Enescence can also be identified by increased expression of senescence-associated biomarkers

Enescence can also be identified by increased expression of senescence-associated biomarkers such as Apo J, CTGF, and fibronectin. All three biomarkers are inducible by oxidative stress [16,18]. In our experiments, exposure of primary human RPE cells to CSE could lead to a significant elevation of Apo J, CTGF, and fibronectin expression. The cellular chaperone Apo J has been previously detected in the RPE of AMD donor eyes, although its role and function in the RPE is still unclear [19,53]. In contrast, CTGF and fibronectin have been found in the Bruch’s membrane, in drusen and in basal linear deposits of AMD eyes [20,21,22]. Furthermore, we could show that treatment of human RPE cells with CSE also increased the secretion of fibronectin and laminin into the culture media. Laminin is a basement membrane protein, which is involved in the formation of basal laminar deposits of the ageing macula [24]. Both laminin and fibronectin have been shown to be secreted by senescent human RPE cells [23]. In the pathogenesis of AMD, it is assumed that cellular senescence and dysfunction of the RPE lead to an increased aggregation of ECM [15,54]. Therefore, CSE-induced levels of CTGF and fibronectinrepresent senescence-associated changes and demonstrate increased ECM synthesis in cultured human RPE cells. A similar effect could also be observed after treatment of RPE cells with hypoxia/reoxygenation [55]. Furthermore, exposure to cigarette smoke could increase the formation of sub-RPE ECM deposits in an experimental mouse model [56,57]. An induction of CTGF levels was previously observed during cutaneous wound healing in smoke-exposed mice [58]. Whether or not CSE is responsible for the ECM accumulation in the RPE of AMD patients awaits further DprE1-IN-2 investigations. Based on these results, we conclude that cigarette smoke may be responsible for the cell loss, senescent changes, and synthesis of ECM components in primary cultured human RPE cells. Therefore, cigarette smoke may induce cellular events, which may resemble pathogenic changes in AMD. Hence, these results may provide one explanation for the adverse effects of cigarette smoke on the pathogenesis and progression of AMD.AcknowledgmentsThe authors thank Katja Obholzer and Jerome Moriniere for excellent technical assistance.Author ContributionsConceived and designed the experiments: ALY KB JB UWL. Performed the experiments: ALY KB JB UWL. Analyzed the data: ALY KB UWL. Contributed reagents/materials/analysis tools: ALY UWL. Wrote the paper: ALY UWL. Obtained permission for the use of cell line: ALY UWL.
Thrombin signaling in platelets is mediated by protease activated receptors (PARs). PARs are G-protein-coupled receptors (GPCR) that are activated via proteolytic cleavage of the extracellular N-terminus to initiate a variety of signaling cascades through activation of heterotrimeric G proteins [1,2]. The expression of PARs in platelets is species specific. Human platelets express PAR1 and PAR4 and cleavage of each receptor initiates signaling cascades [3]. Mouse platelets express PAR3 and PAR4, but 16574785 PAR3 does not signal making PAR4 the signaling receptor [4?6]. PAR1 and PAR4 in human platelets are coupled to Gq and G12/13 [7,8]. The presence of a direct 301353-96-8 site interaction between PARs and Gi is controversial. It has been shown that PAR1 is directly coupled to Gi in human platelets and in COS7 cells transfected with PAR1 [9,10]. However, other studies have shown that PAR1 and PAR4 do not couple directly to Gi, r.Enescence can also be identified by increased expression of senescence-associated biomarkers such as Apo J, CTGF, and fibronectin. All three biomarkers are inducible by oxidative stress [16,18]. In our experiments, exposure of primary human RPE cells to CSE could lead to a significant elevation of Apo J, CTGF, and fibronectin expression. The cellular chaperone Apo J has been previously detected in the RPE of AMD donor eyes, although its role and function in the RPE is still unclear [19,53]. In contrast, CTGF and fibronectin have been found in the Bruch’s membrane, in drusen and in basal linear deposits of AMD eyes [20,21,22]. Furthermore, we could show that treatment of human RPE cells with CSE also increased the secretion of fibronectin and laminin into the culture media. Laminin is a basement membrane protein, which is involved in the formation of basal laminar deposits of the ageing macula [24]. Both laminin and fibronectin have been shown to be secreted by senescent human RPE cells [23]. In the pathogenesis of AMD, it is assumed that cellular senescence and dysfunction of the RPE lead to an increased aggregation of ECM [15,54]. Therefore, CSE-induced levels of CTGF and fibronectinrepresent senescence-associated changes and demonstrate increased ECM synthesis in cultured human RPE cells. A similar effect could also be observed after treatment of RPE cells with hypoxia/reoxygenation [55]. Furthermore, exposure to cigarette smoke could increase the formation of sub-RPE ECM deposits in an experimental mouse model [56,57]. An induction of CTGF levels was previously observed during cutaneous wound healing in smoke-exposed mice [58]. Whether or not CSE is responsible for the ECM accumulation in the RPE of AMD patients awaits further investigations. Based on these results, we conclude that cigarette smoke may be responsible for the cell loss, senescent changes, and synthesis of ECM components in primary cultured human RPE cells. Therefore, cigarette smoke may induce cellular events, which may resemble pathogenic changes in AMD. Hence, these results may provide one explanation for the adverse effects of cigarette smoke on the pathogenesis and progression of AMD.AcknowledgmentsThe authors thank Katja Obholzer and Jerome Moriniere for excellent technical assistance.Author ContributionsConceived and designed the experiments: ALY KB JB UWL. Performed the experiments: ALY KB JB UWL. Analyzed the data: ALY KB UWL. Contributed reagents/materials/analysis tools: ALY UWL. Wrote the paper: ALY UWL. Obtained permission for the use of cell line: ALY UWL.
Thrombin signaling in platelets is mediated by protease activated receptors (PARs). PARs are G-protein-coupled receptors (GPCR) that are activated via proteolytic cleavage of the extracellular N-terminus to initiate a variety of signaling cascades through activation of heterotrimeric G proteins [1,2]. The expression of PARs in platelets is species specific. Human platelets express PAR1 and PAR4 and cleavage of each receptor initiates signaling cascades [3]. Mouse platelets express PAR3 and PAR4, but 16574785 PAR3 does not signal making PAR4 the signaling receptor [4?6]. PAR1 and PAR4 in human platelets are coupled to Gq and G12/13 [7,8]. The presence of a direct interaction between PARs and Gi is controversial. It has been shown that PAR1 is directly coupled to Gi in human platelets and in COS7 cells transfected with PAR1 [9,10]. However, other studies have shown that PAR1 and PAR4 do not couple directly to Gi, r.

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