Three-Dimensionally Printed Skin Substitute Using Human Dermal Fibroblasts and Human Epidermal Keratinocytes

2021 ◽  
Vol 86 (6S) ◽  
pp. S628-S631
Author(s):  
Jason Patel ◽  
Joseph Willis ◽  
Akshay Aluri ◽  
Shadi Awad ◽  
Metta Smith ◽  
...  
Keyword(s):  
Dermal Fibroblasts ◽  
Skin Substitute ◽  
Epidermal Keratinocytes ◽  
Human Dermal Fibroblasts ◽  
Human Epidermal Keratinocytes

Effects of silver nanoparticles on human dermal fibroblasts and epidermal keratinocytes

2016 ◽  
Vol 35 (9) ◽  
pp. 946-957 ◽  
Author(s):  
A Galandáková ◽  
J Franková ◽  
N Ambrožová ◽  
K Habartová ◽  
V Pivodová ◽  
...  
Keyword(s):  
Wound Healing ◽  
Silver Nanoparticles ◽  
Biomedical Application ◽  
Dermal Fibroblasts ◽  
Epidermal Keratinocytes ◽  
Topical Agent ◽  
Human Dermal Fibroblasts ◽  
Human Epidermal Keratinocytes ◽  
Normal Human ◽  
Treatment Of Wounds

Biomedical application of silver nanoparticles (AgNPs) has been rapidly increasing. Owing to their strong antimicrobial activity, AgNPs are used in dermatology in the treatment of wounds and burns. However, recent evidence for their cytotoxicity gives rise to safety concerns. This study was undertaken as a part of an ongoing programme in our laboratory to develop a topical agent for wound healing. Here, we investigated the potential toxicity of AgNPs using normal human dermal fibroblasts (NHDF) and normal human epidermal keratinocytes (NHEK) with the aim of comparing the effects of AgNPs and ionic silver (Ag-I). Besides the effect of AgNPs and Ag-I on cell viability, the inflammatory response and DNA damage in AgNPs and Ag-I–treated cells were examined. The results showed that Ag-I were significantly more toxic than AgNPs both on NHDF and NHEK. Non-cytotoxic concentrations of AgNPs and Ag-I did not induce DNA strand breaks and did not affect inflammatory markers, except for a transient increase in interleukin 6 levels in Ag-I–treated NHDF. The results showed that AgNPs are more suitable for the intended application as a topical agent for wound healing up to the concentration 25 µg/mL.

scholarly journals Fabrication of Bio-Based Gelatin Sponge for Potential Use as A Functional Acellular Skin Substitute

Polymers ◽  
2020 ◽  
Vol 12 (11) ◽  
pp. 2678
Author(s):  
Mior Muhammad Amirul Arif ◽  
Mh Busra Fauzi ◽  
Abid Nordin ◽  
Yosuke Hiraoka ◽  
Yasuhiko Tabata ◽  
...  
Keyword(s):  
Pore Size ◽  
Water Retention ◽  
Dermal Fibroblasts ◽  
Gelatin Sponge ◽  
Skin Substitute ◽  
Epidermal Keratinocytes ◽  
Phase Identification ◽  
High Porosity ◽  
Human Epidermal Keratinocytes ◽  
Crosslinking Degree

