Glycation by glyoxal leads to profound changes in the behavior of dermal fibroblasts

IntroductionDiabetes is a worldwide health problem that is associated with severe complications. Advanced Glycation End products (AGEs) such as Nε-(carboxymethyl)lysine, which result from chronic hyperglycemia, accumulate in the skin of patients with diabetes. The effect of AGEs on fibroblast functionality and their impact on wound healing are still poorly understood.Research design and methodsTo investigate this, we treated cultured human fibroblasts with 0.6 mM glyoxal to induce acute glycation. The behavior of fibroblasts was analyzed by time-lapse monolayer wounding healing assay, seahorse technology and atomic force microscopy. Production of extracellular matrix was studied by transmission electronic microscopy and western blot. Lipid metabolism was investigated by staining of lipid droplets (LDs) with BODIPY 493/503.ResultsWe found that the proliferative and migratory capacities of the cells were greatly reduced by glycation, which could be explained by an increase in fibroblast tensile strength. Measurement of the cellular energy balance did not indicate that there was a change in the rate of oxygen consumption of the fibroblasts. Assessment of collagen I revealed that glyoxal did not influence type I collagen secretion although it did disrupt collagen I maturation and it prevented its deposition in the extracellular matrix. We noted a pronounced increase in the number of LDs after glyoxal treatment. AMPK phosphorylation was reduced by glyoxal treatment but it was not responsible for the accumulation of LDs.ConclusionGlyoxal promotes a change in fibroblast behavior in favor of lipogenic activity that could be involved in delaying wound healing.

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Arg-Gly-Asp-containing peptides expose novel collagen receptors on fibroblasts: implications for wound healing.

Cell Regulation ◽

10.1091/mbc.2.12.1035 ◽

1991 ◽

Vol 2 (12) ◽

pp. 1035-1044 ◽

Cited By ~ 20

Author(s):

M V Agrez ◽

R C Bates ◽

A W Boyd ◽

G F Burns

Keyword(s):

Extracellular Matrix ◽

Wound Healing ◽

Type I Collagen ◽

Regulatory Control ◽

Type I ◽

Collagen Gels ◽

Collagen Matrices ◽

Surface Receptors ◽

Rgd Sequence ◽

Collagen Receptors

Integrins are a family of cell-surface receptors intimately involved in the interactions of cells with their extracellular matrix. These receptors comprise an alpha and beta subunit in noncovalent association and many have been shown to recognize and bind an arginine-glycine-aspartate (RGD) sequence contained within their specific extracellular matrix ligand. Fibroblasts express integrin receptors belonging to two major subfamilies. Some of the members within the subfamily defined by beta 1 (VLA) are receptors for collagen but, perhaps surprisingly, the other major subfamily of integrins on fibroblasts--that defined by the alpha chain of the vitronectin receptor, alpha v--all appear to bind primarily vitronectin and/or fibronectin. In the present study we show that RGD-containing peptides expose cryptic binding sites on the alpha v-associated integrins enabling them to function as collagen receptors. The addition of RGD-containing peptides to fibroblasts cultured on type I collagen induced dramatic cell elongation and, when the cells were contained within collagen matrices, the peptides induced marked contraction of the gels. These processes were inhibited by Fab fragments of a monoclonal antibody against an alpha v integrin. Also, alpha v-associated integrins from cell lysates bound to collagen I affinity columns in the presence, but not in the absence, of RGD-containing peptides. These data suggest a novel regulatory control for integrin function. In addition, because the cryptic collagen receptors were shown to be implicated in the contraction of collagen gels, the generation of such binding forces suggests that this may be the major biological role for these integrins in processes such as wound healing.

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Effects of Tenascin C on the Integrity of Extracellular Matrix and Skin Aging

International Journal of Molecular Sciences ◽

10.3390/ijms21228693 ◽

2020 ◽

Vol 21 (22) ◽

pp. 8693

Author(s):

Young Eun Choi ◽

Min Ji Song ◽

Mari Hara ◽

Kyoko Imanaka-Yoshida ◽

Dong Hun Lee ◽

...

