To assess laser therapies in clinical practice, histologic examinations were commonly used. But histologic examinations were invasive and not real-time in nature. In this work, we validate multiphoton microscopy as a useful modality in evaluating laser-tissue reaction in vivo. Multiphoton microscopy based on femtosecond titanium-sapphire laser system were used to evaluate autoflurescence (AF) and second harmonic generation (SHG). Nude mouse skin was irradiated with Erbium:YAG laser at low to high fluence. High resolutional images can be obtained by multiphoton microscopy. At low fluence, Erbium:YAG laser can selectively loosen compact stratum corneum with minimal injury to basal layer. At high fluence, ablated keratinocytes and residual debris can be imaged. The laser thermal effect on dermis could be measured by SHG signals of collagen fibers. SHG decreased as laser fluence increased. Multiphoton microscopy is a useful in-vivo technique in evaluating ablative and thermal effects of Erbium:YAG laser on nude mouse skin.

The aim of this study is to characterize the ability of multiphoton microscopy in monitoring the transdermal penetration enhancing effect of a depilatory agent and the associated structural alterations of stratum corneum. The result is compared with the electron microscopic findings. Our results show that the penetration of both hydrophilic and hydrophobic agents can be enhanced. The morphology of corneocytes becomes a homogenized pattern with focal detachment of surface corneocytes. In combination with Nile red staining, multiphoton imaging also shows that the regular motar-like distribution of lipid matrix was disrupted into a homogenized pattern of lipid distribution. These results are well correlated with the findings of ultrastructural analysis by electron micrographs showing disintegration of the protein envelope of coenocytes, disruption of intracellular keratin and loss of the regular lamellar packing of intercellular lipids. We conclude that, in addition to quantifying the permeation profiles of model drugs, multiphoton microscopy is able to detect the penetration enhancer-induced structural alterations of stratum corneum.

Multiphoton optical tomography provides non-invasive optical sectioning of skin with high spatial and intracellular resolution. The imaging system DermaInspect based on femtosecond laser pulses and equipped with a SPC 830 card for FLIM analysis was used to perform this technique in vivo in clinical studies on patients with abnormal pigmented tissues. After examination in vivo, biopsies were taken and analyzed again. So far it was possible to distinguish between normal and diseased tissues mainly due to their different morphological appearances in the imaging mode: melanoma tissues show differences in the structure of the epidermal layers and the presence of dendritic cells compared to normal skin. The optical analysis can be improved when using the system in the time-correlated single photon counting mode. Fluorescence lifetime analysis reveals characteristic differences between healthy and melanoma tissues even before the visible morphological changes in human tissue in vivo.

As a main pigment in skin tissues, melanin plays an important role in photo-protecting skin from UV radiation. However, melanogenesis may be altered due to disease or environmental factors; for example, sun exposure may cause damage and mutation of melanocytes and induce melanoma. Imaging pigmentation changes may provide invaluable information to catch the malignant transformation in its early stage and in turn improve the prognosis of patients. We have demonstrated previously that transmission mode, two-photon, one- or two-color absorption microscopy could provide remarkable contrast in imaging melanin in skin. In this report we demonstrate significantly improved sensitivity, so that we are now able to image in epi-mode (or back reflection) in two-photon absorption. This improvement makes possible for us to characterize the different types of pigmentation on the skin in vivo at virtually any location. Another finding is that we can also image transient photothermal dynamics due to the light absorption of melanin. By carefully choosing excitation and probe wavelengths, we might be able to image melanin in different structures under different micro-environments in skin, which could provide useful photochemical and photophysical insights in understanding how pigments are involved in photoprotection and photodamage of cells.

Noninvasive evaluation of dermal degeneration is important for a diagnosis of photoaging. Polarization sensitive optical coherence tomography (PS-OCT) can measure the distribution of phase retardation caused by birefringence property of collagen. The PS-OCT has been reported as the technology of in vivo investigation of collagen structure in the skin. PS-spectral domain (SD)-OCT provides a three-dimensional phase retardation volume, and it provides an en face distribution map of the slopes of cumulative phase retardation. In this research, we measured the lateral orbital area and forehead of 7 young (age = 24.7±1.1) and 6 elder (age = 72.5±1.4) volunteers by a custom-built 840 nm fiber based PS-SD-OCT. This system can measure the skin birefringence three-dimensionally (5.5 sec for a 3-D volume). There was a significant difference in mean of the distribution of phase retardation slopes of the lateral orbital area between young and old cohorts (young: 0.356±0.0255 degree/μm; elder: 0.256±0.0118 degree/μm). However, there was no significant difference in forehead. On the other hand, the shapes of histogram was not normal distribution. Therefore, we calculated skewness and kurtosis of the distributions of slopes for the analysis. In the orbital area, there were also significant differences between young and old cohorts in skewness (y: 0.813±0.135; e: 1.51+/- 0.177) and kurtosis (y: 2.85±0.483; e: 5.23±0.862). In forehead, there was a significant difference between young and old cohorts in skewness (p=0.0289) in contrast with mean (p= 0.065).

Photophysical studies of UV-B sunscreens showed a measurable UV-A emission from padimate O (2-ethylhexyl-4-(dimethylamino)benzoate). Since recent studies associate UV-A to skin cancer induction pathways, as well as skin aging, we studied the effect of padimate O emission when applied to skin. After application of padimate O to skin the emission spectrum of skin showed a large increase in the intensity of 470 nm peak. The 470 nm emission in skin arises from a skin component, possibly collagen, which absorbs at about 360 nm, where padimate O emits. The excitation spectra of skin with padimate O measured at an emission wavelength of 468 nm show a peak at 310 nm with a broad shoulder at about 350 nm to 370 nm, which increased in intensity with time. However, the excitation spectrum of skin with octyl salicylate (another UV-B emitting sunscreen) did not show such a shoulder or increase in intensity. Thus, we attribute the presence of a shoulder in the excitation spectrum of skin and the increase in its intensity as evidence for energy transfer from padimate O to collagen. The transfer mechanism is not clear.

Raman spectroscopy is a non-invasive optical technique that is very sensitive to the structure and conformation of biochemical constitutes. It has been successfully used to study dysplasia and cancer in a variety of human tissues including skin. Researchers have a strong interest in quantitative analysis of the Raman spectra in the hope to find the quantitative molecule changes associated with cancers. Because skin is the largest organ in humans, it is very important to study the variability of Raman spectra for different body sites. In this presentation we report our quantitative analysis of normal skin Raman spectra of twenty-five body sites. Partial least square (PLS) analysis was used to quantify the skin molecules from the in vivo Raman spectra. We find that the absolute Raman intensity differs from person to person and from body site to body site. However the normalized Raman spectra are identical for the same body site. It is particularly interesting to find that the distribution of skin molecules is clustered for the same body regions.

