Capillary Refill-The Key to Assessing Dermal Capillary Capacity and Pathology in Optical Coherence Tomography Angiography.

BACKGROUND/OBJECTIVES: Standard optical coherence tomography angiography (OCTA) has been limited to imaging blood vessels actively undergoing perfusion, providing a temporary picture of surface microvasculature. Capillary perfusion in the skin is dynamic and changes in response to the surrounding tissue's respiratory, nutritional, and thermoregulatory needs. Hence, OCTA often represents a given perfusion state without depicting the actual extent of the vascular network. Here we present a method for obtaining a more accurate anatomic representation of the surface capillary network in human skin using OCTA, along with proposing a new parameter, the Relative Capillary Capacity (RCC), a quantifiable proxy for assessing capillary dilation potential and permeability. METHODS: OCTA images were captured at baseline and after compression of the skin. Baseline images display ambient capillary perfusion, while images taken upon capillary refill display the network of existing capillaries at full capacity. An optimization-based automated vessel segmentation method was used to automatically analyze and compare OCTA image sequences obtained from two volunteers. RCC was then compared with visual impressions of capillary viability. RESULTS: Our OCTA imaging sequence provides a method for mapping cutaneous capillary networks independent of ambient perfusion. Differences between baseline and refill images clearly demonstrate the shortcomings of standard OCTA imaging and produce the RCC biometric as a quantifiable proxy for assessing capillary dilation potential and permeability. CONCLUSION: Future dermatological OCTA diagnostic studies should implement the Capillary Refill Methods over standard imaging techniques and further explore the relevance of RCC to differential diagnosis and dermatopathology. Lasers Surg. Med. © The Authors. Lasers in Surgery and Medicine published by Wiley Periodicals, Inc.

Combined OCT distance and FBG force sensing cannulation needle for retinal vein cannulation: in vivo animal validation.

PURPOSE: Retinal vein cannulation is an experimental procedure during which a clot-dissolving drug is injected into an obstructed retinal vein. However, due to the fragility and minute size of retinal veins, such procedure is considered too risky to perform manually. With the aid of surgical robots, key limiting factors such as: unwanted eye rotations, hand tremor and instrument immobilization can be tackled. However, local instrument anatomy distance and force estimation remain unresolved issues. A reliable, real-time local interaction estimation between instrument tip and the retina could be a solution. This paper reports on the development of a combined force and distance sensing cannulation needle, and its experimental validation during in vivo animal trials. METHODS: Two prototypes are reported, relying on force and distance measurements based on FBG and OCT A-scan fibres, respectively. Both instruments provide an 80 [Formula: see text] needle tip and have outer shaft diameters of 0.6 and 2.3 mm, respectively. RESULTS: Both prototypes were characterized and experimentally validated ex vivo. Then, paired with a previously developed surgical robot, in vivo experimental validation was performed. The first prototype successfully demonstrated the feasibility of using a combined force and distance sensing instrument in an in vivo setting. CONCLUSION: The results demonstrate the feasibility of deploying a combined sensing instrument in an in vivo setting. The performed study provides a foundation for further work on real-time local modelling of the surgical scene. This paper provides initial insights; however, additional processing remains necessary.

[Imaging Blood Flow and Pulsation of Retinal Vessels with Full-Field Swept-Source OCT]. / Darstellung von Blutfluss und Pulsation in retinalen Gefäßen mit Full-Field-Swept-Source-OCT.

Optical coherence tomography (OCT) uses interference to image the retina with high axial resolution. In the last 25 years, new technologies have permitted a steady increase in imaging speed, which made it possible to enlarge the imaged field and to avoid motion artefacts. The speed and precision of retinal imaging is now limited by photodamage of the retina caused by the focused OCT beam and by the speed of the scanning mechanics. Full-field swept-source (FF-SS)-OCT decreases irradiance on the retina and dispenses moving parts by using a camera to acquire the full volume of the retina in parallel. Here we show that FF-SS-OCT is rapid and precise enough to image pulsation in the retina induced by the heart beat. Series of OCT volumes 1.8 × 0.7 mm wide and 1.8 mm deep were recorded in young volunteers over a few cycles of the heart beat. Morphology of the retinal vessels, blood flow and tissue motion as caused by vessel pulsation were calculated from the OCT data. FF-SS-OCT was able to visualise the main structures of the neuronal retina, including vessels and small capillaries and without any motion artefacts. Information on three different dynamic processes was obtained from only one recorded series of OCT volumes: pulsation of blood flow and blood pressure in retinal vessels as well as pulsation of the choroid. Delays between arterial and venous pulse and delay between pulsation in retinal and choroidal vessels were calculated. With a time resolution of 0.5 ms, FF-SS-OCT is able to visualise previously unmeasurably fast changes in the retina, including the propagation of pulse waves.

