Effect of Anesthetic Carrier Gas on In Vivo Circulation Times of Intravenously Administered Phospholipid Oxygen Microbubbles in Rats.

OBJECTIVE: For the treatment of tumor hypoxia, microbubbles comprising oxygen as a majority component of the gas core with a stabilizing shell may be used to deliver and release oxygen locally at the tumor site through ultrasound destruction. Previous work has revealed differences in circulation half-life in vivo for perfluorocarbon-filled microbubbles, typically used as ultrasound imaging contrast agents, as a function of anesthetic carrier gas. These differences in circulation time in vivo were likely due to gas diffusion as a function of anesthetic carrier gas, among other variables. This work has motivated studies to evaluate the effect of anesthetic carrier gas on oxygen microbubble circulation dynamics. METHODS: Circulation time for oxygen microbubbles was derived from ultrasound image intensity obtained during longitudinal kidney imaging. Studies were constructed for rats anesthetized on inhaled isoflurane with either pure oxygen or medical air as the anesthetic carrier gas. RESULTS: Results indicated that oxygen microbubbles were highly visible via contrast-specific imaging. Marked signal enhancement and duration differences were observed between animals breathing air and oxygen. Perhaps counterintuitively, oxygen microbubbles disappeared from circulation significantly faster when the animals were breathing pure oxygen compared with medical air. This may be explained by nitrogen counterdiffusion from blood into the bubble, effectively changing the gas composition of the core, as has been observed in perfluorocarbon core microbubbles. CONCLUSION: Our findings suggest that the apparent longevity and persistence of oxygen microbubbles in circulation may not be reflective of oxygen delivery when the animal is anesthetized breathing air.

Stepwise approach for fundus imaging in the diagnosis and management of posterior uveitis.

An array of retinochoroid imaging modalities aid in comprehensive evaluation of the immunopathological changes in the retina and choroid, forming the core component for the diagnosis and management of inflammatory disorders such as uveitis. The recent technological breakthroughs have led to the development of imaging platforms that can evaluate the layers of retina and choroid and the structural and functional alteration in these tissues. Ophthalmologists heavily rely on imaging modalities such as dye-based angiographies (fluorescein angiography and indocyanine green angiography), optical coherence tomography, fundus autofluorescence, as well as dye-less angiography such as optical coherence tomography angiograph,y for establishing a precise diagnosis and understanding the pathophysiology of the diseases. Furthermore, these tools are now being deployed with a 'multimodal' approach for swift and accurate diagnosis. In this comprehensive review, we outline the imaging platforms used for evaluation of posterior uveitis and discuss the organized, algorithmic approach for the assessment of the disorders. Additionally, we provide an insight into disease-specific characteristic pathological changes and the established strategies to rule out disorders with overlapping features on imaging.

Photoacoustic Vaporization of Endoskeletal Droplets Loaded with Zinc Naphthalocyanine.

Vaporizable endoskeletal droplets are solid hydrocarbons in liquid fluorocarbon droplets in which melting of the hydrocarbon phase leads to the vaporization of the fluorocarbon phase. In prior work, vaporization of the endoskeletal droplets was achieved thermally by heating the surrounding aqueous medium. In this work, we introduce a near-infrared (NIR) optically absorbing naphthalocyanine dye (zinc 2,11,20,29-tetra-tert-butyl-2,3-naphthalocynanine) into the solid hydrocarbon (eicosane, n-C20H42) core of liquid fluorocarbon (C5F12) drops suspended in an aqueous medium. Droplets with a uniform diameter of 11.7 ± 0.7 µm were formed using a flow-focusing microfluidic device. The solid hydrocarbon formed a crumpled spherical structure within the liquid fluorocarbon droplet. The photoactivation behavior of these dye-containing endoskeletal droplets was investigated using NIR laser irradiation. When exposed to a pulsed laser of 720 nm wavelength, the dye-containing droplets vaporized at an average laser fluence of 65 mJ/cm2, whereas blank droplets without the dye did not vaporize at any fluence up to 100 mJ/cm2. Furthermore, dye-loaded droplets with a smaller, polydisperse size distribution were prepared using a simple shaking method and studied in a flow phantom for their photoacoustic signal and ultrasound contrast imaging. These results demonstrate that dye-containing endoskeletal droplets can be made to vaporize by externally applied optical energy. Such droplets may be useful for a variety of photoacoustic applications for sensing, imaging, and therapy.

