scholarly journals Locally Produced IGF-1 Promotes Hypertrophy of the Ligamentum Flavum via the mTORC1 Signaling Pathway

2018 ◽  
Vol 48 (1) ◽  
pp. 293-303 ◽  
Author(s):  
Bin Yan ◽  
Minjun Huang ◽  
Canjun Zeng ◽  
Na Yao ◽  
Jie Zhang ◽  
...  
Keyword(s):  
Protein Expression ◽  
Signaling Pathway ◽  
Ligamentum Flavum ◽  
The Elderly ◽  
Collagen I ◽  
Second Primary ◽  
High Morbidity ◽  
Collagen Iii ◽  
Mtorc1 Signaling ◽  
Lumbar Spinal

Background/Aims: Narrowing of the lumbar spinal canal is a condition called lumbar spinal stenosis (LSS) and is a high-morbidity problem in the elderly. LSS is commonly caused by hypertrophy of the ligamentum flavum (HLF). Previous studies showed that fibrosis of the ligamentum flavum (LF) largely contributed to HLF. However, the underlying pathomechanism remains unclear. Insulin-like growth factor-1 (IGF-1) is known to have an intimate relationship with fibrosis in various tissues. Nevertheless, currently, there are few studies regarding IGF-1 in HLF. In this study, we investigated the role of IGF-1 in HLF and its potential molecular mechanism of action. Methods: First, the IGF-1, phosphorylation of IGF-1 receptor (pIGF-1R), phosphorylation of AKT (pAKT), phosphorylation of S6(pS6), collagen I and collagen III expression levels were examined via immunohistochemistry and Western blotting in LF tissues from patients with LSS or Non-LSS. Second, primary LF cells were isolated from adults with a normal LF thickness and were cultured with different concentrations of IGF-1 with or without NVP-AEW541/rapamycin. Results: The results showed that IGF-1, pIGF-1R, pAKT, pS6, collagen I and collagen III protein expression in the LSS group was significantly higher than that in the Non-LSS group. Meanwhile, pIGF-1R, pAKT, pS6, collagen I and collagen III protein expression was significantly enhanced in LF cells after IGF-1 exposure, which can be notably blocked by NVP-AEW541. In addition, pS6, collagen I and collagen III protein expression was blocked by rapamycin. Conclusions: Enhanced IGF-1 promotes the synthesis of collagen I and collagen III via the mTORC1 signaling pathway, which eventually contributes to hypertrophy of the ligamentum flavum.

scholarly journals Hypertrophy of Ligamentum Flavum in Lumbar Spine Stenosis Is Associated with Increased miR-155 Level

2014 ◽  
Vol 2014 ◽  
pp. 1-8 ◽  
Author(s):  
Jianwei Chen ◽  
Zude Liu ◽  
Guibin Zhong ◽  
Lie Qian ◽  
Zhanchun Li ◽  
...  
Keyword(s):  
Protein Expression ◽  
Type I Collagen ◽  
Ligamentum Flavum ◽  
Lumbar Disc ◽  
Collagen I ◽  
Type Iii Collagen ◽  
Type I ◽  
Collagen Iii ◽  
Mrna And Protein Expression ◽  
Collagen Level

Hypertrophy of ligamentum flavum (LF) contributes to lumbar spinal stenosis (LSS) and is caused mainly by fibrosis. Recent data indicate that miR-155 plays a crucial role in the pathogenesis of different fibrotic diseases. This study aimed to test the hypothesis that miR-155 exerts effects on LF thickness by regulating collagen expression. We found that LF thickness and the expression of collagen I and, collagen III were higher in LF from LSS patients than in LF from lumbar disc herniation (LDH) patients (P<0.01). The expression of miR-155 was significantly higher in LF from LSS group than in LF from LDH group (P<0.01). miR-155 level was positively correlated with LF thickness (r=0.958,P<0.01), type I collagen level (r=0.825,P<0.01), and type III collagen level (r=0.827,P<0.01). miR-155 mimic increased mRNA and protein expression of collagen I and collagen III in fibroblasts isolated from LF, while miR-155 sponge decreased mRNA and protein expression of collagen I and III in fibroblasts. In conclusions, miR-155 is a fibrosis-associated miRNA and may play important role in the pathogenesis of LF hypertrophy.