Microanatomy of the tympanic membrane in chronic myringitis obtained with optical coherence tomography.

A microscope-based optical coherence tomography (OCT) device was used to assess the microanatomy of the tympanic membrane in patients with chronic myringitis. A prospective study was designed for this purpose. OCT measurements of the tympanic membrane were done on 11 patients with myringitis with a microscope-based spectral domain OCT system. The in vivo findings were compared with those findings of a control group consisting of 36 patients with retraction pockets or atrophic tympanic membranes (n = 13), myringosclerosis (n = 12) and perforations (n = 11). In active chronic myringitis, the thickness of the tympanic membrane is increased compared to healthy membranes and to other pathological conditions of the tympanic membrane. Consistent changes of the microanatomy of the tympanic membrane were found in chronic myringitis with OCT. Serial OCT measurements revealed no biofilm suspicious findings in all patients with active chronic myringitis. Intraoperative and in vivo OCT measurements may help to detect microanatomical changes of the tympanic membrane in chronic myringitis and in other conditions of the tympanic membrane.

Investigating recurrent neural networks for OCT A-scan based tissue analysis.

OBJECTIVES: Optical Coherence Tomography (OCT) has been proposed as a high resolution image modality to guide transbronchial biopsies. In this study we address the question, whether individual A-scans obtained in needle direction can contribute to the identification of pulmonary nodules. METHODS: OCT A-scans from freshly resected human lung tissue specimen were recorded through a customized needle with an embedded optical fiber. Bidirectional Long Short Term Memory networks (BLSTMs) were trained on randomly distributed training and test sets of the acquired A-scans. Patient specific training and different pre-processing steps were evaluated. RESULTS: Classification rates from 67.5% up to 76% were archived for different training scenarios. Sensitivity and specificity were highest for a patient specific training with 0.87 and 0.85. Low pass filtering decreased the accuracy from 73.2% on a reference distribution to 62.2% for higher cutoff frequencies and to 56% for lower cutoff frequencies. CONCLUSION: The results indicate that a grey value based classification is feasible and may provide additional information for diagnosis and navigation. Furthermore, the experiments show patient specific signal properties and indicate that the lower and upper parts of the frequency spectrum contribute to the classification.

Intravital multidimensional real-time imaging of the conjunctival immune system.

The conjunctiva, as a peripheral mucosal surface, is dependent on the migration of immune cells to facilitate an orchestrated immune response. So far, only limited data to visualize these dynamics directly have been obtained, mainly due to technical and experimental restrictions. To investigate migration on a cellular level, the following conditions need to be met: (1) intravital investigations need to be facilitated by suitable microscopic techniques; (2) tissues need to be investigated in three spatial dimensions and over time; (3) data need to contain detailed information about the tissue character. Whereas the use of confocal laser scanning microscopy allows high-resolution imaging of the superficial conjunctival immune system and enables the recording of rapid cellular migration, intravital two-photon microscopy further enables tracking of individual cells and characterization of cells and structures with unique optical features using autofluorescence detection, fluorescence lifetime measurements and second harmonic generation in deep tissue. Based on current results and experimental studies, two-photon microscopy has the potential for general use in basic research and clinical practice, and would greatly enhance possibilities for diagnosing and analyzing inflammatory processes of the ocular surface. In particular, inflammation in common diseases, such as allergy and dry eye, and its progress under treatment could be investigated in detail.

Imaging of human brain tumor tissue by near-infrared laser coherence tomography.

