Magnetic resonance imaging of in vitro urine flow in single and tandem stented ureters subject to extrinsic ureteral obstruction.

OBJECTIVE: To quantify the relative volumetric flows in stent and ureter lumina, as a function of stent size and configuration, in both unobstructed and externally obstructed stented ureters. METHODS: Magnetic resonance imaging was used to measure flow in stented ureters using a phantom kidney model. Volumetric flow in the stent and ureter lumina were determined along the stented ureters, for each of four single stent sizes (4.8F, 6F, 7F, and 8F), and for tandem (6F and 7F) configurations. Measurements were made in the presence of a fully encircling extrinsic ureteral obstruction as well as in benchmark cases with no extrinsic ureteral obstruction. RESULTS: Under no obstruction, the relative contribution of urine flow in single stents is 1-10%, while the relative contributions to flow are ~6 and ~28% for tandem 6F and 7F, respectively. In the presence of an extrinsic ureteral obstruction and single stents, all urine passes within the stent lumen near the extrinsic ureteral obstruction. For tandem 6F and 7F stents under extrinsic ureteral obstruction, relative volumetric flows in the two stent lumina are ~73% and ~81%, respectively, with the remainder passing through the ureter lumen. CONCLUSIONS: Magnetic resonance imaging demonstrates that with no extrinsic ureteral obstruction, minimal urine flow occurs within a stent. Stent lumen flow is significant in the presence of extrinsic ureteral obstruction, in the vicinity of the extrinsic ureteral obstruction. For tandem stents subjected to extrinsic ureteral obstruction, urine flow also occurs in the ureter lumen between the stents, which can reduce the likelihood of kidney failure even in the case of both stent lumina being occluded.

Self-Healing and Light-Soaking in MAPbI3 : The Effect of H2 O.

The future of halide perovskites (HaPs) is beclouded by limited understanding of their long-term stability. While HaPs can be altered by radiation that induces multiple processes, they can also return to their original state by "self-healing." Here two-photon (2P) absorption is used to effect light-induced modifications within MAPbI3 single crystals. Then the changes in the photodamaged region are followed by measuring the photoluminescence, from 2P absorption with 2.5 orders of magnitude lower intensity than that used for photodamaging the MAPbI3 . After photodamage, two brightening and one darkening process are found, all of which recover but on different timescales. The first two are attributed to trap-filling (the fastest) and to proton-amine-related chemistry (the slowest), while photodamage is attributed to the lead-iodide sublattice. Surprisingly, while after 2P-irradiation of crystals that are stored in dry, inert ambient, photobrightening (or "light-soaking") occurs, mostly photodarkening is seen after photodamage in humid ambient, showing an important connection between the self-healing of a HaP and the presence of H2 O, for long-term steady-state illumination, practically no difference remains between samples kept in dry or humid environments. This result suggests that photobrightening requires a chemical-reservoir that is sensitive to the presence of H2 O, or possibly other proton-related, particularly amine, chemistry.

Drainage of infected kidneys with ureteral stents: does size matter?

PURPOSE: The purpose of our study was to evaluate the ability of ureteral stents with different diameters to drain pus that accumulates in an obstructed kidney using an in vitro model. METHODS: We developed an in vitro model of an obstructed kidney filled with pus. The model included a silicon kidney unit based on computed tomography (CT) data, a 3D printed ureteral stone based on a real extracted ureteral stone, a latex ureter model, a bladder vessel, and a fluid with qualities resembling pus. Identical printed stones were inserted into four ureter models containing stents with varying diameters (4.8F, 6F, 7F, 8F), each of which was connected to the kidney unit and the bladder vessel. The kidney unit was filled with artificial pus to pressures of 30 cmH2O to simulate an infected and obstructed kidney. The obstruction was relieved with stents in place, while artificial urine was pumped into the kidney; pressure in the kidney and remaining pus were measured continuously. RESULTS: The rate of pressure drop and the final pressure measured in the kidney were unaffected by the diameter of the stent. For all stent diameters, the pressure reached non-obstructed levels within 30 s, final pressure was reached within 90-120 s, and minimal amounts of pus remained in the kidney after 120 min. CONCLUSIONS: In vitro experiments demonstrate that all stent diameters drain pus-filled, obstructed kidneys with the same efficacy. The common perception that larger diameter tubes are more effective under such circumstances should be re-examined.

Gelatin Stabilizes Nebulized Proteins in Pulmonary Drug Delivery against COVID-19.

