Capillary Occlusion Research Articles

Geographic Distribution and Neuropathology of Elaeophora schneideri in Shiras Moose (Alces alces shirasi) in Idaho, USA.

Elaeophorosis, infection by the filarial worm Elaeophora schneideri, is a parasitic disease of wild ungulates in North America; however, our understanding of the relevance of E. schneideri to moose (Alces alces) morbidity and mortality is incomplete. Between March 2020 and July 2022, necropsy and histopathology were performed on 61 Shiras moose (Alces alces shirasi) in Idaho, US. Among the 41 adults (greater than 1 yr old), 21 moose were from northern Idaho, and 20 were from southeastern Idaho. Elaeophorosis was diagnosed in 24% (10 of 41). All 10 infected moose were from southeastern Idaho; none of the 21 moose from northern Idaho were infected. No juvenile moose (nine from northern and 11 from southeastern Idaho) were infected. Microfilariae were detected histologically in 9 of 10 infected moose, most consistently in brain tissue associated with lesions indicative of ischemic injury to the neuroparenchyma attributed to occlusion of arterioles and capillaries by microfilariae or fibrin thrombi, including edema, necrosis, and glial nodules. Microfilariae found in other tissues of the head, including the eye, tongue, and pinnae of some animals, as well as in lung, heart, liver, and kidney, typically were associated with inflammation. Three of the 10 infected moose had cropped ears attributed to elaeophorosis, and four exhibited abnormal behavior, which may have been due to neuropathology associated with E. schneideri microfilariae in the brain.

Evidence from the large VALIGA cohort validates the subclassification of focal segmental glomerulosclerosis in IgA nephropathy

Evidence from the Oxford IgA nephropathy (IgAN) cohort supports the clinical value of subclassifying focal segmental glomerulosclerosis lesions (S1). Using the larger Validation in IgA (VALIGA) study cohort, we investigated the association between podocytopathic changes and higher proteinuria, kidney outcome and response to immunosuppressive therapy. All biopsies were evaluated for glomeruli with segmental capillary occlusion by matrix ("not otherwise specified", NOS lesion), simple capsular adhesion without capillary occlusion (Adh), tip lesions, and podocyte hypertrophy (PH). S1 required a NOS lesion and/or Adh. A Chi-Squared Automatic Interaction Detection method was used to identify subgroups of FSGS lesions associated with distinctive proteinuria at biopsy. We assessed survival from a combined event (kidney failure or 50% decline in estimated glomerular filtration rate). Finally, we evaluated within each subgroup if immunosuppression was associated with a favorable outcome using propensity analysis. In 1147 patients, S1 was found in 70% of biopsies. Subclassification found NOS lesions in 44%, Adh in 59%, PH in 13%, and tip lesions in 3%, with much overlap. Four subgroups were identified with progressively higher proteinuria: from lowest, S1 without NOS, S1 with NOS but without Adh/PH, to highest, S1 with NOS and Adh but without PH, and S1 with NOS and PH. These four subgroups showed progressively worse kidney survival. Immunosuppression was associated with a better outcome only in the two highest proteinuria subgroups. Propensity analysis in these two groups, adjusted for clinical and pathological findings, found a significantly reduced time-dependent hazard of combined outcome with corticosteroids. Podocyte hypertrophy and glomeruli with simple adhesions appeared to reflect active lesions associated with a response to corticosteroids, while other S1 lesions defined chronicity. Thus, our findings support subclassifying S1 lesions in IgAN.

Pulmonary CT perfusion robustly measures cardiac output in the context of multilevel pulmonary occlusion: a porcine study

BackgroundTo validate pulmonary computed tomography (CT) perfusion in a porcine model by invasive monitoring of cardiac output (CO) using thermodilution method.MethodsAnimals were studied at a single center, using a Swan-Ganz catheter for invasive CO monitoring as a reference. Fifteen pigs were included. Contrast-enhanced CT perfusion of the descending aorta and right and left pulmonary artery was performed. For variation purposes, a balloon catheter was inserted to block the contralateral pulmonary vascular bed; additionally, two increased CO settings were created by intravenous administration of catecholamines. Finally, stepwise capillary occlusion was performed by intrapulmonary arterial injection of 75-μm microspheres in four stages. A semiautomatic selection of AFs and a recirculation-aware tracer-kinetics model to extract the first-pass of AFs, estimating blood flow with the Stewart-Hamilton method, was implemented. Linear mixed models (LMM) were developed to calibrate blood flow calculations accounting with individual- and cohort-level effects.ResultsNine of 15 pigs had complete datasets. Strong correlations were observed between calibrated pulmonary (0.73, 95% confidence interval [CI] 0.6–0.82) and aortic blood flow measurements (0.82, 95% CI, 0.73–0.88) and the reference as well as agreements (± 2.24 L/min and ± 1.86 L/min, respectively) comparable to the state of the art, on a relatively wide range of right ventricle-CO measurements.ConclusionsCT perfusion validly measures CO using LMMs at both individual and cohort levels, as demonstrated by referencing the invasive CO.Relevance statementPossible clinical applications of CT perfusion for measuring CO could be in acute pulmonary thromboembolism or to assess right ventricular function to show impairment or mismatch to the left ventricle.Key points• CT perfusion measures flow in vessels.• CT perfusion measures cumulative cardiac output in the aorta and pulmonary vessels.• CT perfusion validly measures CO using LMMs at both individual and cohort levels, as demonstrated by using the invasive CO as a reference standard.Graphical

