Otitis media is one of the most common reasons for pediatrician visits, antibiotic prescription, and surgery in the pediatric population. Visible light pneumatic-otoscopy is considered the best currently available diagnostic tool for otitis media. However, it has various limitations e.g. the disposable speculum cannot create an adequate seal against the external auditory canal to obtain tympanic membrane movement. Also, lack of training for effective pneumatic-otoscopy for most clinicians is another factor. To overcome these limitations, we have recently developed an otoscope sensitive to shortwave infrared (SWIR) wavelengths of light. A SWIR otoscope could help identify middle-ear-effusions based on the strong light absorption by ear fluid. Due to a longer wavelength, light can penetrate deeper through tissue, enabling a better view behind the tympanic membrane. Here we present our preliminary findings on the feasibility of using video rate SWIR imaging in a pediatric population. A total of 74 ear video recordings were obtained in the study from 20 patients. There was an improvement in the ability to see through the tympanic membrane using the SWIR otoscope. Three patients with middle ear effusion, confirmed by pneumatic otoscopy, were all identified using both visible and SWIR otoscopy. The average contrast for visible otoscopy in the presence of middle ear effusion was 0.097 and for SWIR was 0.29. In tympanic membranes with myringosclerosis, neither technique was able to see through affected areas. However, the SWIR otoscope was able to see through dried blood, dried secretions and thin dry areas of cerumen overlying the tympanic membrane.

The tympanic membrane (TM) and ossicular chain play a central role in hearing by providing acoustic impedance matching between the air-filled ear canal and the fluid-filled inner ear. Vibrometric measurement of the ossicles and TM has been critical for advancing our understanding of the hearing mechanics and improving treatments such as middle-ear prosthetics. It also holds promise for diagnosis of ossicular disorders and planning surgical interventions. Phase-sensitive optical coherence tomography (OCT) is a promising tool in hearing research and otology because it can simultaneously image the anatomical structure of the middle ear and measure sound transduction along the TM and ossicular chain with nanometer level sensitivity. Up to now, the demonstrations of OCT-based middle ear vibrometry have been largely focused on vibration magnitude, and vibration phase has been generally overlooked. Here we show OCT vibrography, in which the data acquisition is synchronized with sound excitation and beam scanning, is well suited for volumetric, vibrational imaging of the ossicles and TM. The acquired vibrography data provide intuitive motion pictures of the ossicular chain and how they vary with sound frequency. We investigated the chinchilla ear over 100 Hz to 15 kHz. The vibrography images reveal a previously undescribed mode of motion of the chinchilla ossicles at high frequencies, involving the rotation of the ossicular chain around a secondary axis parallel to the manubrium. We also found evidence of bending and torsion of the manubrium.

Significant technical and optical advancements are required for intraoperative optical coherence tomography (OCT) to perform boundless surgical applications in otology, since the translation of OCT for surgical-microscope facilitates the simultaneous OCT and microscopic visualization of soft tissue structures of the surgical region with a high-resolution in real-time. Herein, we developed an augmented-reality intrasurgical OCT microscope system with an extended 280 mm working distance, which simply provides a sufficient space for the manipulation during surgeries compared to conventional techniques. Ex vivo experiments were initially performed to evaluate the enhanced system performances and secondly, the developed system was well-utilized to clinically assess the preliminary findings of tympanomastoidectomy in six patients with chronic otitis media. The OCT system evaluated the residual inflammation of region of interest in the mastoid bone and most importantly OCT was sufficiently useful for visualizing the connection between the graft and remnant tympanic membrane intraoperatively. Use of this extended-working distance OCT integrated surgical-microscope enables the surgeons to precisely define the aforementioned surgical requirements, while performing intraoperative imaging over the complete range.

Inner ear imaging is important for the assessment of hearing disorders. A major cause of hearing loss is the damage to the sensory hair cells, which are located inside the cochlea, a spiral-shaped bone in the inner ear. Imaging of intracochear hair cells is of high interest because it can provide precise diagnosis and treatment of hearing loss. However, this goal is very challenging because the cochlea is very small and enclosed by a dense bone, thus preventing visualization of intracochlear microanatomy. In this paper, we present a novel technique for imaging cochlear cells through the bone by two-photon microscopy. We optimized the imaging quality by thinning the obscuring scattering bone above the hair cells using a femtosecond laser. We controlled the ablation with an optical coherence tomography system and a bright-field camera for real time visualization. The proposed method enables optical access to the cochlea by thinning the cochlear bone, thus allowing imaging of cells underneath.

