Overexpression of MiR-29b-3p Inhibits Atrial Remodeling in Rats by Targeting PDGF-B Signaling Pathway

Purpose. Studies have found that microRNAs (miRNAs) are closely associated with atrial fibrillation, but their specific mechanism remains unclear. The purpose of this experiment is to explore the function of miR-29b-3p in regulating atrial remodeling by targeting PDGF-B signaling pathway and thereby also explore the potential mechanisms. Methods. We randomly divided twenty-four rats into four groups. Caudal intravenous injections of angiotensin-II (Ang-II) were administered to establish atrial fibrosis models. Expressions of miR-29b-3p and PDGF-B were then tested via RT-PCR, western blot, and immunohistochemistry. Binding sites were then analyzed via the bioinformatics online software TargetScan and verified by Luciferase Reporter. We used Masson staining to detect the degree of atrial fibrosis, while immunofluorescence and western blot were used to detect the expressions of Collagen-I and a-SMA. We used immunohistochemistry and western blot to detect the expression of connexin 43 (Cx43). Results. In comparison with the Ang-II group, miR-29b-3p was seen to lower the degree of atrial fibrosis, decrease the expression of fibrosis markers such as Collagen-I and a-SMA, and increase the protein expression of Cx43. MiR-29b-3p can lower the expression of PDGF-B, while the Luciferase Reporter showed that PDGF-B is the verified target gene of miR-29b-3p. Conclusions. MiR-29b-3p was able to reduce atrial structural and electrical remodeling in the study’s rat fibrosis model. This biological function may be expressed through the targeted regulation of the PDGF-B signaling pathway.

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Profiling of miR-205/P4HA3 Following Angiotensin II-Induced Atrial Fibrosis: Implications for Atrial Fibrillation

Frontiers in Cardiovascular Medicine ◽

10.3389/fcvm.2021.609300 ◽

2021 ◽

Vol 8 ◽

Author(s):

Zezhou Xiao ◽

Desai Pavan Kumar Reddy ◽

Chuqing Xue ◽

Ximao Liu ◽

Xiong Chen ◽

...

Keyword(s):

Angiotensin Ii ◽

Western Blot ◽

Signaling Pathway ◽

Collagen I ◽

Luciferase Reporter ◽

Ang Ii ◽

Atrial Fibrosis ◽

Jnk Signaling ◽

Qrt Pcr ◽

Jnk Signaling Pathway

Objective: Atrial fibroblasts are the main component of atrial fibrosis. Data in previous studies proved the implication of miRNAs in AF progression and the association of miR-205 with cancer associated-fibroblasts, while no evidence supported the implication of miR-205 in atrial fibrosis. Therefore, this study aims to explore the effect and mechanism of miR-205/P4HA3 axis on atrial fibrosis.Methods: Angiotensin II (Ang II) was used to induce atrial fibrosis model in rats, which was verified by H&E staining and Masson staining. qRT-PCR and Western blot were applied to measure the expressions of miR-205, P4HA3, collagen I, and α-SMA. The rat atrial fibroblasts were isolated and then subjected to Ang II treatment or cell transfection for determination of cell biological functions using CCK-8, BrdU assay, TUNEL staining, and cell scratch assay. qRT-PCR and Western blot was applied to analyze the expressions of miR-205, P4HA3, collagen I, α-SMA, JNK, and p-JNK in atrial fibroblasts. Dual-luciferase reporter gene assay and RNA immune-precipitation experiment was employed to verify the binding relationship between miR-205 and P4HA3.Results: Ang II induced rats had disordered arrangement of atrial muscles with uneven nuclear sizes and necrotic atrial myocytes, and increased collagen deposition, in which elevated expressions of P4HA3, collagen I, and α-SMA as well as suppressed expression level of miR-205 were found. In vitro, Ang II treatment in atrial fibroblasts with overexpression of P4HA3 facilitated cellular migration and proliferation, with the induction of JNK signaling pathway. However, these trends were reversed after transfection with miR-205 mimic. P4HA3 is a target gene of miR-205.Conclusion: The miR-205/P4HA3 axis is implicated in atrial fibrosis by inhibition of rat fibroblast proliferation and migration and the inactivation of JNK signaling pathway.

