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Purpose To evaluate the feasibility of a new optical device that measures peripheral blood flow as a diagnostic and monitoring tool for patients with peripheral artery disease (PAD). Materials and Methods In this prospective study, 167 limbs of 90 patients (mean age, 76 y; 53% men) with suspected PAD were evaluated with the FlowMet device, which uses a new type of dynamic light-scattering technology to assess blood flow in real time. Measurements of magnitude and phasicity of blood flow were combined into a single-value flow–waveform score and compared vs ankle–brachial index (ABI), toe–brachial index (TBI), and clinical presentation of patients per Rutherford category (RC). Receiver operating characteristic curves were constructed to predict RC. Area under the curve (AUC), sensitivity, and specificity were compared among flow–waveform score, ABI, and TBI. Results Qualitatively, the FlowMet waveforms were analogous to Doppler velocity measurements, and degradation of waveform phasicity and amplitude were observed with increasing PAD severity. Quantitatively, the flow, waveform, and composite flow–waveform scores decreased significantly with decreasing TBI. In predicting RC ≥ 4, the flow–waveform score (AUC = 0.83) showed a linear decrease with worsening patient symptoms and power comparable to that of TBI (AUC = 0.82) and better than that of ABI (AUC = 0.71). Optimal sensitivity and specificity pairs were found to be 56%/83%, 72%/81%, and 89%/74% for ABI, TBI, and flow–waveform score, respectively. Conclusions The technology tested in this pilot study showed a high predictive value for diagnosis of critical limb ischemia. The device showed promise as a diagnostic tool capable of providing clinical feedback in real time.

Objectives To assess the validity of a bench-top model of an optical tympanometry device to diagnose in vitro model of middle ear effusion (MEE). Methods and materials We illuminated an in vitro model of ear canal and tympanic membrane with broadband light and relayed remitted light to a spectrometer system. We then used our proprietary algorithm to extract spectral features that, together with our logistic regression classifiers, led us to calculate a set of simplified indices related to different middle ear states. Our model included a glass vial covered with a porcine submucosa (representing the tympanic membrane) and filled with air, water, or milk solution (representing different MEE), and a set of cover-glass slips filled with either blood (representing erythema) or cerumen. By interchanging fluid types and cover-glass slips, we made measurements on combinations corresponding to normal healthy ear and purulent or serous MEE. Results Each simulated condition had a distinct spectral profile, which was then employed by our algorithm to discriminate clean and cerumen-covered purulent and serous MEE. Two logistic purulent and serous MEE classifiers correctly classified all in vitro middle ear states with 100% accuracy assessed by leave-one-out and k-fold cross validation. Conclusions This proof-of-concept in vitro study addressed an unmet need by introducing a device that easily and accurately can assess middle ear effusion. Future in vivo studies aimed at collecting data from clinical settings are warranted to further elucidate the validity of the technology in diagnosing pediatric acute otitis media.

Background and Objective Alternative treatments are needed to achieve consistent and more complete port wine stain (PWS) removal, especially in darker skin types; photodynamic therapy (PDT) is a promising alternative treatment. To this end, we previously reported on Talaporfin Sodium (TS)-mediated PDT. It is essential to understand treatment tissue effects to design a protocol that will achieve selective vascular injury without ulceration and scarring. The objective of this work is to assess skin changes associated with TS-mediated PDT with clinically relevant treatment parameters. Study Design/Materials and Methods We performed TS (0.75 mg/kg)-mediated PDT (664 nm) on Sprague Dawley rats. Radiant exposures were varied between 15 and 100 J/cm2. We took skin biopsies from subjects at 9 hours following PDT. We assessed the degree and depth of vascular and surrounding tissue injury using histology and immunohistochemical staining. Results TS-mediated PDT at 0.75 mg/kg combined with 15 and 25 J/cm2 light doses resulted in vascular injury with minimal epidermal damage. At light dose of 50 J/cm2, epidermal damage was noted with vascular injury. At light doses >50 J/cm2, both vascular and surrounding tissue injury were observed in the forms of vasculitis, extravasated red blood cells, and coagulative necrosis. Extensive coagulative necrosis involving deeper adnexal structures was observed for 75 and 100 J/cm2 light doses. Observed depth of injury increased with increasing radiant exposure, although this relationship was not linear. Conclusion TS-mediated PDT can cause selective vascular injury; however, at higher light doses, significant extra-vascular injury was observed. This information can be used to contribute to design of safe protocols to be used for treatment of cutaneous vascular lesions. Lasers Surg. Med. © 2017 Wiley Periodicals, Inc.

