Observation of structural and vascular features of retina and choroid in myopia using ultra-widefield SS-OCTA.

BACKGROUND: To find the relationship between the changes of retinal and choriodal structure/ vascular densities (VD) and the myopia progress. METHODS: 126 eyes of 126 age-matched young participants were divided into three groups: Emmetropia and Low Myopia (EaLM) (33 eyes), Moderate Myopia (MM) (39 eyes), and High Myopia (HM) (54 eyes). Fundus images measuring 12 × 12 mm were captured using ultra-widefield swept-source optical coherence tomography angiography (SS-OCTA). Each image was uniformly divided into nine regions: supra-temporal (ST), temporal (T), infra-temporal (IT), superior (S), central macular area (C), inferior (I), supra-nasal (SN), nasal (N), and infra-nasal (IN). Various structural parameters, including inner retina thickness (IRT), outer retina thickness (ORT), and choroid thickness (CT), were assessed, and the VD of the superficial capillary plexus (SCP), deep capillary plexus (DCP), choriocapillaries (CC), and choroid vessels (ChdV) were quantified. RESULTS: CT in upper fundus exhibited a significant reduction from EaLM to MM. Additionally, ORT (ST, S. SN, C, N, IT, I, IN), CT (ST, S, SN, T, C, N, IT, I, IN) and VDs of SCP (ST, S, C, I, IN), DCP (ST, S, T, C, I) and ChdV (T, N, I, IN) were statistically diminished in EaLM compared to HM. Furthermore, IRT (N), ORT (N, IN), CT (S, SN, T, C, IT, I) and VDs of SCP (I, IN) and DCP (I) exhibited significant decreases as MM progressed towards HM. Intriguingly, there was a notable increase in the VD of CC (ST, S, T, C, N) as myopia progressed from MM to HM. CONCLUSION: Significant changes in retinal and choroid structure and vascular density occur as moderate myopia advances to high myopia. Efforts to curb myopia progression to this stage are essential, as the failure to do so may lead to the development of corresponding retinopathy.

Intraocular lens power calculation for silicone oil-dependent eyes.

Background: Silicone oil tamponade is widely used in vitreoretinal surgery. In some cases, silicone oil may not be extracted for a long time or even permanently and is referred to as silicone oil-dependent eyes. In this study, we aimed to deduce a theoretical formula for calculating intraocular lens power for silicone oil-dependent eyes and compare it with clinical findings. Methods: A theoretical formula was deduced using strict geometric optical principles and the Gullstrand simplified eye model. The preoperative and postoperative refractive statuses of patients with silicone oil-dependent eyes who underwent intraocular lens implantation were studied (Group A, n = 13). To further test our derived theoretical formula, patients with silicone oil tamponade and first-stage intraocular lens implantation were included (Group B, n = 19). In total, 32 patients (32 eyes) were included in the study. Results: In group A, the calculated intraocular lens power based on our formula was 24.96 ± 3.29 diopters (D), and the actual refraction of the patients was 24.02 ± 4.14D. In group B, the theoretical intraocular lens power was 23.10 ± 3.08D, and the clinical intraocular lens power was 22.84 ± 3.42D. There was no significant difference between the theoretical and clinical refractive powers, and the intraclass correlation coefficient was 0.771 for group A and 0.811 for group B (both p ≤ 0.001). The mean absolute error for silicone oil-dependent eyes of the formula was 1.66 ± 2.09D. After excluding data for two patients with a flat cornea (corneal refractive power < 42D), the mean absolute error decreased to 0.83 ± 0.62D. Conclusion: A strong correlation between the theoretical and clinical intraocular lens powers was observed, and the formula we deduced can be used to calculate the intraocular lens power for silicone oil-dependent eyes. This formula will help clinicians select a more appropriate intraocular lens for patients with silicone oil-dependent eyes, especially when the corneal refractive power is ≥42D.

Analysis of retinal and choroidal characteristics in patients with early diabetic retinopathy using WSS-OCTA.

