Accuracy of a novel real-time continuous glucose monitoring system: a prospective self-controlled study in thirty hospitalized patients with type 2 diabetes.

Aims: The present study aimed to investigate the accuracy of the Glunovo® real-time continuous glucose monitoring system (rtCGMS). Methods: We conducted a 14-day interstitial glucose level monitoring using Glunovo® rtCGMS on thirty hospitalized patients with type 2 diabetes. The flash glucose monitoring (FGM) was used as a self-control. Consistency tests, error grid analysis, and calculation of the mean absolute relative difference (MARD) were performed using R software to assess the accuracy of Glunovo® rtCGMS. Results: Glunovo® exhibited an overall MARD value of 8.89% during hospitalization, compared to 10.42% for FGM. The overall percentages of glucose values within ±10%/10, ± 15%/15, ± 20%/20, ± 30%/30, and ±40%/40 of the venous blood glucose reference value were 63.34%, 81.31%, 90.50%, 97.29%, and 99.36% for Glunovo®, respectively, compared with 61.58%, 79.63%, 88.31%, 96.22% and 99.23% for FGM. The Clarke Error Grid Analysis showed that 99.61% of Glunovo® glucose pairs and 100.00% of FGM glucose pairs within zones A and B. Conclusion: Our study confirms the superior accuracy of Glunovo® in monitoring blood glucose levels among hospitalized patients with type 2 diabetes.

Serum bile acid and unsaturated fatty acid profiles of non-alcoholic fatty liver disease in type 2 diabetic patients.

BACKGROUND: The understanding of bile acid (BA) and unsaturated fatty acid (UFA) profiles, as well as their dysregulation, remains elusive in individuals with type 2 diabetes mellitus (T2DM) coexisting with non-alcoholic fatty liver disease (NAFLD). Investigating these metabolites could offer valuable insights into the pathophy-siology of NAFLD in T2DM. AIM: To identify potential metabolite biomarkers capable of distinguishing between NAFLD and T2DM. METHODS: A training model was developed involving 399 participants, comprising 113 healthy controls (HCs), 134 individuals with T2DM without NAFLD, and 152 individuals with T2DM and NAFLD. External validation encompassed 172 participants. NAFLD patients were divided based on liver fibrosis scores. The analytical approach employed univariate testing, orthogonal partial least squares-discriminant analysis, logistic regression, receiver operating characteristic curve analysis, and decision curve analysis to pinpoint and assess the diagnostic value of serum biomarkers. RESULTS: Compared to HCs, both T2DM and NAFLD groups exhibited diminished levels of specific BAs. In UFAs, particular acids exhibited a positive correlation with NAFLD risk in T2DM, while the ω-6:ω-3 UFA ratio demonstrated a negative correlation. Levels of α-linolenic acid and γ-linolenic acid were linked to significant liver fibrosis in NAFLD. The validation cohort substantiated the predictive efficacy of these biomarkers for assessing NAFLD risk in T2DM patients. CONCLUSION: This study underscores the connection between altered BA and UFA profiles and the presence of NAFLD in individuals with T2DM, proposing their potential as biomarkers in the pathogenesis of NAFLD.

Satellite cell-derived exosome-mediated delivery of microRNA-23a/27a/26a cluster ameliorates the renal tubulointerstitial fibrosis in mouse diabetic nephropathy.

