Proteasome subunit-α type-6 protein is post-transcriptionally repressed by the microRNA-4490 in diabetic nephropathy

A common complication of both type I and type II diabetes is nephropathy, characterized by accumulation of extracellular matrix in the glomerular mesangium. This indicates a central role of mesangial cells in the pathophysiology of diabetic nephropathy. Using the proteomic approach, it was earlier elucidated in a rat model that the proteasome subunit-α type-6 protein (PSMA6) is suppressed in the renal cortex in nephropathic kidney. However, the underlying mechanism effecting suppression of PSMA6 protein in the renal cortex is not yet known. Twenty diabetic patients were enrolled and the expression level of PSMA6 in them was detected by immunohistochemistry. The protein and mRNA expression levels of PSMA6 in NRK-52E cells under high glucose condition were determined by Western blot and quantitative real-time PCR, respectively. Dual luciferase assay was used to detect the relationship of PSMA6 and miR-4490. Our results show that PSMA6 protein is down-regulated in patients with diabetic nephropathy compared with healthy control. Using the NRK-52E cell line cultured under high glucose condition as an in vitro model of diabetic nephropathy, we show that loss of PSMA6 protein expression occured independent of changes the in PSMA6 mRNA expression. We next elucidate that PSMA6 mRNA is post-transcriptionally regulated by the microRNA (miRNA)-4490, whose expression is inversely correlated to PSMA6 protein expression. Using reporter assays we show that PSMA6 is a direct target of the miR-4490. Exogenous manipulation of miR-4490 levels modulated expression of PSMA6, indicating that miR-4490 can be tested as a biomarker for nephropathy in diabetic patients.

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The Long Noncoding RNA 150Rik Promotes Mesangial Cell Proliferation via miR-451/IGF1R/p38 MAPK Signaling in Diabetic Nephropathy

Cellular Physiology and Biochemistry ◽

10.1159/000495590 ◽

2018 ◽

Vol 51 (3) ◽

pp. 1410-1428 ◽

Cited By ~ 9

Author(s):

Yajuan Zhang ◽

Yan Sun ◽

Rui Peng ◽

Handeng Liu ◽

Weihao He ◽

...

Keyword(s):

Cell Proliferation ◽

Diabetic Nephropathy ◽

High Glucose ◽

Noncoding Rna ◽

Mapk Signaling ◽

Luciferase Assay ◽

Dependent Manner ◽

High Glucose Condition ◽

P38mapk Pathway ◽

Glucose Condition

Background/Aims: Diabetic nephropathy (DN) as the primary cause of end-stage kidney disease is a common complication of diabetes. However, the initiating molecular events triggering DN are unknown. Recently, long noncoding RNAs (lncRNAs) have been shown to play important roles in DN. Methods: The expression level of lncRNA 1500026H17Rik (150Rik for short) was measured by qRT-PCR (quantitative real-time PCR). Cell proliferation ability was detected by 5-Ethynyl-2’-deoxyuridine (EdU). The relationship between 150Rik and microRNA 451 (miR-451) was examined by luciferase assay and RNA immunoprecipitation (RIP) assay. Finally, the effect of 150Rik on cell proliferation through the miR-451/insulin-like growth factor 1 receptor (IGF1R)/mitogen-activated protein kinases (p38MAPK) pathway was detected by EdU, flow cytometry analysis, western blot. Results: We found that 150Rik, an evolutionarily conserved lncRNA, was significantly upregulated in renal tissue of db/db DN mice and in mesangial cells (MCs) cultured under a high glucose condition. Further, overexpression or knockdown of 150Rik was found to regulate cell proliferation in MCs. Moreover, 150Rik was found to interact with miR-451 in both a direct and argonaute-2 (Ago2)-dependent manner. Results also revealed that overexpression of 150Rik inhibited cell proliferation through the miR-451/IGF1R/p38MAPK pathway in MCs under the high glucose condition, while knockdown of 150Rik increased cell proliferation via the miR-451/IGF1R/p38MAPK pathway. Conclusion: Taken together, these results provide new insight into the association between 150Rik and the miR-451/IGF1R/p38MAPK signaling pathway during DN progression.

