scholarly journals Knockdown of Sestrin2 Increases Lipopolysaccharide-Induced Oxidative Stress, Apoptosis, and Fibrotic Reactions in H9c2 Cells and Heart Tissues of Mice via an AMPK-Dependent Mechanism

2018 ◽  
Vol 2018 ◽  
pp. 1-10 ◽  
Author(s):  
Hwan-Jin Hwang ◽  
Joo Won Kim ◽  
Hye Soo Chung ◽  
Ji A. Seo ◽  
Sin Gon Kim ◽  
...  
Keyword(s):  
Cell Death ◽  
Inflammatory Diseases ◽  
Adenosine Monophosphate ◽  
Collagen Type I ◽  
Heart Tissue ◽  
Type I ◽  
H9c2 Cells ◽  
Antioxidant Genes ◽  
Ampk Phosphorylation

Sestrin2 (sesn2) is an endogenous antioxidant protein that has recently gained attention for its potential to treat various inflammatory diseases. However, the relationship of sesn2 with cardiomyopathy is still unclear. In H9c2 cells, sesn2 knockdown reduced the level of 5′ adenosine monophosphate-activated protein kinase (AMPK) phosphorylation, downregulated antioxidant genes including catalase and superoxide dismutase (SOD2), and increased reactive oxygen species (ROS) production upon lipopolysaccharide (LPS) treatment. LPS-mediated cell death and the expression of matrix metalloproteinase (MMP) 2 and MMP9 were significantly increased by sesn2 knockdown. However, these increases were prevented by treatment with 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR), an AMPK activator. Consistent with the in vitro results, AMPK phosphorylation was decreased in heart tissue from sesn2 knockdown mice compared to heart tissue from control C57BL/6 mice, which was associated with decreased expression of antioxidant genes and increased LPS-mediated cell death signaling. Furthermore, the decrease in AMPK phosphorylation caused by sesn2 knockdown increased LPS-mediated expression of cardiac fibrotic factors, including collagen type I and type III, in addition to MMP2 and MMP9, in heart tissue from C57BL/6 mice. These results suggest that sesn2 is a novel potential therapeutic target for cardiomyopathy under inflammatory conditions.

The neuropeptide neuromedin U activates eosinophils and is involved in allergen-induced eosinophilia

2006 ◽  
Vol 290 (5) ◽  
pp. L971-L977 ◽  
Author(s):  
Maiko Moriyama ◽  
Satoru Fukuyama ◽  
Hiromasa Inoue ◽  
Takafumi Matsumoto ◽  
Takahiro Sato ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Inflammatory Diseases ◽  
Human Peripheral Blood ◽  
Collagen Type I ◽  
Eosinophil Infiltration ◽  
Type I ◽  
Dependent Manner ◽  
Peripheral Tissues ◽  
Eosinophil Cell

Neuromedin U (NMU) is a neuropeptide expressed not only in the central nervous system but also in various organs, including the gastrointestinal tract and lungs. NMU interacts with two G protein-coupled receptors, NMU-R1 and NMU-R2. Although NMU-R2 is expressed in a specific region of the brain, NMU-R1 is expressed in various peripheral tissues, including immune and hematopoietic cells. Our recent study demonstrated an important role of NMU in mast cell-mediated inflammation. In this study, we showed that airway eosinophilia was reduced in NMU-deficient mice in an allergen-induced asthma model. There were no differences in the antigen-induced Th2 responses between wild-type and NMU knockout mice. NMU-R1 was highly expressed in the eosinophil cell line, and NMU directly induced Ca2+mobilization and extracellular/signal-regulated kinase phosphorylation. NMU also induced cell adhesion to components of the extracellular matrix (fibronectin and collagen type I), and chemotaxis in vitro. Furthermore, NMU-R1 was also expressed in human peripheral blood eosinophils, and NMU induced cell adhesion in a dose-dependent manner. These data indicate that NMU promotes eosinophil infiltration into inflammatory sites by directly activating eosinophils. Our study suggests that NMU receptor antagonists could be novel targets for pharmacological inhibition of allergic inflammatory diseases, including asthma.

