Differential response of antioxidative systems of maize (Zea mays L.) roots cell walls to osmotic and heavy metal stress.

An analysis of peroxidase and ascorbate oxidase activity, phenolic content and antioxidant capacity of isolated maize root cell walls was performed in controls and plants stressed with polyethylene glycol (PEG) or heavy metals, zinc or copper. Peroxidase activity (oxidative and peroxidative) was more pronounced in the ionic than in the covalent cell wall fraction. PEG induced an increase and Zn(2+) a decrease of both ionically bound peroxidase activities. In the covalent fraction, Cu(2+) decreased oxidative and increased peroxidative activity of peroxidase. Isoelectric focusing of ionically bound proteins and activity staining for peroxidase demonstrated increased intensities and appearance of new acidic isoforms, especially in Zn(2+) and PEG treatments. Most pronounced basic isoforms (pI ~ 7.5) in controls, decreased in intensity or completely disappeared in stressed plants. Ascorbate oxidase activity was significantly increased by PEG and decreased by Zn(2+) treatments, and highly correlated with peroxidase activity. Antioxidant capacity and total phenolics content increased in heavy metal-treated and decreased in PEG-treated plants. Analysis of individual phenolic components revealed p-coumaric and ferulic acids, as the most abundant, as well as ferulic acid dimers, trimers and tetramers in the cell walls; their quantity increased under stress conditions. Results presented demonstrate the existence of diverse mechanisms of plant response to different stresses.

Effects of conventional versus biocompatible peritoneal dialysis solutions on peritoneal and systemic inflammation, malnutrition and atherosclerosis in CAPD patients.

BACKGROUND: Chronic inflammation, malnutrition and atherosclerosis (MIA syndrome) are important predictors of high mortality in continuous ambulatory peritoneal dialysis (CAPD) patients. We aimed to evaluate the effects of PD solutions (standard vs. biocompatible) on some parameters of MIA syndrome in patients undergoing CAPD. METHODS: 42 stable patients who were on CAPD at least 2.5 years participated in this cross-sectional study. Patients who had severe anemia (Hb < 10 g/l), immunomodulatory therapy, peritonitis or any inflammatory conditions for at least 3 months before the analysis, malignant disease and acute exacerbation of heart failure, were excluded. 21 (50%) patients were treated with standard PD solutions (CAPDP-1), while the remaining 21 (50% of patients) were treated with biocompatible PD solutions (neutral solutions with lower level of glucose degradation products and lower concentration of calcium, CAPDP-2). All patients underwent echocardiography and B-mode ultrasonography of common carotid arteries together with assessments of nutrition status and parameters of systemic and local inflammation. RESULTS: There were no significant differences between the groups concerning age, gender, underlying disease, residual renal function, peritoneal transport characteristics, comorbidity or therapy applied. Patients from group CAPDP-2 had a significantly lower serum level of hs-CRP (3.7 ± 2.6 mg/l vs. 6.3 ± 4.5 mg/l; p = 0.023) and significantly better nutritional status confirmed by mid-arm circumference (p = 0.015), mid-arm muscle circumference (p = 0.002) and subjective global assessment (14.28% of patients in CAPDP-2 vs. 71% of patients in CAPDP-1 were malnourished; p = 0.000). Group CAPD-2 had less frequent left ventricular hypertrophy (p = 0.039), thinner intima-media thickness (p = 0.005), smaller carotid narrowing (p = 0.000) and fewer calcified plaques of common carotide arteries (p = 0.003). No significant difference between the CAPDP groups was observed in serum and effluent levels of inflammatory cytokines (IL-1, IL-6 and TNF-α) and CA-125 effluent level. Logistic regression analysis did not confirm that biocompatibility of PD solutions was an independent predictor of any parameter of MIA syndrome. CONCLUSIONS: According to the present study and logistic regression analysis, the effect of biocompatible CAPD solutions on parameters of malnutrition, inflammation and atherosclerosis have to be confirmed by well-designed and controlled studies in a higher number of patients.

Anti-tumor effect of Coriolus versicolor methanol extract against mouse B16 melanoma cells: in vitro and in vivo study.

