RESUMO
BACKGROUND: One increasingly important trend in plastic, reconstructive, and aesthetic surgery is the use of fat grafts to improve cutaneous wound healing. In clinical practice, lipoaspirates (adipose tissue harvested by liposuction) are re-injected in a procedure called lipofilling. Previous studies, however, mainly evaluated the regenerative effect of isolated adipocytes, adipose-derived stem cells, and excised en bloc adipose tissue on keratinocytes, whereas no study to date has examined the effect of lipoaspirates. OBJECTIVES: The authors aimed to investigate differences in the regenerative property of en bloc adipose tissue and lipoaspirates on keratinocytes. METHODS: Human keratinocytes, lipoaspirates, and en bloc adipose tissue from 36 healthy donors were isolated. In vitro proliferation, differentiation, migration, stratification, and wound healing of keratinocyte monolayers were measured. Furthermore, secreted levels of VEGF, bFGF, IGF-1, MMP-9, and MIF were detected by ELISA. RESULTS: Migration, proliferation, and wound healing of keratinocytes were increased by lipoaspirates. Interestingly, the effect of lipoaspirates on keratinocyte proliferation was significantly higher than by en bloc adipose tissue after 5 days. The differentiation of keratinocytes was equally attenuated by lipoaspirates and en bloc adipose tissue. Stratification of keratinocyte layers was enhanced by lipoaspirates and en bloc fat when compared to controls. Lipoaspirates secrete higher levels of bFGF, whereas higher levels of VEGF and IGF-1 are released by en bloc adipose tissue. CONCLUSION: We show that lipoaspirates and en bloc adipose tissue have a regenerative effect on keratinocytes. One reason for the higher effect of lipoaspirates on keratinocyte proliferation may be the secretion of different cytokines.
Assuntos
Tecido Adiposo/metabolismo , Tecido Adiposo/cirurgia , Queratinócitos/metabolismo , Lipectomia , Adipócitos/metabolismo , Adolescente , Adulto , Idoso , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Adulto JovemRESUMO
INTRODUCTION: Autologous fat grafting has been used for reconstructive and aesthetic surgery for more than a century. Although initial criticism involved fat resorption and later the possible radiological interference in mammographies, the most recent debate focuses on the carcinogenicity of fat grafts. Among malignant skin tumors, melanomas show the highest lethality and lead to significant soft tissue defects after resection with fat grafting representing a feasible therapy option for tissue reconstruction. In contrast to breast carcinoma, little data is available regarding the carcinogenicity of fat grafts in melanoma patients. The present study was designed to investigate the influence of whole adipose tissue on the growth of melanoma cell lines compared to breast cancer cell lines. METHODS: Human subcutaneous adipose tissue was obtained from elective surgeries and cut into pieces ranging from 20 to 500 mg. Adipose tissue samples were co-cultivated with the human breast cancer cell line MCF-7 and the melanoma cell lines MEL-JUSO and IGR-37, and proliferation was measured using the alamarBlue proliferation assay. Additionally, the same experiment was conducted under nutrient-limited conditions. RESULTS: We found that adipose tissue significantly increased the proliferation of the human breast cancer cell line MCF-7 and importantly also of the melanoma cell lines MEL-JUSO and IGR-37 under optimal nutrient supply. Although the impact of adipose tissue on the growth of MCF-7 cells under nutrient deprivation was comparatively weak, the proliferation of MEL-JUSO and IGR-37 cells was clearly stimulated by adipose tissue. DISCUSSION: Our results show that adipose tissue enhances not only the growth of breast cancer but also melanoma cells even under conditions of nutrient limitation by soluble factors secreted by adipose tissue. With respect to our results and the current literature, reconstruction of soft tissue defects after malignancies including breast cancer and melanoma should be conducted with great care and only after complete tumor resection.
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Tecido Adiposo/transplante , Neoplasias da Mama/patologia , Melanoma/patologia , Neoplasias Cutâneas/patologia , Adolescente , Adulto , Idoso , Neoplasias da Mama/terapia , Linhagem Celular Tumoral , Proliferação de Células , Feminino , Humanos , Masculino , Melanoma/terapia , Pessoa de Meia-Idade , Neoplasias Cutâneas/terapia , Adulto JovemRESUMO
In this study we investigated the influence of surface topography on the inflammatory response of human macrophages. We generated different polyvinylidene fluoride (PVDF) surfaces including (i) a smooth surface of PVDF spherulites as a control, (ii) a randomly nanotextured surface with alumina particles, and (iii) a microstructure using laser ablation. The identical chemistry of all PVDF surfaces was demonstrated by X-ray photoelectron spectroscopy. The topography was evaluated by white light interferometry and X-profile analysis. Macrophages were cultured on the different surfaces including lipopolysaccharide (LPS) treatment as an inflammatory activator. Our results demonstrate that the microstructured surface but not the nanotexured significantly affects the activation of primary human macrophages by inducing a specific cytokine and gene expression pattern. This activation resulted in a subtype of macrophages with pro- but also anti-inflammatory properties. Interestingly, the response on the topography differed from that triggered by LPS, pointing to a different activation state of the cells. Our data clearly show that a particular topography induces an inflammatory response. This suggests that the modification of topography could influence the inflammatory potency of a biomaterial and hence could affect the biocompatibility of implants.