Keyword(s):

Extracellular Matrix ◽

Type I Collagen ◽

Transforming Growth Factor ◽

Mrna Level ◽

Skin Aging ◽

Dermal Fibroblasts ◽

Cell Physiology ◽

Type I ◽

Tenascin C ◽

Matrix Metalloproteinase 1

Tenascin C (TNC) is an element of the extracellular matrix (ECM) of various tissues, including the skin, and is involved in modulating ECM integrity and cell physiology. Although skin aging is apparently associated with changes in the ECM, little is known about the role of TNC in skin aging. In this study, we found that the Tnc mRNA level was significantly reduced in the skin tissues of aged mice compared with young mice, consistent with reduced TNC protein expression in aged human skin. TNC-large (TNC-L; 330-kDa) and -small (TNC-S; 240-kDa) polypeptides were observed in conditional media from primary dermal fibroblasts. Both recombinant TNC polypeptides, corresponding to TNC-L and TNC-S, increased the expression of type I collagen and reduced the expression of matrix metalloproteinase-1 in fibroblasts. Treatment of fibroblasts with a recombinant TNC polypeptide, corresponding to TNC-L, induced phosphorylation of SMAD2 and SMAD3. TNC increased the level of transforming growth factor-β1 (TGF-β1) mRNA and upregulated the expression of type I collagen by activating the TGF-β signaling pathway. In addition, TNC also promoted the expression of type I collagen in fibroblasts embedded in a three-dimensional collagen matrix. Our findings suggest that TNC contributes to the integrity of ECM in young skin and to prevention of skin aging.

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Elevated Fibronectin Levels in Profibrotic CD14+ Monocytes and CD14+ Macrophages in Systemic Sclerosis

Frontiers in Immunology ◽

10.3389/fimmu.2021.642891 ◽

2021 ◽

Vol 12 ◽

Author(s):

Michał Rudnik ◽

Amela Hukara ◽

Ievgeniia Kocherova ◽

Suzana Jordan ◽

Janine Schniering ◽

...

Keyword(s):

Systemic Sclerosis ◽

Type I Collagen ◽

Smooth Muscle Actin ◽

Dermal Fibroblasts ◽

Signalling Pathway ◽

Type I ◽

Sequencing Analysis ◽

Enhanced Production ◽

Collagen Secretion ◽

Pulmonary Macrophages

BackgroundSystemic sclerosis (SSc) is an autoimmune disease characterized by overproduction of extracellular matrix (ECM) and multiorgan fibrosis. Animal studies pointed to bone marrow-derived cells as a potential source of pathological ECM-producing cells in immunofibrotic disorders. So far, involvement of monocytes and macrophages in the fibrogenesis of SSc remains poorly understood.Methods and ResultsImmunohistochemistry analysis showed accumulation of CD14+ monocytes in the collagen-rich areas, as well as increased amount of alpha smooth muscle actin (αSMA)-positive fibroblasts, CD68+ and mannose-R+ macrophages in the heart and lungs of SSc patients. The full genome transcriptomics analyses of CD14+ blood monocytes revealed dysregulation in cytoskeleton rearrangement, ECM remodeling, including elevated FN1 (gene encoding fibronectin) expression and TGF-β signalling pathway in SSc patients. In addition, single cell RNA sequencing analysis of tissue-resident CD14+ pulmonary macrophages demonstrated activated profibrotic signature with the elevated FN1 expression in SSc patients with interstitial lung disease. Peripheral blood CD14+ monocytes obtained from either healthy subjects or SSc patients exposed to profibrotic treatment with profibrotic cytokines TGF-β, IL-4, IL-10, and IL-13 increased production of type I collagen, fibronectin, and αSMA. In addition, CD14+ monocytes co-cultured with dermal fibroblasts obtained from SSc patients or healthy individuals acquired a spindle shape and further enhanced production of profibrotic markers. Pharmacological blockade of the TGF-β signalling pathway with SD208 (TGF-β receptor type I inhibitor), SIS3 (Smad3 inhibitor) or (5Z)-7-oxozeaenol (TGF-β-activated kinase 1 inhibitor) ameliorated fibronectin levels and type I collagen secretion.ConclusionsOur findings identified activated profibrotic signature with elevated production of profibrotic fibronectin in CD14+ monocytes and CD14+ pulmonary macrophages in SSc and highlighted the capability of CD14+ monocytes to acquire a profibrotic phenotype. Taking together, tissue-infiltrating CD14+ monocytes/macrophages can be considered as ECM producers in SSc pathogenesis.

