• The Korean Journal of Physiology and Pharmacology
• /
• v.21 no.4
• /
• pp.371-376
• /
• 2017

The caudal subnucleus of the spinal trigeminal nucleus (medullary dorsal horn; MDH) receives direct inputs from small diameter primary afferent fibers that predominantly transmit nociceptive information in the orofacial region. Recent studies indicate that reactive oxygen species (ROS) is involved in persistent pain, primarily through spinal mechanisms. In this study, we aimed to investigate the role of xanthine/xanthine oxidase (X/XO) system, a known generator of superoxide anion ($O_2{^-}$), on membrane excitability in the rat MDH neurons. For this, we used patch clamp recording and confocal imaging. An application of X/XO ($300{\mu}M/30mU$) induced membrane depolarization and inward currents. When slices were pretreated with ROS scavengers, such as phenyl N-tert-butylnitrone (PBN), superoxide dismutase (SOD), and catalase, X/XO-induced responses decreased. Fluorescence intensity in the DCF-DA and DHE-loaded MDH cells increased on the application of X/XO. An anion channel blocker, 4,4-diisothiocyanatostilbene-2,2-disulfonic acid (DIDS), significantly decreased X/XO-induced depolarization. X/XO elicited an inward current associated with a linear current-voltage relationship that reversed near -40 mV. X/XO-induced depolarization reduced in the presence of $La^{3+}$, a nonselective cation channel (NSCC) blocker, and by lowering the external sodium concentration, indicating that membrane depolarization and inward current are induced by influx of $Na^+$ ions. In conclusion, X/XO-induced ROS modulate the membrane excitability of MDH neurons, which was related to the activation of NSCC.

• Journal of Ginseng Research
• /
• v.45 no.5
• /
• pp.565-574
• /
• 2021

Background: Saengmaeksan (SMS) is a traditional Korean medicine composed of three herbs, Panax ginseng, Schisandra chinensis, and Liriope platyphylla. SMS is used to treat respiratory and cardiovascular disorders. However, whether SMS exerts antihyperuricemic effects is unknown. Methods: Effects of the SMS extract in water (SMS-W) and 30% ethanol (SMS-E) were studied in a rat model of potassium oxonate-induced hyperuricemia. Uric acid concentrations and xanthine oxidase (XO) activities were evaluated in the serum, urine, and hepatic tissue. Using renal histopathology to assess kidney function and uric acid excretion, we investigated serum creatinine and blood urea nitrogen concentrations, as well as protein levels of renal urate transporter 1 (URAT1), glucose transporter 9 (GLUT9), and organic anion transporter 1 (OAT1). The effects of SMS on in vitro XO activity and uric acid uptake were also evaluated. The components of SMS were identified using Ultra Performance Liquid Chromatography (UPLC). Results: SMS-E reduced serum uric acid and creatinine concentrations, and elevated urine uric acid excretion. SMS-E lowered XO activities in both the serum and liver, and downregulated the expression of renal URAT1 and GLUT9 proteins. SMS-E reduced renal inflammation and IL-1b levels in both the serum and kidneys. SMS-E inhibited both in vitro XO activity and urate uptake in URAT1-expressing oocytes. Using UPLC, 25 ginsenosides were identified, all of which were present in higher levels in SMS-E than in SMS-W. Conclusion: SMS-E exhibited antihyperuricemic effects by regulating XO activity and renal urate transporters, providing the first evidence of its applicability in the treatment of hyperuricemia and gout.

• Applied Microscopy
• /
• v.31 no.3
• /
• pp.275-285
• /
• 2001

The pulmonary alveolar macrophage is thought to play an important role in the mediation of acute inflammatory lung injury by secretory products including degraded enzymes, cytokines, and reactive oxygen metabolites . This study was conceived to understand the role of alveolar macrophage in oxidative stress induced acute lung injury. To examine the alveolar macrophages and xanthine oxidase (XO) activity in bronchoalveolar lavage fluid (BALF), time-dependent changes of numbers of alveolar macrophages, monocytes and neutrophils in alveolar cavity were counted in association with ultrastructural and cytochemical observations of lung tissue and alveolar cells. The number of monocytes was increased (p<0.001) at 1h after IL-1 treatment compared with that of sham. At 2h after instillation of IL-1, the number of alveolar macrophages was the highest, XO activity in BALF was elevated at 2h after IL-1 instillation and the activity was markedly elevated(p<0.05) at 3h after IL-1 treatment. On the basis of these experimental results, it is suggested that, during early phase of acute lung injury induced by IL-1, alveolar macrophage-derived XO contributes to lung injury earlier than the neutrophilic respiratory burst.

