No abstract

Males are much more susceptible to ischemia/reperfusion (I/R)-induced kidney injury when compared with females. Recently we reported that the presence of testosterone, rather than the absence of estrogen, plays a critical role in gender differences in kidney susceptibility to I/R injury in mice. Although reactive oxygen species and antioxidant defenses have been implicated in I/R injury, their roles remain to be defined. Here we report that the orchiectomized animal had significantly less lipid peroxidation and lower hydrogen peroxide levels in the kidney 4 and 24 h after 30 min of bilateral renal ischemia when compared with intact or dihydrotestosterone-treated orchiectomized males. The post-ischemic kidney expression and activity of manganese superoxide dismutase (MnSOD) in orchiectomized mice was much greater than in intact or dihydrotestosterone-administered orchiectomized mice. Four hours after 30 min of bilateral ischemia, superoxide formation was significantly lower in orchiectomized mice than in intact mice. In Madin-Darby canine kidney cells, a kidney epithelial cell line, 1 mM H 2 O 2 decreased MnSOD activity, an effect that was potentiated by pretreatment with dihydrotestosterone. Orchiectomy prevented the post-ischemic decrease of catalase activity. Treatment of male mice with manganese(III) tetrakis(1-methyl-4-pyridyl)porphyrin (MnTMPyP), a SOD mimetic, reduced the post-ischemic increase of plasma creatinine, lipid peroxidation, and tissue hydrogen peroxide. These results suggest that orchiectomy accelerates the post-ischemic activation of MnSOD and reduces reactive oxygen species and lipid peroxidation, resulting in reduced kidney susceptibility to I/R injury.Gender differences in disease susceptibility have been well characterized in many cardiovascular diseases (1-3). The increased risk for cardiovascular diseases in men and postmenopausal women (4) has been attributed primarily to lack of estrogen-mediated protection (5-7). Recently we demonstrated that male mice are much more susceptible to kidney ischemia/reperfusion (I/R) 5 injury when compared with female mice (8), and that orchiectomy decreases kidney susceptibility to I/R injury (9), whereas ovariectomy has no effect on kidney susceptibility to I/R injury (8). Testosterone administration to orchiectomized male or female mice reverses the protective phenotype, increasing susceptibility to kidney I/R injury. We concluded that the presence of testosterone, rather than the absence of estrogen, plays a critical role in the gender differences in kidney susceptibility to I/R injury. The detailed molecular mechanisms responsible for this testosterone effect remain to be defined.Ischemia/reperfusion markedly increases the production of reactive oxygen species (ROS) including superoxide anions, hydroxyl radicals, hypochlorous acid, hydrogen peroxide, and peroxynitrite (10). The abnormal excessive production of ROS results in lipid peroxidation, leukocyte activation, endothelial cell damage, and cytokine production, all of which contribute to tissue ...

Alzheimer's disease (AD) is characterized by the deposition of aggregated beta-amyloid (Aβ), which triggers a cellular stress response called the unfolded protein response (UPR). The UPR signaling pathway is a cellular defense system for dealing with the accumulation of misfolded proteins but switches to apoptosis when endoplasmic reticulum (ER) stress is prolonged. ER stress is involved in neurodegenerative diseases including AD, but the molecular mechanisms of ER stress-mediated Aβ neurotoxicity still remain unknown. Here, we show that treatment of Aβ triggers the UPR in the SK-N-SH human neuroblastoma cells. Aβ mediated UPR pathway accompanies the activation of protective pathways such as Grp78/Bip and PERK-eIF2α pathway, as well as the apoptotic pathways of the UPR such as CHOP and caspase-4. Knockdown of PERK enhances Aβ neurotoxicity through reducing the activation of eIF2α and Grp8/Bip in neurons. Salubrinal, an activator of the eIF2α pathway, significantly increased the Grp78/Bip ER chaperone resulted in attenuating caspase-4 dependent apoptosis in Aβ treated neurons. These results indicate that PERK-eIF2α pathway is a potential target for therapeutic applications in neurodegenerative diseases including AD.

