Down-regulation of long noncoding RNA HOXA11-AS nullifies the impact of microRNA-506-3p on chondrocytes proliferation and apoptosis in osteoarthritis.

OBJECTIVES: This study was directed towards exploring the impacts of lncRNA HOXA11-AS-mediated microRNA (miR)-506-3p on chondrocytes proliferation and apoptosis in osteoarthritis (OA). METHODS: The articular cartilages were provided by OA patients who received total knee arthroplasty, and Human Chondrocyte (HC)-OA (HCOA) was also attained. The miR-506-3p and HOXA11-AS expressions in articular cartilages from OA patients and HCOA cells were analyzed via qPCR. After gain- and loss-of-function assays in HCOA cells, MTT assay and flow cytometry (FC) were used for assessing cell viability and apoptosis, accordingly. The levels of PIK3CA, AKT, and mTOR as well as AKT and mTOR phosphorylation levels assessed using western blotting (WB). The targeting correlation of HOXA11-AS and miR-506-3p as well as miR-506-3p and PIK3CA was assessed through Dual-Luciferase Reporter gene Assay (DLRA). RESULT: The articular cartilages from OA patients and Human Chondrocyte (HC)-OA (HCOA) cells showed increased HOXA11-AS and decreased miR-506-3p. Mechanistically, HOXA11-AS was capable of binding to miR-506-3p to increase PIK3CA, the target gene of miR-506-3p. miR-506-3p suppression facilitated HCOA cell proliferation and reduced their apoptosis, which was nullified by further silencing HOXA11-AS or silencing PIK3CA. The down-regulation of HOXA11-AS disrupted the PI3K/AKT/mTOR pathway, which was counteracted by further miR-506-3p inhibition. CONCLUSION: The silencing of HOXA11-AS might block the PI3K/AKT/mTOR pathway through miR-506-3p up-regulation, thereby restricting HCOA cell proliferation and provoking apoptosis.

Hypoxia/inflammation-induced upregulation of HIF-1α and C/EBPß promotes nephroblastoma cell EMT by improving HOXA11-AS transcription.

Background: Homeobox (HOX) A11 antisense RNA (HOXA11-AS) has been identified as a cancer promoting lncRNA and is overexpressed in nephroblastoma. However, how HOXA11-AS is regulated in a hypoxic inflammatory environment has not been studied. Methods: In this study, gene expression and epithelial-mesenchymal transition (EMT) ability were detected in the nephroblastoma cell line WiT49 under conditions of hypoxia and inflammation. Next, HOXA11-AS transcription factors were predicted by datasets and subsequently confirmed by CHIP-QPCR, EMSA, and dual-luciferase reporter assays. Moreover, the regulatory relationships of HOXA11-AS and its transcription factors were further confirmed by rescue experiments. Results: Our results showed that a hypoxic microenvironment promoted HOXA11-AS expression and nephroblastoma progression, induced EMT, and activated the Wnt signaling pathway. Combined hypoxia and inflammation had a more substantial effect on nephroblastoma than either hypoxia or inflammation alone. HIF-1α and C/EBPß were confirmed to be the transcription factors for HOXA11-AS. Silencing of HIF-1α or C/EBPß downregulated HOXA11-AS expression and suppressed EMT and the Wnt signaling pathway in nephroblastoma cells exposed to a hypoxic or inflammatory microenvironment. HOXA11-AS overexpression partly reversed the effect of HIF-1α or C/EBPß knockdown. Conclusion: We demonstrated that hypoxia/inflammation-induced upregulation of HIF-1α and C/EBPß promoted nephroblastoma EMT by improving HOXA11-AS transcription. HOXA11-AS might be a therapy target for nephroblastoma.

Propofol improves ischemia reperfusion-induced liver fibrosis by regulating lncRNA HOXA11-AS.