Renal tubulointerstitial fibrosis (TIF) is considered as the final convergent pathway of diabetic nephropathy (DN) without effective therapies currently. MiRNAs play a key role in fibrotic diseases and become promising therapeutic targets for kidney diseases, while miRNA clusters, formed by the cluster arrangement of miRNAs on chromosomes, can regulate diverse biological functions alone or synergistically. In this study, we developed clustered miR-23a/27a/26a-loaded skeletal muscle satellite cells-derived exosomes (Exos) engineered with RVG peptide, and investigated their therapeutic efficacy in a murine model of DN. Firstly, we showed that miR-23a-3p, miR-26a-5p and miR-27a-3p were markedly decreased in serum samples of DN patients using miRNA sequencing. Meanwhile, we confirmed that miR-23a-3p, miR-26a-5p and miR-27a-3p were primarily located in proximal renal tubules and highly negatively correlated with TIF in db/db mice at 20 weeks of age. We then engineered RVG-miR-23a/27a/26a cluster loaded Exos derived from muscle satellite cells, which not only enhanced the stability of miR-23a/27a/26a cluster, but also efficiently delivered more miR-23a/27a/26a cluster homing to the injured kidney. More importantly, administration of RVG-miR-23a/27a/26a-Exos (100 µg, i.v., once a week for 8 weeks) significantly ameliorated tubular injury and TIF in db/db mice at 20 weeks of age. We revealed that miR-23a/27a/26a-Exos enhanced antifibrotic effects by repressing miRNA cluster-targeting Lpp simultaneously, as well as miR-27a-3p-targeting Zbtb20 and miR-26a-5p-targeting Klhl42, respectively. Knockdown of Lpp by injection of AAV-Lpp-RNAi effectively ameliorated the progression of TIF in DN mice. Taken together, we established a novel kidney-targeting Exo-based delivery system by manipulating the miRNA-23a/27a/26a cluster to ameliorate TIF in DN, thus providing a promising therapeutic strategy for DN.

Association of Serum Bile Acid and Unsaturated Fatty Acid Profiles with the Risk of Diabetic Retinopathy in Type 2 Diabetic Patients.

Aim: We aimed to identify the ability of serum bile acids (BAs) and unsaturated fatty acids (UFAs) profiles to predict the development of diabetic retinopathy (DR) in type 2 diabetes mellitus (T2DM) patients. Methods: We first used univariate and multivariate analysis to compare 15 serum BA and 11 UFA levels in healthy control (HC) group (n = 82), T2DM patients with DR (n = 58) and T2DM patients without DR (n = 60). Forty T2DM patients were considered for validation. Then, the receiver operating characteristic curve (ROC) and decision curve analysis were used to assess the diagnostic value and clinical benefit of serum biomarkers alone, clinical variables alone or in combination, and the area under the curve (AUC), integrated discrimination improvement (IDI), and net reclassification improvement (NRI) were used to further assess whether the addition of biomarkers significantly improved the predictive ability of the model. Results: Orthogonal partial least squares-discriminant analysis (OPLS-DA) of serum BAs and UFAs separated the three cohorts including HC, T2DM patients with or without DR. The difference in serum BA and UFA profiles of T2DM patients with or without DR was mainly manifested in the three metabolites of taurolithocholic acid (TLCA), tauroursodeoxycholic acid (TUDCA) and arachidonic acid (AA). Together, they had an AUC of 0.785 (0.918 for validation cohort) for predicting DR in T2DM patients. After adjusting for numerous confounding factors, TLCA, TUDCA, and AA were independent predictors that differentiated T2DM with or without DR. The results of AUC, IDI, and NRI demonstrated that adding these three biomarkers to a model with clinical variables statistically increased their predictive value and were replicated in our independent validation cohort. Conclusion: These findings highlight the association of three metabolites, TLCA, TUDCA and AA, with DR and may indicate their potential value in the pathogenesis of DR.

Fecal microbiota transplantation ameliorates type 2 diabetes via metabolic remodeling of the gut microbiota in db/db mice.

