Triiodothyronine enhances various forms of kidney-specific Klotho protein and suppresses the Wnt/ß-catenin pathway: Insights from in-vitro, in-vivo and in-silico investigations.

Age-related diseases are intricately linked to the molecular processes underlying aging, with the decline of the antiaging protein Klotho being a key factor. Investigating these processes is crucial for developing therapeutic strategies. The age-associated reduction in Klotho expression, coupled with a decline in the endocrine hormone triiodothyronine (T3), prompted a detailed exploration of their potential interplay. Our research, conducted through both in-vitro and in-vivo studies on BALB/c mice, unveiled a significant capacity of T3 to upregulate various forms of Klotho via ATF-3/p-c-Jun transcription factor. This effect was particularly noteworthy in aged individuals, where Klotho expression had waned compared to their younger counterparts. Importantly, T3 demonstrated a promising therapeutic impact in rejuvenating Klotho expression in this context. Further investigations elucidated the molecular mechanisms underlying T3's impact on aging-related pathways. In-vitro and in-vivo experiments established T3's ability to downregulate the Wnt/ß-Catenin pathway by enhancing Klotho expression. In-silico analyses provided insights into Klotho's intricate role, showing its capacity to inhibit Wnt ligands such as Wnt3 and Wnt8a, consequently disrupting their interaction with the Wnt receptor. Additionally, T3 was found to downregulate kidney-specific GSK-3ß expression through the augmentation of Klotho expression. The study also highlighted T3's role in maintaining calcium and phosphate homeostasis via Klotho. This comprehensive investigation not only sheds light on the intricate mechanisms governing aging processes but also presents promising avenues for therapeutic interventions targeting the Wnt/ß-Catenin pathway implicated in various age-associated diseases.

Elucidation of 7,8-dihydroxy flavone in complexing with the oxidative stress-inducing enzymes, its impact on radical quenching and DNA damage: an in silico and in vitro approach.

Oxidative stress (OS) has been attributed to the progression of various disorders, including cancer, diabetes, and cardiovascular diseases. Several antioxidant compounds and free radical quenchers have been shown to mitigate oxidative stress. However, large-scale randomized controlled trials of such compounds on chronic disease aversion have yielded paradoxical and disappointing results due to the constrained cognizance of their oxidative mechanisms and therapeutic targets. The current study sought to identify the potential therapeutic targets of 7,8-Dihydroxyflavone (7,8-DHF) by analyzing its interactions with the enzymes implicated in oxidative stress and also to explore its radicle quenching potential and prophylactic impact on the H2O2-induced DNA damage. Through the in silco approach, we investigated the antioxidant potential of 7,8-DHF by evaluating its interactions with the human oxidative stress-inducing enzymes such as myeloperoxidase (MPO), NADPH oxidase (NOX), nitric oxide synthase (NOS), and xanthine oxidase (XO) and a comparative analysis of those interactions with known antioxidants (Ascorbic acid, Melatonin, Tocopherol) used as controls. The best-scoring complex was adopted for the simulation analysis in investigating protein-ligand conformational dynamics. The in vitro radicle quenching potential was evaluated by performing a spectrum of antioxidant assays, and radical quenching was observed in a dose-dependent fashion with IC50 values of < 60 µM/mL. Further, we probed its anti-hemolytic potential and prophylactic impact in avian erythrocytes subjected to H2O2-induced hemolysis and DNA damage by implementing hemolysis and comet assays. The protective effect was more pronounced at higher concentrations of the drug.Communicated by Ramaswamy H. Sarma.

Molecular modelling and biological evaluation of phyto-molecules as potential activators of gluconolactone oxidase (GULO).

Diabetes, the cause of colossal economic and disease burden, is a key area of research in drug discovery programs. Elevated blood glucose levels in diabetes lead to several adverse consequences due to the formation of advanced glycation end products and free radicals. Vitamin C, a potent antioxidant, protects the body's cells and tissues from oxidative damage and dysfunctions. Glucose is the precursor of Vitamin C synthesis in plants and some mammals. L-gulono lactone oxidase (GULO) is the rate-limiting enzyme in producing Vitamin C. However, it is not synthesized in bats, primates, humans, and guinea pigs because of the pseudogene. Several phytomolecules having antioxidant properties are hypothesized to be promising and selective activators of GULO. Therefore, the present study focused on screening agonists of GULO from phytomolecules as an effective augmentor for Vitamin C synthesis, thereby suppressing the sequela of diabetic events. The 3D structure of GULO was generated by the ab-initio method. Subsequently, molecular docking explored the possible binding patterns of GULO protein with different plant phenolic compounds, followed by supplementation of the potent phytomolecules to diabetic guinea pigs. It is noteworthy that Resveratrol and Hydroxytyrosol showed better binding affinity. The molecular simulation also confirmed that Resveratrol is an activator of the GULO enzyme. Interestingly, it was also established that Vitamin C levels were improved in diabetic guinea pigs supplemented with the phytomolecules and comparatively Resveratrol modulates the concentration of glucose and Vitamin C levels substantially, thereby alleviating hyperglycemia. However, further studies are warranted to study the mechanisms.Communicated by Ramaswamy H. Sarma.

Effect of mutation of NS2B cofactor residues on Dengue 2 NS2B-NS3 protease complex - an insight to viral replication.

