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1.
Chem Commun (Camb) ; 60(14): 1948-1951, 2024 Feb 13.
Article in English | MEDLINE | ID: mdl-38284146

ABSTRACT

Nucleic acids in blood are early indicators of disease that could be detected by point-of-care biosensors if sufficiently sensitive and facile sensors existed. Electrochemical hybridization assays are sensitive and specific but are limited to very short nucleic acids. We have developed a restriction enzyme-assisted electrochemical hybridization (REH) assay for improved nucleic acid detection. By incorporating target-specific restriction enzymes, we detect long nucleic acids, with performance dependent on the location of the cut site relative to the electrode surface. Thus, we have further established guidelines for REH design to serve as a generalizable platform for robust electrochemical detection of long nucleic acids.


Subject(s)
Biosensing Techniques , Nucleic Acids , Electrochemical Techniques , Nucleic Acid Hybridization , Electrodes
2.
Toxics ; 11(11)2023 Nov 13.
Article in English | MEDLINE | ID: mdl-37999580

ABSTRACT

Arsenic pollution is a global environmental concern. Arsenic-induced chronic liver injury and its irreversible outcomes, including liver cirrhosis and liver cancer, threaten the health of residents in arsenic-contaminated areas. Liver fibrosis is a reversible pathological stage in the progression of arsenic-induced chronic liver injury to cirrhosis and liver cancer. The aim of this study is to identify the epigenetic mechanism of arsenic-induced liver fibrosis based on the dedifferentiation of liver sinusoidal endothelial cells (LSECs). Rats were treated with 0.0, 2.5, 5.0, or 10.0 mg/kg sodium arsenite for 36 weeks. Marked fibrotic phenotypes were observed in the rat livers, manifested by hepatic stellate cell activation and an increased extracellular matrix, as well as the deposition of collagen fibers. The reduced fenestrations on the cells' surface and the increased expression of the dedifferentiation marker CD31 corroborated the LSECs' dedifferentiation in the liver tissue, which was also found to be significantly associated with fibrotic phenotypes. We further revealed that arsenic exposure could inhibit the enrichment of histone H3 lysine 18 acetylation (H3K18ac) in the promoters of Fcgr2b and Lyve1, two key genes responsible for maintaining the differentiation phenotype of LSECs. This inhibition subsequently suppressed the genes' expression, promoting LSEC dedifferentiation and subsequent liver fibrosis. In conclusion, arsenic can trigger liver fibrosis by inhibiting H3K18ac-dependent maintenance of LSEC differentiation. These findings uncover a novel mechanism of arsenic-induced liver fibrosis based on a new insight into epigenetically dependent LSEC dedifferentiation.

3.
Nat Commun ; 14(1): 4839, 2023 Aug 10.
Article in English | MEDLINE | ID: mdl-37563116

ABSTRACT

Persistent room temperature phosphorescent materials with unique mechanical properties and robust optical properties have great potential in flexible electronics and photonics. However, developing such materials remains a formidable challenge. Here, we present highly stretchable, lightweight, and multicolored persistent luminescence elastomers, produced by incorporating ionic room temperature phosphorescent polymers and polyvinyl alcohol into a polydimethylsiloxane matrix. These prepared elastomers exhibit high optical transparency in daylight and emit bright persistent luminescence after the removal of 365 nm excitation. The homogeneous distribution of polymers within the matrix has been confirmed by confocal fluorescence microscopy, scanning electron microscopy, and atomic force microscopy. Mechanical property investigations revealed that the prepared persistent luminescence elastomers possess satisfactory stretchability. Impressively, these elastomers maintain robust optical properties even under extensive and repeated mechanical deformations, a characteristic previously unprecedented. These fantastic features make these persistent luminescence elastomers ideal candidates for potential applications in wearable devices, flexible displays, and anti-counterfeiting.

