scholarly journals Improvement of Cell Culture Methods for the Successful Generation of Human Keratinocyte Primary Cell Cultures Using EGF-Loaded Nanostructured Lipid Carriers

Biomedicines ◽  
2021 ◽  
Vol 9 (11) ◽  
pp. 1634
Author(s):  
Jesús Chato-Astrain ◽  
David Sánchez-Porras ◽  
Óscar Darío García-García ◽  
Claudia Vairo ◽  
María Villar-Vidal ◽  
...  

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.

Pharmaceutics ◽  
2020 ◽  
Vol 12 (10) ◽  
pp. 928
Author(s):  
Sara Hernando ◽  
Enara Herran ◽  
Rosa Maria Hernandez ◽  
Manoli Igartua

Neurodegenerative diseases (ND) are one of the main problems of public health systems in the 21st century. The rise of nanotechnology-based drug delivery systems (DDS) has become in an emerging approach to target and treat these disorders related to the central nervous system (CNS). Among others, the use of nanostructured lipid carriers (NLCs) has increased in the last few years. Up to today, most of the developed NLCs have been made of a mixture of solid and liquid lipids without any active role in preventing or treating diseases. In this study, we successfully developed NLCs made of a functional lipid, such as the hydroxylated derivate of docohexaenoic acid (DHAH), named DHAH-NLCs. The newly developed nanocarriers were around 100 nm in size, with a polydispersity index (PDI) value of <0.3, and they exhibited positive zeta potential due to the successful chitosan (CS) and TAT coating. DHAH-NLCs were shown to be safe in both dopaminergic and microglia primary cell cultures. Moreover, they exhibited neuroprotective effects in dopaminergic neuron cell cultures after exposition to 6-hydroxydopamine hydrochloride (6-OHDA) neurotoxin and decreased the proinflammatory cytokine levels in microglia primary cell cultures after lipopolysaccharide (LPS) stimuli. The levels of the three tested cytokines, IL-6, IL-1β and TNF-α were decreased almost to control levels after the treatment with DHAH-NLCs. Taken together, these data suggest the suitability of DHAH-NLCs to attaining enhanced and synergistic effects for the treatment of NDs.


2021 ◽  
Vol 11 (1) ◽  
Author(s):  
James D. Nowotny ◽  
Michael T. Connelly ◽  
Nikki Traylor‑Knowles

An amendment to this paper has been published and can be accessed via a link at the top of the paper.


APOPTOSIS ◽  
2013 ◽  
Vol 18 (4) ◽  
pp. 452-466 ◽  
Author(s):  
Christina Pfister ◽  
Heike Pfrommer ◽  
Marcos S. Tatagiba ◽  
Florian Roser

1988 ◽  
Vol 405 (1) ◽  
pp. 77-103 ◽  
Author(s):  
R C Boucher ◽  
C U Cotton ◽  
J T Gatzy ◽  
M R Knowles ◽  
J R Yankaskas

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