We recorded multiphoton images of human skin biopsies using endogenous sources of nonlinear optical signals. We detected simultaneously two-photon excited fluorescence (2PEF) from intrinsic fluorophores and second harmonic generation (SHG) from collagen. We observed SHG from fibrillar collagens in the dermis, whereas no SHG was detectable from the non fibrillar type IV collagen in the basal laminae. We compared these distinct behaviours of collagens I and IV in SHG microscopy to polarization-resolved surface SHG experiments on thin films of collagens I and IV molecules. We observed similar signals for both types of molecular films, except for the chiroptical contributions which are present only for collagen I and enhance the signal typically by a factor of 2. We concluded that SHG microscopy is a sensitive probe of the micrometer-scale structural organization of collagen in biological tissues. In order to elucidate the origin of the endogenous fluorescence signals, we recorded 2PEF spectra at various positions in the skin biopsies, and compared these data to in vitro spectroscopic analysis. In particular, we studied the keratin fluorescence and determined its 2PEF action cross section. We observed a good agreement between 2PEF spectra recorded in the keratinized upper layers of the epidermis and in a solution of purified keratin. Finally, to illustrate the capabilities of this technique, we recorded 2PEF/SHG images of skin biopsies obtained from patients of various ages.

It is a field of great interest to develop therapies to rejuvenate photoaged skin. However, the treatment response can not be ideally determined due to lack of a reliable non-invasive method to quantify photoaging. In this study, the photoaging process of skin is investigated by use of a multiphoton fluorescence and second harmonic generation microscopy. We obtain the autofluorescence and second harmonic generation images of superficial dermis from facial skin of individuals of different ages. The results show that autofluorescence signals increase with age while second harmonic generation signals decrease with age. The results are consistent with the histological findings in which collagen is progressively replaced by elastic fibers. In the case of severe photoaging, solar elastosis can be clearly demonstrated by the presence of thick curvy autofluorescent materials in the superficial dermis. We propose a second harmonic generation to autofluorescence aging index of dermis to quantify the photoaging changes. This index is shown to be a good indicator of photoaging. Our results suggest that multiphoton fluorescence and second harmonic generation microscopy can be developed into a non-invasive imaging modelity for the clinical evaluation of photoaging.

Multiphoton autofluorescence and second harmonic generation (SHG) microscopy are useful in respectively identifying elastin and collagen fibers within the skin dermis. In this study, we attempt to characterize the degree of skin thermal damage by using multiphoton microscopy to characterize the thermal changes to collagen and elastin fibers. We found that autofluorescence and SHG imaging behave differently in skin dermis treated with different temperatures and that an index reflecting the relative changes in autofluorescence and SHG intensity is useful to identify the degree of dermal thermal damage to the skin. With additional development, our approach can be used to identify the extent of thermal damage in patients.

We report results of clinical trials on flap monitoring in 65 plastic surgeries. Hemoglobin oxygen saturation of flap tissue (StO₂) was monitored non-invasively by using ODISsey^TM tissue oximeter, an infrared spectroscopic device. StO₂ measurements were conducted both intra-operatively and post-operatively. From the intra-operative measurements, we observed that StO₂ values dropped when the main blood vessels supplying the flap were clamped in surgery, and that StO₂ jumped after anastomosis to a value close to its pre-operative value. From post-operative monitoring measurements for the 65 flap cases, each lasted two days or so, we found that the StO₂ values approach to a level close to the baseline if the surgery was successful, and that the StO₂ value dropped to a value below 30% if there is a perfusion compromise, such as vascular thrombosis.

Both reflectance spectroscopy and the determination Young's Modulus of skin have shown promise for identifying skin pathology. At present, these determinations are carried out using separate methodologies. This study demonstrates a new technology combining digital UV/VIS reflectance spectroscopy and vacuum aspiration for simultaneously determining the reflectance spectrum and mechanical properties of human skin tissue. A small hand held prototype device incorporating fiber-optic light guides into a vacuum channel was calibrated using various elastic materials subjected to increments of stress by vacuum from 0 to 25 in Hg. The intensity of a UV/VIS light beam reflected from the material at each vacuum increment was compared to the resulting material strain. The reflected beam was also spectrophotometrically analyzed. Skin types were similarly evaluated comparing normal and scar tissue and skin of various ages and coloration. An exponential relationship between reflected beam intensity and the amount of strain resulting from vacuum increments was observed. Young's Modulus (calculated from Aoki et. al equation) and spectra from normal skin and scar tissue were in agreement with previously published observations. Age related decreases in skin elasticity were also demonstrated. In the reflectance spectra, oxy and deoxy-hemoglobin absorbance bands were detected, becoming significantly enhanced at increased levels of vacuum. Melanin absorbance was also easily detected and appeared to correlate with skin coloration. Since superficial skin pathologies have characteristic spectroscopic and mechanical properties, this technique may provide a promising new approach for rapid, non-invasive method for the evaluation of skin lesions.

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.

Although post-inflammatory pigmentation (PIH) is a common acquired skin disorder, however, its mechanism still remains unclear. In this paper, multimode optical spectroscopic techniques including diffuse reflectance spectroscopy, fluorescence spectroscopy and Raman spectroscopy have been used to probe the morphologic and biochemical changes associated with PIH. Both pigmented skin and the adjacent normal skin were measured for paired analysis. Distinctive spectral features were observed which may provide the potentials for identifying the mechanism of PIH. The diffuse reflectance spectrum was also converted into the CIE standard L*a*b* color system. The melanin and the hemoglobin content in the PIH were quantitatively derived from the diffusive reflectance spectrum using the Stamatas-Kollias algorithm. It was found that PIH always has a lower value in L* and more melanin and hemoglobin content which is consistent with clinical appearance of dark skin as perceived by the human eye.

This paper provides an overview of the recent developments of photonics in dermatology and aesthetic applications. The range of products covers lasers, continuous Xenon lamps, Intense Pulsed Light systems, and LEDs. We will mention several applications and how different photonics systems are used. We will also discuss methods combining photonics with other technologies. For example, in Photo Dynamic Therapy (PDT) this includes a drugs, or equally the combination of intense light pulses with Radio Frequency (RF) for applications like hair removal. We will also describe some new developments in photonics technology that affect the development of new products, showing the direction of market development. Additionally, some examples of new technology are shown.

American and European statistics have shown that 1-2 per cent of the human population is affected by the skin disease psoriasis. Recent research reports promising treatment results when irradiating skin areas affected by psoriasis with high powered excimer lasers with a wavelength of 308 nm. In order to apply the necessary high energy dose without hurting healthy parts of the skin new approaches regarding the system technology must be considered. The aim of the current research project is the development of a sensor-based, automated laser treatment system for psoriasis. In this paper we present the algorithms used to cope with the diffculties of irradiating irregularly shaped areas on curved surfaces with a predefined energy level using a pulsed laser. Patients prefer the treatment to take as little time as possible. This also helps to reduce costs. Thus the distribution of laser pulses on the surface to achieve the given energy level on every point of the surface has to be calculated within a limited time frame. The remainder of the paper will describe in detail an efficient method to plan and optimize the laser pulse distribution. Towards the end, some first results will be presented.

Blood velocity information can be extracted by analyzing, either temporally or spatially, laser speckle (LS) patterns generated when a laser source illuminates the tissue. While a temporal analysis, such as that used for laser Doppler velocimetry (LDV), provides high spatial resolution, the time required to obtain flow data in vivo on large areas of tissue limits its utility. The LS imaging (LSI) technique combines the nonscanning, full-field, LS method and the modified multiple scattering algorithms developed for LDV analysis to retrieve blood velocity parameters. It provides a noninvasive means for realtime, quantitative measurements of subtle changes in the tissue vasculature. This paper describes the use of the LSI technique on free flap measurements of a swine model and compares the results with those obtained using an LDV probe. Both the LSI and the LDV measurements showed similar results - blood velocity and flow decreased about 10%-33% from the tip to the caudal base of the flap, respectively. The difference between the tip and the caudal base is a measure of flap ischemia. However, tissue pigmentation affects the blood flow parameters retrieved from the LDV measurements, it does not affect the blood velocity parameters retrieved from the LSI measurements. Both techniques were also used during free tissue transfer procedures in patients to demonstrate the utility of the LSI for monitoring the status of the graft.

After segmentation of the epidermis from three-dimensional coherence tomography volume, a depth-oriented algorithm provides a segmentation of the infundibulum. In this process, the epidermal thickness, the population and the occupation ratio of the infundibula are provided.

In this paper we present preliminary results obtained from fluorescence lifetime measurements on human skin using time-correlated single photon counting (TCSPC) techniques. Human skin was exposed to light from a pulsed LED of 700 ps pulse width at a wavelength of 375 nm and fluorescence decays were recorded at four different emission wavelengths (442, 460, 478 and 496 nm) using a photomultiplier tube (PMT) coupled to a monochromator. Measurements were carried out on the left and right palms of subjects recruited for the study after obtaining consent using a UCLA IRB approved consent form. The subjects recruited consisted of 18 males and 17 females with different skin complexions and ages ranging from 10 to 70 years. In addition, a set of experiments were also performed on various locations including the palm, the arm and the cheek of a Caucasian subject. The fluorescence decays thus obtained were fit to a three-exponential decay model in all cases and were approximately 0.4, 2.7 and 9.4 ns, respectively. The variations in these lifetimes with location, gender, skin complexion and age are studied. It is speculated that the shorter lifetimes correspond to free and bound NADH while the longer lifetime is due to AGE crosslinks.

