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BackgroundDysfunction or loss of limbal stem cells can result in limbal stem cell deficiency (LSCD), a disease that cause corneal opacity, pain, and loss of vision. Cultivated limbal epithelial transplantation (CLET) can be used to restore stem cell niche homeostasis and replenish the progenitor pool. Transplantation has been reported with high success rate, but there is an unmet need of prognostic markers that correlate with clinical outcomes. To date, the progenitor content in the graft is the only parameter that has been retrospectively linked to success.MethodsIn this study, we investigate extracellular micro RNAs (miRNAs) associated with stem/progenitor cells in cultivated limbal epithelial cells (cLECs). Using micro RNA sequencing and linear regression modelling, we identify a miRNA signature in cultures containing high proportion of stem/progenitor cells. We then develop a robust RNA extraction workflow from culture media to confirm a positive miRNA correlation with stem/progenitor cell proportion.ResultsmiR-6723-5p is associated with cultures containing high proportion of stem/progenitor cells, and is detected in the basal layer of corneal epithelium.ConclusionsThese results indicate that miR-6723-5p could potentially serve as a stem/progenitor cell marker in cLECs.

Purpose: The purpose of this study was to assess trends in microbiology profiles and antibiotic susceptibility among pediatric infectious keratitis cases in Southern California, United States., Methods: This is a retrospective chart review of patients aged 18 years or younger diagnosed with infectious keratitis with a positive microbiology culture between 2006 and 2019 at a tertiary academic institution. Patient demographics, microbiology results, and antibiotic susceptibilities were analyzed., Results: There were 23 patients included in the study period. The mean age (±SD) was 11 ± 7 years, with 60% 10 years and older. 75% were female, and 45% were of Hispanic or Latino descent. A total of 14 patients (61%) presented between June and November. A total of 30 microorganisms were identified from 24 corneal cultures of 23 eyes, and 92% were bacterial. The most common gram-positive and gram-negative isolates were Staphylococcus spp. (5/13, 38%) and Pseudomonas spp. (9/15, 60%), respectively. 92.3% gram-positive isolates were susceptible to vancomycin, but only 7.7% were susceptible to erythromycin. Gram-negative isolates were susceptible to gentamicin (86%), ciprofloxacin (79%), and ceftazidime (55%). Pseudomonas spp. was highly susceptible to gentamicin (100%) and ciprofloxacin (83%), and it was moderately susceptible to tobramycin (44%) and ceftazidime (67%)., Conclusions: Pediatric microbial keratitis cases in Southern California were predominantly bacterial infections, with gram-positive isolates highly susceptible to vancomycin and gram-negative isolates, particularly Pseudomonas spp., showing high susceptibility to gentamicin and ciprofloxacin., Competing Interests: The authors have no funding or conflicts of interest to disclose., (Copyright © 2024 The Author(s). Published by Wolters Kluwer Health, Inc.)

Purpose: To investigate whether neurotrophic keratopathy is present in limbal stem cell deficiency (LSCD), by measuring corneal sensation and characterizing corneal subbasal nerve plexus., Design: Prospective, cross-sectional, case-control comparative study., Methods: A total of 46 eyes with LSCD and 14 normal eyes were recruited from 2019 to 2022. Corneal sensation was measured using a Cochet-Bonnet esthesiometer, and subbasal nerve plexus was imaged using in vivo confocal microscopy (IVCM) at the central cornea and 4 limbal regions. Subbasal nerve density (SND, number of nerves/mm 2 ), subbasal nerve length (SNL, total length of nerves/mm 2 ) and subbasal nerve branch density (SNBD, number of branches/mm 2 ) were quantified. LSCD was graded to stage 1, 2, and 3 using a previously established staging method consisting of clinical scores, basal cell density, central corneal epithelial thickness, and SNL., Results: The mean (±SD) cornea sensation in the central cornea and limbus were 29.2 ± 21.5 and 33.6 ± 15.1 mm in the LSCD group and 57.6 ± 5.8 and 54.3 ± 4.7 mm in the control group, respectively (all P < .001). In sectoral LSCD, the corneal sensation in the affected regions (29.1 ± 17.6 mm) decreased significantly compared to the unaffected regions (41.4 ± 18.2 mm, P < .001). Central corneal SND, SNL, and SNBD were reduced by 84.6%, 82.6%, and 89.2%, respectively, in LSCD compared to controls (all P < 0.05). The central corneal sensation negatively correlated with the severity of LSCD (rho = -0.64, P < .0001) and positively correlated with SND, SNL, and SNBD (rho = 0.63, 0.66, and 0.56, respectively; all P < .001)., Conclusions: Corneal sensation was reduced in eyes with LSCD. The degree of corneal sensation reduction positively correlated with the severity of LSCD. This finding demonstrated the coexistence of neurotropic keratopathy in LSCD., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: We previously reported that a single injection of (R,S)-ketamine or its metabolite (2S,6S)-hydroxynorketamine (HNK) prior to stress attenuated learned fear. However, whether these drugs attenuate learned fear through divergent or convergent effects on neural activity remains to be determined., Methods: 129S6/SvEv male mice were injected with saline, (R,S)-ketamine, or (2S,6S)-HNK 1 week before a 3-shock contextual fear conditioning paradigm. Five days later, mice were re-exposed to the aversive context and euthanized 1 hour later to quantify active cells. Brains were processed for c-fos immunoreactivity, and neural networks were built with a novel, wide-scale imaging pipeline., Results: We found that (R,S)-ketamine and (2S,6S)-HNK attenuated learned fear. Fear-related neural activity was altered in dorsal CA3 following (2S,6S)-HNK; ventral CA3 and CA1, infralimbic and prelimbic regions, insular cortex, retrosplenial cortex, piriform cortex, nucleus reuniens, and periaqueductal gray following both (R,S)-ketamine and (2S,6S)-HNK; and in the paraventricular nucleus of the thalamus (PVT) following (R,S)-ketamine. Dorsal CA3 and ventral hippocampus activation correlated with freezing in the (R,S)-ketamine group, and retrosplenial cortex activation correlated with freezing in both (R,S)-ketamine and (2S,6S)-HNK groups. (R,S)-ketamine increased connectivity between cortical and subcortical regions while (2S,6S)-HNK increased connectivity within these regions., Conclusions: This work identifies novel nodes in fear networks that involve the nucleus reuniens, piriform cortex, insular cortex, periaqueductal gray, and retrosplenial cortex that can be targeted with neuromodulatory strategies or pharmaceutical compounds to treat fear-induced disorders. This approach could be used to optimize target engagement and dosing strategies of existing medications., (Copyright © 2024 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.)

Venous thrombosis (VT) is a common vascular disease associated with reduced survival and a high recurrence rate. VT is initiated by the accumulation of platelets and neutrophils at sites of endothelial cell activation. A role for platelet αIIbβ3 in VT is not established, a task complicated by the increased bleeding risk caused by partial agonists such as tirofiban. Here, we show that m-tirofiban, a modified version of tirofiban, does not agonize αIIbβ3 based on lack of neoepitope expression and the cryo-EM structure of m-tirofiban/full-length αIIbβ3 complex. m-tirofiban abolishes agonist-induced platelet aggregation while preserving clot retraction ex vivo and, unlike tirofiban, it suppresses venous thrombogenesis in a mouse model without increasing bleeding. These findings establish a key role for αIIbβ3 in VT initiation and suggest that m-tirofiban and compounds with a similar structurally-defined mechanism of action merit consideration as potential thromboprophylaxis agents in patients at high risk for VT and hemorrhage., (© 2024. The Author(s).)