• Biomedical Science Letters
• /
• v.27 no.4
• /
• pp.277-282
• /
• 2021

Hydrocotyle is a genus of prostrate, perennial aquatic or semi-aquatic plants formerly classified in the family Apiaceae, now in the family Araliaceae. Lipoxygenases (LOX) are present in the human body and play an important role in the stimulation of inflammatory reactions. Ethanolic extracts of five Hydrocotyle species (H. ramiflora, H. maritima, H. nepalensis, H. sibthorpioides, and H. yabei) showed inhibition of 23.5~50.6% at 2.0 mg/mL. Their extracts showed LOX inhibition in half maximal effective concentration (EC₅₀) range 15.1~15.7 ㎍/mL. Hyaluronic acid is a glycosaminoglycan, a major component of the extracellular matrix Five extracts of these species inhibited less than 23.0% of Hyaluronidase (HAase) activity at a concentration of 2.0 mg/mL Xanthine oxidase (XO) is a form of xanthine oxidoreductase, a type of enzyme that generates reactive oxygen species. Five Hydrocotyle species were found to have inhibitory activity of XO at 2.0 mg/ml, with 65% having greater than 50% inhibition. H. ramiflora exhibited the highest activity with an inhibition of 80.0%. The results suggested that Lipoxygenases, Hyaluronidase, and Busan 47340, Republic of Korea from five Hydrocotyle species might be multifunctional and prevent the degradation of allergic reactions and inflammation.

• International Journal of Oral Biology
• /
• v.41 no.3
• /
• pp.141-147
• /
• 2016

Reactive oxygen species (ROS) and nitrogen species (RNS) are both important signaling molecules involved in pain transmission in the dorsal horn of the spinal cord. Xanthine oxidase (XO) is a well-known enzyme for the generation of superoxide anions ($O_2^{\bullet-}$), while S-nitroso-N-acetyl-DL-penicillamine (SNAP) is a representative nitric oxide (NO) donor. In this study, we used patch clamp recording in spinal slices of rats to investigate the effects of $O_2^{\bullet-}$ and NO on the excitability of substantia gelatinosa (SG) neurons. We also used confocal scanning laser microscopy to measure XO- and SNAP-induced ROS and RNS production in live slices. We observed that the ROS level increased during the perfusion of xanthine and xanthine oxidase (X/XO) compound and SNAP after the loading of 2',7'-dichlorofluorescin diacetate ($H_2DCF-DA$), which is an indicator of intracellular ROS and RNS. Application of ROS donors such as X/XO, ${\beta}-nicotinamide$ adenine dinucleotide phosphate (NADPH), and 3-morpholinosydnomimine (SIN-1) induced a membrane depolarization and inward currents. SNAP, an RNS donor, also induced membrane depolarization and inward currents. X/XO-induced inward currents were significantly decreased by pretreatment with phenyl N-tert-butylnitrone (PBN; nonspecific ROS and RNS scavenger) and manganese(III) tetrakis(4-benzoic acid) porphyrin (MnTBAP; superoxide dismutase mimetics). Nitro-L-arginine methyl ester (NAME; NO scavenger) also slightly decreased X/XO-induced inward currents, suggesting that X/XO-induced responses can be involved in the generation of peroxynitrite ($ONOO^-$). Our data suggest that elevated ROS, especially $O_2^{\bullet-}$, NO and $ONOO^-$, in the spinal cord can increase the excitability of the SG neurons related to pain transmission.