Delivering medication to the lungs via nebulization of pharmaceuticals is a noninvasive and efficient therapy route, particularly for respiratory diseases. The recent worldwide severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) pandemic urges the development of such therapies as an effective alternative to vaccines. The main difficulties in using inhalation therapy are the development of effective medicine and methods to stabilize the biological molecules and transfer them to the lungs efficiently following nebulization. We have developed a high-affinity angiotensin-converting enzyme 2 (ACE2) receptor-binding domain (RBD-62) that can be used as a medication to inhibit infection with SARS-CoV-2 and its variants. In this study, we established a nebulization protocol for drug delivery by inhalation using two commercial vibrating mesh (VM) nebulizers (Aerogen Solo and PARI eFlow) that generate similar mist size distribution in a size range that allows efficient deposition in the small respiratory airway. In a series of experiments, we show the high activity of RBD-62, interferon-α2 (IFN-α2), and other proteins following nebulization. The addition of gelatin significantly stabilizes the proteins and enhances the fractions of active proteins after nebulization, minimizing the medication dosage. Furthermore, hamster inhalation experiments verified the feasibility of the protocol in pulmonary drug delivery. In short, the gelatin-modified RBD-62 formulation in coordination with VM nebulizer can be used as a therapy to cure SARS-CoV-2.

Time-space Fourier κω' filter for motion artifacts compensation during transcranial fluorescence brain imaging.

Intravital imaging of brain vasculature through the intact cranium in vivo is based on the evolution of the fluorescence intensity and provides an ability to characterize various physiological processes in the natural context of cellular resolution. The involuntary motions of the examined subjects often limit in vivo non-invasive functional optical imaging. Conventional imaging diagnostic modalities encounter serious difficulties in correction of artificial motions, associated with fast high dynamics of the intensity values in the collected image sequences, when a common reference cannot be provided. In the current report, we introduce an alternative solution based on a time-space Fourier transform method so-called K-Omega. We demonstrate that the proposed approach is effective for image stabilization of fast dynamic image sequences and can be used autonomously without supervision and assignation of a reference image.

Combined laser speckle imaging and fluorescent intravital microscopy for monitoring acute vascular permeability reaction.

Optical clearing agents (OCAs) and many chemicals are widely used in functional diagnosis of skin tissues. Numerous studies are associated with the transcutaneous diffusion of OCA in epidermal, dermal, and hypodermal tissues, which results in changing their optical properties. In addition, an objective approach that is suitable for screening the influence of utilized OCA, as well as various chemical agents, synthetics, and nanomaterials, on blood and lymph flows is highly desirable. In our study, a highly sensitive laser speckle imaging (LSI) system and fluorescent intravital microscopy (FIM) were used team-wise to inspect the acute skin vascular permeability reaction in mouse ear during the local application of OCA on the skin surface. Fluorescent contrast material administrated intravenously was used for quantitatively assessing the intensity of vascular permeability reaction and the strength of skin irritation. The obtained results suggest that a combined use of LSI and FIM is highly effective for monitoring the cutaneous vascular permeability reaction, with great potential for assessment of allergic reactions of skin in response to interactions with chemical substances.

Dual role of E-cadherin in the regulation of invasive collective migration of mammary carcinoma cells.

In this article, we explore a non-canonical form of collective cell migration, displayed by the metastatic murine mammary carcinoma cell line 4T1. We show here that in sparsely plated 4T1 cells, E-cadherin levels are moderately reduced (~50%), leading to the development of collective migration, whereby cells translocate in loose clusters, interconnected by thin membrane tethers. Knocking down E-cadherin blocked tether formation in these cells, leading to enhancement of migration rate and, at the same time, to suppression of lung metastases formation in vivo, and inhibition of infiltration into fibroblast monolayers ex vivo. These findings suggest that the moderate E-cadherin levels present in wild-type 4T1 cells play a key role in promoting cancer invasion and metastasis.

Self-Healing Inside APbBr3 Halide Perovskite Crystals.

