Optimal imaging windows of indocyanine green‐assisted near‐infrared dental imaging with rat model and its comparison to X‐ray imaging

In this study, we used rat animal model to compare the efficiency of indocyanine green (ICG)‐assisted dental near‐infrared fluorescence imaging with X‐ray imaging, and we optimized the imaging window for both unerupted and erupted molars. The results show that the morphology of the dental structures was observed clearly from ICG‐assisted dental images (especially through the endoscope). A better image contrast was easily acquired at the short imaging windows (<10 minutes) for unerupted and erupted molars. For unerupted molars, there is another optimized imaging window (48‐96 hours) with a prominent glow‐in‐the‐dark effect: only the molars remain bright. This study also revealed that the laser ablation of dental follicles can disrupt the molar development, and our method is able to efficiently detect laser‐treated molars and acquire the precise morphology. Thus, ICG‐assisted dental imaging has the potential to be a safer and more efficient imaging modality for the real‐time diagnosis of dental diseases.     

Computed tomography for in vivo deep over‐1000 nm near‐infrared fluorescence imaging

This study aims to develop a novel cross‐sectional imaging of fluorescence in over‐1000 nm near‐infrared (OTN‐NIR), which allows in vivo deep imaging, using computed tomography (CT) system. Cylindrical specimens of composite of OTN‐NIR fluorophore, NaGdF₄ co‐doped with Yb³⁺ and Ho³⁺ (ex: 980 nm, em: 1150 nm), were embedded in cubic agar (10.5–12 mm) or in the peritoneal cavity of mice and placed on a rotatable stage. When the fluorescence from inside of the samples was serially captured from multiple angles, the images were disrupted by the reflection and refraction of emitted light on the sample‐air interface. Immersing the sample into water filled in a rectangular bath suppressed the disruption at the interface and successfully reconstructed the position and concentration of OTN‐NIR fluorophores on the cross‐sectional images using a CT technique. This is promising as a novel three‐dimensional imaging technique for OTN‐NIR fluorescent image projections of small animals captured from multiple angles.     

Bond‐selective imaging of deep tissue through the optical window between 1600 and 1850 nm

We report the employment of an optical window between 1600 nm and 1850 nm for bond‐selective deep tissue imaging through harmonic vibrational excitation and acoustic detection of resultant pressure waves. In this window where a local minimum of water absorption resides, we found a 5 times enhancement of photoacoustic signal by first overtone excitation of the methylene group CH₂ at 1730 nm, compared to the second overtone excitation at 1210 nm. The enhancement allows 3D mapping of intramuscular fat with improved contrast and of lipid deposition inside an atherosclerotic artery wall in the presence of blood. Moreover, lipid and protein are differentiated based on the first overtone absorption profiles of CH₂ and methyl group CH₃ in this window. (© 2012 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Spectral correction for handheld optoacoustic imaging by means of near‐infrared optical tomography in reflection mode

In vivo imaging of tissue/vasculature oxygen saturation levels is of prime interest in many clinical applications. To this end, the feasibility of combining two distinct and complementary imaging modalities is investigated: optoacoustics (OA) and near‐infrared optical tomography (NIROT), both operating noninvasively in reflection mode. Experiments were conducted on two optically heterogeneous phantoms mimicking tissue before and after the occurrence of a perturbation. OA imaging was used to resolve submillimetric vessel‐like optical absorbers at depths up to 25 mm, but with a spectral distortion in the OA signals. NIROT measurements were utilized to image perturbations in the background and to estimate the light fluence inside the phantoms at the wavelength pair (760 nm, 830 nm). This enabled the spectral correction of the vessel‐like absorbers' OA signals: the error in the ratio of the absorption coefficient at 830 nm to that at 760 nm was reduced from 60%‐150% to 10%‐20%. The results suggest that oxygen saturation (SO ₂) levels in arteries can be determined with <10% error and furthermore, that relative changes in vessels' SO ₂ can be monitored with even better accuracy. The outcome relies on a proper identification of the OA signals emanating from the studied vessels.      

Phase dual‐slopes in frequency‐domain near‐infrared spectroscopy for enhanced sensitivity to brain tissue: First applications to human subjects

We present a first in vivo application of phase dual‐slopes (DS?), measured with frequency‐domain near‐infrared spectroscopy on four healthy human subjects, to demonstrate their enhanced sensitivity to cerebral hemodynamics. During arterial blood pressure oscillations elicited at a frequency of 0.1 Hz, we compare three different ways to analyze either intensity (I) or phase (?) data collected on the subject's forehead at multiple source‐detector distances: Single‐distance, single‐slope and DS. Theoretical calculations based on diffusion theory show that the method with the deepest maximal sensitivity (at about 11 mm) is DS?. The in vivo results indicate a qualitative difference of phase data (especially DS?) and intensity data (especially single‐distance intensity [SDI]), which we assign to stronger contributions from scalp hemodynamics to SDI and from cortical hemodynamics to DS?. Our findings suggest that scalp hemodynamic oscillations may be dominated by blood volume dynamics, whereas cortical hemodynamics may be dominated by blood flow velocity dynamics.     