Purpose: Corneal subbasal nerve parameters have been previously reported using 2-dimensional scans of in vivo laser scanning confocal microscopy (IVCM) in eyes with limbal stem cell deficiency (LSCD). This study aims to develop and validate a method to better quantify corneal subbasal nerve parameters and changes from reconstructed 3-dimensional (3D) images., Methods: IVCM volume scans from 73 eyes with various degrees of LSCD (mild/moderate/severe) confirmed by multimodal anterior segment imaging including IVCM and 20 control subjects were included. Using ImageJ, the scans were manually aligned and compiled to generate a 3D reconstruction. Using filament-tracing semiautomated software (Imaris), subbasal nerve density (SND), corneal nerve fiber length, long nerves (>200 μm), and branch points were quantified and correlated with other biomarkers of LSCD., Results: 3D SND decreased in eyes with LSCD when compared with control subjects. The decrease was significant for moderate and severe LSCD ( P < 0.01). 3D SND was reduced by 3.7% in mild LSCD, 32.4% in moderate LSCD, and 96.5% in severe LSCD. The number of long nerves and points of branching correlated with the severity of LSCD ( P < 0.0001) and with declining SND (R 2 = 0.66 and 0.67, respectively). When compared with 2-dimensional scans, 3D reconstructions yielded significant increases of SND and branch points in all conditions except severe LSCD. 3D analysis showed a 46% increase in long nerves only in mild LSCD ( P < 0.01)., Conclusions: This proof-of-concept study validates the use of 3D reconstruction to better characterize the corneal subbasal nerve in eyes with LSCD. In the future, this concept could be used with machine learning to automate the measurements., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

Purpose: The purpose of this study was to compare the outcomes of manual and automated corneal epithelial thickness (CET) assessments using anterior segment optical coherence tomography in eyes with limbal stem cell deficiency (LSCD)., Methods: Eighty-seven eyes from 64 patients with LSCD and 65 eyes from 39 healthy subjects were included in this cross-sectional study. OCT images of corneas were acquired using a Fourier-domain anterior segment optical coherence tomography. The automated measurement of CET was obtained using the manufacturer's program and the manual measurements were obtained by 2 masked observers. The average CET was obtained from 3 consecutive epithelial profile maps. The average CET of manual measurement was obtained from 3 consecutive cross-line scans using 3-point measurement as previously described., Results: The central CET was thinner in patients with LSCD compared with healthy individuals by both manual and automated measurements. Automated CET measurements were similar to the manually obtained CET in the healthy controls (r = 0.911). However, there is a lower correlation between the automated and manually obtained CET in the LSCD group (r = 0.497). Manual measurements of CET (39.8 ± 11 μm) in eyes with LSCD were significantly lower than the automated measurements (48.8 ± 7.2 μm, P = 0.001, 95% CI -11.0 to -6.9). The correlations between the automated and manually obtained measurements in the mild, moderate, and severe stages of LSCD were 0.623, 0.632, and 0.378, respectively., Conclusions: There is a large discrepancy between the manual and automated measurements in eyes with LSCD but not in normal eyes. Automated measurements in LSCD exhibited a higher rate of errors and nonuniform measurements due to the presence of subepithelial scars. Automated CET measurements appear to be unreliable in eyes with corneal scars in LSCD., Competing Interests: The authors have no conflicts of interest to disclose., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

Background: Wnt proteins are crucial for embryonic development, stem cell growth, and tissue regeneration. Wnt signaling pathway is activated when Wnt proteins bind to cell membrane receptors., Methods and Results: We employed a luciferase reporter assay in HEK293STF cells to measure Wnt protein-induced signaling. We observed that Wnt3a uniquely promotes the Wnt/β-catenin pathway through positive cooperativity. Additionally, MFH-ND, a molecular mimic of Wnt ligands, markedly increased Wnt3a-induced signaling in a dose-responsive manner. This suggests that various Wnt ligands can synergistically enhance Wnt pathway activation., Conclusions: The study suggests the likelihood of various Wnt ligands coexisting in a single signalosome on the cell membrane, providing new insights into the complexities of Wnt signaling mechanisms., (© 2024. The Author(s).)

Venous thrombosis (VT) is a common vascular disease associated with reduced survival and a high recurrence rate. Previous studies have shown that the accumulation of platelets and neutrophils at sites of endothelial cell activation is a primary event in VT, but a role for platelet αIIbβ3 in the initiation of venous thrombosis has not been established. This task has been complicated by the increased bleeding linked to partial agonism of current αIIbβ3 inhibitory drugs such as tirofiban (Aggrastat ® ). Here, we show that m-tirofiban, an engineered version of tirofiban, is not a partial agonist of αIIbβ3. This is based on its cryo-EM structure in complex with human full-length αIIbβ3 and its inability to increase expression of an activation-sensitive epitope on platelet αIIbβ3. m-tirofiban abolished agonist-induced platelet aggregation ex vivo at concentrations that preserved clot retraction and markedly suppressed the accumulation of platelets, neutrophils, and fibrin on thrombin-activated endothelium in real-time using intravital microscopy in a mouse model of venous thrombogenesis. Unlike tirofiban, however, m-tirofiban did not increase bleeding at the thrombosis-inhibitory dose. These findings establish a key role for αIIbβ3 in the initiation of VT, provide a guiding principle for designing potentially safer inhibitors for other integrins, and suggest that pure antagonists of αIIbβ3 like m-tirofiban merit further consideration as potential thromboprophylaxis agents in patients at high-risk for VT and hemorrhage.

Purpose of Review: To highlight the progress and future direction of limbal stem cell (LSC) therapies for the treatment of limbal stem cell deficiency (LSCD)., Recent Findings: Direct LSC transplantation have demonstrated good long-term outcomes. Cultivated limbal epithelial transplantation (CLET) has been an alternative to treat severe to total LSCD aiming to improve the safety and efficacy of the LSC transplant. A prospective early-stage uncontrolled clinical trial shows the feasibility and safety of CLET manufactured under xenobiotic free conditions. Other cell sources for repopulating of the corneal epithelium such as mesenchymal stem cells (MSCs) and induced pluripotent stem cells are being investigated. The first clinical trials of using MSCs showed short-term results, but long-term efficacy seems to be disappointing. A better understanding of the niche function and regulation of LSC survival and proliferation will lead to the development of medical therapies to rejuvenate the residual LSCs found in a majority of eyes with LSCD in vivo. Prior efforts have been largely focused on improving LSC transplantation. Additional effort should be placed on improving the accuracy of diagnosis and staging of LSCD, and implementing standardized outcome measures which enable comparison of efficacy of different LSCD treatments for different severity of LSCD. The choice of LSCD treatment will be customized based on the severity of LSCD in the future., Summary: New approaches for managing different stages of LSCD are being developed. This concise review summarizes the progresses in LSC therapies for LSCD, underlying mechanisms, limitations, and future areas of development., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

Background: We have developed an algorithmic approach to laparoscopic cholecystectomy, including subtotal cholecystectomy, as a bailout strategy when the Critical View of Safety cannot be safely achieved due to significant inflammation and fibrosis of the hepatocystic triangle., Methods: This is a retrospective cohort study comparing postoperative outcomes in patients with severe cholecystitis who underwent laparoscopic cholecystectomy or laparoscopic subtotal cholecystectomy at St. Joseph's Health Centre from May 2016 to July 2021, as well as against a historical cohort. We further stratified laparoscopic subtotal cholecystectomy cases based on fenestrating or reconstituting subtype., Results: The cohort included a total of 105 patients who underwent laparoscopic cholecystectomy and 31 patients who underwent laparoscopic subtotal cholecystectomy. Bile leaks (25.8% vs 1.0%, relative risk 3.5, 95% confidence interval 3.5-208.4) were more common in the laparoscopic subtotal cholecystectomy group. Postoperative endoscopic retrograde cholangiopancreatography (22.6% vs 3.8%, relative risk 5.9, 95% confidence interval 1.9-18.9) and biliary stent insertion (19.4% vs 1.0%, relative risk 20.3, 95% confidence interval 2.5-162.5) were also more common in the laparoscopic subtotal cholecystectomy group. Bile leaks in laparoscopic subtotal cholecystectomy were only documented in the fenestration subtype, most of which were successfully managed with endoscopic retrograde cholangiopancreatography and biliary stenting. Compared to our previous study of laparoscopic cholecystectomy and subtotal cholecystectomy for severe cholecystitis between 2010 and 2016, there has been a decrease in postoperative laparoscopic cholecystectomy complications, subtotal cholecystectomy cases, and no bile duct injuries., Conclusion: Following our algorithmic approach to safe laparoscopic cholecystectomy has helped to prevent bile duct injury. Laparoscopic cholecystectomy remains the gold standard for the management of severe cholecystitis; however, in extreme cases, laparoscopic subtotal cholecystectomy is a safe bailout strategy with manageable morbidity., (Crown Copyright © 2023. Published by Elsevier Inc. All rights reserved.)

