stones positioned at the fiber tip before each trial. Laser energy was applied for 2 seconds, with maximum and net retropulsion recorded in millimeters. Each measurement series was recorded in a database for paired Student t-tests. Net retropulsion was then compared by statistically holding each of the three variables constant (fiber size constant with power and frequency varying; frequency constant with power and fiber size varying; and power constant with fiber size and frequency varying). Most retropulsion occurred with the 365-microm and 550-microm fibers. Most comminution was also noted with these fiber sizes. There was no statistical correlation between observed retropulsion and efficiency of comminution. This self-contained model for laser lithotripsy allowed us to measure retropulsion accurately. Silicate stones are not chemically similar to human uroliths but are of uniform composition. The irregular surface characteristics are similar to human stones, making them ideal for retropulsion investigations.

Endoscopic subcutaneous neurovascular lower-extremity myofascial flap harvesting for genitourinary reconstruction.

Neurovascular myofascial flaps have been widely utilized when healthy tissues outside the anatomic confines of the pelvis are necessary for complex genitourinary reconstructions. Myofascial flaps have great potential in providing support and tone to the bladder neck in a model of total urinary incontinence. Two muscles were evaluated for minimally invasive harvesting utilizing an adult rabbit and human cadaveric model: the gracilis, because of its neurovascular supply and length, and the sartorius, because of its location. Three adult New Zealand White rabbits were prepared and draped in sterile fashion after general anesthesia. Bilateral 2-cm incisions were made over the origins of the gracilis and sartorius muscles. Dissection times were entered in a database on a personal computer for paired Student t-tests. Special techniques for dissection included utilization of a 5-mm subcutaneous dissector/retractor and reticulated scissors and dissector. Once the tissues were harvested, rotational angles of the functional arcs around the origins of the gracilis and sartorius flaps were measured, as were the lengths of the muscular flaps from the proximal pedicle to the free distal end. Initial positioning of the myofascial flap at the bladder neck was investigated in this nonsurvival study. Blood loss was estimated before the animals were humanely sacrificed. The possibilities of minimally invasive genitourinary reconstruction are numerous. Both flaps can be dissected endoscopically and have rotational angles allowing transfer to more cephalad positions in the groin or intra-abdominally. Further studies are needed to establish the efficacy of the myofascial graft for minimally invasive genitourinary reconstructions.

Initial studies of holmium:YAG laser creation of spinal defects in fetal rabbits: model for urologic effects of myelomeningocele.

Myelomeningocele (MMC) is characterized by paraplegia and incontinence, often necessitating surgery. Current models of MMC in sheep and primates create a spinal defect long after anomalous neural tube closure ordinarily occurs. An ideal model of MMC would allow creation of the defect at the earliest age in a low-cost species with a short gestation. We present a method utilizing the holmium laser to create spinal defects in rabbits in utero for the study of the pathophysiology and repair of MMC. Pregnant rabbits of 22 to 23 days' gestational age were prepared and draped in sterile fashion for laparotomy under general anesthesia. The abdomen was opened, and both uterine horns were inspected. Double opposing pursestring sutures were placed to secure the chorioamniotic membranes over the fetal lumbar spine. Amniotic fluid was removed with a needle and saved. Electrocautery was used to open the uterus within the pursestring suture, exposing the fetal dorsum. The spine was exposed by laser dissection of the fetal dorsal musculature. Posterior laminectomy was accomplished with laser incisions of each side of the spinous process, leaving the underlying dura and cord exposed. The pursestring was then cinched, amniotic fluid was returned, and the uterus and trocar sites were closed. Cesarian section was performed at 30 to 31 gestational days, and the pups were examined and then humanely sacrificed for histologic evaluation of the lesion. The rabbit is an inexpensive species with a short gestation (33-35 days), and four or more fetuses may be operated on per litter, with the remainder serving as controls. Utilization of minimally invasive techniques including holmium:YAG laser dissection facilitates creation of spinal defects at an early age in this small-animal model.

Unilateral suprainguinal ectopic scrotum: the role of the gubernaculum in the formation of an ectopic scrotum.

