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The rationale of the therapeutic efficacy of photodynamic therapy is based on the cytotoxic action of products generated by excited photosensitizers. The excited photosensitizer can react directly with tissue constituents through the so-called type I process, yielding radicals or radical ions or through the type II process in which energy is transferred to singlet oxygen. The relation between damage to the normal tissue and that to the malignant tissue is determined by the difference in photosensitizer concentration. In summary, the optimal use of photobleaching enables the selective depth to coincide with the therapeutic depth, whereas the value for the therapeutic depth itself is rather unaffected by bleaching. The retention ratio is of only indirect importance in a procedure optimized for bleaching; the optimization procedure, in principle, requires, only that the retention ratio is larger than 1.

Hyperspectral imaging, image analysis and diffusion theory were used to visualize skin vasculature and to monitor the development of fresh skin bruises. Bruises were inflicted in a porcine model, and the development of the hemorrhage was monitored using white light hyperspectral imaging (400-1000 nm). Hyperspectral images from human volunteers were also included in the study. Statistical image analysis was used to classify bruised regions and to visualize the skin vasculature. Biopsies were collected from the animals to reveal the true depth of the bruising. A three-layer diffusion model and an analytic hemoglobin transport model were used to model the reflectance spectra from the images. The results show that hyperspectral images contain depth information, and that the approximate depth and extent of bruises can be retrieved using a combination of statistical image analysis and diffusion theory. This technique also shows potential to visualize vascular structures in human skin.

Background and Objectives Cryogen spray cooling with tetrafluoroethane (R134a) has been used to enhance epidermal protection during dermatologic laser surgery. However, R134a has a very high global warming potential (GWP = 1300). Our objective was to evaluate the cooling effectiveness of an alternative cryogen with a much lower GWP, namely liquid carbon dioxide (CO2, GWP = 1). Study Design/Materials and Methods A thin-film thermocouple deposited on an epoxy skin phantom was used to measure surface temperature (Ts) variations induced by R134a or CO2 sprays. The temperature distribution in the skin phantom was estimated using Ts and Duhamel's method. Impact pressure and noise level of both cryogen sprays were measured with a dynamic sensor and sound meter, respectively. Consumption of both cryogens was also evaluated. Results For R134a sprays, Ts was kept almost constant after 15 milliseconds. For CO2 sprays, Ts decreased continuously during the entire spurt of 50 milliseconds. The minimum Ts induced by the CO2 sprays was lower than that induced by R134a when the spurt duration was longer than 35 milliseconds. Numerical simulation shows that CO2 sprays were able to induce very similar temperature reductions in the skin phantom as compared to R134a sprays when the spurt duration and delay time were selected appropriately. R134a sprays induced an impact pressure of 3.6 kPa, as compared to 43.1 kPa for CO2 sprays. The maximum noise level for R134a sprays was 109 dBA as compared to 135 dBA for the CO2 sprays. The R134a consumption for a 50 milliseconds spurt is 67 mg as compared to 225 mg for a CO2 spurt of the same duration. Conclusions CO2 sprays are expected to have similar skin cooling efficacy as R134a sprays. Although the CO2 consumption is higher than R134a, its contribution to global warming is still much less than R134a. The effects of varying spurt durations on in vivo human skin and the impact on cutaneous blood flow require further investigation. Lasers Surg. Med. 39:776–781, 2007. © 2007 Wiley-Liss, Inc.

Intravascular thermography has been proposed as a method for detecting vulnerable plaque. A marker of vulnerability in a plaque is inflammation, which is believed to reduce its mechanical stability. It has been hypothesized that this inflammation leads to a higher metabolic rate and therefore higher heat generation, causing increased temperature in the vicinity of the plaque. This temperature increase could be measured intravascularly using a temperature sensor, e.g., a thermistor or a thermocouple. The aim of this study is to present a thorough mathematical and physical analysis of the thermal distribution that can be expected in the plaque under various physiological conditions. To get reasonable predictions on the expected temperature distributions, idealized models with simple geometries are solved analytically. More realistic models, with more complex geometries, are solved numerically using the finite element method (FEM). Based on this analysis, the maximum temperature increase that can be expected in a plaque due to increased metabolism is less than 0.1 K.

This thesis covers a wide field of applications, with an emphasis on applications of reflectance spectroscopy for diagnostic purposes. Reflectance spectroscopy in the visible part of the spectrum has been proved to be a valuable tool in a variety of applications including e. g. port-wine stain diagnostics, diagnostics of liver pathology, neonatal jaundice and age determination of bruises for forensic applications.

Background and Objective Bruises are currently evaluated by visual inspection, and little is known about the first phase after injury. The temporal development of fresh injuries must be accurately described to be able to age bruises in a reliable manner. Color changes in a bruise caused by hemoglobin breakdown products will depend on the severity of the trauma, and thus on the local immune response in the skin. It is therefore important to relate the nature of the impact to the temporal tissue responses. Materials and Methods Controlled injuries were inflicted on anesthetized domestic pigs. Trauma was induced either by a pendulum device, or by paintballs released using pressurized air. The speed of the projectiles was recorded using a high speed camera. Biopsies and reflection spectra (400–850 nm) were collected from normal and bruised skin. The experiments were approved by the national animal research authority. Results The temporal development of the injury was found to depend strongly on the weight and speed of the object. Low speed, blunt objects did not cause persistent skin changes. However, deep muscular bleeding could be found in most cases. High speed, light weight objects caused a rapidly developing bruise. These bruises were fully developed within 15–20 minutes. No deep muscular hemorrhages were observed in those cases. White blood cells (neutrophilic granulocytes) could be found in biopsies from high speed injuries. The amount of white blood cells depended on the time between injury and collection of the biopsies. Conclusion Further investigations utilizing a larger range of object weight and velocities are required to be able to fully classify minor traumatic injuries. Preliminary results indicate that this can be achieved by controlled experiments using a porcine model. Reflectance spectroscopy was found to be a useful tool to study immediate skin reactions to the trauma. Lasers Surg. Med. © 2007 Wiley-Liss, Inc.

