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Cisplatin is one of the most effective chemotherapeutic agents strongly associated with nephrotoxicity. Tubular adult renal progenitor cells (tARPC) can regenerate functional tubules and participate in the repair processes after cisplatin exposition. This study investigated the molecular mechanisms underlying the protective effect of tARPC on renal epithelium during cisplatin nephrotoxicity. By performing a whole-genome transcriptomic analysis, we found that tARPC, in presence of cisplatin, can strongly influence the gene expression of renal proximal tubular cell [RPTEC] by inducing overexpression of CYP1B1, a member of the cytochrome P450 superfamily capable of metabolizing cisplatin and of hypoxia/cancer-related lncRNAs as MIR210HG and LINC00511. Particularly, tARPC exerted renoprotection and regeneration effects via extracellular vesicles (EV) enriched with CYP1B1 and miR-27b-3p, a well-known CYP1B1 regulatory miRNA. The expression of CYP1B1 by tARPC was confirmed by analyzing biopsies of cisplatin-treated renal carcinoma patients that showed the colocalization of CYP1B1 with the tARPC marker CD133. CYP1B1 was also overexpressed in urinary EV purified from oncologic patients that presented nephrotoxicity episodes after cisplatin treatment. Interestingly CYP1B1 expression significantly correlated with creatinine and eGFR levels. Taken together, our results show that tARPC are able to counteract cisplatin-induced nephrotoxicity via CYP1B1 release through EV. These findings provide a promising therapeutic strategy for nephrotoxicity risk assessment that could be related to abundance of renal progenitors.

Postoperative cognitive dysfunction (POCD) is a normal condition that develops after surgery with anesthesia, leading to deterioration of cognitive functions. However, the mechanism of POCD still remains unknown. To elucidate the POCD molecular mechanism, sevoflurane was employed in the present study to generate neuroinflammation mice model. Sevoflurane treatment caused inflammatory markers IL6, IL-10 and TNF-α high expression in primary hippocampal neurons and blood samples. Long non-coding RNA Gm5106 was found to be increased after being stimulated with sevoflurane. Silencing Gm5106 inhibited neuron inflammation. In the meanwhile, Gm5106 was identified as a direct target of miR-27b-3p that was inhibited by sevoflurane and related to inflammation suppression. In addition, transcription factor (TF) Hoxa5 was validated to activate Gm5106 through two binding motifs in the promoter region after sevoflurane exposure. Furthermore, miR-27b-3p also directly targeted Hoxa5 3ʹUTR, which affected nuclear Hoxa5 protein served as TF. Hoxa5 protein instead of 3ʹUTR reduced miR-27b-3p, in which Gm5106 knocking down abrogated this effect. In conclusion, sevoflurane induces neuroinflammation through increasing long non-coding RNA Gm5106, which is transcriptionally activated by Hoxa5 and directly targeted by miR-27-3p. Apart from that, Hoxa5, Gm5106, and miR-27b-3p form a positive feedback loop in sevoflurane stimulation.

Inflammatory reaction exerts a pivotal role in secondary damage after cerebral hemorrhage and spinal cord injury. miRNAs can both promote and inhibit inflammatory actions among microglial cells. The objective of the present paper was to figure out whether miR-27b-3p produced regulatory effects during processes of microglial inflammation. Lipopolysaccharides (LPS) were used to prepare microglial activation models. Following miR-27b-3p overexpression and interference, the RNA and protein levels of tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-1β were subjected to real-time fluorescent quantitative PCR (qPCR) and western blot assays, respectively. Cellular apoptosis was subjected to flow cytometry and miR-27b-3p target genes were visualized using a dual luciferase reporter system for verification. The levels of TNF-α, IL-6, and IL-1β mRNA in miR-27b-3p-overexpressed microglial cells were markedly increased compared to the control. Apoptosis of microglial cells was increased markedly in the overexpressed miR-27b-3p group compared to the negative control. Conversely, a different result was presented in the microglial transfected with miR-27b-3p inhibitors. The downregulation of A20, a miR-27b-3p target gene, mediated levels of TNF-α, IL-6, and IL-1β. Furthermore, A20 reduced microglial apoptosis. These data revealed that miR-27b-3p could mediate not only microglia activation but also neuroinflammation via downregulating A20 expression. Thus, miR-27b-3p is regarded as gene therapy in treating cerebral hemorrhage and spinal cord injury., Graphical abstract

Lei Li, Guohua Lv, Bing Wang, Hong Ma Department of Spine Surgery, The Second Xiangya Hospital, Central South University, Changsha, Hunan 410011, People’s Republic of ChinaCorrespondence: Hong Ma Tel +86 731-85295124Email spinema@csu.edu.cnBackground: Chordoma, a rare bone tumor, occurs most commonly at the sacrococcygeal and skull base region. To date, chemotherapy is used to treat patients with advanced-stage chordoma. However, multidrug resistance (MDR) greatly hinders the effect of chemotherapy in chordoma. Here, we studied the correlation between KCNQ1OT1 and chemotherapy resistance.Methods: RT-PCR assay was used to examine KCNQ1OT1, miR-27b-3p, and ATF2 mRNA expression. CCK8 assay was exercised to detect IC50 values of cisplatin in chordoma cells. ATF2 protein expression was detected by Western blot.Results: KCNQ1OT1 was increased in chemotherapy-resistant patients and cisplatin-resistant cells, and downregulation of KCNQ1OT1 expression weakened MDR in chordoma. In addition, KCNQ1OT1 promoted MDR in chordoma by sponging miR-27b-3p and subsequently increasing ATF2 expression.Conclusion: KCNQ1OT1 is proved to be strikingly raised in the chemotherapy-resistant group and to promote MDR in chordoma. Our findings demonstrated the role of the KCNQ1OT1/miR-27b-3p/ATF2 axis in MDR of chordoma, which provides new insight into the molecular mechanism of chordoma MDR, and may determine the effect of therapy after receiving chemotherapy by detecting the expression of KCNQ1OT1 in serum.Keywords: KCNQ1OT1, MDR, miR-27b-3p, ATF2, chordoma