Yi Mai Jian herbal formula (YMJ) is formulated with Eucommiae Folium, Astragali Radix, Ligustri Lucidi Fructus, and Elaeagnus Fructus to improve bone function in traditional Chinese medicine. The anti-osteoporotic effects of YMJ in bone metabolism were evaluated in ovariectomized (OVX) rats. The skeletal structure of the femur and vertebrae was analyzed after treating OVX rats with YMJ for 114 days. The results showed that YMJ significantly increased the bone mineral density (BMD) and trabecular number (Tb. N) of the femur and 5th lumbar vertebrae and reduced trabecular separation (Tb. Sp). Moreover, trabecular bone volume/total tissue volume (BV/TV), bone stiffness, and maximum femur load were significantly increased. The serum concentrations of NTX1 and PYD were significantly decreased. According to these results, YMJ could ameliorate osteoporosis in ovariectomized rats. Eucommiae Folium and Elaeagnus Fructus inhibited osteoclast differentiation, Ligustri Lucidi Fructus inhibited calcium reabsorption, Astragali Radix stimulated osteoblast proliferation, and Astragali Radix and Eucommiae Folium stimulated mineralization. Therefore, the combination of the four herbs into one formula, YMJ, could alleviate bone remodeling caused by low estrogen levels. We suggest that YMJ could be a healthy food candidate for preventing post-menopausal osteoporosis., Competing Interests: BS, D-SN, and H-WZ, were employed by the Infinitus (China) Company Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Shi, Lin, Lee, Ning, Lin, Zhao, Yang, Deng, Chiu and Wang.)

Machine learning (ML) has been used for many clinical decision-making processes and diagnostic procedures in bioinformatics applications. We examined eight algorithms, including linear discriminant analysis (LDA), logistic regression (LR), k-nearest neighbor (KNN), random forest (RF), gradient boosting machine (GBM), support vector machine (SVM), Naïve Bayes classifier (NB), and artificial neural network (ANN) models, to evaluate their classification and prediction capabilities for four tissue types in Wolfiporia extensa using their monosaccharide composition profiles. All 8 ML-based models were assessed as exemplary models with AUC exceeding 0.8. Five models, namely LDA, KNN, RF, GBM, and ANN, performed excellently in the four-tissue-type classification (AUC > 0.9). Additionally, all eight models were evaluated as good predictive models with AUC value > 0.8 in the three-tissue-type classification. Notably, all 8 ML-based methods outperformed the single linear discriminant analysis (LDA) plotting method. For large sample sizes, the ML-based methods perform better than traditional regression techniques and could potentially increase the accuracy in identifying tissue samples of W. extensa., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

As a major neuron type in the brain, the excitatory neuron (EN) regulates the lifespan in C. elegans. How the EN acquires senescence, however, is unknown. Here, we show that growth differentiation factor 11 (GDF11) is predominantly expressed in the EN in the adult mouse, marmoset and human brain. In mice, selective knock-out of GDF11 in the post-mitotic EN shapes the brain ageing-related transcriptional profile, induces EN senescence and hyperexcitability, prunes their dendrites, impedes their synaptic input, impairs object recognition memory and shortens the lifespan, establishing a functional link between GDF11, brain ageing and cognition. In vitro GDF11 deletion causes cellular senescence in Neuro-2a cells. Mechanistically, GDF11 deletion induces neuronal senescence via Smad2-induced transcription of the pro-senescence factor p21. This work indicates that endogenous GDF11 acts as a brake on EN senescence and brain ageing., (© 2023. The Author(s).)

Limbal stem cells (LSCs) are adult stem cells located at the limbus ensuring the continuous renewal of the corneal epithelium, critical to maintain an optimal visual function. Damages to the LSCs or their niche microenvironment lead to limbal stem cell deficiency (LSCD), a potentially blinding disease. Transplantation of LSCs as a treatment for severe to total LSCD has gained popularity since 1980s, owing to the clinical success of the first direct limbal autograft transplantation. Recent advances in the understanding of the LSCs' molecular identity and regulation have enabled preclinical and clinical advancements of promising LSCs therapies. However, lack of standardization of the diagnostic methods, staging of the disease severity, manufacturing process, and clinical outcome measures have hindered the advancement of the therapy. To move these therapies to the clinic, optimization and standardization of the diagnostic strategy, cell product manufacturing, and assessment of clinical efficacy with potency assays are key points to the development of customized therapies. Recent findings suggest that residual LSCs exist in eyes presenting with clinical signs of total LSCD, which opens new therapeutic strategies for eyes with partial LSCD. Prospective, randomized, multicentric controlled clinical trials are necessary to determine the efficacy of different LSCs therapies for different stages of LSCD using a set of standardized outcome measures., (© The Author(s) 2023. Published by Oxford University Press.)

Introduction: Limbal Stem Cell Deficiency (LSCD) is a blinding corneal disease characterized by the loss of function or deficiency in adult stem cells located at the junction between the cornea and the sclera (i.e., the limbus), namely the limbal stem cells (LSCs). Recent advances in in vivo imaging technology have improved disease diagnosis and staging to quantify several biomarkers of in vivo LSC function including epithelial thickness measured by anterior segment optical coherence tomography, and basal epithelial cell density and subbasal nerve plexus by in vivo confocal microscopy. A decrease in central corneal sub-basal nerve density and nerve fiber and branching number has been shown to correlate with the severity of the disease in parallel with increased nerve tortuosity. Yet, image acquisition and manual quantification require a high level of expertise and are time-consuming. Manual quantification presents inevitable interobserver variability., Methods: The current study employs a novel deep learning approach to classify neuron morphology in various LSCD stages and healthy controls, by integrating images created through latent diffusion augmentation. The proposed model, a residual U-Net, is based in part on the InceptionResNetV2 transfer learning model., Results: Deep learning was able to determine fiber number, branching, and fiber length with high accuracy (R2 of 0.63, 0.63, and 0.80, respectively). The model trained on images generated through latent diffusion on average outperformed the same model when trained on solely original images. The model was also able to detect LSCD with an AUC of 0.867, which showed slightly higher performance compared to classification using manually assessed metrics., Discussion: The results suggest that utilizing latent diffusion to supplement training data may be effective in bolstering model performance. The results of the model emphasize the ability as well as the shortcomings of this novel deep learning approach to predict various nerve morphology metrics as well as LSCD disease severity., Competing Interests: SD is a consultant for Novartis US, Amgen, Cellusion, Kala Pharmaceuticals, and Claris Biotherapeutics, Inc. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Gibson, Tran, Raveendran, Bonnet, Siu, Vinet, Stoddard-Bennett, Arnold, Deng and Speier.)

