Comparison of fluorescence and photodynamic activities of whole hematoporphyrin derivative and its enriched active components.

The in vivo biologic activities of the hematoporphyrin derivative (Photofrin) and the enriched, so-called "active fraction" (Photofrin II) were determined by measuring the necrosis produced in implanted tumors in DBA/2Ha mice exposed to various total doses of light (20-100 J/cm2) after ip administration of 10 mg/kg standard doses of either Photofrin or Photofrin II. Total relative percentage increase in fluorescence in tumor tissue, as compared to fluorescence in control tissue, also was measured for both Photofrin and Photofrin II. In response to total light doses (630 nm) of 40-100 J/cm2, mice that received Photofrin had comparable amounts of tumor necrosis to those mice that received Photofrin II. At doses of 40-60 J/cm2, 80% tumor destruction resulted, and at 80-100 J/cm2, tumor destruction was 100%. However, at a total light dose of 20 J/cm2, the tumors that received Photofrin II exhibited 60-80% tumor necrosis, whereas those animals that received Photofrin had only small areas of patchy necrosis associated with signs of vascular thrombosis and hemorrhage into the surrounding perivascular stroma. A 25.2% total increase in maximal tissue fluorescence over that in controls was observed for animals that received Photofrin II, as compared to 13.9% for those animals that received Photofrin. It is concluded that the greater demonstrable efficacy of treatment with Photofrin II, as compared to treatment with Photofrin, is due to enrichment of those nonpolar hydrophobic components of the hematoporphyrin derivative mixture that are thought to be primarily responsible for the in vivo biologic activities.

Combination cytotoxic chemotherapy with cisplatin or doxorubicin and photodynamic therapy in murine tumors.

This study was designed to evaluate the interaction of photodynamic therapy (PDT) and chemotherapy in an animal model. PDT is based on the interaction of hematoporphyrin derivative and red light of the appropriate wavelength (630 nm) and intensity. Two tumor models were utilized: C3H/Km mice bearing the RIF-1 tumor and BALB/c mice bearing the EMT-6 tumor. Tumor-bearing mice were treated with either cisplatin (DDP), doxorubicin (ADM), PDT, or a combination of drug and PDT. It was demonstrated that the RIF-1 tumor was sensitive to DDP and insensitive to both PDT and ADM. There was no additional antitumor effect when either drug was combined with PDT. The EMT-6 tumor was moderately sensitive to PDT and mildly sensitive to both DDP and ADM. Although the addition of DDP did not potentiate tumor destruction, the addition of ADM significantly enhanced the effect of PDT (P = .01). The enhanced activity of the combination of PDT and ADM appeared to be the result of increased activity of ADM alone, when illuminated with red (630 nm) light. This potentiation may be due to a photochemical process or may be secondary to the mild hyperthermia generated by illumination with the laser. This study demonstrates that PDT combined with cytotoxic chemotherapy is well tolerated in these animals and that certain combinations of PDT and chemotherapy may result in an enhanced tumoricidal effect.

Histopathological comparison of the effects of hematoporphyrin derivative on two different murine tumors using computer-enhanced digital video fluorescence microscopy.

A comparison study of the effects of hematoporphyrin derivative (HPD) photoradiation therapy on two different mouse sarcoma tumor model systems (RIF-1 + EMT-6) was performed. Twenty-four h after i.v. administration of HPD, the responses to total laser light doses of 50-400 J/cm2 were evaluated by histological examination and the uptake and distribution of HPD using a computer enhanced digital video fluorescence microscopy technique. In response to total laser light dose (630 nm) of 50-400 J/cm2, 40 mice with RIF-1 tumor showed only minimal superficial tumor necrosis upon histological examination and a 9-12% increase in maximal tissue fluorescence. In contrast, 40 mice with EMT-6 tumor showed marked areas of patchy coagulation necrosis and vascular hemorrhage at doses as low as 50 J/cm2 and essentially total tumor destruction at total light doses of 150 J/cm2 or more. A 59-74% increase in maximal tissue fluorescence was observed using digital video fluorescence microscopy. It is concluded that the greater efficacy of treatment in the EMT-6 tumor as compared to the RIF-1 tumor was due to the greater localization of HPD as demonstrated by digital video fluorescence microscopy.

Mid-infrared laser ablation of stratum corneum enhances in vitro percutaneous transport of drugs.

