Overcoming the influence of interfering substances in the environment and achieving superior sensing performance are significant challenges in biomarker detection within complex matrices. Herein, an integrated electrochemical sensing platform for sensitive detection of biomarkers in complex biofluids was developed based on a newly designed PEGylated multifunctional peptide (PEG-MPEP). The designed PEG-MPEP contains a poly(serine) sequence (-ssssss-) as the antifouling part and recognition peptide sequence (-avwgrwh) specific for the target human immunoglobulin G (IgG). To improve the peptide stability to protease hydrolysis, d-amino acids were adopted to synthesize the whole peptide. Additionally, the PEGylation can further enhance the stability of the peptide, and the PEG itself was also antifouling, ensuring superstrong antifouling capability of the PEG-MPEP. The designed PEG-MPEP-based biosensor possessed a high sensitivity for the detection of IgG in the range of 1.0 pg mL-1 to 1.0 µg mL-1, with a low limit of detection (0.41 pg mL-1), and it was capable of assaying targets accurately in real serum samples. Compared with conventional peptide-modified biosensors, the PEG-MPEP-modified biosensor exhibited superior antifouling and antihydrolysis properties in complex biofluid, showcasing promising potential for practical assay applications.
Obesity is one of the most common metabolic diseases around the world, which is distinguished by the abnormal buildup of triglycerides within adipose cells. Recent research has revealed that autophagy regulates lipid mobilization to maintain energy balance. TIGAR (Trp53 induced glycolysis regulatory phosphatase) has been identified as a glycolysis inhibitor, whether it plays a role in the metabolism of lipids is unknown. Here, we found that TIGAR transgenic (TIGAR+/+) mice exhibited increased fat mass and trended to obesity phenotype. Non-target metabolomics showed that TIGAR caused the dysregulation of the metabolism profile. The quantitative transcriptome sequencing identified an increased levels of LRRK2 and RAB7B in the adipose tissue of TIGAR+/+ mice. It was confirmed in vitro that TIGAR overexpression increased the levels of LRRK2 by inhibiting polyubiquitination degradation, thereby suppressing macroautophagy and chaperone-mediated autophagy (CMA) while increasing lipid accumulation which were reversed by the LRRK2 inhibitor DNL201. Furthermore, TIGAR drove LRRK2 to interact with RAB7B for suppressing lysosomal degradation of lipid droplets, while the increased lipid droplets in adipocytes were blocked by the RAB7B inhibitor ML282. Additionally, fat-specific TIGAR knockdown of TIGAR+/+ mice alleviated the symptoms of obesity, and adipose tissues-targeting superiority DNL201 nano-emulsion counteracted the obesity phenotype in TIGAR+/+ mice. In summary, the current results indicated that TIGAR performed a vital function in the lipid metabolism through LRRK2-mediated negative regulation of macroautophagy and CMA in adipocyte. The findings suggest that TIGAR has the potential to serve as a viable therapeutic target for treating obesity and its associated metabolic dysfunction.
Spinal cord injury (SCI) triggers a complex cascade of events, including myelin loss, neuronal damage, neuroinflammation, and the accumulation of damaged cells and debris at the injury site. Infiltrating bone marrow derived macrophages (BMDMÏ) migrate to the epicenter of the SCI lesion, where they engulf cell debris including abundant myelin debris to become pro-inflammatory foamy macrophages (foamy MÏ), participate neuroinflammation, and facilitate the progression of SCI. This study aimed to elucidate the cellular and molecular mechanisms underlying the functional changes in foamy MÏ and their potential implications for SCI. Contusion at T10 level of the spinal cord was induced using a New York University (NYU) impactor (5 g rod from a height of 6.25 mm) in male mice. ABCA1, an ATP-binding cassette transporter expressed by MÏ, plays a crucial role in lipid efflux from foamy cells. We observed that foamy MÏ lacking ABCA1 exhibited increased lipid accumulation and a higher presence of lipid-accumulated foamy MÏ as well as elevated pro-inflammatory response in vitro and in injured spinal cord. We also found that both genetic and pharmacological enhancement of ABCA1 expression accelerated lipid efflux from foamy MÏ, reduced lipid accumulation and inhibited the pro-inflammatory response of foamy MÏ, and accelerated clearance of cell debris and necrotic cells, which resulted in functional recovery. Our study highlights the importance of understanding the pathologic role of foamy MÏ in SCI progression and the potential of ABCA1 as a therapeutic target for modulating the inflammatory response, promoting lipid metabolism, and facilitating functional recovery in SCI.
