LINC00342 regulates the PI3K-AKT signaling pathway via the miR-149-5p/FGF11 axis and affects the progression of oral cancer.

BACKGROUND: A large number of long non-coding RNAs (lncRNAs) have been implicated in the progression of oral cancer (OC). This study aimed to investigate the role of a novel lncRNA, LINC00342, in OC and elucidate its molecular mechanism. METHODS: Differential expression of lncRNA/miRNA/mRNA was analyzed using the Gene Expression Omnibus database and validated with RT-qPCR. Additionally, the expression levels of these molecules in OC cells and their effects on cell viability and cell cycle were assessed using the Cell Counting Kit-8 and flow cytometry. RNA bindings was analyzed by dual luciferase, and Western blot was used to detect the activation of relevant pathways. RESULTS: This study showed that, in contrast to miR-149-5p, the expression of LINC00342 and fibroblast growth factor 11 (FGF11) were upregulated in OC cells (LINC00342: 10.00 ± 1.06 (FaDu) and 3.55 ± 0.25 (CAL-27) vs 1.00 ± 0.07 (HOECs), P < 0.05; FGF11: 7.31 ± 0.33 (FaDu) and 3.43 ± 0.08 (CAL-27) vs 1.00 ± 0.10 (HOECs), P < 0.05). Dual-luciferase assays confirmed that LINC00342 bind to miR-149-5p in a direct targeting manner. Furthermore, inhibition of LINC00342 expression resulted in decreased proliferation rate (FaDu: 136.22 ± 22.10% vs 59.36 ± 8.98% (control), P < 0.05; CAL-27: 131.40 ± 11.58% vs 49.83 ± 11.19 (control), P < 0.05) and migration ability of OC cells, cell cycle arrest in G1 phase, and inhibition of PI3K-AKT signaling. Inhibition of miR-149-5p or overexpression of FGF11 reversed the effects of si-LINC00342. CONCLUSIONS: LINC00342 promotes PI3K-AKT signaling by activating FGF11 through adsorption of miR-149-5p, thereby regulating the progression of OC.

Hepatic miR-149-5p upregulation fosters steatosis, inflammation and fibrosis development in mice and in human liver organoids.

Background & Aims: The incidence of metabolic dysfunction-associated steatotic liver disease (MASLD) is increasing worldwide. Alterations of hepatic microRNA (miRNA) expression/activity significantly contribute to the development and progression of MASLD. Genetic polymorphisms of miR-149 are associated with an increased susceptibility to MASLD development in humans. Aberrant expression of miR-149 was also associated with metabolic alterations in several organs, but the impact of hepatic miR-149-5p deregulation in MASLD remains poorly characterized. Methods: MiR-149-5p was downregulated in the livers of mice by in vivo transduction with hepatotropic adeno-associated virus 8 harboring short-hairpin RNAs (shRNAs) specific for miR-149-5p (shmiR149) or scrambled shRNAs (shCTL). MASLD was then induced with a methionine/choline-deficient (MCD, n = 7 per group) diet or a fructose/palmitate/cholesterol-enriched (FPC, n = 8-12 per group, per protocol) diet. The impact of miR-149-5p modulation on MASLD development was assessed in vivo and in vitro using multi-lineage 3D human liver organoids (HLOs) and Huh7 cells. Results: MiR-149-5p expression was strongly upregulated in mouse livers from different models of MASLD (2-4-fold increase in ob/ob, db/db mice, high-fat and FPC-fed mice). In vivo downregulation of miR-149-5p led to an amelioration of diet-induced hepatic steatosis, inflammation/fibrosis, and to increased whole-body fatty acid consumption. In HLOs, miR-149-5p overexpression promoted lipid accumulation, inflammation and fibrosis. In vitro analyses of human Huh7 cells overexpressing miR-149-5p indicated that glycolysis and intracellular lipid accumulation was promoted, while mitochondrial respiration was impaired. Translatomic analyses highlighted deregulation of multiple potential miR-149-5p targets in hepatocytes involved in MASLD development. Conclusions: MiR-149-5p upregulation contributes to MASLD development by affecting multiple metabolic/inflammatory/fibrotic pathways in hepatocytes. Our results further demonstrate that HLOs are a relevant 3D in vitro model to investigate hepatic steatosis and inflammation/fibrosis development. Impact and implications: Our research shows compelling evidence that miR-149-5p plays a pivotal role in the development and progression of MASLD. By employing in vivo and innovative in vitro models using multi-lineage human liver organoids, we demonstrate that miR-149-5p upregulation significantly impacts hepatocyte energy metabolism, exacerbating hepatic steatosis and inflammation/fibrosis by modulating a wide network of target genes. These findings not only shed light on the intricate miR-149-5p-dependent molecular mechanisms underlying MASLD, but also underscore the importance of human liver organoids as valuable 3D in vitro models for studying the disease's pathogenesis.

