PKCδ inhibition normalizes the wound-healing capacity of diabetic human fibroblasts
Mogher Khamaisi, … , Amy Wagers, George L. King
Mogher Khamaisi, … , Amy Wagers, George L. King
Published January 25, 2016
Citation Information: J Clin Invest. 2016;126(3):837-853. https://doi.org/10.1172/JCI82788.
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Research Article Endocrinology

PKCδ inhibition normalizes the wound-healing capacity of diabetic human fibroblasts

Abstract

Abnormal fibroblast function underlies poor wound healing in patients with diabetes; however, the mechanisms that impair wound healing are poorly defined. Here, we evaluated fibroblasts from individuals who had type 1 diabetes (T1D) for 50 years or more (Medalists, n = 26) and from age-matched controls (n = 7). Compared with those from controls, Medalist fibroblasts demonstrated a reduced migration response to insulin, lower VEGF expression, and less phosphorylated AKT (p-AKT), but not p-ERK, activation. Medalist fibroblasts were also functionally less effective at wound closure in nude mice. Activation of the δ isoform of protein kinase C (PKCδ) was increased in postmortem fibroblasts from Medalists, fibroblasts from living T1D subjects, biopsies of active wounds of living T1D subjects, and granulation tissues from mice with streptozotocin-induced diabetes. Diabetes-induced PKCD mRNA expression was related to a 2-fold increase in the mRNA half-life. Pharmacologic inhibition and siRNA-mediated knockdown of PKCδ or expression of a dominant-negative isoform restored insulin signaling of p-AKT and VEGF expression in vitro and improved wound healing in vivo. Additionally, increasing PKCδ expression in control fibroblasts produced the same abnormalities as those seen in Medalist fibroblasts. Our results indicate that persistent PKCδ elevation in fibroblasts from diabetic patients inhibits insulin signaling and function to impair wound healing and suggest PKCδ inhibition as a potential therapy to improve wound healing in diabetic patients.

Authors

Mogher Khamaisi, Sayaka Katagiri, Hillary Keenan, Kyoungmin Park, Yasutaka Maeda, Qian Li, Weier Qi, Thomas Thomou, Danielle Eschuk, Ana Tellechea, Aris Veves, Chenyu Huang, Dennis Paul Orgill, Amy Wagers, George L. King

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Figure 3

Medalist fibroblasts display impaired wound healing in vivo.

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Medalist fibroblasts display impaired wound healing in vivo. (A) Macrosc...
(A) Macroscopic images of wound area surface not covered by an epithelial layer in wounds covered with Integra without human cells, Integra with control cells, or Integra with Medalist fibroblasts. (B) Percentage of open wound areas on days 9 and 15 of the initial wound area. (C) H&E-stained sections of open wound area and granulation tissues on day 15 after initial wounding. D, dermis; E, epidermis. Scale bar: 50 μm. (D) Representative immunoblots of VEGF protein levels and quantification (right panel) in the granulation tissues on day 15 after wounding. Data represent the mean ± SD. n = 12 for wounds treated with Integra without cells; n = 7 for wounds treated with control fibroblasts; and n = 12 for wounds treated with Medalist fibroblasts. The criteria for selecting the cell lines for these experiments were completely random, and the clinical and demographic characteristics of the selected subjects did not differ from those of the rest of the patients. A Student’s t or χ2 test was used for 2-way comparisons based on the distribution and number of observations of the variable.