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BACKGROUND: Uremia-associated immunodeficiency, mainly characterized by T cell dysfunction, exists in patients on maintenance hemodialysis (MHD) and promotes systemic inflammation. However, T cell senescence, one of the causes of T cell dysfunction, has not been clearly revealed yet. In this cross-sectional research, we aimed to study the manifestation of T cell premature senescence in MHD patients and further investigate the associated clinical factors. METHODS: 76 MHD patients including 33 patients with cardiovascular diseases (CVD) and 28 patients with arteriovenous fistula (AVF) event history were enrolled in this study. Complementarity determining region 3 (CDR3) of T cell receptor (TCR) was analyzed by immune repertoire sequencing (IR-Seq). CD28- T cell subsets and expression of senescence marker p16 and p21 genes were detected by multicolor flow cytometry and RT-qPCR, respectively. RESULTS: MHD patients had significantly decreased TCR diversity (P < 0.001), increased CDR3 clone proliferation (P = 0.001) and a left-skewed CDR3 length distribution. The proportion of CD4 + CD28- T cells increased in MHD patients (P = 0.014) and showed a negative correlation with TCR diversity (P = 0.001). p16 but not p21 expression in T cells was up-regulated in MHD patients (P = 0.039). Patients with CVD exhibited increased expression of p16 and p21 genes (P = 0.010 and 0.004, respectively), and patients with AVF events showed further TCR diversity and evenness reduction (P = 0.002 and 0.017, respectively) compared to patients without the comorbidities. Moreover, age, average convection volume, total cholesterol, high-density lipoprotein cholesterol and transferrin saturation were associated with TCR diversity or CD4 + CD28- T cell proportion (P < 0.05). CONCLUSIONS: MHD patients undergo T cell premature senescence characterized by significant TCR diversity reduction and repertoire skew, as well as accumulation of the CD4 + CD28- subset and up-regulation of p16 gene. Patients with CVD or AVF events show higher level of immunosenescence. Furthermore, T cell senescence in MHD patients is associated with blood cholesterol and uremic toxin retention, suggesting potential intervention strategies in the future.
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Senescência Celular , Receptores de Antígenos de Linfócitos T , Diálise Renal , Humanos , Feminino , Masculino , Pessoa de Meia-Idade , Idoso , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/imunologia , Estudos Transversais , Inibidor de Quinase Dependente de Ciclina p21/genética , Linfócitos T/imunologia , Inibidor p16 de Quinase Dependente de Ciclina/genética , Antígenos CD28 , Uremia/imunologia , Regiões Determinantes de Complementaridade/genética , Adulto , Doenças Cardiovasculares/imunologia , Linfócitos T CD4-Positivos/imunologiaRESUMO
BACKGROUND: The relationship between malnutrition-inflammation-atherosclerosis syndrome (MIAS) and self-management ability has not been previously revealed even though both play an important role in the management of peritoneal dialysis (PD) patients. METHODS: In total, 93 patients were enrolled in this study. A self-management questionnaire was used for the evaluation of self-management ability. The identification of MIAS was based on one or more of the following three conditions: C-reactive protein (CRP)≥10 mg/L, malnutrition-inflammation score (MIS)> 7, and the presence of atherosclerosis-related medical records. The possible association between different self-management abilities and MIAS was analyzed with a Spearman correlation analysis. RESULTS: There were 40 (43.0%) patients in the atherosclerosis group, and 38 (40.9%), 38 (40.9%), 10 (10.8%), and 7 (7.5%) patients in the MIAS0, MIAS1, MIAS2, and MIAS3 groups, respectively. The group with a score above the mean score of the Dialysis Effect Evaluation and Monitoring dimension had a fewer number of hospitalizations, higher albumin levels, lower MIS scores, a lower level of IL-6, and a lower number of MIAS factors. The Pearson and Spearman correlation analyses also revealed that this dimension was negatively correlated with the MIAS, MIS, IL-6, BNP, number of hospitalizations, and age and positively associated with albumin and prealbumin. CONCLUSION: The Dialysis Effect Evaluation and Monitoring dimension of the self-management scale for PD patients is closely linked to the MIAS, and a better dialysis effect evaluation and monitoring capacity results in a decreased likelihood of exposure to malnutrition and inflammation. TRIAL REGISTRATION: Chinese Clinical Trial Registry: ChiCTR2000035525 ( http://www.chictr.org.cn/showproj.aspx?proj=58110 ), registered August 13, 2020.
