Phenotypic modulation of vascular smooth muscle in organ culture

Sammanfattning: Contraction of blood vessels for the regulation of blood flow and pressure is dependent on vascular smooth muscle cells (VSMC) located in the medial layer of the vascular wall. Adult, differentiated VSMC have a well developed contractile system and a low rate of proliferation. Under the influence of hormones, growth factors, inflammatory mediators and tissue factors, cells may modulate to a ”synthetic” phenotype, characterized by diminished contractility, a high rate of proliferation, and a tendency to migrate out of the organized tissue. Such cells are believed to form the ”neointima”, a thickening of the innermost layer of the wall characteristic of responses to endothelial injury, and important for the development of atherosclerotic lesions and of restenosis following vascular surgery. To study conditions of importance for maintaining the vessel wall in its contractile state, it is desirable to develop methods to keep blood vessels in organ culture, whereby tissue factors are preserved and experimental conditions can be well controlled. The aim of this thesis work was to study contractile and metabolic properties as well as biochemical composition and receptor expression in an arterial preparation (rat tail artery) kept in organ culture. After 4 days of culture arterial rings the morphology of the tissue was well preserved, but signs of phenotypic modulation were observed, particularly in the areas adjacent to the cut ends of the rings, where PDGF receptors appeared and the differentiation marker calponin was decreased. Culture with fetal calf serum (FCS) as growth promotor caused decreased contractility and ultrastructural signs of tissue damage. Detailed examination of this phenomenon disclosed that it is mainly caused by increased intracellular [Ca 2+ ]i , stimulated by vasoconstrictors present in FCS. Tight control of intracellular [Ca 2+ ]i is thus essential for tissue preservation. Cultured rings showed increased rate of lactate production but decreased O2 consumption, such that ATP production rate was the same as in fresh rings. Culture under hypoxia, however, caused reduced ATP production, suggesting decreased rate of energy consuming processes, such as protein turnover. Cultured rings maintained tension in hypoxic solution, in contrast to fresh rings. Reasons for this might be the increased reliance on glycolysis, and also an altered pattern of intracellular Ca 2+ waves, as revealed by laser scanning confocal microscopy. These results show that organ culture of vascular smooth muscle is useful for studying early events during phenotypic modulation in response to a variety of stimuli.