Tumor-suppressive action of miR-30a-5p in lung adenocarcinoma correlates with ABL2 inhibition and PI3K/AKT pathway inactivation

Introduction ABL2 contributes to the oncogenic potential of cancers, pointing to its inhibition as a possible strategy against malignant diseases. Bioinformatics prediction of upstream effector miR-30a-5p for ABL2 allowed us to hypothesize and then validate mechanistic actions of miR-30a-5p in lung adenocarcinoma (LUAD). Materials and methods The ABL2 expression in LUAD was analyzed in the TCGA data, clinical samples, and cell lines. The shRNA-mediated silencing of ABL2 was introduced to illustrate its effect on malignant phenotypes of LUAD cells. The binding affinity between ABL2 and miR-30a-5p was verified by luciferase activity and RNA pull-down assay. Ectopic expression, knockdown methods, and PI3K inhibitor LY294002 were used to investigate their effects on in vitro biological characteristics and in vivo tumor growth of LUAD cells. Using nude mouse lung adenocarcinoma in situ and brain metastasis models to validate the inhibitory effect of miR-30a-5p on LUAD by regulating the ABL2/PI3K/AKT signaling axis. Results High expression of ABL2 and poor expression of miR-30a-5p were noticed in LUAD tissues and cell lines. Importantly, miR-30a-5p was demonstrated to target and downregulate ABL2, subsequently inactivating the PI3K/AKT pathway. miR-30a-5p inhibited the malignant phenotypes of LUAD cells by inhibiting ABL2 expression and inactivating the PI3K/AKT pathway. For in vivo experiments, miR-30a-5p was substantiated to thwart tumor tumorigenesis by regulating the ABL2/PI3K/AKT axis. In addition, miR-30a-5p suppresses the occurrence and development of in situ lung cancer and brain metastasis via the ABL2/PI3K/AKT signaling pathway. Conclusion This study underscores the inhibitory role of miR-30a-5p in LUAD through the ABL2/PI3K/AKT axis, which may be a viable target for LUAD treatment (AU)

Tumor-suppressive action of miR-30a-5p in lung adenocarcinoma correlates with ABL2 inhibition and PI3K/AKT pathway inactivation.

INTRODUCTION: ABL2 contributes to the oncogenic potential of cancers, pointing to its inhibition as a possible strategy against malignant diseases. Bioinformatics prediction of upstream effector miR-30a-5p for ABL2 allowed us to hypothesize and then validate mechanistic actions of miR-30a-5p in lung adenocarcinoma (LUAD). MATERIALS AND METHODS: The ABL2 expression in LUAD was analyzed in the TCGA data, clinical samples, and cell lines. The shRNA-mediated silencing of ABL2 was introduced to illustrate its effect on malignant phenotypes of LUAD cells. The binding affinity between ABL2 and miR-30a-5p was verified by luciferase activity and RNA pull-down assay. Ectopic expression, knockdown methods, and PI3K inhibitor LY294002 were used to investigate their effects on in vitro biological characteristics and in vivo tumor growth of LUAD cells. Using nude mouse lung adenocarcinoma in situ and brain metastasis models to validate the inhibitory effect of miR-30a-5p on LUAD by regulating the ABL2/PI3K/AKT signaling axis. RESULTS: High expression of ABL2 and poor expression of miR-30a-5p were noticed in LUAD tissues and cell lines. Importantly, miR-30a-5p was demonstrated to target and downregulate ABL2, subsequently inactivating the PI3K/AKT pathway. miR-30a-5p inhibited the malignant phenotypes of LUAD cells by inhibiting ABL2 expression and inactivating the PI3K/AKT pathway. For in vivo experiments, miR-30a-5p was substantiated to thwart tumor tumorigenesis by regulating the ABL2/PI3K/AKT axis. In addition, miR-30a-5p suppresses the occurrence and development of in situ lung cancer and brain metastasis via the ABL2/PI3K/AKT signaling pathway. CONCLUSION: This study underscores the inhibitory role of miR-30a-5p in LUAD through the ABL2/PI3K/AKT axis, which may be a viable target for LUAD treatment.

Extracellular Vesicles Carrying RUNX3 Promote Differentiation of Dental Pulp Stem Cells.