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PPARβ/δ accelerates bone regeneration in diabetic mellitus by enhancing AMPK/mTOR pathway-mediated autophagy

Stem Cell Research & Therapy ◽

10.1186/s13287-021-02628-8 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Miao Chen ◽

Dian Jing ◽

Rui Ye ◽

Jianru Yi ◽

Zhihe Zhao

Keyword(s):

Bone Regeneration ◽

Osteogenic Differentiation ◽

Bone Healing ◽

High Glucose ◽

Diabetic Rats ◽

Diabetic Patients ◽

Regulatory Effect ◽

Compound C ◽

High Glucose Condition ◽

Glucose Condition

Abstract Background Diabetic patients are more vulnerable to skeletal complications. Peroxisome proliferators-activated receptor (PPAR) β/δ has a positive regulatory effect on bone turnover under physiologic glucose concentration; however, the regulatory effect in diabetes mellitus has not been investigated yet. Herein, we explored the effects of PPARβ/δ agonist on the regeneration of diabetic bone defects and the osteogenic differentiation of rat bone marrow mesenchymal stem cells (rBMSCs) under a pathological high-glucose condition. Methods We detected the effect of PPARβ/δ agonist on osteogenic differentiation of rBMSCs in vitro and investigated the bone healing process in diabetic rats after PPARβ/δ agonist treatment in vivo. RNA sequencing was performed to detect the differentially expressed genes and enriched pathways. Western blot was performed to detect the autophagy-related protein level. Laser confocal microscope (LSCM) and transmission electron microscope (TEM) were used to observe the formation of autophagosomes. Results Our results demonstrated that the activation of PPARβ/δ can improve the osteogenic differentiation of rBMSCs in high-glucose condition and promote the bone regeneration of calvarial defects in diabetic rats, while the inhibition of PPARβ/δ alleviated the osteogenic differentiation of rBMSCs. Mechanistically, the activation of PPARβ/δ up-regulates AMPK phosphorylation, yielding mTOR suppression and resulting in enhanced autophagy activity, which further promotes the osteogenic differentiation of rBMSCs in high-glucose condition. The addition of AMPK inhibitor Compound C or autophagy inhibitor 3-MA inhibited the osteogenesis of rBMSCs in high-glucose condition, suggesting that PPARβ/δ agonist promotes osteogenic differentiation of rBMSCs through AMPK/mTOR-regulated autophagy. Conclusion In conclusion, our study demonstrates the potential role of PPARβ/δ as a molecular target for the treatment of impaired bone quality and delayed bone healing in diabetic patients for the first time.

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ERM Complex, a Therapeutic Target for Vascular Leakage Induced by Diabetes

Current Medicinal Chemistry ◽

10.2174/0929867328666210526114417 ◽

2021 ◽

Vol 28 ◽

Author(s):

Olga Simó-Servat ◽

Hugo Ramos ◽

Patricia Bogdanov ◽

Marta García-Ramírez ◽

Jordi Huerta ◽

...

Keyword(s):

High Glucose ◽

Therapeutic Target ◽

Vascular Leakage ◽

Cell Permeability ◽

Diabetic Patients ◽

Diabetic Mice ◽

Erm Proteins ◽

High Glucose Condition ◽

Tnf Α ◽

Glucose Condition

Background: Ezrin, radixin, and moesin (the ERM complex) interact directly with membrane proteins regulating their attachment to actin filaments. ERM protein activation modifies cytoskeleton organization and alters the endothelial barrier function, thus favoring vascular leakage. However, little is known regarding the role of ERM proteins in diabetic retinopathy (DR). Objective: This study aimed to examine whether overexpression of the ERM complex exists in db/db mice and its main regulating factors. Methods: 9 male db/db mice and 9 male db/+ aged 14 weeks were analyzed. ERM proteins were assessed by western blot and by immunohistochemistry. Vascular leakage was determined by the Evans blue method. To assess ERM regulation, HRECs were cultured in a medium containing 5.5 mM D-glucose (mimicking physiological conditions) and 25 mM D-glucose (mimicking hyperglycemia that occurs in diabetic patients). Moreover, treatment with TNF-α, IL-1β, or VEGF was added to a high glucose condition. The expression of ERM proteins was quantified by RT-PCR. Cell permeability was evaluated by measuring movements of FITC-dextran. Results: A significant increase of ERM in diabetic mice in comparison with non-diabetic mice was observed. A high glucose condition alone did not have any effect on ERM expression. However, TNF-α and IL-1β induced a significant increase in ERM proteins. Conclusion: The increase of ERM proteins induced by diabetes could be one of the mechanisms involved in vascular leakage and could be considered as a therapeutic target. Moreover, the upregulation of the ERM complex by diabetes is induced by inflammatory mediators rather than by high glucose itself.

