H1N1 nanobody development and therapeutic efficacy verification in H1N1-challenged mice.

Influenza A virus causes numerous deaths and infections worldwide annually. Therefore, we have considered nanobodies as a potential treatment for patients with severe cases of influenza. We developed a nanobody that was expected to have protective efficacy against the A/California/04/2009 (CA/04; pandemic 2009 flu strain) and evaluated its therapeutic efficacy against CA/04 in mice experiments. This nanobody was derived from the immunization of the alpaca, and the inactivated CA/04 virus was used as an immunogen. We successfully generated a nanobody library through bio-panning, phage ELISA, and Bio-layer interferometry. Moreover, we confirmed that administering nanobodies after lethal doses of CA/04 reduced viral replication in the lungs and influenza-induced clinical signs in mice. These research findings will help to develop nanobodies as viral therapeutics for CA/04 and other infectious viruses.

Antidepressant-Like Activities of Hispidol and Decursin in Mice and Analysis of Neurotransmitter Monoamines.

The antidepressant activities of hispidol and decursin (both potent monoamine oxidase A (MAO-A) inhibitors) were evaluated using the forced swimming test (FST) and the tail suspension test (TST) in mice, and thereafter, levels of neurotransmitter monoamines and metabolites in brain tissues were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Hispidol (15 mg/kg) caused less or comparable immobility than fluoxetine (15 mg/kg; the positive control) in immobility time, as determined by FST (9.6 vs 32.0 s) and TST (53.1 vs 48.7 s), respectively, and its effects were dose-dependent and significant. Decursin (15 mg/kg) also produced immobility comparable to that of fluoxetine as determined by FST (47.0 vs 43.4 s) and TST (55.6 vs 63.4 s), and its effects were also dose-dependent and significant. LC-MS/MS analysis after FST showed that hispidol (15 mg/kg) greatly increased dopamine (DA) and serotonin levels dose-dependently in brain tissues as compared with the positive control. Decursin (15 mg/kg) dose-dependently increased DA level after TST. Slight changes in norepinephrine and 3,4-dihydroxyphenylacetic acid levels were observed after FST and TST in hispidol- or decursin-treated animals. It was observed that hispidol and decursin were effective and comparable to fluoxetine in immobility tests. These immobility and monoamine level results suggest that hispidol and decursin are potential antidepressant agents for the treatment of depression, and that they act mainly through serotonergic and/or dopaminergic systems.

Calycosin and 8-O-methylretusin isolated from Maackia amurensis as potent and selective reversible inhibitors of human monoamine oxidase-B.

Nineteen compounds were isolated from the stems of Maackia amurensis by activity-guided screening for new human monoamine oxidase-B (hMAO-B) inhibitors. Among the compounds isolated, flavonoids calycosin (5) and 8-O-methylretusin (6) were found to potently and selectively inhibit hMAO-B (IC50 = 0.24 and 0.23 µM, respectively) but not hMAO-A with high selectivity index (SI) values (SI = 293.8 and 81.3, respectively). In addition, 5 and 6 reversibly and competitively inhibited hMAO-B with Ki values of 0.057 and 0.054 µM, respectively. A pterocarpan (-)-medicarpin (18) was also observed to strongly inhibit hMAO-B (IC50 = 0.30 µM). Most of the compounds weakly inhibited AChE, except isolupalbigenin (13) (IC50 = 20.6 µM), which suggested 13 be considered a potential dual function inhibitor of MAO-B and AChE. Molecular docking simulation revealed that the binding affinities of 5 and 6 for hMAO-B (both -9.3 kcal/mol) were higher than those for hMAO-A (-7.4 and -7.2 kcal/mol, respectively). Compound 5 was found to interact by hydrogen bonding with hMAO-B at Cys172 residue (distance: 3.250 Å); no hydrogen bonding was predicted between 5 and hMAO-A. These findings suggest that compounds 5 and 6 be considered novel potent, selective, and reversible hMAO-B inhibitors and candidates for the treatment of neurological disorders.

Design, synthesis and biological evaluation of oxygenated chalcones as potent and selective MAO-B inhibitors.

