Oxidative stress induces inactivation of protein phosphatase 2A, promoting proinflammatory NF-κB in aged rat kidney.

The molecular inflammation hypothesis of aging proposes that redox dysregulation causes an age-related activation of NF-κB and its signaling to upregulate various proinflammatory genes. In the present study, we focused on the inactive form of the protein phosphastase 2A (PP2A). More specifically, we aimed to define the correlation between PP2A inactivation and NF-κB activation by age-related oxidative stress. Experimentations were designed to determine the effect of oxidative stress-induced PP2A inactivation on NF-κB activity, utilizing prooxidants t-BHP and AAPH, the PTP inhibitor Na3VO4, and the PP2A inhibitor Calyculin A and PP2A siRNA, in HEK293T cells. We also assessed the phosphorylation of PP2A catalytic subunit (PP2Ac) and the activities of PP2A and NF-κB in aged rat kidney, utilizing aging-retarding 40% calorie restriction (CR) -60% of food intake and inflammation-triggering LPS paradigms. Results revealed that an oxidative stress-induced PTK/PTP imbalance led to phosphorylation of PP2Ac, following exposures to t-BHP, AAPH, and Na3VO4 in HEK293T cells. Subsequently, we found that Calyculin A and PP2A siRNA activates NIK/IKK and MAPKs, leading to upregulation of NF-κB and its dependent oxidative stress. Also, the contrasting relation between PP2A inactivation and NF-κB activation was confirmed by AAPH-induced oxidative status in mice, and non-induced normal status or LPS-induced inflammatory status in aged rats while the antioxidative, anti-inflammatory, anti-aging effects of CR significantly blunted these actions. Thus, we present evidence that PP2A inactivation via PTK/PTP imbalance provoked by oxidative stress causes NF-κB activation, which contributes to the accumulation of oxidative stress in aged rat kidney.

FoxO1 links insulin resistance to proinflammatory cytokine IL-1beta production in macrophages.

OBJECTIVE: Macrophages play an important role in the pathogenesis of insulin resistance via the production of proinflammatory cytokines. Our goal is to decipher the molecular linkage between proinflammatory cytokine production and insulin resistance in macrophages. RESEARCH DESIGN AND METHODS: We determined cytokine profiles in cultured macrophages and identified interleukin (IL)-1ß gene as a potential target of FoxO1, a key transcription factor that mediates insulin action on gene expression. We studied the mechanism by which FoxO1 mediates insulin-dependent regulation of IL-1ß expression in cultured macrophages and correlated FoxO1 activity in peritoneal macrophages with IL-1ß production profiles in mice with low-grade inflammation or insulin resistance. RESULTS: FoxO1 selectively promoted IL-1ß production in cultured macrophages. This effect correlated with the ability of FoxO1 to bind and enhance IL-1ß promoter activity. Mutations of the FoxO1 binding site within the IL-1ß promoter abolished FoxO1 induction of IL-1ß expression. Macrophages from insulin-resistant obese db/db mice or lipopolysaccharide-inflicted mice were associated with increased FoxO1 production, correlating with elevated levels of IL-1ß mRNA in macrophages and IL-1 protein in plasma. In nonstimulated macrophages, FoxO1 remained inert with benign effects on IL-1ß expression. In response to inflammatory stimuli, FoxO1 activity was augmented because of an impaired ability of insulin to phosphorylate FoxO1 and promote its nuclear exclusion. This effect along with nuclear factor-κB acted to stimulate IL-1ß production in activated macrophages. CONCLUSIONS: FoxO1 signaling through nuclear factor-κB plays an important role in coupling proinflammatory cytokine production to insulin resistance in obesity and diabetes.