MicroRNA‑532‑5p regulates oxidative stress and insulin secretion damage in high glucose‑induced pancreatic β cells by downregulating the expression levels of CCND1
- Authors:
- Published online on: September 10, 2021 https://doi.org/10.3892/mmr.2021.12433
- Article Number: 793
-
Copyright: © Zhong et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
Abstract
Introduction
Diabetes is a chronic disease involving metabolic disorders of sugar, protein and fat that is primarily caused by insufficient insulin in the body. β cells are endocrine cells that secrete insulin. Insulin secretion by islet β cells is mainly influenced by blood glucose levels (1,2). A sharp decrease in the number of islet β cells results in insufficient insulin secretion by cells, which in turn induces metabolic disorders of proteins, glucose and other substances. Among others, amino acids, hormones and hyperglycemia are all inducers of the abnormal function of islet β cells, among which hyperglycemia is the most important one (3,4). It is of great significance to investigate the underlying mechanism of damaged islet cells in a high glucose (HG) environment.
MicroRNAs (miRNAs/miRs) are an important regulator of numerous physiological and pathophysiological processes, and serve a key role in a number of biological processes, such as cell proliferation, differentiation, apoptosis and carcinogenesis (5). A previous study demonstrated that miRNAs promote insulin secretion and regulate insulin resistance by acting on multiple pathways, and abnormal miRNA expression may be the underlying pathogenesis of diabetes (6). It has been reported that miR-532-5p expression is downregulated in the plasma of patients with type 2 diabetes (7). miR-532-5p expression is downregulated in H9C2 cells exposed to hypoxia, and in the myocardium of rats with acute myocardial infarction, reducing the apoptosis of H9C2 cells induced by hypoxia (8). However, to the best of our knowledge, the specific role of miR-532-5p in diabetes has not been reported.
By querying the StarBase website, it was identified that miR-532-5p could target cyclin D1 (CCND1). CCND1, a member of the cyclin family, is a regulator of cyclin-dependent kinase. CCND1 expression has been reported to be upregulated in diabetic islets (9). Shen and Zhu (10) analyzed gene expression in type 2 diabetes and identified 124 upregulated differentially expressed genes, including CCND1, in the GSE15653 dataset, and these were associated with fatty acid and glucose metabolic pathways, and oxidation/reduction reactions, and may be involved in the development of obesity-related type 2 diabetes. In addition, in multiple myeloma, CCND1 can control the redox metabolism by producing reactive oxygen species (ROS) to disrupt redox balance (11). Furthermore, CCND1 silencing can damage the repair of DNA double-strand breaks, induce G0/G1 phase cell cycle arrest and inhibit ovarian cancer cell proliferation (12).
The apoptotic activation gene p53 induces cell cycle stagnation at the G0 stage, as well as apoptosis. A previous study demonstrated that p53 serves an important role in the initiation of apoptosis under different physiological conditions (13). Following treatment with HG, the expression levels of p53 in INS-1 cells increased in a concentration-dependent manner, and overexpression of p53 induces apoptosis and reduces insulin secretion (14). Adaptive EGF expression sensitizes pancreatic cancer cells to ionizing radiation via activation of the CCND1/p53/poly(ADP-ribose) polymerase (PARP) signaling pathway (15). Therefore, it was speculated that miR-532-5p may regulate oxidative stress and insulin secretion damage of pancreatic β cells induced by HG by regulating CCND1/p53.
The present study examined the regulatory effects of miR-532-5p on diabetes and explored the underlying mechanisms by inducing islet cells with HG. The present study provided a theoretical basis for the investigation of the underlying mechanisms of diabetes and potential drug targets.