Limbal epithelial stem/progenitor cells (LSCs) are adult stem cells located at the limbus, tightly regulated by their niche involving numerous signaling pathways, such as Wnt. Wnt proteins are secreted morphogens that play critical roles in embryonic development, stem cell proliferation, self-renewal, tissue regeneration, and remodeling in adults. It has been shown that a small molecule Wnt mimic could improve LSCs expansion ex vivo. Damage to the LSCs and/or their niche can lead to limbal stem cell deficiency (LSCD), a condition that can cause corneal blindness and is difficult to treat. This study explored if repopulating residual LSCs in partial LSCD through Wnt activation could be a novel therapeutic approach. To mimic LSCD due to a chemical injury, single cultured LSCs were exposed to various concentrations of sodium hydroxide. A progressive loss of the LSCs phenotype was observed: the percentage of p63 bright cells and cytokeratin (K)14 + cells decreased while the percentage of K12 + increased. Wnt activation was attained by treating the LSCs with lithium chloride (LiCl) and a small-molecule Wnt mimic, respectively. After 18 h of treatment, LSCs proliferation was increased, and the LSCs phenotype was recovered, while the untreated cells did not proliferate and lost their phenotype. The percentage of p63 bright cells was significantly higher in the Wnt mimic-treated cells compared with untreated cells, while the percentage of K12 + cells was significantly lower. These findings suggest that local Wnt activation may rescue LSCs upon alkaline injury., (© 2023. Springer Nature Limited.)

Phosphodiesterase 5 (PDE5) is a cyclic guanosine monophosphate-degrading enzyme involved in numerous biological pathways. Inhibitors of PDE5 are important therapeutics for the treatment of neurodegenerative diseases, including Alzheimer's disease (AD). We previously reported the first generation of quinoline-based PDE5 inhibitors for the treatment of AD. However, the short in vitro microsomal stability rendered them unsuitable drug candidates. Here we report a series of new quinoline-based PDE5 inhibitors. Among them, compound 4b, 8-cyclopropyl-3-(hydroxymethyl)-4-(((6-methoxypyridin-3-yl)methyl)amino)quinoline-6-carbonitrile, shows a PDE5 IC 50 of 20 nM and improved in vitro microsomal stability (t 1/2  = 44.6 min) as well as excellent efficacy in restoring long-term potentiation, a type of synaptic plasticity to underlie memory formation, in electrophysiology experiments with a mouse model of AD. These results provide an insight into the development of a new class of PDE5 inhibitors for the treatment of AD., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: OA is a founder of Neurokine Therapeutics. EZ, EA, HZ, AS and DP declare that they have no competing interests. OA, SXD, DWL and JF have designed and licensed a series of PDE5 inhibitors to Aribio Co., LTD., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Quantifying the contribution of transport processes to air pollution events has been a prominent challenge and an important need in regional air pollution prevention and control. The WRF-Chem model was used to simulate a typical regional ozone (O 3 ) pollution event in Foshan, and the four-dimensional flux method was applied to quantify the transport fluxes of ozone and its precursors from the surrounding areas to Foshan and to clarify the contributions of the direct transport of ozone and transport of precursors. The average ozone flux from the surrounding areas to Foshan was 120.3 t·h -1 , the volatile organic compound (VOCs) flux was 30.2 t·h -1 , and the corresponding ozone formation potential (OFP) was 114.8 t·h -1 . By counting the transport fluxes of each ozone pollution event, it was found that the city with the largest ozone flux into Foshan during the pollution period was Guangzhou (contributed 44%); the city with the largest input VOCs flux was Zhaoqing (contributed 48%). The analysis of ozone generation potential due to transported VOCs emissions found that oxygenated volatile organic compounds (OVOCs) contributed the most to OFP, accounting for 47% of the "maximum input events." OVOCs and aromatic hydrocarbons such as formaldehyde, xylenes, aldehydes, acetone, and phenols were the top five species contributing to the OFP, contributing more than 50% of the total OFP, mainly from industrial solvent sources.

Purpose of Review: To highlight the progress and future direction of stem-cell based regenerative therapies for the treatment of corneal disease., Recent Findings: Corneal stem cell-based therapies, such as limbal stem cell transplantation, corneal stromal stem cell transplantation, endothelial stem cell transplantation, and stem cell-derived extracellular vesicles have demonstrated promising results in the laboratory. Although most are still in preclinical development or early phase clinical trials, these stem cell-based therapies hold potential to facilitate tissue regeneration, restore native function, and inhibit pathologic disease processes such as fibrosis, inflammation, and neovascularization., Summary: Stem cell-based therapy offers a promising therapeutic option that can circumvent several of the challenges and limitations of traditional surgical treatment. This concise review summarizes the progress in stem-cell based therapies for corneal diseases along with their history, underlying mechanisms, limitations, and future areas for development., (Copyright © 2023 Wolters Kluwer Health, Inc. All rights reserved.)