Introduction: Diabetic retinopathy (DR) is one of the most common and destructive microvascular complications of DM, and has become a major cause of irreversible visual impairment. The purpose of this study was to evaluate the changes in fundus microcirculation in non-diabetic retinopathy (NDR) and mild non-proliferative diabetic retinopathy (NPDR) in patients with type 2 diabetic mellitus (T2DM) using widefield swept-source optical coherence tomography angiography (WSSOCTA), and to investigate the correlation with laboratory indices of T2DM. Methods: Eighty nine, 58 and 28 eyes were included in the NDR, NPDR and Control groups, respectively, were enrolled in this study. The 12mm×12mm fundus images obtained by WSS-OCTA were divided into 9 regions (supratemporal, ST; temporal, T; inferotemporal, IT; superior, S; central macular area, C; inferior, I; supranasal, SN; nasal, N; inferonasal, IN) to evaluate changes in vessel density (VD) of the superficial capillary plexus (SCP), deep capillary plexus (DCP), choriocapillaris, and mid-large choroidal vessel (MLCV), as well as changes in inner retinal thickness (IRT), outer retinal thickness (ORT), and choroidal thickness (CT). Results: Compared with control group, MLCV VD (I, N, IN) was significantly decreased in NDR group, SCP VD (IT, C, I) and DCP VD (T, IT, I) were significantly decreased in NPDR group. In NPDR group, DCP VD (IT) was significantly decreased compared with that in NDR group. Compared with control group, CT (ST, T, IT, S, SN, IN) was significantly declined in NDR group, and IRT (ST, IT) and ORT (ST, N) were significantly increased in NPDR group. In NPDR group, IRT (ST) and ORT (T, S) were significantly increased compared with NDR group. Correlation analysis showed that age, body mass index, fasting blood glucose, fasting insulin, fasting C-peptide, and estimated glomerular filtration rate in T2DM patients were statistically correlated with retinal and choroidal thickness/VD. Discussion: Structural and blood flow changes in the choroid occur before the onset of DR and precede changes in the retinal microcirculation, and MLCV thickness/VD is a more sensitive imaging biomarker for the clinical detection of DR. WSS-OCTA enables large-scale non-invasive visual screening and follow-up of the retinal and choroidal vasculature in DR patients, providing a new strategy for the prevention and monitoring of DR in patients with T2DM.

Peptidase Activation by a Leader Peptide-Bound RiPP Recognition Element.

The RiPP precursor recognition element (RRE) is a conserved domain found in many prokaryotic ribosomally synthesized and post-translationally modified peptide (RiPP) biosynthetic gene clusters (BGCs). RREs bind with high specificity and affinity to a recognition sequence within the N-terminal leader region of RiPP precursor peptides. Lasso peptide biosynthesis involves an RRE-dependent leader peptidase, which is discretely encoded or fused to the RRE as a di-domain protein. Here we leveraged thousands of predicted BGCs to define the RRE:leader peptidase interaction through evolutionary covariance analysis. Each interacting domain contributes a three-stranded ß-sheet to form a hydrophobic ß-sandwich-like interface. The bioinformatics-guided predictions were experimentally confirmed using proteins from discrete and fused lasso peptide BGC architectures. Support for the domain-domain interface derived from chemical shift perturbation, paramagnetic relaxation enhancement experiments, and rapid variant activity screening using cell-free biosynthesis. Further validation of selected variants was performed with purified proteins. We developed a p-nitroanilide-based leader peptidase assay to illuminate the role of RRE domains. Our data show that RRE domains play a dual function. RRE domains deliver the precursor peptide to the leader peptidase, and the rate is saturable as expected for a substrate. RRE domains also partially compose the elusive S2 proteolytic pocket that binds the penultimate threonine of lasso leader peptides. Because the RRE domain is required to form the active site, leader peptidase activity is greatly diminished when the RRE domain is supplied at substoichiometric levels. Full proteolytic activation requires RRE engagement with the recognition sequence-containing portion of the leader peptide. Together, our observations define a new mechanism for protease activity regulation.

Heterobivalent Inhibitors of Acetyl-CoA Carboxylase: Drug Target Residence Time and Time-Dependent Antibacterial Activity.

The relationship between drug-target residence time and the post-antibiotic effect (PAE) provides insights into target vulnerability. To probe the vulnerability of bacterial acetyl-CoA carboxylase (ACC), a series of heterobivalent inhibitors were synthesized based on pyridopyrimidine 1 and moiramide B (3) which bind to the biotin carboxylase and carboxyltransferase ACC active sites, respectively. The heterobivalent compound 17, which has a linker of 50 Å, was a tight binding inhibitor of Escherichia coli ACC (Kiapp 0.2 nM) and could be displaced from ACC by a combination of both 1 and 3 but not just by 1. In agreement with the prolonged occupancy of ACC resulting from forced proximity binding, the heterobivalent inhibitors produced a PAE in E. coli of 1-4 h in contrast to 1 and 3 in combination or alone, indicating that ACC is a vulnerable target and highlighting the utility of kinetic, time-dependent effects in the drug mechanism of action.

Long non-coding RNA NCK1-AS1 is overexpressed in esophageal squamous cell carcinoma and predicts survival.