Ming Li,1 Xufeng Guo2 1Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan 430000, Hubei, People’s Republic of China; 2Cancer Center, Renmin Hospital of Wuhan University, Wuhan 430000, Hubei, People’s Republic of ChinaCorrespondence: Xufeng GuoCancer Center, Renmin Hospital of Wuhan University, Wuhan 430000, Hubei, People’s Republic of ChinaEmail guoxufeng007@126.comBackground: An increasing number of studies demonstrate that long non-coding RNAs (lncRNAs) are regulators in cancer biology. Nevertheless, the expression and mechanism of LINC01089 in colorectal cancer (CRC) remain unclear.Methods: Quantitative real-time polymerase chain reaction (qRT-PCR) was taken to investigate the expression levels of LINC01089 and miR-27b-3p in CRC tissues and cells. MTT method and transwell test were employed to assess the proliferation and invasion of CRC cells, respectively. Dual-luciferase activity reporter assay, RNA immunoprecipitation assay, Pearson’s correlation analysis, and Western blot were performed to investigate the regulatory mechanism of LINC01089/miR-27b-3p/HOXA10 axis in CRC.Results: LINC01089 was down-regulated in CRC tissues and cell lines. LINC01089 overexpression impeded the proliferation and invasion of SW620 and LoVo cells, whereas LINC01089 knockdown increased the malignancy of SW480 and HT29 cells. Moreover, LINC01089 directly interacted with miR-27b-3p to repressed its expression and indirectly promoted the expression of HOXA10.Conclusion: LINC01089 impedes the proliferation and invasion of colorectal cancer cells by adsorbing miR-27b-3p and up-regulating the expression of HOXA10.Keywords: LINC01089, CRC, miR-27b-3p, HOXA10

Qingfeng Jiang, Wenqun Xing, Jinhua Cheng, Yongkui Yu Department of Thoracic Surgery, The Affiliated Cancer Hospital of Zhengzhou University, Henan Cancer Hospital, Zhengzhou, Henan, People’s Republic of ChinaCorrespondence: Wenqun XingDepartment of Thoracic Surgery, The Affiliated Cancer Hospital of Zhengzhou University, Henan Cancer Hospital, No. 127 Dongming Road, Zhengzhou 450008, Henan, People’s Republic of ChinaTel +86-0371-65588265Email kcbbgn@163.comPurpose: Long non-coding RNA P73 antisense RNA 1T (TP73-AS1) is a newly discovered lncRNA involved in the occurrence and development of several cancers. However, its role in lung cancer has not been well investigated yet.Methods: The expressions of TP73-AS1, microRNA-27b-3p (miR-27b-3p) and lysosomal-associated protein transmembrane-4 Beta (LAPTM4B) were detected by quantitative real-time polymerase chain reaction (qRT-PCR). The cell proliferation, apoptosis, migration and invasion were detected by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), Annexin V-FITC/PI and transwell assays, respectively. Tumor xenografts were applied to explore the role of TP73-AS1 in vivo. The target relationship was predicted by StarBase v.2.0 or TargetScan and confirmed by luciferase reporter assay. Pearson’s coefficient assay was applied to assess the expression correlation between two groups. Protein expression levels were detected by Western blot.Results: We found that TP73-AS1 was strikingly up-regulated in lung cancer tissues and cells. TP73-AS1 depletion inhibited the growth and metastasis of lung cancer cells in vitro. Furthermore, TP73-AS1 could act as an endogenous sponge by directly binding miR-27b-3p, and a notable inverse correlation between them was also discovered. Importantly, knockdown of miR-27b-3p could reverse the inhibitory effects of TP73-AS1 depletion on the growth and metastasis of lung cancer cells. Besides, LAPTM4B was directly targeted by miR-27b-3p and could be co-regulated by TP73-AS1 and miR-27b-3p in lung cancer cells. Silencing TP73-AS1 hampered tumor growth by regulating miR-27b-3p/LAPTM4B axis in vivo.Conclusion: TP73-AS1 promoted the progression of lung cancer through regulating miR-27b-3p/LAPTM4B axis and it might be a potential target for diagnosis and treatment of lung cancer.Keywords: lung cancer, TP73-AS1, miR-27b-3p, LAPTM4B, proliferation, metastasis