Ethnopharmacological Relevance: Ji-Ming-Shan (JMS) is a traditional herbal prescription consisting of seven herbs including Areca cathechu Burm.f., Citrus reticulata Blanco, Chaenomeles speciosa (Sweet) Nakai, Euodia ruticarpa (A. Juss.) Benth., Perilla frutescens (L.) Britton, Zingiber officinale Roscoe, Platycodon grandiflorus (Jacq.). It was first recorded during the Song dynasty and has been used extensively for protection against rheumatism, treatment of swelling of tendons, relief from foot pain, gout and diuresis and other forms of inflammation., Aim of the Study: The aim of this study is to evaluate the anti-inflammatory and anti-osteoarthritis activity of JMS extracts with the use of different cell lines (RAW 264.7 cells, SW1353 cells and primary cultured rat chondrocytes). MIA-induced rat animal models were used to assess the anti-osteoarthritis activity of the extract., Materials and Methods: This study investigated the anti-inflammatory activity of JMS-95E on LPS-induced RAW 264.7 macrophages and IL-1β-stimulated chondrocytes. For the in vivo study, male Wistar rats were used and they were randomly assigned in different groups: blank, control, positive control and three different JMS-95E treatment groups (200, 400, 800 mg/kg/d). Paw edema, hind-limb weight bearing, serum inflammatory cytokines including hematoxylin and eosin (HE) staining experiments were used to assess the efficacy of the extract in the rat model., Result: JMS 95% ethanol extract (JMS-95E, marker substance: narirutin (5.10 mg/g) and hesperidin (11.33 mg/g) has been identified in the extract using high pressure liquid chromatography. For in vitro assays, JMS-95E did not exhibit cytotoxicity and was able to downregulate the protein expression of iNOS, COX-2 and MMP-13. The production of inflammatory mediators such as NO and PGE 2 were also reduced with an increase in dose-dependent manner in various cell lines. Inhibitory activity on the key enzyme xanthine oxidase was also observed in this study. In rat animal models, JMS-95E reduced the inflammatory responses such as acute swelling, chondrocyte degradation and pain section of paw edema in rat model. Molecular marker studies of inflammation demonstrated that JMS-95E significantly decrease PGE 2 expression in MIA model., Conclusion: JMS-95E inhibited the inflammatory pathway leading to the production or expression levels of NO, iNOS, COX-2 and PGE 2 in macrophage cells. In primary cultured rat chondrocytes iNOS and SW1353 MMP-13 expression were downregulated after JMS-95E treatment. For the in vivo study JMS-95E significantly reduced the paw volume of carrageenan-induced rat paw edema through each dose and significantly inhibited paw volume, counterweight the distribution of hind-paw weight bearing through the MIA model which means JMS-95E could promote recovery of the acute swelling and chondrocyte degradation of the ankle joints. The above results provided the multiple mechanism of JMS-95E in OA treatment of the scientific founding which supported the description of JMS in traditional use., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Limbal epithelial stem/progenitor cells (LSCs) are adult stem cells located at the limbus, tightly regulated by their close microenvironment. It has been shown that Wnt signaling pathway is crucial for LSCs regulation. Previous differential gene profiling studies confirmed the preferential expression of specific Wnt ligands (WNT2, WNT6, WNT11, WNT16) and Wnt inhibitors (DKK1, SFRP5, WIF1, FRZB) in the limbal region compared to the cornea. Among all frizzled receptors, frizzled7 (Fzd7) was found to be preferentially expressed in the basal limbal epithelium. However, the exact localization of Wnt signaling molecules-producing cells in the limbus remains unknown. The current study aims to evaluate the in situ spatial expression of these 4 Wnt ligands, 4 Wnt inhibitors, and Fzd7. Wnt ligands, DKK1, and Fzd7 expression were scattered within the limbal epithelium, at a higher abundance in the basal layer than the superficial layer. SFRP5 expression was diffuse among the limbal epithelium, whereas WIF1 and FRZB expression was clustered at the basal limbal epithelial layer corresponding to the areas of high levels of Fzd7 expression. Quantitation of the fluorescence intensity showed that all 4 Wnt ligands, 3 Wnt inhibitors (WIF1, DKK1, FRZB), and Fzd7 were highly expressed at the basal layer of the limbus, then in a decreasing gradient toward the superficial layer (P < 0.05). The expression levels of all 4 Wnt ligands, FRZB, and Fzd7 in the basal epithelial layer were higher in the limbus than the central cornea (P < 0.05). All 4 Wnt ligands, 4 Wnt inhibitors, and Fzd7 were also highly expressed in the limbal stroma immediately below the epithelium but not in the corneal stroma (P < 0.05). In addition, Fzd7 had a preferential expression in the superior limbus compared to other limbal quadrants (P < 0.05). Taken together, the unique expression patterns of the Wnt molecules in the limbus suggests the involvement of both paracrine and autocrine effects in LSCs regulation, and a fine balance between Wnt activators and inhibitors to govern LSC fate., Competing Interests: Declaration of competing interest SXD is a consultant for Dompe US, Novartis US, and Claris Biotherapeutics, Inc.; other authors: none., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Aims: To evaluate the clinical and visual outcomes of fluid-filled scleral lens devices (SL) wear in patients with limbal stem cell deficiency (LSCD)., Design: Retrospective consecutive case series., Methods: 27 eyes with LSCD confirmed by in vivo confocal microscopy at the Stein Eye Institute and fitted with SL were included. Correlations between corrected distance visual acuity (CDVA) and LSCD stage determined by clinical grading were performed between baseline (after the SL fit) and the last follow-up (the time of discontinuation of SL wear or the last visit in eyes in which SL were continued). In a subset of patients that had worsened LSCD while using SL, anterior segment optical coherence tomography (AS-OCT) and anterior segment fluorescein angiogram (AS-FA) were performed., Results: Baseline LSCD grading was stage I in 12 eyes (44.4%), stage 2 in 12 eyes (44.4%), and stage III in 3 eyes (11.1%). At the last follow-up, CDVA was improved in 7 eyes (25.9%), remained stable in 13 eyes (48.1%) and decreased in 7 eyes (25.9%, P = 0.16). The LSCD stage was improved in 7 eyes (25.9%), remained stable in 8 eyes (29.6%) and worsened in 12 eyes (44.4%, P = 0.10). AS-OCT and AS-FA, performed in 5 eyes, showed limbal compression and delayed fluorescein filling., Conclusion: SL can improve visual acuity and maintain the ocular surface in the majority of eyes. Worsening of the ocular surface might be a result of limbal hypoxia. Close monitoring of SL fit is necessary in these compromised eyes., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021. Published by Elsevier Ltd.)

Purpose: To evaluate safety and efficacy of autologous serum eye drops (AS) in the treatment of limbal stem cell deficiency (LSCD) associated with glaucoma surgery., Methods: Retrospective case series of eyes with glaucoma surgery-induced LSCD treated with AS. Diagnosis of LSCD was confirmed by anterior segment optical coherence tomography, in vivo confocal microscopy, and/or impression cytology. Limbal stem cell deficiency severity was staged using a clinical scoring system (2-10 points). Outcome measures were changes (≥2 points) of the LSCD score and best-corrected visual acuity (BCVA) from the baseline to the last follow-up., Results: Thirteen eyes of 12 consecutive patients treated with 50% AS for at least 3 months were included. The mean age was 78.9±7.5 years and the mean duration of AS use was 20.9±16.8 months. Indications of AS included LSCD progression in eight eyes (61.5%) and visual axis threatening in five eyes (38.5%). The mean LSCD score at baseline (6.7±1.6) was similar to that at last follow-up (6.5±2.2, P =0.625). Two eyes (15.4%) showed improvement, nine eyes (69.2%) were stable, and two eyes (15.4%) worsened. The mean baseline BCVA (0.89±0.64 logMAR) was similar to the mean final BCVA (1.05±0.63 logMAR, P =0.173). There were no serious adverse complications related to AS., Conclusion: AS appears to be well tolerated and may stabilize the progression of LSCD with limited effects. A larger study is necessary to confirm the findings., Competing Interests: The authors have no funding or conflicts of interest to disclose., (Copyright © 2022 Contact Lens Association of Ophthalmologists.)

With significant advancement and development of extracellular vesicle (EV)-based therapies, there is a growing need to understand how their storage affects their physical and functional characteristics. EVs were isolated from the conditioned medium of a corneal stromal stem cell line (imCSSC) using Total Exosome isolation kit (TEI) and ultracentrifugation (UC) combined protocol. Purified EVs were stored at 4°C, - 80°C, room temperature (RT) after lyophilization with or without trehalose for 4 weeks. EVs stored at - 80°C and RT (lyophilization with trehalose) demonstrated a comparable morphology, while the freeze-dried samples without trehalose showed aggregation and degradation under a transmission electron microscope (TEM). Lyophilized samples without trehalose demonstrated a decreased particle concentration, recovery rate and protein concentration, which was remediated by the addition of trehalose. EVs stored at - 80℃ showed no change in the protein expression of CD9, CD63, and CD81. Regardless of the storage condition, all EV samples investigated reduced inflammation, as well as inhibited expression of fibrotic markers in vitro. Lyophilization of EVs with trehalose was a feasible storage method that retained the physical property and in vitro biological activities of EVs after 4 weeks of storage, while - 80°C offered the best retention of imCSSC-derived EV physical properties. For the first time, this data demonstrated a practical and translatable method for the storage of CSSC-derived EVs for clinical use., (© 2022. The Author(s).)