The present study documents the synthesis of oxygenated chalcone (O1-O26) derivatives and their abilities to inhibit monoamine oxidases. All 26 derivatives examined showed potent inhibitory activity against MAO-B. Compound O23 showed the greatest inhibitory activity against MAO-B with an IC50 value of 0.0021 µM, followed by compounds O10 and O17 (IC50 = 0.0030 and 0.0034 µM, respectively). In addition, most of the derivatives potently inhibited MAO-A and O6 was the most potent inhibitor with an IC50 value of 0.029 µM, followed by O3, O4, O9, and O2 (IC50 = 0.035, 0.053, 0.072, and 0.082 µM, respectively). O23 had a high selectivity index (SI) value for MAO-B of 138.1, and O20 (IC50 value for MAO-B = 0.010 µM) had an extremely high SI of >4000. In dialysis experiments, inhibitions of MAO-A and MAO-B by O6 and O23, respectively, were recovered to their respective reversible reference levels, demonstrating both are reversible inhibitors. Kinetic studies revealed that O6 and O23 competitively inhibited MAO-A and MAO-B, respectively, with respective Ki values of 0.016 ±â€¯0.0007 and 0.00050 ±â€¯0.00003 µM. Lead compound are also non-toxic at 200 µg/mL in normal rat spleen cells. Molecular docking simulations and subsequent Molecular Mechanics/Generalized Born Surface Area calculations provided a rationale that explained experimental data.

Ethyl Acetohydroxamate Incorporated Chalcones: Unveiling a Novel Class of Chalcones for Multitarget Monoamine Oxidase-B Inhibitors Against Alzheimer's Disease.

BACKGROUND: Chalcones are considered as the selective scaffold for the inhibition of MAO-B. OBJECTIVES: A previously synthesized ethyl acetohydroxamate-chalcones (L1-L22) were studied for their inhibitory activities against human recombinant monoamine oxidase A and B (hMAO-A and hMAO-B, respectively) and acetylcholinesterase (AChE) as multi-target directed ligands for the treatment of Alzheimer's Disease (AD). METHODS: Enzyme inhibition studies of MAO-A, MAO-B and AChE is carried out. Computational studies such as Molecular docking, Molecular Mechanics/Generalized Born Surface Area calculations, ADMET prediction, and protein target prediction are also performed. RESULTS: Among the screened compounds, compound L3 has most potent hMAO-B inhibition with an IC50 value of 0.028 ± 0.0016 µM, and other compounds, L1, L2, L4, L8, L12, and L21 showed significant potent hMAO-B inhibition with IC50 values of 0.051 ± 0.0014, 0.086 ± 0.0035, 0.036 ± 0.0011, 0.096 ± 0.0061, 0.083 ± 0.0016, and 0.038 ± 0.0021 µM, respectively. On the other hand, among the tested compounds, compound L13 showed highest hMAO-A inhibition with an IC50 value of 0.51± 0.051 µM and L9 has a significant value of 1.85 ± 0.045 µM. However, the compounds L3 and L4 only showed high selectivities for hMAO-B with Selectivity Index (SI) values of 621.4 and 416.7, respectively. Among the substituents in ring A of ethyl acetohydroxamate-chalcones (L1-L9), F atom at p-position (L3) showed highest inhibitory effect against hMAO-B. This result supports the uniqness and bizarre behavior of fluorine. Moreover, chalcones L3, L4, L9, L11, and L12 showed potential AChE inhibitory effect with IC50 values of 0.67, 0.85, 0.39, 0.30, and 0.45 µM, respectively. Inhibitions of hMAO-B by L3 or L4 were recovered to the level of the reversible reference (lazabemide), and were competitive with Ki values of 0.0030 ± 0.0002 and 0.0046 ± 0.0005 µM, respectively. Inhibitions of AChE by L3 and L11 were of the competitive and mixed types with Ki values of 0.30 ± 0.044 and 0.14 ± 0.0054 µM, respectively. CONCLUSION: The studies indicated that L3 and L4 are considered to be promising multitarget drug molecules with potent, selective, and reversible competitive inhibitors of hMAO-B and with highly potent AChE inhibitory effect.

Potent and selective inhibition of human monoamine oxidase-B by 4-dimethylaminochalcone and selected chalcone derivatives.

