Paclitaxel inhibits proliferation by negatively regulating Cdk1-cell cycle axis in rat airway smooth muscle cells.

OBJECTIVE: Paclitaxel exhibits outstanding biological activities in inhibiting cell proliferation and inducing cell apoptosis. However, the effects of paclitaxel on airway smooth muscle cells (ASMCs) have not been reported yet. The purpose of this study is to determine the effects of paclitaxel on the proliferation and apoptosis of ASMCs. METHODS: Rat primary ASMCs were isolated and used in all the experiments. Cell Counting Kit-8 assay and Edu assay were used to analyze the cell viability and proliferation, respectively. Flow cytometry was used to detect the cell cycle and apoptosis. Quantitative real-time PCR (qRT-PCR), western blotting, and immunostaining were used to detect the expression of cyclin-dependent kinase 1 (Cdk1). RESULTS: Our study showed that paclitaxel inhibits the proliferation of ASMCs in a dose- and time-gradient-dependent manner. Further study displayed that cell cycle is arrested at G2/M phase. And Cdk1 was dramatically down-regulated by paclitaxel treatment. Cell morphological analysis showed that ASMCs are elliptical with a larger surface area after paclitaxel treatment. Nucleus morphological analysis showed that the nuclei are in a diffuse state after paclitaxel treatment. However, paclitaxel did not induce the apoptosis of ASMCs. CONCLUSIONS: Our study demonstrated that paclitaxel inhibits the proliferation of ASMCs at least partly by negatively regulating Cdk1-cell cycle axis.

Human beta defensin 3 knockdown inhibits the proliferation and migration of airway smooth muscle cells through regulating the PI3K/AKT signaling pathway.

Asthma, a common pediatric pulmonary disease, significantly affects children's healthy development. This study aimed to investigate the functions of human ß defensin-3 (HBD-3) in asthma progression. For this purpose, blood samples from asthmatic and healthy children were collected. Moreover, the airway smooth muscle cells (ASMCs) were treated with platelet-derived growth factor BB (PDGF-BB) to develop an in vitro asthma model, then evaluated cell viability and migration via CCK-8 and transwell assays. The mRNA levels of interferon γ (INF-γ), interleukin 4 (IL-4), interleukin 10 (IL-10), alpha-smooth muscle actin (α-SMA), HBD-3, and the protein levels of phosphatidylinositol 3-kinase (PI3K) along with protein kinase B (AKT) were detected. Similarly, the N6-methyladenosine (m6A) content in the ASMCs and m6A levels of HBD-3 were also measured. Results indicated an upregulated HBD-3 in the asthmatic children. The ASMCs were found to be stimulated by PDGF-BB, in addition to the promotion of cell viability and migration. The INF-γ, IL-4, and α-SMA levels were reduced, while IL-10 was elevated in PDGF-BB-stimulated ASMCs. Silencing HBD-3 in PDGF-BB stimulated ASMCs was found to exert the opposite effect by inhibiting cell viability and migration, enhancing the levels of INF-γ, IL-4, and α-SMA, while the IL-10 levels were found to decline. PDGF-BB stimulation of ASMCs resulted in activation of the PI3K/AKT signaling pathway, which was blocked post HBD-3 silencing, while the role of si-hBD in PDGF-BB stimulated ASMCs was neutralized post-treatment with IGF-1. Finally, it was found that METTL3 overexpression prominently upregulated the m6A levels of HBD-3 and decreased the mRNA expression and stability of HBD-3 in the PDGF-BB-stimulated ASMCs. The study concluded that METTL3-mediated HBD-3 participates in the progression of asthma through the PI3K/AKT signaling pathway.

Nerolidol attenuates airway inflammation and airway remodeling and alters gut microbes in ovalbumin-induced asthmatic mice.

