Inferring complex evolutionary history of the closely related East Asian wild roses in Rosa sect. Synstylae (Rosaceae) based on genomic evidence from conserved orthologs.

BACKGROUND AND AIMS: The section Synstylae in genus Rosa (Rosaceae) comprises 25-36 species and includes several major progenitors of modern rose cultivars. East Asian Synstylae species have recently diverged and are closely related, and their phylogenetic relationships remain unclear. In the present study, we employed a conserved ortholog set (COS) markers and genome-wide nuclear orthologs to elucidate their phylogenetic relationships and unravel their complex evolutionary history. METHODS: Utilising on eight Rosaceae COS (RosCOS) markers, we analysed a total of 137 accessions representing 15 East Asian Synstylae taxa to establish a robust phylogenetic framework and reconstruct ancestral areas. Furthermore, we constructed the species tree for eight representative species and estimated their divergence times based on 1,683 genome-wide orthologs. The species tree-gene tree coalescence time comparison, Patterson's D, f4-ratio, and f-branch statistics were analysed to identify incomplete lineage sorting (ILS), genetic introgression, and reticulation events using conserved ortholog data. KEY RESULTS: RosCOS markers and genome-wide orthologs effectively resolved the robust phylogeny of East Asian Rosa sect. Synstylae. Species divergence times estimated with genome-wide orthologs indicated that East Asian Synstylae species have recently diverged, with an estimated crown age of approximately 2 Mya. The rampant gene tree discordance indicated the possibility of ILS and/or genetic introgression. In the section Synstylae, deeper coalescence in the gene trees compared to the species tree suggested ILS as a source of gene tree discordance. Further, Patterson's D and f-branch statistics indicated that several lineages in the section were involved in genetic introgression. CONCLUSIONS: We have unravelled the complex evolutionary history of East Asian Rosa sect. Synstylae, including recent species divergences, ILS, and genetic introgression. Coupled with the geographical and ecological complexity of East Asia, ILS and genetic introgression may have contributed to the rapid diversification of East Asian Synstylae species by permitting adaptation to diverse environments.

In vitro modulatory effects of ginsenoside compound K, 20(S)-protopanaxadiol and 20(S)-protopanaxatriol on uridine 5'-diphospho-glucuronosyltransferase activity and expression.

We explored the inhibitory effect of ginsenoside compound K (CK), 20(S)-protopanaxadiol (PPD), and 20(S)-protopanaxatriol (PPT) on six uridine 5'-diphospho-glucuronosyltransferase (UGT) enzyme (UGT1A1, 1A3, 1A4, 1A6, 1A9, and 2B7) activities in human liver microsomes (HLMs) and 10 UGT enzyme (UGT1A1, 1A3, 1A4, 1A6, 1A9, 2B4, 2B7, 2B10, 2B15, and 2B17) activities in recombinant UGT isoforms.PPD was a potent inhibitor of UGT1A3 activity with half-maximal inhibitory concentration values of 5.62 and 3.38 µM in HLMs and recombinant UGT1A3, respectively. UGT1A3 inhibition by CK and PPD was competitive with inhibitory constant (Ki) values of 17.4 and 1.21 µM, respectively, and inhibition by PPT was non-competitive with a Ki value of 8.07 µM in HLMs. PPD exhibited more than 3.4-fold selectivity for UGT1A3 inhibition compared with other UGT isoforms inhibition, while CK and PPT showed more than 2.16- and 2.21-fold selectivity, respectively.PPD did not significantly increase the mRNA expression of UGT1A1, 1A3, 1A4, 1A9, and 2B7 in hepatocytes.Given the low plasma concentrations of PPD in healthy human subjects and the absence of induction potential on UGT isoforms, we conclude that PPD cause no pharmacokinetic interactions with other co-administered drugs metabolised by UGT1A3.

In Vitro Interaction of AB-FUBINACA with Human Cytochrome P450, UDP-Glucuronosyltransferase Enzymes and Drug Transporters.