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Human Adipose-Derived Stem Cell Secreted Extracellular Matrix Incorporated into Electrospun Poly(Lactic-co-Glycolic Acid) Nanofibrous Dressing for Enhancing Wound Healing

Polymers ◽

10.3390/polym11101609 ◽

2019 ◽

Vol 11 (10) ◽

pp. 1609 ◽

Cited By ~ 6

Author(s):

Tang ◽

Yang ◽

Lin ◽

Chen ◽

Lu ◽

...

Keyword(s):

Extracellular Matrix ◽

Wound Healing ◽

Growth Factors ◽

Wound Dressing ◽

Type I Collagen ◽

Healing Process ◽

Wound Dressings ◽

Random Structure ◽

Type I ◽

Enhance Wound Healing

Wound dressing, which prevents dehydration and provides a physical barrier against infection to wound beds, can improve wound healing. The interactions between extracellular matrix (ECM) and growth factors is critical to the healing process. Electrospun nanofibers are promising templates for wound dressings due to the structure similarity to ECM of skin. Otherwise, the ECM secreted by human adipose-derived stem cells (hASCs) is rich in growth factors known to enhance wound healing. Accordingly, we propose that the PLGA nanofibrous template incorporated with hASCs-secreted ECM may enhance wound healing. In this study, PLGA nanofibrous matrixes with an aligned or a random structure were prepared by electrospinning. Human ASCs cultured on the aligned matrix had a better viability and produced a larger amount of ECM relative to that of random one. After 7 days’ cultivation, the hASCs on aligned PLGA substrates underwent decellularization to fabricate cECM/PLGA dressings. By using immunohistochemical staining against F-actin and cell nucleus, the removal of cellular components was verified. However, the type I collagen and laminin were well preserved on the cECM/PLGA nanofibrous matrixes. In addition, this substrate was hydrophilic, with appropriate mechanical strength to act as a wound dressing. The L929 fibroblasts had good activity, survival and proliferation on the cECM/PLGA meshes. In addition, the cECM/PLGA nanofibrous dressings improved the wound healing of surgically created full-thickness skin excision in a mouse model. This hASCs-secreted ECM incorporated into electrospun PLGA nanofibrous could be a promising dressing for enhancing wound healing.

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Antioxidant and Anti-Inflammatory Properties of Anthocyanins Extracted from Oryza sativa L. in Primary Dermal Fibroblasts

Oxidative Medicine and Cellular Longevity ◽

10.1155/2019/2089817 ◽

2019 ◽

Vol 2019 ◽

pp. 1-18 ◽

Cited By ~ 5

Author(s):

Pakhawadee Palungwachira ◽

Salunya Tancharoen ◽

Chareerut Phruksaniyom ◽

Sirinapha Klungsaeng ◽

Ratchaporn Srichan ◽

...

