High Affinity Binding Capacity Research Articles

Integrated computational and experimental evaluation of phenolic constituents of Apricot fruit L. for antiqourum sensing, antibiofilm, antioxidant, and 15-lox inhibitory properties

The present study investigated the antioxidant profile together with the antibacterial potential of Apricot L. with the aim to find a functional food based anti-infective lead. Additionally the study evaluated the biofilm and QS inhibitory potential of the plant using Pseudomonas aeruginosa (ATCC 15442) and Chromo bacterium Violaceum (DSM 30191) respectively. Several fractions of the peel of Apricot were subjected to initial antimicrobial and antibiofilm screening. Among all the fractions, methanol and ethyl acetate fractions displayed significant antimicrobial activity against the strains selected with MIC values 1.25 mg/dL and 1.68 mg/dL respectively. Similarly, while evaluating antiqourum-sensing potential, methanol extract showed remarkable zone of inhibition (14mm) with Violaceum inhibition (58%) while aqueous part presented moderately good inhibition (32%) with zone of inhibition of (4mm). N-hexane fraction was least active in this regard. In case of free radicals scavenging aptitudes, Ethanolic fraction displayed the highest free radicals scavenging potential (IC50μg/mL 13.76 ± 23.61) while Aqueous and ethyl acetate part exhibited moderate to good antioxidant behaviors with IC50μg/mL of 26.74 ± 22.00 and 19.49 ± 2.91 respectively. Then the selected compounds were screened for putative binding sites and molecular docking studies followed by enzyme inhibition assays. The negative binding energies and close proximity to residues in the binding pocket of selected targets including human α- soybean lox (PDB ID 1IK3), quorum sensing regulators LasR (2UV0) were observed which indicated high affinity and tight binding capacity of compounds 1 and 5 towards the active sites of LasR 2UV0 and 15-lipoxygenase. The physicochemical characteristics and toxicity expectation were computationally accomplished. Bioactivity prediction study revealed that all of the selected Phytoconstituents displayed incredible Bioactivity score. None of the selected chemical compound was found to be toxic as discovered by toxicity studies. Compound 4 exhibited the highest inhibition of 15-lipoxygenase in vitro (69%, at 0.037 mM final concentration) and that is accompanied by compound 5 (60%) whereas in the biofilm inhibition assay, compound 1 was most active (IC50 0.05 mM), followed by compound 3 (IC50 0.07 mM). It was therefore determined that compounds 1 and 3 had the highest biofilm inhibitory activity, whereas compounds 4 and 5 were potent 15-lipoxygenase inhibitors with potentially anti-inflammatory properties. Future investigations are suggested for the characterization and formulation development.

In Vitro Biological Evaluation of Aprepitant Based 177Lu-Radioconjugates.

Currently, the search for promising NK1R-positive tumor-targeting radiopharmaceuticals based on the structure of small molecular antagonists of neurokinin-1 receptor can be observed. Following this trend, we continued our evaluation of aprepitant-based 177Lu-radioconjugates in terms of future oncological applications. For this purpose, three novel aprepitant homologues were synthesized to broaden the previously obtained derivative portfolio, functionalized with the DOTA chelator and labeled with 68Ga and 177Lu. The newly evaluated radioconjugates showed the intended significant increase in lipophilicity compared to the previous ones, while maintaining stability in the human serum. Then, in a receptor binding study to the human NK1 receptor, we compared the two series of 177Lu-radioconjugates of aprepitant with each other and with the reference Substance P derivative currently used in glioblastoma therapy, clearly indicating the high affinity and better binding capacity of the novel radioconjugates. The in vitro experimental results included in the presented study, supported by labeling optimization, radioconjugate characterization and docking modeling of new aprepitant-derived radioagents, confirm our assumptions about the usefulness of aprepitant as a NK1R targeting vector and point out the perspectives for the forthcoming first in vivo trials.

