Albena Alexandrova

Effects of proteasome inhibitor, MG132, on proteasome activity and oxidative status of rat liver.

In vivo effects of N‐benzyloxycarbonyl (Cbz)‐Leu–Leu‐leucinal (MG132) on chymotryptic‐like (ChT‐L), tryptic‐like, and post‐glutamyl peptide hydrolytic‐like proteasome activities, protein oxidation, lipid peroxidation (LP), glutathione (GSH) level, as well as on the activity of antioxidant enzymes (superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH‐Px), and glutathione‐reductase) in the rat liver were studied. The possibility of MG132 provoking the formation of free oxygen radicals was also assayed in primary hepatocytes. The following results were obtained: (1) In vivo, MG132 did not change the spontaneous LP, but increased Fe‐induced LP and the amount of oxidized proteins; it decreased the GSH level in liver. From the proteasome activities studied in liver cytosol only ChT‐L activity was significantly decreased after MG132 administration. Furthermore, MG132 increased antioxidant enzyme activities of SOD, CAT, and GSH‐Px. (2) In vitro, MG132 increased free radical oxygen species in hepatocytes; this effect disappeared in the presence of CAT or mannitol. In conclusion, since nowadays proteasome inhibitors are entering into the swing of laboratory and clinical practice, the present data could provide useful information for MG132 action. Consequently, future in vivo experiments with MG132 could highlight the possibility of its use at different pathological conditions. Copyright

In vivo effects of pentoxifylline on enzyme and non-enzyme antioxidant levels in rat liver after carrageenan-induced paw inflammation

The present study aimed to investigate the effects of pentoxifylline (PTX) on the carrageenan (CG)‐induced paw oedema and on the endogenous levels of cell enzyme and non‐enzyme antioxidants in rat liver, 4 and 24 h after CG injection. PTX (50 mg kg−1, i.p.), administered 30 min before CG, decreased the paw oedema, 2–4 h after CG administration. The drug protected CG‐induced decrease of glutathione (non‐enzyme antioxidant) and had no effect on CG‐unchanged activities of superoxide dismutase, glutathione peroxidase (enzyme antioxidants) and glucose‐6‐phosphate dehydrogenase (enzyme, important for the activity of GSH‐conjugated antioxidant enzymes). The drug showed a good antioxidant capacity in chemical systems, generating reactive oxygen species. The present results suggest that the antioxidant activity of PTX might contribute to its beneficial effects in liver injuries. Copyright

Effect of MG132 on proteasome activity and prooxidant/antioxidant status of rat liver subjected to ischemia/reperfusion injury.

Aim:  Previous studies have shown that proteasome inhibitors exerted protective effects against ischemia/reperfusion injury (IRI) of brain, heart, kidney and intestine. The aim of the present study was to investigate: (i) whether the proteasome inhibitor MG132 protects rat liver against IRI; and (ii) whether MG132 modulates prooxidant/antioxidant status of rat liver subjected to warm IRI.

Effects of desipramine on the antioxidant status in rat tissues at carrageenan-induced paw inflammation

The pathogenesis of many diseases and different pathological conditions, including inflammation, is associated with excess production of reactive oxygen species (ROS). The present study aimed to investigate the effects of the antidepressant desipramine (DES) on carrageenan (CG)‐induced inflammation, as well as on the endogenous levels of cell enzyme and non‐enzyme antioxidants in rat liver and spleen, 4 and 24 h after CG injection. The intra‐plantar CG injection into the right hind paw resulted in a time‐dependent increase in the paw volume; the maximum of CG‐induced edema peak was in 2–4 h. A single DES dose of 20 mg·kg−1, administered 30 min before CG, had no effect on paw edema, whereas the higher drug dose used (50 mg·kg−1) suppressed the edematous response to CG. The latter drug dose protected CG‐induced decrease of glutathione (non‐enzyme antioxidant) in the liver; it did not affect CG‐unchanged activities of superoxide dismutase, glutathione peroxidase (enzyme antioxidants) and glucose‐6‐phosphate dehydrogenase (enzyme, important for the activity of glutathione‐conjugated antioxidant enzymes) in both liver and spleen. The drug showed an efficient antioxidant capacity in ROS‐generating chemical systems; it was higher than that of fluoxetine (another type of antidepressant). The present results suggest that the good antioxidant activity of DES might contribute to its beneficial effects in liver injuries. Copyright

Effect of copper intoxication on rat liver proteasome activity: relationship with oxidative stress.