Six synthetic (1-6) and six natural (7-12) chalcones were tested for human monoamine oxidases (hMAOs) and acetylcholinesterase (AChE) inhibitory activities. Compounds 4-dimethylaminochalcone (2), 4'-chloro-4-dimethylaminochalcone (5), and 2,4'-dichloro-4-dimethylaminochalcone (1) potently inhibited hMAO-B with IC50 values of 0.029, 0.061, and 0.075 µM, respectively. 4-Nitrochalcone (4) and 4-chlorochalcone (3) also potently inhibited hMAO-B with IC50 values of 0.066 and 0.082 µM, respectively (2.3- and 2.6-fold less than compound 2). Compound 2 had a high selectivity index (113.1) for hMAO-B over hMAO-A (IC50 = 3.28 µM). Compounds 1 and 2,2'-dihydroxy-4',6'-dimethoxychalcone (12) potently inhibited hMAO-A with IC50 values of 0.18 and 0.39 µM, respectively. In addition, compounds 4 and 2 also effectively inhibited AChE with IC50 values of 1.25 and 6.07 µM, respectively, and thus, exhibited dual-targeting. Compound 2 reversibly and competitively inhibited hMAO-B with a Ki value of 0.0066 µM. Docking simulations showed binding affinities of compounds 1 to 5 for hMAO-B were higher than those for hMAO-A or AChE and suggested these five chalcones form hydrogen bonds with MAO-B at Cys172 but that they do not form hydrogen bonds with hMAO-A or AChE. These findings suggest compound 2 be considered a promising and dual-targeting lead compound for the treatment of Alzheimer's disease.

Potent inhibition of acetylcholinesterase by sargachromanol I from Sargassum siliquastrum and by selected natural compounds.

Six hundred forty natural compounds were tested for acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibitory activities. Of those, sargachromanol I (SCI) and G (SCG) isolated from the brown alga Sargassum siliquastrum, dihydroberberine (DB) isolated from Coptis chinensis, and macelignan (ML) isolated from Myristica fragrans, potently and effectively inhibited AChE with IC50 values of 0.79, 1.81, 1.18, and 4.16 µM, respectively. SCI, DB, and ML reversibly inhibited AChE and showed mixed, competitive, and noncompetitive inhibition, respectively, with Ki values of 0.63, 0.77, and 4.46 µM, respectively. Broussonin A most potently inhibited BChE (IC50 = 4.16 µM), followed by ML, SCG, and SCI (9.69, 10.79, and 13.69 µM, respectively). In dual-targeting experiments, ML effectively inhibited monoamine oxidase B with the greatest potency (IC50 = 7.42 µM). Molecular docking simulation suggested the binding affinity of SCI (-8.6 kcal/mol) with AChE was greater than those of SCG (-7.9 kcal/mol) and DB (-8.2 kcal/mol). Docking simulation indicated SCI interacts with AChE at Trp81, and that SCG interacts at Ser119. No hydrogen bond was predicted for the interaction between AChE and DB. This study suggests SCI, SCG, DB, and ML be viewed as new reversible AChE inhibitors and useful lead compounds for the development for the treatment of Alzheimer's disease.

Osthenol, a prenylated coumarin, as a monoamine oxidase A inhibitor with high selectivity.

Osthenol (6), a prenylated coumarin isolated from the dried roots of Angelica pubescens, potently and selectively inhibited recombinant human monoamine oxidase-A (hMAO-A) with an IC50 value of 0.74 µM and showed a high selectivity index (SI > 81.1) for hMAO-A versus hMAO-B. Compound 6 was a reversible competitive hMAO-A inhibitor (Ki = 0.26 µM) with a potency greater than toloxatone (IC50 = 0.93 µM), a marketed drug. Isopsoralen (3) and bakuchicin (1), furanocoumarin derivatives isolated from Psoralea corylifolia L., showed slightly higher IC50 values (0.88 and 1.78 µM, respectively) for hMAO-A than 6, but had low SI values (3.1 for both). Other coumarins tested did not effectively inhibit hMAO-A or hMAO-B. A structural comparison suggested that the 8-(3,3-dimethylallyl) group of 6 increased its inhibitory activity against hMAO-A compared with the 6-methoxy group of scopoletin (4). Molecular docking simulations revealed that the binding affinity of 6 for hMAO-A (-8.5 kcal/mol) was greater than that for hMAO-B (-5.6 kcal/mol) and that of 4 for hMAO-A (-7.3 kcal/mol). Docking simulations also implied that 6 interacted with hMAO-A at Phe208 and with hMAO-B at Ile199 by carbon hydrogen bondings. Our findings suggest that osthenol, derived from natural products, is a selective and potent reversible inhibitor of MAO-A, and can be regarded a potential lead compound for the design of novel reversible MAO-A inhibitors.

Rhamnocitrin isolated from Prunus padus var. seoulensis: A potent and selective reversible inhibitor of human monoamine oxidase A.