4.
FASEB J ; 37(3): e22794, 2023 03.
Article in English | MEDLINE | ID: mdl-36753399

ABSTRACT

Diabetic kidney disease (DKD) is one of the most serious complications of diabetes mellitus (DM) and the main cause of end-stage renal failure. However, the pathogenesis of DKD is complicated. In this study, we found that miR-124-3p plays a key role in regulating renal mitochondrial function and explored its possible mechanism in DKD progression by performing a series of in vitro and in vivo experiments. Decreased expression of miR-124-3p was found in db/db mice compared to db/m mice. Moreover, miR-124-3p down-regulated FOXQ1 by targeting FOXQ1 mRNA 3'-UTR in NRK-52E cells. Also, an increase in FOXQ1 and down-regulation of Sirt4 were found in db/db mouse kidney and renal tubular epithelial cells cultured with high glucose and high lipid. Overexpression of FOXQ1 could further down-regulate the expression of Sirt4 and aggravate the damage of mitochondria. Conversely, the knockdown of the FOXQ1 gene induced Sirt4 expression and partially restored mitochondrial function. To verify the effects of miR-124-3p on Sirt4 and mitochondria, we found that miR-124-3p mimics could up-regulate Sirt4 and inhibit ROS production and MitoSOX, thus restoring the number and morphology of mitochondria. These results showed that under high-glucose and high-lipid conditions, the down-regulation of miR-124-3p induces FOXQ1 in renal tubular epithelial cells, which in turn suppresses Sirt4 and leads to mitochondrial dysfunction, promoting the development of DKD.


Subject(s)
Diabetic Nephropathies , MicroRNAs , Mice , Animals , MicroRNAs/metabolism , Epithelial Cells/metabolism , Diabetic Nephropathies/metabolism , Mice, Inbred Strains , Glucose/metabolism , Mitochondria/metabolism , Lipids/pharmacology
5.
Bioconjug Chem ; 34(2): 358-365, 2023 02 15.
Article in English | MEDLINE | ID: mdl-36633230

ABSTRACT

Modification of electrodes with biomolecules is an essential first step for the development of bioelectrochemical systems, which are used in a variety of applications ranging from sensors to fuel cells. Gold is often used because of its ease of modification with thiolated biomolecules, but carbon screen-printed electrodes (SPEs) are gaining popularity due to their low cost and fabrication from abundant resources. However, their effective modification with biomolecules remains a challenge; the majority of work to-date relies on nonspecific adhesion or broad amide bond formation to chemical handles on the electrode surface. By combining facile electrochemical modification to add an aniline handle to electrodes with a specific and biocompatible oxidative coupling reaction, we can readily modify carbon electrodes with a variety of biomolecules. Importantly, both proteins and DNA maintain bioactive conformations following coupling. We have then used biomolecule-modified electrodes to generate microbial monolayers through DNA-directed immobilization. This work provides an easy, general strategy to modify inexpensive carbon electrodes, significantly expanding their potential as bioelectrochemical systems.


Subject(s)
Biosensing Techniques , Carbon , Carbon/chemistry , Proteins , DNA , Electrodes , Gold/chemistry , Electrochemical Techniques
6.
Acta Pharmacol Sin ; 44(5): 1051-1065, 2023 May.
Article in English | MEDLINE | ID: mdl-36347997

ABSTRACT

Previous studies have shown mitochondrial dysfunction in various acute kidney injuries and chronic kidney diseases. Lipoic acid exerts potent effects on oxidant stress and modulation of mitochondrial function in damaged organ. In this study we investigated whether alpha lipoamide (ALM), a derivative of lipoic acid, exerted a renal protective effect in a type 2 diabetes mellitus mouse model. 9-week-old db/db mice were treated with ALM (50 mg·kg-1·d-1, i.g) for 8 weeks. We showed that ALM administration did not affect blood glucose levels in db/db mice, but restored renal function and significantly improved fibrosis of kidneys. We demonstrated that ALM administration significantly ameliorated mitochondrial dysfunction and tubulointerstitial fibrotic lesions, along with increased expression of CDX2 and CFTR and decreased expression of ß-catenin and Snail in kidneys of db/db mice. Similar protective effects were observed in rat renal tubular epithelial cell line NRK-52E cultured in high-glucose medium following treatment with ALM (200 µM). The protective mechanisms of ALM in diabetic kidney disease (DKD) were further explored: Autodock Vina software predicted that ALM could activate RXRα protein by forming stable hydrogen bonds. PROMO Database predicted that RXRα could bind the promoter sequences of CDX2 gene. Knockdown of RXRα expression in NRK-52E cells under normal glucose condition suppressed CDX2 expression and promoted phenotypic changes in renal tubular epithelial cells. However, RXRα overexpression increased CDX2 expression which in turn inhibited high glucose-mediated renal tubular epithelial cell injury. Therefore, we reveal the protective effect of ALM on DKD and its possible potential targets: ALM ameliorates mitochondrial dysfunction and regulates the CDX2/CFTR/ß-catenin signaling axis through upregulation and activation of RXRα. Schematic figure illustrating that ALM alleviates diabetic kidney disease by improving mitochondrial function and upregulation and activation of RXRα, which in turn upregulated CDX2 to exert an inhibitory effect on ß-catenin activation and nuclear translocation. RTEC renal tubular epithelial cell. ROS Reactive oxygen species. RXRα Retinoid X receptor-α. Mfn1 Mitofusin 1. Drp1 dynamic-related protein 1. MDA malondialdehyde. 4-HNE 4-hydroxynonenal. T-SOD Total-superoxide dismutase. CDX2 Caudal-type homeobox transcription factor 2. CFTR Cystic fibrosis transmembrane conductance regulator. EMT epithelial mesenchymal transition. α-SMA Alpha-smooth muscle actin. ECM extracellular matrix. DKD diabetic kidney disease. Schematic figure was drawn by Figdraw ( www.figdraw.com ).