Numerous studies have shown immunostimulatory and anti-tumor effects of water and standardized aqueous ethanol extracts derived from the medicinal mushroom, Coriolus versicolor, but the biological activity of methanol extracts has not been examined so far. In the present study we investigated the anti-tumor effect of C. versicolor methanol extract (which contains terpenoids and polyphenols) on B16 mouse melanoma cells both in vitro and in vivo. In vitro treatment of the cells with the methanol extract (25-1600 microg/ml) reduced melanoma cell viability in a dose-dependent manner. Furthermore, in the presence of the methanol extract (200 microg/ml, concentration IC(50)) the proliferation of B16 cells was arrested in the G(0)/G(1) phase of the cell cycle, followed by both apoptotic and secondary necrotic cell death. In vivo methanol extract treatment (i.p. 50 mg/kg, for 14 days) inhibited tumor growth in C57BL/6 mice inoculated with syngeneic B16 tumor cells. Moreover, peritoneal macrophages collected 21 days after tumor implantation from methanol extract-treated animals exerted stronger tumoristatic activity ex vivo than macrophages from control melanoma-bearing mice. Taken together, our results demonstrate that C. versicolor methanol extract exerts pronounced anti-melanoma activity, both directly through antiproliferative and cytotoxic effects on tumor cells and indirectly through promotion of macrophage anti-tumor activity.

Characterization of NAD-dependent malate dehydrogenases from spinach leaves.

Spinach leaves were used to extract isoforms of NAD-dependent malate dehydrogenase (NAD-MDH) (EC 1.1.1.37), either soluble or bound to microsomal, plasma, or chloroplast envelope membranes. All fractions were subjected to isoelectric focusing analysis, which showed that purified chloroplast envelopes contain an NAD-MDH isoform tightly bound to the membranes, since treatment with 0.5 or 1% Triton X-100 was not able to release the enzyme from the envelopes. In contrast, plasma membranes released an isoform with a pI of 3.5 following treatment with 0.5% Triton X-100. The most abundant soluble leaf isoform had a pI of 9, while the chloroplast stroma contained an isoform with a pI of 5.3. Kinetic analysis of oxaloacetate (OAA)-dependent NADH oxidation in different fractions gave different Km values for both substrates, the envelope- and plasma membrane-bound NAD-MDH exhibiting the highest affinities for OAA. Leaf plasma membrane-bound MDH exhibited a high capacity for both reaction directions (malate oxidation and OAA reduction), while the two chloroplast isoforms (stromal and envelope-bound) preferentially reduced OAA. Our results indicate that the chloroplast envelope contains a specifically attached NAD-MDH isoform that could provide direct coupling between chloroplast and cytosol adenylate pools.

Role of fructose in the adaptation of plants to cold-induced oxidative stress.

This work presents findings, which indicate important role of fructose, fructose 6-phosphate (F6P), and fructose 1,6-bisphosphate (FBP) in preservation of homeostasis in plants under low temperature. Cold combined with light is known to incite increased generation of superoxide in chloroplasts leading to photoinhibition, but also an increased level of soluble sugars. In the present study, oxidative stress in pea leaves provoked by cold/light regime was asserted by the observed decrease of the level of oxidized form of PSI pigment P700 (P700+). Alongside, the increased antioxidative status and the accumulation of fructose were observed. The antioxidative properties of fructose and its phosphorylated forms were evaluated to appraise their potential protective role in plants exposed to chilling stress. Fructose, and particularly F6P and FBP exhibited high capacities for scavenging superoxide and showed to be involved in antioxidative protection in pea leaves. These results combined with previously established links implicate that the increase in level of fructose sugars through various pathways intercalated into physiological mechanisms of homeostasis represents important non-enzymatic antioxidative defense in plants under cold-related stress.

Superoxide dismutase, peroxidase, and germin-like protein activity in plasma membranes and apoplast of maize roots.

The analysis of plasma membranes from maize roots by native gel electrophoresis revealed the existence of Mn-containing 120 kDa and CuZn-containing 70, 40, and 15 kDa superoxide dismutase (SOD) isoform activities. Isoelectric focusing of the plasma membranes differentiated anionic SOD isoforms with a pI of about 5 and cationic SOD isoforms at pI 8.6. Solubilization of the plasma membrane proteins further separated the cationic SOD into pI 8.6, 8.2, 8.4, and 7.2 isoforms. Double staining for both SOD and peroxidase activities showed an overlap of these activities only in the case of the high-molecular-mass (ca. 120 kDa) isoforms. High-temperature treatments demonstrated that the 120 kDa isoform was active even at 100 degrees C, indicating that it was a germin-like protein with superoxide-dismutating activity, different from the peroxidase with a similar molecular mass and the lower-molecular-mass CuZn-containing superoxide dismutases. These results are compared to those obtained from whole-tissue extract and apoplastic fluid.

Superoxide synthase and dismutase activity of plasma membranes from maize roots.

