Circ_0099630 knockdown alleviates lipopolysaccharide-induced injuries of human periodontal ligament cells through the inhibition of TLR4 by releasing miR-409-3p
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ABSTRACT:
SUBMITTER: Qi H
PROVIDER: S-EPMC10675886 | biostudies-literature | 2023 Jan
REPOSITORIES: biostudies-literature
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Project description:BackgroundEvidence has confirmed that forkhead box protein A1 (FOXA1) inhibits the osteogenic differentiation of bone marrow mesenchymal stem cells. However, whether FOXA1 regulates the osteogenic differentiation of human periodontal ligament stem cells (hPDLSCs) to participate in periodontitis process is unclear.MethodsLipopolysaccharide (LPS) was used to treat hPDLSCs to mimic inflammation environments. FOXA1 expression was examined by quantitative real-time PCR and western blot. The levels of IL-6 and TNF-α were evaluated by quantitative real-time PCR, ELISA and immunohistochemistry staining. hPDLSCs osteogenic differentiation was assessed by measuring alkaline phosphatase activity, alizarin red S intensity and the levels of osteogenic differentiation-related markers. Besides, the expression of signal transducer and activator of transcription 3 (STAT3) pathway-related markers were examined by western blot and immunofluorescence staining.ResultsFOXA1 was upregulated in the periodontal ligament tissues of periodontitis patients, and its knockdown enhanced osteogenic differentiation of hPDLSCs. Besides, downregulation of FOXA1 suppressed inflammation levels in LPS-induced hPDLSCs. Also, FOXA1 silencing promoted the osteogenic differentiation of LPS-induced hPDLSCs by the inactivation of STAT3 pathway.ConclusionOur data confirmed that knockdown of FOXA1 attenuated inflammation and enhanced osteogenic differentiation of LPS-induced hPDLSCs by regulating STAT3 pathway, indicating that FOXA1 might be a target for periodontitis treatment.
| S-EPMC11613586 | biostudies-literature
Project description:BackgroundPeriodontitis is a common oral inflammatory disease, and lipopolysaccharide (LPS) is a key risk factor in periodontitis pathology. Here, we used LPS-induced periodontal ligament cells (PDLCs) to explore the molecular mechanism of periodontitis.MethodsCell viability, proliferation, and apoptosis were analyzed by Cell Counting Kit-8, 5-ethynyl-20-deoxyuridine (EDU), and flow cytometry assays, respectively. Apart from that, their targeting relationship was validated using dual-luciferase reporter and RNA-pull down.ResultsCircular RNA_0138960 (circ_0138960) was notably upregulated in periodontitis sufferers (p < .001) and LPS-disposed PDLCs (p < .05). LPS exposure dampened PDLC proliferation, and promoted apoptosis and inflammation (p < .05). Circ_0138960 acted as a microRNA sponge for miR-518a-5p to affect histone deacetylase 6 (HDAC6) expression. Circ_0138960 absence-mediated protective effects in LPS-induced PDLCs were largely abrogated via silencing miR-518a-5p or HDAC6 overexpression (p < .05).ConclusionCirc_0138960 promoted LPS-induced dysfunction in PDLCs by targeting miR-518a-5p/HDAC6 axis, which provided novel potential therapeutic targets for periodontitis.
| S-EPMC9639461 | biostudies-literature
Project description:BackgroundTamoxifen (TAM) is a frequently-used treatment for breast cancer (BC). But the TAM resistance seriously affects the patient therapeutic effect. Previous research indicated that circular RNAs (circRNAs) might participate in the regulatory processes of BC. Here, we discovered the parts of circular RNA tripartite motif-containing 28 (circTRIM28) in BC.MethodsCircTRIM28, microRNA-409-3p (miR-409-3p), and high mobility group AT-hook 2 (HMGA2) levels were perceived by qRT-PCR and western blot. Moreover, the biological functions of the cells were examined. Furthermore, dual-luciferase report was employed to reconnoiter the targeted relationship between miR-409-3p and circTRIM28 or HMGA2.ResultsCircTRIM28 and HMGA2 were augmented, and the miR-409-3p was repressed in BC. Silencing circTRIM28 enhanced tamoxifen sensitivity and cell apoptosis, whereas hampered cell development in BC cells. In mechanism, circTRIM28 could sponge miR-409-3p to increase HMGA2. In addition, silencing circTRIM28 impeded tumor growth.ConclusionCircTRIM28 facilitated the BC via miR-409-3p/HMGA2.
