Miniature probe for dual‐modality photoacoustic microscopy and white‐light microscopy for image guidance: A prototype toward an endoscope

In this study, a novel photoacoustic microscopy (PAM) probe integrating white‐light microscopy (WLM) modality that provides guidance for PAM imaging and complementary information is implemented. One single core of an imaging fiber bundle is employed to deliver a pulsed laser for photoacoustic excitation for PAM mode, which provides high resolution with deep penetration. Meanwhile, for WLM mode, the imaging fiber bundle is used to transmit two‐dimensional superficial images. Lateral resolution of 7.2 μm for PAM is achieved. Since miniature components are used, the probe diameter is only 1.7 mm. Imaging of phantom and animals in vivo is conducted to show the imaging capability of the probe. The probe has several advantages by introducing the WLM mode, such as being able to conveniently identify regions of interest and align the focus for PAM mode. The prototype of an endoscope shows potential to facilitate clinical photoacoustic endoscopic applications.     

Dynamic change in optical properties of a nanoparticle embedded tumor phantom for plasmonic photothermal cancer therapeutics

In this study, the temporal dynamic changes in optical properties of gold nanorods (GNR) embedded tumor phantom, during photothermal interaction, are reported for plasmonic photothermal therapeutics. Tumor mimicking bilayer phantoms were prepared by using 1% agarose incorporated with 0.1% coffee powder, 0.3% intralipid solution as epidermis layer; 3% intralipid solution and 0.3% human hemoglobin (Hb) powder as dermis layer. On incorporating GNRs of concentrations 10, 20, and 40 μg/ml within the phantom, the absorption coefficients increases 4–8 times, while there is minimal change in the reduced scattering coefficients. Further the absorption coefficient increased by ~8% with the incorporation of GNRs of concentration 40 μg/ml, while no considerable dynamic change in the optical properties is observed for the phantom embedded with GNRs of concentrations 10, and 20 μg/ml. The discussed results are useful for the selection of GNRs dose for pre-treatment planning of plasmonic photothermal cancer therapeutics.     

Optical fan for single‐cell screening

The single‐cell screening has attracted great attentions in advanced biomedicine and tissue biology, especially for the early disease diagnosis and treatment monitoring. In this work, by using a specific‐designed fiber probe with a flat facet, we propose an “optical fan” strategy to screen K562 cells at the single‐cell level from a populations of RBCs. After the 980‐nm laser beam injected into the fiber probe, the RBCs were blown away but holding target K562 cells in place. Further, multiple leukemic cells can be screened from hundreds of red blood cells, providing an efficient approach for the cell screening. The experimental results were interpreted by the numerical simulation, and the stiffness of optical fan was also discussed.     

Near-infrared emissive polymer-coated IR-820 nanoparticles assisted photothermal therapy for cervical cancer cells

Photothermal therapy (PTT) has attracted wide attention due to its noninvasiveness and its thermal ablation ability. As photothermal agents are crucial factor in PTT, those with the characteristics of biocompatibility, non-toxicity and high photothermal stability have attracted great interest. In this work, new indocyanine green (IR-820) was utilized as a photothermal agent and near-infrared (NIR) fluorescence imaging nanoprobe. To improve the biocompatibility, poly(styrene-co-maleic anhydride) (PSMA) was utilized to encapsulate the IR-820 molecules to form novel IR-820@PSMA nanoparticles (NPs). Then, the optical and thermal properties of IR-820@PSMA NPs were studied in detail. The IR-820@PSMA NPs showed excellent photothermal stability and biocompatibility. The cellular uptaking ability of the IR-820@PSMA NPs was further confirmed in HeLa cells by the NIR fluorescent confocal microscopic imaging technique. The IR-820@PSMA NPs assisted PTT of living HeLa cells was conducted under 793 nm laser excitation, and a high PTT efficiency of 73.3% was obtained.     