BACKGROUND: This study aims to clarify the mechanism underlying dental pulp cells-extracellular vesicles (DPC-EVs) carrying runt-related transcription factor 3 (RUNX3) in mediating odontogenic differentiation of dental pulp stem cells (DPSCs) with the involvement of miR-30a-5p-regulated NOTCH1. METHODS: Extracellular vesicles (EVs) were isolated from human DPSCs, and identified using transmission electron microscopy, and nanoparticle tracking analysis. PBS, EVs, or EV inhibitor GW4869 was added to DPSCs for co-culture, whilst odontogenic differentiation was assessed in terms of ratio of mineralized nodules and expression odontoblast differentiation markers. Dual luciferase reporter gene assay and chromatin immunoprecipitation for binding relation among RUNX3, miR-30a-5p and NOTCH1were employed to evaluate their roles in odontogenic differentiation was determined. Animal experiment was established to confirm the effect of DPC-EVs-loaded RUNX3 on dental pulp. RESULTS: In vitro finding demonstrated that EVs delivered RUNX3 to DPSCs, thereby activated miR-30a-5p expression and inhibited NOTCH1 expression, which was reversed by addition of GW4869. RUNX3 upregulation promoted miR-30a-5p while miR-30a-5p targeted and inhibited NOTCH1. Silencing of RUNX3 in EVs decreased expression of those differentiation markers, downregulated miR-30a-5p and upregulated NOTCH1. CONCLUSION: DPSC-EVs can carry RUNX3 to the DPSCs, promote the transcription of miR-30a-5p, and then inhibit the expression of NOTCH1, and finally promote the odontogenic differentiation of DPSCs.

miR-30a-5p targets ITGA6 to inhibit oral squamous cell carcinoma progression.

OBJECTIVE: Nowadays, many studies focus on the relationship between microRNAs (miRs) and the development of oral squamous cell carcinoma (OSCC). Here, we broaden the understanding of miR-30a-5p in OSCC. METHODS: In silico analysis was implemented to screen differentially expressed genes in OSCC and the related upstream regulatory miR. OSCC SCC9 cells were manipulated with lentivirus-mediated miR-30a-5p mimic, oe-ITGA6 or sh-ITGA6 and LY294002 (the PI3K/AKT pathway inhibitor) for studying their roles in cell biological processes. Tumors were xenografted in nude mouse for in vivo mechanism verification. RESULTS: In silico analysis results depicted that ITGA6 was highly expressed in OSCC, and that miR-30a-5p was the upstream regulatory miR of ITGA6. miR-30a-5p was downregulated and ITGA6 was highly expressed in OSCC tissues and cells. miR-30a-5p targeted and downregulated ITGA6. ITGA6 promoted epithelial-mesenchymal transition, migration and invasion in OSCC cells by activating PI3K/AKT pathway. miR-30a-5p could suppress the in vivo growth and metastasis of OSCC by inhibiting the ITGA6/PI3K/AKT axis. CONCLUSION: Taken together, miR-30a-5p prevents OSCC progression by inhibiting PI3K/AKT pathway through inhibition of ITGA6 expression.

[Retracted] miR­30a­5p inhibits the proliferation, migration and invasion of melanoma cells by targeting SOX4.

Following the publication of this paper, it was drawn to the Editor's attention by a concerned reader that the tumour images shown in Fig. 3B were strikingly similar to data appearing in different form in another article written by different authors at different research institutes. Owing to the fact that the contentious data in the above article were already under consideration for publication prior to its submission to Molecular Medicine Reports, the Editor has decided that this paper should be retracted from the Journal. The authors were asked for an explanation to account for these concerns, but the Editorial Office did not receive a reply. The Editor apologizes to the readership for any inconvenience caused. [Molecular Medicine Reports 18: 2492­2498, 2018; DOI: 10.3892/mmr.2018.9166].

MicroRNA­30a­5p regulates cypermethrin-induced apoptosis of Sertoli cells by targeting KLF9 in vitro.