AB-FUBINACA, a synthetic indazole carboxamide cannabinoid, has been used worldwide as a new psychoactive substance. Because drug abusers take various drugs concomitantly, it is necessary to explore potential AB-FUBINACA-induced drug-drug interactions caused by modulation of drug-metabolizing enzymes and transporters. In this study, the inhibitory effects of AB-FUBINACA on eight major human cytochrome P450s (CYPs) and six uridine 5'-diphospho-glucuronosyltransferases (UGTs) of human liver microsomes, and on eight clinically important transport activities including organic cation transporters (OCT)1 and OCT2, organic anion transporters (OAT)1 and OAT3, organic anion transporting polypeptide transporters (OATP)1B1 and OATP1B3, P-glycoprotein, and breast cancer resistance protein (BCRP) in transporter-overexpressing cells were investigated. AB-FUBINACA inhibited CYP2B6-mediated bupropion hydroxylation via mixed inhibition with Ki value of 15.0 µM and competitively inhibited CYP2C8-catalyzed amodiaquine N-de-ethylation, CYP2C9-catalyzed diclofenac 4'-hydroxylation, CYP2C19-catalyzed [S]-mephenytoin 4'-hydroxylation, and CYP2D6-catalyzed bufuralol 1'-hydroxylation with Ki values of 19.9, 13.1, 6.3, and 20.8 µM, respectively. AB-FUBINACA inhibited OCT2-mediated MPP+ uptake via mixed inhibition (Ki, 54.2 µM) and competitively inhibited OATP1B1-mediated estrone-3-sulfate uptake (Ki, 94.4 µM). However, AB-FUBINACA did not significantly inhibit CYP1A2, CYP2A6, CYP3A4, UGT1A1, UGT1A3, UGT1A4, UGT1A6, or UGT2B7 enzyme activities at concentrations up to 100 µM. AB-FUBINACA did not significantly inhibit the transport activities of OCT1, OAT1/3, OATP1B3, P-glycoprotein, or BCRP at concentrations up to 250 µM. As the pharmacokinetics of AB-FUBINACA in humans and animals remain unknown, it is necessary to clinically evaluate potential in vivo pharmacokinetic drug-drug interactions induced by AB-FUBINACA-mediated inhibition of CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, OCT2, and OATP1B1 activities.

Herb-Drug Interaction of Red Ginseng Extract and Ginsenoside Rc with Valsartan in Rats.

The purpose of this study was to investigate the herb-drug interactions involving red ginseng extract (RGE) or ginsenoside Rc with valsartan, a substrate for organic anion transporting polypeptide (OATP/Oatp) transporters. In HEK293 cells overexpressing drug transporters, the protopanaxadiol (PPD)-type ginsenosides- Rb1, Rb2, Rc, Rd, Rg3, compound K, and Rh2-inhibited human OATP1B1 and OATP1B3 transporters (IC50 values of 7.99-68.2 µM for OATP1B1; 1.36-30.8 µM for OATP1B3), suggesting the herb-drug interaction of PPD-type ginsenosides involving OATPs. Protopanaxatriol (PPT)-type ginsenosides-Re, Rg1, and Rh1-did not inhibit OATP1B1 and OATP1B3 and all ginsenosides tested didn't inhibit OCT and OAT transporters. However, in rats, neither RGE nor Rc, a potent OATP inhibitor among PPD-type ginsenoside, changed in vivo pharmacokinetics of valsartan following repeated oral administration of RGE (1.5 g/kg/day for 7 days) or repeated intravenous injection of Rc (3 mg/kg for 5 days). The lack of in vivo herb-drug interaction between orally administered RGE and valsartan could be attributed to the low plasma concentration of PPD-type ginsenosides (5.3-48.4 nM). Even high plasma concentration of Rc did not effectively alter the pharmacokinetics of valsartan because of high protein binding and the limited liver distribution of Rc. The results, in conclusion, would provide useful information for herb-drug interaction between RGE or PPD-type ginsenosides and Oatp substrate drugs.

Detection of 13 Ginsenosides (Rb1, Rb2, Rc, Rd, Re, Rf, Rg1, Rg3, Rh2, F1, Compound K, 20(S)-Protopanaxadiol, and 20(S)-Protopanaxatriol) in Human Plasma and Application of the Analytical Method to Human Pharmacokinetic Studies Following Two Week-Repeated Administration of Red Ginseng Extract.