The Ultrasound Hepato-Jugular Reflux: Measuring the Hepato-Jugular Reflux with Ultrasound with Comparison to Invasive Right Heart Catheterization

BackgroundUltrasound can overcome barriers to visualizing the internal jugular vein, allowing hepato-jugular reflux and jugular venous pressure measurement. We aimed to determine operating characteristics of the ultrasound hepato-jugular reflux and ultrasound jugular venous pressure predicting right atrial and pulmonary capillary occlusion pressures. MethodsIn a prospective observational cohort at three US academic hospitals the hepato-jugular reflux and jugular venous pressure were measured with ultrasound before right heart catheterization. Receiver operating curves, likelihood ratios, and regression models were utilized to compare the ultrasound hepato-jugular reflux and ultrasound jugular venous pressure to the right atrial and pulmonary capillary occlusion pressures. ResultsIn 99 adults undergoing right heart catheterization, an ultrasound hepato-jugular reflux had a negative likelihood ratio of 0.4 if 0 cm and a positive likelihood ratio of 4.3 if ≥ 1.5 cm for predicting a pulmonary capillary occlusion pressure ≥ 15 mmHg. Regression modeling predicting pulmonary capillary occlusion pressure was not only improved by including the ultrasound hepato-jugular reflux (P < .001), it was the more impactful predictor compared with the ultrasound jugular venous pressure (adjusted odds ratio 2.6 vs 1.2). The ultrasound hepato-jugular reflux showed substantial agreement (kappa 0.76; 95% confidence interval, 0.30-1.21), with poor agreement for the ultrasound jugular venous pressure (kappa 0.11; 95% confidence interval, -0.37-0.58). ConclusionIn patients undergoing right heart catheterization, the ultrasound hepato-jugular reflux is reproducible, has modest impact on the probability of a normal pulmonary capillary occlusion pressure when 0 cm, and more substantial impact on the probability of an elevated pulmonary capillary occlusion pressure when ≥ 1.5 cm.

Relationship between Biochemical Pathways and Non-Coding RNAs Involved in the Progression of Diabetic Retinopathy.

Diabetic retinopathy (DR) is a progressive blinding disease, which affects the vision and quality of life of patients, and it severely impacts the society. This complication, caused by abnormal glucose metabolism, leads to structural, functional, molecular, and biochemical abnormalities in the retina. Oxidative stress and inflammation also play pivotal roles in the pathogenic process of DR, leading to mitochondrial damage and a decrease in mitochondrial function. DR causes retinal degeneration in glial and neural cells, while the disappearance of pericytes in retinal blood vessels leads to alterations in vascular regulation and stability. Clinical changes include dilatation and blood flow changes in response to the decrease in retinal perfusion in retinal blood vessels, leading to vascular leakage, neovascularization, and neurodegeneration. The loss of vascular cells in the retina results in capillary occlusion and ischemia. Thus, DR is a highly complex disease with various biological factors, which contribute to its pathogenesis. The interplay between biochemical pathways and non-coding RNAs (ncRNAs) is essential for understanding the development and progression of DR. Abnormal expression of ncRNAs has been confirmed to promote the development of DR, suggesting that ncRNAs such as miRNAs, lncRNAs, and circRNAs have potential as diagnostic biomarkers and theranostic targets in DR. This review provides an overview of the interactions between abnormal biochemical pathways and dysregulated expression of ncRNAs under the influence of hyperglycemic environment in DR.