Long non-coding RNA LNC-POTEM-4 promotes HCC progression via the LNC-POTEM-4/miR-149-5p/Wnt4 signaling axis.

Information on the potential role of the long non-coding RNA LNC-POTEM-4 in cancer progression is limited. Our preliminary study found that LNC-POTEM-4 was overexpressed in hepatocellular carcinoma (HCC) tissues, which led us to further investigate the biological function and molecular mechanism of LNC-POTEM-4 in HCC development. LNC-POTEM-4 expression in HCC tissues was examined using transcriptome sequencing and quantitative reverse transcription PCR. The relationships between LNC-POTEM-4 and the stage and prognosis of HCC in patient data from the TCGA database were analyzed. The effects of LNC-POTEM-4 on proliferation, invasion/migration, and epithelial-mesenchymal transition marker expression in HCC cells were evaluated in vitro using gain- and loss-of-function assays, while its effects on tumor growth and metastasis were explored through animal experiments. A LNC-POTEM-4/microRNA (miR)-149-5p/Wnt4 regulatory signaling axis was identified using bioinformatics analysis, and dual luciferase reporter, RNA immunoprecipitation, and RNA pull-down assays. Co-transfection of LNC-POTEM-4 and Wnt4 expression plasmids was employed to confirm the new signaling pathway. We found that LNC-POTEM-4 was overexpressed in HCC tissues and was linked to poor staging and prognosis. LNC-POTEM-4 promoted proliferation, invasion, migration, and the epithelial-mesenchymal transition of HCC cells in vitro. Silencing of LNC-POTEM-4 inhibited HCC growth and distant metastasis in vivo. Mechanically, LNC-POTEM-4 was found to function as a competitive endogenous RNA, upregulating Wnt4 by sponging miR-149-5p to promote HCC progression. Wnt4 overexpression may have counteracted the tumor-inhibition effect of LNC-POTEM-4 silencing. In conclusion, LNC-POTEM-4 upregulated Wnt4 to activate the Wnt signaling pathway and stimulate the malignancy tendency of HCC by sponging miR-149-5p, providing a prospective target for the detection and therapy of HCC. However, the effects of LNC-POTEM-4 on the miR-149-5p/Wnt4 signaling axis should be further studied in animal experiments.

Distinct Effects of Respiratory Viral Infection Models on miR-149-5p, IL-6 and p63 Expression in BEAS-2B and A549 Epithelial Cells.

Respiratory viruses cause airway inflammation, resulting in epithelial injury and repair. miRNAs, including miR-149-5p, regulate different pathological conditions. We aimed to determine how miR-149-5p functions in regulating pro-inflammatory IL-6 and p63, key regulators of airway epithelial wound repair, in response to viral proteins in bronchial (BEAS-2B) and alveolar (A549) epithelial cells. BEAS-2B or A549 cells were incubated with poly (I:C, 0.5 µg/mL) for 48 h or SARS-CoV-2 spike protein-1 or 2 subunit (S1 or S2, 1 µg/mL) for 24 h. miR-149-5p was suppressed in BEAS-2B challenged with poly (I:C), correlating with IL-6 and p63 upregulation. miR-149-5p was down-regulated in A549 stimulated with poly (I:C); IL-6 expression increased, but p63 protein levels were undetectable. miR-149-5p remained unchanged in cells exposed to S1 or S2, while S1 transfection increased IL-6 expression in BEAS-2B cells. Ectopic over-expression of miR-149-5p in BEAS-2B cells suppressed IL-6 and p63 mRNA levels and inhibited poly (I:C)-induced IL-6 and p63 mRNA expressions. miR-149-5p directly suppressed IL-6 mRNA in BEAS-2B cells. Hence, BEAS-2B cells respond differently to poly (I:C), S1 or S2 compared to A549 cells. Thus, miR-149-5p dysregulation may be involved in poly (I:C)-stimulated but not S1- or S2-stimulated increased IL-6 production and p63 expression in BEAS-2B cells.

HIF-2α/LINC02609/APOL1-mediated lipid storage promotes endoplasmic reticulum homeostasis and regulates tumor progression in clear-cell renal cell carcinoma.

