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.
RESULTS: 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.
CONCLUSIONS: 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.

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.
CONCLUSIONS: 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.

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/β-catenin pathway, and PTOV1-AS1 recruited SUV39H1 to suppress miR-149-5p transcription, in turn activating Wnt/β-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/β-catenin pathway, which provides insights into a potential novel target for enhancing the effectiveness of chemotherapy in colorectal cancer patients.

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.

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-1β, 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.

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.
CONCLUSIONS: 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.

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.

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.
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.

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.

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.
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.
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.
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.