MicroRNA‑182‑5p contributes to the protective effects of thrombospondin 1 against lipotoxicity in INS‑1 cells

  • Authors:
    • Ying Liu
    • Jiayue Dong
    • Bo Ren
  • View Affiliations

  • Published online on: October 19, 2018     https://doi.org/10.3892/etm.2018.6883
  • Pages: 5272-5279
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Abstract

The dysfunction of beta cells serves an important role in the pathogenesis of type 2 diabetes mellitus (T2DM). An improved understanding of the molecular mechanisms underlying beta cell mass and failure will be useful for identifying novel approaches toward preventing and treating this disease. Recent studies have indicated that free fatty acids (FFAs) can cause beta cell dysfunction. In the present study, palmitate (Pal) was used as a FFA and its functions on cell viability and apoptosis were detected. MTT assay and flow cytometry were used and the results revealed that incubation of INS‑1 cells with Pal significantly decreased cell viability and increased cell apoptosis. However, a co‑incubation with thrombospondin 1 (THBS‑1) protected the cells against Pal‑induced toxicity. Numerous studies have demonstrated that microRNAs (miRs) are involved in fatty acid‑induced beta cell dysfunction. Various studies have reported that miR‑182‑5p is associated with a number of diseases, including cancer, heart disease, and leukemia. However, to the best of our knowledge miR‑182‑5p has never been reported to be associated with diabetes. In the present study, miR‑182‑5p, which is predicted to target the 3'‑untranslated region (UTR) of THBS‑1, was detected using reverse transcription‑quantitative polymerase chain reaction in INS‑1 cells in response to Pal. miR‑182‑5p was significantly increased in Pal‑treated cells compared with the control cells. Furthermore, miR‑182‑5p mimics significantly decreased cell viability and increased Pal‑induced apoptosis in INS‑1 cells. However, cell viability was increased and Pal‑induced apoptosis was decreased in cells that were treated with miR‑182‑5p inhibitors. The present findings also revealed that overexpression of THBS‑1 counteracted the effect of miR‑182‑5p on cell viability and apoptosis. These results suggested that miR‑182‑5p is involved in the mechanism of THBS 1 on the modulation of beta cell survival.

Introduction

Diabetes is one of the most important non-communicable diseases worldwide (1). According to the International Diabetes Federation statistics, the number of patients with diabetes worldwide in 2011 reached 370 million, and this number will be nearly 550 million by 2030. T2DM is a progressive disease caused by insulin resistance and/or beta cell dysfunction, which results in relative insulin deficiency. The reduction in beta cell mass and beta cell dysfunction contribute toward the pathological process (2). A previous study has suggested that beta cell dysfunction in patients with clinical manifestations of T2DM may have begun 15 years previously (2). Increased levels of circulating free fatty acids (FFAs) have been indicated to cause defective beta cell proliferation and increased beta cell apoptosis. Therefore, lipotoxicity has an important role in the pathogenesis of T2DM (3).

THBS-1 is an extracellular matrix-bound factor and was reported as the first naturally occurring inhibitor of angiogenesis (4). It was also revealed to be involved in other processes, including regulation of extracellular matrix function, blood clot formation and the immune response (57). Recently, accumulating research has suggested that THBS-1 is associated with T2DM and beta cell function (810).

MicroRNAs (miRs) are 19 to 22-nucleotide noncoding RNAs that can regulate cell survival, cell function, apoptosis and differentiation by suppressing the transcription of mRNA (1113). Currently, extensive research has suggested that miRs are involved in fatty acid-induced beta cell dysfunction (1416). miR-182-5p, which has been predicted to target THBS-1, has been confirmed to participate in the progression of various diseases, including cancer (17), ischemia-reperfusion injury (18) and leukemia (19). However, to the best of our knowledge miR-182-5p has not been reported to be associated with T2DM or beta cell function.

The present study aimed to identify whether palmitate (Pal) impacted the viability and apoptosis of INS-1 cells. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was performed to assess the THBS-1 mRNA expression levels in Pal-treated cells compared with the control cells. Subsequent MTT and flow cytometry assays were also performed. The present study provided an insight into whether miR-182-5p may participate in the protective effects of THBS-1 against lipotoxicity in INS-1 cells, and whether it may be a novel biomarker for the diagnosis and treatment of T2DM.

