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Disruption of actin cytoskeleton mediates loss of tensile stress induced early phenotypic modulation of vascular smooth muscle cells in organ culture.

ABSTRACT: Aorta organ culture has been widely used as an ex vivo model for studying vessel pathophysiology. Recent studies show that the vascular smooth muscle cells (VSMCs) in organ culture undergo drastic dedifferentiation within the first few hours (termed early phenotypic modulation). Loss of tensile stress to which aorta is subject in vivo is the cause of this early phenotypic modulation. However, no underlying molecular mechanism has been discovered thus far. The purpose of the present study is to identify intracellular signals involved in the early phenotypic modulation of VSMC in organ culture. We find that the drastic VSMC dedifferentiation is accompanied by accelerated actin cytoskeleton dynamics and downregulation of SRF and myocardin. Among the variety of signal pathways examined, increasing actin polymerization by jasplakinolide is the only one hindering VSMC dedifferentiation in organ culture. Moreover, jasplakinolide reverses actin dynamics during organ culture. Latrunculin B (disrupting actin cytoskeleton) and jasplakinolide respectively suppressed and enhanced the expression of VSMC markers, SRF, myocardin, and CArG-box-mediated SMC promoters in PAC1, a VSMC line. These results identify actin cytoskeleton degradation as a major intracellular signal for loss of tensile stress-induced early phenotypic modulation of VSMC in organ culture. This study suggests that disrupting actin cytoskeleton integrity may contribute to the pathogenesis of vascular diseases.

SUBMITTER: Zheng JP

PROVIDER: S-EPMC2815224 | biostudies-literature | 2010 Feb

REPOSITORIES: biostudies-literature

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Disruption of actin cytoskeleton mediates loss of tensile stress induced early phenotypic modulation of vascular smooth muscle cells in organ culture.

Zheng Jian-Pu JP Ju Donghong D Shen Jianbin J Yang Maozhou M Li Li L

Experimental and molecular pathology 20091027 1

Aorta organ culture has been widely used as an ex vivo model for studying vessel pathophysiology. Recent studies show that the vascular smooth muscle cells (VSMCs) in organ culture undergo drastic dedifferentiation within the first few hours (termed early phenotypic modulation). Loss of tensile stress to which aorta is subject in vivo is the cause of this early phenotypic modulation. However, no underlying molecular mechanism has been discovered thus far. The purpose of the present study is to i ...[more]

PMID: 19874818

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Project description:Vascular smooth muscle cell (SMC) switching between differentiated and dedifferentiated phenotypes is reversible and accompanied by morphological and functional alterations that require reconfiguration of cell-cell and cell-matrix adhesion networks. Studies attempting to explore changes in overall composition of the adhesion nexus during SMC phenotype transition are lacking. We have previously demonstrated that T-cadherin knockdown enforces SMC differentiation, whereas T-cadherin upregulation promotes SMC dedifferentiation. This study used human aortic SMCs ectopically modified with respect to T-cadherin expression to characterize phenotype-associated cell-matrix adhesion molecule expression, focal adhesions configuration and migration modes. Compared with dedifferentiated/migratory SMCs (expressing T-cadherin), the differentiated/contractile SMCs (T-cadherin-deficient) exhibited increased adhesion to several extracellular matrix substrata, decreased expression of several integrins, matrix metalloproteinases and collagens, and also distinct focal adhesion, adherens junction and intracellular tension network configurations. Differentiated and dedifferentiated phenotypes displayed distinct migrational velocity and directional persistence. The restricted migration efficiency of the differentiated phenotype was fully overcome by reducing actin polymerization with ROCK inhibitor Y-27632 whereas myosin II inhibitor blebbistatin was less effective. Migration efficiency of the dedifferentiated phenotype was diminished by promoting actin polymerization with lysophosphatidic acid. These findings held true in both 2D-monolayer and 3D-spheroid migration models. Thus, our data suggest that despite global differences in the cell adhesion nexus of the differentiated and dedifferentiated phenotypes, structural actin cytoskeleton characteristics per se play a crucial role in permissive regulation of cell-matrix adhesive interactions and cell migration behavior during T-cadherin-induced SMC phenotype transition.

Project description:Several vector-borne plant pathogens have evolved mechanisms to exploit and to hijack vector host cellular, molecular, and defense mechanisms for their transmission. In the past few years, Liberibacter species, which are transmitted by several psyllid vectors, have become an economically important group of pathogens that have devastated the citrus industry and caused tremendous losses to many other important crops worldwide. The molecular mechanisms underlying the interactions of Liberibacter species with their psyllid vectors are poorly studied. "Candidatus Liberibacter solanacearum," which is associated with important vegetable diseases, is transmitted by the carrot psyllid Bactericera trigonica in a persistent manner. Here, we elucidated the role of the B. trigonica Arp2/3 protein complex, which plays a major role in regulation of the actin cytoskeleton, in the transmission of "Ca Liberibacter solanacearum." "Ca Liberibacter solanacearum" colocalized with ArpC2, a key protein in this complex, and this colocalization was strongly associated with actin filaments. Silencing of the psyllid ArpC2 disrupted the colocalization and the dynamics of F-actin. Silencing of RhoGAP21 and Cdc42, which act in the signaling cascade leading to upregulation of Arp2/3 and F-actin bundling, showed similar results. On the other hand, silencing of ArpC5, another component of the complex, did not induce any significant effects on F-actin formation. Finally, ArpC2 silencing caused a 73.4% reduction in "Ca Liberibacter solanacearum" transmission by psyllids, strongly suggesting that transmission of "Ca Liberibacter solanacearum" by B. trigonica is cytoskeleton dependent and "Ca Liberibacter solanacearum" interacts with ArpC2 to exploit the intracellular actin nucleation process for transmission. Targeting this unique interaction could lead to the development of a novel strategy for the management of Liberibacter-associated diseases.IMPORTANCE Plant diseases caused by vector-borne pathogens are responsible for tremendous losses and threaten some of the most important agricultural crops. A good example is the citrus greening disease, which is caused by bacteria of the genus Liberibacter and is transmitted by psyllids; it has devastated the citrus industry in the United States, China, and Brazil. Here, we show that psyllid-transmitted "Candidatus Liberibacter solanacearum" employs the actin cytoskeleton of psyllid gut cells, specifically the ArpC2 protein in the Arp2/3 complex of this system, for movement and transmission in the vector. Silencing of ArpC2 dramatically influenced the interaction of "Ca Liberibacter solanacearum" with the cytoskeleton and decreased the bacterial transmission to plants. This system could be targeted to develop a novel approach for the control of Liberibacter-associated diseases.

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Disruption of actin cytoskeleton mediates loss of tensile stress induced early phenotypic modulation of vascular smooth muscle cells in organ culture.

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Disruption of actin cytoskeleton mediates loss of tensile stress induced early phenotypic modulation of vascular smooth muscle cells in organ culture.

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