Diffuse large B-cell lymphoma (DLBCL) is commonly treated with R-CHOP, but ~30 to 50% of the patients are poorly responsive to this strategy. Geniposide, an extract from the Gardenia jasminoides Ellis, plays antitumor roles in human gastric cancer, hepatocellular carcinoma, and oral squamous carcinoma. However, the effects of geniposide treatment on DLBCL cells, as well as its underlying mechanism, are still unknown. Here, we found that geniposide inhibited the proliferation of OCI-LY7 and OCI-LY3 cells in a dose-dependent manner. Furthermore, geniposide increased the percentage of apoptotic cells and upregulated the levels of cleaved PARP and cleaved caspase-3 in DLBCL cells. Interestingly, geniposide treatment significantly reduced the expression of the long noncoding RNA HLA complex P5 (lncRNA HCP5) in DLBCL cells. HCP5 expression was revealed to be upregulated in DLBCL tissues and cell lines. Moreover, HCP5 knockdown resulted in proliferation inhibition and apoptosis in OCI-LY7 and OCI-LY3 cells. miR-27b-3p was predicted as a potential target of HCP5 using the lnCAR web tool. Both HCP5 silencing and geniposide treatment increased the level of miR-27b-3p in DLBCL cells. Accordingly, a luciferase reporter assay identified miR-27b-3p as a direct target of HCP5. The expression of miR-27b-3p was upregulated and inversely correlated with the HCP5 level in DLBCL tissues. HCP5 knockdown reduced MET protein expression, which was subsequently rescued by miR-27b-3p silencing in DLBCL cells. Importantly, the restoration of MET partially reversed the geniposide-induced proliferation inhibition and apoptosis of DLBCL cells. In conclusion, geniposide inhibits the proliferation and induces the apoptosis of DLBCL cells at least partially by regulating the HCP5/miR-27b-3p/MET axis, indicating a potential strategy for DLBCL treatment.

Oxaliplatin (OXA) resistance is the major obstacle to the anticancer effects of chemotherapy in colorectal cancer (CRC) patients. MicroRNAs (miRNAs) play an important role in the chemoresistance of various tumors. Our objective is to clarify the underlying mechanism of miRNAs in chemoresistance and provide a potential strategy to improve the response of CRC patients to chemotherapeutics. Methods: MiRNA microarray and Real-time PCR were performed to compare changes in miRNA expression between oxaliplatin-resistant and the parental cells. CCK8, apoptosis assay, immunofluorescence and xenograft studies were used to elucidate the impact of miR-27b-3p on regulating chemoresistance. Luciferase reporter assay and western blot were carried to assess the regulatory role of miR-27b-3p in ATG10 expression. The effects of miR-27b-3p and ATG10 on autophagy were investigated by GFP-LC3 fluorescence microscopy, transmission electron microscopy, and western blot. ChIP assay and luciferase assay were performed to test the c-Myc's occupancy on the miR-27B promoter. Results: We observed that miR-27b-3p expression was significantly downregulated in oxaliplatin-resistant cell lines (SW480-OxR and HCT116-OxR) compared to the corresponding parental cell lines and that miR-27b-3p expression was positively correlated with disease-free survival (DFS) time in colorectal cancer patients. MiR-27b-3p could sensitize colorectal cancer cells to oxaliplatin in vitro and in vivo. Under oxaliplatin treatment, chemoresistant cells showed a higher autophagy level than parental cells. Moreover, we also identified that miR-27b-3p inhibited the expression of ATG10 at the posttranscriptional level, thus inhibiting autophagy. Further study demonstrated that c-Myc can inhibit the expression of miR-27b-3p via binding to the promoter region of miR-27B gene. Conclusions: Our study identifies a novel c-Myc/miR-27b-3p/ATG10 signaling pathway that regulates colorectal cancer chemoresistance. These results suggest that miR-27b-3p is not only a potential indicator for evaluating efficiency of chemotherapy, but also a valuable therapeutic target for CRC, especially for patients with chemoresistance.

Background Metastasis is the main cause of death in colorectal cancer (CRC). Circulating tumour cells (CTCs) are regarded as the precursor cells of metastasis. The CTCs, which underwent epithelial‐mesenchymal transition (EMT), are associated with metastasis and responsible for poor prognosis. EMT cancer cells modulate endothelial permeability in the invasive front and facilitate cancer cell intravasation, resulting in CTCs‐mediated distant metastasis. Exosomes derived from cancer cells are key mediators of cancer‐host intercommunication. However, the mechanism by which EMT‐tumour cells‐derived exosomes modulate vascular permeability and promote CTCs generation has remained unclear. Methods Exosomes isolation and purification were conducted by ultra‐centrifugation. Exosomal miRNA was identified by sequencing followed by quantitative PCR. In vitro co‐culture assay experiments were conducted to evaluate the effect of exosomal miR‐27b‐3p on the permeability of blood vessel endothelium. Dual‐luciferase reporter assay, chromatin immunoprecipitation (ChIP) and RNA immunoprecipitation (RIP) were performed to investigate the underlying mechanism by which miR‐27b‐3p is packaged into exosomes. A mouse model was established to determine the role of exosomal miR‐27b‐3p in blood vessel permeability modulation in vivo. Results We found that EMT‐CRC cells attenuate the blood vessel barrier by transferring miR‐27b‐3p to human umbilical vein endothelial cells (HUVECs) in exosomes. Mechanically, miR‐27b‐3p atteuated the expression of vascular endothelial cadherin (VE‐Cad) and p120 at the post‐transcriptional level by binding to 3′‐untranslated region of VE‐Cad and p120 directly. The packaging of miR‐27b‐3p into exosomes was induced by heterogeneous nuclear ribonucleoprotein A1 (hnRNPA1), which activated by STAT3. Clinically, miR‐27b‐3p up‐regulated in CRC tissues. Plasma exosomal miR‐27b‐3p was positively correlated with malignant progression and CTC count in CRC patients. Our study reveals a novel mechanism by which EMT‐CRC cells promote metastasis, increasing blood vessel permeability and facilitating the generation of CTCs. Conclusion Exosomal miR‐27b‐3p secreted by EMT‐CRC cells increases blood vessel permeability and facilitates the generation of CTCs. Exosomal miR‐27b‐3p may become a promising biomarker for CRC metastasis., Collectively, our findings revealed that EMT CRC cell‐derived exosomal miR‐27b‐3p transferred to endothelial cells, enhanced blood vessel permeability, and thus facilitated CTC generation by targeting VE‐cadherin and p120. Upstream STAT3 upregulated hnRNPA1 by binding its promotor and hnRNPA1 mediated the packaging of miR‐27b‐3p. Our study highlighted that the STAT3/hnRNPA1/miR‐27b‐3p signal cascade and exosomal miR‐27b‐3p may be promising diagnostic biomarkers for CRC metastasis.