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High Glucose Treatment Limits Drosha Protein Expression and Alters AngiomiR Maturation in Microvascular Primary Endothelial Cells via an Mdm2-dependent Mechanism

Cells ◽

10.3390/cells10040742 ◽

2021 ◽

Vol 10 (4) ◽

pp. 742

Author(s):

Brian Lam ◽

Emmanuel Nwadozi ◽

Tara L. Haas ◽

Olivier Birot ◽

Emilie Roudier

Keyword(s):

Skeletal Muscle ◽

Endothelial Cells ◽

Protein Expression ◽

High Glucose ◽

Micro Rna ◽

Microvascular Endothelium ◽

High Glucose Condition ◽

Mdm2 Expression ◽

Dicer Protein ◽

Glucose Condition

Diabetes promotes an angiostatic phenotype in the microvascular endothelium of skeletal muscle and skin. Angiogenesis-related microRNAs (angiomiRs) regulate angiogenesis through the translational repression of pro- and anti-angiogenic genes. The maturation of micro-RNA (miRs), including angiomiRs, requires the action of DROSHA and DICER proteins. While hyperglycemia modifies the expression of angiomiRs, it is unknown whether high glucose conditions alter the maturation process of angiomiRs in dermal and skeletal muscle microvascular endothelial cells (MECs). Compared to 5 mM of glucose, high glucose condition (30 mM, 6–24 h) decreased DROSHA protein expression, without changing DROSHA mRNA, DICER mRNA, or DICER protein in primary dermal MECs. Despite DROSHA decreasing, high glucose enhanced the maturation and expression of one angiomiR, miR-15a, and downregulated an miR-15a target: Vascular Endothelial Growth Factor-A (VEGF-A). The high glucose condition increased Murine Double Minute-2 (MDM2) expression and MDM2-binding to DROSHA. Inhibition of MDM2 prevented the effects evoked by high glucose on DROSHA protein and miR-15a maturation in dermal MECs. In db/db mice, blood glucose was negatively correlated with the expression of skeletal muscle DROSHA protein, and high glucose decreased DROSHA protein in skeletal muscle MECs. Altogether, our results suggest that high glucose reduces DROSHA protein and enhances the maturation of the angiostatic miR-15a through a mechanism that requires MDM2 activity.

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High Glucose Attenuates Cardioprotective Effects of Glucagon-Like Peptide-1 Through Induction of Mitochondria Dysfunction via Inhibition of β-Arrestin-Signaling

Frontiers in Physiology ◽

10.3389/fphys.2021.648399 ◽

2021 ◽

Vol 12 ◽

Author(s):

Xietian Pan ◽

Chengxiang Li ◽

Haokao Gao

Keyword(s):

