Anna Merwid-Ląd

Effect of simvastatin on nitric oxide synthases (eNOS, iNOS) and arginine and its derivatives (ADMA, SDMA) in ischemia/reperfusion injury in rat liver.

Hydroxymethylglutaryl-CoA reductase inhibitors play a role in nitric oxide synthesis. In this study, the impact of simvastatin (SV) on the levels of nitric oxide synthases, and arginine (Arg) and its derivatives was evaluated in rat liver under ischemia-reperfusion (I/R) conditions. Rats received SV (25 mg/kg) (groups S and S-IR) or saline solution (groups C and C-IR) intragastrically for 21 days. The livers of groups C and S were homogenized after treatment while those of groups C-IR and S-IR underwent ischemia and reperfusion before homogenization. Endothelial (eNOS) and inducible (iNOS) nitric oxide synthase concentrations were determined in the homogenates. Alanine and asparagine aminotransferase (ALT, AST, respectively), arginine (Arg), and asymmetric (ADMA) and symmetric (SDMA) methylarginine levels were determined in the blood before I/R and during reperfusion. I/R injury produced significant increases in aminotransferase, ADMA, eNOS, and iNOS, but decreases in Arg and Arg/ADMA levels. Arg concentration increased significantly after warm ischemia in the S-IR group, but decreased significantly during the first 30 minutes of reperfusion in both the S-IR and C-IR groups. eNOS concentration was significantly higher in group S than in group C. Both I/R and SV exerted no influence on SDMA concentration. SV exerted a protective action by increasing eNOS levels under normal conditions and Arg levels after ischemia and by preventing a significant increase in iNOS concentration after I/R. SV had no effect on ADMA concentration under normal and pathological conditions.

The protective effect of the Cornus mas fruits (cornelian cherry) on hypertriglyceridemia and atherosclerosis through PPARα activation in hypercholesterolemic rabbits.

Cornelian cherry (Cornus mas L.) fruits have been used in traditional cuisine and in folk medicine in various countries. This study was conducted to evaluate the constituents and impact of cornelian cherry (C. mas L.) fruits lyophilisate on lipid levels, PPARα protein expression, atheromatous changes in the aorta, oxido-redox state, and proinflammatory cytokines in hypercholesterolemic rabbits. The HPLC-MS method was used for determining active constituents in cornelian cherry. In a subsequent in vivo study the protective effect of the cornelian cherry on diet-induced hyperlipidemia was studied using a rabbit model fed 1% cholesterol. Cornelian cherry (100mg/kg b.w.) or simvastatin (5mg/kg b.w.) were administered orally for 60 days. Two iridoids - loganic acid and cornuside - and five anthocyanins were identified as the main constituents of the cornelian cherry. The administering of the cornelian cherry led to a 44% significant decrease in serum triglyceride levels, as well as prevented development of atheromatous changes in the thoracic aorta. Cornelian cherry significantly increased PPARα protein expression in the liver, indicating that its hypolipidemic effect may stem from enhanced fatty acid catabolism. Simvastatin treatment did not affect PPAR-α expression. Moreover, the cornelian cherry had a significant protective effect on diet-induced oxidative stress in the liver, as well as restored upregulated proinflammatory cytokines serum levels. In conclusion, we have shown loganic acid to be the main iridoid constituent in the European cultivar of the cornelian cherry, and proven that the cornelian cherry could have protective effects on diet-induced hypertriglicerydemia and atherosclerosis through enhanced PPARα protein expression and via regulating oxidative stress and inflammation.

Effects of morin-5′-sulfonic acid sodium salt (NaMSA) on cyclophosphamide-induced changes in oxido-redox state in rat liver and kidney

Cyclophosphamide (CPX) is an anticancer drug with immunosuppressive properties. Its adverse effects are partly connected to the induction of oxidative stress. Some studies indicate that water-soluble derivative of morin—morin-5′-sulfonic acid sodium salt (NaMSA) exhibits strong antioxidant activity. The aim of present study was to evaluate the effect of NaMSA on CPX-induced changes in oxido-redox state in rat. Experiment was carried out on Wistar rats divided in three experimental groups (N = 12) receiving: 0.9% saline, CPX (15 mg/kg) or CPX (15 mg/kg) + NaMSA (100 mg/kg), respectively, and were given intragastrically for 10 days. Malondialdehyde (MDA) and glutathione (GSH) concentrations and superoxide dismutase (SOD) activity were determined in liver and kidneys. Catalase (CAT) activity was assessed only in liver. Treatment with CPX resulted in significant decrease in MDA level in both tissues, which was completely reversed by NaMSA treatment only in liver. In comparison to the control group significant decrease in SOD activity were observed in both tissues of CPX receiving group. In kidneys this parameter was fully restored by NaMSA administration. CPX evoked significant decrease in GSH concentration in kidneys, which was completely reversed by NaMSA treatment. No significant changes were seen in GSH levels and CAT activity between all groups in liver. Results of our study suggest that CPX may exert significant impact on oxido-redox state in both organs. NaMSA fully reversed the CPX-induced changes, especially MDA level in liver, SOD activity and GSH concentration in kidneys and it may be done by enhancement of activity/concentration of endogenous antioxidants.