Materials and methods
Cell lines and culture conditions
The β cell line of the pancreas (MIN6) was obtained from The Cell Bank of Type Culture Collection of the Chinese Academy of Sciences and incubated in DMEM (Gibco; Thermo Fisher Scientific, Inc.) supplemented with 10% FBS (Gibco; Thermo Fisher Scientific, Inc.) at 37°C with 5% CO2. In the present experiment, the normal glucose control (5 mM glucose, NG group) was set as a control group, the mannitol group (MA) was set up to exclude the osmotic pressure effects of HG on cells. The MIN6 cells were incubated in complete medium containing 25 mM glucose (final concentration in the medium) for 24 h, which was called the HG group (16). The HG + mimic-NC group was transfected with a mimic-NC and then induced with 25 mM glucose. The HG + miR-532-5p mimic group was transfected with the miR-532-5p mimic and then induced with 25 mM glucose. The HG + miR-532-5p mimic + pcDNA-NC group were transfected with miR-532-5p mimic and the pcDNA-NC plasmid, and were then induced with 25 mM glucose. The HG + miR-532-5p mimic + pcDNA-CCND1 group was transfected with miR-532-5p mimic and the pcDNA-CCND plasmid, and were then induced with 25 mM glucose.
Database selection and analysis
StarBase (http://starbase.sysu.edu.cn/) was used to identify the binding sites of miR-532-5p and CCND1.
MTT assay
Cells were seeded at a density of 1×104 cells/well in 96-well plates. Following treatment of the cells, 20 µl MTT solution (5 mg/ml; Gen-view Scientific, Inc.) was added to each well and cells were incubated at 37°C with 5% CO2 for 4 h. Subsequently, 100 µl DMSO was added to dissolve the formazan crystals at 37°C for 10 min. The optical density at 490 nm was measured the following day to determine the quantities of formazan formed by cleaving of MTT in living cells.
Reverse transcription-quantitative PCR (RT-qPCR)
Total RNA was extracted from cells using TRIzol® reagent (Invitrogen; Thermo Fisher Scientific, Inc.) and then reverse transcribed into cDNA using the First-Strand cDNA Synthesis kit (Invitrogen; Thermo Fisher Scientific, Inc.). The SuperScript™ III Platinum™ SYBR Green One-Step qRT-PCR kit (Invitrogen; Thermo Fisher Scientific, Inc.) was used according to the manufacturer's protocols as follows: 95°C for 10 min, 40 cycles of 95°C for 10 sec, 55°C for 10 sec and 72°C for 30 sec. Relative expression levels were calculated according to the 2−ΔΔCq method (17). The following primers were used: miR-532-5p forward, 5′-GCGCGCATGCCTTGAGTGTAG-3′ and reverse, 5′-ATCCAGTGCAGGGTCCGAGG-3′; CCND1 forward, 5′-ACGAAGGTCTGCGCGTGTT-3′ and reverse, 5′-CCGCTGGCCATGAACTACCT-3′; p53 forward, 5′-GGCCCACTTCACCGTACTAA-3′ and reverse, 5′-GTGGTTTCAAGGCCAGATGT-3′; U6 forward, 5′-CTCGCTTCGGCAGCACA-3′ and reverse, 5′-AACGCTTCACGAATTTGCGT-3′; Insulin1 forward, 5′-TAGTGACCAGCTATAATCAGAG-3′ and reverse, 5′-ACGCCAAGGTCTGAAGGTCC-3′; Insulin2 forward, 5′-CCCTGCTGGCCCTGCTCTT3-3′ and reverse, 5′-AGGTCTGAAGGTCACCTGCT-3′; and GAPDH forward, 5′-ACCACAGTCCATGCCATCAC-3′ and reverse, 5′-TCCACCACCCTGTTGCTGTA-3′. Insulin1 and Insulin2 were detected as transcripts of the insulin gene, which are markers of pancreatic β cells (18). GAPDH was used as the housekeeping gene for CCND1, Insulin1, Insulin2 and p53. U6 was used as the housekeeping gene for miR-532-5p.