BACKGROUND: The VHL-HIF pathway and lipid droplet accumulation are the main characteristics of clear cell renal cell carcinoma (ccRCC). However, the connection between the two features is largely unknown. METHODS: We used transcriptional sequencing and TCGA database analysis to identify APOL1 as a novel therapeutic target for ccRCC. The oncogenic functions of APOL1 were investigated by cell proliferation, colony formation, migration and invasion assays in ccRCC cells in vitro and xenografts derived from ccRCC cells in vivo. Oil red O staining and quantification were used to detect lipid droplets. Chromatin immunoprecipitation (ChIP) assays and luciferase reporter assays were carried out to identify HIF-2α bound to the promoter of APOL1 and lncRNA LINC02609. RNA-FISH and luciferase reporter assays were performed to determine that LncRNA LINC02609 functions as a competing endogenous RNA to regulate APOL1 expression by sponging miR-149-5p. FINDINGS: RNA-seq data revealed that HIF2α can regulate APOL1 and lncRNA LINC02609 expression. We also found that HIF-2α can bind to the promoter of APOL1 and lncRNA LINC02609 and transcriptionally regulate their expression directly. We further demonstrated that LncRNA LINC02609 functions as a competing endogenous RNA to regulate APOL1 expression by sponging miR-149-5p in ccRCC. Mechanistically, APOL1-dependent lipid storage is required for endoplasmic reticulum (ER) homeostasis and cell viability and metastasis in ccRCC. We also showed that high APOL1 expression correlated with worse clinical outcomes, and knockdown of APOL1 inhibited tumor cell lipid droplet formation, proliferation, metastasis and xenograft tumor formation abilities. Together, our studies identify that HIF2α can regulate the expression of the lipid metabolism related gene APOL1 by direct and indirect means, which are essential for ccRCC tumorigenesis. INTERPRETATION: Based on the experimental data, in ccRCC, the HIF-2α/LINC02609/APOL1 axis can regulate the expression of APOL1, thus interfering with lipid storage, promoting endoplasmic reticulum homeostasis and regulating tumor progression in ccRCC. Together, our findings provide potential biomarkers and novel therapeutic targets for future studies in ccRCC.

CircUCK2 promotes hepatocellular carcinoma development by upregulating UCK2 in a mir-149-5p-dependent manner.

BACKGROUND: Circular RNAs (circRNAs) participate in the regulation of Hepatocellular Carcinoma (HCC) progression. The objective of this study was to explore the function and mechanism of circUCK2 in HCC development. METHODS: The RNA levels of circUCK2, miR-149-5p and uridine-cytidine kinase 2 (UCK2) were examined by quantitative real-time polymerase chain reaction (qRT-PCR). EdU incorporation assay and colony formation assay were respectively performed to analyze cell proliferation and colony formation. Wound healing assay and transwell assay were conducted for cell migration and invasion. Flow cytometry was used for cell apoptosis analysis. Western blot assay was conducted to determine the protein levels of E-cadherin, N-cadherin, matrix metallopeptidase 9 (MMP-9) and UCK2. Dual-luciferase reporter assay, RNA immunoprecipitation (RIP) assay and RNA pull-down assay were conducted to confirm the interaction between miR-149-5p and circUCK2 or UCK2. The xenograft model was established to explore the role of circUCK2 in tumor growth in vivo. RESULTS: CircUCK2 level was elevated in HCC, and circUCK2 depletion suppressed HCC cell proliferation, colony formation, migration and invasion and accelerated cell apoptosis. Mechanistically, circUCK2 could positively modulate UCK2 expression by interacting with miR-149-5p. Furthermore, the repressive effects of circUCK2 knockdown on the malignant behaviors of HCC cells were alleviated by UCK2 overexpression or miR-149-5p inhibition. The promoting effects of circUCK2 overexpression on HCC cell malignancy were alleviated by UCK2 silencing or miR-149-5p introduction. Additionally, circUCK2 knockdown hampered tumor growth in vivo. CONCLUSION: CircUCK2 contributed to HCC malignant progression in vitro and in vivo via targeting miR-149-5p/UCK2 axis, demonstrating that circUCK2 might be a novel therapeutic target for HCC.

PTOV1-AS1 desensitizes colorectal cancer cells to 5-FU through depressing miR-149-5p to activate the positive feedback loop with Wnt/ß-catenin pathway.

5-Fluorouracil is a commonly used chemotherapy drug for colorectal cancer. Resistance to 5-Fluorouracil remains a challenge. This research aimed to explore the mechanism of 5-Fluorouracil resistance in colorectal cancer. RT-qPCR and Western blot were used to determine the RNA and protein expression in both cells and exosome. Assays in vitro and in vivo were performed to measure the role of miR-149-5p in colorectal cancer cells. RIP, luciferase activity report, and RNA pulldown assay were applied to detect the association of PTOV1-AS1, SUV39H1, miR-149-5p, and FOXM1. MiR-149-5p was down-expressed in 5-Fluorouracil-resistant cells. MiR-149-5p enhanced the effectiveness of 5-Fluorouracil both in vitro and in vivo. Sensitive colorectal cancer cells released exosomal miR-149-5p to sensitize resistant cells to chemotherapy. Mechanistically, miR-149-5p targeted the FOXM1 to inactivate Wnt/ß-catenin pathway, and PTOV1-AS1 recruited SUV39H1 to suppress miR-149-5p transcription, in turn activating Wnt/ß-catenin pathway, and forming a positive feedback loop with FOXM1. PTOV1-AS1 inhibits miR-149-5p by a positive feedback loop with FOXM1-mediated Wnt/ß-catenin pathway, which provides insights into a potential novel target for enhancing the effectiveness of chemotherapy in colorectal cancer patients.