Background and Objectives Aging of injuries on a victim's body is an important aspect of forensic medicine. Currently, visual assessment and colorimetry based on empirical criteria are the most common techniques for this task, although the results are uncertain. A trauma causing localized vessel damage will rapidly result in a pool of blood in subcutaneous tissues. The color of the bruise is, however, primarily due to hemoglobin transport into dermis and secondarily to its breakdown products. This transport is analyzed in terms of hemoglobin diffusion followed by clearance by macrophage activity, lymphatic flow, and conversion to breakdown products such as bilirubin. The color of a bruise is caused by hemoglobin and hemoglobin breakdown products. The color will change with time, and such color changes can be recorded using reflectance spectroscopy. The aim of this study was to develop a mathematical model to describe blood diffusion within bruised skin, and to use this method to retrieve the age of a bruise from measured skin reflectance. Study Design/Materials and Methods An analytic model was established to describe the development and fading of bruise color. The model, which is based on Darcy's law of convection flow and Fick's law of diffusion, describes the distribution of blood and hemoglobin breakdown products within a hematoma as a function of time after injury. The initial phase after injury is described by a convective extravascular blood flow in subcutaneous tissues, and further development of the bruise is described by diffusion and breakdown of whole erythrocytes and hemoglobin in dermis. Experimental data were used to verify the model. Reflection spectra in the 400–850 nm wavelength range were collected from normal and bruised skin using an integrating sphere setup. The subjects were adult patients admitted to the Department of cardiothoracic surgery, St. Olav's Hospital, Trondheim, Norway. The skin hematomas were caused by external trauma, cardiothoracic examinations, or surgery. Results Preliminary results show that measured and simulated skin reflectance agrees well. The model predicts the age of a hematoma with an accuracy of approximately 1 day. The accuracy of the method depends on precise information of skin thickness in the injured area. The quality of the estimates from the model will thus be enhanced if a reliable measure of skin thickness is collected concurrently with the reflection measurement. Conclusions The time development of a skin hematoma is described with good accuracy by the implemented model. The analytic method provides a theoretical basis for developing an apparatus to determine the age of injuries in forensic medicine. Lasers Surg. Med. 38:277–289, 2006. © 2006 Wiley-Liss, Inc.

Background and Objectives: Clinical results have demonstrated that dark purple port wine stain (PWS) birthmarks respond favorably to laser induced photothermolysis after the first three to five treatments. Nevertheless, complete blanching is rarely achieved and the lesions stabilize at a red-pink color. In a feasibility study (Part I), we showed that local hypobaric pressure on PWS human skin prior to laser irradiation induced significant lesion blanching. The objective of the present study (Part II) is to investigate the effects of hypobaric pressures on the efficiency of cryogen spray cooling (CSC), a technique that assists laser therapy of PWS and other dermatoses. Study Design/Materials and Methods: Experiments were carried out within a suction cup and vacuum chamber to study the effect of hypobaric pressure on the: (1) interaction of cryogen sprays with human skin; (2) spray atomization; and (3) thermal response of a model skin phantom. A high-speed camera was used to acquire digital images of spray impingement on in vivo human skin and spray cones generated at different hypobaric pressures. Subsequently, liquid cryogen was sprayed onto a skin phantom at atmospheric and 17, 34, 51, and 68 kPa (5, 10, 15, and 20 in Hg) hypobaric pressures. A fast-response temperature sensor measured sub-surface phantom temperature as a function of time. Measurements were used to solve an inverse heat conduction problem to calculate surface temperatures, heat flux, and overall heat extraction at the skin phantom surface. Results: Under hypobaric pressures, cryogen spurts did not produce skin indentation and only minimal frost formation. Sprays also showed shorter jet lengths and better atomization. Lower minimum surface temperatures and higher overall heat extraction from skin phantoms were reached. Conclusions: The combined effects of hypobaric pressure result in more efficient cryogen evaporation that enhances heat extraction and, therefore, improves the epidermal protection provided by CSC. Lasers Surg. Med. 36:130– 135, 2005. 2005 Wiley-Liss, Inc.

Although epidermal melanin content has been quantified non-invasively using visible reflectance spectroscopy (VRS), there is currently no way to determine melanin distribution in the epidermis. We have developed a photoacoustic probe that uses a Q-switched, frequency-doubled Nd:YAG (neodymiurn, yttrium, aluminum, garnet) laser operating at 532 nm to generate acoustic pulses in skin in vivo. The probe contained a piezoelectric element that detected photoacoustic waves that were then analyzed for epidermal melanin content using a photoacoustic melanin index (PAMD). Melanin content was compared between results of photoacoustics and VRS. Spectra from human skin were fitted to a model based on diffusion theory that included parameters for epidermal thickness, melanin content, hair color and density, and dermal blood content. Ten human subjects with skin phototypes I-VI were tested using the photoacoustic probe and VRS. A plot of PAMI v. VRS showed a good linear fit with r2 = 0.85. Photoacoustic and VRS measurements are shown for a human subject with vitiligo, indicating that melanin was almost completely absent. We present preliminary modeling for photoacoustic probe design and analysis necessary for depth profiling of epidermal melanin.

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°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. Lasers Surg. Med. 34:414–419, 2004. © 2004 Wiley-Liss, Inc.

Author(s): Kelly, Kristen M; Kimel, Sol; Smith, Tia; Stacy, Amy; Hammer-Wilson, Marie J; Svaasand, Lars O; Nelson, J Stuart | Abstract: Background and objectivesThe degree of port wine stain (PWS) blanching following pulsed dye laser (PDL) therapy remains variable and unpredictable. Because of the limitations of current PDL therapy, alternative treatment approaches should be explored. The objective was to evaluate a novel methodology for selective vascular damage, combined photodynamic (PDT) and photothermal (PDL) treatment, using the in vivo chick chorioallantoic membrane (CAM) model.Study design/materials and methodsThirty microliters of benzoporphyrin derivative monoacid ring A (BPD) solution was administered intraperitoneally into chick embryos at day 12 of development. Study groups were: (1) control (no BPD, no light); (2) BPD alone; (3) continuous wave irradiation (CW) alone (576 nm, 60 mW/cm2, 125 seconds); (4) CW + PDL; (5) BPD+PDL; (6) PDT (BPD+CW); (7) PDL alone (585 nm, 4 J/cm(2)); and (8) PDT+PDL (BPD + CW followed immediately by PDL). Vessels were videotaped prior to, and at 1 hour post-intervention and then assessed for damage based on the following scale: 0, no damage; 1, coagulation; 1.5, vasoconstriction; 2.0, coagulation+vasoconstriction; 2.5, angiostasis; 3.0, hemorrhage. Damage scores were weighted by vessel "order."ResultsPDT + PDL resulted in significantly (P l 0.01) more severe vascular damage than was observed in any other study group: 127% more than PDT, 47% more than PDL alone.ConclusionsPDT + PDL is a novel and promising approach for selective vascular damage and may offer a more effective method for treatment of PWS and other vascular skin lesions.