Recent advances in optical technology have emerged for measuring blood flow in the extremities using speckleplethysmography (SPG), which may address needs in vascular medicine and other fields. SPG has demonstrated a highly linear response with flow rate, but the susceptibility to differences in skin tone is unclear. Two validation studies using skin-simulating phantoms and a simple clinical protocol were conducted to determine the impact of absorbing skin layers on SPG measurements. Benchtop results demonstrated that the coefficient of determination between known flow rate and SPG was highly linear (R2 = 0.990) and was unaffected by the addition of skin-phantom layers with variable absorption (R2 = 0.996-0.999). Additionally, no significant trend was found between the fit residuals of SPG and flow rate with increasing skin-phantom absorption (R2=0.025, p = 0.29). In clinical testing, no significant difference was found using both a 4-way ANOVA between demographic classifications (F = 0.89, p = 0.45), and a 2-way ANOVA test between lower- and higher-melanin subclassifications (F = 0.4, p = 0.52).

In this work we introduce a modified form of laser speckle imaging (LSI) referred to as affixed transmission speckle analysis (ATSA) that uses a single coherent light source to probe two physiological signals: one related to pulsatile vascular expansion (classically known as the photoplethysmographic (PPG) waveform) and one related to pulsatile vascular blood flow (named here the speckle plethysmographic (SPG) waveform). The PPG signal is determined by recording intensity fluctuations, and the SPG signal is determined via the LSI dynamic light scattering technique. These two co-registered signals are obtained by transilluminating a single digit (e.g. finger) which produces quasi-periodic waveforms derived from the cardiac cycle. Because PPG and SPG waveforms probe vascular expansion and flow, respectively, in cm-thick tissue, these complementary phenomena are offset in time and have rich dynamic features. We characterize the timing offset and harmonic content of the waveforms in 16 human subjects and demonstrate physiologic relevance for assessing microvascular flow and resistance.

Dermatoscopes are commonly utilized by medical professionals for the qualitative visual inspection of skin lesions. While automated image processing techniques and varied illumination strategies can aid in structural analysis of lesions, robust quantification of functional information is largely unknown. To address this knowledge gap, we have developed a compact, handheld dermatoscope that enables real-time blood flow measurements of skin using coherent illumination and laser speckle imaging (LSI). A second color camera attached to the dermatoscope helps with the simultaneous real-time observation of the skin lesions and allows the user to acquire and save color images via a custom Graphical User Interface. In-vitro characterization utilizing a blood flow phantom demonstrated that the dermatoscope is capable of quantifying changes in blood flow across a physiologically relevant range even when used in a handheld manner with ambient lighting. We also demonstrated that the dermatoscope can quantify blood flow in skin lesions in human subjects and that significant differences in blood flow are present among lesion types. There was significantly increased blood flow relative to the surrounding skin in cherry angiomas compared to solar lentigos (p

Author(s): Ozel, Tugba; White, Sean; Nguyen, Elaine; Moy, Austin; Brenes, Nicholas; Choi, Bernard; Betancourt, Tania | Abstract: Background and objectiveNanotechnology offers the possibility of creating multi-functional structures that can provide solutions for biomedical problems. The nanoprobes herein described are an example of such structures, where nano-scaled particles have been designed to provide high specificity and contrast potential for optical detection of cancer. Specifically, enzymatically activated fluorescent nanoprobes (EANPs) were synthesized as cancer-specific contrast agents for optical imaging.Study design/materials and methodsEANPs were prepared by nanoprecipitation of blends of poly(lactic acid)-b-poly(ethylene glycol) and poly(lactic-co-glycolic acid)-b-poly(l-lysine). The lysine moieties were then covalently decorated with the near infrared (NIR) fluorescent molecule AlexaFluor-750 (AF750). Close proximity of the fluorescent molecules to each other resulted in fluorescence quenching, which was reversed by enzymatically mediated cleavage of poly(l-lysine) chains. EANPs were characterized by dynamic light scattering and electron microscopy. Enzymatic development of fluorescence was studied in vitro by fluorescence spectroscopy. Biocompatibility and contrast potential of EANPs were studied in cancerous and noncancerous cells. The potential of the nanoprobes as contrast agents for NIR fluorescence imaging was studied in tissue phantoms.ResultsSpherical EANPs of ∼100 nm were synthesized via nanoprecipitation of polymer blends. Fluorescence activation of EANPs by treatment with a model protease was demonstrated with up to 15-fold optical signal enhancement within 120 minutes. Studies with MDA-MB-231 breast cancer cells demonstrated the cytocompatibility of EANPs, as well as enhanced fluorescence associated with enzymatic activation. Imaging studies in tissue phantoms confirmed the ability of a simple imaging system based on a laser source and CCD camera to image dilute suspensions of the nanoprobe at depths of up to 4 mm, as well as up to a 13-fold signal-to-background ratio for enzymatically activated EANPs compared to un-activated EANPs at the same concentration.ConclusionNanoprecipitation of copolymer blends containing poly(l-lysine) was utilized as a method for preparation of highly functional nanoprobes with high potential as contrast agents for fluorescence based imaging of cancer. Lasers Surg. Med. 47:579-594, 2015. © 2015 Wiley Periodicals, Inc.