Long noncoding RNAs have shown pivotal regulatory roles in tumorigenesis and progression. NCK1-AS1 promotes cervical cancer, while its involvement in esophageal cancer is hardly known. This study enrolled 52 esophageal squamous cell carcinoma (ESCC) patients (30 males and 22 females) at the average age of 56.4 ± 6.6 years in the range from 46 to 70 years, explored the involvement of NCK1-AS1 in ESCC, and analyzed the possible interaction between NCK1-AS1 and TGF-ß signaling. Changes in gene expression were analyzed using RT-qPCR and Western blot. Interactions between gene expressions were analyzed using ESCC cells with transient transfections. Cell invasion and migration were analyzed using Transwell assays. Our data showed that plasma NCK1-AS1 was overexpressed in ESCC patients and positively correlated with NCK1-AS1 expression in tumor tissues but not in non-tumor tissues. Moreover, high plasma NCK1-AS1 levels were accompanied with poor survival. TGF-ß1 expression level was also increased in tumor tissues compared to the adjacent normal tissues and positively correlated with NCK1-AS1 in tumor tissues. TGF-ß1 overexpression in ESCC cells did not affect NCK1-AS1 expression, while NCK1-AS1 overexpression in ESCC cells upregulated TGF-ß1. Moreover, TGF-ß1 and NCK1-AS1 overexpression increased ESCC cell migration and invasion, while TGF-ß inhibitor reduced the effects of NCK1-AS1 overexpression. Overall, NCK1-AS1 may promote ESCC by upregulating TGF-ß1.

Occurrence and Ecological Risk Assessment of Perfluoroalkyl Substances (PFASs) in Water and Sediment from an Urban River in South China.

The chemical substances in urban rivers influence municipal water systems and reflect the recent use of these chemicals by humans or industries around the urban center. In this study, seven perfluoroalkyl substances (PFASs)-perfluorohexanoic acid (PFHxA), perfluoroheptanoic acid (PFHpA), perfluorooctanoic acid (PFOA), perfluorononanoic acid (PFNA), 2-perfluorohexyl ethanol (6:2 FTOH), 2-perfluorooctyl ethanol (8:2 FTOH), and 6:2 chlorinated polyfluoroalkyl ether sulfonic acids (F-53B)-could be detected and quantified in river water and sediment samples collected from one tributary of the Liuxi River, which is part of Pearl River near Guangzhou in Guangdong province, South China. The fluxes of target PFASs into Liuxi River and their related ecological risks were further estimated. The total concentrations of PFASs (ΣPFASs) ranged from 506 to 3.16 × 103 ng/L in water samples and 9.13 to 850 ng/L in sediment samples. The two dominant PFAS compounds were 6:2 FTOH and PFHpA, which accounted for more than 90.0% of ΣPFASs in river water and sediment. Correlation analysis showed that there was significant positive correlation (p < 0.01) between two selected PFASs (e.g., between 6:2 FTOH and PFHpA). Correlation analysis of PFASs in river water and sediment indicated most PFASs in sediment were partitioned from river water. The ecological risk assessment indicated that the detected PFASs have a low risk (HQ < 0.1) in river water and sediment to Daphnia magna in the Liuxi River.

Cell-Free Biosynthesis to Evaluate Lasso Peptide Formation and Enzyme-Substrate Tolerance.

Lasso peptides are ribosomally synthesized and post-translationally modified peptide (RiPP) natural products that display a unique lariat-like, threaded conformation. Owing to a locked three-dimensional structure, lasso peptides can be unusually stable toward heat and proteolytic degradation. Some lasso peptides have been shown to bind human cell-surface receptors and exhibit anticancer properties, while others display antibacterial or antiviral activities. All known lasso peptides are produced by bacteria and genome-mining studies indicate that lasso peptides are a relatively prevalent class of RiPPs; however, the discovery, isolation, and characterization of lasso peptides are constrained by the lack of an efficient production system. In this study, we employ a cell-free biosynthesis (CFB) strategy to address longstanding challenges associated with lasso peptide production. We report the successful use of CFB for the formation of an array of sequence-diverse lasso peptides that include known examples as well as a new predicted lasso peptide from Thermobifida halotolerans. We further demonstrate the utility of CFB to rapidly generate and characterize multisite precursor peptide variants to evaluate the substrate tolerance of the biosynthetic pathway. By evaluating more than 1000 randomly chosen variants, we show that the lasso-forming cyclase from the fusilassin pathway is capable of producing millions of sequence-diverse lasso peptides via CFB. These data lay a firm foundation for the creation of large lasso peptide libraries using CFB to identify new variants with unique properties.

