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BACKGROUND/AIMS: The induction of excessive autophagy by increased levels of oxidative stress is one of the main mechanisms underlying unilateral ureteral obstruction (UUO)-induced vascular endothelial cell dysfunction. Hydrogen sulfide (H2S) has been shown to have an anti-oxidative effect, but its mode of action on excessive autophagy in vascular endothelial cells is unclear. METHODS: Surgery was used to induce UUO in male C57BL/6 mice as an in vivo model. Human renal epithelial cells (HK-2) were treated with H2O2 as an in vitro model. NaHS was used as an exogenous H2S donor. Transmission electron microscopy was applied to observe the structure of renal autophagosomes. The expression of proteins related to autophagy and apoptosis was detected by western blot analysis in vivo and in vitro. Flow cytometry (DCFH-DA) was used to examine the levels of intracellular reactive oxygen species (ROS). The terminal deoxynucleotidyl transferase dUTP nick end labeling assay was used to detect cell apoptosis. Compound C was used to analyze the association of AMPK with autophagy. RESULTS: Compared with the sham group, in which the ureter was exposed but not ligated, the cell apoptosis index, number of autophagosomes, protein expression of microtubule-associated protein 1 light-chain 3 (LC3)-II/I, beclin-1, and p-AMPK/AMPK were significantly increased in the UUO group. On the other hand, p62, cystathionine ß-synthase, and cystathionine γ-lyase protein expression levels and H2S concentration were significantly decreased (p < 0.05). These alterations were ameliorated by the addition of NaHS (p < 0.05). Similar results were observed in vitro. By using the AMPK inhibitor compound C, it was indicated that AMPK was involved in ROS-induced autophagy. In addition, using tissue from patients with obstructive nephropathy, excessive autophagy was observed by an increased LC3-II/LC3-I ratio. CONCLUSION: NaHS-treatment may exert a protective effect on mouse kidney against UUO by suppressing the ROS-AMPK pathway. ROS-AMPK-mediated autophagy may represent a promising therapeutic target for obstructive nephropathy.
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Proteínas Quinases Ativadas por AMP/metabolismo , Autofagia/efeitos dos fármacos , Sulfeto de Hidrogênio/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteínas Quinases Ativadas por AMP/antagonistas & inibidores , Animais , Autofagossomos/metabolismo , Células Cultivadas , Cistationina gama-Liase/metabolismo , Humanos , Túbulos Renais Proximais/química , Túbulos Renais Proximais/efeitos dos fármacos , Túbulos Renais Proximais/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Associadas aos Microtúbulos/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Fator de Transcrição TFIIH , Fatores de Transcrição/metabolismo , Obstrução Ureteral/metabolismo , Obstrução Ureteral/patologia , Obstrução Ureteral/veterináriaRESUMO
This study was aimed to investigate the preventive effects of N-acetyl-L-cysteine (NAC) against renal tubular cell injury induced by oxalate and stone formation and further explore the related mechanism. Transcriptome sequencing combined with bioinformatics analysis were performed to identify differentially expressed gene (DEG) and related pathways. HK-2 cells were pretreated with or without antioxidant NAC/with or silencing DEG before exposed to sodium oxalate. Then, the cell viability, oxidative biomarkers of superoxidase dismutase (SOD) and malondialdehyde (MDA), apoptosis and cell cycle were measured through CCK8, ELISA and flow cytometry assay, respectively. Male SD rats were separated into control group, hyperoxaluria (HOx) group, NAC intervention group, and TGF-ß/SMAD pathway inhibitor group. After treatment, the structure changes and oxidative stress and CaOx crystals deposition were evaluated in renal tissues by H&E staining, immunohistochemical and Pizzolato method. The expression of TGF-ß/SMAD pathway related proteins (TGF-ß1, SMAD3 and SMAD7) were determined by Western blot in vivo and in vitro. CDKN2B is a DEG screened by transcriptome sequencing combined with bioinformatics analysis, and verified by qRT-PCR. Sodium oxalate induced declined HK-2 cell viability, in parallel with inhibited cellular oxidative stress and apoptosis. The changes induced by oxalate in HK-2 cells were significantly reversed by NAC treatment or the silencing of CDKN2B. The cell structure damage and CaOx crystals deposition were observed in kidney tissues of HOx group. Meanwhile, the expression levels of SOD and 8-OHdG were detected in kidney tissues of HOx group. The changes induced by oxalate in kidney tissues were significantly reversed by NAC treatment. Besides, expression of SMAD7 was significantly down-regulated, while TGF-ß1 and SMAD3 were accumulated induced by oxalate in vitro and in vivo. The expression levels of TGF-ß/SMAD pathway related proteins induced by oxalate were reversed by NAC. In conclusion, we found that NAC could play an anti-calculus role by mediating CDKN2B/TGF-ß/SMAD axis.