LINC00460 promotes neuroblastoma tumorigenesis and cisplatin resistance by targeting miR-149-5p/DLL1 axis and activating Notch pathway in vitro and in vivo.

Long noncoding RNAs (lncRNAs) have been demonstrated to participate in neuroblastoma cisplatin resistance and tumorigenesis. LncRNA LINC00460 was previously reported to play a critical regulatory role in many cancer development. Nevertheless, its role in modulating neuroblastoma cisplatin resistance has not been explored till now. Cisplatin-resistant neuroblastoma cell lines were established by exposing neuroblastoma cell lines to progressively increasing concentrations of cisplatin for 6 months. LINC00460, microRNA (miR)-149-5p, and delta-like ligand 1 (DLL1) mRNA expression was measured through RT-qPCR. The protein levels of DLL1, epithelial-to-mesenchymal transition (EMT) markers, and the Notch signaling-related molecules were measured via western blotting. The IC50 value for cisplatin, cell growth, metastasis and apoptosis were analyzed in cisplatin-resistant neuroblastoma cells. The binding between LINC00460 (or DLL1) and miR-149-5p was validated through dual-luciferase reporter assay. The murine xenograft model was established to perform in vivo assays. LINC00460 and DLL1 levels were elevated, while miR-149-5p level was reduced in cisplatin-resistant neuroblastoma cells. LINC00460 depletion attenuated IC50 values for cisplatin, weakened cell growth, metastasis, and EMT, and enhanced apoptosis in cisplatin-resistant neuroblastoma cells. Mechanically, LINC00460 sponged miR-338-3p to increase DLL1 level, thereby activating Notch signaling pathway. DLL1 overexpression antagonized LINC00460 silencing-induced suppression on neuroblastoma cell cisplatin resistance and malignant behaviors, while such effects were further reversed by treatment with DAPT, the inhibitor of Notch pathway. Additionally, LINC00460 knockdown further augmented cisplatin-induced impairment on tumor growth in vivo. LINC00460 contributes to neuroblastoma cisplatin resistance and tumorigenesis through miR-149-5p/DLL1/Notch pathway, providing new directions to improve the therapeutic efficacy of chemotherapy drugs applied in patients with neuroblastoma.

Circ_0008410 contributes to fibroblast-like synoviocytes dysfunction by regulating miR-149-5p/HIPK2 axis.

Circular RNAs (circRNAs) play functional roles in rheumatoid arthritis (RA) progression. Fibroblast-like synoviocytes (RASFs) are the main effectors in RA development. In this study, we explored the function and mechanism of circ_0008410 in RASFs. qRT-PCR was used to detect the expression of circ_0008410, microRNA-149-5p (miR-149-5p), and homeodomain-interacting protein kinase 2 (HIPK2). Cell counting kit-8, EdU assay, flow cytometry, and transwell assay were performed to evaluate cell proliferation, apoptosis, migration, and invasion. Western blot measured the protein levels of related markers and HIPK2. The levels of IL-1ß, TNF-α, and IL-6 were tested by corresponding ELISA kits and Western blot. The combination between miR-149-5p and circ_0008410 or HIPK2 was detected by dual-luciferase reporter assay or RNA immunoprecipitation (RIP) assay. Our data showed that circ_0008410 and HIPK2 were elevated, while miR-149-5p was downregulated in RA synovial tissues and RASFs. Circ_0008410 promoted RASF proliferation, migration, invasion, and inflammation while inhibiting apoptosis. MiR-149-5p was a target of circ_0008410, and its overexpression could reverse the promoting effects of circ_0008410 on RASF dysfunction. Moreover, miR-149-5p could target HIPK2 to suppress RASF proliferation, migration, invasion, and inflammation. Collectively, circ_0008410 promoted RASF dysfunction via miR-149-5p/HIPK2, which might provide a potential target for RA therapy.

The hsa_circ_0004805/hsa_miR-149-5p/TGFB2 axis plays critical roles in the pathophysiology of diabetic retinopathy in vitro and in vivo.