The utility of far‐infrared illumination in oxygenation dynamics as measured with near‐infrared spectroscopy

Near‐infrared spectroscopy (NIRS) is a noninvasive method for measuring the oxygenation in muscle and other tissues in vivo. For quantitative NIRS measurement of oxygenation dynamics, the vessel‐occlusion test was usually applied as physiological intervention. There are several drawbacks of the vessel‐occlusion method that include skin contact, uncomfortable and microcirculation block of patients. Thus, we propose the far‐infrared (FIR) illumination as a new physiological intervention method in this paper. Our preliminary result shows a linear correlation of oxygenation dynamic signals between FIR illumination and arterial‐occlusion test (AOT) that implies the FIR illumination could be applied for hemodynamic response measurement in clinical diagnosis. (© 2012 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Perfusion‐based fluorescence imaging method delineates diverse organs and identifies multifocal tumors using generic near‐infrared molecular probes

Rapid detection of multifocal cancer without the use of complex imaging schemes will improve treatment outcomes. In this study, dynamic fluorescence imaging was used to harness differences in the perfusion kinetics of near‐infrared (NIR) fluorescent dyes to visualize structural characteristics of different tissues. Using the hydrophobic nontumor‐selective NIR dye cypate, and the hydrophilic dye LS288, a high tumor‐to‐background contrast was achieved, allowing the delineation of diverse tissue types while maintaining short imaging times. By clustering tissue types with similar perfusion properties, the dynamic fluorescence imaging method identified secondary tumor locations when only the primary tumor position was known, with a respective sensitivity and specificity of 0.97 and 0.75 for cypate, and 0.85 and 0.81 for LS288. Histological analysis suggests that the vasculature in the connective tissue that directly surrounds the tumor was a major factor for tumor identification through perfusion imaging. Although the hydrophobic dye showed higher specificity than the hydrophilic probe, use of other dyes with different physical and biological properties could further improve the accuracy of the dynamic imaging platform to identify multifocal tumors for potential use in real‐time intraoperative procedures.      

The whither of bacteriophytochrome‐based near‐infrared fluorescent proteins: Insights from two‐photon absorption spectroscopy

We present one‐ and two‐photon‐absorption fluorescence spectroscopic analysis of biliverdin (BV) chromophore–based single‐domain near‐infrared fluorescent proteins (iRFPs). The results of these studies are used to estimate the internal electric fields acting on BV inside iRFPs and quantify the electric dipole properties of this chromophore, defining the red shift of excitation and emission spectra of BV‐based iRFPs. The iRFP studied in this work is shown to fit well the global diagram of the red‐shift tunability of currently available BV‐based iRFPs as dictated by the quadratic Stark effect, suggesting the existence of the lower bound for the strongest red shifts attainable within this family of fluorescent proteins. The absolute value of the two‐photon absorption (TPA) cross section of a fluorescent calcium sensor based on the studied iRFP is found to be significantly larger than the TPA cross sections of other widely used genetically encodable fluorescent calcium sensors.      

Cellular transformations in near‐infrared light‐induced apoptosis in cancer cells revealed by label‐free CARS imaging

Photobiomodulation (PBM) involves light to activate cellular signaling pathways leading to cell proliferation or death. In this work, fluorescence and Coherent anti‐Stokes Raman Scattering (CARS) imaging techniques were applied to assess apoptosis in human cervical cancer cells (HeLa) induced by near infrared (NIR) laser light (808 nm). Using the Caspase 3/7 fluorescent probe to identify apoptotic cells, we found that the pro‐apoptotic effect is significantly dependent of irradiation dose. The highest apoptosis rate was noted for the lower irradiation doses, that is, 0.3 J/cm² (~58%) and 3 J/cm² (~28%). The impact of light doses on proteins/lipids intracellular metabolism and distribution was evaluated using CARS imaging, which revealed apoptosis‐associated reorganization of nuclear proteins and cytoplasmic lipids after irradiation with 0.3 J/cm². Doses of NIR light causing apoptosis (0.3, 3 and 30 J/cm²) induced a gradual increase in the nuclear protein level over time, in contrast to proteins in cells non‐irradiated and irradiated with 10 J/cm². Furthermore, irradiation of the cells with the 0.3 J/cm² dose resulted in lipid droplets (LDs) accumulation, which was apparently caused by an increase in reactive oxygen species (ROS) generation. We suggest that PBM induced apoptosis could be caused by the ability of NIR light to trigger excessive LDs formation which, in turn, induces cellular cytotoxicity.      