Materials and methods

Cell culture

Rat INS-1 cells (Shanghai Fushan Industrial Co., Ltd, Shanghai, China) were cultured in RPMI 1640 medium (Gibco, Thermo Fisher Scientific, Inc., Waltham, MA, USA), containing 11 mmol/l glucose, 10% fetal bovine serum (Gibco, Thermo Fisher Scientific, Inc.), 1 mmol/l sodium pyruvate, 2 mmol/l glutamine, 10 mmol/l HEPES, 55 µmol/l beta-mercaptoethanol, 100 IU/ml penicillin and 100 µg/ml streptomycin at 37°C containing 5% CO2.

Vector constructions and miR transfection

The coding sequence of THBS-1 was cloned into a pcDNA3 vector, prior to the construction of the THBS-1 expression plasmid, pcDNA3-THBS-1. The enhanced green fluorescent (EGFP) coding region from the pEGFP-N2 vector was cloned into pcDNA3 to form pcDNA3-EGFP, and the wild-type or mutant-type THBS-1 3′-UTR was amplified and cloned into the pcDNA3-EGFP vector. A total of 20 µM miR-182-5p mimics, inhibitors and the corresponding control (miR-NC) (Shanghai GenePharma Co., Ltd., Shanghai, China) were transfected into INS-1 cells using Lipofectamine 2000 reagent (Invitrogen; Thermo Fisher Scientific, Inc.) according to the manufacturer's instructions. The sequences used were as follows: miR-182-5p, 5′-UUUGGCAAUGGUAGAACUCACACCG-3′; miR-182-5p inhibitor, 5′-CGGUGUGAGUUCUACCAUUGCCAAA-3′; and miR-NC, 5′-UUGUACUACACAAAAGUAGUC-3′. Following 24 h of transfection, the subsequent experiments were carried out.

FFA treatment

Pal (Sigma-Aldrich; Merck KGaA, Darmstadt, Germany), which was used as the FFA, was dissolved into 0.01 mol/l NaOH to a concentration of 100 mmol/l. Subsequently, the solution was diluted in RPMI-1640 medium and mixed with 0.5% FFA-free bovine serum albumin (Gibco; Thermo Fisher Scientific, Inc.) for 30 min at 55°C. INS-1 cells were cultured and exposed at 37°C to Pal (0.25, 0.5, and 1.0 mol/l) for 8, 24 or 48 h. The control cells were treated with equivalent concentrations of NaOH.

MTT assay

INS-1 cells were transfected with plasmids, miR-182-5p mimics or inhibitors. A total of 8×103 cells were placed into 96-well culture plates for 24 h at 37°C. Cells were incubated with different concentrations of Pal (0.25, 0.5 and 1.0 mmol/l) for a further 24 h. A total of 10 µl of MTT (5 mg/ml; Sigma-Aldrich; Merck KGaA) was added into the wells and the cells were incubated for 4 h at 37°C. The culture plate was centrifuged at 225 × g for 5 min at room temperature and the supernatant was removed. Following this, 150 µl dimethyl sulfoxide (Sigma-Aldrich; Merck KGaA) was added to the medium of each well and the plate was placed on a shaker in the dark for 20 min at room temperature. Finally, the absorbance value of each well at the wavelength of 570 nm (A570) was measured.

Apoptosis assay

An annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) apoptosis detection kit (BD Biosciences Medical Devices Shanghai Co., Ltd., Shanghai, China) was used to detect INS-1 cell apoptosis. Cells were collected following centrifugation at room temperature for 10 min at 225 × g, washed with phosphate buffered saline and resuspended in binding buffer (300 µl). A total of 5 µl annexin V-FITC solution was added, followed by incubation at room temperature for 15 min in the dark. Finally, 5 µl PI was added to the cell suspension. A flow cytometer (BD Bioscience, Shanghai, China) was used to analyze cell apoptosis. Among the analyzed cells, FITC/PI cells represented healthy living cells, FITC+/PI cells indicated early apoptotic cells and FITC+/PI+ cells represented necrosis and late apoptotic cells.