Cardiovascular disease (CVD) is the major cause of death in chronic kidney disease (CKD) and is associated with high circulating fibroblast growth factor (FGF)23 levels. It is unresolved whether high circulating FGF23 is a mere biomarker or pathogenically contributes to cardiomyopathy. It is also unknown whether the C-terminal FGF23 peptide (cFGF23), a natural FGF23 antagonist proteolyzed from intact FGF23 (iFGF23), retards CKD progression and improves cardiomyopathy. We addressed these questions in three murine models with high endogenous FGF23 and cardiomyopathy. First, we examined wild-type (WT) mice with CKD induced by unilateral ischemia-reperfusion and contralateral nephrectomy followed by a high-phosphate diet. These mice were continuously treated with intraperitoneal implanted osmotic minipumps containing either iFGF23 protein to further escalate FGF23 bioactivity, cFGF23 peptide to block FGF23 signaling, vehicle, or scrambled peptide as negative controls. Exogenous iFGF23 protein given to CKD mice exacerbated pathological cardiac remodeling and CKD progression, whereas cFGF23 treatment improved heart and kidney function, attenuated fibrosis, and increased circulating soluble Klotho. WT mice without renal insult placed on a high-phosphate diet and homozygous Klotho hypomorphic mice, both of whom develop moderate CKD and clear cardiomyopathy, were treated with cFGF23 or vehicle. Mice treated with cFGF23 in both models had improved heart and kidney function and histopathology. Taken together, these data indicate high endogenous iFGF23 is not just a mere biomarker but pathogenically deleterious in CKD and cardiomyopathy. Furthermore, attenuation of FGF23 bioactivity by cFGF23 peptide is a promising therapeutic strategy to protect the kidney and heart from high FGF23 activity.NEW & NOTEWORTHY There is a strong correlation between cardiovascular morbidity and high circulating fibroblast growth factor 23 (FGF23) levels, but causality was never proven. We used a murine chronic kidney disease (CKD) model to show that intact FGF23 (iFGF23) is pathogenic and contributes to both CKD progression and cardiomyopathy. Blockade of FGF23 signaling with a natural proteolytic product of iFGF23, C-terminal FGF23, alleviated kidney and cardiac histology, and function in three separate murine models of high endogenous FGF23.
Cardiomyopathies , Renal Insufficiency, Chronic , Animals , Mice , Fibroblast Growth Factor-23 , Disease Models, Animal , Renal Insufficiency, Chronic/metabolism , Fibroblast Growth Factors/pharmacology , Fibroblast Growth Factors/metabolism , Biomarkers , Phosphates , Cardiomyopathies/drug therapy , Cardiomyopathies/complications
Biomolecular condensates have been proposed to mediate cellular signaling transduction. However, the mechanism and functional consequences of signal condensates are not well understood. Here we report that LATS2, the core kinase of the Hippo pathway, responds to F-actin cytoskeleton reduction and forms condensates. The proline-rich motif (PRM) of LATS2 mediates its condensation. LATS2 partitions with the main components of the Hippo pathway to assemble a signalosome for LATS2 activation and for its stability by physically compartmentalizing from E3 ligase FBXL16 complex-dependent degradation, which in turn mediates yes-associated protein (YAP)-transcriptional coactivator with PDZ-binding motif (TAZ) recruitment and inactivation. This oncogenic FBXL16 complex blocks LATS2 condensation by binding to the PRM region to promote its degradation. Disruption of LATS2 condensation leads to tumor progression. Thus, our study uncovers that the signalosomes assembled by LATS2 condensation provide a compartmentalized and reversible platform for Hippo signaling transduction and protein stability, which have potential implications in cancer diagnosis and therapeutics.