The precise removal of stratum corneum from cadaveric swine skin by a mid-infrared erbium:yttrium scandium gallium garnet laser (lambda = 2.79 microns; 250 microseconds pulse width) was assessed by electrical resistance measurements and documented by histology. The effects of stratum corneum removal by laser ablation and by adhesive tape-stripping on the in vitro penetration of 3H-hydrocortisone and 125I-gamma-interferon were determined. Excised swine skin was irradiated with laser (1 J/cm2; 31 mJ/pulse; 1 Hz; 2 mm spot diameter). For skin penetration studies, laser pulses were delivered to discrete 2-mm areas to ablate up to 12.6% of the total 3-cm2 stratum corneum diffusional area. Franz in vitro skin penetration chambers were used to measure the cumulative 48-h penetration of 3H-hydrocortisone and 125I-gamma-interferon in laser-treated and tape-stripped skin. Electrical resistance measurements and histologic studies demonstrated that 10-14 laser pulses at the above energy density were required to abolish skin resistance and selectively ablate stratum corneum without damage to adjacent dermal structures. Laser ablation of 12.6% of the surface area of stratum corneum produced a 2.8 and 2.1-times increase in permeability constant (kp) for 3H-hydrocortisone and 125I-gamma-interferon, respectively. These studies demonstrate that a pulsed mid-infrared laser can reliably and precisely remove the stratum corneum, facilitating penetration of large molecules such as 125I-gamma-interferon that cannot penetrate intact skin. This new technique may be useful for basic and clinical investigation of skin barrier properties.

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.

The relationship between attachment height and attachment area of teeth using a digitizer and a digital computer.

Nine lower first bicuspid teeth were mounted in blocks of plastic and their roots were serially sectioned and photographed. Data from the photographs were compiled by a digitizer and a digital computer, which calculated the cumulative root surface area for each millimeter of root height. From these data, three variables, which relate root surface area to root height, were calculated. It was found that if one-half of the height of attachment to the root were lost due to periodontal disease, a mean of 61.5% of the actual attachment area to the root is lost. If a mean of 5.72 mm of root attachment height is lost, or if a mean of 60.6% of the same root height remains, only one-half of the total root attachment area remains to provide tooth support.

Effects of oral hygiene on the results of periodontal surgery in beagle dogs with artificially created defects.

The effect of oral hygiene on periodontal surgery was evaluated clinically, radiographically and histologically during 24 months. In 13 beagle dogs artificial periodontal defects were created around the second, third and fourth premolars of the mandible. One week after removal of the plaque retention ligatures, periodontal surgery consisting of mucoperiosteal flaps and osseous recontouring was performed. Postoperatively, plaque accumulation was prevented by daily application of a 2% chlorhexidine solution for 2 weeks and daily toothbrushing for the next 2 weeks. Toothbrushing was continued in seven dogs; oral hygiene was discontinued in the other six. All dogs were scaled and polished at 1, 6, 12 and 18 months after surgery. Plaque accumulation in the nonbrushed dogs resulted in a heavy inflammation of the periodontium. Probing depths that had been reduced by the surgery increased and subsequently the clinical attachment level changed into a more apical level compared to the brushed dogs. Histometrically, however, no difference in attachment level could be recorded, indicating that oral hygiene did not affect the attachment level in the 2 years after surgery. It appears that oral hygiene stimulated bone formation and that the periodontal tissues had not matured in the 2 years after surgery.

A consideration of economic impact of schistosomiasis.

An attempt is made to estimate the economic effects of schistosomiasis, a disease known to be endemic in 71 countries or islands with a total population of about 1 362 million persons, of whom approximately 124 905 800 are infected. These data are based on prevalence rates representing, for the most part, single stool or urine examinations; the actual number of cases is undoubtedly much greater. This analysis refers only to resource loss attributable to reduced productivity. The annual loss from complete and partial disability is estimated to be US $445 866 945 in Africa, US $755 480 in Mauritius, US $16 527 275 in South-West Asia, US $118 143 675 in South-East Asia, and US $60 496 755 for the Americas. The total estimated annual world loss amounts to US $641 790 130 but this sum does not include the cost of public health programmes, medical care, or compensation for illness.

Parametric study of the forces on microspheres held by optical tweezers.

Optical-trapping forces exerted on polystyrene microspheres are predicted and measured as a function of sphere size, laser spot size, and laser beam polarization. Axial and transverse forces are in good and excellent agreement, respectively, with a ray-optics model when the sphere diameter is ≥ 10 µm. Results are compared with results from an electromagnetic model when the sphere size is ≤ 1 µm. Axial trapping performance is found to be optimum when the numerical aperture of the objective lens is as large as possible, and when the trapped sphere is located just below the chamber cover slip. Forces in the transverse direction are not as sensitive to parametric variations as are the axial forces. These results are important as a first-order approximation to the forces that can be applied either directly to biological objects or by means of microsphere handles attached to the biological specimen.