Narasimhan Rajaram

Lookup table–based inverse model for determining optical properties of turbid media

We present a lookup table (LUT)-based inverse model for determining the optical properties of turbid media from steady-state diffuse reflectance spectra that is valid for fiber-based probe geometries with close source-detector separations and tissue with low albedo. The lookup table is based solely on experimental measurements of calibration standards. We used tissue-simulating phantoms to validate the accuracy of the LUT inverse model. Our results show excellent agreement between the expected and extracted values of the optical parameters. In addition, the LUT represents a significant improvement in accuracy at short source-detector separations (300 microm) and low albedo (approximately 0.35). We also present in vivo data from clinically normal and malignant nonmelanoma skin cancers fit to the LUT-based model.

Pilot clinical study for quantitative spectral diagnosis of non-melanoma skin cancer

Several research groups have demonstrated the non‐invasive diagnostic potential of diffuse optical spectroscopy (DOS) and laser‐induced fluorescence (LIF) techniques for early cancer detection. By combining both modalities, one can simultaneously measure quantitative parameters related to the morphology, function and biochemical composition of tissue and use them to diagnose malignancy. The objective of this study was to use a quantitative reflectance/fluorescence spectroscopic technique to determine the optical properties of normal skin and non‐melanoma skin cancers and the ability to accurately classify them. An additional goal was to determine the ability of the technique to differentiate non‐melanoma skin cancers from normal skin.

In Vivo Detection of Gold Nanoshells in Tumors Using Diffuse Optical Spectroscopy

This study demonstrates the use of diffuse optical spectroscopy (DOS) for the noninvasive measurement of gold nanoshell concentrations in tumors of live mice. We measured the diffuse optical spectra (500-800 nm) using an optical fiber probe placed in contact with the tissue surface. We performed in vitro studies on tissue phantoms illustrating an accurate measurement of gold-silica nanoshell concentration within 12.6% of the known concentration. In vivo studies were performed on a mouse xenograft tumor model. DOS spectra were measured at preinjection, immediately postinjection, 1 and 24 h postinjection times, and the nanoshell concentrations were verified using neutron activation analysis.

Probe pressure effects on human skin diffuse reflectance and fluorescence spectroscopy measurements

Diffuse reflectance and fluorescence spectroscopy are popular research techniques for noninvasive disease diagnostics. Most systems include an optical fiber probe that transmits and collects optical spectra in contact with the suspected lesion. The purpose of this study is to investigate probe pressure effects on human skin spectroscopic measurements. We conduct an in-vivo experiment on human skin tissue to study the short-term (<2 s) and long-term (>30 s) effects of probe pressure on diffuse reflectance and fluorescence measurements. Short-term light probe pressure (P0<9 mN∕mm2) effects are within 0 ± 10% on all physiological properties extracted from diffuse reflectance and fluorescence measurements, and less than 0±5% for diagnostically significant physiological properties. Absorption decreases with site-specific variations due to blood being compressed out of the sampled volume. Reduced scattering coefficient variation is site specific. Intrinsic fluorescence shows a large standard error, although no specific pressure-related trend is observed. Differences in tissue structure and morphology contribute to site-specific probe pressure effects. Therefore, the effects of pressure can be minimized when the pressure is small and applied for a short amount of time; however, long-term and large pressures induce significant distortions in measured spectra.

Design and validation of a clinical instrument for spectral diagnosis of cutaneous malignancy

We report a probe-based portable and clinically compatible instrument for the spectral diagnosis of melanoma and nonmelanoma skin cancers. The instrument combines two modalities--diffuse reflectance and intrinsic fluorescence spectroscopy--to provide complementary information regarding tissue morphology, function, and biochemical composition. The instrument provides a good signal-to-noise ratio for the collected reflectance and laser-induced fluorescence spectra. Validation experiments on tissue phantoms over a physiologically relevant range of albedos (0.35-0.99) demonstrate an accuracy of close to 10% in determining scattering, absorption and fluorescence characteristics. We also demonstrate the ability of our instrument to collect in vivo diffuse reflectance and fluorescence measurements from clinically normal skin, dysplastic nevus, and malignant nonmelanoma skin cancer.

Clinical study of noninvasive in vivo melanoma and nonmelanoma skin cancers using multimodal spectral diagnosis.

Abstract. The goal of this study was to determine the diagnostic capability of a multimodal spectral diagnosis (SD) for in vivo noninvasive disease diagnosis of melanoma and nonmelanoma skin cancers. We acquired reflectance, fluorescence, and Raman spectra from 137 lesions in 76 patients using custom-built optical fiber-based clinical systems. Biopsies of lesions were classified using standard histopathology as malignant melanoma (MM), nonmelanoma pigmented lesion (PL), basal cell carcinoma (BCC), actinic keratosis (AK), and squamous cell carcinoma (SCC). Spectral data were analyzed using principal component analysis. Using multiple diagnostically relevant principal components, we built leave-one-out logistic regression classifiers. Classification results were compared with histopathology of the lesion. Sensitivity/specificity for classifying MM versus PL (12 versus 17 lesions) was 100%/100%, for SCC and BCC versus AK (57 versus 14 lesions) was 95%/71%, and for AK and SCC and BCC versus normal skin (71 versus 71 lesions) was 90%/85%. The best classification for nonmelanoma skin cancers required multiple modalities; however, the best melanoma classification occurred with Raman spectroscopy alone. The high diagnostic accuracy for classifying both melanoma and nonmelanoma skin cancer lesions demonstrates the potential for SD as a clinical diagnostic device.

