In Vitro Scratch Assay to Demonstrate Effects of Arsenic on Skin Cell Migration

Bronson I. Pinto, Nathan D. Cruz, Oscar R. Lujan, Catherine R Propper, Robert S Kellar

Research output: Contribution to journalArticle

Abstract

Understanding the physiologic mechanisms of wound healing has been the focus of ongoing research for many years. This research directly translates into changes in clinical standards used for treating wounds and decreasing morbidity and mortality for patients. Wound healing is a complex process that requires strategic cell and tissue interaction and function. One of the many critically important functions of wound healing is individual and collective cellular migration. Upon injury, various cells from the blood, surrounding connective, and epithelial tissues rapidly migrate to the wound site by way of chemical and/or physical stimuli. This migration response can largely dictate the outcomes and success of a healing wound. Understanding this specific cellular function is important for translational medicine that can lead to improved wound healing outcomes. Here, we describe a protocol used to better understand cellular migration as it pertains to wound healing, and how changes to the cellular environment can significantly alter this process. In this example study, dermal fibroblasts were grown in media supplemented with fetal bovine serum (FBS) as monolayer cultures in tissue culture flasks. Cells were aseptically transferred into tissue culture treated 12-well plates and grown to 100% confluence. Upon reaching confluence, the cells in the monolayer were vertically scratched using a p200 pipet tip. Arsenic diluted in culture media supplemented with FBS was added to individual wells at environmentally relevant doses ranging 0.1-10 M. Images were captured every 4 hours (h) over a 24 h period using an inverted light microscope to observe cellular migration (wound closure). Images were individually analyzed using image analysis software, and percent wound closure was calculated. Results demonstrate that arsenic slows down wound healing. This technique provides a rapid and inexpensive first screen for evaluation of the effects of contaminants on wound healing.

Original languageEnglish (US)
JournalJournal of visualized experiments : JoVE
Issue number144
DOIs
StatePublished - Feb 23 2019

Fingerprint

Arsenic
Wound Healing
Cell Movement
Assays
Tissue culture
Skin
Monolayers
Tissue
Fibroblasts
Wounds and Injuries
Cell culture
Image analysis
Medicine
Culture Media
Microscopes
Blood
Impurities
Translational Medical Research
In Vitro Techniques
Serum

ASJC Scopus subject areas

  • Neuroscience(all)
  • Chemical Engineering(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)

Cite this

In Vitro Scratch Assay to Demonstrate Effects of Arsenic on Skin Cell Migration. / Pinto, Bronson I.; Cruz, Nathan D.; Lujan, Oscar R.; Propper, Catherine R; Kellar, Robert S.

In: Journal of visualized experiments : JoVE, No. 144, 23.02.2019.

Research output: Contribution to journalArticle

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