scholarly journals The Long Non-Coding RNA LncRNA8975-1 is Upregulated in Hypertrophic Scar Fibroblasts and Controls Collagen Expression

2016 ◽  
Vol 40 (1-2) ◽  
pp. 326-334 ◽  
Author(s):  
Jun Li ◽  
Ling Chen ◽  
Chunyu Cao ◽  
Hui Yan ◽  
Bei Zhou ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Hypertrophic Scar ◽  
Smooth Muscle Actin ◽  
Scar Formation ◽  
Dermal Fibroblasts ◽  
Alpha Smooth Muscle Actin ◽  
Protein Levels ◽  
Reverse Transcription Pcr ◽  
Non Coding Rna ◽  
Long Non Coding Rna

Background/Aims: Long non-coding RNAs (lncRNAs) are thought to play crucial roles in human diseases. However, the function of lncRNAs in hypertrophic scar formation remains poorly understood. Methods: In this study, we investigated the expression of lncRNA8975-1 in hypertrophic scar tissues and fibroblasts by quantitative reverse transcription PCR (qRT-PCR). To investigate its function, overexpression and knockdown of lncRNA8975-1 were performed using lentivirus infection and Stealth RNAi transfection, respectively. Cell proliferation was detected by CCK-8 assay. The protein levels of collagens and alpha-smooth muscle actin (α-SMA) were analysed by western blot. Results: We found that lncRNA8975-1 was overexpressed in hypertrophic scar tissues and dermal fibroblasts. Overexpression of lncRNA8975-1 inhibited cell proliferation and reduced the protein expression levels of COL1A2, COL1A1, COL3A1 and α-SMA in hypertrophic scar fibroblasts, whereas knockdown of lncRNA8975-1 had the opposite effect. Conclusion: Our results show that the long non-coding RNA lncRNA8975-1 is upregulated in hypertrophic scar fibroblasts; furthermore, it inhibits fibroblast proliferation and reduces collagen and α-SMA expression. Further studies on the mechanisms regulated by lncRNA8975-1 would lead to a better understanding of the pathogenesis of hypertrophic scar formation.

scholarly journals O30 Long non-coding RNA HOTAIR induces beta catenin expression in systemic sclerosis through EZH2 dependent repression of Axin-2

Rheumatology ◽  
2020 ◽  
Vol 59 (Supplement_2) ◽  
Author(s):  
Christopher W Wasson ◽  
Giuseppina Abignano ◽  
Rebecca Ross ◽  
Francesco Del Galdo
Keyword(s):  
Systemic Sclerosis ◽  
Smooth Muscle Actin ◽  
Specific Inhibitor ◽  
Dermal Fibroblasts ◽  
Wnt Signalling ◽  
Beta Catenin ◽  
Cultured Fibroblasts ◽  
Non Coding Rna ◽  
Long Non Coding Rna

Abstract Background Fibroblasts explanted from affected tissues in systemic sclerosis (SSc) maintain their pro-fibrotic phenotype in vitro. This includes increased secretion of collagens and other extracellular matrix proteins and increased proportion of α-Smooth Muscle Actin (α-SMA) positive cells (myofibroblasts). It has been previously shown that among their profibrotic features, myofibroblasts display activation of WNT signalling, which is linked to a decreased basal expression of AXIN2. Here we aimed to determine whether specific long non-coding RNA (lncRNAs) expressed in myofibroblasts could drive the epigenetically stable decreased expression of Axin 2 in vitro. Methods Full thickness skin biopsies were surgically obtained from the forearms of twelve adult patients with SSc of recent onset. Fibroblasts were isolated and cultured in monolayers and protein and RNA extracted from the fibroblast cultures. Laser capture was performed to isolate cells expressing or not α-SMA as a marker of myofibroblast differentiation. LncRNA HOTAIR was overexpressed in healthy dermal fibroblasts by lentiviral induction. EZH2 was blocked in cultured fibroblasts with the specific inhibitor GSK126. Results HOTAIR expression was increased in SSc patients’ skin (100 fold) and in SSc explanted fibroblasts (5 fold; p < 0.001 for both). Further, laser captured α-SMA expressing fibroblasts expressed in average 2.5 fold higher HOTAIR RNA levels compared to α -SMA negative cells from the same donors (P < 0.05). In vitro, lentiviral induced stable overexpression of HOTAIR in healthy dermal fibroblasts led to their profibrotic activation, including significantly increased expression of Col1A1 and α-SMA both at mRNA and protein levels (2.8 and 1.8 fold respectively, p < 0.05). We further showed that HOTAIR-induced profibrotic activation was due to EZH2 dependent spread of H3k27me3 methylation marker, as demonstrated by complete inhibition by treatment with GSK126. HOTAIR driven EZH2 histone methylation suppressed the expression of Axin 2 in SSc fibroblasts. The reduced Axin 2 levels led to stabilisation of beta catenin and WNT signalling pathway activation. Conclusion Here we show that the epigenetically stable activation of SSc dermal fibroblasts is due to HOTAIR. We also identified a major downstream target of HOTAIR is Axin-2. The results of these studies identify a new venue to modulate fibroblasts biology which could inform clinical research to resolve chronic fibrosis and re-establish tissue homeostasis in SSc. Disclosures C.W. Wasson None. G. Abignano None. R. Ross None. F. Del Galdo Consultancies; AstraZeneca, GSK, Boehringer-Ingelheim, Actelion, Capella Biosciences, Chemomab.

