RESUMO
Single-cell RNA sequencing (scRNAseq) is a crucial tool in kidney research. These technologies cluster cells based on transcriptome similarity, irrespective of the anatomical location and order within the nephron. Thus, a transcriptome cluster may obscure the heterogeneity of the cell population within a nephron segment. Elevated dietary fructose leads to salt-sensitive hypertension, in part, through fructose reabsorption in the proximal tubule (PT). However, the organization of the four known fructose transporters in apical PTs (SGLT4, SGLT5, GLUT5, and NaGLT1) remains poorly understood. We hypothesized that cells within each subsegment of the proximal tubule exhibit complex, heterogeneous fructose transporter expression patterns. To test this hypothesis, we analyzed rat kidney transcriptomes and proteomes from publicly available scRNAseq and tubule microdissection databases. We found that microdissected PT-S1 segments consist of 81% ± 12% cells with scRNAseq-derived transcriptional characteristics of S1, whereas PT-S2 express a mixture of 18% ± 9% S1, 58% ± 8% S2, and 19% ± 5% S3 transcripts, and PT-S3 consists of 75% ± 9% S3 transcripts. The expression of all four fructose transporters was detectable in all three PT segments, but key fructose transporters SGLT5 and GLUT5 progressively increased from S1 to S3, and both were significantly upregulated in S3 vs. S1/S2 (Slc5a10: 1.9 log2FC, p < 1 × 10-299; Scl2a5: 1.4 log2FC, p < 4 × 10-105). A similar distribution was found in human kidneys. These data suggest that S3 is the primary site of fructose reabsorption in both humans and rats. Finally, because of the multiple scRNAseq transcriptional phenotypes found in each segment, our findings also imply that anatomical labels applied to scRNAseq clusters may be misleading.
Assuntos
Frutose , Transcriptoma , Humanos , Ratos , Animais , Frutose/metabolismo , Néfrons/metabolismo , Rim/metabolismo , Túbulos Renais Proximais/metabolismo , Proteínas de Membrana Transportadoras/metabolismoRESUMO
Long non-coding RNAs are considered to be key regulatory factors of oncogenesis and tumor progression. It is reported that LINC00460 plays the role of oncogene in some tumors. However, LINC00460's role and mechanism of action in pancreatic cancer have not yet been fully elucidated. We identified LINC00460 by analyzing data from the Gene Expression Omnibus database. The role of LINC00460 in proliferation and metastasis was examined using CCK8, colony formation, wound healing, and transwell assays. The potential mechanisms of LINC00460 in regulating mRNA levels were elucidated by RNA pull-down, RNA immunoprecipitation, Chromatin immunoprecipitation, Co-immunoprecipitation, and Immunofluorescence assays. The results showed that LINC00460 was upregulated in pancreatic cancer cells and tissues. Highly expressed LINC00460 is significantly related to short survival of pancreatic cancer patients. Inhibition of LINC00460 attenuated pancreatic cancer cell proliferation and metastasis, whereas its overexpression reversed this effect. Mechanically, LINC00460 is induced by hypoxia, through binding of the hypoxia-inducible factor 1-α in the promoter region of LINC00460. Furthermore, LINC00460 functioned as an miR-4689 sponge to regulate the downstream target gene UBE2V1, enhancing the stability of mutant p53 in pancreatic cancer cells. LINC00460 also further promotes pancreatic cancer development by sequestering USP10, a cytoplasmic ubiquitin-specific protease that deubiquitinates p53 and enhances its stability. Collectively, our study demonstrated that LINC00460 is a hypoxia-induced lncRNA that plays the role of oncogene in pancreatic cancer by modulating the miR-4689/UBE2V1 axis, sequestering USP10, and ultimately enhancing the stability of mutant p53.