RT-qPCR analysis

TRIzol reagent (Invitrogen; Thermo Fisher Scientific, Inc) and the mirVana miRNA Isolation Kit (Ambion; Thermo Fisher Scientific.) were used to extract total RNA and miRs. The oligo-dT or stem-loop reverse transcriptase primers were used to obtain cDNA. PCR was performed using the SYBR Premix Ex Taq (Takara Biotechnology Co., Ltd., Dalian, China) to detect the expression levels of THBS-1 mRNA and miR-182-5p. β-actin and U6 were used as the corresponding controls. The primer sequences used in the present study are indicated in Table I. The reaction conditions used were as follows: 94°C for 5 min, followed by 40 cycles of 94°C for 1 min, 56°C for 1 min and 72°C for 1 min. The relative expression levels were calculated using the 2−ΔΔCq method (20).

Table I.

Primer sequences used in reverse transcription-quantitative polymerase chain reaction.

Table I.

Primer sequences used in reverse transcription-quantitative polymerase chain reaction.

NamePrimer sequence
miR-182-5p RT primer 5′-GTCGTATCCAGTGCAGGGTCCGAGGTGCACTGGATACGACAAGTGTG-3′
U6 RT primer 5′-GTCGTATCCAGTGCAGGGTCCGAGGTATTCGCACTGGATACGACAAAATATGGAAC-3′
miR-182-5p forward 5′-TCGGGTTTGGCAATGGTAGAAC-3′
U6 forward 5′-TGCGGGTGCTCGCTTCGGCAGC-3′
Reverse 5′-CCAGTGCAGGGTCCGAGGT-3′
THBS1 forward 5′-TTTGCTGCGTTTGTGGAA-3′
THBS1 reverse 5′-GAGGAGGTATCTGTAATGC-3′
β-actin forward 5′-CGTGACATTAAGGAGAAGCTG-3′
β-actin reverse 5′-CTAGAAGCATTTGCGGTGGAC-3′
Western blot analysis

Cells were transfected with miR-182-5p mimics or inhibitors. Protein concentration was determined by bicinchoninic acid assay. Cell protein samples were obtained using radioimmunoprecipitation assay lysis buffer (Beijing BLKW Biotechnology Co., Ltd, Beijing, China), separated by 8% SDS-PAGE (20 µg/lane) and transferred to nitrocellulose membranes. Following this, samples were blocked with 5% skimmed milk for 2 h at room temperature. Membranes were incubated with anti-THBS-1 antibody (ab88529; 1:200; Abcam, Cambridge, MA, USA) and anti-GAPDH antibody (ab8245; 1:500; Abcam) overnight at 4°C. Subsequently, the horseradish peroxidase-conjugated corresponding immunoglobulin G (ab7090; 1:1,000; Abcam) was added for incubation at room temperature for 2 h. The protein expression level was assessed using an enhanced chemiluminescence regent (Santa Cruz Biotechnology, Inc., Dallas, TX, USA). GAPDH was used as the corresponding control. The relative protein expression levels were determined relative to GAPDH.

miR target prediction

The possible miRs that can target THBS-1 were predicted using miRanda (http://microrna.sanger.ac.uk/), TargetScan (genes.mit.edu/targetscan) and PicTar (http://pictar.mdc-berlin.de).

EGFP reporter assay

INS-1 cells were transfected with miR-182-5p mimics, miR-182-5p inhibitor and reporter vectors bearing either THBS 1 3′UTR wild-type or THBS 1 3′UTR mutant-type. An F-4500 fluorescence spectrophotometer (Hitachi, Ltd., Tokyo, Japan) was used to detect the intensities of fluorescence. Red fluorescent protein (RFP) expression vector was used as a reporter control. Relative EGFP fluorescence intensity was normalized to the RFP fluorescence intensity.

Statistical analysis

Data were analyzed using GraphPad Prism 6.0 statistical software (GraphPad Software, Inc., La Jolla, CA, USA) and were presented as the mean ± standard deviation. A two-tailed Student's t-test was performed to compare differences between two groups. One-way analysis of variance followed by Tukey's post hoc test was used to compare differences between groups. P<0.05 was considered to indicate a statistically significant difference.