Keyword(s):

Wound Healing ◽

Oryza Sativa ◽

Group Treatment ◽

High Performance ◽

Type I Collagen ◽

Oryza Sativa L ◽

Antioxidant Properties ◽

Dermal Fibroblasts ◽

Type I ◽

Anti Inflammatory

Flavonoids are naturally active substances that form a large class of phenolic compounds abundant in certain foods. Black rice (Oryza sativa L.) contains high levels of anthocyanin polyphenols, which have beneficial effects on health owing to their antioxidant properties. The breakdown of collagenous networks with aging or skin deterioration results in the impairment of wound healing in the skin. Accordingly, reviving stagnant collagen synthesis can help maintain dermal homeostasis during wound healing. This study presents an assessment of the cellular activity of anthocyanins (ANT) extracted from Oryza sativa L., providing information necessary for the development of new products that support natural healing processes. The relative composition of ANT from Oryza sativa L. was determined by high-performance liquid chromatography/diode array detection. ANT promoted the migration of rat dermal fibroblasts (RDFs) and demonstrated antioxidant properties. ANT increased the mRNA expression of collagen type I alpha 2 (COL1A2) and upregulated type I collagen protein levels in H2O2-stimulated RDFs without cytotoxicity. Compared with the untreated group, treatment of RDFs with ANT in the presence of H2O2 led to the activation of signaling pathways, including the extracellular signal-regulated protein kinases 1 and 2 (ERK1/2) and Akt, whereas it significantly (p<0.001) inhibited the phosphorylation of IκBα and suppressed the activation of the nuclear factor-kappa B (NF-κB) subunits, p50 and p65, which are transcription factors responsible for inflammation. Taken together, our findings suggest that ANT from Oryza sativa L. have anti-inflammatory properties and antiaging potential by modulating type I collagen gene expression and suppressing H2O2-induced NF-κB activation in skin fibroblasts.

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Platelet-derived growth factors-AA and -BB regulate collagen and collagenase gene expression differentially in human fibroblasts

Biochemical Journal ◽

10.1042/bj3100585 ◽

1995 ◽

Vol 310 (2) ◽

pp. 585-588 ◽

Cited By ~ 28

Author(s):

E M L Tan ◽

H Qin ◽

S H Kennedy ◽

S Rouda ◽

J W Fox ◽

...

Keyword(s):

Gene Expression ◽

Time Course ◽

Type I Collagen ◽

Biological Activities ◽

Human Fibroblasts ◽

Dermal Fibroblasts ◽

Type I ◽

Mrna Levels ◽

Interstitial Collagenase ◽

Collagenase Gene

Platelet-derived growth factor (PDGF) is a mitogen associated with tissue repair, a process involving collagen synthesis and remodelling by interstitial collagenase. This study examines and compares the regulation of interstitial collagenase and collagen gene expression by PDGF-AA and -BB in human fibroblasts. Time-course analysis showed that neither PDGF-AA or -BB had a consistent effect on the expression of pro-alpha 1(I) or pro-alpha 2(I) type-I collagen genes. In contrast, interstitial collagenase gene expression was found to be consistently up-regulated severalfold by PDGF-BB. Enhanced expression of the collagenase gene was not apparently due to up-regulation of its promoter activity in human dermal fibroblasts, as indicated by transient and stable transfection experiments. Unlike PDGF-BB, PDGF-AA did not alter collagenase mRNA levels under low-serum culture conditions. Thus, the biological activities of the PDGF homodimers are different, with PDGF-BB being clearly more potent than PDGF-AA in its regulation of collagenase gene expression.

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Potential Wound Healing Activities of Galla Rhois in Human Fibroblasts and Keratinocytes

The American Journal of Chinese Medicine ◽

10.1142/s0192415x15500925 ◽

2015 ◽

Vol 43 (08) ◽

pp. 1625-1636 ◽

Cited By ~ 8

Author(s):

Hyo-Hyun Park ◽

Na-Young Park ◽

Sun-Gun Kim ◽

Kyu-Tae Jeong ◽

Eu-Jin Lee ◽

...