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Hiperoxalúria , Oxalatos , Animais , Masculino , Ratos , Acetilcisteína/farmacologia , Oxalato de Cálcio/metabolismo , Células Epiteliais/metabolismo , Hiperoxalúria/induzido quimicamente , Hiperoxalúria/metabolismo , Oxalatos/metabolismo , Ratos Sprague-Dawley , Superóxido Dismutase/metabolismo , Fator de Crescimento Transformador beta1/metabolismoRESUMO
BACKGROUNDDue to the long time interval between exposure and outcome, it is difficult to infer the causal relationship between educational attainment (EA) and common chronic diseases. Therefore, we utilized Mendelian randomization (MR) to predict the causal relationships of EA with hypertension and type-2 diabetes (T2DM). METHODSA two-sample MR analysis was conducted using genome-wide association studies (GWASs) combined with inferential measurements. A GWAS meta-analysis including 1,131,881 European individuals was used to identify instruments for EA. Hypertension and T2DM data were obtained from a Finnish database. MR analyses were performed using inverse-variance weighted meta-analysis (IVW), weighted median regression, MRâEgger regression, simple mode regression, weighted mode regression and the MR-Pleiotropy RESidual Sum and Outlier test. Sensitivity analyses were further performed using the leave-one-out method to test the robustness of our findings. RESULTSUsing the MR approach, our results showed that EA was significantly associated with a reduced risk of hypertension (OR = 0.63; P = 2.94 × 10-47; [95% CI: 0.59, 0.67]) and type-2 diabetes (OR = 0.59; P = 1.25 × 10-16; [95% CI: 0.52, 0.67]). CONCLUSIONSThis study showed that EA is causally linked to the risk of chronic diseases, including high blood pressure and T2DM.
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Calcium oxalate (CaOx) crystals can activate autophagy, causing damage to renal tubular epithelial cells (TECs). Puerarin has been shown to have protective and therapeutic effects against a variety of diseases by inhibiting autophagy activation. However, the protective effect of puerarin against CaOx crystals and the underlying molecular mechanisms are unclear. Cell Counting Kit-8 (CCK-8) assays were used to evaluate the effects of puerarin on cell viability. Intracellular reactive oxygen species (ROS) levels were measured by the cell-permeable fluorogenic probe 2,7-dichlorofluorescein diacetate (DCFH-DA). Immunofluorescence, immunohistochemistry, and western blotting were used to examine the expression of SIRT1, Beclin1, p62, and LC3, and explore the underlying molecular mechanisms in vivo and in vitro. Puerarin treatment significantly attenuated CaOx crystal-induced autophagy of TECs and CaOx cytotoxicity to TECs by altering SIRT1 expression in vitro and in vivo, whereas the SIRT1-specific inhibitor EX527 exerted contrasting effects. In addition, we found that the protective effect of puerarin was related to the SIRT1/AKT/p38 signaling pathway. The findings suggest that puerarin regulates CaOx crystal-induced autophagy by activating the SIRT1-mediated signaling pathway, and they suggest a series of potential therapeutic targets and strategies for treating nephrolithiasis.
