Cellular Dysfunction in the Diabetic Fibroblast

Abstract

Although it is known that systemic diseases such as diabetes result in impaired wound healing, the mechanism for this impairment is not understood. Because fibroblasts are essential for wound repair, we compared the in vitro behavior of fibroblasts cultured from diabetic, leptin receptor-deficient (db/db) mice with wild-type fibroblasts from mice of the same genetic background in processes important during tissue repair. Adult diabetic mouse fibroblast migration exhibited a 75% reduction in migration compared to normal fibroblasts (P < 0.001) and was not significantly stimulated by hypoxia (1% O(2)), whereas wild-type fibroblast migration was up-regulated nearly twofold in hypoxic conditions (P < 0.05). Diabetic fibroblasts produced twice the amount of pro-matrix metalloproteinase-9 as normal fibroblasts, as measured by both gelatin zymography and enzyme-linked immunosorbent assay (P < 0.05). Adult diabetic fibroblasts exhibited a sevenfold impairment in vascular endothelial growth factor (VEGF) production (4.5 +/- 1.3 pg/ml versus 34.8 +/- 3.3 pg/ml, P < 0.001) compared to wild-type fibroblasts. Moreover, wild-type fibroblast production of VEGF increased threefold in response to hypoxia, whereas diabetic fibroblast production of VEGF was not up-regulated in hypoxic conditions (P < 0.001). To address the question whether these differences resulted from chronic hyperglycemia or absence of the leptin receptor, fibroblasts were harvested from newborn db/db mice before the onset of diabetes (4 to 5 weeks old). These fibroblasts showed no impairments in VEGF production under basal or hypoxic conditions, confirming that the results from db/db fibroblasts in mature mice resulted from the diabetic state and were not because of alterations in the leptin-leptin receptor axis. Markers of cellular viability including proliferation and senescence were not significantly different between diabetic and wild-type fibroblasts. We conclude that, in vitro, diabetic fibroblasts show selective impairments in discrete cellular processes critical for tissue repair including cellular migration, VEGF production, and the response to hypoxia. The VEGF abnormalities developed concurrently with the onset of hyperglycemia and were not seen in normoglycemic, leptin receptor-deficient db/db mice. These observations support a role for fibroblast dysfunction in the impaired wound healing observed in human diabetics, and also suggest a mechanism for the poor clinical outcomes that occur after ischemic injury in diabetic patients.

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

“…Thus, this unique cohort with an extreme duration of disease and well-characterized micro-and macrovascular complications of diabetes enables analysis of the distinct contribution of each complication to fibroblast function and wound-healing efficiency. Our experiments confirmed previous studies showing that fibroblasts derived from individuals with diabetes migrate less in response to various growth factors including PDGF and insulin (37)(38)(39). Interestingly, fibroblasts from Medalists did not exhibit resistance to PDGF-BB ( Figure 5B), indicating that the inhibition of insulin actions by PKCδ was limited to selective signaling pathways.…”

“…Interestingly, fibroblasts from Medalists did not exhibit resistance to PDGF-BB ( Figure 5B), indicating that the inhibition of insulin actions by PKCδ was limited to selective signaling pathways. In addition, the ability of Medalist fibroblasts to express VEGF in response to insulin and hypoxia was decreased, confirming previous reports (37). Pulmonary analysis suggested that abnormalities in the fibroblasts from Medalists showed some correlation with a history of CVD and amputation, however, more detailed studies will be needed.…”

Differential Evaluation of Excisional Non-occluded Wound Healing in db/db Mice

“…Several studies reported in the literature have elucidated the function of various genes in the healing process, revealing the role of growth factors, cytokines and their downstream effectors in wound repair. The expression of platelet-derived growth factor (PDGFs) and its receptors is reduced in db/db mice (Beer et al 1997), as well as that of fibroblast growth factors (FGF) 1 and 2 (Werner et al 1994) and vascular endothelial growth factor (VEGF) (Lerman et al 2003). Also, Wetzler et al (2000) showed that prolonged persistence of neutrophils and macrophages in the wounds of db/db mice correlates with a large and sustained induction of the CC chemokine, macrophage chemoattractant protein (MCP-1/CCL2), and macrophage inflammatory protein 2 (MIP-2).…”

The Role of Myeloid Lineage Cells on Skin Healing and Skin Regeneration

“…In deficient healing, the prolonged inflammatory response causes insufficient re-epithelialization, a common outcome in diabetic patients and the elderly [24]. In diabetic ulcer wounds, there is a significant decrease in the TGF-β signaling and fibroblasts become unresponsive to growth factors thus reducing their migration to the wound bed [25][26][27][28]. The persistence of pro-inflammatory cytokines such as TNF-α and IL-1β further inhibits the proliferation and migration of fibroblasts [29].…”