Assuntos
MicroRNAs , Neoplasias Pancreáticas , RNA Longo não Codificante , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Linhagem Celular Tumoral , Neoplasias Pancreáticas/genética , Proliferação de Células/genética , Hipóxia , Regulação Neoplásica da Expressão Gênica , Movimento Celular/genética , Fatores de Transcrição/genética , Enzimas de Conjugação de Ubiquitina/genética , Ubiquitina Tiolesterase/genética , Ubiquitina Tiolesterase/metabolismoRESUMO
Ventricular arrhythmia is the most common cause of sudden cardiac death in patients with myocardial infarction (MI). Fibroblast growth factor 21 (FGF21) has been shown to play an important role in cardiovascular and metabolic diseases. However, the effects of FGF21 on ventricular arrhythmias following MI have not been addressed yet. The present study was conducted to investigate the pharmacological action of FGF21 on ventricular arrhythmias after MI. Adult male mice were administrated with or without recombinant human basic FGF21 (rhbFGF21), and the susceptibility to arrhythmias was assessed by programmed electrical stimulation and optical mapping techniques. Here, we found that rhbFGF21 administration reduced the occurrence of ventricular tachycardia (VT), improved epicardial conduction velocity and shorted action potential duration at 90% (APD90) in infarcted mouse hearts. Mechanistically, FGF21 may improve cardiac electrophysiological remodeling as characterized by the decrease of INa and IK1 current density in border zone of infarcted mouse hearts. Consistently, in vitro study also demonstrated that FGF21 may rescue oxidant stress-induced dysfunction of INa and IK1 currents in cultured ventricular myocytes. We further found that oxidant stress-induced down-regulation of early growth response protein 1 (EGR1) contributed to INa and IK1 reduction in post-infarcted hearts, and FGF21 may recruit EGR1 into the SCN5A and KCNJ2 promoter regions to up-regulate NaV1.5 and Kir2.1 expression at transcriptional level. Moreover, miR-143 was identified as upstream of EGR1 and mediated FGF21-induced EGR1 up-regulation in cardiomyocytes. Collectively, rhbFGF21 administration effectively suppressed ventricular arrhythmias in post-infarcted hearts by regulating miR-143-EGR1-NaV1.5/Kir2.1 axis, which provides novel therapeutic strategies for ischemic arrhythmias in clinics.
Assuntos
Antiarrítmicos/administração & dosagem , Proteína 1 de Resposta de Crescimento Precoce/metabolismo , Fatores de Crescimento de Fibroblastos/administração & dosagem , Sistema de Condução Cardíaco/efeitos dos fármacos , Frequência Cardíaca/efeitos dos fármacos , MicroRNAs/metabolismo , Infarto do Miocárdio/tratamento farmacológico , Miocárdio/metabolismo , Taquicardia Ventricular/prevenção & controle , Potenciais de Ação/efeitos dos fármacos , Animais , Células Cultivadas , Modelos Animais de Doenças , Proteína 1 de Resposta de Crescimento Precoce/genética , Sistema de Condução Cardíaco/metabolismo , Sistema de Condução Cardíaco/fisiopatologia , Masculino , Camundongos Endogâmicos C57BL , MicroRNAs/genética , Infarto do Miocárdio/genética , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/fisiopatologia , Miocárdio/patologia , Canal de Sódio Disparado por Voltagem NAV1.5/genética , Canal de Sódio Disparado por Voltagem NAV1.5/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização/genética , Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Proteínas Recombinantes/administração & dosagem , Transdução de Sinais , Taquicardia Ventricular/genética , Taquicardia Ventricular/metabolismo , Taquicardia Ventricular/fisiopatologiaRESUMO
BACKGROUND: Recreational water contaminated with fecal pollution poses a great public health concern, as fecal waste may cause serious waterborne illnesses. Current recreational water standards using fecal indicator bacteria (FIB) have their limitations for human protection especially in developing countries such as China. METHODS: To explore the potential use of enteric viruses as a potential indicator of fecal contamination, four viruses: norovirus geno-groups I and II, enteroviruses, and adenoviruses were tested in this study using molecular detection methods and sensitive RT-PC developed in the University of Hawaii. Water samples were also tested for FIB in order to determine their association with enteric virus detection. RESULTS: All sample sites tested positive for four enteric viruses. Human enterovirus (58%) and adenovirus (67%) were more frequently detected from these six sites, followed by norovirus I (50%) and norovirus II (38%). Six sampling sites all met the level-I water quality of GB3838-2002 criteria in microbiological level, but they all tested positive for enteric viruses. CONCLUSION: These findings indicate the current sewage contamination of Poyang Lake and also support the essential need of additional indicator such as human enteric viruses for enhanced monitoring of water quality since the presence of enteric viruses does not always correlate with fecal bacterial indicator detection.