INTRODUCTION: Gut microbiome (GM) deregulation has been implicated in major conditions such as obesity and type 2 diabetes (T2DM). Our previous prospective study indicated that fecal microbiota transplantation (FMT) successfully improved patients with T2DM. We hypothesized that FMT may be a potential therapeutic method for T2DM, but its precise mechanisms in T2DM remains to be elucidated. RESEARCH DESIGN AND METHODS: Eight db/m mice were FMT donors and control mice, and 16 genetically diabetic db/db mice were equally divided into two groups (db/db+phosphate-buffered saline (PBS) group, db/db+FMT group). The db/db+FMT group was administered fresh fecal suspension (0.2 mL/mice) daily for 4 weeks. Analysis of the GM and serum metabolome was carried out by 16S ribosomal RNA sequencing and liquid chromatogram-mass spectrometry, respectively. Effects of FMT on the gut barrier and pancreas were assessed using protein assays, messenger RNA, immunohistology and clinical indicators testing. RESULTS: Our results showed that FMT treatment of db/db mice relieves a series of clinical indicators, including fasting plasma glucose, serum insulin and oral glucose tolerance test among others. Compared with non-diabetic control mice, db/db+PBS mice exhibited decreased abundance of Ruminococaceae, Porphyromonadaceae and increased abundance of Rikenellaceae and Lactobacillaceae. FMT treatment reversed this effect on the microbiome. Eleven metabolites were changed between the db/db+PBS and db/db+FMT groups. Correlation analysis showed that the structural changes of the GM were correlated with host metabolite levels. We further showed that FMT treatment of db/db mice improved intestinal barrier function, reduced inflammation and caused an alteration in the number of circulating immune cells. CONCLUSIONS: FMT-mediated changes in the GM, serum metabolites, intestinal epithelial barrier, inflammation and circulating immune cells play an important role in the efficacy of FMT on T2DM disease progression.

Prospective Study Reveals Host Microbial Determinants of Clinical Response to Fecal Microbiota Transplant Therapy in Type 2 Diabetes Patients.

Background: Increasing evidence shows that alterations in gut microbiome (GM) contribute to the development of type 2 diabetes mellitus (T2DM), and fecal microbiota transplantation (FMT) successfully treats various human diseases. However, the benefits of FMT therapy to T2DM patients remain unknown. Methods: We enrolled 17 patients with T2DM for nonblinded, one-armed intervention trial of FMT. A total of 20 healthy individuals were recruited as the baseline control. HbA1c% and metabolic parameter change were evaluated in 17 T2DM patients 12 weeks after they received FMT from healthy donors. The GM composition was characterized by 16S rRNA gene amplicon sequencing from fecal samples prior to and 12 weeks after FMT treatment. Results: We found that the GM of T2DM patients was reconstituted by FMT. We observed a statistically significant decrease in HbA1c% (from 7.565 ± 0.148 to 7.190 ± 0.210, p<0.01), blood glucose (from 8.483 ± 0.497 to 7.286 ± 0.454 mmol/L, p<0.01), and uric acid (from 309.4 ± 21.5 to 259.1 ± 15.8 µmol/L, p<0.01) while a significant increase in postprandial C-peptide (from 4.503 ± 0.600 to 5.471 ± 0.728 ng/ml, p<0.01) at 12 weeks after FMT. Closely evaluating the changes in these assays, we found individual variability in response to FMT treatment. Out of 17 T2DM patients, 11 were found to significantly improve T2DM symptoms. The FMT responders have significantly higher levels of the family Rikenellaceae and the genus Anaerotruncus (family Ruminococcaceae) in their pretreated fecal in comparison to nonresponders, which could predict the clinical response with an area under the curve of 0.83. Conclusion: Our findings suggest that certain T2DM patients can potentially benefit from FMT, and the pretreated abundance of Rikenellaceae and Anaerotruncus in the fecal of patients may serve as potential biomarkers for selecting T2DM patients to receive FMT.

Silencing of Long Non-coding RNA ENST00000606790.1 Inhibits the Malignant Behaviors of Papillary Thyroid Carcinoma through the PI3K/AKT Pathway.