In laser therapy of port wine stain (PWS) birthmarks, measurement of maximum temperature rise is important to determine the maximum permissible light dose for PWS laser therapy. To measure maximum temperature rise on arbitrary PWS skin site, we developed a handheld pulsed photothermal radiometry (PPTR) system, which overcomes in vivo measurement limitations of bench-top PPTR systems. The developed PPTR system consists of an IR lens, an AC-coupled thermoelectrically cooled IR detector, a laser handpiece holder, and a fixed distance measurement port. With system characterization, experimental results were in good agreement with theoretical calculations. Preliminary results for maximum temperature rise demonstrate the feasibility of the PPTR system for PWS skin characterization in the clinic.

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.

Skin inflammation is often accompanied by edema and erythema. While erythema is the result of capillary dilation and subsequent local increase of oxygenated hemoglobin (oxy-Hb) concentration, edema is characterized by an increase in extracellular fluid in the dermis leading to local tissue swelling. Edema and erythema are typically graded visually. In this work we tested the potential of spectral imaging as a non-invasive method for quantitative documentation of both the erythema and the edema reactions. As examples of dermatological conditions that exhibit skin inflammation we imaged patients suffering from acne, herpes zoster, and poison ivy rashes using a hyperspectral-imaging camera. Spectral images were acquired in the visible and near infrared part of the spectrum, where oxy-Hb and water demonstrate absorption bands. The values of apparent concentrations of oxy-Hb and water were calculated based on an algorithm that takes into account spectral contributions of deoxy-hemoglobin, melanin, and scattering. In each case examined concentration maps of oxy-Hb and water can be constructed that represent quantitative visualizations of the intensity and extent of erythema and edema correspondingly. In summary, we demonstrate that spectral imaging can be used in dermatology to quantitatively document parameters relating to skin inflammation. Applications may include monitoring of disease progression as well as efficacy of treatments.

Port wine stained patients need multiple laser treatments which each are paused for 3 months per treatment. For clinical evaluation and better understanding of the laser settings used during these treatments, a special developed multispectral dermatoscope is used to quantify the micro vascularization within the port wine stains. The multispectral dermatoscope captures high resolution images of skin tissue with a surface of approximately one square centimeter. For more accurate treatment evaluation of the port wine stain, recapturing of the exact same location is desired. For multispectral image-analyses an elastic match can be performed so that per pixel spectral analyses can be performed.

We present a novel spectro-polarimetric instrument based on hemispherical backscattering for the assessment of superficial skin lesions. The system is capable of capturing polarized light images non-invasively. The effect of the rough skin backscattering is eliminated with the use of out-of-plane illumination. A glass slide with an index matching fluid, commonly used in polarized light imaging, is no longer necessary. The system is composed of sixteen polarized light sources that provide red, green, or blue illumination. The light sources are distributed on a hemispherical shell, and each source produces a collimated beam incident on the center of the hemisphere. A Stokes vector imaging system is mounted on the shell at an oblique angle to the sample normal and consists of a 12-bit scientific camera, two liquid crystal variable retarders, and a fixed polarizer. Stokes vector images of light scattered towards the camera direction are generated for each source. A useful decomposition of the Stokes vector is presented. Examples of images generated by the system are presented.

Melanin is essentially a nonfluoresent material under ultra-violet (UV) and short wavelength visible light excitation. However, fluorescence emission from in vivo cutaneous melanin has recently been detected spectroscopically under near-infrared (NIR) excitation by our group. The objective of this study is to develop an in vivo NIR autofluorescence imaging system for direct observation and characterization of melanin distribution in human skin. In the imaging system, light coming from a 785 nm diode laser is coupled into a ring light guide to uniformly illuminate the skin surface. The fluorescence or reflectance light is collected by an NIR-sensitive CCD camera with and without long-pass filters. Both reflectance and autofluorescence images of nevi from three volunteers were obtained with exposure time of less than 1 second. In NIR autofluorescence images the pigmented areas showed higher fluorescence than adjacent normal skin, thus confirmed the previous spectroscopic results and demonstrated great promises for using NIR autofluorescence for evaluating pigmented skin lesions.

We successfully delivered therapeutic gene, Hepatocyte Growth Factor (HGF), to rat skin in vivo. The level of hHGF protein increased by the application of LISWs when compared with that of control samples without LISW application.

High level of light scattering in mammalian skin significantly deteriorates the effectiveness and precision of light delivery to intra- and subcutaneous targets in therapeutic and diagnostic applications. Optical immersion based on the refractive index matching of tissue scattering centers with surrounding matter via introduction of an exogenous indexmatching agent is capable to alleviate the problem. But this process goes slowly enough because of sluggish diffusion of various immersion agents through a stratum corneum (SC) barrier, making a practical realization of this approach difficult. In order to increase the rate of the process we designed the method of accelerating the penetration of optical clearing agents (OCAs) by enhancing skin permeability through a creation of lattice-like pattern of localized thermal damage (ablation) islets in the SC. OCAs, such as glucose, propylene glycol, and glycerol solutions, were applied. Experimental results show a remarkable increase in the rate of optical clearing. The most of results were obtained on in vivo human skin.

Background. Laser tissue repair usually relies on haemoderivate solders, based on serum albumin. These solders have intrinsic limitations that impair their widespread use, such as limited repair strength, high solubility, brittleness and viral transmission. Furthermore, the solder activation temperature (65-70 °C) can induce significant damage to tissue. In this study, a new laser-activated biomaterial for tissue repair was developed and tested in vitro and in vivo to overcome some of the shortcomings of traditional solders. Materials and Methods. Flexible and insoluble strips of chitosan adhesive (surface area ~34 mm², thickness ~20 μm) were developed and bonded on sheep intestine with a laser fluence and irradiance of 52 ± 2 J/cm² and ~15 W/cm² respectively. The temperature between tissue and adhesive was measured using small thermocouples. The strength of repaired tissue was tested by a calibrated tensiometer. The adhesive was also bonded in vivo to the sciatic nerve of rats to assess the thermal damage induced by the laser (fluence = 65 ± 11 J/cm², irradiance = 15 W/cm²) four days post-operatively. Results. Chitosan adhesives successfully repaired intestine tissue, achieving a repair strength of 0.50 ± 0.15 N (shear stress = 14.7 ± 4.7 KPa, n=30) at a temperature of 60-65 °C. The laser caused demyelination of axons at the operated site; nevertheless, the myelinated axons retained their normal morphology proximally and distally.

In this study, a NIR erbium fiber laser tuned to a water vibrational overtone absorption band at 1455 nm was used to weld directly, in vitro, seventy-six porcine aorta tissues without the need for extrinsic solder materials. The tissues were divided into eleven groups based on the multiple and variable parameters that were used to weld the tissues. The effectiveness of the parameters used in each of the weld groups was evaluated directly at the time of the weld and also by tensile strength measurements done at the termination of the weld. Management of heat produced in tissues is of critical importance for good laser tissue welding (LTW). To address heat management issues, we report LTW using a transparent cover over the tissue specimen as a heat sink. Multiple scanning helps distribute the laser-generated heat and allows the tissue to cool between scans, reducing thermal damage. Better heat management using a transparent cover slide enhances the welding success. It reduces collateral damage and limits water evaporation and control the buckling of tissue around the line of apposition so that the two pieces that are welded do not move apart along the line of apposition due to buckling pressure and ensure a full-length weld.

Laser tissue soldering is a technique for the closure of incisions, which provides an immediate air- and watertight bond. Previous studies have shown that this method can bond tracheal incisions with the aid of stay sutures or solid albumin strips. In this study we investigated whether soldering using a diode laser and indocyanine green (ICG) dyed liquid albumin solution as solder, was efficacious for the repair of tracheal incisions without the need for additional strengthening aids. A transverse incision was made in isolated pig tracheas. The incision was smeared with liquid solder composed of 42% bovine albumin and 0.1 mg/ml ICG, and soldered using a temperature-controlled fiberoptic diode laser system. The soldered tracheal ends were sealed and the burst pressure measured. In a series of experiments the burst pressure was found to be higher than 382 mm Hg. These preliminary results demonstrate that diode laser tissue soldering of tracheal incisions provides better results than with a CO2 laser, and that it is possible to achieve considerable soldering strength without the use of any additional strengthening procedures.

In this study, tissue welding with 980-nm laser system, which is first-time in the literature, was performed. Hence, a preliminary study was done to determine optimal parameters for further studies. 1 cm long incisions done on the Wistar rat's dorsal skin were welded. Tissue welding with 980-nm wavelength depends on the degree of photothermal interaction. Thus, different power levels and exposure schedule were investigated. Dorsal sides of all animals were photographed from the date of surgery until they were sacrificed. The clinical examination - opening of wound and presence of infection - was noted. The rats did not show any abnormality on their health, behavior and nutrition manner. As a result, 980-nm diode laser was concluded to be a good candidate for tissue welding applications.

