Superoxide-producing thermostable associate from the small intestines of control and alloxan-induced diabetic rats: quantitative and qualitative changes.

BACKGROUND: NADPH oxidase 1 (Nox1), which is highly expressed in the colon, is thought to play a potential role in host defense as a physical and innate immune barrier against commensal or pathogenic microbes in the gastrointestinal epithelium. Diabetes can be caused by several biological factors, including insulin resistance is one of them. Alloxan is widely used to induce insulin-dependent diabetes in experimental animals. Alloxan increases the generation of reactive oxygen species as a result of metabolic reactions in the body, along with a massive increase in cytosolic calcium concentration. METHODS: Using a universal method, a superoxide radical (О2-)-thermostable associate between NADPH-containing lipoprotein (NLP) and NADPH oxidase (Nox)- NLP-Nox was isolated and purified from the small intestine (SI) of control (C) and alloxan-induced diabetic (AD) albino rats. RESULTS: In comparison to the C indices, in AD in the SI, an increase in the specific content of NLP-Nox associate and a decrease in the stationary concentration of produced О2- in liquid phase (in solution) and gas phase (during blowing by oxygen of the NLP-Nox solution) were observed. The NLP-Nox of SI associate in C and AD rats produced О2- by an immediate mechanism, using NLP as a substrate. The phenomenon of the hiding of the optical absorption maxima of the Nox in oxidized states at pH10,5 was observed in the composition of these SI associates of the C and AD rat groups. The characteristic absorption maxima of the «hidden¼ Nox were observed under these conditions after reduction by potassium dithionite. CONCLUSION: Thus, at AD, the decrease in the stationary concentration of produced О2- in the solution and gas phase was compensated for by an increase in the specific amount of associate. In addition,  the decrease in the stationary concentration of produced О2- by NLP-Nox associates at AD can be linked to a decrease in the level of NADPH in NLP-Nox composition. This could be used as a new mechanism of AD pathogenesis.

Superoxide-producing lipoprotein fraction from Stevia leaves: definition of specific activity.

BACKGROUND: Stevia rebaudiana Bertoni has various pharmacological actions, which includes antidiabetic, antioxidant, anti-inflammatory activities. The superoxide and consequently NADPH oxidase (Nox) are relevant targets involved in biological effects of Stevia. The presence of NADPH-containing superoxide-producing lipoprotein (suprol) in Stevia leaves has not yet been tested. The mechanism of producing superoxide radicals (O2-) by suprol was determined in vitro, which is associated with the electron transfer from NADPH in the composition of suprol by traces of transition metal ions (Fe3+ or Cu2+) to molecular oxygen, turning it into O2-. It is expected that the therapeutic efficacy of Stevia leaves is caused by specific activity of superoxide-producing lipoprotein fraction. METHODS: For the first time, from the dry leaves of Stevia the NADPH-containing superoxide-producing lipoprotein was isolated and purified. The specific content of suprol (milligrams in 1 g of Stevia leaves- mg/g) was determined after desalination of suprol and lyophilization. RESULTS: According to the method provided, the specific content of the isolated suprol from Stevia's leaves was up to 4.5 ± 0.2 mg / g (yields up to 68.5 ± 4.7%, p < 0.05, n = 6). Nox forms a stable complex with suprol. The optical absorption spectrum of the Nox-suprol complex represents the overlapping suprol and Nox spectra, with a certain background increase and characteristic features of optical absorption for Nox. Due to O2- producing capacity suprol-Nox complex discolors KMnO4 solutions, Coomassie brilliant blue, restores nitrotetrazolium blue to formazan and oxidizes epinephrine to adrenochrome. The oxidation activity of adrenaline is 50.3 ± 5.1 U / mg / ml (p < 0.05, n = 6). CONCLUSION: Superoxide-producing lipoprotein fraction-Nox complex from Stevia leaves (membranes) can modulate redox regulated signaling pathways and may play a positive role in type-2 diabetes by means of adrenaline oxidation mechanism.

Effects of stevia on synaptic plasticity and NADPH oxidase level of CNS in conditions of metabolic disorders caused by fructose.

BACKGROUND: Excess dietary fructose intake associated with metabolic syndrome and insulin resistance and increased risk of developing type 2 diabetes. Previous animal studies have reported that diabetic animals have significantly impaired behavioural and cognitive functions, pathological synaptic function and impaired expression of glutamate receptors. Correction of the antioxidant status of laboratory rodents largely prevents the development of fructose-induced plurimetabolic changes in the nervous system. We suggest a novel concept of efficiency of Stevia leaves for treatment of central diabetic neuropathy. METHODS: By in vivo extracellular studies induced spike activity of hippocampal neurons during high frequency stimulation of entorhinal cortex, as well as neurons of basolateral amygdala to high-frequency stimulation of the hippocampus effects of Stevia rebaudiana Bertoni plant evaluated in synaptic activity in the brain of fructose-enriched diet rats. In the conditions of metabolic disorders caused by fructose, antioxidant activity of Stevia rebaudiana was assessed by measuring the NOX activity of the hippocampus, amygdala and spinal cord. RESULTS: In this study, the characteristic features of the metabolic effects of dietary fructose on synaptic plasticity in hippocampal neurons and basolateral amygdala and the state of the NADPH oxidase (NOX) oxidative system of these brain formations are revealed, as well as the prospects for development of multitarget and polyfunctional phytopreparations (with adaptogenic, antioxidant, antidiabetic, nootropic activity) from native raw material of Stevia rebaudiana. Stevia modulates degree of expressiveness of potentiation/depression (approaches but fails to achieve the norm) by shifting the percentage balance in favor of depressor type of responses during high-frequency stimulation, indicating its adaptogenic role in plasticity of neural networks. Under the action of fructose an increase (3-5 times) in specific quantity of total fraction of NOX isoforms isolated from the central nervous system tissue (amygdala, hippocampus, spinal cord) was revealed. Stevia exhibits an antistress, membrane-stabilizing role reducing the level of total fractions of NOX isoforms from central nervous system tissues and regulates NADPH-dependent O2- -producing activity. CONCLUSION: Generally, in condition of metabolic disorders caused by intensive consumption of dietary fructose Stevia leaves contributes to the control of neuronal synaptic plasticity possibly influencing the conjugated NOX-specific targets.

Proline rich polypeptide (PRP-1) increases the superoxide-producing and ferrihemoglobin reducing activities of cytochrome B(558) isoforms from human lymphosarcoma tissue cells.

The two cytochromes (cyt) b(558) of acidic nature, one-95-100 kDa and another one, 60-70 kDa were isolated for the first time from the human's lymphosarcoma tissue cells using gel filtration and ion exchange chromatography. These hemoproteins possess NADPH dependent O(2)(-)-producing and ferrihemoglobin-reducing activities. The incubation of neuropeptide PRP-1 (5 µg) with cytochrome b(558), caused elevation of these activities. The gel filtration results indicated possible binding of PRP-1 to these cytochromes b(558). PRP-1 activated both NADPH dependent O(2)(-)-producing and ferriHb-reducing activities of the cyt b(1)(558) and cyt b(2)(558), obtained from human lymphosarcoma tissue cells. One can assume that PRP-1 associated with cyt b(558) on the surface of the tumor cells by increasing both NADPH dependent O(2)(-)-producing and ferriHb-reducing activities of cyt b(558), increases the oxidation- reduction status. Changing the oxidation-reduction status and oxygen homeostasis of the tumor cells by PRP-1 can serve as one of the possible explanation of antitumorigenic effect of this cytokine.