PURPOSE: This study aimed to investigate the role and mechanism of lncRNA ENST00000606790.1 (ENST) in promoting the progression of papillary thyroid carcinoma (PTC). METHODS: The expression of ENST in human PTC and normal para-cancerous thyroid (NPTC) tissues or cell lines was determined by RT-qPCR. Cell growth was determined by CCK8 assay. Cell colony formation was determined by cell colony formation assay. Cell cycle analysis was performed by staining cells with PI (Propidium Iodide). Cell invasion was assessed by transwell assay. Protein expression was examined by western-blot. siRNA was constructed to inhibit the expression of ENST. 740-Y-P was used to activate PI3K. The correlation between ENST expression and clinical outcomes was analyzed. RESULTS: ENST was significantly up-regulated in PTC tissues or PTC cell lines (PTC and IHH4 cell lines), compared to NPTC tissues or normal cell lines, respectively. High expression of ENST was strongly correlated to lymph node metastasis and tumor size at diagnosis. Silencing of ENST significantly inhibited cell growth and colony formation, arrested the cell cycle at G2/M phase, upregulated the expression of CHK1, downregulated the expression of CDC25C, and inhibited cell invasion. Silencing of ENST significantly down-regulated the expression of PI3K, p-PI3K, AKT, and p-AKT in IHH4 cells. Furthermore, treatment with the PI3K activator  740-Y-P partially abolished the effect of silencing of ENST on PTC cells. CONCLUSIONS: Overall, our results demonstrated that ENST can promote PTC progression by activating the PI3K/AKT signaling pathway, suggesting that ENST can serve as a potential biomarker and new therapeutic target for patients with PTC.

Resveratrol Modulates the Gut Microbiota and Inflammation to Protect Against Diabetic Nephropathy in Mice.

Oral administration of resveratrol is able to ameliorate the progression of diabetic nephropathy (DN); however, its mechanisms of action remain unclear. Recent evidence suggested that the gut microbiota is involved in the metabolism therapeutics. In the current study, we sought to determine whether the anti-DN effects of resveratrol are mediated through modulation of the gut microbiota using the genetic db/db mouse model of DN. We demonstrate that resveratrol treatment of db/db mice relieves a series of clinical indicators of DN. We then show that resveratrol improves intestinal barrier function and ameliorates intestinal permeability and inflammation. The composition of the gut microbiome was significantly altered in db/db mice compared to control db/m mice. Dysbiosis in db/db mice characterized by low abundance levels of Bacteroides, Alistipes, Rikenella, Odoribacter, Parabacteroides, and Alloprevotella genera were reversed by resveratrol treatment, suggesting a potential role for the microbiome in DN progression. Furthermore, fecal microbiota transplantation, derived from healthy resveratrol-treated db/m mice, was sufficient to antagonize the renal dysfunction, rebalance the gut microbiome and improve intestinal permeability and inflammation in recipient db/db mice. These results indicate that resveratrol-mediated changes in the gut microbiome may play an important role in the mechanism of action of resveratrol, which provides supporting evidence for the gut-kidney axis in DN.

Overexpression of novel long intergenic non­coding RNA LINC02454 is associated with a poor prognosis in papillary thyroid cancer.

It has been revealed from microarray data analysis that long intergenic non­coding RNA 02454 (LINC02454) is highly expressed in papillary thyroid cancer (PTC). The aim of the present study was to explore the potential role of LINC02454 in the tumorigenesis of PTC. The mRNA expression levels of LINC02454 were assessed using data from The Cancer Genome Atlas (TCGA) and the GSE66783 cohort in thyroid cancer, and were validated using reverse transcription­quantitative PCR in 104 patients with PTC recruited in the present study. The association between the LINC02454 mRNA expression levels and the clinicopathological features of the 104 patients with PTC were also analyzed. Functional enrichment analyses were conducted on the differentially expressed genes in the high and low LINC02454 expression groups that were identified from the TCGA cohort. RNA interference, using short interfering (si)RNA against LINC02454, was used to investigate the role of LINC02454 in the biological functions of PTC cells in vitro. The expression level of LINC02454 was significantly increased in PTC tissues (P=0.0011) and was significantly associated with a larger tumor size, T stage, an advanced TNM stage and an increased lymph node metastasis (P<0.05), which was consistent with that in the TCGA and GSE66783 cohort. High expression levels of LINC02454 were observed in patients with PTC that also had BRAF mutations (P<0.001), and were significantly associated with a poorer disease­free survival in the TCGA cohort (P<0.05). Functional enrichment analysis indicated that LINC02454­related genes were significantly enriched in Gene Ontology terms, such as 'positive regulation of cell proliferation', 'positive regulation of cell division' and 'cell adhesion', and the following Kyoto Encyclopedia of Genes and Genomes pathways: 'Pathways in cancer' 'proteoglycans in cancer' and 'ECM­receptor interaction'. In vitro, the knockdown of LINC02454 markedly arrested the cells in the G0/G1 phase of the cell cycle, and also led to an overall increase in apoptosis, as well as to an unexpected decrease in cell proliferation. LINC02454 may thus potentially function as an oncogene, which inhibits the apoptosis and enhances proliferation of PTC cells. Thus, as suggested by the findings of the present study, LINC02454 may be used as a diagnostic and prognostic biomarker for PTC in the future.