Understanding the regulation of DNA repair mechanisms is of utmost importance to identify altered cellular processes that lead to diseases such as cancer through genomic instability. In this sense, miRNAs have shown a crucial role. Specifically, miR-27b-3 biogenesis has been shown to be induced in response to DNA damage, suggesting that this microRNA has a role in DNA repair. In this work, we show that the overexpression of miR-27b-3p reduces the ability of cells to repair DNA lesions, mainly double-stranded breaks (DSB), and causes the deregulation of genes involved in homologous recombination repair (HRR), base excision repair (BER), and the cell cycle. DNA damage was induced in BALB/c-3T3 cells, which overexpress miR-27b-3p, using xenobiotic agents with specific mechanisms of action that challenge different repair mechanisms to determine their reparative capacity. In addition, we evaluated the expression of 84 DNA damage signaling and repair genes and performed pathway enrichment analysis to identify altered cellular processes. Taken together, our results indicate that miR-27b-3p acts as a negative regulator of DNA repair when overexpressed.

Wen Sun,1 Liwen Zhang,2 Ranran Yan,2 Ying Yang,2 Xiangli Meng31Teaching Administration Office, Affiliated Hospital of Jining Medical University, Jining, People’s Republic of China; 2Intensive Care Unit, Affiliated Hospital of Jining Medical University, Jining, People’s Republic of China; 3Nursing Department, Affiliated Hospital of Jining Medical University, Jining, People’s Republic of ChinaBackground: Non-small cell lung cancer (NSCLC) has a significant impact on human health. The aim of this study was to explore the role of long non-coding RNA DLX6-AS1 in the proliferation, migration, and invasion of NSCLC cells.Methods: The expression of DLX6-AS1 in NSCLC tumor tissues and cell lines was examined by qRT-PCR. The effects of DLX6-AS1 knockdown on cell proliferation, migration, and invasion were assessed by Cell Counting Kit-8, wound healing, and transwell assays, respectively. Bioinformatics analyses, luciferase reporter assays, and RNA pull-down assays were employed to examine the mechanism by which DLX6-AS1 exerted its oncogenesis effects in NSCLC. The anti-tumor effect of silencing DLX6-AS1 in vivo was also evaluated.Results: DLX6-AS1 was over-expressed in NSCLC tumor tissues and cell lines and its level of expression was found to be associated with tumor size and advanced clinical stage in patients with NSCLC. Downregulation of DLX6-AS1 inhibited cell proliferation, cell clone formation, migration, and invasion of NSCLC cells. DLX6-AS1 was found to interact with miR-27b-3p/GSPT1. DLX6-AS1 expression was negatively correlated with miR-27b-3p expression, but positively correlated with GSPT1 expression in NSCLC samples. DLX6-AS1 knockdown also effectively suppressed tumor growth in an in vivo xenograft model.Conclusion: DLX6-AS1 regulated NSCLC progression by targeting the miR-27b-3p/GSPT1 axis, which may provide novel insights for NSCLC prognosis and therapy.Keywords: DLX6-AS1, NSCLC, miR-27b-3p, GSPT1, invasion

Oral lichen planus (OLP) is considered a precancerous lesion with no known cure. Recent studies reported that abnormal regulation of apoptosis was involved in the pathogenesis of OLP. Next generation sequencing was used to screen the candidate microRNAs and genes in biopsies from patients with OLP and healthy mucosa. Human oral keratinocytes were transfected into the related oligonucleotides of miR‐27b‐3p/cyclophilin D and their control groups. Apoptosis was detected by TdT‐mediated dUTP nick end labelling and flow cytometry. The levels of mRNA and protein were detected by quantitative PCR, Western blots, and enzyme‐linked immunosorbent assays, respectively. Luciferase assays were performed to detect the luciferase activities of miR‐27b‐3p and cyclophilin D. Here, we showed that basal epithelium apoptosis was reduced and the miR‐27b‐3p levels were decreased in clinical OLP samples. We also found that down‐regulation of miR‐27b‐3p inhibited epithelial keratinocyte apoptosis by up‐regulating cyclophilin D expression. Moreover, cyclophilin D increased the protein stability of Bcl2 through direct binding, and Bcl2 suppressed caspase9/3 activation and cytochrome C release. Taken together, these data showed that miR‐27b‐3p regulated keratinocyte apoptosis through cyclophilin D/Bcl2 signalling, suggesting the miR‐27b‐3p regulated the pathogenesis of OLP.