scholarly journals Mechanical Stress-Induced IGF-1 Facilitates col-I and col-III Synthesis via the IGF-1R/AKT/mTORC1 Signaling Pathway

2021 ◽  
Vol 2021 ◽  
pp. 1-11
Author(s):  
Bin Yan ◽  
Canjun Zeng ◽  
Yuhui Chen ◽  
Minjun Huang ◽  
Na Yao ◽  
...  
Keyword(s):  
Mechanical Stress ◽  
Signaling Pathway ◽  
Ligamentum Flavum ◽  
Neutralizing Antibody ◽  
Underlying Mechanism ◽  
Mrna Levels ◽  
Stress Exposure ◽  
Collagen Iii ◽  
Mtorc1 Signaling ◽  
The Relationship

Mechanical stress promotes human ligamentum flavum cells (LFCs) to synthesize multitype collagens, leading to ligamentum flavum hypertrophy (LFH). However, the mechanism of mechanical stress in the formation of collagen remains unclear. Therefore, we investigated the relationship between mechanical stress and collagen synthesis in the present study. First, LFCs were isolated from 9 patients and cultured with or without mechanical stress exposure for different times. IGF-1, collagen I (col-I), and collagen III (col-III) protein and mRNA levels were then detected via ELISA and qPCR, respectively. Moreover, the activation of pIGF-1R, pAKT, and pS6 was examined by Western blot analysis. To further explore the underlying mechanism, an IGF-1 neutralizing antibody, NVP-AEW541, and rapamycin were used. IGF-1, col-I, and col-III were significantly increased in stressed LFCs compared to nonstressed LFCs. In addition, the activation of pIGF-1R, pAKT, and pS6 was obviously enhanced in stressed LFCs. Interestingly, col-I protein, col-I mRNA, col-III protein, col-III mRNA, and IGF-1 protein, but not IGF-1 mRNA, were inhibited by IGF-1 neutralizing antibody. In addition, col-I and col-III protein and mRNA, but not IGF-1, were inhibited by both NVP-AEW541 and rapamycin. Moreover, the activation of pIGF-1R, pAKT, and pS6 was reduced by the IGF-1 neutralizing antibody and NVP-AEW541, and the activation of pS6 was reduced by rapamycin. In summary, these results suggested that mechanical stress promotes LFCs to produce IGF-1, which facilitates col-I and col-III synthesis via the IGF-1R/AKT/mTORC1 signaling pathway.

scholarly journals MiRNA-1202 promotes the TGF-β1-induced proliferation, differentiation and collagen production of cardiac fibroblasts by targeting nNOS

PLoS ONE ◽  
2021 ◽  
Vol 16 (8) ◽  
pp. e0256066
Author(s):  
Jingwen Xiao ◽  
Yan Zhang ◽  
Yuan Tang ◽  
Hengfen Dai ◽  
Yu OuYang ◽  
...  
Keyword(s):  
Protein Expression ◽  
Myocardial Fibrosis ◽  
Cardiac Fibroblasts ◽  
Collagen I ◽  
Luciferase Reporter ◽  
Control Group ◽  
Protein Levels ◽  
Collagen Iii ◽  
Human Cardiac Fibroblasts ◽  
Tgf Β1

Background Atrial fibrillation (AF) is a clinically common arrhythmia that affects human health. Myocardial fibrosis serves as an important contributor to AF. Recently, miRNA-1202 have been reported to be up-regulated in AF. However, the role of miRNA-1202 and its mechanism in myocardial fibrosis remain unclear. Methods Human cardiac fibroblasts (HCFs) were used to construct a fibrosis model by TGF-β1 induction. The expression of miR-1202 was measured by qRT-PCR. Cell proliferation was assessed by CCK-8 assays. Protein expression levels were measured by western blot. Collagen accumulation was measured by ELISA. The relationship between miR-1202 and nNOS was investigated by luciferase reporter assays. Results MiR-1202 expression was obviously increased in HCFs and was both time- and dose-independent. MiR-1202 could increase the proliferation and collagen I, collagen III, and α-SMA levels with or without TGF-β1. MiR-1202 could also increase TGF-β1 and p-Smad2/3 protein levels in comparison to the control group. However, they were obviously decreased after inhibitor transfection. MiR-1202 targets nNOS for negative regulation of HCFs fibrosis by decreasing cell differentiation, collagen deposition and the activity of the TGF-β1/Smad2/3 pathway. Co-transfection of miR-1202 inhibitor and siRNA of nNOS inhibited nNOS protein expression, thereby enhancing the HCFs proliferation. Furthermore, co-transfection of the miR-1202 inhibitor and siRNA of nNOS significantly promoted collagen I, collagen III, TGF-β1, Smad2/3 and α-SMA protein expression and Smad2/3 protein phosphorylation. These findings suggested that miR-1202 promotes HCFs transformation to a pro-fibrotic phenotype by targeting nNOS through activating the TGF-β1/Smad2/3 pathway.