The impact of elevated lipoprotein a levels on retinal vein occlusion in young individual: Case report with 7 months follow‐up

Aims/Purpose: Branch retinal vein occlusion (BRVO) occurs when a branch of the retinal vein becomes obstructed. The aim of this study case was to provide insights into the potential role of Lipoprotein (Lp) (a) in the pathogenesis of BRVO.Methods: Case of a 47‐year‐old woman with no significant ophthalmological or general medical history, except the use of oral contraception and smoking habits. Presented to the emergency care after 5 days of experiencing a sudden vision loss in her right eye.Results: The ophthalmological examination revealed a visual acuity of 10/10 on the left (OS) and VBLM/10 on the right eye (OD). Slit lamp and Fundus examination showed on OD a relative afferent pupillary defect and the classic features of acute retinal branch vein occlusion, including tortuous veins, capillary occlusion, papillary and macular oedema and haemorrhages. While all was normal in OS. A comprehensive ophthalmic examination was conducted, including fluorescein angiography and optical coherent tomography that showed no abnormal vessel growth at this stage. Then, the indication for a dexamethasone intravitreal implant has been made along with panretinal photocoagulation (PRP) in the ischaemic areas. More, discontinuing oral contraception, the serum concentrations of specific antibodies, as anticardiolipin and antiphospholipids, as well as homocysteine and Lp (a) levels. The blood tests yielded negative results except for the abnormal plasma level of lipoprotein (a), which was measured at 900 mg/L. After 2 months of follow‐up. The visual acuity in OD was measured at 9/10. Moreover, the restoration of normal macular thickness after 4 months. However, at 6 months follow‐up, the appearance of a neovascularization adjacent to the previously laser‐treated area. As a result, a series of closely spaced PRP was performed along with an intravitreal anti‐vascular endothelial growth factor. After 1 month, a complete disappearance of the neovascularization was observed.Conclusions: Identification of independent and highly predictive risk factors for BRVO is still a big challenge. The main goals are the preservation of visual acuity and the prevention of relapses. This case report emphasizes the importance of incorporating a systematic Lp (a) testing into clinical practice, even in young individual.

Ultrastructural insights from myocardial ablation lesions from microsecond pulsed field vs radiofrequency energy

BackgroundUltrastructural findings immediately after pulsed field ablation (PFA) of the myocardium have not been described. ObjectivesThe purpose of this study was to elucidate ultrastructural characteristics and differences between microsecond PFA at the 1- and 4-hour timepoints after pulse delivery and to compare them to irrigated radiofrequency ablation (RFA) lesions. MethodsHealthy swine underwent endocardial PFA or RFA followed by necropsy. Discrete microsecond PFA and irrigated RFA lesions were created in the ventricles with a lattice tip ablation catheter. Lesions were delivered in a manner so as to allow sampling to occur 1 and 4 hours after ablation. All lesions were located at necropsy, and samples were carefully obtained from within the lesion core, lesion periphery, and adjacent healthy myocardium. Transmission electron microscopic assessment was performed after fixation using paraformaldehyde and glutaraldehyde. ResultsOne hour after microsecond PFA delivery, myocytes were noted to be significantly and uniformly disrupted. Clustered, misaligned, swollen mitochondria coupled with degenerating nuclei and condensed chromatin were visualized. These findings progressed over the subsequent few hours with worsening edema. Similar changes were seen with RFA but reduced in severity. However, there was prominent extravasation of red blood cells with occlusion of capillaries that was not seen in PFA. At the lesion periphery, an abrupt change in the degree of myocyte damage was observed with PFA but not RFA. ConclusionTransmission electron microscopy demonstrates evidence of widespread destruction of myocytes as early as an hour after PFA and corroborates known histologic features such as sparing of vessels and sharp lesion margins.