There can be a large variation in response between laser treatments of vascular malformations like port-wine stains even in one patient. This could be ascribed to variations in the skin properties like tint (melanin) and perfusion (redness) which will influence the effectiveness of the laser dosimetry. To obtain a better understanding of the relation between skin properties just before treatment, laser dosimetry and clinical response, a multi-spectral dermatoscope is applied. A sequence of calibrated images is captured from 400 to 720 nm. Images at the treatment laser wavelength (532 nm) show the absorbing structures during laser exposure. Images of different treatment sessions of one patient were matched with dedicated registration software to quantify the results of the laser treatment (change in blood vessels structure, effect on pigment). For feasibility, images were collected from 5 patients and used to determine the optimal wavelength combination strategies. The image matching software gives an objective impression of the improvement, e.g. the clearing of the port-wine stain over time or pigment reactions, which will facilitate the discussion with the patient about the end point of treatment. The multi-spectral dermatoscope and software developed enables the evaluation of large patient series which will result in objective data to advise the dermatologist on the optimal laser dosimetry in future in relation to the skin properties.

A compact optical hyperspectral imager that can detect both spectral and polarization signatures was used for passive noninvasive imaging of human skin. This vibration-insensitive imager uses an acousto-optic tunable filter (AOTF) as a spectral selection element and an electronically tunable liquid crystal variable retarder (LCVR) as a polarization device. Such an imager is ideally suited to provide both agile spectral and polarization signatures and can be readily used for real time in vivo medical imaging applications. Operation of this imager and image acquisition is fully computer controlled. This imager covers visible to near-infrared (VNIR) region from 400 to 800 nm with a 10 nm spectral resolution at 600 nm and uses a TeO₂ AOTF with a 15×15 mm² linear aperture and a 4.2° angular aperture. At each wavelength 640×480 images with two orthogonal polarization are captured and a total of 41 spectral images are collected to form an image cube. A commercial Si CCD camera was used along with off-the-shelf lenses, mirrors and irises. We carried out experiments with a human subject and controlled the blood perfusion in the individual arm and finger by using a pressure cuff and a rubber band, respectively. Images were captured by illuminating the subject with a white light lamp source and imaging it from a distance. When the hyperspectral image analysis was performed we could observe the effects of skin deoxygenation. In this paper we will described our instrument, the experimental setup, the images obtained and the analysis results.

Laser speckle imaging modality is one of widely used methods to evaluate blood flow because of its simplicity. However, laser speckle image has a limitation in the evaluation of subcutaneous blood flow due to its low contrast perfusion image. Various methods have been tried to enhance the perfusion image contrast. Such methods presented positive results in some degree. However, it could not be fundamental solutions due to low penetration depth of lasers restricted by optical tissue scattering property. This study suggests a method to enhance the perfusion image contrast of laser speckle imaging modality by increasing the penetration depth of lasers. An optical clearing agent (glycerol) was topically applied on skin treated with micro-needle roller in order to reduce the time period of optical tissue clearing and therefore, enhance the penetration depth of laser. In this study, we investigated the effect of glycerol and micro-needling methods in the contrast enhancement of laser speckle perfusion skin image and presented the results of in-vitro and in-vivo animal experiment.

The local microcirculatory dynamics underlying phenomenon of urticarial dermographism (UD) are not yet sufficiently elucidated in dermatological patients. A fiber optic laser Doppler flowmeter (LDF) was used to monitor skin blood flow (SBF) changes on the back of the patients with UD before and after application of the series of pressure stimuli (9.8×10⁴, 14.7×10⁴, 19.6×10⁴ and 24.5×10⁴ Pa). All patients acted as self-controls to assess their disease activity by means of SBF values based on response to pressure stimuli before and after treatment with antihistamines, when compared to baseline SBF. Throughout 30 minutes evaluation inter-subject SBF values at pressure-tested sites were noticeably distinguished as high, moderate and low. By LDF we could differentiate the highest development of vascular dynamics after 5 minutes, coming back to normal within about 30 minutes in one group of patients, and the vascular dynamics reaching its maximum in 15 minutes, but with no fade after 30 minutes, in another group of patients. All treatment regimens in both groups of patients by LDF produced a measurable reduction already during 1-2 days of therapy, accompanied by a reduction in SBF baseline values in patients with severe and moderate symptoms of UD.

A polarized light camera (polCAM) is being used to guide surgical excision of skin cancers. Preliminary results of a clinical pilot study are presented. The polCAM uses linearly polarized light and acquires parallel- and cross-polarized images, then the difference image (par - cross) subtracts the background diffusely scattered light and yields an image the unveils the fabric of the skin. Cancer disrupts the complex pattern of this fabric, revealing the cancer margin.

Polarized fluorescence has shown some promising results in assessment of skin cancer margins. Researchers have used tetracycline and cross polarization imaging for nonmelanoma skin cancer demarcation as well as investigating endogenous skin polarized fluorescence. In this paper we present a new instrument for polarized fluorescence imaging, able to calculate the full fluorescence Stokes vector in one snapshot. The core of our system is a multi-aperture camera constructed with a two by two lenslet array. Three of the lenses have polarizing elements in front of them, oriented at 0°, + 45°and 90° with respect to light source polarization. A flash lamp combined with a polarizer parallel to the source-camera-sample plane and a UV filter is used as an excitation source. A blue filter in front of the camera system is used to collect only the fluorescent emission of interest and filter out the incident light. In-vitro tests of endogenous and exogenous polarized fluorescence on collagen rich material like bovine tendon were performed and Stokes vector of polarized fluorescence calculated. The system has the advantage of eliminating moving artifacts with the collection of different polarization states and stoke vector in a single snap shot.

In dermatology, various digital imaging modalities have been used as an important tool to quantitatively evaluate the treatment effect of skin lesions. Cross-polarization color image was used to evaluate skin chromophores (melanin and hemoglobin) information and parallel-polarization image to evaluate skin texture information. In addition, UV-A induced fluorescent image has been widely used to evaluate various skin conditions such as sebum, keratosis, sun damages, and vitiligo. In order to maximize the evaluation efficacy of various skin lesions, it is necessary to integrate various imaging modalities into an imaging system. In this study, we propose a multimodal digital color imaging system, which provides four different digital color images of standard color image, parallel and cross-polarization color image, and UV-A induced fluorescent color image. Herein, we describe the imaging system and present the examples of image analysis. By analyzing the color information and morphological features of facial skin lesions, we are able to comparably and simultaneously evaluate various skin lesions. In conclusion, we are sure that the multimodal color imaging system can be utilized as an important assistant tool in dermatology.