Signaling Pathway ◽

High Glucose ◽

Ischemia Reperfusion Injury ◽

Ischemia Reperfusion ◽

Diabetic Patients ◽

High Glucose Condition ◽

Normal Glucose ◽

Cardioprotective Effects ◽

Glucagon Like Peptide 1 ◽

Glucose Condition

An increased vulnerability has been detected after ischemia/reperfusion injury in cardiomyocytes in diabetic patients. Glucagon-like peptide-1 (GLP-1) has been proven to have a notable cardioprotective effect in cardiomyocytes. However, in diabetic patients, the cardioprotective effects of GLP-1 are compromised, which is called GLP-1 resistance. β-arrestin is one of the two main downstream effectors of GLP-1 and β-arrestin signaling pathway exerts cardioprotective effects upon activation of GLP-1R. Our hypothesis is that the increased vulnerability of cardiomyocytes in diabetic patients is partly due to disruption of the β-arrestin signaling pathway. To test this, we analyzed cardiomyocyte viability and survival in high glucose and normal glucose condition after hypoxia/reoxygenation injury in vitro, additional GLP-1 was used to determine whether β-arrestin signaling pathway was involved. We also investigated the role of mitochondrial dysfunction in GLP-1 resistance. Our results showed that cardioprotective effects of GLP-1 were reduced in high glucose cultured H9C2 cells compared to normal glucose cultured H9C2, verifying the existence of GLP-1 resistance in high glucose cultured H9C2 cells. Further study suggested that β-arrestin plays a key role in GLP-1 resistance: β-arrestin expression is notably downregulated in high glucose condition and cardioprotective effects of GLP-1 can be diminished by downregulation of β-arrestin in normal glucose condition while upregulation of β-arrestin can restore cardioprotective effects of GLP-1 in high glucose condition. Then we explore how β-arrestin affects the cardioprotective effects of GLP-1 and found that β-arrestin exerts cardioprotective effects by improving mitochondria quality control via the PI3K/Akt signaling pathway. Thus, our study found out a new mechanism of GLP-1 resistance of cardiomyocytes in high glucose conditions that impaired β-arrestin expression, caused mitochondria dysfunction and eventually cell death. Our study provided a new perspective in treating myocardial ischemia/reperfusion injury in diabetic patients.

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Abstract 563: Enhanced Renal Angiotensinogen Upregulation Under High Glucose Condition is Mediated by TRPA1

Hypertension ◽

10.1161/hyp.62.suppl_1.a563 ◽

2013 ◽

Vol 62 (suppl_1) ◽

Author(s):

Ryousuke Satou ◽

Andrea Zsombok ◽

Kayoko Miyata ◽

Akemi Katsurada ◽

Andrei V Derbenev

Keyword(s):

Oxidative Stress ◽

Hydrogen Peroxide ◽

Diabetic Nephropathy ◽

High Glucose ◽

Glucose Medium ◽

High Glucose Medium ◽

High Glucose Condition ◽

Normal Glucose ◽

Intrarenal Ras ◽

Glucose Condition

Clinical and basic studies have revealed that the renin-angiotensin system (RAS) plays crucial roles in the development of kidney injury during progression of type 1 diabetes mellitus (T1DM) whereas the activity of systemic RAS is suppressed in T1DM. Thus, intrarenal RAS is increasingly being recognized as an important factor in the development of diabetic nephropathy. We recently demonstrated expression of TRPA1, an oxidative stress-sensitive calcium channel, in renal proximal tubular cells (PTC) and TRPA1 mediates angiotensinogen (AGT) augmentation in the cells. However, regulation and function of the intrarenal TRPA1 in T1DM have not been elucidated. Therefore, this study was performed to evaluate intrarenal TRPA1 expression and its contribution to AGT augmentation under high glucose condition. Streptozotocin (STZ, 200 mg/kg) was intraperitoneally injected to male C57BL/6 mice. Intrarenal TRPA1 expression, kidney AGT, urinary AGT, urinary 8-isoprostane, a maker of oxidative stress, and blood glucose levels were determined one week after STZ injection. Furthermore, contribution of TRPA1 to AGT regulation was tested by using mouse PTC which was cultured with normal (1.0 g/L) or high (4.5 g/L) glucose media. Under these conditions, PTC was treated with 100 μM hydrogen peroxide. Blood glucose and urinary 8-isoprostane levels were higher in the STZ-treated mice compared with control mice. TRPA1 mRNA levels were augmented in the kidney of STZ-treated mice (2.26 ± 0.2, ratio to control mice) accompanied by increases in kidney AGT and urinary AGT levels. TPRA1 upregulation was also observed in PTC which was cultured with the high glucose medium (1.55 ± 0.11, ratio to the normal glucose). Hydrogen peroxide stimulated AGT expression in PTC. The AGT elevation was enhanced in the cultured cells with the high glucose (1.52 ± 0.13-fold in the normal glucose medium vs. 1.97 ± 0.29-fold in the high glucose medium). Treatment with a TRPA1 antagonist attenuated the enhanced AGT augmentation in PTC. These results demonstrate that intrarenal TRPA1 mediates enhanced AGT upregulation under high glucose condition and suggest that intrarenal TRPA1 is implicated in intrarenal RAS activation leading to the development of T1DM-associated diabetic nephropathy.