Gelatin possesses biological properties that resemble native skin and can potentially be fabricated as a skin substitute for full-thickness wound treatment. The native property of gelatin, whereby it is easily melted and degraded at body temperature, could prevent its biofunctionality for various applications. This study aimed to fabricate and characterise buffalo gelatin (Infanca halal certified) crosslinked with chemical type crosslinker (genipin and genipin fortified with EDC) and physicaly crosslink using the dihydrothermal (DHT) method. A porous gelatin sponge (GS) was fabricated by a freeze-drying process followed by a complete crosslinking via chemical—natural and synthetic—or physical intervention using genipin (GNP), 1-ethyl-3-(3-dimethylaminopropyl) (EDC) and dihydrothermal (DHT) methods, respectively. The physicochemical, biomechanical, cellular biocompatibility and cell-biomaterial interaction of GS towards human epidermal keratinocytes (HEK) and dermal fibroblasts (HDF) were evaluated. Results showed that GS had a uniform porous structure with pore size ranging between 60 and 200 µm with high porosity (>78.6 ± 4.1%), high wettability (<72.2 ± 7.0°), high tensile strain (>13.65 ± 1.10%) and 14 h of degradation rate. An increase in the concentration and double-crosslinking approach demonstrated an increment in the crosslinking degree, enzymatic hydrolysis resistance, thermal stability, porosity, wettability and mechanical strength. The GS can be tuned differently from the control by approaching the GS via a different crosslinking strategy. However, a decreasing trend was observed in the pore size, water retention and water absorption ability. Crosslinking with DHT resulted in large pore sizes (85–300 µm) and low water retention (236.9 ± 18.7 g/m2·day) and a comparable swelling ratio with the control (89.6 ± 7.1%). Moreover no changes in the chemical content and amorphous phase identification were observed. The HEK and HDF revealed slight toxicity with double crosslinking. HEK and HDF attachment and proliferation remain similar to each crosslinking approach. Immunogenicity was observed to be higher in the double-crosslinking compared to the single-crosslinking intervention. The fabricated GS demonstrated a dynamic potential to be tailored according to wound types by manipulating the crosslinking intervention.

scholarly journals Extracellular Vesicles Derived from Senescent Fibroblasts Attenuate the Dermal Effect on Keratinocyte Differentiation

2020 ◽  
Vol 21 (3) ◽  
pp. 1022 ◽  
Author(s):  
Eun-Jeong Choi ◽  
In Sup Kil ◽  
Eun-Gyung Cho
Keyword(s):  
Extracellular Vesicles ◽  
Proinflammatory Cytokine ◽  
Sandwich Elisa ◽  
Dermal Fibroblasts ◽  
Keratinocyte Differentiation ◽  
Epidermal Keratinocytes ◽  
Human Dermal Fibroblasts ◽  
Human Epidermal Keratinocytes ◽  
Lysosomal Activity ◽  
Epidermal Homeostasis

The skin is a multilayered and primary defensive organ. Intimate intercellular communication in the skin is necessary to ensure effective surveillance. Extracellular vesicles (EVs) are being explored for their involvement in intercellular skin communication. The aim of this study was to evaluate how human dermal fibroblasts (HDFs) accelerate EV production during senescence and the effects of senescence-associated EVs on epidermal homeostasis. Replicative senescent HDFs were assessed with senescence-associated β-galactosidase staining and the expression of senescence-related markers. Isolated EVs were characterized by dynamic light scattering and EV marker expression. EVs secreted from untreated young or senescent HDFs, or from those treated with a nSMase inhibitor, antioxidant, and lysosomal activity regulators, were determined by sandwich ELISA for CD81. Human epidermal keratinocytes were treated with young- and senescent HDF-derived EVs. Compared to young HDFs, senescent HDFs produced relatively high levels of EVs due to the increased nSMase activity, oxidative stress, and altered lysosomal activity. The nSMase inhibitor, antioxidant, and agents that recovered lysosomal activity reduced EV secretion in senescent HDFs. Relative to young HDF-derived EVs, senescent HDF-derived EVs were less supportive in keratinocyte differentiation and barrier function but increased proinflammatory cytokine IL-6 levels. Our study suggests that dermis-derived EVs may regulate epidermal homeostasis by reflecting cellular status, which provides insight as to how the dermis communicates with the epidermis and influences skin senescence.

Keratinocyte growth regulation in fibroblast cocultures via a double paracrine mechanism

1999 ◽  
Vol 112 (12) ◽  
pp. 1843-1853 ◽  
Author(s):  
N. Maas-Szabowski ◽  
A. Shimotoyodome ◽  
N.E. Fusenig
Keyword(s):  
Neutralizing Antibodies ◽  
Growth Regulation ◽  
Keratinocyte Growth Factor ◽  
Dermal Fibroblasts ◽  
Tissue Homeostasis ◽  
Epidermal Keratinocytes ◽  
Human Epidermal Keratinocytes ◽  
Keratinocyte Proliferation ◽  
Normal Human ◽  
Kinetics Of