Asthma is a common respiratory disease associated with airway inflammation. Nerolidol is an acyclic sesquiterpenoid with anti-inflammatory properties. BALB/C mice were sensitized with ovalbumin (OVA) to induce asthma symptoms and given different doses of Nerolidol. We found that Nerolidol reduced OVA-induced inflammatory cell infiltration, the number of goblet cells and collagen deposition in lung tissue. Nerolidol reduced the OVA-specific IgE levels in serum and alveolar lavage fluid in an asthma model. Immunohistochemical staining of α-SMA (the marker of airway smooth muscle) showed that Nerolidol caused bronchial basement membrane thinning in asthmatic mice. The hyperplasia of airway smooth muscle cells (ASMCs) is an important feature of airway remodeling in asthma. ASMCs were treated with 10 ng/mL TGF-ß to simulate the pathological environment of asthma in vitro and then treated with different doses of Nerolidol. Nerolidol inhibited the activity of TGF-ß/Smad signaling pathway both in the lung tissue of OVA-induced mouse and TGF-ß-stimulated ASMCs. 16s rRNA sequencing was performed on feces of normal mice, the changes of intestinal flora in OVA-induced asthmatic mice and Nerolidol-treated asthmatic mice were studied. The results showed that Nerolidol reversed the reduced gut microbial alpha diversity in asthmatic mice. Nerolidol changed the relative abundance of gut bacteria at different taxonomic levels. At the phylum level, the dominant bacteria were Bacteroidota, Firmicutes, and Proteobacteria. At the genus level, the dominant bacteria were Lactobacillus, Muribaculaceae, Bacteroides, and Lachnospiraceae. We conclude that Nerolidol attenuates OVA-induced airway inflammation and alters gut microbes in mice with asthma via TGF-ß/Smad signaling.

PM2.5 induces lung inflammation and fibrosis via airway smooth muscle cell expression of the Wnt5a/JNK pathway.

Background: In recent years, particulate matter 2.5 (PM2.5) exposure has been considered a key dangerous factor in chronic obstructive pulmonary disease (COPD). The dysfunction of airway smooth muscle cells (ASMCs) facilitates lung inflammation and fibrosis in COPD. Therefore, we explored whether PM2.5 could promote the inflammatory response and fibrosis in ASMCs in vivo and in vitro via the wingless-related integration site 5a (Wnt5a)/c-Jun N-terminal kinase (JNK)/nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathway. Methods: Wnt5a expression in the bronchoalveolar lavage fluid (BALF) of COPD patients exposed to PM2.5 was measured by enzyme-linked immunosorbent assay (ELISA). Mice were intratracheally injected with PM2.5 and a Wnt5a antagonist (BOX5). ASMCs were transfected with Wnt5a small interfering RNA (siRNA), BOX5 and the JNK inhibitor SP600125 before PM2.5 stimulation. Hematoxylin and eosin (H&E) staining was performed to measure the inflammatory response and airway fibrosis. The production of Wnt5a/JNK/NF-KB pathway factors was analyzed by Western blotting. The secretion of interleukin-6 (IL-6), IL-8 and tumor necrosis factor-α (TNF-α) was measured by ELISA. The expression levels of alpha smooth muscle actin (α-SMA), collagen I and collagen III were assessed by quantitative real time polymerase chain reaction (qRT-PCR) and Western blotting. Results: We found that the increase in Wnt5a expression in the BALF of COPD patients was positively correlated with the levels of PM2.5 exposure. The Wnt5a/JNK/NF-κB pathway was activated in the lung samples of PM2.5-induced model mice and PM2.5-exposed ASMCs, which promoted the production of α-SMA, collagen I and collagen III and increased the secretion of IL-6, IL-8 and TNF-α. Furthermore, our results showed that BOX5 could prevent these effects. Wnt5a siRNA blocked the activation of the Wnt5a/JNK/NF-κB pathway and inhibited the effects of PM2.5 on fibrosis and inflammation in ASMCs. SP600125 blocked the phosphorylation of NF-κB and inhibited inflammation and fibrosis in PM2.5-exposed ASMCs. Conclusions: These findings suggest that PM2.5 stimulation of ASMCs induces pulmonary inflammatory factor expression and collagen deposition during COPD via the Wnt5a/JNK pathway, which indicates that modulating the Wnt5a/JNK pathway could be a promising therapeutic strategy for PM2.5-induced COPD.

MiR-182/Sestrin2 affects the function of asthmatic airway smooth muscle cells by the AMPK/mTOR pathway.