INTRODUCTION: Intraoperative detection of residual tumor remains an important challenge in surgery to treat gliomas. New developments in optical techniques offer non-invasive high-resolution imaging that may integrate well into the workflow of neurosurgical operations. Using an intracranial glioma model, we have recently shown that time domain optical coherence tomography (OCT) allows discrimination of normal brain, diffusely invaded brain tissue, and solid tumor. OCT imaging allowed acquisition of 2D and 3D data arrays for multiplanar analysis of the tumor to brain interface. In this study we have analyzed biopsy specimens of human brain tumors and we present the first feasibility study of intraoperative OCT and post-image acquisition processing for non-invasive imaging of the brain and brain tumor. METHODS: We used a Sirius 713 Tomograph with a superluminescence diode emitting light at a near infrared central wavelength of 1,310 nm and a coherence length of 15 microm. The light is passed through an optical mono mode fiber to a modified OCT adapter containing a lens system with a working distance of 10 cm and an integrated pilot laser. Navigation-registered tumor biopsies were imaged ex vivo and the intraoperative site of optical tissue analysis was registered by marker acquisition using a neuronavigation system. RESULTS: Optical coherence tomography non-contact measurements of brain and brain tumor tissue produced B-scan images of 4 mm in width and 1.5-2.0 mm in depth at an axial and lateral optical resolution of 15 microm. OCT imaging demonstrated a different microstructure and characteristic signal attenuation profiles of tumor versus normal brain. Post-image acquisition processing and automated detection of the tissue to air interface was used to realign A-scans to compensate for image distortions caused by pulse- and respiration-induced movements of the target volume. Realigned images allowed monitoring of intensity changes within the scan line and facilitated selection of areas for the averaging of A-scans and the calculation of attenuation coefficients for specific regions of interest. CONCLUSION: This feasibility study has demonstrated that OCT analysis of the tissue microstructure and light attenuation characteristics discriminate normal brain, areas of tumor infiltrated brain, solid tumor, and necrosis. The working distance of the OCT adapter and the A-scan acquisition rate conceptually allows integration of the OCT applicator into the optical path of the operating microscopes. This would allow a continuous analysis of the resection plain, providing optical tomography, thereby adding a third dimension to the microscopic view and information on the light attenuation characteristics of the tissue.

[Optical coherence tomography in middle ear surgery]. / Optische Kohärenztomographie in der Mittelohrchirurgie.

The application of optical coherence tomography (OCT) in middle ear surgery has a high diagnostic potential, especially for intraoperative evaluation of the cause of stapes fixation, intraoperative assessment of the morphology of the stapes footplate in revision stapes surgery, and as an orientation guide in cochlear implantation in congenital anomalies. OCT displays the middle and inner ear structures precisely. This technology enables the surgeon to use this information for further specification of the intraoperative modus operandi. Considering our own in vivo and ex vivo investigations and animal experiments, potential areas of application can be defined: visualization of the oval window niche in revision stapesplasty and reconstructive middle ear surgery, as well as during explorative tympanotomy for intraoperative assessment of perilymph fistula, and demonstration of structures of the exposed but not opened inner ear.

Intra-operative application of optical coherence tomography with an operating microscope.

OBJECTIVE: To introduce the use of optical coherence tomography with an operating microscope for intra-operative evaluation of the human larynx. METHODS: A specially equipped operating microscope with integrated spectral domain optical coherence tomography apparatus was used during microlaryngoscopy. RESULTS: Technical improvements in optical coherence tomography equipment (e.g. pilot beam, variable focal distance, improved image quality and integration into an operating microscope) have enabled greater sensitivity and imaging speed and a non-contact approach. Spectral domain optical coherence tomography now enables a better correlation between optical coherence tomography images and histological findings. With this new technology, the precision of biopsy can be improved during microlaryngoscopy. CONCLUSIONS: Use of this new optical coherence tomography technology, integrated into an operating microscope, enables the surgeon to define the biopsy site location and resection plane precisely, while the optical zoom of the operating microscope can be used over the complete range.

Optical coherence tomography of the oval window niche.

