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Abstract Background Diabetic retinopathy (DR), the principal cause of acquired blindness among the working-age population, is the most frequent microvascular complication of diabetes. Although metabolic disorders are hypothesized to play a role in its pathogenesis, the underlying mechanism remains largely elusive. Methods To elucidate the mechanism, we initially compared metabolite profiles of vitreous fluid between 23 patients with DR and 12 non-diabetic controls using liquid chromatography/tandem mass spectrometry, identifying the distinct metabolite indoxyl sulfate (IS). Subsequently, streptozotocin (STZ)-induced diabetic and IS-injected rat models were established to examine the effects of IS on retinal microvasculature. RNA sequencing was conducted to identify potential regulatory mechanisms in IS-treated human retinal endothelial cells (HREC). Finally, target gene knockdown in HREC and treatment of IS-injected rats with inhibitors (targeting IS production or downstream regulators) were employed to elucidate the detailed mechanisms and identify therapeutic targets for DR. Results Metabolomics identified 172 significantly altered metabolites in the vitreous humor of diabetics, including the dysregulated tryptophan metabolite indoxyl sulfate (IS). IS was observed to breach the blood-retinal barrier and accumulate in the intraocular fluid of diabetic rats. Both in vivo and in vitro experiments indicated that elevated levels of IS induced endothelial apoptosis and disrupted cell junctions. RNA sequencing pinpointed prostaglandin E2 (PGE2) synthetase-cyclooxygenase 2 (COX-2) as a potential target of IS. Validation experiments demonstrated that IS enhanced COX-2 expression, which subsequently increased PGE2 secretion by promoting transcription factor EGR1 binding to COX-2 DNA following entry into cells via organic anion transporting polypeptides (OATP2B1). Furthermore, inhibition of COX-2 in vivo or silencing EGR1/OATP2B1 in HREC mitigated IS-induced microcapillary damage and the activation of COX-2/PGE2. Conclusion Our study demonstrated that indoxyl sulfate (IS), a uremic toxin originating from the gut microbiota product indole, increased significantly and contributed to retinal microvascular damage in diabetic retinopathy (DR). Mechanistically, IS impaired retinal microvascular integrity by inducing the expression of COX-2 and the production of PGE2. Consequently, targeting the gut microbiota or the PGE2 pathway may offer effective therapeutic strategies for the treatment of DR. Graphical Abstract

Abstract. Background:. Ischemic acute kidney injury (AKI) is a common syndrome associated with considerable mortality and healthcare costs. Up to now, the underlying pathogenesis of ischemic AKI remains incompletely understood, and specific strategies for early diagnosis and treatment of ischemic AKI are still lacking. Here, this study aimed to define the transcriptomic landscape of AKI patients through single-cell RNA sequencing (scRNA-seq) analysis in kidneys. Methods:. In this study, scRNA-seq technology was applied to kidneys from two ischemic AKI patients, and three human public scRNA-seq datasets were collected as controls. Differentially expressed genes (DEGs) and cell clusters of kidneys were determined. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis, as well as the ligand–receptor interaction between cells, were performed. We also validated several DEGs expression in kidneys from human ischemic AKI and ischemia/reperfusion (I/R) injury induced AKI mice through immunohistochemistry staining. Results:. 15 distinct cell clusters were determined in kidney from subjects of ischemic AKI and control. The injured proximal tubules (PT) displayed a proapoptotic and proinflammatory phenotype. PT cells of ischemic AKI had up-regulation of novel pro-apoptotic genes including USP47, RASSF4, EBAG9, IER3, SASH1, SEPTIN7, and NUB1, which have not been reported in ischemic AKI previously. Several hub genes were validated in kidneys from human AKI and renal I/R injury mice, respectively. Furthermore, PT highly expressed DEGs enriched in endoplasmic reticulum stress, autophagy, and retinoic acid-inducible gene I (RIG-I) signaling. DEGs overexpressed in other tubular cells were primarily enriched in nucleotide-binding and oligomerization domain (NOD)-like receptor signaling, estrogen signaling, interleukin (IL)-12 signaling, and IL-17 signaling. Overexpressed genes in kidney-resident immune cells including macrophages, natural killer T (NKT) cells, monocytes, and dendritic cells were associated with leukocyte activation, chemotaxis, cell adhesion, and complement activation. In addition, the ligand–receptor interactions analysis revealed prominent communications between macrophages and monocytes with other cells in the process of ischemic AKI. Conclusion:. Together, this study reveals distinct cell-specific transcriptomic atlas of kidney in ischemic AKI patients, altered signaling pathways, and potential cell–cell crosstalk in the development of AKI. These data reveal new insights into the pathogenesis and potential therapeutic strategies in ischemic AKI.