Core–shell bioprinting as a strategy to apply differentiation factors in a spatially defined manner inside osteochondral tissue substitutes

One of the key challenges in osteochondral tissue engineering is to define specified zones with varying material properties, cell types and biochemical factors supporting locally adjusted differentiation into the osteogenic and chondrogenic lineage, respectively. Herein, extrusion-based core–shell bioprinting is introduced as a potent tool allowing a spatially defined delivery of cell types and differentiation factors TGF-β3 and BMP-2 in separated compartments of hydrogel strands, and, therefore, a local supply of matching factors for chondrocytes and osteoblasts. Ink development was based on blends of alginate and methylcellulose, in combination with varying concentrations of the nanoclay Laponite whose high affinity binding capacity for various molecules was exploited. Release kinetics of model molecules was successfully tuned by Laponite addition. Core–shell bioprinting was proven to generate well-oriented compartments within one strand as monitored by optical coherence tomography in a non-invasive manner. Chondrocytes and osteoblasts were applied each in the shell while the respective differentiation factors (TGF-β3, BMP-2) were provided by a Laponite-supported core serving as central factor depot within the strand, allowing directed differentiation of cells in close contact to the core. Experiments with bi-zonal constructs, comprising an osteogenic and a chondrogenic zone, revealed that the local delivery of the factors from the core reduces effects of these factors on the cells in the other scaffold zone. These observations prove the general suitability of the suggested system for co-differentiation of different cell types within a zonal construct.

A peptide-based vaccine ACP derived from antigens of Mycobacterium tuberculosis induced Th1 response but failed to enhance the protective efficacy of BCG in mice

BackgroundTuberculosis (TB) is a global infectious disease, but there is no ideal vaccine against TB except the Bacille Calmette-Guérin (BCG) vaccine. MethodsHerein, 25 candidate peptides were predicted from four antigens of Mycobacterium tuberculosis based on their high-affinity binding capacity for the human leukocyte antigen (HLA) DRB1∗0101. Three T-helper 1 (Th1) immunodominant peptides (Ag85B12-26, CFP2112-26, and PPE18149-163) were identified by ELISPOT assays in the humanized C57BL/6 mice. They resulted in a novel Th1 peptide-based vaccine ACP named by the first letter of the three peptides. In addition, the protective efficacy was evaluated in humanized or wild-type C57BL/6 mice and the humoral and cellular immune responses were confirmed in vitro. ResultsCompared with the PBS group, the ACP vaccinated mice showed slight decreases in colony-forming units (CFUs) and pathological lesions. However, when using it as a booster, the ACP vaccine did not significantly enhance the protective efficacy of BCG in humanized or wild-type mice. Interestingly, we found that ACP vaccination significantly increased the number of interferon-γ positive (IFN-γ+) T lymphocytes and the levels of IFN-γ cytokines as well as antibodies. Furthermore, the IL-2 level was significantly higher in humanized mice prime-boosted with BCG and ACP. ConclusionsOur results suggested that ACP vaccination could stimulate higher levels of cytokines and antibodies but failed to improve the protective efficacy of BCG in mice, indicating that the secretion level of IFN-γ may not be positively correlated with the protection efficiency of the vaccine. These findings provided important information on the feasibility of a peptide vaccine as a booster for enhancing the protective efficacy of BCG.

Peptides-Based Vaccine MP3RT Induced Protective Immunity Against Mycobacterium Tuberculosis Infection in a Humanized Mouse Model.

BackgroundTuberculosis (TB) is still a global infectious disease that seriously threatens human beings. The only licensed TB vaccine Bacille Calmette-Guérin (BCG)’s protective efficacy varies significantly among populations and regions. It is very urgent to develop more effective vaccines.MethodsIn this study, eleven candidate proteins of Mycobacterium tuberculosis were selected to predict peptides with high-affinity binding capacity for the HLA-DRB1*01:01 molecule. The immunodominant peptides were identified with the enzyme-linked immunospot assay (ELISPOT) and linked in silico to result in a novel polypeptide vaccine in Escherichia coli cells. The vaccine’s protective efficacy was evaluated in humanized and wild-type C57BL/6 mice. The potential immune protective mechanisms were explored with Enzyme-linked Immunosorbent Assay (ELISA), flow cytometry, and ELISPOT.ResultsSix immunodominant peptides screened from 50 predicted peptides were used to construct a new polypeptide vaccine named MP3RT. After challenge with M. tuberculosis, the colony-forming units (CFUs), lung lesion area, and the number of inflammatory cells in humanized mice rather than wild-type mice vaccinated with MP3RT were significantly lower than these in mice immunized with PBS. The humanized mice vaccinated with MP3RT revealed significant increases in IFN-γ cytokine production, IFN-γ+ T lymphocytes, CD3+IFN-γ+ T lymphocytes, and the MP3RT-specific IgG antibody.ConclusionsTaken together, MP3RT is a promising peptides-based TB vaccine characterized by inducing high levels of IFN-γ and CD3+IFN-γ+ T lymphocytes in humanized mice. These new findings will lay a foundation for the development of peptides-based vaccines against TB.