microRNAs play an important role in the growth and development of chicken embryos, including the regulation of skeletal muscle genesis, myoblast proliferation, differentiation, and apoptosis. Our previous RNA-seq studies showed that microRNA-27b-3p (miR-27b-3p) might play an important role in regulating the proliferation and differentiation of chicken primary myoblasts (CPMs). However, the mechanism of miR-27b-3p regulating the proliferation and differentiation of CPMs is still unclear. In this study, the results showed that miR-27b-3p significantly promoted the proliferation of CPMs and inhibited the differentiation of CPMs. Then, myostatin (MSTN) was confirmed to be the target gene of miR-27b-3p by double luciferase reporter assay, RT-qPCR, and Western blot. By overexpressing and interfering with MSTN expression in CPMs, the results showed that overexpression of MSTN significantly inhibited the proliferation and differentiation of CPMs. In contrast, interference of MSTN expression had the opposite effect. This study showed that miR-27b-3p could promote the proliferation of CPMs by targeting MSTN. Interestingly, both miR-27b-3p and MSTN can inhibit the differentiation of CPMs. These results provide a theoretical basis for further understanding the function of miR-27b-3p in chicken and revealing its regulation mechanism on chicken muscle growth.

Breast cancer is a prevalent malignancy with high mortality and poor prognosis. Ropivacaine is a widely used local anesthetic and presents potential anti-tumor activity. Nevertheless, the function of ropivacaine in breast cancer development remains elusive. Here, we tried to investigate the impact of ropivacaine on breast cancer progression and the underlying mechanism. Significantly, we revealed that ropivacaine was able to reduce the proliferation and induce the apoptosis of breast cancer cells. Ropivacaine could attenuate the invasion and migration in the cells. Mechanically, ropivacaine could enhance the miR-27b-3p expression and miR-27b-3p inhibited breast cancer progression in breast cancer cells. MiR-27b-3p targeted YAP in the breast cancer cells. Ropivacaine decreased the breast cancer progression by modulating miR-27b-3p/YAP axis in vitro. Ropivacaine could inhibit tumor growth in vivo. In conclusion, we discovered that the local anesthetic ropivacaine inhibits the progression of breast cancer via the miR-27b-3p/YAP axis. Our finding presents novel insights into the mechanism of ropivacaine inhibiting the development of breast cancer. Ropivacaine may potentially serve as an anti-tumor candidate in the therapeutic strategy of breast cancer.

Background Long noncoding RNA nuclear-enriched abundant transcript 1 (NEAT1) exhibits an oncogenic role in multiple cancers, including gastric cancer (GC). But, the functions of NEAT1 in modulating radio-sensitivity of GC and its potential molecular mechanisms have not been totally elucidated. Methods qRT-PCR was performed to detect the expressions of NEAT1 and microRNA-27b-3p (miR-27b-3p). Kaplan–Meier survival curves for NEAT1 expression in GC created using KM Plotter. Colony formation assay was used to determine the survival fraction. Cell apoptosis was evaluated by flow cytometry. Luciferase reporter assay was used to verify the relationship between miR-27b-3p and NEAT1. Results NEAT1 was highly expressed while miR-27b-3p was downregulated in GC tissues and cells. NEAT1 was negatively correlated with that of miR-27b-3p and associated with poor overall survival. Moreover, NEAT1 and miR-27b-3p varied inversely after radiation in GC tissues and cells. Loss of NEAT1 or upregulation of miR-27b-3p increased the effect of radiation on cell survival fraction inhibition and apoptosis promotion. In addition, NEAT1 negatively regulated the expression of miR-27b-3p in GC cells. Interestingly, the depletion of miR-27b-3p dramatically attenuated the NEAT1 knockdown-mediated function in AGS and MKN-45 cells treated with radiation in vitro. Similarly, downregulation of NEAT1 enhanced the radiation-mediated inhibition of tumor growth, which was mitigated by decrease of miR-27b-3p. Conclusion NEAT1 depletion enhanced radio-sensitivity of GC by negatively regulating miR-27b-3p in vitro and in vivo.

