Thoracoscopic laser ablation of pulmonary bullae. Radiographic selection and treatment response.

The purpose of this study was to develop objective preoperative selection methods for predicting outcome in patients undergoing thoracoscopic laser ablation of emphysematous pulmonary bullae. Initial radiographic presentation was correlated with physiologic function both before and after the operation in 24 patients entered into a prospective clinical protocol for evaluation of carbon dioxide laser treatment of emphysematous pulmonary bullae. Nineteen surviving patients underwent follow-up evaluation 1 to 3 months after the operation. Pulmonary function test results showed improvements in spirometry (forced vital capacity increased 0.82 +/- 0.125 L, forced expiratory volume in 1 second increased 0.36 +/- 0.07 L, and maximum voluntary ventilation increased 11.69 +/- 2.6 L/m; p < 0.002); airway resistance decreased by 0.9 +/- 0.35 cm of water/L per second, and specific conductance increased 0.019 +/- 0.006 L/cm H2O per second (p < 0.02). Lung volumes improved (residual volume decreased 1.25 +/- 0.23 L, p < 0.001) without significant change in resting gas exchange. Quantitative radiographic grading of extent of preoperative pulmonary bullae correlated well with response to laser treatment in patients with preoperative and postoperative studies. Patients with large bullae accompanied by crowding of adjacent lung structures, upper lobe predominance, and minimal underlying emphysema had greatest improvement in pulmonary function results with laser bullae ablation (p < 0.05). However, some patients with multiple smaller bullae and diffuse emphysema also demonstrated objective improvement after operation. Quantitative radiographic analysis of the extent of bullous disease and the degree of associated emphysema can be used to determine short-term postoperative pulmonary response and may be useful in selecting future thoracoscopic laser bullae ablation candidates. Additional follow-up will be necessary to further improve selection criteria and help define the long-term role of thoracoscopic laser treatment of bullous emphysema.

Electrical cutting device for laparoscopic removal of tissue from the abdominal cavity.

The extraction of large tissue masses from the abdominal cavity during laparoscopic surgery is a time-consuming, complicated process. A new prototype instrument is introduced that allows quick laparoscopic removal of fibromas, ovaries, or other tissues from the abdomen. A cylinder with a coning knife at its intra-abdominal end is placed inside the trocar sleeve and is rotated by an electrical micro-engine attached to the trocar. Cylindrical tissue blocks are cut step by step out of the main specimen and removed from the peritoneal cavity through the sleeve with a grasping forceps. Tissue removed is suitable for histologic examination. The principal application is morcellation of fibromas, whereas use in ovarian disease is limited. This new device provides a safe and effective approach inside the abdominal cavity.

New instrumentation and technique for laparoscopic carbon dioxide laser operations: a preliminary report.

Laparoscopic surgery, the accepted procedure for several indications, still has some limitations, mainly due to the risk of uncontrollable bleeding and damage to the surrounding tissues that may result from the procedure. With the advent of the surgical carbon dioxide laser apparatus and its advantage in producing a bloodless incision, the authors have modified their approach toward laparoscopic surgery. A variety of special straight laser channels for introduction into the abdominal cavity have been developed; use of these channels requires special care to protect surrounding tissues from damage during the procedure. A Sharplan laser delivery system comprising an articulated arm was used, and the operative area visualized and controlled by single- and double-puncture laparoscope systems. The techniques were first applied in rabbits and, after several improvements, used in humans for various indications. The laser laparoscope can be particularly useful in the treatment of endometriosis and lysis of pelvic adhesions.

Intrauterine light probe for photodynamic ablation therapy.