| S-EPMC9524098 | biostudies-literature
Project description:Background Periodontitis is an inflammatory destructive bone disease and is the most critical cause of tooth loss in adults. Recent studies have reported that circular RNAs (circRNAs) are essential in periodontitis. However, the influence and mechanism of hsa_circ_0099630 on periodontitis are not clear. Methods Normal periodontal tissues and inflammatory periodontal tissues were obtained from healthy patients and patients with periodontitis, respectively. Hsa_circ_0099630 was 1st identified by polymerase chain reaction (PCR) and sanger sequencing, and hsa_circ_0099630 expression was determined by real-time (RT)-quantitative PCR in periodontitis. Porphyromonas gingivalis-lipopolysaccharide (Pg-LPS) was used to construct an inflammation model in vitro. Next, cell proliferation, apoptosis, and osteogenic differentiation were monitored using Cell Counting Kit-8, flow cytometry, and western blot in the Pg-LPS-induced human periodontal ligament fibroblasts (HPLFs). The microRNA (miRNA)/messenger RNA (mRNA) axis of hsa_circ_0099630 was predicted and screened, and the function of the target genes was analyzed by a Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses. Results The identified hsa_circ_0099630 was upregulated in the gingival tissue of patients with periodontitis. Next, an inflammation model was constructed using Pg-LPS in the HPLFs. We discovered that Pg-LPS or hsa_circ_0099630 overexpression suppressed cell proliferation and osteogenic differentiation, and induced apoptosis in HPLFs. Additionally, hsa_circ_0099630 knockdown induced proliferation and osteogenic differentiation and prevented apoptosis in the Pg-LPS-induced HPLFs. We also screened the vast miRNA/mRNA axis associated with hsa_circ_0099630. Conclusions The current study uncovered the crucial role of hsa_circ_0099630 in periodontitis.
| S-EPMC9577728 | biostudies-literature
Project description:Pneumonia is an inflammatory disease that occurs in the lungs associated with pathogens or other factors. It has been well established that long noncoding RNA X inactivate-specific transcript (XIST) is involved in several cancers. The present study focused on the effect and detailed mechanism of XIST in lipopolysaccharide (LPS)-induced injury in pneumonia. Here, XIST was silenced by transfection with XIST-targeted siRNA, and then, mRNA expression, cell viability, apoptosis, and protein expression were, respectively, assessed by qRT-PCR, CCK-8, flow cytometry, and Western blotting. Luciferase reporter, RIP, and RNA pull-down assays were used to detect the combination of miR-370-3p and XIST. Besides, the tested proinflammatory factors were analysed by qRT-PCR and Western blot, and their productions were quantified by ELISA. The results showed that XIST expression was robustly increased in serum of patients with acute-stage pneumonia and LPS-induced WI-38 human lung fibroblasts cells. Functional analyses demonstrated that knockdown of XIST remarkably alleviated LPS-induced cell injury through increasing cell viability and inhibiting apoptosis and inflammatory cytokine levels. Mechanistically, XIST functioned as a competitive endogenous RNA (ceRNA) by effectively binding to miR-370-3p and then restoring TLR4 expression. More importantly, miR-370-3p inhibitor abolished the function of XIST knockdown on cell injury and JAK/STAT and NF-κB pathways. Taken together, XIST may be involved in progression of cell inflammatory response, and XIST/miR-370-3p/TLR4 axis thus may shed light on the development of novel therapeutics to the treatment of acute stage of pneumonia. SIGNIFICANCE OF THE STUDY: Our study demonstrated that XIST was highly expressed in patients with acute stage of pneumonia. Knockdown of XIST remarkably alleviated LPS-induced cell injury through increasing cell viability and inhibiting apoptosis and inflammatory cytokine levels through regulating JAK/STAT and NF-κB pathways.