Cypermethrin (CYP) has been identified as one kind of endocrine-disrupting chemicals (EDCs) to induce male reproduction damage. This study aimed to investigate the effects and mechanisms of miR-30a-5p on CYP induced apoptosis of TM4 mouse Sertoli cells in vitro. In the present study, 0 µM, 10 µM, 20 µM, 40 µM and 80 µM CYP were used to treat TM4 cells for 24 h. The apoptosis of TM4 cells, the expression level of miR-30a-5p, the protein expressions and the interaction between miR-30a-5p and KLF9 were detected by flow cytometry, quantitative Real-Time PCR, Western blot and luciferase reporter assays. CYP induced apoptosis of TM4 cells, inhibited expression of miR-30a-5p in TM4 cells, and overexpression of miR-30a-5p partially recovered CYP induced cells apoptosis. Furthermore, KLF9 was a potential downstream target of miR-30a-5p predicted by publicly available databases. KLF9 expression level in TM4 cells was significantly elevated after treatment with CYP, and the induction was inhibited by miR-30a-5p mimics transfection. Meanwhile, dual-luciferase reporter assay demonstrated that miR-30a-5p directly targeted KLF9-3'UTR. Moreover, in the presence of CYP, the apoptosis regulator p53 expression was also increased in TM4 cells. Overexpression miR-30a-5p or down-regulation of KLF9 both attenuated the induction of CYP on p53 expression. Overall, the present study demonstrated that miR-30a-5p regulated CYP induced TM4 cells apoptosis by targeting KLF9/p53 axis.

Cardioprotective effects of circ_0002612 in myocardial ischemia/reperfusion injury correlate with disruption of miR-30a-5p-dependent Ppargc1a inhibition.

INTRODUCTION: Novel mechanistic insights into the effects of circular RNAs (circRNAs) on the physiology and pathology of cardiovascular diseases are under increasingly active investigation. This study defined the cardioprotective role and mechanistic actions of circ_0002612 in myocardial ischemia/reperfusion injury (MI/RI). METHODS: MI/RI was induced in mice by ligation of the left anterior descending (LAD) artery followed by reperfusion, and the in vitro model was established in cultured cardiomyocytes under hypoxia/reoxygenation (H/R) conditions. Interaction among circ_0002612, miR-30a-5p, Ppargc1a, and NLRP3 was predicted by bioinformatics analysis and further experimentally identified. Gain- and loss-of-function experiments were performed to evaluate the effect of the circ_0002612/miR-30a-5p/Ppargc1a/NLRP3 axis on the cardiac function and myocardial infarction of I/R-injured mice, as well as viability and apoptosis of H/R-challenged cardiomyocytes. RESULTS: In the myocardial tissues of MI/RI mice, miR-30a-5p was negatively correlated with circ_0002612 or Ppargc1a, but circ_0002612 was positively correlated with the expression of Ppargc1a. circ_0002612 competitively bound to miR-30a-5p to release expression of its target gene Ppargc1a. circ_0002612 promoted cardiomyocyte viability while suppressing the apoptosis by impairing the miR-30a-5p-mediated inhibition of Ppargc1a. Additionally, Ppargc1a inhibited the expression of NLRP3 and consequently facilitated cardiomyocyte proliferation while suppressing cell apoptosis. By inhibiting the expression of NLRP3, circ_0002612 protected mice from MI/RI. CONCLUSION: Overall, this study demonstrates the cardioprotective role of circ_0002612 against MI/RI, which may be a viable target for MI/RI.

MicroRNA-30a-5p Promotes Chronic Heart Failure in Rats by Targeting Sirtuin-1 to Activate the Nuclear Factor-κB/NOD-Like Receptor 3 Signaling Pathway.

OBJECTIVE: MicroRNA-30a-5p (miR-30a-5p) has been identified as a marker of heart failure; however, its functional mechanisms in chronic heart failure (CHF) remain unknown. We aim to investigate the role of miR-30a-5p targeting sirtuin-1 (SIRT1) in myocardial remodeling in CHF via the nuclear factor-κB/NOD-like receptor 3 (NF-κB/NLRP3) signaling pathway. METHODS: CHF rat models were established using aortic coarctation. The expression of miR-30a-5p, SIRT1, and the NF-κB/NLRP3 signaling pathway-related factors in CHF rats was determined. The CHF rats were then respectively treated with altered miR-30a-5p or SIRT1 to explore their roles in cardiac function, myocardial function, inflammatory response, pathological changes, and cardiomyocyte apoptosis. The binding relation between miR-30a-5p and SIRT1 was confirmed. RESULTS: MiR-30a-5p was upregulated whereas SIRT1 was downregulated in myocardial tissues of CHF rats. MiR-30a-5p inhibition or SIRT1 overexpression improved cardiac and myocardial function, and suppressed the inflammatory response, alleviated pathological changes and inhibited cardiomyocyte apoptosis in CHF rats. MiR-30a-5p targeted SIRT1 to regulate the NF-κB/NLRP3 signaling pathway. In CHF rats, downregulated miR-30a-5p and silenced SIRT1 could reverse the beneficial effects of downregulated miR-30a-5p. CONCLUSION: Inhibited miR-30a-5p inhibits CHF progression via the SIRT1-mediated NF-κB/NLRP3 signaling pathway.