Background and Objectives: Asthma is a chronic inflammatory airway disease and brings heavy economic and spiritual burdens to patients' families and the society. Airway smooth muscle cells (ASMCs) afect the development of asthma by secreting cytokines, growth factors, and prostates. The stress-inducing protein, Sestrin2, plays a vital role in antioxidant defense. The aim of this study is to investigate the role of Sestrin2 in asthma and its corresponding molecular mechanism. Materials and Methods: Airway remodeling was induced by construction of asthma rat model. Primary ASMCs were isolated through combining tissue block adherence and enzymatic digestion and identified by immunofluorescence staining. Gene expression was measured by quantitative real-time PCR (qPCR) and western blot (WB) experiments. Cell viability, proliferation, migration, and calcium flow of ASMCs were measured by Cell Counting Kit-8 (CCK-8), 5-ethynyl-deoxyuridine (EdU), Transwell, and Fluo-3AM, respectively. The binding of miR-182 and Sestrin2 3'-untranslated region (3'-UTR) was measured by luciferase reporter system and RNA-binding protein immunoprecipitation (RIP) analysis. Results: Sestrin2 expression was upregulated in asthma rat model and cell model. Overexpression of Sestrin2 enhanced the growth, migration, and calcium flow, and inversely, repression of Sestrin2 was reduced in ASMCs from the asthma group. MiR-182, one of the microRNAs (miRNAs) that possesses the potential to regulate Sestrin2, was downregulated in ASMCs from the asthma group. Further experiments revealed that Sestrin2 was inhibited by miR-182 and that overexpression of Sestrin2 reversed the miR-182-induced inhibition of the cellular progression of ASMCs from the asthma group. This study further investigated the downstream signaling pathway of Sestrin2 and found that increased expression of Sestrin2 activated 5'-adenosine monophosphate-activated protein kinase (AMPK), leading to the inactivation of mammalian target of rapamycin (mTOR) and thus promoting the growth, migration, and calcium flow of ASMCs from the asthma group. Conclusion: This study investigated the role of Sestrin2 for the first time and further dissected the regulatory factor of Sestrin2, ultimately elucidating the downstream signaling pathway of Sestrin2 in asthma, providing a novel pathway, and improving the understanding of the development and progression of asthma.

LncRNA DGCR5/miR-204-5p/SRSF7 axis regulates PDGF-BB-induced proliferation and migration of airway smooth muscle cells with potential role in asthma.

Increased proliferation and migration of abnormal airway smooth muscle cells (ASMCs) are significantly associated with asthma. Recently, the function of long non-coding RNAs (lncRNAs) in ASMCs has been identified. Our research attempted to resolve the function of the lncRNA DGCR5 in ASMCs and focus on its in-depth mechanisms of proliferation and migration. Quantitative real-time PCR and western blotting were used to evaluate the expressions of RNA and proteins, respectively. The CCK-8 assay was used to detect cell proliferation. Transwell assays were used to assess migration. Luciferase and RNA pull-down assays were used to verify the targeting between miR-204-5p and lncRNA DGCR5 or SRSF7. LncRNA DGCR5 was upregulated in ASMCs stimulated with PDGF-BB. Knockdown of DGCR5 reversed the effect of platelet-derived growth factor BB (PDGF-BB) on the proliferation and migration of ASMCs. The results of this study showed that lncRNA DGCR5 binds to miR-204-5p, and SRSF7 is the target of miR-204-5p. Rescue experiments verified the interaction between miR-204-5p and DGCR5, as well as that between miR-204-5p and SRSF7. Overall, this study revealed that the lncRNA DGCR5/miR-204-5p/SRSF7 signalling axis has key regulatory effects on the proliferation and migration of ASMCs.

Serum from acupuncture-treated asthmatic rats regulates p38-MAPK activation in airway smooth muscle cells.

OBJECTIVES: This study aimed to investigate the regulation function of acupuncture on airway smooth muscle cells (ASMCs) of asthmatic rats. METHODS: Male Sprague-Dawley rats were challenged using inhalational Ovalbumin (OVA) to establish an asthma model. The acupuncture points (GV14 for Dazhui, bilateral BL12 for Fengmen, and bilateral BL13 for Feishu) were stimulated for asthma relief. The ASMCs isolated from asthmatic rats were incubated in medium containing the serum obtained from asthmatic rats treated with acupuncture. The expression levels of p38 MAPK and p-p38 MAPK were determined by immunocytochemical and western blot. RESULTS: ASMCs were successfully isolated and cultured. The 20% acupuncture treatment of asthmatic rat serum had the least effect on the proliferation ability of asthmatic ASMCs. The serum from asthmatic rats treated with acupuncture could decrease the expression of p-p38 MAPK in asthmatic rat ASMCs. CONCLUSIONS: The serum from acupuncture-treated asthmatic rats has an effect on treating asthma in rats, and the mechanism of action may be by regulating the p38 pathway.