This study tests the hypothesis that repeat steam sterilization will result in an increase in the rate of holmium:YAG laser fiber failure during bench testing in a series of commercially available single use and reusable small core sized (200-272 μm) holmium:YAG laser fibers. Single use and reusable small core-sized holmium laser fibers were tested. Single use fibers included the Dornier Lightguide Super 200, Dornier DUR Laser Fiber Single Use, Optical Integrity Scopesafe 272, IQinc. LLF200TG-D and LLF273TG-D, Boston Scientific AccuFlex 200 and AccuFlex 273, and Lumenis Slimline EZ 200. Reusable fibers included the Dornier DUR Laser Fiber Reusable, Lumenis Slimline 200, Sharplan 200, Laser Peripherals RBLF-200, IQinc. LFT273NT, and Convergent Optiview SMH1020F. A Lumenis VersaPulse 100 watt and a PowerSuite 20 watt holmium:YAG lasers were used. Fibers were bent to 180 degrees at a diameter of 1.5 cm. The laser was fired at 1.5J, 10Hz for 30 minutes or until fiber fracture. Reusable fibers were sterilized for a total of twenty cycles using the manufacturers' specifications and retested after every five completed cycles. No fiber fractured with bending alone. Two of three Dornier Super 200 single use fibers fractured repeatedly within several laser pulses. The Laser Peripherals RBLF-200 fiber fractured during initial but not repeat testing. One of three reusable Dornier DUR Laser Fibers failed during testing after the twentieth steam sterilization cycle. Five of five Boston Scientific Accuflex 200 failed at the SMA connector and did not couple well with the Lumenis Powersuite laser. The test hypothesis was not supported.

Stone retropulsion during Ho:YAG (λ = 2.12 μm) laser lithotripsy with various pulse durations (τ_p: 250 ~ 495 μsec) was investigated. Depending on pulse energy, optical pulse durations were divided into two regimes: short pulse (250~350 μsec) and long pulse (315~495 μsec). Retropulsion distance was measured as a function of pulse energy from 0.4 J to 1.2 J. Calculus phantoms made from plaster of Paris were ablated with a free running Ho:YAG laser using various optical fibers (200, 400, 600 μm) in water. In order to examine the ablation efficiency of two different pulse durations, a single pulse was applied, and the dynamics of the recoil action of a calculus phantom was monitored using a high-speed camera. The correlation among laser-induced topography, ablation volume, and retropulsion was evaluated. Higher pulse energy and larger fibers resulted in larger ablation volume and retropulsion. At a given pulse energy, optical pulses with different durations yielded comparable ablation volumes. The shorter duration pulses induced more retropulsion than longer pulses did at the same pulse energy. Larger retropulsion with the shorter pulse is thought to be induced by higher temperature at the vapor-solid interface, subsequently resulting in faster plume ejection with higher recoil momentum. The results suggest that a longer pulse could minimize retropulsion of the stone during lithotripsy.

The objective was to prove the advantage of endoscopic laser-urethroplasty over internal urethrotomy in acquired urethral strictures. Patients and Method: From January, 1996 to June, 2005, 35 patients with a mean age of 66 years were submitted to endoscopic laser-urethroplasty for strictures of either the bulbar (30) or membranous (5) urethra. The operations were carried out under general anesthesia. First of all, the strictures were incised at the 4, 8 and 12 o'clock position by means of a Sachse-urethrotom. Then the scar flap between the 4 and 8 o'clock position was vaporized using a Nd:YAG laser, wavelength 1060 nm and a 600 pm bare fiber, the latter always being in contact with the tissue. The laser worked at 40W power in continuous mode. The total energy averaged 2574 J. An indwelling catheter was kept in place overnight and the patients were discharged the following day. Urinalysis, uroflowmetry and clinical examination were performed at two months after surgery and from then on every six months. Results: No serious complications were encountered. Considering a mean follow-up of 18 months, the average peak flow improved from 7.3 ml/s preoperatively to 18.7 mVs postoperatively. The treatment faded in 5 patients ( 14.3% ) who finally underwent open urethroplasty. Conclusions: Endoscopic laser-urethroplasty yields better short-term results than internal visual urethrotomy. Long-term follow-up has yet to confirm its superiority in the treatment of acquired urethral strictures.

Recent advances in Thulium fiber laser technology have resulted in the availability of a compact, inexpensive, tunable, mid-infrared laser for potential use in laser surgery. The objective of this study was to tune the Thulium fiber laser wavelength and corresponding optical penetration depth to match the tissue thickness, and thus produce full-thickness, watertight tissue closure during microsurgical laser welding of urinary tissues. 1-cm-length incisions were made, ex vivo, in porcine ureters. Thulium fiber laser radiation with a wavelength of 1873 nm, power of 550-650 mW, and 750-μm-diameter spot was delivered to the tissue in continuous-wave mode through a 600-μm silica optical fiber. The fiber was scanned over the weld site once at a rate of 0.1 mm/s using a motion controller and linear stage controlled by a PC. Optical coherence tomography, histology, flow rates, and temperature measurements were used to optimize and evaluate laser welding success. Histologic analysis demonstrated full-thickness welding of the ureteral wall. Weld success rates ranged from 67% (8/12) at an incident laser power of 550 mW to 91% (10/11) at 650 mW. Peak flow rates greater than 200 ml/min were measured, however, mean flow rates were only about 50 ml/min. Average tissue temperatures increased with incident laser power from 59-89^oC. The tunable Thulium fiber laser may be useful for surgical applications requiring variable control of thermal coagulation depth, such as microsurgical laser tissue welding.

Bladder cancer is the fourth common malignant disease worldwide, accounting for 4% of all cancer cases. In Singapore, it is the ninth most common form of cancer. The high mortality rate can be reduced by early treatment following precancerous screening. Currently, the gold standard for screening bladder tumors is histological examination of biopsy specimen, which is both invasive and time-consuming. In this study ex vivo urine fluorescence cytology is investigated to offer a timely and biopsy-free means for detecting bladder cancers. Sediments in patients' urine samples were extracted and incubated with a novel photosensitizer, hypericin. Laser confocal microscopy was used to capture the fluorescence images at an excitation wavelength of 488 nm. Images were subsequently processed to single out the exfoliated bladder cells from the other cells based on the cellular size. Intensity histogram of each targeted cell was plotted and feature vectors, derived from the histogram moments, were used to represent each sample. A difference in the distribution of the feature vectors of normal and low-grade cancerous bladder cells was observed. Diagnostic algorithm for discriminating between normal and low-grade cancerous cells is elucidated in this paper. This study suggests that the fluorescence intensity profiles of hypericin in bladder cells can potentially provide an automated quantitative means of early bladder cancer diagnosis.

Photodynamic diagnosis (PDD) has become popular in bladder cancer detection. Several studies have however shown an increased false positive biopsies rate under PDD guidance compared to conventional cystoscopy. Raman spectroscopy is an optical technique that utilizes molecular specific, inelastic scattering of light photons to interrogate biological tissues, which can successfully differentiate epithelial neoplasia from normal tissue and inflammations in vitro. This investigation was performed to show the feasibility of NIR Raman spectroscopy in vitro on biopsies obtained under guidance of 5-ALA induced PPIX fluorescence imaging. Raman spectra of a PPIX solution was measured to obtain a characteristic signature for the photosensitzer without contributions from tissue constituents. Biopsies were obtained from patients with known bladder cancer instilled with 50ml, 5mg 5-ALA two hours prior to trans-urethral resection of tumor (TURT). Additional biopsies were obtained at a fluorescent and non-fluorescent area, snap-frozen in liquid nitrogen and stored at -80 °C. Each biopsy was thawed before measurements (10sec integration time) with a confocal Raman system (Renishaw Gloucestershire, UK). The 830 nm excitation (300mW) source is focused on the tissue by a 20X ultra-long-working-distance objective. Differences in fluorescence background between the two groups were removed by means of a special developed fluorescence subtraction algorithm. Raman spectra from ALA biopsies showed different fluorescence background which can be effectively removed by a fluorescence subtraction algorithm. This investigation shows that the interaction of the ALA induced PPIX with Raman spectroscopy in bladder samples. Combination of these techniques in-vivo may lead to a viable method of optical biopsies in bladder cancer detection.

Er:YAG laser (wavelength 2.94 μm) operating both in free-running and Q-switched regime was designed and developed for the purpose of ureter wall perforation or incision, and urinary stones fragmentation. Component of this system was a special transfer part consisted of a cyclic olefin polymer-coated silver (COP/Ag) hollow glass waveguide (inner/outer diameter 700/850 μm or 320/450 μm) with a sealed cap for a contact treatment. Maximum pulse interaction energy and length for free-running Er:YAG laser were 100 mJ and 200 μs, respectively (corresponding intensity was 130 kW/cm² for the 700 μm waveguide and 500 kW/cm² for the 320 μm waveguide). Maximum interaction pulse energy and length in Q-switched regime were 30 mJ and 70 ns, respectively (corresponding intensity was 111 MW/cm² for the 700 μm waveguide and 357 MW/cm² for the 320 μm waveguide). Basic interaction characteristics and parameters of ureter wall perforation and urinary stones fragmentation were found. For that reason the number of pulses needed for the perforation of the ureter wall tissue (thickness ~1mm), ablation threshold and ablation rate were measured for free-running and Q-switched Er:YAG laser radiation. Subsequently, the investigated tissue samples were histologically evaluated after the interaction. The ablation rate of the Q-switched Er:YAG laser radiation was higher compared to the free-running radiation. The application of Q-switched Er:YAG laser radiation on ureter tissue resulted in minimum tissue alteration (up to 50 μm from the surface) without any influence on the deeper layers. The possibility of urinary stones perforation with free-running Er:YAG laser radiation (with maximum interaction energy) was also demonstrated.

