Akt Regulates TNFα synthesis downstream of RIP1 kinase activation during necroptosis.

Necroptosis is a regulated form of necrotic cell death that has been implicated in the pathogenesis of various diseases including intestinal inflammation and systemic inflammatory response syndrome (SIRS). In this work, we investigated the signaling mechanisms controlled by the necroptosis mediator receptor interacting protein-1 (RIP1) kinase. We show that Akt kinase activity is critical for necroptosis in L929 cells and plays a key role in TNFα production. During necroptosis, Akt is activated in a RIP1 dependent fashion through its phosphorylation on Thr308. In L929 cells, this activation requires independent signaling inputs from both growth factors and RIP1. Akt controls necroptosis through downstream targeting of mammalian Target of Rapamycin complex 1 (mTORC1). Akt activity, mediated in part through mTORC1, links RIP1 to JNK activation and autocrine production of TNFα. In other cell types, such as mouse lung fibroblasts and macrophages, Akt exhibited control over necroptosis-associated TNFα production without contributing to cell death. Overall, our results provide new insights into the mechanism of necroptosis and the role of Akt kinase in both cell death and inflammatory regulation.

Small-molecule inhibitors of the PI3K signaling network.

The phosphoinositide 3-kinase (PI3K) signaling pathway controls a wide variety of cellular processes including cell death and survival, cell migration, protein synthesis and metabolism. Aberrant PI3K-dependent signaling, mediated by Akt kinase, has been implicated in many human diseases including cancer, inflammation, cardiovascular disease and metabolic diseases, making this pathway a principle target for drug development. In this article we will summarize the PI3K signaling network and discuss current strategies for pathway inhibition. We will also explore the importance and emerging relevance of Akt-independent PI3K signaling pathways and discuss attempts being made to harness these pathways by inhibiting the binding of a product of PI3K, phosphatidylinositol-(3,4,5)-trisphosphate, to effector pleckstrin homology domains.

Inhibition of the cation channel TRPV4 improves bladder function in mice and rats with cyclophosphamide-induced cystitis.

Reduced functional bladder capacity and concomitant increased micturition frequency (pollakisuria) are common lower urinary tract symptoms associated with conditions such as cystitis, prostatic hyperplasia, neurological disease, and overactive bladder syndrome. These symptoms can profoundly affect the quality of life of afflicted individuals, but available pharmacological treatments are often unsatisfactory. Recent work has demonstrated that the cation channel TRPV4 is highly expressed in urothelial cells and plays a role in sensing the normal filling state of the bladder. In this article, we show that the development of cystitis-induced bladder dysfunction is strongly impaired in Trpv4(-/-) mice. Moreover, we describe HC-067047, a previously uncharacterized, potent, and selective TRPV4 antagonist that increases functional bladder capacity and reduces micturition frequency in WT mice and rats with cystitis. HC-067047 did not affect bladder function in Trpv4(-/-) mice, demonstrating that its in vivo effects are on target. These results indicate that TRPV4 antagonists may provide a promising means of treating bladder dysfunction.

TRPA1 mediates formalin-induced pain.

The formalin model is widely used for evaluating the effects of analgesic compounds in laboratory animals. Injection of formalin into the hind paw induces a biphasic pain response; the first phase is thought to result from direct activation of primary afferent sensory neurons, whereas the second phase has been proposed to reflect the combined effects of afferent input and central sensitization in the dorsal horn. Here we show that formalin excites sensory neurons by directly activating TRPA1, a cation channel that plays an important role in inflammatory pain. Formalin induced robust calcium influx in cells expressing cloned or native TRPA1 channels, and these responses were attenuated by a previously undescribed TRPA1-selective antagonist. Moreover, sensory neurons from TRPA1-deficient mice lacked formalin sensitivity. At the behavioral level, pharmacologic blockade or genetic ablation of TRPA1 produced marked attenuation of the characteristic flinching, licking, and lifting responses resulting from intraplantar injection of formalin. Our results show that TRPA1 is the principal site of formalin's pain-producing action in vivo, and that activation of this excitatory channel underlies the physiological and behavioral responses associated with this model of pain hypersensitivity.