Results

FFA induces cell toxicity in INS-1 cells

To determine the effect of FFA on cell viability and apoptosis, Pal was selected as the FFA. INS-1 cells were cultured and incubated with different concentrations of Pal. The subsequent MTT assay revealed that Pal increased INS-1 cell viability following 8 h of incubation and significantly decreased cell viability at 24 or 48 h of incubation. These effects occurred in a dose-dependent manner (Fig. 1A). Results of flow cytometry suggested that incubations for 24 or 48 h with Pal (0.5 mmol/l) increased the cell apoptosis rate by 2.8- or 3.2- fold, respectively (Fig. 1B). These results suggested that Pal can induce cell toxicity in INS-1 cells.

THBS-1 protects INS-1 cells from Pal-induced cell toxicity

It has previously been indicated that THBS-1 is associated with beta cell function and T2DM (8). To further explore whether THBS-1 is involved in the lipotoxicity of INS-1 cells, the THBS-1 expression plasmid, pcDNA3-THBS-1, was transfected into INS-1 cells, which were incubated with different concentrations of Pal. The efficiency of pcDNA3-THBS 1 was detected by RT-qPCR (Fig. 2A). Data indicated that THBS-1 significantly reversed the changes in cell viability and apoptosis that were induced by Pal following incubation for 24 or 48 h. These data suggested that THBS 1 protects INS-1 cells from Pal-induced lipotoxicity (Fig. 2).

Bioinformatics prediction

Previous studies have reported that a number of miRs are involved in regulating the occurrence and progression of T2DM (1216). To further determine whether THBS-1 is regulated by other molecules in its actions on the lipotoxicity of INS-1 cells, miRanda, Targetscan and PicTar were used to predict the possible miRs that may target THBS-1. The prediction results from these bioinformatics software were combined and several miRs were selected. Among these, miR-182-5p was of interest. To the best of our knowledge, miR-182-5p has not been reported to be associated with T2DM. A total of 7 bases from the 3′-UTR region of THBS-1 were identified to be complementary to the seed sequence of miR-182-5p (Fig. 3). Considering these observations, miR-182-5p was selected for further study.

miR-182-5p directly targets the 3′-UTR of THBS-1 and downregulates its expression

miRs are well known to exert their function by targeting the 3′-UTR of target genes. To determine whether or not miR-182-5p directly targets THBS-1, EGFP reporter analysis was performed. Reporter vectors bearing either wild-type or mutant-type THBS-1 3′-UTR were co-transfected with miR-182-5p mimics or inhibitor into INS-1 cells. The efficiency of miR-182-5p mimics and inhibitors were detected by RT-qPCR (Fig. 4A). The fluorescence intensity of each group was measured and the results indicated miR-182-5p decreased the intensity of wild-type 3′-UTR by ~69%, while the intensity of the wild-type 3′-UTR was increased by ~1.8-fold when miR-182-5p was inhibited. However, the intensity of mutant-type 3′-UTR was not significantly affected when miR-182-5p was overexpressed or inhibited (Fig. 4B).

The majority of miRs can negatively regulate their target genes by targeting the 3′-UTR. To further determine the regulation mode of miR-182-5p in the present study, RT-qPCR and western blot analysis were performed. The results demonstrated that miR-182-5p mimics decreased the mRNA and protein expression level of THBS-1 by ~65 or 70%, respectively, whereas the expression of THBS-1 was significantly increased by ~1.8- or 2.5-fold when miR-182-5p was inhibited (Fig. 4C and D). These results suggested that miR-182-5p can directly bind to the 3′-UTR of THBS-1 and negatively regulate its expression at mRNA and protein levels.

