Characterization of the optical properties of color pastes for the design of optical phantoms mimicking biological tissue

Clinicians need a way to rapidly and reliably test the correct functioning of near‐infrared spectroscopy (NIRS)–based oximeters. Therefore, optical phantoms for quality assessment of NIRS oximeters are needed. The fabrication of such phantoms that mimic the optical properties of biological tissue in the NIR range represents a challenge. To enable their development, the aim was to characterize the absorption and scattering spectra of different dyes. The optical properties of silicone SILPURAN 2420 with 11 color pastes of type ELASTOSIL were measured in the 500 to 1000 nm range by a spectrometer with an integrating sphere. In addition, two commercial frequency‐domain NIRS devices, the ISS OxiplexTS and the ISS Imagent, were used to assess the optical properties at specific wavelengths. The evaluated colors present mostly features in the visible range below 650 nm, but two colors include peaks in the near‐infrared region, simulating low tissue oxygenation values. These colors were used to create an optical phantom, which matched the designed StO₂ value within an error of only 4%. This set of dyes already enables simulating many different spectra, thus achieving a first step on the way to a long‐term stable comparison and validation method.      

Transmission in near‐infrared optical windows for deep brain imaging

Near‐infrared (NIR) radiation has been employed using one‐ and two‐photon excitation of fluorescence imaging at wavelengths 650–950 nm (optical window I) for deep brain imaging; however, longer wavelengths in NIR have been overlooked due to a lack of suitable NIR‐low band gap semiconductor imaging detectors and/or femtosecond laser sources. This research introduces three new optical windows in NIR and demonstrates their potential for deep brain tissue imaging. The transmittances are measured in rat brain tissue in the second (II, 1,100–1,350 nm), third (III, 1,600–1,870 nm), and fourth (IV, centered at 2,200 nm) NIR optical tissue windows. The relationship between transmission and tissue thickness is measured and compared with the theory. Due to a reduction in scattering and minimal absorption, window III is shown to be the best for deep brain imaging, and windows II and IV show similar but better potential for deep imaging than window I.

     

Diffuse optical assessment of cerebral‐autoregulation in older adults stratified by cerebrovascular risk

Diagnosis of cerebrovascular disease (CVD) at early stages is essential for preventing sequential complications. CVD is often associated with abnormal cerebral microvasculature, which may impact cerebral‐autoregulation (CA). A novel hybrid near‐infrared diffuse optical instrument and a finger plethysmograph were used to simultaneously detect low‐frequency oscillations (LFOs) of cerebral blood flow (CBF), oxy‐hemoglobin concentration ([HbO₂]), deoxy‐hemoglobin concentration ([Hb]) and mean arterial pressure (MAP) in older adults before, during and after 70° head‐up‐tilting (HUT). The participants with valid data were divided based on Framingham risk score (FRS, 1‐30 points) into low‐risk (FRS ≤15, n = 13) and high‐risk (FRS >15, n = 11) groups for developing CVD. The LFO gains were determined by transfer function analyses with MAP as the input, and CBF, [HbO₂] and [Hb] as the outputs (CA ∝ 1/Gain) . At resting‐baseline, LFO gains in the high‐risk group were relatively lower compared to the low‐risk group. The lower baseline gains in the high‐risk group may attribute to compensatory mechanisms to maintain stronger steady‐state CAs. However, HUT resulted in smaller gain reductions in the high‐risk group compared to the low‐risk group, suggesting weaker dynamic CAs. LFO gains are potentially valuable biomarkers for early detection of CVD based on associations with CAs.     

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.     

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).      

Real‐time optical spectroscopic monitoring of nonirrigated lesion progression within atrial and ventricular tissues

Despite considerable advances in guidance of radiofrequency ablation (RFA) therapy for the treatment of cardiac arrhythmias, success rates have been hampered by a lack of tools for precise intraoperative evaluation of lesion extent. Near‐infrared spectroscopic (NIRS) techniques are sensitive to tissue structural and biomolecular properties, characteristics that are directly altered by radiofrequency (RF) treatment. In this work, a combined NIRS‐RFA catheter is developed for real‐time monitoring of tissue reflectance during RF energy delivery. An algorithm is proposed for processing NIR spectra to approximate nonirrigated lesion depth in both atrial and ventricular tissues. The probe optical geometry was designed to bias measurement influence toward absorption enabling enhanced sensitivity to changes in tissue composition. A set of parameters termed “lesion optical indices” are defined encapsulating spectral differences between ablated and unablated tissue. Utilizing these features, a model for real‐time tissue spectra classification and lesion size estimation is presented. Experimental validation conducted within freshly excised porcine cardiac specimens showed strong concordance between algorithm estimates and post‐hoc tissue assessment.      