Characteristics of the intestinal flora in patients with peripheral neuropathy associated with type 2 diabetes.

OBJECTIVE: To study the characteristics of the intestinal flora in patients with diabetic peripheral neuropathy (DPN) and analyze the association between the intestinal flora and clinical indicators. METHODS: We classified 80 subjects into three groups: patients with DPN (n = 45), patients type 2 diabetes without DPN (n = 21), and healthy controls (n = 14). The intestinal flora composition was compared among the three groups, and the correlation between the intestinal flora and clinical indicators was analyzed. RESULTS: At the phylum level, the richness of Firmicutes and Actinobacteria was elevated in the DN group, and that of Bacteroidetes was decreased. At the genus level, the richness of Bacteroides and Faecalibacterium was significantly decreased in the DPN group, whereas that of Escherichia-Shigella, Lachnoclostridium, Blautia, Megasphaera, and Ruminococcus torques group was increased. The homeostasis model assessment insulin resistance index was positively correlated with Megasphaera richness. Glycine ursodeoxycholic acid was positively correlated with Ruminococcus gnavus group and Phascolarctobacterium richness. Tauroursodeoxycholic acid was positively correlated with Ruminococcus gnavus group and Parabacteroides richness. CONCLUSION: There was obvious intestinal microbiota disorder in patients with DPN, which may be related to insulin resistance. These changes may have important roles in the development of DPN.

Opposite effects of vitamin C and vitamin E on the antifungal activity of honokiol.

The aim of the present study was to evaluate the effects of two well-known natural antioxidants vitamin C (VC) and vitamin E (VE) on the antifungal activity of honokiol against Candida albicans. The broth microdilution method was employed to test the antifungal activities of honokiol with or without antioxidants in the medium against C. albicans strain. Intracellular reactive oxygen species (ROS) and lipid peroxidation were determined by fluorescence staining assay. Mitochondrial dysfunction was assessed by detecting the mitochondrial DNA and the mitochondrial membrane potential. We observed that VC could significantly potentiate the antifungal activities of honokiol while VE reduced the effectiveness of honokiol against C. albicans. In addition, VC accelerated honokiol-induced mitochondrial dysfunction and inhibited glycolysis leading to a decrease in cellular ATP. However, VE could protect against mitochondrial membrane lipid peroxidation and rescue mitochondrial function after honokiol treatment. Our research provides new insight into the understanding of the action mechanism of honokiol and VC combination against C. albicans.

LncRNA ENST00000539653 acts as an oncogenic factor via MAPK signalling in papillary thyroid cancer.

BACKGROUND: Papillary thyroid cancer (PTC) is the most frequent type of thyroid malignancy. In this study, we investigated the mechanisms whereby long non-coding RNAs (lncRNAs) are associated with PTC pathogenesis. METHODS: Microarray analysis was used to determine differentially expressed lncRNAs between paired PTC tissues and normal adjacent thyroid tissues. Quantitative RT-PCR was used for validation in 86 PTC cases. Small interfering RNA (siRNA) transfection assays were then performed to assess how a novel lncRNA affected key proliferation and cell death pathways in IHH4 PTC cells. RESULTS: We identified 1878 differentially expressed lncRNAs versus matched control samples (fold change ≥2.0, P < 0.05), of which 429 were upregulated and 1449 were downregulated. ENST00000539653.1 (ENS-653), one of the top hits in this microarray, was selected for further study. Higher ENS-653 expression was observed in PTC tissue samples versus adjacent normal tissues, and was associated with a larger tumor size and a more advanced clinical stage. In the Cancer Genome Atlas (TCGA) PTC cohort, higher ENS-653 expression was correlated with more frequent BRAF (V600E) mutation and poorer disease-free survival. Furthermore, ENS-653 downregulation reduced the proliferation of PTC cells and led to G1-S arrest, but had no impact on apoptosis. ENS-653 downregulation also inactivated ERK1/2 and ERK5, causing partial MAPK cascade suppression. CONCLUSION: ENS-653 exhibits oncogenic properties in PTC, and could be a diagnostic and/or prognostic PTC biomarker, in addition to possibly being a future target for therapy.