MiR-30a-5p Promotes Vein Graft Restenosis by Inhibiting Cell Autophagy through Targeting ATG5.

OBJECTIVE: The aim of the study was to investigate the role of miR-30a-5p in restenosis of rats following vein grafting and the underlying mechanism. METHODS: Vein graft rat models were established and perfused with miR-30a-5p antagomir and si-ATG5 to probe the regulation of miR-30a-5p/ATG5 on intimal hyperplasia. Human saphenous vein smooth muscle cells (HSVSMCs) were obtained from the great saphenous veins of patients undergoing coronary artery bypass grafting and subjected to assays for autophagy, proliferation, and migration after gain and loss of function of miR-30a-5p and/or ATG5. The binding of miR-30a-5p and ATG5 was confirmed by RIP and dual-luciferase reporter assays. RESULTS: MiR-30a-5p expression gradually increased, ATG5 expression gradually decreased, and the intima was increasingly thickened during restenosis of grafted veins. Knockdown of miR-30a-5p in rats repressed the restenosis of vein grafts, while a deficiency of ATG5 reversed the effect of miR-30a-5p inhibition. Upregulation of miR-30a-5p enhanced the proliferation and migration of HSVSMCs and inhibited the autophagy, while downregulation of miR-30a-5p or overexpression of ATG5 showed opposite effects. ATG5 is a target gene of miR-30a-5p. CONCLUSION: MiR-30a-5p exacerbates vein graft restenosis by repressing ATG5 expression and inhibiting autophagy.

LINC00488 Induces Tumorigenicity in Retinoblastoma by Regulating microRNA-30a-5p/EPHB2 Axis.

OBJECTIVE: LINC00488 confers oncogenic activity in the progression of some tumors. Hence, the target of the study was about to specify LINC00488-mediated network in retinoblastoma (RB). METHODS: LINC00488 expression was tested in RB clinical tissues. siRNA targeting LINC00488 or miR-30a-5p mimic was introduced into RB cell line (Y79) to observe cellular biological functions. The relationship between LINC00488, miR-30a-5p and EPHB2 was verified. Afterward, the role of miR-30a-5p involved in RB through targeted regulation of EPHB2 was probed in vitro and in vivo. RESULTS: LINC00488 was induced in RB tissue and cells. LINC00488 knockdown or miR-30a-5p upregulation depressed the malignant activities of Y79 cells. LINC00488 could sponge miR-30a-5p that targeted EPHB2. EPHB2, and EPHB2 overexpression counteracted miR-30a-5p restoration-induced inhibition of Y79 cell development in vitro and in vivo. CONCLUSION: LINC00488 induces tumorigenicity in RB by binding to miR-30a-5p to target EPHB2, which may offer a new clue of RB treatment from an lncRNA-miRNA-mRNA network.

The urinary exosomes derived from premature infants attenuate cisplatin-induced acute kidney injury in mice via microRNA-30a-5p/ mitogen-activated protein kinase 8 (MAPK8).

Acute kidney injury (AKI) is a susceptible factor for chronic kidney disease (CKD). There is still a lack of effective prevention methods in clinical practice. This study investigated the protective effect of the urinary exosomes from premature infants on cisplatin-induced acute kidney injury. Here we isolated exosomes from the fresh urine of premature infants. A C57BL/6 mice model of cisplatin-induced acute kidney injury was given 100 ug urinary exosomes 24 hours after model establishment. The kidneys were collected for pathological examination and the evaluation of renal tubular damage and apoptosis. In the in vitro experiment, human renal cortex/proximal tubular cells (HK-2) were induced by cisplatin to assess the effect of the urine exosomes from premature infants. Exosome microRNA (miRNA) sequencing technology was applied to investigate the miRNAs enriched in exosomes and the dual-luciferase gene reporter system to examine the targeting relationship of the miRNA with target genes. The results indicated that the urinary exosomes could decrease the serum creatinine level and the apoptosis of renal tubular cells, and reduce mice mortality. In addition, miR-30a-5p was the most abundant miRNA in the exosomes. It protected HK-2 cells from cisplatin-induced apoptosis by targeting and down-regulating the mitogen-activated protein kinase 8 (MAPK8). Together, our findings identified that the urinary exosomes derived from premature infants alleviated cisplatin-induced acute kidney injury and inhibited the apoptosis of HK-2 via miR-30a-5p, which could target MAPK8. These findings implied that urinary exosomes from premature infants riched in miR-30a-5p might become a potential treatment for AKI.