A Mouse Model of CD19 CAR - T Cell Therapy Reveals Mechanism of Cytopenia

Immunotherapy with Chimeric Antigen Receptor (CAR) T cells against CD19 can cure patients with refractory B cell lymphoma and leukemia. However, complications of CAR-T cell therapy can cause morbidity. Among the complications of CAR-T cell therapy is cytopenia, i.e. decreased peripheral blood counts of platelets, leukocytes and / or erythrocytes. The exact mechanism of this side - effect is not known, retrospective analysis showed it affects 10-20% of patients and is linked with the severity of cytokine release syndrome (Juluri et al, PMID34666344). We have previously developed a mouse model of CAR-T cell therapy using C56BL/6 mouse expressing human CD19 under a B- Cell specific promoter (Pennell et al, PMID29735365). The recipient mice are heterozygous for human hCD19 on CD45.2 background. Congenic C56BL/6 CD45.1 mice served as T cell donors. The aim of this study was to investigate if the mouse model recapitulates cytopenia seen in patients after CAR-T cell therapy. C56BL/6 recipient mice were treated with lymphodepleting chemotherapy (Cyclophosphamide (CY) 300 mg/kg) and infused with 1) no T cells 2) control T cells transfected with hEGFR 3) T cells transfected with CD19 CAR with hCD137 and hCD3zeta signaling domains. The chosen dose of infused T cells at 0.3E+6 per animal was low, approximately 10x below the lethal dose and 3x less than IC50 dose, to reduce early mortality. The animals were weighed and examined regularly, and appearance and activity were scored from 0 (healthy) to 8 (moribund). Total of 30 animals per group were harvested at 5 time points, 6 mice each, on days 5, 12, 26, 40 and 54. Peripheral blood counts were measured using HemaVet CBC counter. Flow cytometry was performed on blood and bone marrow. Spleens, long bones (femur, tibia) and sternum were harvested for histopathologic assessment. Multiparametric immunohistochemistry (IHC) was performed using Codex (Phenocycler, Akoya). The mice receiving CAR-19 T cells showed significant, but transient loss of weight and transient increase in morbidity score during the first 2 weeks. Mortality was not significantly different between groups: 0, 1 and 2 deceased animals in CY, control T and CAR-T cell groups, respectively. We observed transient significant cytopenia in the CAR-T cell treated mice, with significantly lower platelet counts on day 5 (p&amp;lt;0.05) and lower White Blood Cell counts on days 12 (p&amp;lt;0.06) and day 26 (p&amp;lt;0.05). These changes were associated with 100-fold larger expansion of CAR-T cells in blood as compared to the control hEGFR T cells and 20-fold higher relative expansion of CAR-T cells in the bone marrow. Multiparametric IHC showed loss of B cell follicles and increased density of donor T cells in the CAR-T cells recipients compared to hEGFR T cell control. Pathologic evaluation of sternums revealed individual segments (sternebra) with necrosis of the hematopoietic elements at the first time point (day 5). At subsequent time points there were progressive changes reminiscent of evolution of infarct with organization, proliferation of fibroblasts and fibrosis with increased osteogenesis and finally with regression of mesenchymal elements and reestablishment of hematopoiesis (Figure 1). The number of segments with necrosis and sequalae was significantly higher in the CAR-T cell group as compared to the hEGFR T cell control (p=0.0005). The necrosis of hematopoietic elements within individual sternum segments involved at the most 3 segments out of 6. The involved segments appeared to distribute randomly: either adjacent or separated by viable segments. The necrotic segments showed paucity of erythrocytes with foci of recent erythrocyte extravasation consistent with ischemic infarct. The mouse model of CAR-T cell therapy against CD19 recapitulates cytopenia in early post - treatment period. The observed pattern of necrosis of the hematopoietic elements in entire segments suggests compromised blood supply. We hypothesize sternebra are more susceptible to necrosis due to limited blood supply, separate for each segment. We hypothesize that occlusion of arteries or capillaries in the bone marrow may contribute to the mechanism of cytopenia following CAR-T cell therapy. Prior retrospective analysis linked the cytopenia with increased levels of inflammatory mediators of Cytokine Release Syndrome. The present findings highlight a role of local factors in situ, which may be influenced by systemic cytokines.

Microfluidic Assessment of T Cell Deformability and Capillary Network Occlusion

Introduction: Adoptive T cell therapies offer a new frontier of cancer treatment that disrupts the conventional chemotherapy paradigm. Naïve T cells can be genetically modified to directly target cancerous cells and have proven to be aggressively effective in fighting cancers resistant to chemotherapies. A major challenge that adoptive T cell therapies face is successfully delivering the cell products to the location of the target cells, which is particularly challenging in the case of solid tumors. A complete characterization of the biomechanical properties of T cell products would help researchers gain insight into the capacity of these engineered cells to navigate and extravasate through vasculature and tumor microenvironments. Previous studies have found that T cell volume, cortical tension, and dynamic resistance increase with activation (Waugh et al. Front Bioeng Biotechnol. 2023). A simple and reproducible method to easily quantify and assess such cellular properties has yet to be established. Presently, we describe a microfluidic assay for the rapid and robust characterization of the mechanical changes of T cells upon activation. Our assay models the microcapillaries through a gradient of micropillar arrays, which allows for the evaluation of T cells perfused through the network. The spacing between pillars is 12 µm at the first array, similar to small arterioles, and incrementally decreases to 3 µm spacing in the last array, mimicking small capillaries ( Figure 1A). Trafficking of adoptive T cell therapies through the microvasculature networks of this scale into their target tissues represents a persistent and major challenge facing these therapies (Marofi et al. Stem Cell Res. Ther. 2021). Methods: The microfluidic devices were fabricated by casting polydimethylsiloxane (PDMS) on a silicone wafer fabricated through photolithographic protocols. Devices contain arrays of pillars measuring 20 µm x 10 µm x 12 µm with gradient spacing beginning at 12 µm at the inlet and 3 µm at the outlet. Jurkat T cells were cultured in RPMI 1640 medium with 10% fetal bovine serum and used in a naïve state or activated via antibody stimulation of CD3 and CD28. T cells were then suspended in growth media at 1x10 6 cells/mL. Microchannels were wetted and cleaned with 100% ethanol perfused at 10 µL/min for 15 minutes followed by cell culture media at 10 µL/min for 15 minutes to block the non-specific binding sites on the PDMS and to create a biocompatible environment for the cell sample. The devices were connected to a digital pneumatic pressure-based flow pump system. The cell sample was perfused at 100 mBar for 7 minutes and a subsequent wash process with growth media at 100 mBar for 7 minutes was performed to flush non-occluded cells. Upon completion of the experiment, the channel was scanned using Phase and DAPI filters with an Olympus IX83 inverted motorized microscope, and occluded T cells were quantified using ImageJ. Results: Our assay showed distinct levels of T cell occlusion upon activation through CD3 and CD28 stimulation. Naïve T cells showed lower levels of occlusion in each of the first five arrays compared to activated T cells, while both groups showed high levels of occlusion in the final 3 µm array. Activated T cells showed approximately three times the amount of occlusion in the second through fifth arrays. To assess overall occlusion, we use the Occlusion Index (OI), which considers the density of pillars and the number of cells caught in each array. We found that activated T cells had a significantly higher OI when compared to non-activated T cell samples (p &amp;lt; 0.05, unpaired t-test). Naïve T cells had an OI mean of 24.4 ± 5.0 while activated T cells had a mean of 42.5 ± 5.6 ( Figure 1B). Conclusions: Our results show that there is a significant increase in resistance of activated T cells through a microvasculature-modeled environment compared to their naïve counterparts. As adoptive T cell therapies increase in popularity, a complete understanding of the biomechanical properties of these cell products will be necessary for effective design. Our microfluidic assay mimics the narrowing channels of the microvasculature and permits the study of how naïve and engineered T cells travel through these mechanically challenging spaces.

