title: Comparison of interleukin 10 homologs on dermal wound healing using a novel human skin ex vivo organ culture model.
abstract: BACKGROUND
  Anti-inflammatory cytokine interleukin (IL)-10 has been shown to induce regenerative healing in postnatal wounds. A viral homolog of IL-10 produced by human cytomegalovirus (CMV IL-10) similarly generates potent immunoregulatory effects, but its effects on wound healing have not been investigated. Currently, there are limited cost-effective methods of screening vulnerary therapeutics. Taken together, we aim to develop and validate a novel human ex vivo dermal wound model and hypothesize that CMV IL-10 will enhance dermal wound healing.
  
  
  METHODS
  Full-thickness circular (6-mm) explants were taken from surgical skin samples and 3-mm full-thickness wounds were created. Explants were embedded in collagen I matrix and maintained in specially formulated media with the epidermis at air-liquid interface, and treated with human IL-10 or CMV IL-10 (200 ng/mL). The viability of cultured explants was validated by histology and lactate dehydrogenase (LDH) activity. Epithelial gap, epithelial height, basal keratinocyte migration, vascular endothelial growth factor levels, and neovascularization were measured at days 3 and 7 to determine IL-10 effects on wound healing.
  
  
  RESULTS
  Culture explants at day 7 appeared similar to fresh skin in morphology, cell, and vessel density. By day 14, the epidermis separated from the dermis and the cell density diminished. Day 7 wounds appeared viable with advancing epithelial and basal keratinocyte migration with no evidence of necrosis. Cytotoxicity analysis via the quantification of LDH revealed no differences between controls and treated groups. There was a slight increase in the quantity of LDH in media at day 3; however, this decreased at day 5 and continued to decline up to day 21. CMV IL-10 treatment resulted in a significant decrease in the epithelial gap and an increase in epithelial height. There were no differences in the rates of basal keratinocyte migration at day 7 between treated and control groups. Interestingly, human IL-10 increased vascular endothelial growth factor expression and neovascularization compared with controls.
  
  
  CONCLUSIONS
  The human ex vivo wound model provides a simple and viable design to study dermal wound healing. Both IL-10 homologs demonstrate vulnerary effects. The viral homolog demonstrates enhanced effects on wound closure compared with human IL-10. These data represent a novel tool that can be used to screen therapeutics, such as CMV IL-10, before preclinical studies.
authors.affiliation: Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio.
authors.forename: Swathi
authors.initials: S
authors.lastname: Balaji
authors.affiliation: Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio.
authors.forename: Chad M
authors.initials: CM
authors.lastname: Moles
authors.affiliation: Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio.
authors.forename: Sukanta S
authors.initials: SS
authors.lastname: Bhattacharya
authors.affiliation: Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio.
authors.forename: Maria
authors.initials: M
authors.lastname: LeSaint
authors.affiliation: Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio.
authors.forename: Yashu
authors.initials: Y
authors.lastname: Dhamija
authors.affiliation: Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio.
authors.forename: Louis D
authors.initials: LD
authors.lastname: Le
authors.affiliation: Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio.
authors.forename: Alice
authors.initials: A
authors.lastname: King
authors.affiliation: Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio.
authors.forename: Mykia
authors.initials: M
authors.lastname: Kidd
authors.affiliation: Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio.
authors.forename: Muhammad F
authors.initials: MF
authors.lastname: Bouso
authors.affiliation: Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio.
authors.forename: Aimen
authors.initials: A
authors.lastname: Shaaban
authors.affiliation: Children's Hospital Colorado, Aurora, Colorado.
authors.forename: Timothy M
authors.initials: TM
authors.lastname: Crombleholme
authors.affiliation: Stanford University, Palo Alto, California.
authors.forename: Paul
authors.initials: P
authors.lastname: Bollyky
authors.affiliation: Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio. Electronic address: sundeepkeswani@gmail.com.
authors.forename: Sundeep G
authors.initials: SG
authors.lastname: Keswani
mesh_terms: D002465:Cell Movement / Q000187:drug effects; D003587:Cytomegalovirus / Q000737:chemistry*; D006801:Humans; D016753:Interleukin-10 / Q000494:pharmacology*; D007770:L-Lactate Dehydrogenase / Q000032:analysis; D018919:Neovascularization, Physiologic / Q000187:drug effects; D009924:Organ Culture Techniques; D012867:Skin / Q000293:injuries*; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics; D014764:Viral Proteins / Q000494:pharmacology*; D014945:Wound Healing / Q000187:drug effects*
chemical_list: D042461:Vascular Endothelial Growth Factor A; D014764:Viral Proteins; D016753:Interleukin-10; D007770:L-Lactate Dehydrogenase
keywords: IL-10 homologs; Organ culture; Regenerative medicine; Wound healing