Three flavanones and two flavones were isolated from the leaves of Prunus padus var. seoulensis by the activity-guided screening for new monoamine oxidase (MAO) inhibitors. Among the compounds isolated, rhamnocitrin (5) was found to potently and selectively inhibit human MAO-A (hMAO-A, IC50 = 0.051 µM) and effectively inhibit hMAO-B (IC50 = 2.97 µM). The IC50 value of 5 for hMAO-A was the lowest amongst all natural flavonoids reported to date, and the potency was 20.2 times higher than that of toloxatone (1.03 µM), a marketed drug. In addition, 5 reversibly and competitively inhibited hMAO-A and hMAO-B with Ki values of 0.030 and 0.91 µM, respectively. Genkwanin (4) was also observed to strongly inhibit hMAO-A and hMAO-B (IC50 = 0.14 and 0.35 µM, respectively), and competitively inhibit hMAO-A and hMAO-B (Ki = 0.097 and 0.12 µM, respectively). Molecular docking simulation reveals that the binding affinity of 5 with hMAO-A (-18.49 kcal/mol) is higher than that observed with hMAO-B (0.19 kcal/mol). Compound 5 interacts with hMAO-A at four possible residues (Asn181, Gln215, Thr336, and Tyr444), while hMAO-B forms a single hydrogen bond at Glu84. These findings suggest that compound 5 as well as 4 can be considered as novel potent and reversible hMAO-A and/or hMAO-B inhibitors or useful lead compounds for future development of hMAO inhibitors in neurological disorder therapies.

Characterization of truncated endo-ß-1,4-glucanases from a compost metagenomic library and their saccharification potentials.

A gene encoding an endo-ß-1,4-glucanase (Cel6H-f481) was cloned from a compost metagenomic library. The gene, cel6H-f481, was composed of 1446 bp to encode a fused protein of 481 amino acid residues (50,429 Da), i.e., 445 residues (Cel6H-445) from the metagenome, and 36 residues from the pUC19 vector at N-terminus. Cel6H-445 belonged to glycosyl hydrolase (GH) family 6 and showed 71% identity with Actinotalea fermentans endoglucanase with low coverage. Several active bands of truncated forms were observed by activity staining of the crude extract. Major truncated enzymes of 35 (Cel6H-p35) and 23 kDa (Cel6H-p23) were separated by HiTrap Q chromatography. The two enzymes had the same optimum temperature (50 °C) and pH (5.5), but Cel6H-p35 was more thermostable than Cel6H-p23 and other GH6 endoglucanases reported. Both enzymes efficiently hydrolyzed carboxymethyl-cellulose (CMC) and barley ß-glucan, but hardly hydrolyzed other substrates tested. The Vmax of Cel6H-p35 for CMC was 1.4 times greater than that of Cel6H-p23. The addition of the crude enzymes to a commercial enzyme set increased the saccharification of pretreated rice straw powder by up to 30.9%. These results suggest the N-terminal region of Cel6H-p35 contributes to thermostability and specific activity, and that the enzymes might be a useful additive for saccharification.

Characterization of Three Extracellular ß-Glucosidases Produced by a Fungal Isolate Aspergillus sp. YDJ14 and Their Hydrolyzing Activity for a Flavone Glycoside.

A cellulolytic fungus, YDJ14, was isolated from compost and identified as an Aspergillus sp. strain. Three extracellular ß-glucosidases, BGL-A1, BGL-A2, and BGL-A3, were separated using ultrafiltration, ammonium sulfate fractionation, and High-Q chromatography. The molecular masses of the three enzymes were estimated to be 100, 45, and 40 kDa, respectively, by SDS-PAGE. The optimum pH and temperature of BGL-A3 were 5.0 and 50°C, respectively, whereas the optimum pH and temperature of BGL-A1 and BGL-A2 were identical (4.0 and 60°C, respectively). The half-life of BGL-A3 at 70°C (2.8 min) was shorter than that of BGL-A1 and BGL-A2 (12.1 and 8.8 min, respectively). All three enzymes preferred p-nitrophenyl-ß-D-glucopyranoside (pNPG) and hardly hydrolyzed cellobiose, suggesting that these enzymes were aryl ß-glucosidases. The Km of BGL-A3 (1.26 mM) for pNPG was much higher than that of BGL-A1 and BGL-A2 (0.25 and 0.27 mM, respectively). These results suggested that BGL-A1 and BGL-A2 were similar in their enzymatic properties, whereas BGL-A3 differed from the two enzymes. When tilianin (a flavone glycoside of acacetin) was reacted with the three enzymes, the inhibitory activity for monoamine oxidase, a target in the treatment of neurological disorders, was similar to that shown by acacetin. We conclude that these enzymes may be useful in the hydrolysis of flavone glycosides to improve their inhibitory activities.