Assuntos
Enterovirus/isolamento & purificação , Monitoramento Ambiental/métodos , Lagos/virologia , Esgotos/virologia , Poluição da Água , Qualidade da Água/normas , Adenoviridae/isolamento & purificação , China , Humanos , Norovirus/isolamento & purificação , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
BACKGROUND: A fructose high-salt (FHS) diet increases systolic blood pressure and Ang II (angiotensin II)-stimulated proximal tubule (PT) superoxide (O2-) production. These increases are prevented by scavenging O2- or an Ang II type 1 receptor antagonist. SGLT4 (sodium glucose-linked cotransporters 4) and SGLT5 are implicated in PT fructose reabsorption, but their roles in fructose-induced hypertension are unclear. We hypothesized that PT fructose reabsorption by SGLT5 initiates a genetic program enhancing Ang II-stimulated oxidative stress in males and females, thereby causing fructose-induced salt-sensitive hypertension. METHODS: We measured systolic blood pressure in male and female Sprague-Dawley (wild type [WT]), SGLT4 knockout (-/-), and SGLT5-/- rats. Then, we measured basal and Ang II-stimulated (37 nmol/L) O2- production by PTs and conducted gene coexpression network analysis. RESULTS: In male WT and female WT rats, FHS increased systolic blood pressure by 15±3 (n=7; P<0.0027) and 17±4 mmâ Hg (n=9; P<0.0037), respectively. Male and female SGLT4-/- had similar increases. Systolic blood pressure was unchanged by FHS in male and female SGLT5-/-. In male WT and female WT fed FHS, Ang II stimulated O2- production by 14±5 (n=6; P<0.0493) and 8±3 relative light units/µg protein/s (n=7; P<0.0218), respectively. The responses of SGTL4-/- were similar. Ang II did not stimulate O2- production in tubules from SGLT5-/-. Five gene coexpression modules were correlated with FHS. These correlations were completely blunted in SGLT5-/- and partially blunted by chronically scavenging O2- with tempol. CONCLUSIONS: SGLT5-mediated PT fructose reabsorption is required for FHS to augment Ang II-stimulated proximal nephron O2- production, and increases in PT oxidative stress likely contribute to FHS-induced hypertension.
Assuntos
Pressão Sanguínea , Frutose , Hipertensão , Túbulos Renais Proximais , Estresse Oxidativo , Proteínas de Transporte de Sódio-Glucose , Animais , Feminino , Masculino , Ratos , Angiotensina II , Pressão Sanguínea/efeitos dos fármacos , Pressão Sanguínea/fisiologia , Modelos Animais de Doenças , Frutose/efeitos adversos , Frutose/farmacologia , Hipertensão/metabolismo , Hipertensão/genética , Hipertensão/induzido quimicamente , Hipertensão/fisiopatologia , Túbulos Renais Proximais/metabolismo , Túbulos Renais Proximais/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Ratos Sprague-Dawley , Cloreto de Sódio na Dieta/efeitos adversos , Proteínas de Transporte de Sódio-Glucose/genética , Proteínas de Transporte de Sódio-Glucose/metabolismoRESUMO
Pancreatic cancer (PCa) is one of the most fatal human malignancies. The enhanced infiltration of stromal tissue into the PCa tumor microenvironment limits the identification of key tumor-specific transcription factors and epigenomic abnormalities in malignant epithelial cells. Integrated transcriptome and epigenetic multiomics analyses of the paired PCa organoids indicate that the basic helix-loop-helix transcription factor 40 (BHLHE40) is significantly upregulated in tumor samples. Increased chromatin accessibility at the promoter region and enhanced mTOR pathway activity contribute to the elevated expression of BHLHE40. Integrated analysis of chromatin immunoprecipitation-seq, RNA-seq, and high-throughput chromosome conformation capture data, together with chromosome conformation capture assays, indicate that BHLHE40 not only regulates sterol regulatory element-binding factor 1 (SREBF1) transcription as a classic transcription factor but also links the enhancer and promoter regions of SREBF1. It is found that the BHLHE40-SREBF1-stearoyl-CoA desaturase axis protects PCa cells from ferroptosis, resulting in the reduced accumulation of lipid peroxidation. Moreover, fatostatin, an SREBF1 inhibitor, significantly suppresses the growth of PCa tumors with high expressions of BHLHE40. This study highlights the important roles of BHLHE40-mediated lipid peroxidation in inducing ferroptosis in PCa cells and provides a novel mechanism underlying SREBF1 overexpression in PCa.