The aim of this study was to investigate the mechanism underlying the role of a recently identified hsa_circ_0004805/hsa_miR-149-5p/transforming growth factor beta 2 (TGFB2) axis in the progression of diabetic retinopathy (DR). Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis suggested that hsa_circ_0004805 was highly expressed in aqueous humor samples of patients with DR, whereas hsa_miR-149-5p showed the opposite trend. Meanwhile, the results of a dual-luciferase reporter assay indicated that hsa_miR-149-5p directly interacted with both hsa_circ_0004805 and TGFB2. Using a variety of assays (Cell Counting Kit-8, EdU-labeling, Transwell, flow cytometric, wound healing, tube formation assays), we found that the overexpression of hsa_circ_0004805 significantly downregulated the level of hsa_miR-149-5p and promoted DNA synthesis, proliferation, migration, and tube formation in human retinal microvascular epithelial cells (hRECs) cultivated in a high-glucose environment. In contrast, hsa_miR-149-5p mimics inhibited DNA synthesis, proliferation, migration, and tube formation in hRECs by reducing the expression of its downstream target TGFB2 as well as the levels of phosphorylated SMAD2; however, these effects were reversed by the overexpression of hsa_circ_0004805. In a streptozotocin-induced Sprague-Dawley rat model of DR, retinal vascular leakage, capillary decellularization, loss of pericytes, fibrosis, and gliosis were evident, which could be reversed by vitreous microinjection of rat miR-149-5p mimics (rno-miR-149-5p agomir). Combined, our findings indicated that, under hyperglycemia, the hsa_circ_0004805/hsa_miR-149-5p/TGFB2 axis plays a critical role in the retinal pathophysiology associated with the development of DR, and has potential as a therapeutic target in the treatment of this condition.

Circular RNA 0000157 depletion protects human bronchial epithelioid cells from cigarette smoke extract-induced human bronchial epithelioid cell injury through the microRNA-149-5p/bromodomain containing 4 pathway.

BACKGROUND: Circular RNA (circRNA) has been reported to regulate respiratory diseases. In the study, we aimed to elucidate the role of circ_0000157 in smoke-related chronic obstructive pulmonary disease (COPD) and the inner mechanism. METHODS: COPD-like cell injury was induced by treating human bronchial epithelioid cells (16HBE) with cigarette smoke extract (CSE). The expression of circ_0000157, miR-149-5p, bromodomain containing 4 (BRD4), BCL2-associated x protein (Bax) and B-cell lymphoma-2 (Bcl-2) was analyzed by quantitative real-time polymerase chain reaction (qRT-PCR) or Western blotting. Enzyme-linked immunosorbent assay was performed to detect interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) levels. Malondialdehyde (MDA) production was detected by a lipid peroxidation MDA assay kit. Superoxide dismutase (SOD) activity was analyzed by a SOD activity assay kit. RESULTS: Circ_0000157 and BRD4 expression were upregulated, while miR-149-5p expression was downregulated in the blood of smokers with COPD and CSE-induced 16HBE cells compared with control groups. CSE treatment inhibited 16HBE cell proliferation and induced cell apoptosis, inflammation, and oxidative stress; however, these effects were remitted when circ_0000157 expression was decreased. In addition, circ_0000157 acted as a miR-149-5p sponge and regulated CSE-caused 16HBE cell damage by targeting miR-149-5p. The overexpression of BRD4, a target gene of miR-149-5p, attenuated the inhibitory effects of miR-149-5p introduction on CSE-induced cell damage. Further, circ_0000157 modulated BRD4 expression by associating with miR-149-5p in CSE-treated 16HBE cells. CONCLUSION: Circ_0000157 knockdown ameliorated CSE-caused 16HBE cell damage by targeting the miR-149-5p/BRD4 pathway, providing a potential therapeutic strategy for clinic intervention in COPD.

Two antisense RNAs-AFAP1-AS1 and MLK7-AS1-promote colorectal cancer progression by sponging miR-149-5p and miR-485-5p.

Colorectal cancer (CRC) is one of the leading causes of cancer-related deaths. Antisense RNAs (asRNAs) are closely associated with cancer malignancy. This study aimed to identify the action mechanism of asRNAs in controlling CRC malignancy. Analysis of the RNA sequencing data revealed that AFAP1-AS1 and MLK7-AS1 were upregulated in CRC patients and cell lines. High levels of both asRNAs were associated with poor prognosis in patients with CRC. Both in vitro and in vivo experiments revealed that the knockdown of the two asRNAs decreased the proliferative and metastatic abilities of CRC cells. Mechanistically, AFAP1-AS1 and MLK7-AS1 decreased the levels of miR-149-5p and miR-485-5p by functioning as ceRNAs. Overexpression of miRNAs by introducing miRNA mimics suppressed the expression of SHMT2 and IGFBP5 by directly binding to the 3' UTR of their mRNA. Knockdown of both asRNAs decreased the expression of SHMT2 and IGFBP5, which was reversed by inhibition of both miRNAs by miRNA inhibitors. In vivo pharmacological targeting of both asRNAs by small interfering RNA-loaded nanoparticles showed that knockdown of asRNAs significantly reduced tumor growth and metastasis. Our findings demonstrate that AFAP1-AS1 and MLK7-AS1 promote CRC progression by sponging the tumor-suppressing miRNAs miR-149-5p and miR-485-5p, thus upregulating SHMT2 and IGFBP5.