Superoxide synthase and superoxide dismutase activity have been monitored in isolated maize ( Zea mays) root plasma membranes spectrophotometrically by determination of nitro-blue tetrazolium and cytochrome c reduction, respectively. Superoxide production was induced by NADH and NADPH, with similar kinetics and approaching saturation at 0.06 mM in the case of NADPH and 0.1 mM in the case of NADH, with rates of 18.6 +/- 5.0 and 21.8 +/- 7.2 nmol/min. mg of protein, respectively. These activities exhibited a broad pH optimum between pH 6.5 and 7.5. Diphenylene iodonium inhibited about 25% (10 microM DPI) and 40% (100 microM DPI) of this activity, imidazole inhibited about 20%, while KCN, a peroxidase inhibitor, did not show any significant inhibition. Superoxide-dismutating activity was shown to occur in the same isolates and depended on the quantity of plasma membrane protein present. Growth of plants on salicylic acid prior to membrane isolation induced a rise in the activity of both of the enzymes by 20-35%, suggesting their coordinated action.

[Successful surgical removal of a cardiac myxoma from the left ventricular outflow tract]. / Uspesno hirurski odstranjen miksom srca u izlaznom trakty leve komore.

The precondition for successful, exclusively surgical treatment in the extracorporeal circulation, is the precise diagnosis of heart myxoma, particularly in rare locations such as the left ventricle. We present a case of myxoma in the outflow tract of the left ventricle as the exceptionally rare location, successfully diagnosed and surgically treated at the Clinic for Cardiac and Thoracic Surgery of the Military Medical Academy. A female patient, aged 46 years, was sent from another hospital with misdiagnosis of idiopathic hypertrophic subaortic stenosis. The patient was successfully operated after transthoracic and transesophageal echocardiography as the main diagnostic procedures. Myxoma that completely obstructed the aortic opening if pulled, was completely removed through aortic valve in the extracorporeal circulation. Its pedicle was arising from the ventricular side of the great mitral cusp. Postoperative course was uneventful and the patient was released from the hospital on the tenth postoperative day.

[A severe form of falciparum malaria associated with staphylococcal endocarditis]. / Teska forma faliciparum malarije udruzene sa stafilokoknim endokarditisom.

A case is presented of a patient, aged 56 years, with severe form of imported malaria caused by Plasmodia falciparum. Hyperparasitemia of erythrocytes > 30% was registered, and during the course of the disease CNS dysfunction, severe anemia, acute renal failure, disseminated intravenous coagulation with manifest hemorrhagic syndrome, icterus, enterocolitis, pneumonia and staphylococcal endocarditis were developed Due to hyperparasitemia and numerous complications, antimalarial drugs such as quinidine (1,200 mg/day) and artemether (160 mg/day) were administered parenterally. Infected erythrocytes were exchanged with 2.5 litres of healthy erythrocytes suspension. Hemodialysis was also performed as well as nine-week antistaphylococcal therapy. During the treatment preparation of deplasmated blood, concentrated thrombocytes, fresh frozen plasma, cryoprecipitates, human albumins and immunoglobulins were applied, along with the correction of electrolytic dysbalance, administration of diuretic, cardiotonic, antiarrhythmic, anxiolytic, antipsychotic and antidepressive drugs. Two months after the admission the patient was released from the Clinic in good condition, with normal clinical-laboratory findings.

Factors influencing the degree and outcome of acute beta-blockers poisoning.

Since severe and fatal poisoning with beta-blockers due only to beta-receptor blockade is unlikely, a prospective and partly retrospective analysis of 67 patients with beta-blockers poisoning was done in five-year period in order to determine the factors influencing the degree and outcome of acute poisoning. According to pharmacological properties of drugs, the patients were divided in groups: group I--50 patients with propranolol, group II-A--10 patients with atenolol and group II-B--7 patients with metoprolol poisoning. Electrocardiogram (ECG), 24-h ECG monitoring, toxicological screening (determination of beta-blockers in blood, urine and lavage by high performance liquid chromatography) and biochemical analysis were performed in all patients. Significantly smaller number of patients with serious poisoning was observed in group II-B. Patient's age did not correlate with the degree of poisoning, but significant correlation was found between preexisting disease, ingested dose and the time elapsed before the treatment started. Analysis of pharmacological properties showed that membrane stabilizing activity and lipophilicity of the drug might be the important determinants of the toxicity, while the role of cardioselectivity was lost in an overdose.

[The effect of cell phones on pacemaker function]. / Uticaj mobilnih telefona na funkciju pejsmejkera.