Preconditioning tissues with an initial mild thermal stress, thereby eliciting a stress response, can serve to protect tissue from subsequent stresses. Patients at risk for impaired healing, such as diabetics, can benefit from therapeutic methods which enhance wound repair. We present a laser thermal preconditioning protocol that accelerates cutaneous wound repair in a murine model. A pulsed diode laser (λ = 1.86 μm, τ_p = 2 ms, 50 Hz, H = 7.64 mJ/cm²) was used to precondition mouse skin before incisional wounds were made. The preconditioning protocol was optimized in vitro and in vivo using hsp70 expression, cell viability, and temperature measurements as benchmarks. Hsp70 expression was non-invasively monitored using a transgenic mouse strain with the hsp70 promoter driving luciferase expression. Tissue temperature recordings were acquired in real time using an infrared camera. Wound repair was assessed by measuring hsp70 expression, biomechanical properties, and wound histology for up to 24 d. Bioluminescence (BLI) was monitored with the IVIS 200 System (Xenogen) and tensile properties with a tensiometer (BTC-2000). The in vivo BLI studies indicated that the optimized laser preconditioning protocol increased hsp70 expression by 15-fold. The tensiometer data revealed that laser preconditioned wounds are ~40% stronger than control wounds at 10 days post surgery. Similar experiments in a diabetic mouse model also enhanced wound repair strength. These results indicate that 1) noninvasive imaging methods can aid in the optimization of novel laser preconditioning methods; 2) that optimized preconditioning with a 1.86 μm diode laser enhances early wound repair.

Local recurrence of cancer after cryosurgery is related to the inability to monitor and predict destruction of cancer (temperatures > -40°C) within an iceball. We previously reported that a cytokine adjuvant TNF-α could be used to achieve complete cancer destruction at the periphery of an iceball (0 to -40°C). This study is a further development of that work in which cryosurgery was performed using cryoprobes operating at temperatures > -40°C. LNCaP Pro 5 tumor grown in a dorsal skin fold chamber (DSFC) was frozen at -6°C after TNF-α incubation for 4 or 24 hours. Tumors grown in the hind limb were frozen with a probe tip temperature of -40°C, 4 or 24 hours after systemic injection with TNF-α. Both cryosurgery alone or TNF-α treatment alone caused only a minimal damage to the tumor tissue at the conditions used in the study. The combination of TNF-α and cryosurgery produced a significant damage to the tumor tissue in both the DSFC and the hind limb model system. This augmentation in cryoinjury was found to be time-dependent with 4-hour time period between the two treatments being more effective than 24-hour. These results suggests the possibility of cryotreatment at temperatures > -40°C with the administration of TNF-α.

This study investigates the potential of using gold nanoshells to mediate a thermally induced modulation of tumor vasculature in experimental prostate tumors. We demonstrate that after passive extravasation and retention of the circulating nanoshells from the tumor vasculature into the tumor interstitium, the enhanced nanoshells absorption of near-infrared irradiation over normal vasculature, can be used to increase tumor perfusion or shut it down at powers which result in no observable affects on tissue without nanoshells. Temperature rise was monitored in real time using magnetic resonance temperature imaging and registered with perfusion changes as extrapolated from MR dynamic contrast enhanced (DCE) imaging results before and after each treatment. Results indicate that nanoshell mediated heating can be used to improve perfusion and subsequently enhance drug delivery and radiation effects, or be used to shut down perfusion to assist in thermal ablative therapy delivery.

Minimally invasive thermal therapy is gaining ground as a new treatment modality of prostate tumors. However, further understanding of molecular events like HSP70 expression is required for treatment planning and coordination with chemotherapy and radiation. Metastatic prostate tumor (PC3 MM2) cells, transduced with reporter genes, were utilized to study the expression of HSP70 induced by normal and nanoshell-mediated heating. A correlation was noted between HSP70 expression, cellular viability and heating temperatures. This imaging paradigm can be developed into a PET-MR thermal treatment regimen, which would include dosimetry planning, real time temperature monitoring and post treatment assessment of tumor response at a cellular level.

Cryoablation is a prostate cancer treatment which uses extreme cold to kill prostate cancer cells. Unfortunately, cryoablation is not specific to cancer cells and normal cells within and surrounding the ice ball may also die which often includes those in the neurovascular bundles (NVBs). The NVBs contain neuronal and vascular components essential for erectile function and continence, and damage can cause impotence and incontinence. Therefore, agents that protect the NVBs during cryoablative procedures are extremely important for the next generation of this technology. The purpose of these studies was to analyze various agents for their ability to reduce cellular toxicity upon freezing of fibroblasts and neuronal cells. The agents were compared against DMSO, a well accepted but toxic cryoprotective agent (CPA). Various concentrations of trehalose, PVP, PEG, and DMSO were incubated with fibroblast or neuronal cells for 45 minutes at 37°C, frozen in liquid nitrogen for one hour, thawed in a 37°C water bath for 5 minutes, and then plated in 6 well plates. After 18 hours, live cells were counted using the trypan blue exclusion assay. No cells survived in DMEM alone, however, cell viability was observed in the trehalose and DMSO treated samples in both cell types and in the fibroblast cells only for PEG treated samples. To mimic the general cryoablation procedure consisting of 2 freeze/thaw cycles, neuronal cells were frozen and thawed twice. Cell viability was observed in the some trehalose and DMSO treated cells. Our experiments demonstrate that trehalose is capable of acting as a CPA of both fibroblasts and nerve cells. Its overall ability to act as a CPA is comparable to DMSO, a widely used and well accepted CPA, thus warranting further experimentation with this agent.

Purpose: To evaluate outcomes among a matched cohort of prostate cancer patients treated with radical retropubic prostatectomy (RRP) and robot assisted radical prostatectomy (RARP). Materials and methods: Between 2002 and 2005, 294 patients underwent RARP at our institution. Comparison RRP patients were matched 2:1 for surgical year, age, PSA, clinical stage, and biopsy grade (n=588). Outcomes among groups were compared. From an oncologic standpoint, pathologic features among groups were assessed and Kaplan-Meier estimates of PSA recurrence free survival were compared. Results: Overall margin positivity was not significantly different between groups (RARP, 15.6%, RRP, 17%), yet risk of apical margin was significantly less with RARP. RARP was associated with significantly shorter hospitalization (p<0.01) and lower incidence of blood transfusion (p < 0.01). Early complications were higher in the RARP group (16% vs 10%, p<0.01). Among late complications, risk of bladder neck contracture was lower with RARP (1.2%, p=0.02). Adjuvant hormonal therapy was significantly higher in the RRP group (6.6% p<0.01). Continence at 1 year among groups was equivalent (p=0.15). Potency at 1 year was better among RARP patients (p=0.02). At a median followup of 1.3 years, PSA recurrence free estimates were not significantly different (92% vs 92%, p=0.69). Conclusions: Early complications were higher in this RARP group, but this experience includes cases performed in the learning curve. Oncologic, quality of life, and functional data in this study revealed encouraging results for RARP when compared to RRP.