Pathogenesis of psoriasis via miR-149-5p/AKT1axis by long noncoding RNA BLACAT1.

BACKGROUND: Psoriasis is a chronic, complicated, and recurrent inflammatory skin disease, whose precise molecular mechanisms need to be further explored. The lncRNA bladder cancer-associated transcript 1 (BLACAT1) is aberrantly expressed in many cancers and associated with cellular hyperproliferation and may play a role in the pathogenesis of psoriasis. Thus, this study aimed at identifying the primary mechanism associated with BLACAT1 in psoriasis pathogenesis. MATERIALS AND METHODS: Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) was performed to detect the expression of BLACAT1 in psoriasis tissues. Cell proliferation and apoptosis were assessed using cell counting kit-8 and apoptosis assays, respectively. In vivo experiments and histopathological examinations were performed to investigate the effects of BLACAT1 on psoriasis. Dual-luciferase Reporter and RNA immunoprecipitation assays were used to evaluate the relationship among BLACAT1 and miR-149-5p and AKT1. RESULTS: BLACAT1 was upregulated in psoriasis tissues. Overexpression exacerbated the clinical manifestation of psoriasis and increased the epidermal thickness in imiquimod-induced mice. BLACAT1 could promote proliferation and inhibit apoptosis of keratinocytes. Further studies demonstrated that BLACAT1 positively regulated AKT1 expression, functioning as a competing endogenous RNA (ceRNA) by sponging miR-149-5p. CONCLUSIONS: The combination of lncRNA BLACAT1 and miR-149-5p regulates AKT1 expression and promotes psoriasis formation thus may provide a new direction for psoriasis treatment.

Effect of miR-149-5p on intramuscular fat deposition in pigs based on metabolomics and transcriptomics.

As one of the important traits in pig production, meat quality has important research significance and value. Intramuscular fat (IMF) content is one of the most important factors affecting pork quality. Many experimental studies have shown that IMF content is closely related to the flavor, tenderness, and juiciness of pork. Therefore, it is of great significance to study the mechanism of porcine IMF deposition. Previous research indicated that miR-149-5p promoted the proliferation of porcine intramuscular (IM) preadipocytes and decreased their ability to differentiate, albeit the exact mechanism of action is unknown. In vitro, foreign pigs showed increased miR-149-5p expression and reduced fat deposition when compared to Queshan Black pigs. This study conducted metabolomics and transcriptomics analyses of porcine IM preadipocytes overexpressing miR-149-5p to verify their effects on lipid formation. According to metabolomics analysis, the overexpression of miR-149-5p has significantly altered the lipid, organic acid, and organic oxygen metabolites of porcine IM preadipocytes. Specially speaking, it has changed 115 metabolites, including 105 up-regulated and 10 down-regulated ones, as well as the composition of lipid, organic acid, and organic oxygen metabolism-related metabolites. RNA-seq analysis showed that overexpression of miR-149-5p significantly altered 857 genes, of which 442 were up-regulated, and 415 were down-regulated, with enrichment to MAPK, IL-17, PI3K-Akt, and ErbB signaling pathways. We found that overexpression of miR-149-5p inhibited adipogenic differentiation by changing cAMP signaling pathway in porcine IM preadipocytes. In addition, the overexpression of miR-149-5p may affect the transport of Cu2+ by targeting ATP7A and inhibiting adipogenic differentiation. These findings elucidate the regulatory function of miR-149-5p in porcine IM preadipocytes, which may be a key target for controlling pork quality.

Hsa_circ_0001162 Inhibition Alleviates High Glucose-Induced Human Podocytes Injury by the miR-149-5p/MMP9 Signaling Pathway.