Vipera berus bites in the Region of Southwest Poland--a clinical analysis of 26 cases.

OBJECTIVE Vipera berus is the only naturally occurring venomous snake in Poland. Its venom is primarily vasculotoxic and evokes both local and systemic findings. The aim of the study was to review a series of clinical cases of V berus bites occurring in southwest Poland. METHODS The charts of 26 patients (age range, 16-66 years; mean, 42 years) hospitalized with V berus bites were retrospectively analyzed using a data collection tool. Demographic and clinical data were extracted. RESULTS The most common local findings of envenomation were edema of the bitten limb with associated extravasations observed in 24 (92.3%) patients, but in only 1 (3.8%) case did the edema spread to the trunk. In 22 (84.6%) cases edema disappeared within 2 weeks after the bite. Systemic disturbances observed in the patients were: shock (1 case), mild transient hypotension (1 case), prolonged hypotension (3 cases), bronchospasm and laryngeal edema (1 case), diarrhea (1 case), transient supraventricular arrhythmias (2 cases), neutrophilic hyperleukocytosis (2 cases), and thrombocytopenia below 50000 cells/microL (5 cases). In 16 patients (61.5%) the envenomation was classified as moderate and this type was predominant. Six cases were classified as severe. No fatal case was reported. Treatment included the administration of specific antivenom in 14 cases (in all severe and half of moderate cases) and symptomatic treatment applied in all cases. CONCLUSIONS Moderate envenomation prevailed among the patients analyzed in the study. Antivenom treatment is primarily necessary in cases of severe (grade 3) and in some cases of moderate (grade 2) envenomation, especially in patients with persistent or recurring hypotension.

Influence of ezetimibe on selected parameters of oxidative stress in rat liver subjected to ischemia/reperfusion.

Introduction Ischemia/reperfusion (I/R) is considered to be one of the main causes of liver damage after transplantation. The authors evaluated the effect of ezetimibe on selected oxidative stress parameters in ischemic/reperfused (I/R) rat liver. Material and methods Rats were administered ezetimibe (5 mg/kg) (groups E and E-I/R) or saline solution (groups C and C-I/R) intragastrically for 21 days. Livers of animals in groups C-I/R and E-I/R were subjected to 60 min of partial ischemia (left lateral and median lobes) followed by 4 h of reperfusion. Alanine and asparagine aminotransferase (ALT, AST) activity was determined in blood before I/R and during reperfusion (at 15 and 240 min). After the reperfusion period, malondialdehyde (MDA), superoxide dismutase (SOD), glutathione (GSH) and glutathione peroxidase (GPx) were determined in liver homogenates using colorimetric methods. Results Ezetimibe caused a significant increase in GSH level in groups subjected to I/R (E-I/R (99.91 ±9.01) vs. C-I/R (90.51 ±8.87), p < 0.05). Additionally, under I/R the decrease of GPx activity in the drug-treated group was lower compared to the non-treated group (E-I/R (3.88 ±1.11) vs. E (5.31 ±1.83), p = 0.076). Neither ezetimibe nor I/R affected SOD or MDA levels. I/R produced a significant increase in aminotransferase levels (ALT240-0: C-I/R (42.23 ±43.56) vs. C (9.75 ±11.09), and E-I/R (39.85 ±26.53) vs. E (4.38 ±1.36), p < 0.05 in both cases; AST 240-0: E-I/R (53.87 ±17.23) vs. E (24.10 ±9.66), p < 0.05) but no effect of ezetimibe on those enzymes was found. Conclusions Ezetimibe demonstrates antioxidant properties in rat livers subjected to I/R. However, neither a hepatoprotective nor a hepatotoxic effect of ezetimibe was demonstrated, regardless of I/R.