Femtosecond pulsed laser ablation to enhance drug delivery across the skin

Laser poration of the skin locally removes its outermost, barrier layer, and thereby provides a route for the diffusion of topically applied drugs. Ideally, no thermal damage would surround the pores created in the skin, as tissue coagulation would be expected to limit drug diffusion. Here, a femtosecond pulsed fiber laser is used to porate mammalian skin ex vivo. This first application of a hollow core negative curvature fiber (HC‐NCF) to convey a femtosecond pulsed, visible laser beam results in reproducible skin poration. The effect of applying ink to the skin surface, prior to ultra‐short pulsed ablation, has been examined and Raman spectroscopy reveals that the least, collateral thermal damage occurs in inked skin. Pre‐application of ink reduces the laser power threshold for poration, an effect attributed to the initiation of plasma formation by thermionic electron emission from the dye in the ink. Poration under these conditions significantly increases the percutaneous permeation of caffeine in vitro. Dye‐enhanced, plasma‐mediated ablation of the skin is therefore a potentially advantageous approach to enhance topical/transdermal drug absorption. The combination of a fiber laser and a HC‐NCF, capable of emitting and delivering femtosecond pulsed, visible light, may permit a compact poration device to be developed.

Using a femtosecond pulsed, visible laser beam to create an array of micropores in dyed mammalian skin, with little collateral, thermal damage, leads to an enhancement in the percutaneous permeation of caffeine in vitro.     

One Minute,Sub-One-Watt Photothermal Tumor Ablation Using Porphysomes,Intrinsic Multifunctional Nanovesicles

We recently developed porphysomes as intrinsically multifunctional nanovesicles. A photosensitizer, pyropheophorbide α, was conjugated to a phospholipid and then self-assembled to liposome-like spherical vesicles. Due to the extremely high density of porphyrin in the porphyrin-lipid bilayer, porphysomes generated large extinction coefficients, structure-dependent fluorescence self-quenching, and excellent photothermal efficacy. In our formulation, porphysomes were synthesized using high pressure extrusion, and displayed a mean particle size around 120 nm. Twenty-four hr post-intravenous injection of porphysomes, the local temperature of the tumor increased from 30 °C to 62 °C rapidly upon one minute exposure of 750 mW (1.18 W/cm²), 671 nm laser irradiation. Following the complete thermal ablation of the tumor, eschars formed and healed within 2 weeks, while in the control groups the tumors continued to grow and all reached the defined end point within 3 weeks. These data show how porphysomes can be used as potent photothermal therapy (PTT) agents.     

A readily usable two‐photon fluorescence lifetime microendoscope

A two‐photon fluorescence lifetime (2P‐FLIM) microendoscope, capable of energetic metabolism imaging through the intracellular nicotinamide adenine dinucleotide (NADH) autofluorescence, at sub‐cellular resolution, is demonstrated. It exhibits readily usable characteristics such as convenient endoscope probe diameter (≈2 mm), fiber length (>5 m) and data accumulation rate (16 frames per second (fps)), leading to a FLIM refreshing rate of ≈0.1 to 1 fps depending on the sample. The spiral scanning image formation does not influence the instrument response function (IRF) characteristics of the system. Near table‐top microscope performances are achieved through a comprehensive system including a home‐designed spectro‐temporal pulse shaper and a custom air‐silica double‐clad photonic crystal fiber, which enables to reach up to 40 mW of ≈100 fs pulses @ 760 nm with a 80 MHz repetition rate. A GRadient INdex (GRIN) lens provides a lateral resolution of 0.67 μm at the focus of the fiber probe. Intracellular NADH fluorescence lifetime data are finally acquired on cultured cells at 16 fps.      

Noninvasive multimodal imaging by integrating optical coherence tomography with autofluorescence imaging for dental applications

We report the development of an integrated multifunctional imaging system capable of providing anatomical (optical coherence tomography, OCT), functional (OCT angiography, OCTA) and molecular imaging (light‐induced autofluorescence, LIAF) for in vivo dental applications. Blue excitation light (405 nm) was used for LIAF imaging, while the OCT was powered by a 1310 nm swept laser source. A red‐green‐blue digital camera, with a 450 nm cut‐on broadband optical filter, was used for LIAF detection. The exciting light source and camera were integrated directly with the OCT scanning probe. The integrated system used two noninvasive imaging modalities to improve the speed of in vivo OCT data collection and to better target the regions of interest. The newly designed system maintained the ability to detect differences between healthy and hypomineralized teeth, identify dental biofilm and visualize the microvasculature of gingival tissue. The development of the integrated OCT‐LIAF system provides an opportunity to conduct clinical studies more efficiently, examining changes in oral conditions over time.     

