Abnormal cytoskeletal protein expression in cultured skin fibroblasts from type 1 diabetes mellitus patients with nephropathy: A proteomic approach

Abstract

Diabetic nephropathy (DN) develops in about 40% of insulin-dependent type 1 diabetes mellitus (T1DM) patients, and is associated not only with diabetes duration and metabolic control, but also with a genetic predisposition. Constitutive alterations of cytoskeletal proteins may play a role in the development of DN. We investigated the expression of these proteins in cultured skin fibroblasts, obtained from long-term T1DM patients with and without DN but comparable metabolic control, and from matched healthy subjects, by means of 2-DE electrophoresis and MS-MALDI analyses. In T1DM with DN, compared to the other two groups, quantitative analyses revealed an altered expression of 17 spots (p<0.05-p<0.01), corresponding to 12 unique proteins. In T1DM with DN, beta-actin and three isoforms of tubulin beta-2 chain, tropomodulin-3, and LASP-1 were decreased, whereas two tubulin beta-4 chain isoforms, one alpha actinin-4 isoform, membrane-organizing extension spike protein (MOESIN), FLJ00279 (corresponding to a fragment of myosin heavy chain, non-muscle type A), vinculin, a tropomyosin isoform, and the macrophage capping protein were increased. A shift in caldesmon isoforms was also detected. These results demonstrate an association between DN and the constitutive expression of cytoskeleton proteins in cultured skin fibroblasts from T1DM with DN, which may retain pathophysiologycal implications.

Związek polimorfizmu rs 3807337 genu CALD1 z nefropatią cukrzycową w przebiegu cukrzycy typu 1 — wstępne wyniki badania rodzin

“…We found only one study (a casecontrol one), performed by Conway et al, examining the association of polymorphism of this candidate gene with diabetic nephropathy development in type 1 diabetes, and our family-based association study confirms their findings [17]. Evidence also exists that there are some proteins differentially expressed in cultured fibroblast in patients with type 1 diabetes with established nephropathy, compared to the ones without that complication and, typically, altered protein classes consist of the actin-associated protein, including caldesmon [21]. This information is consistent with the results of in vivo and in vitro studies pointing out the importance of genes that regulate the actin filament organisation in diabetic nephropathy development.…”

Improving Power of Genome-Wide Association Studies with Weighted False Discovery Rate Control and Prioritized Subset Analysis

“…In this way, we obtained an estimated permutation-based FDR [18] at 3.64%, still less than the FDR control level of 5%. In fact, several studies published later than the WTCCC paper [17] also supported the association of these three genes with T1D ( INS [19]–[22], CALD1 [23], [24], and PD-1 [25], [26]). …”

Skin fibroblasts as a tool for identifying the risk of nephropathy in the type 1 diabetic population

“…Protein identification was performed by matrix‐assisted laser desorption/ionization mass spectrometry. We showed that several proteins are differentially expressed between cultured skin fibroblasts from T1DM patients with and without established diabetic nephropathy 55–57, evidencing an association between diabetic nephropathy and the constitutive expression of some proteins, thus suggesting their involvement in the development of diabetic nephropathy. We showed alterations in the abundance of nine molecular chaperones controlling protein turnover and folding in cultured skin fibroblasts from T1DM patients with diabetic nephropathy 57.…”

“…Recently, we performed a comparative proteomic analysis of cultured skin fibroblasts from T1DM patients with and without diabetic nephropathy and from healthy controls 15, 55–58. Proteins were separated by two‐dimensional electrophoresis and the gel images were qualitatively and quantitatively analysed.…”

“…Most structural changes observed in diabetic nephropathy, such as increased mesangial matrix accumulation, tubular damage and regeneration, and interstitial fibrosis, have been associated with the de novo expression of a wide range of cytoskeletal and cytoskeleton‐related proteins, including a‐smooth muscle actin and the intermediate filament protein vimentin 59. Using cultured skin fibroblasts, we also identified 12 cytoskeletal‐related proteins the expression of which was different in cells from patients with diabetic nephropathy with respect to those from T1DM patients without diabetic nephropathy and controls 56. Yip et al 60 reported that T1DM patients with diabetic nephropathy were more insulin resistant than T1DM patients without diabetic nephropathy who were matched for age and duration of diabetes.…”