Ultrasonic fragmentation for removing thick organized clots during diabetic vitrectomy - A novel technique.

A 65-year-old male with proliferative diabetic retinopathy (PDR) and non-clearing vitreous hemorrhage underwent 25G pars plana vitrectomy (PPV). A large disk of thick organized blood of 5 disk diameter (DD) size was encountered in subhyaloid space. All attempts including lower cut rates to remove this disk using a 25G cutter turned futile. We used a 20G fragmatome to safely remove this hard clot from vitreous cavity in 50 s. Surgical time for removal of similar clot of 3 DD by 25G cutter in another eye was 5 min. Removal of thick clotted subhyaloid blood by ultrasonic fragmentation during diabetic vitrectomy is a safe, faster, and useful maneuver.

Epiretinal Membrane from Uncontrolled Gliosis After Inverted ILM Flap Technique for Idiopathic Macular Hole.

We describe two cases with epiretinal membrane (ERM) from uncontrolled gliosis seen after multilayered inverted internal limiting membrane (ILM) flap technique for full thickness macular hole (FTMH). Two patients with FTMH who had undergone surgery with inverted ILM flap technique were examined by serial optical coherence tomography scans to evaluate the course of multilayered ILM flaps seen as foveal hyperreflective lesions postoperatively. We observed excessive uncontrolled gliosis over these hyperreflective ILM flaps with ERM formation, along with worsening metamorphopsia and best-corrected visual acuity. Case 1 underwent a repeat surgery for ERM. We report excessive uncontrolled gliosis as a rare complication of multilayered inverted ILM flap technique for FTMH. [Ophthalmic Surg Lasers Imaging Retina. 2021;52:663-665.].

Detecting insulitis in type 1 diabetes with ultrasound phase-change contrast agents.

Type 1 diabetes (T1D) results from immune infiltration and destruction of insulin-producing ß cells within the pancreatic islets of Langerhans (insulitis). Early diagnosis during presymptomatic T1D would allow for therapeutic intervention prior to substantial ß-cell loss at onset. There are limited methods to track the progression of insulitis and ß-cell mass decline. During insulitis, the islet microvasculature increases permeability, such that submicron-sized particles can extravasate and accumulate within the islet microenvironment. Ultrasound is a widely deployable and cost-effective clinical imaging modality. However, conventional microbubble contrast agents are restricted to the vasculature. Submicron nanodroplet (ND) phase-change agents can be vaporized into micron-sized bubbles, serving as a microbubble precursor. We tested whether NDs extravasate into the immune-infiltrated islet microenvironment. We performed ultrasound contrast-imaging following ND infusion in nonobese diabetic (NOD) mice and NOD;Rag1ko controls and tracked diabetes development. We measured the biodistribution of fluorescently labeled NDs, with histological analysis of insulitis. Ultrasound contrast signal was elevated in the pancreas of 10-wk-old NOD mice following ND infusion and vaporization but was absent in both the noninfiltrated kidney of NOD mice and the pancreas of Rag1ko controls. High-contrast elevation also correlated with rapid diabetes onset. Elevated contrast was also observed as early as 4 wk, prior to mouse insulin autoantibody detection. In the pancreata of NOD mice, infiltrated islets and nearby exocrine tissue were selectively labeled with fluorescent NDs. Thus, contrast ultrasound imaging with ND phase-change agents can detect insulitis prior to diabetes onset. This will be important for monitoring disease progression, to guide and assess preventative therapeutic interventions for T1D.

Contrast-Enhanced Sonography with Biomimetic Lung Surfactant Nanodrops.