Direct Measurement of Aqueous Mercury(II): Combining DNA-Based Sensing with Diffusive Gradients in Thin Films.

A highly specific DNA-functionalized hydrogel sensing layer was integrated with the diffusive gradients in thin films (DGT) technique for the direct determination of aqueous mercury(II). The DNA-functionalized layer in the DGT unit exhibited both high affinity (complexation constant Kc = 1019.8 at 25 °C) and high binding capacity (9.5 mg Hg disk-1) toward Hg2+. The diffusion coefficient for Hg2+ complexed with common inorganic ligands was an order of magnitude higher than that for Hg2+ complexed with natural dissolved organic matter: 9.0 × 10-6 versus 9.8 × 10-7 cm2 s-1 at 25 °C. The performance of the DNA-DGT sensor was further assessed under variable pH (3-10) and temperature (5-40 °C) conditions, as well as across a range of hydrochemically diverse artificial and natural freshwaters. The observed effects of the environmental and solution compositional variables on Hg2+ binding to the DNA in the sensing layer were successfully accounted for by equilibrium speciation calculations and temperature-corrected, multicomponent diffusion coefficients for aqueous Hg(II). The results therefore support the use of the DNA-DGT sensor as an alternative to traditional sampling and analysis methods for measuring aqueous Hg(II) concentrations down to the nanomolar level in freshwater environments.

Sesquiterpene Coumarins from Ferula narthex 15-LOX, α-Glucosidase Inhibition and Molecular Docking Studies

Ferula narthex Boiss., Apiaceae, is commonly used in Pakistan and India as spice and it is associated with diverse biological activities including antidiabetic and anti-inflammatory. Research has suggested that the genus Ferula is rich in sesquiterpene coumarins. During this project, both in vitro and in silico models were adopted to investigate its traditional use. Based on structural diversity, selected compounds isolated from the exudate were processed for molecular docking studies. The negative binding energies and close proximity to residues in the binding pocket of selected targets including human α-glucosidase (PDB ID 3TOP) and soybean LOX (PDB ID 1IK3) were recorded, which indicated high affinity and tight binding capacity of 10′-R-acetyl-karatavacinol and 10′-R-karatavacinol towards the active sites of α-glucosidase and 15-lipoxygenase. 10′-R-acetyl-karatavacinol exhibited the highest inhibition of α-glucosidase in vitro (IC50 0.05 mM), whereas in the 15-lipoxygenase inhibition assay, 10′-R-karatavacinol was most active (70%, at 0.023 mM final concentration). It was therefore concluded that 10′-R-acetyl-karatavacinol had the highest α-glucosidase inhibitory activity, whereas 10′-R-karatavacinol was a 15-lipoxygenase inhibitor with potentially anti-inflammatory properties.

Didymin, a dietary citrus flavonoid exhibits anti-diabetic complications and promotes glucose uptake through the activation of PI3K/Akt signaling pathway in insulin-resistant HepG2 cells