Background: MRI-based breast cancer screening for high-risk women has been associated with false positives. This study explored the benefits and drawbacks of MRI-based screening using in-depth patient interviews., Methods: This was a qualitative study of interviews with women participating in the High Risk Ontario Breast Screening Program. Women enrolled at two centers who had completed at least one round of screening were invited to participate. Recruitment was suspended when thematic saturation was reached. Semi-structured telephone interviews were conducted and transcribed verbatim. Emergent themes were identified and a coding framework established., Results: 21 women (median age 41 years) participated in telephone interviews. Women had been in the program a median of 4 years (IQR 1-5), and 71% had experienced at least one abnormal screen. Eight participants (38%) had undergone biopsies. Six women (29%) were BRCA mutation carriers. MRIs were described as intimidating, uncomfortable, and claustrophobic. Participants were concerned about long-term exposure to gadolinium contrast. Compared to MRI, mammography alone was viewed as painful, less sensitive, and a "useless…waste of time." MRI provided a "psychological safety net" that outweighed the distress associated with abnormal screens. Many women accepted this trade-off as a "two-edged sword" that was "worth it" and provided a sense of control. Suggestions for improvement included more information regarding the risks of MRI, and access to counselling., Conclusions: Women participating in MRI-based screening strongly value reassurance from a highly sensitive screening test. This outweighed the distress of abnormal screens. There are areas for improvement around patient communication and psychosocial support., Competing Interests: Competing interests None to declare., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Laparoscopic subtotal cholecystectomy (LSC) is a bailout strategy to prevent bile duct injury in difficult gallbladder cases. It is associated with acceptable morbidity that is readily managed with postoperative interventions. Here we share our techniques for LSC. We begin with landmarking, which includes the line of safety, a theoretical line the sulcus of Rouvière and the junction of the cystic and hilar plates. If the fundus can be grasped, then the gallbladder is dissected off the cystic plate using the top-around approach. The gallbladder is then amputated, creating a short cuff of proximal gallbladder. This cuff can be left patent (2A) or cinched close with an ENDOLOOP (Ethicon) if it is small, ideally less than 1 cm (1A). If the fundus cannot be grasped, then an inverted T incision is made on the anterior gallbladder wall. The longitudinal incision is extended toward the fundus, and the transverse incision is extended superiorly along the cystic plate edge. Two "bunny ears" are developed and ultimately resected to excise the anterior gallbladder wall at an oblique angle while leaving the posterior wall intact (2B). If the remaining cuff is small, then it can be sutured closed against the gallbladder back wall (1B). In the setting of extensive bowel adhesion to the anterior gallbladder, we perform a fundectomy, from which we extend two incisions along the cystic plate to open the gallbladder like a clamshell. Our paper describes and illustrates our St Joseph's Health Centre institutional LSC approach and subtype classification (1A, 1B, 2A, and 2B)., (Copyright © 2022 by the American College of Surgeons. Published by Wolters Kluwer Health, Inc. All rights reserved.)

Endothelial keratoplasty has become the standard for the treatment of endothelial dysfunction. In Descemet membrane endothelial keratoplasty (DMEK), only the endothelium and Descemet membrane are transplanted, providing superior outcomes compared to Descemet stripping endothelial keratoplasty (DSEK). A substantial subset of patients who require DMEK have comorbid glaucoma. Even in eyes with complex anterior segment such as eyes with previous trabeculectomy or tube shunts, DMEK can restore meaningful vision and outperforms DSEK in terms of visual recovery, decreased rejection rate, and the need for high dose of topical steroids. However, accelerated endothelial cell loss and secondary graft failure have been described in eyes with previous glaucoma surgery, namely trabeculectomy and drainage device. During DMEK and DSEK procedures, raised intraocular pressure is required to attach the graft, which could worsen preexisting glaucoma or cause de novo glaucoma. Mechanisms of postoperative ocular hypertension include delayed air clearance, pupillary block, steroid response, and damage to angle structures. Medically treated glaucoma has increased risk for postoperative ocular hypertension. By understanding these additional complications and making appropriate modifications in surgical techniques and postoperative management, DMEK can be performed successfully and achieve very good visual outcome in eyes with glaucoma. Such modifications include precisely controlled unfolding technique, iridectomies that can help avoid pupillary block, tube shunts that can be trimmed to facilitate graft unfolding, air fill tension that can be adjusted, and postoperative steroid regimens that can be modified to decrease the risk for steroid response. Long-term survival of the DMEK graft, however, is shorter in eyes with previous glaucoma surgery than those without, as observed after other types of keratoplasty., Competing Interests: The authors declare that there are no conflicts of interests of this paper., (Copyright: © 2022 Taiwan Journal of Ophthalmology.)

In addition to their therapeutic potential in regenerative medicine, human corneal stromal stem cells (CSSCs) could serve as a powerful tool for drug discovery and development. Variations from different donors, their isolation method, and their limited life span in culture hinder the utility of primary human CSSCs. To address these limitations, this study aims to establish and characterize immortalized CSSC lines (imCSSC) generated from primary human CSSCs. Primary CSSCs (pCSSC), isolated from human adult corneoscleral tissue, were transduced with ectopic expression of hTERT, c-MYC, or the large T antigen of the Simian virus 40 (SV40T) to generate imCSSC. Cellular morphology, proliferation capacity, and expression of CSSCs specific surface markers were investigated in all cell lines, including TNFAIP6 gene expression levels in vitro, a known biomarker of in vivo anti-inflammatory efficacy. SV40T-overexpressing imCSSC successfully extended the lifespan of pCSSC while retaining a similar morphology, proliferative capacity, multilineage differentiation potential, and anti-inflammatory properties. The current study serves as a proof-of-concept that immortalization of CSSCs could enable a large-scale source of CSSC for use in regenerative medicine.

Purpose: The purpose of this study was to determine longitudinal trends in prevalence and resistance profiles for infectious keratitis at referral centers in Southern California., Methods: Cultured infectious keratitis cases from January 1, 2006, through December 31, 2009, and January 1, 2016, through December 31, 2019, at the University of California, Los Angeles, were evaluated. Outcome measures included microbial isolate prevalence and antibiotic/antifungal susceptibility and resistance patterns., Results: One hundred thirty-nine and 315 culture-positive isolates were obtained between 2006-2009 and 2016-2019, respectively. Gram-positive organisms accounted for 65% (2006-2009) and 74% (2016-2019) of bacterial isolates ( P = 0.076). Staphylococcus infections, the most common gram-positive and bacterial isolate in both study epochs, demonstrated increased prevalence from 2006-2009 to 2016-2019 (41% vs. 53%, P = 0.019). Although coagulase-negative Staphylococcus (CoNS) increased from 40% to 58% ( P = 0.0012), the prevalence of methicillin-resistant Staphylococcus aureus was unchanged (28% vs. 28%, P = 0.99). Pseudomonas aeruginosa , the most common gram-negative organism, demonstrated decreased prevalence from 18% to 10% ( P = 0.027). Candida species comprised 3.5% of culture-positive isolates in both epochs. All gram-positive isolates were susceptible to vancomycin, and all Staphylococcus isolates were susceptible to linezolid. Pseudomonas aeruginosa remained susceptible to tested fluoroquinolones (>93%) and aminoglycosides (100%) over time., Conclusions: In southern California between 2006 and 2019, there was a shift toward Staphylococcus species, with increased CoNS, decreased methicillin-sensitive Staphylococcus aureus , and decreased prevalence of P. aeruginosa . Empiric therapy of vancomycin and a fluoroquinolone or aminoglycoside provides effective antibacterial coverage for predominant bacterial species when culture sensitivities are pending., Competing Interests: S. X. Deng received grant support from the National Eye Institute (R01 EY021797 and R01 EY028557) and California Institute for Regenerative Medicine (CLIN1-08686 and CLIN2-11650) to study limbal stem cells. S. X. Deng is a consultant for Dompe US. The remaining authors have no conflicts of interest to disclose., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)

Purpose: The aim of this study was to investigate basal epithelial cell morphology (CM) in the central cornea and limbal areas of eyes with limbal stem cell deficiency (LSCD)., Methods: This was a prospective, cross-sectional comparative study. We developed a CM scoring system based on basal epithelial cell phenotypes graded from 0 (normal) to 3 (severe morphologic alterations); this system was evaluated by 2 independent masked observers. The CM score was compared with the LSCD clinical score, mean best-corrected visual acuity, and in vivo laser scanning confocal microscopy parameters used to stage LSCD (ie, basal epithelial cell density, basal epithelial thickness, and subbasal corneal nerve fiber length density)., Results: One hundred sixty-eight eyes with LSCD and 63 normal eyes were included. Compared with the control group, the LSCD group had significantly higher mean (±SD) CM scores in the central cornea (1.8 ± 0.7 vs. 0.5 ± 0.4, respectively; P = 0.01) and limbal areas (1.6 ± 0.2 vs. 1.3 ± 0.0, respectively; P < 0.05). The mean CM score in the central cornea was positively correlated with the clinical score ( P < 0.01, r = 0.66) and negatively correlated with the best-corrected visual acuity ( P < 0.01, r = 0.42). The CM scores were positively correlated with all other in vivo laser scanning confocal microscopy parameters in the central cornea and limbal areas (all P < 0.001)., Conclusions: Basal epithelial CM is altered in the central cornea and limbus of eyes with LSCD and thus can be used to stage the clinical severity of the disease., Competing Interests: S. X. Deng is a consultant for Dompe US and Claris Biotherapeutics, Inc. The remaining authors have no conflicts of interest to disclose., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)