AMPK-autophagy-mediated inhibition of microRNA-30a-5p alleviates morphine tolerance via SOCS3-dependent neuroinflammation suppression.

BACKGROUND: The development of morphine tolerance is a clinical challenge for managing severe pain. Studies have shown that neuroinflammation is a critical aspect for the development of analgesic tolerance. We found that AMPK-autophagy activation could suppress neuroinflammation and improve morphine tolerance via the upregulation of suppressor of cytokine signaling 3 (SOCS3) by inhibiting the processing and maturation of microRNA-30a-5p. METHODS: CD-1 mice were utilized for the tail-flick test to evaluate morphine tolerance. The microglial cell line BV-2 was utilized to investigate the mechanism of AMPK-autophagy-mediated posttranscriptional regulation of SOCS3. Proinflammatory cytokines were measured by western blotting and real-time PCR. The levels of SOCS3 and miRNA-processing enzymes were evaluated by western blotting, real-time PCR and immunofluorescence staining. RESULTS: Based on experimental verification, miRNA-30a-5p could negatively regulate SOCS3. The AMPK activators AICAR, resveratrol and metformin downregulated miRNA-30a-5p. We found that AMPK activators specifically inhibited the processing and maturation of miRNA-30a-5p in microglia by degrading DICER and AGO2 via autophagy. Furthermore, a miRNA-30a-5p inhibitor significantly improved morphine tolerance via upregulation of SCOS3 in mice. It markedly increased the level of SOCS3 in the spinal cord of mice and subsequently inhibited morphine-induced phosphorylation of NF-κB p65. In addition, a miRNA-30a-5p inhibitor decreased the levels of IL-1ß and TNF-α caused by morphine in microglia. CONCLUSION: AMPK-autophagy activation suppresses neuroinflammation and improves morphine tolerance via the upregulation of SOCS3 by inhibiting miRNA-30a-5p.

LncRNA HOX transcript antisense RNA mitigates cardiac function injury in chronic heart failure via regulating microRNA-30a-5p to target KDM3A.

Long noncoding RNA HOX transcript antisense RNA (HOTAIR) has been studied in multiple diseases, but the role of HOTAIR on chronic heart failure (CHF) through the regulation of microRNA (miR)-30a-5p and lysine-specific demethylase 3A (KDM3A) remains unexplored. This research aims to probe the effects of HOTAIR on CHF progression via modulating miR-30a-5p to target KDM3A. CHF mouse model was established by intraperitoneal injection of doxorubicin. The CHF mice were then injected with high-expressed HOTAIR, miR-30a-5p or KDM3A adenovirus vectors to determine the cardiac function, oxidative stress, inflammatory response, pathological change and cardiomyocyte apoptosis. HOTAIR, miR-30a-5p, KDM3A and Bcl-2/adenovirus E1B 19kDa interacting protein 3 (BNIP3) expression in CHF mice was detected. The binding relations among HOTAIR, miR-30a-5p and KDM3A were validated. HOTAIR and KDM3A were depleted, while miR-30a-5p was augmented in CHF mice. The elevated HOTAIR or KDM3A or could improve cardiac function, mitigate oxidative stress and pathological change, reduce inflammatory factor levels and cardiomyocyte apoptosis, while the increased miR-30a-5p exerted opposite effects. The miR-30a-5p elevation could reverse the effects of enriched HOTAIR, while BNIP3 reduction abrogated the effects of KDM3A on CHF. HOTAIR sponged miR-30a-5p that targeted KDM3A. HOTAIR improves cardiac injury in CHF via modulating miR-30a-5p to target KDM3A. This study provides novel therapeutic strategies for CHF treatment.

Curcumin inhibits prostate cancer progression by regulating the miR-30a-5p/PCLAF axis.