Didymin is a naturally occurring orally active flavonoid glycoside (isosakuranetin 7-O-rutinoside) found in various citrus fruits, which has been previously reported to possess a wide variety of pharmacological activities including anticancer, antioxidant, antinociceptive, neuroprotective, hepatoprotective, inflammatory, and cardiovascular. However, there have not been any reports concerning its anti-diabetic potential until now. Therefore, we evaluated the anti-diabetic potential of didymin via inhibition of α-glucosidase, protein tyrosine phosphatase 1B (PTP1B), rat lens aldose reductase (RLAR), human recombinant AR (HRAR), and advanced glycation end-product (AGE) formation inhibitory assays. Didymin strongly inhibited PTP1B, α-glucosidase, HRAR, RLAR, and AGE in the corresponding assays. Kinetic study revealed that didymin exhibited a mixed type inhibition against α-glucosidase and HRAR, while it competitively inhibited PTP1B and RLAR. Docking simulations of didymin demonstrated negative binding energies and close proximity to residues in the binding pocket of HRAR, RLAR, PTP1B and α-glucosidase, indicating that didymin have high affinity and tight binding capacity towards the active site of these enzymes. Furthermore, we also examined the molecular mechanisms underlying the anti-diabetic effects of didymin in insulin-resistant HepG2 cells which significantly increased glucose uptake and decreased the expression of PTP1B in insulin-resistant HepG2 cells. In addition, didymin activated insulin receptor substrate (IRS)-1 by increasing phosphorylation at tyrosine 895 and enhanced the phosphorylations of phosphoinositide 3-kinase (PI3K), Akt, and glycogen synthasekinase-3(GSK-3). Interestingly, didymin reduced the expression of phosphoenolpyruvate carboxykinase and glucose 6-phosphatase, two key enzymes involved in the gluconeogenesis and leading to a diminished glucose production. The results of the present study clearly demonstrated that didymin will be useful for developing multiple target-oriented therapeutic modalities for treatment of diabetes, and diabetes-associated complications.

Chalcone derivatives from the root bark of Morus alba L. act as inhibitors of PTP1B and α-glucosidase

As part of our continuing research to obtain pharmacologically active compounds from Morus alba L. (Moraceae), four Diels-Alder type adducts (DAs) [morusalbins A−D], one isoprenylated flavonoid [albanin T], together with twenty-one known phenolic compounds were isolated from its root bark. The chemical structures were established using NMR, MS, and ECD spectra. The DAs including morusalbins A−D, albasin B, macrourin G, yunanensin A, mulberrofuran G and K, and albanol B exhibited strong inhibitory activities against both protein tyrosine phosphatase 1B (PTP1B) (IC50, 1.90−9.67 μM) and α-glucosidase (IC50, 2.29−5.91 μM). In the kinetic study, morusalbin D, albasin B, and macrourin G showed noncompetitive PTP1B inhibition, with Ki values of 0.33, 1.00, and 1.09 μM, respectively. In contrast, these DAs together with yunanensin A produced competitive inhibition of α-glucosidase, with Ki values of 0.64, 0.42, 2.42, and 1.19 μM, respectively. Furthermore, molecular docking studies revealed that these active DAs have high affinity and tight binding capacity towards the active site of PTP1B and α-glucosidase.

In Silico Docking and In Vitro Approaches towards BACE1 and Cholinesterases Inhibitory Effect of Citrus Flavanones.

Alzheimer’s disease (AD) is the most prevalent neurodegenerative disease, distinctively characterized by senile plaques, neurofibrillary tangles, and synaptic loss, finally resulting in neuronal death. β-Site amyloid precursor protein (APP) cleaving enzyme 1 (BACE1) and cholinesterases have been identified as therapeutic targets for AD, and the discovery of their inhibitors is of critical importance for developing preventive strategies for AD. To discover natural multi-target compounds possessing BACE1, acetylcholinesterase (AChE), and butyrylcholinesterase (BChE) inhibitory properties, major citrus flavanones including hesperetin, naringenin, and hesperidin were evaluated. In vitro anti-AD activities were performed via BACE1 and cholinesterases inhibition assays, as well as enzyme kinetic predictions. For the design of potential inhibitors of AD-related enzymes, molecular docking analysis was performed. Based on the biological evaluation, hesperidin demonstrated the best inhibitory properties toward BACE1, AChE, and BChE, with IC50 values of 10.02 ± 1.12, 22.80 ± 2.78, and 48.09 ± 0.74 µM, respectively. Kinetic studies revealed that all tested compounds were found to be noncompetitive inhibitors against BACE1 and cholineseterases. In addition, molecular docking studies of these compounds demonstrated negative binding energies for BACE1, AChE, and BChE, indicating high affinity and tight binding capacity for the target enzymes. The present study suggested that the selected citrus flavanones could act together as multiple inhibitors of BACE1, AChE, and BChE, indicating preventive and therapeutic potential against AD.

Development of a Phototoxicity Testing Strategy for Accurate Photosafety Evaluation of Pharmaceuticals Based on the Assessment of Possible Melanin-Binding Effects.

