LncRNA MYLK antisense RNA 1 activates cell division cycle 42/Neutal Wiskott-Aldrich syndrome protein pathway via microRNA-101-5p to accelerate epithelial-to-mesenchymal transition of colon cancer cells.

Long noncoding RNA MYLK antisense RNA 1 (MYLK-AS1) is the crux in multiple diseases. Therefore, the purpose of this study was to investigate the possible mechanism of MYLK-AS1. A total of 62 colon cancer (CC) specimens and paired adjacent normal tissues were collected, and the expression of MYLK-AS1, microRNA (miR)-101-5p/cell division cycle 42 (CDC42) was detected. CC cell lines were transfected with MYLK-AS1, miR-101-5p, CDC42-related plasmids, and the biological functions and markers of epithelial-mesenchymal transition (EMT) were analyzed. The binding relationship between MYLK-AS1, miR-101-5p, and CDC42 was evaluated. In CC tissues and cell lines, MYLK-AS1 and CDC42 were highly expressed, and miR-101-5p was lowly expressed. Inhibition of MYLK-AS1 or upregulation of miR-101-5p can inhibit CC cell growth and EMT. miR-101-5p inhibited CDC42/N-wasp axis activation in CC cells by targeting CDC42. Knockdown of CDC42 or upregulation of miR-101-5p partially reversed the effects caused by upregulation of MYLK-AS1. MYLK-AS1, which is significantly upregulated in CC, may be a molecular sponge for miR-101-5p, and MYLK-AS1 promotes the activation of the CDC42/N-wasp axis in CC cells by targeting CDC42 through miR-101-5p, which in turn promotes tumor development. MYLK-AS1 may be a potential biomarker and target for CC therapy.

Aging impairs recovery from stress-induced depression in male rats possibly by alteration of microRNA-101 expression and Rac1/RhoA pathway in the prefrontal cortex.

Along with altering brain responses to stress, aging may also impair recovery from depression symptoms. In the present study, we investigated depressive-like behaviors in young and aged rats and assayed the levels of microRNA-101 (miR-101), Rac1/RhoA, PSD-95, and GluR1 in the prefrontal cortex (PFC) after stress cessation and after a recovery period. Young (3 months old) and aged (22 months old) male Wistar rats were divided into six groups; Young control (YNG), young rats received chronic stress for four weeks (YNG + CS), young rats received chronic stress for four weeks followed by a 6-week recovery period (YNG + CS + REC), Aged control (AGED), aged rats received chronic stress for four weeks (AGED + CS), and aged rats received chronic stress for four weeks followed by a 6-week recovery period (AGED + CS + REC). Stress-induced depression, evaluated by the sucrose preference test (SPT) and forced swimming test (FST), was yet observed after the recovery period in aged but not in young rats, which were accompanied by unchanged levels of miR-101, Rac1/RhoA, GluR1, and PSD-95 in the PFC of aged rats. These data suggested that impaired synaptic plasticity of glutamatergic synapses via the miR-101/Rac1/RhoA pathway may contribute to the delayed behavioral recovery after stress exposure observed in aging animals.

Aberrantly Expressed MicroRNAs in Cancer-Associated Fibroblasts and Their Target Oncogenic Signatures in Hepatocellular Carcinoma.

Cancer-associated fibroblasts (CAFs) contribute to tumor progression, and microRNAs (miRs) play an important role in regulating the tumor-promoting properties of CAFs. The objectives of this study were to clarify the specific miR expression profile in CAFs of hepatocellular carcinoma (HCC) and identify its target gene signatures. Small-RNA-sequencing data were generated from nine pairs of CAFs and para-cancer fibroblasts isolated from human HCC and para-tumor tissues, respectively. Bioinformatic analyses were performed to identify the HCC-CAF-specific miR expression profile and the target gene signatures of the deregulated miRs in CAFs. Clinical and immunological implications of the target gene signatures were evaluated in The Cancer Genome Atlas Liver Hepatocellular Carcinoma (TCGA_LIHC) database using Cox regression and TIMER analysis. The expressions of hsa-miR-101-3p and hsa-miR-490-3p were significantly downregulated in HCC-CAFs. Their expression in HCC tissue gradually decreased as HCC stage progressed in the clinical staging analysis. Bioinformatic network analysis using miRWalks, miRDB, and miRTarBase databases pointed to TGFBR1 as a common target gene of hsa-miR-101-3p and hsa-miR-490-3p. TGFBR1 expression was negatively correlated with miR-101-3p and miR-490-3p expression in HCC tissues and was also decreased by ectopic miR-101-3p and miR-490-3p expression. HCC patients with TGFBR1 overexpression and downregulated hsa-miR-101-3p and hsa-miR-490-3p demonstrated a significantly poorer prognosis in TCGA_LIHC. TGFBR1 expression was positively correlated with the infiltration of myeloid-derived suppressor cells, regulatory T cells, and M2 macrophages in a TIMER analysis. In conclusion, hsa-miR-101-3p and hsa-miR-490-3p were substantially downregulated miRs in CAFs of HCC, and their common target gene was TGFBR1. The downregulation of hsa-miR-101-3p and hsa-miR-490-3p, as well as high TGFBR1 expression, was associated with poor clinical outcome in HCC patients. In addition, TGFBR1 expression was correlated with the infiltration of immunosuppressive immune cells.