Background and Objective Treatment of port wine stains (PWS) by photothermolysis can be improved by optimizing laser parameters on an individual patient basis. We have studied the critical role of pulse duration (tp) on the treatment efficacy. Study Design/Materials and Methods The V-beam laser (Candela) allowed changing tp over user-specified discrete values between 1.5 and 40 milliseconds by delivering a series of 100 microsecond spikes. For the 1.5 and 3 millisecond pulses, three spikes were observed at intervals tp/2 and for tp ≥ 6 milliseconds, four spikes separated by tp/3. The ScleroPlus laser (Candela) has a smooth output over its fixed 1.5 milliseconds duration. Blood vessels in the chick chorioallantoic membrane (CAM) were irradiated at fixed wavelength (595 nm), spot size (7 mm), radiant exposure (15 Jcm−2), and at variable tp. The CAM contains an extensive microvascular network ranging from capillaries with diameter D 0.05). Conclusions The difference between initial arteriole and venule damage could be explained by the threefold higher absorption coefficient at 595 nm in (oxygen-poor!) arterioles. In human patients, PWS consist of ectatic venules (characterized by higher absorption), so that these considerations favor the use of 595-nm irradiation for laser photothermolysis. For optimal treatment of PWS it is proposed that tp be between 0.1 and 1.5 milliseconds. This is based on a modified relaxation time τd′, defined as the time required for heat conduction into the full thickness of the vessel wall, which is assumed to have a thickness ΔD ≈ 0.1D. The corresponding τd′ will be a factor of about six smaller than given in the literature. For vessels with D between 30 and 300 μm, τd′ ranges from 0.1 to 1.5 milliseconds. Lasers Surg. Med. 30:160–169, 2002. © 2002 Wiley-Liss, Inc.

Objective: Our study was designed to investigate 5-aminolevulinic acid (ALA) as a candidate for intraperitoneal photodynamic therapy (IP-PDT). The toxicity of IP-PDT and the effects of IP-PDT on abdominal and pelvic organs, particularly the small intestine, were investigated after ALA administration and illumination with violet laser light. Study design and results: The toxicity of IP-PDT was evaluated in Fischer 344 rats in two ways. In the first part of the study local PDT effects on the intestine were analyzed histologically. Violet laser light (lambda: 406-415 nm) was applied as a 2 cm diameter spot on the intestine 3 h after intraperitoneal (i.p.) administration of 50 mg/kg ALA. (A) Histological tissue samples were taken 0 min, 6 h and 1, 2 and 3 days after treatment (optical dose 3.2 J/cm(2)). Immediately after local PDT (3.2 J/cm(2), 50 mg/kg ALA) showed no effect on the intestine. However, 6 h post PDT there was complete destruction of the mesothelial lining and the outer (longitudinal) smooth muscle. Ganglion cells of the myenteric (Auerbach) plexus were also destroyed. The inner circular smooth muscle, the muscularis mucosa and the lamina propria were unharmed. Marked lymphectasia was present at this time. (B) To determine the threshold light dose of tissue destruction caused by PDT, different optical doses (1.6, 3.2, 6.4 J/cm(2)) were administered and histologic analysis of tissue samples were obtained I day post treatment. Destruction of the entire external musculature, submucosal structures and muscularis mucosa of the intestine at the illumination site could be observed above 1.6 J/cm(2) (50 mg/ka ALA). In the second part of the study whole peritoneal cavity PDT (WPC-PDT) was performed by illumination of the whole peritoneal cavity with 1.6 j/cm(2) Violet light 3 h after ALA administration using different drug doses (200, 100 and 50 mg/kg). WPC-PDT showed lethal toxicity with a drug dose above 50 mg/kg ALA at 1.6 j/cm(2). The probable cause of death in the first 3 days after IP-PDT was rhabdomyolysis, whereas when death occurred at longer time intervals, megaintestine associated with significant damage could be observed; however, without perforation of the intestinal wall. Conclusion: In rats WPC-PDT with 50 mg/kg ALA, 1.6 J/cm2 at lambda=415 nm was the maximum tolerable light dose. This dose is likely to be above the threshold of destruction of ovarian cancer micrometastasis. (C) 2002 Elsevier Science BY All rights reserved

Background and Objective: Cryogen sprays are used for cooling human skin during various laser treatments. Since characteristics of such sprays have not been completely understood, the optimal atomizing nozzle design and operating conditions for cooling human skin remain to be determined. Materials and Methods: Two commercial cryogenic spray nozzles are characterized by imaging the sprays and the resulting areas on a substrate, as well as by measurements of the average spray droplet diameters, velocities, temperatures, and heat transfer coefficients at the cryogen-substrate interface; all as a function of distance from the nozzle tip. Results: Size of spray cones and sprayed areas vary with distance and nozzle. Average droplet diameter and velocity increase with distance in the vicinity of the nozzle, slowly decreasing after a certain maximum is reached. Spray temperature decreases with distance due to the extraction of latent heat of vaporization. At larger distances, temperature increases due to complete evaporation of spray droplets. These three variables combined determine the heat transfer coefficient, which may also initially increase with distance, but eventually decreases as nozzles are moved far from the target. Conclusion: Sprayed areas and heat extraction efficiencies produced by current commercial nozzles may be significantly modified by varying the distance between the nozzle and the sprayed surface. Lasers Surg. Med. 28:113‐120, 2001. fl 2001 Wiley-Liss, Inc.

Background and Objective: Dynamics of cryogen spray deposition, water condensation and frost formation is studied in relationship to cooling rate and efficiency of cryogen spray cooling (CSC) in combination with laser dermatologic surgery. Study Design/Materials and Methods: A high-speed video camera was used to image the surface of human skin during and after CSC using a commercial device. The influence of ambient humidity on heat extraction dynamics was measured in an atmosphere-controlled chamber using an epoxy block with embedded thermocouples. Results: A layer of liquid cryogen may remain on the skin after the spurt termination and prolong the cooling time well beyond that selected by the user. A layer of frost starts forming only after the liquid cryogen retracts. Condensation of ambient water vapor and subsequent frost formation deposit latent heat to the target site and may significantly impair the CSC cooling rate. Conclusion: Frost formation following CSC does not usually affect laser dosage delivered for therapy of subsurface targets. Moreover, frost formation may reduce the risk of cryo-injury associated with prolonged cooling. The epidermal protection during CSC assisted laser dermatologic surgery can be further improved by eliminating the adverse influence of ambient humidity. Lasers Surg. Med. 28:469‐476, 2001. fl 2001 Wiley-Liss, Inc.

