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Serum Amyloid A Stimulates Atherogenesis and Renal Dysfunction in Apolipoprotein E-Deficient Mice Fed a Normal Chow Diet
modification to its structure and functional properties. These changes may contribute to endothelial dysfunction and rupture of atherosclerotic lesions.
doi: 10.1016/j.freeradbiomed.2016.10.404 364 Superior Targeted Anti-Platelet Activity of Nitrite Nadeem Wajih1, Anuj Jailwala1, Mark Gladwin2, David L Caudell3, and Daniel B Kim-Shapiro1 1 Wake Forest University, Winston-Salem, USA, 2Heart, Lung, Blood and Vascular Medicine Institute and the Department of Medicine, University of Pittsburgh, Pittsburgh, USA, 3Wake Forest University Health Sciences, Winston-Salem, USA Nitric Oxide (NO) is known to inhibit platelet activation and aggregation, primarily through activation of soluble guanylyl cyclase. Several NO donor drugs that take advantage of this pathway have been proposed to decrease platelet activation and thrombosis These include the S-nitrosothiols such as Snitrosoglutathione (GSNO) and spontaneous NO releasing drugs such as diazeniumdiolates (also known as NONOates). More recently, the simple anion nitrite (NO2–) has been shown to inhibit platelet activation and aggregation in vitro and in vivo in rodents and humans. In this work, we compare the efficacy of one micromolar nitrite to that of one micromolar GSNO and one micromolar Diethylamine NONOate (DEANO) in inhibiting platelet activation and aggregation. Under normoxic conditions, nitrite had no effect on platelet activation while GSNO and DEANO significantly inhibited platelet activation. In the absence of red blood cells (that is in platelet rich plasma) nitrite again showed no activity whereas one micromolar GSNO and DEANO inhibited platelet activation. However, under hypoxic conditions, and in the presence of red blood cells, nitrite was as or more effective at inhibiting platelet activation than GSNO and DEANO. Nitrite was also as effective as GSNO and DEANO in inhibiting platelet aggregation in mixed venous blood. Whereas the presence of red blood cells is necessary for nitrite’s anti-platelet action, red blood cells reduced the efficacy of GSNO and DEANO. Importantly, physiological concentrations of NO and GSNO are three orders of magnitude below those used in these studies (one micromolar), but one micromolar nitrite is only 3-10 times resting physiological plasma levels and equal to that obtained after consumption of high nitrate containing foods or beverages. Thus, nitrite is an effective anti-platelet agent that is activated by red blood cells and has enhanced activities with physiological hypoxia. A nitrite-red blood cell-NO pathway may contribute to maintenance of normal hemostasis.
doi: 10.1016/j.freeradbiomed.2016.10.405 365 Laminar Flow Promotes Mitochondrial Functions in Endothelial Cells
We examined the dynamic changes of mitochondria and its regulatory proteins associated with mitochondrial functions in flowtreated ECs. Cultured ECs were subjected to a constant shear stress (12 dynes/cm2) with a flow chamber system and mitochondrial biogenesis, dynamics and function were analyzed. Shear-treated ECs increased mitochondrial biogenesis as indicated by the elevated levels of mitochondrial transcriptional factors (TFAM, PGC1-, NRF1) and proteins (SDHA and COX4). Shear flow triggered an increment of mitochondrial fusion that was coupled by an increased expression of fusion proteins (MFN2, OPA1) but decreased fission protein (FIS1). Consistently, an increase of phosphorylation at S637 but decrease at S616 on Drp1 and the reduced mitochondria-associated Drp1 were observed in sheared ECs. Shear flow increased the expression of mitochondrial antioxidant enzymes MnSOD2, TRX2, PRX3 and PRX5. Thus, Sheared ECs improve mitochondrial function as revealed by a decreased mitochondrial ROS and an increase of mitochondrial membrane potential and ATP production. Our results suggest that steady flow promotes mitochondrial homeostasis in ECs.