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

2019 ◽  
Vol 2019 ◽  
pp. 1-13 ◽  
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.

scholarly journals Leptin-induced inflammation by activating IL-6 expression contributes to the fibrosis and hypertrophy of ligamentum flavum in lumbar spinal canal stenosis

2018 ◽  
Vol 38 (2) ◽  
Author(s):  
Chao Sun ◽  
Zhen Wang ◽  
Ji-Wei Tian ◽  
Yun-Hao Wang
Keyword(s):  
Spinal Canal ◽  
Ligamentum Flavum ◽  
Spinal Canal Stenosis ◽  
Collagen I ◽  
Inflammatory Factor ◽  
Fibrosis Score ◽  
Lumbar Spinal Canal Stenosis ◽  
Canal Stenosis ◽  
Lumbar Spinal Canal ◽  
Lumbar Spinal

The ongoing chronic inflammation and subsequent fibrosis play an important role in ligamentum flavum (LF) fibrosis and hypertrophy in patients with lumbar spinal canal stenosis (LSCS). Leptin is a chronic inflammatory mediator and involved in the fibrotic process in multiple organ systems. The present study aimed to investigate the role of leptin in LF fibrosis and its related regulatory mechanisms. The LF specimens were obtained during the surgery from 12 patients with LSCS (LSCS group) and 12 control patients with lumbar disc herniation (LDH) group. The morphologic changes and fibrosis score of LF were assessed by Hematoxylin and eosin (H&E) and Masson’s trichrome staining respectively. The location and expression of leptin in LF tissues were determined. Then, the LF cells were cultured and exposed to recombinant human leptin (rhleptin). Collagen I and III were used as fibrosis markers and IL-6 was used as the inflammatory factor. As a result, the LF thickness and fibrosis score in the LSCS group were significantly higher than those in the LDH group (P<0.05). Leptin was detected in the hypertrophied LF and its expression was substantially increased in the LSCS group and positively correlated with LF thickness and fibrosis score (P<0.05). Moreover, our in vitro experiments revealed that rhleptin treated LF cells elevated the expression of collagen I and III. Finally, leptin administration induced IL-6 expression via nuclear factor-κB (NF-κB) pathway in LF cell (P<0.05). Our study demonstrated novel molecular events for leptin-induced inflammation in LF tissue by promoting IL-6 expression and thus might have potential implications for clarifying the mechanism underlying LF fibrosis and hypertrophy.

scholarly journals Citrus Alkaline Extract Delayed the Progression of Pulmonary Fibrosis by Inhibiting p38/NF-κB Signaling Pathway-Induced Cell Apoptosis

2019 ◽  
Vol 2019 ◽  
pp. 1-13 ◽  
Author(s):  
Qi Wu ◽  
Yao Zhou ◽  
Xian-mei Zhou
Keyword(s):  
Pulmonary Fibrosis ◽  
Signaling Pathway ◽  
Cell Apoptosis ◽  
The Body ◽  
Collagen I ◽  
Intragastric Administration ◽  
Group Model ◽  
Pulmonary Tissue ◽  
Alkaline Extract ◽  
Collagen Iii