The role of phospholipase A2 (PLA2) in lung injury in humans is unclear. Previous studies have failed to identify an increase in PLA2 activity in bronchoalveolar lavage fluids (BALF) of patients with the adult respiratory distress syndrome (ARDS). In this study, increased phospholipase A2 (PLA2) activity was detected in BALF from patients with ARDS. PLA2 levels in BALF correlated positively with lung injury score in patients with lung disease. BALF PLA2 activity in patients with ARDS was resolved into heparin binding and nonbinding activities. Both PLA2 activities were increased in BALF of ARDS patients. The PLA2 activity that bound to heparin was identified as a group II PLA2 by its chromatographic characteristics, its inhibition by dithiothreitol, its substrate specificity, and its approximate molecular mass of 14 kDa. The second PLA2 activity was further purified and found to require Ca2+ at a concentration > 2 x 10(-4) M for activity. This form of PLA2 exhibited a neutral and broad pH optimum (pH 6.0-8.0) and hydrolyzed both phosphatidylethanolamine and phosphatidylcholine effectively. Its apparent molecular mass was estimated to be 80-90 kDa. Neither anti-pancreatic PLA2 antiserum nor anti-pig spleen cytosolic 100-kDa PLA2 antiserum immunoprecipitated the enzymatic activity. Thus at least two forms of PLA2 are increased in activity in BALF of patients with ARDS, a group II PLA2 and a biochemically and immunochemically form distinct from group I, group II, and cytosolic PLA2. Increased lung PLA2 activity may be important for the pathophysiology of ARDS.

Under low oxygen tension, cells increase the transcription of specific genes involved in angiogenesis, erythropoiesis, and glycolysis. Hypoxia-induced gene expression depends primarily on stabilization of the ␣ subunit of hypoxia-inducible factor-1 (HIF-1␣), which acts as a heterodimeric trans-activator with the nuclear protein known as the aryl hydrocarbon receptor nuclear translocator (Arnt). The resulting heterodimer (HIF-1␣/Arnt) interacts specifically with the hypoxia-responsive element (HRE), thereby increasing transcription of the genes under HRE control. Our results indicate that the 90-kDa heat-shock protein (Hsp90) inhibitor radicicol reduces the hypoxia-induced expression of both endogenous vascular endothelial growth factor (VEGF) and HRE-driven reporter plasmids. Radicicol treatment (0.5 g/ml) does not significantly change the stability of the HIF-1␣ protein and does not inhibit the nuclear localization of HIF-1␣. However, this dose of radicicol significantly reduces HRE binding by the HIF-1␣/Arnt heterodimer. Our results, the first to show that radicicol specifically inhibits the interaction between the HIF-1␣/Arnt heterodimer and HRE, suggest that Hsp90 modulates the conformation of the HIF-1␣/Arnt heterodimer, making it suitable for interaction with HRE. Furthermore, we demonstrate that radicicol reduces hypoxia-induced VEGF expression to decrease hypoxia-induced angiogenesis.Cells adapt to hypoxia by up-regulating the transcription of specific genes involved in angiogenesis, erythropoiesis, and glycolysis. Pathologically, tumor hypoxia contributes directly to enhanced glucose metabolism and angiogenesis, which are major features of malignant progression. The genes up-regulated during hypoxia include vascular endothelial growth factor (VEGF), erythropoietin, and several glycolytic enzymes. These diverse, targeted genes are induced by a common trans-activator, hypoxia-inducible factor 1 (HIF-1) (Iyer et al., 1998;Bruick and McKnight, 2001b;Semenza, 2002).HIF-1 was first identified as a heterodimeric trans-activator composed of two subunits, HIF-1␣ and -␤, both of which belong to the growing family of basic-helix-loop-helix-PAS (bHLH-PAS) proteins, including period (Per), Arnt, and single-minded (Sim). The bHLH-PAS proteins share common characteristics: first, a bHLH-PAS protein dimerizes with a specific partner protein through the HLH-PAS domain. Second, a partner such as the aryl hydrocarbon receptor (AhR) or HIF-1␣ is activated by specific stimuli (i.e., xenobiotics or low oxygen tension, respectively) before translocating to the nucleus, where it heterodimerizes with a partner protein. Alternatively, Arnt, another bHLH-PAS protein, is constitutively located in the nucleus and interacts with several