A rare case of ectopic scrotum is described together with a review of the literature and a discussion of the embryological role of the gubernaculum in the formation and location of normal and ectopic scrota. We identified 16 reported cases of a suprainguinal ectopic scrotum, 4 cases of a femoral ectopic scrotum, 26 cases of penoscrotal transposition, and 19 cases of a perineal (accessory) scrotum. Although the gubernaculum is a prerequisite for the ultimate location of both the testis and scrotum, its role is complicated by the subsequent differential growth of the labioscrotal folds in which the gubernaculum is stabilized. If this interaction is disturbed, the result may be a suprainguinal ectopia, penoscrotal transposition or a perineal scrotum. A femoral ectopic scrotum, unlike the above, is the result of an aberrant gubernacular stabilization. While the etiology of these malformations is likely to be multifactorial, the existence of an inbred strain of rats characterized by a high incidence of an ectopic scrotum suggests a genetic component to this anomaly.

Stationary mechanical-assist technology for single-surgeon laparoscopic bilateral varicocelectomy.

One potential detraction from the continued application of laparoscopy in the management of varicoceles is the requirement for a skilled assistant, which increases the cost of performing this surgery. This report describes the clinical application of a simple stationary mechanical-assist device to allow a single surgeon to perform bilateral varicocelectomies. The device is attached to the surgical table with a rotary adjusting stem-arm. A cross-mounted sidearm stretches over the patient and forms the attachment to the friction-jointed elbow and wrist attachment to the laparoscope, providing the range of motions similar to a human arm (shoulder, elbow, and wrist). By adjusting the tension at all three levels, changes of the laparoscopic camera portal are possible. Both of the surgeon's hands are then free to work through two operating trocars. The Laprotract arm (Minnesota Scientific, St. Paul, MN) costs $3000 and is autoclavable for quick reutilization. The average time needed to set up the device for bilateral varicocelectomy during eight cases was 2 minutes. The mean procedure time was 65.0 (+/-23.1) minutes, reflecting its ease of use. The electronic image obtained from the stationary mechanical assistant was always steady, and there was no inadvertent wandering from the surgical field. Statistical comparison with 63 open left and 22 open bilateral varicocelectomies during the same time period demonstrated no significant differences in the procedural times. Mechanical-assist technologies can facilitate laparoscopic bilateral varicocelectomies, allowing a single surgeon to perform this operation as quickly as left and bilateral open procedures.

Minimally traumatic techniques for in utero access and fetal surgery.

BACKGROUND AND OBJECTIVES: Prenatal interventions may prevent some sequelae of congenital anomalies, yet open fetal surgery is limited by pre-term labor. We are developing amnioscopic strategies to reduce risks for in utero surgery. METHODS: Seven fetal sheep were accessed percutaneously under ultrasound guidance, via maternal laparoscopy/transuterine trocars, or via laparoscopically assisted partial uterine exteriorization (mini-hysterotomy). Three fetal immobilization methods were investigated: 2 mm grasping forceps, 2-0 absorbable Roeder loop lasso, and detachable metal T-fasteners. RESULTS: Percutaneous access with 2 mm trocars required accessible amniotic fluid pockets. The laparoscopically assisted method enabled 5 mm trocar placement under direct fetal visualization. Mini-hysterotomy avoided trocars and was leak-proof. 2 mm graspers enabled fetal manipulation with repeatable applications. Roeder loop sutures were not readily repositioned, and required two 3 mm or larger trocars. T-fasteners were easily deployed into fetal skin via the self-contained needle applicator with minimal hemorrhage. Cutaneous marks were present immediately from the grasper and T-fastener, but not at postoperative day 10. The Roeder loop produced no observable effects. All devices demonstrated adequate intrauterine performance. CONCLUSIONS: Minimally invasive fetal surgery promises to lower maternal-fetal risks. All strategies permitted prolonged amnioscopy and fetal manipulation. The 2 mm grasper was easiest to use, producing no observable lasting trauma.

The incidence of renal anomalies at full term in fetal rats is synergistically increased by estradiol (but not testosterone) supplementation on day 18 of alcoholic gestation.

