RT Journal Article
T1 Improvement of Cell Culture Methods for the Successful Generation of Human Keratinocyte Primary Cell Cultures Using EGF-Loaded Nanostructured Lipid Carriers
A1 Chato-Astrain, Jesus
A1 Sanchez-Porras, David
A1 Garcia-Garcia, Oscar Dario
A1 Vairo, Claudia
A1 Villar-Vidal, Maria
A1 Villullas, Silvia
A1 Sanchez-Montesinos, Indalecio
A1 Campos, Fernando
A1 Garzon, Ingrid
A1 Alaminos, Miguel
K1 EGF
K1 nanoparticles
K1 NLC
K1 keratinocytes
K1 tissue engineering
K1 cell culture
K1 Epidermal-growth-factor
K1 In-vitro
K1 Factor receptor
K1 Nanoparticles
K1 Rhegf
K1 Differentiation
K1 Proliferation
K1 Dressings
K1 Delivery
K1 Wounds
AB Human skin keratinocyte primary cultures can be established from skin biopsies with culture media containing epithelial growth factor (EGF). Although current methods are efficient, optimization is required to accelerate the procedure and obtain these cultures in less time. In the present study, we evaluated the effect of novel formulations based on EGF-loaded nanostructured lipid carriers (NLC). First, biosafety of NLC containing recombinant human EGF (NLC-rhEGF) was verified in immortalized skin keratinocytes and cornea epithelial cells, and in two epithelial cancer cell lines, by quantifying free DNA released to the culture medium. Then we established primary cell cultures of human skin keratinocytes with basal culture media (BM) and BM supplemented with NLC-rhEGF, liquid EGF (L-rhEGF), or NLC alone (NLC-blank). The results showed that cells isolated by enzymatic digestion and cultured with or without a feeder layer had a similar growth rate regardless of the medium used. However, the explant technique showed higher efficiency when NLC-rhEGF culture medium was used, compared to BM, L-rhEGF, or NLC-blank. Gene expression analysis showed that NLC-rhEGF was able to increase EGFR gene expression, along with that of other genes related to cytokeratins, cell-cell junctions, and keratinocyte maturation and differentiation. In summary, these results support the use of NLC-rhEGF to improve the efficiency of explant-based methods in the efficient generation of human keratinocyte primary cell cultures for tissue engineering use.
PB Mdpi
YR 2021
FD 2021-11-01
LK https://hdl.handle.net/10668/28038
UL https://hdl.handle.net/10668/28038
LA en
DS RISalud
RD Apr 10, 2025