Comparison of spectroscopy technologies for improved monitoring of cell culture processes in miniature bioreactors

Cell culture process development requires the screening of large numbers of cell lines and process conditions. The development of miniature bioreactor systems has increased the throughput of such studies; however, there are limitations with their use. One important constraint is the limited number of offline samples that can be taken compared to those taken for monitoring cultures in large‐scale bioreactors. The small volume of miniature bioreactor cultures (15 mL) is incompatible with the large sample volume (600 µL) required for bioanalysers routinely used. Spectroscopy technologies may be used to resolve this limitation. The purpose of this study was to compare the use of NIR, Raman, and 2D‐fluorescence to measure multiple analytes simultaneously in volumes suitable for daily monitoring of a miniature bioreactor system. A novel design‐of‐experiment approach is described that utilizes previously analyzed cell culture supernatant to assess metabolite concentrations under various conditions while providing optimal coverage of the desired design space. Multivariate data analysis techniques were used to develop predictive models. Model performance was compared to determine which technology is more suitable for this application. 2D‐fluorescence could more accurately measure ammonium concentration (RMSE_CV 0.031 g L^?1) than Raman and NIR. Raman spectroscopy, however, was more robust at measuring lactate and glucose concentrations (RMSE_CV 1.11 and 0.92 g L^?1, respectively) than the other two techniques. The findings suggest that Raman spectroscopy is more suited for this application than NIR and 2D‐fluorescence. The implementation of Raman spectroscopy increases at‐line measuring capabilities, enabling daily monitoring of key cell culture components within miniature bioreactor cultures. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:337–346, 2017     

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     

In‐situ monitoring of Saccharomyces cerevisiae ITV01 bioethanol process using near‐infrared spectroscopy NIRS and chemometrics

The application feasibility of in‐situ or in‐line monitoring of S. cerevisiae ITV01 alcoholic fermentation process, employing Near‐Infrared Spectroscopy (NIRS) and Chemometrics, was investigated. During the process in a bioreactor, in the complex analytical matrix, biomass, glucose, ethanol and glycerol determinations were performed by a transflection fiber optic probe immersed in the culture broth and connected to a Near‐Infrared (NIR) process analyzer. The NIR spectra recorded between 800 and 2,200 nm were pretreated using Savitzky‐Golay smoothing and second derivative in order to perform a partial least squares regression (PLSR) and generate the calibration models. These calibration models were tested by external validation and then used to predict concentrations in batch alcoholic fermentations. The standard errors of calibration (SEC) for biomass, ethanol, glucose and glycerol were 0.212, 0.287, 0.532, and 0.296 g/L and standard errors of prediction (SEP) were 0.323, 0.369, 0.794, and 0.507 g/L, respectively. Calibration and validation criteria were defined and evaluated in order to generate robust and reliable models for an alcoholic fermentation process matrix. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:510–517, 2016     

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.

     

Brain structure and spatial sensitivity profile assessing by near‐infrared spectroscopy modeling based on 3D MRI data

The goal of this study is to prove that the light propagation in the head by used the 3‐D optical model from in vivo MRI data set can also provide significant characteristics on the spatial sensitivity of cerebral cortex folding geometry based on Monte Carlo simulation. Thus, we proposed a MRI based approach for 3‐D brain modeling of near‐infrared spectroscopy (NIRS). In the results, the spatial sensitivity profile of the cerebral cortex folding geometry and the arrangement of source‐detector separation have being necessarily considered for applications of functional NIRS. The optimal choice of source‐detector separation is suggested within 3–3.5 cm by the received intensity with different source‐detector separations and the ratio of received light from the gray and white matter layer is greater than 50%. Additionally, this study has demonstrated the capability of NIRS in not only assessing the functional but also detecting the structural change of the brain by taking advantage of the low scattering and absorption coefficients observed in CSF of sagittal view. (© 2013 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Investigation of the source‐detector separation in near infrared spectroscopy for healthy and clinical applications