Purpose: Catenin Beta 1 (CTNNB1) is a key regulator of cell proliferation and invasion in endometriosis; however, its upstream factor is not clear. Long noncoding RNAs may participate in endometriosis. The aim of this study was to investigate the mechanism of interaction between LINC02381 and CTNNB1 in endometriosis. Method: Screening and validation of RNAs were completed by whole transcriptional sequencing and qRT-PCR. The subcellular localization of LINC02381 was determined by RNA in situ hybridization and nucleo-cytoplasmic separation. Plasmids were transfected for functional experiments. Luciferase assay was used to verify the binding relationship. Results: The expression of LINC02381 and CTNNB1 was significantly increased in ovarian ectopic endometrial tissues (OSAs) and ectopic endometrial stromal cells (ESCs). When LINC02381 was downregulated in ESCs, the expression of CTNNB1, metallopeptidase 9 (MMP9) and cyclinD1, as well as ESCs invasion and proliferation, decreased. LINC02381 was mainly present in the cytoplasm of ESCs, indicating that it may act as a competitive endogenous RNA. Bioinformatic analysis revealed that microRNA-27b-3p (miR-27b-3p) is a downstream target of LINC02381. miR-27b-3p decreased in OSAs and ESCs. Moreover, when miR-27b-3p was upregulated in ESCs, the expression of CTNNB1, MMP9 and cyclinD1, as well as the invasion and proliferation ability of ESCs, were reduced. Additionally, rescue experiments demonstrated that the expression of CTNNB1, MMP9 and cyclinD1, as well as the invasion and proliferation ability, were significantly increased in the group transfected with both sh-LINC02381 and a miR-27b-3p inhibitor. Conclusion: LINC02381 upregulated CTNNB1 by adsorbing miR-27b-3p, causing increased proliferation and invasion of ESCs.

Gastric cancer (GC) is one of the most common aggressive cancers. The discovery of an effective biomarker is necessary for GC diagnosis. In this study, we confirmed that Paired box gene 4 (PAX4) is up-regulated in GC tissues and cells via quantitative real time polymerase chain reaction (qRT-PCR), western blot and immunohistochemical staining. It was also identified that PAX4 contributed to GC cell proliferation, migration and invasion through Cell Counting Kit-8, BrdU, flow cytometry assay, colony formation assay, transwell assays, and wound healing assay. miR-27b-3p was confirmed with the binding site with PAX4 using ChIP assay and served as a tumor suppressor that inhibiting GC cell growth and metastasis, and reversed the effect of PAX4. Bioinformatics prediction and dual luciferase assay results demonstrated that miR-27b-3p targeted Grb2, which could alter the function of miR-27b-3p. Furthermore, the transcriptional control of PAX4-regulated miR-27b-3p activated the Ras-ERK pathway. Taken together, the PAX4/miR-27b-3p/Grb2 loop is known to be involved in GC cell promotion, and can be seen as a promising target for GC therapy.

Lei Li, Guohua Lv, Bing Wang, Hong Ma Department of Spine Surgery, The Second Xiangya Hospital, Central South University, Changsha, Hunan 410011, People’s Republic of ChinaCorrespondence: Hong Ma Tel +86 731-85295124Email spinema@csu.edu.cnBackground: Chordoma, a rare bone tumor, occurs most commonly at the sacrococcygeal and skull base region. To date, chemotherapy is used to treat patients with advanced-stage chordoma. However, multidrug resistance (MDR) greatly hinders the effect of chemotherapy in chordoma. Here, we studied the correlation between KCNQ1OT1 and chemotherapy resistance.Methods: RT-PCR assay was used to examine KCNQ1OT1, miR-27b-3p, and ATF2 mRNA expression. CCK8 assay was exercised to detect IC50 values of cisplatin in chordoma cells. ATF2 protein expression was detected by Western blot.Results: KCNQ1OT1 was increased in chemotherapy-resistant patients and cisplatin-resistant cells, and downregulation of KCNQ1OT1 expression weakened MDR in chordoma. In addition, KCNQ1OT1 promoted MDR in chordoma by sponging miR-27b-3p and subsequently increasing ATF2 expression.Conclusion: KCNQ1OT1 is proved to be strikingly raised in the chemotherapy-resistant group and to promote MDR in chordoma. Our findings demonstrated the role of the KCNQ1OT1/miR-27b-3p/ATF2 axis in MDR of chordoma, which provides new insight into the molecular mechanism of chordoma MDR, and may determine the effect of therapy after receiving chemotherapy by detecting the expression of KCNQ1OT1 in serum.Keywords: KCNQ1OT1, MDR, miR-27b-3p, ATF2, chordoma

Renal fibrosis is a pathologic change in chronic kidney disease (CKD). MicroRNAs (miRNAs) have been shown to play an important role in the development of renal fibrosis. However, the biological role of miR-27b-3p in renal fibrosis remains unclear. Thus, this study aimed to investigate the role of miR-27b-3p in the progression of renal fibrosis. In this study, HK-2 cells were stimulated with transforming growth factor (TGF)-β1 for mimicking fibrosis progression in vitro. The unilateral ureteric obstruction (UUO)-induced mice renal fibrosis in vivo was established as well. The results indicated that the overexpression of miR-27b-3p significantly inhibited epithelial-to-mesenchymal transition (EMT) in TGF-β1-stimulated HK-2 cells, as shown by the decreased expressions of α-SMA, collagen III, Fibronectin and Vimentin. In addition, overexpression of miR-27b-3p markedly decreased TGF-β1-induced apoptosis in HK-2 cells, as evidenced by the decreased levels of Fas, active caspase 8 and active caspase 3. Meanwhile, dual-luciferase assay showed that miR-27b-3p downregulated signal transducers and activators of transcription 1 (STAT1) expression through direct binding with the 3′-UTR of STAT1. Furthermore, overexpression of miR-27b-3p attenuated UUO-induced renal fibrosis via downregulation of STAT1, α-SMA and collagen III. In conclusion, miR-27b-3p overexpression could alleviate renal fibrosis via suppressing STAT1 in vivo and in vitro. Therefore, miR-27b-3p might be a promising therapeutic target for the treatment of renal fibrosis.