OBJECTIVE: Optical coherence tomography was used to study the stapes footplate, both in cadaveric temporal bones and during middle-ear surgery. MATERIALS AND METHODS: Optical coherence tomography was conducted on five temporal bone preparations (from two children and three adults) and in eight patients during middle-ear surgery. A specially equipped operating microscope with integrated spectral domain optical coherence tomography apparatus was used for standard middle-ear surgical procedures. RESULTS: This optical coherence tomography investigation enabled in vivo visualisation and documentation of the annular ligament, the different layers of the footplate and the inner-ear structures, both in non-fixed and fixed stapes footplates. In cases of otosclerosis and tympanosclerosis, an inhomogeneous and irregularly thickened footplate was found, in contrast to the appearance of non-fixed footplates. In both fixed and non-fixed footplates, there was a lack of visualisation of the border between the footplate and the otic capsule. CONCLUSIONS: Investigation of the relatively new technology of optical coherence tomography indicated that this imaging modality may assist the ear surgeon to assess the oval window niche intra-operatively.

[Optical coherence tomography: from retina imaging to intraoperative use - a review]. / Ubersicht der apparativen Entwicklungen in der optischen Kohärenztomografie: von der Darstellung der Retina zur Unterstützung therapeutischer Eingriffe.

BACKGROUND: Optical coherence tomography (OCT) is new diagnostic procedure that has rapidly evolved in the last years. The recently developed spectral domain OCT allows one to increase the imaging speed by a hundred times compared to the first generation time domain OCT and enables three-dimensional imaging as well as real-time imaging of fast moving structures. Volumetric imaging improves the quantitative measurement of morphology and the evaluation of temporal changes. In addition, an exact correlation with images acquired with other imaging modalities is possible. Real-time imaging enables also the use of OCT during examinations with the slit-lamp and during ophthalmological surgery. METHODS: A spectral domain OCT was adapted to a slit lamp. 70 patients (91 eyes) were examined at the anterior or posterior segment of the eye. Images of healthy structures and different pathologies were compared to OCT images obtained with Stratus 3, Spectralis, and the SL-OCT. To demonstrate the feasibility of OCT during surgery, spectral domain OCT devices working with 20,000 and 210,000 A scans per second were coupled by specially developed optics to the camera port of a surgical microscope. The device was tested with phantoms and enucleated pig eyes. RESULTS AND CONCLUSIONS: A 5 kHz spectral domain OCT can image the retina during slit lamp-based indirect ophthalmoscopy with a quality similar to that of the Stratus 3. In addition, relevant structures of the anterior segment were imaged. Here, compared to commercially available devices, the imaged field was smaller and the angle of the anterior chamber was not directly visible due to the 830 nm wavelength, which had to be used for retinal imaging. Through the surgical microscope, a volumetric imaging of epithelium, Bowman's, Descemet's membranes, limbus, iris, lens, conjunctiva and sclera was demonstrated with several tens of centimetre working distance. Instruments and incisions in the cornea were visualised with 20 microm precision. Real-time imaging and visualisation of volumetric OCT data were also demonstrated. In principle, all technical problems of an intraoperative use of OCT have been solved and a clinical trial will start in the near future. OCT has the potential to improve the precision of surgical interventions and may even enable new interventions.

[Intraoperative application of optical coherence-tomography (OCT) for visualization of the oval window niche]. / Intraoperative Anwendung der Optischen Kohärenz-Tomografie (OCT) zur Darstellung der ovalen Fensternische.

BACKGROUND: CT and MRI of the windows of the temporal bone have become an important tool in the analysis of malformation, trauma and chronic otitis media. Optical Coherence-Tomography (OCT) provides optical cross-sections of the tissue, comparable to ultrasound. In a study on temporal bone specimens and during middle ear surgery we tested, whether OCT provides information about the oval window niche. MATERIAL AND METHODS: OCT was performed ex vivo on five human temporal bone preparations, in which the oval window niche was exposed and in vivo in eight patients who underwent middle ear surgery. A new OCT device, which based on spectrally resolved detection of the interference signals, was used. This OCT technology was fully integrated into an operating microscope. For direct correlation between OCT-scans and histology, three temporal bones were used. RESULTS: On all ex and in-vivo scans OCT supplies information about morphology of the stapes footplate. This OCT investigation documents the possibility to visualize in vivo annular ligament, different layers of the FP and inner ear structures. CONCLUSIONS: Intra-operative OCT application will help to visualize FP anomalies. Our study provides morphological information of the FP that may help in stapes surgery of the patients concerned.