Subject(s)
Diabetes Mellitus, Type 2 , Diabetic Nephropathies , Thioctic Acid , Animals , Mice , Rats , beta Catenin/metabolism , Cystic Fibrosis Transmembrane Conductance Regulator/metabolism , Diabetes Mellitus, Type 2/complications , Diabetes Mellitus, Type 2/drug therapy , Diabetes Mellitus, Type 2/metabolism , Diabetic Nephropathies/drug therapy , Diabetic Nephropathies/pathology , Epithelial-Mesenchymal Transition , Fibrosis/drug therapy , Fibrosis/metabolism , Glucose/metabolism , Kidney/pathology , Mitochondria/drug effects , Mitochondria/metabolism , Thioctic Acid/pharmacology , Thioctic Acid/therapeutic use , Retinoid X Receptor alpha/drug effects , Retinoid X Receptor alpha/metabolism
7.
Water Res ; 224: 119045, 2022 Oct 01.
Article in English | MEDLINE | ID: mdl-36108396

ABSTRACT

Microplastics are widely present in global ecosystems, threatening both marine and freshwater species. Given this problem, it is vital to research where land-based microplastics originate and how they are transmitted to receiving waters in urban agglomerations. Research results should inform systemic mitigation efforts to prevent future contamination. This study established the multi-directional transmission network of a microplastic mass balance system using a source-pathway-receptor framework, and involving annual source stocks and pathway flows with considerable variations under dry and wet weather patterns. Details of a baseline scenario quantifying the occurrence and spread of microplastics in an urban agglomeration were also determined in the context of current environmental management practices. We demonstrated that the total stock of the six major pollution sources amounted to 5317.7 ± 2175.3 and 3320.1 ± 953.6 tons/a in dry and wet weather, respectively; and 2347.8 ± 766.9 and 1991.8 ± 701.8 tons/a flows directly entered the sewer system and receiving water in Shanghai, China, respectively. Prominent microplastic stocks were found in atmospheric fallout, industrial wastewater, and domestic sewage. These stocks were much higher compared to crop farming wastewater, aquacultural wastewater, and livestock and poultry breeding wastewater. Total microplastic flows entering receiving water reached 3207.4 ± 1071.6 tons/a; the largest contributions were from wet weather overflow (23.7%), direct atmospheric fallout (21.7%), wastewater treatment plant effluent (14.2%), industrial wastewater (14.1%), and surface runoff (10.4%). Weather patterns led to divergent microplastic transmission pathways and mass flows, revealing a lagging timeline mode and illustrating the basic spatiotemporal features of microplastic contamination in urban agglomerations. Terminal disposal practices retained about two-fifths of the microplastic flows that would have otherwise been transmitted into receiving water. Of these, land surface sweep contributed half of the retained flow. Improvements in WWTP removal efficiency, storm sewage interception rate, industrial wastewater collection rate, and sewer sediment dredge rate could further enhance the systemic benefits.


Subject(s)
Microplastics , Water Pollutants, Chemical , China , Conservation of Natural Resources , Ecosystem , Environmental Monitoring , Plastics , Sewage/analysis , Wastewater , Water , Water Pollutants, Chemical/analysis , Weather
8.
Front Chem ; 10: 911678, 2022.
Article in English | MEDLINE | ID: mdl-35769443

ABSTRACT

The field of infectious disease diagnostics is burdened by inequality in access to healthcare resources. In particular, "point-of-care" (POC) diagnostics that can be utilized in non-laboratory, sub-optimal environments are appealing for disease control with limited resources. Electrochemical biosensors, which combine biorecognition elements with electrochemical readout to enable sensitive and specific sensing using inexpensive, simple equipment, are a major area of research for the development of POC diagnostics. To improve the limit of detection (LOD) and selectivity, signal amplification strategies have been applied towards these sensors. In this perspective, we review recent advances in electrochemical biosensor signal amplification strategies for infectious disease diagnostics, specifically biosensors for nucleic acids and pathogenic microbes. We classify these strategies into target-based amplification and signal-based amplification. Target-based amplification strategies improve the LOD by increasing the number of detectable analytes, while signal-based amplification strategies increase the detectable signal by modifying the transducer system and keep the number of targets static. Finally, we argue that signal amplification strategies should be designed with application location and disease target in mind, and that the resources required to produce and operate the sensor should reflect its proposed application, especially when the platform is designed to be utilized in low-resource settings. We anticipate that, based on current technologies to diagnose infectious diseases, incorporating signal-based amplification strategies will enable electrochemical POC devices to be deployed for illnesses in a wide variety of settings.