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Oxalato de Cálcio , Cálculos Renais , Autofagia , Oxalato de Cálcio/metabolismo , Células Epiteliais/metabolismo , Humanos , Isoflavonas , Cálculos Renais/metabolismo , Estresse Oxidativo , Transdução de Sinais , Sirtuína 1/metabolismo , Sirtuína 1/farmacologiaRESUMO
PURPOSE: We evaluated the possible involvement of phospholipid transporters and reactive oxygen species in the oxalate induced redistribution of renal epithelial cell phosphatidylserine. MATERIALS AND METHODS: Madin-Darby canine kidney cells were labeled with the fluorescent phospholipid NBD-PS in the inner or outer leaflet of the plasma membrane and then exposed to oxalate in the presence or absence of antioxidant. This probe was tracked using a fluorescent quenching assay to assess the bidirectional transmembrane movement of phosphatidylserine. Surface expressed phosphatidylserine was detected by annexin V binding assay. The cell permeable fluorogenic probe DCFH-DA was used to measure the intracellular reactive oxygen species level. RESULTS: Oxalate produced a time and concentration dependent increase in phosphatidylserine, which may have resulted from impaired aminophospholipid translocase mediated, inward directed phosphatidylserine transport and from enhanced phosphatidylserine outward transport. Adding the antioxidant N-acetyl-L-cysteine significantly attenuated phosphatidylserine externalization by effectively rescuing aminophospholipid translocase activity. CONCLUSIONS: To our knowledge our findings are the first to show that oxalate induced increased reactive oxygen species generation impairs aminophospholipid translocase activity and decreased aminophospholipid translocase activity has a role in hyperoxaluria promoted calcium oxalate urolithiasis by facilitating phosphatidylserine redistribution in renal epithelial cells.
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Oxalato de Cálcio/metabolismo , Células Epiteliais/metabolismo , Rim/citologia , Estresse Oxidativo , Proteínas de Transferência de Fosfolipídeos/metabolismo , Urolitíase/etiologia , Animais , Células Cultivadas , CãesRESUMO
Arsenic trioxide has shown remarkable biological activity against bladder cancer in some clinical studies. However, the mechanism of its action is unknown. Our aim was to find the relationship between miRNAs and arsenic trioxide treatment by using T24 human bladder carcinoma cells. By performing microRNA microarray and quantitative real-time PCR after ATO treatment, we found that expression levels of several miRNAs, in particular, miRNA-19a, were significantly decreased in T24 cell line. Furthermore, cell proliferation assay, flow cytometry analysis, prediction of miRNA targets, Western blot analysis, and luciferase reporter assay were performed to determine the role of mir-19a in affecting the biological behaviors of T24 cells. Several miRNAs were up-regulated or down-regulated in T24 cells treated with arsenic trioxide compared to their controls. If only changes above two folds were considered, two miRNAs were identified, miRNA-19a was down-regulated, while miRNA-222* was up-regulated. Among them, knockdown of miRNA-19a by anti-miRNA-19a transfection showed a positive therapeutic effect in bladder cancer cells by inhibiting cell growth and inducing cell apoptosis targeting PTEN through the PTEN/Akt pathway. Besides this, a synergy effect was detected between knockdown of miRNA-19a and arsenic trioxide. Arsenic trioxide altered miRNA expression profile in T24 cells. It seems miRNA-19a plays a critical role in the mechanism of arsenic trioxide treatment in bladder cancer. The synergy effect between miRNA-19a and arsenic trioxide that advocates targeting the mir-19a may represent a potential approach to enhance the efficacy and safety of ATO to treat bladder cancer by a decrease in dose.