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HADC regulates the diabetic vascular endothelial dysfunction by targetting MnSOD

Bioscience Reports ◽

10.1042/bsr20181042 ◽

2018 ◽

Vol 38 (5) ◽

Cited By ~ 2

Author(s):

Qian Hou ◽

Ke Hu ◽

Xiaofeng Liu ◽

Jiao Quan ◽

Zehao Liu

Keyword(s):

Oxidative Stress ◽

Endothelial Dysfunction ◽

High Glucose ◽

Vascular Dysfunction ◽

Diabetic Patients ◽

Vascular Endothelial ◽

Mechanism Study ◽

Histone Deacetylase 2 ◽

High Glucose Condition ◽

Glucose Condition

Vascular dysfunction is a common result of diabetes in humans. However, the mechanism underlying diabetic vascular dysfunction is not fully understood. Here in the present study, we showed that the histone deacetylase 2 (HDAC2) promoted the endothelial dysfunction induced by diabetes. The expression and activity of HDAC2 were up-regulated in vascular endothelial cells (ECs) from diabetic patients and mice. The expression of HDAC2 was also increased by high glucose stress in isolated human ECs. HDAC2 knockdown repressed the proliferation rate and promoted high glucose-induced apoptosis of ECs, which was associated with the activation of apoptotic pathways (Bcl-2, Caspase 3, and Bax). By contrast, HDAC2 overexpression led to opposing results. Significantly, we observed that HDAC2 regulated the accumulation of reactive oxygen species (ROS) induced by high glucose in ECs, which accounted for the effects of HDAC2 on proliferation and apoptosis because antioxidants, N-acetyl-l-cysteine (NAC) or MnTBAP treatment blocked the effects of HDAC2 on apoptosis of ECs under high glucose condition. Mechanism study revealed that HDAC2 bound to the promoter of MnSOD and repressed the expression of MnSOD by regulating the level of acetylated H3K9 and H3K27, which led to the promotion of oxidative stress and contributed to the function of HDAC2 in ECs under high glucose condition. Altogether, our evidence demonstrated that HDAC2-MnSOD signaling was critical in oxidative stress and proliferation as well as the survival of ECs under high glucose condition.

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Effects of High Glucose Concentration on Pericyte-Like Differentiated Human Adipose-Derived Mesenchymal Stem Cells

International Journal of Molecular Sciences ◽

10.3390/ijms22094604 ◽

2021 ◽

Vol 22 (9) ◽

pp. 4604

Author(s):

Giuliana Mannino ◽

Anna Longo ◽

Florinda Gennuso ◽

Carmelina Daniela Anfuso ◽

Gabriella Lupo ◽

...

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Mrna Expression ◽

Inflammatory Cytokines ◽

High Glucose ◽

Angiogenic Factors ◽

Diabetic Patients ◽

Ros Production ◽

Migration Ability ◽

And Migration

A pericyte-like differentiation of human adipose-derived mesenchymal stem cells (ASCs) was tested in in vitro experiments for possible therapeutic applications in cases of diabetic retinopathy (DR) to replace irreversibly lost pericytes. For this purpose, pericyte-like ASCs were obtained after their growth in a specific pericyte medium. They were then cultured in high glucose conditions to mimic the altered microenvironment of a diabetic eye. Several parameters were monitored, especially those particularly affected by disease progression: cell proliferation, viability and migration ability; reactive oxygen species (ROS) production; inflammation-related cytokines and angiogenic factors. Overall, encouraging results were obtained. In fact, even after glucose addition, ASCs pre-cultured in the pericyte medium (pmASCs) showed high proliferation rate, viability and migration ability. A considerable increase in mRNA expression levels of the anti-inflammatory cytokines transforming growth factor-β1 (TGF-β1) and interleukin-10 (IL-10) was observed, associated with reduction in ROS production, and mRNA expression of pro-inflammatory cytokines interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α), and angiogenic factors. Finally, a pmASC-induced better organization of tube-like formation by retinal endothelial cells was observed in three-dimensional co-culture. The pericyte-like ASCs obtained in these experiments represent a valuable tool for the treatment of retinal damages occurring in diabetic patients.

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