We aimed to develop a sensitive method for detecting 13 ginsenosides using liquid chromatography-tandem mass spectrometry and to apply this method to pharmacokinetic studies in human following repeated oral administration of red ginseng extract. The chromatograms of Rb1, Rb2, Rc, Rd, Re, Rf, Rg1, Rg3, Rh2, F1, compound K (CK), protopanaxadiol (PPD), and protopanaxatriol (PPT) in human plasma were well separated. The calibration curve range for 13 ginsenosides was 0.5-200 ng/mL and the lower limit of quantitation was 0.5 ng/mL for all ginsenosides. The inter- and intra-day accuracy, precision, and stability were less than 15%. Among the 13 ginsenosides tested, nine ginsenosides (Rb1, Rb2, Rc, Rd, Rg3, CK, Rh2, PPD, and PPT) were detected in the human plasma samples. The plasma concentrations of Rb1, Rb2, Rc, Rd, and Rg3 were correlated with the content in red ginseng extract; however, CK, Rh2, PPD, and PPT were detected although they are not present in red ginseng extract, suggesting the formation of these ginsenosides through the human metabolism. In conclusion, our analytical method could be effectively used to evaluate pharmacokinetic properties of ginsenosides, which would be useful for establishing the pharmacokinetic-pharmacodymic relationship of ginsenosides as well as ginsenoside metabolism in humans.

Phylogenetic relationships and genetic diversity of the Korean endemic Phedimus latiovalifolius (Crassulaceae) and its close relatives.

Phedimus latiovalifolius (Y.N.Lee) D.C.Son & H.J.Kim is exclusively distributed in the high mountains in the Korean Peninsula, mainly along the Baekdudaegan mountain range. Despite its morphological and distributional distinction from other Phedimus Raf. species, its taxonomic identity and phylogenetic relationship with congeneric species remain unclear. This study employs genotyping-by-sequencing-derived genome-wide single nucleotide polymorphisms to establish the monophyly of P. latiovalifolius and its relationship with closely related species. Genetic diversity and population differentiation of P. latiovalifolius are also assessed to provide baseline genetic information for future conservation and management strategies. Our phylogenetic analyses robustly demonstrate the monophyletic nature of P. latiovalifolius, with P. aizoon (L.) 't Hart identified as its closest sister lineage. There is no genetic evidence supporting a hybrid origin of P. latiovalifolius from P. aizoon involving either P. ellacombeanus (Praeger) 't Hart or P. kamtschaticus (Fisch.) 't Hart. Population genetic analyses reveal two major groups within P. latiovalifolius. A higher genetic variation is observed in P. ellacombeanus than in the congeneric species. Notably, most of the genetic variation exists within P. latiovalifolius populations. Given its distribution and the potential role of Baekdudaegan as an East Asian Pleistocene refugia, P. latiovalifolius could be considered rare and endemic, persisting in the refugium across glacial/interglacial cycles.

Pharmacokinetics and Enterohepatic Circulation of 2-(Quinoline-8-carboxamido)benzoic Acid (2-QBA) in Mice.

The quinoline alkaloid 2-(quinoline-8-carboxamido)benzoic acid (2-QBA), which is isolated from Aspergillus sp. SCSIO06786, a deep sea-derived fungus, has been suggested as a therapeutic candidate for the treatment of Parkinson's disease. We developed an analytical method for 2-QBA using a liquid chromatography-tandem mass spectrometry (LC-MS/MS) in mouse plasma, in which a protein precipitation method for the sample preparation of 2-QBA in mouse plasma was used due to its simplicity and good extraction recovery rates (80.49-97.56%). The linearity of the calibration standard sample, inter- and intraday precision and accuracy, and stability of three quality control samples were suitable based on the assessment criteria and the lower limit of quantification (LLOQ) of the 2-QBA was 1 ng/mL. A pharmacokinetic study of 2-QBA was performed in mice divided into oral (2.0, 5.0, and 15 mg/kg) and intravenous (0.5 and 1.0 mg/kg) administration groups. The absolute oral bioavailability (BA) range of 2-QBA was calculated as 68.3-83.7%. Secondary peaks were observed at approximately 4-8 h after the oral administration of 2-QBA at all doses. The elimination half-life of the orally administered 2-QBA was significantly longer than that of the intravenous 2-QBA. In addition, glucuronide metabolites of 2-QBA were identified. They were transformed into 2-QBA using the ß-glucuronidase treatment. Furthermore, the 2-QBA was readily absorbed from the jejunum to lower ileum. Taken together, the secondary peaks could be explained by the enterohepatic circulation of 2-QBA. In conclusion, the reabsorption of orally administered 2-QBA could contribute to the high oral BA of 2-QBA and could be beneficial for the efficacy of 2-QBA. Moreover, the simple and validated analytical method for 2-QBA using LC-MS/MS was applied to the pharmacokinetic study and BA assessments of 2-QBA in mice and would be helpful for subsequent pharmacokinetic studies, as well as for evaluations of the toxicokinetics and pharmacokinetic-pharmacodynamic correlation of 2-QBA to assess its potential as a drug.