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Antineoplásicos/farmacologia , Arsenicais/farmacologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , MicroRNAs/fisiologia , Óxidos/farmacologia , PTEN Fosfo-Hidrolase/genética , Neoplasias da Bexiga Urinária/tratamento farmacológico , Neoplasias da Bexiga Urinária/genética , Apoptose/efeitos dos fármacos , Trióxido de Arsênio , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Western Blotting , Proliferação de Células/efeitos dos fármacos , Perfilação da Expressão Gênica , Humanos , Luciferases/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , PTEN Fosfo-Hidrolase/metabolismo , RNA Mensageiro/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células Tumorais Cultivadas , Neoplasias da Bexiga Urinária/patologiaRESUMO
BACKGROUND: Chronic low back pain (CLBP) is 1 of the common clinical diseases, and many treatment methods can only improve the symptoms of pain in the short term. Traditional Chinese sports - Baduanjin has been proven to have a positive effect on chronic low back pain. However, the quality of the research is low, the sample size is small, and safety observations are lacking. We describe the protocol of a randomized controlled trial to study the efficacy and safety of Baduanjin chronic low back pain. METHODS: This randomized, controlled, evaluator-blind, two-arm, parallel clinical trial will include 90 outpatients with chronic low back pain recruited from the First Hospital of Nanping City, Fujian Province. The patients were randomly assigned to the intervention group (Baduanjin exercise training) and the control group (not receiving any special exercise training) at a ratio of 1:1. Patients in the intervention group will receive Baduanjin exercise training 3 times a week for 24âweeks. The 2 groups received a 4- week follow-up observation at 24âweeks. The main result from the intervention before intervention to 24âweeks later, and the follow-up of 4 changes the visual analog scale score at weeks, and by independent t are tested groups. It will also review the Pain-related disability index, The Quebec Back Pain Disability Scale, Health-related quality of life, Roland Morris (Roland Morris) Disability Questionnaire, Overall Perceived Effect (OPE) and safety Compare. Cost data for cost-benefit and cost-benefit analysis will be collected. DISCUSSION: This will be the first study to compare the effectiveness and safety of Baduanjin for patients with chronic low back pain. The results may help healthcare professionals make clinical decisions and may reduce the cost of treatment for this disease. TRIAL REGISTRATION: ChiCTR2000033908.
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Dor Crônica/terapia , Técnicas de Exercício e de Movimento/métodos , Dor Lombar/terapia , Medicina Tradicional Chinesa/métodos , Adulto , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Medição da Dor , Ensaios Clínicos Controlados Aleatórios como Assunto , Método Simples-Cego , Resultado do Tratamento , Adulto JovemRESUMO
Of all pathological types of renal cell cancer (RCC), clear cell renal cell carcinoma (ccRCC) has the highest incidence. Cyclovirobuxine (CVB), a triterpenoid alkaloid isolated from Buxus microphylla, exhibits antitumour activity against gastric cancer and breast cancer; however, the mechanism by which CVB inhibits ccRCC remains unclear. The aim of our study was to explore the antitumour effects of CVB on ccRCC and to elucidate its exact mechanism. Cell viability, proliferation, cell cycle distribution, apoptosis, wound healing and invasion were evaluated. Furthermore, Western blotting, immunofluorescence staining, immunohistochemical staining, and bioinformatics analyses were utilized to comprehensively probe the molecular mechanisms. The in vivo curative effect of CVB was explored using a 786-O xenograft model established in nude mice. CVB reduced cell viability, proliferation, angiogenesis, the epithelial-mesenchymal transition (EMT), migration and invasion. In addition, CVB induced cell cycle arrest in S phase and promoted apoptosis. The expression of the EMT-related transcription factor Snail was significantly downregulated by CVB via the inhibition of the AKT, STAT3 and MAPK pathways. We revealed that insulin-like growth factor binding protein 3 (IGFBP3) was the true therapeutic target of CVB. CVB exerted anti-ccRCC effects by blocking the IGFBP3-AKT/STAT3/MAPK-Snail pathway. Targeted inhibition of IGFBP3 with CVB treatment may become a promising therapeutic regimen for ccRCC.