scholarly journals Long non-coding RNA PVT1 regulates the migration of hepatocellular carcinoma HepG2 cells via miR-3619-5p/MKL1 axis

2020 ◽  
Author(s):  
Hua Liu ◽  
Yan Yin ◽  
Ting Liu ◽  
Yanying Gao ◽  
Qing Ye ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Migration ◽  
Serum Response Factor ◽  
Migration And Invasion ◽  
Protein Levels ◽  
Reverse Transcription Pcr ◽  
Non Coding Rna ◽  
And Migration ◽  
Long Non Coding Rna ◽  
Hcc Cells

Hepatocellular carcinoma (HCC) is the third most common malignant tumor of the digestive system. Plasma cell tumor heterotopic gene 1 (PVT1) is an intergenic long non-coding RNA that is aberrantly expressed in different cancers. Myocardin related transcription factor A (MKL1) is a transcriptional coactivator of serum response factor that has been shown to promote cancer cell migration and invasion. In this study, we investigated the relationship between PVT1 and MKL1 as a novel regulatory mechanism underlying HCC progression. We used HepG2 and Cos‑7 cell lines. Transfection experiments with miR-3619-5p mimics/inhibitor, PVT1, siRNA-PVT1, MKL1, or siRNA-MKL1 were performed. RNA and protein levels were analyzed by quantitative reverse transcription PCR and Western blot, respectively. Cell migration was assessed by Transwell assay. Luciferase assays, RNA-FISH, RNA immunoprecipitation, and chromatin immunoprecipitation assays were performed to confirm the interaction between PVT1, miR-3619-5p, and MKL1 in HCC cells. Overexpression of PVT1 was positively correlated with MKL1 upregulation, which promoted HepG2 cell migration. miR-3619-5p inhibited MKL1 expression in HCC cells by acting on its 3' UTR. Furthermore, PVT1 promoted MKL1 expression and migration in HCC cells by directly binding to miR-3619-5p. In a positive feedback loop, MKL1 could activate PVT1 transcription by binding to the CArG box in the promoter region. Our findings may provide a basis for the development of novel targeted therapies in HCC.

scholarly journals Long non-coding RNA PART1 predicts a poor prognosis and promotes the malignant progression of pancreatic cancer by sponging miR-122

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Xibao Hu ◽  
Lei Zhang ◽  
Jingjing Tian ◽  
Junhong Ma
Keyword(s):  
Pancreatic Cancer ◽  
Cell Proliferation ◽  
Overall Survival ◽  
Clinical Stage ◽  
Malignant Progression ◽  
Luciferase Reporter ◽  
Poor Overall Survival ◽  
Pancreatic Cancer Cells ◽  
Non Coding Rna ◽  
Long Non Coding Rna

Abstract Background and objectives Long non-coding RNA (lncRNA) prostate androgen-regulated transcript 1 (PART1) was previously shown to exert an oncogenic role in several human cancers. However, whether PART1 is associated with the malignant progression of pancreatic cancer remains unclear. In the current study, we aimed to identify the role and potential mechanism of PART1 in pancreatic cancer. Methods qRT-PCR was applied to detect PART1 expression in 45 cases of pancreatic cancer patients. The chi-square test was performed to assess the association between PART1 expression and clinicopathologic features, and Kaplan-Meier method was applied to evaluate overall survival. In vitro CCK-8, transwell invasion, and flow cytometry assays were applied to detect the effects of PART1 on cell proliferation, invasion, and apoptosis, respectively. Luciferase reporter and RNA immunoprecipitation assays were used to identify the regulatory mechanism between PART1 and miR-122. Results PART1 expression was upregulated in pancreatic cancer tissues and cell lines. High PART1 expression was closely correlated with tumor size, T classification, clinical stage, and vascular invasion, and predicted a poor overall survival. PART1 knockdown significantly suppressed cell proliferation and invasion abilities of pancreatic cancer but promoted cell apoptosis. PART1 was found to serve as a molecular sponge of miR-122, and miR-122 inhibition partially reversed the inhibitory phenotypes of PART1 knockdown on pancreatic cancer cells. Conclusions PART1 promotes the malignant progression of pancreatic cancer by sponging miR-122. The PART1/miR-122 axis might be a promising target for anticancer therapy in patients with pancreatic cancer.

scholarly journals P149 The intracellular chloride channel 4 (CLIC4) plays an important role in systemic sclerosis fibroblast activation