GLI1+ cells are a source of repair-supportive mesenchymal cells (RSMCs) during airway epithelial regeneration.

Repair-supportive mesenchymal cells (RSMCs) have been recently reported in the context of naphthalene (NA)-induced airway injury and regeneration. These cells transiently express smooth muscle actin (Acta2) and are enriched with platelet-derived growth factor receptor alpha (Pdgfra) and fibroblast growth factor 10 (Fgf10) expression. Genetic deletion of Ctnnb1 (gene coding for beta catenin) or Fgf10 in these cells using the Acta2-Cre-ERT2 driver line after injury (defined as NA-Tam condition; Tam refers to tamoxifen) led to impaired repair of the airway epithelium. In this study, we demonstrate that RSMCs are mostly captured using the Acta2-Cre-ERT2 driver when labeling occurs after (NA-Tam condition) rather than before injury (Tam-NA condition), and that their expansion occurs mostly between days 3 and 7 following NA treatment. Previous studies have shown that lineage-traced peribronchial GLI1+ cells are transiently amplified after NA injury. Here, we report that Gli1 expression is enriched in RSMCs. Using lineage tracing with Gli1Cre-ERT2 mice combined with genetic inactivation of Fgf10, we show that GLI1+ cells with Fgf10 deletion fail to amplify around the injured airways, thus resulting in impaired airway epithelial repair. Interestingly, Fgf10 expression is not upregulated in GLI1+ cells following NA treatment, suggesting that epithelial repair is mostly due to the increased number of Fgf10-expressing GLI1+ cells. Co-culture of SCGB1A1+ cells with GLI1+ cells isolated from non-injured or injured lungs showed that GLI1+ cells from these two conditions are similarly capable of supporting bronchiolar organoid (or bronchiolosphere) formation. Single-cell RNA sequencing on sorted lineage-labeled cells showed that the RSMC signature resembles that of alveolar fibroblasts. Altogether, our study provides strong evidence for the involvement of mesenchymal progenitors in airway epithelial regeneration and highlights the critical role played by Fgf10-expressing GLI1+ cells in this context.

Effect of the BMPR-II-SMAD3/MRTF pathway on proliferation and migration of ASMCs and the mechanism in asthma.

BACKGROUND: A variety of smooth muscle-specific genes and proteins, including SMAD3, BMPR-II, and MRTF, are involved in airway remodeling in asthma. As a receptor of bone morphogenetic protein (BMP) signaling, BMPR-II has important roles in airway remodeling in asthma. However, the underlying mechanism of BMPR-II in airway smooth muscle cells (ASMCs) in asthma remains incomplete. METHODS: Wistar rats were intraperitoneally injected with ovalbumin antigen suspension and aluminium hydroxide and, stimulated with ovalbumin nebulized inhalation to constructed asthma model. Primary ASMCs were isolated with collagenase I and identified by testing the α-SMA expression. Quantitative polymerase chain reaction (qPCR) and western blot assay were employed to detect the gene expression. CCK8, Transwell and Fluo-4 A assays were introduced to measure the cell viability, migration and intracellular Ca2+. Co-Immunoprecipitation (Co-IP) assay was applied to test the interaction among proteins. RESULTS: First, we observed significant increases in BMPR-II in asthmatic rat model and ASMCs at both the mRNA and protein levels. Second, we observed that silencing of siBMPR-II inhibited proliferation, migratory capacity and intracellular Ca2+ concentration in ASMCs. Furthermore, our study demonstrated that siBMPR-II inhibited the Smad3 expression and overexpression promoted the bioactivity of ASMCs. In addition, this study showed that p-Smad3 could interacted with MRTF and siMRTF inhibits the bioactivity of ASMCs. Finally, our results revealed BMPR-II-SMAD3/MRTF pathway affected the bioactivity of ASMCs. CONCLUSIONS: This study indicates that the BMPR-II-SMAD3/MRTF signaling pathway is involved in the process of ASMCs remodeling, providing novel avenues for the identification of new therapeutic modalities.