Liver ischemia reperfusion (IR) injury induces hepatic stellate cell (HSC) activation and liver fibrosis. Propofol (PRO) possesses a positive protective effect on liver ischemia reperfusion injury. We aimed to investigate PRO function and mechanism in IR-induced liver fibrosis. A mice model of liver IR was established. Hematoxylin-eosin (HE) staining was utilized to evaluate liver tissue's pathological changes. Masson staining was applied to evaluate liver fibrosis. The expression level of α-SMA was measured by immunohistochemical (IHC). The expressions of lncRNA HOXA11-AS (HOXA11-AS), PTBP1, HDAC4, α-SMA, COL1A1 and Fibronectin were tested by qRT-PCR or Western blot. The commercial kits detected alanine aminotransferase (ALT) and aspartate aminotransferase (AST) concentrations in serum. Enzyme-linked immunosorbent assay (ELISA) measured TNF-α and IL-6 levels. The binding relationship between HOXA11-AS, PTBP1 and HDAC4 was verified by RNA immunoprecipitation (RIP). Our results showed that PRO alleviated liver fibrosis and the inflammation in IR-induced mice. PRO decreased the expression levels of HOXA11-AS, PTBP1 and HDAC4. Furthermore, HOXA11-AS overexpression abolished the protective effect of PRO against liver fibrosis in mice with IR-disposed. HOXA11-AS interacted with PTBP1 to regulate HDAC4 level and prevented its degradation in JS-1 cells. HDAC4 silencing eliminated the regulatory of HOXA11-AS overexpression on fibrosis and inflammation in IR-induced mice PRO inhibited HOXA11-AS expression to regulate HDAC4, thereby influencing liver fibrosis and inflammation induced by IR. It suggesting that PRO plays a protective role in liver fibrosis induced by ischemia-reperfusion in mice by regulating HOXA11-AS/PTBP1/HDAC4 axis.

lncRNA HOXA11-AS maintains the stemness of oral squamous cell carcinoma stem cells and reduces the radiosensitivity by targeting miR-518a-3p/PDK1.

OBJECTIVE: Oral squamous cell carcinoma (OSCC) is the most prevailing oral malignancy. The lncRNA HOXA11-AS shows prominent roles in OSCC. This study explored the effects of lncRNA HOXA11-AS on regulating OSCC stem cell stemness and radiosensitivity by targeting miR-518a-3p/PDK1. METHODS: Human OSCC cell lines SCC9 and SCC15 were selected. CD133+ cancer stem cells (CSCs) were sorted by immunomagnetic beads. CD133 expression in cells and HOXA11-AS expression in SCC9, SCC15, and CD133+ SCC9, CD133+ SCC15 cells were assessed by flow cytometry and RT-qPCR. HOXA11-AS was silenced/overexpressed in SCC9, SCC15, CD133+ SCC9, and CD133+ SCC15 cells. Cell proliferation, radiosensitivity, invasion, and stem cell sphere formation ability were examined by CCK-8, colony formation, Transwell, and stem cell sphere formation. The levels of stemness-related genes (Oct4, Nanog, Sox2), miR-518a-3p, epithelial-mesenchymal transition (EMT)-related proteins (E-cadherin, Vimentin, N-cadherin), and PDK1 were assessed by RT-qPCR and Western blot assay. RESULTS: HOXA11-AS was up-regulated in SCC9, SCC15, CD133+ SCC9, and CD133+ SCC15 cells. HOXA11-AS silencing inhibited OSCC proliferation and invasion and enhanced radiosensitivity. HOXA11-AS maintained CSC stemness in OSCC. HOXA11-AS silencing reduced CD133+ SCC9 and CD133+ SCC15 stem cell sphere formation ability, reduced stem cell stemness-related gene levels, and inhibited EMT. HOXA11-AS regulated OSCC stem cell stemness and radiosensitivity by targeting miR-518a-3p. PDK1 overexpression annulled the regulatory effects of HOXA11-AS silencing on OSCC cell stem cell stemness and radiosensitivity. CONCLUSION: In vitro lncRNA HOXA11-AS silencing inhibited OSCC stem cell stemness by targeting the miR-518a-3p/PDK1 axis, thus enhancing OSCC cell radiosensitivity.

nc-RNA-mediated high expression of CDK6 correlates with poor prognosis and immune infiltration in pancreatic cancer.

BACKGROUND: Emerging evidence manifests that cyclin-dependent kinase 6 (CDK6) plays an essential part in the initiation and progression of several types of human cancer, and its descending expression is correlated with an adverse prognosis. However, the precise role of CDK6 in Pancreatic cancer (PC) remains obscure. AIMS: To identify the potential ceRNA regulatory axis of CDK6 in PC and explore its relationship with immune cells and immune checkpoints. MATERIALS & METHODS: Using The Cancer Genome Atlas TCGA and GTEx data analyze the expression and survival of CDK6 in patients in pan-cancer, and cellular experiments were performed to verify the effect of CDK6 on cell function. Using GEPIA and STARBASE databases to analyze prognosis, expression and survival, and identify non coding RNA (ncRNA) that mediates CDK6 overexpression. The TIMER 2.0 database was used for immune correlation analysis. RESULTS: We revealed CDK6 might be an oncogene in PC, and the HOXA11-AS /NR2F1-AS1- miR-454-3p axis was identified as the possible upstream ncRNA-associated pathway of CDK6 in PC. In addition, CDK6 show significant association with three immune checkpoints (PD-L1, PD-L2, and HAVCR2), the infiltration level of immune cells, and immunity biomarkers. DISCUSSION: We discussed some applications of CDK6 in breast cancer, melanoma, and hemorrhagic malignancies. The role of miR-15a-5p, HOXA11-AS and NR2F1-AS1 in tumor development was also discussed based on existing studies. The potential mechanism of CDK6 affecting immune cells in pancreatic cancer was discussed. CONCLUSIONS: Overall, these results established that nc-RNA-mediated high expression of CDK6 is associated with patient outcomes and immune invasion in pancreatic cancer.