Cell transfection
Cells (1×105 cells/well) were seeded into 6-well plates and cultured for 24 h at 37°C with 5% CO2. Cell transfection was performed when the cells reached 80% confluency. The miR-532-5p mimic and mimic-negative control (mimic-NC; Invitrogen; Thermo Fisher Scientific, Inc.) were transfected directly into cells. For overexpression of miR-532-5p, the cells were transfected with mimic at a final concentration of 25 nM for 48 h at 37°C. For pcDNA-NC (empty vector, Invitrogen; Thermo Fisher Scientific, Inc.) and pcDNA-CCND1, full length transcript of CCND1 was amplified from cDNA obtained from 293T cells by PCR using PrimeSTAR® HS DNA polymerase (Takara Bio, Inc.), and was transfected at a final concentration of 500 ng for 48 h at 37°C. The PCR amplification product was inserted into the KpnI and BamHI sites of the pcDNA vector (Invitrogen; Thermo Fisher Scientific, Inc.), which was termed pcDNA-CCND1. Transfection was performed using Lipofectamine® 2000 (Invitrogen; Thermo Fisher Scientific, Inc.) according to the manufacturer's protocol. Transfection efficiency was determined by RT-qPCR 48 h after transfection.
ELISA
Cytokine concentration normalized to total insulin was detected using ELISA kits (ELISA MAX™ Deluxe Set Human IGFALS; cat. no. 445904 BioLegend, Inc.) (16). After the cells were centrifuged (300 × g, 10 min, 37°C), cell-free supernatant was diluted to 100 µl (1:20) and incubated with the specific capture antibody and detection antibody. Measurements were made at an optical density of 450 nm.
ROS assay
ROS levels of cells were detected using the fluorescent probe 2′,7′-dichlorodihydrofluorescein diacetate (Sigma-Aldrich; Merck KGaA), which could be rapidly oxidized into the fluorescent 2′,7′-dichlorofluorescein (DCF) in the presence of intracellular ROS. Fluorescence was monitored with a laser scanning confocal microscope (Leica Microsystems GmbH) at 488 nm (magnification, ×200). The amount of ROS was quantified as the relative fluorescence intensity of DCF per cell in the scanned area.
TUNEL assay
A total of 3×104 cells were seeded in 24-well plates and incubated overnight. The cells were treated after fusion. Cells were fixed with 4% paraformaldehyde for 30 min at room temperature and then washed with PBS. Subsequently, 0.3% Triton X-100 in PBS was added and incubated for 5 min. The cells were collected and washed with PBS three times, and treated with 50 µl TUNEL assay solution (Roche Diagnostics GmbH) at 37°C in the dark for 60 min, followed by the addition of stop solution. Subsequently, cells were incubated with DAB solution and stained with hematoxylin and eosin for 5 min at room temperature according to the manufacturer's protocol. Stained apoptotic cells were visualized at ×20 magnification under an LSM 710 laser scanning confocal microscope (Carl Zeiss AG).
Western blotting
Cells were collected, lysed with RIPA lysis buffer (Beyotime Institute of Biotechnology) and incubated for 30 min on ice. Subsequently, proteins were detected using a BCA protein assay kit (Bio-Rad Laboratories, Inc.). A total of 40 µg protein was loaded onto 10% SDS-polyacrylamide gels to separate proteins, which were subsequently transferred to PVDF membranes. The membranes were blocked with 10% skimmed milk for 2 h at room temperature, followed by incubation overnight at 4°C with the following primary antibodies: Anti-Bax (1:1,000; cat. no. 14796S; Cell Signaling Technology, Inc.), anti-caspase-3 (1:1,000; cat. no. 700182; Thermo Fisher Scientific, Inc.), anti-cleaved caspase-3 (1:1,000; cat. no. PA5-17913; Thermo Fisher Scientific, Inc.), anti-Bcl-2 (1:1,000; cat. no. 15071S; Cell Signaling Technology, Inc.), anti-CCND1 (1:1,000; cat. no. MA5-14512; Thermo Fisher Scientific, Inc.), anti-P53 (1:1,000; cat. no. MA5-12557; Thermo Fisher Scientific, Inc.) and anti-GAPDH (1:1,000; cat. no. 5174S; Cell Signaling Technology, Inc.). Subsequently, the membranes were incubated with goat anti-rabbit horseradish peroxidase-conjugated IgG secondary antibodies (1:5,000; cat. nos. A32731 and A11032; Thermo Fisher Scientific, Inc.) at room temperature for 1 h. The signals were detected using enhanced chemiluminescence reagent (GE Healthcare), and ImageJ software (version 146; National Institutes of Health) was used to analyze the fold changes of protein levels.
