Increased Aortic Calpain-1 Activity Mediates Age-Associated Angiotensin II Signaling of Vascular Smooth Muscle Cells

Angiotensin II (Ang II) signaling, including matrix metalloproteinase type II (MMP2) activation, has been linked to an age-associated increase in migration capacity of vascular smooth muscle cells (VSMC), and to other proinflammatory features of arterial aging. Calpain-1 activation is required for MMP2 expression in fibroblasts and is induced in cardiomyocytes by Ang II. The consequences of engagement of calpain-1 with its substrates, however, in governing the age-associated proinflammatory status within the arterial wall, remains unknown.The present findings demonstrate that transcription, translation, and activity of calpain-1 are significantly up-regulated in rat aortae or early-passage aortic VSMC from old (30-mo) rats compared to young (8-mo). Dual immunolabeling of the arterial wall indicates that colocalization of calpain-1 and Ang II increases within the aged arterial wall. To further explore the relationship of calpain-1 to Ang II, we chronically infused Ang II into young rats, and treated cultured aortic rings or VSMC with Ang II. We also constructed adenoviruses harboring calpain-1 (CANP1) or its endogenous inhibitor calpastatin (CAST) and infected these into VSMC. Ang II induces calpain-1 expression in the aortic walls in vivo and ex vivo and VSMC in vitro. The Ang II mediated, age-associated increased MMP2 activity and migration in VSMC are both blocked by calpain inhibitor 1 or CAST. Over-expression of calpain-1 in young VSMC results in cleavage of intact vimentin, and an increased migratory capacity mimicking that of old VSMC, which is blocked by the MMP inhibitor, GM6001.Calpain-1 activation is a pivotal molecular event in the age-associated arterial Ang II/MMP2 signaling cascade that is linked to cytoskeleton protein restructuring, and VSMC migration. Therefore, targeting calpain-1 has the potential to delay or reverse the arterial remodeling that underlies age-associated diseases i.e. atherosclerosis

Reduced vasorin enhances angiotensin II signaling within the aging arterial wall

The glycosylated protein vasorin physically interacts with the transforming growth factor-beta1 (TGF-β1) and functionally attenuates its fibrogenic signaling in the vascular smooth muscle cells (VSMCs) of the arterial wall. Angiotensin II (Ang II) amplifies TGF-β1 activation in the VSMCs of the arterial wall with aging. In this study, we hypothesized that a reduced expression of the protein vasorin plays a contributory role in magnifying Ang II-associated fibrogenic signaling in the VSMCs of the arterial wall with aging. The current study shows that vasorin mRNA and protein expression were significantly decreased both in aortic wall and VSMCs from old (30 mo) vs. young (8 mo) FXBN rats. Exposing young VSMCs to Ang II reduced vasorin protein expression to the levels of old untreated cells while treating old VSMCs with the Ang II type AT1 receptor antagonist Losartan upregulated vasorin protein expression up to the levels of young. The physical interaction between vasorin and TGF-β1 was significantly decreased in old vs. young VSMCs. Further, exposing young VSMCs to Ang II increased the levels of matrix metalloproteinase type II (MMP-2) activation and TGF-β1 downstream molecules p-SMAD-2/3 and collagen type I production up to the levels of old untreated VSMCs, and these effects were substantially inhibited by overexpressing vasorin. Administration of Ang II to young rats (8 mo) for 28 days via an osmotic minipump markedly reduced the expression of vasorin. Importantly, vasorin protein was effectively cleaved by activated MMP-2 both in vitro and in vivo. Administration of the MMP inhibitor, PD 166793, for 6 mo to young adult (18 mo) via a daily gavage markedly increased levels of vasorin in the aortic wall. Thus, reduced vasorin amplifies Ang II profibrotic signaling via an activation of MMP-2 in VSMCs within the aging arterial wall

A Local Proinflammatory Signalling Loop Facilitates Adverse Age-Associated Arterial Remodeling

Background: The coincidence of vascular smooth muscle cells (VSMC) infiltration and collagen deposition within a diffusely thickened intima is a salient feature of central arterial wall inflammation that accompanies advancing age. However, the molecular mechanisms involved remain undefined. Methodology/Principal Findings: Immunostaining and immunoblotting of rat aortae demonstrate that a triad of proinflammatory molecules, MCP-1, TGF-b1, and MMP-2 increases within the aortic wall with aging. Exposure of VSMC isolated from 8-mo-old rats (young) to MCP-1 effects, via CCR-2 signaling, both an increase in TGF-b1 activity, up to levels of untreated VSMC from 30-mo-old (old) rats, and a concurrent increase in MMP-2 activation. Furthermore, exposure of young VSMC to TGF-b1 increases levels of MCP-1, and MMP-2 activation, to levels of untreated VSMC from old rats. This autocatalytic signaling loop that enhances collagen production and invasiveness of VSMC is effectively suppressed by si-MCP-1, a CCR2 antagonist, or MMP-2 inhibition. Conclusions/Significance: Threshold levels of MCP-1, MMP-2, or TGF-b1 activity trigger a feed-forward signaling mechanism that is implicated in the initiation and progression of adverse age-associated arterial wall remodeling. Intervention that suppressed this signaling loop may potentially retard age-associated adverse arterial remodeling

Democracy in trade unions, democracy through trade unions?

