High glucose inhibits human epidermal keratinocyte proliferation for cellular studies on diabetes mellitus

Abstract

In order to more clarify the delayed wound healing in diabetes mellitus, we cultured the human epidermal keratinocytes in both 6 mM (control group) and 12 mM glucose (high-glucose group) of "complete" MCDB 153 medium. Hyperglycaemia slowed the rate of their proliferation and inhibited their DNA synthesis and the production of total proteins. By 1 month after primary seeding in high-glucose group, the cells ceased their proliferation, whereas the cells in control group grew for more than 40 days. Mean population doublings in high-glucose group was 5.27 (vs. 7.25 in control, P = 0.001), and mean population doubling time during 1 month in high glucose group was 5.43 days (vs. 3.65 days in control, P = 0.02). They indicate that prolonged exposure to high glucose decreases the replicative life span of human epidermal keratinocytes in vitro. Furthermore, analysis of fatty acid contents in membrane phospholipids with thin-layer and gas chromatography showed no difference between the cultured keratinocytes in both conditions. Immunocytochemical staining of glucose transporter 1 shows that 28.1% of cells in high-glucose group were almost twice positive of those in control group (13.2%, P = 0.008). The mechanism of the ill effects of high glucose on epidermal keratinocytes is not so far clear, but it indicates the possibility of any direct effect of hyperglycaemia on glucose metabolism without changing lipid metabolism on cell membrane. The high-glucose group presented in this report can be available as an in vitro valuable study model of skin epidermal condition on diabetes mellitus.

Ganglioside GM3 Depletion Reverses Impaired Wound Healing in Diabetic Mice by Activating IGF-1 and Insulin Receptors

“…Pre-incubation of WT primary mouse KCs in 20 mM glucose for 72 h, traditionally used to simulate the hyperglycemic diabetic environment in vitro (Ingram et al , 2008; Suzuki et al , 2011), stopped migration in in vitro wounds (Figure 3a and b) and reduced cell proliferation (Figure 3c) (Lan et al , 2008; Spravchikov et al , 2001; Terashi et al , 2005). GM3S −/− cells migrated faster than WT cells by 24 h after wounding in normoglycemic conditions.…”

Peripheral Blood Fibrocytes

“…19 Prior studies revealed that chronic wound states in diabetes are related to the deficiency of functional stem cells, dysfunctional cellular response, and insufficient cell proliferation at the injury site, in which diabetic fibroblasts show selective impairments in discrete cellular processes critical for tissue repair including cellular migration, VEGF production, and the response to hypoxia. [23][24][25][26][27][28][29] In the genetically diabetic mice experimental model, the tail vein-injected fibrocytes was effective in improving wound closure. It was probably that the injected fibrocytes migrated into the wounds and recruited to compensate the dysfunctional resident dermal fibroblasts; in addition, it appeared to provide an additional, renewable source of functional progenitor cells to promote wound healing.…”

Human Adipose-Derived Stromal Cells Accelerate Diabetic Wound Healing: Impact of Cell Formulation and Delivery

“…For example, both fibroblasts and endothelial progenitor cells from diabetic ulcers have been shown to exhibit impaired migration, proliferation, adhesion, and growth factor production. [4][5][6] This has led some to propose the application of fresh, unimpaired cells to nonhealing wounds as a means to reinitiate or replace the host tissue's wound-healing cascade. 7 Emerging work suggests that adipose-derived stromal cells (ASCs) may be useful for therapeutic wound-healing applications.…”