Methods to study differences in cell mobility during skin wound healing in vitro

Monsuur, Hanneke N.; Boink, Mireille A.; Weijers, Ester M.; Roffel, Sanne; Breetveld, Melanie; Gefen, Amit; Broek, Lenie J. van den; Gibbs, Susan

doi:10.1016/j.jbiomech.2016.01.040

Cited by 63 publications

(48 citation statements)

References 42 publications

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“…Wound closure is one of the vital steps of wound healing. The in vitro scratch wound model is a commonly used model to assess cell migration and wound closure because of its simplicity, reproducibility, and cost effectiveness . Using this model, we found that CSG enhanced the mobility of both L929 and HDFa and increased the speed of closure by more than 20%.…”

Section: Discussionmentioning

confidence: 87%

In vitro cellular viability studies on a concentrated surfactant‐based wound dressing

Chen¹,

Salisbury²,

Percival³

2019

International Wound Journal

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In this study, three cellular cytotoxic assays (direct contact assay, extraction assay, and cell insert assay) were applied to evaluate the effects of a concentrated surfactant gel preserved with antimicrobials and a concentrated surfactant gel with 1% silver sulfadiazine on both the mouse fibroblast cell line L929 and human dermal fibroblasts (HDFa). Also, the in vitro wound model was wounded by a 100 μL pipette tip and used to assess cell migration and wound closure after treatment with both gels. A needle‐scratched membrane disruption model was used to preliminarily evaluate membrane stabilisation and the membrane‐resealing effects of concentrated surfactant gels. It was demonstrated that the concentrated surfactant gel preserved with antimicrobials was not toxic to both L929 and HDFa. However, the concentrated surfactant gel with 1% silver sulfadiazine demonstrated a degree of cytotoxicity to both cell types. After treatment with a concentrated surfactant gel preserved with antimicrobials, cell movement to close the scratch gap was enhanced at 24 and 48 hours. The results also showed that cells treated with the concentrated surfactant gel preserved with antimicrobials decreased cell necrosis and improved cell resistance of the f‐actin rearrangement after a needle scratch. The results demonstrated that a concentrated surfactant gel preserved with antimicrobials is non‐cytotoxic and has ability to accelerate wound closure by enhancing cell mobility. Furthermore, the concentrated surfactant gel appeared to stabilise the plasma membrane and demonstrated a resealing ability and helped to retain the plasma membrane integrity and enhanced wound healing.

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Section: Discussionmentioning

confidence: 87%

In vitro cellular viability studies on a concentrated surfactant‐based wound dressing

Chen¹,

Salisbury²,

Percival³

2019

International Wound Journal

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“…A number of recent quantitative studies have analysed in vitro wounding assays to investigate aspects of cell migration for various cell types . Human in vitro models were used in a study to investigate human skin integrity, wound closure, and scar formation . These methods included, cell culture, fibroblast and endothelial cell migration scratch assay, fibroblast chemotaxis assay, endothelial cell in vitro tube formation assay, and skin equivalent.…”

Section: Resultsmentioning

confidence: 99%

“…Additionally, in vitro models have had a significant role in the study of wound healing and scar formation . This had led to a better understanding of biomarker expression, the role of keratinocytes and fibroblast activity in wound and scar development . Although the disadvantage is that, it can be difficult to extrapolate the findings of in vitro research to the in vivo wound scenario.…”

Section: Discussionmentioning

confidence: 99%

Non‐animal models of wound healing in cutaneous repair: In silico, in vitro, ex vivo, and in vivo models of wounds and scars in human skin