UNLABELLED: The aim of the study was to examine if there existed a risk from interference of cellular phones on patients with implanted permanent pacemakers. The study comprised 144 patients (134 VVI/VVIR and 10 DDD/DDDR) with permanent pacemakers of different manufactures. METHODS: During the routine pacemaker check, cellular phone aerial (Nokia 1610; GSM Standard) was placed against the skin of patients above the spot of pacemaker implantation, while the phone calls were repeated and ECG was continuously monitored. The effect of cellular phone on pacemaker was established upon: a) preexisting parameters of electric pacemaker stimulation; b) minimal ventricular rate of 90 beats/min, while electric stimulation parameters were set to their most sensitive values (MSV). RESULTS: Only in 9 (6.25%) patients was observed intermittent pacemaker inhibition, when pacemaker settings were on preexisting values of electric stimulation, while in 17 patients (11.8%) inhibition was noticed, when pacemaker settings were changed to their MSV. Besides, programmed values of electric stimulation in our patients remained unchanged. CONCLUSION: Although the electromagnetic interference interactions of cellular phones on pacemaker function were observed in relatively small number of our patients, we were of the opinion that pacemaker-dependent patients should avoid the use of cellular phones.

Involvement of plasma membrane redox system in the generation of trans-root electrical potential difference in excised maize root.

Possible involvement of the plasma membrane bound redox system in the generation of the trans-root electrical potential difference (TRP) arising across 8 day old maize (Zea mays L. hybrid ZPSC704) roots was studied. Excised roots were exposed to artificial impermeable electron acceptors (potassium hexacyanoferrate III and potassium hexachloroiridate IV) in external solution, and TRP response, oxygen consumption rate, proton efflux and reduction of the electron acceptors were analyzed. The effect of hexacyanoferrate III (HCF III) was tested at three concentrations (0.1; 0.5 and 1.0 mmol/l), and hexachloroiridate IV (HCI IV) in the concentration range 10(-7)-5.10(-4) mol/l. Both electron acceptors depolarized the trans-root potential, an order of magnitude lower concentrations of hexachloroiridate producing a much more rapid depolarization of greater magnitude. The roots had a higher capacity to reduce 0.1 mmol/l hexachloroiridate than 1 mmol/l hexacyanoferrate. Also, an increased level of acidification induced by HCI IV than HCF III could be observed. The rate of oxygen consumption showed an increase of about 20% in both cases. These results prove that electron transplasma membrane transport process(es) contribute to the total trans-root electrical potential difference across an excised maize root.

Photosynthetic oxygen evolution and chlorophyll fluorescence in intact isolated chloroplasts on a solid support: the influence of orthophosphate.

We devised recently a method to trap intact isolated chloroplasts on a solid support consisting of membrane filters made of cellulose nitrate (Cerovic et al., 1987, Plant Physiol. 84, 1249-1251). The addition of alkaline phosphatase to the reaction medium enabled continuous photosynthesis by spinach (Spinacia oleracea L.) chloroplasts to be sustained by hydrolysis of newly produced and exported triose phosphates and recycling of orthophosphate. In this system, simultaneous measurements of chlorophyll fluorescence and oxygen evolution were performed and their dependence on orthophosphate concentration was investigated. Optimal photosynthesis was obtained at a much higher initial orthophosphate concentration (2-4 mM) compared to intact chloroplasts in suspension. Secondary kinetics of chlorophyll fluorescence yield were observed and were shown to depend on the initial orthophosphate concentration.

Effect of propagators and inhibitors on the ultraweak luminescence from maize roots.

This study has investigated the kinetics and mechanism of ultraweak luminescence in maize roots. Mannitol induced the second maximum and enhanced the main maximum of the relative intensity of luminescence from the roots. Hydroquinone and quinone enhanced the relative intensity of the luminescence. Catalase enhanced the maximum of the luminescence and changed the kinetics of the light emission. The effect of catalase on the kinetics was abolished by superoxide dismutase. Ascorbate in the presence of catalase on the kinetics was abolished by superoxide dismutase. Ascorbate in the presence of catalase reduced the luminescence maximum, but did not alter the kinetics. In the presence of catalase only, or in the combination with superoxide dismutase, or ascorbate, the luminescence intensity in the stationary phase was significantly lower compared to the control. The results support the participation of superoxide-radical, singlet oxygen, electron transfer and the role of peroxidase in the reactions generating ultraweak luminescence in the roots. Ascorbate, catalase and superoxide dismutase have a protective role in the luminescent reactions.

Electrogenic proton pump in Nitella and the effect of calcium.

The hyperpolarisation of the membrane potential in Characeae above that of the diffusion potential is explained by the operation of the electrogenic proton pump. We studied the interaction of calcium with the functioning of the pump. The membrane potential was measured using the standard microelectrode technique. An increase in the calcium concentration resulted in depolarisation, its magnitude increasing with lower proton concentrations. Calcium-induced membrane potential changes, tested in the concentration range of 0.25 mmol/l to 25 mmol/l, were greatest at 0.25 mmol/l CaCl2 and decreased with the increasing calcium concentration. Light-induced initial changes in the membrane potential also showed a dependence on the presence of calcium in the external medium. We conclude that calcium has a role in the regulation of the proton pump in Nitella.