Cerebral edema, as an acute head injury, has been of great interest over decades in clinical neurosurgery research. Many patients exhibited extensor motor abnormalities in the acute phase because their cerebral lose control of brainstem and spinal cord. These phenomenon, which is called “decerebrate rigidity”, showed up spontaneously or when their bodies were stimulated by external forces. Neuro imaging methods provide a new perspective in cerebral edema with decerebrate rigidity. Functional near infrared spectroscopy (fNIRS), as a novel optical detection method, has been widely used in monitoring changes of cerebral cortex blood oxygen concentration. Here, we utilized a self-made fNIRS detecting instrument with 4 channels fixed in forehead to monitor cerebral hemodynamic changes of patients with cerebral edema. The instrument has three waveforms (735nm, 805nm and 850nm) to detect concentration changes of oxygenated hemoglobin (HbO, deoxy hemoglobin (Hb), and blood volume (HbT). During the monitoring period, we captured two typical decerebrate rigidity in a patient (caused by external stimulation). At the beginning of decerebrate rigidity, Hb and HbT increased rapidly along with HbO decreased. The stage of this change lasts about 3s. Then the three hemodynamic parameters keep stable for about 6s. Finally, the three hemodynamic parameters return to the initial stage slowly (about 10s). The change amplitude of decerebrate rigidity caused by external stimulation were significantly higher than spontaneous situation. The results indicate that decerebrate rigidity caused acute hypoxia and congestion in patients’ cerebral cortex.

Mechanical ventilation is a critical intervention given to intensive care unit (ICU) patients who need airway support. However, this intervention with an endotracheal tube (ETT) is associated with complications such as ventilator-associated pneumonia (VAP). VAP is reported to develop within 48 hours after intubation and is associated with a mortality rate between 20 to 50%. The formation of bacterial biofilms within these ETT tubes provides a niche for infectious bacteria to become resistant to antibiotics. Suctioning of the ETT is believed to prevent airway colonization by pathogens, reduce resistance to airflow, and decrease biofilm formation. However, reports have shown that standard-of-care suctioning is not adequate to eliminate secretions from the ETT, and additional measures aiming to reduce the formation of ETT biofilms have been proposed to reduce VAP. We have recently demonstrated the use of catheter-based 3-D OCT imaging to identify the presence of in vivo biofilms within the ETTs of intubated human subjects in the ICU. In this study, we quantify the volume of mucus and biofilm in ETTs in intubated ICU patients using 3-D OCT, and define the efficacy of suctioning. Longitudinal OCT imaging was performed daily before and after suctioning at approximately 24-hour intervals until extubation. Extubated ETTs were subsequently imaged for further analysis. OCT image analysis results were correlated with clinical data and fluorescence microscopy/Gram stain images to verify the presence of bacteria and biofilm. In vivo catheter-based 3-D OCT offers the potential to rapidly determine the efficacy of ETT suctioning in order to effectively compare suctioning and brushing strategies in an effort to reduce the incidence of VAP.

Ciliary motion in the upper airway is the primary mechanism by which the body transports foreign particulate out of the respiratory system in order to maintain proper respiratory function. The ciliary beating frequency (CBF) is often disrupted with the onset of disease as well as other conditions, such as changes in temperature or in response to drug administration. Current imaging of ciliary motion relies on microscopy and high speed cameras, which cannot be easily adapted to in-vivo imaging. M-mode optical coherence tomography (OCT) imaging is capable of visualization of ciliary activity, and phase-resolved Doppler (PRD) algorithm can be integrated to measure the ciliary beating direction and amplitude with nanometer sensitivity. However, since ciliary activity naturally happens on the tissue surface, enface imaging modalities should be more suitable than cross-sectional ones such as OCT. We report on the development of a spectrally encoded interferometric microscopy (SEIM) system using a phase-resolved Doppler (PRD) algorithm to measure and map the ciliary beating frequency within an en face region. This novel high speed, high resolution system allows for visualization of both temporal and spatial ciliary motion patterns with nanometer sensitivity. Rabbit tracheal CBF ranging from 9 to 13 Hz have been observed under different temperature conditions, and the effects of using lidocaine and albuterol have also been measured. This study is the stepping stone to in-vivo studies and the translation of imaging spatial CBF in clinics.