The long noncoding RNA HOXA11-AS promotes lung adenocarcinoma proliferation and glycolysis via the microRNA-148b-3p/PKM2 axis.

BACKGROUND: Lung cancer is the most common malignancy in the world and a growing number of researches have focused on its metabolic characteristics. Studies have shown that the long non-coding RNA (lncRNA) HOXA11-AS is aberrantly expressed in many tumors. However, the role of HOXA11-AS in lung adenocarcinoma (LUAD) glycolysis and other energy metabolism pathways has not been characterized. METHOD: The mRNA levels of HOXA11-AS, microRNA-148b-3p (miR-148b-3p), and pyruvate kinase M2 (PKM2) were detected using qRT-PCR. The expression levels of proteins were measured using immunohistochemistry and western blot. The CCK-8, EdU, and colony formation assays were used to assess proliferation. Glycolytic changes were assessed by measuring lactate production, ATP production, and 18 F-FDG uptake. Bioinformatics analysis and dual-luciferase reporter assays were used to characterize the relationship between HOXA11-AS, miR-148b-3p, and PKM2. Proliferation and glycolytic changes were analyzed in xenograft tumor experiments using Micro-PET imaging after downregulation of HOXA11-AS in vivo. RESULTS: The expression of HOXA11-AS was markedly increased in LUAD, and was strongly associated with a poor prognosis. In addition, HOXA11-AS promoted proliferation and glycolysis in LUAD, and miR-148b-3p inhibited proliferation and glycolysis in LUAD. Mechanistically, HOXA11-AS positively regulated PKM2 expression by binding to miR-148b-3p, thereby promoting LUAD proliferation and glycolysis. In addition, HOXA11-AS inhibited LUAD xenograft growth and glycolysis via upregulation of miR-148b-3p expression and downregulation of PKM2 expression in vivo. CONCLUSIONS: These results showed that HOXA11-AS enhanced LUAD proliferation and glycolysis via the miR-148b-3p/PKM2 axis. The findings in this paper expanded our understanding of the molecular mechanisms of LUAD tumorigenesis and glycolysis and showed that HOXA11-AS could be useful as a diagnostic and prognostic marker for LUAD. 18 F-FDG PET/CT can be used to visually evaluate the therapeutic effect of targeting HOXA11-AS.

Long non-coding RNA HOXA11-AS contributes to the formation of keloid by relieving the inhibition of miR-182-5p on ZNF217.

BACKGROUND: Long non-coding RNA (lncRNA) dysregulation is demonstrated to be associated with disease progression. Mounting studies show that lncRNA promotes or inhibits the development of keloid. We aimed to disclose the role of homebox A11 antisense RNA (HOXA11-AS) in the formation of keloid. METHODS: Quantitative real-time PCR (qPCR) was adopted for expression analysis of HOXA11-AS, miR-182-5p and zinc finger protein 217 (ZNF217) mRNA, and the expression of ZNF protein and marker proteins was detected by western blot. Cell proliferation, cell migration and cell apoptosis were investigated using CCK-8 assay, wound healing assay and flow cytometry assay, respectively. The potential interplay between miR-182-5p and HOXA11-AS or ZNF217 was verified by dual-luciferase reporter assay, RIP assay and pull-down assay. The role of HOXA11 in vivo was studied by establishing animal models. RESULTS: HOXA11-AS was highly expressed in tissues and fibroblasts of keloid. Deficiency of HOXA11-AS blocked the proliferation and migration of keloid fibroblasts and induced fibroblast apoptosis. HOXA11-AS directly combined to miR-182-5p whose downregulation reversed the effects of HOXA11-AS knockdown. ZNF217 was a target of miR-182-5p, and HOXA11-AS indirectly promoted ZNF217 expression by binding to miR-182-5p. MiR-182-5p enrichment also blocked keloid fibroblast proliferation, survival and migration, while further ZNF217 overexpression abolished these effects. HOXA11-AS knockdown also hindered the growth of keloid in mouse models. CONCLUSION: High expression of HOXA11-AS promoted the formation and growth of keloid through the upregulation of ZNF217 by targeting miR-182-5p, and the inhibition of HOXA11-AS might be a novel strategy to prevent keloid development.