Introduction: Early detection of bladder carcinoma is very important clinical problem. Diagnostic yield of white light cystoscopy with random biopsies remains poor. The use of exogenous fluorescence significantly increases the sensitivity, but specificity remains low. We analyzed diagnostic efficacy of OCT during white light cystoscopy and combined use of OCT and fluorescence cystoscopy. Materials and methods: An OCT device (1280 nm wavelength with 3 mW power, 8 Fr endoscopic probe, in-depth resolution 15 μm in tissue, lateral resolution 30 μm, acquisition time 1.5 sec for a 200x200 pixels image) was used in combination with a standard Karl Storz fluorescence cystoscope. A 3% solution of 5-ALA was instilled intravesically for 2 hours before the procedure. Initial examination was made under white light. OCT imaging and biopsy of all fluorescence zones were performed in blue light. 20 patients were studied. The study is ongoing. Results: 80 fluorescence zones (16 exophytic and 64 flat lesions) were analyzed with OCT. All exophytic zones were correctly detected by OCT and white light cystoscopy. Out of 64 flat fluorescent areas, 56 had benign histopathology readings, with 45 of them having the benign type of OCT images. Of 8 fluorescent zones with neoplastic histopathology, OCT correctly detected all 8. Based on this preliminary data, OCT could help to avoid 80% of unnecessary biopsies/resections. Conclusion: Combined use of OCT imaging and fluorescence cystoscopy can substantially improve diagnostic yield of bladder neoplasia detection.

One of the important problems of modern medicine is treatment of urogenital diseases.^1-2 There is a set of the treatment methods for such problems, but any of them does not obey the modern physicians completely.^3-4 Our aim is to present the new combined therapeutic apparatus called "Yarovit" (produced in Russia, in collaboration between Bauman Moscow State University of Technology and Scientific Production Association and Medical Center "Yarovit") which successfully applied in clinics for cure the patients with copulative dysfunction diseases.^5-6 At this apparatus "Yarovit" (description model have abbreviation AMVL-0 1) there is a combination of vacuum decompression (0.1-0.4 kgs/cm²) and light emitting diodes matrix system (660 nm, 1-3 mW/cm²). In treatment procedure apparatus can be applied together with expanded module "Intratherm" (39 °C on average), which has rectal heating elements. The latest clinical studies were made together with volunteer participation of more then one hundred patients, and received results showed the good dynamic of healing. That let to conclude these combinations of physical therapeutic methods supplement each other and in conjunction provides a significant clinical effect. The further developments of such apparatuses are discussed.

Vocal cords disorders constitute an important problem for people suffering from them. Particularly the reduction of mucosal wave movement is not appropriately treated by conventional therapies, like drugs administration or surgery. In this work, an alternative therapy, consisting in controlled temperature increases by means of optical sources is proposed. The distribution of heat inside vocal cords when an optical source illuminates them is studied. Optical and thermal properties of tissue are discussed, as a basis for the appropriate knowledge of its behaviour. Propagation of light is shown using the Radiation Transfer Theory (RTT) and a numerical Monte Carlo model. A thermal transfer model, that uses the results of the propagation of radiation, determines the distribution of temperature in the tissue. Two widely used lasers are considered, Nd:YAG (1064 nm) and KTP (532 nm). Adequate amounts of radiation, resulting in temperature rise, must be achieved in order to avoid damage in vocal cords and so to assure an improvement in the vocal functions of the patient. The limits in temperature should be considered with a combined temperature-time and Arrhenius analysis.

We present the results in optical coherence tomography (OCT) visualization of structural changes in human oral cavity and pharyngeal mucosa for 14 patients in the process of radiation and chemoradiation therapy. Typical mucosal changes are seen as a decrease in the tissue layer contrast progressing to a complete contrast loss as severe mucosal damage occurs. Similar evolution in OCT images was observed for all patients. OCT changes can be seen prior to visual mucosal changes and increased as more mucosal damage occurs. Moreover, OCT was able to obtain information on the specific features of the patient response depending on the irradiation method and the individual radiosensitivity.

Objectives: 1. To investigate preventive effect of LLL on gentamicin-induced vestibular ototoxicity. 2. To evaluate the effectiveness of lower level laser (LLL) in the treatment of tinnitus. Methods: 1. Twenty guinea pigs were divided into control and laser groups. Vestibular ototoxicity was induced by intratympanic injection of gentamicin into left ear. LLL was irradiated into left ear canal of animals in laser group. Vestibular function of the animals was evaluated with vertical and off-vertical axis rotation testing. 2. Forty patients with tinnitus were treated with ginkgo biloba orally and randomly divided into control and laser groups. The 20 patients of laser group received 80.4 J/cm² of 830 nm laser, 3 times per week for 4 weeks, via transmeatal irradiation. Tinnitus was evaluated by visual analogue scale (VAS) and tinnitus handicap inventory (THI). Results: 1. Preventive effect of LLL to gentamicin induced vestibular ototoxicity was demonstrated by preventing reduction of gain in slow harmonic acceleration test and modulation in the off-vertical axis rotation test. 2. Eleven of 20 laser group patients have shown significant improvement in VAS and THI compared to those of the control group. Conclusions: 1. LLL therapy may have preventive effect to vestibular ototoxicity. 2. LLL therapy in combination with ginkgo biloba seems to be worth trying on patients with tinnitus.

The current standard procedure to ensure the diagnosis, if tissue is malignant, is still an invasive one. Optical coherence tomography (OCT) is a new non-invasive method to investigate biological tissue. In this study OCT was used on porcine and on human vocal folds. The optical penetration depth of the used radiation is up to 2 mm. Three different OCT application systems were used. The first is a high resolution OCT, which works in contact mode. It was used to examine porcine vocal folds ex vivo. Porcine vocal folds were assigned to defined areas and examined by OCT in contact mode followed by traditional histo-morphological analysis. The second OCT is fiber based. It also works in contact mode. Images of human vocal folds were done in contact mode. They were compared with a typical histo-morphological image of a human vocal fold. The third application system works in non contact to the tissue. It was integrated in a conventional laryngoscope. Human vocal folds were examined in vivo. Single layers of the vocal folds could be distinguished from each other with all used systems. Pathological alterations could be seen. Imaging is possible in real time. General anaesthesia is not necessary. OCT makes it possible to get a view under the surface of the vocal fold without being invasive.

It is known that electrical current injected from cochlear implant contacts spreads within the cochlea, causing overlapping stimulation fields and possibly limiting the performance of cochlear implant users. We have investigated an alternative mechanism to stimulate auditory neurons in the gerbil cochlea using a laser, rather than electrical current. With the laser, it is possible to direct the light to a selected, known volume of tissue that is smaller than the electrically stimulated population of cells. In the present experiments, a transiently expressed transcription factor, c-FOS, was used to stain activated nerve cells. Immunohistochemical staining for c-FOS in the cochlea shows a small area of optical stimulation, which occurs directly opposite to the optical fiber. Additionally, masking data indicate that the laser can stimulate a small population of cells similar to an acoustic toneburst. Smaller populations of stimulated cells could reduce the amount of overlap in stimulation fields and allow more stimulation contacts in a neuroprothesis.

Cigarette smoking is the most preventable cause of death in the United States. Researchers have extensively studied smoking in regards to its association with cancer, cardiovascular, and pulmonary disease. In contrast, the impact of cigarette smoking on skin has received much less attention. To provide a better understanding of the effect of cigarette smoking on the human dermal layer, this study used multi-photon microscopy (MPM) to examine collagen in organotypic skin models exposed to cigarette smoke condensate (CSC). Adult and neonatal organotypic tissue-engineered artificial skin models (RAFTs) were constructed and exposed to varying concentrations of CSC. Imaging of the RAFTs was performed using MPM and second-harmonic generation signals (SHG), which allowed for collagen structure to be viewed and analyzed as well as for collagen density to be assessed from derived depth-dependent decay (DDD) values. RAFT contraction as related to exposure concentration was monitored as well. Results indicated a dose dependent between contraction rates and CSC concentration. Collagen structure showed more preservation of its original structure at a greater depth in RAFTs with higher concentrations of CSC. No clear trends could be drawn from analysis of derived DDD values.