Characterization of a Multimodular Endo-ß-1,4-Glucanase (Cel9K) from Paenibacillus sp. X4 with a Potential Additive for Saccharification.

An endo-ß-1,4-glucanase gene, cel9K, was cloned using the shot-gun method from Paenibacillus sp. X4, which was isolated from alpine soil. The gene was 2,994 bp in length, encoding a protein of 997 amino acid residues with a predicted signal peptide composed of 32 amino acid residues. Cel9K was a multimodular enzyme, and the molecular mass and theoretical pI of the mature Cel9K were 103.5 kDa and 4.81, respectively. Cel9K contains the GGxxDAGD, PHHR, GAxxGG, YxDDI, and EVxxDYN motifs found in most glycoside hydrolase family 9 (GH9). The protein sequence showed the highest similarity (88%) with the cellulase of Bacillus sp. BP23 in comparison to the enzymes with reported properties. The enzyme was purified by chromatography using HiTrap Q, CHT-II, and HiTrap Butyl HP. Using SDS-PAGE/activity staining, the molecular mass of Cel9K was estimated to be 93 kDa, which is a truncated form produced by the proteolytic cleavage of its C-terminus. Cel9K was optimally active at pH 5.5 and 50°C and showed a half-life of 59.2 min at 50°C. The CMCase activity was increased to more than 150% in the presence of 2 mM Na⁺, K⁺, and Ba²âº, but decreased significantly to less than 50% by Mn²âº and Co²âº. The addition of Cel9K to a commercial enzyme set (Celluclast 1.5L + Novozym 188) increased the saccharification of the pretreated reed and rice straw powders by 30.4 and 15.9%, respectively. The results suggest that Cel9K can be used to enhance the enzymatic conversion of lignocellulosic biomass to reducing sugars as an additive.

Characterization of two extracellular ß-glucosidases produced from the cellulolytic fungus Aspergillus sp. YDJ216 and their potential applications for the hydrolysis of flavone glycosides.

A cellulolytic fungus YDJ216 was isolated from a compost and identified as an Aspergillus sp. strain. Two extracellular ß-glucosidases, BGL1 and BGL2, were purified using ultrafiltration, ammonium sulfate fractionation, and High-Q chromatography. Molecular masses of BGL1 and BGL2 were estimated to be 97 and 45 kDa, respectively, by SDS-PAGE. The two enzymes eluted as one peak at 87 kDa by Sephacryl S-200 chromatography, and located at similar positions in a zymogram after intact gel electrophoresis, suggesting BGL1 and BGL2 might be monomeric and dimeric, respectively. Both enzymes showed similar enzymatic properties; they were optimally active at pH 4.0-4.5 and 60 °C, and had similar half-lives at 70 °C. Two enzymes also preferred p-nitrophenyl glucose (pNPG) with the same Km and hardly hydrolyzed cellobiose, suggesting both enzymes are aryl ß-glucosidases. However, Vmax for pNPG of BGL1 (953.2 U/mg) was much higher than those of BGL2 (66.5U/mg) and other ß-glucosidases reported. When tilianin (a flavone glycoside of acacetin) was reacted with both enzymes, inhibitory activity for monoamine oxidase, relating to oxidation of neurotransmitter amines, was increased closely to the degree obtained by acacetin. These results suggest that BGL1 and BGL2 could be used to hydrolyze flavone glycosides to improve their inhibitory activities.

Roles of Carbohydrate-Binding Module (CBM) of an Endo-ß-1,4-Glucanase (Cel5L) from Bacillus sp. KD1014 in Thermostability and Small-Substrate Hydrolyzing Activity.