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Periplaneta Americana Extract may Attenuate 2,4,6-Trinitrobenzenesulfonic Acid-Induced Intestinal Fibrosis by Inhibiting TGF-β/Smad Signaling Pathway

10.21203/rs.3.rs-174235/v1 ◽

2021 ◽

Author(s):

Jing Liu ◽

Pin Lv ◽

Xiang Rao ◽

Jiajia Wang

Keyword(s):

Western Blot ◽

Signaling Pathway ◽

Real Time Pcr ◽

Periplaneta Americana ◽

Collagen I ◽

Smad Signaling ◽

Intestinal Fibrosis ◽

Smad Signaling Pathway

Abstract PurposeIntestinal fibrosis is an incurable digestive disease accompanied by stricture formation, and it has an increasing incidence in recent years. Periplaneta americana is one of the medicinal insects with a long history. There are few reports on the effect of intestinal fibrosis. This study aims to evaluate the inhibitory effect of PA treatment on intestinal fibrosis. MethodsTNBS was used to establish intestinal fibrosis model by enema in BALB/c mice. The mice were treated with PA (50, 100, 200 mg/kg body weight) and 5-aminosalicylic acid (5-ASA) (40mg/kg) by gavage once a day for 6 weeks. At the end of the last week, the mice were sacrificed. Colon samples were collected for H&E and Masson staining. The mRNA and protein expression of α-smooth muscle actin (α-SMA), collagen I and the transforming growth factor-β (TGF-β) / Smad signaling pathway were conducted by real-time PCR and western blot analysis. In vitro, TGF-β1 was used to induce intestinal fibrosis at human colon fibroblasts (CCD-18Co). And using real-time PCR and western blot methods to detect the expression of α-SMA and collagen I. ResultsPA inhibited the expression of α-SMA and collagen I in vivo and in vitro. But the difference was that PA inhibited the TGF-β/Smad signaling pathway in vivo, and the same results had not been obtained in vitro. Conclusion: PA may attenuate intestinal fibrosis by inhibiting TGF-β/Smad signaling pathway, but more experiments were needed to prove it in vitro. ConclusionsPA has potential pharmacological effects in inhibiting intestinal fibrosis, and the TGF-β/Smad signaling pathway seemed promising.

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P936Atrial remodeling in rat hearts after exposure to high-intensity infrasound

EP Europace ◽

10.1093/europace/euaa162.014 ◽

2020 ◽

Vol 22 (Supplement_1) ◽

Author(s):

A Lousinha ◽

G Pereira ◽

G Borrecho ◽

J Brito ◽

A Oliveira De Carvalho ◽

...

Keyword(s):