Cryogen spray cooling (CSC) has been used for selective epidermal cooling of human skin during laser therapy of patients with port wine stain (PWS) birthmarks. Unfortunately, current commercial CSC devices do not provide optimal cooling selectivity and, therefore, provide insufficient epidermal protection for some PWS patients. To assist in the development of improved atomizing nozzle designs, a reliable method to quantify the CSC heat flux is needed. We introduce a novel method to determine the heat flux (q/sub s/) and heat transfer coefficient (h) at the surface of a sprayed object, based on measurements of steady-state temperature gradients along a thin copper rod during continuous cryogen spraying. For an atomizing nozzle of inner diameter d/sub N/ = 0.7 mm, we found that q/sub s/ varies from 15 to 130 W/cm/sup 2/ and h increases nonlinearly from 15000 to 35000 W/m/sup 2/.K in the explored range of surface temperatures (T/sub s/, from -32 to -7/spl deg/C). Values of q/sub s/ obtained with a wider diameter nozzle (d/sub N/ = 1.4 mm) are approximately twice as large than those of the narrow nozzle. The corresponding values of h are significantly higher (32000-40000 W/m/sup 2/.K) and almost independent of T/sub s/ within the same temperature range. When combined with fast flashlamp photography (FFLP) of spray shapes and sprayed surfaces, the results demonstrate that the liquid cryogen layer, as deposited by finely atomized sprays from narrower nozzles, can significantly impair q/sub s/. In contrast, the higher-momentum impact of coarser sprays from wider nozzles reduces the thickness of the liquid layer in the impact area and/or enhances convection within it, yielding a larger q/sub s/.

The basic principles of a non-contact, near-infrared technique for the mapping of layered tissues are discussed theoretically and verified experimentally. The propagation properties of diffuse photon-density waves in tissues depend on the optical properties of the tissue. When a layered medium is irradiated by amplitude modulated light, the difference in optical properties between the layers is evident in the phase and amplitude of the diffuse reflection coefficient, which is a result of the interference of the partial waves propagating in the different layers. Thus, diffuse photon- density waves are applicable to the analysis of the structure of layered tissue. The probing depth is determined by the modulation frequency of the incident light. For modulation frequencies between several hundred megahertz and a few gigahertz, this allows us to analyze the properties of muscle tissue of up to 4-8 mm below the surface. Experimental results based on chicken breast muscle are given. As an example, the technique might be of use for evaluating the depth of necrosis and the blood volume fraction in deep burns.

The epidermal melanin content affects most dermatologic treatments involving light, and can limit the therapeutic success significantly. Therefore, knowledge of the optical properties of skin is required. This study investigates how the concentration of melanin influences visible reflectance spectra of skin and the relationship to threshold radiant energy fluence for melanosomal or melanocyte destruction. Reflectance spectra were measured at 28 pigmented human skin sites in vivo. For Asian and Caucasian subjects, measured reflectance values varied over the same range, while significantly lower values were recorded for African individuals. Epidermal melanin absorption coefficients measured at 694 nm were about 2500 m-1 for African, and 300–1200 m-16 for Caucasian and Asian skin. Twenty-five skin sites were exposed to ruby laser pulses (694 nm), where the pulse duration was long enough to allow heat diffusion between melanosomes. Hypopigmentation occurred, on average, at 12 and 26 J cm-2 for sun-exposed and sun-protected white skin, respectively, while slightly lower threshold values resulted from the measured spectra. As visible reflectance spectra reveal information regarding skin pigmentation and individual threshold doses for melanosomal damage, a use as a diagnostic tool in various dermatological laser treatments is apparent.

Photodynamic therapy (PDT) based on topical application of photosensitizers is currently in clinical use for the treatment of basal cell carcinoma of the skin, and it has been evaluated in animal models for photo-ablation of the endometrium. This paper presents a dosimetry model which indicates that a limiting factor in treating thick tumours will be the transport of the drug into the tumour rather than depletion of the optical distribution. The model predicts that an optical irradiation of 100 mW cm−2 at 635 nm for 20 min, ie well below the threshold for hyperthermic reaction, will give an adequate light dose to a depth of 3 mm. The time required for photosensitizers to diffuse to this depth is in the range of 3–15 h, dependent on the diffusion properties of the tissue.

Thirty patients were treated with a flashlamp-pumped pulsed dye laser, with 0.45 ms pulse width and 585 nm wavelength. Punch biopsies were taken prior to treatment, and the biopsies were examined morphometrically. Three different test sites were exposed to laser light of fluence 5.25, 6.50 and 7.75 J/cm2. The degree of blanching was examined 6-8 weeks after treatment, and each site was retreated four times. Six patients (20%) achieved poor blanching, eight patients (27%) obtained moderate lightening and 16 patients (53%) showed good response. The vessels of the good responders were located significantly more superficially than the vessels of the moderate and poor responders (P < 0.05). The poor responders had significantly smaller vessels than the moderate and good responders (P < 0.01). The moderate responders had deeper, but larger vessels, than the poor responders. Hence, an increasing vessel diameter reduces the negative outcome of increasing vessel depth. The vessel diameter was correlated to the colour (P < 0.01), e.g. the mean vessel diameter was increasing from 16.5 microns in pink lesions to 51.2 microns in purple lesions. The vessel depth was partly reflected in the lesional colour, as the pink and purple lesions had significantly deeper vessels than the red ones (P = 0.02). These results indicate that pink lesions predict poor blanching due to deeply located small vessels, while red lesions predict a good therapeutic result because of more superficially located vessels.

We are involved in the development of photodynamic therapy (PDT) as a minimally invasive method for treating dysfunctional uterine bleeding, one of the primary clinical indications for hysterectomy. In this paper, we analyse light propagation through the uterus in order to specify the requirements for a light delivery system capable of effectively performing endometrial PDT. Our approach involves developing an analytical model based on diffusion theory to predict optical fluence rate distributions when cylindrical and spherical optical applicators are placed in the uterine cavity. We apply the results of our model calculations to estimate the thermal effects of optical irradiation and the effective photodynamic optical dose. Theoretical fluence rate calculations are compared to fluence rate measurements made in fresh, surgically removed human uteri. Our results show that a trifurcated cylindrical optical applicator inserted into the human uterus can provide a light dose that is sufficient to cause photodynamic destruction of the entire endometrium. When the optical power per unit length of each cylindrical applicator is 100 mW cm-1 (at 630 nm), a fluence rate of 40 mW cm-2 is delivered to the boundary layer between the endometrium and the myometrium (a depth of about 4-6 mm). The optical fluence delivered to the boundary layer after 20 min of exposure is 50 J cm-2, a level that is generally accepted to cause tissue damage throughout the endometrium in most patients.