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Carcinoma de Células Renais/tratamento farmacológico , Medicamentos de Ervas Chinesas/uso terapêutico , Neoplasias Renais/tratamento farmacológico , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Animais , Carcinoma de Células Renais/metabolismo , Linhagem Celular Tumoral , Medicamentos de Ervas Chinesas/farmacologia , Células Endoteliais da Veia Umbilical Humana , Humanos , Proteína 3 de Ligação a Fator de Crescimento Semelhante à Insulina/metabolismo , Neoplasias Renais/metabolismo , Camundongos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Distribuição Aleatória , Fator de Transcrição STAT3/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
AIMS: Telmisartan (TLM), a highly selective angiotensin II type 1 receptor blocker (ARB) and partial PPAR-γ agonist, has versatile beneficial effects against oxidative stress, apoptosis, inflammatory responses and epithelial-mesenchymal transition (EMT). However, its underlying mechanism of inhibiting oxalate and calcium oxalate (CaOx) crystal-induced EMT by activating the PPAR-γ pathway remains unclear. MAIN METHODS: CCK-8 assays were used to evaluate the effects of TLM on cell viability. In addition, intracellular reactive oxygen species (ROS) levels were measured by the cell-permeable fluorogenic probe 2,7-dichlorofluorescein diacetate (DCFH-DA). Wound-healing and Transwell assays were used to evaluate the migration ability of HK2 cells exposed to oxalate. Moreover, immunofluorescence, immunohistochemistry and western blotting were used to examine the expression of E-cadherin, N-cadherin, vimentin and α-SMA and explore the underlying molecular mechanisms in HK2 cells and a stone-forming rat model. KEY FINDINGS: Our results showed that TLM treatment could protect HK2 cells from oxalate-induced cytotoxicity and oxidative stress injury. Additionally, TLM prevented EMT induction by oxalate and CaOx crystals via the PPAR-γ-AKT/STAT3/p38 MAPK-Snail pathway in vitro and in vivo. However, knockdown of PPAR-γ with small interfering RNA or the PPAR-γ-specific antagonist GW9662 abrogated these protective effects of TLM. SIGNIFICANCE: As a PPAR-γ agonist, TLM can ameliorate oxalate and CaOx crystal-induced EMT by exerting an antioxidant effect through the PPAR-γ-AKT/STAT3/p38 MAPK-Snail signaling pathway. Therefore, TLM can block EMT progression and could be a potential therapeutic agent for preventing and treating calcium oxalate urolithiasis formation and recurrence.
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Transição Epitelial-Mesenquimal/efeitos dos fármacos , Oxalatos/toxicidade , PPAR gama/metabolismo , Telmisartan/farmacologia , Animais , Oxalato de Cálcio/toxicidade , Linhagem Celular , Transição Epitelial-Mesenquimal/fisiologia , Humanos , Túbulos Renais/citologia , Masculino , Estresse Oxidativo/efeitos dos fármacos , PPAR gama/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos Sprague-Dawley , Fator de Transcrição STAT3/metabolismo , Fatores de Transcrição da Família Snail/metabolismo , Fator de Crescimento Transformador beta1/toxicidade , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
AIMS: Oxymatrine (OMT) has been identified to possess immunomodulatory, antiinflammatory and anticancer properties. This study aimed to investigate its precise function and the underlying molecular mechanisms in renal cell carcinoma progression. METHODS: The antineoplastic effect of oxymatrine was investigated by CCK-8 assay, cell cycle analysis, apoptosis assay, wound healing experiment, transwell assay, and drug-sensitivity analysis in renal cancer cells following oxymatrine treatment. The modulation of oxymatrine on ß-catenin was analyzed through western blot and immunofluorescence assay. ß-catenin overexpression was employed to determine the key role of ß-catenin in oxymatrine-inhibited renal cell carcinoma in vitro. In addition, animal model was established to investigate the effect of oxymatrine on tumor growth in vivo. RESULTS: Oxymatrine inhibited renal cell carcinoma progression in vitro, including cell proliferation, apoptosis, migration, invasion and chemotherapy sensitivity. Further mechanistic studies demonstrated that oxymatrine exerted its antineoplastic effect through suppressing the expression of ß-catenin. Moreover, in nude mice model, oxymatrine exhibited remarkable inhibition of tumor growth, which was consistent with our in vitro results. CONCLUSIONS: Our findings illuminate oxymatrine as an effective antitumor agent in renal cell carcinoma, and suggest it a promising therapeutic application in renal cell carcinoma treatment.