Experimental analysis of PM2.5 reduction characteristics between Korean red pine (Pinus densiflora) and sawtooth oak (Quercus acutissima) saplings under different densities and arrangement structures.

As global air pollution, particularly fine particulate matter (PM2.5), has become a major environmental problem, various PM2.5 mitigation technologies including green infrastructure have received significant attention. However, owing to spatial constraints on urban greening, there is a lack of management plans for urban forests to efficiently mitigate PM2.5. In this study, we assessed the PM2.5 reduction capabilities of Pinus densiflora (Korean red pine) and Quercus acutissima (sawtooth oak) by measuring the changes of PM2.5 concentrations using an experimental chamber system. In addition, the PM2.5 reduction efficiency in 90 min (PMRE90) and the amount of PM2.5 reduction per leaf area (PMRLA) were compared based on arrangement structures and density levels. The results showed that the PM2.5 reduction by plants was significantly greater than that of the control experiment without any plants, and an additional reduction effect of approximately 1.38 times was induced by a 1.5 m s-1 air flow. The PMRE90 of Korean red pine was the highest at medium density. In contrast, the PMRE90 of sawtooth oak was the highest at high density. The PMRLA of both species was highest at low densities. The different responses of the species to total reduction were well explained by total leaf area (TLA). The PMRE90 of both species was positively correlated with TLA. The PMRLA of sawtooth oak was approximately 2.3 times greater than that of Korean red pine. However, there were no significant differences in both PMRE90 and PMRLA between the arrangement structures. Our findings reveal the potential mechanisms of vegetation in reducing PM2.5 according to arrangement structure and density. This highlights the importance of efficiently using urban green spaces with spatial constraints on PM2.5 mitigation in the future.

Evaluation of the Drug-Induced Liver Injury Potential of Saxagliptin through Reactive Metabolite Identification in Rats.

A liver injury was recently reported for saxagliptin, which is a dipeptidyl peptidase-4 (DPP-4) inhibitor. However, the underlying mechanisms of saxagliptin-induced liver injury remain unknown. This study aimed to evaluate whether saxagliptin, a potent and selective DPP-4 inhibitor that is globally used for treating type 2 diabetes mellitus, binds to the nucleophiles in vitro. Four DPP-4 inhibitors, including vildagliptin, were evaluated for comparison. Only saxagliptin and vildagliptin, which both contain a cyanopyrrolidine group, quickly reacted with L-cysteine to enzyme-independently produce thiazolinic acid metabolites. This saxagliptin-cysteine adduct was also found in saxagliptin-administered male Sprague-Dawley rats. In addition, this study newly identified cysteinyl glycine conjugates of saxagliptin and 5-hydroxysaxagliptin. The observed metabolic pathways were hydroxylation and conjugation with cysteine, glutathione, sulfate, and glucuronide. In summary, we determined four new thiazoline-containing thiol metabolites (cysteine and cysteinylglycine conjugates of saxagliptin and 5-hydroxysaxagliptin) in saxagliptin-administered male rats. Our results reveal that saxagliptin can covalently bind to the thiol groups of cysteine residues of endogenous proteins in vivo, indicating the potential for saxagliptin to cause drug-induced liver injury.

Comparisons of PM2.5 mitigation with stand characteristics between evergreen Korean pine plantations and deciduous broad-leaved forests in the Republic of Korea.

Owing to industrialization and urbanization in recent decades, fine particulate matter (PM2.5) in the atmosphere has become a major environmental problem worldwide. This environmental issue pushed the use of forests as air filtering tools. However, there is a lack of continuous and long-term forest management to efficiently mitigate PM2.5. In this study, we assessed the potential of different forest types to control air pollution by measuring the seasonal PM2.5 concentrations inside and outside the forest for one year. In addition, the PM2.5 reduction efficiencies (PMREs) of two forest types were compared, and their relationship with stand characteristics was analyzed. The results showed that the average PMRE inside the forests was approximately 18.2%; the seasonal PMRE was highest in winter (approximately 28.1%) and lowest in summer (approximately 9.6%). The average PMRE of the Taehwa deciduous broad-leaved forest (TDF) (approximately 18.8%) was significantly higher than that of the Taehwa coniferous forest (TCF) (approximately 17.5%) (P < 0.001); differences were also observed seasonally. The PMRE in the TCF was higher in spring and summer (P < 0.001), while that in the TDF was higher in autumn and winter (P < 0.001). Furthermore, the PMRE in the TDF was negatively correlated with stand density (P = 0.003) and positively correlated with the average diameter at breast height (DBH) (P = 0.028). However, the PMRE in the TCF did not significantly correlate with stand characteristics. As such, the results of this study revealed the differences in PM2.5 mitigation according to stand characteristics, which should be considered in urban forest management.