Since the Webbs published Industrial Democracy at the end of the nineteenth century, the principle that workers have a legitimate voice in decision-making in the world of work – in some versions through trade unions, in others at least formally through separate representative structures – has become widely accepted in most west European countries. There is now a vast literature on the strengths and weaknesses of such mechanisms, and we review briefly some of the key interpretations of the rise (and fall) of policies and structures for workplace and board-level representation. We also discuss the mainly failed attempts to establish broader processes of economic democracy, which the eclipse of nationally specific mechanisms of class compromise makes again a salient demand. Economic globalization also highlights the need for transnational mechanisms to achieve worker voice (or more radically, control) in the dynamics of capital-labour relations. We therefore examine the role of trade unions in coordinating pressure for a countervailing force at European and global levels, and in the construction of (emergent?) supranational industrial relations. However, many would argue that unions cannot win legitimacy as democratizing force unless manifestly democratic internally. We therefore revisit debates on and dilemmas of democracy within trade unions, and examine recent initiatives to enhance democratization

Characterization of diverse populations of sinoatrial node cells and their proliferation potential at single nucleus resolutionResearch in context

Background: Each heartbeat is initiated in the sinoatrial node (SAN), and although a recent study (GSE130710) using single nucleus RNA-seq had discovered different populations of cell types within SAN tissue, the distinct potential functions of these cell types have not been delineated. Methods: To infer some special potential functions of different SAN cell clusters, we applied principal component analysis (PCA), t-distributed stochastic neighbor embedding (t-SNE) and uniform manifold approximation and projection (UMAP) to the GSE130710 dataset to reduce dimensions, followed by Pseudotime trajectory and AUCell analyses, ANOVA and Hurdle statistical models, and Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichments to determine functional potential of cell types. Nuclear EdU immuno-labeling of SAN tissue confirmed cell type proliferation. Findings: We identified elements of a coupled clock system known to drive SAN cell pacemaking within the GSE130710 sinus node myocyte cluster, which, surprisingly, manifested signals of suppressed fatty acid and nitrogen metabolism and reduced immune gene expression. Proliferation signaling was enriched in endocardial, epicardial, epithelial cells, and macrophages, in which, fatty acid and nitrogen metabolic signals were also suppressed, but immune signaling was enhanced. EdU labeling was rare in pacemaker cells but was robust in interstitial cells. Interpretation: Pacemaker cells that initiate each heartbeat manifest suppressed fatty acid and nitrogen metabolism and limited immune signaling and proliferation potential. In contrast, other populations of SAN cells not directly involved in the initiation of heartbeats, manifest robust proliferation and immune potential, likely to ensure an environment required to sustain healthy SAN tissue pacemaker function

Milk fat globule protein epidermal growth factor-8. a pivotal relay element within the angiotensin II and monocyte chemoattractant protein-1 signaling cascade mediating vascular smooth muscle cells invasion

Advancing age induces aortic wall thickening that results from the concerted effects of numerous signaling proteins, many of which have yet to be identified. To search for novel proteins associated with aortic wall thickening, we have performed a comprehensive quantitative proteomic study to analyze aortic proteins from young (8 months) and old (30 months) rats and identified 50 proteins that significantly change in abundance with aging. One novel protein, the milk fat globule protein epidermal growth factor 8 (MFG-E8), increases 2.3-fold in abundance in old aorta. Transcription and translation analysis demonstrated that aortic MFG-E8 mRNA and protein levels increase with aging in several mammalian species including humans. Dual immunolabeling shows that MFG-E8 colocalizes with both angiotensin II and monocyte chemoattractant protein (MCP)-1 within vascular smooth muscle cells (VSMCs) of the thickened aged aortic wall. Exposure of early passage VSMCs from young aorta to angiotensin II markedly increases MFG-E8 and enhances invasive capacity to levels observed in VSMCs from old rats. Treatment of VSMCs with MFG-E8 increases MCP-1 expression and VSMCs invasion that are inhibited by the MCP-1 receptor blocker vCCI. Silencing MFG-E8 RNA substantially reduces MFG-E8 expression and VSMCs invasion capacity. The data indicate that arterial MFG-E8 significantly increases with aging and is a pivotal relay element within the angiotensin II/MCP-1/VSMC invasion signaling cascade. Thus, targeting of MFG-E8 within this signaling axis pathway is a potential novel therapy for the prevention and treatment of the age-associated vascular diseases such as atherosclerosis

A remarkable adaptive paradigm of heart performance and protection emerges in response to marked cardiac-specific overexpression of ADCY8

Adult (3 month) mice with cardiac-specific overexpression of adenylyl cyclase (AC) type VIII (TGAC8) adapt to an increased cAMP-induced cardiac workload (~30% increases in heart rate, ejection fraction and cardiac output) for up to a year without signs of heart failure or excessive mortality. Here, we show classical cardiac hypertrophy markers were absent in TGAC8, and that total left ventricular (LV) mass was not increased: a reduced LV cavity volume in TGAC8 was encased by thicker LV walls harboring an increased number of small cardiac myocytes, and a network of small interstitial proliferative non-cardiac myocytes compared to wild type (WT) littermates; Protein synthesis, proteosome activity, and autophagy were enhanced in TGAC8 vs WT, and Nrf-2, Hsp90α, and ACC2 protein levels were increased. Despite increased energy demands in vivo LV ATP and phosphocreatine levels in TGAC8 did not differ from WT. Unbiased omics analyses identified more than 2,000 transcripts and proteins, comprising a broad array of biological processes across multiple cellular compartments, which differed by genotype; compared to WT, in TGAC8 there was a shift from fatty acid oxidation to aerobic glycolysis in the context of increased utilization of the pentose phosphate shunt and nucleotide synthesis. Thus, marked overexpression of AC8 engages complex, coordinate adaptation "circuity" that has evolved in mammalian cells to defend against stress that threatens health or life (elements of which have already been shown to be central to cardiac ischemic pre-conditioning and exercise endurance cardiac conditioning) that may be of biological significance to allow for proper healing in disease states such as infarction or failure of the heart