Pericyte contractility in vitro: a physiological, morphological and biochemical study
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Abstract
Pericytes are cells of the microvascular wall that are thought to be contractile and thereby influence capillary hemodynamics and permeability. Investigations were conducted to characterize pericyte contractility in vitro. Contraction of collagen lattices containing bovine retinal pericytes (RP) , vascular smooth muscle cells (VSMC) , pulmonary microvessel endothelial cells (PMEC) , or aortic endothelial cells (AEC) was quantitated. The contractile activity of these cells was VSMC > RP > PMEC; AEC did not contract. A second assay involving contraction of a silicone rubber substratum was also used. The results were consistent with those obtained using the collagen lattice assay except that VSMC were less contractile than RP. Rhodamine phalloidin staining of contracting RP revealed microfilament bundle orientations that suggested their association with the applied contractile force. Also, RP contraction was inhibited by cytochalasin B. Contraction of all three cell types was stimulated by fetal calf serum. RP contraction was greater in calf serum than calf plasma-derived serum, indicating that RP respond to substances that appear continuously and episodically in blood. The contractile responses of RP to selected vasoactive agents were examined. Histamine and serotonin stimulated contraction, epinephrine had no effect, whereas isoproterenol elicited relaxation of RP. These effects were reversible. Since hormone-induced relaxation of smooth muscle involves cAMP, the modulation of RP contraction by cAMP was investigated. Dibutyryl cAMP (dBcAMP), forskolin, and 1-isobutyl-3-methylxanthine ( IBHX) induced RP relaxation. Relaxation was associated with elevated intracellular cAMP levels and stress fiber disassembly. Myosin light chain (MLC) phosphorylation, which is considered a prerequisite for contraction, is partly under the control of the adenylate cyclase system. Studies were performed to correlate MLC phosphorylation with the contractile state of RP and to define the role of cAMP in the relaxation event. Densitometry of 32P autoradiograms of myosin immunoprecipitated from pericytes revealed that the heavy and 20 kDa light chains of myosin were phosphorylated and that stimulation of RP with dBcAMP or forskolin resulted in a decrease in MLC phosphorylation. Collectively, these findings support the theory that pericytes are contractile, that their contraction can be regulated by vasoactive agonists, and that changes in cAMP , MLC phosphorylation, and stress fiber arrangement mediate the regulation.
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Dissertation (Ph.D.)--Boston University
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