Luciferase reporter assays
To validate the direct targeting of miR-532-5p, the 3′ untranslated region (3′UTR) of the putative target gene CCND1 was cloned into the psiCHECK2 vector (Promega Corporation), according to the manufacturer's instructions. The mutated or wild-type CCND1 cells were divided into mimic-NC + CCND1 group and miR-532-5p mimic + CCND1 group. Vectors containing the respective 3′UTRs were co-transfected with miRNA mimic (5′-CAUGCCUUGAGUGUAGGACCGU-3′) into cells (1×106 cells) at a final concentration of 500 ng for 48 h at 37°C using Lipofectamine 2000 according to the manufacturer's protocol. Mutations in each of the predicted target sites in CCND1 3′UTRs were generated by site-directed mutagenesis using the QuikChange II Site-Directed Mutagenesis kit (Agilent Technologies, Inc.) according to the manufacturer's protocols. Subsequently, the cells were washed with PBS and lysed with cell lysis buffer (Beyotime Institute of Biotechnology) after transfection. The luciferase activity was measured using a plate reader (BD Biosciences) and was normalized to Renilla luciferase activity (pRL-TK) using the Luc-Screen™ Extended-Glow Luciferase Reporter Gene Assay system (cat. no. T1033; Thermo Fisher Scientific, Inc.). All procedures were conducted according to the manufacturers' instructions. Luciferase gene plasmid (containing WT and Mut UTRs) was constructed by Thermo Fisher Scientific, Inc.
Statistical analysis
Data are presented as the mean ± standard deviation. Each experiment was repeated three times. SPSS version 19.0 software (IBM Corp.) was used to perform statistical analysis. Comparisons among multiple groups were analyzed using one-way ANOVA followed by Tukey's post hoc test. P<0.05 was considered to indicate a statistically significant difference.
Results
Overexpression of miR-532-5p improves the impaired functions of secreted insulin in HG-induced cells
The MTT assay results demonstrated that compared with that of cells in the NG and MA groups, the survival rate of cells in the HG group was decreased (Fig. 1A), and this was accompanied by a decrease in the expression levels of miR-532-5p (Fig. 1B). This suggested that miR-532-5p served an important role in HG-induced cells. Subsequently, miR-532-5p was overexpressed using the cell transfection technique, and transfection efficiency was measured by RT-qPCR (Fig. 2A). Furthermore, cells were divided into NG, MA, HG, HG + mimic-NC and HG + miR-532-5p mimic groups. ELISA was used to detect the functions of secreted insulin. Compared with those in the NG and MA groups, the insulin secretion levels in the HG group were decreased. Compared with those of cells in the HG + mimic-NC group, the insulin secretion levels of cells in the HG + miR-532-5p mimic group were increased (Fig. 2B). In addition, RT-qPCR was used to detect the gene transcription levels of Insulin1 and Insulin2, and the trend was consistent with that of the total insulin level (Fig. 2C and D). The results demonstrated that overexpression of miR-532-5p improved the impaired functions of secreted insulin in HG-induced cells.
Overexpression of miR-532-5p inhibits oxidative stress levels in HG-induced cells
Subsequently, the levels of ROS were detected. Compared with those in the NG and MA groups, the ROS levels in the HG group were increased. Additionally, following overexpression of miR-532-5p, the ROS levels were decreased compared with the HG + mimic-NC group (Fig. 3A and B). TUNEL staining was subsequently used to detect apoptosis. Compared with the NG and MA groups, the HG group exhibited an increased apoptosis rate (Fig. 4A), and this was accompanied by increased expression levels of Bax and cleaved caspase-3, and decreased expression levels of Bcl-2 (Fig. 4B). Additionally, compared with that of cells in the HG + mimic-NC group, the apoptosis rate of cells in the HG + miR-532-5p mimic group exhibited a significant decline, and this was accompanied by decreased expression levels of Bax and cleaved caspase-3, and increased expression levels of Bcl-2. The results revealed that overexpression of miR-532-5p could inhibit oxidative stress levels in HG-induced cells.
