Dataset Information

TGF?-1 Induced Cross-Linking of the Extracellular Matrix of Primary Human Dermal Fibroblasts.

ABSTRACT: Excessive cross-linking is a major factor in the resistance to the remodelling of the extracellular matrix (ECM) during fibrotic progression. The role of TGF? signalling in impairing ECM remodelling has been demonstrated in various fibrotic models. We hypothesised that increased ECM cross-linking by TGF? contributes to skin fibrosis in Systemic Sclerosis (SSc). Proteomics was used to identify cross-linking enzymes in the ECM of primary human dermal fibroblasts, and to compare their levels following treatment with TGF?-1. A significant upregulation and enrichment of lysyl-oxidase-like 1, 2 and 4 and transglutaminase 2 were found. Western blotting confirmed the upregulation of lysyl hydroxylase 2 in the ECM. Increased transglutaminase activity in TGF?-1 treated ECM was revealed from a cell-based assay. We employed a mass spectrometry-based method to identify alterations in the ECM cross-linking pattern caused by TGF?-1. Cross-linking sites were identified in collagens I and V, fibrinogen and fibronectin. One cross-linking site in fibrinogen alpha was found only in TGF?-treated samples. In conclusion, we have mapped novel cross-links between ECM proteins and demonstrated that activation of TGF? signalling in cultured dermal fibroblasts upregulates multiple cross-linking enzymes in the ECM.

SUBMITTER: Semkova ME

PROVIDER: S-EPMC7863744 | biostudies-literature | 2021 Jan

REPOSITORIES: biostudies-literature

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TGFβ-1 Induced Cross-Linking of the Extracellular Matrix of Primary Human Dermal Fibroblasts.

Semkova Mariya E ME Hsuan J Justin JJ

International journal of molecular sciences 20210120 3

Excessive cross-linking is a major factor in the resistance to the remodelling of the extracellular matrix (ECM) during fibrotic progression. The role of TGFβ signalling in impairing ECM remodelling has been demonstrated in various fibrotic models. We hypothesised that increased ECM cross-linking by TGFβ contributes to skin fibrosis in Systemic Sclerosis (SSc). Proteomics was used to identify cross-linking enzymes in the ECM of primary human dermal fibroblasts, and to compare their levels follow ...[more]

PMID: 33498156

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Project description:The dermis of human skin contains large numbers of fibroblasts that are responsible for the production of the extracellular matrix (ECM) that supporting skin integrity, elasticity and wound healing. Previously, an in vivo study demonstrated that dermal fibroblasts siting in the lower dermis are capable to convert into skin adipose layer and hence fibroblast lipogenesis may vary the structure and elasticity of dermis. In the present study, Hs68 human dermal fibroblasts were utilized as an in vitro model to study the lipogenesis via using adipogenic differentiation medium (ADM). Baicalein, isolated from Scutellaria baicalensis, is one of the flavonoids to inhibit adipocyte differentiation due to high antioxidant activity in vitro. In order to develop a suitable formulation for baicalein (a poorly water-soluble drug), soybean phosphatidylcholine (SPC) was used to prepare baicalein-loaded liposomes to enhance drug bioavailability. Our results demonstrated that liposome-encapsulated baicalein protected cell viability and increased cellular uptake efficiency of Hs68 fibroblasts. Lipid accumulation, triglyceride synthesis and gene expressions of lipogenesis enzymes (FABP4 and LPL) were significantly increased in ADM-stimulated Hs68 fibroblasts but subsequently suppressed by liposome-encapsulated baicalein. In addition, ADM-induced TNF-? expression and related inflammatory factors was down-regulated by liposome-encapsulated baicalein. Through ADM-induced lipogenesis, the protein expression of elastin, type I and type III collagens increased remarkably, whereas liposome-encapsulated baicalein can down-regulate ADM-induced ECM protein synthesis. Taken together, we found that liposome-encapsulated baicalein can inhibit ADM-induced lipid accumulation and ECM formation in Hs68 fibroblasts through the suppression of lipogenesis enzymes and inflammatory responses. Liposome-encapsulated baicalein may have the potential to improve wound healing and restore skin structure after skin injury.

Project description:BackgroundAlginate matrix 3-dimensional culture offers the opportunity for the development and maturation of human secondary follicles in vitro. However, alginate may not be the most suitable culture system for human primordial/primary follicles in vitro. Thus, the innovation of alginate matrix 3-dimensional culture systems for human primordial/primary follicles could hold promise as an ideal approach to restoring fertility.MethodsWe extracted primordial/primary follicles from ovarian tissues collected from patients with non-ovarian benign gynecological conditions. Fibroblasts were isolated from dermal tissue from 1 male patient who had undergone posthectomy. The isolated human follicles were randomly divided into 2 groups and encapsulated within fibroblast-alginate-hydrogels or alginate hydrogels. The survival and growth of human primordial/primary follicles were measured after 21 days of in vitro culture.ResultsThe dermal fibroblasts in alginate hydrogel microcapsules were round in shape, and were distributed as uniform clouds on the surface and gaps of the alginate. After 21 days of culture, the survival rate of follicles in the fibroblast-alginate group was higher than that of the alginate group (P<0.05). The diameter of follicles in the fibroblast-alginate group and the alginate group after 21 days of culture was 152.80±13.64 and 129.14±9.95 μm, respectively (P<0.05). After 21-day culture, the mean cpm (log-converted) for 3H-thymidine incorporated by granulosa cells in the fibroblast-alginate and alginate groups was 6.87±0.24 and 4.63±0.38, respectively (P<0.05). After 21 days of culture, the messenger RNA expression levels of growth differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15) were significantly higher in oocytes in fibroblast-alginate hydrogels than in those in alginate hydrogels (P<0.05).ConclusionsHuman fibroblasts are beneficial to the development of human follicles in 3-dimensional culture alginate gel systems over a long period of time. More studies are required to investigate the molecular biological mechanisms of human fibroblasts that promote follicle growth in vitro.

Dataset Information

TGF?-1 Induced Cross-Linking of the Extracellular Matrix of Primary Human Dermal Fibroblasts.

Publications

TGFβ-1 Induced Cross-Linking of the Extracellular Matrix of Primary Human Dermal Fibroblasts.

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