Cellular sources of airway smooth muscle cells in asthmatic airway remodeling and their clinical relevance: a narrative review.

Background and Objective: Airway remodeling in asthma refers to numerous structural changes in the airway in asthmatic patients, with thickening of the airway smooth muscle layer as its core feature. However, the nature and sources of the abnormally increased airway smooth muscle cells (ASMCs) in airway remodeling remain unclear. ASMCs play a key role in the pathogenesis of fatal asthma; therefore, it is important to clarify the properties and sources of these ASMCs responsible for asthmatic airway remodeling, which may provide a new direction for the precise treatment for asthma. Methods: We performed a narrative review of the literature on PubMed, Web of Science, and Google Scholar databases searching for the cellular sources of ASMCs in asthmatic airway remodeling and their clinical relevance. Key Content and Findings: It has long been thought that ASMCs are the result of abnormal proliferation of the native ASMCs in asthma; however, increasing evidence suggests that increased "ASMCs" may be due to the differentiation/transdifferentiation of other cells including mesenchymal stem cells (MSCs), myofibroblasts (MYFs), pericytes, and epithelial-mesenchymal transition (EMT). Recently, several pharmacological and biological therapies aimed at "reducing asthmatic ASMCs" have been developed, among which gallopamil, JQ1 [an inhibitor of the bromodomain and extra-terminal domain (BET) protein family], and histone deacetylase (HDAC) inhibitors can alleviate asthma airway remodeling and hyperresponsiveness and improve asthma symptoms in both mouse models and preclinical experiments. Conclusions: As one of the core features of asthma, ASMCs are an important effector of airway remodeling. It has become extremely important to develop therapies for the reduction and prevention of the "ASMCs" on the basis of the properties and sources of "ASMCs". Many studies have shown that epigenetic regulation is closely related to the abnormal increase of ASMCs in asthma, and interfering with epigenetic regulation factors can reduce the increased smooth muscle cells. Although the epigenetic regulation of asthma is still in its nascent stage, epigenetic therapy targeting "ASMCs" may become another new strategy for asthma prevention and treatment.

JNJ0966 inhibits PDGF-BB-induced airway smooth muscle cell proliferation and extracellular matrix production by regulating MMP-9.

The increased proliferation and extracellular matrix (ECM) production of airway smooth muscle cells (ASMCs) are crucial factors in asthma progression. JNJ0966, one of the metalloproteinase-9 (MMP-9)-specific inhibitors, has been demonstrated to be involved in the progression and development of diversified diseases. Nevertheless, the function of JNJ0966 in ASMCs remains unclear. This study aimed at investigating the effects of JNJ0966 on asthma progression. In our study, the platelet-derived growth factor BB (PDGF-BB) was first utilized to stimulate the cell model for asthma. Results demonstrated that the cell viability of ASMCs was increased by PDGF-BB (0, 10, 20, and 30 ng/mL) in a dose-dependent manner. Further investigation revealed that JNJ0966 inhibited the cell activity and migration ability of PDGF-BB-induced ASMCs. In addition, JNJ0966 relieved ECM deposition in PDGF-BB-induced ASMCs. Finally, through rescue assays, the results showed that overexpression of MMP-9 reversed the inhibitory effects of JNJ0966 on cell viability and ECM deposition in ASMCs. In conclusion, our findings suggested that JNJ0966 inhibited PDGF-BB-induced ASMC proliferation and ECM production by modulating MMP-9. These findings might provide novel insight for the treatment of asthma.

RAB11A aggravates PDGF-BB-stimulated proliferation, migration, and inflammation of airway smooth muscle cells via affecting the NF-κB and PI3K/AKT pathways.