Concomitant Administration of Red Ginseng Extract with Lactic Acid Bacteria Increases the Plasma Concentration of Deglycosylated Ginsenosides in Healthy Human Subjects.

With the increased frequency of red ginseng extract (RGE) and lactic acid bacteria (LAB) co-administration, we aimed to investigate the interactions between RGE and LAB with regard to in vitro and in vivo deglycosylation metabolism and the pharmacokinetics of ginsenosides. As a proof-of-concept study, five healthy humans were administered RGE (104.1 mg of total ginsenosides/day) with or without co-administration of LAB (2 g, 1 billion CFU/day) for 2 weeks, and the plasma concentrations of ginsenosides in human plasma were monitored. The plasma exposure to compound K (CK), ginsenoside Rh2 (GRh2), protopanaxadiol (PPD), and protopanaxatriol (PPT) in the concomitant administration RGE and LAB groups increased by 2.7-, 2.1-, 1.6-, and 3.5-fold, respectively, compared to those in the RGE administration group, without a significant change in Tmax. The plasma concentrations of GRb1, GRb2, and GRc remained unchanged, whereas the AUC values of GRd and GRg3 significantly decreased in the concomitant administration RGE and LAB groups. To understand the underlying mechanism, the in vitro metabolic activity of ginsenosides was measured during the fermentation of RGE or individual ginsenosides in the presence of LAB for 1 week. Consistent with the in vivo results, co-incubation with RGE and LAB significantly increased the formation rate of GRh2, CK, PPD, and PPT. These results may be attributed to the facilitated deglycosylation of GRd and GRg3 and the increased production of GRh2, CK, PPD, and PPT by the co-administration of LAB and RGE. In conclusion, LAB supplementation increased the plasma concentrations of deglycosylated ginsenosides, such as GRh2, CK, PPD, and PPT, through facilitated deglycosylation metabolism of ginsenosides in the intestine.

Simultaneous Analysis of a Combination of Anti-Hypertensive Drugs, Fimasartan, Amlodipine, and Hydrochlorothiazide, in Rats Using LC-MS/MS and Subsequent Application to Pharmacokinetic Drug Interaction with Red Ginseng Extract.

Fimasartan, amlodipine, and hydrochlorothiazide are commonly used in combination therapies as antihypertensive drugs. This study aimed to develop and validate an analytical method for fimasartan, its active and major metabolite fimasartan-amide, amlodipine, and hydrochlorothiazide in rat plasma using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The standard calibration curves for fimasartan (1−500 ng/mL), its active and major metabolite fimasartan-amide (0.3−100 ng/mL), amlodipine (0.5−200 ng/mL), and hydrochlorothiazide (5−5000 ng/mL) were linear with R2 > 0.9964, and the inter- and intra-day accuracy and precision and stability were within the acceptable criteria. Using this validated analytical method, the pharmacokinetic interaction of these triple combination drugs between single administration and concomitant administration of the triple combination was investigated; the results did not reveal a significant difference in any of the pharmacokinetic parameters. Based on these results, we investigated the effects of red ginseng extract (RGE) on the pharmacokinetics of fimasartan, fimasartan-amide, amlodipine, and hydrochlorothiazide after oral administration of the combination in rats. No significant difference was observed in the pharmacokinetic parameters of fimasartan, fimasartan-amide, amlodipine, and hydrochlorothiazide, except for the Tmax values of amlodipine. The delayed Tmax value of amlodipine was attributed to its decreased intestinal permeability after repeated RGE treatments. In conclusion, using a combination of antihypertensive drugs and simultaneous analytical methods, we established efficient drug interaction and toxicokinetic studies using a small number of animals.

Enhanced bioavailability and hepatoprotective effect of silymarin by preparing silymarin-loaded solid dispersion formulation using freeze-drying method.

