Purpose:Corneal guttae are the hallmark pathologic phenotype of Fuchs endothelial corneal dystrophy (FECD). Tools performing quantitative analysis of guttae and their changes over time are needed as medical therapies advance. We developed and validated a novel automated software platform that reports the guttae area ratio (GAR) in FECD specular microscopy images. Methods:Patients with FECD with modified Krachmer grade (mKg) 2 to 5 disease and no prior keratoplasty were enrolled and imaged prospectively with one specular microscope unit. In a validation study, expert manual grading and automated software grading of GAR were compared for agreement. Additional images were analyzed to correlate automated GAR with disease grade and subclinical edema by Scheimpflug imaging. Results:Nineteen images were included in the validation study. Mean manual GAR (0.400 ± 0.213) and mean automated GAR (0.395 ± 0.182) demonstrated equivalence (P = 0.003). Additionally, 446 images (109 eyes) demonstrated that automated GAR increases with worsening disease in mKg 2 to 5. Most mKg 4 images (n = 172, 93.5%; 49 eyes) and mKg 5 images (n = 59, 55.1%; 19 eyes) had no evidence of definite subclinical edema. Repeat image analysis demonstrated high repeatability of GAR (intraclass correlation = 0.85; 95% confidence interval, 0.78-0.90). Conclusions:This study validates that automated software for guttae quantification in FECD specular microscopy images is equivalent to manual expert grading. GAR, an objective metric that increases in direct proportion to mKg, may precede development of subclinical edema. Translational Relevance:This software supports precise quantification of FECD disease secondary to guttae, allowing for objective measurement of disease progression and therapeutic efficacy.

There is limited information on the relationship between multidisciplinary bundle compliance and long-term outcomes. We explored the association between bundle compliance and outcomes at 3 and 12 months after hospital discharge. We conducted secondary analyses of a randomized controlled trial comparing haloperidol, ziprasidone, and placebo for delirium in patients with critical illness. From 2011 to 2017, adult patients with respiratory failure and/or shock were recruited from 16 US medical centers. Participants received a daily ABCDE bundle during their critical illness. We assessed cognition, disability, mental health, quality of life, and survival over 12 months of follow-up. We analyzed data using linear mixed effects regression and Cox proportional hazards models. Among 566 participants, 418 survived to hospital discharge, 304 were assessed for 3-month outcomes, and 251 for 12-month outcomes. Median proportional bundle compliance was 97 https://clinicaltrials.gov/ct2/show/NCT01211522 .

PURPOSE. Evidence suggests that corneal endothelial cell (CEC) death in Fuchs endothelial corneal dystrophy (FECD) is due to ferroptosis, an iron-mediated cell death. Iron-sulfur cluster (ISC)-containing aconitases and the iron responsive element binding proteins IREBP1 and IREBP2 are known mediators of iron homeostasis. This study investigates mechanisms underlying iron dysregulation in CECs and proposes a role for ISCs and IREBPs in the context of FECD pathogenesis. METHODS. We studied gene expression of proteins responsible for ISC synthesis and iron homeostasis in human and mouse CECs and analyzed published RNA sequencing datasets. We validated a subset of transcriptional changes between FECD and control tissues using microfluidic Western blotting with human CEC tissues. Finally, we silenced proteins involved in ISC synthesis or iron homeostasis in cell cultures and assessed ferroptosis susceptibility. RESULTS. RNA-seq and qPCR data demonstrated significantly decreased transcription of genes required for ISC synthesis in FECD tissues (P < 0.05). Protein quantification revealed a significant decrease in mitochondrial aconitase (P < 0.05), ferredoxin 1 (P < 0.001), and mitofusin (P < 0.05), and a significant increase in cysteine desulfurase (P < 0.05), cytosolic aconitase/IREBP1, and IREBP2 (P < 0.05) in FECD tissues. Silencing studies revealed increased susceptibility to ferroptosis upon siRNA knockdown of ferredoxin 1 (P < 0.05). CONCLUSIONS. We identified differential gene expression of proteins responsible for ISC synthesis, ISC-containing proteins, IREBPs that mediate cellular iron homeostasis, and mitofusin, which promotes mitochondrial fusion in FECD. We also identified increased susceptibility to ferroptosis after ferredoxin 1 knockdown in CECs. These results advance an ISC-and IREBP-mediated mechanism of iron accumulation in FECD CECs.