Oxygenation dynamics of sepsis patients undergoing far‐infrared intervention based on near‐infrared spectroscopy

Near‐infrared spectroscopy (NIRS; continuous wave type) is a noninvasive tool for detecting the relative change of oxyhemoglobin and deoxyhemoglobin. To make this change, intervention methods must be applied. This study determined the hemodynamics of 44 healthy participants and 35 patients with sepsis during exposure to FIR as a novel physical intervention approach. Local microcirculation of their brachioradialis was monitored during exposure and recovery through NIRS. The variations in blood flow and microvascular reaction were determined by conducting paired and unpaired t tests. The oxyhemoglobin levels of the healthy participants increased continuously, even during recovery. In contrast to expextations, the oxyhemoglobin levels of the patients plateaued after only 5 min of FIR illumination. The proposed method has potential applications for ensuring efficient treatment and facilitating doctors in diagnosing the functions of vessels in intensive care units.

Mapping diagrams of HbO₂ in healthy males and males with sepsis illustrated unique scenarios during the process.     

Gender differences in brain networks during verbal Sternberg tasks: A simultaneous near‐infrared spectroscopy and electro‐encephalography study

Gender differences in psychological processes have been of great interest in a variety of fields including verbal fluency, emotion processing and working memory. Previous studies suggested that women outperform men in verbal working memory (VWM). However, the inherent mechanisms are still unclear. To obtain a deeper insight into the gender differences in brain networks in VWM, this study used near‐infrared spectroscopy (NIRS) and electro‐encephalography (EEG) simultaneously to investigate gender‐related brain networks during verbal Sternberg tasks. NIRS results confirmed that women surpass men in VWM from the perspective of both brain activation and connectivity. Results of EEG (effective connectivity and event‐related spectral power) showed that men tend to use a more visuospatial strategy to encode memory. In addition, novel analysis methods of brain networks can provide useful information about the gender specifics of brain functions. Gender‐related pseudo‐color maps constructed from all channels of average HbO₂ activity during low‐ and high‐load tasks (from 0 to 6 seconds after beginning).      

Linear polarized near‐infrared irradiation stimulates mechanical expansion of the rat sagittal suture

Mechanical sutural separation has been carried out in clinical orthodontics for controlling the growth of the craniofacial skeleton. This study was designed to evaluate the effects of linear polarized near‐infrared ray irradiation on the sutural expansion of rat sagittal suture. Twenty 8‐week‐old Wistar strain male rats were equally divided into experimental and control groups. Suture expansion was carried out for 5 days for all animals using an expansion appliance. The experimental animals were subjected to linear polarized near‐infrared irradiation. This study has demonstrated that linear polarized near‐infrared irradiation stimulates sutural expansion without any pathological changes. Copyright © 2002 John Wiley & Sons, Ltd.     

Activity assessment of root caries lesions with thermal and near‐IR imaging methods

The purpose of this study was to evaluate thermal and near‐infrared (NIR) reflectance imaging methods for the assessment of the activity of root caries lesions. In addition, changes in the lesion structure were monitored with polarization sensitive optical coherence tomography (PS‐OCT). Artificial bovine and natural root caries lesions were imaged with PS‐OCT, and their dehydration rate was measured with thermal and NIR cameras. The lesion activity of the natural root caries samples was also assessed by two clinicians by conventional means according to ICDAS II guidelines. The thickness of the highly mineralized transparent surface layer measured using PS‐OCT increased and the area enclosed by the time‐temperature curve, ΔQ, measured with thermal imaging decreased significantly with longer periods of remineralization in simulated dentin lesions, but the NIR reflectance intensity differences, ΔI, failed to show any significant relationship with the degree of remineralization. The PS‐OCT algorithm for the automated assessment of remineralization successfully detected the highly mineralized surface layer on both natural and simulated lesions. Thermal imaging provided the most accurate diagnosis of root caries lesion activity. These results demonstrate that thermal imaging and PS‐OCT may be ideally suited for the nondestructive root caries lesion activity during a clinical examination.