It is well known that the relation between the partial pressure of oxygen in blood (PO₂) and the hemoglobin oxygen saturation in blood (SO₂) is given by the oxyhemoglobin dissociation curve. In this study, we investigate if a similar relation exists in tissue. The PO₂ in tissue was approximated by the transcutaneous partial pressure of oxygen (TcPO₂) measured by TCM3^TM Transcutanous pO₂/pCO₂ Monitoring System; and the SO₂ in tissue (StO₂) was measured by ODISsey^TM Tissue Oximeter. The study showed that the TcPO₂ versus StO₂ relation is similar to the dissociation curve in blood, as expected.

The early stage development studies of novel implantable continuous metabolite sensor systems for glucose, lactate and fatty acids are discussed. These sensors utilize non-enzymatic "reagentless" sensor systems based on NIR fluorophore-labeled binding proteins. For in vivo applications, NIR fluorescence based systems (beyond 600 nm) have the added benefit of reduced interference from background scattering, tissue and serum absorption and cell auto-fluorescence. The long wavelength emission facilitates implanted sensor disks to transmit fluorescence to an external reader through wireless connections and the resulting fluorescence signals can be correlated to metabolite concentrations. We have developed a prototype optical system that uses a bifurcated optical fiber to transmit excitation and read emission at the surface of the skin. With this system, fluorescence signals were read over time through animal skin. The changes in glucose concentration were studied using immobilized sensor proteins and were compared to non-immobilized sensors in solution. For sensors in solution, no response delay was observed. For immobilized systems, the fluorescence response showed a delay corresponding to the diffusion time for the metabolite to equilibrate within the sensor.

Identifying injuries, deformities, and diseases by non-invasive instrumental means has been a major innovation in medicine. Diagnostic and imaging medical devices have revolutionized diagnosis and surgery, providing more efficient way to identify injuries and diseased or damaged tissues. In this paper, identification of different animal tissues using a miniature near-infrared (NIR) spectrometer will be demonstrated. Each tissue type contains different amounts of moisture and proteins, and by using this miniature spectrometer, a miniature fiber-optic probe and chemometrics; the ability to recognize tissues spectral differences is established.

An apparatus for minimally-invasive in vivo measurements of the ciliary beat frequency (CBF) has been developed. The instrument is based on speckle interferometry technique. To be able to distinguish speckle fluctuations caused by motion of the detection probe with respect to the epithelium from the oscillations due to ciliary beat, an image fiber bundle with a position sensitive CCD detector is used. A fast CCD camera taking images with a frame rate of 500 fps provides position and timing resolution to simultaneously detect the CBF and the probe displacement. The probe can be inserted into the working channel of a standard bronchoscope. Two prototypes with different observation angles (straight and 90 degree) were constructed and are now subject to testing. First results of CBF measurements in a human nose are presented.

Previous ex vivo tissue studies utilizing deep laser heating combined with contact cooling of the tissue surface produced noninvasive thermal destruction of subsurface tissue structures in skin and liver samples. This study describes the design and preliminary in vivo testing of two integrated laser/cooling probes for simultaneous Nd:YAG laser irradiation and sapphire contact cooling of liver and skin tissues in an in vivo, acute porcine model for potential use in laparoscopic and endoscopic surgery. Nd:YAG laser radiation with a wavelength of 1.06 μm, power of 20 W, 7.5-mm-diameter spot, 500-ms pulse length, and repetition rate of 0.625 Hz, was delivered to the tissue with a total irradiation time of 16 s. The tissue surface was continuously cooled with a sapphire plate maintained at -5 ^oC, and with pre- and post-ablation cooling times measuring 120 s and 30 s, resulting in a total operation time of 166 s per a lesion. Thermal lesions were created in liver and skin at a 1-mm depth below the tissue surface and with a 3-4 mm diameter. The laser parameters and lesion dimensions were comparable to previous ex vivo tissue studies. Preliminary in vivo animal studies demonstrate noninvasive creation of subsurface thermal lesions in tissue using Nd:YAG laser irradiation in conjunction with sapphire contact cooling. Chronic wound healing studies will be necessary to optimize the laser and cooling parameters. Potential clinical applications include endoscopic laser treatment of female stress urinary incontinence and thermal coagulation of early stage bladder tumors.

Applications of lasers (light amplification by stimulated emission of radiation) in various disciplines of medicine including Urology are well developed. Urology is among the medical specialties that apply many different types of laser systems to treat a broad spectrum of clinical conditions ranging from genital, bladder and urethral tumors to the treatment of benign prostate hyperplasia (BPH), urethral strictures, and stones. The specific application of various laser systems depends on the characteristics of the laser itself, delivery media for the beams, laser-tissue interaction and the desired effect. These complex conditions require an intensive and continuous exchange of information between non-medical researchers and physicians to verify "what is currently technically possible and what is medically needed". Only this exchange can lead to the development of new laser systems. While lasers have become the treatment of choice in some conditions, they could not, despite excellent clinical results, replace conventional therapy options in others. Nonetheless, the use and the introduction of lasers of different wavelengths forces urologists to keep step with the fast developing laser technology. This paper reviews current indications for clinical laser applications relevant to urology and the advantages and disadvantages of using lasers for the management of various urological lesions.

In 1991 the World Health Organization (WHO) defined osteoporosis as a "loss of bone mass and micro architectural deterioration of the skeleton leading to increased risk of fracture."^1,2 Since microarchitecture can not be measured directly, a panel of the WHO recommended that the diagnosis be made according to a quantifiable surrogate marker, calcium mineral, in bone. Subsequently in 1994, the definition focused on the actual bone "density," giving densitometric technology a central place in establishing the diagnosis of osteoporosis.^3,4 But soon it became obvious that there was only limited correlation between bone mineral density (BMD) and actual occurrence of fractures and that decreases in bone mass account for only about 50% of the deterioration of bone strength with aging. In other words only about 60% of bone strength is related to BMD.⁵ Recent developments in bone research have shown that bone mineral density in itself is not sufficient to accurately predict fracture risk. Bone is composed of inorganic calcium apatite crystals that mineralize an organic type I collagen matrix. The degree of mineralization, the properties of the collagen matrix, crystal size, trabecular orientation, special distribution of the different components and many more factors are all impacting bone strength.^6-14 Human cadaver studies have confirmed the correlation between bone density and bone.²⁶ strength.^5,15-20 Changes in cancellous bone morphology appear to lead to a disproportionate decrease in bone strength.^21-26 When postmenopausal women are stratified by age, obvious differences between BMD and actual fracture risk are observed.²⁴ Felsenberg eloquently summarizes what he calls the "Bone Quality Framework." In great detail he talks about the geometry and micro- architecture of bone and how the different components are related to functional stability.²⁷ Are our current testing modalities appropriately addressing these structural factors? Are we keeping in mind that in screening for osteoporosis the key variable is fragility, not bone density itself? All currently FDA approved and commercially available equipments for the evaluation of bone status claim that they - at least indirectly - assess the biological fracture risk. This review summarizes an extensive current literature research covering FDA approved as well as experimental devices for the evaluation of bone. The pros and cons of the different techniques are discussed in the context of diagnostic accuracies and practical implications.

A lot of industry workers all over the world have dealings with a strong mechanical vibration as with daily technology processes. Very often such long-time professional vibration causes the so-called professional "vibration disease", in English literature "white fingers syndrome", caused by a local vibration of hands. Among different clinical features of the vibration disease a leader's part of them consists of different cardiovascular and trophic disorders of tissues. The objects of the present study were the peripheral blood microcirculation, peripheral blood oxygenation and tissues hypoxia state in a finger skin under vibration disease. For this purpose we have used a combined noninvasive spectrophotometry diagnostic technique consisting of Laser-Doppler Flowmetry (LDF), Laser Fluorescent Diagnostics (LFD) and Tissues Reflectance Oximetry (TRO). The results show good possibilities of all mentioned above diagnostic methods in estimation of different vascular disorders. A good correlation between persistent microcirculation disorders and trophic disturbances revealed in tissues of distal ends of upper extremities of the patients with vibration disease was estimated. Additionally, in present study with the use of real and long-time TRO and LDF methods a good correlation between LDF and TRO data including correlation in detected rhythms of blood microcirculation was estimated as well.

We at Fibersense and Signals are pleased to report the research and development of novel multiple-wavelength and multi-spectral techniques and devices to distinguish glucose from a comprehensive panel of other spectrally evident and interactive blood components such as: albumin, salts, urea and amino acids. A highly selective, glucose-specific bio-spectroscopic technique has been developed to provide clinically-relevant information regarding glucose concentrations. Furthermore, this work has resulted in novel multi-wavelength technology and techniques for the non-invasive detection and continuous non-invasive monitoring of blood glucose levels. As a developer of fiber optic, photonic and laser-related products as well as optical test and measurement instrumentation, Fibersense has used an essentially problem-solving approach to address practical issues that have long hindered progress in non-invasive glucose detection and monitoring, and has overcome the limitations of previous techniques. We have fabricated and optimized optical sources for this application, specifically with respect to useful wavelength ranges, transmittance rates, reflectance, absorption and scattering/dispersion. We have also characterized a multi-wavelength combination and procedure to address and screen out the spectral overlapping of glucose and common blood compounds within physiological criteria. Furthermore, attention has been paid to the viable optical launching of effective wavelengths and the ultra-sensitive optical pick up and detection within very tight wavelength margins to generate useful blood glucose information. The culmination of these techniques has been the design and fabrication of miniaturized, non-invasive multi-channel glucose sensors with dynamic self-calibration features for use in in vitro and experimental in vivo applications.