This study aimed to develop a solid dispersion formulation of silymarin (Silymarin-SD) using freeze-drying method to enhance its oral bioavailability (BA) by inhibiting the intestinal first-pass effect and increasing its solubility and permeability. Silymarin-SD formulation (i.e., silymarin:tween 80:hydroxypropyl cellulose (HPC) = 1:1:3 (w/w/w) significantly increased silymarin permeability in the duodenum, jejunum, and ileum by decreasing the efflux ratio of silymarin and by inhibiting silymarin-glucuronidation activity, in which tween 80 played a crucial role. As a result, orally administered Silymarin-SD formulation increased plasma silymarin concentrations and decreased silymarin-glucuronide in rats compared with silymarin alone and silymmarin:D-α-tocopherol polyethylene glycol 1000 succinate (1:1, w/w) formulation. In addition to modulating intestinal first-pass effect, Silymarin-SD formulation showed a significantly higher cumulative dissolution for 120 min compared with that of silymarin from the physical mixture (PM) of the same composition as Silymarin-SD and silymarin alone; the relative BA of silymarin-SD increased to 215% and 589% compared with silymarin-PM and silymarin alone, respectively. This could be attributed to the amorphous status of the Silymarin-SD formulation without chemical interaction with excipients, such as tween 80 and HPC. Moreover, the hepatoprotective effect of Silymarin-SD in acetaminophen-induced acute hepatotoxicity, as estimated from the alanine aminotransferase and aspartate aminotransferase values, was superior to that of silymarin. In conclusion, the increase in the dissolution rate and intestinal permeability of silymarin, and the inhibition of silymarin-glucuronidation by the Silymarin-SD formulation, prepared using tween 80 and HPC, increased its plasma concentration and resulted in a superior hepatoprotective effect compared to silymarin.

Pharmacokinetics and Tissue Distribution of Enavogliflozin in Mice and Rats.

This study investigated the pharmacokinetics and tissue distribution of enavogliflozin, a novel sodium-glucose cotransporter 2 inhibitor that is currently in phase three clinical trials. Enavogliflozin showed dose-proportional pharmacokinetics following intravenous and oral administration (doses of 0.3, 1, and 3 mg/kg) in both mice and rats. Oral bioavailability was 84.5-97.2% for mice and 56.3-62.1% for rats. Recovery of enavogliflozin as parent form from feces and urine was 39.3 ± 3.5% and 6.6 ± 0.7%, respectively, 72 h after its intravenous injection (1 mg/kg), suggesting higher biliary than urinary excretion in mice. Major biliary excretion was also suggested for rats, with 15.9 ± 5.9% in fecal recovery and 0.7 ± 0.2% in urinary recovery for 72 h, following intravenous injection (1 mg/kg). Enavogliflozin was highly distributed to the kidney, which was evidenced by the AUC ratio of kidney to plasma (i.e., 41.9 ± 7.7 in mice following its oral administration of 1 mg/kg) and showed slow elimination from the kidney (i.e., T1/2 of 29 h). It was also substantially distributed to the liver, stomach, and small and large intestine. In addition, the tissue distribution of enavogliflozin after single oral administration was not significantly altered by repeated oral administration for 7 days or 14 days. Overall, enavogliflozin displayed linear pharmacokinetics following intravenous and oral administration, significant kidney distribution, and favorable biliary excretion, but it was not accumulated in the plasma and major distributed tissues, following repeated oral administration for 2 weeks. These features may be beneficial for drug efficacy. However, species differences between rats and mice in metabolism and oral bioavailability should be considered as drug development continues.

Canopy height affects the allocation of photosynthetic carbon and nitrogen in two deciduous tree species under elevated CO2.

Down-regulation of leaf N and Rubisco under elevated CO2 (eCO2) are accompanied by increased non-structural carbohydrates (NSC) due to the sink-source imbalance. Here, to investigate whether the canopy position affects the down-regulation of Rubisco, we measured leaf N, NSC and N allocation in two species with different heights at maturity [Fraxinus rhynchophylla (6.8 ± 0.3 m) and Sorbus alnifolia (3.6 ± 0.2 m)] from 2017 to 2019. Since 2009, both species were grown at three different CO2 concentrations in open-top chambers: ambient CO2 (400 ppm; aCO2); ambient CO2 × 1.4 (560 ppm; eCO21.4); and ambient CO2 × 1.8 (720 ppm; eCO21.8). Leaf N per unit mass (Nmass) decreased under eCO2, except under eCO21.8 in S. alnifolia and coincided with increased NSC. NSC increased under eCO2 in F. rhynchophylla, but the increment of NSC was greater in the upper canopy of S. alnifolia. Conversely, Rubisco content per unit area was reduced under eCO2 in S. alnifolia and there was no interaction between CO2 and canopy position. In contrast, the reduction of Rubisco content per unit area was greater in the upper canopy of F. rhynchophylla, with a significant interaction between CO2 and canopy position. Rubisco was negatively correlated with NSC only in the upper canopy of F. rhynchophylla, and at the same NSC, Rubisco was lower under eCO2 than under aCO2. Contrary to Rubisco, chlorophyll increased under eCO2 in both species, although there was no interaction between CO2 and canopy position. Finally, photosynthetic N content (Rubisco + chlorophyll + PSII) was reduced and consistent with down-regulation of Rubisco. Therefore, the observed Nmass reduction under eCO2 was associated with dilution due to NSC accumulation. Moreover, down-regulation of Rubisco under eCO2 was more sensitive to NSC accumulation in the upper canopy. Our findings emphasize the need for the modification of the canopy level model in the context of climate change.