PURPOSE: Bladder neck contracture (BNC) is a rare but intolerant complication after transurethral surgery of prostate. The present study aims to investigate the incidence and risk factors of BNC in patients diagnosed benign prostate hyperplasia (BPH) and following transurethral resection or enucleation of the prostate (TURP/TUEP). METHODS: This retrospective study included 1008 BPH individuals who underwent transurethral surgery of the prostate between January 2017 and January 2022. Patients' demographics, medical comorbidities, urologic characteristics, perioperative parameters, and the presence of BNC were documented. Univariate and multivariate analyses were conducted to identify the risk factors. RESULTS: A total of 2% (20/1008) BPH patients developed BNC postoperatively and the median occurring time was 5.8 months. Particularly, the incidences of BNC were 4.7% and 1.3% in patients underwent Bipolar-TURP and TUEP respectively. Preoperative urinary tract infection (UTI), elevated PSA, smaller prostate volume (PV), bladder diverticulum (BD), and B-TURP were significantly associated with BNC in the univariate analysis. Further multivariate logistic regression demonstrated preoperative UTI (OR 4.04, 95% CI 2.25 to 17.42, p < 0.001), BD (OR 7.40, 95% CI 1.83 to 31.66, p < 0.001), and B-TURP (OR 3.97, 95% CI 1.55 to 10.18, p = 0.004) as independent risk factors. All BNC patients were treated with transurethral incision of the bladder neck (TUIBN) combined with local multisite injection of betamethasone. During a median follow-up of 35.8 months, 35% (7/20) of BNC patients recurred at a median time of 1.8 months. CONCLUSION: BNC was a low-frequency complication following transurethral surgery of prostate. Preoperative UTI, BD, and B-TURP were likely independent risk factors of BNC. TUIBN combined with local multisite injection of betamethasone may be promising choice for BNC treatment.
Contracture , Prostatic Hyperplasia , Male , Humans , Urinary Bladder , Prostate , Retrospective Studies , Prostatic Hyperplasia/surgery , Contracture/epidemiology , Contracture/etiology , Betamethasone
The activation of YAP/TAZ, a pair of paralogs of transcriptional coactivators, initiates a dysregulated transcription program, which is a key feature of human cancer cells. However, it is not fully understood how YAP/TAZ promote dysregulated transcription for tumor progression. In this study, we employed the BioID method to identify the interactome of YAP/TAZ and discovered that YAP/TAZ interact with multiple components of SRCAP complex, a finding that was further validated through endogenous and exogenous co-immunoprecipitation, as well as immunofluorescence experiments. CUT&Tag analysis revealed that SRCAP complex facilitates the deposition of histone variant H2A.Z at target promoters. The depletion of SRCAP complex resulted in a decrease in H2A.Z occupancy and the oncogenic transcription of YAP/TAZ target genes. Additionally, the blockade of SRCAP complex suppressed YAP-driven tumor growth. In a genetically engineered lung adenocarcinoma mouse model and non-small cell lung cancer patients, SRCAP complex and H2A.Z deposition were found to be upregulated. This upregulation was statistically correlated with YAP expression, pathological stages, and poor survival in lung cancer patients. Together, our study uncovers that SRCAP complex plays a critical role in YAP/TAZ oncogenic transcription by coordinating H2A.Z deposition during cancer progression, providing potential targets for cancer diagnosis and prevention.
Carcinoma, Non-Small-Cell Lung , Lung Neoplasms , Animals , Mice , Humans , Lung Neoplasms/metabolism , Adaptor Proteins, Signal Transducing/genetics , Adaptor Proteins, Signal Transducing/metabolism , Signal Transduction/genetics , Protein Serine-Threonine Kinases/genetics , Protein Serine-Threonine Kinases/metabolism , YAP-Signaling Proteins , Histones/metabolism , Phosphoproteins/genetics , Phosphoproteins/metabolism , Adenosine Triphosphatases/metabolism
Background: The objective of this study was to explore the perioperative outcomes of single-plane posterior retroperitoneoscopic adrenalectomy (SPRA) guided by indocyanine green dye (ICG) fluorescence imaging. Methods: A retrospective analysis of patients who underwent SPRA from April to September 2023 in our center was conducted. Patients were divided into the ICG group and the non-ICG group, based on whether they received intraoperative ICG fluorescence guided or not. Baseline and perioperative data were recorded and analyzed by R software (R 4.3.1). Results: A total of 23 patients were enrolled in the study, with 12 in the ICG group and 11 in the non-ICG group. The demographics including age, gender, body mass index, or American Society of Anesthesiologists classification showed no significant differences between groups. There were obvious advantages in shortening adrenal gland localization time and total operative time, as well as reducing estimated blood loss in the ICG group compared with the non-ICG group (5.58 ± 0.36 minutes vs 7.55 ± 0.62 minutes, p < 0.001; 27.50 ± 5.46 minutes vs 45.00 ± 10.99 minutes, p < 0.001; 22.91 ± 7.57 mL vs 54.54 ± 18.90 mL, p < 0.001; respectively). Furthermore, patients in the ICG group exhibited significantly lower visual analog pain scale scores at 24 hours postoperatively and at discharge (p = 0.001 and p = 0.006, respectively). The oral intake intervals, hospital stays, and perioperative complications were comparable between groups. Conclusions: ICG-guided SPRA could be a safe and effective procedure for patients with adrenal tumors. This technique improves the accuracy and efficacy of adrenal gland localization and has shown benefits in perioperative outcomes. The use of ICG fluorescence guidance represents a promising clinical application.