Fecal microbiota transplantation relieve painful diabetic neuropathy: A case report.

RATIONALE: Fecal microbiota transplantation (FMT) has been used in a wide variety of diseases. In this article, we reported a 46-year-old female with diabetic neuropathy (DN) achieved remission by the treatment of FMT. PATIENT CONCERNS: The patient with an 8-year history of diabetes and hypertension was admitted to hospital due to sensitive pain of her right thigh and poor blood glucose control. The traditional hypoglycemic and analgesic treatment were useless to her symptoms. DIAGNOSIS: Diabetic-induced neuropathy was considered. INTERVENTIONS: This patient received twice FMTs for 3 months. OUTCOMES: After twice FMTs, the clinical response of patient was pleasant. The glycemic control was improved, with a remarkable relief of the symptoms of painful DN in particular. No obvious adverse effects were observed during the FMTs and follow-up observation-testing. LESSONS: We proposed that FMT could be a promising treatment in patients with diabetes or diabetes-related complications like DN. FMT also appeared to be definitely safer and more tolerable than the pharmacologic treatment in patients with DN.

Dysregulation of lncRNAs GM5524 and GM15645 involved in high­glucose­induced podocyte apoptosis and autophagy in diabetic nephropathy.

Diabetic nephropathy (DN) is an important microvascular complication of diabetes, and one of the leading causes of end­stage kidney disease. However, the mechanism of the DN pathogenic process remains unclear. Recently, long non­coding (lnc)RNA dysregulation has been regarded to cause the occurrence and development of various human diseases, although the functions of lncRNAs in human DN are poorly understood. The authors' previous study using microarray analysis identified hundreds of dysregulated lncRNAs in DN, although the functions of these lncRNAs were not demonstrated. Out of those dysregulated lncRNAs, Gm5524 was significantly upregulated in response to DN, while Gm15645 was significantly downregulated in response to DN. In the present study, this result was further validated by reverse transcription­quantitative polymerase chain reaction assays, and downregulating or overexpressing Gm5524 and Gm15645 in mouse podocytes. Notably, knockdown of Gm5524 and overexpression of Gm15645 induced mouse podocyte apoptosis and decreased cell autophagy in high­glucose culture conditions. In conclusion, the results of the present study revealed the roles of lncRNAs Gm5524 and Gm15645 in high­glucose induced podocyte apoptosis and autophagy during DN, which may further the understanding of the involvement of lncRNAs in DN, and provide a potential novel therapeutic target for this disease.

Integrated bioinformatics analysis reveals that the expression of cathepsin S is associated with lymph node metastasis and poor prognosis in papillary thyroid cancer.

The prognosis of the majority of patients with papillary thyroid cancer (PTC) is excellent, although there are patients who experience disease recurrence and progression. The aim of the present study was to identify potential prognostic risk markers in PTC. Differentially expressed genes (DEGs), identified from four Genome Expression Omnibus cohorts were subjected to functional enrichment analyses with Gene Ontology terms and the Kyoto Encyclopedia of Genes and Genome pathways. Hub genes, filtered from cytoHubba, were validated using the The Cancer Genome Atlas (TCGA) cohort, and their associations with clinicopathological features and prognosis were analyzed. A total of 277 DEGs were identified following data preprocessing. DEGs were primarily enriched in 'small cell lung cancer', 'ECM-receptor interaction', 'pathways in cancer'and 'tyrosine metabolism'. Hub genes [APOE, cathepsin S (CTSS), insulin receptor substrate 1 (IRS1), KIT, LGALS3, RUNX2 and TGFBR1] were extracted from cytoHubba. Their expression in the TCGA cohort was consistent with that in the GEO cohorts. CTSS (P=0.006) and IRS1 (P=0.005) were associated with disease­free survival, as determined using the Kaplan-Meier analysis. CTSS was an independent risk factor for poor disease­free survival (HR, 2.649; 95% CI, 1.095-6.409; P=0.031). Patients with high expression of CTSS exhibited different histological types (increased tall-cell subtype and reduced follicular subtype; P<0.001), more frequent lymph node metastasis (P<0.001) and advanced tumor-node-metastasis stages (P=0.049) compared with the low-expression group. High expression of CTSS was independently associated with lymph node metastasis (OR, 2.015; 95% CI, 1.225-3.315; P=0.006). Therefore, CTSS may serve as a predictive risk marker for the progression and prognosis of PTC.