Optical properties of tissues and tissue components are important parameters in biomedical optics. We report measurements of tissue refractive index n, and the attenuation coefficient μ_t using optical coherence tomography of individual vascular wall layers and plaque components. Moreover, since the temperature dependence of optical properties is widely known, we compared measurements at room and body temperatures. A decrease of n and μ_t was observed in all samples, with most profound effect on samples with high lipid content. The sample temperature is of influence on the quantitative measurements within OCT images. For extrapolation of ex vivo experimental results, especially for structures with high lipid content, this effect should be taken into account.

In this paper we develop the concept of a tissue-engineered optical phantom that uses engineered tissue as a phantom for calibration and optimization of biomedical optics instrumentation. With this method, the effects of biological processes on measured signals can be studied in a well controlled manner. To demonstrate this concept, we attempted to investigate how the cellular remodeling of a collagen matrix affected the optical properties extracted from optical coherence tomography (OCT) images of the samples. Tissue-engineered optical phantoms of the vascular system were created by seeding smooth muscle cells in a collagen matrix. Four different optical properties were evaluated by fitting the OCT signal to 2 different models: the sample reflectivity ρ and attenuation parameter μ were extracted from the single scattering model, and the scattering coefficient μ_s and root-mean-square scattering angle θ_rms were extracted from the extended Huygens-Fresnel model. We found that while contraction of the smooth muscle cells was clearly evident macroscopically, on the microscopic scale very few cells were actually embedded in the collagen. Consequently, no significant difference between the cellular and acellular samples in either set of measured optical properties was observed. We believe that further optimization of our tissue-engineering methods is needed in order to make the histology and biochemistry of the cellular samples sufficiently different from the acellular samples on the microscopic level. Once these methods are optimized, we can better verify whether the optical properties of the cellular and acellular collagen samples differ.

Current evidence indicates that most plaques classified as vulnerable or ruptured plaques do not lead to unstable angina or myocardial infarction. Improved methods are needed to risk stratify plaques to identify those which lead to most acute coronary syndromes. Collagen depletion in the intima overlying lipid collections appears to be a critical component of unstable plaques. In this study, we use polarization sensitive optical coherence tomography (PS-OCT) for the assessment of coronary plaque collagen. Collagen is birefringent, meaning that different polarization states travel through it at different velocities. Changes in PS-OCT images are a measure of tissue birefringence. Twenty-two coronary artery segments were imaged with PS-OCT and analyzed by picrosirius staining (a measure of collagen intensity and fiber size) and trichrome blue. The regression plot between PS-OCT changes and measured collagen yielded a correlation coefficient value of 0.475 (p<0.002). Good correlation was noted between two blinded investigators both with respect to PS-OCT measurements as well as luminosity as assessed by picrosirius. The predictive value of a PS-OCT measurement of negligible birefringence (less than 33% change) for minimal collagen was 93% while the predictive value of high birefringence (greater than 66% change) for high collagen concentrations was 89%. The effect of fiber type (chemical composition) was minimal relative to the effect due to fiber concentration. The capability of PS-OCT to assess plaque collagen content, in addition to its ability to generate high resolution structural assessments, make it a potentially powerful technology for identifying high risk plaques.

Atherosclerosis is traditionally viewed as a disease of uncontrolled plaque growth leading to arterial occlusion. More recently, however, occlusion of the arterial lumen is being viewed as an acute event triggered by plaque rupture and thrombosis. An atheromatous plaque becomes vulnerable to sudden activation and/or rupture when a constellation of processes are activated by various trigger mechanisms. There is growing evidence that the vulnerability (i.e. susceptibility to rupture) and thrombogenic nature of the plaque need to be taken into account in the planning and treatment of the disease. X-ray fluoroscopy and intravascular ultrasound, the current clinical diagnostic tools are not capable of the providing a complete histological picture of the plaque region. Intravascular diagnostic imaging of coronary atherosclerotic plaques by optical means to assess plaque, patient risk and assist in planning treatment strategies represents the future in angioplasty treatment by interventional cardiologists. The techniques which will enable a clinically acceptable and reliable intravascular diagnostic platform are currently being investigated and compared to the clinical standard of histology. Currently, we are investigating the use of a number of optical and imaging techniques for biochemical analysis of arterial tissue including Raman, near infrared and fluorescence spectroscopies. Biochemical imaging will provide compositional information on collagen, elastin, lipid and thrombogenic by-products as well as gauging inflammation and tissue remodeling activity levels. To complement the functional biochemical imaging, optical coherence tomography will be provide structural morphological imaging. The synergistic combination of functional and structural imagery will provide the interventional cardiologist with a complete clinical picture of the atherosclerotic plaque region. The clinician can use this diagnostic information to plan a personalized treatment procedure based on the entire clinical presentation.

The present paper introduces an integrated sensing and processing spectroscopic system that combines Multivariate Factor Analysis (MFA) and Molecular Factor Computation (MFC) with a broad band light source, incorporating a rotary, multichannel optical device to produce real-time, spectroscopic analysis. The system is able to distinguish samples of cholesterol and collagen/elastin within a time range of 25 ms, by using near-infrared reflectance spectroscopy. The potential for the use of the device for in-vivo coronary angiography will be discussed. The system presents several key features that make it unique as a spectroscopic tool: it is compact and rugged with multilayer interference wavelength selection, and capable of operation in high vibration environments. The system is also fast, with the capacity to reduce the measurement delay even further; it is broad band, with a bandwidth limited by the optical source and the sensitivity of the photodetector used; it will allow quantitative, simultaneous, multispecies detection; and it is amenable to different optical delivery/detection schemes. Standard telecommunications-grade parts comprise the system, bridging the gap between the prototype and manufactured product; these include: optical fibers, electrically actuated optical switches, and free-space optics elements. A detailed description of the data analysis algorithm will be presented, with special emphasis on its use for multispecies detection.

This study introduces new methods of time-resolved laser-induced fluorescence spectroscopy (TR-LIFS) data analysis for tissue characterization. These analytical methods were applied for the detection of atherosclerotic vulnerable plaques. Upon pulsed nitrogen laser (337 nm, 1 ns) excitation, TR-LIFS measurements were obtained from carotid atherosclerotic plaque specimens (57 endarteroctomy patients) at 492 distinct areas. The emission was both spectrally- (360-600 nm range at 5 nm interval) and temporally- (0.3 ns resolution) resolved using a prototype clinically compatible fiber-optic catheter TR-LIFS apparatus. The TR-LIFS measurements were subsequently analyzed using a standard multiexponential deconvolution and a recently introduced Laguerre deconvolution technique. Based on their histopathology, the lesions were classified as early (thin intima), fibrotic (collagen-rich intima), and high-risk (thin cap over necrotic core and/or inflamed intima). Stepwise linear discriminant analysis (SLDA) was applied for lesion classification. Normalized spectral intensity values and Laguerre expansion coefficients (LEC) at discrete emission wavelengths (390, 450, 500 and 550 nm) were used as features for classification. The Laguerre based SLDA classifier provided discrimination of high-risk lesions with high sensitivity (SE>81%) and specificity (SP>95%). Based on these findings, we believe that TR-LIFS information derived from the Laguerre expansion coefficients can provide a valuable additional dimension for the diagnosis of high-risk vulnerable atherosclerotic plaques.

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.

Wavefront synchronization is an important aspect preceding the termination of ventricular fibrillation (VF). We designed a novel approach using multi-electrode pacing that could reduce the energy consumption of successful defibrillation by more than 100-fold. We compared the efficacy of a synchronized pacing algorithm using optical recording-guided pacing at the right ventricle and the posterior left ventricle in isolated rabbit hearts. Two modes of pacing were used in the study. The electrodes were individually controlled (independent mode, ISyncP) or fired together (simultaneous mode, SSyncP) when triggered by tissue polarization at a reference site separate from the pacing sites. The hearts were stained with voltage-sensitive dye and illuminated with laser for epifluorescence imaging during pacing. The number of phase singularities of VF propagation was used to estimate the complexity of VF. A decrease in the number of PS signified a higher degree of VF organization. We found that the pacing algorithm was more effective if the pacing was applied to the posterior left ventricle than to the right ventricle. Pacing in the posterior left ventricle resulted in a 25.3% decrease in the number of PS for ISyncP and a 31% decrease for SSyncP. Pacing in the right ventricle resulted in a 29.0% decrease for ISyncP and a 2.3% increase for SSyncP. The 5mA current reflected a 27.8% decrease for ISyncP and a 32.2% decrease for SSyncP, whereas the 10mA current reflected a 27.2% decrease for ISyncP and a 5.2% increase for SSyncP. We conclude that synchronized pacing can induce VF organization, and the efficacy is higher when pacing in the left ventricle with 5 mA.

