Premature senescence of skin fibroblasts from insulin-dependent diabetic patients with kidney disease

Abstract

Diabetic glomerulopathy develops in a subset only of patients with insulin-dependent diabetes (IDDM) and early, in its course, is characterized by cell hypertrophy and by excessive extracellular matrix production. These observations suggest that an alteration in the control of cell growth processes may contribute to its pathogenesis and be related to the susceptibility to kidney disease. We therefore investigated whether the development of diabetic nephropathy is associated with abnormalities of cell growth and morphology. Cultured skin fibroblasts from 14 IDDM patients with nephropathy (DN) were compared with those of 10 IDDM patients without nephropathy (D) and of 14 control non-diabetic subjects (C). Cell volume (in arbitrary units) and total protein content (microgram/10, 000 cells) were increased in serially passaged skin fibroblasts of IDDM patients with nephropathy (DN = 809.5 +/- 33.1 and 1.93 +/- 0.38 vs. D = 764.4 +/- 31.5 and 1.5 +/- 0.37, P = 0.005 and P = 0.03, respectively; vs. C = 756.2 +/- 36.3 and 1.5 +/- 0.38, P = 0.0006 and P = 0.03, respectively). These hypertrophic cells had a tendency to a slower duplication rate and exhibited a dissociation of the DNA and cytoplasmic cell-cycles, resulting in a higher proportion of tetraploid cells (DN = 25 +/- 15% vs. D = 6 +/- 4%, P = 0.005; and vs. C = 10 +/- 8%, P = 0.04). The frequency of terminally differentiated post-mitotic fibrocytes, cells specialized for extracellular matrix production, was higher in patients with nephropathy compared to that of patients without nephropathy and normal controls (DN = 34 +/- 14% vs. D = 21 +/- 10%, P = 0.02; and vs. C = 19 +/- 12%, P = 0.008). That early differentiation was a specific feature of cells derived from patients with diabetic nephropathy was confirmed by the study of cell life-span which demonstrated that these cells aged prematurely (log rank test, chi 2 = 10,012; P = 0.0067). We conclude that an acceleration of cell aging is a peculiar feature of diabetic kidney disease and may contribute to its pathological tissue changes.

Premature cell ageing and evolution of diabetic nephropathy

“…These results confirm our previous observations [3], that patients with diabetic nephropathy, as a group, show features of early cell ageing, which seem to be independent of the diabetic state. Moreover these data suggest that, in addition to the degree of hyperglycaemia, early cell differentiation is associated with a faster onset of proteinuria and a more rapid decline of renal function in IDDM patients.…”

“…The assumption that abnormalities measured in the skin fibroblasts may apply to other cell types is speculative, but there are clear similarities between cell types of mesodermic origin, to which mesangial cells, renal and skin fibroblasts belong, in their function of producing extracellular matrix. Skin fibroblasts have proved useful as a model to explain cell functional abnormalities of other cell types in a variety of metabolic and genetic disorders [3,10]. The degree of cellular abnormalities is unlikely to be secondary to the degree of renal failure in that we found no correlation between the actual level of uraemia or GFR and cellular changes [3] and have observed these cell abnormalities in patients with microalbuminuria who have normal renal function (unpublished observation).…”

“…The reasons for these differences in the evolution and progression of diabetic kidney disease are poorly understood, even though a number of environmental factors, including levels of blood pressure [6], blood glucose [7], serum cholesterol [8] and age of onset of diabetes have been implicated. At the cellular level early cell differentiation could be involved [3]. In this study we have compared cell volume and the percentage of PMF in IDDM patients with and without nephropathy.…”

“…Fibroblasts were obtained from forearm skin biopsies and experiments conducted between the 6 th and 8 th passage. Cell volume in arbitrary units was measured on quiescent fibroblasts by flow cytometry [3]. The percentage of terminally differentiated PMF was evaluated on sparse mass cell cultures, as previously described, and scored by one of us, H. P. R., who was unaware of the patients' clinical details [3].…”

“…We have recently described that serially cultured skin fibroblasts from IDDM patients with nephropathy exhibit hypertrophy, delayed replication and an increased percentage of terminally differentiated post-mitotic fibrocytes (PMF) [3], cells specialized for the production of extracellular matrix components. Diabetologia (1997) Summary The rate of development and progression of renal disease varies greatly in insulin-dependent diabetic (IDDM) patients.…”