Self-healing, where a modification in some parameter is reversed with time without any external intervention, is one of the particularly interesting properties of halide perovskites. While there are a number of studies showing such self-healing in perovskites, they all are carried out on thin films, where the interface between the perovskite and another phase (including the ambient) is often a dominating and interfering factor in the process. Here, self-healing in perovskite (methylammonium, formamidinium, and cesium lead bromide (MAPbBr3 , FAPbBr3 , and CsPbBr3 )) single crystals is reported, using two-photon microscopy to create damage (photobleaching) ≈110 µm inside the crystals and to monitor the recovery of photoluminescence after the damage. Self-healing occurs in all three perovskites with FAPbBr3 the fastest (≈1 h) and CsPbBr3 the slowest (tens of hours) to recover. This behavior, different from surface-dominated stability trends, is typical of the bulk and is strongly dependent on the localization of degradation products not far from the site of the damage. The mechanism of self-healing is discussed with the possible participation of polybromide species. It provides a closed chemical cycle and does not necessarily involve defect or ion migration phenomena that are often proposed to explain reversible phenomena in halide perovskites.

Regulatory Forum Opinion Piece*: Imaging Applications in Toxicologic Pathology-Recommendations for Use in Regulated Nonclinical Toxicity Studies.

Available imaging systems for use in preclinical toxicology studies increasingly show utility as important tools in the toxicologic pathologist's armamentarium, permit longitudinal evaluation of functional and morphological changes in tissues, and provide important information such as organ and lesion volume not obtained by conventional toxicology study parameters. Representative examples of practical imaging applications in toxicology research and preclinical studies are presented for ultrasound, positron emission tomography/single-photon emission computed tomography, optical, magnetic resonance imaging, and matrix-assisted laser desorption ionization-imaging mass spectrometry imaging. Some of the challenges for making imaging systems good laboratory practice-compliant for regulatory submission are presented. Use of imaging data on a case-by-case basis as part of safety evaluation in regulatory submissions is encouraged.

Brown-adipose-tissue macrophages control tissue innervation and homeostatic energy expenditure.

Tissue macrophages provide immunological defense and contribute to the establishment and maintenance of tissue homeostasis. Here we used constitutive and inducible mutagenesis to delete the nuclear transcription regulator Mecp2 in macrophages. Mice that lacked the gene encoding Mecp2, which is associated with Rett syndrome, in macrophages did not show signs of neurodevelopmental disorder but displayed spontaneous obesity, which was linked to impaired function of brown adipose tissue (BAT). Specifically, mutagenesis of a BAT-resident Cx3Cr1+ macrophage subpopulation compromised homeostatic thermogenesis but not acute, cold-induced thermogenesis. Mechanistically, malfunction of BAT in pre-obese mice with mutant macrophages was associated with diminished sympathetic innervation and local titers of norepinephrine, which resulted in lower expression of thermogenic factors by adipocytes. Mutant macrophages overexpressed the signaling receptor and ligand PlexinA4, which might contribute to the phenotype by repulsion of sympathetic axons expressing the transmembrane semaphorin Sema6A. Collectively, we report a previously unappreciated homeostatic role for macrophages in the control of tissue innervation. Disruption of this circuit in BAT resulted in metabolic imbalance.

The retinal pigment epithelium as a gateway for monocyte trafficking into the eye.

The choroid plexus epithelium within the brain ventricles orchestrates blood-derived monocyte entry to the central nervous system under injurious conditions, including when the primary injury site is remote from the brain. Here, we hypothesized that the retinal pigment epithelium (RPE) serves a parallel role, as a gateway for monocyte trafficking to the retina following direct or remote injury. We found elevated expression of genes encoding leukocyte trafficking determinants in mouse RPE as a consequence of retinal glutamate intoxication or optic nerve crush (ONC). Blocking VCAM-1 after ONC interfered with monocyte infiltration into the retina and resulted in a local pro-inflammatory cytokine bias. Live imaging of the injured eye showed monocyte accumulation first in the RPE, and subsequently in the retina, and peripheral leukocytes formed close contact with the RPE Our findings further implied that the ocular milieu can confer monocytes a phenotype advantageous for neuroprotection. These results suggest that the eye utilizes a mechanism of crosstalk with the immune system similar to that of the brain, whereby epithelial barriers serve as gateways for leukocyte entry.

Quantification of the Blood Platelet Reactivity in the ADP-Induced Model of Non-Lethal Pulmonary Thromboembolism in Mice with the Use of Laser Doppler Flowmetry.

