In Silico and In Vitro multiple analysis approach for screening naturally derived ligands for red seabream aryl hydrocarbon receptor.

The aryl hydrocarbon receptor (AHR) is a key ligand-dependent transcription factor that mediates the toxic effects of compounds such as dioxin. Recently, natural ligands of AHR, including flavonoids, have been attracting physiological and toxicological attention as they have been reported to regulate major biological functions such as inflammation and anti-cancer by reducing the toxic effects of dioxin. Additionally, it is known that natural AHR ligands can accumulate in wildlife tissues, such as fish. However, studies in fish have investigated only a few ligands in experimental fish species, and the AHR response of marine fish to natural AHR ligands of various other structures has not been thoroughly investigated. To explore various natural AHR ligands in marine fish, which make up the most fish, it is necessary to develop new screening methods that consider the specificity of marine fish. In this study, we investigated the response of natural ligands by constructing in vitro and in silico experimental systems using red seabream as a model species. We attempted to develop a new predictive model to screen potential ligands that can induce transcriptional activation of red seabream AHR1 and AHR2 (rsAHR1 and rsAHR2). This was achieved through multiple analyses using in silico/ in vitro data and Tox21 big data. First, we constructed an in vitro reporter gene assay of rsAHR1 and rsAHR2 and measured the response of 10 representatives natural AHR ligands in COS-7 cells. The results showed that FICZ, Genistein, Daidzein, I3C, DIM, Quercetin and Baicalin induced the transcriptional activity of rsAHR1 and rsAHR2, while Resveratrol and Retinol did not induce the transcriptional activity of rsAHR isoforms. Comparing the EC50 values of the respective compounds in rsAHR1 and rsAHR2, FICZ, Genistein, and Daidzein exhibited similar isoform responses, but I3C, Baicalin, DIM and Quercetin show the isoform-specific responses. These results suggest that natural AHR ligands have specific profiling and transcriptional activity for each rsAHR isoform. In silico analysis, we constructed homology models of the ligand binding domains (LBDs) of rsAHR1 and rsAHR2 and calculated the docking energies (U_dock values) of natural ligands with measured in vitro transcriptional activity and dioxins reported in previous studies. The results showed a significant correlation (R2=0.74(rsAHR1), R2=0.83(rsAHR2)) between docking energy and transcriptional activity (EC50) value, suggesting that the homology model of rsAHR1 and rsAHR2 can be utilized to predict the potential transactivation of ligands. To broaden the applicability of the homology model to diverse compound structures and validate the correlation with transcriptional activity, we conducted additional analyses utilizing Tox21 big data. We calculated the docking energy values for 1860 chemicals in both rsAHR1 and rsAHR2, which were tested for transcriptional activation in Tox21 data against human AHR. By comparing the U_dock energy values between 775 active compounds and 1085 inactive compounds, a significant difference (p<0.001) was observed between the U_dock energy values in the two groups, suggesting that the U_dock value can be applied to distinguish the activation of compounds. Furthermore, we observed a significant correlation (R2=0.45) between the AC50 of Tox21 database and U_dock values of human AHR model. In conclusion, we calculated equations to translate the results of an in silico prediction model for ligand screening of rsAHR1 and rsAHR2 transactivation. This ligand screening model can be a powerful tool to quantitatively estimate AHR transactivation of major marine agents to which red seabream may be exposed. The study introduces a new screening approach for potential natural AHR ligands in marine fish, based on homology model-docking energy values of rsAHR1 and rsAHR2, with implications for future agonist development and applications bridging in silico and in vitro data.

Molecular Markers for Detecting a Wide Range of Trichoderma spp. that Might Potentially Cause Green Mold in Pleurotus eryngii.