BACKGROUND: Endometrial ablation by means of photodynamic therapy is currently being evaluated as an outpatient treatment for dysfunctional uterine bleeding. Photodynamic therapy requires the activation of a photosensitizer by laser light. We describe a new device specifically designed to provide light delivery to the uterus for endometrial photodynamic therapy. INSTRUMENT: The intrauterine light probe consists of the three flexible optical fibers converging to one bundle resembling the shape of the uterine cavity. Each of the fibers contains a cylindrical light diffuser. EXPERIENCE: The intrauterine light probe was tested in removed human uteri for its capability to distribute light in a tissue-simulating scattering medium and to deliver sufficient light throughout the endometrium. The light distribution of the intrauterine light probe in the scattering medium is uniform on eight axes tested around the diffusing fibers. The pattern of light distribution in human uteri is similar to that in the medium. At the endomyometrial junction, there is still one third of the light applied to the endometrial surface whereas deeper in the myometrium, the light power drops to less than 10%. CONCLUSION: We propose a device that will deliver light to the uterine cavity to induce endometrial ablation by means of photodynamic therapy.

Benzoporphyrin derivative: a potent photosensitizer for photodynamic destruction of rabbit endometrium.

OBJECTIVES: To determine the optimal pharmacokinetic characteristics for photodynamic endometrial destruction using topically applied benzoporphyrin derivative and to evaluate long-term morphologic effects in a rabbit model. METHODS: Benzoporphyrin derivative pharmacokinetics were measured by analyzing frozen tissue sections 1.5-12 hours following topical administration. Photodynamic therapy was induced intraluminally 1.5 hours after drug application, and tissue morphology was evaluated by light and scanning electron microscopy 1 and 4 weeks after treatment. RESULTS: The highest glandular and stromal fluorescence was observed 1.5 hours after topical administration. Relative fluorescence of the endometrial glands significantly exceeded that of stroma and myometrium mainly at 1.5 and 3 hours (P < .0001). Histology revealed persistent epithelial destruction with minimal regeneration. Stroma resembled scar tissue in the majority of sections. The bordering myometrium was loosened and invaded by connective tissue. CONCLUSION: Topically applied benzoporphyrin derivative can be used for highly effective, long-lasting photodynamic destruction of rabbit endometrium. However, optical dosimetry can vary, particularly in the rabbit model, and this appears to affect long-term reepithelialization.

Thoracoscopic treatment of spontaneous pneumothorax using carbon dioxide laser.

In an effort to improve the success rate of the previously described thoracoscopic electrocautery ablation technique of spontaneous pneumothorax, the carbon dioxide laser was evaluated in 12 patients. The recurrent (5 patients) or persistent (7 patients) spontaneous pneumothoraces were caused by rupture of (1) blebs in 6 patients, (2) intrapulmonary apical type II bullae in 3 patients, and (3) diffuse bullous emphysema, type III, in 3. The air leaks were successfully sealed in all but 1 patient with ruptured type II bulla. Surgical specimen from this single failure suggested that the entire inner lining of the bullae must be thermocoagulated. This technical modification led to successful outcome in 2 subsequent cases. With the use of carbon dioxide laser, it was possible to treat not only small blebs but all types of bullae causing spontaneous pneumothorax. Laser thoracoscopy is effective and safe in treating spontaneous pneumothorax.

Actual effective CO2 laser power on tissue in endoscopic surgery.

To elucidate the cause of power reduction in endoscopic carbon dioxide (CO2) laser surgery and to define the actual effective power on tissue of a beam delivered through different endoscopes, we designed an experimental model. In passing a CO2 laser beam through a CO2 gas-pressurized chamber, no significant power loss was noted. A correlation between the internal diameter of an endoscope and the power output became evident. Effective power of 92%, 85%, and 48% of the potential 100% was measured when a beam was delivered through single- and double-puncture laparoscopes and a hysteroscope, respectively. These figures are relevant in predicting CO2 laser effect used endoscopically.

Cutaneous graft-versus-host-like reaction after paternal lymphocyte immunization for prevention of recurrent abortion.

