Effect of insulin and growth hormone on the synthesis of radiolabelled proteoglycans from cultured human arterial smooth-muscle cells

Abstract

The present study focuses on the pathogenesis of increased frequency of large-vessel disease in diabetes. The diabetic arterial wall displays characteristic alterations of the extracellular matrix secreted by arterial smooth-muscle cells. The effects of insulin and growth hormone on the synthesis of sulphate-labelled proteoglycans and heparan sulphate proteoglycan were studied. Proteoglycans and heparan sulphate proteoglycan were obtained from the medium and the cell layer of cultured human arterial smooth-muscle cells grown in 5% human serum. Heparan sulphate proteoglycan was quantified using ethanol precipitation combined with specific enzyme degradation. Addition of insulin (0.01, 0.05 and 0.10 mU/ml) induced a significant accumulation of 35S-labelled proteoglycans in the cell layer (2p < 0.005 and 2p < 0.001). The relative amount of cell-associated heparan sulphate proteoglycan increased during insulin stimulation (2p < 0.05). Growth hormone stimulation (5.0 and 10.0 ng/ml) caused a significant decrease in the ratio between heparan sulphate proteoglycan and proteoglycan in the cell layer (2p < 0.02 and 2p < 0.01), whereas the distribution of proteoglycans between the cell layer and the medium was unaltered.

The SARS-CoV-2 Entry Inhibition Mechanisms of Serine Protease Inhibitors, OM-85, Heparin and Soluble HS Might Be Linked to HS Attachment Sites

“…Free D-xylose molecules significantly stimulate the biosynthesis of HS, CS, and DS [ 54 , 55 , 56 , 57 , 58 ]. Thus, insulin stimulates the biosynthesis of cell-layer GAGs, since it helps transport more D-xylose into the cell, which then stimulates the biosynthesis of GAGs, as confirmed by a previous study [ 59 ]. In addition, in 2009, Han et al [ 60 ], in a study on changes in HS of endothelial cells in diabetes, showed that high glucose or insulin levels alone reduced endothelial GAGs, but the presence of both simultaneously, i.e., hyperglycaemia + insulin, preserved GAGs [ 60 ].…”

Amphiregulin is expressed in primary cultures of prostate myofibroblasts, fibroblasts, epithelial cells, and in prostate tissue

“…Smooth muscle cells from the aorta and collagenase released human umbilical vein endothelial cells were grown as described. 16,17 All primary cell cultures were grown to semi-con¯uence. The medium was removed and the cells were washed with ice-cold phosphate buffered saline, and harvested by scraping off the cells.…”

“…The concentration and purity of the RNA preparations were analysed by measurement of the optical densities at wavelengths of 260 and 280 nm (Shimadzu, model UV-160A). 0.8 mg of the puri®ed RNA was used for cDNA synthesis in 20 mL of a buffer containing 50 mM KCl, 100 mM Tris-HCl (pH 8.3), 5 mM MgCl 2 , 1 unit/mL RNA H ase inhibitor, 1 mM of each of the four deoxynucleotides, 2.5 mM of a 16 mer d(T) 16 oligonucleotide primer, and 2.5 units/mL reverse transcriptase (all reaction components were from Perkin Elmer, Denmark). The reaction was incubated at 42 C for 30 min, and the cDNA was employed for the duplex PCR reactions.…”

Up-regulation of mitochondrial transcription factor 1 mRNA levels by GH in VSMC

“…Several lines of evidence suggest that GH may regulate the proliferation of VSMC (Bengtsson et al, 1988). However, the mechanism of GH action in VSMC remains unknown (Boger, 1999;Thogersen et al, 1996).…”