MicroRNA-101-3p inhibits nasopharyngeal carcinoma cell proliferation and cisplatin resistance through ZIC5 down-regulation by targeting SOX2.

This study aims to explore the mechanism of microRNA (miR)-101-3p-mediated SOX2/ZIC5 axis in the progression of cisplatin resistance of nasopharyngeal carcinoma (NPC). ZIC5 expression was analyzed with a bioinformatics database and detected in NPC cell lines. Cisplatin-resistant cells (HNE-1/DDP and C666-1/DDP) were transfected with sh-ZIC5, sh-SOX2, sh-SOX2 + pcDNA3.1-ZIC5, or miR-101-3p Agomir + pcDNA3.1-SOX2. MiR-101-3p, SOX2, and ZIC5 expression was assessed after transfection, and cancer associated phenotypes were evaluated after cisplatin treatment. The potential relationships among miR-101-3p, SOX2, and ZIC5 were analyzed. A xenograft mouse model of NPC was established with HNE-1 cells stably transfected or not transfected with oe-ZIC5 and subjected to tail vein injection of miR-101-3p Agomir and intraperitoneal injection of cisplatin. Overexpression of ZIC5 was found in cisplatin-resistant NPC cells. Downregulating ZIC5 in NPC cells decreased cell viability, promoted apoptosis, and reduced cisplatin resistance. SOX2 had a binding site on ZIC5, and SOX2 promoted proliferation, migration, and cisplatin resistance and inhibited cell apoptosis by up-regulating ZIC5. Mechanistically, miR-101-3p was decreased in cisplatin-resistant NPC cells and negatively targeted SOX2. Overexpression of miR-101-3p inhibited tumor growth and cisplatin resistance in xenograft mouse model, which was reversed by ZIC5 overexpression. In conclusion, the miR-101-3p/SOX2/ZIC5 axis was implicated in cancer associated phenotypes and cisplatin resistance in NPC.

miR-101-3p contributes to the progression of preeclampsia by suppressing WDR5-mediated proliferation and invasion of trophoblast.

BACKGROUND: Decreased proliferation and invasion of trophoblast were proven to be involved in the pathogenesis of preeclampsia (PE). However, the regulatory network has not been clarified yet. This study aimed to explore the role of miR-101-3p in the progression of PE. METHODS: miR-101-3p expression in placentas of pregnant women with or without PE was analyzed by real-time quantitative PCR (RT-qPCR). Trophoblastic HTR-8/SVneo and HPT-8 cell lines were cultured and underwent hypoxia/reoxygenation (H/R) treatment to mimic PE in vitro. Cell proliferation and invasion were analyzed in gain-of and loss-of-function assays. Finally, we undertook in vivo studies to explore effects of miR-101-3p in the PE model. RESULTS: Compared to placentas from patients without PE, miR-101-3p expressed significantly higher in placentas from PE patients, and its level was positively correlated with the severity of patients. In vitro studies found that overexpression of miR-101-3p significantly suppressed cell proliferation and invasion, while knockdown of miR-101-3p reversed the impacts of H/R treatment. Further research showed that the expression of WD repeat domain 5 (WDR5) was significantly lower in placentas from patients with PE, and its level was negatively associated with the severity of patients. In vitro and in vivo studies confirmed that miR-101-3p promoted PE progression through the regulation of WD WDR5 expression. CONCLUSION: Increased expression of miR-101-3p in placenta contributes to the development of PE by suppressing WDR5-mediated proliferation and invasion of trophoblast.