Enrichment of lycopene in watermelon juice by ultrafiltration: technical assessment and fouling analysis

SummaryWatermelon juice is proven to be a potential source of lycopene, but its exploitation and consumption are currently constrained. The research focused on lycopene enrichment in watermelon juice by ultrafiltration (UF) with 1000 Da of molecular weight cut‐off (MWCO). The selectivity of UF membrane for the constituents in watermelon juice was investigated from 2 to 8 bar by dead‐end mode combined with stirring. The 1000 Da UF membrane showed a high potential to recover and separate lycopene from other components. However, membrane deposition and capillary occlusion could occur, affecting the overall efficiency of the enrichment process. To better understand kinetics, phenomena and efficiency, the study was evaluated in cross‐flow mode with full recirculation of retentate. The enrichment efficiency was found to increase the concentration and purity of lycopene by several times. The concentration polarisation was used to describe permeate flux decline, while the Hermia model was developed to explain fouling mechanisms. The utilised models were highly compatible; additionally, pore blocking was thought to be the main mechanism of fouling in lycopene enrichment by 1000 Da UF. The built mathematical model was in good agreement with the experimental observations of permeate flux, and lycopene, TS and ash concentrations. The 1000 Da UF of watermelon juice is a feasible approach for lycopene enrichment.

Balanced Fast-SpaciMS capillary configurations provide practically noninvasive channel-average measurements in catalytic monoliths

Spatially resolved capillary-inlet mass spectrometry (SpaciMS) provides a detailed picture of the spatiotemporal evolution of reaction network in catalytic monoliths. In the present work, we combine the SpaciMS experiments with a newly developed non-isothermal 3D CFD model for heterogeneously catalyzed reactive flows, including diffusion and permeation through the coated catalyst and channel wall. We explore how the capillary size and sampling rate can be balanced to minimize the impact on probed-channel conversions and provide species concentration measurements representative of the free channel average. In all studied configurations, the balanced sampling-rate fraction is noticeably higher than the corresponding capillary occlusion fraction. For a typical 350-micron capillary that occupies 11% of the channel cross- section, the balanced sampling rate represents 43% of the channel flow. The present work shows that together, a balanced SpaciMS configuration and numerical simulation provide accurate channel-averaged information of a monolithic catalyst under realistic operation conditions.

Red Blood Cell Deformability Is Expressed by a Set of Interrelated Membrane Proteins.

Red blood cell (RBC) deformability, expressing their ability to change their shape, allows them to minimize their resistance to flow and optimize oxygen delivery to the tissues. RBC with reduced deformability may lead to increased vascular resistance, capillary occlusion, and impaired perfusion and oxygen delivery. A reduction in deformability, as occurs during RBC physiological aging and under blood storage, is implicated in the pathophysiology of diverse conditions with circulatory disorders and anemias. The change in RBC deformability is associated with metabolic and structural alterations, mostly uncharacterized. To bridge this gap, we analyzed the membrane protein levels, using mass spectroscopy, of RBC with varying deformability determined by image analysis. In total, 752 membrane proteins were identified. However, deformability was positively correlated with the level of only fourteen proteins, with a highly significant inter-correlation between them. These proteins are involved in membrane rafting and/or the membrane-cytoskeleton linkage. These findings suggest that the reduction of deformability is a programmed (not arbitrary) process of remodeling and shedding of membrane fragments, possibly mirroring the formation of extracellular vesicles. The highly significant inter-correlation between the deformability-expressing proteins infers that the cell deformability can be assessed by determining the level of a few, possibly one, of them.