Alternative treatments are needed to achieve consistent and more complete port wine stain (PWS) removal, especially in darker skin types; photodynamic therapy (PDT) is a promising alternative treatment. To this end, we previously reported on Talaporfin Sodium (TS)-mediated PDT. It is essential to understand treatment tissue effects to design a protocol that will achieve selective vascular injury without ulceration and scarring. The objective of this work is to assess skin changes associated with TS-mediated PDT with clinically relevant treatment parameters.We performed TS (0.75 mg/kg)-mediated PDT (664 nm) on Sprague Dawley rats. Radiant exposures were varied between 15 and 100 J/cmTS-mediated PDT at 0.75 mg/kg combined with 15 and 25 J/cmTS-mediated PDT can cause selective vascular injury; however, at higher light doses, significant extra-vascular injury was observed. This information can be used to contribute to design of safe protocols to be used for treatment of cutaneous vascular lesions. Lasers Surg. Med. 49:767-772, 2017. © 2017 Wiley Periodicals, Inc.

We report a methodology for computing functional vascular density within a rodent dorsal window chamber model based on long-exposure laser speckle imaging (LSI). This technique relies on the presence of flow to create detailed vasculature maps. Employing this contrast mechanism is not possible using conventional imaging methods. Additionally, a freeware algorithm for computing functional vascular density (FVD) from images acquired using long-exposure LSI is also described to facilitate ease in adopting this method. We demonstrate that together these tools can be used to compute FVD nearly twelve times faster than manual computation, yet with with comparable accuracy.

© 2015 IEEE. Here, we review our current knowledge on the etiology and treatment of port-wine stain (PWS) birthmarks. Current treatment options have significant limitations in terms of efficacy. With the combination of 1) a suitable preclinical microvascular model, 2) laser speckle imaging (LSI) to evaluate blood-flow dynamics, and 3) a longitudinal experimental design, rapid preclinical assessment of new phototherapies can be translated from the lab to the clinic. The combination of photodynamic therapy (PDT) and pulsed-dye laser (PDL) irradiation achieves a synergistic effect that reduces the required radiant exposures of the individual phototherapies to achieve persistent vascular shutdown. PDL combined with antiangiogenic agents is a promising strategy to achieve persistent vascular shutdown by preventing reformation and reperfusion of photocoagulated blood vessels. Integration of LSI into the clinical workflow may lead to surgical image guidance that maximizes acute photocoagulation, which is expected to improve PWS therapeutic outcome. Continued integration of noninvasive optical imaging technologies and biochemical analysis collectively are expected to lead to more robust treatment strategies.

Alginate encapsulation reduces the risk of transplant rejection by evading immune-mediated cell injury and rejection; however, poor vascular perfusion results in graft failure. Since existing imaging models are incapable of quantifying the vascular response to biomaterial implants after transplantation, in this study, we demonstrate the use of invivo laser speckle imaging (LSI) and wide-field functional imaging (WiFI) to monitor the microvascular environment surrounding biomaterial implants. The vascular response to two islet-containing biomaterial encapsulation devices, alginate microcapsules and a high-guluronate alginate sheet, was studied and compared after implantation into the mouse dorsal window chamber (N=4 per implant group). Images obtained over a 14-day period using LSI and WiFI were analyzed using algorithms to quantify blood flow, hemoglobin oxygen saturation and vascular density. Using our method, we were able to monitor the changes in the peri-implant microvasculature noninvasively without the use of fluorescent dyes. Significant changes in blood flow, hemoglobin oxygen saturation and vascular density were noted as early as the first week post-transplant. The dorsal window chamber model enables comparison of host responses to transplanted biomaterials. Future experiments will study the effect of changes in alginate composition on the vascular and immune responses. © 2013 Elsevier Ltd.

© Copyright 2014, Mary Ann Liebert, Inc. Achieving adequate vascularization within implanted engineered tissues is a significant obstacle to maintaining viability and functionality. In vitro prevascularization of engineered tissues has been explored as a potential solution to this challenge. The traditional paradigm of in vitro prevascularization is to implant an engineered tissue with a preformed vascular network that is perfused after anastomosis with the host circulation. We investigated the efficacy of this strategy by implanting cell-dense prevascularized tissues created via cell-mediated contraction and composed of collagen and a collagen-fibrin mixture into dorsal window chambers surgically prepared on immunocompromised mice. We found that host-implant anastomosis takes place in 2-6 days and that perfusion of vessels within the implants is subsequently restricted by thrombosis. However, by day 7, a functional vascular network composed of host and implant vessels developed. Prevascularization enhanced intra-implant pO2significantly as early as 2 days postimplantation, reaching a maximum of 55 mmHg by day 8, which was significantly greater than the maximum within cellularized control tissues (18 mmHg). By day 14, collagen tissues supported ∼0.51 × 109implanted and host-derived cells per mL. Our findings elucidate key features of in vitro prevascularization that can be used toward the design of larger and more functionally complex engineered tissues.