High Intensity ◽

Connexin 43 ◽

Noise Exposure ◽

Experimental Studies ◽

Blue Staining ◽

P Value ◽

Atrial Remodeling ◽

Atrial Fibrosis ◽

Rat Hearts ◽

Arrhythmogenic Substrate

Abstract Background Noise is an important environmental risk factor. Industrial environments are rich in high-intensity infrasound (hi-IFS), which we have found to induce myocardial and coronary perivascular fibrosis in rats. Recently, a significant association between noise exposure and the incidence of atrial fibrillation (AF) was found in large cohort studies but the pathophysiology is unclear. Atrial fibrosis remains the cornerstone of atrial pathology in AF. Purpose: We hypothesized that rats exposed to hi-IFS develop atrial remodeling involving fibrosis and connexin 43, which we sought to evaluate. Material and Methods: Seventy-two Wistar rats, half exposed to hi-IFS (120dB, <20Hz) during a maximum period of 12 weeks and half age-matched controls, were studied. Atrial fibrosis was analyzed by Chromotrope-aniline blue staining. The immunohistochemical evaluation of Cx43 was performed using the polyclonal antibody connexin-43m diluted 1:1000 at 4ºC overnight. Digitized images were obtained with an optical microscope using 400× magnifications. The measurements were performed using image J software. A two-way ANOVA model was used to compare the groups. Results: The mean values of the ratio "atrial fibrosis / cardiomyocytes" increased to a maximum of 0,1095 ± 0,04 and 0,5408 ± 0,01, and of the ratio "CX43 / cardiomyocytes" decreased to 0,0834 ± 0,03 and 0,0966 ± 0,03, respectively in IFS-exposed rats and controls. IFS-exposed rats exhibited a significantly higher ratio of fibrosis (p < 0,001) and lower ratio of Cx43 (p = 0,009). Conclusion: High-intensity infrasound exposure triggers atrial remodeling in rat hearts. Whether this finding correlates to arrhythmogenic substrate in noise induced-AF is not known and reinforces the need for further experimental studies. Table 1 Time of exposure (weeks) Group IFS (n = 36) Group CTL (n = 36) P value Ratio of atrial fibrosis / cardiomyocytesMean ± SD 1 0,0896 ± 0,04 0,0460 ± 0,03 0,007 6 0,0936 ± 0,03 0,0491 ± 0,01 0,001 12 0,1095 ± 0,04 0,0541 ± 0,01 0,001 Ratio of atrial CX43 / cardiomyocytesMean ± SD 1 0,1100 ± 0,03 0,1371 ± 0,03 0,047 6 0,0829 ± 0,04 0,1036 ± 0,03 0,170 12 0,0834 ± 0,03 0,0966 ± 0,03 0,259 Mean ± SD of the two measured outcomes in the two groups. IFS – Infrasound; CTL – Control; SD – standard deviation; Cx43 – Connexin 43 Abstract Figure. Atrial fibrosis and Cx43 /cardiomyocytes

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Cimifugin Suppresses NF-κB Signaling to Prevent Osteoclastogenesis and Periprosthetic Osteolysis

Frontiers in Pharmacology ◽

10.3389/fphar.2021.724256 ◽

2021 ◽

Vol 12 ◽

Author(s):

Juan Duan ◽

Xuantao Hu ◽

Tao Li ◽

Gen Wu ◽

Pengcheng Dou ◽

...

Keyword(s):

Bone Resorption ◽

Western Blot ◽

Signaling Pathway ◽

Osteoclast Differentiation ◽

Mapk Signaling ◽

Luciferase Reporter ◽

Pcr Analysis ◽

Periprosthetic Osteolysis

Background: Aseptic loosening of prosthesis (ALP) is one of the most common long-term complications of knee and hip arthroplasty. Wear particle-induced osteoclastogenesis and subsequent periprosthetic osteolysis account for the morbidity of ALP. Here, we investigate the potential of cimifugin (CIM), a natural extract from Cimicifuga racemosa and Saposhnikovia divaricata, as a bone-protective drug in the treatment of ALP.Method: First, we performed cell viability and osteoclast formation assays to assess the effect of noncytotoxic CIM on osteoclast differentiation in vitro. Bone slice resorption and F-actin ring immunofluorescence assays were adopted to assess the effects of CIM on bone-resorption function. Then, quantitative real-time polymerase chain reaction (qRT–PCR) analysis was performed to further assess the repressive effects of CIM on osteoclastogenesis at the gene expression level. To elucidate the mechanisms underlying the above findings, Western blot and luciferase reporter gene assays were used to assess the regulatory effects of CIM on the NF-κB and MAPK signaling pathways. Moreover, a Ti particle-induced murine calvarial osteolysis model and subsequent histomorphometric analysis via micro-CT and immunohistochemical staining were used to elucidate the effect of CIM on periprosthetic osteolysis in vivo.Result: CIM dose-dependently inhibited both bone marrow-derived macrophage (BMM)- and RAW264.7 cell-derived osteoclastogenesis and bone resorption pit formation in vitro, which was further supported by the reduced expression of F-actin and osteoclast-specific genes. According to the Western blot analysis, inhibition of IκBα phosphorylation in the NF-κB signaling pathway, not the phosphorylation of MAPKs, was responsible for the suppressive effect of CIM on osteoclastogenesis. Animal experiments demonstrated that CIM alleviated Ti particle-induced bone erosion and osteoclast accumulation in murine calvaria.Conclusion: The current study suggested for the first time that CIM can inhibit RANKL-induced osetoclastogenesis by suppressing the NF-κB signaling pathway in vitro and prevent periprosthetic osteolysis in vivo. These findings suggest the potential of CIM as a therapeutic in ALP.