Background and Objective: Photodynamic therapy (PDT) based on topical application of photosensitizers has been under development over the last years. Typical applications are treatment of basal cell carcinoma of the skin and photoablation of the endometrium. The dosimetry for topically administered photosensitizers must take a time-dependent inhomogenous drug distribution into account together with the conventional parameters such as optical scattering, absorption, and photobleaching. Study DesignlMuterials and Methods: This study presents a dosimetry model where the cytotoxic dose is calculated in a stepwise procedure. The first step calculates the time-dependent distribution of 5-aminolevulinic acid GALA) from diffusion theory. In skin this distribution is dependent on drug permeability through the stratum corneum, on the diffusivity of dermis and epidermis, on the drug clearance time, and on the conversion rate from 5-ALA to protoporphyrin IX (PpIX). In the second step the distribution of PpIX is calculated from the 5-ALA distribution found in the first step taking the dynamics of the biosynthesis of 5-ALA to PpIX and the clearance time of PpIX into account. In the third step the generation of cytotoxic singlet oxygen is calculated from the optical distribution during irradiation, taking a photobleaching mechanism into account. Results: The distribution of cytotoxic oxygen is predicted from the optical dose, the drug dose, and the time between the application of the drug and the irradiation. Conclusion: The presented dosimetry model is made as simple as possible, yet composite enough to enable all relevant parameters to be taken into account. The model that is based on a linear theory in a semi-infinite medium can, if required, be extended to take nonuniform and nonlinear phenomena into account. 0 1996 Wiley-Liss, Inc.

When a cryogen spurt is applied to the skin surface for an appropriately short period of time (on the order of tens of milliseconds), the spatial distribution of cooling remains localized in the normal overlying epidermis, while leaving the temperature of the deeper port wine stain (PWS) blood vessels unchanged. Furthermore, cooling continues after pulsed laser exposure as cryogen remaining on the surface evaporates and removes heat deposited by light absorption in epidermal melanin. An additional advantage of dynamic cooling is a reduction in the level of pain and discomfort associated with flashlamp-pumped pulsed dye laser therapy of PWS. Preliminary clinical studies and supporting theoretical calculations demonstrate the feasibility of selective epidermal cooling while achieving photothermolysis of blood vessels during pulsed laser treatment of PWS.

We describe a non-contact method using infrared radiometry to determine lateral physical dimensions of laser heated subsurface chromophores in biological materials. An imaging equation is derived that relates measured radiometric temperature change to the reduced two-dimensional temperature increase of laser heated chromophores. From measured images of radiometric temperature change, the lateral physical dimensions of chromophores positioned in an in vitro model of human skin are determined by deconvolution of the derived imaging equation using a non-negative constrained conjugate gradient algorithm. Conditions for optimum spatial resolution are found by analysis of a derived radiometric transfer function and correspond to superficial chromophores and/or weak infrared absorption in a laser irradiated biological material. Analysis indicates that if the infrared attenuation coefficient is sufficiently small (i.e., less than 10mm-1), infrared radiometry in combination with a deconvolution algorithm allows estimation of lateral physical dimensions of laser heated subsurface chromophores in human skin.

Monte Carlo simulation of laser energy deposition in a port-wine stain (PWS) skin model and numerical solution of the thermal diffusion equation have been used to calculate threshold energies for thermal injury of PWS blood vessels for different vessel sizes and laser pulse durations. It has been assumed that an average vessel temperature rise of 65 ‡C causes thermal injury to the blood vessel. The result is that for a certain combination of wavelength, pulse duration and incident energy density, only a limited range of blood vessel sizes can be injured optimally. Higher energy densities are required to injure smaller vessels with the same pulse duration, spot size and wavelength. This gives support to the mechanisms of selective photothermolysis suggested previously by Anderson and Parrish, although their model was based on the cooling behaviour of instantaneously heated vessels. The authors hypothesize that different laser parameter settings that match the individual PWS vessel anatomy during treatment will be used in the future, instead of many treatments with the same laser parameters. This could lead to less treatment sessions and to an improved predictability of clinical results.

Selective photothermolysis with pulsed lasers is presumably the most successful therapy for port-wine stain birthmarks (flammeus nevi). Selectivity is obtained by using an optical wavelength corresponding to high absorption in blood, together with small absorption in tissues. Further on, the pulse length is selected to be long enough to allow heat to diffuse into the vessel wall, but simultaneously short enough to prevent thermal damage to perivascular tissues. The optical wavelength and pulse length are therefore dependent on vessel diameter, vessel wall thickness and depth in dermis. The present work demonstrates that in the case of a 0.45 ms long pulse at 585 nm wavelength, vessels of 40–60Μm require minimum optical fluence. Smaller vessels require higher fluence because the amount of heat needed to heat the wall becomes a substantial fraction of the absorbed optical energy. Larger vessels also require a higher dose because the attenuation of light in blood prevents the blood in the centre of the lumen from participating in the heating process. It is shown that the commonly used optical dose in the range of 6–7 J cm−2 is expected to inflict vessel rupture rather than thermolysis in superficially located vessels. The present analysis might serve to draw guidelines for a protocol where the optical energy, wavelength and pulse length are optimized with respect to vessel diameter and depth in dermis.

Photodynamic therapy is currently being evaluated as a minimally invasive procedure for endometrial ablation not requiring anaesthesia. Light penetration depths at 630, 660 and 690 nm and the optimal configuration of intrauterine light-diffusing fibres were determined in 14 human uteri to assist in the design of a light intrauterine device. Post-menopausal ex-vivo uteri showed a significantly lower light penetration depth than pre-menopausal uteri. With a single central diffusing fibre inserted, the fluence rate measured in the uterine wall at the most remote point of the cavity decreased to 1.1 +/- 0.4% of that measured at closest proximity, whereas it decreased to only 40.0 +/- 9.0% with three fibres. Distension of the uterine cavity with 2 ml of an optically clear fluid increased the fluence rate at the fundus between the fibres at a depth of 2 mm by a factor of 4. We conclude that in normal-sized pre-menopausal uterine cavities, three diffusing fibres will deliver an optical dose above the photodynamic threshold level at a depth of 4 mm, even in the most remote areas, in < 30 min without causing thermal damage. For distorted and elongated cavities, either slight distension of the cavity or the insertion of a fourth diffusing fibre is required.