9.
Cell Death Dis ; 13(3): 254, 2022 03 21.
Article in English | MEDLINE | ID: mdl-35314669

ABSTRACT

Tubulointerstitial fibrosis (TIF) is involved in the development of diabetic kidney disease (DKD). Transforming growth factor ß1 (TGF-ß1) is involved in the extensive fibrosis of renal tissue by facilitating the partial epithelial-mesenchymal transition (EMT), increasing the synthesis of extracellular matrix (ECM), inhibiting degradation, inducing apoptosis of renal parenchyma cells, and activating renal interstitial fibroblasts and inflammatory cells. Recent studies indicated that bone morphogenetic protein-7 (BMP-7) upregulated the expression of endogenous SnoN against renal TIF induced by TGF-ß1 or hyperglycemia. Nevertheless, the mechanisms underlying the BMP-7-mediated restoration of SnoN protein level remains elusive. The present study demonstrated the increased expression of BMP-7 in diabetic mellitus (DM) mice by hydrodynamic tail vein injection of overexpressed BMP-7 plasmid, which attenuated the effects of DM on kidney in mice. Partial tubular EMT and the accumulation of Collagen-III were resisted in DM mice that received overexpressed BMP-7 plasmid. Similar in vivo results showed that BMP-7 was competent to alleviate NRK-52E cells undergoing partial EMT in a high-glucose milieu. Furthermore, exogenous BMP-7 activated the Smad1/5 pathway to promote gene transcription of SnoN and intervened ubiquitination of SnoN; both effects repaired the SnoN protein level in renal tubular cells and kidney tissues of DM mice. Therefore, these findings suggested that BMP-7 could upregulate SnoN mRNA and protein levels by activating the classical Smad1/5 pathway to refrain from the partial EMT of renal tubular epithelial cells and the deposition of ECM in DKD-induced renal fibrosis.


Subject(s)
Diabetes Mellitus , Diabetic Nephropathies , Animals , Bone Morphogenetic Protein 7/metabolism , Diabetes Mellitus/pathology , Diabetic Nephropathies/pathology , Epithelial Cells/metabolism , Epithelial-Mesenchymal Transition/genetics , Fibrosis , Kidney Tubules/pathology , Mice , Smad1 Protein/metabolism , Transforming Growth Factor beta1/metabolism
10.
Front Pharmacol ; 13: 819787, 2022.
Article in English | MEDLINE | ID: mdl-35222033

ABSTRACT

Atorvastatin is a classical lipid-lowering drug. It has been reported to have renoprotective effects, such as reducing urinary protein excretion and extracellular matrix aggregation. The present study aimed to investigate the specific mechanism of action of Atorvastatin in type 1 diabetic mice (T1DM) in inhibiting renal tubular epithelial cell injury following treatment with high glucose and high fat. The anti-injury mechanism of Atorvastatin involved the inhibition of miR-21 expression and the upregulation of the transcription and expression of its downstream gene Peroxisome proliferator-activated receptors-α(PPARα). An increase in blood glucose and lipid levels was noted in the T1DM model, which was associated with renal fibrosis and inflammation. These changes were accompanied by increased miR-21 levels, downregulation of PPARα and Mfn1 expressions, and upregulation of Drp1 and IL6 expressions in renal tissues. These phenomena were reversed following the administration of Atorvastatin. miR-21 targeted PPARα by inhibiting its mRNA translation. Inhibition of miR-21 expression or Fenofibrate (PPARα agonist) administration prevented the decrease of PPARα in renal tubular epithelial cells under high glucose (HG) and high fat (Palmitic acid, PA) conditions, alleviating lipid metabolism disorders and reducing mitochondrial dynamics and inflammation. Consistent with the in vivo results, the in vitro findings also demonstrated that mRTECs administered with Atorvastatin in HG + PA increased PPARα expression and restored the normal expression of Mfn1 and Drp1, and effectively increasing the number of biologically active mitochondria and ATP content, reducing ROS production, and restoring mitochondrial membrane potential following Atorvastatin intervention. In addition, these effects were noted to the inhibition of FN expression and tubular cell inflammatory response; however, in the presence of miR-21mimics, the aforementioned effects of Atorvastatin were significantly diminished. Based on these observations, we conclude that Atorvastatin inhibits tubular epithelial cell injury in T1DM with concomitant induction of lipid metabolism disorders by a mechanism involving inhibition of miR-21 expression and consequent upregulation of PPARα expression. Moreover, Atorvastatin regulated lipid metabolism homeostasis and PPARα to restore mitochondrial function. The results emphasize the potential of Atorvastatin to exhibit lipid-regulating functions and non-lipid effects that balance mitochondrial dynamics.