Effect of simvastatin treatment on rat livers subjected to ischemia/reperfusion.

We evaluated the effect of simvastatin (SV) on the oxido-redox state in rat livers submitted to ischemia-reperfusion (I/R). Rats received SV (groups: S, S-IR) or saline solution (groups: C, C-IR) intragastrically (25 mg/kg) for 21 days. Before homogenization, rat livers (C-IR, S-IR) underwent ischemia (40 min) and reperfusion (60 min). Activities of such antioxidative enzymes as superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase (CAT) as well as lipid peroxides (LPO) level as indicator of oxidative stress were then estimated in the homogenates. All these parameters were measured spectrophotometrically. Additionally, alanine and asparagine aminotransferase (ALT, AST) levels were estimated in the blood before and after I/R. In groups C and S all examined parameters were similar regardless of SV-treatment. I/R produced significant increases in GPx and CAT activities only in the C-IR group. Conversely, GPx activity was significantly decreased and ALT and AST increased significantly in the S-IR group. SV did not evoke any noticeable protective changes in rat livers after 3 weeks of treatment. After I/R, some of the observed properties could suggest that SV may have even made liver function and the oxidative state worse.

Age-related changes in ADMA–DDAH–NO pathway in rat liver subjected to partial ischemia followed by global reperfusion

BACKGROUND Liver function is affected during ischemia/reperfusion (IR). We evaluated the effect of the aging process on selected parameters determining the NO level in rat liver subjected to IR. METHODS The animals were divided into the C-2 and the IR-2 group of young rats (2-4 months old) and the C-12 and the IR-12 group of older rats (12-14 months old). Livers belonging to the IR-2 and the IR-12 group were subjected to partial ischemia (60 min) and reperfusion (4 h). Blood samples were obtained after surgeries to estimate the activity of aminotransferases, as well as just before ischemia and during reperfusion (15, 120, and 240 min) to estimate concentration of arginine (Arg) and its derivatives: asymmetric and symmetric dimethylarginine (ADMA, SDMA). After IR, dimethylarginine dimethylaminohydrolase (DDAH) activity and protein concentration of inducible nitric oxide synthase (iNOS) were measured in liver homogenates. RESULTS In the IR-2 group ADMA level increased the most between 15 and 120 min of reperfusion and was the highest of all the groups (0.72±0.2 μmol/l). In the IR-12 group ADMA level decreased significantly and was lower compared to all the other groups at 15 min (0.42±0.2 μmol/l) and to IR-2 at 120 (0.52±0.1 μmol/l) and 240 min (0.38±0.1 μmol/l) of reperfusion. Only the IR-2 group SDMA level increased significantly between 15 (0.75±0.9 μmol/l) and 240 min (1.0±1.2 μmol/l) of reperfusion. At the beginning of the surgery the Arg level was significantly higher in young rats (C-2: 102.1±35.7 μmol/l; IR-2: 114.63±28.9 μmol/l) than in older ones (C-12: 41.88±44.7 μmol/l; IR-12: 28.64±30.6 μmol/l). In the C-2 group the Arg level (77.41±37.5 μmol/l) and Arg/ADMA (A/A) ratio (138.03±62.8 μmol/l) were significantly higher compared to the ischemic groups at 15 min and to all the other groups at 120 (Arg: 47.17±31.7 μmol/l; A/A: 88.28±66.2 μmol/l) and 240 min (Arg: 43.87±21.9 μmol/l; A/A: 118.02±106.3 μmol/l). In the IR-2 group Arg level (11.4±12.0 μmol/l) and A/A ratio (16.11±16.2 μmol/l) decreased significantly at 15 min and during the next phase of reperfusion the levels of those parameters were low, comparably to those in IR-12. As a result of IR, a decrease in DDAH activity and an increase in iNOS protein concentration were observed only in the young rats. CONCLUSIONS We found that in the non-ischemic groups the Arg level may be affected by the aging process. Under IR conditions, important changes in DDAH-ADMA-NO pathway were observed only in young livers.

