In this paper we present images derived from measurements of diffuse photon density waves in turbid media. After a brief introduction and description of the experimental set-up, this document is divided into two sections. In the first, we present an analytic solution for the scattering of diffuse photon density waves by spherical and cylindrical inhomogeneities within turbid media and discuss the application of this solution to systems containing multiple objects and multiple sources. This exact solution is a useful tool for simulating the distortion of a diffuse photon density wave due to simple inhomogeneities. Until recently, simulations of heterogeneous media were achieved by Monte-Carlo methods, finite element analysis, or other time consuming techniques. More recently, Feng et al. [1] presented a perturbative solution for a single, point-like heterogeneity. The exact calculation we present is not limited to small objects, and numerical calculations using this solution are fast compared to Monte Carlo simulations[2]. Finally, we experimentally demonstrate that this analytic solution enables us to fit for the optical and geometric properties of arbitrary spherical and cylindrical objects, and we discuss the detector sensitivity required to characterize such objects. In the second section we derive images of more general inhomogeneous scattering media. Our method is based on the diffusion approximation, but is not an iterative approach; it is a direct inversion that is exact to first order in variations of the sample absorption. An increase in sensitivity is achieved through the use of multiple-source/single-detector units placed around the turbid medium. In our particular case, each unit consisted of a set of two modulated diffuse light sources phased 180° out of phase with respect to one another, and a single detector located at positions equidistant from each source. In this report we will describe the general theoretical approach, provide experimental evidence of its utility, and discuss potential applications.

In anaerobic, uncoupled pigeon-heart mitochondria treated with oxidizable substrate, the cyto- chrome b566 remains largely oxidized. In the presence of antimycin A, addition of oxygen induces a reduction of this cytochrome. The rate of cytochrome b566 reduction is comparable to and dependent on the rate of cytochrome ci oxidation. Kinetic data suggest that either ubiquinone or another donor of similar potential provides electrons for the reduction of cytochrome b566. It is postulated that the aerobic reduction of cytochrome b566 is directly related to the energy conservation at site II.

This investigation aimed to test all tumor-bearing patients who undergo biopsy to see if angiogenesis and hypoxia can detect cancer. We used continuous-wave near-infrared spectroscopy (NIRS) to measure blood hemoglobin concentration to obtain blood volume or total hemoglobin [Hbtot] and oxygen saturation for the angiogenesis and hypoxic biomarkers. The contralateral breast was used as a reference to derive the difference from breast tumor as a difference in total hemoglobin Δ[HBtot] and a difference in deoxygenation Δ([Hb]-[HbO2]). A total of 91 invasive cancers, 26 DCIS, 45 fibroblastomas, 96 benign tumors excluding cysts, and 67 normal breasts were examined from four hospitals. In larger-size tumors, there is significantly higher deoxygenation in invasive and ductal carcinoma in situ (DCIS) than in that of benign tumors, but no significant difference was seen in smaller tumors of ≤ 1 cm. With the two parameters of high total hemoglobin and hypoxia score, the sensitivity and specificity of cancer detection were 60.3 % and 85.3 %, respectively. In summary, smaller-size tumors are difficult to detect with NIRS, whereas DCIS can be detected by the same total hemoglobin and hypoxic score in our study.

Currently, the gold standard to establish benign vs. malignant breast tissue diagnosis requires an invasive biopsy followed by tissue fixation for subsequent histopathological examination. This process takes at least 24 h resulting in tissues that are less suitable for molecular, functional, or metabolic analysis. We have recently conducted redox scanning (cryogenic NADH/flavoprotein fluorescence imaging) on snap-frozen breast tissue biopsy samples obtained from human breast cancer patients at the time of their breast cancer surgery. The redox state was readily determined by the redox scanner at liquid nitrogen temperature with extraordinary sensitivity, giving oxidized flavoproteins (Fp) an up to tenfold discrimination of cancer to non-cancer of breast in our preliminary data. Our finding suggests that the identified metabolic parameters could discriminate between cancer and non-cancer breast tissues without subjecting tissues to fixatives. The remainder of the frozen tissue is available for additional analysis such as molecular analysis and conventional histopathology. We propose that this novel redox scanning procedure may assist in tissue diagnosis in ex vivo tissues.