Purpose: The purpose of this study was to evaluate the expression of cytokeratin (K) 13 on the corneal surface and to validate its application in the diagnosis of limbal stem cell deficiency (LSCD)., Methods: This prospective comparative study included 26 corneal impression cytology (IC) specimens from patients diagnosed with LSCD. Twenty-three IC specimens from normal donors served as controls. K12 and K13 expression were detected on the IC specimens by immunohistochemistry study. The number of K12 + or K13 + cells in all areas of the IC was quantified using ImageJ software., Results: The epithelial cells harvested from IC specimens from control corneas were all K12 + . In eyes with LSCD, K13 + and K12 + /K13 + cells accounted for 93.8% and 2.6%, respectively, in the cornea. In eyes with sectoral LSCD, the median number of K13 + cells in the clinically affected area was higher than that in the unaffected area (810.0 vs. 115.0 cells/mm 2 ; P < 0.001). No significant correlation was found between the LSCD severity and the number of K12 + cells (r = -0.284, P = 0.16) or K13 + cells (r = -0.011, P = 0.95). The presence of at least 16 K13 + cells/mm 2 was suggestive of LSCD., Conclusions: Identification of K13 + cells on IC specimens provides a simple and reliable method to detect conjunctival epithelial cells on the cornea. K13 is a marker for diagnosing LSCD and localizing the involved area in sectoral LSCD., Competing Interests: S. X. Deng is a consultant for Dompe US and Claris Biotherapeutics, Inc. The remaining authors have no conflicts of interest to disclose., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)

Background and Objectives: Total kidney volume is a validated prognostic biomarker for autosomal dominant polycystic kidney disease. Total kidney volume by magnetic resonance imaging (MRI) and manual segmentation is considered the "reference standard," but it is time consuming and not readily accessible. By contrast, three-dimensional (3D) ultrasound provides a promising technology for total kidney volume measurements with unknown potential. Here, we report a comparative study of total kidney volume measurements by 3D ultrasound versus the conventional methods by ultrasound ellipsoid and MRI ellipsoid., Design, Setting, Participants, & Measurements: This single-center prospective study included 142 patients who completed a standardized 3D ultrasound and MRI. Total kidney volumes by 3D ultrasound and ultrasound ellipsoid were compared with those by MRI. We assessed the agreement of total kidney volume measurements by Bland-Altman plots and misclassification of the Mayo Clinic imaging classes between the different imaging methods, and we assessed prediction of Mayo Clinic imaging classes 1C-1E by average ultrasound kidney length >16.5 cm., Results: Compared with MRI manual segmentation, MRI ellipsoid, 3D ultrasound, and ultrasound ellipsoid underestimated total kidney volume (mean difference: -3%, -9%, and -11%, respectively), with Mayo Clinic imaging classes misclassified in 11%, 21%, and 22% of patients, respectively; most misclassified cases by MRI ellipsoid (11 of 16), 3D ultrasound (23 of 30), and ultrasound ellipsoid (26 of 31) were placed into a lower Mayo Clinic imaging class. Predictions of the high-risk Mayo Clinic imaging classes (1C-1E) by MRI ellipsoid, 3D ultrasound, and ultrasound ellipsoid all yielded high positive predictive value (96%, 95%, and 98%, respectively) and specificity (96%, 96%, and 99%, respectively). However, both negative predictive value (90%, 88%, and 95%, respectively) and sensitivity (88%, 85%, and 94%, respectively) were lower for 3D ultrasound and ultrasound ellipsoid compared with MRI ellipsoid. An average ultrasound kidney length >16.5 cm was highly predictive of Mayo Clinic imaging classes 1C-1E only in patients aged ≤45 years., Conclusions: Total kidney volume measurements in autosomal dominant polycystic kidney disease by 3D ultrasound and ultrasound ellipsoid displayed similar bias and variability and are less accurate than MRI ellipsoid. Prediction of high-risk Mayo Clinic imaging classes (1C-1E) by all three methods provides high positive predictive value, but ultrasound ellipsoid is simpler to use and more readily available., (Copyright © 2022 by the American Society of Nephrology.)

A system concept for online alignment verification of millimeter-wave, corneal reflectometry is presented. The system utilizes beam scanning to generate magnitude-only reflectivity maps of the cornea at 650 GHz and compares these images to a precomputed/measured template map to confirm/reject sufficient alignment. A system utilizing 5 off-axis parabolic mirrors, a thin film beam splitter, and 2-axis galvanometric mirror was designed, simulated, and evaluated with geometric and physical optics. Simulation results informed the construction of a demonstrator system which was tested with a reference reflector. Similarity metrics computed with the aligned template and 26 misaligned positions, distributed on a 0.5 mm x 0.5 mm x 0.5 mm mesh, demonstrated sufficient misalignment detection sensitivity in 23 out of 26 positions. The results show that positional accuracy on the order of 0.5 mm is possible using 0.462 mm wavelength radiation due to the perturbation of coupling efficiency via beam distortion and beam walk-off.

Pterygium is an ocular surface disorder with high prevalence that can lead to vision impairment. As a pathological outgrowth of conjunctiva, pterygium involves neovascularization and chronic inflammation. Here, we developed a 3D multicellular in vitro pterygium model using a digital light processing (DLP)-based 3D bioprinting platform with human conjunctival stem cells (hCjSCs). A novel feeder-free culture system was adopted and efficiently expanded the primary hCjSCs with homogeneity, stemness and differentiation potency. The DLP-based 3D bioprinting method was able to fabricate hydrogel scaffolds that support the viability and biological integrity of the encapsulated hCjSCs. The bioprinted 3D pterygium model consisted of hCjSCs, immune cells, and vascular cells to recapitulate the disease microenvironment. Transcriptomic analysis using RNA sequencing (RNA-seq) identified a distinct profile correlated to inflammation response, angiogenesis, and epithelial mesenchymal transition in the bioprinted 3D pterygium model. In addition, the pterygium signatures and disease relevance of the bioprinted model were validated with the public RNA-seq data from patient-derived pterygium tissues. By integrating the stem cell technology with 3D bioprinting, this is the first reported 3D in vitro disease model for pterygium that can be utilized for future studies towards personalized medicine and drug screening., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Introduction: Anesthetic management of morbidly obese patients is challenging, particularly in those undergoing bariatric surgery. Dexmedetomidine is a α2-adrenergic receptor agonist that is increasingly used in the perioperative setting for its beneficial properties including sedation, anxiolysis, analgesia with opioid-sparing effects, and minimal impact on respiration. The objective of this study was to evaluate the effect of dexmedetomidine on postoperative analgesia and recovery-related outcomes among patients undergoing bariatric surgery., Evidence Acquisition: We conducted a systematic review and meta-analysis of MEDLINE, EMBASE, and CENTRAL databases from conception to September 2021 for randomized controlled trials (RCTs) using dexmedetomidine in bariatric patients on postoperative outcomes. Outcomes were pooled using random effects model and presented as relative risks (RR) or mean differences (MD) with 95% confidence intervals (CI)., Evidence Synthesis: In total, 20 RCTs with 665 patients in the dexmedetomidine group and 671 patients in the control groups were included. Among RCTs, the dexmedetomidine group had significantly lower opioid usage at 24-hours postoperatively (MD: -5.14, 95% CI: -10.18 to -0.10; moderate certainty), reduced pain scores on a 10-point scale at PACU arrival (MD: -1.69, 95% CI: -2.79 to -0.59; moderate certainty) and six hours postoperatively (MD: -1.82, 95% CI: -3.00 to -0.64; low certainty), and fewer instances of nausea (RR: 0.59, 95% CI: 0.45 to 0.75; moderate certainty) and vomiting (RR: 0.25, 95% CI: 0.15 to 0.43; moderate certainty), compared to control groups., Conclusions: Dexmedetomidine is an efficacious anesthesia adjunct in patients undergoing bariatric surgery. These benefits of dexmedetomidine may be considered in the multi-modal analgesic management and enhanced recovery pathways in this high-risk population.