Laser-assisted nucleic acid delivery: A systematic review

Gene therapy has become an effective treatment modality for some conditions. Laser light may augment or enhance gene therapy through photomechanical, photothermal, and photochemical. This review examined the evidence base for laser therapy to enhance nucleic acid transfection in mammalian cells. An electronic search of MEDLINE, Scopus, EMBASE, Web of Science, and Google Scholar was performed, covering all available years. The preferred reporting items for systematic reviews and meta-analyses guideline for systematic reviews was used for designing the study and analyzing the results. In total, 49 studies of laser irradiation for nucleic acid delivery were included. Key approaches were optoporation, photomechanical gene transfection, and photochemical internalization. Optoporation is better suited to cells in culture, photomechanical and photochemical approaches appear well suited to in vivo use. Additional studies explored the impact of photothermal for enhancing gene transfection. Each approach has merits and limitations. Augmenting nucleic acid delivery using laser irradiation is a promising method for improving gene therapy. Laser protocols can be non-invasive because of the penetration of desirable wavelengths of light, but it depends on various parameters such as power density, treatment duration, irradiation mode, etc. The current protocols show low efficiency, and there is a need for further work to optimize irradiation parameters.     

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

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

Sialolithiasis: Application parameters for an optimal laser therapy

In‐vitro experimental parametric studies of laser ablation using natural sialoliths and artificial stones have been performed toward an efficient laser treatment of sialolithiasis. Surface microstructure and water adsorption become critical for coupling high power pulsed Ho:YAG laser radiation (λ = 2080 nm, τ ～250 μsec), inducing ablative interactions and stone fragmentation. Results reveal a generic trend, with single pulse laser energy density threshold for sialolith ablative erosion at ～200 J cm^?2 (corresponding to intensity ～800 kW cm^?2) and fragmentation rates reaching ～1 mm/pulse at ～2400 J cm^?2. This process shows no saturation, suggesting that very high energy density irradiation at low pulse repetition rate is an efficient approach. Such operation facilitates rapid cooling and minimal thermal loading of the oral and maxillofacial area, thus causing negligible adverse effects. The method is expected to contribute to the establishment of an easy and optimal therapeutic protocol for sialolithiasis pathology.     

Stereotaxic endoscopy for the ocular imaging of awake,freely moving animal models

Stereotaxic instruments are increasingly used in research animals for the study of disease, but typically require restraints and anesthetic procedures. A stereotaxic head mount that enables imaging of the anterior chamber of the eye in alert and freely mobile mice is presented in this study. The head mount is fitted based on computed tomography scans and manufactured using 3D printing. The system is placed noninvasively using temporal mount bars and a snout mount, without breaking the skin or risking suffocation, while an instrument channel stabilizes the ocular probes. With a flexible micro‐endoscopic probe and a confocal scanning laser microscopy system, <20 μm resolution is achieved in vivo with a field of view of nearly 1 mm. Discomfort is minimal, and further adaptations for minimally invasive neuroscience, optogenetics and auditory studies are possible.     