scholarly journals Collagen-Based Electrospun Materials for Tissue Engineering: A Systematic Review

2021 ◽  
Vol 8 (3) ◽  
pp. 39
Author(s):  
Britani N. Blackstone ◽  
Summer C. Gallentine ◽  
Heather M. Powell
Keyword(s):  
Systematic Review ◽  
Tissue Engineering ◽  
Collagen Type I ◽  
The Body ◽  
Pool Data ◽  
Type I ◽  
High Concentrations ◽  
Increased Strength

Collagen is a key component of the extracellular matrix (ECM) in organs and tissues throughout the body and is used for many tissue engineering applications. Electrospinning of collagen can produce scaffolds in a wide variety of shapes, fiber diameters and porosities to match that of the native ECM. This systematic review aims to pool data from available manuscripts on electrospun collagen and tissue engineering to provide insight into the connection between source material, solvent, crosslinking method and functional outcomes. D-banding was most often observed in electrospun collagen formed using collagen type I isolated from calfskin, often isolated within the laboratory, with short solution solubilization times. All physical and chemical methods of crosslinking utilized imparted resistance to degradation and increased strength. Cytotoxicity was observed at high concentrations of crosslinking agents and when abbreviated rinsing protocols were utilized. Collagen and collagen-based scaffolds were capable of forming engineered tissues in vitro and in vivo with high similarity to the native structures.

scholarly journals Relative rates of biosynthesis of collagen type I, type V and type VI in calf cornea

1991 ◽  
Vol 274 (2) ◽  
pp. 615-617 ◽  
Author(s):  
P Kern ◽  
M Menasche ◽  
L Robert
Keyword(s):  
Type I Collagen ◽  
Collagen Type ◽  
Corneal Stroma ◽  
Collagen Type I ◽  
Type I ◽  
Type Vi Collagen ◽  
Sds Page ◽  
Proline Incorporation ◽  
Type V

The biosynthesis of type I, type V and type VI collagens was studied by incubation of calf corneas in vitro with [3H]proline as a marker. Pepsin-solubilized collagen types were isolated by salt fractionation and quantified by SDS/PAGE. Expressed as proportions of the total hydroxyproline solubilized, corneal stroma comprised 75% type I, 8% type V and 17% type VI collagen. The rates of [3H]proline incorporation, linear up to 24 h for each collagen type, were highest for type VI collagen and lowest for type I collagen. From pulse-chase experiments, the calculated apparent half-lives for types I, V and VI collagens were 36 h, 10 h and 6 h respectively.

scholarly journals Plant Recombinant Human Collagen Type I Hydrogels for Corneal Regeneration

2021 ◽  
Author(s):  
Michel Haagdorens ◽  
Elle Edin ◽  
Per Fagerholm ◽  
Marc Groleau ◽  
Zvi Shtein ◽  
...  
Keyword(s):  
Collagen Type ◽  
Collagen Type I ◽  
Collagen Fibrils ◽  
Type I ◽  
Safe Alternative ◽  
Human Donor ◽  
Human Collagen ◽  
Donor Corneas