R-(+)-WIN55212-2 protects pericytes from ischemic damage and restores retinal microcirculatory patency after ischemia/reperfusion injury

Background and purposeCannabinoids are vasoactive substances that act as key regulators of arterial tone in the blood vessels supplying peripheral tissues and the central nervous system. This study aimed to investigate the potential of R-(+)-WIN55212-2 (WIN), a cannabinoid receptor 1 agonist (CB1), as a treatment for retinal ischemia/reperfusion (I/R) injury. Experimental approachMale Wistar rats were subjected to retinal I/R injury by increasing intraocular pressure in the anterior chamber. The rats were randomly divided into four groups: normal control, I/R, vehicle (pre-treated with dimethyl sulfoxide [DMSO] via intraperitoneal injection), and experimental (pre-treated with WIN at a dose of 1 ml/kg via intraperitoneal injection). The rats were sacrificed at different time points of reperfusion (1 hour, 3 hours, 6 hours, and 1 day) after inducing retinal I/R injury, and their retinas were collected for analysis. Oxygen-glucose deprived/reperfusion (OGD/R) was performed by initially perfusing the retinas with oxygenated artificial cerebrospinal fluid (ACSF), then switching to an OGD solution to simulate ischemia, followed by another perfusion with ACSF. Pericyte contraction and the "no-reflow" phenomenon were observed using infrared differential interference contrast (IR-DIC) microscopy and immunohistochemistry. Western blot, enzyme-linked immunosorbent assay (ELISA), and nitric oxide (NO) detection were used to explore the potential mechanism. Key resultsIn both the OGD/R and I/R models, retinal pericytes exhibited persistent contraction even after reperfusion. The ability of WIN to regulate the tone of retinal pericytes and capillaries was specifically blocked by the BKCa inhibitor iberiotoxin (100 nM). WIN demonstrated a protective effect against retinal I/R injury by preserving blood flow in vessels containing pericytes. Pretreatment with WIN alleviated the persistent contraction and apoptosis of retinal pericytes in I/R-induced rats, accompanied by a reduction in intracellular calcium ion (Ca2+) concentration. The expression of CB1 decreased in a time-dependent manner in the I/R group. After I/R injury, endothelium-derived nitric oxide (eNOS) levels were reduced at all time points, which was successfully reversed by WIN therapy except for the 1 day group. Additionally, the downregulation of cyclic guanosine monophosphate (cGMP) and BKCa expression at 3 hours, 6 hours, and 1 day after I/R injury was restored by pretreatment of WIN. Conclusions & implicationsWIN exerted its protective effects on retinal I/R injury by inhibiting the contraction and apoptosis of pericytes through the CB1-eNOS-cGMP-BKCa signaling pathway, thus ameliorated the occlusion of retinal capillaries.

Single-microvessel occlusion produces lamina-specific microvascular flow vasodynamics and signs of neurodegenerative change

Recent studies have highlighted the importance of understanding the architecture and function of microvasculature, and dysfunction of these microvessels may underlie neurodegenerative disease. Here, we utilize a high-precision ultrafast laser-induced photothrombosis (PLP) method to occlude single capillaries and then quantitatively study the effects on vasodynamics and surrounding neurons. Analysis of the microvascular architecture and hemodynamics after single-capillary occlusion reveals distinct changes upstream vs. downstream branches, which shows rapid regional flow redistribution and local downstream blood-brain barrier (BBB) leakage. Focal ischemia via capillary occlusions surrounding labeled target neurons induces dramatic and rapid lamina-specific changes in neuronal dendritic architecture. Further, we find that micro-occlusion at two different depths within the same vascular arbor results in distinct effects on flow profiles in layers 2/3 vs layer 4. The current results reveal laminar-scale regulation distinctions in microinfarct response and raise the possibility that relatively greater impacts on microvascular function contribute to cognitive decline in neurodegenerative disease.

Abstract 141: Lung Thrombosis Upregulates Endothelial Pla2g2d To Inhibit Inflammatory Lung Injury