RGD1564534 represses NLRP3 inflammasome activity in cerebral injury following ischemia-reperfusion by impairing miR-101a-3p-mediated Dusp1 inhibition.

BACKGROUND: Mitochondrial autophagy, the elimination of damaged mitochondria through autophagy, contributes to neuron survival in cerebral ischemia. Long non-coding RNAs (lncRNAs)/microRNAs (miRNAs)/mRNAs are important regulatory networks implicated in various biological processes, including cerebral ischemia-reperfusion (I/R) injury. Therefore, this work clarifies a novel RGD1564534-mediated regulatory network on mitochondrial autophagy in cerebral I/R injury. METHODS: Differentially expressed lncRNAs in cerebral I/R injury were predicted by bioinformatics analysis. Expression of RGD1564534 was examined in the established middle cerebral artery occlusion (MCAO) rats and oxygen glucose deprivation/reoxygenation (OGD/R)-exposed neurons. We conducted luciferase activity, RNA pull-down and RIP assays to illustrate the interaction among RGD1564534, miR-101a-3p and Dusp1. Gain- or loss-of-function approaches were used to manipulate RGD1564534 and Dusp1 expression. The mechanism of RGD1564534 in cerebral I/R injury was evaluated both in vivo and in vitro. RESULTS: RGD1564534 was poorly expressed in the MCAO rats and OGD/R-treated cells, while its high expression attenuated nerve damage, cognitive dysfunction, brain white matter and small vessel damage in MCAO rats. In addition, RGD1564534 promoted mitochondrial autophagy and inhibited NLRP3 inflammasome activity. RGD1564534 competitively bound to miR-101a-3p and attenuated its binding to Dusp1, increasing the expression of Dusp1 in neurons. By this mechanism, RGD1564534 enhanced mitochondrial autophagy, reduced NLRP3 inflammasome activity and suppressed the neuron apoptosis induced by OGD/R. CONCLUSION: Altogether, RGD1564534 elevates the expression of Dusp1 by competitively binding to miR-101a-3p, which facilitates mitochondrial autophagy-mediated inactivation of NLRP3 inflammasome and thus retards cerebral I/R injury.

Extracellular vesicles derived from M2-polarized tumor-associated macrophages promote immune escape in ovarian cancer through NEAT1/miR-101-3p/ZEB1/PD-L1 axis.

Evidence has been presented demonstrating that CD8+ T cells confer anti-cancer effects, which offers a promising approach to enhance immunotherapy. M2-polarized tumor-associated macrophages (TAMs) could transfer RNA to cancer cells by secreting extracellular vesicles (EVs) and stimulate immune escape of cancer cells. Thus, the current study aimed at exploring how EVs derived from M2-polarized TAMs (M2-TAMs) affected the proliferation of ovarian cancer (OC) cells and apoptosis of CD8+ T cells. M2-TAMs were observed in OC tissues, which promoted proliferation of OC cells and CD8+ T cell apoptosis by secreting EVs. OC-associated differentially expressed gene NEAT1 was screened by bioinformatics analysis. The in vitro and in vivo effects of TAM-EVs-NEAT1 and its regulatory mechanism were assessed using gain- and loss-of-function assays in co-culture systems of TAMs-derived EVs, OC cells, and CD8+ T cells and in tumor-bearing mice. NEAT1 was highly expressed in M2-derived EVs and OC cells co-cultured with M2-derived EVs. NEAT1 sponged miR-101-3p to increase ZEB1 and PD-L1 expression. In vitro and in vivo assays confirmed the tumor-supporting effects of NEAT1 delivered by M2-derived EVs on OC cell proliferation and CD8+ T cell apoptosis as well as tumor growth. Collectively, M2-derived EVs containing NEAT1 exerted a tumor-promoting role in OC via the miR-101-3p/ZEB1/PD-L1 axis.

Morinda officinalis polysaccharide enable suppression of osteoclastic differentiation by exosomes derived from rat mesenchymal stem cells.

