@misc{10481/71365,
year = {2021},
url = {http://hdl.handle.net/10481/71365},
abstract = {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 explantbased methods in the efficient generation of human keratinocyte primary cell cultures for tissue
engineering use.},
organization = {This study was supported by the NanoGSkin project of EuroNanoMed-III (ERA-NET Cofund action of the Horizon 2020 Research and Innovation Framework Programme, EU), and cofunded by the Instituto de Salud Carlos III (ISCIII) through AES 2017 (ref. AC17/00013) and the
Centro para el Desarrollo Tecnológico Industrial (CDTI) (ref. 00108589), Spanish Ministry of Science
and Innovation. Support was also provided through projects PE-0395-2019 from the Consejería de
Salud y Familias, Junta de Andalucía, Spain, and B-CTS-450-UGR20 (proyectos de I+D+i en el marco
del Programa Operativo FEDER Andalucía 2014-2020, University of Granada and Consejería de
Transformación Económica, Industria, Conocimiento y Universidades). Additional support was
from the Spanish Plan Nacional de Investigación Científica, Desarrollo e Innovación Tecnológica
(I+D+i) of the Spanish Ministry of Science and Innovation through grants FIS PI20/0317 (ISCIII), FIS
PI21/0980 (ISCIII) and RTC-2017-6696-1 (Nanogrow), co-financed by the European Regional Development Fund (ERDF) through the “Una manera de hacer Europa” program.},
publisher = {MDPI},
keywords = {EGF},
keywords = {Nanoparticles},
keywords = {NLC},
keywords = {Keratinocytes},
keywords = {Tissue engineering},
keywords = {Cell culture},
title = {Improvement of Cell Culture Methods for the Successful Generation of Human Keratinocyte Primary Cell Cultures Using EGF-Loaded Nanostructured Lipid Carriers},
doi = {10.3390/biomedicines9111634},
author = {Chato Astrain, Jesús and Sánchez Porras, David and García García, Óscar Darío and Vairo, Claudia and Villar Vidal, María and Villullas, Silvia and Sánchez-Montesinos García, Indalecio and Campos Sánchez, Fernando and Garzón Bello, Ingrid Johanna and Alaminos Mingorance, Miguel},
}