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Materiais Biocompatíveis/administração & dosagem , Citocinas/imunologia , Regulação da Expressão Gênica/imunologia , Ativação de Macrófagos/imunologia , Macrófagos/imunologia , Polivinil/administração & dosagem , Agregação Celular/efeitos dos fármacos , Agregação Celular/imunologia , Células Cultivadas , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Teste de Materiais , Propriedades de SuperfícieRESUMO
To allow for a better implementation of external volume expansion to clinical applications for soft tissue regeneration, it is necessary to comprehensively understand the underlying mechanisms. As human adipose-derived stem cells (hASCs) play a crucial role in soft tissue enlargement, we investigated the impact of cyclic stretch on gene expression, proliferation rate and adipogenic differentiation of these cells. After cyclic stretching, RNA was extracted and subjected to DNA microarray analysis and reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Also, the expression of FABP4 mRNA was analysed by RT-qPCR to test whether mechanical stretch affected adipogenic differentiation of hASCs. The proliferation rate was assessed using the alamarBlue assay and Ki-67 staining. A cell cycle analysis was performed with flow cytometry and Western blot. We found that cyclic stretch significantly induced the expression of CYP1B1 mRNA. Furthermore, the adipogenic differentiation of hASCs was impaired, as was the proliferation. This was partly due to a decrease in extracellular signal-regulated kinase (ERK) 1/2 and histone H3 phosphorylation, suggesting a growth arrest in the G2 /M phase of the cell cycle. Enrichment analyses demonstrated that stretch-regulated genes were over-represented in pathways and biological processes involved in extracellular matrix organization, vascular remodelling and responses to cell stress. Taken together, mechanical stress impaired both proliferation and adipogenic differentiation, but led to a tissue-remodelling phenotype of hASCs. These data suggest that extracellular matrix remodelling and neoangiogenesis may play a more important role in external volume expansion than proliferation and adipogenesis of hASCs. Copyright © 2017 John Wiley & Sons, Ltd.
Assuntos
Adipogenia , Tecido Adiposo/citologia , Regulação da Expressão Gênica , Células-Tronco/citologia , Células-Tronco/metabolismo , Estresse Mecânico , Ciclo Celular , Proliferação de Células , Citocromo P-450 CYP1B1/genética , Citocromo P-450 CYP1B1/metabolismo , Proteínas de Ligação a Ácido Graxo/genética , Proteínas de Ligação a Ácido Graxo/metabolismo , Ontologia Genética , HumanosRESUMO
Surface modifications of implants are frequently done using bioactive peptides. However, immune cells such as macrophages might evoke a rejection of an implant due to an undesired activation by the materials. Here, the influence of different strategies for peptide immobilization onto (poly)-vinylidene fluoride (PVDF) on inflammation and angiogenesis is studied. The inflammatory response of human primary macrophages is investigated by analyzing inflammatory cytokine expression. Surface roughness and adsorptive coupling have only minor effects on macrophage activation. Acrylic acid (AAc)-based covalent RGD-coupling leads to the most favorable cellular reaction, indicated by increased VEGF release. Chemical vapor deposition treated surfaces are inert, but additional covalent coupling of RGD induces a pronounced proinflammatory reaction. An in vivo angiogenesis study reveals that covalent coupling of RGD results in delayed but increased angiogenesis. It is concluded that for implant decoration with peptides, the substrate material has to be selected carefully to prevent inflammatory immune responses.
Assuntos
Proteínas Imobilizadas/química , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Neovascularização Fisiológica/efeitos dos fármacos , Oligopeptídeos/química , Acrilatos/química , Animais , Materiais Biocompatíveis , Adesão Celular , Embrião de Galinha , Membrana Corioalantoide/irrigação sanguínea , Membrana Corioalantoide/efeitos dos fármacos , Humanos , Proteínas Imobilizadas/farmacologia , Interleucina-1beta/metabolismo , Macrófagos/citologia , Macrófagos/metabolismo , Oligopeptídeos/farmacologia , Polivinil/química , Cultura Primária de Células , Propriedades de Superfície/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
Adipose tissue engineering aims to grow fat tissue for soft tissue reconstruction after tumour resection or trauma. However, insufficient progenitor cell differentiation and poor vascularization compromise the generation of clinically applicable adipose tissue. The desired process of neo-adipogenesis seems to be difficult to mimic, even though it takes place in all of us, inevitably and rapidly, as soon as we start consuming high-caloric diets. It has previously been proposed that inflammation and its key regulator, nitric oxide (NO), may play a relevant part in neo-adipogenesis. We here discuss how a controlled activation of the nitric oxide system on various levels may represent a cure for several current shortcomings in adipose tissue engineering.
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Tecido Adiposo , Óxido Nítrico/fisiologia , Engenharia Tecidual , Animais , HumanosRESUMO
In this study, we investigated the influence of different perfluoropolyether (PFPE) microstructures on the inflammatory response of human macrophages. We generated four different microstructured PFPE surfaces by replica molding from silicon masters. The function-associated surface markers 27E10 and CD163 were monitored using flow cytometry to measure the pro- and anti-inflammatory reactions. Inflammatory mediator expression was measured at the protein and mRNA level. Lipopolysaccharide treatment served as positive control for pro-inflammatory activation. We observed that each micropattern induced a specific morphology, phenotype and mediator profile. A microstructure of regular grooves induced a pro-inflammatory phenotype (M1) which was not accompanied by release of pro-inflammatory mediators. However, the larger cylindrical posts induced an anti-inflammatory phenotype (M2) with a remarkable down-regulation of CXCL10. Smaller posts with a shorter distance exhibited a stronger pro-inflammatory response than those with a longer distance, on the levels of both phenotype and mediator release. Regression analysis suggests that the geometrical parameters of the microstructures, specifically the period of structures, may play an important role in macrophage response. Optimization of such microstructures may provide a method to invoke a predictable response of macrophages to implants and control the mediator release.