Upregulation of miR-182-5p in Pal-treated cells

The aforementioned results indicated that miR-182-5p could target THBS-1 in INS-1 cells. To further investigate whether or not miR-182-5p was involved in Pal-induced cytotoxicity, RT-qPCR was used to detect the alteration in miR-182-5p expression in response to Pal. INS-1 cells were incubated with Pal (0.5 mmol/l) for 24 h. The results of RT-qPCR indicated that miR-182-5p expression level was increased by 1.7-fold in Pal-amended cells compared with the control cells (Fig. 5). This result suggested that miR-182-5p may serve a role in Pal-induced lipotoxicity in INS-1 cells.

miR-182-5p promotes Pal-induced lipotoxicity of INS-1 cells by directly targeting THBS-1

To determine the exact effect of miR-182-5p on Pal-induced cytotoxicity in INS-1 cells, MTT and flow cytometric analysis were also performed. As indicated in Fig. 6, miR-182-5p mimics significantly decreased THBS-1 cell viability (Fig. 6A and B), while increasing Pal-induced apoptosis in INS-1 cells following incubation with Pal (Fig. 6C). However, the cell viability was increased (Fig. 6A and B) and Pal-induced apoptosis was decreased (Fig. 6C) in cells that were transfected with miR-182-5p inhibitors. To further confirm that miR-182-5p affected Pal-induced lipotoxicity by regulating THBS-1 directly, a rescue experiment was performed. miR-182-5p and THBS-1 expression plasmid without the 3′-UTR (pcDNA3-THBS 1) were co-transfected into INS-1 cells. As expected, overexpression of THBS-1 counteracted the effect of miR-182-5p on cell viability (Fig. 6D and E) and apoptosis (Fig. 6F). Taken together, these results indicated that miR-182-5p affects Pal-induced lipotoxicity by directly regulating THBS-1.

Discussion

More than 415 million people worldwide are estimated to have diabetes, with an ever-increasing morbidity and mortality (21). It is reported that ~95% of all diabetes cases are T2DM (22,23). T2DM is a complex disorder characterized by insulin resistance and defects in pancreatic beta cell function. Numerous experiments have reported that the failure of beta cells to increase mass and function is central in T2DM (24,25). Nutrients are essential for the maintenance of beta cell function and mass. However, an excess of nutrients such as glucose or FFAs can induce injurious effects on beta cell mass and function that contribute toward the progressive loss of functional beta cell mass (26).

In the present study, Pal, which was used as an FFA, increased INS-1 cell viability after a short exposure time (8 h). However, Pal significantly decreased cell viability following 24 or 48 h of incubation. These results indicated that FFAs exert stimulatory effects acutely, but inhibitory effects chronically on beta cell survival. The subsequent flow cytometry assay suggested that Pal (0.5 mmol/l) markedly increased the cell apoptosis rate following 24 or 48 h of exposure.

THBS-1 is a multifunctional glycoprotein released from various types of cell and exerts its diverse biological effects through binding to extracellular matrix proteins and cell surface receptors (8). Recent studies have suggested that THBS-1 is involved in the pathogenesis of insulin resistance and is important for beta cell function (27,28). However, the exact effect of THBS 1 on FFA-induced lipotoxicity is not thoroughly understood. The present study revealed that THBS-1 significantly reversed the changes of cell viability and apoptosis that were induced by Pal exposure. These results suggested that THBS-1 can protect INS-1 cells from Pal-induced lipotoxicity.

Additionally, the precise mechanisms of THBS-1 that caused lipotoxic effects, and whether or not THBS-1 is regulated by other molecules, were further investigated. In recent years, it has been determined that an increasing number of miRs participate in various pathological processes (2931), including lipotoxicity (32,33). Subsequent experiments focused on the potential participation of miRs in Pal-induced lipotoxicity. To begin with, combining the prediction results of three bioinformatic analyses, miR-182-5p was selected. To further identify whether or not miR-182-5p directly targets THBS-1, an EGFP reporter assay was performed, which revealed that miR-182-5p could negatively regulate the intensity of wild-type 3′-UTR. However, miR-182-5p could not affect the intensity of mutant-type 3′-UTR. Furthermore, miR-182-5p was indicated to negatively regulate the expression of THBS-1 at the mRNA and protein levels.