Emerging evidences suggested that circular RNAs (circRNAs) are involved in diabetic nephropathy (DN). Accumulating evidence had suggested that the degree of podocyte is a major prognostic determinant of DN progression. However, the function and in-depth mechanisms of hsa_circ_0001162 in podocyte injury of DN remain unclear. Hsa_circ_0001162 expression was detected by real-time quantitative PCR (RT-qPCR) in peripheral blood of DN patients and high glucose-induced podocytes injury model. The cell counting kit 8, 5-ethynyl-2'-deoxyuridine, flow cytometry with Annexin V-FITC/PI staining, caspase-3 activity assay Kit, enzyme linked immunosorbent assay (ELISA), RT-qPCR and western blotting were used to evaluate the effect of hsa_circ_0001162 / miR-149-5p / MMP9 axis on high glucose-induced podocyte injury. Mechanistically, dual luciferase reporter was used to confirm the relationship of miR-149-5p and hsa_circ_0001162 or MMP9. Furthermore, RNA-pull down and immunoprecipitation assay were implemented to verify the potential regulatory effects of EIF4A3 on biogenesis of hsa_circ_0001162. Our results showed that hsa_circ_0001162 was highly expressed in peripheral blood of DN patients and high glucose-induced podocytes injury model, and the knockdown of hsa_circ_0001162 increased the proliferation, inhibited the apoptosis, and suppressed inflammatory response in high glucose-induced podocytes injury. Mechanism studies demonstrated that EIF4A3 bound with flanking sequences of hsa_circ_0001162 to promote hsa_circ_0001162 expression, upregulated hsa_circ_0001162 increased the MMP9 expression via sponging miR-149-5p, thus aggravating the high glucose-induced podocytes injury. Overall, our data demonstrated that knockdown of hsa_circ_0001162 inhibited high glucose-induced podocytes injury by regulating miR-149-5p/MMP9 axis, and intervention of hsa_circ_0001162/miR-149-5p/MMP9 axis may be a potentially promising therapeutic strategy for podocyte injury in DN patients.

MiR-149-5p inhibits cell proliferation, promotes cell apoptosis and retards cell cycle of IL-22-stimulated HaCaT and NHEK keratinocytes via regulating PDE4D.

BACKGROUND: Psoriasis is a chronic autoimmune skin disease with unclear pathogenesis. It was found that miR-149-5p was significantly decreased in psoriatic lesion tissues. In this study, we aims to investigate the role and related molecular mechanism of miR-149-5p on psoriasis. METHOD: IL-22 was used to stimulate HaCaT and NHEK cells to establish psoriasis model in vitro. The miR-149-5p and phosphodiesterase 4D (PDE4D) expression levels were detected by quantitative real-time PCR. HaCaT and NHEK cells proliferation was determined by Cell Couting Kit-8 assay. The cell apoptosis and cell cycle were detected by flow cytometry. The cleaved Caspase-3, Bax and Bcl-2 protein expressions were detected by western blot. The targeting relationship between PDE4D and miR-149-5p was predicted and confirmed by Starbase V2.0 and dual-luciferase reporter assay, respectively. RESULT: There was a low expression level of miR-149-5p and a high expression of PDE4D in psoriatic lesion tissues. MiR-149-5p could target PDE4D. IL-22 promoted HaCaT and NHEK cells proliferation, while inhibited cell apoptosis and accelerated cell cycle. Moreover, IL-22 decreased the expressions of cleaved Caspase-3 and Bax, and increased the expression of Bcl-2. And the overexpressed miR-149-5p promoted HaCaT and NHEK cells apoptosis, inhibited cell proliferation and retarded cell cycle, meanwhile increased the cleaved Caspase-3 and Bax expressions, decreased the Bcl-2 expression. In addition, PDE4D overexpression has the opposite effect as miR-149-5p. CONCLUSION: The overexpressed miR-149-5p inhibits IL-22-stimulated HaCaT and NHEK keratinocytes proliferation, promotes cell apoptosis and retards cell cycle by down-regulating the expression of PDE4D, which could be the promising therapeutic target of psoriasis.

microRNA-149-5p mediates the PM2.5-induced inflammatory response by targeting TAB2 via MAPK and NF-κB signaling pathways in vivo and in vitro.

Epidemiological evidence has shown that fine particulate matter (PM2.5)-triggered inflammatory cascades are pivotal causes of chronic obstructive pulmonary disease (COPD). However, the specific molecular mechanism involved in PM2.5-induced COPD has not been clarified. Herein, we found that PM2.5 significantly downregulated miR-149-5p and activated the mitogen-activated protein kinase (MAPK) and nuclear factor-kappa B (NF-κB) signaling pathways and generated the inflammatory response in COPD mice and in human bronchial epithelial (BEAS-2B) cells. We determined that increased expression of interleukin-1ß (IL-1ß), IL-6, IL-8, and tumor necrosis factor-α (TNF-α) induced by PM2.5 was associated with decreased expression of miR-149-5p. The loss- and gain-of-function approach further confirmed that miR-149-5p could inhibit PM2.5-induced cell inflammation in BEAS-2B cells. The double luciferase reporter assay showed that miR-149-5p directly targeted TGF-beta-activated kinase 1 binding protein 2 (TAB2), which regulates the MAPK and NF-κB signaling pathways. We showed that miR-149-5p mediated the inflammatory response by targeting the 3'-UTR sequence of TAB2 and that it subsequently weakened the TAB2 promotor effect via the MAPK and NF-κB signaling pathways in BEAS-2B cells exposed to PM2.5. Thus, miR-149-5p may be a key factor in PM2.5-induced COPD. This study improves our understanding of the molecular mechanism of COPD.