Opposite functions of RapA and RapC in cell adhesion and migration in Dictyostelium.

There are three Rap proteins in Dictyostelium. RapA is a key regulator of cell adhesion and cytoskeletal rearrangement. Recently, RapC has been reported to be involved in cytokinesis, cell migration, and multicellular development. Here, we compare the functions of RapA and RapC using cells expressing or lacking Rap proteins, and confirm that RapA and RapC have opposite functions in cell spreading, adhesion, and migration. On the other hand, RapC has a unique function in cytokinesis and multicellular development. Activated RapA appears to stimulate spreading and adhesion of the cells to the substrate, possibly resulting in a decrease in the migration speed of the cells during chemotaxis without affecting the directionality, whereas RapC suppresses cell spreading and adhesion, thereby increasing the migration speed. Cells lacking RapC were defective in cytokinesis and multicellular development and showed multinucleation and formation of multiple tips from a mound during development. At the C-terminus, RapC has an additional stretch of amino acids, which is not found in RapA. The mechanism through which RapA and RapC perform their opposite functions in diverse cellular processes should be characterized further to understand the Rap signaling pathways in detail. ABBREVIATIONS: GAP; GTPase-activating proteins; GEF; guanine nucleotide exchanging factor; WT; wild type; CA; constitutively active; DN; dominantly negative.

Enhanced Bioavailability and Efficacy of Silymarin Solid Dispersion in Rats with Acetaminophen-Induced Hepatotoxicity.

We evaluated the bioavailability, liver distribution, and efficacy of silymarin-D-α-tocopherol polyethylene glycol 1000 succinate (TPGS) solid dispersion (silymarin-SD) in rats with acetaminophen-induced hepatotoxicity (APAP) compared with silymarin alone. The solubility of silybin, the major and active component of silymarin, in the silymarin-SD group increased 23-fold compared with the silymarin group. The absorptive permeability of silybin increased by 4.6-fold and its efflux ratio decreased from 5.5 to 0.6 in the presence of TPGS. The results suggested that TPGS functioned as a solubilizing agent and permeation enhancer by inhibiting efflux pump. Thus, silybin concentrations in plasma and liver were increased in the silymarin-SD group and liver distribution increased 3.4-fold after repeated oral administration of silymarin-SD (20 mg/kg as silybin) for five consecutive days compared with that of silymarin alone (20 mg/kg as silybin). Based on higher liver silybin concentrations in the silymarin-SD group, the therapeutic effects of silymarin-SD in hepatotoxic rats were evaluated and compared with silymarin administration only. Elevated alanine aminotransferase, aspartate aminotransferase, and alkaline phosphatase levels were significantly decreased by silymarin-SD, silymarin, and TPGS treatments, but these decreases were much higher in silymarin-SD animals than in those treated with silymarin or TPGS. In conclusion, silymarin-SD (20 mg/kg as silybin, three times per day for 5 days) exhibited hepatoprotective properties toward hepatotoxic rats and these properties were superior to silymarin alone, which may be attributed to increased solubility, enhanced intestinal permeability, and increased liver distribution of the silymarin-SD formulation.

Genome-Wide Differential Methylation Profiles from Two Terpene-Rich Medicinal Plant Extracts Administered in Osteoarthritis Rats.