Keyword(s):

Wound Healing ◽

Tissue Injury ◽

Human Fibroblasts ◽

Ethanol Extract ◽

Collagen Type I ◽

Dermal Fibroblasts ◽

Type I ◽

Collagen Deposition ◽

Soluble Collagen ◽

Ldh Release

Wound healing is a complex process orchestrated by the regeneration of the epithelium and the remodeling of the extracellular matrix through processes like collagen deposition. Galla Rhois has been widely used in traditional Korean medicine for its various pharmacological effects, including an anticoccidial effect, however, little is known about its healing activity. The purpose of this study was to determine the effects of Galla Rhois ethanol extract (GRE) on wound healing activities, including H2O2-induced oxidative stress, cell migration, and lactate dehydrogenase (LDH) release assays using human keratinocyte (HaCaT) and dermal fibroblasts (CCD-986SK). In addition, total soluble collagen deposition and collagen gene expression for Type I and III collagen were evaluated in CCD-986SK. Total tannin and flavonoid contents for GRE were measured. GRE induced a significant increase in the number and migration of cells, along with a decrease in cell death and LDH release. In addition, it also induced the over-expression of collagen Type I and III mRNA and caused increased synthesis of total soluble collagen. The contents of total tannin and flavonoid for GRE were 55.7% ([Formula: see text][Formula: see text]mg/g) and 62.9% ([Formula: see text][Formula: see text]mg/g), respectively. The results suggest that GRE can cause accelerated wound healing by increasing cell survival, proliferation, migration, and collagen synthesis along with a potential anti-oxidant property. This evidence provides novel insight into natural therapy for tissue injury.

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Viability and collagen secretion by fibroblasts on titanium surfaces with different acid-etching protocols

International Journal of Implant Dentistry ◽

10.1186/s40729-019-0192-4 ◽

2019 ◽

Vol 5 (1) ◽

Cited By ~ 1

Author(s):

Vilton Zimmermann de Souza ◽

Rafael Manfro ◽

Júlio César Joly ◽

Carlos Nelson Elias ◽

Daiane Cristina Peruzzo ◽

...

Keyword(s):

Cell Proliferation ◽

Type I Collagen ◽

Human Fibroblasts ◽

Surface Treatments ◽

Blue Staining ◽

Type I ◽

Acid Solutions ◽

Collagen Secretion ◽

Significant Difference ◽

Titanium Surfaces

Abstract Background From the consolidation of surface treatments of dental implants and knowledge on the cellular mechanisms of osseointegration, studies have highlighted the importance of a connective tissue seal against the implant to prevent contamination from the oral environment and consequent biofilm formation. Objective This in vitro study aimed to evaluate whether different titanium surface treatments using acid solutions promoted an increase in collagen secretion, proliferation, and viability of fibroblasts. Material and methods Commercially pure grade-4 titanium disks (6 × 2 mm) were treated with different acid solutions (hydrochloric, nitric, and sulfuric) for 20 and 60 min, respectively, obtaining mean surface roughness of 0.1 to 0.15 μm and 0.5 to 0.7 μm. Human fibroblasts were seeded onto different surfaces and assessed after 24 h, 48 h, and 72 h for cell proliferation and viability using Trypan blue staining and MTT, respectively, as well as the secretion of type I collagen on to such surfaces using ELISA. Machined titanium surfaces were used as controls. Data were statistically analyzed using one-way ANOVA and Fisher's LSD test for multiple comparisons, adopting a significance level of 5%. Results No significant difference was observed in cell proliferation for the different surfaces analyzed. Cell viability was significantly lower on the machined surface, after 48 h, when compared to the groups treated with acid for 20 or 60 min, which did not differ from each other. The expression of type I collagen was lowest on the acid-treated surfaces. Conclusion The results showed that the acid treatment proposed did not promote fibroblast proliferation and viability nor favor type I collagen synthesis.