Functional elucidation of TfuA in peptide backbone thioamidation.

YcaO enzymes catalyze several post-translational modifications on peptide substrates, including thioamidation, which substitutes an amide oxygen with sulfur. Most predicted thioamide-forming YcaO enzymes are encoded adjacent to TfuA, which when present, is required for thioamidation. While activation of the peptide amide backbone is well established for YcaO enzymes, the function of TfuA has remained enigmatic. Here we characterize the TfuA protein involved in methyl-coenzyme M reductase thioamidation and demonstrate that TfuA catalyzes the hydrolysis of thiocarboxylated ThiS (ThiS-COSH), a proteinaceous sulfur donor, and enhances the affinity of YcaO toward the thioamidation substrate. We also report a crystal structure of a TfuA, which displays a new protein fold. Our structural and mutational analyses of TfuA have uncovered conserved binding interfaces with YcaO and ThiS in addition to revealing a hydrolase-like active site featuring a Ser-Lys catalytic pair.

Positron Emission Tomography Imaging of Staphylococcus aureus Infection Using a Nitro-Prodrug Analogue of 2-[18F]F-p-Aminobenzoic Acid.

Deep-seated bacterial infections caused by pathogens such as Staphylococcus aureus are difficult to diagnose and treat and are thus a major threat to human health. In previous work we demonstrated that positron emission tomography (PET) imaging with 2-[18F]F-p-aminobenzoic acid (2-[18F]F-PABA) could noninvasively identify, localize, and monitor S. aureus infection with excellent sensitivity and specificity in a rodent soft tissue infection model. However, 2-[18F]F-PABA is rapidly N-acetylated and eliminated, and in an attempt to improve radiotracer accumulation in bacteria we adopted a prodrug strategy in which the acid was protected by an ester and the amine was replaced with a nitro group. Metabolite analysis indicated that the nitro group of ethyl 2-[18F]fluoro-4-nitrobenzoate (2-[18F]F-ENB) is converted to the corresponding amine by bacteria-specific nitroreductases while the ester is hydrolyzed in vivo into the acid. PET/CT imaging of 2-[18F]F-ENB and the corresponding acid 2-[18F]F-NB in a rat soft tissue infection model demonstrated colocalization of the radiotracer with the bioluminescent signal arising from S. aureus Xen29, and demonstrated that the tracer could differentiate S. aureus infection from sterile inflammation. Significantly, the accumulation of both 2-[18F]F-ENB and 2-[18F]F-NB at the site of infection was 17-fold higher than at the site of sterile inflammation compared to 8-fold difference observed for 2-[18F]F-PABA, supporting the proposal that the active radiotracer in vivo is 2-[18F]F-NB. Collectively, these data suggest that 2-[18F]F-ENB and 2-[18F]F-NB have the potential for translation to humans as a rapid, noninvasive diagnostic tool to identify and localize S. aureus infections.

First genetic assessment of brackish water polychaete Tylorrhynchus heterochaetus: mitochondrial COI sequences reveal strong genetic differentiation and population expansion in samples collected from southeast China and north Vietnam.

Tylorrhynchus heterochaetus is a widespread benthic polychaete worm found in coastal brackish waters of the west Pacific. It has high ecological and economic value as a biomarker of water quality and as a high-quality feed in aquaculture and fisheries and is considered a delicacy in some areas of Asia. However, it has experienced a marked reduction in recent years due to overexploitation as well as changes in the environment and climate. Here, to comprehensively understand its genetic background and thus provide insights for better conservation and utilization of this species, we assessed the genetic variability and demographic history of T. heterochaetus individuals sampled from eight locations along the coasts of southeast China and north Vietnam based on mitochondrial cytochrome c oxidase I ( COI) sequences. We observed high haplotype diversity ( Hd), with an average of 0.926, but relatively low nucleotide diversity ( π), with a mean of 0.032 across all samples. A total of 94 polymorphic sites and 85 haplotypes were identified among 320 individuals. The pairwise genetic distances among haplotypes ranged from 0.001 to 0.067, with the high intraspecific divergence possibly reflecting geographic isolation and gene pool fragmentation. Significant genetic structures were revealed among the studied locations; specifically, the eight locations could be treated as six genetically different populations based on pairwise Φ ST results (0.026-0.951, P<0.01). A significant pattern of isolation-by-distance was detected between the genetic and geographic distances ( r=0.873, P=0.001). Three geographic lineages were defined based on phylogenetic tree and network analyses of COI haplotypes. AMOVA results indicated that genetic variations mainly occurred among the three lineages (89.96%). Tests of neutrality and mismatch distribution suggested that T. heterochaetus underwent recent population expansion. These results provide the first report on the genetic status of T. heterochaetus and will be valuable for the management of genetic resources and better understanding of the ecology and evolution in this species.