The aforementioned results have indicated that miR-182-5p could target THBS-1 in INS-1 cells. To further investigate whether or not miR-182-5p was involved in Pal-induced cytotoxicity, miR-182-5p expression was increased in Pal-amended cells and it was hypothesized that miR-182-5p may participate in Pal-induced lipotoxicity. The subsequent cell phenotype experiments suggested that miR-182-5p decreased cell viability and increased Pal-induced apoptosis in INS-1 cells that were incubated with Pal. To further confirm that THBS-1 was a direct functional target gene of miR-182-5p, a rescue experiment was designed. The results suggested that restoration of THBS-1 could counteract the effects of miR-182-5p on cell viability and apoptosis. Taken together, the results of the present study suggested that miR-182-5p affects Pal-induced cell viability and apoptosis by directly targeting THBS-1.

In conclusion, the present findings suggested that THBS-1 can protect INS-1 cells from FFA-induced lipotoxicity. Furthermore, miR-182-5p, which is able to directly target the 3′-UTR of THBS-1, can increase FFA-induced cytotoxicity. These findings demonstrated that miR-182-5p may be useful for identifying novel therapeutic approaches that may improve beta cell mass and function. Additionally, miR-182-5p may be a novel biomarker for the pathogenesis and progression of T2DM.

Acknowledgements

Not applicable.

Funding

No funding was received.

Availability of data and materials

The datasets used and/or analyzed during the present study are available from the corresponding author on reasonable request.

Authors' contributions

RB conceived and designed the study, acquired data and had an important role in interpreting the results. LY performed the experiments, contributed significantly to analysis and manuscript preparation. DJ assisted in performing the data analysis with constructive discussions. All authors read and approved the final manuscript.

Ethics approval and consent to participate

Not applicable.

Patient consent for publication

Not applicable.

Competing interests

The authors declare that they have no competing interests.

Glossary

Abbreviations

Abbreviations:

miRs

microRNAs

T2DM

type 2 diabetes

RT-qPCR

reverse transcription-quantitative polymerase chain reaction

FFAs

free fatty acids

References

1 

Camlio A, Cecilia SP, Valeria H and Marcos S: Analysis of the relationship between the referral and evolution of patients with type 2 diabetes mellitus. Int J Environ Res Public Health. 15:15342018. View Article : Google Scholar

2 

Mashitisho ML and Mashitisho BG: Early insulin therapy in patients with type 2 diabetes mellitus. J Endocrinol Metab Diab South Africa. 21:13–15. 2016. View Article : Google Scholar

3 

Gu J, Wei Q, Zheng H, Meng X, Zhang J and Wang D: Exendin-4 promotes survival of mouse pancreatic β-cell line in lipotoxic conditions, through the extracellular signal-related kinase 1/2 pathway. J Diabetes Res. 2016:52940252016. View Article : Google Scholar : PubMed/NCBI

4 

Good DJ, Polverini PJ, Rastinejad F, Le Beau MM, Lemons RS, Frazier WA and Bouck NP: A tumor suppressor-dependent inhibitor of angiogenesis is immunologically and functionally indistinguishable from a fragment of thrombospondin. Proc Natl Acad Sci USA. 87:6624–6628. 1990. View Article : Google Scholar : PubMed/NCBI

5 

Adams JC and Lawler J: The thrombospondins. Int J Biochem Cell Biol. 36:961–968. 2004. View Article : Google Scholar : PubMed/NCBI

6 

Savill J, Fadok V, Henson P and Haslett C: Phagocyte recognition of cells undergoing apoptosis. Immunol Today. 14:131–136. 1993. View Article : Google Scholar : PubMed/NCBI

7 

Ribeiro SM, Poczatek M, Schultz-Cherry S, Villain M and Murphy-Ullrich JE: The activation sequence of thrombospondin-1 interacts with the latency-associated peptide to regulate activation of latent transforming growth factor-beta. J Biol Chem. 274:13586–13593. 1999. View Article : Google Scholar : PubMed/NCBI

8 

Matsuo Y, Tanska M, Yamakage H, Sasaki Y, Muranaka K, Hata H, Ikai I, Shimatsu A, Inoue M, Chun TH and Satoh-Asahara N: Thrombospondin 1 as a novel biological marker of obesity and metabolic syndrome. Metabolism. 64:1490–1499. 2015. View Article : Google Scholar : PubMed/NCBI

