Browsing by Subject "keratinocyte"

Now showing 1 - 3 of 3
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    Adipose stromal cells enhance keratinocyte survival and migration in vitro, and graft revascularization in mouse wound healing model
    (2013-12-11) Knowles, Kellen Alexander; Berbari, Edward J.; March, Keith Leonard, 1963-; Ji, Julie
    In the US, more than 1 million burn injuries are reported annually. About 45,000 injuries due to fires and burns result in hospitalization and ten percent of these result in death every year. Advances in burn treatment have led to a reduction in mortality rate over the last decades. Since more patients are surviving the initial resuscitation phase even with very large areas of skin being burned away, wound care has become increasingly important to ensure continued patient survival and improvement. While currently a common treatment for third degree burn wounds, skin grafts have several drawbacks. The availability of donor sites for autografts may be limited, especially in incidences of extensive skin loss. The rejection associated with the use of allografts and xenografts may render them inadequate or undesirable. Even if a suitable graft is found, poor retention due to infection, hematoma, and low vascularity at the recipient site are other drawbacks associated with the use of skin grafts as a primary treatment for severe burn wounds. As such, research has been done into alternative treatments, which include but are not limited to artificial skin, cell therapy, and growth factor application. We propose the delivery of adipose derived stem cells (ASC) in combination with endothelial progenitor cells (EC) via Integra Dermal Regenerative Template (DRT) to promote faster graft vascularization and thus faster healing of wounds. Integra DRT is an acellular skin substitute that consists of a dermal layer composed of bovine collagen and chondroitin-6-sulfate glycosaminoglycan, and an "epidermal" layer, which consists of silicone polymer. This silicone layer is removed after the collagen matrix is adequately vascularized (usually takes 2-3 weeks), and then a thin layer autograft is applied to the top of the neo-dermis. ASC are derived from the stromal-vascular fraction (SVF) of adipose tissue and are a readily available, pluripotent, mesenchymal cell known to promote angiogenesis. They are being explored as a treatment for a myriad of diseases and conditions, including wound healing. In combination with ECs, they form stable microvessel networks in vitro and in vivo. In our work, we found that ASC+EC form stable microvessel networks when cultured on Integra DRT. Also, ASC and ASC+EC conditioned media promoted both survival and migration of human epidermal keratinocytes compared to control medium. In a full thickness wound healing model, using healthy NSG mice, the ASC+EC case showed a significantly higher rate of wound closure compared to control. Based on best linear unbiased estimates (BLUE), the difference between the healing rates of ASC alone treatment and the Control treatment group is -0.45 +/- 0.22 mm²/day (p=0.041), which is not less than 0.025 and thus not statistically significant (Bonferroni Adjusted). However, the BLUE for the difference between the ASC+EC group and the Control group healing rates is -0.55 +/- 0.28 mm²/day (p = 0.017<0.025, Bonferroni Adjusted), which is statistically significant. Histology revealed a significantly higher number of vessels compared to control in both ASC alone and ASC+EC case. CD31 staining revealed the presence of human vessels in ASC+EC treatment scaffolds. We conclude that the combination of ASC and EC can be used to accelerate healing of full-thickness wounds when delivered to site of the wound via Integra. This result is especially compelling due to the fact that the mice used were all healthy. Thus our treatment shows an improvement in healing rate even compared to normal wound healing.
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    Human keratinocyte differentiation requires translational control by the eIF2α kinase GCN2
    (Elsevier, 2017) Collier, Ann E.; Wek, Ronald C.; Spandau, Dan F.; Department of Dermatology, IU School of Medicine
    Appropriate and sequential differentiation of keratinocytes is essential for all functions of the human epidermis. While transcriptional regulation has proven to be important for keratinocyte differentiation, little is known about the role of translational control. A key mechanism for modulating translation is through phosphorylation of the α subunit of eIF2. A family of different eIF2α kinases function in the integrative stress response to inhibit general protein synthesis coincident with preferential translation of select mRNAs that participate in stress alleviation. Here we demonstrate that translational control through eIF2α phosphorylation is required for normal keratinocyte differentiation. Analyses of polysome profiles revealed that key differentiation genes, including involucrin, are bound to heavy polysomes during differentiation, despite decreased general protein synthesis. Induced eIF2α phosphorylation by the GCN2 protein kinase facilitated translational control and differentiation-specific protein expression during keratinocyte differentiation. Furthermore, loss of GCN2 thwarted translational control, normal epidermal differentiation, and differentiation gene expression in organotypic skin culture. These findings underscore a previously unknown function for GCN2 phosphorylation of eIF2α and translational control in the formation of an intact human epidermis.
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    Mechanisms for Alternaria alternata Function in the Skin During Induction of Peanut Allergy in Neonatal Mice With Skin Barrier Mutations
    (Frontiers, 2021-09) Buelow, Lauren M.; Hoji, Akihiko; Tat, Kiet; Schroeder-Carter, Lindsay M.; Carroll, Daniela J.; Cook-Mills, Joan M.; Pediatrics, School of Medicine
    Neonatal mice with heterozygous mutations in genes encoding the skin barrier proteins filaggrin and mattrin (flaky tail mice [FT+/−]) exhibit oral peanut-induced anaphylaxis after skin sensitization. As we have previously reported, sensitization in this model is achieved via skin co- exposure to the environmental allergen Alternaria alternata (Alt), peanut extract (PNE), and detergent. However, the function of Alt in initiation of peanut allergy in this model is little understood. The purpose of this study was to investigate candidate cytokines induced by Alt in the skin and determine the role of these cytokines in the development of food allergy, namely oncostatin M (Osm), amphiregulin (Areg), and IL-33. RT-qPCR analyses demonstrated that skin of FT+/− neonates expressed Il33 and Osm following Alt or Alt/PNE but not PNE exposure. By contrast, expression of Areg was induced by either Alt, PNE, or Alt/PNE sensitization in FT+/− neonates. In scRNAseq analyses, Osm, Areg, and Il33 were expressed by several cell types, including a keratinocyte cluster that was expanded in the skin of Alt/PNE-exposed FT+/− pups as compared to Alt/PNE-exposed WT pups. Areg and OSM were required for oral PNE-induced anaphylaxis since anaphylaxis was inhibited by administration of neutralizing anti-Areg or anti-OSM antibodies prior to each skin sensitization with Alt/PNE. It was then determined if intradermal injection of recombinant IL33 (rIL33), rAreg, or rOSM in the skin could substitute for Alt during skin sensitization to PNE. PNE skin sensitization with intradermal rIL33 was sufficient for oral PNE-induced anaphylaxis, whereas skin sensitization with intradermal rAreg or rOSM during skin exposure to PNE was not sufficient for anaphylaxis to oral PNE challenge. Based on these studies a pathway for IL33, Areg and OSM in Alt/PNE sensitized FT+/− skin was defined for IgE induction and anaphylaxis. Alt stimulated two pathways, an IL33 pathway and a pathway involving OSM and Areg. These two pathways acted in concert with PNE to induce food allergy in pups with skin barrier mutations.