Intratumor heterogeneity is of great research interest for understanding cancer pathology and developing diagnostic and therapeutic methods. The low-temperature 3D NADH/Fp (oxidized flavoproteins including FAD) fluorescence imaging or the redox scanning provides an effective tool for imaging intratumor heterogeneity in mitochondrial redox state at a high spatial resolution (down to 50 × 50 × 20 μm3). Previously, we have shown mitochondrial redox state and its heterogeneity in tumor tissue provide sensitive and potentially diagnosis-useful characteristics for differentiating among five human melanoma and two breast cancer mouse xenografts of different metastatic potential. Here, we report the preliminary results of imaging the in vivo mitochondrial redox state of the entire tumor for three human breast cancer lines having ascending order of aggressiveness, i.e., MCF-7 < MDA-MB-468 < MDA-MB-231 xenografted in athymic nude mice. The tumor-bearing mice were anesthetized and snap-frozen in liquid N2 so that the in vivo mitochondrial redox state was maintained for ex vivo redox scanning. The entire excised tumors were scanned section by section at different depths with 400 μm spacing, total of 11–14 sections per tumor. We obtained the 3D distribution of nominal concentrations of oxidized flavoproteins (Fp) and NADH in tissue and calculated the Fp redox ratio Fp/(Fp + NADH) representing the mitochondrial redox state. The results revealed that both aggressive tumors (MDA-MB-231 and MDA-MB-468) displayed heterogeneity in the distributions of NADH, Fp and Fp redox ratio, with a localized area exhibiting significantly higher Fp redox ratio than other regions; the indolent MCF-7 tumor displayed a relatively uniform distribution in both Fp and NADH, thus Fp redox ratio. The results suggest possible novel imaging biomarkers on the basis of NADH, Fp and Fp redox ratio images to differentiate among these tumors. Potentially, these biomarkers may be useful for cancer diagnosis and therapy.

Tumor stratification on the basis of metabolic phenotypes may provide useful information for cancer diagnosis and treatment. Higher than normal glucose uptake/metabolism by cancer cells is an established hallmark for cancer staging by fluorinated deoxyglucose positron emission tomography (FDG-PET). High-resolution (down to 50 × 50 × 20 μm3) biomarkers of mitochondrial redox state provided by the redox scanning or the low-temperature 3-D NADH/Fp (reduced nicotinamide adenine dinucleotide/oxidized flavoproteins including FAD, i.e., flavin adenine dinucleotide) fluorescence imaging can differentiate human melanoma and breast cancer mouse xenografts of different metastatic potentials. In this project, by injecting into breast tumor bearing mice a near-infrared fluorescent glucose-analogue Pyro-2DG (pyropheophorbide 2-deoxyglucosamide), we were able to simultaneously image both the redox state and glucose uptake by mouse xenografts of human breast cancer MDA-MB-231 with an in-plane resolution of 200 μm. The preliminary results showed heterogeneity in the distribution of both Pyro-2DG uptake and the mitochondrial redox state, with Pyro-2DG uptake tending to correlate more with NADH distribution in tumor rim and also exhibiting pronounced uptake in localized areas of the tumor core. The co-registered images of mitochondrial redox state and glucose uptake may provide relevant information for understanding tumor metabolism and its role in cancer progression.