Abstract Purpose To determine feasibility of plant-derived recombinant human collagen type I (RHCI) for use in corneal regenerative implants Methods RHCI was crosslinked with 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS) to form hydrogels. Application of shear force to liquid crystalline RHCI aligned the collagen fibrils. Both aligned and random hydrogels were evaluated for mechanical and optical properties, as well as in vitro biocompatibility. Further evaluation was performed in vivo by subcutaneous implantation in rats and corneal implantation in Göttingen minipigs. Results Spontaneous crosslinking of randomly aligned RHCI (rRHCI) formed robust, transparent hydrogels that were sufficient for implantation. Aligning the RHCI (aRHCI) resulted in thicker collagen fibrils forming an opaque hydrogel with insufficient transverse mechanical strength for surgical manipulation. rRHCI showed minimal inflammation when implanted subcutaneously in rats. The corneal implants in minipigs showed that rRHCI hydrogels promoted regeneration of corneal epithelium, stroma, and nerves; some myofibroblasts were seen in the regenerated neo-corneas. Conclusion Plant-derived RHCI was used to fabricate a hydrogel that is transparent, mechanically stable, and biocompatible when grafted as corneal implants in minipigs. Plant-derived collagen is determined to be a safe alternative to allografts, animal collagens, or yeast-derived recombinant human collagen for tissue engineering applications. The main advantage is that unlike donor corneas or yeast-produced collagen, the RHCI supply is potentially unlimited due to the high yields of this production method. Lay Summary A severe shortage of human-donor corneas for transplantation has led scientists to develop synthetic alternatives. Here, recombinant human collagen type I made of tobacco plants through genetic engineering was tested for use in making corneal implants. We made strong, transparent hydrogels that were tested by implanting subcutaneously in rats and in the corneas of minipigs. We showed that the plant collagen was biocompatible and was able to stably regenerate the corneas of minipigs comparable to yeast-produced recombinant collagen that we previously tested in clinical trials. The advantage of the plant collagen is that the supply is potentially limitless.

scholarly journals Structural basis for distinct inflammasome complex assembly by human NLRP1 and CARD8

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Qin Gong ◽  
Kim Robinson ◽  
Chenrui Xu ◽  
Phuong Thao Huynh ◽  
Kelvin Han Chung Chong ◽  
...  
Keyword(s):  
Cell Death ◽  
Inner Core ◽  
Inflammatory Diseases ◽  
Structural Features ◽  
Structural Basis ◽  
Cultured Human Cells ◽  
Structural Insight ◽  
Sensor Proteins ◽  
Insight Into

AbstractNod-like receptor (NLR) proteins activate pyroptotic cell death and IL-1 driven inflammation by assembling and activating the inflammasome complex. Closely related sensor proteins NLRP1 and CARD8 undergo unique auto-proteolysis-dependent activation and are implicated in auto-inflammatory diseases; however, their mechanisms of activation are not understood. Here we report the structural basis of how the activating domains (FIINDUPA-CARD) of NLRP1 and CARD8 self-oligomerize to assemble distinct inflammasome complexes. Recombinant FIINDUPA-CARD of NLRP1 forms a two-layered filament, with an inner core of oligomerized CARD surrounded by an outer ring of FIINDUPA. Biochemically, self-assembled NLRP1-CARD filaments are sufficient to drive ASC speck formation in cultured human cells—a process that is greatly enhanced by NLRP1-FIINDUPA which forms oligomers in vitro. The cryo-EM structures of NLRP1-CARD and CARD8-CARD filaments, solved here at 3.7 Å, uncover unique structural features that enable NLRP1 and CARD8 to discriminate between ASC and pro-caspase-1. In summary, our findings provide structural insight into the mechanisms of activation for human NLRP1 and CARD8 and reveal how highly specific signaling can be achieved by heterotypic CARD interactions within the inflammasome complexes.

scholarly journals WNT-5A regulates TGF-β-related activities in liver fibrosis

2017 ◽  
Vol 312 (3) ◽  
pp. G219-G227 ◽  
Author(s):  
Leonie Beljaars ◽  
Sara Daliri ◽  
Christa Dijkhuizen ◽  
Klaas Poelstra ◽  
Reinoud Gosens
Keyword(s):  
Liver Fibrosis ◽  
Functional Role ◽  
Collagen Type ◽  
Collagen Type I ◽  
Matrix Proteins ◽  
Type I ◽  
Human Livers