Chronic-rhinosinusitis (CRS) is one of the most common conditions affecting ~14.2% (29.2-million) of US adults leading to estimated 18-22 million-physician office visits. It causes significant physical symptoms, negatively affects the quality-of-life and can substantially impair daily functioning. Various factors including microorganisms, allergies, and other inflammatory triggers play role in CRS. Lack of a universal marker and acknowledged difficulty in establishing the causes for the condition contributes to the poor treatment strategies and outcomes associated with CRS. Utilizing panel of sensitive markers associated with inflammatory responses in the nasal area can provide clinicians valuable information about the disease at the molecular level. The present study aims at identifying spectrochemical markers associated with the onset of CRS using data-driven Raman imaging. By combining high-resolution Raman imaging and machine learning we have developed a novel approach to obtain an integrated insight. Our findings are suggestive of differential changes in the biochemical composition of nasal tissues with CRS onset. A regression-based framework has been developed to link the inflammation score with spectral features. Support vector machine has been employed to explore the feasibility of classification. Successful recognition of these markers in nasal tissues will be helpful not only in designing automated diagnosis platforms but can also be used for identifying novel treatment strategies. Findings of this study will also serve as the foundation of our future research work on evaluating the applicability of nasal lavage for a minimally invasive method for objective CRS diagnosis.

Cartilage is the best nature material for transplantation. Laser reshaping of cartilage allows obtaining any desirable shape of the implant. This work continues the interdisciplinary studies on laser-induced development of stable costal cartilage implants, the most promising biological object for the manufacture of implants (grafts) used in otolaryngology and maxillofacial surgery, in particular, in recovering tracheal defects for the treatment of larynx stenosis. We used 1560 nm fiber laser with feedback control system measuring tissue temperature in the course of laser irradiation.

The thermomechanical IR-laser effect on reshaping process for ENT is studied. The possibility and conditions of the laser reshaping of costal cartilage for stable human implants are investigated. The causes of nonlinear thermomechanical behavior of cartilage and its influence on controlled reshaping and stability of costal cartilage for ENT are discussed. Clinical trials for 5 patients with one and a half year follow-up observation demonstrate stable positive results.

Optical coherence tomography (OCT) enables non-invasive depth-resolved investigation of laryngeal tissue. However, with conventional systems, OCT cross-sectional images of vibrating vocal cords always suffer from motion artifacts. This is the case even at low phonation frequencies of about 100 Hz. Motion artifacts of predictable repetitive movements can be avoided with carefully timed acquisitions. Irregular, non-repetitive movements, e.g. disturbed vocal cord vibration caused by laryngeal disorders, require different strategies, such as the use of high frame rates. We present a novel concept for dynamic vocal cord imaging with a high speed 1.6 MHz swept-source OCT system. Due to the high image rate, a graphics processing unit (GPU) based signal processing software has been developed in order to obtain real time OCT images. To demonstrate the feasibility of our approach on vibrating samples, we present a laboratory setup which includes a MHz swept source for OCT. To enable the transfer of our setup to clinical applications a concept for a curved rigid laryngoscope design, integrating the optical components for high-speed OCT, is proposed.

Studies investigating relationship between tobacco withdrawal/discontinuation (tobacco-quitting habit) and risk of cancer-progression are usually retrospective. Hamster-buccal-pouch (HBP) develops tumors in 14 weeks on topicalapplication (painting) of carcinogen 7,12-Dimethylbenz[a]anthracene (DMBA). In this study HBP model was employed to investigate cancer-progression post 8-week-DMBA-application employing in vivo Raman spectroscopy (RS). 20 animals (6–8 weeks) were randomized into 4 sets; spectra were acquired at the end of 0, 8 and 14 weeks from each animal. Set1: DMBA-painting was carried out for 14-weeks. Set2: DMBA-painting was discontinued after 8-weeks of painting (analogous to tobacco-quitting). Set 3: Vehicle control—only vehicle was applied. Set 4: Independent set of animals with DMBA-painting for 14-weeks; to compare spectra from sets 1-3. DMBA painting involved application of 0.5% DMBA dissolved in vegetable oil (vehicle). Multivariate analysis revealed cancer-progression in spectra from 8 and 14 weeks of continuous DMBA-painting however, in case of DMBA painting discontinued after 8 weeks, but spectra acquired after 14 weeks, only 28% spectra classified with tumors. Visible tumours were not observed but histopathological evaluation suggested higher pathologies (carcinoma-in-situ) in Set 2 animals. Observations suggest a cause-and-consequence effect between DMBA-discontinuation and reduced cancer-progression over a defined timeperiod. Findings can be extrapolated to tobacco habitués to suggest reduced risk of cancer progression on quitting primary consumption of tobacco. Findings further suggest RS as a prospective label-free and noninvasive candidate tool to screen risk-prone population.