     

Head exposure system for a human provocation study to assess the possible influence of UMTS‐like electromagnetic fields on cerebral blood circulation using near‐infrared imaging

A head exposure setup for efficient and precisely defined exposure of human subjects equipped with a near‐infrared imaging (NIRI) sensor is presented. In a partially shielded anechoic chamber the subjects were exposed to Universal Mobile Telecommunications System (UMTS)‐like electromagnetic fields (EMF) by using a patch antenna at a distance of 4 cm from the head. The non‐contact design of the exposure setup enabled NIRI sensors to easily attach to the head. Moreover, different regions of the head were chosen for localised exposure and simultaneous NIRI investigation. The control software enabled the simple adaptation of the test parameters during exploratory testing as well as the performance of controlled, randomised, crossover and double‐blind provocation studies. Four different signals with a carrier frequency of 1900 MHz were chosen for the exposure: a simple continuous wave signal and three different UMTS signals. Furthermore, three exposure doses were available: sham, low (spatial peak specific absorption rate (SAR) = 0.18 W/kg averaged over 10 g) and high (spatial peak SAR = 1.8 W/kg averaged over 10 g). The SAR assessment was performed by measurement and simulation. Direct comparison of measurement and numerical results showed good agreement in terms of spatial peak SAR and SAR distribution. The variability analysis of the spatial peak SAR over 10 g was assessed by numerical simulations. Maximal deviations of ?22% and +32% from the nominal situation were observed. Compared to other exposure setups, the present setup allows for low exposure uncertainty, combined with high SAR efficiency, easy access for the NIRI sensor and minimal impairment of test subjects. Bioelectromagnetics 33:124–133, 2012. © 2011 Wiley Periodicals, Inc.     

Controlling the near‐infrared transparency of costal cartilage by impregnation with clearing agents and magnetite nanoparticles

Penetration depth of near‐infrared laser radiation to costal cartilage is controlled by the tissue absorption and scattering, and it is the critical parameter to provide the relaxation of mechanical stress throughout the whole thickness of cartilage implant. To enhance the penetration for the laser radiation on 1.56 μm, the optical clearing solutions of glycerol and fructose of various concentrations are tested. The effective and reversible tissue clearance was achieved. However, the increasing absorption of radiation should be concerned: 5°C‐8°C increase of tissue temperature was detected. Laser parameters used for stress relaxation in cartilage should be optimized when applying optical clearing agents. To concentrate the absorption in the superficial tissue layers, magnetite nanoparticle (NP) dispersions with the mean size 95 ± 5 nm and concentration 3.9 ± 1.1 × 10¹¹ particles/mL are applied. The significant increase in the tissue heating rate was observed along with the decrease in its transparency. Using NPs the respective laser power can be decreased, allowing us to obtain the working temperature locally with reduced thermal effect on the surrounding tissue.      

Rhodobacter sphaeroides,a novel tumor‐targeting bacteria that emits natural near‐infrared fluorescence

Several optical imaging techniques have been used to monitor bacterial tropisms for cancer. Most such techniques require genetic engineering of the bacteria to express optical reporter genes. This study investigated a novel tumor‐targeting strain of bacteria, Rhodobacter sphaeroides 2.4.1 (R. sphaeroides), which naturally emits near‐infrared fluorescence, thereby facilitating the visualization of bacterial tropisms for cancer. To determine the penetration depth of bacterial fluorescence, various numbers of cells (from 10⁸ to 10¹⁰ CFU) of R. sphaeroides and two types of Escherichia coli, which stably express green fluorescent protein (GFP) or red fluorescent protein (RFP), were injected s.c. or i.m. into mice. Bacterial tropism for cancer was determined after i.v. injection of R. sphaeroides (10⁸ CFU) into mice implanted s.c. with eight types of tumors. The intensity of the fluorescence signal in deep tissue (muscle) from R. sphaeroides was much stronger than from E. coli‐expressing GFP or RFP. The near‐infrared fluorescence signal from R. sphaeroides was visualized clearly in all types of human or murine tumors via accumulation of bacteria. Analyses of C‐reactive protein and procalcitonin concentrations and body weights indicated that i.v. injection of R. sphaeroides does not induce serious systemic immune reactions. This study suggests that R. sphaeroides could be used as a tumor‐targeting microorganism for the selective delivery of drugs to tumor tissues without eliciting a systemic immune reaction and for visualizing tumors.     