The clinical objective in laser treatment of port-wine stains (PWS) is to induce selective photothermolysis of subsurface blood vessels without damaging the overlying epidermis. This paper investigates the effectiveness of ‘dynamic’ cooling, where a cryogen is sprayed on the skin surface for an appropriately short period of time, to eliminate epidermal thermal injury during laser treatment of PWS. Comparative measurements of radiometric surface temperature from cooled and uncooled laser irradiated (585 nm) PWS sites, and theoretical predictions of temperature distributions within skin in response to dynamic cooling in conjunction with laser irradiation are presented. Rapid reduction of skin surface temperature and localization of cooling the epidermis are obtained when a cryogen is sprayed on skin prior to laser irradiation. Successful blanching of the PWS without thermal injury to the overlying epidermis is accomplished.

Port-wine stain is a congenital birthmark consisting of an abnormal density of blood vessels in the upper dermis. The enlarged blood volume gives the lesion a red to purple colour. The aim of the treatments is to destroy the vessels to the extent necessary for obtaining a normal skin coloration. Thus, in principle, all relevant information about the lesion should be contained in a reflectance spectrum in the visible. However, the relation between the reflectance spectrum and tissue parameters such as scattering, melanin content and blood distribution is somewhat composite. This work tries to enlighten this relation in terms of a very simple analytical mathematical model, and it is demonstrated that such a model at least will contribute to a qualitative understanding of the relevance of the various parameters.

The relationship between photothermal damage to blood vessels of diameter,d, and laser pulse duration,t p, was verified in a series of studies using the chick chorioallantoic membrane (CAM). A total of 879 individual CAM blood vessels (d=50–130μm) was irradiated, using a laser pulse duration of 0.45 or 10 ms. Laser-induced vascular damage was observed in real time, recorded on videotape, and evaluated in a double-blind fashion. Permanent damage was confirmed by inspection 24 h after laser exposure. Under the conditions of this experiment, only when laser pulse durations are approximately equal to the estimated thermal relaxation times (τ) of the CAM microvessels can the critical core intravascular temperature, necessary to destroy vessels irreversibly, be achieved and sustained for sufficient time. Shorter pulse durations are more effective for damaging smaller blood vessels; conversely, longer pulse durations are more effective for damaging larger diameter vessels.

The ability to control the degree and spatial distribution of cooling in biological tissues during a thermally mediated therapeutic procedure would be useful for several biomedical applications of lasers. The authors present a theory based on the solution of the heat conduction equation that demonstrates the feasibility of selectively cooling biological tissues. Model predictions are compared with infrared thermal measurements of in vivo human skin in response to cooling by a cryogen spurt. The presence of a boundary layer, undergoing a liquid-vapour phase transition, is associated with a relatively large thermal convection coefficient ( approximately=40 kW m-2 K-1), which gives rise to the observed surface temperature reductions (30-40 degrees C). The degree and the spatial temporal distribution of cooling are shown to be directly related to the cryogen spurt duration.

Individual blood vessels in the chick choriallantoic membrane were selectively coagulated through photothermolysis, using pulsed laser irradiation at 585 nm. Pulse durations were chosen to be 0.45 ms and 10 ms, which correspond to the thermal relaxation times in blood vessels of 30 microns and 150 microns diameter, respectively. The short pulses, at a light fluence F = 3 Jcm-2, caused permanent occlusion of vessels of 40 microns diameter or less, whereas larger caliber vessels (60-120 microns) required F = 4-5 Jcm-2. The long-duration pulses, at F = 7 Jcm-2, caused coagulation of the larger diameter vessels; the small-caliber vessels and capillaries showed resistance to photothermolysis and required multiple exposures to achieve coagulation. The fluence versus diameter (F versus d) relationship for coagulation was calculated for the two pulse durations. The energy deposited in a cylindrical absorber of diameter d by an optical field, incident perpendicular to the vessel, was expressed analytically and compared with the energy required to coagulate a blood vessel of the same lumen dimeter. When thermal diffusion is incorporated into the model, our findings can be accounted for quantitatively. This information will be of use for improving the laser treatment of port wine stains and other vasculopathies. A surprising observation was that arterioles were damaged at lower incident energy densities than venules having the same lumen diameter, despite the fact that absorbance in oxygen-rich and oxygen-poor blood is the same at 585 nm.

Vulnerable plaques constitute a risk for serious heart problems, and are difficult to identify using existing methods. Hyperspectral imaging combines spectral- and spatial information, providing new possibilities for precise optical characterization of atherosclerotic lesions. Hyperspectral data were collected from excised aorta samples (n = 11) using both white-light and ultraviolet illumination. Single lesions (n = 42) were chosen for further investigation, and classified according to histological findings. The corresponding hyperspectral images were characterized using statistical image analysis tools (minimum noise fraction, K-means clustering, principal component analysis) and evaluation of reflectance/fluorescence spectra. Image analysis combined with histology revealed the complexity and heterogeneity of aortic plaques. Plaque features such as lipids and calcifications could be identified from the hyperspectral images. Most of the advanced lesions had a central region surrounded by an outer rim or shoulder-region of the plaque, which is considered a weak spot in vulnerable lesions. These features could be identified in both the white-light and fluorescence data. Hyperspectral imaging was shown to be a promising tool for detection and characterization of advanced atherosclerotic plaques in vitro. Hyperspectral imaging provides more diagnostic information about the heterogeneity of the lesions than conventional single point spectroscopic measurements. © (2011) Society of Photo-Optical Instrumentation Engineers (SPIE)

Orthotopic bladder cancer model in rats mimics human bladder cancer with respect to urothelial tumorigenesis and progression. Utilizing this model at pT1 (superficial stage), we analyze the tissue responses to hexyl 5-aminolevulinate-induced photodynamic therapy (HAL-PDT). In comparison to untreated rats, HAL-PDT causes little change in tumor-free rat bladder but induces inflammatory changes with increased lymphocytes and mononuclear cell infiltration in rat bladders with tumor. Immunohistochemistry reveals that HAL-PDT is without effect on proliferating cell nuclear antigen expression within the tumor and increases caspase-3 expression in both normal urothelium and the tumor. Transmission electron microscopy reveals severe mitochondrial damage, formations of apoptotic bodies, vacuoles, and lipofuscin bodies, but no microvillus-formed niches in HAL-PDT-treated bladder cancer rats. Bioinformatics analysis of the gene expression profile indicates an activation of T-cell receptor signaling pathway in bladder cancer rats without PDT. HAL-PDT increases the expression of CD3 and CD45RA in the tumor (determined by immunohistochemistry). We suggest that pathways of action of HAL-PDT may include, at least, activations of mitochondrial apoptosis and autophagy, breakdown of cancer stem cell niches, and importantly, enhancement of T-cell activation. Open Access

Experimental results for the optical distribution and temperature rise during laser irradiation of tumors are presented. The experimental conditions are chosen to simulate laser irradiation of ocular tumors. The tumor models are human retinoblastoma heterotransplanted in athymic mice, murine mammary carcinoma in C3H/HEJ mice, and B16 melanotic melanoma in C57/BL6 mice. The experimental results are discussed in terms of a mathematical model where the thermal distribution is calculated from the bioheat transfer equation, and the optical distribution is determined according to diffusion theory.