Drug-induced phototoxicity occurs when drugs absorb natural sunlight, leading to chemical reactions causing cellular damage. Distribution to light-exposed tissues is critical and is enhanced by binding to melanin. The International Council on Harmonization S10 guidance document on photosafety evaluation of pharmaceuticals states that although nonpigmented skin tends to be more sensitive than pigmented skin, pigmented skin models should be considered for drugs that bind significantly to melanin. In this study, an in vitro melanin-binding assay was evaluated as prescreening tool for animal model selection. Binding of various structurally diverse phototoxic drugs to synthetic melanin was investigated in vitro and the high-affinity binder sparfloxacin (SPX), moderate-affinity binder 8-methoxypsoralen (8-MOP), and low-affinity binder pirfenidone (PIF) were selected for in vivo studies. Pigmented Brown Norway (BN) rats were compared with nonpigmented Wistar Albino rats to evaluate their sensitivity for the assessment of phototoxicity and skin concentrations of the drugs were measured. For SPX, the onset of phototoxic symptoms was faster for BN rats and drug concentrations were significantly higher in skin of BN rats. For 8-MOP, both models showed comparable sensitivity and skin concentrations did not differ. For the low-affinity binder PIF, no phototoxic effects were observed and skin concentrations in both models were similar. A combined in vitro/in vivo approach was developed that can be applied for accurate photosafety evaluation of pharmaceuticals based on the assessment of possible melanin-binding effects. In view of the presented data, the pigmented model could be considered for compounds showing a high-affinity binding capacity in vitro.

Inhibitor discovery from pomegranate rind for targeting human salivary α-amylase

This study explored the effects of the main active compounds of the pomegranate (Punica granatum L.) rind extract on the activity of the human salivary α-amylase and their molecular inhibitory mechanisms. Four compounds exhibited remarkable inhibitory activities against α-amylase, including (1) rutin, (2) luteolin, (3) quercetin, and (4) kaempferol. The IC50 values were found to be 265.65, 59.67, 99.56, and 139.72 μM for rutin, luteolin, quercetin, and kaempferol, respectively. The kinetic study using the Lineweaver–Burk revealed the four compounds showed a non-competitive inhibition against α-amylase. However, the exact localization of the binding site and the potentiation mechanism at the molecular level are presently unknown. We have performed the “blind docking” of four compounds on the human salivary α-amylase. The molecular modeling demonstrated a high affinity and tight binding capacity of these compounds at the binding site of α-amylase, where the Glu 233 was supposed to play a key role in exerting the inhibition activity of these compounds. The results may provide an important insight for the applications of computational methods in the drug design with treating disorders of carbohydrate metabolism.

Dihydroxanthyletin-type coumarins from Angelica decursiva that inhibits the formation of advanced glycation end products and human recombinant aldose reductase.

The formation of advanced glycation end-products (AGE) and aldose reductase activity have been implicated in the development of diabetic complications. The present study was aimed to evaluate human recombinant aldose reductase (HRAR) and AGE inhibitory activity of seven natural dihydroxanthyletin-type coumarins, 4-hydroxy Pd-C-III (1), 4'-methoxy Pd-C-I (2), Pd-C-I (3), Pd-C-II (4), Pd-C-III (5), decursidin (6), and (+)-trans-decursidinol (7) from Angelica decursiva. Coumarins 1-7 showed potent HRAR and AGE inhibitory activities with ranges of IC50 values of 1.03-21.31 and 0.41-5.56µM, respectively. In the kinetic study for HRAR enzyme inhibition, coumarins 1, 3, 4, and 7 were competitive-type inhibitors, 6 was a mixed-type inhibitor, whereas 2 and 5 were noncompetitive-type inhibitors. Furthermore, we also predicted the docking interactions of HRAR with coumarins 1-7 using AutoDock Vina, and as a result, the simulated enzyme-inhibitor complexes exhibited negative binding energies (Autodock Vina=-9.6 to -8.1kcal/mol for HRAR), indicating a high affinity and tight binding capacity for the HRAR active site. Our results clearly indicate the potential HRAR and AGE formation inhibitory activities of dihydroxanthyletin-type coumarins, which could be further explored to develop therapeutic modalities for the treatment of diabetes and related complications.

α-Glucosidase and Protein Tyrosine Phosphatase 1B Inhibitory Activity of Plastoquinones from Marine Brown Alga Sargassum serratifolium.