High performance functional imaging is needed for dynamic measurements of neural processing in retina. Emerging techniques of visual prosthesis also require advanced methodology for reliable validation of electromagnetic stimulation of the retina. Imaging of fast intrinsic optical responses associated with neural activation promises a variety of technical advantages over traditional single and multi-channel electrophysiological techniques for these purposes, but the application of fast optical signals for neural imaging has been limited by low signal to noise ratio and high background light intensity. However, using optimized near infrared probe light and improved optical systems, we have improved the optical signals substantially, allowing single pass measurements. Fast photodiode measurements typically disclose dynamic transmitted light changes of whole retina at the level of 10^-4 dI/I, where dI is the dynamic optical change and I is the baseline light intensity. Using a fast high performance CCD, we imaged fast intrinsic optical responses from isolated retina activated by a visible light flash. Fast, high resolution imaging disclosed larger local optical responses, and showed evidence of multiple response components with both negative- and positive-going signals, on different timescales. Darkfield imaging techniques further enhanced the sensitivity of optical measurements. At single cell resolution, brightfield imaging disclosed maxima of optical responses ~5% dI/I, while darkfield imaging showed maxima of optical responses exceeding 10% dI/I. In comparison with simultaneous electrophysiological recording, optical imaging provided much better localized patterns of response over the activated area of the retina.

As a molecular probe of tissue composition, infrared spectroscopic imaging serves as an adjunct to histopathology in detecting and diagnosing disease. In the past it was demonstrated that the IR spectra of brain tumors can be discriminated from one another according to their grade of malignancy. Although classification success rates up to 93% were observed one problem consists in the variation of the models depending on the number of samples used for the development of the classification model. In order to open the path for clinical trials the classification has to be validated. A series of classification models were built using a k-fold cross validation scheme and the classification predictions from the various models were combined to provide an aggregated prediction. The validation highlights instabilities in the models, error rates, sensitivity as well as specificity of the classification and allows the determination of confidence intervals. Better classification models could be achieved by an aggregated prediction. The validation shows that brain tumors can be classified by infrared spectroscopy and the grade of malignancy corresponds reasonably to the histopathological assignment.

Background: Oral application of 20 mg/kg body weight of 5-aminolevulinic acid (ALA) results in a highly specific accumulation of fluorescent Protoporphyrin IX (PpIX) in malignant glioma tissue. In a clinical phase II study, Photodynamic Therapy (PDT) to postsurgical tumor-remnants was monitored by fluorescence spectroscopy and photobleaching of PpIX was detected. Methods: Fluorescence spectra were measured from the tumor remnant pre and at intervals during PDT with 100, 150 and 200 J/cm² and from adjacent normal cortex. The spectra obtained were fitted with spectra from pure PpIX, autofluorescence and photoproducts, the fit-parameters quantifying the contributions of these components to the measured spectrum. Results: The PpIX-fluorescence bleached to 8%, 16% and 1% of the initial intensity for the 100, 150 and 200 J/cm²-groups (median values). Photoproduct formation was minimal. PpIX-fluorescence of normal cortex was 6% of tumor remnant fluorescence (median, range: 0% to 49%).

Introduction: High grade gliomas are characterised by rapid and invasive growth, that cause massive tissue destruction at both the tumour- brain boarder as well as in regions remote from the tumor core. Eradication or inhibition of infiltrating glioma cells poses a significant clinical challenge that is unlikely to be solved using conventional treatment regimens consisting of ionizing radiation and chemotherapeutic agents. In this study we evaluated the effects of ALA mediated photodynamic therapy (PDT) on the invesivness of human glioma cells migrating from implanted multicell tumor spheroids. Materials and method 3-400nm diameter tumor spheroids, derived from the human glioma cell line ACBT, were implanted into a gel matrix of collagen type I. 24 hours following implantation there was a significant invasion of the surrounding gel by individual tumor cells to an average distance of 400nm. The cultures were incubated in increasing concentrations of ALA (10-1000 ug/ml) for four hours and then exposed to 635nm laser light in a titration of both fluence level and fluence rate. Results Fluences of 25J/cm² were clearly cytotoxic for both the infiltrating cells as well as the spheroids at all ALA concentrations. Fluence levels of 6J did not stop the spheroid growth or prove cytotoxic to the glioma cells that had previously migrated into the gel, in a majority of cultures but inhibited further migration of the cells by 80-90% compared to control. Conclusion: Measurement of cell survival and cell proliferation indices seemed to indicate a direct migratory inhibition effect on the invading cells and not cytotoxicity as the most likely mechanism for this observation.

A significant contributory factor to the poor prognosis of patients with glioblastoma multiforme is the inability of conventional treatments to eradicate infiltrating glioma cells. A syngeneic rat brain tumor model is used to investigate the effects of aminolevulinic acid (ALA)-mediated photodynamic therapy (PDT) on small clusters of tumor cells sequestered in normal brain. The intrinsic sensitivity of rat glioma cells to PDT was investigated by exposing ALA-incubated cells to a range of radiant exposures and irradiances using 635 nm light. Biodistribution studies were undertaken on tumor-bearing animals in order to determine the tumor selectivity of the photosensitizer following systemic administration (i.p.) of ALA. Effects of ALA-PDT on normal brain and gross tumor were evaluated using histopathology. Effects of PDT on isolated glioma cells in normal brain were investigated by treating animals 48 h after tumor cell implantation: a time when the micro-clusters of cells are protected by an intact blood-brain-barrier (BBB). Rat glioma cells in monolayer are susceptible to ALA-PDT - lower irradiances are more effective than higher ones. Fluorescence microscopy of frozen tissue sections showed that photosensitizer is produced with better than 200:1 tumor-to-normal tissue selectivity following i.p. ALA administration. ALA-PDT resulted in significant damage to both gross tumor and normal brain, however, the treatment failed to prolong survival of animals with newly implanted glioma cells compared to non-treated controls if the drug was delivered either i.p. or directly into the brain. In contrast, animals inoculated with tumor cells pre-incubated in vitro with ALA showed a significant survival advantage in response to PDT.

Optical coherence tomography (OCT) is a non-invasive imaging technique with a micrometer resolution. It allows non-contact / non-invasive analysis of central nervous system tissues with a penetration depth of 1-3,5 mm reaching a spatial resolution of approximately 4-15 μm. We have adapted spectral-domain OCT (SD-OCT) and time-domain OCT (TD-OCT) for intraoperative detection of residual tumor during brain tumor surgery. Human brain tumor tissue and areas of the resection cavity were analyzed during the resection of gliomas using this new technology. The site of analysis was registered using a neuronavigation system and biopsies were taken and submitted to routine histology. We have used post image acquisition processing to compensate for movements of the brain and to realign A-scan images for calculation of a light attenuation factor. OCT imaging of normal cortex and white matter showed a typical light attenuation profile. Tumor tissue depending on the cellularity of the specimen showed a loss of the normal light attenuation profile resulting in altered light attenuation coefficients compared to normal brain. Based on this parameter and the microstructure of the tumor tissue, which was entirely absent in normal tissue, OCT analysis allowed the discrimination of normal brain tissue, invaded brain, solid tumor tissue, and necrosis. Following macroscopically complete resections OCT analysis of the resection cavity displayed the typical microstructure and light attenuation profile of tumor tissue in some specimens, which in routine histology contained microscopic residual tumor tissue. We have demonstrated that this technology may be applied to the intraoperative detection of residual tumor during resection of human gliomas.

An in-vivo tumor model composed of multicellular human glioma spheroids implanted on a shell-less chorioallantoic membrane (CAM), has been developed. Following removal of a portion of the ectodermal epithelium layer of the CAM, human glioma spheroids were implanted on day 7 of embryonic development. Tumor invasion, rapid growth and vasculature formation were observed 7 days post implantation. Single tumor cell migration towards blood vessels, angiogenesis and satellite tumor growth were also evident. The human tumor/CAM model is being used to examine the effects of combined ALA PDT and anti-angiogenic agents. The shell-less CAM is well suited for topical, i.p. and i.v. photosensitizer and/or drug application.

To investigate the correlation between light scattering and tissue viability of brains, we performed measurements of multiwavelength diffuse reflectance and autofluorescence for brains during loss of tissue viability. As a model temporally losing tissue viability of brain, a perfused brain in a rat was used. Diffuse reflectance at 620 nm, which is an isosbestic point of cytochrome oxidase (cyt. ox.), showed that a triphasic, drastic change in light scattering occurred 220 - 300 s after starting perfusion. After this event, light scattering stayed at a higher level than that before this event. We observed a different behavior between the reflectance at 620 nm and that at 605 nm, showing that the tissue absorption at 605 nm was increasing. This is attributable to the reduction of heme a+a₃ in cyt. ox., which preceded the triphasic change in light scattering. An important difference was also found between the reflectance at 620 nm and that at 800 nm, indicating that the absorption decrease at 800 nm presumably due to the reduction of copper A in cyt. ox. coincided with the triphasic change in light scattering; the reduction of copper A in cyt. ox. indicates the decrease in ATP production. Change in fluorescence of NADH did not correlate either with change in light scattering or with the reduction of cytochrome oxidase. These findings suggest that light scattering is useful as an indicator of brain tissue viability.