BACKGROUND: Pediatric asthma is an usual disease and a kind of fearful health threat for children. Airway smooth muscle cells (ASMCs) with increased cell proliferation and migration abilities serve as important features in the progression of asthma. RAB11A has been shown to aggravate cancer progression and is closely associated with inflammation. Gene analysis discovered that RAB11A exhibited higher expression in asthmatic patients. However, the detailed regulatory function of RAB11A in asthma still needs further investigation. METHOD: The mRNA and protein expressions of genes were examined through RT-qPCR and western blot. Cell proliferation was examined through MTT and BrdU assays. Cell apoptosis was tested through flow cytometry. The cell migration ability was detected through wound healing and transwell assays. The levels of TNF-α, IL-1ß, IL-8, and IL-6 were measured through ELISA. RESULT: In this study, the mRNA and protein expressions of RAB11A were increased with PDGF-BB treatment in a dose-dependent manner. Additionally, the silencing of RAB11A suppressed the proliferation ability of PDGF-BB-mediated ASMCs. Moreover, it was uncovered that the knockdown of RAB11A inhibited the migration ability of PDGF-BB-stimulated ASMCs. Besides, suppression of RAB11A relieved the inflammatory response in PDGF-BB-stimulated ASMCs. Lastly, inhibition of RAB11A retarded the NF-κB and PI3K/AKT pathways. CONCLUSION: Our results revealed that RAB11A aggravated PDGF-BB-stimulated proliferation, migration, and inflammation of ASMCs through modulating NF-κB and PI3K/AKT signaling pathways. This finding implied that the RAB11A may be deemed as a novel and prospective biomarker for asthma treatment.

JNJ0966 inhibits PDGF-BB-induced airway smooth muscle cell proliferation and extracellular matrix production by regulating MMP-9

The increased proliferation and extracellular matrix (ECM) production of airway smooth muscle cells (ASMCs) are crucial factors in asthma progression. JNJ0966, one of the metal-loproteinase-9 (MMP-9)-specific inhibitors, has been demonstrated to be involved in the pro-gression and development of diversified diseases. Nevertheless, the function of JNJ0966 in ASMCs remains unclear. This study aimed at investigating the effects of JNJ0966 on asthma progression. In our study, the platelet-derived growth factor BB (PDGF-BB) was first utilized to stimulate the cell model for asthma. Results demonstrated that the cell viability of ASMCs was increased by PDGF-BB (0, 10, 20, and 30 ng/mL) in a dose-dependent manner. Further investigation revealed that JNJ0966 inhibited the cell activity and migration ability of PDGF-BB-induced ASMCs. In addition, JNJ0966 relieved ECM deposition in PDGF-BB-induced ASMCs. Finally, through rescue assays, the results showed that overexpression of MMP-9 reversed the inhibitory effects of JNJ0966 on cell viability and ECM deposition in ASMCs. In conclusion, our findings suggested that JNJ0966 inhibited PDGF-BB-induced ASMC proliferation and ECM pro-duction by modulating MMP-9. These findings might provide novel insight for the treatment of asthm (AU)

RAB11A aggravates PDGF-BB-stimulated proliferation, migration, and inflammation of airway smooth muscle cells via affecting the NF-kB and PI3K/AKT pathways

Background: Pediatric asthma is an usual disease and a kind of fearful health threat for chil-dren. Airway smooth muscle cells (ASMCs) with increased cell proliferation and migration abil-ities serve as important features in the progression of asthma. RAB11A has been shown to aggravate cancer progression and is closely associated with inflammation. Gene analysis dis-covered that RAB11A exhibited higher expression in asthmatic patients. However, the detailed regulatory function of RAB11A in asthma still needs further investigation.Method: The mRNA and protein expressions of genes were examined through RT-qPCR and western blot. Cell proliferation was examined through MTT and BrdU assays. Cell apoptosis was tested through flow cytometry. The cell migration ability was detected through wound healing and transwell assays. The levels of TNF-α, I L-1β, IL-8, and IL-6 were measured through ELISA.Result: In this study, the mRNA and protein expressions of RAB11A were increased with PDGF-BB treatment in a dose-dependent manner. Additionally, the silencing of RAB11A sup-pressed the proliferation ability of PDGF-BB-mediated ASMCs. Moreover, it was uncovered that the knockdown of RAB11A inhibited the migration ability of PDGF-BB-stimulated ASMCs. Besides, suppression of RAB11A relieved the inflammatory response in PDGF-BB-stimulated ASMCs. Lastly, inhibition of RAB11A retarded the NF-κB and PI3K/AKT pathways.Conclusion: Our results revealed that RAB11A aggravated PDGF-BB-stimulated proliferation, migration, and inflammation of ASMCs through modulating NF-κB and PI3K/AKT signaling path-ways. This finding implied that the RAB11A may be deemed as a novel and prospective bio-marker for asthma treatment (AU)

Differential Expression of lncRNA CASC2 in the Serum of Childhood Asthma and Its Role in Airway Smooth Muscle Cells Proliferation and Migration.