The treatment of partial nefrectomies is usually performed under a warm ischemic period. Recently, various treatment modalities have become available to perform a partial nefrectomy without clamping off the blood circulation. We have studied three devices in laboratory setting, investigating the thermal and high speed imaging techniques in tissue models and consequently, applying the instruments in the clinic during open procedures especially looking at efficacy and blood loss. The continuous wave 2.0 micron laser of 70 W (Revolix, LISA laser) is used as a fiber delivered knife cutting through circulated tissue with controlled hemostasis for vessels up to 3 mm diameter. The 2 μm wavelength effectively vaporizes tissue water and coagulates the smaller vessels. The Hydro-Jet (ERBE, Germany) uses high pressure (20-80 bar) to ejects a water jet of 40 um diameter at high velocity (10-30 m/s). The parenchyma is resected while vessels are preserved. Consequently, the exposed vessels can be coagulated in a controlled way with minimal blood loss. The water jet showed to induce cavitation bubbles that resect the soft tissue from the matrix leaving the elastic microvessels intact. Various systems are based on bipolar RF technology. We are using the Habib device (Rita 1500X RF generator) to create a coagulation zone around the tumor. Subsequently, the tumor can be resected along the coagulation zone with minimal bleeding. The treatment modalities investigated, have their own advantages and, stand-alone or in combination, can facilitate laparoscopic partial nephrectomies without an ischemic period.

Laser therapy for obstructive benign prostatic hyperplasia (BPH) has gained broad adoption due to effective tissue removal, immediate hemostasis, and minor complications. The aim of this study is to quantitatively compare ablation characteristics of PV (Photoselective Vaporization) and the newly introduced HPS (High Performance System) 532 nm lasers. Bovine prostatic tissues were ablated in vitro, using a custom-made scanning system. Laser-induced volume produced by two lasers was quantified as a function of applied power, fiber working distance (WD), and treatment speed. Given the same power of 80 W and speed of 4 mm/s, HPS created up to 50 % higher tissue ablation volume than PV did. PV induced a rapid decrease of ablation volume when WD increased from 0.5 mm to 3 mm while HPS yielded almost constant tissue removal up to 3 mm for both 80 W and 120 W. As the treatment speed increased, both lasers reached saturation in tissue ablation volume. Lastly, both PV and HPS lasers exhibited approximately 1 mm thick heat affected zone (HAZ) in this study although HPS created twice deeper ablation channels with a depth of up to 4 mm. Due to a smaller beam size and a higher output power, HPS maximized tissue ablation rate with minimal thermal effects to the adjacent tissue. Furthermore, more collimated beam characteristics provides more spatial flexibility and may even help to decrease the rate of fiber degradation associated with thermal damage from debris reattachment to the tip.

Direct studies comparing different lasers for treatment of BPH are lacking. This preliminary study compares continuous-wave (CW) vs. pulsed prostate tissue vaporization for the Thulium fiber laser and Holmium:YAG laser, both operating near the 1940 nm water absorption peak in tissue. A 50-W Thulium fiber laser (λ= 1908 nm) delivered CW laser radiation through a 600-μm silica fiber in non-contact mode with a 5-mm-diameter spot at the tissue surface. A Holmium:YAG laser (λ= 2120 nm) operated with an energy of 2 J, pulse rate of 25 Hz, and average power of 50 W, and delivered pulsed laser radiation through a 600-μm silica fiber with a 5-mm-diameter laser spot to achieve similar irradiances at the tissue surface. Tissue vaporization was performed in air with the prostate kept hydrated in saline. Tissue vaporization efficiency of both lasers was compared (n = 10 canine prostates for each laser group). Mean vaporization efficiency measured 5.30 ± 0.48 kJ/g vs. 4.13 ± 0.46 kJ/g for Thulium fiber and Holmium lasers (P < 0.05). Tissue vaporization rates measured 0.57 ± 0.05 g/min vs. 0.73 ± 0.07 g/min (P < 0.05). The Holmium:YAG laser vaporizes prostate tissue at a higher rate than the Thulium fiber laser, for the same average power delivered to the tissue. Both the Thulium fiber laser and Holmium:YAG lasers are capable of vaporizing prostate tissue at a rate > 1 g/min if operated at the high powers (100-W) typically used in the clinic.

Photoselective vaporization of the Prostate (PVP) using the 80W-Greenlight-PV^(R) Laser System (Laserscope^(R), San Jose, USA) has been established as a treatment option for patients suffering from obstructive symptoms caused by benign prostatic hyperplasia. However, longer operation time compared to standard trans-urethral resection of the prostate (TURP) and the high costs of the laser fibers are specific problems of this technique. In addition, many clinicians performing PVP complain about a reduced effectiveness of vaporization during treatment. Therefore, power measurement was performed during PVP using the 80W-Greenlight-PV^(R) Laser System. Power output was measured at the beginning and also regularly throughout the operation. A total of 40 fibers were investigated in 35 patients. Damage to the tip of the fibers was regularly visible and increased as more energy was supplied. Additionally, in 90% of all fibers a decrease of power output was detectable during the operation. This became pronounced after the application of 200kJ, resulting in an end of lifespan (i.e. 275kJ) median power output of only 20% of the starting value. Corresponding to the clinical observations the impressive damage to the emission window was associated with a substantial decrease of power output during PVP. These observations might explain the impaired vaporization during PVP and a longer operation time compared to conventional TURP. Hence, improvements in the quality of the laser fibers are necessary to advance the efficiency of this promising technology.

Endoscopic surgical applications of Erbium:YAG and Erbium:YSGG lasers have been limited due to the lack of a suitable mid-IR optical fiber delivery system. Germanium oxide fibers are currently used in the clinic for non-contact tissue ablation, but are not recommended for contact tissue ablation applications typically required during endoscopic surgery in a fluid environment. This study describes the assembly and characterization of hybrid mid-IR fibers consisting of germanium oxide trunk fibers and sapphire fiber tips, and side-firing germanium oxide fibers with either 45° angled fiber tips or micro-mirrors. Average powers up to 8.5 W (850 mJ at 10 Hz) and transmission up to 70% was demonstrated through 450-μm-core, 1.5-meter-long fibers, sufficient for endoscopic laser ablation of soft and hard tissues in contact mode.

Laser nerve stimulation has recently been studied as an alternative to electrical stimulation in neuroscience. Advantages include non-contact stimulation, improved spatial selectivity, and elimination of electrical stimulation artifacts. This study explores laser stimulation of the rat cavernous nerves, as a potential alternative to electrical nerve mapping during nerve-sparing radical prostatectomy. The cavernous nerves were surgically exposed in a total of 10 male rats. A Thulium fiber laser stimulated the nerves, with a wavelength of 1870 nm, pulse energy of 7.5 mJ, radiant exposure of 1 J/cm², pulse duration of 2.5 ms, pulse rate of 10 Hz, and 1-mm laser spot diameter, for a stimulation time of 60 s. A significant increase in the intracavernosal pressure was detected upon laser stimulation, with pressure returning to baseline levels after stimulation. This study demonstrates the feasibility of non-contact laser stimulation of the cavernous nerves using near-infrared laser radiation.