Bioimaging of fluorescent probes provides a fantastic tool to visualize different cellular responses from various cells and to monitor target/probe interaction in cells. Previously, we introduced a small molecule named 3,6-bis(1-methyl-4- vinylpyridinium)carbazole diiodide (BMVC) as a fluorescent marker to light up live cancer cells for cancer diagnosis. Here we used time-gated fluorescence lifetime imaging microscopy (FLIM) of a BMVC isomer, 3,6-bis(1-methyl-2- vinylpyridinium)carbazole diiodide (o-BMVC) as a fluorescent probe to distinguish fixed cancer cells for cancer diagnosis. The time-gated FLIM results showed many more number of o-BMVC foci, characterized by the longer fluorescent decay time of o-BMVC (≥2.4 ns), in fixed cancer cells (HeLa, H1299, CL1-0, MCF-7, MCF-7/ADR, and SAS) than in fixed normal cells (MRC-5, IMR-90, and BJ). Further study of tissue biopsy, a total of 50 head and neck cancer (HNC) samples obtained during surgery and 20 normal oral samples collected from healthy volunteers showed that o-BMVC foci are hardly detectable in the normal oral epithelial cells. The average numbers of o-BMVC foci in tumors and in normal oral epithelial cells are 28.3 and 2.2, respectively. The receiver operating characteristic curve analysis showed the area under curve was 0.992, indicating that this method provides a very high accuracy for clinical detection of HNC cancers. We propose that the combination of BMVC test for live cells and o-BMVC test for fixed cells could provide a powerful tool for screening of human cancers.

We present our 2nd generation handheld simultaneous multispectral frequency-domain FLIM endoscopic system for label-free metabolic imaging of oral cancer, with enhanced optical performance and system usability. Our custom-designed and 3D-printed handheld endoscope consists of an enclosure (6 x 3 x 3 cm3) with a rigid probe (1 cm diameter, 9 cm length) that weighs less than 125 g with all the system components, which, compared to our previous system, is significantly smaller and lighter, and has improved ergonomics and usability. The enclosure has mounts for a dual axes bi-directional MEMS scanner and a dichroic mirror, and plug-and-play ports for excitation, emission collection and rigid probe optics. The rigid probe used for oral mucosa imaging contains a three-lens imaging system that, compared to our previous system, has: an increased field of view (FOV) (6 x 6 mm2 vs. 16 x 16 mm2), improved lateral resolution (36 μm in the center and 65 μm at the edge, diffraction-limited performance across a central ± 5.5 mm field), and an extended working distance (10 mm vs. 40 mm). A 375 nm CW laser is used as the excitation source, and fluorescence emission is spectrally divided into three emission bands (405±10 nm, 440±20 nm, and 525±25 nm) targeting collagen, NADH, and FAD, which are relevant for early detection of oral cancer. Fluorescence emission is then detected by three APDs and further processed onboard an FPGA. Our clinically compatible handheld endoscope allows for noninvasive and fast in situ clinical metabolic imaging of the oral mucosa.

Head and neck cancers overwhelmingly overexpress epidermal growth factor receptor (EGFR). This overexpression has been utilized for head and neck cancers using molecular targeted agents for therapy and cancer cell detection. Significant progress has been made in using EGFR-targeted fluorescent antibody and Affibody molecule agents for fluorescent guided surgery in head and neck cancers. Although success in achieving tumor-to-background ratio of 3-5 have been achieved, the field is limited by the non-specific fluorescence in normal tissues as well as EGFR specific fluorescence in the oral cavity. We propose that paired-agent imaging (PAI) could improve the contrast between tumor and normal tissue by removing the fluorescent signal arising from non-specific binding. Here, ABY-029 – an anti-EGFR Affibody molecule labeled with IRDye 800CW – and IRDye 680RD conjugated to Affibody Control Imaging Agent molecule (IR680-Aff_ctrl) are used as targeted and untargeted control agents, respectively, in a panel of head and neck squamous cell carcinomas (HNSCC) to test the ability of PAI to increase tumor detection. Initial results demonstrate that binding potential, a value proportional to receptor concentration, correlates well to EGFR expression but experimental limitations prevented pixel-by-pixel analysis that was desired. Although promising, a more rigorous and well-defined experimental protocol is required to align ex vivo EGFR immunohistochemistry with in vivo binding potential and fluorescence intensity. Additionally, a new set of paired-agents, ABY-029 and IRDye 700DX, are successfully tested in naïve mice and will be carried forward for clinical translation.