Epithelial-mesenchymal interactions play an important role in regulating tissue homeostasis and repair. For skin, the regulatory mechanisms of epidermal-dermal interactions were studied in cocultures of normal human epidermal keratinocytes (NEK) and dermal fibroblasts (HDF) rendered postmitotic by alpha-irradiation (HDFi). The expression kinetics of different cytokines and their receptors with presumed signalling function in skin were determined at the RNA and protein level in mono- and cocultured NEK and HDFi. In cocultured HDFi, mRNA and protein synthesis of keratinocyte growth factor (KGF) (FGF-7) was strongly enhanced, whereas in cocultured keratinocytes interleukin (IL)-1alpha and -1beta mRNA expression increased compared to monocultures. Thus we postulated that IL-1, which had no effect on keratinocyte proliferation, induced in fibroblasts the expression of factors stimulating keratinocyte proliferation, such as KGF. The functional significance of this reciprocal modulation was substantiated by blocking experiments. Both IL-1alpha and -1beta-neutralizing antibodies and IL-1 receptor antagonist significantly reduced keratinocyte proliferation supposedly through abrogation of KGF production, because IL-1 antibodies blocked the induced KGF production. These data indicate a regulation of keratinocyte growth by a double paracrine mechanism through release of IL-1 which induces KGF in cocultured fibroblasts. Thus IL-1, in addition to its proinflammatory function in skin, may play an essential role in regulating tissue homeostasis.

scholarly journals Reduced serum methods for contact-based coculture of human dermal fibroblasts and epidermal keratinocytes

BioTechniques ◽  
2020 ◽  
Vol 69 (5) ◽  
pp. 347-355
Author(s):  
Snehal Kadam ◽  
Madhusoodhanan Vandana ◽  
Karishma S Kaushik
Keyword(s):  
Wound Closure ◽  
Fetal Bovine Serum ◽  
Cell Types ◽  
High Serum ◽  
Dermal Fibroblasts ◽  
Epidermal Keratinocytes ◽  
Human Dermal Fibroblasts ◽  
Fetal Bovine ◽  
High Concentrations ◽  
Serum Free Conditions

Direct contact-based coculture of human dermal fibroblasts and epidermal keratinocytes has been a long-standing and challenging issue owing to different serum and growth factor requirements of the two cell types. Existing protocols employ high serum concentrations (up to 10% fetal bovine serum), complex feeder systems and a range of supplemental factors. These approaches are technically demanding and labor intensive, and pose scientific and ethical limitations associated with the high concentrations of animal serum. On the other hand, serum-free conditions often fail to support the proliferation of one or both cell types when they are cultured together. We have developed two reduced serum approaches (1–2% serum) that support the contact-based coculture of human dermal fibroblasts and immortalized keratinocytes and enable the study of cell migration and wound closure.

scholarly journals Persea americanaMill. Seed: Fractionation, Characterization, and Effects on Human Keratinocytes and Fibroblasts

2013 ◽  
Vol 2013 ◽  
pp. 1-12 ◽  
Author(s):  
Maria del R. Ramos-Jerz ◽  
Socorro Villanueva ◽  
Gerold Jerz ◽  
Peter Winterhalter ◽  
Alexandra M. Deters
Keyword(s):  
Chlorogenic Acid ◽  
Dermal Fibroblasts ◽  
Persea Americana ◽  
Epidermal Keratinocytes ◽  
Hacat Keratinocytes ◽  
Human Epidermal Keratinocytes ◽  
Residual Solvent ◽  
Keratinocyte Proliferation ◽  
Normal Human

Methanolic avocado (Persea americanaMill., Lauraceae) seed extracts were separated by preparative HSCCC. Partition and HSCCC fractions were principally characterized by LC-ESI-MS/MS analysis. Theirin vitroinfluence was investigated on proliferation, differentiation, cell viability, and gene expression on HaCaT and normal human epidermal keratinocytes (NHEK) and normal human dermal fibroblasts (NHDF). The methanol-water partition (M) from avocado seeds and HSCCC fraction 3 (M.3) were mostly composed of chlorogenic acid and its isomers. Both reduced NHDF but enhanced HaCaT keratinocytes proliferation. HSCCC fractionM.2composed of quinic acid among chlorogenic acid and its isomers inhibited proliferation and directly induced differentiation of keratinocytes as observed on gene and protein level. Furthermore,M.2increased NHDF proliferation via upregulation of growth factor receptors. Salidrosides and ABA derivatives present in HSCCC fractionM.6increased NHDF and keratinocyte proliferation that resulted in differentiation. The residual solvent fractionM.7contained among low concentrations of ABA derivatives high amounts of proanthocyanidins B1 and B2 as well as an A-type trimer and stimulated proliferation of normal cells and inhibited the proliferation of immortalized HaCaT keratinocytes.