Abstract Metabolism of mitochondria can be monitored by measuring thermogenic activity. A linear increase of heating rate vs. rate of oxygen utilization for NADH oxidation has been shown [Poe, Arch. Biochem. Biophy., 1967]. Moreover, the heat generation increases four-fold in the uncoupled apoptotic state compared to resting state of mitochondria, as a successful anticancer drug opens the ion channel pore or increase membrane permeability [Estabrook, 1967, Johnstone, Cell 2002]. During apoptosis, more numerous and smaller mitochondria are yielded, which might also explain increase of heat generation. In this study we explore these issues: Diffuse Optical Spectroscopy Imaging (DOSI) monitors absolute temperature of a breast with infiltrating ductal carcinoma (IDC), as well as oxy- and deoxy-hemoglobin concentration. DOSI employs low power near-infrared light to quantify tissues. The diffusion model affords quantitative determination of tissue chromophore absorption spectra by separating scattering and absorption effects. Especially, the water peak appear at 935-1000nm provides information about tissue water concentration, state and absolute temperature as shown by Chung et al [PMB 2008, 2010]. In this case study, we measured a 63 year old subject who had IDC in both breasts. She received neoadjuvant chemotherapy with two different regimens: Adriamycin+Cytoxan (AC, one and half months) and Carboplatin+Abraxane+Avastin (CAA, 3 months). The patient was measured 19 times along the course of therapy: at pre-treatment, 8 points during AC, 9 points during CAA, and after completion of the therapy. She was a pathological complete responder. All points on a spectroscopic image of the lesion breast were used for the average and standard deviation of each quantity at each time point. After some fluctuations in the early stage, the temperature was observed to increase up to 39.5±1.2°C by the last phase of AC dose, showing 4.6% increase/day. During CAA, the temperature decreased eventually at a rate of -1%/day. After completion of the therapy, the temperature changed only about -0.6% to 36.9±0.7°C. In two time windows just before the end of AC and in the middle of CAA, oxy-hemoglobin increased with the temperature. The four fold increase of temperature during AC suggests the uncoupling of mitochondria due to apoptosis, an effect which appears as both an increase of temperature and deoxy-hemoglobin concentration. Then, possible massive cell death is indicated by the decrease of temperature. During AC therapy, possible aerobic glycolysis followed metabolic increase due to apoptosis, and then massive cell death followed. Overall, although preliminary, this study suggests the potential of DOSI measured temperature and other quantitative physiological components as a non-invasive and longitudinal monitor of apoptosis process during neoadjuvant chemotherapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4147. doi:10.1158/1538-7445.AM2011-4147

Redox state mediates embryonic stem cell (ESC) differentiation and thus offers an important complementary approach to understanding the pluripotency of stem cells. NADH redox ratio (NADH/(Fp + NADH)), where NADH is the reduced form of nicotinamide adenine dinucleotide and Fp is the oxidized flavoproteins, has been established as a sensitive indicator of mitochondrial redox state. In this paper, we report our redox imaging data on the mitochondrial redox state of mouse ESC (mESC) colonies and the implications thereof. The low-temperature NADH/Fp redox scanner was employed to image mESC colonies grown on a feeder layer of gamma-irradiated mouse embryonic fibroblasts (MEFs) on glass cover slips. The result showed significant heterogeneity in the mitochondrial redox state within individual mESC colonies (size: ~200–440 μm), exhibiting a core with a more reduced state than the periphery. This more reduced state positively correlates with the expression pattern of Oct4, a well-established marker of pluripotency. Our observation is the first to show the heterogeneity in the mitochondrial redox state within a mESC colony, suggesting that mitochondrial redox state should be further investigated as a potential new biomarker for the stemness of embryonic stem cells.

Near-infrared spectroscopy (NIRS) has been shown to be one of the tools that can measure oxygenation in muscle and other tissues in vivo. This review paper highlights the progress, specifically in this decade, that has been made for evaluating skeletal muscle oxygenation and oxidative energy metabolism in sport, health and clinical sciences. Development of NIRS technologies has focused on improving quantification of the signal using multiple wavelengths to solve for absorption and scattering coefficients, multiple pathlengths to correct for the influence of superficial skin and fat, and time-resolved and phase-modulated light sources to determine optical pathlengths. In addition, advances in optical imaging with multiple source and detector pairs as well as portability using small wireless detectors have expanded the usefulness of the devices. NIRS measurements have provided information on oxidative metabolism in various athletes during localized exercise and whole-body exercise, as well as training-induced adaptations. Furthermore, NIRS technology has been used in the study of a number of chronic health conditions. Future developments of NIRS technology will include enhancing signal quantification. In addition, advances in NIRS imaging and portability promise to transform how measurements of oxygen utilization are obtained in the future.

AIM To evaluate the novel nanoparticle reconstituted bacteriochlorin e6 bisoleate low-density lipoprotein (r-Bchl-BOA-LDL) for its efficacy as a photodynamic therapy agent delivery system in xenografts of human hepatoblastoma G2 (HepG2) tumors. MATERIALS & METHODS Bchl-BOA was encapsulated in the nanoparticle low-density lipoprotein (LDL), a native particle whose receptor's overexpression is a cancer signature for a number of neoplasms. Evaluation of r-Bchl-BOA-LDL as a potential photosensitizer was performed using a tumor response and foot response assay. RESULTS & DISCUSSION When compared with controls, tumor regrowth was significantly delayed at injected murine doses of 2 µmole/kg r-Bchl-BOA-LDL after illumination at fluences of 125, 150 or 175 J/cm(2). Foot response assays showed that although normal tissue toxicity accompanied the higher fluences it was significantly reduced at the lowest fluence tested. CONCLUSION This research demonstrates that r-Bchl-BOA-LDL is an effective photosensitizer and a promising candidate for further investigation.

