Abnormalities in Proliferation and Protein Synthesis in Skin Fibroblast Cultures from Patients with Diabetes Mellitus

Abstract

Several aspects of in-vitro cell growth and protein synthesis were assessed in cultures of skin fibroblasts from subjects with juvenile-onset diabetes mellitus (JODM) or adult-onset diabetes mellitus (AODM) and from age-matched nondiabetic controls (C). There was an inverse correlation between increasing age and both the log-phase doubling rate and saturation density at confluence in C fibroblasts. JODM and AODM cells had a reduction in both indices of cell population growth in comparison with age-matched C fibroblasts. Fibroblasts grown in the presence of 0.3 micronM hydrocortisone were stimulated to grow more rapidly and to a greater saturation density. Stimulation of cell division by hydrocortisone accentuated the abnormalities in growth of JODM and AODM fibroblasts. Total protein and collagen synthesis was measured whtn the fibroblasts had grown to confluency in medium with or without hydrocorticone. Hydrocorticone did not produce a significant change in total protein and collagen synthesis per cell by C fibroblasts. Fibroblasts from AODM had a 180 per cent increase in total protein and collagen synthesis in the presence of hydrocortisone. In contrast, total protein and collagen synthesis decreased 40 per cent in fibroblasts from JODM when grown in the hydrocortisone medium. These studies indicate that skin fibroblast cultures from patients with diabetes exhibit abnormalities in cell proliferation. Furthermore, hydrocortisone appears to unmask diffeerences in protein synthesis that distinguish JODM and AODM fibroblasts in culture.

Insulin-stimulated glucose uptake, leucine incorporation into protein, and uridine incorporation into RNA in skin fibroblast cultures from patients with diabetes mellitus

“…The other significant difference, IDD vs. NIDD, occurred only at insulin 4000 ng/ml. Alterations in total protein and collagen synthesis have already been defined in fibroblasts from diabetic donors in a prior study in which basal protein and collagen synthesis rates per cell were greater in IDD than NIDD cells [6]. The addition of hydrocortisone caused a decrease in synthesis by IDD cells and an increase by NIDD cells [6].…”

“…Previously demonstrated abnormalities in plating efficiency, growth rate, population density at confluence, replicative lifespan and protein synthesis in fibroblasts from patients with diabetes mellitus suggest that intrinsic cellular abnormalities are present [1][2][3][4][5][6][7]. It was, therefore, reasoned that if differences in insulin action were demonstrable in fibroblasts from patients with diabetes, this would represent further evidence for intrinsic cellular changes in diabetes.…”

“…Fibroblasts were established in tissue culture from minced skin biopsies taken from the deltoid skin of human donors, using previously reported methods [6,12]. All studies were performed on cells that had been stored in liquid nitrogen and had not undergone more than 17 in vitro passages (one passage = 1 to 2 split = one population doubling).…”

“…These results provide additional evidence for intrinsic cellular abnormalities in diabetes mellitus. Whether the differences in basal or insulin-stimulated response between fibroblasts of different donor types are attributable to alterations in protein or RNA synthesis, metabolite pool size or turnover have yet to be determined.Key words: Diabetes mellitus, insulin-dependent diabetes, non-insulin-dependent diabetes, human fibroblasts, insulin, glucose uptake, RNA, protein, cellular abnormalities, cell culture, uridine incorporation, leucine incorporation Alterations in plating efficiency, growth rate, population density at confluence, replicative lifespan and protein synthesis [1][2][3][4][5][6][7] in fibroblast cultures established from diabetic patients suggest that intrinsic cellular abnormalities do occur in diabetes. These changes are said to resemble aging and, therefore, may have pathogenic relevance to the complications of diabetes [2].…”

“…Alterations in plating efficiency, growth rate, population density at confluence, replicative lifespan and protein synthesis [1][2][3][4][5][6][7] in fibroblast cultures established from diabetic patients suggest that intrinsic cellular abnormalities do occur in diabetes. These changes are said to resemble aging and, therefore, may have pathogenic relevance to the complications of diabetes [2].…”