Forensic medicine is a field of medicine where technology plays an increasingly important role in securing and evaluating evidence in, for example, child abuse cases and cases of domestic violence. Methods from chemistry and biological sciences have found a wide application within forensic medicine. Optical technologies like microscopy are also widely used. Despite this, in vivo or post mortem optical diagnostics by spectroscopy have traditionally not had an important role in clinical or forensic examinations. Forensic medical optics as a field might include all kinds of optical analysis for use within forensic science. This includes everything from microscopic techniques to methods for examination of evidence from a crime scene. This chapter will, however, focus on the use of optical diagnostics for examining skin, with a focus on identification, characterization and age determination of minor traumatic injuries like skin bruises.

Imaging modalities like hyperspectral imaging create large amounts of data. Time efficient, automated analytic techniques are therefore required to enjoy the power of such methods. In this study it was investigated if hyperspectral imaging followed by automated noise filtering and statistical image analysis is a suitable method for characterization of the macroscopic structure of atherosclerotic lesions. Ten human aorta samples (6x8 cm) were collected during autopsy. Hyperspectral white light and fluorescence images and 5 – 6 biopsies were collected from each sample. The biopsies were stained (HES, Sudan red), and grouped according to histology. All images were noise filtered and normalized. Fluorescence spectra were collected from all biopsied regions, and used to compute average spectra for each histological group. Supervised classification was performed using Spectral angle mapping (SAM) with the average spectra as endmembers. K-means- and ISO-data clustering was used for unsupervised classification. The results show that noise filtering and normalization is essential for reliable classification. Supervised classification was in general found to perform better than unsupervised classification. However, the SAM results strongly depend on the variation in the spectra used to compute the average endmember spectra. The analysis show that fatty deposits, calcifications, connective tissue and hemoglobin can be identified. The lesions were found to have a complex structure where vulnerable regions could be found next to stabile regions within the same lesion. In conclusion hyperspectral imaging, automated filtering and -analysis was found to be a suitable tool to classify advanced atherosclerotic lesions.

Hyperspectral imaging is a modality which combines spatial resolution and spectroscopy in one technique. Analysis of hyperspectral data from biological samples is a demanding task due to the large amount of data, and due to the complex optical properties of biological tissue. In this study it was investigated if depth information could be revealed from hyperspectral images using a combination of image analysis and analytic simulations of skin reflectance. It was also investigated if hyperspectral imaging could be utilized to monitor changes in the distribution of hemoglobin species after smoking. Hyperspectral data in the wavelength range 400-1000nm were collected from the forearm of 15 non-smokers and 5 smokers. The hyperspectral images were analyzed with respect to the distribution of hemoglobin species and vascular structures. Changes in the vascular system due to smoking were also evaluated. Principal component analysis (PCA), Spectral angle mapping (SAM), and Mixture tuned matched filtering (MTMF) were used to enhance vascular structures. Emphasis was put on identifying apparent and true absorption spectra for the present chromophores by combining image analysis and an analytical photon transport model. The results show that the depth resolution of hyperspectral images can be better understood using analytical simulations. Modulation of the chromophore spectra by the optical properties of overlying tissue was found to be an important mechanism causing the depth resolution in hyperspectral images. It was also found that hyperspectral imaging and image analysis can be successfully applied to quantify skin changes following smoking.

Collimated light sources in turbid media are difficult to describe within the diffusion approximation, because they do not meet the requirement of near isotropy. For precise calculation of light intensities close to the source, alternative descriptions of the light source are necessary. In this paper the transition of collimated light into diffusivity is studied by Monte Carlo simulations. On the basis of these simulations and the diffusion approximation a hybrid approach is designed and used to analyze approaches based on analytic source terms. The influence of boundaries to air is studied. The benefits of increased approximation orders are investigated. It is shown that, even in the presence of strong absorption, the diffusion approach can give satisfactory results if only the source terms are suitably chosen.

The rationale of hyperthermic treatment of tumours is based on the phenomenon that several tumours have a reduced tolerance for exposure to temperatures in the region 42–47 °C for 20–30 min. Hyperthermia may also be used as an adjunct to other modalities such as chemotherapy, conventional radiation therapy or photodynamic therapy.

Cardiovascular disease is the primary cause of death in the United States; the majority of these deaths are caused by the rupture of vulnerable plaques. An important feature of vulnerable plaques is the thickness of the fibrous cap that covers the necrotic core. A thickness of less than 65 microm has been proposed as a value that renders the plaque prone to rupture. This work shows that multiphoton microscopy (MPM) can image the plaque with microm resolution to a depth deeper than 65 microm. The fibrous cap emits primarily second harmonic generation due to collagen, in contrast to the necrotic core and healthy artery, which emits primarily two-photon excited fluorescence from elastin. This gives a good demarcation of the fibrous cap from underlying layers, facilitating the measurement of the fibrous cap thickness. Based on a measure of the collagen/elastin ratio, plaques were detected with a sensitivity of 65% and specificity of 81%. Furthermore, the technique gives detailed information on the structure of the collagen network in the fibrous cap. This network ultimately determines the mechanical strength of the plaque. A mechanical model based on this information could yield a measure of the propensity of the plaque to rupture.

Multiphoton microscopy is a techniques that fascilitates three dimensional imaging of intact, unstained tissue. Especially connective tissue has a relatively strong nonlinear optical response and can easily be imaged. Atherosclerosis is a disease where lipids accumulate in the vessel wall and there is a thickening of the intima by growth of a cap of connective tissue. The mechanical strength of this fibrous cap is of clinically importance. If the cap ruptures a thrombosis forms which can block a coronary vessel and therby causing myocardial infarction. Multiphoton microscopy can be used to image the fibrous cap and thereby determine the thickness of the cap and the structure of the connective fibres. This could possibly be developed into a diagnostic and clincal tool to monitor the vulnerability of a plaque and also to better understand the development of a plaque and effects of treatment. We have collected multiphoton microscopy images from atherosclerotic plaque in human aorta, both two photon excited fluorescens and second harmonic generated signal. The feasability of using this technique to determine the state of the plaque is explored.