9 

Delić D, Eisele C, Schmid R, Baum P, Wiech F, Gerl M, Zimdahl H, Pullen SS and Urquhart R: Urinary exosomal miRNA signature in type II diabetic nephropathy patients. PLoS One. 11:e01501542016. View Article : Google Scholar : PubMed/NCBI

10 

Olerud J, Mokhtari D, Johansson M, Christoffersson G, Lawler J, Welsh N and Carlsson PO: Thrombospondin-1: An islet endothelial cell signal of importance for β-cell function. Diabetes. 60:1946–1954. 2011. View Article : Google Scholar : PubMed/NCBI

11 

Fu X, Li Y, Alvero A, Li J, Wu Q, Xiao Q, Peng Y, Hu Y, Li X, Yan W, et al: MicroRNA-222-3p/GNAI2/AKT axis inhibits epithelial ovarian cancer cell growth and associates with good overall survival. Oncotarget. 7:80633–80654. 2016. View Article : Google Scholar : PubMed/NCBI

12 

Li Y, Zhang T, Zhou Y, Sun Y, Cao Y, Chang X, Zhu Y and Han X: A presenilin/Notch1 pathway regulated by miR-375, miR-30a, and miR-34a mediates glucotoxicity induced-pancreatic beta cell apoptosis. Sci Rep. 6:361362016. View Article : Google Scholar : PubMed/NCBI

13 

Chen X, Shi C, Wang C, Liu W, Chu Y, Xiang Z, Hu K, Dong P and Han X: The role of miR-497-5p in myofibroblast differentiation of LR-MSCs and pulmonary fibrogenesis. Sci Rep. 7:409582017. View Article : Google Scholar : PubMed/NCBI

14 

Lin X, Guan H, Huang Z, Liu J, Li H, Wei G, Cao X and Li Y: Downregulation of Bcl-2 expression by miR-34a mediates palmitate-induced Min6 cells apoptosis. J Diabetes Res. 2014:2586952014. View Article : Google Scholar : PubMed/NCBI

15 

Li Y, Xu X, Liang Y, Liu S, Xiao H, Li F, Cheng H and Fu Z: miR-375 enhances palmitate-induced lipoapoptosis in insulin-secreting NIT-1 cells by repression myotrophin (V1) protein expression. Int J Clin Exp Pathol. 3:254–264. 2010.PubMed/NCBI

16 

Lovis P, Roggli E, Laybutt DR, Gattesco S, Yang JY, Widmann C, Abderrahmani A and Regazzi R: Alterations in microRNA expression contribute to fatty acid-induced pancreatic beta-cell dysfunction. Diabetes. 57:2728–2736. 2008. View Article : Google Scholar : PubMed/NCBI

17 

Yao J, Xu C, Fang Z, Li Y, Liu H, Wang Y, Xu C and Sun Y: Androgen receptor regulated microRNA miR-182-5p promotes prostate cancer progression by targeting the ARRDC3/ITGB4 pathway. Biochem Biophys Res Commun. 474:213–219. 2016. View Article : Google Scholar : PubMed/NCBI

18 

Jiang W, Liu G and Tang W: MicroRNA-182-5p ameliorates liver ischemia-reperfusion injury by suppressing Toll-like receptor 4. Transplant Proc. 48:2809–2814. 2016. View Article : Google Scholar : PubMed/NCBI

19 

Blume CJ, Hotz-Wagenblatt A, Hüllein J, Sellner L, Jethwa A, Stolz T, Slabicki M, Lee K, Sharathchandra A, Benner A, et al: p53-dependent non-coding RNA networks in chronic lymphocytic leukemia. Leukemia. 29:2015–2023. 2015. View Article : Google Scholar : PubMed/NCBI

20 

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

21 

International Diabetes Federation. IDF diabetes atlas. 7:(Brussels). International Diabetes Federation. 2015.