Antibacterial Activity and Mode of Action of a Sulfonamide-Based Class of Oxaborole Leucyl-tRNA-Synthetase Inhibitors.

Benzoxaboroles are a class of boron-containing compounds with a broad range of biological activities. A subset of benzoxaboroles have antimicrobial activity due primarily to their ability to inhibit leucyl-tRNA synthetase (LeuRS) via the oxaborole tRNA-trapping mechanism, which involves the formation of a stable tRNALeu-benzoxaborole adduct in which the boron atom interacts with the 2'- and 3'-oxygen atoms of the terminal 3' tRNA adenosine. We sought to identify other antibacterial targets for this promising class of compounds by means of mode-of-action studies, and we selected a nitrophenyl sulfonamide based oxaborole (PT638) as a probe molecule because it had potent antibacterial activity (MIC of 0.4 µg/mL against methicillin-resistant Staphylococcus aureus) but did not inhibit LeuRS (IC50 > 100 µM). Analogues of PT638 were synthesized to explore the importance of the sulfonamide linker and the impact of altering the functionalization of the phenyl ring. These structure-activity-relationship studies revealed that the nitro substituent was essential for activity. To identify the target for PT638, we raised resistant strains of S. aureus, and whole-genome sequencing revealed mutations in leuRS, suggesting that the target for this compound was indeed LeuRS, despite the lack of enzyme inhibition. Subsequent analysis of PT638 metabolism demonstrated that bacterial nitroreductases readily converted this compound into the amino analogue, which inhibited LeuRS with an IC50 of 3.0 ± 1.2 µM, demonstrating that PT638 is thus a prodrug.

Complete genome sequence of Paracoccus denitrificans ATCC 19367 and its denitrification characteristics.

Studies show that Paracoccus denitrificans can denitrify nitrogen sources under aerobic conditions. However, the lack of data on its genome sequence has restricted molecular studies and practical applications. In this study, the complete genome of P. denitrificans ATCC 19367 was sequenced and its nitrogen metabolism properties were characterized. The size of the whole genome is 5 242 327 bp, with two chromosomes and one plasmid. The average G + C content is 66.8%, and it contains 5308 protein-coding genes, 54 tRNA genes, and nine rRNA operons. Among the protein-coding genes, 71.35% could be assigned to the Gene Ontology (GO) pathway, 86.66% to the Clusters of Orthologous Groups (COG) pathway, and 50.57% to the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway. Comparative genome analysis between P. denitrificans ATCC 19367 and P. denitrificans PD1222 revealed that there are 428 genes specific to ATCC 19367 and 4738 core genes. Furthermore, the expression of genes related to denitrification, biofilm formation, and nitrogen metabolism (nar, nir, and nor) by P. denitrificans ATCC 19367 under aerobic conditions was affected by incubation time and shaking speed. This study elucidates the genomic background of P. denitrificans ATCC 19367 and suggests the possibility of controlling nitrogen pollution in the environment by using this bacterium.

Structure-Based Design, Synthesis, and Biological Evaluation of Non-Acyl Sulfamate Inhibitors of the Adenylate-Forming Enzyme MenE.

N-Acyl sulfamoyladenosines (acyl-AMS) have been used extensively to inhibit adenylate-forming enzymes that are involved in a wide range of biological processes. These acyl-AMS inhibitors are nonhydrolyzable mimics of the cognate acyl adenylate intermediates that are bound tightly by adenylate-forming enzymes. However, the anionic acyl sulfamate moiety presents a pharmacological liability that may be detrimental to cell permeability and pharmacokinetic profiles. We have previously developed the acyl sulfamate OSB-AMS (1) as a potent inhibitor of the adenylate-forming enzyme MenE, an o-succinylbenzoate-CoA (OSB-CoA) synthetase that is required for bacterial menaquinone biosynthesis. Herein, we report the use of computational docking to develop novel, non-acyl sulfamate inhibitors of MenE. A m-phenyl ether-linked analogue (5) was found to be the most potent inhibitor (IC50 = 8 µM; Kd = 244 nM), and its X-ray co-crystal structure was determined to characterize its binding mode in comparison to the computational prediction. This work provides a framework for the development of potent non-acyl sulfamate inhibitors of other adenylate-forming enzymes in the future.