Understanding near infrared light propagation in tissue is vital for designing next generation optical brain imaging devices. Monte Carlo (MC) simulations provide a controlled mechanism to characterize and evaluate contributions of diverse near infrared spectroscopy (NIRS) sensor configurations and parameters. In this study, we developed a multilayer adult digital head model under both healthy and clinical settings and assessed light‐tissue interaction through MC simulations in terms of partial differential pathlength, mean total optical pathlength, diffuse reflectance, detector light intensity and spatial sensitivity profile of optical measurements. The model incorporated four layers: scalp, skull, cerebrospinal‐fluid and cerebral cortex with and without a customizable lesion for modeling hematoma of different sizes and depths. The effect of source‐detector separation (SDS) on optical measurements' sensitivity to brain tissue was investigated. Results from 1330 separate simulations [(4 lesion volumes × 4 lesion depths for clinical +3 healthy settings) × 7 SDS × 10 simulation = 1330)] each with 100 million photons indicated that selection of SDS is critical to acquire optimal measurements from the brain and recommended SDS to be 25 to 35 mm depending on the wavelengths to obtain optical monitoring of the adult brain function. The findings here can guide the design of future NIRS probes for functional neuroimaging and clinical diagnostic systems.      

Evaluation of the bony repair in rat cranial defect using near infrared reflectance spectroscopy and discriminant analysis

We set out to determine whether near infrared reflectance spectroscopy (NIRS) combined with principal component analysis–linear discriminant analysis (LDA) or, variable selection techniques employing successive projection algorithm or genetic algorithm (GA) could evaluate the bone repair in cranial critical‐size (5 mm) defect after stimulation with collagen sponge scaffold and/or infrared low‐level laser therapy directly on the local. Forty‐five Winstar rats were divided into nine groups of five each, namely: group H – healthy, n = 5 (without treatment and without cranial critical‐size defect), (GI positive control – n = 5, 21 days or n = 5, 30 days) without treatment and with cranial critical‐size defect; (GII‐n = 5, 21 days or n = 5, 30 days) cranial critical‐size defect filled with collagen sponge scaffold; (GIII–n = 5, 21 days or n = 5, 30 days) cranial critical‐size defect submitted to low‐level laser therapy; (GIV–n = 5, 21 days or n = 5, 30 days) cranial critical‐size defect submitted to combined collagen sponge scaffold + low‐level laser therapy treatment. In relation to the histological analysis, the collagen sponge scaffold + low‐level laser therapy treatment group (GIV) 30 days showed the best result with the presence of secondary bone, immature bone (osteoid) and newly formed connective tissue (periosteum). GA–LDA model also successfully classified control class of the others classes. Thus, the results provided by the good‐quality classification model revealed the feasibility of NIRS for application to evaluation of the wound healing in rat cranial defect, thanks to the short analysis time of a few seconds and nondestructive advantages of NIRS as an alternative approach for bone repair purposes. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1160–1168, 2017     

The nuclear transporter SAD2 plays a role in calcium‐ and H2O2‐mediated cell death in Arabidopsis

In response to pathogens, plant cells exhibit a rapid increase in the intracellular calcium concentration and a burst of reactive oxygen species (ROS). The cytosolic increase in Ca²⁺ and the accumulation of ROS are critical for inducing programmed cell death (PCD), but the molecular mechanism is not fully understood. We screened an Arabidopsis mutant, sad2‐5, which harbours a T‐DNA insertion in the 18^th exon of the importin beta‐like gene, SAD2. The H₂O₂‐induced increase in the [Ca²⁺]_cyt of the sad2‐5 mutant was greater than that of the wild type, and the sad2‐5 mutant showed clear cell death phenotypes and abnormal H₂O₂ accumulation under fumonisin‐B1 (FB1) treatment. CaCl₂ could enhance the FB1‐induced cell death of the sad2‐5 mutant, whereas lanthanum chloride (LaCl₃), a broad‐spectrum calcium channel blocker, could restore the FB1‐induced PCD phenotype of sad2‐5. The sad2‐5 fbr11‐1 double mutant exhibited the same FB1‐insensitive phenotype as fbr11‐1, which plays a critical role in novo sphingolipid synthesis, indicating that SAD2 works downstream of FBR11. These results suggest the important role of nuclear transporters in calcium‐ and ROS‐mediated PCD response as well as provide an important theoretical basis for further analysis of the molecular mechanism of SAD2 function in PCD and for improvement of the resistance of crops to adverse environments.     