Objective. To investigate the intervention effect and functioning mechanism of citrus alkaline extract (CAE) on bleomycin- (BLM-) induced pulmonary fibrosis in mice. Methods. 42 C57BL/6 male mice were assigned randomly to the normal group, model group, low (16mg/kg), medial (32mg/kg) and high (64mg/kg) CAE dosage groups, prednisone group (6mg/kg), and pirfenidone group (100mg/kg), respectively. One day after model construction, intragastric administration was applied to the mice once a day for 28 days and then killed. Body weights of mice were recorded. Their pulmonary tissues were subjected to HE staining and Masson’s staining and then their degree of pulmonary alveolitis as well as pulmonary fibrosis was scored. Contents of hydroxyproline (HYP) and prostaglandin E2 (PGE2) in pulmonary tissues and levels of interleukin-17 (IL-17) in serum and bronchoalveolar lavage fluid (BALF) were determined by ELISA method. Expression of collagen I, collagen III, and Prosurfactant protein C (Pro-SPC) proteins in pulmonary tissue were measured immunohistochemically and that of nuclear transcription factor κB (NF-κB) and vimentin was determined by the immunofluorescence method. Apoptosis of pulmonary tissue was tested by the Tunel staining method, while the expression of MAPK-related protein was recorded by Western Blot assay. Results. After CAE treatment, the body weight, PGE2 level, and Pro-SPC protein expression of pulmonary fibrosis mice were increased, while the score of pulmonary alveolitis and pulmonary fibrosis, levels of HYP and cell apoptosis, IL-17 contents of serum and BALF in pulmonary tissues, and expression of collagen I, collagen III, vimentin, NF-κB, and p-p38 were reduced. Conclusion. CAE effectively delayed the progression of BLM-induced pulmonary fibrosis in pulmonary fibrosis mice and a possible mechanism is the inhibition of cell apoptosis of NF-κB/p38-mediated signaling pathway.

scholarly journals The study of lumbar ligamentum flavum hypertrophy induced in bipedal mice

2019 ◽  
Author(s):  
Zhenyu Zheng ◽  
Lei Qian ◽  
Xiang Ao ◽  
Peng Li ◽  
Yongxing Peng ◽  
...  
Keyword(s):  
Finite Element Analysis ◽  
Finite Element ◽  
Mouse Model ◽  
Ligamentum Flavum ◽  
The Elderly ◽  
Elastic Fibers ◽  
Related Factors ◽  
Element Analysis ◽  
Lumbar Spinal

AbstractLumbar spinal stenosis (LSS) is a common degenerative disease among the elderly. The role that mechanical stress-induced hypertrophic ligamentum flavum (HLF) plays in patients with LSS remains unclear. Here, we used a finite element analysis to investigate the stress characteristics on the ligamentum flavum (LF) and evaluate the feasibility of a mouse model of HLF. First, we induced a bipedal posture in mice by taking advantage of their hydrophobia. A micro-CT scan was performed to examine their spinal change during bipedal posture. A finite element analysis showed that the stress and strain on the upright posture were significantly increased compared with those on the sprawling posture. Tissue staining showed that the degeneration degree of the LF in bipedal standing group gradually increased over the modeling period. The amount of elastic fibers decreased under HLF, whereas the amount of collagen fibers, the number of the LF cells, and the expression of fibrosis-related factors increased. Compared with aged group, LF degeneration was more severe in the bipedal standing group. Our findings demonstrate that the increased stress caused by a posture change causes HLF and that a bipedal mouse model can be used to study HLF in vivo.

Expression of the JAK/STAT signaling pathway in the ligamentum flavum of patients with lumbar spinal canal stenosis

2017 ◽  
Vol 22 (2) ◽  
pp. 190-196 ◽  
Author(s):  
Hitoshi Yamahata ◽  
Koji Osuka ◽  
Tatsuro Aoyama ◽  
Muneyoshi Yasuda ◽  
Hiroshi Tokimura ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Spinal Canal ◽  
Ligamentum Flavum ◽  
Spinal Canal Stenosis ◽  
Lumbar Spinal Canal Stenosis ◽  
Canal Stenosis ◽  
Stat Signaling ◽  
Lumbar Spinal Canal ◽  
Lumbar Spinal

Abstract 17416: miR-1468-3p Regulates Development of Cardiac Fibrosis

Circulation ◽  
2018 ◽  
Vol 138 (Suppl_1) ◽  
Author(s):  
Ruizhu Lin ◽  
Lea Rahtu-Korpela ◽  
Johanna Magga ◽  
Lasse Pakanen ◽  
Katja Porvari ◽  
...  
Keyword(s):  
Protein Expression ◽  
Cardiac Fibrosis ◽  
Cardiac Fibroblasts ◽  
Collagen I ◽  
Loss Of Function ◽  
Collagen Iii ◽  
Approaches To Study ◽  
Human Cardiac Fibroblasts ◽  
Tgf Β1