Overexpression of miR-532-5p downregulates CCND1 expression in HG-induced cells
StarBase was used to identify that miR-532-5p could target the 3′UTR of CCND1 (Fig. 5A). Additionally, a luciferase reporter assay was used to verify the targeted binding of miR-532-5p and CCND1. The results demonstrated that before CCND1 mutation, luciferase activity was significantly decreased in the miR-532-5p mimic + CCND1 group compared with that in the Vector + CCND1 group. After CCND1 mutation, the luciferase activity was not significantly altered between the Vector + CCND1 and miR-532-5p mimic + CCND1 groups (Fig. 5B). Expression levels of CCND1 in cells were detected following overexpression of miR-532-5p. As shown in Fig. 5C and D, compared with those in the NG and MA groups, the expression levels of CCND1 in the HG group were significantly increased. Compared with those in the HG + mimic-NC group, the expression levels of CCND1 in the HG + miR-532-5p mimic group were decreased.
Overexpression of miR-532-5p improves the impaired functions of secreted insulin and inhibits oxidative stress levels in HG-induced cells by downregulating CCND1 expression
Compared with those in the pcDNA-NC group, the expression levels of CCND1 in the pcDNA-CCND1 group were significantly increased, indicating successful overexpression (Fig. 6A and B). Subsequently, the cells were divided into NG, HG, HG + mimic-NC, HG + miR-532-5p mimic, HG + miR-532-5p mimic + pcDNA-NC and HG + miR-532-5p mimic + PCDNA-CCND1 groups. The secretion of insulin by the cells was detected using ELISA. Compared with those in the HG + miR-532-5p mimic + pcDNA-NC group, the total insulin levels in the HG + miR-532-5p mimic + pcDNA-CCND1 group were decreased (Fig. 6C), as were the levels of Insulin1 (Fig. 6D) and Insulin2 (Fig. 6E). Subsequently, ROS levels were detected, and the trend was the opposite of that detected for insulin (Fig. 7A and B). Apoptosis was detected using a TUNEL assay. Compared with that in the HG + miR-532-5p mimic + pcDNA-NC group, apoptosis in the HG + miR-532-5p mimic + pcDNA-CCND1 group increased (Fig. 7C and D), and this was accompanied by increased expression levels of Bax and cleaved caspase-3, and decreased expression levels of Bcl-2 (Fig. 7E). These results indicated that overexpression of miR-532-5p improved the impaired functions of secreted insulin and inhibited oxidative stress levels in HG-induced cells by downregulating CCND1 expression.
Overexpression of miR-532-5p regulates the expression levels of p53 in HG-induced cells by downregulating CCND1 expression
During the study, abnormal expression levels of p53 were observed in the cells (Fig. 8A and B). Compared with those in the NG group, the expression levels of p53 in the cells were increased following HG induction, whereas the expression levels of p53 in the cells were decreased following overexpression of miR-532-5p. Compared with that in the HG + miR-532-5p mimic + pcDNA-NC group, p53 expression in the HG + miR-532-5p mimic + pcDNA-CCND1 group was increased. Therefore, it was preliminarily concluded that overexpression of miR-532-5p regulated the expression levels of p53 in HG-induced cells by downregulating CCND1 expression.
Discussion
miRNAs have been considered as potential biomarkers of tissue-specific origin, which affect the occurrence and development of diabetes by participating in processes such as collective oxidative stress and the inflammatory response (19). miR-21-5p expression in extracellular vesicles is increased during inflammatory responses and serves as a biomarker for type 1 diabetes (20). Overexpression of miR-22 can reduce oxidative stress injury in diabetic cardiomyopathy via sirtuin 1 (21). miR-365 promotes diabetic retinopathy by inhibiting TIMP metallopeptidase inhibitor 3 and increasing oxidative stress (22). It has been reported that miR-532-5p expression is downregulated in patients with type 2 diabetes (6). In the present study, the expression levels of miR-532-5p were also markedly decreased in HG-induced MIN6 cells.