[Imaging of Cochlear Structures by Optical Coherence Tomography (OCT). Temporal bone experiments for an OCT-guided cochleostomy technique]. / Darstellung cochleärer Binnenstrukturen durch Optische Kohärenz-Tomografie (OCT).

BACKGROUND: Optical Coherence Tomography (OCT) is a relatively new imaging technique, which provides scans similar to sonography on an optical base. We questioned whether OCT may be helpful in optimizing anatomical orientation in cochlear implant (CI) surgery. In a study on temporal bone specimens we tested, whether OCT provides information about the cochlear topography, particularly in situations, when for cochleostomy the bony otic capsule is already opened but the membranous endosteal layer is still intact. MATERIAL AND METHODS: OCT was performed on five human temporal bone preparations, in which the cochleostomy was carried out still leaving the endosteum covering the fluid-filled inner ear intact. A prototype of operating microscope was used, in which a spectral-domain OCT (SD-OCT) with a central wavelength of 840 nm was integrated. RESULTS: On all scans, OCT supplies information about inner ear structures, such as the lateral attachment of the basilar membrane dividing the scalae. Even delicate structures like the Reissner's membrane could be identified in one case. CONCLUSIONS: This pilot study clearly documents the possibility to identify inner ear structures, especially the site of the scala tympani while its enveloping membranes are still intact. These findings may have an impact on cochlear implant surgery, especially as an orientation guide to localize the scala tympani precisely before opening the fluid-filled inner ear.

Optically controlled thermal management on the nanometer length scale.

The manipulation of polymers and biological molecules or the control of chemical reactions on a nanometer scale by means of laser pulses shows great promise for applications in modern nanotechnology, biotechnology, molecular medicine or chemistry. A controllable, parallel, highly efficient and very local heat conversion of the incident laser light into metal nanoparticles without ablation or fragmentation provides the means for a tool like a 'nanoreactor', a 'nanowelder', a 'nanocrystallizer' or a 'nanodesorber'. In this paper we explain theoretically and show experimentally the interaction of laser radiation with gold nanoparticles on a polymethylmethacrylate (PMMA) layer (one-photon excitation) by means of different laser pulse lengths, wavelengths and pulse repetition rates. To the best of our knowledge this is the first report showing the possibility of highly local (in a 40 nm range) regulated heat insertion into the nanoparticle and its surroundings without ablation of the gold nanoparticles. In an earlier paper we showed that near-infrared femtosecond irradiation can cut labeled DNA sequences in metaphase chromosomes below the diffraction-limited spot size. Now, we use gold as well as silver-enhanced gold nanoparticles on DNA (also within chromosomes) as energy coupling objects for femtosecond laser irradiation with single-and two-photon excitation. We show the results of highly localized destruction effects on DNA that occur only nearby the nanoparticles.

Chromophore-assisted light inactivation of pKi-67 leads to inhibition of ribosomal RNA synthesis.

OBJECTIVES: Expression of the nuclear Ki-67 protein (pKi-67) is strongly associated with cell proliferation. For this reason, antibodies against this protein are widely used as prognostic tools for the assessment of cell proliferation in biopsies from cancer patients. Despite this broad application in histopathology, functional evidence for the physiological role of pKi-67 is still missing. Recently, we proposed a function of pKi-67 in the early steps of ribosomal RNA (rRNA) synthesis. Here, we have examined the involvement of pKi-67 in this process by photochemical inhibition using chromophore-assisted light inactivation (CALI). MATERIALS AND METHODS: Anti-pKi-67 antibodies were labelled with the fluorochrome fluorescein 5(6)-isothiocyanate and were irradiated after binding to their target protein. RESULTS: Performing CALI in vitro on cell lysates led to specific cross-linking of pKi-67. Moreover, the upstream binding factor (UBF) necessary for rRNA transcription was also partly subjected to cross-link formation, indicating a close spatial proximity of UBF and pKi-67. CALI in living cells, using micro-injected antibody, caused a striking relocalization of UBF from foci within the nucleoli to spots located at the nucleolar rim or within the nucleoplasm. pKi-67-CALI resulted in dramatic inhibition of RNA polymerase I-dependent nucleolar rRNA synthesis, whereas RNA polymerase II-dependent nucleoplasmic RNA synthesis remained almost unaltered. CONCLUSIONS: Our data presented here argue for a crucial role of pKi-67 in RNA polymerase I-dependent nucleolar rRNA synthesis.