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Aterosclerose/etiologia , Inflamação/etiologia , Desnutrição/etiologia , Diálise Peritoneal/efeitos adversos , Autogestão , Idoso , Estudos Transversais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , SíndromeRESUMO
BACKGROUND: Renin-angiotensin system inhibitors (RASi) treatment is the basic therapy for IgA nephropathy (IgAN) patients. However, there is few of biomarker that can predict the efficacy of RASi. This study aimed to find urinary exosomal mRNAs related to the therapeutic effect of RASi in the treatment of proteinuria in IgAN patients. METHODS: We divided IgAN patients in screening cohort into A1 (proteinuria increase at 3 months), B1 (proteinuria decrease less than 50 % at 3 months), C1 (proteinuria decrease more than 50 % at 3 months) groups according to changes of proteinuria after treatment. The urinary exosomes were collected before biopsy, RNAs were extracted and analyzed with the microarray assay. The candidate genes were screened by differentially expressed genes (DEGs) analysis and then validated by quantitative real-time polymerase chain reaction (qPCR) in a validation cohort. A receiver operating characteristic (ROC) curve was used to evaluate gene performance in predicting therapeutic effect on RASi reducing proteinuria in IgAN patients. RESULTS: ECE1 and PDE1A mRNAs were significantly different among the three groups, and were gradually decreased among A1, B1 and C1 groups. In the validation cohort, the level of urinary exosomal ECE1 and PDE1A mRNAs were also significantly lower in A2 group compared with C2 group(ECE1, P < 0.001;PDE1A, P < 0.01). Besides, the level of ECE1 mRNA was also lower in B2 group compared with C2 group (P < 0.01). The ROC curve verified that urinary exosomal ECE1 and PDE1A gene level predicted RASi efficacy in IgAN patients with area under curve (AUC) 0.68 and 0.63 respectively. CONCLUSION: Urinary exosomal ECE1 and PDE1A mRNAs expression can serve as potential biomarkers for predicting the RASi efficacy to reduce proteinuria in IgAN patients.
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Biomarcadores , Exossomos , Glomerulonefrite por IGA , RNA Mensageiro , Sistema Renina-Angiotensina , Humanos , Glomerulonefrite por IGA/tratamento farmacológico , Glomerulonefrite por IGA/urina , Glomerulonefrite por IGA/genética , Masculino , RNA Mensageiro/genética , RNA Mensageiro/urina , Adulto , Feminino , Exossomos/genética , Exossomos/metabolismo , Biomarcadores/urina , Sistema Renina-Angiotensina/efeitos dos fármacos , Sistema Renina-Angiotensina/genética , Pessoa de Meia-Idade , Inibidores da Enzima Conversora de Angiotensina/uso terapêuticoRESUMO
AIMS: Parietal epithelial cells (PECs) are potential stem cells within the glomerulus, migrating into site of podocytes loss to differentiate into podocytes. Little is known about the mechanism mediating differentiation of PECs into podocytes. RESULTS: In vitro differentiation of PECs into podocytes led to upregulation of podocyte markers such as Wilms' tumour gene 1 (WT-1), FOXC1, synaptopodin, and podocin, accompanied by increased mitochondrial abundance. Preincubation with a mitochondrial reactive oxygen species (ROS) inhibitor prevented all these events in PECs. In vivo, adriamycin (ADR)-treated mice exhibited albuminuria, decreased WT1 positive cells, and claudin-1 expressed in glomerular capillary tuft, as well as PGC-1α overproduction in PECs. Expression of the ROS-related molecule nuclear factor erythroid 2-related factor 2 (Nrf2) and its target protein Brahma-related gene 1 (Brg1) increased during differentiation of PECs into podocytes. Suppressing Nrf2 or Brg1 reduced the differentiation of PECs, while overexpression had the opposite effect. Brg1 directly regulated WT-1 transcription in PECs. Activation of Nrf2 with Bardoxolone-methyl (CDDO-Me) resulted in less proteinuria and more WT1 positive cells in ADR mice. PECs conditional human Nrf2 knock-in (cKI) mice showed increased WT1 cell numbers. CONCLUSION: It concluded that mitochondria-derived ROS mediated differentiation of PECs into podocytes via Nrf2 and Brg1 signaling.