At present, studies on miR-532-5p are concerned with its role in other diseases. miR-532-5p expression is downregulated in H9C2 cells exposed to hypoxia, and in the myocardium of rats with acute myocardial infarction, to reduce the apoptosis of H9C2 cells (8). In addition, miR-532-5p expression is downregulated in LPS-induced inflammation of RAW264.7 macrophages (23). It has been revealed that miR-532-5p serves an important role in the inflammatory response (24), oxidative stress (25) and apoptosis (8). However, to the best of our knowledge, the role of miR-532-5p in diabetes has not yet been reported. In the present study, it was revealed that overexpression of miR-532-5p could increase the secretion of insulin in HG-induced cells. Furthermore, miR-532-5p could inhibit the levels of oxidative stress in cells, and could inhibit the apoptosis of HG-induced cells.
The targeted binding of miR-532-5p to CCND1 was identified using the StarBase website, and this targeting relationship was further verified in the present study using a luciferase reporter assay. Studies have reported that the expression of CCND1 in diabetes is upregulated (9,26). However, to the best of our knowledge, the specific role of CCND1 in diabetes has not yet been reported. In the present study, it was revealed that the expression levels of CCND1 were upregulated in HG-induced cells, and overexpression of miR-532-5p inhibited CCND1 expression, suggesting that miR-532-5p targeting CCND1 served a regulatory role in diabetes. It has been reported that inhibition of CCND1 expression can block the cell cycle and inhibit cell proliferation (27). Furthermore, in multiple myeloma, CCND1 can control redox metabolism by producing ROS to disrupt the redox balance (11). In the present study, upregulation of miR-532-5p was proposed to regulate insulin secretion, oxidative stress and apoptosis in HG-induced cells by downregulating CCND1 expression. The experimental step of knocking down the expression of CCND1 will be further explored in our future research.
During the experiment, it was revealed that overexpression of miR-532-5p regulated the expression levels of p53 in HG-induced MIN6 cells by downregulating CCND1 expression. A previous study reported that adaptive EGF expression sensitized pancreatic cancer cells to ionizing radiation by activating the CCND1/p53/PARP signaling pathway (15). Following glucose treatment, the expression levels of p53 increase in a concentration-dependent manner, and the upregulation of p53 induces apoptosis and reduces insulin secretion (14). Therefore, it was preliminarily concluded that miR-532-5p regulated oxidative stress and insulin secretion of pancreatic β cells induced by HG by downregulating CCND1 expression, which may be regulated via the modulation of p53 expression levels. Further experimental verification of this preliminary conclusion will be carried out in subsequent research.
Due to the length of the article, only in vitro experiments were performed and described in the present study. In addition, our research group is conducting the relevant in vivo experiments to further verify the conclusion obtained from these in vitro experiments.
In conclusion, the present study revealed that miR-532-5p regulated oxidative stress and insulin secretion in HG-induced pancreatic β cells by downregulating the expression levels of CCND1, which was involved in the regulation of p53 expression.
Acknowledgements
Not applicable.
Funding
No funding was received.
Availability of data and materials
The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.
Authors' contributions
ZZ and WS made substantial contributions to the conception and design of the study, and the acquisition of data. HC made substantial contributions to analysis and interpretation of data. ZZ and WS confirm the authenticity of all the raw data. All authors read and approved the final manuscript.
Ethics approval and consent to participate
Not applicable.
Patients consent for publication
Not applicable.
Competing interests
The authors declare that they have no competing interests.