Photothermal lipolysis accelerates ECM production via macrophage-derived ALOX15-mediated p38 MAPK activation in fibroblasts

Skin and subcutaneous tissue tightening is usually treated by noninvasive photothermal treatment for medical esthetics purpose, while the underlying mechanism remains to be elucidated. Here, we hypothesized that adipocyte injury, as a stimulator, may regulate extracellular matrix (ECM) production by increasing ALOX15 in macrophages, which could lead to fibroblast activation. In this study, we show that lipolysis was induced by laser heating (45°C for 15 min) in patients and rats, and adipocyte thermal injury stimulates the ECM production in fibroblasts by ALOX15 that was increased in cocultured macrophages. These phenomena were evidenced by the ALOX15 knockdown. In addition, ALOX15 metabolite 12(S)-HETE activated p38 MAPK signaling pathway that mediated the production of ECM in fibroblast. In summary, the results of this study demonstrate that the mechanisms of adipose photothermal injury-induced skin and/or subcutaneous tissue tightening may have clinical relevance for noninvasive or minimally invasive photothermal therapeutics.     

A flexible wide‐field FLIM endoscope utilising blue excitation light for label‐free contrast of tissue

Fluorescence lifetime imaging (FLIM) has previously been shown to provide contrast between normal and diseased tissue. Here we present progress towards clinical and preclinical FLIM endoscopy of tissue autofluorescence, demonstrating a flexible wide‐field endoscope that utilised a low average power blue picosecond laser diode excitation source and was able to acquire ～mm‐scale spatial maps of autofluorescence lifetimes from fresh ex vivo diseased human larynx biopsies in ～8 seconds using an average excitation power of ～0.5 mW at the specimen. To illustrate its potential for FLIM at higher acquisition rates, a higher power mode‐locked frequency doubled Ti:Sapphire laser was used to demonstrate FLIM of ex vivo mouse bowel at up to 2.5 Hz using 10 mW of average excitation power at the specimen. (© 2014 WILEY‐VCH Verlag GmbH &Co. KGaA, Weinheim)     

Photoacoustic imaging of the human placental vasculature

Minimally invasive fetal interventions require accurate imaging from inside the uterine cavity. Twin‐to‐twin transfusion syndrome (TTTS), a condition considered in this study, occurs from abnormal vascular anastomoses in the placenta that allow blood to flow unevenly between the fetuses. Currently, TTTS is treated fetoscopically by identifying the anastomosing vessels, and then performing laser photocoagulation. However, white light fetoscopy provides limited visibility of placental vasculature, which can lead to missed anastomoses or incomplete photocoagulation. Photoacoustic (PA) imaging is an alternative imaging method that provides contrast for hemoglobin, and in this study, two PA systems were used to visualize chorionic (fetal) superficial and subsurface vasculature in human placentas. The first system comprised an optical parametric oscillator for PA excitation and a 2D Fabry‐Pérot cavity ultrasound sensor; the second, light emitting diode arrays and a 1D clinical linear‐array ultrasound imaging probe. Volumetric photoacoustic images were acquired from ex vivo normal term and TTTS‐treated placentas. It was shown that superficial and subsurface branching blood vessels could be visualized to depths of approximately 7 mm, and that ablated tissue yielded negative image contrast. This study demonstrated the strong potential of PA imaging to guide minimally invasive fetal therapies.      

Photobiomodulation increases intrusion tooth movement and modulates IL‐6, IL‐8 and IL‐1β expression during orthodontically bone remodeling

This study investigated the effects of photobiomodulation (PBM) on upper molar intrusion movement, regarding acceleration of orthodontic movement and its molecular effects. The sample consisted of 30 patients with indication of tooth intrusion for oral rehabilitation. Teeth were divided into three different groups: G1 (n = 10) pre‐molars without force or laser application (control); G2 (n = 10) upper molar intrusion; and G3 (n = 10) upper molar intrusion and PBM. On PBM treated molars, the teeth were irradiated with a low‐power diode laser (808 nm, 100 mW), receiving 1 J per point, density of 25 J/cm², with application of 10 s per point, 10 points (5 per vestibular and 5 per palatal region). Orthodontic force of intrusion applied every 30 days and PBM was performed immediately, 3 and 7 days after force application for 3 months. Gingival crevicular fluid was collected at the same time periods as the laser applications and interleukins (IL) 1‐β, ‐6 and ‐8 were evaluated by enzyme‐linked immunosorbent assay. Clinical measures were performed monthly to verify the amount of intrusion. The levels of IL‐6, IL‐8 and IL‐1β increased under orthodontic force (G2 and G3) when compared to control group (G1), however, the cytokines levels were significantly higher after PBM (G3). The mean intrusion velocity was 0.26 mm/month in the irradiated group (G3), average duration of 8 months vs 0.17 mm/month for the non‐irradiated group (G2), average duration of 12 months. This study suggests that PBM accelerates tooth movement during molar intrusion, due to modulation of IL‐6, IL‐8 and IL‐1β during bone remodeling.      