Studies of immediate skin reactions are important to understand the underlying biological mechanisms involved in traumatic or chemical damage to the skin. In this study the spatial and spectral information provided by hyperspectral images was used to identify and characterize non-penetrating skin injuries in a porcine model. A hyperspectral imaging system (Hyspex, Norsk Elektro Optikk AS) was used to monitor the temporal development of minor skin injuries in an anesthetized Norwegian domestic pig. Hyperspectral data were collected in the wavelength range 400-1000 nm (VNIR), with a spectral sampling interval of 3.7 nm. The measurements were initiated immediately after inflicting the injury, and were repeated at least five times at each site with irregular frequency. The last measurement was performed 4 hours after injury. Punch biopsies (5 mm), were collected from adjacent normal skin, and at the center and the margin of each injury. The study was approved by the national animal research authority. The hyperspectral data were analyzed with respect to oxy- and deoxyhemoglobin, and erythema index. The skin biopsies were examined to determine the extent of skin damage in the bruised zones. Preliminary results show that hyperspectral imaging allows discrimination between traumatized skin and normal skin in an early phase. The extent and location of the hemorrhages can be determined from hyperspectral images. These findings might contribute to a better understanding of immediate skin reactions to minor trauma, and thereby the development of a better diagnostic modality for non-penetrating skin injuries in forensic medicine.

Determining the cause of an injury and the force behind the impact may be of crucial importance in a court case. For non-penetrating soft tissue injuries there is a lack of information available in the literature. In this study controlled bruises were inflicted on an anesthetized pig by high speed, low-weight paintball projectiles (diameter 17.1 mm, weight 3.15 g). The speed of the object and the impact itself were monitored using a high speed camera. Punch biopsies (5 mm) were collected from the injury sites. A red and purple ring with a diameter of 1.5 cm appeared on the skin within 30 seconds after the paintball impact. The ring was visually fully established after 5-10 minutes. Numerical finite element simulations performed with ABAQUS\Explicit showed a build up of shear stresses in the skin where the ring formed. Biopsies demonstrated severe dermal vessel damage in the same area. It is concluded that considerable shear stresses during the impact will create dermal vessel damage and thereby cause a visible bruise. Larger forces are required for compressive stresses to inflict equivalent vascular damage.

Photodynamic therapy (PDT) is a treatment modality which has been shown to be effective for both malignant and non-malignant diseases. New photosensitizers such as hexyl-aminolevulinate (HAL) may increase the efficiency of PDT. HAL penetrates into the cell where the photosensitizer protoporphyrin IX (PPIX) is produced endogenously. In a previous study on HAL based PDT treatment of rat bladder cancer (AY-27 transitional cell carcinoma), a depression of the optical reflectance spectra after treatment was observed in some of the animals. This depression of the spectra was caused by metHemoglobin (metHb). MetHb is an indication of oxidative stress, and can be formed as a result of for instance UV-radiation and heating of blood. The aim of this study was to identify if metHb can be formed in vitro as a result of oxidative stress caused by singlet oxygen and ROS produced during PDT. Methemoglobin formed during PDT might thus be used as an indirect measure of the photochemical processes. This may help predict the PDT treatment outcome. Red blood cells mixed with AY-27 cells exposed to HAL, or PPIX received light treatment, and the changes in the absorption spectra were measured spectrophotometrically. The methemoglobin absorbance spectrum was also studied, and found to be strongly dependant on pH. Hemolysis of erythrocytes by PDT was found, however no metHb was formed in vitro.

Photodynamic therapy (PDT) is a treatment modality which has been shown to be effective for both malignant and non-malignant diseases. New photosensitizers such as 5-aminolevulinic acid hexylester (hALA) may increase the efficiency of PDT. Monitoring of the tissue response provides important information for optimizing factors such as drug and light dose for this treatment modality. Optical spectroscopy may be suited for this task. To test the efficacy of hALA induced PDT, a study on rats with a superficial bladder cancer model, in which a bladder cancer cell line (AY-27) is instilled, will be performed. Preliminary studies have included a PDT feasibility study on rats, fluorescence spectroscopy on AY-27 cell suspensions, and optical reflection and fluorescence spectroscopy in rat bladders in vivo. The results from the preliminary studies are promising, and the study on hALA induced PDT treatment of bladder cancer will be continued.

When performing intravascular spectroscopy there is always the possibility that there will be a thin layer of blood between the probe and the vessel wall. This will affect measurements espescially below 600 nm where blood absorption is strong (important for fluorescence and Raman measurements) and above 1400 nm where the water absorption is strong. Between these two regions (the theraputic window) the absorption is fairly low and can be neglected. This article explores the possibility of performing measurements outside the optical window by minimizing blood layer and increasing the excitation signal.

Bruises can be important evidence in legal medicine, for example in cases of child abuse. Optical techniques can be used to discriminate and quantify the chromophores present in bruised skin, and thereby aid dating of an injury. However, spectroscopic techniques provide only average chromophore concentrations for the sampled volume, and contain little information about the spatial chromophore distribution in the bruise. Hyperspectral imaging combines the power of imaging and spectroscopy, and can provide both spectroscopic and spatial information. In this study a hyperspectral imaging system developed by Norsk Elektro Optikk AS was used to measure the temporal development of bruised skin in a human volunteer. The bruises were inflicted by paintball bullets. The wavelength ranges used were 400 - 1000 nm (VNIR) and 900 - 1700 nm (SWIR), and the spectral sampling intervals were 3.7 and 5 nm, respectively. Preliminary results show good spatial discrimination of the bruised areas compared to normal skin. Development of a white spot can be seen in the central zone of the bruises. This central white zone was found to resemble the shape of the object hitting the skin, and is believed to develop in areas where the impact caused vessel damage. These results show that hyperspectral imaging is a promising technique to evaluate the temporal and spatial development of bruises on human skin.

An objective method for dating of soft tissue injuries is highly desired in forensic medicine. To achieve this, obtaining a fundamental understanding of extravascular blood behavior is necessary. Porcine blood mixed with fluorescein was inserted subcutaneously on an anaesthetized pig. The fluorescence build-up was measured as a function of time, using a fiber probe setup. Fluorescence measurements proved to be a suitable tool for monitoring the temporal distribution of fluid in a subcutaneous hemorrhage. Reasonable values for the fluid velocity was obtained by monitoring the fluorescein distribution around the infusion point. Further theoretical work and development of the experimental technique is necessary to obtain a good understanding of subcutaneous transport of blood.