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UBQLN4 is activated by C/EBPβ and exerts oncogenic effects on colorectal cancer via the Wnt/β-catenin signaling pathway

Cell Death Discovery ◽

10.1038/s41420-021-00795-4 ◽

2021 ◽

Vol 7 (1) ◽

Author(s):

Xiaolong Tang ◽

Yahang Liang ◽

Guorui Sun ◽

Qingsi He ◽

Hui Qu ◽

...

Keyword(s):

Colorectal Cancer ◽

Western Blot ◽

Signaling Pathway ◽

Lymphatic Metastasis ◽

Core Promoter ◽

Enrichment Analysis ◽

Gene Set Enrichment Analysis ◽

Luciferase Reporter ◽

Migration And Invasion

AbstractUbiquilin 4 (UBQLN4) is an important member of the ubiquitin-like protein family. An increasing number of studies have shown that UBQLN4 is an important regulator of tumorigenesis. Nevertheless, the biological function and detailed mechanisms of UBQLN4 in colorectal cancer (CRC) development and progression remain unclear. Here, we identified UBQLN4 upregulation in CRC tissues and it is positively associated with CRC size, TNM stage, and lymphatic metastasis. Patients with high UBQLN4 expression had a poor prognosis. Functionally, overexpression of UBQLN4 significantly promoted CRC cell proliferation, migration, and invasion, while UBQLN4 silencing elicited the opposite effect. This result was consistent with the conclusion that UBQLN4 expression correlated positively with the CRC size and lymphatic metastasis. In vivo, UBQLN4 silencing also inhibited tumor growth. Mechanistically, using gene set enrichment analysis (GSEA) and western blot experiments, we identified that UBQLN4 activated the Wnt/β-catenin signaling pathway to upregulate β-catenin and c-Myc expression, thereby promoting CRC proliferation, migration and invasion. A rescue experiment further verified this conclusion. Dual luciferase reporter, real-time quantitative PCR (RT-qPCR), western blot and chromatin immunoprecipitation (ChIP) assays indicated that the transcription factor CCAAT/enhancer-binding protein beta (C/EBPβ) directly bound to the UBQLN4 core promoter region and activated its transcription, upregulating β-catenin and c-Myc expression to promote CRC progression. Thus, our findings suggest that UBQLN4 is a key oncogene in CRC and may be a promising target for the diagnosis and treatment of patients with CRC.

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Regulation of NSCLC Cell Proliferation By MARCH7 Via the NF-κB and Wnt/β-Catenin Signaling Pathways

10.21203/rs.3.rs-1212966/v1 ◽

2022 ◽

Author(s):

Xiaomi Lu ◽

Lili Shao ◽

Ye Qian ◽

Sixun Zhong ◽

Jinhong Chen ◽

...

Keyword(s):