Changes in the anterior segment of the eye due to type 2 diabetes mellitus (T2DM) are not wellcharacterized, in part due to the lack of a reliable animal model. This study evaluated changes in the anterior segment, including crystalline lens health, corneal endothelial cell density, aqueous humor metabolites, and ciliary body vasculature, in a rat model of T2DM compared with human eyes. Male Sprague-Dawley rats were fed a high-fat diet (45% fat) or normal diet, and rats fed the high-fat diet were injected with streptozotocin intraperitoneally to generate a model of T2DM. Cataract formation and corneal endothelial cell density were assessed using microscopic analysis. Diabetes-related rat aqueous humor alterations were assessed using metabolomics screening. Transmission electron microscopy was used to assess qualitative ultrastructural changes ciliary process microvessels at the site of aqueous formation in the eyes of diabetic rats and humans. Eyes from the diabetic rats demonstrated cataracts, lower corneal endothelial cell densities, altered aqueous metabolites, and ciliary body ultrastructural changes, including vascular endothelial cell activation, pericyte degeneration, perivascular edema, and basement membrane reduplication. These findings recapitulated diabetic changes in human eyes. These results support the use of this model for studying ocular manifestations of T2DM and support a hypothesis postulating blood-aqueous barrier breakdown and vascular leakage at the ciliary body as a mechanism for diabetic anterior segment pathology. (Am J Pathol 2024, 194: 1090-1105; https://doi.org/10.1016/j.ajpath.2024.02.004)

Fuchs endothelial corneal dystrophy (FECD), the leading indication for corneal transplantation in the U.S., causes loss of corneal endothelial cells (CECs) and corneal edema leading to vision loss. FECD pathogenesis is linked to impaired response to oxidative stress and environmental ultraviolet A (UVA) exposure. Although UVA is known to cause nonapoptotic oxidative cell death resulting from iron-mediated lipid peroxidation, ferroptosis has not been characterized in FECD. We investigated the roles of genetic background and UVA exposure in causing CEC degeneration in FECD. Using ungenotyped FECD patient surgical samples, we found increased levels of cytosolic ferrous iron (Fe2+) and lipid peroxidation in end-stage diseased tissues compared with healthy controls. Using primary and immortalized cell cultures modeling the TCF4 intronic trinucleotide repeat expansion genotype, we found altered gene and protein expression involved in ferroptosis compared to controls including elevated levels of Fe2+, basal lipid peroxidation, and the ferroptosis-specific marker transferrin receptor 1. Increased cytosolic Fe2+ levels were detected after physiologically relevant doses of UVA exposure, indicating a role for ferroptosis in FECD disease progression. Cultured cells were more prone to ferroptosis induced by RSL3 and UVA than controls, indicating ferroptosis susceptibility is increased by both FECD genetic background and UVA. Finally, cell death was preventable after RSL3 induced ferroptosis using solubilized ubiquinol, indicating a role for anti-ferroptosis therapies in FECD. This investigation demonstrates that genetic background and UVA exposure contribute to iron-mediated lipid peroxidation and cell death in FECD, and provides the basis for future investigations of ferroptosis-mediated disease progression in FECD.

Prolonged mechanical ventilation (MV) subjects multiple trauma patients to ventilator-induced diaphragmatic dysfunction. There is limited evidence on the predictive role of diaphragm ultrasound (DUS) for weaning success in multiple trauma patients. Therefore, we evaluated Ultrasound of the diaphragm as a valuable indicator of weaning outcomes, in trauma patients. This prospective cohort study included 50 trauma patients from September 2018 to February 2019. DUS was performed twice: upon ICU admission and the first weaning attempt. The diagnostic accuracy of indexes was evaluated by ROC curves. The study included patients with a mean age of 35.4 ± 17.37, and 78