Adrenal Gland Neoplasms , Laparoscopy , Humans , Indocyanine Green , Adrenalectomy/methods , Retrospective Studies , Adrenal Gland Neoplasms/surgery , Length of Stay , Laparoscopy/methods
BACKGROUND: Molecular understanding of muscle-invasive (MIBC) and non-muscle-invasive (NMIBC) bladder cancer is currently based primarily on transcriptomic and genomic analyses. OBJECTIVE: To conduct proteogenomic analyses to gain insights into bladder cancer (BC) heterogeneity and identify underlying processes specific to tumor subgroups and therapeutic outcomes. DESIGN, SETTING, AND PARTICIPANTS: Proteomic data were obtained for 40 MIBC and 23 NMIBC cases for which transcriptomic and genomic data were already available. Four BC-derived cell lines harboring FGFR3 alterations were tested with interventions. INTERVENTION: Recombinant tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), second mitochondrial-derived activator of caspases mimetic (birinapant), pan-FGFR inhibitor (erdafitinib), and FGFR3 knockdown. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Proteomic groups from unsupervised analyses (uPGs) were characterized using clinicopathological, proteomic, genomic, transcriptomic, and pathway enrichment analyses. Additional enrichment analyses were performed for FGFR3-mutated tumors. Treatment effects on cell viability for FGFR3-altered cell lines were evaluated. Synergistic treatment effects were evaluated using the zero interaction potency model. RESULTS AND LIMITATIONS: Five uPGs, covering both NMIBC and MIBC, were identified and bore coarse-grained similarity to transcriptomic subtypes underlying common features of these different entities; uPG-E was associated with the Ta pathway and enriched in FGFR3 mutations. Our analyses also highlighted enrichment of proteins involved in apoptosis in FGFR3-mutated tumors, not captured through transcriptomics. Genetic and pharmacological inhibition demonstrated that FGFR3 activation regulates TRAIL receptor expression and sensitizes cells to TRAIL-mediated apoptosis, further increased by combination with birinapant. CONCLUSIONS: This proteogenomic study provides a comprehensive resource for investigating NMIBC and MIBC heterogeneity and highlights the potential of TRAIL-induced apoptosis as a treatment option for FGFR3-mutated bladder tumors, warranting a clinical investigation. PATIENT SUMMARY: We integrated proteomics, genomics, and transcriptomics to refine molecular classification of bladder cancer, which, combined with clinical and pathological classification, should lead to more appropriate management of patients. Moreover, we identified new biological processes altered in FGFR3-mutated tumors and showed that inducing apoptosis represents a new potential therapeutic option.
Non-Muscle Invasive Bladder Neoplasms , Proteogenomics , Urinary Bladder Neoplasms , Humans , Proteomics , Ligands , Urinary Bladder Neoplasms/drug therapy , Urinary Bladder Neoplasms/genetics , Urinary Bladder Neoplasms/pathology , Apoptosis , Tumor Necrosis Factor-alpha , TNF-Related Apoptosis-Inducing Ligand/genetics , TNF-Related Apoptosis-Inducing Ligand/pharmacology , Receptor, Fibroblast Growth Factor, Type 3/genetics
Developing a generalized strategy for the nonfouling detection of biomarkers in diverse biological fluids presents a significant challenge. Herein, a polyhydroxyproline helical peptide (PHHP) was designed and adopted to fabricate electrochemical microsensors capable of detecting targets in various biological media. The PHHP possessed unique properties such as strong hydrophilicity, rigid structure, and lack of ionizable side-chain groups. Compared with common zwitterionic peptides (ZIPs), the PHHP exhibited similar antifouling capability but exceptional stability, allowing its antifouling performance to be unaffected by environmental alteration. The PHHP can prevent biofouling even in fluctuating pH conditions, high ionic strength environments, and the presence of high-valence ions and resist the protease hydrolysis. The PHHP-modified carbon fiber microelectrode was further immobilized with an aptamer to construct an antifouling microsensor for cortisol detection across diverse biofluids, and the microsensor exhibited acceptable accuracy and higher sensitivity than the ELISA method. In addition, different biological samples of mice were collected in situ using a microsensing device, and cortisol levels were analyzed in each specifically tailored region. This nonfouling sensing strategy based on PHHP allows a comprehensive assessment of biomarkers in both spatial and temporal dimensions in diverse biological environments, holding promising potential for early disease diagnosis and real-time health monitoring.