Curcumin has been shown to inhibit the growth of a variety of tumor cells. However, the biological functions of curcumin in prostate cancer (PCa) have not yet fully elucidated. The objective of the present study was to investigate the role of curcumin on the proliferation, migration, invasion and apoptosis of PCa cells and the underlying mechanism. Cell Counting Kit-8 and flow cytometry were used to detect the effects of curcumin at different concentrations on the proliferation and apoptosis of PCa cell lines, PC-3 and DU145. BrdU and Transwell assays, western blotting and reverse transcription-quantitative PCR were used to determine the effect of curcumin on cell proliferation, migration and invasion, apoptosis-related protein expression, and microRNA (miR)-30a-5p and PCNA clamp associated factor (PCLAF) expression, respectively. In addition, bioinformatics analysis and Pearson's correlation test were used to verify the relationship between miR-30a-5p and PCLAF. Curcumin was observed to impede the proliferation, migration and invasion of PCa cells, and promote their apoptosis in a time- and dose-dependent manner. Curcumin enhanced miR-30a-5p expression and inhibited PCLAF expression; furthermore, there was a negative correlation between miR-30a-5p and PCLAF expression in PCa tissues. In addition, transfection of miR-30a-5p inhibitors partially reversed the function of curcumin on cell proliferation, migration, invasion and apoptosis. Overall, curcumin suppressed the malignant biological behaviors of PCa cells by regulating the miR-30a-5p/PCLAF axis.

miR­30a­5p mitigates autophagy by regulating the Beclin­1/ATG16 pathway in renal ischemia/reperfusion injury.

Renal ischemia/reperfusion (I/R) injury often occurs during multiple organ failure and sepsis, and autophagy may serve a role in I/R injury. The aim of the present study was to explore the effect of microRNA (miR)­30a­5p on autophagy in renal I/R injury. miR­30a­5p and autophagy­related protein expression levels in renal I/R injury mouse models and in hypoxia/re­oxygenation HK­2 cell models were determined using reverse transcription­quantitative PCR or western blotting; apoptosis was analyzed using flow cytometry. The effects of miR­30a­5p, Beclin­1 and autophagy­related gene 16 (ATG16) on the proliferation and autophagy of HK­2 cells were analyzed through gain­ and loss­of­function studies. miR­30a­5p expression was significantly decreased after renal I/R injury in the in vivo and in vitro experiments. Renal I/R injury led to upregulated expression of autophagy­related proteins microtubule­associated protein light chain 3 (LC3)­â…¡ and Beclin­1, and downregulated expression of p62. miR­30a­5p overexpression decreased the number of LC3 punctae, decreased HK­2 cell apoptosis, increased p62 expression and decreased LC3­â…¡ and Beclin­1 expression. Inhibition of miR­30a­5p exhibited the opposite effects. A luciferase reporter assay demonstrated that miR­30a­5p targeted Beclin­1. Beclin­1 overexpression led to a significant increase in LC3­â…¡ expression and a decrease in p62 expression, as well as a significant increase in apoptosis. Beclin­1 overexpression also increased the protein expression level of ATG16. Downregulation of Beclin­1 decreased the expression of LC3­â…¡, elevated the p62 level and decreased apoptosis. ATG16 knockdown showed similar effects as those of Beclin­1 downregulation. In conclusion, miR­30a­5p was increased in renal I/R injury and might mitigate autophagy by regulating the Beclin­1/ATG16 pathway.

H3K14 hyperacetylation­mediated c­Myc binding to the miR­30a­5p gene promoter under hypoxia postconditioning protects senescent cardiomyocytes from hypoxia/reoxygenation injury.

Our previous study reported that microRNA (miR)­30a­5p upregulation under hypoxia postconditioning (HPostC) exert a protective effect on aged H9C2 cells against hypoxia/reoxygenation injury via DNA methyltransferase 3B­induced DNA hypomethylation at the miR­30a­5p gene promoter. This suggests that miR­30a­5p may be a potential preventative and therapeutic target for ischemic heart disease in aged myocardium. The present study aimed to investigate the underlying mechanisms of miR­30a­5p transcription in aged myocardium in ischemic heart disease. Cardiomyocytes were treated with 8 mg/ml D­galactose for 9 days, and then exposed to hypoxic conditions. Cell viability was determined using a cell viability assay. Expression levels of histone deacetylase 2 (HDAC2), LC3B­II/I, beclin­1 and p62 were detected via reverse transcription­quantitative PCR and western blotting. Chromatin immunoprecipitation­PCR and luciferase reporter assays were performed to evaluate the effect of c­Myc binding and activity on the miR­30a­5p promoter in senescent cardiomyocytes following HPostC. It was found that HPostC enhanced the acetylation levels of H3K14 at the miR­30a­5p gene promoter in senescent cardiomyocytes, which attributed to the decreased expression of HDAC2. In addition, c­Myc could positively regulate miR­30a­5p transcription to inhibit senescent cardiomyocyte autophagy. Mechanically, it was observed that increased H3K14 acetylation level exposed to romidepsin facilitated c­Myc binding to the miR­30a­5p gene promoter region, which led to the increased transcription of miR­30a­5p. Taken together, these results demonstrated that HDAC2­mediated H3K14 hyperacetylation promoted c­Myc binding to the miR­30a­5p gene promoter, which contributed to HPostC senescent cardioprotection.