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OBJECTIVES: Calcium oxalate (CaOx) crystals can activate inflammatory cytokines by triggering inflammasomes, which cause damage to the adhered epithelium, a dysfunctional microenvironment and even renal failure. However, a comprehensive and in-depth understanding of the mechanisms underlying the effects of these crystals on damage and cytokine function in renal tubular epithelial cells (TECs) remains limited and to be explored. MATERIALS AND METHODS: We detected the pyroptosis of TECs induced after exposure to CaOx crystals and demonstrated the significance of cytokine activation in the subsequent inflammatory processes through a proteomic study. We then conducted animal and cell experiments to verify relevant mechanisms through morphological, protein, histological and biochemical approaches. Human serum samples were further tested to help explain the pathophysiological mechanism of H3 relaxin. RESULTS: We verified that crystal-induced extracellular adenosine triphosphate (ATP) upregulation via the membrane purinergic 2X7 receptor (P2X7 R) promotes ROS generation and thereby activates NLRP3 inflammasome-mediated interleukin-1ß/18 maturation and gasdermin D cleavage. Human recombinant relaxin-3 (H3 relaxin) can act on the transmembrane receptor RXFP1 to produce cAMP and subsequently improves crystal-derived damage via ATP consumption. Additionally, endogenous relaxin-3 was found to be elevated in patients with renal calculus and can thus serve as a biomarker. CONCLUSIONS: Our results provide previously unidentified mechanistic insights into CaOx crystal-induced inflammatory pyroptotic damage and H3 relaxin-mediated anti-inflammatory protection and thus suggest a series of potential therapeutic targets and methods for but not limited to nephrocalcinosis.
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Anti-Inflamatórios/farmacologia , Oxalato de Cálcio/farmacologia , Piroptose/efeitos dos fármacos , Relaxina/farmacologia , Trifosfato de Adenosina/metabolismo , Animais , Oxalato de Cálcio/química , Linhagem Celular , AMP Cíclico/metabolismo , Humanos , Inflamassomos/metabolismo , Interleucina-1beta/metabolismo , Rim/efeitos dos fármacos , Rim/metabolismo , Rim/patologia , Túbulos Renais Proximais/citologia , Túbulos Renais Proximais/metabolismo , Masculino , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Ratos , Ratos Sprague-Dawley , Espécies Reativas de Oxigênio/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Receptores de Peptídeos/metabolismo , Receptores Purinérgicos P2X7/metabolismo , Relaxina/sangueRESUMO
Peroxisome proliferator-activated receptor- (PPAR-) γ is a ligand-dependent transcription factor, and it has become evident that PPAR-γ agonists have renoprotective effects, but their influence and mechanism during the development of calcium oxalate (CaOx) nephrolithiasis remain unknown. Rosiglitazone (RSG) was used as a representative PPAR-γ agonist in our experiments. The expression of transforming growth factor-ß1 (TGF-ß1), hepatocyte growth factor (HGF), c-Met, p-Met, PPAR-γ, p-PPAR-γ (Ser112), Smad2, Smad3, pSmad2/3, and Smad7 was examined in oxalate-treated Madin-Darby canine kidney (MDCK) cells and a stone-forming rat model. A CCK-8 assay was used to evaluate the effects of RSG on cell viability. In addition, intracellular reactive oxygen species (ROS) levels were monitored, and lipid peroxidation in renal tissue was detected according to superoxide dismutase and malondialdehyde levels. Moreover, the location and extent of CaOx crystal deposition were evaluated by Pizzolato staining. Our results showed that, both in vitro and in vivo, oxalate impaired PPAR-γ expression and phosphorylation, and then accumulative ROS production was observed, accompanied by enhanced TGF-ß1 and reduced HGF. These phenomena could be reversed by the addition of RSG. RSG also promoted cell viability and proliferation and decreased oxidative stress damage and CaOx crystal deposition. However, these protective effects of RSG were abrogated by the PPAR-γ-specific inhibitor GW9662. Our results revealed that the reduction of PPAR-γ activity played a critical role in oxalate-induced ROS damage and CaOx stone formation. RSG can regulate TGF-ß1 and HGF/c-Met through PPAR-γ to exert antioxidant effects against hyperoxaluria and alleviate crystal deposition. Therefore, PPAR-γ agonists may be expected to be a novel therapy for nephrolithiasis, and this effect is related to PPAR-γ-dependent suppression of oxidative stress.