Emerging roles of long non-coding RNAs in keloids.

Keloids are pathologic wound healing conditions caused by fibroblast hyperproliferation and excess collagen deposition following skin injury or irritation, which significantly impact patients by causing psychosocial and functional distress. Extracellular matrix (ECM) deposition and human fibroblast proliferation represents the main pathophysiology of keloid. Long non-coding RNAs (LncRNAs) play important roles in many biological and pathological processes, including development, differentiation and carcinogenesis. Recently, accumulating evidences have demonstrated that deregulated lncRNAs contribute to keloids formation. The present review summarizes the researches of deregulated lncRNAs in keloid. Exploring lncRNA-based methods hold promise as new effective therapies against keloid.

An Axis between the Long Non-Coding RNA HOXA11-AS and NQOs Enhances Metastatic Ability in Oral Squamous Cell Carcinoma.

Long non-coding RNAs (lncRNAs) play critical roles in human cancers. HOXA11 anti-sense RNA (HOXA11-AS) is an lncRNA belonging to the homeobox (HOX) gene cluster that promotes liver metastasis in human colon cancer. However, its role and mechanism of action in human oral squamous cell carcinoma (OSCC) are unclear. In this study, we investigated HOXA11-AS expression and function in human OSCC tissues and cell lines, as well as a mouse model of OSCC. Our analyses showed that HOXA11-AS expression in human OSCC cases correlates with lymph node metastasis, nicotinamide adenine dinucleotide (NAD)(P)H: quinone oxidoreductase 1 (NQO1) upregulation, and dihydronicotinamide riboside (NRH): quinone oxidoreductase 2 (NQO2) downregulation. Using the human OSCC cell lines HSC3 and HSC4, we demonstrate that HOXA11-AS promotes NQO1 expression by sponging microRNA-494. In contrast, HOXA11-AS recruits zeste homolog 2 (EZH2) to the NQO2 promoter to suppress its expression via the trimethylation of H3K27. The upregulation of NQO1 enzymatic activity by HOXA11-AS results in the consumption of flavin adenine dinucleotide (FAD), which reduces FAD-requiring glyceraldehyde-3-phosphate dehydrogenase (GAPDH) activity and suppresses glycolysis. However, our analyses show that lactic acid fermentation levels are preserved by glutaminolysis due to increased malic enzyme-1 expression, promoting enhanced proliferation, invasion, survival, and drug resistance. In contrast, suppression of NQO2 expression reduces the consumption of NRH via NQO2 enzymatic activity and increases NAD levels, which promotes enhanced stemness and metastatic potential. In mouse tumor models, knockdown of HOXA11-AS markedly suppressed tumor growth and lung metastasis. From these findings, targeting HOXA11-AS may strongly suppress high-grade OSCC by regulating both NQO1 and NQO2.

HOXA11-AS1 Promotes PD-L1-Mediated Immune Escape and Metastasis of Hypopharyngeal Carcinoma by Facilitating PTBP1 and FOSL1 Association.

BACKGROUND: The metastatic characteristics of hypopharyngeal squamous cell carcinoma (HSCC) lead to many diagnostic and therapeutic challenges, while functional long non-coding RNAs (lncRNAs) can provide effective strategies for its diagnosis and treatment. METHODS: RT-qPCR, Western blot, immunohistochemistry, and an immunofluorescence assay were used to detect the related gene expression. Flow cytometry was used to measure the percentage of CD8+ and CD4+ T cells. CCK-8 and transwell assays were performed to analyze the role of HOXA11-AS1. The targeted relationship of the FOSL1/PD-L1 promoter was measured by ChIP and dual-luciferase reporter assays. RNA pulldown and RIP assays were used to measure the interaction between HOXA11-AS1, FOSL1, and PTBP1. A tumor xenograft study was used to analyze HOXA11-AS1 function in vivo. RESULTS: HOXA11-AS1, PD-L1, and FOSL1 were upregulated in HSCC, and HOXA11-AS1 positively correlated with PD-L1. HOXA11-AS1 knockdown upregulated CD8+ T cells through an increase in IFN-γ concentration while decreasing the proliferation, migration, and invasion of HSCC cells. FOSL1 bound the PD-L1 promoter, increasing gene expression. HOXA11-AS1 enhanced the stability of FOSL1 mRNA by binding to PTBP1. HOXA11-AS1 or PTBP1 overexpression increased FOSL1 and PD-L1 expression. PD-L1 knockdown arrested the inhibiting function of HOXA11-AS1 overexpression on CD8+ T cell content. HOXA11-AS1 knockdown inhibited immune escape and metastasis through PD-L1 regulation by downregulating FOSL1 in vivo. CONCLUSION: HOXA11-AS1 promoted PD-L1 expression by upregulating FOSL1 levels through PTBP1, thereby facilitating immune escape, proliferation, and metastasis of HSCC cells.