11.
Front Med (Lausanne) ; 9: 1018298, 2022.
Article in English | MEDLINE | ID: mdl-36760880

ABSTRACT

Introduction: Diabetic kidney disease (DKD) is one of the complications of diabetes; however, the pathogenesis is not yet clear. A recent study has shown that senescence is associated with the course of DKD. In the present study, we explored whether senescent renal tubular cells promote renal tubulointerstitial fibrosis by secreting Sonic hedgehog (Shh) which mediates fibroblast activation and proliferation in DKD. Methods: A 36-week-old db/db mice model and the renal tubular epithelial cells were cultured in high glucose (HG, 60 mmol/L) medium for in vivo and in vitro experiments. Results: Compared to db/m mice, blood glucose, microalbuminuria, serum creatinine, urea nitrogen, and UACR (microalbuminuria/urine creatinine) were markedly increased in db/db mice. Collagen III, monocyte chemoattractant protein-1 (MCP-1), and tumor necrosis factor-alpha (TNF-α) were also increased in db/db mice kidneys, suggesting fibrosis and inflammation in the organ. Moreover, the detection of SA-ß-galactosidase (SA-ß-Gal) showed that the activity of SA-ß-Gal in the cytoplasm of renal tubular epithelial cells increased, and the cell cycle inhibition of the expression of senescence-related gene cell cycle inhibitor p16 INK4A protein and p21 protein increased, indicating that renal fibrosis in db/db mice was accompanied by cell senescence. Furthermore, Shh is highly expressed in the injured renal tubules and in the kidney tissue of db/db mice, as detected by enzyme-linked immunosorbent assay (ELISA). The results of immunofluorescence staining showed increased positive staining for Shh in renal tubular epithelial cells of db/db mice and decreased positive staining for Lamin B1, but increased positive staining for γH2A.X in cells with high Shh expression; similar results were obtained in vitro. In addition, HG stimulated renal tubular epithelial cells to secrete Shh in the supernatant of the medium. D-gal treatment of renal tubular epithelial cells increased the protein levels of Shh and p21. We also found enhanced activation and proliferation of fibroblasts cultured with the supernatant of renal tubular epithelial cells stimulated by HG medium but the proliferative effect was significantly diminished when co-cultured with cyclopamine (CPN), an inhibitor of the Shh pathway. Discussion: In conclusion, HG induces renal tubular epithelial cell senescence, and the secretion of senescence-associated proteins and Shh mediates inflammatory responses and fibroblast activation and proliferation, ultimately leading to renal fibrosis.

12.
Exp Cell Res ; 408(1): 112856, 2021 11 01.
Article in English | MEDLINE | ID: mdl-34597680

ABSTRACT

Renal interstitial fibrosis (RIF) is the common irreversible pathway by which chronic kidney disease (CKD) progresses to the end stage. The transforming growth factor-ß (TGF-ß)/signal transducer and activator of transcription 3 (STAT3) signaling pathway is a common factor leading to inflammation-mediated RIF, but its downstream regulatory mechanism is still unclear. Bioinformatics analysis predicted that serum amyloid A protein 1 (SAA1) was one of the target genes for transcriptional activation of STAT3 signaling. As an acute phase reaction protein, SAA1 plays an important role in many inflammatory reactions, and research has suggested that SAA1 is significantly elevated in the serum of patients with CKD. In this research, multiple experiments were performed to investigate the role of SAA1 in the process of RIF. SAA1 was abnormally highly expressed in kidney tissue from individuals who underwent unilateral ureteral obstruction (UUO) and TGF-ß-induced HK2 cells, and the abnormal expression was directly related to the transcriptional activation of STAT3. Additionally, SAA1 can directly target and bind valosin-containing protein (VCP)-interacting membrane selenoprotein (VIMP) to inhibit the function of the Derlin-1/VCP/VIMP complex, preventing the transportation and degradation of the misfolded protein, resulting in endoplasmic reticulum (ER) stress characterized by an increase in glucose-regulated protein 78 (GRP78) levels and ultimately promoting the occurrence and development of RIF.