| S-EPMC6618287 | biostudies-literature
Project description:Osteoarthritis is a chronic degenerative joint disease. Long non‑coding RNA plasmacytoma variant translocation 1 (PVT1) is involved in the progression of osteoarthritis and exosomes serve a central role in intercellular communication. However, whether PVT1 can be mediated by exosomes in osteoarthritis has not been reported. Whole blood was drawn from osteoarthritis patients and healthy volunteers. Lipopolysaccharide (LPS) was used to stimulate human normal chondrocytes (C28/I2) to construct a cell damage model in vitro. Protein levels were examined via western blot analysis. eThe expression of PVT1, microRNA (miR)‑93‑5p and high mobility groupprotein B1 (HMGB1) was evaluated through reverse transcription‑quantitative PCR. Cell viability and apoptosis were determined through CCK‑8 or flow cytometric assay. Inflammatory cytokines were measured via ELISA. The relationship between PVT1 or HMGB1 and miR‑93‑5p was confirmed by dual‑luciferase reporter assay. PVT1, HMGB1 and exosomal PVT1 were upregulated while miR‑93‑5p was downregulated in osteoarthritis patient serum and LPS‑induced C28/I2 cells. Exosomes from osteoarthritis patient serum and LPS‑treated C28/I2 cells increased PVT1 expression in C28/I2 cells. PVT1 depletion reversed the decrease of viability and the increase of apoptosis, inflammation responses and collagen degradation of C28/I2 cells induced by LPS. PVT1 regulated HMGB1 expression via sponging miR‑93‑5p. miR‑93‑5p inhibition abolished PVT1 silencing‑mediated viability, apoptosis, inflammation responses and collagen degradation of LPS‑stimulated C28/I2 cells. HMGB1 increase overturned miR‑93‑5p upregulation‑mediated viability, apoptosis, inflammation responses and collagen degradation of LPS‑stimulated C28/I2 cells. Furthermore, PVT1 modulated the Toll‑like receptor 4/NF‑κB pathway through an miR‑93‑5p/HMGB1 axis. In summary, exosome‑mediated PVT1 regulated LPS‑induced osteoarthritis progression by modulating the HMGB1/TLR4/NF‑κB pathway via miR‑93‑5p, providing a new route for possible osteoarthritis treatment.
| S-EPMC7646997 | biostudies-literature
Project description:ObjectiveEphrinA2-EphA2 and ephrinB2-EphB4 critically engage in bidirectional signalling to modulate alveolar bone remodelling. The present study aimed to investigate the effects of lipopolysaccharides (LPS) derived from Porphyromonas gingivalis on ephrin/Eph signalling in periodontal ligament fibroblasts (PDLFs).Material and methodsThe primary cultured PDLFs were incubated in the absence (as a control) or presence of P. gingivalisLPS at 0.001-10 μg/mL for 24 hours. The PDLFs were then stimulated with P. gingivalisLPS at the optimal concentration (0.1 μg/mL) for different periods (6-48 hours). The expression of ephrinA2, ephrinB2, EphA2 and EphB4 was assessed by quantitative reverse-transcription real-time polymerase chain reaction and western blotting. The osteoblastic markers alkaline phosphatase, osteocalcin and Runt-related transcription factor 2 (Runx2), and the osteoclastogenesis-related factors receptor activator of nuclear factor kappa-B ligand (RANKL) and osteoprotegerin were also evaluated.ResultsThe ephrinA2 and EphA2 expression was upregulated and EphB4 expression was downregulated by stimulation of P. gingivalisLPS. EphrinA2 mRNA expression in the PDLFs was significantly upregulated from 12 to 48 hours (P<.05), whereas EphA2 exhibited no change for the first 24 hours, after which there was a significant increase at 48 hours (P<.05). EphB4 exhibited lower mRNA expression at 12 and 24 hours than did the control (P<.05), but the change was insignificant at 48 hours. In contrast, the expression of ephrinB2 remained unchanged. The expressions of ephrinA2, EphA2, ephrinB2 and EphB4 at the protein level showed a similar pattern to that at the mRNA level. The expression of Runx2 and osteocalcin significantly decreased, whereas that of RANKL/osteoprotegerin increased.ConclusionThe present study suggest that P. gingivalisLPS would contribute to a dysregulation of bone remodelling, whereby ephrinA2/EphA2 expression is stimulated and EphB4 expression is inhibited.