Solamargine Inhibits the Development of Hypopharyngeal Squamous Cell Carcinoma by Decreasing LncRNA HOXA11-As Expression.

Hypopharyngeal squamous cell carcinoma (HSCC) is one of the high mortality cancers with a poor prognosis, which is driving the development of new chemotherapeutic agents. We identified the anticancer effects of a natural compound, solamargine (SM), on FaDU cells and explored its mechanism in terms of non-coding RNA. It was observed that SM inhibited the proliferation of FaDU cells with an IC50 of 5.17 µM. High-throughput sequencing data revealed that lncRNA HOXA11-AS was significantly downregulated in cells co-incubated with SM. Further assays demonstrated that SM-induced downregulation of lncRNA HOXA11-AS showed important implications for apoptosis. Given the properties of HOXA11-AS as a miR-155 sponge, we further confirmed that SM upregulated the expression of miR-155 in FaDU cells. C-Myc is a transcription factor that regulates cell differentiation and apoptosis, whose mRNA is considered to be targeted by miR-155. We showed that c-Myc expression was downregulated by SM and accompanied by increased apoptosis, which was consistent with the findings of transcriptome sequencing. Furthermore, SM administration suppressed xenograft tumor growth in a xenograft mouse model in vivo. In the light of the aforementioned findings, our results suggested that SM downregulated the expression of HOXA11-AS, which in turn induces apoptosis by downregulating c-Myc in FaDU, providing evidence for the anticancer effect of SM on HSCC and uncovering the effect of SM on non-coding RNAs as, at least partly, a mechanism of action.

Long non-coding RNA HOXA11-AS knockout inhibits proliferation and overcomes drug resistance in ovarian cancer.

In ovarian carcinogenesis and progression, long non-coding RNAs (lncRNAs) have been shown to have a role, although the underlying processes remain a mystery. By modulating the degree of autophagy in ovarian cancer cells, we sought to learn more about the function lncRNA HOXA11-AS plays in the development of ovarian cancer. The expression of HOXA11-AS in ovarian normal cells and ovarian cancer cell lines was measured using R package and qRT-PCR. Ovarian cancer cells expressed HOXA11-AS substantially higher than normal cells, while cisplatin-resistant cells expressed HOXA11-AS significantly higher than ovarian cancer cells. Next, we studied the prognostic data of HOXA11-AS in ovarian cancer in the Tissue Cancer Genome Atlas (TCGA). In the next step, lentiviral transfection of ovarian cancer cells A2780, OVCAR3, and A2780/DDP (cisplatin-resistant) were performed, and HOXA11-AS knockdown was found to significantly inhibit cell viability, migration, and invasion of A2780 and OVCAR3 cells, and promote apoptosis by CCK-8 assay, transwell assay, cell cycle, and apoptosis assay, and promoted the sensitivity of A2780/DDP cells to cisplatin. It has been shown by the western blot test that HOXA11-AS knockdown increases the amount of cellular autophagy in cells. In contrast, adding the autophagy inhibitor 3-methyladenine (3-MA) to HOXA11-AS cells knocked down in vivo reduced its anti-tumor properties. As a whole, this study found that HOXA11-AS knockdown increased the expression of autophagy-related proteins and improved cisplatin sensitivity, decreased ovarian cancer cell proliferation, and promoted cell apoptosis. This study provides new insights into the role of HOXA11-AS in ovarian cancer regulation.

The minor allele of rs17427875 in long non-coding RNA-HOXA11-AS influences the prognosis of subarachnoid hemorrhage (SAH) via modulating miR-15a and STAT3 expression.