CONTEXT: Morinda officinalis F.C. How. (MO) (Rubiaceae) can strengthen bone function. OBJECTIVE: To examine the functional mechanism and effect of MO polysaccharides (MOPs) in rats with glucocorticoid-induced osteoporosis (GIOP). MATERIALS AND METHODS: Rats with GIOP were treated with 5, 15 or 45 mL/kg of MOP [n = 15 for each dose, intraperitoneal (i.p.) injection every other day for 8 weeks]. The body weight of rats and histomorphology of bone tissues were examined. Bone marrow mesenchymal stem cells (BMSCs)-derived exosomes (Exo) were collected and identified. Bone marrow-derived macrophages (BMMs) were induced to differentiate into osteoclasts and treated with BMSC-Exo for in vitro studies. RESULTS: MOP reduced the body weight (5, 15, or 45 mg/kg MOP vs. phosphate-buffered saline: 8%, 15% and 25%, p < 0.01), elevated the bone volume to tissue volume (BV/TV), mean trabecular thickness (Tb.Th), mean trabecular number (Tb.N) and mean connectivity density (Conn.D) (40-86%, p < 0.01), decreased the mean trabecular separation/spacing (Tb.Sp) (22-37%, p < 0.01), increased the cortical bone continuity (35-90%, p < 0.01) and elevated RUNX family transcription factor 2 and RANK levels (5-12%, p < 0.01), but suppressed matrix metallopeptidase 9 and cathepsin K levels (9-20%, p < 0.01) in femur tissues. BMSC-Exo from MOP-treated rats (MOP-Exo) suppressed osteoclastic differentiation and proliferation of BMMs. The downregulation of microRNA-101-3p (miR-101-3p) or the upregulation of prostaglandin-endoperoxide synthase 2 (PTGS2) blocked the functions of MOP-Exo. DISCUSSION AND CONCLUSIONS: MOP inhibits osteoclastic differentiation and could potentially be used for osteoporosis management. This suppression may be enhanced by the upregulation of miR-101-3p or the inhibition of PTGS2.

CCDC88A Post-Transcriptionally Regulates VEGF via miR-101 and Subsequently Regulates Hepatocellular Carcinoma.

Background: miR-101 is one of the most abundantly expressed microRNA (miRNA) and exerst a critical role in hepatocellular carcinoma (HCC) by targeting to 3' -untranslated region (UTR) of Girders of actin filaments (CCDC88A) and Vascular endothelial growth factor (VEGF) mRNA, but the underlying molecular mechanism remains to be elucidated. This study aimed to investigate the potential role of CCDC88A on malignancies and stemness by regulating VEGF via miR-101 in HCC. Methods: Gene Expression Profiling Interactive Analysis (GEPIA) was employed to analyze the relevance of CCDC88A expression with prognosis in HCC. Tissue slides were performed to confirm the protein level of CCDC88A in HCC. Correlation between CCDC88A and VEGF was transcriptionally and post-transcriptionally detected, followed by evaluation of malignancies. Results: By employing Immunohistochemistry, we found CCDC88A protein was upregulated in HCC tissues, which is closely correlated to poor prognosis and survival rate. Employment of GEPIA revealed the positive correlation between CCDC88A and VEGF in HCC, but not in liver tissue. Silencing of CCDC88A in Huh-7 and SK-HEP-1 cells significantly decreased proliferation, cell cycle phases, migration, invasion, colony formation, and tumor formation. Introduction of miR-101 mimics specifically targeting CCDC88A and VEGF decreased protein levels of both CCDC88A and VEGFA. Notably, inhibition of miR-101 reversed the correlation between CCDC88A and VEGFA protein levels, indicating that CCDC88A and VEGF may exert as a miR-101 sponge. The addition of SKLB1002, a VEGFR2 inhibitor inhibited malignant behaviors, which was further inhibited by the introduction of miR-101 mimics, indicating that CCDC88A regulates malignant behaviors partially via regulating VEGF. Moreover, CCDC88A also promotes the stemness of cancer stem-like cells derived from HCC cells depending on VEGF modification. Conclusion: Taken together, our findings suggested that the miR-101/CCDC88A/VEGF axis could be a potential therapeutic target of HCC treatment.

[Retracted] MicroRNA­101 inhibits the proliferation and invasion of bladder cancer cells via targeting c­FOS.

Following the publication of this paper, it was drawn to the Editors' attention by a concerned reader that certain of the Transwell cell migration assay data shown in Fig. 5A were strikingly similar to data appearing in different form in other articles by different authors. Owing to the fact that the contentious data in the above article had already been published elsewhere, or 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 any reply. The Editor apologizes to the readership for any inconvenience caused. [the original article was published in Molecular Medicine Reports 14: 2651-2656, 2016; DOI: 10.3892/mmr.2016.5534].