We have presented an immunocompetent patient who, after paternal lymphocyte immunization, developed a GVH-like skin reaction; she recovered spontaneously. This complication is rarely seen. This was the only case observed in our series of over 70 patients immunized. A similar phenomenon was not reported in a larger series by Carp and colleagues (1). However, the case deserves special attention because this reaction can be life-threatening. Therefore, we suggest that paternal lymphocyte immunization should be recommended only for those patients who have undergone a thorough work-up and in whom all causes, other than immunological, for recurrent abortion have been excluded. Even then, this treatment should be given with utmost caution. In view of the above described complication, we believe that more data are still required to justify offering such a treatment modality on a routine basis.

Tissue effects of a new laparoscopic carbon dioxide laser probe.

The effects of CO2 laser beam delivered through a new laparoscopic probe on rabbit uteri is presented. This device has an outer diameter of 4.9 mm and can be delivered through most of the standard operative laparoscopic accessories. Two probes with miniature zinc selenide lenses of 6.5 mm and 17 mm back focal length attached at the distal end were tested. Through each probe, constant 0.5-second pulses and power levels of 5, 10, and 15 W were applied in three different lens-to-target distances. Measurements of beam spot diameters and microscopic crater characteristics revealed significant differences as a result of minor shifting of lens-target distance. From 7 to 9 mm beyond the focal plane and at power levels of 5 to 10 W, negligible tissue effects could be detected. This was defined as an "optical back-stop" effect. A new application mode using an easily maneuverable probe for cutting and coagulating [focusing/defocusing] effects increases the options available for operative laparoscopy.

Limb-body wall complex with complete absence of external genitalia after in vitro fertilization.

The first case of limb-body wall complex (a combination of a body wall defect, neural tube defect, and limb abnormalities) after IVF is described. This is the third reported patient with this complex anomaly associated with complete absence of external genitalia. The possible pathogenic mechanisms are discussed.

Micromanipulation of sperm by a laser generated optical trap.

The force generated by the radiation pressure of a low power laser beam induces an optical trap which may be used to manipulate sperm. We studied the effect of the optical trap on sperm motility. A Nd:YAG laser beam was coupled to a conventional microscope and focused into the viewing plane by the objective lens. Sperm were caught in the trap and manipulated by a joy stick controlled motorized stage. After different exposure periods, the velocity and patterns were analysed by a computerized image processor. There were minor changes in sperm velocity when exposed to the trap for 30 seconds or less. A gradual decrease in the mean linear velocity was observed after 45 seconds of exposure. This optical micromanipulator may also be useful for studying the force generated by a single spermatozoa and evaluating the influence of drugs on motility.

Force generated by human sperm correlated to velocity and determined using a laser generated optical trap.

The development of the single beam gradient force optical trap has made it possible to manipulate cells solely by laser light. A continuous wave Nd:YAG (1.06 microns) laser beam was directed into a conventional microscope and focused onto the viewing plane by the objective lens. The laser beam power at which human sperm were released from the trap was measured and correlated to the sperm's linear velocity before trapping. The mean trapping power readings for slow, medium, and fast motile sperm were 57, 73, and 84 mW, respectively. The analysis of measurements over the total population demonstrated that zig-zag motile sperm had significantly higher mean power readings when compared with straight motile sperm with similar mean linear velocities. In two cases, specimens required significantly less trapping power when the measurements were repeated 24 hours later.

Relative force of human epididymal sperm.