Near-infrared spectroscopy (NIRS) studies have revealed that performing mental arithmetic tasks have associated event-related hemodynamic responses that are detectable. Thus NIRS-based Brain Computer Interface (BCI) has the potential for investigating how to best teach mathematics in a classroom setting. This paper presents a novel computational intelligent method of applying rough set-based neuro-fuzzy system (RNFS) in NIRS-based BCI for assessing numerical cognition. A study is performed on 20 healthy subjects to measure 32 channels of hemoglobin responses in performing three difficulty levels of mental arithmetic. The accuracy is then presented using 5×5-fold cross-validations on the data collected. The results of applying RNFS and its Mutual Information-based Rough Set Reduction (MIRSR) for feature selection is then compared against the Naïve Bayesian Parzen Window classifier and other MI-based feature selection algorithms. The results of applying RNFS yielded significantly better accuracy of 75.7% compared to the other methods, thus demonstrating the potential of RNFS in NIRS-based BCI for assessing numerical cognition.

The dietary supplement co-enzyme Q10 (CoQ10) is a potent antioxidant and a component of the electron transport chain which has been show to improve muscle metabolism and blood flow. PURPOSE: The purpose of this study was to determine the impact of CoQ10 supplementation on oxidative stress, muscle metabolism and muscle blood flow in older adults. METHODS: Fourteen older individuals (68.5±7.0 years) were tested 2 weeks post placebo (week 2) and 2 weeks post CoQ10 supplementation (week 4). During each testing session the participants performed a 12 s dynamic plantar flexion exercise during which medial gastrocnemius muscle PCr and pH were measured using 31P magnetic resonance spectroscopy. Muscle oxygen saturation and blood flow were simultaneously measured using near infrared spectroscopy. Post exercise venous blood samples were drawn to determine malondialdehyde level as an oxidative stress indicator. The placebo and CoQ10 dosage was 200 mg/day. Paired t-test was used to determine differences between trials. RESULTS: After 2 weeks of CoQ10 supplementation, no differences were observed in resting PCr (40.0±1.4 vs. 39.9±2.2 mM), or intracellular muscle pH (7.03±0.02 vs. 7.01±0.02). However, trends for improvements in metabolic, hemodynamic and oxidative stress status were observed (PCr recovery rate after exercise: 26.4±10.6 vs. 27.5±11.9 mM/kg/min; NIRT50: 55.8±43.1 vs. 50.7±30.6 s; post exercise malondialdehyde level: 1.7±0.8 vs. 1.4±0.4 uM/l). CONCLUSIONS: Our results show that there may be alterations in oxidative stress level as well as muscle metabolic and hemodynamic status with CoQ10 supplementation. Two weeks of CoQ10 supplementation at 200 mg/day, however, did not significantly impact these oxidative stress status, metabolic or hemodynamic measures. Based on the trends that were observed, higher doses or a longer supplementation period may be advisable for future research. (Support: Diabetes Action Research and Education Foundation)

Predicting tumor metastatic potential remains a challenge in cancer research and clinical practice. Our goal was to identify novel biomarkers for differentiating human breast tumors with different metastatic potentials by imaging the in vivo mitochondrial redox states of tumor tissues. The more metastatic (aggressive) MDA-MB-231 and less metastatic (indolent) MCF-7 human breast cancer mouse xenografts were imaged with the low-temperature redox scanner to obtain multi-slice fluorescence images of reduced nicotinamide adenine dinucleotide (NADH) and oxidized flavoproteins (Fp). The nominal concentrations of NADH and Fp in tissue were measured using reference standards and used to calculate the Fp redox ratio, Fp(NADH+Fp). We observed significant core-rim differences, with the core being more oxidized than the rim in all aggressive tumors but not in the indolent tumors. These results are consistent with our previous observations on human melanoma mouse xenografts, indicating that mitochondrial redox imaging potentially provides sensitive markers for distinguishing aggressive from indolent breast tumor xenografts. Mitochondrial redox imaging can be clinically implemented utilizing cryogenic biopsy specimens and is useful for drug development and for clinical diagnosis of breast cancer.