Objective: There are no effective treatments for sepsis-induced acute lung injury (ALI) or acute respiratory distress syndrome (ARDS). ALI/ARDS patients have an increased risk of thrombosis, but the impact of lung thrombosis on ALI/ARDS is ambiguous. Our aim was to target thrombosis-induced signaling to treat ALI/ARDS. Approach and Results: First, we used a mouse model of lung capillary occlusion to quantitatively induce lung thrombosis in mice with sepsis-induced ALI (i.p. LPS and CLP models). A severe level of lung thrombosis exacerbated ALI in sepsis mice (i.e., raised permeability, inflammation, and apoptosis), but an innocuous level of lung thrombosis suppressed ALI. RNA sequencing identified distinct gene profiles in lungs of sepsis mice with innocuous or severe lung thrombosis. A panel of survival factors was increased by innocuous but not severe lung thrombosis, and a subset of these factors was increased in lung endothelial cells (ECs), including A2 phospholipase group IID (Pla2g2d). We then used our new mouse model of EC gene knockout via i.v. injection of biodegradable nanoparticles mixed with plasmid DNA expressing Cas9 driven by the CDH5 promoter and Pla2g2d guide RNA driven by the U6 promotor. The protective impact of innocuous lung thrombosis on ALI was absent in EC-specific Pla2g2d knockout mice. We next administered the biodegradable nanoparticles with plasmid DNA expressing Pla2g2d driven by the CDH5 promoter to overexpress Pla2g2d in ECs. The damaging impact of severe lung thrombosis on ALI was reversed in mice overexpressing EC Pla2g2d. Finally, we explored clinical relevance by data mining to show that circulating Pla2g2d levels are greater in surviving versus non-surviving sepsis patients. Conclusions: EC Pla2g2d protects against sepsis-induced ALI in mice. Circulating Pla2g2d is increased in surviving versus non-surviving sepsis patients. Activation of Pla2g2d signaling represents a novel and plausible therapeutic approach for ALI/ARDS.

Understanding occipital pressure sores in UK military casualties: a pilot study in healthy military personnel

IntroductionThe high prevalence of occipital ulcers in UK military casualties observed during the conflict in Afghanistan is a multifactorial phenomenon. However, the consensus is that ulceration is triggered by excessive...

Comparison of Normal Saline and Alkalinized 2% Lignocaine to Reduce Emergence Phenomenon and Post-Intubation Morbidities: A Prospective, Double-Blind, Randomized Study.

Endotracheal intubation for airway management in general anesthesia is associated with post-intubation morbidities due to tracheal mucosa injury caused by endotracheal tube (ETT) cuff. Nitrous oxide (N2O) diffuses into tracheal tube cuffs filled with air. The rate of diffusion of N2O through the membrane is proportional to its concentration gradient. High-volume low-pressure cuffs expand with only a slight increase in pressure until fully inflated. At this point, owing to the inelasticity of the material, the cuff pressure rises rapidly. This increased pressure can damage the tracheal mucosa. This phenomenon can be avoided, if we inflate the cuff with either a liquid or a gas mixture identical to the inspired gas and monitor the cuff pressure and volume at regular intervals. When lignocaine is used to inflate the ETT cuff, it diffuses to the underlying tracheal mucosa. Thus reducing local irritation and inflammation of the airway through its local anesthetic action. Alkalinization of lignocaine increases its rate of diffusion across the ETT cuff. It also reduces the dose of local anesthetic required to achieve the desired result. We sought to determine the benefits of filling the ETT cuff with alkalinized lignocaine 2% over normal saline, to prevent ETT-induced emergence phenomenon and reduce the incidence of post-intubation morbidities like sore throat, hoarseness, and nausea. This prospective, randomized, double-blind, and comparative study was done at a multispecialty hospital. A total of 120 individuals ofAmerican Society of Anesthesiologists (ASA) physical status 1 and 2, posted for surgery under general anesthesia, were randomly selected and divided into two groups: alkalinized 2% lignocaine group (group L) and normal saline group (group S). After induction of general anesthesia, the airway was secured with appropriate-sized ETT. The ETT cuff was inflated with either of the study media. Continuous cuff pressure monitoring was done to keep cuff pressure below 30 centimeters of water (cm of H2O), at all times. At extubation, the response was evaluated in terms of percentage change in heart rate (HR) and blood pressure from baseline, coughing, bucking, and restlessness. All the surgeries lasted more than two hours. Post-operatively, the patients were evaluated for sore throat and hoarseness, at regular intervals of up to 24 hours. ETT cuff pressure was initially less in group S, which rose to a significantly higher level at extubation, compared to group L (p <0.001). At extubation, there was a significant increase in HR and systolic blood pressure (SBP) from baseline, in group S than in group L (p <0.001 and p=0.001, respectively).The incidence of cough and restlessness was less in group L, compared to group S (p<0.001 and p=0.002, respectively). Mean extubation time and emergence time was more in group S than in group L (p<0.001). Post-operatively, the incidence and severity of sore throat were significantly higher in group S than in group L (p<0.001). Meanwhile, the incidence of hoarseness and nausea was comparable in the two groups. Continuous ETT cuff pressure monitoring helps to keep cuff pressure below tracheal mucosa capillary occlusion pressure. Filling the ETT cuff with alkalinized lignocaine further reduces extubation response and post-intubation morbidities.

Inflammation in diabetic retinopathy: possible roles in pathogenesis and potential implications for therapy.