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BACKGROUND: The relationship between Bowman's capsule thickening and progression of diabetic kidney disease (DKD) remains uncertain. METHODS: Renal biopsy specimens from 145 DKD patients and 20 control subjects were evaluated for Bowman's capsule thickness. Immunohistochemical staining assessed col4α2, laminin ß1, and albumin expression. In a discovery cohort of 111 DKD patients with eGFR ≥ 30 ml/min/1.73 m2, thickening was classified as fibrotic or exudative. The composite endpoint included CKD stage 5, dialysis initiation, and renal disease-related death. Prognosis was analyzed using Kaplan-Meier and Cox regression analyses. Two validation cohorts were included. RESULTS: Three types of thickening were observed: fibrotic, exudative, and periglomerular fibrosis. Parietal epithelial cell matrix protein accumulation contributed to fibrotic thickening, while albumin was present in exudative thickening. Bowman's capsule was significantly thicker in DKD patients (5.74 ± 2.09 µm) compared to controls (3.38 ± 0.43 µm, P < 0.01). In discovery cohort, the group of exudative thickning had a poorer prognosis(median time 20 months vs 57 months, P = 0.000). Cox multivariate analysis revealed that exudative thickening of Bowman's capsule were associated with a poor prognosis. The validation cohorts confirmed the result. CONCLUSIONS: Various mechanisms contribute to Bowman's capsule thickening in DKD. The proportion of exudative thickening may serve as a valuable prognostic indicator for DKD patients.
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Diabetes Mellitus , Nefropatias Diabéticas , Falência Renal Crônica , Humanos , Cápsula Glomerular/metabolismo , Cápsula Glomerular/patologia , Nefropatias Diabéticas/patologia , Falência Renal Crônica/patologia , Diálise Renal , Albuminas , Diabetes Mellitus/patologiaRESUMO
Diabetic nephropathy is one of the leading causes of end-stage renal disease worldwide. In our study we found that Adenosine triphosphate (ATP) content was significantly increased in the urine of diabetic mice. We examined the expression of all purinergic receptors in the renal cortex and found that only purinergic P2X7 receptor (P2X7R) expression was significantly increased in the renal cortex of wild-type diabetic mice and that the P2X7R protein partially co-localized with podocytes. Compared with P2X7R(-/-) non-diabetic mice, P2X7R(-/-) diabetic mice showed stable expression of the podocyte marker protein podocin in the renal cortex. The renal expression of microtubule associated protein light chain 3 (LC-3II) in wild-type diabetic mice was significantly lower than in wild-type controls, whereas the expression of LC-3II in the kidneys of P2X7R(-/-) diabetic mice was not significantly different from that of P2X7R(-/-) non-diabetic mice. In vitro, high glucose induced an increase in p-Akt/Akt, p-mTOR/mTOR and p62 protein expression along with a decrease in LC-3II levels in podocytes, whereas after transfection with P2X7R siRNA, Phosphorylated protein kinase B (p-Akt)/Akt, Phosphorylated mammalian target of rapamycin (p-mTOR)/mTOR, and p62 expression were restored and LC-3II expression was increased. In addition, LC-3II expression was also restored after inhibition of Akt and mTOR signaling with MK2206 and rapamycin, respectively. Our results suggest that P2X7R expression is increased in podocytes in diabetes, and that P2X7R is involved in the inhibition of podocyte autophagy by high glucose, at least in part through the Akt-mTOR pathway, thereby exacerbating podocyte damage and promoting the onset of diabetic nephropathy. Targeting P2X7R may be a potential treatment for diabetic nephropathy.