Western Blot ◽

Signaling Pathway ◽

Reporter Gene ◽

Nude Mice ◽

A549 Cells ◽

Luciferase Reporter ◽

Clinicopathological Parameters ◽

G1 Phase ◽

Gene Assay ◽

Nsclc Patients

Abstract The aim of the study was to explore the role of the E3 ubiquitin ligase MARCH7 in the development of non-small-cell lung cancer (NSCLC) and to explore the underlying molecular mechanism.Western blot and immunohistochemistry results showed that the expression of MARCH7 in NSCLC cancer tissues was higher than that in paracancerous tissues. Tissue microarray staining results and clinicopathological parameters of NSCLC patients revealed that MARCH7 expression was closely related to TNM stage, degree of tumor differentiation and lymph node metastasis of NSCLC patients. Furthermore, univariate and multivariate analyses and survival curve analysis showed that high expression of MARCH7 was associated with poor prognosis.In vitro, siRNA was constructed and transfected into A549 cells to inhibit the expression of MARCH7. The CCK-8 assay indicated that the growth rate of tumor cells in the interference group was reduced. The number of colonies and cells in the interference group decreased in the plate clone formation experiment. Flow cytometry showed that G0/G1 phase cells were predominantly increased after blocking endogenous MARCH7 expression, and G0/G1 phase arrest occurred in A549 cells. The reporter gene activity of the NF-κB signaling pathway and Wnt/β-catenin signaling pathway was reduced, as validated by a double luciferase reporter gene assay. Western blot analysis showed that the expression of NF-κB P50, NF-κB P65 and β-catenin was decreased, while the expression of E-cadherin was elevated.In vivo, MARCH7-overexpressing virus was constructed and transfected into A549 cells and then subcutaneously injected into nude mice. It was demonstrated that the tumor volume was significantly larger in the MARCH7 overexpression group than in the control nude mice during the same period. Elevated expression of PCNA and Ki-67 was observed in the tumor mass of the MARCH7 overexpression group, as measured by immunohistochemical analysis, accompanied by enhanced levels of NF-κB P50, NF-κB P65 and β-catenin, as detected by Western blot. These results provide a new idea for the experimental basis for the treatment of NSCLC in the future.

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circRNA LDLRAD3 Enhances the Malignant Behaviors of NSCLC Cells via the miR-20a-5p-SLC7A5 Axis Activating the mTORC1 Signaling Pathway

Journal of Healthcare Engineering ◽

10.1155/2022/2373580 ◽

2022 ◽

Vol 2022 ◽

pp. 1-10

Author(s):

Yu Li ◽

Guangle Qin ◽

Jinyun Du ◽

Peng Yue ◽

Yanling Zhang ◽

...

Keyword(s):

Western Blot ◽

Signaling Pathway ◽

Circular Rna ◽

Luciferase Reporter ◽

Migration And Invasion ◽

Potential Therapeutic Target ◽

Expression Levels ◽

Mtorc1 Signaling ◽

Mtorc1 Pathway ◽

Related Proteins

Circular RNA LDLRAD3 behaved as an oncogene in several malignancies, but its effects in NSCLC and the involvement of downstream molecules and activation of signaling pathways had not been fully reported. We planned to explore how LDLRAD3 facilitated the malignancy of NSCLC. QRT-PCR was performed to evaluate the expression levels of LDLRAD3, miR-20a-5p, and SLC7A5 in NSCLC tissues and cells. si-LDLRAD3 was transfected to A549 and H1299 cells to knock down intrinsic LDLRAD3 to determine its oncogenic roles. CCK-8 assay and transwell assay were executed to assess cell proliferative, migrative, and invasive abilities. Dual-luciferase reporter (DLR) assay was manipulated to verify the ENCORI-predicted relationships between LDLRAD3 and miR-20a-5p and between miR-20a-5p and SLC7A5. Western blot, immunofluorescent assay, and immunohistochemistry were applied to explore the expression levels of SLC7A5, and the levels of mTORC1 pathway-related proteins were evaluated using western blot. Rescue experiments were conducted by transfecting si-LDLRAD3, miR-20a-5p inhibitor, and si-SLC7A5 to explore the influence of the LDLRAD3-miR-20a-5p-SLC7A5 axis on the malignant behaviors of NSCLC cells. The expression levels of LDLRAD3 and SLC7A5 were boosted, whereas miR-20a-5p was impeded in NSCLC tissues and cell lines. Knockdown of LDLRAD3 weakened the proliferation, migration, and invasion of A549 and H1299 cells. LDLRAD3 was verified to sponge miR-20a-5p and miR-20a-5p targeted SLC7A5. LDLRAD3 activated the mTORC1 singling pathway via the miR-20a-5p-SLC7A5 axis to strengthen the malignant properties of A549 and H1299 cells. We concluded that LDLRAD3 exerted oncogenic effects via the miR-20a-5p-SLC7A5 axis to activate the mTORC1 signaling pathway in NSCLC. Our findings enlightened that LDLRAD3 could become a potential therapeutic target in the treatment and management of NSCLC.

