Keratinocytes: Barrier to the Outside World
The epidermis is the outermost layer of the skin, which covers the entire body and provides an essential barrier against the external environment. The most common epidermal cell type is the keratinocyte. Keratinocytes are born in the basal layer of the epidermis and migrate to the outer layers as they mature, becoming terminally differentiated. These post-mitotic keratinocytes form the stratum corneum, or the most external dead layer of skin. Importantly, keratinocytes produce keratin, a structural protein that maintains the integrity of the skin. As with many other cell types, keratinocyte dysregulation can lead to disease. Of these, psoriasis is a very common skin condition caused by hyperproliferation of keratinocytes.
Studies Using Lifeline® Keratinocytes to Study Wound Healing and Psoriasis
Keratinocytes are highly involved in wound healing. In a 2013 study, Dooley et al. set out to develop an implantable resorbable surgical membrane composed of chitosan, a polysaccharide derived from the tough exoskeleton of crustaceans. Although chitosan has been used safely as a biomaterial in humans before, it has not been adapted for use as a surgical membrane. Here the authors generated chitosan membranes that were denser than previously developed chitosan membranes and tested its properties in vitro and in vivo. Compared to commercial membranes, the chitosan membranes displayed similar hydration properties, increased density, increased tensile strength, and increased suture pull-out strength. Furthermore, the in vitro properties of the chitosan membranes included: semi-permeability, ability to be degraded by cellular proteases, and dissolution at low pH. Using Lifeline® human primary keratinocytes, MC3T3 pre-osteoblasts, and human primary dermal foreskin fibroblasts, the researchers demonstrated that their chitosan membranes were biocompatible and sustained cell growth and viability. Finally, the group illustrated that in a rat model of epidermal wounding, chitosan membranes were biocompatible and were resorbed by 12 weeks. Together, the results of this study demonstrate that the dense chitosan membranes developed by the group are superior to current commercial surgical membranes and are applicable to multiple surgical uses.
Psoriasis is a common skin condition in which an inflammatory environment develops within the epidermis and keratinocytes become hyperproliferative. In a 2015 study in Nature Communications, Yan et al. investigated the role of the miR-31 microRNA and NF-kB signaling in psoriasis. They first examined human psoriatic skin lesions and found that miR-31 was upregulated in psoriasis patients, as well as in two different mouse models of psoriasis. Using Lifeline® normal human epidermal keratinocytes (NHEK), the group found that multiple inflammatory cytokines stimulated miR-31 expression. Additionally, in a mouse model of psoriasis, these same cytokines activated NF-kB signaling. Moreover, the group demonstrated that p65 (the transcriptionally active subunit of NF-kB) bound to the miR-31 promoter, suggesting that miR-31 is a target of NF-kB signaling.
To determine whether miR-31 has an active role in the pathogenesis of psoriasis, the authors generated a transgenic mouse in which miR-31 was overexpressed and compared the severity of induced psoriasis to a mouse model in which miR-31 was lost. They found that overexpression of miR-31 increased the severity of psoriatic lesions, while loss of miR-31 decreased lesion severity and onset, suggesting that miR-31 overexpression enhances the pathogenesis of psoriasis. Finally, the authors identified protein phosphatase 6 (ppp6c) as a direct target of miR-31. They showed that ppp6c knockdown in NHEK cells and in vivo resulted in increased keratinocyte proliferation. They also demonstrate that NF-kB activation decreases ppp6c, suggesting that the inflammatory environment present in psoriasis activates NF-kB signaling, increasing miR-30, which targets ppp6c, and increases keratinocyte proliferation.
The inflammatory environment within psoriatic lesions leads to upregulation of cytokines and chemokines, such as IL-17A and IL-22, which induce the production of cathelicidin (hCAP18), an antimicrobial peptide that when cleaved to produce the resultant LL37 peptide, can attract immune cells to the lesion. Vitamin D3 analogues are commonly used as topical treatments for psoriasis and act by suppressing keratinocyte cytokine production. In a 2014 study, Sakabe and colleagues set out to determine the effects of the Vitamin D3 analogue calcipotriol on hCAP18 and LL37. The authors first found that stimulation of Lifeline® normal human epidermal keratinocyte cells with IL-17A and IL-22 increased hCAP18 protein and mRNA. Treatment of stimulated NHEK cells with calcipotriol increased hCAP18 transcript expression further, while also decreasing transcript expression of IL-8, a neutrophil attractant. Furthermore, the researchers found that LL37 secretion into the medium of stimulated NHEK cells increased and calcipotriol treatment decreased this effect. Finally, the researchers examined hCAP18 and LL37 protein localization within NHEK cells. They found that cytokine-stimulated NHEK cells had increased hCAP18/LL37 expression, and while calcipotriol further increased hCAP18/LL37, LL37 was found within the cell, suggesting that calcipotriol prevents LL37 secretion. Together, the results of this study suggest that the therapeutic effects of calcipotriol stem from its ability to decrease IL-8 (thereby reducing neutrophil infiltration) and reducing LL37 release from stimulated keratinocytes.
Lifeline® Keratinocytes for Your Research Needs
Lifeline® keratinocytes are optimized for growth in DermaLife K® cell culture medium and are available from multiple primary sites:
Let us know how you are using Lifeline® cells or medium to answer your scientific questions and your study could be featured here on our blog!