Current methods used to assess gingivitis are qualitative and subjective. We hypothesized that gingival perfusion measurements could provide a quantitative metric of disease severity. We constructed a compact laser speckle imaging (LSI) system that could be mounted in custom-made oral molds. Rigid fixation of the LSI system in the oral cavity enabled measurement of blood flow in the gingiva. In vitro validation performed in controlled flow phantoms demonstrated that the compact LSI system had comparable accuracy and linearity compared to a conventional bench-top LSI setup. In vivo validation demonstrated that the compact LSI system was capable of measuring expected blood flow dynamics during a standard postocclusive reactive hyperemia and that the compact LSI system could be used to measure gingival blood flow repeatedly without significant variation in measured blood flow values (p

The functionality of vascular networks within implanted prevascularized tissues is difficult to assess using traditional analysis techniques, such as histology. This is largely due to the inability to visualize hemodynamics in vivo longitudinally. Therefore, we have developed dynamic imaging methods to measure blood flow and hemoglobin oxygen saturation in implanted prevascularized tissues noninvasively and longitudinally. Using laser speckle imaging, multispectral imaging, and intravital microscopy, we demonstrate that fibrin-based tissue implants anastomose with the host (severe combined immunodeficient mice) in as short as 20h. Anastomosis results in initial perfusion with highly oxygenated blood, and an increase in average hemoglobin oxygenation of 53%. However, shear rates in the preformed vessels were low (20.8±12.8s-1), and flow did not persist in the vast majority of preformed vessels due to thrombus formation. These findings suggest that designing an appropriate vascular network structure in prevascularized tissues to maintain shear rates above the threshold for thrombosis may be necessary to maintain flow following implantation. We conclude that wide-field and microscopic functional imaging can dynamically assess blood flow and oxygenation in vivo in prevascularized tissues, and can be used to rapidly evaluate and improve prevascularization strategies. © Copyright 2012, Mary Ann Liebert, Inc.

The rodent dorsal window chamber is a widely used in vivo model of the microvasculature. The model consists of a 1cm region of exposed microvasculature in the rodent dorsal skin that is immobilized by surgically implanted titanium frames, allowing the skin microvasculature to be visualized. We describe a detailed protocol for surgical implantation of the dorsal window chamber which enables researchers to perform the window chamber implantation surgery. We further describe subsequent wide-field functional imaging of the chamber to obtain hemodynamic information in the form of blood oxygenation and blood flow on a cm size region of interest. Optical imaging techniques, such as intravital microscopy, have been applied extensively to the dorsal window chamber to study microvascular-related disease and conditions. Due to the limited field of view of intravital microscopy, detailed hemodynamic information typically is acquired from small regions of interest, typically on the order of hundreds of μm. The wide-field imaging techniques described herein complement intravital microscopy, allowing researchers to obtain hemodynamic information at both microscopic and macroscopic spatial scales. Compared with intravital microscopy, wide-field functional imaging requires simple instrumentation, is inexpensive, and can give detailed metabolic information over a wide field of view.

In this paper, we report experimental results of a novel SPS on demand based on a single CdSe QD suspended in a liquid crystal host self-assembled in a 1D chiral photonic bandgap structure. This structure provides not only spontaneous emission enhancement and a diminishing of the fluorescence lifetime, but also circular polarization of definite handedness even for emitters without a dipole moment. 1D chiral photonic bandgap structures possess an advantage over conventional 1D photonic bandgap technologies. Because the refractive index n varies gradually in chiral structures rather than abruptly, there are no losses into the waveguide modes, which arise from total internal reflection at the border between two consecutive layers with different n. We report here for the first time fluorescence antibunching of a QD doped in a liquid crystal photonic bandgap material. Earlier we reported both fluorescence antibunching and deterministic polarization of dye fluorescence in aligned liquid crystal hosts.

A single-photon source based on single CdSe quantum-dot fluorescence in a chiral-photonic-bandgap liquid-crystal host manifests itself in observed fluorescence antibunching. Chiral-photonic-bandgap structures will provide deterministically handed, circular-polarized fluorescence, even for emitters without a dipole moment.

Quantum optics teaching laboratory consists of four experiments for undergraduates: (1) entanglement and Bell’s inequalities, (2) single-photon interference, (3) confocal microscope imaging of single-emitter fluorescence, (4) Hanbury Brown and Twiss setup. Fluorescence antibunching.