Copper toxicity is associated with formation of reactive oxygen species, which are capable to oxidize proteins. The selective removal of the latter by the 20S proteasome is considered an essential part of the cell antioxidant defense system. The aim of the present study was to investigate whether peptidase activities of rat liver proteasomes were affected by chronic (40 mg CuSO4/rat/daily with the drinking water for 2 weeks) and acute (20 mg/kg CuSO4, s.c.) copper treatment. To evaluate the role of proteasome, its inhibitor MG132 was also used. The degree of copper‐induced oxidative stress (OS), established by measuring lipid peroxidation, protein oxidation, and cellular glutathione level, as well as activities of antioxidant enzymes—catalase, superoxide dismutase, and gultathionine peroxidase, depended on the mode of copper administration. Chronic copper administration (mild oxidative stress) did not affect proteasome activities, whereas acute copper treatment (severe oxidative stress) caused a decline in chymotryptic‐ and tryptic‐like activities. The treatment of copper‐loaded animals with MG132 did not change copper‐induced alterations in the tested indices, except an additional increase in protein oxidation and inhibition of glutathionine peroxidase activity. The results suggested that the in vivo copper‐induced oxidative stress was associated with changes in the catalytic activity of proteasome.

In vivo effects of N/OFQ(1–13)NH2 and its structural analogue [ORN9]N/OFQ(1–13)NH2 on carrageenan-induced inflammation: rat-paw oedema and antioxidant status

The effects of nociceptin(1–13)NH2 (N/OFQ(1–13)NH2) and its structural analogue [Orn9]N/OFQ(1–13)NH2 on acute carrageenan (CG)-induced peripheral inflammation and paw antioxidant status were studied. CG was injected intraplantarly in the right hind paw of rats and the volume of the inflamed paw was measured each 30 min for a period of 4h. When administered simultaneously with CG, N/OFQ(1–13)NH2 decreased the paw volume, whereas if injected 15 min before CG it had no effect. [Orn9]N/OFQ(1–13)NH2 produced the opposite effects at the same time-intervals of its administration. We also investigated whether these neuropeptides influence CG-induced changes in cell antioxidant system, especially at the 4th hour of CG administration. CG alone decreased the glutathione level and superoxide dismutase activity, as measured in post-nuclear homogenate of the inflamed paw. However, CG injection increased glutathione peroxidase and glucose-6-phospate dehydrogenase activities, while the activity of glutathione reductase was unchanged. The peptides themselves did not change all measured parameters. Moreover, neither N/OFQ(1–13)NH2 nor [Orn9]N/OFQ(1–13)NH2 modified CG-induced changes in the antioxidant status, regardless of the time of their injection (simultaneously or 15 min before CG). The present results suggest that N/OFQ(1–13)NH2 and [Orn9]N/OFQ(1–13)NH2 most likely affect the neuronal inflammation, rather than act as pro- or antioxidants.

Role of Trace Elements for Oxidative Status and Quality of Human Sperm

Background: Oxidative stress affects sperm quality negatively. To maintain the pro/antioxidant balance, some metal ions (e.g. copper, zink, iron, selenium), which are co-factors of the antioxidant enzymes, are essential. However, iron and copper could act as prooxidants inducing oxidative damage of spermatozoa. Aims: To reveal a possible correlation between the concentrations of some metal ions (iron, copper, zinc, and selenium) in human seminal plasma, oxidative stress, assessed by malondialdehyde and total glutathione levels, and semen quality, assessed by the parameters count, motility, and morphology. Study Design: Descriptive study. Methods: The semen analysis for volume, count, and motility was performed according to World Health Organization (2010) guidelines, using computer-assisted semen analysis. For the determination of spermatozoa morphology, a SpermBlue staining method was applied. Depending on their parameters, the sperm samples were categorized into normozoospermic, teratozoospermic, asthenoteratozoospermic, and oligoteratozoospermic. The seminal plasma content of iron, copper, zinc, and selenium was estimated by atomic absorption spectroscopy. The malondialdehyde and total glutathione levels were quantified spectrophotometrically. Results: In the groups with poor sperm quality, the levels of Fe were higher, whereas those of Zn and Se were significantly lower than in the normozoospermic group. In all groups with poor sperm quality, increased levels of malondialdehyde and decreased glutathione levels were detected as evidence of oxidative stress occurrence. All these differences are most pronounced in the asthenoteratozoospermic group where values differ nearly twice as much compared to the normozoospermic group. The Fe concentration correlated positively with the malondialdehyde (r=0.666, p=0.018), whereas it showed a negative correlation with the level of total glutathione (r=-0.689, p=0.013). The total glutathione level correlated positively with the sperm motility (r=0.589, p=0.044). Conclusion: The elevated levels of Fe and the reduced Se levels are associated with sperm damage. The changes in the concentrations of the trace elements in human seminal plasma may be related to sperm quality since they are involved in the maintenance of the pro-/antioxidative balance in ejaculate.