Purpose: Dysregulation of microRNAs (miRNAs) contributes to tumor progression via the regulation of the expression of specific oncogenes and tumor suppressor genes. One such example, miR-27b-3p, has reportedly been involved in tumor progression in many types of cancer. The aim of the present study was to delve into the role and the underlying mechanism of miR-27b-3p in colorectal cancer (CRC) cells. Methods: In the present study, we detected the expression level of miR-27b-3p by RT-PCR. The effect of miR-27b-3p overexpression on cell proliferation in CRC cells was evaluated by cell counting and Edu assays. Transwell migration and invasion assays were used to examine the effects of cell migration and invasion. Bioinformatics, luciferase reporter assay and western blot assay were performed to identify the target of miR-27b-3p. Results: Here, we have demonstrated that although miR-27b-3p can affect cell morphology, it has no observable effect on the proliferation of CRC cells. However, it significantly promotes the migration and invasion of CRC cells. We discovered that HOXA10 was a newly identified target of miR-27b-3p in CRC cells, as confirmed by bioinformatics, western blots and dual luciferase reporter assay. Furthermore, the overexpression of miR-27b-3p or the suppression of HOXA10 can activate the integrin β1 signaling pathway. In conclusion, our results reveal a new function of miR-27b-3p that demonstrates its ability to promote CRC cell migration and invasion by targeting the HOXA10/integrin β1 cell signal axis. Conclusion: This may provide a mechanism to explain why miR-27b-3p promotes CRC cell migration and invasion.

Long non‑coding RNAs (lncRNAs) contribute to the tumorigeneses of numerous types of cancer, including glioma. The present study was designed to unveil a novel lncRNA functioning in glioma and explore the underlying mechanisms. lncRNA titin‑antisense RNA1 (TTN‑AS1), miR‑27b‑3p and Runt‑related transcription factor 1 (RUNX1) expression in glioma tissues and cell lines was estimated by RT‑qPCR. Si‑TTN‑AS1 was transfected into glioma cell lines (U251 and LN229), and CCK‑8 assay, flow cytometry, wound healing and Transwell assays were applied to estimate the function of TTN‑AS1 in glioma cells. miR‑27b‑3p inhibitor was used to explore the mechanisms. The results revealed that TTN‑AS1 was highly expressed in glioma specimens and cell lines. Downregulation of TTN‑AS1 inhibited the proliferation, migration and invasion of the glioma cells, as well as increased the rate of apoptosis. In vivo, the tumor growth was also inhibited by TTN‑AS1 depletion in nude mice. Furthermore, we revealed that TTN‑AS1 exerted oncogenic effects via sponging miR‑27b‑3p and thereby positively regulating RUNX1 expression. In conclusion, the present study supported that TTN‑AS1 acts as an oncogene in glioma by targeting miR‑27b‑3p to release RUNX1. This finding may contribute to gene therapy of glioma.

Qingfeng Jiang, Wenqun Xing, Jinhua Cheng, Yongkui Yu Department of Thoracic Surgery, The Affiliated Cancer Hospital of Zhengzhou University, Henan Cancer Hospital, Zhengzhou, Henan, People’s Republic of ChinaCorrespondence: Wenqun XingDepartment of Thoracic Surgery, The Affiliated Cancer Hospital of Zhengzhou University, Henan Cancer Hospital, No. 127 Dongming Road, Zhengzhou 450008, Henan, People’s Republic of ChinaTel +86-0371-65588265Email kcbbgn@163.comPurpose: Long non-coding RNA P73 antisense RNA 1T (TP73-AS1) is a newly discovered lncRNA involved in the occurrence and development of several cancers. However, its role in lung cancer has not been well investigated yet.Methods: The expressions of TP73-AS1, microRNA-27b-3p (miR-27b-3p) and lysosomal-associated protein transmembrane-4 Beta (LAPTM4B) were detected by quantitative real-time polymerase chain reaction (qRT-PCR). The cell proliferation, apoptosis, migration and invasion were detected by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), Annexin V-FITC/PI and transwell assays, respectively. Tumor xenografts were applied to explore the role of TP73-AS1 in vivo. The target relationship was predicted by StarBase v.2.0 or TargetScan and confirmed by luciferase reporter assay. Pearson’s coefficient assay was applied to assess the expression correlation between two groups. Protein expression levels were detected by Western blot.Results: We found that TP73-AS1 was strikingly up-regulated in lung cancer tissues and cells. TP73-AS1 depletion inhibited the growth and metastasis of lung cancer cells in vitro. Furthermore, TP73-AS1 could act as an endogenous sponge by directly binding miR-27b-3p, and a notable inverse correlation between them was also discovered. Importantly, knockdown of miR-27b-3p could reverse the inhibitory effects of TP73-AS1 depletion on the growth and metastasis of lung cancer cells. Besides, LAPTM4B was directly targeted by miR-27b-3p and could be co-regulated by TP73-AS1 and miR-27b-3p in lung cancer cells. Silencing TP73-AS1 hampered tumor growth by regulating miR-27b-3p/LAPTM4B axis in vivo.Conclusion: TP73-AS1 promoted the progression of lung cancer through regulating miR-27b-3p/LAPTM4B axis and it might be a potential target for diagnosis and treatment of lung cancer.Keywords: lung cancer, TP73-AS1, miR-27b-3p, LAPTM4B, proliferation, metastasis