Sargassum serratifolium C. Agardh (Phaeophyceae, Fucales) is a marine brown alga that belongs to the family Sargassaceae. It is widely distributed throughout coastal areas of Korea and Japan. S. serratifolium has been found to contain high concentrations of plastoquinones, which have strong anti-cancer, anti-inflammatory, antioxidant, and neuroprotective activity. This study aims to investigate the anti-diabetic activity of S. serratifolium and its major constituents through inhibition of protein tyrosine phosphatase 1B (PTP1B), α-glucosidase, and ONOO−-mediated albumin nitration. S. serratifolium ethanolic extract and fractions exhibited broad PTP1B and α-glucosidase inhibitory activity (IC50, 1.83~7.04 and 3.16~24.16 µg/mL for PTP1B and α-glucosidase, respectively). In an attempt to identify bioactive compounds, three plastoquinones (sargahydroquinoic acid, sargachromenol and sargaquinoic acid) were isolated from the active n-hexane fraction of S. serratifolium. All three plastoquinones exhibited dose-dependent inhibitory activity against PTP1B in the IC50 range of 5.14–14.15 µM, while sargachromenol and sargaquinoic acid showed dose-dependent inhibitory activity against α-glucosidase (IC50 42.41 ± 3.09 and 96.17 ± 3.48 µM, respectively). In the kinetic study of PTP1B enzyme inhibition, sargahydroquinoic acid and sargaquinoic acid led to mixed-type inhibition, whereas sargachromenol displayed noncompetitive-type inhibition. Moreover, plastoquinones dose-dependently inhibited ONOO−-mediated albumin nitration. Docking simulations of these plastoquinones demonstrated negative binding energies and close proximity to residues in the binding pocket of PTP1B and α-glucosidase, indicating that these plastoquinones have high affinity and tight binding capacity towards the active site of the enzymes. These results demonstrate that S. serratifolium and its major plastoquinones may have the potential as functional food ingredients for the prevention and treatment of type 2 diabetes.

Th1 epitope peptides induce protective immunity against Rickettsia rickettsii infection in C3H/HeN mice

Rickettsia rickettsii is the causative pathogen of Rocky Mountain spotted fever (RMSF). Adr2, YbgF and OmpB are protective antigens of R. rickettsii. In this study, 90 candidate peptides were selected from these antigens based on their high-affinity binding capacity for the MHC class II molecule H2 I-A or H2 I-E using bioinformatic methods. Six peptides were determined using ELISPOT assay to be immunodominant based on the IFN-γ recall responses of CD4+ T cells from mice immunized with R. rickettsii. Six nucleotide sequences encoding the immunodominant peptides were linked in series and inserted into a plasmid for expression in Escherichia coli cells, resulting in a new, recombinant polypeptide termed GWP. After immunization and challenge, the rickettsial load or histopathological lesions in the organs of mice immunized with GWP or pooled peptides was significantly lower than that in organs of mice immunized with PBS or the individual peptide OmpB399. An in vitro neutralization test revealed that sera from mice immunized with GWP, OmpB399, or pooled peptides reduced R. rickettsii adherence to, and invasion of, vascular endothelial cells. Furthermore, significantly higher levels of IgG, IgG1, or IgG2a were detected in sera from mice immunized with GWP or pooled peptides, and significantly higher levels of IFN-γ or TNF-α secreted by CD4+ T cells from R. rickettsii-infected mice were detected after immunization with GWP. Altogether, our results indicated that polypeptides, especially GWP, could induce a Th1-type immune response against R. rickettsii infection, which might contribute to the rational design of peptide-based vaccines for RMSF.

Spasmolytic Mechanism of Aqueous Licorice Extract on Oxytocin-Induced Uterine Contraction through Inhibiting the Phosphorylation of Heat Shock Protein 27.