Experimental validation of the effects of microvasculature pigment packaging on in vivo diffuse reflectance spectroscopy.

Diffuse reflectance spectroscopy (DRS) uses the steady‐state diffuse reflectance measured from the tissue surface to determine absorption and scattering properties of sampled tissue. Many inverse models used to determine absorber properties have assumed a homogeneous distribution of blood. However, blood in tissue is confined to blood vessels that occupy a small fraction of the overall volume. This simplified assumption can lead to large errors when measuring optical properties. The objective of this study was to examine the effect of confining absorbers to small volumes, such as the microvasculature, on in vivo DRS.

Performance of a lookup table-based approach for measuring tissue optical properties with diffuse optical spectroscopy

Diffuse optical spectroscopy (DOS) provides a powerful tool for fast and noninvasive disease diagnosis. The ability to leverage DOS to accurately quantify tissue optical parameters hinges on the model used to estimate light-tissue interaction. We describe the accuracy of a lookup table (LUT)-based inverse model for measuring optical properties under different conditions relevant to biological tissue. The LUT is a matrix of reflectance values acquired experimentally from calibration standards of varying scattering and absorption properties. Because it is based on experimental values, the LUT inherently accounts for system response and probe geometry. We tested our approach in tissue phantoms containing multiple absorbers, different sizes of scatterers, and varying oxygen saturation of hemoglobin. The LUT-based model was able to extract scattering and absorption properties under most conditions with errors of less than 5 percent. We demonstrate the validity of the lookup table over a range of source-detector separations from 0.25 to 1.48 mm. Finally, we describe the rapid fabrication of a lookup table using only six calibration standards. This optimized LUT was able to extract scattering and absorption properties with average RMS errors of 2.5 and 4 percent, respectively.

Rapid and accurate determination of tissue optical properties using least-squares support vector machines

Diffuse reflectance spectroscopy (DRS) has been extensively applied for the characterization of biological tissue, especially for dysplasia and cancer detection, by determination of the tissue optical properties. A major challenge in performing routine clinical diagnosis lies in the extraction of the relevant parameters, especially at high absorption levels typically observed in cancerous tissue. Here, we present a new least-squares support vector machine (LS-SVM) based regression algorithm for rapid and accurate determination of the absorption and scattering properties. Using physical tissue models, we demonstrate that the proposed method can be implemented more than two orders of magnitude faster than the state-of-the-art approaches while providing better prediction accuracy. Our results show that the proposed regression method has great potential for clinical applications including in tissue scanners for cancer margin assessment, where rapid quantification of optical properties is critical to the performance.

Radiation induces aerobic glycolysis through reactive oxygen species

BACKGROUND AND PURPOSE Although radiation induced reoxygenation has been thought to increase radiosensitivity, we have shown that its associated oxidative stress can have radioprotective effects, including stabilization of the transcription factor hypoxia inducible factor 1 (HIF-1). HIF-1 is known to regulate many of the glycolytic enzymes, thereby promoting aerobic glycolysis, which is known to promote treatment resistance. Thus, we hypothesized that reoxygenation after radiation would increase glycolysis. We previously showed that blockade of oxidative stress using a superoxide dismutase (SOD) mimic during reoxygenation can downregulate HIF-1 activity. Here we tested whether concurrent use of this drug with radiotherapy would reduce the switch to a glycolytic phenotype. MATERIALS AND METHODS 40 mice with skin fold window chambers implanted with 4T1 mammary carcinomas were randomized into (1) no treatment, (2) radiation alone, (3) SOD mimic alone, and (4) SOD mimic with concurrent radiation. All mice were imaged on the ninth day following tumor implantation (30 h following radiation treatment) following injection of a fluorescent glucose analog, 2-[N-(7-nitrobenz-2-oxa-1,3-diaxol-4-yl)amino]-2-deoxyglucose (2-NBDG). Hemoglobin saturation was measured by using hyperspectral imaging to quantify oxygenation state. RESULTS Mice treated with radiation showed significantly higher 2-NBDG fluorescence compared to controls (p=0.007). Hemoglobin saturation analysis demonstrated reoxygenation following radiation, coinciding with the observed increase in glycolysis. The concurrent use of the SOD mimic with radiation demonstrated a significant reduction in 2-NBDG fluorescence compared to effects seen after radiation alone, while having no effect on reoxygenation. CONCLUSIONS Radiation induces an increase in tumor glucose demand approximately 30 h following therapy during reoxygenation. The use of an SOD mimic can prevent the increase in aerobic glycolysis when used concurrently with radiation, without preventing reoxygenation.