Fast and sensitive delineation of brain tumor with clinically compatible moxifloxacin labeling and confocal microscopy

Delineation of brain tumor margins during surgery is critical to maximize tumor removal while preserving normal brain tissue to obtain optimal clinical outcomes. Although various imaging methods have been developed, they have limitations to be used in clinical practice. We developed a high‐speed cellular imaging method by using clinically compatible moxifloxacin and confocal microscopy for sensitive brain tumor detection and delineation. Moxifloxacin is a Food and Drug Administration (FDA) approved antibiotic and was used as a cell labeling agent through topical administration. Its strong fluorescence at short visible excitation wavelengths allowed video‐rate cellular imaging. Moxifloxacin‐based confocal microscopy (MBCM) was characterized in normal mouse brain specimens and visualized their cytoarchitecture clearly. Then, MBCM was applied to both brain tumor murine models and two malignant human brain tumors of glioblastoma and metastatic cancer. MBCM detected tumors in all the specimens by visualizing dense and irregular cell distributions, and tumor margins were easily delineated based on the cytoarchitecture. An image analysis method was developed for automated detection and delineation. MBCM demonstrated sensitive delineation of brain tumors through cytoarchitecture visualization and would have potentials for human applications, such as a surgery‐guiding method for tumor removal.      

In situ near infrared spectroscopy for analyte‐specific monitoring of glucose and ammonium in streptomyces coelicolor fermentations

There are many challenges associated with in situ collection of near infrared (NIR) spectra in a fermentation broth, particularly for highly aerated and agitated fermentations with filamentous organisms. In this study, antibiotic fermentation by the filamentous bacterium Streptomyces coelicolor was used as a model process. Partial least squares (PLS) regression models were calibrated for glucose and ammonium based on NIR spectra collected in situ. To ensure that the models were calibrated based on analyte‐specific information, semisynthetic samples were used for model calibration in addition to data from standard batches. Thereby, part of the inherent correlation between the analytes could be eliminated. The set of semisynthetic samples were generated from fermentation broth from five separate fermentations to which different amounts of glucose, ammonium, and biomass were added. This method has previously been used off line but never before in situ. The use of semisynthetic samples along with validation on an independent batch provided a critical and realistic evaluation of analyte‐specific models based on in situ NIR spectroscopy. The prediction of glucose was highly satisfactory resulting in a RMSEP of 1.1 g/L. The prediction of ammonium based on NIR spectra collected in situ was not satisfactory. A comparison with models calibrated based on NIR spectra collected off line suggested that this is caused by signal attenuation in the optical fibers in the region above 2,000 nm; a region which contains important absorption bands for ammonium. For improved predictions of ammonium in situ, it is suggested to focus efforts on enhancing the signal in that particular region. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2010     

Targeting the ROS/PI3K/AKT/HIF‐1α/HK2 axis of breast cancer cells: Combined administration of Polydatin and 2‐Deoxy‐d‐glucose

It is well established that cancer cells depend upon aerobic glycolysis to provide the energy they need to survive and proliferate. However, anti‐glycolytic agents have yielded few positive results in human patients, in part due to dose‐limiting side effects. Here, we discovered the unexpected anti‐cancer efficacy of Polydatin (PD) combined with 2‐deoxy‐D‐glucose (2‐DG), which is a compound that inhibits glycolysis. We demonstrated in two breast cell lines (MCF‐7 and 4T1) that combination treatment with PD and 2‐DG induced cell apoptosis and inhibited cell proliferation, migration and invasion. Furthermore, we determined the mechanism of PD in synergy with 2‐DG, which decreased the intracellular reactive oxygen (ROS) levels and suppressed the PI3K/AKT pathway. In addition, the combined treatment inhibited the glycolytic phenotype through reducing the expression of HK2. HK2 deletion in breast cancer cells thus improved the anti‐cancer activity of 2‐DG. The combination treatment also resulted in significant tumour regression in the absence of significant morphologic changes in the heart, liver or kidney in vivo. In summary, our study demonstrates that PD synergised with 2‐DG to enhance its anti‐cancer efficacy by inhibiting the ROS/PI3K/AKT/HIF‐1α/HK2 signalling axis, providing a potential anti‐cancer strategy.