| S-EPMC5600103 | biostudies-literature
Project description:Periodontitis is a chronic infectious disease that affects the oral health of adults. Long non-coding RNA OIP5 antisense RNA 1 (OIP5-AS1) has been reported to downregulated in the periodontal tissue of patients with periodontitis. Therefore, the study sought to look at the possible functions of OIP5-AS1 in periodontitis and the associated underlying mechanisms. In the present study, the expression level of OIP5-AS1 and microRNA-92a-3p were analyzed using reverse transcription-quantitative PCR. The levels of osteogenic proteins were determined using western blotting and inflammatory cytokines and oxidative stress were also examined. The proliferation of human periodontal ligament stem cells (hPDLSCs) was evaluated using MTT assays. Assay of osteogenic differentiation was undertaken by means of Alkaline phosphatase staining. The possible association between OIP5-AS1 and miR-92a-3p was determined applying dual-luciferase reporter assays and verified by RNA immunoprecipitation assay. We found that OIP5-AS1 was expressed at low levels in lipopolysaccharide (LPS)-stimulated hPDLSCs. OIP5-AS1 overexpression promoted proliferation and osteogenic differentiation ability and reduced LPS-induced inflammation in hPDLSCs. Furthermore, OIP5-AS1 directly targeted and reduced miR-92a-3p expression. The overexpression of miR-92a-3p partly abolished the effects of OIP5-AS1 on LPS-induced cell proliferation and osteogenic differentiation as well as inflammation in hPDLSCs. Collectively, the results indicated that OIP5-AS1 overexpression inhibited the LPS-induced inflammatory response and promoted the osteogenic differentiation of hPDLSCs by sponging miR-92a-3p. Thus, OIP5-AS1 is probably an essential objective for research during periodontitis treatment.
| S-EPMC9276041 | biostudies-literature
Project description:The upregulation of circ_0001679 was reported in lipopolysaccharide (LPS)-induced lung injury mouse model, but its functional roles and mechanisms in LPS-induced lung injury remain to be investigated. In this study, we aimed to explore the potential role of circ_0001679 in septic acute lung injury. We initially established an in vitro lung cell injury model using LPS-treated MLE-12 cells. siRNAs targeting circRNA_0001679 were employed to stably knock down circRNA_0001679, followed by functional assays to investigate the effect of circRNA_0001679 silencing. The levels of inflammatory cytokines such as IL-6, IL-β and TNF-α (Tumor necrosis factor-α) were detected by ELISA (Enzyme-linked immunosorbent assay). Meanwhile, protein levels of Bcl-2, cleaved-caspase 3, Bax, and MAPK1 (Mitogen-Activated Protein Kinase 1) proteins expression level were measured by Western blot. We found that Circ_0001679 was upregulated in LPS-induced MLE-12 cells, and silencing circ_0001679 attenuated the growth inhibition and suppressed apoptosis induced by LPS. Circ_0001679 knockdown also lowered levels of IL-6, IL-β and TNF-α, and prevent the activation of cleaved-caspase 3 protein. We further revealed that circ_0001679 functioned as a sponge of miR-338-3p to negatively regulate miR-338-3p activity. miR-338-3p downregulated its downstream target MAPK1, while the upregulation of circ_0001679 maintained a high-level expression of MAPK1 by suppressing miR-338-3p. Collectively, our study indicates that circ_0001679/miR-338-3p/MAPK1 axis may play an important role in the pathogenesis of acute lung injury (ALI).
| S-EPMC8973724 | biostudies-literature
Project description:BackgroundCircular RNAs have emerged as important regulators in the pathogenesis of human diseases, including infantile pneumonia (IP). In this study, we aimed to explore the effects of circ_0035292 on lipopolysaccharide (LPS)-treated Wistsar Institute (WI)-38 cells.MethodsQuantitative real-time polymerase chain reaction and western blot were executed to detect the levels of circ_0035292, microRNA-370-3p (miR-370-3p) and transducin β-like 1X related protein 1 (TBL1XR1). Cell counting kit-8, 5-ethynyl-2'-deoxyuridine, and flow cytometry assessed cell proliferation and apoptosis. Concentrations of inflammatory factors were examined with enzyme linked immunosorbent assay kits. Dual-luciferase reporter assay and RNA immunoprecipitation were adopted to analyze binding between miR-370-3p and circ_0035292 or TBL1XR1.ResultsCirc_0035292 level was increased in IP patients and LPS-triggered WI-38 cells. Circ_0035292 knockdown rescued LPS-mediated WI-38 cell proliferation suppression and WI-38 cell apoptosis and inflammation promotion. Circ_0035292 interacted with miR-370-3p and miR-370-3p directly targeted TBL1XR1. Moreover, miR-370-3p overexpression alleviated LPS-induced WI-38 cell apoptosis and inflammatory injury, which was abrogated via TBL1XR1 upregulation. Circ_0035292 absence inhibited the NF-κB pathway.ConclusionKnockdown of circ_0035292 rescued LPS-triggered WI-38 cell injury via miR-370-3p/TBL1XR1 axis and NF-κB pathway.
| S-EPMC10266152 | biostudies-literature