Uddin

Bayat

2017

Wound Repair Regeneration

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Tissue repair models are essential to explore the pathogenesis of wound healing and scar formation, identify new drug targets/biomarkers and to test new therapeutics. However, no animal model is an exact replicate of the clinical situation in man as in addition to differences in the healing of animal skin; the response to novel therapeutics can be variable when compared to human skin. The aim of this review is to evaluate currently available non-animal wound repair models in human skin, including: in silico, in vitro, ex vivo, and in vivo. The appropriate use of these models is extremely relevant to wound-healing research as it enables improved understanding of the basic mechanisms present in the wound healing cascade and aid in discovering better means to regulate them for enhanced healing or prevention of abnormal scarring. The advantage of in silico models is that they can be used as a first in virtue screening tool to predict the effect of a drug/stimulus on cells/tissues and help plan experimental research/clinical trial studies but remain theoretical until validated. In vitro models allow direct quantitative examination of an effect on specific cell types alone without incorporating other tissue-matrix components, which limits their utility. Ex vivo models enable immediate and short-term evaluation of a particular effect on cells and its surrounding tissue components compared with in vivo models that provide direct analysis of a stimulus in the living human subject before/during/after exposure to a stimulus. Despite clear advantages, there remains a lack of standardisation in design, evaluation and follow-up, for acute/chronic wounds and scars in all models. In conclusion, ideal models of wound healing research are desirable and should mimic not only the structure but also the cellular and molecular interactions, of wound types in human skin. Future models may also include organ/skin-on-a-chip with potential application in wound healing research.

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“…Therefore quality controls during culture that confirmed fibroblast viability (>50% confluent after approximately 1 week of culture) at the time point when fibroblasts are combined with the epithelial sheet growing on the dermis were accepted. During the following 2 weeks (3 weeks of culture total), fibroblasts migrated into the dermis (Monsuur et al, 2016). Many fibroblasts remain in the lower regions of the dermis, with few cells migrating into the upper regions.…”

Section: Discussionmentioning

confidence: 99%

“…3450, Costar, Corning Inc., New York, NY, USA) and the adherent fibroblast cells were cultured for 7–10 days in DMEM containing 5% Fetal‐Clone III and 50 μg/ml gentamycin. After 7–10 days of culturing the primary fibroblasts and epithelial sheet apart, the acellular donor dermis containing the epithelial sheet was placed on to the fibroblasts in order to allow fibroblast migration into the reticular side of the dermis (Monsuur et al, 2016). The SS and GS were further cultured air exposed in keratinocyte medium (see above), but with only 1% Fetal‐Clone III and without gentamicin.…”

Section: Methodsmentioning

confidence: 99%

Comparison of advanced therapy medicinal product gingiva and skin substitutes and their in vitro wound healing potentials

Boink

Roffel

Breetveld

et al. 2017

J Tissue Eng Regen Med

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Skin and oral mucosa substitutes are a therapeutic option for closing hard‐to‐heal skin and oral wounds. Our aim was to develop bi‐layered skin and gingiva substitutes, from 3 mm diameter biopsies, cultured under identical conditions, which are compliant with current European regulations for advanced therapy medicinal products. We present in vitro mode of action methods to (i) determine viability: epithelial expansion, proliferation (Ki‐67), metabolic activity (MTT assay); (ii) characterize skin and gingiva substitutes: histology and immunohistochemistry; and (iii) determine potency: soluble wound healing mediator release (enzyme‐linked immunosorbent assay). Both skin and gingiva substitutes consist of metabolically active autologous reconstructed differentiated epithelium expanding from the original biopsy sheet on a fibroblast populated connective tissue matrix (donor dermis). Gingival epithelium expanded 1.7‐fold more than skin epithelium during the 3 week culture period. The percentage of proliferating Ki‐67‐positive cells located in the basal layer of the gingiva substitute was >1.5‐fold higher than in the skin substitute. Keratins 16 and 17, which are upregulated during normal wound healing, were expressed in both the skin and gingiva substitutes. Notably, the gingiva substitute secreted higher amounts of key cytokines involved in mitogenesis, motogenesis and chemotaxis (interleukin‐6 > 23‐fold, CXCL8 > 2.5‐fold) as well as higher amounts of the anti‐fibrotic growth factor, hepatocyte growth factor (>7‐fold), compared with the skin substitute. In conclusion, while addressing the viability, characterization and potency of the tissue substitutes, important intrinsic differences between skin and gingiva were discovered that may explain in part the superior quality of wound healing observed in the oral mucosa compared with skin.

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Methods to study differences in cell mobility during skin wound healing in vitro

Cited by 63 publications

References 42 publications

In vitro cellular viability studies on a concentrated surfactant‐based wound dressing

In vitro cellular viability studies on a concentrated surfactant‐based wound dressing

Non‐animal models of wound healing in cutaneous repair: In silico, in vitro, ex vivo, and in vivo models of wounds and scars in human skin

Comparison of advanced therapy medicinal product gingiva and skin substitutes and their in vitro wound healing potentials

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