Sodium 4-phenylbutyrate (4-PBA) is a low molecular weight fatty acid that has been used for treatment of urea cycle disorders in children, sickle cell disease, and thalassemia. It has been demonstrated recently that 4-PBA can act as a chemical chaperone by reducing the load of mutant or mislocated proteins retained in the endoplasmic reticulum (ER) under conditions associated with cystic fibrosis and liver injury. In the present study, we evaluated the neuroprotective effect of 4-PBA on cerebral ischemic injury. Pre-or post-treatment with 4-PBA at therapeutic doses attenuated infarction volume, hemispheric swelling, and apoptosis and improved neurological status in a mouse model of hypoxia-ischemia. Moreover, 4-PBA suppressed ER-mediated apoptosis by inhibiting eukaryotic initiation factor 2␣ phosphorylation, CCAAT/enhancerbinding protein homologous protein induction, and caspase-12 activation. In neuroblastoma neuro2a cells, 4-PBA reduced caspase-12 activation, DNA fragmentation, and cell death induced by hypoxia/reoxygenation. It protected against ER stress-induced but not mitochondria-mediated cell death. Additionally, 4-PBA inhibited the expression of inducible nitricoxide synthase and tumor necrosis factor-␣ in primary cultured glial cells under hypoxia/reoxygenation. These results indicate that 4-PBA could protect against cerebral ischemia through inhibition of ER stress-mediated apoptosis and inflammation. Therefore, the multiple actions of 4-PBA may provide a strong effect in treatment of cerebral ischemia, and its use as a chemical chaperone would provide a novel approach for the treatment of stroke.

Leptin is an important circulating signal for inhibiting food intake and body weight gain. In recent years, "leptin resistance" has been considered to be one of the main causes of obesity. However, the detailed mechanisms of leptin resistance are poorly understood. Increasing evidence has suggested that stress signals, which impair endoplasmic reticulum (ER) function, lead to an accumulation of unfolded proteins, which results in ER stress. In the present study, we hypothesized that ER stress is involved in leptin resistance. Tunicamycin, thapsigargin, or brefeldin A was used to induce ER stress. The activation status of leptin signals was measured by Western blotting analysis using a phospho-(Tyr705) signal transducer and activator of transcription 3 (STAT3) antibody. We observed that ER stress markedly inhibited leptin-induced STAT3 phosphorylation. In contrast, ER stress did not affect leptin-induced c-Jun NH 2 -terminal kinase activation. These results suggest that ER stress induces leptin resistance. ER stress-induced leptin resistance was mediated through protein tyrosine phosphatase 1B but not through suppressors of cytokine signaling 3. It is noteworthy that a chemical chaperone, which could improve the proteinfolding capacity, reversed ER stress-induced leptin resistance. Moreover, homocysteine, which induces ER stress, caused leptin resistance both in vitro and in vivo. Together, these findings suggest that the pathological mechanism of leptin resistance is derived from ER stress.

Leptin is a circulating molecule for the regulation of food intake and body weight suggested to be mediated in the hypothalamus via Ob-Rb receptor, which activates Janus kinase-signal transducer and activator of transcription (STAT) pathways. Although leptin receptors exist in many regions of the brain, there have been few in vivo functional studies of leptin's target site other than the hypothalamus. We report here that peripherally applied leptin increased STAT3 phosphorylation not only in the hypothalamus but also in the brain stem as assessed by Western blotting. Moreover, administration of leptin induced expression of the suppressor of cytokine signaling 3 mRNA, a negative feedback regulator of leptin signaling, in the brain stem as well as in the hypothalamus. Using immunohistochemistry, we observed phosphorylated STAT3-immunoreactive cells in the arcuate nucleus, ventromedial hypothalamus, lateral hypothalamic area of the hypothalamus, and the nucleus of the tractus solitarius, dorsal motor nucleus of the vagus nerve, lateral parabrachial nucleus, and central gray of the brain stem of leptin-injected mice. These findings represent physiologically functional leptin Ob-Rb receptor in the brain stem as well as in the hypothalamus. It is suggested that circulating leptin may directly act in the brain stem to elicit autonomic and neuroendocrine control of food intake and energy expenditure.