MicroRNA-101-3p Modulates Mitochondrial Metabolism via the Regulation of Complex II Assembly.

MicroRNA-101-3p (miR-101-3p) is a tumour suppressor that regulates cancer proliferation and apoptotic signalling. Loss of miR-101-3p increases the expression of the Polycomb Repressive Complex 2 (PRC2) subunit enhancer of zeste homolog 2 (EZH2), resulting in alterations to the epigenome and enhanced tumorigenesis. MiR-101-3p has also been shown to modulate various aspects of cellular metabolism, however little is known about the mechanisms involved. To investigate the metabolic pathways that are regulated by miR-101-3p, we performed transcriptome and functional analyses of osteosarcoma cells transfected with miR-101-3p. We found that miR-101-3p downregulates multiple mitochondrial processes, including oxidative phosphorylation, pyruvate metabolism, the citric acid cycle and phospholipid metabolism. We also found that miR-101-3p transfection disrupts the transcription of mitochondrial DNA (mtDNA) via the downregulation of the mitochondrial transcription initiation complex proteins TFB2M and Mic60. These alterations in transcript expression disrupt mitochondrial function, with significant decreases in both basal (54%) and maximal (67%) mitochondrial respiration rates. Native gel electrophoresis revealed that this diminished respiratory capacity was associated with reduced steady-state levels of mature succinate dehydrogenase (complex II), with a corresponding reduction of complex II enzymatic activity. Furthermore, miR-101-3p transfection reduced the expression of the SDHB subunit, with a concomitant disruption of the assembly of the SDHC subunit into mature complex II. Overall, we describe a new role for miR-101-3p as a modulator of mitochondrial metabolism via its regulation of multiple mitochondrial processes, including mtDNA transcription and complex II biogenesis.

MiR-101-3p Suppresses Progression of Cervical Squamous Cell Carcinoma by Targeting and Down-Regulating KPNA2.

Objective We explored mechanism of microRNA-101-3p/Karyopherin α2 (KPNA2) axis in cervical squamous cell carcinoma. Methods: Bioinformatics methods were applied to identify genes for the study. Cell functional assays were implemented to examine the role of the genes in malignant progression of cervical squamous cell carcinoma. Targeting relationship between genes was verified by dual-luciferase assay. Results: MicroRNA-101-3p was lowly expressed in cervical squamous cell carcinoma, while KPNA2 was highly expressed. Dual-luciferase assay identified direct targeting relationship between microRNA-101-3p and KPNA2. Functional assays manifested that highly expressed microRNA-101-3p suppressed cervical squamous cell carcinoma cell growth by targeting KPNA2. Conclusion: Overall, microRNA-101-3p/KPNA2 axis can play an important part in progression of cervical squamous cell carcinoma.

[Retracted] MicroRNA­101 induces apoptosis in cisplatin­resistant gastric cancer cells by targeting VEGF­C.

Following the publication of this paper, it was drawn to the Editors' attention by a concerned reader that certain of the cell apoptotic assay data shown in Figs. 3C and 5B were strikingly similar to data appearing in different form in other articles by different authors. Owing to the fact that the contentious data in the above article had already been published elsewhere, or 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 any reply. The Editor apologizes to the readership for any inconvenience caused. [the original article was published in Molecular Medicine Reports 13: 572­578, 2016; DOI: 10.3892/mmr.2015.4560].

miR­101a­3p overexpression prevents acetylcholine­CaCl2­induced atrial fibrillation in rats via reduction of atrial tissue fibrosis, involving inhibition of EZH2.