INTRODUCTION: The paper describes an alternative method for quantification of in vivo ADP-induced thromboembolism. The aim of the studies was to develop a method of quantification which would not require either extravasation or labelling of platelets. Our proposed approach is based on the monitoring of changes of blood flow with the use of laser Doppler flowmetry. MATERIALS AND METHODS: Mice of C57Bl strain were used in the study. ADP was injected to the vena cava and blood flow was monitored with the use of a laser Doppler flowmeter in the mesentery. Measurements in platelet-depleted mice, mice pretreated with cangrelor, an ADP receptor antagonist, and eptifibatide, a blocker of fibrinogen binding to GPIIbIIIa, were conducted as the proof-of-concept in the performed experiments. Intravital microscopy and ex vivo imaging of organs was performed to identify the sites of aggregate formation resulting from ADP injection. RESULTS: The injection of ADP resulted in a dose-dependent reduction of the blood flow in the mesentery. These responses were fully attributable to blood platelet aggregation, as shown by the lack of the effect in platelet-depleted mice, and significantly reduced responses in mice pretreated with cangrelor and eptifibatide. No platelet aggregate formation in mesenteric vessels was revealed by intravital microscopy, while ex vivo imaging showed accumulation of fluorescent labelled platelets in the lung. CONCLUSIONS: Injection of ADP to the venous system results in the formation of platelet aggregates predominantly in the lung. This results in reversible blood flow cessation in peripheral blood vessels. The measurement of this blood flow cessation in the mesentery allows indirect measurement of ADP-induced pulmonary thromboembolism. We suggest that this approach can be useful for in vivo screening for antiplatelet drug candidates.

Study of the cortical representation of whisker frequency selectivity using voltage-sensitive dye optical imaging.

The facial whiskers of rodents act as a high-resolution tactile apparatus that allow the animal to detect the finest details of its environment. Previously it was shown that whisker-sensitive neurons in the somatosensory cortex show frequency selectivity to small amplitude stimuli, An intravital voltage-sensitive dye optical imaging (VSDi) method in combination with the different frequency whisker stimulation was used in order to visualize neural activity in the mice somatosensory cortex in response to the stimulation of a single whisker by different frequencies. Using the intravital voltage-sensitive dye optical imaging (VSDi) method in combination with the different frequency whisker stimulation we visualized neural activity in the mice somatosensory cortex in response to the stimulation of a single whisker by different frequencies. We found that whisker stimuli with different frequencies led to different optical signals in the barrel field. Our results provide evidence that different neurons of the barrel cortex have different frequency preferences. This supports prior research that whisker deflections cause responses in cortical neurons within the barrel field according to the frequency of the stimulation. Many studies of the whisker frequency selectivity were performed using unit recording but to map spatial organization, imaging methods are essential. In the work described in the present paper, we take a serious step toward detailed functional mapping of the somatosensory cortex using VSDi. To our knowledge, this is the first demonstration of whisker frequency sensitivity and selectivity of barrel cortex neurons with optical imaging methods.

Self-Assembled Organic Nanocrystals with Strong Nonlinear Optical Response.

Facile molecular self-assembly affords a new family of organic nanocrystals that, unintuitively, exhibit a significant nonlinear optical response (second harmonic generation, SHG) despite the relatively small molecular dipole moment of the constituent molecules. The nanocrystals are self-assembled in aqueous media from simple monosubstituted perylenediimide (PDI) molecular building blocks. Control over the crystal dimensions can be achieved via modification of the assembly conditions. The combination of a simple fabrication process with the ability to generate soluble SHG nanocrystals with tunable sizes may open new avenues in the area of organic SHG materials.

A Novel Intravital Imaging Window for Longitudinal Microscopy of the Mouse Ovary.

The ovary is a dynamic organ that undergoes dramatic remodeling throughout the ovulatory cycle. Maturation of the ovarian follicle, release of the oocyte in the course of ovulation as well as formation and degradation of corpus luteum involve tightly controlled remodeling of the extracellular matrix and vasculature. Ovarian tumors, regardless of their tissue of origin, dynamically interact with the ovarian microenvironment. Their activity in the tissue encompasses recruitment of host stroma and immune cells, attachment of tumor cells to mesothelial layer, degradation of the extracellular matrix and tumor cell migration. High-resolution dynamic imaging of such processes is particularly challenging for internal organs. The implementation of a novel imaging window as reported here enabled longitudinal microscopy of ovarian physiology and orthotopic tumor invasion.

A simple approach for non-invasive transcranial optical vascular imaging (nTOVI).