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β-catenin/LEF-1 Transcription Complex is Responsible for the Transcriptional Activation of LINC01278

10.21203/rs.3.rs-113487/v1 ◽

2020 ◽

Author(s):

Shaojian Lin ◽

Weiwei Zhang ◽

Ziwen Shi ◽

Langping Tan ◽

Yue Zhu ◽

...

Keyword(s):

Western Blot ◽

Signaling Pathway ◽

Transcriptional Activation ◽

Feedback Regulation ◽

Luciferase Reporter ◽

Regulation Mechanism ◽

Putative Promoter ◽

Negative Feedback Regulation ◽

Pathway Activation ◽

Rna Immunoprecipitation

Abstract Background: Our previous study shows that LINC01278 inhibits the development of papillary thyroid carcinoma (PTC) by regulating miR-376c-3p/DNM3 axis. However, the regulation mechanism of LINC01278 expression in PTC cells is still unclear. Methods: The luciferase reporter and ChIP assays were used to confirme the binding of LEF-1 to the putative promoter site of LINC01278. The RNA immunoprecipitation was used the enrichment of LINC01278 in β-catenin protein. Western blot was used to detected the expression of target proteins. Results: Firstly, the online PROMO algorithm determined a putative LEF-1 binding site on LINC01278 promoter. Then, the luciferase reporter and ChIP assays confirmed the binding of LEF-1 to the putative promoter site of LINC01278. Furthermore, the overexpression of β-catenin increased the binding of LEF-1 to the LINC01278 promoter, and the knockdown or overexpression of LEF-1 or β-catenin can affect the expression level of LINC01278. In addition, RNA immunoprecipitation showed that LINC01278 was enriched in β-catenin protein. RNA pulldown and western blot also confirmed that LINC01278 precipitated β-catenin in TPC-1 and BCPAP cells. Furthermore, the knockdown or overexpression of LINC01278 significantly affected the expression of β-catenin and targets of Wnt/β-catenin signaling pathway (CCND2, CyclinD1, MYC, and SOX4). Conclusion: In summary, we found the transcriptional activation of LINC01278 by the β-catenin/LEF-1 transcription factor, and the negative feedback regulation of LINC01278 on Wnt/β-catenin signaling pathway activation.

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Immunoproteasome Subunit β5i Promotes Ang II (Angiotensin II)–Induced Atrial Fibrillation by Targeting ATRAP (Ang II Type I Receptor–Associated Protein) Degradation in Mice

Hypertension ◽

10.1161/hypertensionaha.118.11813 ◽

2019 ◽

Vol 73 (1) ◽

pp. 92-101 ◽

Cited By ~ 16

Author(s):

Jing Li ◽

Shuai Wang ◽

Yun-Long Zhang ◽

Jie Bai ◽

Qiu-Yue Lin ◽

...

Keyword(s):