BACKGROUND: HOAX11-AS was reported to promote the progression of liver cancer via the signaling pathway of miR-15a-3p/STAT3. In this study, we investigated the effect of rs17427875 on the prognosis of subarachnoid hemorrhage (SAH) and its underlying molecular mechanisms. METHODS: 158 SAH patients were recruited and grouped according to their genotypes rs17427875. Peripheral blood and cerebrospinal fluid (CSF) samples were collected for subsequent analysis. Quantitative real-time PCR, luciferase assays, Western blot and ELISA were performed to analyze the correlations between the expression of lncRNA-HOXA11-AS, miR-15a, TNF-α and NF-κB. RESULTS: The survival rate was remarkably higher in SAH patients carrying the AA genotype of rs17427875 when compared with those carrying the AT genotype. The expression of miR-15a was significantly repressed in the peripheral blood and CSF of SAH patients carrying the AT allele when compared with that in patients carrying the AA allele. MiR-15a showed a remarkable efficacy in inhibiting the luciferase activity of wild type lncRNA-HOXA11-AS and STAT3 in THP-1 cells. P-HOXA11-AS-T showed a stronger ability to suppress the expression of miR-15a and activate the expression of STAT3, TNF-α and NF-κB in THP-1 cells when compared with P-HOXA11-AS-A. CONCLUSIONS: The findings demonstrated that the presence of the minor allele of rs17427875 in lncRNA-HOXA11-AS could increase the expression level of lncRNA-HOXA11-AS, thus elevating the expression level of STAT3 via down-regulating miR-15a, and increased STAT3 expression could aggravate inflammation to cause poor prognosis of SAH. Therefore, the rs17427875 polymorphism can be used as a potential biomarker for the prognosis of SAH.

LncRNA HOXA11-AS promotes vascular endothelial cell injury in atherosclerosis by regulating the miR-515-5p/ROCK1 axis.

AIMS: Long non-coding RNA HOXA11-AS participated in heart disease. In this study, we aim to evaluate the potential roles of HOXA11-AS in atherosclerosis and its underlying mechanisms. METHODS AND RESULTS: The expression levels of HOXA11-AS in ox-LDL-treated HUVECs and arch tissues of high-fat diet-fed ApoE-/- mice (n = 10) were assessed by qRT-PCR. The effects of HOXA11-AS knockdown on the development of atherosclerosis were evaluated using in vitro and in vivo models. Luciferase reporter and RNA immunoprecipitation (RIP) assays verified the potential relationships between HOXA11-AS or ROCK1 and miR-515-5p. The interactive roles between HOXA11-AS and miR-515-5p and between miR-515-5p and ROCK1 were further characterized in ox-LDL-treated HUVECs. Our data showed that HOXA11-AS was significantly up-regulated (P < 0.001), whereas miR-515-5p was dramatically down-regulated in AS mice tissues (P < 0.001) and ox-LDL-treated HUVECs (P < 0.01). Ox-LDL could induce endothelial injuries by inhibiting cell proliferation (P < 0.001) and SOD synthesis (P < 0.001), promoting apoptosis (P < 0.01), ROS (P < 0.001), and MDA production (P < 0.001), increasing Bax (P < 0.001) and cleaved Caspase-3 (P < 0.001), and decreasing Bcl-2 (P < 0.001) and phosphorylated eNOS (P < 0.01). HOXA11-AS knockdown attenuated endothelial injuries via increasing eNOS phosphorylation. Luciferase assay and RIP results confirmed that miR-515-5p is directly bound to HOXA11-AS and ROCK1. HOXA11-AS promoted ox-LDL-induced HUVECs injury by directly inhibiting miR-515-5p from increasing ROCK1 expression and subsequently decreasing the expression and phosphorylation of eNOS. MiR-515-5p mimics could partially reverse the effects of HOXA11-AS knockdown. CONCLUSIONS: HOXA11-AS contributed to atherosclerotic injuries by directly regulating the miR-515-5p/ROCK1 axis. This study provided new evidence that HOXA11-AS might be a candidate for atherosclerosis therapy.

The Haoqin-Huaban formula alleviates UVB-induced skin damage through HOXA11-AS-mediated stabilization of EZH2.

Exposure of skin to ultraviolet B (UVB) irradiation induces oxidative damage, immune suppression, inflammation, and skin cancer. Recently, an increase in the use of traditional Chinese medicine decoction with antioxidant properties has emerged as protection for skin tissues against UVB-induced damage. The aim of this study was to investigate mechanisms of the protective effect of the Haoqin-Huaban formula (HQHB) on UVB-induced skin damage. First, cell survival, apoptosis, and oxidative stress were evaluated upon UVB irradiation in the presence of HQHB using HaCaT cells and mice as model systems. Subsequently, bioinformatic analyses, RNA pulldown assays, RNA immunoprecipitation, luciferase reporter assays, and chromatin immunoprecipitation were conducted to verify the regulation among HQHB, hypoxia-inducible factor 1α (HIF-1α), HOXA11-AS and enhancer of zeste homolog 2 (EZH2) in HaCaT cells. In this study, we found that administration of HQHB inhibited, in a dose-dependent manner, UVB-induced skin damage by eliminating oxidative stress. HQHB was found to upregulate HOXA11-AS expression by activating HIF-1α. Furthermore, HOXA11-AS stabilized the EZH2 protein by inhibiting its ubiquitination and proteasomal degradation. Consequently, rescue assays demonstrated that HOXA11-AS promoted proliferation and inhibited apoptosis in HaCaT cells by reducing oxidative stress. Taken together, our results help to elucidate the function and regulatory mechanism of HQHB in reducing UVB-induced skin damage.