The Lithoclast Ultra^(R) is a combined ultrasonic and pneumatic lithotrite shown to speed the fragmentation and clearance of renal calculi. We constructed an ex vivo cystolitholapaxy model to investigate the optimal pneumatic frequency for the Lithoclast Ultra^(R). Bego and U-30 test stones were used. Ten trials of each stone type were performed at each setting. Each trial consisted of ultrasonic lithotripsy alone, and then in combination with pneumatic lithotripsy at frequencies of 2Hz, 4Hz, 8Hz, and 12 Hz. Ultrasonic lithotripsy alone was the control group. The ultrasound settings remained constant at 100% power and 90% duty. Time to 5mm fragments and time to stone clearance were recorded. Mean time to 5mm fragments for Bego stones using ultrasound alone, and with the pneumatic lithotrite at 2Hz, 4Hz, 8Hz, and 12Hz was 21min, 14min, 10min, 13min, and 14min respectively. Mean time to stone clearance for Bego stones was 29min, 25min, 23min, 22min, and 22min respectively. Mean time to 5mm fragments for U-30 stones was 101s, 89s, 68s, 64s, and 66s respectively. Mean time to stone clearance for U-30 stones was 162s, 148s, 133s, 137s, and 127s respectively. The combination of pneumatic and ultrasonic lithotrites decreases the time to stone fragmentation and clearance compared to ultrasound alone. The optimal pneumatic frequency appears to reach a plateau at 4Hz.

The Lithoclast Ultra^(R) combines ultrasonic and pneumatic lithotripsy with a single handpiece, while the Cyberwand^(R) combines two concentric ultrasonic lithotrites. We tested each device in a cystolitholapaxy model. An ex vivo bladder model was constructed. The test stones used were the Bego and U-30. For the Lithoclast Ultra, 10 trials of each stone type were performed. The ultrasound was tested at 90% duty and 100% power and the pneumatic tested at 12Hz. For the Cyberwand, the 'Large' stone and 'Small' stone setting were tested. Twenty trials with the U-30 stone were performed at each setting. Only 1 Bego stone finished Cyberwand testing. The time to clear all stone fragments was recorded. For the U-30 stones, the total time to stone clearance was 127 seconds for the Lithoclast Ultra, 41 seconds for the Cyberwand (Large setting) (p=0.001), and 69 seconds for the Cyberwand (Small setting)(p=0.12). For the Bego stones, the time to 5mm fragments and the total time to stone clearance was 14 and 22 minutes for the Lithoclast Ultra. The Cyberwand probe failed repeatedly during Bego stone testing. Four probes were required to completely eliminate a single Bego stone. For the softer U-30 stones, the Cyberwand time to stone clearance was significantly faster than the Lithoclast Ultra (p<0.05). For the harder Bego stones, the Cyberwand was not able to complete the trial due to repeated probe failure, and was outperformed by the Lithoclast Ultra.

Our long-term activity in the development of fluorescence imaging for the detection of early superficial bladder cancer aimed at optimizing the selective production and accumulation of photoactivable porphyrins (PaP), mainly protoporphyrin IX (PpIX), after the instillation of derivatives of aminolevulinic acid (ALA) within cancerous tissues. This research eventually led to the approval of hexylaminolevulinate (HAL, Hexvix^(R)) in 27 European countries. Although the selective production of PpIX and the sensitivity of this procedure are outstanding, its specificity is limited due to false positive lesions that are mainly associated with inflammations of the bladder mucosa. Therefore, our current research focuses on the improvement of the specificity of this detection method. New methods, using high magnification (HM) endoscopy, are being investigated by our group in order to discriminate false from true positive findings, and hopefully resulting in a reduced number of biopsies. In this study, we are using a dedicated magnification cystoscope, allowing conventional magnification during "macroscopic" white light and fluorescence observation, as well as image acquisition with HM when the endoscope is in contact with the tissue. This is realized by an optical setup directly integrated in the cystoscope. The diameter of the field of view of the images is 500 microns in the HM mode and the resolution is about 3 microns. With this optical setup, our on going study is aimed at observing and characterizing the neo*-vascularization of the flat fluorescing sites in order to distinguish (pre-)cancerous tissue from inflammation. Thirty nine biopsies were taken on fluorescence-positive sites. The vascular patterns observed on CIS (n = 7) were significantly altered in 5 of them (71%), as compared to normal and inflamed mucosa where such alterations were never observed.

Hypericin (HYP) is used after instillation as a diagnostic tool for the fluorescence detection of CIS in the human bladder. In this study the in vitro cellular accumulation and intraspheroidal distribution of HYP and three analogues (OH1, OH2, OH3) with gradually increasing hydrophilicity were studied. E-cadherin negative (T24-C1^-) and E-cadherin positive (T24-H3⁺⁺) human bladder cancer cells were used. We report that in the presence of FBS all compounds were taken up by the monolayer cells to the same limited extent, whereas the overall intracellular accumulation was substantially higher when the incubation of the different dyes took place using cell medium not supplemented with FBS. The results of this study therefore confirm the competition between cellular uptake of HYP and analogues and binding to FBS constituents. Investigating the permeation of the compounds in spheroids, it was found that all HYP analogues diffused dramatically better through the three-dimensional cell layers than HYP itself. This enhanced ability of hydrophilic HYP analogues to permeate through the cell layers in the presence of FBS can be explained in terms of a preferred binding to HDL as compared to LDL. The results further show that all compounds, including LDL-binding HYP, substantially permeated better in T24-C1^- spheroids than in T24-H3⁺⁺ spheroids. The data therefore support the hypothesis that a lowered cellular cohesion is the key to understand the selective uptake of hypericin and its analogues in malignant urothelial cells.

A novel micro computed tomography (μCT) image processing method was implemented to measure anatomical features of the gerbil and chinchilla cochleas, taking into account the bent modailosis axis. Measurements were made of the scala vestibule (SV) area, the scala tympani (SV) area, and the basilar membrane (BM) width using prepared cadaveric temporal bones. 3-D cochlear structures were obtained from the scanned images using a process described in this study. It was necessary to consider the sharp curvature of mododailosis axis near the basal region. The SV and ST areas were calculated from the μCT reconstructions and compared with existing data obtained by Magnetic Resonance Microscopy (MRM), showing both qualitative and quantitative agreement. In addition to this, the width of the BM, which is the distance between the primary and secondary osseous spiral laminae, is calculated for the two animals and compared with previous data from the MRM method. For the gerbil cochlea, which does not have much cartilage in the osseous spiral lamina, the μCT-based BM width measurements show good agreement with previous data. The chinchilla BM, which contains more cartilage in the osseous spiral lamina than the gerbil, shows a large difference in the BM widths between the μCT and MRM methods. The SV area, ST area, and BM width measurements from this study can be used in building an anatomically based mathematical cochlear model.

Understanding the biomechanics of the middle ear is important for surgical reconstructions. As the output of the middle ear, the stapes plays a key role in transferring acoustic vibrations to the cochlea. In order to develop anatomically-based mathematical models, which are needed to improve our understanding of stapes dynamics, detailed morphometry of the stapes is required. High-resolution micro-CT imaging techniques were used to generate three-dimensional reconstructions of cadaveric temporal bones from 5 species commonly used in experimental middle ear research: the chinchilla, human (relatively mid-frequency hearing limit), cat, guinea pig, and gerbil (relatively high-frequency hearing limit). From the standard discretizations of micro-CT images and corresponding 3-D volume reconstructions, the centers of mass, principle axes, stapes head areas and stapes footplate areas were calculated. Mechanical relationships were estimated between the capitulum area and the footplate area and inter-species comparisons were performed between the cross-sectional shapes of the anterior and posterior crura. Quantitative dynamic properties were estimated from the rigid body motion calculations. The parameters estimated in this study will be useful for building biocomputational models of the stapes for a variety of species.