Atrial fibrillation (AF), a clinically common heart arrhythmia, can result in left ventricular hypofunction, embolism and infarction. MicroRNA (miR)­101a­3p is lowly expressed in atrial tissues of patients with AF, but its role in AF remains unknown. In the present study, an AF model in rats was established via intravenous injection of acetylcholine (Ach)­CaCl2. The downregulation of miR­101a­3p and upregulation of enhancer of zeste 2 homolog 2 (EZH2) were observed in AF model rats, indicating the involvement of miR­101a­3p and EZH2 in AF development. To study the effect of miR­101a­3p on AF in vivo, AF model rats were intramyocardially injected with lentivirus expressing miR­101a­3p. Electrocardiogram analysis identified that miR­101a­3p overexpression restored disappeared P wave and R­R interphase changes in Ach­CaCl2­induced rats. Overexpression of miR­101a­3p also increased the atrial effective refractory period, reduced AF incidence and shortened duration of AF. Histological changes in atrial tissues were observed after H&E and Masson staining, which demonstrated that miR­101a­3p reduced atrial remodeling and fibrosis in AF model rats. Moreover, EZH2 expression was downregulated in atrial tissues by miR­101a­3p induction. Immunohistochemistry for collagen Ⅰ and collagen III revealed a reduction in atrial collagen synthesis following miR­101a­3p overexpression in AF model rats. Additionally, miR­101a­3p lowered the expression of pro­fibrotic biomarkers, including TGF­ß1, connective tissue growth factor, fibronectin and α­smooth muscle actin. The luciferase reporter assay results also indicated that EZH2 was a target gene of miR­101a­3p. Taken together, it was found that miR­101a­3p prevented AF in rats possibly via inhibition of collagen synthesis and atrial fibrosis by targeting EZH2, which provided a potential target for preventing AF.

Knockdown of Myeloid Cell Leukemia-1 by MicroRNA-101 Increases Sensitivity of A549 Lung Cancer Cells to Etoposide.

Background: Studies have shown that myeloid cell leukemia-1 (Mcl-1) is the target gene for microRNA -101 (miRNA-101), and decreased levels of miRNA-101 are associated with elevated levels of Mcl-1 and lung cancer survival. The objective of the present study was to investigate the effect of miRNA-101 on the sensitivity of A549 lung cancer cells to etoposide. Methods: The study was conducted during 2018 and 2019 at Arak University of Medical Sciences, Arak, Iran. The effect of miRNA-101 on Mcl-1 expression was assessed using reverse transcription-quantitative polymerase chain reaction 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT), and trypan blue exclusion assays were performed to determine the effect of treatments on cell survival and proliferation, respectively. The interaction between miRNA-101 and etoposide was evaluated using the combination index analysis of Chou-Talalay. Apoptosis was quantified using ELISA cell death assay. ANOVA and Bonferroni's tests were used to determine statistical differences between the groups (P<0.05). GraphPad Prism software (version 6.01) was used for data analysis. Results: The results showed that miRNA-101 clearly inhibited the expression of Mcl-1 and reduced the growth of A549 cells, relative to blank control and negative control miRNA (P<0.05). Transfection of miRNA-101 synergistically enhanced the sensitivity of the A549 cells to etoposide. Apoptosis assay data also showed that miRNA-101 triggered apoptosis and augmented the etoposide-mediated apoptosis. Conclusion: Up-regulation of miRNA-101 inhibited cell survival and proliferation, and sensitized A549 cells to etoposide by suppressing Mcl-1 expression. miRNA-101 replacement therapy can be considered as an effective therapeutic strategy in non-small cell lung cancer.

LINC00665 promotes the viability, migration and invasion of T cell acute lymphoblastic leukemia cells by targeting miR-101 via modulating PI3K/Akt pathway.

T cell acute lymphoblastic leukemia (T-ALL) is a high-risk malignancy. The effects of cancer growth-related Long Intergenic Non-Protein Coding RNA 665 (LINC00665) in T-ALL remained obscure, and therefore further exploration was conducted on that in this study. The expression of LINC00665 in acute myeloid leukemia (LAML) tissues and myeloid tissues was analyzed using Gene Expression Profiling Interactive Analysis (GEPIA) 2. The target microRNA (miR) of LINC00665 was predicted by LncBase Predicted v.2 and verified using dual-luciferase reporter assay. After LINC00665 and miR-101 in T-ALL cells were overexpressed or silenced, the viability, migration and invasion of cell were detected using cell counting kit-8 and Transwell assays. The expressions of LINC00665, miR-101, Cyclin D1, Matrix metalloproteinases (MMP)-2, MMP-9, phosphorylated (p)-phosphatidylinositol 3-kinase (PI3K), PI3K, p-Akt, Akt were detected by quantitative real-time Polymerase Chain Reaction (qRT-PCR) and western blot. High expression of LINC00665 was presented in LAML tissues, the peripheral blood samples from patients with T-ALL and T-ALL cells. Overexpression of LINC00665 promoted the viability, migration and invasion of T-ALL cells and downregulated miR-101 expression, whereas silencing of LINC00665 did oppositely. MiR-101 could competitively bind to LINC00665, and was low-expressed in T-ALL. MiR-101 mimic inhibited viability, migration and invasion of T-ALL cells, and reversed effects of overexpressed LINC00665, whilst miR-101 inhibitor reversed the effects of LINC00665 silencing. Besides, overexpressed LINC00665 upregulated the expressions of Cyclin D1 MMP-2, and MMP-9 and the ratios of p-PI3K/PI3K and p-Akt/Akt, which were reversed by miR-101 mimic. LINC00665 could enhance the viability, migration and invasion abilities of T-ALL cells by targeting miR-101 via activating PI3K/Akt pathway.