• Journal of Physiology & Pathology in Korean Medicine
• /
• v.17 no.4
• /
• pp.1008-1012
• /
• 2003

In order to evaluate the cytotoxic effect of reactive oxygen species(AOS), the cell viability was measured by MTT assay after cultured mouse hippocampal neurons were treated with various concentrations of xanthine oxidase(XO) and hypoxanthine (HX) for 5 hours. And also, the protective effect of Salviae Mutiorrhizae Radix(SMR) on XO/HX-induced neurotoxicity was examined in these cultures. XO/HX significantly decreased cell viability in dose-and time dependent manners when cultured mouse hippocampal neurons were treated with 5～40 mU/ml XO for 5 hours. In the protective effect of SMA, SMR increased cell viability dose-dependently after cultured mouse hippocampal neurons were preincubated with 30～120 ㎍/ml SMR for 2 hours. From these results, it is suggested that XO/HX is toxic on cultured mouse hippocampal neurons, and herbe medicine such as SMR is very effective in blocking the cytotoxicity induced by AOS.

• Journal of Plant Biology
• /
• v.30 no.2
• /
• pp.95-108
• /
• 1987

Effects of allopurinol (2mM), a specific inhibitor of xanthine oxidase, on the growth and metabolism of llantoin in dark grown Chinese cabbage (Brassica campestris L.) seedlings were investigated. Allopurinol treatment maintained the fresh and dry weights of cotyledons at higher levels, but inhibited the elongation of hypocotyls and roots of the seedlings. Total nitrogen content in the cotyledons decreased at slower rate by allopurinol. Accordingly, the levels of total nitrogen contents in the hypocotyls and roots, were depressed by the inhibitor. In the cotyledons, allopurinol began to elevate RNA levels after day 3, which it did not affect DNA level throughout the experiment. Activities of xanthine oxidase (XO:EC 1.2.3.2), uricase (UO:EC 1.7.3.3) and allantoinase (AL:EC 3.5.2.5) in the cotyledons were examined. The activity of XO was not detected, but the accumulation of xanthine by allopurinol treatment presented an indirect evidence of the existence of XO in the organ. Allopurinol kept UO activity high up to day 2 after sowing and depressed AL activity throughout the experiment. By allopurinol treatment, allantoin content was kept high over the control both in cotyledons and roots, but it was kept low in hypocotyls. The level of allantoic acid in the 3 organs were shown to be depressed by allopurinol. These results suggest that allantoin and allantoic acid produced by the degradation of stored and newly synthesized RNA are transported from the storage tissue to hypocotyls and roots as important nitrogen sources for the development of Chinese cabbage seedlings.

• International Journal of Oral Biology
• /
• v.39 no.4
• /
• pp.229-236
• /
• 2014

Reactive oxygen species (ROS) and nitrogen species (RNS) are implicated in cellular signaling processes and as a cause of oxidative stress. Recent studies indicate that ROS and RNS are important signaling molecules involved in nociceptive transmission. Xanthine oxidase (XO) system is a well-known system for superoxide anions ($O{_2}^{{\cdot}_-}$) generation, and sodium nitroprusside (SNP) is a representative nitric oxide (NO) donor. Patch clamp recording in spinal slices was used to investigate the role of $O{_2}^{{\cdot}_-}$ and NO on substantia gelatinosa (SG) neuronal excitability. Application of xanthine and xanthine oxidase (X/XO) compound induced membrane depolarization. Low concentration SNP ($10{\mu}M$) induced depolarization of the membrane, whereas high concentration SNP (1 mM) evoked membrane hyperpolarization. These responses were significantly decreased by pretreatment with phenyl N-tert-butylnitrone (PBN; nonspecific ROS and RNS scavenger). Addition of thapsigargin to an external calcium free solution for blocking synaptic transmission, led to significantly decreased X/XO-induced responses. Additionally, X/XO and SNP-induced responses were unchanged in the presence of intracellular applied PBN, indicative of the involvement of presynaptic action. Inclusion of GDP-${\beta}$-S or suramin (G protein inhibitors) in the patch pipette decreased SNP-induced responses, whereas it failed to decrease X/XO-induced responses. Pretreatment with n-ethylmaleimide (NEM; thiol-alkylating agent) decreased the effects of SNP, suggesting that these responses were mediated by direct oxidation of channel protein, whereas X/XO-induced responses were unchanged. These data suggested that ROS and RNS play distinct roles in the regulation of the membrane excitability of SG neurons related to the pain transmission.