WNT-5A is a secreted growth factor that belongs to the noncanonical members of the Wingless-related MMTV-integration family. Previous studies pointed to a connection between WNT-5A and the fibrogenic factor TGF-β warranting further studies into the functional role of WNT-5A in liver fibrosis. Therefore, we studied WNT-5A expressions in mouse and human fibrotic livers and examined the relation between WNT-5A and various fibrosis-associated growth factors, cytokines, and extracellular matrix proteins. WNT-5A gene and protein expressions were significantly increased in fibrotic mouse and human livers compared with healthy livers. Regression or therapeutic intervention in mice resulted in decreased hepatic WNT-5A levels paralleled by lower collagen levels. Immunohistochemical analysis showed WNT-5A staining in fibrotic septa colocalizing with desmin staining indicating WNT-5A expression in myofibroblasts. In vitro studies confirmed WNT-5A expression in this cell type and showed that TGF-β significantly enhanced WNT-5A expression in contrast to PDGF-BB and proinflammatory cytokines IL-1β and TNF-α. Additionally, TGF-β induces the expression of the WNT receptors FZD2 and FZD8. After silencing of WNT-5A, reduced levels of collagen type I, vimentin, and fibronectin in TGF-β-stimulated myofibroblasts were measured compared with nonsilencing siRNA-treated controls. Interestingly, the antifibrotic cytokine IFNγ suppressed WNT-5A in vitro and in vivo. IFNγ-treated fibrotic mice showed significantly less WNT-5A expression compared with untreated fibrotic mice. In conclusion, WNT-5A paralleled collagen I levels in fibrotic mouse and human livers. WNT-5A expression in myofibroblasts is induced by the profibrotic factor TGF-β and plays an important role in TGF-β-induced regulation of fibrotic matrix proteins, whereas its expression can be reversed upon treatment, both in vitro and in vivo. NEW & NOTEWORTHY This study describes the localization and functional role of WNT-5A in human and mouse fibrotic livers. Hepatic WNT-5A expression parallels collagen type I expression. In vivo and in vitro, the myofibroblasts were identified as the key hepatic cells producing WNT-5A. WNT-5A is under control of TGF-β and its activities are primarily profibrotic.

In Vitro Chondrotoxicity of Nonsteroidal Anti-inflammatory Drugs and Opioid Medications

2017 ◽  
Vol 45 (14) ◽  
pp. 3345-3350 ◽  
Author(s):  
Geoffrey D. Abrams ◽  
Wenteh Chang ◽  
Jason L. Dragoo
Keyword(s):  
Cell Death ◽  
Single Dose ◽  
Joint Surface ◽  
Saline Control ◽  
Type I ◽  
Dose Equivalent ◽  
Inflammatory Drugs ◽  
Anti Inflammatory ◽  
Opioid Medications

Background: A variety of medications are administered to the intra-articular space for the relief of joint pain. While amide-type local anesthetics have been extensively studied, there is minimal information regarding the potential chondrotoxicity of nonsteroidal anti-inflammatory drugs (NSAIDs) and opioid medications. Purpose: To investigate the in vitro chondrotoxicity of single-dose equivalent concentrations of ketorolac, morphine, meperidine, and fentanyl on human chondrocytes. Study Design: Controlled laboratory study. Methods: Human cartilage was arthroscopically harvested from the intercondylar notch and expanded in vitro. Gene expression of cultured chondrocytes before treatment was performed with quantitative polymerase chain reaction for type I collagen, type II collagen, aggrecan, and SOX9. Chondrocytes were then exposed to 0.01%, 0.02%, and 0.04% morphine sulfate; 0.3% and 0.6% ketorolac tromethamine; 0.5%, 1.0%, and 1.5% meperidine hydrochloride; 0.0005% and 0.001% fentanyl citrate; and saline. A custom bioreactor was used to constantly deliver medications, with the dosage of each medication and the duration of exposure based on standard dose equivalents, medication half-lives, and differences in the surface area between the 6-well plates and the native joint surface. After treatment, a live/dead assay was used to assess chondrocyte viability and if minimal cell death was detected. A subset of samples after treatment was maintained to analyze for possible delayed cell death. Results: All tested concentrations of ketorolac and meperidine caused significantly increased cell death versus the saline control, demonstrating a dose-response relationship. The morphine and fentanyl groups did not show increased chondrotoxicity compared with the saline group, even after 2 weeks of additional culture. Conclusion: In vitro exposure of chondrocytes to single-dose equivalent concentrations of either ketorolac or meperidine demonstrated significant chondrotoxicity, while exposure to morphine or fentanyl did not lead to increased cell death.