Aptamers, Nucleotide , Biofouling , Biosensing Techniques , Animals , Mice , Biofouling/prevention & control , Hydrocortisone , Biosensing Techniques/methods , Peptides/chemistry , Aptamers, Nucleotide/chemistry , Electrochemical Techniques/methods , Biomarkers
Impaired wound healing and ulcer complications are major causes of morbidity in patients with diabetes. Impaired wound healing is associated with increased inflammation and poor angiogenesis in diabetes patients. Here, we demonstrate that topical administration of a secreted recombinant protein (Meteorin-like [Metrnl]) accelerates wound epithelialization and angiogenesis in mice. We observed a significant increase in Metrnl expression during physiological wound healing; however, its expression remained low during diabetic wound healing. Functionally, the recombinant protein Metrnl significantly accelerated wound closure in normal and diabetic mice models including db/db, high-fat diet/streptozotocin (HFD/STZ), and STZ mice. Mechanistically, keratinocytes secrete quantities of Metrnl to promote angiogenesis; increase endothelial cell proliferation, migration, and tube formation; and enhance macrophage polarization to the M2 type. Meanwhile, M2 macrophages secrete Metrnl to further stimulate angiogenesis. Moreover, the keratinocyte- and macrophage-produced cytokine Metrnl drives postinjury angiogenesis and reepithelialization through activation of AKT phosphorylation (S473) in a KIT receptor tyrosine kinase (c-Kit)-dependent manner. In conclusion, our study suggests that Metrnl has a biological effect in accelerating wound closure through c-Kit-dependent angiogenesis and epithelialization.
Objective: Increasing evidence suggests that gut microbiota is involved in the occurrence and progression of urinary system diseases such as clear cell renal cell carcinoma (ccRCC). However, the mechanism of how alteration of gut metagenome promotes ccRCC remains unclear. Here we aim to elucidate the association of specific gut bacteria and their metabolites with ccRCC. Methods: In a pilot case-control study among 30 ccRCC patients (RCC group) and 30 healthy controls (Control group), 16S ribosomal RNA (rRNA) gene sequencing were analyzed from fecal samples collected prior to surgery or hospitalization. Alpha diversity and beta diversity analysis of the gut microbiota were performed, and differential taxa were identified by multivariate statistics. Meanwhile, serum metabolism was measured by UHPLC-MS, and differential genes were identified based on the TCGA database. Results: Alpha diversity found there were no significant microbial diversity differences of gut microbiota between the RCC group and the Control group. However, beta diversity analysis showed that the overall structures of the two groups were significantly separated (p = 0.008). Random Forests revealed the relative abundances of 20 species differed significantly between the RCC group and the Control group, among which nine species were enriched in the RCC group such as Desulfovibrionaceae, and 11 species were less abundant such as four kinds of Lactobacillus. Concomitantly, serum level of taurine, which was considered to be consumed by Desulfovibrionaceae and released by Lactobacillus, has decreased in the RCC group. In addition, macrophage-related genes such as Gabbr1 was upregulated in ccRCC patients. Conclusion: Reduction of protective bacteria, proliferation of sulfide-degrading bacteria Desulfovibrionaceae, reduction of taurine, and enrichment of macrophage related genes might be the risk predictors of ccRCC.