Developmental dyslexia, a kind of prevalent psychological disease, represents that dyslexic children have unexpected difficulties in phonological processing and recognition test of Chinese characters. Some functional imaging technologies, such as fMRI and PET, have been used to study the brain activities of the children with dyslexia whose first language is English. In this paper, a portable, 16-channel, continuous-wave (CW) NIRS instrument was used to monitor the concentration changes of each hemoglobin species when Chinese children did the task of phonological processing and recognition test. The NIRS recorded the hemodynamic changes in the left prefrontal cortex of the children. 20 dyslexia-reading children (10~12 years old) and 20 normal-reading children took part in the phonological processing of Chinese characters including the phonological awareness section and the phonological decoding section. During the phonological awareness section, the changed concentration of deoxy-hemoglobin in dyslexia-reading children were significantly higher (p<0.05) than normal-reading children in the left ventrolateral prefrontal cortex (VLPFC). While in the phonological decoding section, both normal and dyslexic reading children had more activity in the left VLPFC, but only normal-reading children had activity in the left middorsal prefrontal cortex. In conclusion, both dyslexic and normal-reading children have activity in the left prefrontal cortex, but the degree and the areas of the prefrontal cortex activity are different between them when they did phonological processing.

Photodynamic therapy (PDT) is ideally suited for the treatment of head and neck cancer (HNC) in humans. Developing useful PDT protocols for HNC is challenging due to the expense of Phase I and II clinical trials. Moreover, the often-poor predictive value of murine models means that photosensitizers may proceed far into development before problems are noted. Dogs and cats with spontaneous oral squamous cell carcinoma (SCC) share striking similarities with humans affected with oral SCC. These similarities include viral and environmental tobacco smoke as risk factors, location-dependent prognoses, and relative resistance to chemotherapy. The relatively large oral cancers encountered in veterinary patients allow for light and drug dosimetry that are directly applicable to humans. The irregular shape of oral SCC allows a rigorous evaluation of novel photodynamic therapy protocols under field conditions. Because spontaneous tumors in dogs and cats arise in an outbred animal population it is possible to observe an intact host response to PDT. The shorter lifespan of dogs and cats allows rapid accrual of endpoint data. External beam radiation therapy and chemotherapy are commonplace in veterinary medicine, making dogs and cats with spontaneous SCC a useful resource to study the interactions with PDT and other cancer treatment modalities. Our preliminary results demonstrate that PDT is well-tolerated by dogs with oral cancer, and a Phase II clinical trial of zinc-phthalocyanine-based photodynamic therapy is underway in dogs with oral SCC. The usefulness of 5-aminolevulinic acid methyl ester-based PDT is being investigated in cats with oral SCC.

Two major carotenoids species found in salmonids muscle tissues are astaxanthin and cantaxanthin. They are taken up from fish food and are responsible for the attractive red-orange color of salmon filet. Since carotenoids are powerful antioxidants and biomarkers of nutrient consumption, they are thought to indicate fish health and resistance to diseases in fish farm environments. Therefore, a rapid, accurate, quantitative optical technique for measuring carotenoid content in salmon tissues is of economic interest. We demonstrate the possibility of using fast, selective, quantitative detection of astaxanthin and cantaxanthin in salmon muscle tissues, employing resonance Raman spectroscopy. Analyzing strong Raman signals originating from the carbon-carbon double bond stretch vibrations of the carotenoid molecules under blue laser excitation, we are able to characterize quantitatively the concentrations of carotenoids in salmon muscle tissue. To validate the technique, we compared Raman data with absorption measurements of carotenoid extracts in acetone. A close correspondence was observed in absorption spectra for tissue extract in acetone and a pure astaxanthin solution. Raman results show a linear dependence between Raman and absorption data. The proposed technique holds promise as a method of rapid screening of carotenoid levels in fish muscle tissues and may be attractive for the fish farm industry to assess the dietary status of salmon, risk for infective diseases, and product quality control.

Introduction: This study evaluated the ability to fragment and remove naturally occurring uroliths in dogs using a holmium: YAG laser. Methods: Twenty four dogs with naturally occurring uroliths including 10 spayed females and 14 neutered males. The dogs were 8.7 ± 2.8 years old and weighed 13.7 ± 8.0 kg. All dogs had bladder stones and 5 male dogs also had urethral stones. In female dogs, cystoscopy was performed using a rigid cystoscope with sheath diameter of 14 to 19 french. Cystoscopy was performed in males dogs using a 7.5 french diameter pediatric ureteroscope. Uroliths were fragmented using a 20 watt Holmium: YAG laser and the fragments were removed by basket extraction and voiding urohydropropulsion. Results: Average laser parameters for urolith fragmentation were 0.7 Joules at 8 Hertz (range: 0.5 to 1.3 Joules at 5 to 13 Hertz). All urolith fragments were successfully removed in all 10 female dogs and 11 of 14 male dogs. In one male dog, the urethra was too small to allow passage of the ureteroscope. In one of the male dogs, the urethral stones were successfully removed by laser lithotripsy, but removal of the bladder stones was performed by cystotomy. There was one complication of urethral perforation during attempts to pass an access sheath transurethrally in a dog with extensive proliferative urethritis. Conclusions: Laser lithotripsy is a safe and effective method of removing bladder and urethral stones in dogs provided the dog is large enough to permit transurethral passage of a cystoscope or ureteroscope.

The use of lasers by both the military and civilian community is rapidly expanding. Thus, the potential for and severity of laser eye injury and retinal damage is increasing. Sensitive and accurate methods to evaluate and follow laser retinal damage are needed. The multifocal electroretinogram (mfERG) has the potential to meet these criteria. In this study, the mfERG was used to evaluate changes to retinal function following laser exposure. Landolt C contrast acuity was also measured in the six behaviorally trained Rhesus monkeys. The monkeys then received Nd:YAG laser lesions (1064 nm, 9 ns pulse width) in each eye. One eye received a single foveal lesion of approximately 0.13 mJ total intraocular exposure (TIE) and the other received six parafoveal lesions which varied in TIE from 0.13 to 4 mJ. mfERGs and behavioral data were collected both pre- and post-exposure. mfERGs were recorded using stimuli that contained 103, 241, and 509 hexagons. Landolt C contrast acuity was measured with five sizes of Landolt C (0.33 to 11.15 cycles/degree) of varying contrast. mfERG response densities were sensitive to the functional retinal changes caused by the laser insult. In general, larger lesions showed greater mfERG abnormalities than smaller laser lesions. Deficits in contrast acuity were found to be more severe in the eyes with foveal injuries. Although the mfERG and contrast acuity assess different areas of the visual system, both are sensitive to laser-induced retinal damage and may be complementary tests for laser eye injury triage.

Approximately 12,000 people are diagnosed with invasive transitional cell carcinoma of the urinary bladder (InvTCC) each year in the United States. Surgical removal of the bladder (cystectomy) and regional lymph node dissection are considered frontline therapy. Cystectomy causes extensive acute morbidity, and 50% of patients with InvTCC have occult metastases at the time of diagnosis. Better staging procedures for InvTCC are greatly needed. This study was performed to evaluate an intra-operative near infrared fluorescence imaging (NIRF) system (Frangioni laboratory) for identifying sentinel lymph nodes draining InvTCC. NIRF imaging was used to map lymph node drainage from specific quadrants of the urinary bladder in normal dogs and pigs, and to map lymph node drainage from naturally-occurring InvTCC in pet dogs where the disease closely mimics the human condition. Briefly, during surgery NIR fluorophores (human serum albumen-fluorophore complex, or quantum dots) were injected directly into the bladder wall, and fluorescence observed in lymphatics and regional nodes. Conditions studied to optimize the procedure including: type of fluorophore, depth of injection, volume of fluorophore injected, and degree of bladder distention at the time of injection. Optimal imaging occurred with very superficial injection of the fluorophore in the serosal surface of the moderately distended bladder. Considerable variability was noted from dog to dog in the pattern of lymph node drainage. NIR fluorescence was noted in lymph nodes with metastases in dogs with InvTCC. In conclusion, intra-operative NIRF imaging is a promising approach to improve sentinel lymph node mapping in invasive urinary bladder cancer.

Transendoscopic laser surgery has been performed in horses since 1984. It is used to treat many upper respiratory disorders, as well as urogenital diseases. Initially, the Nd:YAG laser was the laser of choice until the early 1990s, when smaller, more compact diode lasers entered the veterinary field. In the mid 1980s, several attempts were made to transmit CO₂ laser energy transendoscopically. True success was not obtained until 2004 when the OmniGuide CO₂ Fiber was fabricated. Although there is attenuation of energy, this very flexible fiber allows the CO₂ laser to be used transendoscopically for incision and ablation of tissue. Intermittent dorsal displacement of the soft palate has more recently been treated using a diode laser and contact fiber to scarify the caudal border of the soft palate. This procedure was initially reported as being performed in combination with a myectomy. The CO₂ laser's fiber was used in eight cases. It offered no touch technique and allowed improved visualization of the target tissue. Both healing and recuperation time were reduced, compared to other wavelengths transmitted through solid quartz fiber. The OmniGuide Fiber can be coupled to the output port of CO₂ lasers commonly used in veterinary medicine. Transendoscopic application of the CO₂ laser is advantageous in that there is no endoscopic white-out, no volume heating of tissue, and it provides an accurate means of performing upper respiratory surgery in the standing horse.