scholarly journals A gelatin/collagen/polycaprolactone scaffold for skin regeneration

PeerJ ◽  
2019 ◽  
Vol 7 ◽  
pp. e6358 ◽  
Author(s):  
Lin-Gwei Wei ◽  
Hsin-I Chang ◽  
Yiwei Wang ◽  
Shan-hui Hsu ◽  
Lien-Guo Dai ◽  
...  
Keyword(s):  
Stem Cells ◽  
Wound Healing ◽  
Dermal Fibroblasts ◽  
Skin Regeneration ◽  
Collagen Content ◽  
Wound Dressings ◽  
Skin Substitute ◽  
Epidermal Keratinocytes ◽  
Adipose Derived Stem Cells

Background A tissue-engineered skin substitute, based on gelatin (“G”), collagen (“C”), and poly(ε-caprolactone) (PCL; “P”), was developed. Method G/C/P biocomposites were fabricated by impregnation of lyophilized gelatin/collagen (GC) mats with PCL solutions, followed by solvent evaporation. Two different GC:PCL ratios (1:8 and 1:20) were used. Results Differential scanning calorimetry revealed that all G/C/P biocomposites had characteristic melting point of PCL at around 60 °C. Scanning electron microscopy showed that all biocomposites had similar fibrous structures. Good cytocompatibility was present in all G/C/P biocomposites when incubated with primary human epidermal keratinocytes (PHEK), human dermal fibroblasts (PHDF) and human adipose-derived stem cells (ASCs) in vitro. All G/C/P biocomposites exhibited similar cell growth and mechanical characteristics in comparison with C/P biocomposites. G/C/P biocomposites with a lower collagen content showed better cell proliferation than those with a higher collagen content in vitro. Due to reasonable mechanical strength and biocompatibility in vitro, G/C/P with a lower content of collagen and a higher content of PCL (GCLPH) was selected for animal wound healing studies. According to our data, a significant promotion in wound healing and skin regeneration could be observed in GCLPH seeded with adipose-derived stem cells by Gomori’s trichrome staining. Conclusion This study may provide an effective and low-cost wound dressings to assist skin regeneration for clinical use.

PREPARATION AND EVALUATION OF GAG-INCORPORATED SKIN SUBSTITUTE: AN IN VITRO STUDY

2006 ◽  
Vol 18 (03) ◽  
pp. 153-157 ◽  
Author(s):  
TZU-WEI WANG ◽  
HSI-CHIN WU ◽  
YI-CHAU HUANG ◽  
JUI-SHENG SUN ◽  
FENG-HUEI LIN
Keyword(s):  
Lower Layer ◽  
In Vitro Study ◽  
Dermal Fibroblasts ◽  
Skin Equivalent ◽  
Skin Substitute ◽  
Epidermal Keratinocytes ◽  
Different Temperatures ◽  
Air Liquid Interface ◽  
Epidermis Structure

A bi-layered gelatin-C6S-HA membrane with different pore sizes was prepared by freeze-drying at different temperatures - 20°C and -196°C, respectively Glycosaminoglycans (GAGs) were incorporated within the gelatin matrices to mimic the dermal composition and to create an appropriate environment for cell growth. The gelatin-C6S-HA membrane was cross-linked by 1-ethyl-3(3-dimethylaminopropryl) carbodiimide (EDC) to resist rapidly biodegradation by matrix enzymes. In this study, the lower layer of the sponge was inoculated with dermal fibroblasts for dermis development and as the feeder layer for epidermal keratinocytes. The upper layer was seeded with keratinocytes for epidermalization. After cultured for a period of time in air-liquid interface, the upper layer was developed into an epidermis structure with stratified epidermal layers. The lower part was developed into dermis-like structure synthesized by dermal fibroblasts surrounding with its own secreted extracellular matrix. In brief, the bi-layered skin equivalent with biological dermal analog and epidermal analog would be a suitable tool for autologous skin equivalent tissue engineering.

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