Greater understanding of tympanic membrane (TM) biomechanics has the potential to guide future advances in medical technology related to its surgical repair (myringoplasty). The pars tensa of the TM is a composite structure with two collagen fiber layers that provide the main scaffolding for the TM. The external layer is arranged in an approximately radial configuration, and the other is arranged in an approximately circumferential configuration. A more detailed knowledge of collagen fiber orientation and volume fraction could greatly improve existing mechanical simulations of the TM. To address this, we employed multiphoton microscopy (MPM) imaging of the TM in two modalities: second harmonic generation (SHG) and two-photon fluorescence (TPF). The unique spectral signature of SHG allows selective imaging of collagen fibers. TPF also produces images of fibrillar-type collagen but lacks the specificity of SHG. Both the SHG and TPF images show patterns of collagen organization in the TM that match expected results with respect to both orientation and size. Through MPM, we intend to accurately determine the collagen fiber layer thickness, density, and orientation as a function of radial position and quadrant location.

A laser has been developed with the aim of being a microsurgical tool for ear surgery. Its emission at 2.7 - 2.8 μm is readily absorbed by water in tissue. This makes it ideal for ablation of the stapes, while minimizing transmission to the inner ear. Slices of porcine otic capsule bone to represent the stapes were ablated with the laser. The mechanical stress imparted to the stapes during ablation was measured using a piezoelectric film. The Er:YAG laser has similar optical properties, but this laser offers the possibility of a more compact surgical tool.

To assess the potential use of optical coherence tomography (OCT) in cochlear implant surgery, OCT was applied in human temporal bones before cochleostomy. The question was whether OCT might provide information about the cochlear topography, especially about the site of the scala tympani. OCT was carried out on human temporal bone preparations, in which the cochleostomy was performed leaving the membranous labyrinth and the fluid-filled inner ear intact. A specially equipped operating microscope with integrated OCT prototype was used. Spectral-domain (SD)-OCT was used for all investigations. On all scans, OCT supplied information about inner ear structures, such as scala tympani, scala vestibuli while the membranous labyrinth was still intact. In the fresh temporal bone the scala media, basilar membrane and the Reissner's membrane were identified. This OCT study clearly documents the possibility to identify inner ear structures, especially the scala tympani without opening its enveloping membranes. These findings may have an impact on cochlear implant surgery, especially as an orientation guide to localize the scala tympani precisely before opening the fluid filled inner ear.

Innovative laser-based approaches to laryngeal cancer include the clinical applications of two new technologies, photoangiolysis using a 532nm wavelength pulsed-KTP laser and fiber-based cutting using a 2μm wavelength thulium laser. Photoangiolysis is well-suited for treatment of minimally invasive glottic cancer and allows maximum preservation of phonatory surfaces needed for optimal voicing. The thulium laser offers an alternative to the carbon dioxide laser as an endolaryngeal cutting tool due to its enhanced hemostatic properties and fiber-based delivery. Clinical examples of pulsed-KTP laser involution of early glottic cancer will be presented in order to highlight the concept of targeting tumor angiogenesis in treating laryngeal cancer. The surgical experience using the thulium laser for complex endoscopic endolaryngeal excisions of large laryngeal cancers is presented to demonstrate the expanded clinical applications of endolaryngeal cutting offered by this laser. The laryngeal tissue effects of various laser power and pulse width (PW) settings, mode of delivery, active cooling to reduce thermal trauma, and wavelength selection have been extensively studied for the KTP and thulium lasers in both ex-vivo and live-perfusing models. The results from these studies, included herein, determine the clinical efficacy and safety of these innovative laser-based approaches to laryngeal cancer.

Anatomical optical coherence tomography (aOCT) is an endoscopic optical technique that enables continuous, quantitative assessment of hollow organ size and shape in three dimensions. It is a powerful alternative to X-ray computed tomography, magnetic resonance imaging, and video endoscopy for the assessment of gross hollow-organ anatomy. This paper reviews our instrument and its application to the upper and lower airway, and includes a number of new results.

Optical coherence tomography (OCT) is a new promising imaging modality in laryngology as yet of undetermined value. The objective of the present study was to evaluate the ability of this noninvasive method to make a reliable prediction of diagnosis in laryngeal disease. In a prospective study, 225 benign and malignant laryngeal lesions were examined with a fiber-based OCT system in contact mode during elective microlaryngoscopy. Intraoperative OCT findings were compared to conventional histopathology supplying a correct specific diagnosis in about 90% of cases. With increasing experience, a reliable prediction of invasive tumor growth and often the exact grade of dysplasia were possible. Due to the current spatial resolution of OCT, microinvasive cancer could not be safely delineated from severe dysplasia or carcinoma in situ. In contrast, benign lesions made no essential diagnostic difficulties due to their characteristic OCT image. In conclusion, OCT has proved to be very useful in the diagnostic investigation and the intraoperative monitoring of laryngeal disease.

In this report, the diagnostic ability of near-infrared (NIR) Raman spectroscopy for identifying the malignant tumors from normal tissues in the larynx was studied. A rapid NIR Raman system was utilized. Multivariate statistical techniques were employed to develop effective diagnostic algorithms. Raman spectra in the range of 800-1,800 cm-1 differed significantly between normal and malignant tumor tissues. The diagnostic algorithms can yielded a diagnostic sensitivity of 92.9% and specificity 83.3% for separating malignant tumors from normal laryngeal tissues. NIR Raman spectroscopy with multivariate statistical techniques has a potential for the non-invasive detection of malignant tumors in the larynx.

One sequela of skull base surgery is the iatrogenic damage to cranial nerves. Devices that stimulate nerves with electric current can assist in the nerve identification. Contemporary devices have two main limitations: (1) the physical contact of the stimulating electrode and (2) the spread of the current through the tissue. In contrast to electrical stimulation, pulsed infrared optical radiation can be used to safely and selectively stimulate neural tissue. Stimulation and screening of the nerve is possible without making physical contact. The gerbil facial nerve was irradiated with 250-μs-long pulses of 2.12 μm radiation delivered via a 600-μm-diameter optical fiber at a repetition rate of 2 Hz. Muscle action potentials were recorded with intradermal electrodes. Nerve samples were examined for possible tissue damage. Eight facial nerves were stimulated with radiant exposures between 0.71-1.77 J/cm², resulting in compound muscle action potentials (CmAPs) that were simultaneously measured at the m. orbicularis oculi, m. levator nasolabialis, and m. orbicularis oris. Resulting CmAP amplitudes were 0.3-0.4 mV, 0.15-1.4 mV and 0.3-2.3 mV, respectively, depending on the radial location of the optical fiber and the radiant exposure. Individual nerve branches were also stimulated, resulting in CmAP amplitudes between 0.2 and 1.6 mV. Histology revealed tissue damage at radiant exposures of 2.2 J/cm², but no apparent damage at radiant exposures of 2.0 J/cm².