MicroRNA-134-5p promotes high glucose-induced podocyte apoptosis by targeting bcl-2.

Podocyte apoptosis is a typical early feature of diabetic nephropathy (DN), with loss of nephrin integrity contributing to increased proteinuria in patients with DN. Emerging evidence shows that microRNAs (miRNAs) play vital roles in the pathogenesis of DN. Thus, we aimed to further elucidate the role of miRNAs in podocyte apoptosis in DN. We used db/db and db/m mice maintained under a continuous feeding regime for 12 weeks. Using microarray analysis, we found several miRNAs potentially related to podocyte apoptosis. In addition, we cultured a conditionally immortalized human podocyte cell line in 30 mM D-glucose and found that miR-134-5p was upregulated in both db/db mice and high-glucose (HG)-treated podocytes. Upregulation of miR-134-5p was accompanied by podocyte apoptosis and downregulation of nephrin. Inhibition of miR-134-5p produced the opposite effect. Dual-luciferase reporter assays showed that miR-134-5p directly targeted the 3'-untranslated region of the B-cell lymphoma-2 gene (BCL2), and further study confirmed an increase in bcl-2 protein level in HG-treated podocytes transfected with anti-miR-134-5p. Knockdown of BCL2 impeded the antiapoptotic effect of anti-miR-134-5p. Finally, we found that miR-134-5p might regulate apoptosis in db/db mice and podocytes by targeting BCL2. Taken together, our findings suggest that miR-134-5p promotes podocyte apoptosis under HG conditions by targeting BCL2. Our study provides a meaningful approach to interpret the mechanisms of action of miRNAs involved in DN.

Hyperoside alleviates adriamycin-induced podocyte injury via inhibiting mitochondrial fission.

Podocyte injury underlies many forms of glomerular diseases. Our previous study showed that hyperoside, a naturally occurring flavonoid, could decrease albuminuria at the early stage of diabetic nephropathy by ameliorating renal damage and podocyte injury. However, its protective mechanism against podocyte injury is unknown. A previous study demonstrated that hyperoside might inhibit amyloid ß-protein-induced neurotoxicity by suppressing mitochondrial dysfunction. Both mitochondrial dysfunction and its upstream determinant mitochondrial fission were closely related to podocyte injury. Thus, in the current study, we tested the effect of hyperoside on mitochondrial dysfunction and mitochondrial fission in adriamycin (ADR)-induced podocyte injury. In the mice model of ADR-induced nephropathy, hyperoside treatment inhibited ADR-induced albuminuria and podocyte injury. Meanwhile, hyperoside also blocked ADR-induced mitochondrial dysfunction and mitochondrial fission. Consistently, in cultured human podocytes, hyperoside suppressed ADR-induced podocyte injury, mitochondrial dysfunction and mitochondrial fission. All these results indicated that hyperoside might inhibit ADR-induced mitochondrial dysfunction and podocyte injury through suppressing mitochondrial fission both in vivo and in vitro. The underlying mechanisms which we revealed support the therapeutic effects of hyperoside for a broad range of glomerular diseases.

Resveratrol protects podocytes against apoptosis via stimulation of autophagy in a mouse model of diabetic nephropathy.