Chronic urotensin-II infusion induces diastolic dysfunction and enhances collagen production in rats

2010 ◽  
Vol 298 (2) ◽  
pp. H608-H613 ◽  
Author(s):  
Lavinia Tran ◽  
Andrew R. Kompa ◽  
Will Kemp ◽  
Arintaya Phrommintikul ◽  
Bing H. Wang ◽  
...  
Keyword(s):  
Diastolic Dysfunction ◽  
Intracellular Signaling ◽  
Cardiac Myocyte ◽  
Collagen Type I ◽  
Left Ventricular ◽  
High Dose ◽  
Type I ◽  
Urotensin Ii ◽  
Rock Inhibitor

The vasoactive peptide urotensin-II (U-II) is likely to play a key causal role in cardiac remodeling that ultimately leads to heart failure. Its contribution, specifically to the development of diastolic dysfunction and the downstream intracellular signaling, however, remains unresolved. This study interrogates the effect of chronic U-II infusion in normal rats on cardiac structure and function. The contribution of Rho kinase (ROCK) signaling to these pathophysiological changes is evaluated in cell culture studies. Chronic high-dose U-II infusion over 4 wk significantly impaired diastolic function in rats on echocardiography-derived Doppler indexes, including E-wave deceleration time (vehicle 56.7 ± 3.3 ms, U-II 118.0 ± 21.5 ms; P < 0.01) and mitral valve annulus peak early/late diastolic tissue velocity (vehicle 2.01 ± 0.19 ms, U-II 1.04 ± 0.25 ms; P < 0.01). A lower dose of U-II infusion (1 nmol·kg−1·h−1) yielded comparable changes. Diastolic dysfunction was accompanied by molecular [significant increases in procollagen-α1(I) gene expression on real-time PCR] and morphological (increases in total collagen, P < 0.05, and collagen type-I protein deposition, P < 0.001) evidence of left ventricular (LV) fibrosis following high-dose U-II infusion. The ROCK inhibitor GSK-576371 (10−7 to 10−5 M) elicited concentration-dependent inhibition of U-II (10−7 M)-stimulated cardiac fibroblast collagen synthesis and cardiac myocyte protein synthesis. Chronic U-II infusion causes diastolic dysfunction, caused by fibrosis of the LV. The in vitro data suggest that this may be in part occurring via a ROCK-dependent pathway.

Asporin competes with decorin for collagen binding, binds calcium and promotes osteoblast collagen mineralization

2009 ◽  
Vol 423 (1) ◽  
pp. 53-59 ◽  
Author(s):  
Sebastian Kalamajski ◽  
Anders Aspberg ◽  
Karin Lindblom ◽  
Dick Heinegård ◽  
Åke Oldberg
Keyword(s):  
Matrix Protein ◽  
Collagen Type I ◽  
Full Length ◽  
Extracellular Matrix Protein ◽  
Type I ◽  
Similar System ◽  
Collagen Binding ◽  
Osteoblastic Activity ◽  
Collagen Mineralization

The interactions of the ECM (extracellular matrix) protein asporin with ECM components have previously not been investigated. Here, we show that asporin binds collagen type I. This binding is inhibited by recombinant asporin fragment LRR (leucine-rich repeat) 10–12 and by full-length decorin, but not by biglycan. We demonstrate that the polyaspartate domain binds calcium and regulates hydroxyapatite formation in vitro. In the presence of asporin, the number of collagen nodules, and mRNA of osteoblastic markers Osterix and Runx2, were increased. Moreover, decorin or the collagen-binding asporin fragment LRR 10–12 inhibited the pro-osteoblastic activity of full-length asporin. Our results suggest that asporin and decorin compete for binding to collagen and that the polyaspartate in asporin directly regulates collagen mineralization. Therefore asporin has a role in osteoblast-driven collagen biomineralization activity. We also show that asporin can be expressed in Escherichia coli (Rosetta-gami™) with correctly positioned cysteine bridges, and a similar system can possibly be used for the expression of other SLRPs (small LRR proteoglycans/proteins).

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