Nanodrops comprising a perfluorocarbon liquid core can be acoustically vaporized into echogenic microbubbles for ultrasound imaging. Packaging the microbubble in its condensed liquid state provides some advantages, including in situ activation of the acoustic signal, longer circulation persistence, and the advent of expanded diagnostic and therapeutic applications in pathologies which exhibit compromised vasculature. One obstacle to clinical translation is the inability of the limited surfactant present on the nanodrop to encapsulate the greatly expanded microbubble interface, resulting in ephemeral microbubbles with limited utility. In this study, we examine a biomimetic approach to stabilize an expanding gas surface by employing the lung surfactant replacement, beractant. Lung surfactant contains a suite of lipids and proteins that provide efficient shuttling of material from bilayer folds to the monolayer surface. We hypothesized that beractant would improve stability of acoustically vaporized microbubbles. To test this hypothesis, we characterized beractant surface dilation mechanics and revealed a novel biophysical phenomenon of rapid interfacial melting, spreading, and resolidification. We then harnessed this unique functionality to increase the stability and echogenicity of microbubbles produced after acoustic droplet vaporization for in vivo ultrasound imaging. Such biomimetic lung surfactant-stabilized nanodrops may be useful for applications in ultrasound imaging and therapy.

Acoustic Nanodrops for Biomedical Applications.

Acoustic nanodrops are designed to vaporize into ultrasound-responsive microbubbles, which presents certain challenges nonexistent for conventional nano-emulsions. The requirements of biocompatibility, vaporizability and colloidal stability has focused research on perfluorocarbons (PFCs). Shorter PFCs yield better vaporizability via their lower critical temperature, but they also dissolve more easily owing to their higher vapor pressure and solubility. Thus, acoustic nanodrops have required a tradeoff between vaporizability and colloidal stability in vivo. The recent advent of vaporizable endoskeletal droplets, which are both stable and vaporizable, may have solved this problem. The purpose of this review is to justify this premise by pointing out the beneficial properties of acoustic nanodrops, providing an analysis of vaporization and dissolution mechanisms, and reviewing current biomedical applications.

The effect of size range on ultrasound-induced translations in microbubble populations.

Microbubble translations driven by ultrasound-induced radiation forces can be beneficial for applications in ultrasound molecular imaging and drug delivery. Here, the effect of size range in microbubble populations on their translations is investigated experimentally and theoretically. The displacements within five distinct size-isolated microbubble populations are driven by a standard ultrasound-imaging probe at frequencies ranging from 3 to 7 MHz, and measured using the multi-gate spectral Doppler approach. Peak microbubble displacements, reaching up to 10 µm per pulse, are found to describe transient phenomena from the resonant proportion of each bubble population. The overall trend of the statistical behavior of the bubble displacements, quantified by the total number of identified displacements, reveals significant differences between the bubble populations as a function of the transmission frequency. A good agreement is found between the experiments and theory that includes a model parameter fit, which is further supported by separate measurements of individual microbubbles to characterize the viscoelasticity of their stabilizing lipid shell. These findings may help to tune the microbubble size distribution and ultrasound transmission parameters to optimize the radiation-force translations. They also demonstrate a simple technique to characterize the microbubble shell viscosity, the fitted model parameter, from freely floating microbubble populations using a standard ultrasound-imaging probe.

Microbubble Radiation Force-Induced Translation in Plane-Wave Versus Focused Transmission Modes.

Due to the primary radiation force, microbubble displacement has been observed previously in the focal region of single-element and array ultrasound probes. This effect has been harnessed to increase the contact between the microbubbles and targeted endothelium for drug delivery and ultrasound molecular imaging. In this study, microbubble displacements associated with plane-wave (PW) transmission are thoroughly investigated and compared to those obtained in focused-wave (FW) transmission over a range of pulse repetition frequencies, burst lengths (BLs), peak negative pressures, and transmission frequencies. In PW mode, the displacements, depending upon the experimental conditions, are in some cases consistently higher (e.g., by 28%, when the longest BL was used at PRF = 4 kHz), and the axial displacements are spatially more uniform compared to FW mode. Statistical analysis on the measured displacements reveals a slightly different frequency dependence of statistical quantities compared to transient peak microbubble displacements, which may suggest the need to consider the size range within the tested microbubble population.

Hyperacute spontaneous closure of a traumatic macular hole in a colobomatous eye.

A 12 year old boy presented to us with complaints of diminution of vision following blunt injury to his left eye. Examination revealed a grade 5 retina-choroidal coloboma in left eye with haemorrhage at its inferior border. A macular hole was seen clinically which was confirmed on ocular coherence tomography (OCT). Repeat fundus examination & OCT after 10 days revealed spontaneous closure of the macular hole with improvement in the visual acuity.

Microbubble Formulations: Synthesis, Stability, Modeling and Biomedical Applications.