References
Yang X, Zhang Y, Xu W, Deng R, Liu Y, Li F, Wang Y, Ji X, Bai M, Zhou F, et al: Potential role of Hsp90 in rat islet function under the condition of high glucose. Acta Diabetol. 53:621–628. 2016. View Article : Google Scholar : PubMed/NCBI | |
Zhao YC, Zhu J, Song GY and Li XS: Relationship between thioredoxin-interacting protein (TXNIP) and islet β-cell dysfunction in patients with impaired glucose tolerance and hypertriglyceridemia. Int J Clin Exp Med. 8:4363–4368. 2015.PubMed/NCBI | |
Nelson P, Smith N, Ciupe S, Zou W, Omenn GS and Pietropaolo M: Modeling dynamic changes in type 1 diabetes progression: Quantifying beta-cell variation after the appearance of islet-specific autoimmune responses. Math Biosci Eng. 6:753–778. 2009. View Article : Google Scholar : PubMed/NCBI | |
Dahan T, Ziv O, Horwitz E, Zemmour H, Lavi J, Swisa A, Leibowitz G, Ashcroft FM, In't Veld P, Glaser B and Dor Y: Pancreatic beta-cells express the fetal islet hormone gastrin in rodent and human diabetes. Diabetes. 66:426–436. 2017. View Article : Google Scholar : PubMed/NCBI | |
Saliminejad K, Khorram Khorshid HR, Soleymani Fard S and Ghaffari SH: An overview of microRNAs: Biology, functions, therapeutics, and analysis methods. J Cell Physiol. 234:5451–5465. 2019. View Article : Google Scholar : PubMed/NCBI | |
Jones A, Danielson KM, Benton MC, Ziegler O, Shah R, Stubbs RS, Das S and Macartney-Coxson D: MiRNA signatures of insulin resistance in obesity. Obesity (Silver Spring). 25:1734–1744. 2017. View Article : Google Scholar : PubMed/NCBI | |
Ortega FJ, Mercader JM, Moreno-Navarrete JM, Rovira O, Guerra E, Esteve E, Xifra G, Martínez C, Ricart W, Rieusset J, et al: Profiling of circulating microRNAs reveals common microRNAs linked to type 2 diabetes that change with insulin sensitization. Diabetes Care. 37:1375–1383. 2014. View Article : Google Scholar : PubMed/NCBI | |
Ma J, Zhang J, Wang Y, Long K, Wang X, Jin L, Tang Q, Zhu L, Tang G, Li X and Li M: MiR-532-5p alleviates hypoxia-induced cardiomyocyte apoptosis by targeting PDCD4. Gene. 675:36–43. 2018. View Article : Google Scholar : PubMed/NCBI | |
Taneera J, Fadista J, Ahlqvist E, Zhang M, Wierup N, Renström E and Groop L: Expression profiling of cell cycle genes in human pancreatic islets with and without type 2 diabetes. Mol Cell Endocrinol. 375:35–42. 2013. View Article : Google Scholar : PubMed/NCBI | |
Shen J and Zhu B: Integrated analysis of the gene expression profile and DNA methylation profile of obese patients with type 2 diabetes. Mol Med Rep. 17:7636–7644. 2018.PubMed/NCBI | |
Bustany S, Bourgeais J, Tchakarska G, Body S, Hérault O, Gouilleux F and Sola B: Cyclin D1 unbalances the redox status controlling cell adhesion, migration, and drug resistance in myeloma cells. Oncotarget. 7:45214–45224. 2016. View Article : Google Scholar : PubMed/NCBI | |
Zhong Q, Hu Z, Li Q, Yi T, Li J and Yang H: Cyclin D1 silencing impairs DNA double strand break repair, sensitizes BRCA1 wildtype ovarian cancer cells to olaparib. Gynecol Oncol. 152:157–165. 2019. View Article : Google Scholar : PubMed/NCBI | |
Pistritto G, Trisciuoglio D, Ceci C, Garufi A and D'Orazi G: Apoptosis as anticancer mechanism: Function and dysfunction of its modulators and targeted therapeutic strategies. Aging (Albany NY). 8:603–619. 2016. View Article : Google Scholar : PubMed/NCBI | |
Liu RX, Ma Y, Hu XL, Liao YP, Hu XN, He BC and Sun WJ: Pioglitazone/metformin adduct regulates insulin secretion and inhibits high glucose-induced apoptosis via p21-p53-MDM2 signaling in INS-1 cells. J Cell Biochem. 119:5449–5459. 2018. View Article : Google Scholar : PubMed/NCBI | |