HOXA11-AS facilitates the proliferation, cell cycle process and migration of keloid fibroblasts through sponging miR-188-5p to regulate VEGFA.

BACKGROUND: Abnormal expression of long non-coding RNA (lncRNA) has been proved to be related to the formation of keloid. Homeobox A11 antisense (HOXA11-AS) has been found to be a significantly upregulated lncRNA in keloid tissues. OBJECTIVE: To explore the mechanism of HOXA11-AS regulates keloid formation. METHODS: Primary fibroblasts were isolated from keloid tissues and normal skin tissues. The expression of HOXA11-AS, microRNA (miR)-188-5p and vascular endothelial growth factor A (VEGFA) was determined using quantitative real-time PCR (qRT-PCR). Cell counting kit 8 (CCK8) assay, EdU staining, flow cytometry and wound healing assay were performed to assess the proliferation, cell cycle process, apoptosis and migration of keloid fibroblasts. Importantly, some marker protein levels and VEGFA protein level were examined by western blot (WB) analysis. The interaction between miR-188-5p and HOXA11-AS or VEGFA was confirmed using dual-luciferase reporter assay, RNA pull-down assay and RIP assay. Animal experiments were performed to further confirm the role of HOXA11-AS in keloid growth. RESULTS: HOXA11-AS was markedly upregulated in keloid tissues and fibroblasts. Knockdown of HOXA11-AS repressed the proliferation, cell cycle process, migration and promoted the apoptosis of keloid fibroblasts. Further analysis suggested that HOXA11-AS could sponge miR-188-5p to positively regulate VEGFA. The inhibition of HOXA11-AS silencing on the biological functions of keloid fibroblasts could be reversed by miR-188-5p inhibitor. In addition, VEGFA overexpression also abolished the suppressive effect of miR-188-5p on the biological functions of keloid fibroblasts. Interferences of HOXA11-AS suppressed keloid growth in vivo. CONCLUSION: HOXA11-AS might regulate the miR-188-5p/VEGFA axis to promote keloid formation.

Construction of a HOXA11-AS-Interact Ed Network in Keloid Fibroblasts Using Integrated Bioinformatic Analysis and in Vitro Validation.

Background: Expression of the long noncoding RNA (lncRNA) HOXA11-AS significantly increased in keloids by unclarified molecular regulation mechanisms. Methods: Using successfully primary cultured keloid-derived fibroblasts from central region of chronic keloid tissues (sample 0), small interfering RNAs were designed and transfected into two keloid fibroblast samples (samples 1 and 2) to knockdown HOXA11-AS. One nonspecific transfection control (sample 3) and one blank control (sample 4) were used to remove nonspecific overlap from the studied group. The lncRNAs, messenger RNAs (mRNAs), and microRNAs (miRNAs) of five samples were sequenced to identify differentially expressed (DE) profiles in HOXA11-AS-knockdown keloid fibroblasts in samples 1 and 2 (by intersection), which facilitated removal of overlap with the nonspecific controls (samples 3 and 4, by union). Using stepwise bioinformatic analysis, a HOXA11-AS-interacted competing endogenous network (ceRNA) was screened based on three DE profiles. Results: Keloid fibroblasts with or without HOXA11-AS as well as with or without nonspecific interferences were successfully constructed respectively. A total of 1,396 mRNAs and 39 lncRNAs were significantly changed in keloid fibroblast with HOXA11-AS knockdown. Simultaneously, 1,626 mRNAs and 99 lncRNAs were significantly changed in keloid fibroblast with nonspecific interference. With removal of nonspecific overlap, a lncRNA-mRNA interactive network characterized by close natural/intronic antisense relationship was initially constructed in keloid fibroblast with HOXA11-AS knockdown. Based on this network, a lncRNA-mRNA-protein interaction network was extended by integration of the human protein-protein interaction network. Significant functional genes were screened using PageRank algorithm in the extended network. Three genes, including SNED1, NIPAL3, and VTN, were validated by real-time PCR in HOXA11-AS-knockdown keloid fibroblasts. Only NIPAL3 was predicted to be a target gene for HOXA11-AS via three competing endogenous miRNAs (hsa-miRNA-19a-3p, hsa-miR-141-3p, and hsa-miR-140-5p). Conclusion: An interactive network of HOXA11-AS-three miRNAs-NIPAL3 was predicted in keloid fibroblasts by integrative bioinformatic analysis and in vitro validation.