Influence of ezetimibe on ADMA-DDAH-NO pathway in rat liver subjected to partial ischemia followed by global reperfusion

BACKGROUND We evaluated effect of ezetimibe on selected parameters determining NO level in rat liver subjected to ischemia reperfusion (IR). METHODS Rats received ezetimibe (5 mg/kg) (groups E0 and EIR) or saline solution (groups C0 and CIR) intragastrically for 21 days. Then, the livers of CIR and EIR underwent ischemia (60 min) and reperfusion (4 h). Blood samples were obtained before surgery to estimate activities of aminotransferases, and just before ischemia and during reperfusion to estimate asymmetric and symmetric dimethylarginine (ADMA, SDMA) and arginine (Arg) levels. After IR, dimethylarginine dimethylaminohydrolase (DDAH) activity and endothelial nitric oxide synthase (eNOS) protein concentration were measured in liver homogenates. DDAH and protein arginine methyltransferase (PRMT) mRNA were quantified by real-time PCR in liver tissue samples. RESULTS In CIR, the ADMA level was significantly higher compared to all other groups in 30 min and to E0 group in 120 min of reperfusion. In EIR, ADMA was low, compared to non-ischemic groups. At 30 and 120 min of reperfusion, in non-ischemic groups the level of Arg and Arg/ADMA ratio were significantly higher than in ischemic groups and E0 was the group with the highest levels of those parameters of all. In CIR, eNOS protein concentration was significantly lower than in ezetimibe-treated groups. Activity of DDAH was significantly higher in E0 than in non-treated groups. In ischemic groups, DDAH mRNA expression was significantly higher than in non-ischemic ones and PRMT mRNA expression was significantly higher in C0 than in all other groups. CONCLUSIONS Influence of ezetimibe on ADMA/DDAH/NO pathway demonstrated in this work may suggest protective properties of this drug on rat livers injured by IR and, to a lower extent, on livers non-subjected to IR.

Omeprazole does not change the oral bioavailability or pharmacokinetics of vinpocetine in rats

Previous studies proved that food strongly enhanced the bioavailability of vinpocetine. Food may change the pharmacokinetics of a drug by affecting various factors, including gastrointestinal pH. However, the influence of proton pump inhibitor-induced pH alterations on vinpocetine pharmacokinetics is not known. The aim was to evaluate the influence of omeprazole on the pharmacokinetics of oral vinpocetine. One group of male Wistar rats received single oral doses of vinpocetine (2 mg/kg - regimen V). In the second group, omeprazole (10 mg/kg) was administered intraperitoneally for 5 days before vinpocetine administration (regimen OV). For analysis of vinpocetine pharmacokinetics, blood samples were obtained before and 0.25, 0.5, 1, 1.5, 2, 3, 4, 5, 6, 8, 10 and 12 h after vinpocetine administration. Vinpocetine concentrations were measured by high performance liquid chromatography (HPLC). The mean values of AUC(0-t), AUC(0-inf) and C(max) in regimen V were very similar to respective values in regimen OV. The mean T(max) in both regimens was estimated for 1.5 h. There were no statistically significant differences between both regimens. In conclusion, omeprazole did not affect the pharmacokinetic profile of vinpocetine.

The influence of bexarotene, a selective agonist of the retinoid receptor X (RXR), and tazarotene, a selective agonist of the retinoid acid receptor (RAR), on bone metabolism in rats

PURPOSE The purpose of this study was to investigate the influence of selective agonists of the retinoid receptor X (RXR) and the retinoid acid receptor (RAR) on bone metabolism in rats. METHODS Thirty six male Wistar rats were divided into three groups: receiving bexarotene, or tazarotene, or to control group. Serum biochemical markers of bone turnover (osteocalcin - OC, tartrate resistant acid phosphatase 5 - TRACP5b and osteoprotegerin - OPG) and mechanical properties of bones were analyzed. RESULTS There was a significant decrease in the femur index value in groups receiving tazarotene and bexarotene on Day 14 (8% and 20% respectively, p=0.0039). On Day 28, 14 days after discontinuation of tazarotene and bexarotene, the difference in femur indexes was still significant (4% for T1-6 and B1-6, p=0.0270). In the bexarotene group an increase in mean plasma osteocalcin level and mean plasma TRACP5b level was detected. In the tazarotene group the mean osteocalcin level remained unchanged and the mean plasma TRACP5b level decreased. An increased yield stress was detected in groups receiving retinoids comparing to controls after 14 days of tazarotene and bexarotene administration. CONCLUSION Although bexarotene and tazarotene administration caused decrease in the femur index, mechanisms responsible for that effect seem to be different. Our results suggest that bexaroten increases bone turnover. On the contrary, tazaroten seems to have inhibitory effect on bone turnover. A counter influence of selective RAR and RXR agonists on the bone turnover might be the reason for inconsistency in results from published research concerning the influence of retinoids on bone metabolism.