Studies on the growth of smooth muscle from the airways of hyperresponsive rats

The aim of this work was to study proliferative growth of airway smooth muscle (ASM) cells from Fisher rats, a highly inbred strain known to have an increased amount of ASM in vivo, and airway hyperresponsiveness, when compared to control Lewis rats. Airway hyperresponsiveness is a hallmark of asthma, and ASM provides the contractile force which narrows or closes the airways during asthmatic episodes. Growth of ASM in asthma and animal models of asthma is well documented, and is likely to be a crucial determinant of an excessive airway response to contractile agonists. However, the mechanisms of ASM growth in asthma are unknown. Because ASM cells proliferate when exposed to inflammatory mediators, ASM growth in asthma has been assumed to be a sequel of inflammation of the airways. This thesis now presents data supporting an alternative hypothesis for the evolution of increased smooth muscle mass in asthma. Studies on highly inbred rats suggest that enhanced responsiveness of ASM cells to mitogens is a genetically determined phenotype, and may thus predispose to an increased mass and contractility of ASM in vivo. Thus, an inborn susceptibility for enhanced ASM growth may lead to airway hyperresponsiveness. The proliferative response to mitogenic stimulation in ASM cells from hyperresponsive Fisher rats was enhanced when compared to the response of ASM cells from control Lewis rats. This enhancement was inherent to ASM cells, persisting over multiple passages in culture, and the mechanisms responsible were operative in G₀ or G₁ of the cell cycle. An enhanced proliferative response of Fisher ASM cells was selectively induced by a subset of the growth factors examined. These included fetal bovine serum (FBS), epidermal growth factor (EGF), and the AA or AB isoforms of platelet-derived growth factor (PDGF). The growth response of Fisher ASM cells to other mitogens tested was not enhanced with respect to Lewis ASM cells, indicating that the mechanisms responsible for enhanced Fisher ASM cell growth are not at a point in growth signalling pathways which is converged upon by all mitogens. The data suggest that several mechanisms can be ruled out as potential explanations of enhanced Fisher ASM cell growth. These include differences in basal levels of mitogen activated protein (MAP) kinases or PDGF receptors, as well as differences in phospholipase C (PLC)-mediated growth signals, intracellular calcium responses, and tyrosine phosphorylation of MAP kinases, PDGF receptors, or a number of other cellular proteins. Instead it appears that enhanced Fisher ASM cell growth is mediated by phorbol ester-sensitive isoforms, of the serine/threonine specific protein kinase C (PKC), and is associated with enhanced serine/threonine phosphorylation of a 115 kD protein. These findings lay the groundwork for characterizing a model of genetic predisposition to enhanced ASM growth. This model may prove a valuable tool for elucidating the mechanisms of ASM growth, and may thus contribute to the development of new strategies for the treatment of asthma.