Impaired protein N-glycosylation leads to the endoplasmic reticulum (ER) stress, which triggers adaptive survival or maladaptive apoptosis in renal tubules in diabetic kidney disease (DKD). Therapeutic strategies targeting ER stress are promising for the treatment of DKD. Here, we report a previously unappreciated role played by ENTPD5 in alleviating renal injury by mediating ER stress. We found that ENTPD5 was highly expressed in normal renal tubules; however, ENTPD5 was dynamically expressed in the kidney and closely related to pathological DKD progression in both human patients and mouse models. Overexpression of ENTPD5 relieved ER stress in renal tubular cells, leading to compensatory cell proliferation that resulted in hypertrophy, while ENTPD5 knockdown aggravated ER stress to induce cell apoptosis, leading to renal tubular atrophy and interstitial fibrosis. Mechanistically, ENTPD5-regulated N-glycosylation of proteins in the ER to promote cell proliferation in the early stage of DKD, and continuous hyperglycemia activated the hexosamine biosynthesis pathway (HBP) to increase the level of UDP-GlcNAc, which driving a feedback mechanism that inhibited transcription factor SP1 activity to downregulate ENTPD5 expression in the late stage of DKD. This study was the first to demonstrate that ENTPD5 regulated renal tubule cell numbers through adaptive proliferation or apoptosis in the kidney by modulating the protein N-glycosylation rate in the ER, suggesting that ENTPD5 drives cell fate in response to metabolic stress and is a potential therapeutic target for renal diseases.
Endoplasmic Reticulum Stress , Kidney Tubules , Kidney , Animals , Humans , Mice , Glycosylation , Oncogene Proteins , Pyrophosphatases
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.
Diabetic Nephropathies , MicroRNAs , Mice , Animals , MicroRNAs/metabolism , Epithelial Cells/metabolism , Diabetic Nephropathies/metabolism , Mice, Inbred Strains , Glucose/metabolism , Mitochondria/metabolism , Lipids/pharmacology
Ectopic lipid accumulation in renal tubules is closely related to the pathogenesis of diabetic kidney disease (DKD), and mitochondrial dysfunction is thought to play a key role in lipid accumulation. Therefore, maintaining mitochondrial homeostasis holds considerable promise as a therapeutic strategy for the treatment of DKD. Here, we report that the Meteorin-like (Metrnl) gene product mediates lipid accumulation in the kidney and has therapeutic potential for DKD. We confirmed the reduced expression of Metrnl in renal tubules, which was inversely correlated with DKD pathological changes in human patients and mouse models. Functionally, pharmacological administration of recombinant Metrnl (rMetrnl) or Metrnl overexpression could alleviate lipid accumulation and inhibit kidney failure. In vitro, rMetrnl or Metrnl overexpression attenuated palmitic acid-induced mitochondrial dysfunction and lipid accumulation in renal tubules accompanied by maintained mitochondrial homeostasis and enhanced lipid consumption. Conversely, shRNA-mediated Metrnl knockdown diminished the protective effect on the kidney. Mechanistically, these beneficial effects of Metrnl were mediated by the Sirt3-AMPK signaling axis to maintain mitochondrial homeostasis and through Sirt3-uncoupling protein-1 to promote thermogenesis, consequently alleviating lipid accumulation. In conclusion, our study demonstrates that Metrnl regulated lipid metabolism in the kidney by modulating mitochondrial function and is a stress-responsive regulator of kidney pathophysiology, which sheds light on novel strategies for treating DKD and associated kidney diseases. ARTICLE HIGHLIGHTS: Metrnl is expressed in renal tubules and is reduced under diabetic conditions. The concentration of Metrnl in the kidney is correlated with lipid accumulation and serum creatinine. Metrnl-specific overexpression in the kidney or recombinant Metrnl administration alleviates renal injuries in diabetic mice. Metrnl regulates renal tubules lipid metabolism through Sirt3-AMPK/UCP1 signaling axis-mediated mitochondrial homeostasis.