BACKGROUND:Delirium is common during critical illness and is associated with long-term cognitive impairment and disability. Antipsychotics are frequently used to treat delirium, but their effects on long-term outcomes are unknown. We aimed to investigate the effects of antipsychotic treatment of delirious, critically ill patients on long-term cognitive, functional, psychological, and quality-of-life outcomes. METHODS:This prespecified, long-term follow-up to the randomised, double-blind, placebo-controlled phase 3 MIND-USA Study was conducted in 16 hospitals throughout the USA. Adults (aged ≥18 years) who had been admitted to an intensive care unit with respiratory failure or septic or cardiogenic shock were eligible for inclusion in the study if they had delirium. Participants were randomly assigned-using a computer-generated, permuted-block randomisation scheme with stratification by trial site and age-in a 1:1:1 ratio to receive intravenous placebo, haloperidol, or ziprasidone for up to 14 days. Investigators and participants were masked to treatment group assignment. 3 months and 12 months after randomisation, we assessed survivors' cognitive, functional, psychological, quality-of-life, and employment outcomes using validated telephone-administered tests and questionnaires. This trial was registered with ClinicalTrials.gov, NCT01211522, and is complete. FINDINGS:Between Dec 7, 2011, and Aug 12, 2017, we screened 20 914 individuals, of whom 566 were eligible and consented or had consent provided to participate. Of these 566 patients, 184 were assigned to the placebo group, 192 to the haloperidol group, and 190 to the ziprasidone group. 1-year survival and follow-up rates were similar between groups. Cognitive impairment was common in all three treatment groups, with a third of survivors impaired at both 3-month and 12-month follow-up in all groups. More than half of the surveyed survivors in each group had cognitive or physical limitations (or both) that precluded employment at both 3-month and 12-month follow-up. At both 3 months and 12 months, neither haloperidol (adjusted odds ratio 1·22 [95% CI 0·73-2.04] at 3 months and 1·12 [0·60-2·11] at 12 months) nor ziprasidone (1·07 [0·59-1·96] at 3 months and 0·94 [0·62-1·44] at 12 months) significantly altered cognitive outcomes, as measured by the Telephone Interview for Cognitive Status T score, compared with placebo. We also found no evidence that functional, psychological, quality-of-life, or employment outcomes improved with haloperidol or ziprasidone compared with placebo. INTERPRETATION:In delirious, critically ill patients, neither haloperidol nor ziprasidone had a significant effect on cognitive, functional, psychological, or quality-of-life outcomes among survivors. Our findings, along with insufficient evidence of short-term benefit and frequent inappropriate continuation of antipsychotics at hospital discharge, indicate that antipsychotics should not be used routinely to treat delirium in critically ill adults. FUNDING:National Institutes of Health and the US Department of Veterans Affairs.

Optimal oxygenation in the intensive care unit requires adequate pulmonary gas exchange, oxygen-carrying capacity in the form of hemoglobin, sufficient delivery of oxygenated hemoglobin to the tissue, and an appropriate tissue oxygen demand. In this Case Study in Physiology, we describe a patient with COVID-19 whose pulmonary gas exchange and oxygen delivery were severely compromised by COVID-19 pneumonia requiring extracorporeal membrane oxygenation (ECMO) support. His clinical course was complicated by a secondary superinfection with staphylococcus aureus and sepsis. This case study is provided with two goals in mind (1) We outline how basic physiology was used to address life-threatening consequences of a novel infection-COVID-19. (2) We describe a strategy of whole-body cooling to lower the cardiac output and oxygen consumption, use of the shunt equation to optimize flow to the ECMO circuit, and transfusion to improve oxygen-carrying capacity when ECMO alone failed to provide sufficient oxygenation.

Diabetes mellitus is a multiorgan systemic disease impacting numerous ocular structures that results in significant ocular morbidity and often results in more frequent corneal and glaucoma surgeries for affected individuals. We hypothesize that the systemic metabolic and proteomic derangement observed in the progression of diabetes influences the composition of the aqueous humor (AH), which ultimately impacts the anterior segment health of the eye. To identify changes associated with diabetes progression, we mapped the metabolite profile and proteome of AH samples from patients with varying severities of type II diabetes (T2DM). Patients were classified as nondiabetic (ND or control), non-insulin-dependent diabetic without advanced features of disease (NAD-ni), insulin-dependent diabetic without advanced features (NAD-i), or diabetic with advanced features (AD). AH samples collected from the anterior chamber during elective ophthalmic surgery were evaluated for metabolite and protein expression changes associated with diabetic severity via gas chromatography/mass spectrometry and ultra-high performance liquid chromatography tandem mass spectrometry, respectively. Metabolic and proteomic pathway analyses were conducted utilizing MetaboAnalyst 4.0 and Ingenuity Pathway Analysis. A total of 14 control, 12 NAD-ni, 4 NAD-I, and 14 AD samples were included for analysis. Elevated levels of several branched amino acids (e.g., valine, leucine, isoleucine), and lipid metabolites (e.g., palmitate) were found only with increasing diabetic severity (i.e., the AD group). Similar proteomic trends were noted in amino acid and fatty acid metabolism and the unfolded protein/stress response. These results represent the first report of both metabolomic and proteomic evaluation of aqueous humor. Diabetes results in metabolic and proteomic perturbations detectable in the AH, and unique changes become manifest as T2DM severity worsens. Changes in AH composition may serve as an indicator of disease severity, risk assessment of anterior segment cells and structures, and potential future therapies.