Licorice derived from the roots and rhizomes of Glycyrrhiza uralensis Fisch. (Fabaceae), is one of the most widely-used traditional herbal medicines in China. It has been reported to possess significant analgesic activity for treating spastic pain. The aim of this study is to investigate the spasmolytic molecular mechanism of licorice on oxytocin-induced uterine contractions and predict the relevant bioactive constituents in the aqueous extract. The aqueous extraction from licorice inhibited the amplitude and frequency of uterine contraction in a concentration-dependent manner. A morphological examination showed that myometrial smooth muscle cells of oxytocin-stimulated group were oval-shaped and arranged irregularly, while those with a single centrally located nucleus of control and licorice-treated groups were fusiform and arranged orderly. The percentage of phosphorylation of HSP27 at Ser-15 residue increased up to 50.33% at 60 min after oxytocin stimulation. Furthermore, this increase was significantly suppressed by licorice treatment at the concentration of 0.2 and 0.4 mg/mL. Colocalization between HSP27 and α-SMA was observed in the myometrial tissues, especially along the actin bundles in the oxytocin-stimulated group. On the contrary, the colocalization was no longer shown after treatment with licorice. Additionally, employing ChemGPS-NP provided support for a preliminary assignment of liquiritigenin and isoliquiritigenin as protein kinase C (PKC) inhibitors in addition to liquiritigenin, isoliquiritigenin, liquiritin and isoliquiritin as MAPK-activated protein kinase 2 (MK2) inhibitors. These assigned compounds were docked with corresponding crystal structures of respective proteins with negative and low binding energy, which indicated a high affinity and tight binding capacity for the active site of the kinases. These results suggest that licorice exerts its spasmolytic effect through inhibiting the phosphorylation of HSP27 to alter the interaction between HSP27 and actin. Furthermore, our results provide support for the prediction that potential bioactive constituents from aqueous licorice extract inhibit the relevant up-stream kinases that phosphorylate HSP27.

Rosuvastatin Attenuates High-Salt and Cholesterol Diet Induced Neuroinflammation and Cognitive Impairment via Preventing Nuclear Factor KappaB Pathway

Recent attention is focused on the impact of diet on health and mental well-being. High-salt and cholesterol diet (HSCD) is known to be associated with neuroinflammation which is the predominant factor for neurodegenerative disease like Alzheimer disease (AD). In the present study, we examined the neuroprotective potential of rosuvastatin, an HMG-CoA reductase inhibitor against HSCD induced neuroinflammation and cognitive impairment. Our results demonstrated that HSCD-induced cognitive impairment as determined by Morris water maze (MWM) task. HSCD also activated nuclear factor kappaB (NF-kB) signaling pathway. The cytokine response was measured using a cytometric bead-based assay quantified by flow cytometry. Treatment with rosuvastatin decreased the production of nitric oxide (NO), tumor necrosis factor alpha (TNF-α) and increased interleukin-10 (IL-10) in a dose-dependent manner. Our results also demonstrated that the rosuvastatin modulates neuronal cell death by inhibiting the overexpression of NF-kB in the CA1 region of hippocampus. In addition, molecular docking study of rosuvastatin indicated high affinity and tighter binding capacity for the active site of the NF-kB. These results suggest that HSCD-triggered inflammatory response and cognitive impairment may be associated with NF-κB signaling pathway. Therefore, treatment with rosuvastatin could be a potential new therapeutic strategy for sporadic dementia of AD.

Poor glycemic control attributable to insulin antibody in a fulminant type 1 diabetes patient under hemodialysis: successful treatment with double filtration plasmapheresis and prednisolone

BackgroundInsulin antibody appears in approximately 40% of insulin users using human insulin and insulin analog formulations. Insulin antibody, characterized by high affinity and low insulin antibody binding capacity (% bound), rarely causes clinical problems including poor glycemic control. Recently, the insulin antibody against human insulin causing deterioration of glycemic control, characterized by low affinity and high insulin antibody binding capacity similar to those observed in insulin autoimmune syndrome, has been reported.Case presentationA 62-year-old male developing fulminant type 1 diabetes showing rapid hyperglycemia (602 mg/dL) and normal range of glycoalbumin (GA) (18.4%) in the introductory stage of hemodialysis (HD) was transferred to our hospital 3 weeks after the onset of diabetes. He had been administered insulin intensification therapy in other medical clinic; however, his glycemic control remained poor, showing ketoacidosis. After hospitalization, he was initially administered intravenous insulin for 2 days and then switched to subcutaneous intensification therapy (24 U/day); the dose was gradually increased maximum of 72 U/day within 2 months. His glycemic control deteriorated despite high-dose insulin administration and change in the insulin type. Insulin antibody was detected, insulin antibody binding capacity (% bound) was of a high value (89.5%), GA was 32.9%, and casual blood glucose widely changed in continuous glucose monitor, suggesting that unstable blood glucose fluctuations caused by insulin antibody; therefore, treatment with double filtration plasmapheresis (DFPP) and prednisolone (PSL) was performed. The insulin dose reduced, and glycemic control improved from 1 month after the commencement of treatment. Subsequently, insulin antibody level remained high; however, after 6 months, % bound declined (20.9%), and glycemic control improved (GA 23.7%) by PSL (gradually reduced to 5 mg/day). The Scatchard plots also indicated the effectiveness with DFPP and PSL therapy.ConclusionsThe authors present a unique case of fulminant type 1 diabetes developing during the HD introduction period. Although this is a rare complication, physicians specializing in dialysis therapy should also keep in mind that this is a life-threatening complication if the treatment is delayed and that concomitant therapy with apheresis such as DFPP and long-term steroids is effective in some cases when glycemic control deteriorates attributable to insulin antibody.