Deficiency of miRNA-149-3p shaped gut microbiota and enhanced dextran sulfate sodium-induced colitis.

Genetic predisposition and disruption of host gut microbiota and immune system can result in inflammatory bowel disease (IBD). Here, we show that miRNA-149-5p (miR-149-5p) and miRNA-149-3p (miR-149-3p) play crucial roles in IBD. Mice lacking miR-149-3p were considerably more susceptible to dextran sulfate sodium (DSS)-induced colitis than wild-type (WT) mice, accompanied by more serious inflammatory symptoms and increased gene expression of certain inflammatory cytokines. Both miR-149-5p and miR-149-3p suppressed colon inflammatory response in vitro and in vivo. Furthermore, we found significant differences in the composition of the gut microbiota between WT and miR-149-3p-/- mice by 16S rRNA sequencing. Co-housing endowed susceptibility to WT mice against DSS-induced colitis compared with the WT control group. However, susceptibility of miR-149-3p-/- mice against DSS-induced colitis was still present after antibiotic treatment. These findings suggest that the deletion of miR-149-3p altered gut microbiota and influenced pathogenesis of intestinal inflammation, but sensitivity of miR-149-3p-/- mice to DSS-induced colitis is not conferred by microbiota. In addition, we identified the roles of miR-149-5p and miR-149-3p in colon inflammation, which may serve as an attractive therapeutic tool for colitis or IBD, and even colitis-associated carcinoma.

circBGN accelerates gastric cancer cell proliferation and invasion via activating IL6/STAT3 signaling pathway.

Circular RNAs participate in the pathogenesis of various tumors, including gastric cancer (GC). In this study, we investigated the role of circBGN in regulating proliferation and invasion of GC cells and elucidated the mechanism. The expression of circBGN was assessed by quantitative reverse-transcription PCR and in situ hybridization. In addition, loss- and gain-of-function investigations in vitro and in vivo were performed to determine the biological functions of circBGN. Luciferase reporter assays and rescue experiments were applied to investigate the interaction between circBGN and miR-149-5p as well as the relationship between miR-149-5p and IL6. Our results showed that circBGN expression was significantly elevated in GC tissues and cells. Knockdown of circBGN dramatically suppressed GC cell proliferation and invasion in vitro. Xenograft experiments revealed that knockdown of circBGN delayed tumor growth in vivo. Furthermore, circBGN can directly bind to miR-149-5p, thereby preventing miR-149-5p from binding to its target mRNA [IL6 mRNA], thus activating IL6/STAT3 signaling pathway. Rescue assays indicated that circBGN regulates GC cell proliferation and invasion by upregulating miR-149-5p/IL6 axis output. Taken together, our investigation indicates that circBGN supports GC progression by activating IL6/STAT3 signaling pathway, thus pointing to a new possible therapeutic target in GC.

Alpha-Synuclein Clearance through Inhibiting Akt/mTOR Pathway by Microfluidic Encapsulated Induced Conjunctival MSCs in a Parkinsonian Model.

Background & Objective: Parkinson's disease (PD) is a progressive neurodegenerative disorder in which the cause is attributed to the alpha-synuclein (α-Syn) accumulation due to the decreased rate of autophagy. Due to the many advantages, mesenchymal stem cells (MSCs), such as the secretion of neurotrophic factors, have been proposed for PD cell therapy. The present study, in continuation of the previous study, aimed to investigate the therapeutic effect of human-derived Conjunctival MSCs (CJ-MSCs) on the clearance of α-Syn by the microRNA-149(miR-149)/Akt/mTOR/ pathway. Methods: Stereotaxic 6-hydroxy dopamine (6-OHDA) was injected directly into the medial forebrain bundle (MFB) to induce Parkinson's disease. An apomorphine-induced rotation test was used to confirm the model establishment. CJ-MSCs were encapsulated in alginate microgel using a microfluidic system. The green fluorescent protein (GFP) labeled CJ-MSCs were encapsulated, and free cells were transplanted into the rats' right striatum. Behavioral and molecular analyses evaluated the potency of CJ-MSCs (encapsulated and free cells) in PD rats. Real-Time Quantitative Reverse Transcription PCR (qRT-PCR) was performed to investigate the expression of the miR-149-5p, Akt, mTOR, and α-Syn. Results: Our obtained results indicated that transplantation of CJ-MSCs leads to a decrease in the number of rotations while raising the balance and motor abilities. The gene expression evaluation showed a significant reduction in Akt, mTOR, and α-Syn mRNA levels and a significant increase in the level of miR-149-5p compared to the control group. Conclusion: It seems that CJ-MSCs can promote the degradation of intracellular α-Syn by miR-149-5p/Akt/mTOR pathway and improve rats' motor functions.