Curcumol inhibits the viability and invasion of colorectal cancer cells via miR-30a-5p and Hippo signaling pathway.

MicroRNA-30a-5p (miR-30a-5p), which functions as a tumor suppressor, has been reported to be downregulated in colorectal cancer (CRC) tissues and to be associated with cancer invasion. However, the detailed regulatory mechanism of curcumol in the malignant progression of CRC remains unknown. MTT, Transwell, scratch, western blotting and reverse transcription-quantitative PCR assays were performed to examine how curcumol inhibited CRC cell viability, invasion and migration, and to detect the role of miR-30a-5p and curcumol in the invasion and Hippo signaling pathways of CRC cells. The present study revealed that miR-30a-5p expression was downregulated in human CRC tissues and cells. The results demonstrated that miR-30a-5p downregulation was accompanied by the inactivation of the Hippo signaling pathway, which was demonstrated to promote CRC cell viability, invasion and migration. Curcumol treatment was identified to increase miR-30a-5p expression and to activate the Hippo signaling pathway, which in turn inhibited the invasion and migration of CRC cells. Overexpression of miR-30a-5p enhanced the effects of curcumol on cell invasion and migration, and the Hippo signaling pathway in CRC cells. Furthermore, downregulation of miR-30a-5p reversed the effects of curcumol on cell invasion and migration, and the Hippo signaling pathway in CRC cells. These findings identified novel signaling pathways associated with miR-30a-5p and revealed the effects of curcumol on miR-30a-5p expression. Therefore, curcumol may serve as a potential therapeutic strategy to delay CRC progression.

MicroRNA-30a-5p silencing polarizes macrophages toward M2 phenotype to alleviate cardiac injury following viral myocarditis by targeting SOCS1.

Viral myocarditis (VMC) is a life-threatening disease characterized by severe cardiac inflammation generally caused by coxsackievirus B3 (CVB3) infection. Several microRNAs (miRNAs or miRs) are known to play crucial roles in the pathogenesis of VMC. The study aimed to decipher the role of miR-30a-5p in the underlying mechanisms of VMC pathogenesis. We first quantified miR-30a-5p expression in a CVB3-induced mouse VMC model. The physiological characteristics of mouse cardiac tissues were then detected by hematoxylin and eosin (HE) and Picrosirius red staining. We established the correlation between miR-30a-5p and SOCS1, using dual-luciferase gene assay and Pearson's correlation coefficient. The expression of inflammatory factors (IFN-γ, IL-6, IL-10, and IL-13), M1 polarization markers [TNF-α, inducible nitric oxide synthase (iNOS)], M2 polarization markers (Arg-1, IL-10), and myocardial hypertrophy markers [atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP)] was detected by RT-qPCR and Western blot analysis. miR-30a-5p was found to be highly expressed in VMC mice. Silencing of miR-30a-5p improved the cardiac function index and reduced heart weight-to-body weight ratio, myocardial tissue pathological changes and fibrosis degree, serological indexes, as well as proinflammatory factor levels, while enhancing anti-inflammatory factor levels in VMC mice. Furthermore, silencing of miR-30a-5p inhibited M1 polarization of macrophages while promoting M2 polarization in vivo and in vitro. SOCS1 was a target gene of miR-30a-5p, and the aforementioned cardioprotective effects of miR-30a-5p silencing were reversed upon silencing of SOCS1. Overall, this study shows that silencing of miR-30a-5p may promote M2 polarization of macrophages and improve cardiac injury following VMC via SOCS1 upregulation, constituting a potential therapeutic target for VMC treatment.NEW & NOTEWORTHY We found in this study that microRNA (miR)-30a-5p inhibition might improve cardiac injury following viral myocarditis (VMC) by accelerating M2 polarization of macrophages via SOCS1 upregulation. Furthermore, the anti-inflammatory mechanisms of miR-30a-5p inhibition may contribute to the development of new therapeutic strategies for VMC.