Extracts from the plants Phlomis umbrosa and Dipsacus asperoides-which are widely used in Korean and Chinese traditional medicine to treat osteoarthritis and other bone diseases-were used to treat experimental osteoarthritis (OA) rats. Genome-wide differential methylation regions (DMRs) of these medicinal-plant-treated rats were profiled as therapeutic evidence associated with traditional medicine, and they need to be investigated further using detailed molecular research to extrapolate traditional practices to modern medicine. In total, 49 protein-encoding genes whose expression is differentially regulated during disease progression and recovery have been discovered via systematic bioinformatic analysis and have been approved/proposed as druggable targets for various bone diseases by the US food and drug administration. Genes encoding proteins involved in the PI3K/AKT pathway were found to be enriched, likely as this pathway plays a crucial role during OA progression as well as during the recovery process after treatment with the aforementioned plant extracts. The four sub-networks of PI3K/AKT were highly regulated by these plant extracts. Overall, 29 genes were seen in level 2 (51-75%) DMRs and were correlated highly with OA pathogenesis. Here, we propose that these genes could serve as targets to study OA; moreover, the iridoid and triterpenoid phytochemicals obtained from these two plants may serve as potential therapeutic agents.

Effect of Lactic Acid Bacteria on the Pharmacokinetics and Metabolism of Ginsenosides in Mice.

This study aims to investigate the effect of lactic acid bacteria (LAB) on in vitro and in vivo metabolism and the pharmacokinetics of ginsenosides in mice. When the in vitro fermentation test of RGE with LAB was carried out, protopanaxadiol (PPD) and protopanaxadiol (PPD), which are final metabolites of ginsenosides but not contained in RGE, were greatly increased. Compound K (CK), ginsenoside Rh1 (GRh1), and GRg3 also increased by about 30%. Other ginsenosides with a sugar number of more than 2 showed a gradual decrease by fermentation with LAB for 7 days, suggesting the involvement of LAB in the deglycosylation of ginsenosides. Incubation of single ginsenoside with LAB produced GRg3, CK, and PPD with the highest formation rate and GRd, GRh2, and GF with the lower rate among PPD-type ginsenosides. Among PPT-type ginsenosides, GRh1 and PPT had the highest formation rate. The amoxicillin pretreatment (20 mg/kg/day, twice a day for 3 days) resulted in a significant decrease in the fecal recovery of CK, PPD, and PPT through the blockade of deglycosylation of ginsenosides after single oral administrations of RGE (2 g/kg) in mice. The plasma concentrations of CK, PPD, and PPT were not detectable without change in GRb1, GRb2, and GRc in this group. LAB supplementation (1 billion CFU/2 g/kg/day for 1 week) after the amoxicillin treatment in mice restored the ginsenoside metabolism and the plasma concentrations of ginsenosides to the control level. In conclusion, the alterations in the gut microbiota environment could change the ginsenoside metabolism and plasma concentrations of ginsenosides. Therefore, the supplementation of LAB with oral administrations of RGE would help increase plasma concentrations of deglycosylated ginsenosides such as CK, PPD, and PPT.

Down-regulation of photosynthesis and its relationship with changes in leaf N allocation and N availability after long-term exposure to elevated CO2 concentration.

Down-regulation of photosynthesis under elevated CO2 (eCO2) concentrations could be attributed to the depletion of nitrogen (N) availability after long-term exposure to eCO2 (progressive nitrogen limitation, PNL) or leaf N dilutions due to excessive accumulation of nonstructural carbohydrates. To determine the mechanism underlying this down-regulation, we investigated N availability, photosynthetic characteristics, and N allocation in leaves of Pinus densiflora (shade-intolerant species, evergreen tree), Fraxinus rhynchophylla (intermediate shade-tolerant species, deciduous tree), and Sorbus alnifolia (shade-tolerant species, deciduous tree). The three species were grown under three different CO2 concentrations in open-top chambers, i.e., ambient 400 ppm (aCO2); ambient × 1.4, 560 ppm (eCO21.4); and ambient × 1.8, 720 ppm (eCO21.8), for 11 years. Unlike previous studies that addressed PNL, after 11 years of eCO2 exposure, N availability remained higher under eCO21.8, and chlorophyll and photosynthetic N use efficiency increased under eCO2. In the case of nonstructural carbohydrates, starch and soluble sugar showed significant increases under eCO2. The maximum carboxylation rate, leaf N per mass (Nmass), and ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) were low under eCO21.8. The ratio of RuBP regeneration to the carboxylation rate as well as that of chlorophyll N to Rubisco N increased with CO2 concentrations. Based on the reduction in Nmass (not in Narea) that was diluted by increase in nonstructural carbohydrate, down-regulation of photosynthesis was found to be caused by the dilution rather than PNL. The greatest increases in chlorophyll under eCO2 were observed in S. alnifolia, which was the most shade-tolerant species. This study could help provide more detailed, mechanistically based processes to explain the down-regulation of photosynthesis by considering two hypotheses together and showed N allocation seems to be flexible against changes in CO2 concentration.