Diabetic retinopathy, characterized as a microangiopathy and neurodegenerative disease, is the leading cause of visual impairment in diabetic patients. Many clinical features observed in diabetic retinopathy, such as capillary occlusion, acellular capillaries and retinal non-perfusion, aggregate retinal ischemia and represent relatively late events in diabetic retinopathy. In fact, retinal microvascular injury is an early event in diabetic retinopathy involving multiple biochemical alterations, and is manifested by changes to the retinal neurovascular unit and its cellular components. Currently, intravitreal anti-vascular endothelial growth factor therapy is the first-line treatment for diabetic macular edema, and benefits the patient by decreasing the edema and improving visual acuity. However, a significant proportion of patients respond poorly to anti-vascular endothelial growth factor treatments, indicating that factors other than vascular endothelial growth factor are involved in the pathogenesis of diabetic macular edema. Accumulating evidence confirms that low-grade inflammation plays a critical role in the pathogenesis and development of diabetic retinopathy as multiple inflammatory factors, such as interleukin-1β, monocyte chemotactic protein-1 and tumor necrosis factor -α, are increased in the vitreous and retina of diabetic retinopathy patients. These inflammatory factors, together with growth factors such as vascular endothelial growth factor, contribute to blood-retinal barrier breakdown, vascular damage and neuroinflammation, as well as pathological angiogenesis in diabetic retinopathy, complicated by diabetic macular edema and proliferative diabetic retinopathy. In addition, retinal cell types including microglia, Müller glia, astrocytes, retinal pigment epithelial cells, and others are activated, to secrete inflammatory mediators, aggravating cell apoptosis and subsequent vascular leakage. New therapies, targeting these inflammatory molecules or related signaling pathways, have the potential to inhibit retinal inflammation and prevent diabetic retinopathy progression. Here, we review the relevant literature to date, summarize the inflammatory mechanisms underlying the pathogenesis of diabetic retinopathy, and propose inflammation-based treatments for diabetic retinopathy and diabetic macular edema.

Selective Activation of the Wnt-Signaling Pathway as a Novel Therapy for the Treatment of Diabetic Retinopathy and Other Retinal Vascular Diseases.

Retinal ischemia, often associated with various disorders such as diabetic retinopathy (DR), retinal vein occlusion, glaucoma, optic neuropathies, stroke, and other retinopathies, is a major cause of visual impairment and blindness worldwide. As proper blood supply to the retina is critical to maintain its high metabolic demand, any impediment to blood flow can lead to a decrease in oxygen supply, resulting in retinal ischemia. In the pathogenesis of DR, including diabetic macular edema (DME), elevated blood glucose leads to blood-retina barrier (BRB) disruptions, vascular leakage, and capillary occlusion and dropouts, causing insufficient delivery of oxygen to the retina, and ultimately resulting in visual impairment. Other potential causes of DR include neuronal dysfunction in the absence of vascular defect, genetic, and environmental factors. The exact disease progression remains unclear and varies from patient to patient. Vascular leakage leading to edema clearly links to visual impairment and remains an important target for therapy. Despite recent advances in the treatment of DME and DR with anti-VEGFs, effective therapies with new mechanisms of action to address current treatment limitations regarding vessel regeneration and reperfusion of ischemic retinal areas are still needed. The Wnt signaling pathway plays a critical role in proper vascular development and maintenance in the retina, and thus provides a novel therapeutic approach for the treatment of diabetic and other retinopathies. In this review, we summarize the potential of this pathway to address treatment gaps with current therapies, its promise as a novel and potentially disease modifying therapy for patients with DR and opportunities in other retinal vascular diseases.

Neuregulin-1/ErbB4 signaling modulates Plasmodium falciparum HRP2-induced damage to brain cortical organoids

SummaryHuman cerebral malaria (HCM) is a severe complication of Plasmodium falciparum (P.f.) infection that is characterized by capillary occlusions, rupture of the blood-brain barrier (BBB), perivascular cellular injury, and brain swelling. P.f.histidine-rich protein 2 (HRP2), a byproduct of parasitized red blood cell (pRBC) lysis, crosses the BBB when compromised to cause brain injury. We hypothesized that HRP2-induced neuronal damage can be attenuated by Neuregulin-1 (NRG1), an anti-inflammatory neuroprotective factor. Using brain cortical organoids, we determined that HRP2 upregulated cell death and inflammatory markers and disorganized brain organoid tissue. We identified toll-like receptors (TLR1 and 2) as potential mediators of HRP2-induced cellular damage and inflammation. Exogenous acute treatment of organoids with NRG1 attenuated HRP2 effects. The results indicate that HRP2 mediates malaria-associated HRP2-induced brain injury and inflammation and that NRG1 may be an effective therapy against HRP2 effects in the brain.