MicroRNA­101 inhibits renal tubular epithelial­to­mesenchymal transition by targeting TGF­ß1 type I receptor.

MicroRNAs (miRNAs/miRs) are key regulators of renal interstitial fibrosis (RIF). The present study was designed to identify miRNAs associated with the development of RIF, and to explore the ability of these identified miRNAs to modulate the renal tubular epithelial­to­mesenchymal transition (EMT) process. To this end, miRNAs that were differentially expressed between normal and fibrotic kidneys in a rat model of mercury chloride (HgCl2)­induced RIF were detected via an array­based approach. Bioinformatics analyses revealed that miR­101 was the miRNA that was most significantly downregulated in the fibrotic renal tissue samples, and this was confirmed by RT­qPCR, which also demonstrated that this miRNA was downregulated in transforming growth factor (TGF)­ß1­treated human proximal tubular epithelial (HK­2) cells. When miR­101 was overexpressed, this was sufficient to reverse TGF­ß1­induced EMT in HK­2 cells, leading to the upregulation of the epithelial marker, E­cadherin, and the downregulation of the mesenchymal marker, α­smooth muscle actin. By contrast, the downregulation of miR­101 using an inhibitor exerted the opposite effect. The overexpression of miR­101 also suppressed the expression of the miR­101 target gene, TGF­ß1 type I receptor (TßR­I), and thereby impaired TGF­ß1/Smad3 signaling, while the opposite was observed upon miR­101 inhibition. To further confirm the ability of miR­101 to modulate EMT, the HK­2 cells were treated with the TßR­I inhibitor, SB­431542, which significantly suppressed TGF­ß1­induced EMT in these cells. Notably, miR­101 inhibition exerted a less pronounced effect upon EMT­related phenotypes in these TßR­I inhibitor­treated HK­2 cells, supporting a model wherein miR­101 inhibits TGF­ß1­induced EMT by suppressing TßR­I expression. On the whole, the present study demonstrates that miR­101 is capable of inhibiting TGF­ß1­induced tubular EMT by targeting TßR­I, suggesting that it may be an important regulator of RIF.

miR­101­3p negatively regulates inflammation in systemic lupus erythematosus via MAPK1 targeting and inhibition of the NF­κB pathway.

Systemic lupus erythematosus (SLE) is an autoimmune disease often used as a model in genomics research. The downregulation of microRNA­101­3p (miR­101­3p) participates in the progression of SLE, although the underlying mechanisms remain to be elucidated. The present study aimed to evaluate the specific roles of miR­101­3p in the SLE inflammatory response and its potential mechanisms. Reverse transcription­quantitative (RT­q) PCR was used to profile miR­101­3p expression in the peripheral blood mononuclear cells (PBMCs) from 40 female patients with SLE and 20 female healthy volunteers. The interactions between miR­101­3p and MAPK1 were identified and evaluated using dual­luciferase reporter and RNA pull­down assays. The levels of IL­10 and IFN­Î³ were evaluated by enzyme­linked immunosorbent assay. The expression of NF­κB p65 and phosphorylated IκBα were evaluated using western blotting. miR­101­3p expression was demonstrated to be downregulated in SLE PBMCs. miR­101­3p negatively regulated IL­10 and IFN­Î³ expression in SLE samples and was demonstrated to target MAPK1. Increases in MAPK1 expression eliminated miR­101­3p inhibition of IL­10 and IFN­Î³. MAPK1 activated the NF­κB pathway in SLE PBMCs and this activation was inhibited when miR­101­3p was overexpressed. In addition, treatment with BAY11­7085 (NF­κB activator) was demonstrated to reverse the inhibitory effects of miR­101­3p expression on both IL­10 and IFN­Î³ in SLE PBMCs. BAY11­7082 also markedly reduced MAPK1­induced increases in IL­10 and IFN­Î³ in SLE PBMCs. miR­101­3p overexpression attenuated the inflammatory response in SLE PBMCs by inhibiting the expression of MAPK1 and blocking the NF­κB pathway. The results revealed a novel regulatory mechanism in SLE inflammation and offer a new direction for the development of SLE treatments.