Diabetes Mellitus, Experimental , Diabetic Nephropathies , Sirtuin 3 , Humans , Mice , Animals , Diabetic Nephropathies/metabolism , Diabetes Mellitus, Experimental/metabolism , Sirtuin 3/genetics , AMP-Activated Protein Kinases/metabolism , Mitochondria/metabolism , Lipids , Homeostasis
BACKGROUND: Bladder cancer (BCa) is more common in men and presents differences in molecular subtypes based on sex. Fibroblast growth factor receptor 3 (FGFR3) mutations are enriched in the luminal papillary muscle-invasive BCa (MIBC) and non-MIBC subtypes. OBJECTIVE: To determine whether FGFR3 mutations initiate BCa and impact BCa male sex bias. DESIGN, SETTING, AND PARTICIPANTS: We developed a transgenic mouse model expressing the most frequent FGFR3 mutation, FGFR3-S249C, in urothelial cells. Bladder tumorigenesis was monitored in transgenic mice, with and without carcinogen exposure. Mouse and human BCa transcriptomic data were compared. INTERVENTION: Mutant FGFR3 overexpression in mouse urothelium and siRNA knockdown in cell lines, and N-butyl-N(4-hydroxybutyl)-nitrosamine (BBN) exposure. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Impact of transgene dosage on tumor frequency, synergy with BBN treatment, and FGFR3 pathway activation were analyzed. The sex-specific incidence of FGFR3-mutated tumors was evaluated in mice and humans. FGFR3 expression in FGFR3-S249C mouse urothelium and in various human epithelia was measured. Mutant FGFR3 regulation of androgen (AR) and estrogen (ESR1) receptor activity was evaluated, through target gene expression (regulon) and reporter assays. RESULTS AND LIMITATIONS: FGFR3-S249C expression in mice induced low-grade papillary BCa resembling human luminal counterpart at histological, genomic, and transcriptomic levels, and promoted BBN-induced basal BCa formation. Mutant FGFR3 expression levels impacted tumor incidence in mice, and mutant FGFR3-driven human tumors were restricted to epithelia presenting high normal FGFR3 expression levels. BCa male sex bias, also found in our model, was even higher in human FGFR3-mutated tumors compared with wild-type tumors and was associated with higher AR and lower ESR1 regulon activity. Mutant FGFR3 expression inhibited both ESR1 and AR activity in mouse tumors and human cell lines, demonstrating causation only between FGFR3 activation and low ESR1 activity in tumors. CONCLUSIONS: Mutant FGFR3 initiates luminal papillary BCa formation and favors BCa male sex bias, potentially through FGFR3-dependent ESR1 downregulation. Patients with premalignant lesions or early-stage BCa could thus potentially benefit from FGFR3 targeting. FGFR3 expression level in epithelia could account for FGFR3-driven carcinoma tissue specificity. PATIENT SUMMARY: By developing a transgenic mouse model, we showed that gain-of-function mutations of FGFR3 receptor, among the most frequent genetic alterations in bladder cancer (BCa), initiate BCa formation. Our results could support noninvasive detection of FGFR3 mutations and FGFR3 targeting in early-stage bladder lesions.
Receptor, Fibroblast Growth Factor, Type 3 , Urinary Bladder Neoplasms , Female , Humans , Male , Mice , Animals , Receptor, Fibroblast Growth Factor, Type 3/genetics , Urinary Bladder/pathology , Sexism , Urinary Bladder Neoplasms/pathology , Mutation , Mice, Transgenic , Androgens/adverse effects
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 ).
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
Vascular endothelial growth factor (VEGF) and its cognate receptor (VEGFR2) system are crucial for cell functions associated with angiogenesis and vasculogenesis. Klotho contributes to vascular health maintenance in the kidney and other organs in mammals, but it is unknown whether renoprotection by Klotho is dependent on VEGF/VEGFR2 signaling. We used heterozygous VEGFR2-haploinsufficient (VEGFR2+/-) mice resulting from heterozygous knockin of green fluorescent protein in the locus of fetal liver kinase 1 encoding VEGFR2 to test the interplay of Klotho, phosphate, and VEGFR2 in kidney function, the vasculature, and fibrosis. VEGFR2+/- mice displayed downregulated VEGF/VEGFR2 signaling in the kidney, lower density of peritubular capillaries, and accelerated kidney fibrosis, all of which were also found in the homozygous Klotho hypomorphic mice. High dietary phosphate induced higher plasma phosphate, greater peritubular capillary rarefaction, and more kidney fibrosis in VEGFR2+/- mice compared with wild-type mice. Genetic overexpression of Klotho significantly attenuated the elevated plasma phosphate, kidney dysfunction, peritubular capillary rarefaction, and kidney fibrosis induced by a high-phosphate diet in wild-type mice but only modestly ameliorated these changes in the VEGFR2+/- background. In cultured endothelial cells, VEGFR2 inhibition reduced free VEGFR2 but enhanced its costaining of an endothelial marker (CD31) and exacerbated phosphotoxicity. Klotho protein maintained VEGFR2 expression and attenuated high phosphate-induced cell injury, which was reduced by VEGFR2 inhibition. In conclusion, normal VEGFR2 function is required for vascular integrity and for Klotho to exert vascular protective and antifibrotic actions in the kidney partially through the regulation of VEGFR2 function.NEW & NOTEWORTHY This research paper studied the interplay of vascular endothelial growth factor receptor type 2 (VEGFR2), high dietary phosphate, and Klotho, an antiaging protein, in peritubular structure and kidney fibrosis. Klotho protein was shown to maintain VEGFR2 expression in the kidney and reduce high phosphate-induced cell injury. However, Klotho cytoprotection was attenuated by VEGFR2 inhibition. Thus, normal VEGFR2 function is required for vascular integrity and Klotho to exert vascular protective and antifibrotic actions in the kidney.