Kinetics and molecular docking studies of loganin, morroniside and 7-O-galloyl-D-sedoheptulose derived from Corni fructus as cholinesterase and β-secretase 1 inhibitors.

We evaluated the major active components isolated from Corni Fructus: loganin, morroniside, and 7-O-galloyl-D-sedoheptulose as inhibitors of acetylcholinesterase (AChE), butyrylcholinesterase (BChE), and β-site amyloid precursor protein (APP) cleaving enzyme 1 (BACE1) for use in Alzheimer's disease treatment. These compounds exhibited predominant cholinesterase (ChEs) inhibitory effects with IC50 values of 0.33, 3.95, and 10.50±1.16µM, respectively, for AChE, and 33.02, 37.78, and 87.94±4.66µM, respectively, for BChE. Kinetics studies revealed that loganin and 7-O-galloyl-D-sedoheptulose inhibited AChE with characteristics typical of mixed inhibitors, while morroniside was found to be a noncompetitive inhibitor against AChE and also exerted mixed BChE inhibitory activities. For BACE1, loganin showed noncompetitive type inhibitory effects, while morroniside and 7-O-galloyl-D-sedoheptulose were found to be mixed inhibitors. Furthermore, these compounds exhibited dose-dependent inhibitory activity with ONOO(-)-mediated protein tyrosine nitration. Molecular docking simulation of these compounds demonstrated negative binding energies for ChEs, and BACE1, indicating high affinity and tighter binding capacity for the active site of the enzyme. Loganin was the most potent inhibitor against both ChEs and BACE1. The data suggest that these compounds together can act as a triple inhibitor of AChE, BChE, and BACE1, providing a preventive and therapeutic strategy for Alzheimer's disease treatment.

Promising anti-diabetic potential of capillin and capillinol isolated from Artemisia capillaris.

Caffeoylquinic acids, flavonoids, and coumarins isolated from Artemisia capillaris have recently emerged as therapeutic candidates for diabetes and diabetic complications; however, there have been very few studies of the anti-diabetic potential of polyacetylenes. In the present study, we investigated the anti-diabetic potential of two polyacetylenes isolated from A. capillaris, namely capillin and capillinol by investigating their ability to inhibit α-glucosidase, protein tyrosine phosphatase 1B (PTP1B), and rat lens aldose reductase (RLAR). Capillin displayed potent inhibitory activity against α-glucosidase, PTP1B, and RLAR, while capillinol showed moderate inhibitory activity against α-glucosidase and PTP1B at the concentrations tested. In addition, a kinetic study revealed that capillin inhibited α-glucosidase and RLAR in a noncompetitive manner, while inhibited PTP1B in a mixed-type manner. Capillinol inhibited α-glucosidase and PTP1B in a mixed-type manner. Docking simulations of these compounds demonstrated negative binding energies and close proximity to residues in the binding pocket of PTP1B, indicating that these polyacetylenes have a high affinity and tight binding capacity for the active site of the enzyme. Furthermore, capillin dose-dependently inhibited peroxynitrite (ONOO(-))-mediated tyrosine nitration. The results clearly demonstrate the promising potential of capillin and capillinol as therapeutic interventions for the management of diabetes as well as diabetes-associated complications.