Subject(s)
Endoplasmic Reticulum Stress/physiology , Fibrosis/metabolism , STAT3 Transcription Factor/metabolism , Serum Amyloid A Protein/metabolism , Animals , Endoplasmic Reticulum Chaperone BiP , Fibrosis/pathology , Humans , Inflammation/metabolism , Mice , Renal Insufficiency, Chronic/metabolism , Signal Transduction/physiology , Ureteral Obstruction/metabolism
13.
Biosci Rep ; 40(6)2020 06 26.
Article in English | MEDLINE | ID: mdl-32484208

ABSTRACT

Diabetic nephropathy (DN) commonly causes end-stage renal disease (ESRD). Increasing evidence indicates that abnormal miRNA expression is tightly associated with chronic kidney disease (CKD). This work aimed to investigate whether miR-27a can promote the occurrence of renal fibrosis in DN by suppressing the expression of secreted frizzled-related protein 1 (Sfrp1) to activate Wnt/ß-catenin signalling. Therefore, we assessed the expression levels of miR-27a, Sfrp1, Wnt signalling components, and extracellular matrix (ECM)-related molecules in vitro and in vivo. Sfrp1 was significantly down-regulated in a high-glucose environment, while miR-27a levels were markedly increased. A luciferase reporter assay confirmed that miR-27a down-regulated Sfrp1 by binding to the 3' untranslated region directly. Further, NRK-52E cells under high-glucose conditions underwent transfection with miR-27a mimic or the corresponding negative control, miR-27a inhibitor or the corresponding negative control, si-Sfrp1, or combined miR-27a inhibitor and si-Sfrp1. Immunoblotting and immunofluorescence were performed to assess the relative expression levels of Wnt/ß-catenin signalling and ECM components. The mRNA levels of Sfrp1, miR-27a, and ECM-related molecules were also detected by quantitative real-time PCR (qPCR). We found that miR-27a inhibitor inactivated Wnt/ß-catenin signalling and reduced ECM deposition. Conversely, Wnt/ß-catenin signalling was activated, while ECM deposition was increased after transfection with si-Sfrp1. Interestingly, miR-27a inhibitor attenuated the effects of si-Sfrp1. We concluded that miR-27a down-regulated Sfrp1 and activated Wnt/ß-catenin signalling to promote renal fibrosis.


Subject(s)
Diabetic Nephropathies/metabolism , Extracellular Matrix/metabolism , Intercellular Signaling Peptides and Proteins/metabolism , Kidney/metabolism , Membrane Proteins/metabolism , MicroRNAs/metabolism , Wnt Signaling Pathway , Animals , Blood Glucose/metabolism , Cell Line , Diabetes Mellitus, Experimental/blood , Diabetes Mellitus, Experimental/chemically induced , Diabetic Nephropathies/etiology , Diabetic Nephropathies/genetics , Diabetic Nephropathies/pathology , Extracellular Matrix/drug effects , Extracellular Matrix/pathology , Fibrosis , Glucose/toxicity , Intercellular Signaling Peptides and Proteins/genetics , Kidney/drug effects , Kidney/pathology , Male , Membrane Proteins/genetics , MicroRNAs/genetics , Rats, Sprague-Dawley , Streptozocin , Wnt Signaling Pathway/drug effects
14.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 36(3): 212-219, 2020 Mar.
Article in Chinese | MEDLINE | ID: mdl-32389168

ABSTRACT

Objective To investigate the effect of enhancer of zeste 2 polycomb repressive complex 2 subunit (EZH2) on the development of renal tubular epithelial-mesenchymal transformation (EMT) and diabetic nephropathy (DN). Methods Mouse model of type 1 diabetic nephropathy (T1DN) was established by intraperitoneal injection of streptozotocin (STZ) (55 mg/kg) and randomly divided into normal control and diabetic group. The mice were killed, and their biochemical indexes (blood glucose, creatinine, microalbumin and total protein in urine) of blood and urine were recorded. The kidneys were subjected to HE and Masson staining to observe morphological changes. Immunohistochemical staining was used to observe the expression and localization of type IV collagen (Col4) and EZH2. The mRNA and protein expressions of E-cadherin, α-smooth muscle actin (α-SMA), Col4 and EZH2 were detected by real-time fluorescent quantitative PCR and Western blotting. Rat renal tubular epithelial cells were cultured with high glucose and transfected with small interfering RNA (siRNA) of EZH2.The protein levels of E-cadherin, α-SMA, Col4 and EZH2 were detected by Western blot analysis. Results Compared with the normal group, blood glucose, serum creatinine, microalbuminuria and total urine protein significantly increased in the diabetic group. The mRNA and protein levels of E-cadherin, α-SMA, Col4 and EZH2 went up; the tubular lumen collapsed; the basement membrane of glomerulus thickened and there was a large amount of collagen deposition in the renal interstitium. Compared with normal sugar, high glucose stimulation promoted EMT and significantly up-regulated EZH2 expression. Compared with high glucose group, the transfection of EZH2 siRNA in high glucose inhibited EMT. Conclusion EZH2 can effectively promote the EMT process of renal tubular epithelial cells and may participate in the occurrence and development of DN through this role.