Dengue fever is an endemic virus-borne disease that causes many severe ailments, including dengue hemorrhagic fever and dengue shock syndrome. NS2B-NS3 protease is present in all four strains of the dengue virus. NS2B-NS3 is a non-structural protein that performs three distinct functions: protease activity, helicase activity, and nucleoside triphosphatase activity. NS2B-NS3 pro-complex plays a crucial role in viral replication, and NS2B interacts with NS3 protease at a flat active site with an amino acid of the N-terminal region. NS2B acts as a cofactor for NS3 protease. In the current study, the conserved residues of NS2B were identified. Dengue virus-2 NS2B was mutated at the identified conserved amino acid region to investigate the role of NS2B on activation of NS3 pro. Molecular dynamics simulations were performed to investigate the mutated complex's changes in stability, conformation, and free energy. The EAG mutant complex exhibited more unstable conformation, less hydrogen bond formation, and high binding energy than wild type. This result suggests a vital role of E63, A65, G69 mutation in NS2B for the interruption of activation of the NS3.Communicated by Ramaswamy H. Sarma.

Uracil derivatives as HIV-1 capsid protein inhibitors: design, in silico, in vitro and cytotoxicity studies.

A series of novel uracil derivatives such as bispyrimidine dione and tetrapyrimidine dione derivatives were designed based on the existing four-point pharmacophore model as effective HIV capsid protein inhibitors. The compounds were initially docked with an HIV capsid protein monomer to rationalize the ideas of design and to find the potential binding modes. The successful design and computational studies led to the synthesis of bispyrimidine dione and tetrapyrimidine dione derivatives from uracil and aromatic aldehydes in the presence of HCl using novel methodology. The in vitro evaluation in HIV p24 assay revealed five potential uracil derivatives with IC50 values ranging from 191.5 µg ml-1 to 62.5 µg ml-1. The meta-chloro substituted uracil compound 9a showed promising activity with an IC50 value of 62.5 µg ml-1 which is well correlated with the computational studies. As expected, all the active compounds were noncytotoxic in BA/F3 and Mo7e cell lines highlighting the thoughtful design. The structure activity relationship indicates the position priority and lower log P values as the possible cause of inhibitory potential of the uracil compounds.

Molecular docking and simulation of IcaC protein as O-succinyltransferase function in staphylococcus epidermidis biofilm formation.

Intercellular adhesion (IcaADBC) operon is necessary for PNAG (Polyß-1,6-N-acetyl-D-glucosamine) biosynthesis of biofilm formation in Staphylococcus epidermidis. IcaC protein has a wide range of functions in terms of growth phase variation, migration, transposon insertion, PNAG modification, biofilm formation. Unusual TTTA signature motifs were identified from nucleotide sequence. Asparagine-linked glycosylation consensus motifs were identified at position 169 and 240. S. epidermidis was a close evolutionary association with S. haemolyticus and other Staphylococcus spp. Due to the non-availability of crystal structure, protein threading procedure was selected for constructing a full length IcaC three-dimensional structure. QMEANBrane structure quality assessment with model scores -100000 range within predicted integral membrane structure. IcaC motif constitutes 18 transmembrane helix, 37 helix-helix interaction, 8 beta turn, 2 gamma turn. Binding free energy was calculated with their succinate ligand docking form hydrogen bond with critical amino acids showed ΔG score -2.574 â€‹kJ/mol using Schrödinger. Serine (Ser96), Glutamic acid (Glu99), Tryptophan (Trp191) were active site amino acids form the catalytic core required for O-succinyltransferase function. Molecular dynamics simulation (MDS) was performed to evaluate the stability of IcaC protein and IcaC-Succinate binding complexes with the active site amino acids throughout trajectories captured with time scale 100 ns simulation period using GROMACS 4.5.

First-in-class pyrido[2,3-d]pyrimidine-2,4(1H,3H)-diones against leishmaniasis and tuberculosis: Rationale, in vitro, ex vivo studies and mechanistic insights.

Pyrido[2,3-d]pyrimidine-2,4(1H,3H)-diones were synthesized, for the first time, from indole chalcones and 6-aminouracil, and their ability to inhibit leishmaniasis and tuberculosis (Tb) infections was evaluated. The in vitro antileishmanial activity against promastigotes of Leishmania donovani revealed exceptional activities of compounds 3, 12 and 13, with IC50 values ranging from 10.23 ± 1.50 to 15.58 ± 1.67 µg/ml, which is better than the IC50 value of the standard drug pentostam of 500 µg/ml. The selectivity of the compounds towards Leishmania parasites was evaluated via ex vivo studies in Swiss albino mice. The efficiency of these compounds against Tb infection was then evaluated using the in vitro anti-Tb microplate Alamar Blue assay. Five compounds, 3, 7, 8, 9 and 12, showed MIC100 values against the Mycobacterium tuberculosis H37 Rv strain at 25 µg/ml, and compound 20 yielded an MIC100 value of 50 µg/ml. Molecular modelling of these compounds highlighted interactions with binding sites of dihydrofolate reductase, pteridine reductase and thymidylate kinase, thus establishing the rationale of their pharmacological activity against both pathogens, which is consistent with the in vitro results. From the above results, it is clear that compounds 3 and 12 are promising lead candidates for Leishmania and Mycobacterium infections and may be promising for coinfections.