Introduction: Being practical, efficient and inexpensive, fibre guided diode laser systems are preferable over others for endonasal applications. A new medical 1470 nm diode laser system is expected to offer good ablative and coagulative tissue effects. Methods: The new 1470 nm diode laser system was compared to a conventional 940 nm system with regards to laser tissue effects (ablation, coagulation, carbonization zones) in an ex vivo setup using fresh liver and muscle tissue. The laser fibres were fixed to a computer controlled stepper motor, and the light was applied using comparable power settings and a reproducible procedure under constant conditions. Clinical efficacy and postoperative morbidity was evaluated in two groups of 10 patients undergoing laser coagulation therapy of hyperplastic nasal turbinates. Results: In the experimental setup, the 1470 nm laser diode system proved to be more efficient in inducing tissue effects with an energy factor of 2-3 for highly perfused hepatic tissue to 30 for muscular tissue. In the clinical case series, the higher efficacy of the 1470 nm diode laser system led to reduced energy settings as compared to the conventional system with comparable clinical results. Postoperative crusting was less pronounced in the 1470 nm laser group. Conclusion: The 1470 nm diode laser system offers a highly efficient alternative to conventional diode laser systems for the coagulation of hyperplastic nasal turbinates. According to the experimental results it can be furthermore expected that it disposes of an excellent surgical potential with regards to its cutting abilities.

In this research, we have used an algorithm to extract four different optical parameters, including the scattering coefficient (μ_s), effective anisotropy factor (g_off), birefringence coefficient (Δn), and fast-axis angle (β) from polarization-sensitive optical coherence tomography (PS-OCT) images. The proposed method was evaluated from a set of phantoms and it yielded valid results. We demonstrate that the optical characteristic of normal and different kinds of atherosclerotic lesions in human vessel can be quantitatively described. Whether change in these quantified optical properties could be used to discriminate between different types of human atherosclerotic plaques was examined.

InfraReDx has developed a spectroscopic cardiac catheter system capable of acquiring near-infrared (NIR) reflectance spectra from coronary arteries in vivo for identification of lipid-rich plaques of interest (LRP). The spectral data are analyzed with a chemometric model, producing a hyperspectral image (a chemogram) used to identify LRP in the interrogated region. In this paper, we describe a FT-IR microscopy system for measurement of the NIR scattering and absorption properties of healthy and diseased regions of human coronary arteries in small volumes (~10 μl). Scattering and absorption coefficients are obtained from sequential 140 um x 140 um regions of interest across the face of 500-micron thick, saline-irrigated fresh coronary artery sections. A customized FTIR microscope, measurement protocol, and inversion algorithm are used for optical property determination, and the system is calibrated using measurements of tissue-simulating phantoms having well-characterized optical properties. Tissue optical properties are co-registered with brightfield transmission images as well as with stained histologic thin sections (H&E, Movat Pentachrome, and Oil Red O) acquired from an immediately-adjacent section. The ultimate goal of these experiments is to establish a mechanistic link between the multivariate model predictions displayed on the InfraReDx chemogram and the light-tissue interactions that govern the measured NIR reflectance spectra.

To improve the efficacy of tumor resection in image guided neurosurgery, two new types of feedback were investigated. Firstly, sound feedback to give the surgeons a warning signal when the instrument tip approaches normal/non-planned brain tissue. Secondly, workflow feedback by logging and tagging all instrument positions in image space. Results from laboratory and clinic suggest that sound feedback seems to be useful but needs more fine tuning before it will be practical for clinical use. Workflow feedback, on the other hand, appears to be a simple, cheap and efficient method to give the surgeon insight in the progress of resection, to log locations of interest, and to give insight in brain deformations occurring during surgery.

Optical spectroscopy for in vivo tissue diagnosis is performed traditionally in a static manner; a snap shot of the tissue biochemical and morphological characteristics is captured through the interaction between light and the tissue. This approach does not capture the dynamic nature of a living organ, which is critical to the studies of brain disorders such as epilepsy. Therefore, a time-dependent diffuse reflectance spectroscopy system with a fiber-optic probe was designed and developed. The system was designed to acquire broadband diffuse reflectance spectra (240 ~ 932 nm) at an acquisition rate of 33 Hz. The broadband spectral acquisition feature allows simultaneous monitoring of various physiological characteristics of tissues. The utility of such a system in guiding pediatric epilepsy surgery was tested in a pilot clinical study including 13 epilepsy patients and seven brain tumor patients. The control patients were children undergoing suregery for brain tumors in which measurements were taken from normal brain exposed during the surgery. Diffuse reflectance spectra were acquired for 12 seconds from various parts of the brain of the patients during surgery. Recorded spectra were processed and analyzed in both spectral and time domains to gain insights into the dynamic changes in, for example, hemodynamics of the investigated brain tissue. One finding from this pilot study is that unsynchronized alterations in local blood oxygenation and local blood volume were observed in epileptogenic cortex. These study results suggest the advantage of using a time-dependent diffuse reflectance spectroscopy system to study epileptogenic brain in vivo.

Following surgical removal of malignant brain tumors 80% of all cases develop tumor recurrence within 2 cm of the resected margin. The aim of postoperative therapy is therefore elimination of nests of tumor cells remaining in the margins of the resection cavity. Light attenuation in tissue makes it difficult for adequate light fluences to reach depths of 1-2 cm in the resection margin making it difficult for standard intraoperative photodynamic therapy (PDT) to accomplish this goal. Thus additional agents are required that either increase the efficacy of low fluence PDT or inhibit cellular repair, to enhance effectiveness of PDT in the tumor resection cavity. Motexafin gadolinium (MGd) is one such agent previously reported to enhance the cytotoxic potential of radiation therapy, as well as several chemotherapeutic agents by causing redox stress to cancerous cells. MGd is well tolerated with tumor specific uptake in clinical studies. The authors evaluated MGd as a potential PDT enhancing agent at low light fluences using an in vitro model. Multicellular Glioma spheroids (MGS) of approximately 300 micron diameter, obtained from ACBT cell lines were subjected to acute PDT treatments at 6J, 12J, and 18J light fluences. Growth was determined by measuring diameters in two axes. At four weeks a dose dependent inhibition of spheroid growth was seen in 33%, 55%, and 83% of the MGS at 6J, 12J, and 18J respectively, while inhibition followed by a partial reversal of growth was seen in 17%, 33%, and 17% respectively. This study provides a rationale for the use of this drug as a PDT enhancer in the management of brain tumors.