OBJECTIVE: To assess the relative escape force of human epididymal sperm using a laser generated optical trap and compare it with that of human ejaculated sperm. DESIGN: Evaluation of the relative force generated by epididymal and ejaculated sperm using an 800-nm laser-generated optical trap system (titanium-sapphire, model 899-01; Coherent Innova, Palo Alto, CA). SETTING: University-based facility at the Beckman Laser Institute and Medical Clinic and Center for Reproductive Health, University of California, Irvine. INTERVENTIONS: A total of 2,720 sperm from 28 samples were randomly analyzed. Fifteen were ejaculated samples (1,650 sperm) obtained from men with proven fertilization, and 13 were epididymal samples (1,070 sperm) aspirated microsurgically from patients with obstructive azoospermia. An optical trap equipped with the 100x Neofluar objective was used to analyze an average of 100 sperm per patient. MAIN OUTCOME MEASURES: Determination of mean relative escape force values in milliwatts for epididymal and ejaculated sperm samples. RESULTS: The mean relative escape force for epididymal sperm was 32.4 mW, significantly lower than ejaculated sperm, which was 85.1 mW. By correlating epididymal sperm relative force with fertilization in vitro at an arbitrary cutoff value of 30 mW, it was found that no fertilization occurred if a sample had < 13% of sperm at that value. CONCLUSIONS: [1] The average relative escape force of the epididymal sperm was found to be 60% weaker than that of ejaculated sperm. [2] It is demonstrated that the noncontact laser optical trap is a sensitive tool that can evaluate single sperm force as a new physiological parameter.

The relationship between sperm ultrastructural features and fertilizing capacity in vitro.

OBJECTIVE: To evaluate the relationship between ultramorphological features of the human sperm and its fertilizing capacity in vitro. DESIGN: The study was performed retrospectively. Ultrastructural features were assessed using scanning and transmission electron microscopes in sperm samples of individuals who underwent an in vitro fertilization (IVF) treatment cycle no more than 6 months before the study. SETTING: Institutional clinical care. PATIENTS: Fifty-six infertile couples in whom mechanical infertility was diagnosed in the female partner. Patients were categorized as fertilizing when fertilization of at least 30% of the oocytes occurred (n = 27) and nonfertilizing when none of the oocytes fertilized in at least two consecutive IVF treatment cycles (n = 29). RESULTS: The two groups differed significantly only in ultramorphological parameters of the sperm head and acrosome (head, F(8,36) = 2.8, P less than 0.02; acrosome, F(4,40) = 2.8, P less than 0.04), and especially in the following malformation patterns: hyperelongated head, acrosome deficiency, and acrosome damage. The suggested score based on these findings was able to predict 90% and 76% of the cases with and without fertilizing potential, respectively. CONCLUSION: The ultrastructural morphology of the sperm head components is a key parameter for assessing the sperm fertilizing capacity in vitro.

Effect of freezing on the relative escape force of sperm as measured by a laser optical trap.

OBJECTIVE: To determine the possible effect of freezing on sperm escape force as measured by laser trap. DESIGN: Controlled clinical study. SETTING: Normal volunteers, academic setting. PATIENTS: Normal, healthy volunteers. INTERVENTION: Sperm selection, freezing and thawing. Measurement of relative (sperm) escape force before and after freezing. MAIN OUTCOME MEASURE: Comparison of escape force as a measurement of freezing effects. RESULTS: Wide individual variations noted, with some individuals showing significant increases or decreases. The relative escape force in fresh (76.1 +/- 31.1 mW) and in frozen-thawed samples (75.6 +/- 40.0 mW) were similar. CONCLUSION: Freezing does not affect the relative escape force of normal sperm. Any possible detrimental effect of freezing on sperm may be related to other sperm functions.

The relationship between in vitro fertilization and naturally occurring antibodies: evidence for increased production of antiphospholipid autoantibodies.

OBJECTIVE: Assessment of possible effects of ovarian stimulation during in vitro fertilization (IVF) treatment cycles on circulating levels of antiphospholipid and antinuclear autoantibodies. DESIGN: The study was performed prospectively. Sera were obtained at three time points along IVF treatment cycle. Levels of autoantibodies directed against nuclear components, mitochondrial antigens, and phospholipids were determined using enzyme-linked immunosorbent assay. PATIENTS: Thirty-five patients, who underwent at least one previous IVF attempt, and 36 age- and sex-matched controls were analyzed. All participants were randomly selected. RESULTS: The mean levels of antiphospholipid (but not antinuclear) autoantibodies in sera from IVF-treated patients were found to be significantly higher than the corresponding values of the control group (for immunoglobulin [Ig]M isotype: anticardiolipin, antiphosphatidyl L-serine; for IgG isotype: anticardiolipin, antiphosphatidyl L-serine, and antiphosphatidylcholine; P less than 0.0001, assessed by Mann-Whitney test). The autoantibody levels remained more or less constant at different time points along the treatment cycle. No correlation with age and number of previous IVF cycles was demonstrated. CONCLUSIONS: Serum levels of antiphospholipid (but not antinuclear) autoantibodies increase after IVF treatment. Based on these preliminary data, it is not yet possible to estimate if the observed changes in autoantibody levels might have any future clinical influence on infertile patients undergoing IVF treatment.