Long Non-coding RNA HOXA11-AS Facilitates Proliferation of Lung Adenocarcinoma Cells via Targeting the Let-7c-5p/IGF2BP1 Axis.

Objective: This study investigates the relationship between the HOXA11-AS/let-7c-5p/IGF2BP1 regulatory axis and lung adenocarcinoma. Methods: The expression levels of HOXA11-AS, let-7c-5p, and IGF2BP1 were evaluated in LUAD tissue and cell lines. Subcellular fractionation detection assay was adopted to verify the HOXA11-AS distribution in LUAD cells. The interaction relationship between let-7c-5p and HOXA11-AS or IGF2BP1 was validated by dual-luciferase reporter detection. In RNA binding protein immunoprecipitation assay, the binding relationship between HOXA11-AS and let-7c-5p was identified. The cell viability of transfected cells was tested by the Cell Counting Kit-8 assay. The mouse xenograft model was used to identify the effect of HOXA11-AS on tumor growth in vivo. Results: Upregulation of lncRNA HOXA11-AS was found in LUAD, and suppression of HOXA11-AS could suppress the proliferative ability of LUAD cells. The let-7c-5p was expressed to be downregulated, which played an inhibitory role in LUAD cell proliferation. Let-7c-5p was negatively regulated by HOXA11-AS. HOXA11-AS promoted LUAD cell proliferation, while let-7c-5p had an inverse effect. Besides, IGF2BP1, regulated by let-7c-5p, had a positive relation with HOXA11-AS, while overexpression of IGF2BP1 could suppress the inhibition of silencing HOXA11-AS on LUAD cell proliferation. Experiments on mice confirmed that HOXA11-AS facilitated LUAD cell growth in vivo through regulating the let-7c-5p/IGF2BP1 axis. Conclusion: HOXA11-AS promoted LUAD cell proliferation by targeting let-7c-5p/IGF2BP1, which could be potential molecular targets for LUAD.

The lncRNA HOXA11-AS acts as a tumor promoter in breast cancer through regulation of the miR-125a-5p/TMPRSS4 axis.

BACKGROUND: Long non-coding RNAs (lncRNAs) play vital roles in tumorigenesis. Here, we explored how lncRNA HOXA11-AS functions in the progression of breast cancer (BC). METHODS: HOXA11-AS and miR-125a-5p levels were measured by a quantitative real-time polymerase chain reaction, whereas western blotting determined TMPRSS4 levels in BC tumor tissues, adjacent normal tissues and BC cell lines. The roles of HOXA11-AS, miR-125a-5p and TMPRSS4 in BC proliferation were investigated using cell counting kit-8, colony formation and flow cytometry assays, whereas scratch and transwell assays were used to measure metastasis. RNA pull-down assays and dual-luciferase assays assessed direct interactions between HOXA11-AS and miR-125a-5p. The effects of HOXA11-AS in vivo were investigated in a BC xenograft model. RESULTS: HOXA11-AS was upregulated in tumor tissues of 56 BC patients compared to adjacent non-tumor tissues, with high levels being associated with worse overall survival. Silencing of HOXA11-AS inhibited the proliferation and metastasis of BC cells, leading to cell cycle arrest in G0/G1 and induction of apoptosis. We identified miR-125a-5p as a target of HOXA11-AS, with miR-125a-5p inhibitors partially restoring the reduction of cell proliferation and metastasis induced by HOXA11-AS silencing. We also determined that miR-125a-5p targeted TMPRSS4 mRNA, with HOXA11-AS knockdown and miR-125a-5p mimics suppressing TMPRSS4. Overexpression of TMPRSS4 partially compensated for the reduction of cell proliferation and metastasis induced by HOXA11-AS silencing. Finally, we confirmed the mechanism of HOXA11-AS in the regulation of tumorigenesis in the mouse model. CONCLUSIONS: HOXA11-AS regulates the tumorigenic ability of BC via the miR-125a-5p/TMPRSS4 axis. This provides insights for regulatory mechanisms involved in BC progression, and may enable new treatment strategies in the clinical setting.