PURPOSE: Asthma is a common chronic disease in children. Abnormal expression of lncRNAs can be used as biomarkers for early diagnosis of asthma. The present study aimed to explore the expression change and clinical value of lncRNA CASC2 in asthma, and further investigate its potential mechanism. PATIENTS AND METHODS: Seventy asthma children and 66 healthy controls were recruited. Levels of mRNAs were detected using qRT-PCR. Receiver operating characteristic (ROC) curves were drawn for diagnostic value evaluation. Asthma cell models were established using PDGF-BB in Human airway smooth muscle cells (ASMCs). Levels of Th1/Th2 related cytokines were detected using ELISA. Lipofectamine 3000 was used for cell transfection. The target relationship was verified using luciferase activity assay. RESULTS: CASC2 was at a low level in asthma children in comparison with the healthy controls. Serum CASC2 can distinguish healthy individuals from asthma children. Overexpression of CASC2 inhibited PDGF-BB induced cell proliferation and migration. CASC2 upregulation inhibited the release of Th2 related cytokines (IL-4 and IL-10), but promoted the release of Th1-related cytokine (IFN-γ). In PDGF-BB treated ASMCs, the reduced expression of contractile phenotype marker (α-SMA) was detected, but the trend was reversed by CASC2 upregulation. LncRNA CASC2 serves as a ceRNA of miR-31-5p, overexpression of miR-31-5p reversed the influence of CASC2 on asthma in vitro. CONCLUSION: Serum CASC2 can distinguish healthy individuals from asthma children. CASC2 may be involved in childhood asthma through inhibiting ASMCs proliferation, migration and inflammation via sponging miR-31-5p.

MiR-20b-5p contributes to the dysfunction of vascular smooth muscle cells by targeting MAGI3 in hypertension.

INTRODUCTION: MicroRNAs (miRNAs), have been frequently reported to regulate various diseases including hypertension. However, the biological role and regulatory mechanism of miR-20b-5p are unclear in hypertension. The current study aimed to investigate the role of miR-20b-5p in hypertension. METHODS: Bioinformatics analysis (starBase: http://starbase.sysu.edu.cn ) and a wide range of experiments including blood pressure detection, morphometric sampling by electron microscopy, real-time quantitative polymerase chain reaction (RT-qPCR), CCK-8, western blot, luciferase reporter, hematoxylin and eosin (H&E) staining and Masson trichrome staining assays were used to explore the function and mechanism of miR-20b-5p in hypertension. RESULTS: MiR-20b-5p level was significantly upregulated in Spontaneously hypertensive rats' (SHRs') thoracic aortic vascular tissues. In function, miR-20b-5p silencing inhibited the proliferation and migration of aortic smooth muscle cells (ASMCs) of SHRs. In mechanism, we predicted 10 potential target mRNAs for miR-20b-5p. After prediction by bioinformatics, MAGI3 was validated to bind with miR-20b-5p. Rescue assays showed that MAGI3 silencing reversed the inhibitive influence of miR-20b-5p depletion on cell proliferation and migration. CONCLUSIONS: MiR-20b-5p contributed to the dysfunction of ASMCs by targeting MAGI3 in hypertension. This new discovery provided a potential novel insight for hypertension treatment.

Scopoletin inhibits PDGF-BB-induced proliferation and migration of airway smooth muscle cells by regulating NF-kB signaling pathway

Background: Asthma is a common chronic inflammatory disease of the airway, and airway remodeling and the proliferation mechanism of airway smooth muscle cells (ASMCs) is of great significance to combat this disease.Objective: To assess possible effects of scopoletin on asthma and the potential signaling pathway.Materials and methods: ASMCs were treated PDGF-BB and scopoletin and subjected to cell viability detection by CCK-8 assay. Cell migration of ASMCs was determined by a wound closure assay and transwell assay. The protein level of MMP2, MMP9, calponin and α-SMA were measured using western blot. The levels of NF-κB signaling pathway were detected by Western blotting.Results: Scopoletin inhibited proliferation of PDGF-BB - induced ASMCs. Also it suppressed the migration and invasion of PDGF-BB - induced ASMCs. We further showed that Scopoletin regulated phenotypic transition of ASMCs. Mechanically, Scopoletin inhibited proliferation and invasion of ASMCs by regulating NF-κB signaling pathway (AU)

Scopoletin inhibits PDGF-BB-induced proliferation and migration of airway smooth muscle cells by regulating NF-κB signaling pathway.