Atrial Fibrillation ◽

Angiotensin Ii ◽

Connexin 43 ◽

Transforming Growth Factor ◽

Negative Regulator ◽

Atrial Remodeling ◽

Type I ◽

Ang Ii ◽

Adeno Associated Virus ◽

Virus Serotype

Atrial fibrillation (AF) is the most common type of cardiac arrhythmia and increases the risk of stroke, heart failure, and death. Ang II (angiotensin II) triggers AF, mainly through stimulation of the AT1R (Ang II type I receptor). The immunoproteasome is a highly efficient proteolytic machine derived from the constitutive proteasome, but the role it plays in regulating AT1R activation and triggering AF remains unknown. Here, we show that among the catalytic subunits, β5i (PSMB8) expression, and chymotrypsin-like activity were the most significantly upregulated in atrial tissue of Ang II–infused mice or serum from patients with AF. β5i KO (β5i knockout) in mice markedly attenuated Ang II-induced AF incidence, atrial fibrosis, inflammatory response, and oxidative stress compared with WT (wild type) animals, but injection with recombinant adeno-associated virus serotype 9–β5i increased these effects. Moreover, we found that ATRAP (AT1R-associated protein) was a target of β5i. Overexpression of ATRAP significantly attenuated Ang II-induced atrial remodeling and AF in recombinant adeno-associated virus serotype 9–β5i-injected mice. Mechanistically, Ang II upregulated β5i expression to promote ATRAP degradation, which resulted in activation of AT1R-mediated NF-κB signaling, increased NADPH oxidase activity, increased TGF (transforming growth factor)-β1/Smad signaling, and altered the expression of Kir2.1 and CX43 (connexin 43) in the atria, thereby affecting atrial remodeling and AF. In summary, this study identifies β5i as a negative regulator of ATRAP stability that contributes to AT1R activation and to AF, highlighting that targeting β5i activity may represent a potential therapeutic approach for the treatment of hypertensive AF.

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Overexpression of miR-27b-3p Targeting Wnt3a Regulates the Signaling Pathway of Wnt/β-Catenin and Attenuates Atrial Fibrosis in Rats with Atrial Fibrillation

Oxidative Medicine and Cellular Longevity ◽

10.1155/2019/5703764 ◽

2019 ◽

Vol 2019 ◽

pp. 1-13 ◽

Cited By ~ 3

Author(s):

Xiangwei Lv ◽

Jinyi Li ◽

Yisen Hu ◽

Shirong Wang ◽

Chengye Yang ◽

...

Keyword(s):

Atrial Fibrillation ◽

Signaling Pathway ◽

Collagen I ◽

Immunohistochemical Method ◽

Atrial Fibrosis ◽

Cx43 Expression ◽

Adeno Associated Virus ◽

Tail Vein ◽

Tail Vein Injection ◽

Collagen Iii

MicroRNAs (miRNAs) are regarded as a potential method for the treatment of atrial fibrillation (AF) although its molecular mechanism remains unknown. We found in our previous study that the level of peripheral blood miR-27b-3p and the expression of atrial tissue CX43 were both significantly downregulated in AF patients. In the present study, we propose and test this hypothesis that overexpression of miR-27b-3p attenuates atrial fibrosis, increases CX43 expression, and regulates the signaling pathway of Wnt/β-Catenin by targeting Wnt3a. miR-27b-3p overexpression was induced by rat tail vein injection of adeno-associated virus. Two weeks after transfection of adeno-associated virus, the rat AF model was established by tail vein injection of acetylcholine- (ACh-) CaCl2 for 7 days, and 1 ml/kg was injected daily. The incidence and duration of AF were recorded with an electrocardiogram. Cardiac function was monitored by cardiac ultrasound. Serum cardiac enzyme was detected by ELISA. The expression of atrial miR-27b-3 and Wnt3a was assayed by quantitative RT-PCR. Atrial fibrosis was determined by Masson’s trichrome staining. Expression of atrial Collagen-I and Collagen-III was tested by the immunohistochemical method. Expression of CX43 was measured by immunofluorescence. The expression of Collagen-I, a-SMA, Collagen-III, TGF-β1, CX43, Wnt3a, β-Catenin, and p-β-Catenin was assayed by western blot. Our results showed that miR-27b-3p overexpression could reduce the incidence and duration of AF, alleviate atrial fibrosis, increase atrial CX43 expression, and decrease the expression of Collagen-I, a-SMA, Collagen-III, TGF-β1, Wnt3a, and p-β-Catenin. In addition, the results of luciferase activity assay showed that Wnt3a is a validated miR-27b-3p target in HEK 293T cells. Our results provide a new evidence that miR-27b-3p regulates the signaling pathway of Wnt/β-Catenin by targeting Wnt3a, which may play an important role in the development of atrial fibrosis and AF.

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