Modification of extracorporeal photopheresis technology with porphyrin precursors. Comparison between 8-methoxypsoralen and hexaminolevulinate in killing human T-cell lymphoma cell lines in vitro.

BACKGROUND: Extracorporeal photopheresis that exposes isolated white blood cells to 8-methoxypsoralen (8-MOP) and ultraviolet-A (UV-A) light is used for the management of cutaneous T-cell lymphoma and graft-versus-host disease. 8-MOP binds to DNA of both tumor and normal cells, thus increasing the risk of carcinogenesis of normal cells; and also kills both tumor and normal cells with no selectivity after UV-A irradiation. Hexaminolevulinate (HAL)-induced protoporphyrin-IX is a potent photosensitizer that localizes at membranous structures outside of the nucleus of a cell. HAL-mediated photodynamic therapy selectively destroys activated/transformed lymphocytes and induces systemic anti-tumor immunity. The aim of the present study was to explore the possibility of using HAL instead of 8-MOP to kill cells after UV-A exposure. METHODS: Human T-cell lymphoma Jurkat and Karpas 299 cell lines were used to evaluate cell photoinactivation after 8-MOP and/or HAL plus UV-A light with cell proliferation and long term survival assays. The mode of cell death was also analyzed by fluorescence microscopy. RESULTS: Cell proliferation was decreased by HAL/UV-A, 8-MOP/UV-A or HAL/8-MOP/UV-A. At sufficient doses, the cells were killed by all the regimens; however, the mode of cell death was dependent on the treatment conditions. 8-MOP/UV-A produced apoptotic death exclusively; whereas both apoptosis and necrosis were induced by HAL/UV-A. CONCLUSION: 8-MOP can be replaced by HAL to inactivate the Jurkat and Karpas 299 T-cell lymphoma cells after UV-A irradiation via apoptosis and necrosis. This finding may have an impact on improved efficacy of photopheresis.

Influence of crystal modification on the photoinduced color change in riboflavin.

Two different qualities of riboflavin (RF) i.e., synthetic (RFs) and biosynthetic riboflavin (RFbs) have been investigated with respect to photoinduced color change in the solid state. Several methods (XRD, FT-IR, VIS-, NIR- and fluorescence spectroscopy) were employed to elucidate the properties of the crystalline structure of RFs and RFbs and the influence of irradiation on the color and structural changes of the samples in the solid state. It was shown that RFs an RFbs represent two different crystal modifications of riboflavin and that RFbs can easily be transformed into a dihydrate upon exposure to humidity. Based on the observed irreversible color change and reduction in fluorescence intensity upon irradiation, an irreversible photoreduction of the molecule was assumed in case of RFs. A more pronounced, reversible color change and reversible reduction in fluorescence intensity indicated a reversible photoreduction process in the case of RFbs. The mechanism of these processes was further investigated by means of NIR and FT-IR spectrophotometry. It is apparent from the current study that the crystal modification of RF can strongly influence the solid state photochemistry of this molecule.

Methemoglobin formation during laser induced photothermolysis of vascular skin lesions.

BACKGROUND AND OBJECTIVE: Monitoring dynamic changes during laser induced photothermolysis of vascular skin lesions is essential for obtaining an optimal therapeutic result. Rapid photoinduced thermal damage occurs at a threshold temperature of about 70 degrees C. It is therefore, relevant to identify markers to indicate if this threshold temperature has been reached. Methemoglobin, which is formed by a photo-induced oxidation of hemoglobin, indicates that the temperature has reached this threshold value. This study presents a proof of concept of a method for monitoring the in vivo presence of methemoglobin immediately after laser exposure. STUDY DESIGN/MATERIALS AND METHODS: The present study was designed to investigate the in vivo temperature dependence of hemoglobin absorption in the 450-800 nm spectrum range. In vivo diffuse reflectance measurements of port-wine stain (PWS) and telangiectasia were performed prior to, and immediately after, laser treatment with a pulsed dye laser (PDL) at 585 nm wavelength. RESULTS: In vivo measurements following laser treatment of vascular skin lesions showed an immediate increase in the optical absorption of blood. This effect, caused by thermal stress, is a result of an increased dermal blood volume fraction and methemoglobin formation. The effect is light dose dependent, and reflectance spectra revealed methemoglobin formation in patients treated with fluences above 5 J/cm2 at 585 nm wavelength. CONCLUSIONS: It was proved that methemoglobin can be measured in vivo by reflectance spectroscopy. Measurements of the average methemoglobin concentrations immediately after laser exposure may be a valuable diagnostic tool to verify that the blood temperature has been sufficiently high to induce thermal damage to the vessel wall.

Cooling efficiency of cryogen spray during laser therapy of skin.

BACKGROUND AND OBJECTIVES: Cryogen spray cooling (CSC) is used extensively for epidermal protection during laser-induced photothermolysis of port wine stains and other vascular skin lesions. The efficacy of CSC depends critically on the heat transfer coefficient (H) at the skin surface for which, however, no reliable values exist. Reported values for H, based on tissue phantoms, vary from 1,600 to 60,000 W/m(2) K. STUDY DESIGN/MATERIALS AND METHODS: A simple experimental model was designed and constructed, consisting of a pure silver-measuring disk (diameter 10 mm, thickness approximately 1 mm), embedded in a thermal insulator. The disk was covered with a 10 microm thick stratum corneum layer, detached from in vivo human skin. The heat transfer coefficient of the stratum corneum/cryogen interface was measured during CSC with short spurts of atomized tetrafluoroethane. RESULTS: H was found to be dependent on the specific design of the cryogen valve and nozzle. With nozzles used in typical clinical settings, H was 11,500 W/m(2) K, when averaged over a 100 ms spurt, and 8,000 W/m(2) K when averaged over a 200 ms spurt. CONCLUSIONS: The presented model enables accurate prediction of H and thus improve control over temperature depth profile and cooling efficiency during laser therapy. Thereby, it may contribute to improvement of therapeutic outcome.