Cytoprotection , Kidney Diseases , Kidney , Klotho Proteins , Microvascular Rarefaction , Vascular Endothelial Growth Factor Receptor-2 , Animals , Mice , Endothelial Cells/metabolism , Fibrosis , Kidney/blood supply , Kidney/pathology , Kidney Diseases/pathology , Microvascular Rarefaction/pathology , Phosphates/metabolism , Vascular Endothelial Growth Factor A/metabolism , Vascular Endothelial Growth Factor Receptor-2/deficiency , Klotho Proteins/genetics , Klotho Proteins/metabolism
Bladder cancer (BLCA) is a common malignancy of the urinary system. The gut microbiome produces various metabolites that play functional roles in tumorigenesis and tumor progression. However, the integrative analysis of the gut microbiome and metabolome in BLCA has still been lacking. Thus, the aim of this study was to identify microbial and functional characteristics and metabolites in BLCA in a Chinese population. Metagenomics, targeted metabolomics, bioinformatics, and integrative analysis were used in fecal samples of BLCA patients and healthy individuals. We found gut microbiomes were significantly dysregulated in BLCA patients, including Bifidobacterium, Lactobacillus, Streptococcus, Blautia, and Eubacterium. We also found 11Z-eicosenoic acid, 3-methoxytyrosine, abrine, aniline-2-sulfonate, arachidic acid, conjugated linoleic acids, elaidic acid, glycylleucine, glycylproline, leucyl-glycine, linoelaidic acid, linoleic acid, nicotinamide hypoxanthine dinucleotide, oleic acid, petroselinic acid, and ricinoleic acid to be significantly decreased, while cholesterol sulfate was significantly increased in BLCA patients. Integration of metagenomics and metabolomics revealed interactions between gut microbiota and metabolites and the host. We identified the alterations of gut microbiomes and metabolites in BLCA in a Chinese population. Moreover, we preliminarily revealed the associations between specific gut microbiomes and metabolites. These findings determined potential causative links among gut dysbiosis, dysregulated metabolites, and BLCA.
Metagenome , Urinary Bladder Neoplasms , Humans , Urinary Bladder Neoplasms/genetics , Metabolome , Metabolomics , China
BACKGROUND: Patients with urothelial bladder carcinoma (UBC) have a poor prognosis and a high risk of progression. Recently, tRNA-derived small RNAs (tsRNAs), a novel type of noncoding RNA, have been identified. In our previous study, we found tiRNA-Gly-GCC-1 was significantly upregulated in UBC tissue and might target the predicted target gene toll-like receptor 4 (TLR4) to play a regulatory role in UBC. Thus, the aim of this study was to identify the functional roles of tiRNA-Gly-GCC-1 and the relationship between tiRNA-Gly-GCC-1 and TLR4. METHODS: After lentiviral transfection in 5637 and T24 cell lines, quantitative reverse transcription-PCR, Cell Counting Kit-8, IncuCyte ZOOM™ live cell imaging, flow cytometry, Transwell assays, scratch assay, and luciferase assay were performed. RESULTS: The results showed down-regulation of tiRNA-Gly-GCC-1 inhibits cell proliferation, migration and invasion, promotes cell apoptosis, and affects the cell cycle. Besides, tiRNA-Gly-GCC-1 was found to inhibit TLR4 expression by directly targeting its 3'UTR. CONCLUSIONS: Our study demonstrated that tiRNA-Gly-GCC-1 promotes the progression of UBC and directly targets TLR4. This study provides novel insights for future investigations to explore the mechanisms and therapeutic targets for UBC.