doi: 10.1016/j.freeradbiomed.2016.10.406 366 Serum Amyloid A Stimulates Atherogenesis and Renal Dysfunction in Apolipoprotein E-Deficient Mice Fed a Normal Chow Diet Paul Witting1, Belal Chami1, Thomas Hambly2, XiaoSuo Wang1, Genevieve Fong1, and Joanne Dennis1 1 University of Sydney, Australia, 2University of New England, Armidale, Australia Elevated serum amyloid A (SAA) can cause endothelial dysfunction via enhanced oxidative stress that advances inflammation/thrombosis in cardiovascular and renal pathology. We investigated the impact of SAA on aortic vessels and renal function in apolipoprotein E-deficient (ApoE-/-) mice. Male ApoE-/mice received vehicle-(control), low-level lipopolysaccharide (LPS) or recombinant human SAA by i.p. injection every third day (over 2-weeks). Heart, aorta and kidney were harvested after 3 or 5 days, or after a total of 4 weeks (designated as young) or 18 weeks (old) mice. SAA-mediated-renal injury was evident in young mice manifesting as increased plasma blood nitrogen products, and urinary levels of protein and kidney injury molecule-1 (KIM-1), tissue content of oxidized lipids and several cytokines/chemokines, relative to controls. SAA stimulated NFB activation and macrophage recruitment in kidneys as early as 3 days after administration. SAA increased vascular cell adhesion molecule (VCAM)-1 in aortic vessels and plasma monocyte chemotactic protein (MCP)-1 levels in young mice relative to their respective controls. Aortic gene expression of nuclear-factor-kappa-beta (NFB-p50) and tissue factor (TF) also increased in young mice confirming that SAA stimulated pro-inflammatory/pro-thrombotic responses. These data suggest that SAA promotes renal impairment and endothelial dysfunction in ApoE-/- mice in the absence of a high-fat diet.
doi: 10.1016/j.freeradbiomed.2016.10.407
Ling D Wang1 1 Tzu-Chi University, Hualian,Taiwan Hemodynamic shear stress exerts atheroprotection in endothelial cells (ECs) have been intensively studied. However, the effects of shear flow on mitochondrial homeostasis in ECs remain unclear.
S154
SfRBM / SFRRI 2016
doi: 10.1016/j.freeradbiomed.2016.10.404 364 Superior Targeted Anti-Platelet Activity of Nitrite Nadeem Wajih1, Anuj Jailwala1, Mark Gladwin2, David L Caudell3, and Daniel B Kim-Shapiro1 1 Wake Forest University, Winston-Salem, USA, 2Heart, Lung, Blood and Vascular Medicine Institute and the Department of Medicine, University of Pittsburgh, Pittsburgh, USA, 3Wake Forest University Health Sciences, Winston-Salem, USA Nitric Oxide (NO) is known to inhibit platelet activation and aggregation, primarily through activation of soluble guanylyl cyclase. Several NO donor drugs that take advantage of this pathway have been proposed to decrease platelet activation and thrombosis These include the S-nitrosothiols such as Snitrosoglutathione (GSNO) and spontaneous NO releasing drugs such as diazeniumdiolates (also known as NONOates). More recently, the simple anion nitrite (NO2–) has been shown to inhibit platelet activation and aggregation in vitro and in vivo in rodents and humans. In this work, we compare the efficacy of one micromolar nitrite to that of one micromolar GSNO and one micromolar Diethylamine NONOate (DEANO) in inhibiting platelet activation and aggregation. Under normoxic conditions, nitrite had no effect on platelet activation while GSNO and DEANO significantly inhibited platelet activation. In the absence of red blood cells (that is in platelet rich plasma) nitrite again showed no activity whereas one micromolar GSNO and DEANO inhibited platelet activation. However, under hypoxic conditions, and in the presence of red blood cells, nitrite was as or more effective at inhibiting platelet activation than GSNO and DEANO. Nitrite was also as effective as GSNO and DEANO in inhibiting platelet aggregation in mixed venous blood. Whereas the presence of red blood cells is necessary for nitrite’s anti-platelet action, red blood cells reduced the efficacy of GSNO and DEANO. Importantly, physiological concentrations of NO and GSNO are three orders of magnitude below those used in these studies (one micromolar), but one micromolar nitrite is only 3-10 times resting physiological plasma levels and equal to that obtained after consumption of high nitrate containing foods or beverages. Thus, nitrite is an effective anti-platelet agent that is activated by red blood cells and has enhanced activities with physiological hypoxia. A nitrite-red blood cell-NO pathway may contribute to maintenance of normal hemostasis.