Purpose There is a pressing need to investigate the impact of type II diabetes mellitus on the posterior cornea in donor tissues given its increasing prevalence and potential impact on endothelial keratoplasty surgical outcomes. Methods Immortalized human cultured corneal endothelial cells (CECs; HCEC-B4G12) were grown in hyperglycemic media for 2 weeks. Extracellular matrix (ECM) adhesive glycoprotein expression and advanced glycation end products (AGEs) in cultured cells and corneoscleral donor tissues, as well as the elastic modulus for the Descemet membrane (DMs) and CECs of diabetic and nondiabetic donor corneas, were measured. Results In CEC cultures, increasing hyperglycemia resulted in increased transforming growth factor beta-induced (TGFBI) protein expression and colocalization with AGEs in the ECM. In donor corneas, the thicknesses of the DM and the interfacial matrix (IFM) between the DM and stroma both increased from 8.42 ± 1.35 µm and 0.504 ± 0.13 µm in normal corneas, respectively, to 11.13 ± 2.91 µm (DM) and 0.681 ± 0.24 µm (IFM) in non-advanced diabetes (P = 0.013 and P = 0.075, respectively) and 11.31 ± 1.76 µm (DM) and 0.744 ± 0.18 µm (IFM) in advanced diabetes (AD; P = 0.0002 and P = 0.003, respectively). Immunofluorescence in AD tissues versus controls showed increased AGEs (P < 0.001) and markedly increased labeling intensity for adhesive glycoproteins, including TGFBI, that colocalized with AGEs. The elastic modulus significantly increased between AD and control tissues for the DMs (P < 0.0001) and CECs (P < 0.0001). Conclusions Diabetes and hyperglycemia alter human CEC ECM structure and composition, likely contributing to previously documented complications of endothelial keratoplasty using diabetic donor tissue, including tearing during graft preparation and reduced graft survival. AGE accumulation in the DM and IFM may be a useful biomarker for determining diabetic impact on posterior corneal tissue.

Marine oils may offer cardiovascular benefits, but inhibition of prostaglandin E and prostaglandin F synthesis by fish oil has been found in animal studies, and such effects could alter physiological responses in man to a clinically significant degree. Since greater amounts of E and F-type prostaglandins are made in human seminal vesicles than in the rest of the body combined, the influence of n-3 supplements upon semen prostaglandins was assessed in 10 subjects before and after one month of taking 50 ml menhaden oil daily. Prostaglandins E1, E2 and their 19-hydroxy derivatives were measured by HPLC-UV as PGB's, and prostaglandin E3, 19-OH PGE3, and analogous PGF's by gas chromatography/mass spectrometry. Fish oil ingestion reduced concentrations of one- and two series prostaglandins (mean reduction in PGE's = 37%, in PGF's = 20%, p less than 0.05), while more than doubling the low amounts of PGE3 and PGF3 alpha, and their previously undescribed 19-hydroxy derivatives. Semen phospholipids were enriched in eicosapentaenoic acid after dietary fish oil, but sperm counts and motility were not altered during the study. Since dietary fish oil reduces prostaglandin concentration in semen, clinical trials of n-3 fatty acids should also evaluate other possible results of in vivo cyclooxygenase inhibition.