Introduction: Failure of treatment for high grade gliomas is usually due to local recurrence at the site of surgical resection indicating that a more aggressive form of local therapy, such as PDT, could be of benefit. PDT causes damage to tumor cells as well as degradation of the blood brain barrier (BBB). We have evaluated the ability of ALA mediated PDT to open the BBB in rats. This will permit access of chemotherapeutic agents to brain tumor cells remaining in the resection cavity wall, but limit their penetration into normal brain remote from the site of illumination. Materials and Methods: ALA-PDT was performed on non tumor bearing inbred Fisher rats at increasing fluence levels. T2 weighted MRI scans were used to evaluate edema formation and post-contrast T₁ MRI scans were used to monitor the degree BBB disruption which could be inferred from the intensity and volume of the contrast agent visualized. Results. PDT at increasing fluence levels between 9J and 26J demonstrated an increasing contrast flow rate. No effect on the BBB was observed if 26J of light were given in the absence of ALA. A similar increased contrast volume was observed with increasing fluence rates. The BBB was found to be disrupted 2hrs. following PDT and 80-100% restored 72hrs later. Conclusion: PDT was highly effective in opening the BBB in a limited region of the brain. The degradation of the BBB was temporary in nature, opening rapidly following treatment and significantly restored during the next 72 hrs.

Post-operative verification of the specificity and sensitivity of photodynamic therapy (PDT) is most pressing for deeply placed lesions such as brain tumors. We wish to determine whether Dynamic Contrast Enhanced-Magnetic Resonance Imaging (DCE-MRI) can provide a non-invasive and unambiguous quantitative measure of the specificity and sensitivity of brain tumor PDT. Methods: 2.5 x 10⁵ U87 cells were injected into the brains of six athymic nude rats. After 5-6 days, the animals received 0.5 mg/kg b.w. of the phthalocyanine photosensitizer Pc 4 via tail-vein injection. On day 7 peri-tumor DCE-MRI images were acquired on a 7T microMRI scanner before and after tail-vein administration of 100 μL gadolinium and 400 μL saline. After this scan the animals received a 30 J/cm² dose of 672-nm light from a diode laser (i.e., PDT). The DCE-MRI scan protocol was repeated on day 13. Next, the animals were euthanized and their brains were explanted for Hematoxylin and Eosin (H&E) histology. Results: No tumor was found in one animal. The DCE-MRI images of the other five animals demonstrated significant tumor enhancement increase (p < 0.053 two-sided t-test and p < 0.026 one-sided t-test) following PDT. H&E histology presented moderate to severe tumor necrosis. Discussion: The change in signal detected by DCE-MRI appears to be due to PDT-induced tumor necrosis. This DCE-MRI signal appears to provide a quantitative, non-invasive measure of the outcome of PDT in this animal model and may be useful for determining the safety and effectiveness of PDT in deeply placed tumors (e.g., glioma).

The aim of this experimental study was to determine whether Motexafin Gadolinium (MGd) could serve as an efficient intraoperative contrast agent avoiding problems that arise with surgically-induced intracranial enhancement. F98 orthotopic brain tumors or surgical lesions were induced in Fisher rats. T1-weighted MRI studies were performed with either a single or multiple daily doses of MGd. The last contrast dose was administered either 7 or 24 h prior to scanning in both tumor-bearing and surgically treated animals. Animals receiving either 30 or 60 mg/kg MGd i.v. developed clinical signs of impaired motor activity, and increasing lethargy. MGd given i.p. was tolerated up to a dose of 140 mg/kg. Despite multiple dosages, and several administration modes (i.p. and i.v.), no significant enhancement was observed if the scans were performed 7 or 24 h following the last MGd dose. Clear enhancement was observed if the scans were performed 30 min. following MGd administration. Scans of necrotic lesions were positive 7 h post MGd injection. MGd scans showed no significant enhancement following surgically-induced lesions while scans with conventional contrast agents showed both meningeal and intraparenchymal enhancement. This study suggests that MGd is not sequestered in viable tumor for the necessary time interval required to allow delayed imaging in this model. The agent does seem to remain in necrotic tissue for longer time intervals. MGd therefore would not be suitable as a contrast agent in iMRI for the detection of residual tumor tissue during surgery.

Recent years have seen rising interest in optical system-on-a-chip sensors for biological applications. Vertical cavity surface emitting lasers (VCSELs) are a natural choice for array-based sensors requiring high power and low noise. However, much of the noise characterization of VCSELs has been performed in frequency ranges on the order of 10⁸ to 10¹⁰ Hz, whereas many physiological phenomena occur in frequency bands in the hertz to kilohertz range where 1/f and 1/f² noise is dominant. In this work we characterize the relative intensity noise (RIN) of commercial VCSEL devices and evaluate their feasibility for use in an integrated semiconductor optical sensor for functional brain imaging using Intrinsic Optical Signals (IOS). Our results show RIN on the order of -196 to -174 dB/Hz at an offset of 10 Hz. This is well below the signal-to-background and dynamic range requirements of 6 dB and 86 dB, respectively, for this application.

We used spatially modulated near-infrared (NIR) light to detect and map chromophore changes during cerebral edema in the rat neocortex. Cerebral edema was induced by intraperitoneal injections of free water (35% of body weight). Intracranial pressure (ICP) was measured with an optical fiber based Fabry-Perot interferometer sensor inserted into the parenchyma of the right frontal lobe during water administration. Increase in ICP from a baseline value of 10 cm-water to 145 cm-water was observed. Following induction of cerebral edema, there was a 26±1.7% increase in tissue concentration of deoxyhemoglobin and a 47±4.7%, 17±3% and 37±3.7% decrease in oxyhemoglobin, total hemoglobin concentration and cerebral tissue oxygen saturation levels, respectively. To the best of our knowledge, this is the first report describing the use of NIR spatial modulation of light for detecting and mapping changes in tissue concentrations of physiologic chromophores over time in response to cerebral edema.

The aim of this study was to evaluate antibacterial effects of PDT on common bacteria causing otitis media with effusion (OME). In vitro study was carried out using a hematoporphyrin derivative sensitizer (photogem) and 632 nm diode laser on H. influenzae, M. catarrhalis, and S. pneumoniae. One ml of each bacterial suspension was incubated for 3 hours and various concentrations of photogem were administered into the suspension. The suspensions were irradiated with 632 diode laser (15 J/cm²). The presence of colony forming units of the bacteria was examined, microscopic structures of bacteria were examined by TEM, and cytometry of bacteria was performed. The PDT was effective in killing all 3 kinds of bacteria. TEM showed damaged bacterial cell membrane and cytoplasmic structures and the flow cytometry showed lower number of viable bacteria in PDT group comparing to the control group. In vivo PDT study was performed using gerbil. S. pneumoniae or H. influenzae was injected into bullae. Photogem was injected into bullae in 2 days by when OME was developed and transcanal irradiation of 632 nm diode laser (90 J) was performed with a fiber perforated through an ear drum into a middle ear cavity and bulla. Four days after PDT, middle ear and bulla were washed with DPBS and the washed DPBS was cultured. The presence of bacterial colonies was examined. PDT was effective in killing S. pneumoniae in 87 % of the infected bullae with OME while it was effective to eradicate H. influenzae in 50 % of the infected bullae with OME. The results of these studies demonstrated that PDT may be effective to treat otitis media. It may have clinical implication to treat otitis media that is resistant to antibiotic therapy.