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Oxalato de Cálcio/metabolismo , Células Epiteliais/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Fator de Crescimento de Hepatócito/biossíntese , Rim/metabolismo , Estresse Oxidativo/efeitos dos fármacos , PPAR gama/metabolismo , Rosiglitazona/farmacologia , Transdução de Sinais/efeitos dos fármacos , Fator de Crescimento Transformador beta1/metabolismo , Animais , Cães , Células Epiteliais/patologia , Hiperoxalúria/tratamento farmacológico , Hiperoxalúria/metabolismo , Hiperoxalúria/patologia , Rim/patologia , Células Madin Darby de Rim Canino , Masculino , Nefrolitíase/tratamento farmacológico , Nefrolitíase/metabolismo , Nefrolitíase/patologia , Ratos Sprague-DawleyRESUMO
The aim of the present study was to identify potential key genes and single nucleotide variations (SNVs) in prostate cancer. RNA sequencing (RNA-seq) data, GSE22260, were downloaded from the Gene Expression Omnibus database, including 4 prostate cancer samples and 4 normal tissues samples. RNA-Seq reads were processed using Tophat and differentially-expressed genes (DEGs) were identified using the Cufflinks package. Gene Ontology enrichment analysis of DEGs was performed. Subsequently, Seqpos was used to identify the potential upstream regulatory elements of DEGs. SNV was analyzed using Genome Analysis Toolkit. In addition, the frequency and risk-level of mutant genes were calculated using VarioWatch. A total of 150 upregulated and 211 downregulated DEGs were selected and 25 upregulated and 17 downregulated potential upstream regulatory elements were identified, respectively. The SNV annotations of somatic mutations revealed that 65% were base transition and 35% were base transversion. At frequencies ≥2, a total of 17 mutation sites were identified. The mutation site with the highest frequency was located in the folate hydrolase 1B (FOLH1B) gene. Furthermore, 20 high-risk mutant genes with high frequency were identified using VarioWatch, including ribosomal protein S4 Y-linked 2 (RPS4Y2), polycystin 1 transient receptor potential channel interacting (PKD1) and FOLH1B. In addition, kallikrein 1 (KLK1) and PKD1 are known tumor suppressor genes. The potential regulatory elements and high-frequency mutant genes (RPS4Y2, KLK1, PKD1 and FOLH1B) may have key functions in prostate cancer. The results of the present study may provide novel information for the understanding of prostate cancer development.
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OBJECTIVE: To investigate the possible involvement of multidrug resistance-associated protein 1 (MRP-1) and breast cancer resistance protein (BCRP) in the oxalate-induced redistribution of phosphatidylserine (PS) in renal epithelial cell membranes. METHODS: A western blot analysis was used to examine the MRP-1 and BCRP expression levels. Surface-expressed PS was detected by the annexin V-binding assay. The cell-permeable fluorogenic probe 2,7-dichlorofluorescein diacetate was used to measure the intracellular reactive oxygen species (ROS) level. A rat model of hyperoxaluria was obtained using 0.5% ethylene glycol and 1.0% ammonium chloride. In addition, certain animals received verapamil (50 mg/kg body weight), which is a common inhibitor of MRP-1 and BCRP. The degree of nephrolithiasis was assessed histomorphometrically using sections stained by Pizzolato method and by measuring the calcium oxalate crystal content in the renal tissue. RESULTS: Oxalate produced a concentration-dependent increase in the synthesis of MRP-1 and BCRP. Treatment with MK571 and Ko143 (MRP-1- and BCRP-specific inhibitors, respectively) significantly attenuated the oxalate-induced PS externalization. Adding the antioxidant N-acetyl-l-cysteine significantly reduced MRP-1 and BCRP expression. In vivo, markedly decreased nephrocalcinosis was observed compared with that in the rat model of hyperoxaluria without verapamil treatment. CONCLUSION: Oxalate induces the upregulation of MRP-1 and BCRP, which act as phospholipid floppases causing PS externalization in the renal epithelial cell membrane. The process is mediated by intracellular ROS production. The ROS-mediated increase in the synthesis of MRP-1 and BCRP can play an important role in hyperoxaluria-promoted calcium oxalate urolithiasis by facilitating phosphatidylserine redistribution in renal epithelial cells.