Fuchs endothelial corneal dystrophy (FECD), a progressive polygenic disease that causes degeneration of the corneal endothelium, is the leading indication for corneal transplantation in the U.S. Characteristically, loss of corneal endothelial cells and formation of degenerative extracellular “guttae” result in failure to maintain appropriate corneal hydration through active ion pumping to counter the passive leakage of aqueous humor. FECD pathogenesis is linked to oxidative stress and environmental exposure to ultraviolet A (UVA), and impaired endogenous response to oxidative stress is common to the disease across multiple genotypes. UV irradiation is also known to cause cellular damage by ferroptosis, a nonapoptotic oxidative cell death resulting from iron-mediated lipid peroxidation. Although a possible role for ferroptosis in FECD has been postulated to account for the increased susceptibility to oxidative damage and lipid peroxidation, this has not been explored systematically. In this study, we investigated the roles of genetic background and UVA exposure in causing lipid membrane damage and endothelial cell degeneration in FECD. We first sought clinical evidence of iron-mediated lipid peroxidation in surgical samples obtained from FECD patients undergoing endothelial keratoplasty, and found increased levels of cytosolic ferrous iron (Fe2+) and evidence of lipid peroxidation were present in end-stage diseased tissues compared with healthy human donor corneas. Next, using immortalized (F35T) and primary cell cultures modeling the TCF4 intronic trinucleotide repeat expansion genotype, we found that alterations in gene and protein expression involved in ferroptosis were conserved in both cell culture and tissue models of FECD disease compared to controls, including elevated levels of the ferroptosis-specific marker transferrin receptor 1. F35T immortalized cells showed significantly higher basal lipid peroxidation than control HCEC-B4G12 cells, indicating higher susceptibility to ferroptosis attributable to genetic background. Then, we tested the impact of physiologically relevant doses of UVA irradiation on F35T cell cultures and found increased cytosolic Fe2+ iron levels indicating a role for ferroptosis in FECD disease progression. Finally, we tested the effects of inhibitory ferroptosis molecules. We found that F35T cells were more prone to RSL3 induced ferroptosis than healthy controls with deferoxamine chelation which indicated increased susceptibility in FECD cells, and cell death could be prevented after experimentally-induced ferroptosis using solubilized antioxidant ubiquinol indicating a role for anti-ferroptosis therapies. This investigation demonstrates that FECD genetic background and UVA exposure contribute to basal and incidental iron-mediated lipid peroxidation and cell death in FECD, and provides the basis for future investigations of ferroptosis-mediated oxidative damage and disease progression in FECD.

In patients with severe respiratory failure or severe cardiogenic shock, extracorporeal support is necessary to support organ function. To provide adequate organ perfusion, the intensivist needs to maintain an adequate forward flow of oxygenated blood, while also minimizing venous congestion. Since right ventricular dysfunction is common in such patients, monitoring and managing this aspect of hemodynamics are vitally important. Optimizing volume overload, and decreasing right ventricular preload as tolerated, is an important hemodynamic goal in these situations. However, overaggressive diuresis or ultrafiltration-guided fluid removal could lead to decreased flow through the extracorporeal circuit, hence decreasing crucial support to the cardiorespiratory systems. The establishment of V-V ECMO might decrease RV strain and improve cardiac function. For some patients with severe cardiogenic shock, a transition from veno-venous extracorporeal membrane oxygenation (V-V ECMO) to veno-arterial extracorporeal membrane oxygenation (V-A ECMO) might be required. In those with clear signs of acute cardiac failure, the establishment of right ventricular support might be essential right at the outset. Monitoring circuit parameters, as well as clinical and laboratory parameters, is crucial for managing volume status and right ventricular function. Point-of-care ultrasonography, including cardiac, respiratory, and abdominal evaluation, is particularly useful in tailoring therapy to patient-specific goals.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a respiratory virus that gains entry via angiotensin-converting enzyme 2 (ACE2) within airway epithelium. Patients exhibit a spectrum of respiratory symptoms from asymptomatic to respiratory failure. Patient factors including obesity, tobacco use, and black race are all associated with increased ACE2 expression and may contribute to increased complications. Consolidation and ground-glass opacities on chest imaging are typical but not specific for coronavirus disease 2019 (COVID-19). Venous thromboembolism occurs infrequently when prophylactic anticoagulation is provided. However, capillary microthrombosis is nearly ubiquitous, suggesting that it contributes to hypoxemia. Remdesivir and glucocorticoids may benefit some hospitalized patients. Many of those afflicted remain symptomatic two weeks following diagnosis and continue to require health care. Total lung capacity, diffusion capacity, and maximal oxygen consumption may be reduced for months in some survivors. Lung transplant offers chronically critically ill patients new hope, and this option may have increasing potential for outpatients with COVID-19-associated fibrosis.