Background: Accumulation of extracellular matrix disturbs the electrical conduction and stiffens myocardium, leading to higher risk for arrhythmias and diastolic dysfunction. MicroRNAs (miRNAs) function in post-translational gene regulation, and aberrant alteration of miRNAs level has been implicated in cardiac pathologies. Methods and Results: RNA sequencing of RNA samples of sudden cardiac death (SCD) victims with idiopathic myocardial fibrosis (IMF) for differentially expressed miRNAs identified miR-1468-3p. qPCR analysis validated that expression of miR-1468-3p is upregulated in hearts of SCD victims with IMF comparing to control subjects. However, the role and molecular function of miR-1468-3p in cardiac fibrosis are not known. We utilized human cardiac fibroblasts (hCFs) and gain- and loss-of-function approaches to study the role of miR-1468-3p in modulating fibroblast function. Overexpressing miR-1468-3p in hCFs resulted in an increase in expression of several fibrotic genes compared with hCFs transfected with control mir-mimic. Western blot analysis showed that miR-1468-3p mimic was sufficient to drive expression of collagen I and CTGF protein expression. Treatment of hCFs with miR-1468-3p antagomir did not alter expression of fibrosis-related gene at basal level, whereas miR-1468-3p inhibition significantly attenuated TGF-β1-induced collagen I and collagen III expression. Treatment of hCFs with miR-1468-3p antagomir blunted TGF-β1-induced collagen I and CTGF protein expression, but not TGF-β1-induced αSMA expression. Employing Sirius Red/Fast Green assay, we validated that depletion of miR-1468-3p antagonized both TGF-β1-triggered collagen and non-collagen protein production. Finally, we found that miR-1468-3p antagomir downregulated TGF-β1-induced collagen expression partially through the interference of TGF-β1/MAPK signals (p38 and JNK) and Integrin signaling. Conclusions: Our data indicate a pro-fibrotic role of miR-1468-3p in modulating cardiac fibrosis, and manipulating the expression of miR-1468-3p may provide a therapeutic strategy for treatment of cardiac fibrosis.

scholarly journals Mice overexpressing latent TGF-β1 are protected against renal fibrosis in obstructive kidney disease

2008 ◽  
Vol 295 (1) ◽  
pp. F118-F127 ◽  
Author(s):  
Xiao R. Huang ◽  
Arthur C. K. Chung ◽  
Xiao J. Wang ◽  
Kar Neng Lai ◽  
Hui Y. Lan
Keyword(s):  
Kidney Disease ◽  
Signaling Pathway ◽  
Renal Fibrosis ◽  
Transforming Growth Factor ◽  
Smooth Muscle Actin ◽  
Critical Factor ◽  
Collagen I ◽  
Collagen Iii ◽  
Renal Histology ◽  
Tgf Β1

Transforming growth factor (TGF)-β1, once activated, binds to its receptors and mediates renal fibrosis via the downstream Smad signaling pathway. We reported here that mice overexpressing latent TGF-β1 in keratinocytes were protected against renal fibrosis in a model of obstructive kidney disease. In normal mice, both transgenic (Tg) and wild-type (WT) mice had normal renal histology and function, despite a 10-fold increase in plasma latent TGF-β1 in Tg mice. A severe renal fibrosis was developed in WT mice at 7 days after urinary obstruction. Unexpectedly, renal fibrosis was prevented in Tg mice, although levels of latent TGF-β1 in both circulation and renal tissues remained high. Compared with the WT mice, quantitative real-time PCR showed that upregulation of renal α-smooth muscle actin (SMA), collagen I, and collagen III mRNA was inhibited in Tg mice (60–70% reduced, all P < 0.01). These were further confirmed by immunohistochemistry with a marked inhibition of tubulointerstitial accumulation of α-SMA+ fibroblasts, collagen I, and collagen III matrix in Tg mice (all P < 0.001). Further studies showed that inhibition of renal fibrosis in Tg mice was associated with a significant reduction in renal TGF-β1 and CTGF (60% reduced, P < 0.05), an increase in renal Smad7, a suppression of TSP-1 (a critical factor for TGF-β1 activation), and an inhibition of Smad2/3 activation (all P < 0.001). In conclusion, latent TGF-β may play a protective role in renal fibrosis. Inhibition of renal TGF-β1 expression and activation, thereby blocking the downstream TGF-β signaling pathway, may be a critical mechanism by which latent TGF-β1 protects against renal fibrosis.

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