Evaluation of cyclophotocoagulation effects with 1310-nm contact optical coherence tomography.

PURPOSE: The aim of this pilot study was to evaluate whether contact optical coherence tomography (OCT) allows visualization of the effects of cyclophotocoagulation. METHODS: In this pilot study, transscleral contact OCT images (1310-nm wavelength) were generated prior to and immediately after conventional transscleral diode laser cyclophotocoagulation in three eyes of three patients who were suffering from uncontrolled glaucoma. RESULTS: In the region of the ciliary body, transscleral contact OCT revealed two layers: (i) a superficial thick hyperreflective complex representing conjunctiva, the Tenon capsule, episclera, and sclera; and (ii) a thinner hyporeflective layer representing the ciliary body. The ciliary body could be differentiated from the overlying sclera by its marked drop in reflectivity. After cyclophotocoagulation, a marked increase of reflectivity in the treated area of the ciliary body was identifiable. After treatment, the distinct border between the hyperreflective scleral complex disappeared, and the region of the ciliary body appeared hyperreflective. The optical properties of the overlying sclera remained unchanged. On corresponding averaged A-scan images, scleral thickness appeared to be slightly increased, whereas ciliary body thickness remained unchanged. CONCLUSIONS: This pilot study demonstrates the capability of contact OCT for allowing visualization of changes in the ciliary body after transscleral cyclophotocoagulation (TSCPC). Further investigations are planned to clarify the complete significance of these data.

Real-time imaging of transscleral diode laser cyclophotocoagulation by optical coherence tomography.

PURPOSE: To present optical coherence tomography (OCT) for real-time imaging of cyclophotocoagulation effects. METHODS: In a pilot study, real-time transscleral OCT images were generated during diode laser cyclophotocoagulation in four eyes of four patients suffering from uncontrolled glaucoma using a specially designed contact applicator containing the OCT fiber, a focussing fiber optic and the fiber of the diode laser. RESULTS: When the contact system was used, two layers could be differentiated: a superficial thick hyperreflective complex representing conjunctiva, Tenon's capsule, episclera and sclera, and a thinner hyporeflective layer representing the ciliary body. During cyclophotocoagulation, real-time OCT showed a clear and sudden thickening of the ciliary body in the treated area. CONCLUSION: This new OCT device represents a first step towards visual, real-time imaging of cyclophotocoagulation. After further adaptation of the delivery system, further trials are needed to correlate OCT findings with aqueous production and intraocular pressure.

Optical coherence tomography for experimental neuroendoscopy.

Optical coherence tomography (OCT) is a non-invasive and non-contact imaging technology that has been applied to several biomedical applications. We have recently demonstrated that OCT allows discrimination of tumor adjacent brain, diffuse and solid tumor tissue and that this technology may be used to detect residual tumor within the resection cavity during resection of intrinsic brain tumors. Here we show that an OCT integrated endoscope can image the endoventricular anatomy and other endoscopically accessible structures in a human brain specimen. A Sirius 713 optical coherence tomography device was mounted to a modified rigid endoscope. A formalin-fixed human brain specimen was used to simulate endoscopic visualization of brain anatomy and two specimens of fixed malignant tumors with endoventricular growth patterns. Simultaneous OCT imaging and endoscopic video imaging of the visible spectrum was possible using a graded index rod endoscope. OCT imaging of a human brain specimen in water allowed an in-depth view into structures like the walls of the ventricular system, the choroid plexus or the thalamostriatal vein. OCT further allowed imaging of structures beyond tissue barriers or opaque media. In this fixed specimen OCT allowed discrimination of vascular structures down to a diameter of 50 mum. In vessels larger that 100 mum the lumen could be discriminated and within larger blood vessels a layered structure of the vascular wall as well as endovascular plaques could be visualized. This in vitro pilot study has demonstrated that OCT integrated into neuroendoscopes may add information that cannot be obtained by the video imaging alone. This technology may provide an extra margin of safety by providing cross-sectional images of tissue barriers within optically opaque conditions.