doi: 10.1016/j.freeradbiomed.2016.10.405 365 Laminar Flow Promotes Mitochondrial Functions in Endothelial Cells
We examined the dynamic changes of mitochondria and its regulatory proteins associated with mitochondrial functions in flowtreated ECs. Cultured ECs were subjected to a constant shear stress (12 dynes/cm2) with a flow chamber system and mitochondrial biogenesis, dynamics and function were analyzed. Shear-treated ECs increased mitochondrial biogenesis as indicated by the elevated levels of mitochondrial transcriptional factors (TFAM, PGC1-, NRF1) and proteins (SDHA and COX4). Shear flow triggered an increment of mitochondrial fusion that was coupled by an increased expression of fusion proteins (MFN2, OPA1) but decreased fission protein (FIS1). Consistently, an increase of phosphorylation at S637 but decrease at S616 on Drp1 and the reduced mitochondria-associated Drp1 were observed in sheared ECs. Shear flow increased the expression of mitochondrial antioxidant enzymes MnSOD2, TRX2, PRX3 and PRX5. Thus, Sheared ECs improve mitochondrial function as revealed by a decreased mitochondrial ROS and an increase of mitochondrial membrane potential and ATP production. Our results suggest that steady flow promotes mitochondrial homeostasis in ECs.
doi: 10.1016/j.freeradbiomed.2016.10.406 366 Serum Amyloid A Stimulates Atherogenesis and Renal Dysfunction in Apolipoprotein E-Deficient Mice Fed a Normal Chow Diet Paul Witting1, Belal Chami1, Thomas Hambly2, XiaoSuo Wang1, Genevieve Fong1, and Joanne Dennis1 1 University of Sydney, Australia, 2University of New England, Armidale, Australia Elevated serum amyloid A (SAA) can cause endothelial dysfunction via enhanced oxidative stress that advances inflammation/thrombosis in cardiovascular and renal pathology. We investigated the impact of SAA on aortic vessels and renal function in apolipoprotein E-deficient (ApoE-/-) mice. Male ApoE-/mice received vehicle-(control), low-level lipopolysaccharide (LPS) or recombinant human SAA by i.p. injection every third day (over 2-weeks). Heart, aorta and kidney were harvested after 3 or 5 days, or after a total of 4 weeks (designated as young) or 18 weeks (old) mice. SAA-mediated-renal injury was evident in young mice manifesting as increased plasma blood nitrogen products, and urinary levels of protein and kidney injury molecule-1 (KIM-1), tissue content of oxidized lipids and several cytokines/chemokines, relative to controls. SAA stimulated NFB activation and macrophage recruitment in kidneys as early as 3 days after administration. SAA increased vascular cell adhesion molecule (VCAM)-1 in aortic vessels and plasma monocyte chemotactic protein (MCP)-1 levels in young mice relative to their respective controls. Aortic gene expression of nuclear-factor-kappa-beta (NFB-p50) and tissue factor (TF) also increased in young mice confirming that SAA stimulated pro-inflammatory/pro-thrombotic responses. These data suggest that SAA promotes renal impairment and endothelial dysfunction in ApoE-/- mice in the absence of a high-fat diet.
doi: 10.1016/j.freeradbiomed.2016.10.407
Ling D Wang1 1 Tzu-Chi University, Hualian,Taiwan Hemodynamic shear stress exerts atheroprotection in endothelial cells (ECs) have been intensively studied. However, the effects of shear flow on mitochondrial homeostasis in ECs remain unclear.
S154
SfRBM / SFRRI 2016