• Journal of Applied Biological Chemistry
• /
• v.57 no.1
• /
• pp.53-59
• /
• 2014

To evaluate the inhibitory effect of xanthine oxidase (XO) and aldehyde oxidase (AO), and antihyperuricemic effect by Aspergillus oryzae fermented Smilax china L. leaf extracts and fractions, we observed extracted yield by each solvent, the content of total polyphenol and total flavonoid (TF), the activities of XO and AO, and serum uric acid level. Extracted yield (g/kg) by 80% ethanol (EtOH) was 13.56, those of n-hexane, dichloromethane (DICM), ethylacetate (EtOAc) and n-butanol fraction (BuOH) were 1.35-3.33. Furthermore, total polyphenol content (mg/g-extract) of EtOAc fraction, BuOH fraction, DICM fraction and EtOH fraction is 478.07-501.26, 259.49-289.02, 165.03-232.27, 134.02-196.54, respectively. Those of fermented EtOAc and DICM fraction was 4.85 and 40.74% higher than that of non-fermented fraction, respectively, while the other fermented fractions were lower than those of non-fermented fractions. And total flavonoid content (mg/g-extract) of EtOAc fraction was higher than those of other fractions. Additionally, TF of fermented EtOAc and BuOH fraction is 10.56 and 60.17% higher, than that of fermented fraction, respectively, although those of the other fermented fractions was lower than that of non-fermented fractions. On the other hand, XO inhibitory activities of all fermented fractions was significantly higher than that of all non-fermented fraction, while those of fermented EtOAc (75.02%) and BuOH fraction (65.59%) was markedly higher than that of non-fermented fraction (39.42 and 5.34%), respectively. In addition, AO inhibitory activities of DICM and EtOAc fraction was 81.82 and 77.93% higher, respectively, than those of the other fractions, and those of fermented fractions as with XO were significantly higher than that of non-fermented fractions. Meanwhile, serum uric acid level (SU) of hyperuricemic control mice (HC, 6.98 mg/dL) was 1.83 folds higher than that of normal control (NC, 3.82 mg/dL). Furthermore, SU in the group treated with EtOAc fraction decreased in a dose dependent manner compared with the allopurinol control group, although those of fermented fractions were significantly lower than those of non-fermented fractions. This study suggests that fermented Smilax china L. leaf extracts may regulate the XO and AO inhibitory activities and antihyperuricemic effect due to aglycone components from glycoside form flavonoids by fermentation of A. oryzae.

• Journal of Physiology & Pathology in Korean Medicine
• /
• v.16 no.2
• /
• pp.353-358
• /
• 2002

To test the protective effect of herbal medicine against oxygen free radical-induced myocardiotoxicity, cytotoxicity of xanthine oxidase/hypoxanthine (XO/HX) was examined using MTT, TBARS, and beating rate assay in the presence of water extract of Sujeom-san(SJS) or single consituents of its prescription. Myocardial toxicity was evaluated in neonatal rat myocardiocytes in cultures. In the present paper, XO/HX resulted in a decrease in viability and beating rate and increases in lipid peroxidation in cultured myocardial cells. In the effect of SJS water extract, it showed effects from the cardiocytotoxicity induced by XO/HX treatment such as increases in beating rate and decreases in lipid peroxidation. In the effect of Rhizoma Corydalis (RC), Faeces Trogopterori (FT), Fructus Amomi Tsaoko (FAT) and Myrrha on the cardiocytotoxicity, they were significantly effective in blocking the XO/HX-induced cardiocytotoxicity by increase of beating rate in FAT and FT group as well as decrease of lipid peroxidation in FT and RC group. These results show that oxygen free radical elicits toxic effects in cultured myocardial cells derived from neonatal rat, and suggest that water extract of Sujeomsan, Rhizoma Corydalis, Faeces Trogopterori, Fructus Amomi Tsaoko or Myrrha is very effective in the prevention of xanthine oxidase/hypoxanthine- induced cardiotoxicity.