An endo-ß-1,4-glucanase gene, cel5L, was cloned using the shot-gun method from Bacillus sp.. The gene, which contained a predicted signal peptide, encoded a protein of 496 amino acid residues, and the molecular mass of the mature Cel5L was estimated to be 51.8 kDa. Cel5L contained a catalytic domain of glycoside hydrolase (GH) family 5 and a carbohydrate-binding module family 3 (CBM_3). Chromatography using HiTrap Q and CHT-II resulted in the isolation of two truncated forms corresponding to 50 (Cel5L-p50) and 35 kDa (Cel5L-p35, CBM_3-deleted form). Both enzymes were optimally active at pH 4.5 and 55°C, but had different half-lives of 4.0 and 22.8 min, respectively, at 70°C. The relative activities of Cel5L-p50 and Cel5L-p35 for barley ß-glucan were 377.0 and 246.7%, respectively, compared to those for carboxymethyl-cellulose. The affinity and hydrolysis rate of pNPC by Cel5L-p35 were 1.7 and 3.3 times higher, respectively, than those by Cel5L-p50. Additions of each to a commercial enzyme set increased saccharification of pretreated rice straw powder by 17.5 and 21.0%, respectively. These results suggest CBM_3 is significantly contributing to thermostability, and to affinity and substrate specificity for small substrates, and that these two enzymes could be used as additives to enhance enzymatic saccharification.

Association between Doppler flow of atrial fibrillatory contraction and recurrence of atrial fibrillation after electrical cardioversion.

BACKGROUND: Left atrial fibrillatory contraction (Afc) flow can be frequently observed interspersed between two successive mitral E waves in patients with atrial fibrillation (AF). The aim of this study was to test the hypothesis that Afc is related to the maintenance of sinus rhythm after electrical cardioversion for AF. METHODS: In this retrospective study, the records of a total of 137 patients with AF who underwent successful electrical cardioversion were examined. Conventional echocardiographic measurements, including left atrial volume index (LAVI), were obtained, and the appearance of Afc flow was also evaluated before cardioversion. Patients were followed to a clinical end point defined as recurrent AF during the study period. RESULTS: AF recurrence was noted in 100 patients (73%) over a mean follow-up period of 5 months. The patients with recurrent AF had greater LAVI and left atrial dimensions and had a lower frequency of Afc flow (57.0% vs 86.5%, P < .001): both the velocity and velocity-time integral (VTI) of Afc flow significantly decreased. Receiver operating characteristic curve analysis showed that the Afc flow VTI and velocity had stronger associations with AF recurrence than did LAVI (areas under the curve: VTI, 0.96; velocity, 0.86; LAVI, 0.71). A VTI of 3.1 cm and velocity of 32 cm/sec for Afc flow were the best cutoff values for AF recurrence. Afc flow VTI (hazard ratio, 0.70; 95% confidence interval, 0.51-0.96) and velocity (hazard ratio, 0.97; 95% confidence interval, 0.94-0.99) were significantly related to AF recurrence in a multivariate Cox regression analysis. CONCLUSIONS: Return of AF after successful electrical cardioversion may be associated with Afc Doppler flow velocity and VTI measured immediately before cardioversion.

The efficacy of the cystatin C based glomerular filtration rate in the estimation of safe contrast media volume.

BACKGROUND AND OBJECTIVES: The risk of contrast-induced nephropathy (CIN) is significantly influenced by baseline renal function and the amount of contrast media (CM). We evaluated the usefulness of the cystatin C (CyC) based estimated glomerular filtration rate (eGFRCyC) in the prediction of CIN and to determine the safe CM dosage. SUBJECTS AND METHODS: We prospectively enrolled a total of 723 patients who received percutaneous coronary intervention (PCI) and investigated the clinical factors associated with the development of CIN. Renal function was calculated as eGFRCyC and a modified diet in the renal disease (MDRD) equation, respectively. Systemic exposure of CM was calculated as CM volume to eGFR ratio. We conducted a regression analysis to evaluate the predictive role of CM volume to eGFRCyC for the risk of CIN. RESULTS: The incidence of CIN was 4.0% (29/723). The patients with CIN had a lower hemoglobin level, decreased renal function, and a higher CyC value, and had greater CM exposure. Through multivariate regression analyses, hemoglobin {odds ratio (OR) 0.743, p=0.032}, CM volume/eGFRCyC (OR 1.697, p=0.006) and CM volume/MDRD (OR 2.275, p<0.001) were found to be independent predictors for CIN. In the receiver operating characteristic curve analysis, fair discrimination for CIN was found at a CM volume/eGFRCyC level of 4.493 (C-statics=0.814), and at this value, the sensitivity and specificity were 79.3% and 80.0%, respectively. CONCLUSION: Both the CM volume/MDRD and CM volume/eGFRCyC method would be simple, useful indicators for determining the safe CM-dose based on eGFR value before PCI. However, there was no significantly different predictive value between creatinine and CyC based GFR estimations.