In Pleurotus sp., green mold, which is considered a major epidemic, is caused by several Trichoderma species. To develop a rapid molecular marker specific for Trichoderma spp. that potentially cause green mold, eleven Trichoderma species were collected from mushroom farms and the Korean Agricultural Culture Collection (KACC). A dominant fungal isolate from a green mold-infected substrate was identified as Trichoderma pleuroticola based on the sequences of its internal transcribed spacer (ITS) and translation elongation factor 1-α (tef1) genes. In artificial inoculation tests, all Trichoderma spp., including T. atroviride, T. cf. virens, T. citrinoviride, T. harzianum, T. koningii, T. longibrachiatum, T. pleurotum, and T. pleuroticola, showed pathogenicity to some extent, and the observed symptoms were soaked mycelia with a red-brown pigment and retarded mycelium regeneration. A molecular marker was developed for the rapid detection of wide range of Trichoderma spp. based on the DNA sequence alignment of the ITS1 and ITS2 regions of Trichoderma spp. The developed primer set detected only Trichoderma spp., and no cross reactivity with edible mushrooms was observed. The detection limits for the PCR assay of T. harzianum (KACC40558), T. pleurotum (KACC44537), and T. pleuroticola (CAF-TP3) were found to be 500, 50, and 5 fg, respectively, and the detection limit for the pathogen-to-host ratio was approximately 1:10,000 (wt/wt).

Severe Hypophosphatemia-Induced Acute Toxic-Metabolic Encephalopathy in Continuous Renal Replacement Therapy.

Acute toxic-metabolic encephalopathy (TME) is an acute condition of global cerebral dysfunction in the absence of primary structural brain disease. Severe hypophosphatemia leads to muscle weakness and involves the diaphragm but hypophosphatemia-induced TME is very rare. Herein, we report the case of a 43-year-old woman with encephalopathy with severe hypophosphatemia during continuous renal replacement therapy. She presented with features of oliguric acute kidney injury on diabetic kidney disease due to volume depletion. At admission, her mental status was alert but gradually changed to stupor mentation during continuous renal replacement therapy. Her phosphate level was less than 0.41 mEq/L and Glasgow coma scale decreased from 15 to 5. After phosphate intravenous replacement and administration of phosphate-containing replacement solution, the phosphate level increased to 2.97 mEq/L and mental state returned to alert state. This case demonstrates that the level of phosphorus should be observed during continuous renal replacement therapy.

The use of fermented buckwheat to produce L-carnitine enriched oyster mushroom.

L-Carnitine is an essential compound that shuttles long chain fatty acids into mitochondria. The objective of this study was to produce L-carnitine enriched oyster mushroom (Pleurotus ostreatus) using common buckwheat fermented by Rhizopus oligosporus. Mushroom grown on common buckwheat medium contained 9.9-23.9% higher L-carnitine (186.3 mg/kg) than those grown on basal medium without any buckwheat addition. Those grown on fermented common buckwheat medium contained the highest L-carnitine content (201.2 mg/kg). Size index and lightness of mushroom pileus (L*) were also the highest (100.7 and 50.6, respectively) for those grown in medium added with fermented common buckwheat (20%, w/w). Antioxidant activities of both mushroom extracts (1.5 mg/mL) showed the same level as 38.7% for mushroom grown in media added with common buckwheat or fermented common buckwheat. At the treatment concentration of 300 µg/mL, viabilities of murine macrophage cell line Raw 264.7 cells treated with ethanol extract of oyster mushroom grown on buckwheat medium ranged from 58.9 to 67.8%. The oyster mushroom grown on buckwheat and fermented buckwheat medium can be used as one of the substitutes for meat based diets.

Zirconium-Formate Macrocycles and Supercage: Molecular Packing versus MOF-like Network for Water Vapor Sorption.