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Nefropatias Diabéticas , Podócitos , Camundongos , Animais , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Nefropatias Diabéticas/genética , Receptores Purinérgicos P2X7/genética , Receptores Purinérgicos P2X7/metabolismo , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo , Glucose/metabolismo , Autofagia , Mamíferos/metabolismoRESUMO
Background: Ultrafiltration (UF) volume and peritoneal solute transport rate (PSTR) are common parameters used to evaluate the efficacy of peritoneal dialysis (PD) on individual patients. It is unclear whether the level of exosomal microRNA (miRNA) in peritoneal dialysis effluent (PDE) can predict UF or PSTR. This study was designed to investigate if there is a correlation between PDE exosomal miRNA (miR-432-5p) levels and various UF volumes and PSTRs in PD patients. It also aimed to explore the underlying mechanism of water and dialytic sodium removal (DSR). Methods: The PSTR was quantified using the 4-hour (4 h) 3.86% dialysate to plasma creatinine ratio. The PDE exosomes (PDE-exo) were isolated by ultracentrifugation. An miRNA assay was used to identify the different miRNA in the PDE-exo of patients in a high (H; PSTR >0.65, n=5) and low (L; PSTR <0.65, n=5) group. We focused on miR-432-5p as bioinformatic analysis had shown that it could be involved in sodium transport. We used mimic/inhibitor transfection and dual luciferase reporter assay to verify the target genes of miR-432-5p. We used PKH-67 stained PDE-exo to observe their interaction with human MeT-5A mesothelial cells. Results: Our results showed that the PDE-exo-miR-432-5p level was higher in group H than in group L. The levels of PDE-exo-miR-432-5p were positively correlated with PSTR (r=0.391; P<0.05; n=40) and negatively correlated with the 4 h UF volume (r=-0.376; P<0.05; n=40) and 4 h DSR (r=-0.535; P<0.01; n=24). Epithelial sodium channel α subunit (α-ENaC) was revealed as a direct target gene of miR-432-5p and expressed on both human peritoneum and MeT-5A cells. Furthermore, we found the PKH67 labeled-PDE-exo could be internalized into MeT-5A cells. Conclusions: A high PDE-exo-miR-432-5p level was associated with poor UF volume and DSR. It may be that PDE-exo-miR-432-5p affects DSR through downregulating α-ENaC expression.
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Background: Extracellular vesicles (EVs) from peritoneal dialysis effluent (PDE), containing molecules such as proteins and microRNAs (miRNAs), may be potential biological markers to monitor peritoneal function or injury. Peritoneal inflammation is an important determinant of peritoneal solute transport rate (PSTR). Thus, the aim of this study is to determine whether the specific proteins capable of evaluating the PSTR could be found in PDE-EVs, and explore the underlying mechanism for the association between PSTR and peritoneal inflammation. Methods: Sixty patients undergoing peritoneal dialysis (PD) were divided into two groups: high/high average transport (H/A) group (PET >0.65) and low/low average transport (L/A) group (PET <0.65). EVs derived from PDE (PDE-EVs) were isolated by ultracentrifugation. Proteomic analysis was performed to explore the differentially expressed proteins and identify the potential biomarkers in PDE-EVs from the two groups, and we focused on glycoprotein 96 (GP96) as it could be involved in the inflammatory process. The expression of GP96 in PDE-EVs and inflammatory cytokines was quantified by real-time PCR and enzyme-linked immunosorbent assay. The infiltration of macrophages and neutrophils into the peritoneum was detected using immunohistochemistry in a PD rat model. Results: The expression of PDE-EVs-GP96 was significantly higher in the H/A group, and was positively correlated with the PSTR and the level of the inflammatory factor interleukin (IL)-6. GP96-enriched EVs enhanced the secretion of proinflammatory cytokines IL-1ß, IL-6, tumor necrosis factor (TNF)-α, and IL-8 in macrophages, which was reversed by a pharmacological GP96-specific inhibitor (PU-WS13). The GP96 inhibitor also reduced local peritoneal inflammation by decreasing the infiltration of inflammatory cells and levels of proinflammatory cytokines (IL-6 and TNF-α) and chemokines (CCL2, CXCL1, and CXCL2) in a PD rat model. Conclusions: PDE-EVs-GP96 is a new promising tool to evaluate the status of peritoneal inflammation and PSTR, and the mechanism may be related to affecting the inflammatory properties of macrophages.