In vivo monitoring blood‐brain barrier permeability using spectral imaging through optical clearing skull window

The blood‐brain barrier (BBB) plays a key role in the health of the central nervous system. Opening the BBB is very important for drug delivery to brain tissues to enhance the therapeutic effect on brain diseases. It is necessary to in vivo monitor the BBB permeability for assessing drug release with high resolution; however, an effective method is lacking. In this work, we developed a new method that combined spectral imaging with an optical clearing skull window to in vivo dynamically monitor BBB opening caused by 5‐aminolevulinic acid (5‐ALA)‐mediated photodynamic therapy (PDT), in which the Evans blue dye (EBd) acted as an indicator of the BBB permeability. Using this method, we effectively monitored the cerebrovascular EBd leakage process. Moreover, the analysis of changes in the vascular and extravascular EBd concentrations demonstrated that the PDT‐induced BBB opening exhibited spatiotemporal differences in the cortex. This spectral imaging method based on the optical clearing skull window provides a low‐cost and simply operated tool for in vivo monitoring BBB opening process. This has a high potential for the visualization of drug delivery to the central nervous system. Thus, it is of tremendous significance in brain disease therapy. Monitoring the changes in PDT‐induced BBB permeability by evaluating the EBd concentration using an optical clearing skull window. (A) Entire brains and coronal sections following treatment of PDT with/without an optical clearing skull window after injection of EBd. (B) Typical EBd distribution maps before and after laser irradiation captured by the spectral imaging method. (Colorbar represents the EBd concentration).      

Advanced near‐infrared monitor for stable real‐time measurement and control of Pichia pastoris bioprocesses

Near‐infrared spectroscopy is considered to be one of the most promising spectroscopic techniques for upstream bioprocess monitoring and control. Traditionally the nature of near‐infrared spectroscopy has demanded multivariate calibration models to relate spectral variance to analyte concentrations. The resulting analytical measurements have proven unreliable for the measurement of metabolic substrates for bioprocess batches performed outside the calibration process. This paper presents results of an innovative near‐infrared spectroscopic monitor designed to follow the concentrations of glycerol and methanol, as well as biomass, in real time and continuously during the production of a monoclonal antibody by a Pichia pastoris high cell density process. A solid state instrumental design overcomes the ruggedness limitations of conventional interferometer‐based spectrometers. Accurate monitoring of glycerol, methanol, and biomass is demonstrated over 274 days postcalibration. In addition, the first example of feedback control to maintain constant methanol concentrations, as low as 1 g/L, is presented. Postcalibration measurements over a 9‐month period illustrate a level of reliability and robustness that promises its adoption for online bioprocess monitoring throughout product development, from early laboratory research and development to pilot and manufacturing scale operation. © 2014 American Institute of Chemical Engineers Biotechnol. Prog., 30:749–759, 2014     

Altered near‐infrared spectroscopy response to breath‐holding in patients with fibromyalgia

Fibromyalgia (FM) is a complex syndrome characterized by chronic widespread pain and a heightened response to pressure. Most medical researches pointed out that FM patients with endothelial dysfunction and arterial stiffness. A continuous‐wave near‐infrared spectroscopy (NIRS) system is used in present study to measure the hemodynamic changes elicited by breath‐holding task in patients with FM. Each patient completed a questionnaire survey including demographics, characteristics of body pain, associated symptoms, headache profiles and Hospital Anxiety and Depression Scale. A total of 27 FM patients and 26 health controls were enrolled. In comparison with healthy controls, patients with FM showed lower maximal and averaged change of oxyhemoglobin concentration in both the left (1.634 ±0.890 and 0.810 ±0.525 μM) and the right (1.576 ±0.897 and 0.811 ±0.601 μM) prefrontal cortex than healthy controls (P < .05 for both sides) during the breath‐holding task. In conclusion, FM is associated with altered cerebrovascular reactivity measured by NIRS and breath‐holding task, which may reflect endothelial dysfunction or arterial stiffness. Oxygenated hemoglobin concentration changes of healthy controls and FM patients.      

Fast noninvasive functional diffuse optical tomography for brain imaging

Advances in epilepsy studies have shown that specific changes in hemodynamics precede and accompany seizure onset and propagation. However, it has been challenging to noninvasively detect these changes in real time and in humans, due to the lack of fast functional neuroimaging tools. In this study, we present a functional diffuse optical tomography (DOT) method with the guidance of an anatomical human head atlas for 3‐dimensionally mapping the brain in real time. Central to our DOT system is a human head interface coupled with a technique that can incorporate topological information of the brain surface into the DOT image reconstruction. The performance of the DOT system was tested by imaging motor tasks‐involved brain activities on N = 6 subjects (3 epilepsy patients and 3 healthy controls). We observed diffuse areas of activations from the reconstructed [HbT] images of patients, relative to more focal activations for healthy subjects. Moreover, significant pretask hemodynamic activations were also seen in the motor cortex of patients, which indicated abnormal activities persistent in the brain of an epilepsy patient. This work demonstrates that fast functional DOT is a valuable tool for noninvasive 3‐dimensional mapping of brain hemodynamics.