In vivo imaging of cerebral vasculature is highly vital for clinicians and medical researchers alike. For a number of years non-invasive optical-based imaging of brain vascular network by using standard fluorescence probes has been considered as impossible. In the current paper controverting this paradigm, we present a robust non-invasive optical-based imaging approach that allows visualize major cerebral vessels at the high temporal and spatial resolution. The developed technique is simple to use, utilizes standard fluorescent dyes, inexpensive micro-imaging and computation procedures. The ability to clearly visualize middle cerebral artery and other major vessels of brain vascular network, as well as the measurements of dynamics of blood flow are presented. The developed imaging approach has a great potential in neuroimaging and can significantly expand the capabilities of preclinical functional studies of brain and notably contribute for analysis of cerebral blood circulation in disorder models. An example of 1 × 1.5 cm color-coded image of brain blood vessels of mouse obtained in vivo by transcranial optical vascular imaging (TOVI) approach through the intact cranium.

Transcranial optical vascular imaging (TOVI) of cortical hemodynamics in mouse brain.

In vivo imaging of cerebral vasculature and blood flow provides highly valuable information for clinicians as well as researchers. Nevertheless, currently available methods are complex, time-consuming and expensive. Here, we present a novel, minimally invasive method for vascular imaging through the sufficiently transparent intact skull of young mice. Our method combines laser speckle and fluorescent imaging with dynamic color mapping and image fusion. Quickly generated wide-field images present clear visual information on blood flow and perfusion in the cerebral cortex and meninges. The ability of the method to visualize hemodynamic changes is demonstrated by induced occlusion of the middle cerebral artery. The compact and easily operated system comprises of several pieces of standard and affordable laboratory equipment. This simple, robust and inexpensive method may become an important tool for assessment of brain hemodynamics in preclinical studies.

Ear swelling test by using laser speckle imaging with a long exposure time.

Laser speckle imaging with long exposure time has been applied noninvasively to visualize the immediate reaction of cutaneous vessels in mice in response to a known primary irritant and potential allergen­methyl salicylate. The compound has been used topically on the surface of the pinna and the reaction of the vascular network was examined. We demonstrate that irritant-induced acute vascular reaction can be effectively and accurately detected by laser speckle imaging technique. The current approach holds a great promise for application in routine screening of the cutaneous vascular response induced by contact agents, screenings of mouse ear swelling test, and testing the allergenic potential of new synthetic materials and healthcare pharmaceutical products.

Dynamic imaging reveals promiscuous crosspresentation of blood-borne antigens to naive CD8+ T cells in the bone marrow.

The bone marrow (BM) hosts memory lymphocytes and supports secondary immune responses against blood-borne antigens, but it is unsettled whether primary responses occur there and which cells present the antigen. We used 2-photon microscopy in the BM of live mice to study these questions. Naïve CD8(+) T cells crawled rapidly at steady state but arrested immediately upon sensing antigenic peptides. Following infusion of soluble protein, various cell types were imaged ingesting the antigen, while antigen-specific T cells decelerated, clustered, upregulated CD69, and were observed dividing in situ to yield effector cells. Unlike in the spleen, T-cell responses persisted when BM-resident dendritic cells (DCs) were ablated but failed when all phagocytic cells were depleted. Potential antigen-presenting cells included monocytes and macrophages but not B cells. Collectively, our results suggest that the BM supports crosspresentation of blood-borne antigens similar to the spleen; uniquely, alongside DCs, other myeloid cells participate in crosspresentation.

Recruitment of beneficial M2 macrophages to injured spinal cord is orchestrated by remote brain choroid plexus.

Monocyte-derived macrophages are essential for recovery after spinal cord injury, but their homing mechanism is poorly understood. Here, we show that although of common origin, the homing of proinflammatory (M1) and the "alternatively activated" anti-inflammatory (M2) macrophages to traumatized spinal cord (SC) was distinctly regulated, neither being through breached blood-brain barrier. The M1 macrophages (Ly6c(hi)CX3CR1(lo)) derived from monocytes homed in a CCL2 chemokine-dependent manner through the adjacent SC leptomeninges. The resolving M2 macrophages (Ly6c(lo)CX3CR1(hi)) derived from monocytes trafficked through a remote blood-cerebrospinal-fluid (CSF) barrier, the brain-ventricular choroid plexus (CP), via VCAM-1-VLA-4 adhesion molecules and epithelial CD73 enzyme for extravasation and epithelial transmigration. Blockage of these determinants, or mechanical CSF flow obstruction, inhibited M2 macrophage recruitment and impaired motor-function recovery. The CP, along with the CSF and the central canal, provided an anti-inflammatory supporting milieu, potentially priming the trafficking monocytes. Overall, our finding demonstrates that the route of monocyte entry to central nervous system provides an instructional environment to shape their function.