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Vesículas Extracelulares , Diálise Peritoneal , Peritonite , Animais , Biomarcadores/metabolismo , Citocinas/metabolismo , Vesículas Extracelulares/metabolismo , Glicoproteínas/metabolismo , Humanos , Inflamação/metabolismo , Interleucina-6/metabolismo , Diálise Peritoneal/efeitos adversos , Peritônio/metabolismo , Peritonite/metabolismo , Proteômica , RatosRESUMO
Background: The Tp-e/QT (peak to end of T-wave duration/QT interval) ratio is a promising marker of myocardial repolarization and ventricular arrhythmogenesis. Its elevation is associated with sudden cardiac death in different clinical conditions. This study was designed to assess the possible association between increased Tp-e/QT ratio and clinical factors in peritoneal dialysis patients. Materials & methods: We devised a prospective cross-sectional study, which included 107 patients who were divided into groups according to their Tp-e/QT ratio. The association of an increased Tp-e/QT ratio with related factors was analyzed with multivariate logistic regression. Results: Thirty-one patients, who had an elevated Tp-e/QT ratio, showed higher values of IL-6, left ventricular end-systolic diameter, Tp-e, percentage of diabetes mellitus, coronary artery calcification, and left ventricular ejection fraction. Multivariate analysis revealed that IL-6 was an independent risk factor for a higher Tp-e/QT ratio after adjustments. Conclusion: Our study revealed that a high serum IL-6 level in peritoneal dialysis patients increased the risk of a higher Tp-e/QT ratio, which indicated a potentially hazardous interplay between inflammation and arrhythmogenesis.
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Arritmias Cardíacas/sangue , Eletrocardiografia , Inflamação/sangue , Interleucina-6/sangue , Falência Renal Crônica/sangue , Diálise Peritoneal , Idoso , Biomarcadores/sangue , Estudos Transversais , Fenômenos Eletrofisiológicos , Feminino , Seguimentos , Humanos , Falência Renal Crônica/complicações , Falência Renal Crônica/terapia , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Fatores de RiscoRESUMO
BACKGROUND: Wharton's jelly-derived mesenchymal stem cells (WJ-MSCs) from the human umbilical cord have been studied extensively due to their immunomodulatory functions. Large-conductance Ca2+-activated K+ (BKCa channels) channels are involved in many inflammatory responses, but their involvement in the anti-inflammatory activity of WJ-MSCs is unknown. The underlying molecular mechanism, through which BKCa channels mediate the immunomodulation of WJ-MSC, which may include changes in exosomes proteomics, has not yet been clarified. METHODS: Alizarin staining, Oil Red O staining, and flow cytometry were used to identify WJ-MSCs, which were isolated from human umbilical cord Wharton's jelly. BKCa channels were detected in WJ-MSCs using western blotting, real-time polymerase chain reaction (real-time PCR), and electrophysiology, and cytokine expression was examined using real-time PCR and enzyme-linked immunosorbent assays (ELISAs). Exosomes were characterized using transmission electron microscopy and nanoparticle tracking analysis. Proteomics analysis was performed to explore exosomal proteomic profiles. RESULTS: The cells derived from human umbilical cord Wharton's jelly were identified as MSCs. BKCa channels were detected in the isolated WJ-MSCs, and the expression of these channels increased after lipopolysaccharide (LPS) stimulation. BKCa channels blockade in LPS-treated WJ-MSCs induced apoptosis and decreased interleukin-6 (IL-6) expression. Furthermore, THP-1 cells (human monocytic cell line) stimulated with LPS/interferon gamma (IFN-γ) produced more anti-inflammatory cytokines after treatment with exosomes derived from BKCa channel-knockdown WJ-MSCs (si-exo). We also observed altered expression of mitochondrial ATP synthase alpha subunit (ATP5A1), filamin B, and other proteins in si-exo, which might increase the anti-inflammatory activity of macrophages. CONCLUSIONS: Our study described the functional expression of BKCa channels in WJ-MSCs, and BKCa channels regulated the immunomodulatory properties of WJ-MSCs by affecting the exosomal protein profiles during the inflammatory response.