Subject(s)
Diabetes Mellitus, Experimental , Diabetic Nephropathies/pathology , Enhancer of Zeste Homolog 2 Protein/physiology , Epithelial Cells/metabolism , Epithelial-Mesenchymal Transition , Animals , Disease Progression , Kidney Tubules/cytology , Mice , Random Allocation
15.
J Pharm Sci ; 109(6): 2009-2017, 2020 06.
Article in English | MEDLINE | ID: mdl-32113978

ABSTRACT

Multiple pharmaceutical powder processes operate at stresses lower than utilized in typical lab-scale shear cell testing. To bridge this gap, we developed a method to determine intrinsic powder flow properties, in particular, flow function (FFc), under such low stresses. A simple, commercially-available flow-through-orifice device (Flodex™ apparatus) was selected. By developing a theoretical framework using Jenike's radial stress field analysis, the major principal stress and FFc of the tested powder can be derived from the otherwise empirical "Flodex" experiment. This method was applied to 10 distinct pharmaceutical powders. The major principal stresses associated with the test were estimated to be in the order of 100 Pa, significantly lower than what is achievable using shear cell-based methods. The resulting FFc values are generally consistent with the data extrapolated from ring shear testing. We showed that for pharmaceutical powders, FFc decreases with decreasing consolidation stress, but the values are always greater than 1. Therefore, the threshold for poor/acceptable flowability (by FFc) should be used with caution at low-stress conditions. Through this work, we showed that by integrating the radial stress field theory with a simple flow-through-orifice experiment, intrinsic powder flow properties under low stresses could be reliably determined.


Subject(s)
Excipients , Technology, Pharmaceutical , Particle Size , Physical Phenomena , Powders
16.
Int J Clin Exp Pathol ; 13(12): 3021-3032, 2020.
Article in English | MEDLINE | ID: mdl-33425103

ABSTRACT

OBJECTIVE: To investigate the expression of Stim1 in the kidneys of mice with lupus, and the effect of Stim1 on the progression of renal interstitial fibrosis. METHODS: Mice (MRL/lpr) with spontaneous lupus nephritis (LN) and normal control mice (C57/BL) were selected. Immunohistochemistry and Masson staining were used to determine the degree of renal interstitial fibrosis in kidney tissues. The expression of Stim1 and fibronectin in tissues was measured by qRT-PCR, western blotting, and immunohistochemistry. Urine protein, blood urea nitrogen, and serum creatinine levels in the mice were analyzed, and Spearman analysis was conducted to determine the correlation with Stim1 expression levels. Mouse renal tubular epithelial cells (mRTECs) were chosen as the experimental objects. After various treatments, the cells were divided into the blank control group, lipopolysaccharide (LPS) treatment group, LPS+siRNA-NC group and LPS+siRNA-Stim1 group. Western blotting and immunofluorescence were used to measure epithelial-mesenchymal transition (EMT)-related protein levels. RESULTS: There was significant interstitial fibrosis in the kidneys of LN mice. Compared with that in normal mice, the expression of Stim1 in the kidney tissues of LN mice was significantly increased, and Stim1 expression was positively correlated with fibronectin, urine protein, blood urea nitrogen and serum creatinine levels. LPS induced the expression of Stim1, fibronectin, and α-SMA in mRTECs and decreased the protein level of E-CA, while silencing Stim1 effectively alleviated the effects of LPS. CONCLUSION: Stim1 is significantly increased in the kidneys of lupus mice, and it is possible to promote EMT in renal tubular epithelial cells and renal interstitial fibrosis by elevating fibronectin, which ultimately contributes to renal damage.