Endoplasmic reticulum (ER) stress is defined as an accumulation of unfolded proteins in the endoplasmic reticulum. 4-phenylbutyrate (4-PBA) has been demonstrated to promote the normal trafficking of the DF508 cystic fibrosis transmembrane conductance regulator (CFTR) mutant from the ER to the plasma membrane and to restore activity. We have reported that 4-PBA protected against cerebral ischemic injury and ER stress-induced neuronal cell death. In this study, we revealed that 4-PBA possesses chemical chaperone activity in vitro, which prevents the aggregation of denatured a-lactalbumin and bovine serum albumin (BSA). Furthermore, we investigated the effects of 4-PBA on the accumulation of Parkin-associated endothelin receptor-like receptor (Pael-R) pathologically relevant to the loss of dopaminergic neurons in autosomal recessive juvenile parkinsonism (AR-JP). Interestingly, 4-PBA restored the normal expression of Pael-R protein and suppressed ER stress induced by the overexpression of Pael-R. In addition, we showed that 4-PBA attenuated the activation of ER stress-induced signal transduction pathways and subsequent neuronal cell death. Moreover, 4-PBA restored the viability of yeasts that fail to induce an ER stress response under ER stress conditions. These results suggest that 4-PBA suppresses ER stress by directly reducing the amount of misfolded protein, including Pael-R accumulated in the ER.

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Various stresses, which impair ER (endoplasmic reticulum) function, lead to an accumulation of unfolded or misfolded proteins. ER stress triggers many rescuer responses, including a UPR (unfolded protein response). Increasing evidence has suggested that ER stress is involved in neurodegenerative diseases (Alzheimer's disease, Parkinson's disease and cerebral ischaemic insults), cancer, obesity and diabetes. In the present review, we consider the importance of ER stress under pathological conditions in mammals. Furthermore, we discuss the therapeutic potential for treatment targeting ER stress.

Possible roles of the afferent vagus nerve in regulation of interleukin (IL)-1beta expression in the brain and hypothalamic-pituitary-adrenal (HPA) axis were examined in anesthetized rats. Levels of IL-1beta mRNA and protein in the brain were measured by comparative RT-PCR and ELISA. Direct electrical stimulation of the central end of the vagus nerve was performed continuously for 2 h. The afferent stimulation of the vagus nerve induced increases in the expression of mRNA and protein levels of IL-1beta in the hypothalamus and the hippocampus. Furthermore, expression of corticotropin-releasing factor mRNA was increased in the hypothalamus 2 h after vagal stimulation. Plasma levels of ACTH and corticosterone were also increased by this stimulation. The present results indicate that activation of the afferent vagus nerves itself can induce production of IL-1beta in the brain and activate the HPA axis. Therefore, the afferent vagus nerve may play an important role in transmitting peripheral signals to the brain in the infection and inflammation.

BackgroundMultiple lines of evidence suggest innate immune response pathways to be involved in the development of obesity-associated diabetes although the molecular mechanism underling the disease is unknown. Recent observations suggest that saturated fatty acids can act as a ligand for toll-like receptor (TLR) 4, which is thought to mediate obesity-associated insulin resistance. Myeloid differentiation factor 88 (MyD88) is an adapter protein for TLR/IL-1 receptor signaling, which is involved in the activation of inflammatory pathways. To evaluate molecular mechanisms linking obesity-associated diabetes down-stream of TLR4, we investigated physiological role of MyD88 in high-fat diet (HFD)-induced obesity.Methodology/Principal FindingsIn the present study, we found MyD88-deficient mice fed a HFD had increased circulating levels of insulin, leptin and cholesterol, as well as liver dysfunction (increased induction of ALT levels, increased activation of JNK and cleavage of PARP), which were linked to the onset of severe diabetes. On the other hand, TNF-α would not be involved in HFD-induced diabetes in MyD88-deficient mice, because TNF-α level was attenuated in MyD88-deficient mice fed with HFD.Conclusions/SignificanceThe present finding of an unexpected role for MyD88 in preventing diabetes may provide a potential novel target/strategy for treating metabolic syndrome.