Liu X, Chen H, Hou Y, Ma X, Ye M, Huang R, Hu B, Cao H, Xu L, Liu M, et al: Adaptive EGF expression sensitizes pancreatic cancer cells to ionizing radiation through activation of the cyclin D1/P53/PARP pathway. Int J Oncol. 54:1466–1480. 2019.PubMed/NCBI | |
Ruan D, Liu Y, Wang X, Yang D and Sun Y: MiR-149-5p protects against high glucose-induced pancreatic beta cell apoptosis via targeting the BH3-only protein BIM. Exp Mol Pathol. 110:1042792019. View Article : Google Scholar : PubMed/NCBI | |
Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods. 25:402–408. 2001. View Article : Google Scholar : PubMed/NCBI | |
Abouzaripour M, Pasbakhsh P, Atlasi N, Shahverdi AH, Mahmoudi R and Kashani IR: In vitro differentiation of insulin secreting cells from mouse bone marrow derived stage-specific embryonic antigen 1 positive stem cells. Cell J. 17:701–710. 2016.PubMed/NCBI | |
Ye D, Zhang T, Lou G, Xu W, Dong F, Chen G and Liu Y: Plasma miR-17, miR-20a, miR-20b and miR-122 as potential biomarkers for diagnosis of NAFLD in type 2 diabetes mellitus patients. Life Sci. 208:201–207. 2018. View Article : Google Scholar : PubMed/NCBI | |
Lakhter AJ, Pratt RE, Moore RE, Doucette KK, Maier BF, DiMeglio LA and Sims EK: Beta cell extracellular vesicle miR-21-5p cargo is increased in response to inflammatory cytokines and serves as a biomarker of type 1 diabetes. Diabetologia. 61:1124–1134. 2018. View Article : Google Scholar : PubMed/NCBI | |
Tang Q, Len Q, Liu Z and Wang W: Overexpression of miR-22 attenuates oxidative stress injury in diabetic cardiomyopathy via Sirt 1. Cardiovasc Ther. 36:2018. View Article : Google Scholar | |
Wang J, Zhang J, Chen X, Yang Y, Wang F, Li W, Awuti M, Sun Y, Lian C, Li Z, et al: MiR-365 promotes diabetic retinopathy through inhibiting Timp3 and increasing oxidative stress. Exp Eye Res. 168:89–99. 2018. View Article : Google Scholar : PubMed/NCBI | |
Cheng Y, Kuang W, Hao Y, Zhang D, Lei M, Du L, Jiao H, Zhang X and Wang F: Downregulation of miR-27a* and miR-532-5p and upregulation of miR-146a and miR-155 in LPS-induced RAW264.7 macrophage cells. Inflammation. 35:1308–1313. 2012. View Article : Google Scholar : PubMed/NCBI | |
Yan X, Zeng D, Zhu H, Zhang Y, Shi Y, Wu Y, Tang H and Li D: MiRNA-532-5p regulates CUMS-induced depression-like behaviors and modulates LPS-induced proinflammatory cytokine signaling by targeting STAT3. Neuropsychiatr Dis Treat. 16:2753–2764. 2020. View Article : Google Scholar : PubMed/NCBI | |
Cai X, Wang S, Hong L, Yu S, Li B, Zeng H, Yang X, Zhang P and Shao L: Long noncoding RNA taurine-upregulated gene 1 knockdown protects cardiomyocytes against hypoxia/reoxygenation-induced injury through regulating miR-532-5p/Sox8 axis. J Cardiovasc Pharmacol. 76:556–563. 2020. View Article : Google Scholar : PubMed/NCBI | |
Gurke J, Schindler M, Pendzialek SM, Thieme R, Grybel KJ, Heller R, Spengler K, Fleming TP, Fischer B and Navarrete Santos A: Maternal diabetes promotes mTORC1 downstream signalling in rabbit preimplantation embryos. Reproduction. 151:465–476. 2016. View Article : Google Scholar : PubMed/NCBI | |
Li N, Zeng J, Sun F, Tong X, Meng G, Wu C, Ding X, Liu L, Han M, Lu C and Dai F: p27 inhibits CDK6/CCND1 complex formation resulting in cell cycle arrest and inhibition of cell proliferation. Cell Cycle. 17:2335–2348. 2018. View Article : Google Scholar : PubMed/NCBI |