BACKGROUND: Asthma is a common chronic inflammatory disease of the airway, and airway remodeling and the proliferation mechanism of airway smooth muscle cells (ASMCs) is of great significance to combat this disease. OBJECTIVE: To assess possible effects of scopoletin on asthma and the potential signaling pathway. MATERIALS AND METHODS: ASMCs were treated PDGF-BB and scopoletin and subjected to cell viability detection by CCK-8 assay. Cell migration of ASMCs was determined by a wound closure assay and transwell assay. The protein level of MMP2, MMP9, calponin and α-SMA were measured using western blot. The levels of NF-κB signaling pathway were detected by Western blotting. RESULTS: Scopoletin inhibited proliferation of PDGF-BB - induced ASMCs. Also it suppressed the migration and invasion of PDGF-BB - induced ASMCs. We further showed that Scopoletin regulated phenotypic transition of ASMCs. Mechanically, Scopoletin inhibited proliferation and invasion of ASMCs by regulating NF-κB signaling pathway. CONCLUSIONS: We therefore thought Scopoletin could serve as a promising drug for the treatment of asthma.

LncRNA RP5-857K21.7 inhibits PDGF-BB-induced proliferation and migration of airway smooth muscle cells through the miR-508-3p/PI3K/AKT/mTOR axis.

The continuous increase in the prevalence of asthma poses a threat to human health. Despites numerous researches, the understanding of asthma development still remain elusive, hindering the development of effective treatment. Here, we explored the role of lncRNA RP5-857K21.7 (RP5-857K21.7) in the development of asthma and its potential molecular mechanism of regulation. Airway smooth muscle cells (ASMCs) were isolated and cultured after which some of the cells were induced with PDGF-BB to build an asthma cell model, and then, qRT-PCR analysis was used to measure the expression level of RP5-857K21.7 in the cell model. Result shows that the RP5-857K21.7 is significantly downregulated in PDGF-BB-induced ASMCs cells. Through CCK-8, transwell, and flow cytometry assay, we examined the functional impact of RP5-857K21.7 on the proliferation, migration, and apoptosis of the ASMCs, respectively, and found that the overexpression of RP5-857K21.7 markedly inhibit PDGF-BB-induced ASMCs cell proliferation, migration and induce apoptosis. Bioinformatics analysis predicted that the RP5-857K21.7 could sponge miR-508-3p and result was validated through a dual-luciferase reporter assay, biotinylated RNA pull-down assay, and RIP-qRT-PCR analysis. Mechanistically, RP5-857K21.7 regulates the PI3K/AKT/mTOR pathway by endogenously sponging miR-508-3p to inhibit PDGF-BB-induced ASMCs cell proliferation, migration and induce apoptosis. The current research suggests that the RP5-857K21.7 and its associated molecular pathway (miR-508-3p/PI3K/AKT/mTOR axis) might be a useful therapeutic target for the treatment of asthma disease.

Trehalose inhibits proliferation while activates apoptosis and autophagy in rat airway smooth muscle cells.

Trehalose is a disaccharide with multiple important biological activities. In many cell types, Trehalose regulates the physiological behaviors of proliferation, apoptosis and autophagy. But the effects of trehalose on ASMCs have never been reported. Here, we showed that trehalose activated autophagy of ASMCs at low dose, inhibited proliferation and induced apoptosis of ASMCs at high dose. Further study, we found the cell cycle was arrested in S and G2\M phases, the expression of CyclinA1 and CyclinB1 decreased. Then, we investigated the ratio of Bcl-2/Bax was drastically reduced. Next, we detected an important transcription factor TFEB, which is closely related to autophagy. We found TFEB was highly activated with trehalose treatment. And many downstream autophagy-related genes of TFEB were also up-regulated. In summary, trehalose plays an important role on the regulation of proliferation, apoptosis and autophagy of ASMCs.