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Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/genética , Proteínas Associadas à Resistência a Múltiplos Medicamentos/genética , Nefrolitíase/metabolismo , Oxalatos/farmacologia , Fosfatidilserinas/metabolismo , RNA/genética , Urotélio/efeitos dos fármacos , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/biossíntese , Animais , Western Blotting , Células Cultivadas , Modelos Animais de Doenças , Citometria de Fluxo , Imuno-Histoquímica , Masculino , Proteínas Associadas à Resistência a Múltiplos Medicamentos/biossíntese , Nefrolitíase/tratamento farmacológico , Nefrolitíase/patologia , Ratos , Ratos Sprague-Dawley , Espécies Reativas de Oxigênio/metabolismo , Urotélio/patologiaRESUMO
OBJECTIVES: We investigated the possible involvement of multidrug resistance protein 1 P-glycoprotein (MDR1 P-gp) in the oxalate-induced redistribution of phosphatidylserine in renal epithelial cell membranes. METHODS: Real-time PCR and western blotting were used to examine MDR1 expression in Madin-Darby canine kidney cells at the mRNA and protein levels, respectively, whereas surface-expressed phosphatidylserine was detected by the annexin V-binding assay. RESULTS: Oxalate treatment resulted in increased synthesis of MDR1, which resulted in phosphatidylserine (PS) externalization in the renal epithelial cell membrane. Treatment with the MDR1 inhibitor PSC833 significantly attenuated phosphatidylserine externalization. Transfection of the human MDR1 gene into renal epithelial cells significantly increased PS externalization. CONCLUSIONS: To our knowledge, this study is the first to show that oxalate increases the synthesis of MDR1 P-gp, which plays a key role in hyperoxaluria-promoted calcium oxalate urolithiasis by facilitating phosphatidylserine redistribution in renal epithelial cells.
Assuntos
Membro 1 da Subfamília B de Cassetes de Ligação de ATP/genética , Regulação da Expressão Gênica , Nefrolitíase/genética , Oxalatos/efeitos adversos , Fosfatidilserinas/metabolismo , RNA Mensageiro/genética , Urotélio/metabolismo , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Animais , Western Blotting , Membrana Celular/metabolismo , Membrana Celular/patologia , Células Cultivadas , Ciclosporinas/farmacologia , Cães , Resistência a Múltiplos Medicamentos , Citometria de Fluxo , Humanos , Nefrolitíase/tratamento farmacológico , Nefrolitíase/metabolismo , Fosfatidilserinas/antagonistas & inibidores , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Urotélio/efeitos dos fármacos , Urotélio/patologiaRESUMO
Cephalopods (octopus, squid and cuttlefish) are some of the most intriguing molluscs, and they represent economically important commercial marine species for fisheries. Previous studies have shown that cephalopods are sensitive to underwater particle motion, especially at low frequencies in the order of 10 Hz. The present paper deals with quantitative modeling of the statocyst system in three cephalopod species: Octopus vulgaris, Sepia officinalis and Loligo vulgaris. The octopus's macula/statolith organ was modeled as a 2nd-order dynamic oscillator using parameter values estimated from scanning electron micrograph images. The modeling results agree reasonably well with experimental data (acceleration threshold) in the three cephalopod species. Insights made from quantitative modeling and simulating the particle motion sensing mechanism of cephalopods elucidated their underwater particle motion detection capabilities. Sensitivity to emerging environmental issues, such as low frequency noise caused by near-shore wind farms and increasing levels of carbon dioxide in the ocean, and sensitivity to sounds produced by impending landslides were investigated in octopus using the model.