Objective: Long-term dysregulation of energy balance is the key component of the obesity epidemic. Given the harm of central obesity and the discovery that beige cells appear within white adipose tissue (WAT), enhancing the energy-expending or “browning” ability of visceral adipose tissue (VAT) has become of therapeutic interest. In this study, we focused on the regulating role of microRNA (miRNA)-27b-3p in mice epididymal white adipose tissue (eWAT) browning. Methods: High-fat diet (HFD) induced obese mice model was constructed. Expression of miR-27b-3p and Ucp1 in eWAT was measured during the course of HFD. Through tail vein injection of antimiR-27b-3p, miR-27b-3p expression was inhibited to analyze the potential role of miR-27b-3p in fat browning and metabolism. Results: miR-27b-3p was predominantly expressed in eWAT and browning ability of eWAT in HFD induced obese mice was impaired. Inhibition of miR-27b-3p enhanced browning capacity of eWAT in mice fed an HFD and led to weight loss and insulin sensitivity improvement. Conclusions: High expression of miR-27b-3p in eWAT inhibits browning ability and leads to visceral fat accumulation. It is suggested miR-27b-3p may become a potential therapeutic option for visceral obesity and its associated diseases.

Background Epigenetics play a vital role in the initiation and development of cancers, including gastric cancer (GC). In the present study, we aimed to explore potential up- and downstream mechanisms of miR-27b-3p in GC. Methods The expression level of miR-27b-3p in GC cells and tissues (n = 80) was measured by quantitative RT-PCR. The mimics, inhibitors, and negative controls of miR-27b-3p were transfected into cell lines to perform the gain and loss of function study. Cell proliferation, migration, and invasion assays were utilized to assess biological behaviors caused by miR-27b-3p in vitro. Common target genes were predicted using four biological software programs and used for gene functional enrichment analysis. GSPT1 was selected for target gene verification using dual luciferase assays and its expression level was detected by western blot. The MKN-45 cell line was treated with 5-aza-2′-deoxycytidine (5-Aza-dC) and the methylation level was measured by methylation-specific PCR (MSP). Results miR-27b-3p was significantly downregulated in the GC cell lines and tissues compared with the normal group. The expression of miR-27b-3p was determined to be negatively associated with TNM stage and tumor size using statistical analysis. Overexpression of miR-27b-3p inhibited MKN-45 and SGC-7901 cell proliferation, invasion, and migration. Gene functional enrichment analysis indicated that the target genes were involved in several signaling pathways. Dual luciferase assays showed that miR-27b-3p combined with the 3′-untranslated region of GSPT1 mRNA. MSP demonstrated that miR-27b-3p promoter CpG island was hyper-methylated and 5-Aza-dC was able to partially reverse the methylation. Conclusions Our study data indicated that miR-27b-3p is downregulated by aberrant DNA methylation in GC. In addition, miR-27b-3p suppresses GC cell proliferation, invasion, and migration via negative expression regulation of GSPT1, which could be a potential therapeutic target.

Background The receptor tyrosine kinase-like orphan receptors (ROR) family contains the atypical member ROR1, which plays an oncogenic role in several malignant tumors. However, the clinical potentials and underlying mechanisms of ROR1 in gastric cancer progression remain largely unknown. In this study, we validated the microRNA-mediated gene repression mechanism involved in the role of ROR1. Methods Bioinformatic prediction, luciferase reporter assay, quantitative real-time PCR (qRT-PCR) and western blotting were used to reveal the regulatory relationship between miR-27b-3p and ROR1. The expression patterns of miR-27b-3p and ROR1 in human gastric cancer (GC) specimens and cell lines were determined by microRNA RT-PCR and western blotting. Cell proliferation, colony formation assay in soft agar in vitro and tumorigenicity in vivo were performed to observe the effects of downregulation and upregulation miR-27b-3p expression on GC cell phenotypes. Results miR-27b-3p suppressed ROR1 expression by binding to the 3’UTR of ROR1 mRNA in GC cells. miR-27b-3p was significantly downregulated and reversely correlated with ROR1 protein levels in clinical samples. Analysis of the clinicopathological significance showed that miR-27b-3p and ROR1 were closely correlated with GC characteristics. Ectopic miR-27b-3p expression suppressed cell proliferation, colony formation in soft agar, xenograft tumors of GC cells. By contrast, miR-27b-3p knockdown enhanced these malignant behaviors. Our studies further revealed that the c-Src/STAT3 signaling pathway was involved in miR-27b-3p-ROR1-mediated cell proliferation regulation. Conclusions These results show that miR-27b-3p suppresses ROR1 expression through the binding site in the 3’UTR inhibiting the cell proliferation. These findings indicate that miR-27b-3p exerts tumor-suppressive effects in GC through the suppression of oncogene ROR1 expression and suggest a therapeutic application of miR-27b-3p in GC.