Assuntos
Ferroptose , Neoplasias Pancreáticas , Humanos , Proteínas de Homeodomínio/genética , Ferroptose/genética , Fatores de Transcrição/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Neoplasias Pancreáticas/genética , Microambiente Tumoral , Proteína de Ligação a Elemento Regulador de Esterol 1/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismoRESUMO
A novel photocatalytic system of Cu/TiO2 for activation the C-H bond in the dehydrogenation of ethane to ethylene at room temperature is proposed. The optimized 1%-Cu/TiO2 catalyst achieved C2H6 conversion of 1.70%, C2H4 selectivity of 98.41%, and exhibited excellent stability. The active site Cuδ+ showed high dispersion on the TiO2 surface. Theoretical calculations and in situ diffuse reflectance infrared Fourier transform spectroscopy revealed a reaction mechanism: C2H6 is first activated by adsorption over the Cu4C/TiO2 catalyst with elongation of the C-H bond, attacked by h+/ËOH to form ethyl radicals, which are then converted to C2H4.
RESUMO
Single-cell RNA sequencing (scRNAseq) is a crucial tool in kidney research. These technologies cluster cells according to transcriptome similarity, irrespective of the anatomical location and ordering within the nephron. Thus, a cluster transcriptome may obscure heterogeneity of the cell population within a nephron segment. Elevated dietary fructose leads to salt-sensitive hypertension, in part by fructose reabsorption in the proximal tubule (PT). However, organization of the four known fructose transporters in apical PTs (SGLT4, SGLT5, GLUT5 and NaGLT1) remains poorly understood. We hypothesized that cells within each subsegment of the proximal tubule exhibit complex, heterogenous fructose transporter expression patterns. To test this hypothesis we analyzed rat and kidney transcriptomes and proteomes from publicly available scRNAseq and tubule microdissection databases. We found that microdissected PT-S1 segments consist of 81±12% cells with scRNAseq-derived transcriptional characteristics of S1, whereas PT-S2 express a mixture of 18±9% S1, 58±8% S2, and 19±5% S3 transcripts, and PT-S3 consists of 75±9% S3 transcripts. The expression of all four fructose transporters was detectable in all three PT segments, but key fructose transporters SGLT5 and GLUT5 progressively increased from S1 to S3, and both were significantly upregulated in S3 vs. S1/S2 (Slc5a10: 1.9 log 2 FC, p<1×10 -299 ; Scl2a5: 1.4 log 2 FC, p<4×10 -105 ). A similar distribution was found in human kidneys. These data suggest that S3 is the primary site of fructose reabsorption in both humans and rats. Finally, because of the multiple scRNAseq transcriptional phenotypes found in each segment our findings also imply that anatomic labels applied to scRNAseq clusters may be misleading.
RESUMO
The extensively activated Notch signaling pathway in pancreatic cancer cells is important in carcinogenesis, chemoresistance, and recurrence. Targeting this pathway is a promising therapeutic strategy for pancreatic cancer; however, few successful approaches have been reported, and currently used molecular inhibitors of this pathway exhibit limited clinical benefits. In this study, we identified a previously uncharacterized microprotein, Notch1 degradation-associated regulatory polypeptide (N1DARP), encoded by LINC00261. N1DARP knockout accelerated tumor progression and enhanced stem cell properties in pancreatic cancer organoids and LSL-Kras, LSL-Trp53, and Pdx1-Cre (KPC) mice. Mechanistically, N1DARP suppressed canonical and non-canonical Notch1 pathways by competitively disrupting the interaction between N1ICD and ubiquitin-specific peptidase 10 (USP10), thereby promoting K11- and K48-linked polyubiquitination of N1ICD. To evaluate the therapeutic potential of N1DARP, we designed a cell-penetrating stapled peptide, SAH-mAH2-5, with a helical structure similar to that of N1DARP that confers remarkable physicochemical stability. SAH-mAH2-5 interacted with and promoted the proteasome-mediated degradation of N1ICD. SAH-mAH2-5 injection provided substantial therapeutic benefits with limited off-target and systemic adverse effects in Notch1-activated pancreatic cancer models. Taken together, these findings confirm that N1DARP acts as a tumor suppressor and chemosensitizer by regulating USP10-Notch1 oncogenic signaling, and suggest a promising therapeutic strategy targeting the N1DARP-N1ICD interaction in Notch1-activated pancreatic cancer.