Miniaturized all fiber probe for optical coherence tomography and pH detection of biological tissue

We present a novel all-fiber probe with 710-μm outside diameter for combined optical coherence tomography and pH detection. In cancer surgery, a significant challenge is how to completely remove the malignant tumor without cutting too much normal tissue. The difference between cancer tissue and normal tissue not only lies in morphology and structure but also in tissue pH, where malignant tissue has a lower pH. This dual-modality probe combined optical coherence tomography and pH detection of biological tissue, is expected to determine whether the tissue is cancerous quickly and accurately. The probe utilizes a typical three-segment structure (double-clad fiber - no-core fiber - graded-index fiber). We obtained a lateral resolution of ~10.6 μm, a working distance of ~506 μm and a pH measurement accuracy of 0.01 pH unit for the probe. The performance of the all-fiber probe was verified through an ex vivo experiment using the porcine brain specimen.     

Is oblique scanning laser ophthalmoscope applicable to human ocular optics? A feasibility study using an eye model for volumetric imaging

Oblique scanning laser ophthalmoscopy (oSLO) is a novel imaging modality to provide volumetric retinal imaging without depth sectioning over a large field of view (FOV). It has been successfully demonstrated in vivo in rodent eyes for volumetric fluorescein angiography (vFA). However, engineering oSLO for human retinal imaging is challenging because of the low numerical aperture (NA) of human ocular optics. To overcome this challenge, we implement optical designs to (a) increase the angle of the intermediate image under Scheimpflug condition, and (b) expand the magnification in the depth dimension with cylindrical lens to enable sufficient sampling density. In addition, we adopt a scanning‐and‐descaning strategy, resulting in a compact oSLO system. We experimentally show that the current setup can achieve a FOV of ~3 × 6 × 0.8 mm³, and the transverse and axial resolutions of 7 and 41 μm, respectively. This feasibility study serves an important step for future in vivo human retinal imaging.     

Front Cover: Implantable self‐aligning fiber‐optic optomechanical devices for in vivo intraocular pressure‐sensing in artificial cornea

This work addresses an unmet clinical need, that of glaucoma monitoring through intraocular pressure (IOP) interrogation in patients with artificial corneas (keratoprost hesis). We demonstrate direct integration of a low‐drift, fiber‐optic Fabry‐Perot pressure sensor embedded in the keratoprosthesis via rapid, non‐contact micromagnetic fiber alignment. IOP interrogation is achieved by using white‐light interferometry, which provides sub‐mmHg IOP sensitivity. Further details can be found in the article by Pui‐Chuen Hui, Katia Shtyrkova, Chengxin Zhou, et al. ( e202000031 ).

     

Development of hyperthermia measurable fiber radiometric thermometer for thermotherapy

Temperature monitoring is extremely important during thermotherapy. Fiber‐optic temperature sensors are preferred because of their flexibility and immunity to electromagnetic interference. Although many types of fiber‐optic sensors have been developed, clinically adopting them remains challenging. Here, we report a silica fiber‐based radiometric thermometer using a low‐cost extended InGaAs detector to detect black body radiation between 1.7 and 2.4 μm. For the first time, this silica fiber‐based thermometer is capable of measuring temperatures down to 35°C, making it suitable for monitoring hyperthermia during surgery. In particular, the thermometer has potential for seamless integration with current silica fiber catheters, which are widely used in laser interstitial thermotherapy. The feasibility, capability and sensitivity of tracking tissue temperature variation were proved through ex vivo tissue studies. After further improvement, the technology has the potential to be translated into clinics for monitoring tissue temperature.