17.
Life Sci ; 238: 116957, 2019 Dec 01.
Article in English | MEDLINE | ID: mdl-31655195

ABSTRACT

Epithelial-mesenchymal transition (EMT) and extracellular matrix (ECM) deposition in renal tubular epithelial cells are critical to diabetic nephropathy (DN) pathogenesis, but the underlying mechanisms remain undefined. Bone morphogenetic protein 7 (BMP-7) inhibits EMT and ECM accumulation in renal tubular epithelial cells cultured in presence of high glucose. Meanwhile, miRNA-21 (miR-21) downregulates Smad7, promoting EMT and ECM deposition. However, the association of BMP-7 with miR-21/Smad7 in DN is unknown. Here, NRK-52E cells incubated in presence of high glucose and STZ-induced C57BL diabetic mice were considered in vitro and in vivo models of DN, respectively. In both models, BMP-7 (mRNA/protein) amounts were decreased as well as Smad7 protein expression, while miR-21 expression and TGF-ß1/Smad3 pathway activation were enhanced, accompanied by enhanced EMT and ECM deposition. Further, addition of BMP-7 human recombinant cytokine (rhBMP-7) and injection of the BMP-7 overexpression plasmid in diabetic mice markedly downregulated miR-21 and upregulated Smad7, reduced Smad3 activation without affecting TGF-ß1 amounts, and prevented EMT and ECM accumulation. MiR-21 overexpression in the in vitro model downregulated Smad7, promoted EMT and ECM accumulation without affecting BMP-7 amounts, and miR-21 downregulation reversed it. By interfering with BMP-7 and miR-21 expression in high glucose conditions, miR-21 amounts and Smad3 phosphorylation were further decreased. Smad7 was then upregulated, and EMT and ECM deposition were inhibited; these effects were reversed after miR-21 overexpression. These findings suggest that BMP-7 decreases renal fibrosis in DN by regulating miR-21/Smad7 signaling, providing a theoretical basis for the development of novel and effective therapeutic drugs for DN.


Subject(s)
Bone Morphogenetic Protein 7/metabolism , Diabetes Mellitus, Experimental/physiopathology , Diabetic Nephropathies/complications , Fibrosis/prevention & control , Gene Expression Regulation , Kidney Tubules/metabolism , MicroRNAs/antagonists & inhibitors , Animals , Bone Morphogenetic Protein 7/genetics , Cells, Cultured , Epithelial-Mesenchymal Transition , Extracellular Matrix/metabolism , Fibrosis/etiology , Fibrosis/pathology , Kidney Tubules/pathology , Male , Mice , Mice, Inbred C57BL , MicroRNAs/genetics , Phosphorylation , Signal Transduction , Smad7 Protein/genetics , Smad7 Protein/metabolism , Transforming Growth Factor beta1/genetics , Transforming Growth Factor beta1/metabolism
18.
J Cell Physiol ; 234(10): 17925-17936, 2019 08.
Article in English | MEDLINE | ID: mdl-30847937

ABSTRACT

Unveiling the mechanisms that drive the pathological phenotypes of diabetic nephropathy (DN) could help develop new effective therapeutics for this ailment. Transforming growth factor-ß1 (TGF-ß1)/Smad3 signaling is aberrantly induced in DN, leading to elevated microRNA-21 (miR-21) expression and tissue fibrosis. Ski-related novel protein (SnoN) negatively regulates the TGF-ß pathway, but the relationship between SnoN and miR-21 has not been described in the context of DN. In this study, this association was investigated in vivo (streptozotocin-induced rat model of diabetes) and in vitro (NRK-52E model system under high glucose conditions). In both model systems, we observed reduced amounts of the SnoN protein and elevated miR-21 amounts, indicative of an inverse relationship. These changes in SnoN and miR-21 amounts were accompanied by reduced E-cadherin and elevated α-smooth muscle actin and collagen III levels, consistent with epithelial to mesenchymal transition (EMT). In vitro overexpression of SnoN in NRK-52E cells downregulated miR-21 at the transcriptional and posttranscriptional levels and repressed EMT and extracellular matrix (ECM) deposition. In contrast, knockdown of SnoN resulted in miR-21 upregulation, particularly at the transcriptional level. We further demonstrated that overexpression and inhibition of miR-21 promoted and suppressed EMT and ECM deposition, respectively, without affecting SnoN levels. Our results indicated that SnoN suppresses the development of DN as well as renal fibrosis by downregulating miR-21, and therefore represents a novel and promising therapeutic target for DN.


Subject(s)
Diabetes Mellitus, Experimental/metabolism , Diabetic Nephropathies/prevention & control , Kidney/metabolism , MicroRNAs/metabolism , Nerve Tissue Proteins/metabolism , Transcription Factors/metabolism , Transforming Growth Factor beta1/metabolism , Animals , Cell Line , Diabetes Mellitus, Experimental/chemically induced , Diabetes Mellitus, Experimental/genetics , Diabetic Nephropathies/etiology , Diabetic Nephropathies/genetics , Diabetic Nephropathies/metabolism , Epithelial-Mesenchymal Transition , Fibrosis , Gene Expression Regulation , Kidney/pathology , Male , MicroRNAs/genetics , Nerve Tissue Proteins/genetics , Rats, Sprague-Dawley , Signal Transduction , Smad3 Protein/metabolism , Streptozocin , Transcription Factors/genetics , Transforming Growth Factor beta1/genetics
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