Inhibition of the 26S and 20S Proteasomes by the Site-specific Inhibitors

OBJECTIVE: The study was performed to establish the purification processes of both 26S and 20S proteasomes, also to investigate the inhibitory properties and patterns of two different proteasome inhibitors on the isolated proteasomes. METHODS: The 26S and 20S proteasomes were purified respectively using liquid chromatographies and glycerol density gradient fractionation. The inhibitory patterns and kinetics of two different proteasome inhibitors were investigated using purified 26S and 20S proteasomes. RESULTS: The purity of the isolated proteasomes were determined by their biochemical properties and electrophoretic patterns. 3-nitro-4-hydroxy-5-indophenylacetyl-leucyl-leucyl-leucyl-vinylsulfone (Nip-L₃-VS) inhibited exclusively the chymotrypsin-like peptidase activities of the 26S and 20S proteasomes. On the other hand, dansyl-phenylyl-leucyl-boronic acid (DFLB) inhibited chymotrpsin-like, trypsin-like, and caspase-like peptidase activities of both proteasomes with different sensitivity. CONCLUSION: The proposed purification method provides efficient separation and isolation of the 26S and 20S proteasomes. Nip-L₃-VS and DFLB were shown to have different inhibitory effects and kinetics on the peptidase activities of the isolated proteasomes. These studies are suggested to be applied to the researches on proteasome inhibitors as therapeutic reagents for many related diseases.

Relationship between Pulmonary Surfactant Protein and Lipid Peroxidation in Lung Injury due to Paraquat Intoxication in Rats

BACKGROUND: Pulmonary damage resulting from lipid peroxidation is a principal effect of paraquat intoxication. The host-defense functions of surfactant are known to be mediated by the surfactant proteins A and D (SP-A and SP-D, respectively). The primary objective of this study was to evaluate the variations over time in levels of surfactant protein and lipid peroxidation (LPO) in lung tissue following free-radical-induced injury. METHODS: 42 adult, male, Sprague-Dawley rats were administered intraperitoneal injections of paraquat (35 mg/kg body weight). SP-A and SP-D levels were determined via Western blot. LPO in the left lung homogenate was measured via analyses of the levels of thiobarbituric acid-reactive substances. RESULTS: LPO levels peaked at 6 hours, with no associated histological changes. SP-D levels increased until hour 12 and declined until hour 48; SP-D levels subsequently began to increase again, peaking at hour 72. SP-A levels peaked at hour 6, declining thereafter. CONCLUSIONS: We suggest that in the early phase of paraquat injury, SP-D levels reflect alveolar damage and that de novo synthesis of SP-D takes 72 hours. Levels of SP-A, on the other hand, reflect abnormalities in the surfactant system in the late stage of paraquat intoxication. Surfactant proteins may play a role in protecting the lungs from reactive oxygen injury. A time-dependent variation has been observed in the levels of surfactant proteins A and D following paraquat injury, and it has been suggested that these proteins play a role in the protection of lung tissue against ROS-induced injuries.

Body Weight, Body Mass Index, Plasma Leptin, Insulin and Fasting Glucose Level in Schizophrenic Patients Receiving Olanzapine / 신경정신의학

OBJECTIVE: The aim this study was to investigate possible mechanisms behind olanzapine-associated weight gain. METHOD: Thirteen psychiatric inpatients (all meeting DSM-IV criteria for schizophrenia or schizophreniform disorder) receiving olanzapine in Soonchunhyang university Chun-an hospital were studied. Body weight, plasma leptin, insulin and fasting glucose levels were measured five-times over 6 weeks. Body mass index (BMI) was calculated. RESULT: At the end of sixth week, body weight, BMI, plasma leptin, insulin and fasting glucose levels were significantly increased from baseline. Both insulin and leptin plasma levels were significantly increased from baseline at the end of the sixth week of treatment. Both body weight and BMI were significantly increased from baseline at the end of the first week, the fourth week and sixth week, respectively. Fasting glucose was significantly increased from baseline at the end of the fourth week and sixth week, respectively. Between baseline and sixth week, the body weight, BMI, plasma leptin and fasting glucose levels were significantly correlated to insulin level. There was a tendency toward a correlation between BMI and leptin level, whereas no correlation was found between body weight and leptin. CONCLUSION: Olanzapine treatment was associated with weight gain and elevated level of leptin, insulin and fasting glucose. Elevated insulin levels and hyperleptinemia may be mechanisms behind olanzapine-induced weight gain.

Nickel (II)-induced apoptosis and G2/M enrichment

Treatment with certain DNA-damaging agents induce a complex cellular response comprising pertubation of cell cycle progression and/or apoptosis on proliferating mammalian cells. Our studies were focused on the cellular effects of nickel (II) acetate, DNA-damaging agent, on Chinese hamster ovary (CHO) cells. Fragmented DNAs were examined by agarose gel electrophoresis and cell cycle was determined by DNA flow cytometry using propidium iodide fluorescence. Apparent DNA laddering was observed in cells treated with 240 microM nickel (II) and increased with a concentration-dependent manner. Treatment of nickel (II) acetate resulted in apoptosis which was accompanied by G2/M cell accumulation. Proportion of CHO cells in G2/M phase was also significantly increased in cells exposed to at least 480 microM nickel (II) from 57.7% of cells in the G0/G1 phase, 34.7% in the S phase, and 7.6% in the G2/M1 phase for 0 microM nickel (II), to 58.6%, 14.5%, and 26.9% for 640 microM nickel (II). These findings suggest that nickel (II) can modulate cellular response through some common effectors involving in both apoptotic and cell cycle regulatory pathways.