Knockdown of lncRNA HOTTIP Inhibits Retinoblastoma Progression by Modulating the miR-101-3p/STC1 Axis.

OBJECTIVE: Retinoblastoma (RB) is a frequent eye cancer in children. Long non-coding RNA (LncRNA) HOXA transcript at the distal tip (HOTTIP) is aberrantly expressed in cancer tissues. This study explores the underlying mechanism of lncRNA HOTTIP in RB. METHODS: HOTTIP expression in normal retinal cells and RB cell lines was detected using qRT-PCR. The proliferation of RB cells was measured using CCK-8 and EdU assays, and apoptosis was detected using flow cytometry and Western blotting after the transfection of si-HOTTIP into Y79 cells and pc-HOTTIP into HXO-RB-44 cells. The target relationships between HOTTIP and miR-101-3p, and miR-101-3p and STC1 were predicted by bioinformatics website and verified using dual-luciferase reporter gene assay. The binding of HOTTIP and miR-101-3p was verified using RNA pull-down assay. STC1 mRNA and protein in RB cells were measured using qRT-PCR and Western blotting. Moreover, si-HOTTIP and in-miR-101-3p/in-NC, and si-HOTTIP and pc-STC1/pcDNA were co-transfected into Y79 cells respectively to evaluate cell proliferation and apoptosis. Xenograft study was conducted, and Ki67-positive expression was detected using immunohistochemical staining. RESULTS: HOTTIP expression was promoted in RB tissues and cells. Downregulation of HOTTIP inhibited proliferation and promoted apoptosis of Y79 cells, while upregulation of HOTTIP promoted proliferation and inhibited apoptosis of HXO-RB-44 cells. There were target relationships between HOTTIP and miR-101-3p, and miR-101-3p and STC1. Inhibition of miR-101-3p or overexpression of STC1 reversed the effect of si-HOTTIP on the proliferation and apoptosis of RB cells. Xenograft study showed that knockdown of HOTTIP suppressed the growth of RB in vitro. CONCLUSION: It could be concluded that HOTTIP sponged miR-101-3p to upregulate STC1 expression, thereby promoting RB cell proliferation and inhibiting apoptosis.

Role of enhancer of zeste homolog 2 in osteoclast formation and periodontitis development by downregulating microRNA-101-regulated VCAM-1.

The enhancer of zeste homolog 2 (EZH2) represents a potential target for periodontitis treatment; however, its role in the development of periodontitis remains unclear. The current study aimed to elucidate the role of EZH2 in osteoclasts (OCs) growth as well as the mechanism underpinning the related process. The potential interaction among EZH2, microRNA-101 (miR-101), and vascular cell adhesion molecule 1 (VCAM-1) was evaluated using chromatin immunoprecipitation and dual-luciferase reporter gene assay. The expressions of EZH2 and miR-101 in OCs were examined by Western blot analysis and reverse transcription squantitative polymerase chain reaction. Loss- and gain-function assays were then performed to determine the role of EZH2/miR-101/VCAM-1 in periodontitis and OCs proliferation, followed by OC growth and proliferation detected using tartrate resistant acid phosphatase (TRAP) and 5-ethynyl-2'-deoxyuridine staining. Enzyme-linked immunoassay was conducted to determine the expression of interleukin 1ß (IL-1ß) and tumor necrosis factor-α (TNF-α). A periodontitis rat model was established to investigate the effect of EZH2 and VCAM-1 in vivo. EZH2 was overexpressed, while miR-101 was downregulated in the OCs of periodontitis. Silencing of EZH2, VCAM-1 repression, or miR-101 elevation suppressed the growth and proliferation of OC while acting to encumber the release of IL-1ß and TNF-α. EZH2 negatively targeted miR-101, while miR-101 negatively targeted VCAM-1. Moreover, silencing of EZH2 or VCAM-1 was observed to attenuate periodontitis which was evidenced by an increase in BMD, BV/TV, and BS/BV as well as reduction in TRAP and cathepsin K in vivo. Taken together, the key findings of the current study demonstrate that EZH2 knockdown inhibited OC formation by elevating the expression of miR-101 via suppression of VCAM-1, ultimately attenuating periodontitis.