BACKGROUND/PURPOSE: Fetal alcohol syndrome is characterized by facial dysmorphology, mental and growth retardation, and somatic anomalies including hydronephrosis. The authors sought to determine the influence of exogenous testosterone or estradiol on the incidence of hydronephrosis in a rodent model of fetal alcohol syndrome (FAS). METHODS: Pregnant rats were fed a liquid diet containing 35% ethanol-derived calories from gestation day 6 through 15, with exogenous testosterone or estradiol supplementation on day 18. On day 20, fetal kidneys were examined for evidence of hydronephrosis, and fetal serum estradiol concentrations were determined by radioimmunoassay. RESULTS: Maternal estrogen supplementation resulted in very high fetal serum estradiol levels that were not additionally increased by alcoholism. Despite this fact, the expression of renal malformations was highest in the alcoholic, estradiol-supplemented offspring. Additionally, the rate of renal malformations was significantly higher in the estrogen-supplemented alcoholic group than in the strictly estradiol animals, yet the fetal serum estradiol concentrations did not differ between the two groups. CONCLUSIONS: This suggests that ethanol may act synergistically with estradiol to increase the rate of renal anomalies including hydronephrosis. Such damage may persist via a suppression of normal testosterone-stimulated renal growth and development. FAS includes significant renal anomalies characterized by hydronephrosis in both animal models and affected children. Although the long-term functional sequelae of hydronephrosis and reflux are well known, the progression of renal disease in FAS children remains to be documented.

XX sex reversal: molecular analysis of the SRY/ZFY regions.

PURPOSE: The mammalian sex determining gene, sex region Y chromosome (SRY), is now firmly established as the testis determining locus. The SRY locus is close to the short arm Y terminus and just distal to zinc finger Y region (ZFY), a locus previously thought to be involved in testicular differentiation and the male phenotype. We report on XX sex reversal, a rare sex chromosomal disorder in humans. MATERIALS AND METHODS: Routine amniocentesis revealed an XX fetal karyotype, although at birth the neonate was phenotypically male. Radiographic evaluation showed a normal male urethra and the absence of any female internal genitalia. Subsequent molecular analysis with polymerase chain reaction amplified sequences of the SRY and ZFY loci were positive. RESULTS: This case is the fourth in our series of XX sex reversed male individuals and to our knowledge the first to be diagnosed perinatally. In all cases the SRY and ZFY loci are present, presumably on the paternal X chromosome, as well as a Klinefelter phenotype. These sex reversing translocations are thought to be due to an unequal meiotic recombination of the distal X and Y short arms during male gametogenesis. The tendency for XY translocations to break between the SRY and ZFY loci was not seen in these apparent microtranslocation cases. CONCLUSIONS: These 4 cases demonstrate the usefulness of molecular followup of clinically perplexing sexual discordance. We conclude that SRY and ZFY polymerase chain reaction amplification studies should be performed when sexual discrepancies are noted on prenatal ultrasound and karyotype analysis.

Amnioscopic endofetal illumination with infrared-guided fiber.

Minimally invasive amnioscopic surgery has several potential advantages over traditional open hysterotomy, including the reduction of the risk of preterm labor and spontaneous abortion. Adequate visibility of the fetal target organ is obligate to the success of in utero procedures. This is a preliminary report on the use of an end-emitting infrared fiber (750 microns; 810-nm wavelength) that allows image fusion with Infravision videocamera systems (Gabriel Medical, Lafayette, LA). Once placed, this fiber can serve as a homing beacon to identify the fetal bladder in the surgical creation of an amnioscopic vesicostomy for the relief of obstructive uropathies. Under general endotracheal halothane anesthesia, amnioscopic access was established in time-dated pregnant ewes using one 3.7-mm and two radially dilating 2-mm to 5-mm trocars. This access allowed the investigation of several methods of intravesicular infrared fiber placement. Ultrasound-guided direct needle puncture is the quickest method of accessing the fetal bladder but is unreliable when the bladder is empty. Accurate placement of the access needle directly over the suprapubic region was accomplished with amnioscopic assistance. The fiber was also placed transurethrally in one female and one male fetus, with subsequent perforation of the female's bladder. Successful infrared bladder illumination was accomplished in all animals. The fusion integration of this camera system allows simultaneous viewing by visible spectrum and near-infrared wavelengths. We believe that this system provides a further degree of safety for amnioscopic procedures.

Assessment of access strategies for fetoscopic urologic surgery: preliminary results.

Attempted endoscopic fetal interventions have been increasingly reported, given the knowledge that minimizing the trauma to the uterus by a small access portal can allow prolonged amnioscopic-guided surgery. Numerous fetal anomalies have been targeted for these interventions, including congential obstructive uropathies. This preliminary investigation explored innovative access strategies utilizing time-dated pregnant ewes, 95 to 100 days' gestation (term 145 days). Two strategies were specifically investigated: percutaneous, ultrasonographically guided microamnisocopy and ultrasonographically aided, laparoscopically guided amnioscopy. Both strategies provided excellent fetal visibility and were facilitated by the use of small trocars and microlaparoscopic instrumentation, which promoted prolonged amnioscopy with minimal access trauma.