In-vivo effects of nociceptin and its structural analogue [Orn9] nociceptin on the antioxidant status of rat blood and liver after carrageenan-induced paw inflammation

The production of reactive oxygen species (ROS) in cells is well balanced with their elimination by the antioxidant defence system. This balance is essential for maintenance of physiological conditions, and its disturbance (oxidative stress) has been suggested as a potential pathogenic mechanism in a variety of diseases, accompanied by inflammation. In this study, the in-vivo effects of nociceptin (N/OFQ(1–13)NH2) and its structure analogue [Orn9]N/OFQ(1–13)NH2 were studied on markers of oxidative stress in erythrocytes and liver of rats 4 hours after subplantar administration of carrageenan (CG) (1%, 100 µl) in the right hind paw. A considerable inflammatory oedema of the paw was observed. CG did not change blood haemoglobin content, hematocrit value, glutathione level and antioxidant enzyme activities in the erythrocytes, but there was an increase in lipid peroxidation. In liver, CG-induced imbalance was manifested by an increase in lipid peroxidation and a decrease in glutathione level. Both peptides (20 µg, i.p.), when administered alone, had no effect on all parameters tested. When either [Orn9]N/OFQ(1–13)NH2 or N/OFQ(1–13)NH2 was injected simultaneously with CG or 15 minutes before it, they did not affect the CG-induced changes in the antioxidant status of the erythrocytes and liver. Our results suggest that the peptides tested did not play a role in the free radical processes that accompany CG-induced paw inflammation.

Are nociceptin(1-13)NH2 and its structural analogue [ORN9]nociceptin(1-13)NH2 able to affect brain antioxidant status in control and kainic acid-treated rats?

In‐vivo effects of nociceptin (N/OFQ(1‐13)NH2) on the levels of lipid peroxidation and cell enzyme (superoxide dismutase, glutathione peroxidase and glutathione reductase) and non‐enzyme (glutathione) antioxidants in brain of control and kainic acid‐treated rats were studied. N/OFQ(1‐13)NH2 effects were compared with those of its structural analogue [Orn9]N/OFQ(1‐13)NH2. Kainic acid (25 µg, i.c.v) increased the lipid peroxidation (4 and 24 h after kainic acid treatment) and decreased the glutathione level (1 h after kainic acid injection). We failed to find, any changes in antioxidant enzyme activities, independently of the time of kainic acid treatment. At the background of kainic acid‐effects, N/OFQ(1‐13)NH2 and [Orn9] N/OFQ(1‐13)NH2, injected 30 min before kainic acid, had no effects on all parameters, tested in brain. In addition, the neuropeptides did not change the antioxidant status in brain of control animals. It might be concluded that N/OFQ(1‐13)NH2 and [Orn9]N/OFQ(1‐13)NH2 have neither pro‐ nor anti‐oxidant activity. Copyright

Effects of structural analogues of nociceptin(1–13)NH2 on brain antioxidant status in kainic acid‐treated rats

The in vivo effects of nociceptin (N/OFQ(1–13)NH2) and its structural analogues ([Dab9]N/OFQ(1–13)NH2, [Dap9]N/OFQ(1–13)NH2 and [Cav9]N/OFQ(1–13)NH2) on the levels of lipid peroxidation and cell antioxidants (enzyme and non‐enzyme) in brain of control and kainic acid (KA)‐treated rats were studied. In control animals, [Dab9]N/OFQ(1–13)NH2 and [Dap9]N/OFQ(1–13)NH2, unlike N/OFQ(1–13)NH2 and [Cav9]N/OFQ(1–13)NH2, slightly increased the brain lipid peroxidation; the rest of the parameters were unchanged by all neuropeptides tested. KA (0.25 µg in 0.5 µl, i.c.v) increased the lipid peroxidation (4 and 24 h after KA‐injection) and decreased the glutathione level (1 h after KA‐administration). One hour after KA‐administration, the neuropeptides (2 µg in 0.5 µl, injected 30 min before KA) showed the following effects: a slight decrease in the KA‐induced lipid peroxidation by all nociceptin analogues and an enhancement of the KA‐decreased GSH level, but by [Cav9]N/OFQ(1–13)NH2 only. The brain antioxidant enzyme activities were unchanged in all used experimental groups. In addition, the nociceptin analogues, especially [Can9]N/OFQ(1–13)NH2, showed a good antioxidant capacity in chemical systems, generating reactive oxygen species. In conclusion, the substitution of lysin (Lys) in N/OFQ(1–13)NH2 molecule with other amino acids might contribute to changes in its antioxidant properties. Copyright