Abstract. Background:. Since 2019, a novel coronavirus named 2019 novel coronavirus (2019-nCoV) has emerged worldwide. Apart from fever and respiratory complications, acute kidney injury has been observed in a few patients with coronavirus disease 2019. Furthermore, according to recent findings, the virus has been detected in urine. Angiotensin-converting enzyme II (ACE2) has been proposed to serve as the receptor for the entry of 2019-nCoV, which is the same as that for the severe acute respiratory syndrome. This study aimed to investigate the possible cause of kidney damage and the potential route of 2019-nCoV infection in the urinary system. Methods:. We used both published kidney and bladder cell atlas data and new independent kidney single-cell RNA sequencing data generated in-house to evaluate ACE2 gene expression in all cell types in healthy kidneys and bladders. The Pearson correlation coefficients between ACE2 and all other genes were first generated. Then, genes with r values larger than 0.1 and P values smaller than 0.01 were deemed significant co-expression genes with ACE2. Results:. Our results showed the enriched expression of ACE2 in all subtypes of proximal tubule (PT) cells of the kidney. ACE2 expression was found in 5.12%, 5.80%, and 14.38% of the proximal convoluted tubule cells, PT cells, and proximal straight tubule cells, respectively, in three published kidney cell atlas datasets. In addition, ACE2 expression was also confirmed in 12.05%, 6.80%, and 10.20% of cells of the proximal convoluted tubule, PT, and proximal straight tubule, respectively, in our own two healthy kidney samples. For the analysis of public data from three bladder samples, ACE2 expression was low but detectable in bladder epithelial cells. Only 0.25% and 1.28% of intermediate cells and umbrella cells, respectively, had ACE2 expression. Conclusion:. This study has provided bioinformatics evidence of the potential route of 2019-nCoV infection in the urinary system.

Avian pathogenic Escherichia coli (APEC) strains belong to a category that is associated with colibacillosis, a serious illness in the poultry industry worldwide. Additionally, some APEC groups have recently been described as potential zoonotic agents. In this work, we compared APEC strains with extraintestinal pathogenic E. coli (ExPEC) strains isolated from clinical cases of humans with extra-intestinal diseases such as urinary tract infections (UTI) and bacteremia. PCR results showed that genes usually found in the ColV plasmid (tsh, iucA, iss, and hlyF) were associated with APEC strains while fyuA, irp-2, fepC sitDchrom, fimH, crl, csgA, afa, iha, sat, hlyA, hra, cnf1, kpsMTII, clpVSakai and malX were associated with human ExPEC. Both categories shared nine serogroups (O2, O6, O7, O8, O11, O19, O25, O73 and O153) and seven sequence types (ST10, ST88, ST93, ST117, ST131, ST155, ST359, ST648 and ST1011). Interestingly, ST95, which is associated with the zoonotic potential of APEC and is spread in avian E. coli of North America and Europe, was not detected among 76 APEC strains. When the strains were clustered based on the presence of virulence genes, most ExPEC strains (71.7%) were contained in one cluster while most APEC strains (63.2%) segregated to another. In general, the strains showed distinct genetic and fingerprint patterns, but avian and human strains of ST359, or ST23 clonal complex (CC), presented more than 70% of similarity by PFGE. The results demonstrate that some "zoonotic-related" STs (ST117, ST131, ST10CC, ST23CC) are present in Brazil. Also, the presence of moderate fingerprint similarities between ST359 E. coli of avian and human origin indicates that strains of this ST are candidates for having zoonotic potential.

BACKGROUND: The exacerbation of IgA nephropathy (IgAN) is related to respiratory tract infection with hemolytic streptococcus (HS), but the mechanism is unknown. In this study we investigated the role of chemokine ligand 20 (CCL20) in response to the effect of T helper 17 (Th17) cells in the pathogenesis of IgAN associated with HS. METHODS: Thirty mice were randomly divided into five groups: control mice (control), IgAN mice (IgAN), HS-infected IgAN mice (HS-IgAN), CCL20-treated IgAN mice (CCL20-IgAN), and CCL20-treated HS infected IgAN mice (CCL20-HS-IgAN). IgAN mice were induced with lipopolysaccharide, carbon tetrachloride and bovine serum albumin. Then the mice were sensitized with CCL20 antibody and infected with alpha-hemolytic streptococcus (α-HS) isolated from tonsils in sequence. Urine Albumin-Creatinine ratio and sediments were measured. The pathological changes in kidney and lung tissues were observed under microscopy. Th17 cells and regulatory T cells (Tregs) in kidneys were tested by flow cytometry. CCL20, IL-17A, IL-6 and IL-21 in the kidneys were detected by ELISA. RESULTS: The IgAN mice had albuminuria and microscopic hematuria, renal mesangial proliferation, IgA deposition, high electron dense deposition in glomerular mesangial region, decreased frequency of Tregs, increased frequency of Th17 and Th17-Treg ratio. Furthermore, Th17-related cytokines CCL20, IL-17A, IL-6 and IL-21 were all increased in the kidneys of IgAN mice. Compared with IgAN mice, the manifestations in HS-IgAN mice were more severe, but alleviated in CCL20-treated groups. CONCLUSION: α-HS may exacerbate kidney damage in IgAN through CCL20 response to the effect of Th17 cells.