Microbubbles are increasingly being used in biomedical applications such as ultrasonic imaging and targeted drug delivery. Microbubbles typically range from 0.1 to 10 µm in size and consist of a protective shell made of lipids or proteins. The shell encapsulates a gaseous core containing gases such as oxygen, sulfur hexafluoride or perfluorocarbons. This review is a consolidated account of information available in the literature on research related to microbubbles. Efforts have been made to present an overview of microbubble synthesis techniques; microbubble stability; microbubbles as contrast agents in ultrasonic imaging and drug delivery vehicles; and side effects related to microbubble administration in humans. Developments related to the modeling of microbubble dissolution and stability are also discussed.

Microbubble-Mediated Enhanced Delivery of Curcumin to Cervical Cancer Cells.

The major bottleneck in the current chemotherapy treatment of cancer is the low bioavailability and high cytotoxicity. Targeted delivery of drug to the cancer cells can reduce the cytotoxicity and increase the bioavailability. In this context, microbubbles are currently being explored as drug-delivery vehicles to effectively deliver drug to the tumors or cancerous cells. Microbubbles when used along with ultrasound can enhance drug uptake and inhibit the growth of tumor cells. Several potential anticancer molecules exhibit poor water solubility, which limits their use in therapeutic applications. Such poorly water soluble molecules can be coadministered with microbubbles or encapsulated within or loaded on the microbubbles surface, to enhance the effectiveness of these molecules against cancer cells. Curcumin is one of such potential anticancer molecules obtained from the rhizome of herbal spice, turmeric. In this work, curcumin-loaded protein microbubbles were synthesized and examined for effective in vitro delivery of curcumin to HeLa cells. Microbubbles in the size range of 1-10 µm were produced using perfluorobutane as core gas and bovine serum albumin (BSA) as shell material and were loaded with curcumin. The amount of curcumin loaded on the microbubble surface was estimated using UV-vis spectroscopy, and the average curcumin loading was found to be ∼54 µM/108 microbubbles. Kinetics of in vitro curcumin release from microbubble surface was also estimated, where a 4-fold increase in the rate of curcumin release was obtained in the presence of ultrasound. Sonication and incubation of HeLa cells with curcumin-loaded BSA microbubbles enhanced the uptake of curcumin by ∼250 times. Further, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay confirmed ∼71% decrease in cell viability when HeLa cells were sonicated with curcumin-loaded microbubbles and incubated for 48 h.

Effect of PEGylation on performance of protein microbubbles and its comparison with lipid microbubbles.

Aqueous suspensions of microbubbles are being used in various biomedical applications such as contrast imaging, targeted drug and gene delivery, delivery of drugs through blood brain barrier (BBB) and IV O2 delivery etc. Major microbubble formulations use either proteins or lipids as their shell material. Protein microbubble formulations mainly consist of serum albumin, lysozyme etc., while lipid microbubble formulations consist of phospholipids like 1,2-Distearoyl-sn-glycero-3-phosphocholine (DSPC), 1,2-dipalmitoyl-sn-glycero-3 phosphocholine (DPPC), etc. This work focuses on comparing performance of protein and lipid microbubbles in terms of their shelf life and in vitro performance. Protein microbubbles were produced using Bovine serum albumin (BSA) as main ingredient and N-acetyl-dl-tryptophan (Tryp) as an additive. Lipid microbubbles were produced using a mixture of DSPC as main ingredient and PEG40S (90:10molar ratio) as emulsifier. A narrow sized range (3-5µm) microbubble suspension was produced using sonication method followed by size isolation using centrifugation. These microbubbles were stored in a (PGO) solution containing 10% 1,2 propanediol (P), 10% glycerol (G) and 80% original solution used to make microbubbles (O) and were studied for their shelf stability, in vitro stability, immunogenicity and ability to produce contrast. For a 4weeks of observation period, a least reduction in concentration of around 18% was observed for PEGylated BSA microbubbles whereas highest reduction of 38% was observed for DSPC-PEG40S microbubbles. In-vitro persistence performance for PEGylated BSA microbubbles was found to be better than non-PEGylated BSA microbubbles as well as DSPC-PEG40S microbubbles. Non-PEGylated BSA microbubbles were found to be immunogenic whereas PEGylated BSA and DSPC-PEG40S microbubles were found to be non-immunogenic.