Phenotype modulation in primary cultures of arterial smooth-muscle cells. Dual effect of prostaglandin E1

The effects of prostaglandin E1 (PGE1) on the phenotypic state of enzymatically isolated arterial smooth-muscle cells in primary culture were studied by transmission electron microscopy, thymidine autoradiography, and cell counting. Early in culture (day 0-2), PGE1 stimulated conversion of the cells from contractile (less euchromatic nucleus and cytoplasm dominated by myofilament bundles) to synthetic state (more euchromatic nucleus and cytoplasm dominated by cisternae of rough endoplasmic reticulum and a large Golgi complex). The rate of entrance of the cells into DNA synthesis and mitosis was also increased at this time. Later on (day 3-6), when the majority of the cells had entered synthetic state, PGE1 inhibited DNA synthesis and cellular proliferation. These observations indicate that the effect of prostaglandins on arterial smooth muscle is dual in nature and dependent on the state of differentiation of the cells.     

The influence of alkyl pyridinium sponge toxins on membrane properties, cytotoxicity, transfection and protein expression in mammalian cells

The ability of two alkyl pyridinium sponge toxin preparations (poly-APS and halitoxin) to form transient pores/lesions in cell membranes and allow transfection of plasmid cDNA have been investigated using HEK 293 cells. Poly-APS and halitoxin preparations caused a collapse in membrane potential, reductions in input resistance and increased Ca²⁺ permeability. At least partial recovery was observed after poly-APS application but recovery was more rarely seen with halitoxin. The transfection with plasmid cDNAs for an enhanced green fluorescent protein (EGFP) and human tumour necrosis factor receptor 2 (TNFR2) was assessed for both toxin preparations and compared with lipofectamine. Stable transfection was achieved with poly-APS although it was less efficient than lipofectamine. These results show that viable cells transfected with alien cDNA can be obtained using novel transient pore-forming alkyl pyridinium sponge toxins and a simple pre-incubation protocol. This provides the first proof of principle that pore-forming alkyl pyridinium compounds can be used to deliver cDNA to the intracellular environment without permanently compromising the plasma membrane.     

In vivo protein synthesis from 14C-substrates in porcine tissues during the transition to postnatal development

[2-14C] leucine, [1-14C] alanine, [1-14C] glucose, [1-14C] lactate and [1-14C] pyruvate utilization in the protein synthesis has been studied in vivo at early stages of postnatal development of piglets. It has been established, that during the first 24 hours after birth the protein synthesis intensity, judging by [2-14C] leucine incorporation, in liver, skeletal muscle, duodenal wall and subcutaneous tissue of piglets increases 5, 7, 6.5 and 2.1 times respectively. At the age of 1-2 h the radioactive carbon incorporation from [1-14C] glucose into the brain proteins is more pronounced than into the proteins of liver and skeletal muscle. During the first days of life the intensity of the label incorporation from [1-14C] glucose into liver and skeletal muscle proteins of piglets is enhanced, whereas in brain it remains at the same level. The degree of 14C carbon incorporation from [1-14C]-alanine, [1-14C] pyruvate and [1-14C] lactate into the liver and skeletal muscle proteins of 5-days-old piglets is approximately the same, 14C substrates of protein synthesis in brain and subcutaneous adipose tissue having some peculiarities.     

Density-dependent inhibitory effect of transforming growth factor-β on human fibroblasts involves the down-regulation of platelet-derived growth factor α-receptors

We have previously found that transforming growth factor-β1 (TGF-β1) inhibits the mitogenic activity of platelet-derived growth factor (PDGF) in cultures of human neonatal fibroblasts in a density-dependent fashion. In the present investigation we determined the effect of TGF-β1 on the PDGF α-receptor, which binds all PDGF isoforms, as well as on the β-receptor, which binds only PDGF-BB with high affinity. We found that the inhibitory effect of TGF-β1 on PDGF-AA-induced mitogenesis was density-dependent; when dense cell cultures were preincubated with TGF-β1, there was an complete inhibition of ³H-thymidine incorporation, whereas the effect was less in sparse cultures. A similar density-dependent effect of TGF-β1 was seen in PDGF-BB treated cells, although less pronounced. The binding of ¹²⁵I-labeled PDGF-AA and PDGF-BB to the α-receptor was significantly reduced after treatment with TGF-β1 in dense cultures, whereas the sparse cultures were less affected. A decrease of α-receptor mRNA was also seen. The levels of β-receptor protein and mRNA were unaffected. We conclude that the growth inhibitory effect of TGF-β1 is cell density-dependent and involves down-regulation of PDGF α-receptors. © 1993 Wiley-Liss, Inc.     

Effect of some antidiuretic agents on the polyuria-polydipsia provoked by alcohol]

The authors have studied in the dog--normal or diabetic insipidus after hypothalamic lesion--submitted to oral ingestion of ethanol the effects of a few antidiuretic drug: chlorpropamid, atromid-S, carbamazepin and carisoprodol. The carisoprodol only can reduce the diuresis and the drinking provoked by ethanol.