RESUMO
Proximal tubule fructose metabolism is key to fructose-induced hypertension, but the roles of sex and stress are unclear. We hypothesized that females are resistant to the salt-sensitive hypertension caused by low amounts of dietary fructose compared to males and that the magnitude of the increase in blood pressure (BP) depends, in part, on amplification of the stress response of renal sympathetic nerves. We measured systolic BP (SBP) in rats fed high salt with either no sugar (HS), 20% glucose (GHS) or 20% fructose (FHS) in the drinking water for 7-8 days. FHS increased SBP in both males (Δ22 ± 9 mmHg; p < 0.046) and females (Δ16 ± 3 mmHg; p < 0.0007), while neither GHS nor HS alone induced changes in SBP in either sex. The FHS-induced increase in SBP as measured by telemetry in the absence of added stress (8 ± 2 mmHg) was significantly lower than that measured by plethysmography (24 ± 5 mmHg) (p < 0.014). However, when BP was measured by telemetry simulating the stress of plethysmography, the increase in SBP was significantly greater (15 ± 3 mmHg) than under low stress (8 ± 1 mmHg) (p < 0.014). Moderate-stress also increased telemetric diastolic (p < 0.006) and mean BP (p < 0.006) compared to low-stress in FHS-fed animals. Norepinephrine excretion was greater in FHS-fed rats than HS-fed animals (Male: 6.4 ± 1.7 vs.1.8 ± 0.4 nmole/kg/day; p < 0.02. Female 54 ± 18 vs. 1.2 ± 0.6; p < 0.02). We conclude that fructose-induced salt-sensitive hypertension is similar in males and females unlike other forms of hypertension, and the increase in blood pressure depends in part on an augmented response of the sympathetic nervous system to stress.
Assuntos
Água Potável , Hipertensão , Animais , Pressão Sanguínea/fisiologia , Feminino , Frutose/efeitos adversos , Glucose/farmacologia , Hipertensão/induzido quimicamente , Hipertensão/metabolismo , Masculino , Norepinefrina/farmacologia , Ratos , Ratos Sprague-Dawley , Cloreto de Sódio/farmacologia , Cloreto de Sódio na Dieta/efeitos adversosRESUMO
Infarcted myocardium is predisposed to cause lethal ventricular arrhythmias that remain the main cause of death in patients suffering myocardial ischemia. Liver-derived fibroblast growth factor 21 (FGF21) is an endocrine regulator, which exerts metabolic actions by favoring glucose and lipids metabolism. Emerging evidence has shown a beneficial effect of FGF21 on cardiovascular diseases, but the role of FGF21 on ventricular arrhythmias following myocardial infarction (MI) in humans has never been addressed. This study was conducted to investigate the pharmacological effects of FGF21 on cardiomyocytes after MI in humans. Patients with arrhythmia in acute MI and healthy volunteers were enrolled in this study. Serum samples were collected from these subjects on day 1 and days 7-10 after the onset of MI for measuring FGF21 levels using ELISA. Here, we found that the serum level of FGF21 was significantly increased on day 1 after the onset of MI and it returned to normal on days 7-10, relative to the Control samples. In order to clarify the regulation of FGF21 on arrhythmia, two kinds of arrhythmia animal models were established in this study, including ischemic arrhythmia model (MI rat model) and nonischemic arrhythmia model (ouabain-induced guinea pig arrhythmia model). The results showed that the incidence and duration time of ischemic arrhythmias in rhbFGF21-treated MI rats were significantly reduced at different time point after MI compared with normal saline-treated MI rats. Moreover, the onset of the first ventricular arrhythmias was delayed and the numbers of VF and maintenance were attenuated by FGF21 compared to the rhbFGF21-untreated group in the ouabain model. Consistently, in vitro study also demonstrated that FGF21 administration was able to shorten action potential duration (APD) in hydrogen peroxide-treated AC16 cells. Mechanically, FGF21 can ameliorate the electrophysiological function of AC16 cells, which is characterized by rescuing the expression and dysfunction of cardiac sodium current (I Na) and inward rectifier potassium (I k1) in AC16 cells induced by hydrogen peroxide. Moreover, the restorative effect of FGF21 on NaV1.5 and Kir2.1 was eliminated when FGF receptors were inhibited. Collectively, FGF21 has the potential role of ameliorating transmembrane ion channels remodeling through the NaV1.5/Kir2.1 pathway by FGF receptors and thus reducing life-threatening postinfarcted arrhythmias, which provides new strategies for antiarrhythmic therapy in clinics.