In systematic efforts toward a new type of molecule-based porous materials, facile and efficient synthetic methods have been established to obtain macrocyclic [Zr6]6 and supercage-like {[Zr6]6}8, where [Zr6] represents [Zr6O4(OH)4(CO2) n] building unit commonly found in Zr-based metal-organic frameworks. The reactions involve in situ hydrolysis of DMF solvent to produce formate linkers and thus do not require any organic ligand. A minor variation in the composition of two cyclic hexamers thus obtained results in dramatic differences in crystal packing which in turn lead to distinctive and selective sorption behavior for water vapor. It is shown that the high heat of water adsorption and unrestricted uptake under high humidity are consequences of the highly polar surface and flexible crystal packing. The reversibility of water adsorption is demonstrated by cyclic measurements of uptake and regeneration under dynamic flow conditions.

Application of the modified clavien classification system to 402 cases of holmium laser enucleation of the prostate for benign prostatic hyperplasia.

PURPOSE: We attempted to evaluate the perioperative complications of holmium laser enucleation of the prostate (HoLEP) for benign prostatic hyperplasia by using the modified Clavien classification system (MCCS). MATERIALS AND METHODS: Targeting 402 patients who underwent HoLEP for benign prostatic hyperplasia performed by a single surgeon between July 2008 and January 2011, we investigated complications that occurred during and within 1 month after surgery and classified them into grade I to grade V on the basis of the MCCS. If two or more complications occurred in one patient, each complication was graded and counted. RESULTS: The mean age, prostate volume, operation time, hospital stay, and average follow-up period of 402 patients who underwent HoLEP were 68.8 years (range, 52-84 years), 53.2 g (range, 23-228 g), 58.2 minutes (range, 20-230 minutes), 4.5 days (range, 2-7 days), and 9 months (range, 4-27 months), respectively; 78 complications occurred in 71 of the patients (morbidity rate, 17.6%). In MCCS grade I, complications occurred in 54 cases (69.2%); in grade II, complications occurred in 19 cases (24.3%); in grade III, complications occurred in 4 cases (5.1%); and in grade IV, 1 patient required intensive care unit care because of cerebral infarction (1.2%). There were no grade V complications. CONCLUSIONS: The HoLEP-based MCCS complications classification was performed very quickly. However, MCCS, when compared with other measures of endoscopic prostate surgery experiences, including HoLEP, exposed the lack of accuracy in low grade classification and the inability to include late complications.

Holmium laser enucleation of the prostate: modified morcellation technique and results.

PURPOSE: We developed an inverse technique for tissue morcellation by modifying the conventional upward technique and then examined its safety and efficiency. MATERIALS AND METHODS: From July 2008 to December 2010, a total of 389 consecutive patients treated with holmium laser enucleation of the prostate (HoLEP) were enrolled in this study. For tissue morcellation, we used an upward technique for an initial series of 84 patients and an inverse technique for a consecutive series of 305 patients. We compared efficiency and safety between the inverse technique and the upward technique. RESULTS: There were no significant differences in mean age or prostate volume between the two groups. The mean morcellation efficiency was higher in the inverse technique group. The incidence of severe bladder injury was significantly higher in the upward technique group. Regarding the site of bladder injury, 7 and 4 cases of bladder injury occurred in the bladder dome and posterior wall, respectively, in the upward technique group. In the inverse technique group, however, the site of bladder injury was limited to the trigone. We divided our clinical series of patients into the upward technique group and three inverse technique groups on the basis of the timing. The mean morcellation efficiency was significantly higher in all three inverse technique groups than in the upward technique group. However, there was no significant difference in mean morcellation efficiency between the three inverse technique groups. CONCLUSIONS: In conclusion, the inverse technique might be a more effective, safer, and more excellent method of morcellation than the conventional upward technique.

Acute Urinary Retention due to Aseptic Meningitis: Meningitis-Retention Syndrome.

Acute urinary retention in aseptic meningitis is rarely encountered, and the diagnosis of aseptic meningitis may be less than straightforward, because its symptoms and neurological signs are occasionally mild or absent. We report a case in which acute urinary retention provided an appropriate indication for the diagnosis of aseptic meningitis as the cause of an undiagnosed fever.