22 

American Diabetes Association: Diagnosis and classification of diabetes mellitus. Diabetes Care. 36((Suppl 1)): S67–S74. 2013.PubMed/NCBI

23 

American Diabetes Association: Diagnosis and classification of diabetes mellitus. Diabetes Care. 37((Suppl 1)): S81–S90. 2014.PubMed/NCBI

24 

Prentki M and Nolan CJ: Islet beta cell failure in type 2 diabetes. J Clin Invest. 116:1802–1812. 2006. View Article : Google Scholar : PubMed/NCBI

25 

Hull RL, Kodama K, Utzschneider KM, Carr DB, Prigeon RL and Kahn SE: Dietary-fat-induced obesity in mice results in beta cell hyperplasia but not increased insulin release: Evidence for specificity of impaired beta cell adaptation. Diabetologia. 48:1350–1358. 2005. View Article : Google Scholar : PubMed/NCBI

26 

Poitout V, Amyot J, Semache M, Zarrouki B, Hagman D and Fontés G: Glucolipotoxicity of the pancreatic beta cell. Biochim Biophys Acta. 1801:289–298. 2010. View Article : Google Scholar : PubMed/NCBI

27 

Inoue M, Jiang Y, Barnes RH II, Tokunaga M, Martinez-Santibañez G, Geletka L, Lumeng CN, Buchner DA and Chun TH: Thrombospondin 1 mediates high-fat diet-induced muscle fibrosis and insulin resistance in male mice. Endocrinology. 154:4548–4559. 2013. View Article : Google Scholar : PubMed/NCBI

28 

Kong P, Gonzalez-Quesada C, Li N, Cavalera M, Lee DW and Frangogiannis NG: Thrombospondin-1 regulates adiposity and metabolic dysfunction in diet-induced obesity enhancing adipose inflammation and stimulating adipocyte proliferation. Am J Physiol Endocrinol Metab. 305:E439–E450. 2013. View Article : Google Scholar : PubMed/NCBI

29 

Terracciano D, Terreri S, de Nigris F, Costa V, Celin GA and Cimmino A: The role of a new class of long noncoding RNAs transcribed from ultraconserved regions in cancer. Biochim Biophys Acta Rev Cancer. 1868:449–495. 2017. View Article : Google Scholar : PubMed/NCBI

30 

Yin J, Ren W, Huang X, Li T and Yin Y: Protein restriction and cancer. Biochim Biophys Acta Rev Cancer. 1869:256–262. 2018. View Article : Google Scholar : PubMed/NCBI

31 

Zhi F, Shao N, Xue L, Xu Y, Kang X, Yang Y and Xia Y: Characteristic microRNA expression induced by δ-opioid receptor activation in the rat liver under prolonged hypoxia. Cell Physiol Biochem. 44:2296–2309. 2017. View Article : Google Scholar : PubMed/NCBI

32 

Cazanave SC, Mott JL, Elmi NA, Bronk SF, Masuoka HC, Charlton MR and Gores GJ: A role for miR-296 in the regulation of lipoapoptosis by targeting PUMA. J Lipid Res. 52:1517–1525. 2011. View Article : Google Scholar : PubMed/NCBI

33 

Han YB, Wang MN, Li Q, Guo L, Yang YM, Li PJ, Wang W and Zhang JC: MicroRNA-34a contributes to the protective effects of glucagon-like peptide-1 against lipotoxicity in INS-1 cell. Chin Med J (Engl). 125:4202–4208. 2012.PubMed/NCBI

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Liu Y, Dong J and Ren B: MicroRNA‑182‑5p contributes to the protective effects of thrombospondin 1 against lipotoxicity in INS‑1 cells . Exp Ther Med 16: 5272-5279, 2018
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Liu, Y., Dong, J., & Ren, B. (2018). MicroRNA‑182‑5p contributes to the protective effects of thrombospondin 1 against lipotoxicity in INS‑1 cells . Experimental and Therapeutic Medicine, 16, 5272-5279. https://doi.org/10.3892/etm.2018.6883
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Liu, Y., Dong, J., Ren, B."MicroRNA‑182‑5p contributes to the protective effects of thrombospondin 1 against lipotoxicity in INS‑1 cells ". Experimental and Therapeutic Medicine 16.6 (2018): 5272-5279.
Chicago
Liu, Y., Dong, J., Ren, B."MicroRNA‑182‑5p contributes to the protective effects of thrombospondin 1 against lipotoxicity in INS‑1 cells ". Experimental and Therapeutic Medicine 16, no. 6 (2018): 5272-5279. https://doi.org/10.3892/etm.2018.6883