Podocyte apoptosis coincides with albuminuria onset and precedes podocytopenia in diabetic nephropathy. However, there is a lack of effective therapeutic drugs to protect podocytes from apoptosis. Here, we demonstrated that resveratrol relieved a series of indicators of diabetic nephropathy and attenuated apoptosis of podocytes in db/db diabetic model mice. In addition, resveratrol induced autophagy in both db/db mice and human podocytes. Furthermore, inhibition of autophagy by 3-methyladenine (3-MA) and autophagy gene 5 (Atg5) short hairpin RNA (shRNA) reversed the protective effects of resveratrol on podocytes. Finally, we found that resveratrol might regulate autophagy and apoptosis in db/db mice and podocytes through the suppression of microRNA-383-5p (miR-383-5p). Together, our results indicate that resveratrol effectively attenuates high glucose-induced apoptosis via the activation of autophagy in db/db mice and podocytes, which involves miR-383-5p. Thus, this study reveals a new possible strategy to treat diabetic nephropathy.

Microarray analysis of long noncoding RNA expression patterns in diabetic nephropathy.

AIMS: Long noncoding RNAs (lncRNAs) are implicated in various biological processes and human diseases. Diabetic nephropathy (DN) is the leading cause of end-stage renal disease (ESRD). We explored the potential functions of lncRNAs in DN. METHODS: We established a mouse model of DN and compared lncRNA expression patterns between DN model and db/m control mouse kidney tissues using microarray analysis. lncRNA function was predicted by gene ontology enrichment and KEGG pathway analyses of lncRNAs-coexpressed mRNAs. Quantitative reverse-transcription PCR was used for validation. Cis- and trans-regulation analyses were conducted to reveal potential relationships between lncRNAs and their target genes. RESULTS: In DN, 311 lncRNAs were dysregulated. LncRNA-coexpressed mRNAs were mainly targeted to golgi apparatus (ontology: cellular component), catalytic activity (ontology: molecular function), and mitotic nuclear division (ontology: biological process), and were mostly enriched in glutathione metabolism signaling. One hundred forty-seven lncRNAs were regarded as cis-regulatory. Several groups of lncRNAs may participate in biological pathways related to DN via trans-regulation of protein-coding genes. CONCLUSION: Hundreds of lncRNAs are dysregulated in DN. These lncRNAs might be involved in the pathogenesis of DN by modulating multiple molecular pathways. Our findings provide potential candidate biomarkers for predicting or diagnosing DN.

Heme oxygenase-1 enhances autophagy in podocytes as a protective mechanism against high glucose-induced apoptosis.

Injury and loss of podocytes play vital roles in diabetic nephropathy progression. Emerging evidence suggests autophagy, which is induced by multiple stressors including hyperglycemia, plays a protective role. Meanwhile, heme oxygenase-1 (HO-1) possesses powerful anti-apoptotic properties. Therefore, we investigated the impact of autophagy on podocyte apoptosis under diabetic conditions and its association with HO-1. Mouse podocytes were cultured in vitro; apoptosis was detected by flow cytometry. Transmission electron microscopy and biochemical autophagic flux assays were used to measure the autophagy markers microtubule-associated protein 1 light chain 3-II (LC3-II) and beclin-1. LC3-II and beclin-1 expression peaked 12-24h after exposing podocytes to high glucose. Inhibition of autophagy with 3-methyladenine or Beclin-1 siRNAs or Atg 5 siRNAs sensitized cells to apoptosis, suggesting autophagy is a survival mechanism. HO-1 inactivation inhibited autophagy, which aggravated podocyte injury in vitro. Hemin-induced autophagy also protected podocytes from hyperglycemia in vitro and was abrogated by HO-1 siRNA. Adenosine monophosphate-activated protein kinase phosphorylation was higher in hemin-treated and lower in HO-1 siRNA-treated podocytes. Suppression of AMPK activity reversed HO-1-mediated Beclin-1 upregulation and autophagy, indicating HO-1-mediated autophagy is AMPK dependent. These findings suggest HO-1 induction and regulation of autophagy are potential therapeutic targets for diabetic nephropathy.