Effect of pentoxifylline on the intrinsic swimming forces of human sperm assessed by optical tweezers.

It is still controversial whether in vitro exposure of sperm to pentoxifylline increases sperm motility and force, which is defined as the product of velocity by beat frequency of the tail. Laser optical tweezers have been successfully used in the past to evaluate sperm force in basal conditions. The aim of this prospective study was to determine whether exposure of human sperm to pentoxifylline has any effect on sperm intrinsic forces. Twelve healthy subjects undergoing routine semen analysis were enrolled in the study. Ten exhibited normal semen parameters, 2 exhibited asthenozoospermia. Each semen specimen was washed and, after swim-up, resuspended in human tubal fluid (HTF) and divided into 2 aliquots. One aliquot was incubated with pentoxifylline for 30 minutes (final concentration = 3.6 mM); the second aliquot, without pentoxifylline, served as a control. After 30 minutes the pentoxifylline-treated aliquot was divided into 2 portions, 1 of which was washed to remove the pentoxifylline, the other was left in prolonged coincubation with the chemical. The main outcome was the measurement of sperm intrinsic force in milliwatts (mW), which was assessed by means of a noninvasive infrared laser optical trap created by a continuous wave, 1064-nm Nd:YAG laser beam directed in an inverted microscope. Exposure of sperm to pentoxifylline consistently increased sperm relative escape force from the laser optical trap. The increase ranged from 33% to 154% over baseline force compared with controls. The average absolute increase in sperm force rose from 37 mW to 79 mW (P < .05). Specimens with sperm having an initial low relative escape force gained the highest relative increase. The effect of pentoxifylline on sperm force, already apparent after 5 minutes, reached a peak at 30 minutes and persisted for up to 3 hours in sperm that were left in coincubation and in those on which the pentoxifylline had been washed off. In conclusion, pentoxifylline significantly increases sperm intrinsic relative force in normozoospermic and asthenozoospermic samples. This experiment confirms that optical tweezers can provide an accurate determination of sperm force in in vitro conditions. Clinical data must now establish whether a documented increase in sperm force is an important parameter for assessing sperm fertilizing capacity.

PEG-m-THPC-mediated photodynamic effects on normal rat tissues.

Photodynamic therapy (PDT) of malignancies uses light to activate a photosensitizer preferentially accumulated in cancer cells. The first pegylated photosensitizer, tetrakis-(m-methoxypolyethylene glycol) derivative of 7,8-dihydro-5,10,15,20-tetrakis(3-hydroxyphenyl)-21-23-[H]-porphyrin (PEG-m-THPC), was evaluated in non-tumor-bearing rats. The aim of this study was to assess the photodynamic threshold for damage and its sequelae in normal rat tissue. Thirty-five Fischer rats were sensitized with 3, 9 or 30 mg/kg body weight PEG-m-THPC. Colon, vagina and perineum were irradiated with laser light of 652 nm wavelength and an optical dose of 50, 150 or 450 J/cm fiber length. Temperature in the pelvis was measured during PDT. Three days following PDT the effect on skin, vagina, colon, striated muscle, connective tissue, nerves and blood vessels was assessed by histology. The healing of the above-mentioned tissues was assessed on two rats 3 and 8 weeks after PDT using 9 mg/kg PEG-m-THPC activated with 450 J/cm laser light. No dark toxicity was observed. PDT using 30 mg/kg PEG-m-THPC induced severe necrosis irrespective of the optical dose. Body weight of 9 or 3 mg/kg activated with less than 450 J/cm induced moderate or no damage. No substantial increase in body temperature was seen during PDT. Tissues with severe PDT-induced damage seem to have a good tendency to regenerate. We conclude that within the dose required for tumor treatment PEG-m-THPC is a safe photosensitizer with promising properties. PDT of the colon mucosa below 9 mg/kg PEG-m-THPC and 150 J/cm seems to be safe. All other tissues can be exposed to 9 mg/kg PEG-m-THPC activated with less than 450 J/cm laser light with little side effects.