DDP-resistant ovarian cancer cells-derived exosomal microRNA-30a-5p reduces the resistance of ovarian cancer cells to DDP.

Exosomes carrying microRNAs (miRNAs) have been demonstrated to play critical roles in the regulation of development, growth and metastasis of cancer. Bioinformatic predictions identified differentially expressed SRY-box 9 (SOX9) in OC, and the regulatory miRNA miR-139-5p. Here, we aim to evaluate the function of exosomal miR-139-5p in the sensitivity of ovarian cancer (OC) cells to cis-diamminedichloroplatinum(II) (DDP). Expression pattern of miR-139-5p and SOX9 in ovarian cancer cells (SKOV3) and DDP-resistant cells (SKOV3/DDP) was identified using reverse transcription quantitative polymerase chain reaction and western blot analysis. The relationship between miR-139-5p and SOX9 was validated using a dual-luciferase reporter assay. SKOV3/DDP cell line was developed and introduced with miR-30a-5p mimic to analyse the effects of miR-30a-5p on resistance to DDP. The in vitro and in vivo effects of exosomal miR-30a-5p on resistance of SKOV3 cells to DDP were assessed in a co-culture system of exosomes and OC cells as well as in tumour-bearing nude mice. High expression of SOX9 and low expression of miR-30-5p were witnessed in OC. Furthermore, miR-30-5p, a downregulated miRNA in SKOV3/DDP cells, increased the rate of cell apoptosis and enhanced the sensitivity of SKOV3/DDP cells to DDP by targeting SOX9. Moreover, exosomes carrying miR-30a-5p were identified to sensitize SKOV3/DDP cells to DDP both in vitro and in vivo. These data together supported an important conclusion that DDP-resistant OC cell-derived exosomal miR-30a-5p enhanced cellular sensitivity to DDP, highlighting a potential strategy to overcome drug resistance.

MiR-30a-5p Inhibits Epithelial Mesenchymal Transition Function in Human Cervical Cancer Cells by Targeting Ubiquitin-specific Protease 22 / 中山大学学报(医学科学版)

@# 【Objective】To investigate the inhibitory effect and mechanism of microRNA-30a-5p（miR-30a-5p）on epithelial mesenchymal transition in cervical cancer Hela cells.【Methods】Hela cervical cancer cell lines were transfected with miR-30a-5p mimics or negative control mimic，respectively，as 30a-5p or NC group. Control group was established with untreated Hela cervical cancer cells. miR-30a-5p content in each group was detected by RT-PCR assay. Transwell assay was used to detect the invasion ability of the 3 groups. Western-blot assay was used to detect the expressions of N- cadherin，α-Catenin and ubiquitin specific processing peptidase 22（USP22）protein in the 3 groups. Prediction target genes of miR-30a-5p by bioinformatics methods. Antagonistic effect of USP22 over-expression on miR-30a-5p inhibi⁃tion of EMT was detected by western blot assay. The relationship between miR-30a-5p and USP22 was detected by dual luciferase assay. Subcutaneous transplantation tumor model established，and the effect of miR-30a-5p in vivo was ob⁃ served.【Results】The miR-30a-5p intracellular quantity in 30a-5p group Hela cells was up-regulated，and the expres⁃ sion level of miR-30a-5p was 853.82（862.26~843.11）times higher than that of Control group（P＜0.01）. The number of invasive cells in 30a-5p group was 8.17（8.32~8.03），which was significantly lower than that of Control group 62.33 （63.52~60.19）（P＜0.01）. USP22 may be the target gene of miR-30a-5p. In 30a-5p group，the intracellular quantity of N-cadherin protein was decreased，the intracellular quantity of α-Catenin protein was increased，and the intracellular quantity of USP22 protein was decreased. The intracellular quantity of N-cadherin protein in 30a-5p group was down-reg⁃ ulated，the intracellular quantity of α-Catenin protein was increased，and the intracellular quantity of USP22 protein was reduced. After USP22 over-expression，the intracellular quantity of N-cadherin protein in cervical cancer cells of 30a-5p group was up-regulated，and the intracellular quantity of α-Catenin protein were down-regulated. Dual luciferase assay showed that USP22 is a downstream target gene of miR-30a-5p（P＜0.01）. Subcutaneous transplantation tumors in 30a- 5p group were significantly smaller than those in Control group.【Conclusion】miR-30a-5p may inhibit the expression of EMT related protein through the downstream target gene USP22，and the epithelial mesenchymal transition function of cervical cancer Hela cells.