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Platelet-Released Growth Factors Induce Genes Involved in Extracellular Matrix Formation in Human Fibroblasts

International Journal of Molecular Sciences ◽

10.3390/ijms221910536 ◽

2021 ◽

Vol 22 (19) ◽

pp. 10536

Author(s):

Andreas Bayer ◽

Bernard Wijaya ◽

Franziska Rademacher ◽

Lena Möbus ◽

Mark Preuß ◽

...

Keyword(s):

Extracellular Matrix ◽

Growth Factors ◽

Growth Factor ◽

Transforming Growth Factor Beta ◽

Transforming Growth Factor ◽

Lysyl Oxidase ◽

Human Fibroblasts ◽

Dermal Fibroblasts ◽

Type I ◽

Cellular Mechanisms

Platelet concentrate products are increasingly used in many medical disciplines due to their regenerative properties. As they contain a variety of chemokines, cytokines, and growth factors, they are used to support the healing of chronic or complicated wounds. To date, underlying cellular mechanisms have been insufficiently investigated. Therefore, we analyzed the influence of Platelet-Released Growth Factors (PRGF) on human dermal fibroblasts. Whole transcriptome sequencing and gene ontology (GO) enrichment analysis of PRGF-treated fibroblasts revealed an induction of several genes involved in the formation of the extracellular matrix (ECM). Real-time PCR analyses of PRGF-treated fibroblasts and skin explants confirmed the induction of ECM-related genes, in particular transforming growth factor beta-induced protein (TGFBI), fibronectin 1 (FN1), matrix metalloproteinase-9 (MMP-9), transglutaminase 2 (TGM2), fermitin family member 1 (FERMT1), collagen type I alpha 1 (COL1A1), a disintegrin and metalloproteinase 19 (ADAM19), serpin family E member 1 (SERPINE1) and lysyl oxidase-like 3 (LOXL3). The induction of these genes was time-dependent and in part influenced by the epidermal growth factor receptor (EGFR). Moreover, PRGF induced migration and proliferation of the fibroblasts. Taken together, the observed effects of PRGF on human fibroblasts may contribute to the underlying mechanisms that support the beneficial wound-healing effects of thrombocyte concentrate products.

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Denervation does not change the ratio of collagen I and collagen III mRNA in the extracellular matrix of muscle

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.00483.2006 ◽

2007 ◽

Vol 292 (2) ◽

pp. R983-R987 ◽

Cited By ~ 19

Author(s):

Ellen M. Arruda ◽

Kevin Mundy ◽

Sarah Calve ◽

Keith Baar

Keyword(s):

Extracellular Matrix ◽

Type I Collagen ◽

Lysyl Oxidase ◽

Muscle Fibers ◽

Fold Increase ◽

Collagen I ◽

Type I ◽

Collagen Concentration ◽

Collagen Iii ◽

Picrosirius Red

Denervation or inactivity is known to decrease the mass and alter the phenotype of muscle and the mechanics of tendon. It has been proposed that a shift in the collagen of the extracellular matrix (ECM) of the muscle, increasing type III and decreasing type I collagen, may be partially responsible for the observed changes. We directly investigated this hypothesis using quantitative real-time PCR on muscles and tendons that had been denervated for 5 wk. Five weeks of denervation resulted in a 2.91-fold increase in collagen concentration but no change in the content of collagen in the muscle, whereas in the tendon there was no change in either the concentration or content of collagen. The expression of collagen I, collagen III, and lysyl oxidase mRNA in the ECM of muscle decreased (76 ± 1.6%, 73 ± 2.3%, and 83 ± 3.2%, respectively) after 5 wk of denervation. Staining with picrosirius red confirmed the earlier observation of a change in staining color from red to green. Taken with the observed equivalent decreases in collagen I and III mRNA, this suggests that there was a change in orientation of the ECM of muscle becoming more aligned with the axis of the muscle fibers and no change in collagen type. The change in collagen orientation may serve to protect the smaller muscle fibers from damage by increasing the stiffness of the ECM and may partly explain why the region of the tendon closest to the muscle becomes stiffer after inactivity.

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