Rheumatology ◽  
2021 ◽  
Vol 60 (Supplement_1) ◽  
Author(s):  
Christopher Wasson ◽  
Rebecca Ross ◽  
Ruth Morton ◽  
Jamel Mankouri ◽  
Francesco Del Galdo
Keyword(s):  
Systemic Sclerosis ◽  
Ion Channel ◽  
Smooth Muscle Actin ◽  
Chloride Ion ◽  
Dermal Fibroblasts ◽  
Cancer Associated Fibroblasts ◽  
Alpha Smooth Muscle Actin ◽  
Direct Role ◽  
Fibroblast Activation ◽  
Intracellular Chloride

Abstract Background/Aims  The intracellular chloride ion channel CLIC4 mediates the activation of cancer associated fibroblasts. Interestingly, systemic sclerosis (SSc) fibroblasts display a number of similar properties to cancer associated fibroblasts. Tissue fibrosis in SSc is driven by active fibroblasts (myofibroblasts). Therefore in this study we investigated the role of CLIC4 in SSc fibroblast activation. Methods  Dermal fibroblasts were obtained from full thickness skin biopsies from SSc patients (early-diffuse). RNA and protein were collected from the fibroblasts and CLIC4 transcript and protein levels were assessed by qPCR and western blot. SSc patient fibroblasts were treated with the chloride ion channel inhibitors NPPB and IAA-94. Results  CLIC4 was found to be expressed at significantly higher levels in SSc patients fibroblasts compared to healthy controls, at both the transcript (3.7 fold) and protein (1.7 fold) levels. Inhibition of the TGF-β signalling pathway led to reduced CLIC4 expression in SSc fibroblasts, confirming this pathway as the main driver of CLIC4 expression. Finally, treatment of SSc fibroblasts with small molecule inhibitors that target the channel led to reduced expression of the myofibroblast markers collagen type 1 and alpha-smooth muscle actin, suggesting a direct role for CLIC4 in SSc associated skin fibrosis. Conclusion  We have identified a novel role for CLIC4 in SSc myofibroblast activation, which further strengthen the similarities between SSc fibroblasts and cancer associated fibroblasts. Furthermore this study highlights this channel as a novel target for therapeutic intervention. Disclosure  C. Wasson: None. R. Ross: None. R. Morton: None. J. Mankouri: None. F. Del Galdo: None.

scholarly journals Long non-coding RNA FOXD2-AS1 promotes cell proliferation, metastasis and EMT in glioma by sponging miR-506-5p

Open Medicine ◽  
2020 ◽  
Vol 15 (1) ◽  
pp. 921-931
Author(s):  
Juan Zhao ◽  
Xue-Bin Zeng ◽  
Hong-Yan Zhang ◽  
Jie-Wei Xiang ◽  
Yu-Song Liu
Keyword(s):  
Cell Proliferation ◽  
Epithelial Mesenchymal Transition ◽  
Human Cancer ◽  
Glioma Cells ◽  
Suppressor Gene ◽  
Luciferase Reporter ◽  
Migration And Invasion ◽  
Mesenchymal Transition ◽  
Non Coding Rna ◽  
Long Non Coding Rna

AbstractLong non-coding RNA forkhead box D2 adjacent opposite strand RNA 1 (FOXD2-AS1) has emerged as a potential oncogene in several tumors. However, its biological function and potential regulatory mechanism in glioma have not been fully investigated to date. In the present study, RT-qPCR was conducted to detect the levels of FOXD2-AS1 and microRNA (miR)-506-5p, and western blot assays were performed to measure the expression of CDK2, cyclinE1, P21, matrix metalloproteinase (MMP)7, MMP9, N-cadherin, E-cadherin and vimentin in glioma cells. A luciferase reporter assay was performed to verify the direct targeting of miR-506-5p by FOXD2-AS1. Subsequently, cell viability was analyzed using the CCK-8 assay. Cell migration and invasion were analyzed using Transwell and wound healing assays, respectively. The results demonstrated that FOXD2-AS1 was significantly overexpressed in glioma cells, particularly in U251 cells. Knockdown of FOXD2-AS1 in glioma cells significantly inhibited cell proliferation, migration, invasion and epithelial–mesenchymal transition (EMT) and regulated the expression of CDK2, cyclinE1, P21, MMP7 and MMP9. Next, a possible mechanism for these results was explored, and it was observed that FOXD2-AS1 binds to and negatively regulates miR-506-5p, which is known to be a tumor-suppressor gene in certain human cancer types. Furthermore, overexpression of miR-506-5p significantly inhibited cell proliferation, migration, invasion and EMT, and these effects could be reversed by transfecting FOXD2-AS1 into the cells. In conclusion, our data suggested that FOXD2-AS1 contributed to glioma proliferation, metastasis and EMT via competitively binding to miR-506-5p. FOXD2-AS1 may be a promising target for therapy in patients with glioma.

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