Monitoring tumor response during photodynamic therapy using near-infrared photon-migration spectroscopy.

Benzoporphyrin-derivative (BPD)-monoacid-ring A photodynamic therapy (PDT) was performed on subcutaneous tumor implants in a rat ovarian cancer model. In order to assess PDT efficacy the tumor and normal tissue optical properties were measured noninvasively prior to and during PDT using frequency-domain photon migration (FDPM). FDPM data were used to quantify tissue absorption and reduced scattering properties (given by the parameters mu a and mu's, respectively) at four near-infrared (NIR) wavelengths (674, 811, 849 and 956 nm). Tissue physiologic properties, including the in vivo concentration of BPD, deoxy-hemoglobin (Hb), oxy-hemoglobin (HbO2), total hemoglobin (TotHb), water (H2O) and percent tissue hemoglobin oxygen saturation (%StO2), were calculated from optical property data. PDT efficacy was also determined from morphometric analysis of tumor necrosis in histologic specimens. All the measured tumor properties changed significantly during PDT. [Hb] increased by 9%, while [HbO2], [TotHb] and %StO2 decreased by 18, 7 and 12%, respectively. Using histologic data we show that long-term PDT efficacy is highly correlated to mean BPD concentration in tumor and PDT-induced acute changes in [HbO2], [TotHb] and %StO2 (correlation coefficients of 0.829, 0.817 and 0.953, respectively). Overall, our results indicate that NIR FDPM spectroscopy is able to quantify noninvasively and dynamically the PDT-induced physiological effects in vivo that are highly correlated with therapeutic efficacy.

Highly selective targeting of ovarian cancer with the photosensitizer PEG-m-THPC in a rat model.

Photodynamic therapy (PDT) uses light to activate a photosensitizer that has been absorbed or retained preferentially by cancer cells after systemic administration. The first pegylated photosensitizer, tetrakis-(m-methoxypolyethylene glycol) derivative of 7,8-dihydro-5,10,15,20-tetrakis(3-hydroxyphenyl)-21,23-[H]-porphyrin (PEG-m-THPC), was evaluated to target selectively unresectable pelvic ovarian cancer bulks. Our goals were two-fold: (1) to establish an ovarian cancer model suitable for the development of debulking techniques and (2) to characterize the pharmacokinetics and tumor selectivity of PEG-m-THPC by fluorescence microscopy. NuTu-19 ovarian cancer cells were injected into the caudal part of the right psoas muscle of Fisher rats. Five weeks later, 30 mg/kg body weight of PEG-m-THPC was injected intravenously. Necropsy was performed between 4 and 10 days following drug application, and fluorescence of the tumor and various abdominal organs was measured. All rats developed bulky pelvic tumors with an average diameter of 2.6 cm (+/- 0.6 SD). Tumor masses were encompassing and infiltrating pelvic organs in a similar manner to ovarian cancers in humans. Fluorescence of cancer tissue was maximal 8-10 days following drug application. At 8 days, the tumor-to-tissue ratio was 40:1 (+/- 12 SE) for most abdominal organs. We conclude that this tumor model may be used for the study of new pelvic debulking techniques, and that the tumor selectivity of PEG-m-THPC is exceptionally high 8 days after drug application. Based on these data, we are currently developing a PDT-based minimally invasive debulking technique for advanced ovarian cancer.