Etodolac is an FDA-approved nonsteroidal anti-inflammatory drug (NSAID) used to treat a variety of inflammatory diseases. The drug is administered as a racemate (50/50 mixture of R- and S- enantiomers), however, studies have shown that the two enantiomers have distinct biologic and pharmacokinetic differences. Wnt signaling, which plays key roles in cell proliferation, polarity, and differentiation, has been shown to be inhibited by R-etodolac; however, comparative analyses of R- and S-etodolac in this function have not been conducted. We used in silico molecular docking and TOPflash functional biologic assays to compare R- and S-enantiomers effect on Wnt signaling inhibition. Further, we used a cultivated limbal stem epithelial cell (cLSCs) model to investigate enantiospecific changes in the colony-forming efficiency (CFE) of cLSCs. The data shows that R-etodolac is a more potent inhibitor of Wnt signaling. In addition, consistently, while both enantiomers demonstrate a dose-dependent decrease in CFE of cLSCs, R-etodolac is a more potent inhibitor., Competing Interests: The authors declare no conflict of interest., (© 2022 Published by Elsevier B.V.)

Volatile organic compounds (VOCs) are the main precursors of tropospheric O 3 and secondary organic aerosol (SOA), which can enhance atmospheric oxidation, promote the formation of secondary pollutants, and affect regional air quality and human health. In order to gain insights on VOCs characteristics and their potentials for O 3 and SOA formation, the volume fraction of 102 VOCs in autumn and winter in the urban area of Tongchuan were monitored using the TH-300B online monitoring system. The maximum incremental reactivity (MIR) coefficient and the fractional aerosol coefficient (FAC) were used to estimate the ozone formation potential (OFP) and SOA formation potential (SOAFP), respectively. The φ (TVOC, total VOCs) were (50.52±16.81)×10 -9 in autumn and (63.21±35.24)×10 -9 in winter. The OFPs were 138.43×10 -9 in autumn and 137.123×10 -9 in winter, and the SOAFPs were 3.098 μg·m -3 and 0.612 μg·m -3 , respectively. Alkanes (26.19%) and aromatics (26.04%) were the most abundant species in autumn, and alkanes (48.88%) were the most abundant species in winter. Trans-2-pentene, toluene, and m/p -xylene were the most reactive species in terms of OFPs in autumn, and ethylene, acetylene, and propylene were the top three species contributing to the total OFPs in winter. Toluene, m/p -xylene, and ethylbenzene contributed the most to the total SOAFPs in both of autumn and winter. Traffic emissions were considered as the major source of VOCs in both seasons. VOCs from biomass/coal combustion emissions showed seasonal differences, which were more prominent in winter. The results can provide references for the "one city, one policy" to mitigate regional VOCs pollution and improve ambient air quality.

Purpose: To evaluate in vivo parameters as biomarkers of limbal stem cell function and to establish an objective system that detects and stage limbal stem cell deficiency (LSCD)., Methods: A total of 126 patients (172 eyes) with LSCD and 67 normal subjects (99 eyes) were included in this observational cross-sectional comparative study. Slit-lamp biomicroscopy, in vivo laser scanning confocal microscopy (IVCM), and anterior segment optical coherence tomography (AS-OCT) were performed to obtain the following: clinical score, cell morphology score, basal cell density (BCD), central corneal epithelial thickness (CET), limbal epithelial thickness (LET), total corneal nerve fiber length (CNFL), corneal nerve fiber density (CNFD), corneal nerve branch density (CNBD), and tortuosity coefficient. Their potential correlations with the severity of LSCD were investigated, and cutoff values were determined., Results: An increase clinical score correlated with a decrease in central cornea BCD, limbal BCD, CET, mean LET, maximum LET, CNFL, CNFD, CNBD, and tortuosity coefficient. Regression analyses showed that central cornea BCD, CET and CNFL were the best parameters to differentiate LSCD from normal eyes (Coef = 3.123, 3.379, and 2.223; all p < 0.05). The rank correlation analysis showed a similar outcome between the clinical scores and the central cornea BCD (ρ = 0.79), CET (ρ = 0.82), and CNFL (ρ = 0.71). A comprehensive LSCD grading formula based on a combination of these parameters was established., Conclusions: A comprehensive staging system combining clinical presentation, central cornea BCD, CET, and CNFL is established to accurately and objectively diagnose LSCD and stage its severity., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Limbal epithelial stem/progenitor cells (LSCs) reside in a niche that contains finely tuned balances of various signaling pathways including Wnt, Notch, BMP, Shh, YAP, and TGFβ. The activation or inhibition of these pathways is frequently dependent on the interactions of LSCs with various niche cell types and extracellular substrates. In addition to receiving molecular signals from growth factors, cytokines, and other soluble molecules, LSCs also respond to their surrounding physical structure via mechanotransduction, interaction with the ECM, and interactions with other cell types. Damage to LSCs or their niche leads to limbal stem cell deficiency (LSCD). The field of LSCD treatment would greatly benefit from an understanding of the molecular regulation of LSCs in vitro and in vivo. This review synthesizes current literature around the niche factors and signaling pathways that influence LSC function. Future development of LSCD therapies should consider all these niche factors to achieve improved long-term restoration of the LSC population.

The corneal epithelium is continuously renewed by limbal stem/progenitor cells (LSCs), a cell population harbored in a highly regulated niche located at the limbus. Dysfunction and/or loss of LSCs and their niche cause limbal stem cell deficiency (LSCD), a disease that is marked by invasion of conjunctival epithelium into the cornea and results in failure of epithelial wound healing. Corneal opacity, pain, loss of vision, and blindness are the consequences of LSCD. Successful treatment of LSCD depends on accurate diagnosis and staging of the disease and requires restoration of functional LSCs and their niche. This review highlights the major advances in the identification of potential LSC biomarkers and components of the LSC niche, understanding of LSC regulation, methods and regulatory standards in bioengineering of LSCs, and diagnosis and staging of LSCD. Overall, this review presents key points for researchers and clinicians alike to consider in deepening the understanding of LSC biology and improving LSCD therapies., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

The corneal epithelium is consistently regenerated by limbal stem/progenitor cells (LSCs), a very small population of adult stem cells residing in the limbus. Several Wnt ligands, including Wnt6, are preferentially expressed in the limbus. To investigate the role of Wnt6 in regulating proliferation and maintenance of human LSCs in an in vitro LSC expansion setting, we generated NIH-3T3 feeder cells to overexpress different levels of Wnt6. Characterization of LSCs cultured on Wnt6 expressing 3T3 cells showed that high level of Wnt6 increased proliferation of LSCs. Medium and high levels of Wnt6 also increased the percentage of small cells (diameter ≤ 12 µm), a feature of the stem cell population. Additionally, the percentage of cells expressing the differentiation marker K12 was significantly reduced in the presence of medium and high Wnt6 levels. Although Wnt6 is mostly known as a canonical Wnt ligand, our data showed that canonical and non-canonical Wnt signaling pathways were activated in the Wnt6-supplemented LSC cultures, a finding suggesting that interrelationships between both pathways are required for LSC regulation., (© 2021. The Author(s).)