Investigation of the insecticidal potential of curcumin derivatives that target the Helicoverpa armigera sterol carrier protein-2.

Cotton bollworm (Helicoverpa armigera) is a highly polyphagous, widely prevalent, and persistent Old World insect pest that affects numerous important crops that are directly consumed by people, including tomato, cotton, pigeon pea, chickpea, rice, sorghum, and cowpea. Insects do not synthesize steroids but obtain them from their diet. Inhibition of dietary uptake of steroids by insects is a potentially effective insecticidal mechanism that should not be toxic to humans and other mammals, who synthesize their steroids. Ten curcumin derivatives were tested against H. armigera sterol carrier protein-2 (HaSCP-2) for their potential as insecticidal agents. Curcumin derivatives were initially docked at the binding site of HaSCP-2 to determine their binding affinities and plausible binding modes. The binding modes predominantly show hydrophobic interactions of derivatives with Phe53, Phe110, and Phe89 as core interacting residues in the active site. Validation of in silico results was carried out by performing a fluorescence binding and displacement assay to determine the binding affinities of curcumin derivatives. Among a collection of curcumin derivatives tested, Cur10 showed the lowest IC50 value of 9.64 µM, while Cur07 was 19.86 µM, and Cur06 was 20.79 µM. There was a significant negative correlation between the ability of the curcumin derivatives tested to displace the fluorescent probe from the sterol binding site of HaSCP-2 and to inhibit Sf9 insect cell growth in culture, which is consistent with the curcumin derivatives acting by the novel mechanism of blocking sterol uptake. Then molecular dynamics simulation studies validated the predicted binding modes and the interactions of curcumin derivatives with HaSCP-2 protein. In conclusion, these studies support the potential use of curcumin derivatives as insecticidal agents.

RETRACTED: Ahmad et al. Neuroprotection against Aluminum Chloride-Induced Hippocampus Damage in Albino Wistar Rats by Leucophyllum frutescens (Berl.) I.M. Johnst. Leaf Extracts: A Detailed Insight into Phytochemical Analysis and Antioxidant and Enzyme Inhibition Assays. Front. Biosci. (Landmark Ed) 2023, 28(8), 184.

The Editors-in-Chief have retracted the article titled "[Neuroprotection against Aluminum Chloride-Induced Hippocampus Damage in Albino Wistar Rats by Leucophyllum frutescens (Berl.) I.M. Johnst. Leaf Extracts: A Detailed Insight into Phytochemical Analysis and Antioxidant and Enzyme Inhibition Assays]" ([1]) due to significant concerns regarding the reliability and integrity of the data presented. After the publication of the article, several issues were brought to our attention regarding the originality and authenticity of the visual data within the manuscript. Specifically, Figure 4 of the article contains images that are identical to those in the previously published papers [2, 3]. This duplication of images raises serious questions about the validity of the results and the adherence to ethical standards of research. Despite multiple attempts to contact the authors for an explanation and an opportunity to address these concerns, no satisfactory response was provided. Given the lack of accountability and the serious nature of the academic misconduct implied, the Editor-in-Chief, after careful consideration and in accordance with the publication's ethical guidelines, has decided to retract the article.

Mellein: Production in culture by Macrophomina phaseolina isolates from soybean plants exhibiting symptoms of charcoal rot and its role in pathology.

Macrophomina phaseolina (Mp) is a fungal pathogen proposed to enter host roots by releasing toxins that induce local necrosis in roots allowing entry of hyphae. Mp is reported to produce several potent phytotoxins, including (-)-botryodiplodin and phaseolinone, but isolates that do not produce these phytotoxins retain virulence. One hypothesis explaining these observations is that some Mp isolates may produce other unidentified phytotoxin(s) responsible for virulence. A previous study of Mp isolates from soybean found 14 previously unreported secondary metabolites using LC-MS/MS, including mellein, which has various reported biological activities. This study was conducted to investigate the frequency and amounts of mellein produced in culture by Mp isolates from soybean plants exhibiting symptoms of charcoal rot and to investigate the role of mellein in any observed phytotoxicity. LC-MS/MS analysis of cell-free culture filtrates (CCFs) from 89 Mp isolates revealed that 28.1% produced mellein (49-2,203 µg/L). In soybean seedlings in hydroponic culture, Mp CCFs diluted to 25% (vol/vol) in hydroponic growth medium induced phytotoxic symptoms with frequencies of 73% chlorosis, 78% necrosis, 7% wilting, and 16% death, and at 50% (vol/vol) induced phytotoxicity with frequencies of 61% chlorosis, 82% necrosis, 9% wilting, and 26% death. Commercially-available mellein (40-100 µg/mL) in hydroponic culture medium induced wilting. However, mellein concentrations in CCFs exhibited only weak, negative, insignificant correlations with phytotoxicity measures in soybean seedlings, suggesting that mellein does not contribute substantially to observed phytotoxic effects. Further investigation is needed to determine if mellein plays any role in root infection.

Neuroprotection against Aluminum Chloride-Induced Hippocampus Damage in Albino Wistar Rats by Leucophyllum frutescens (Berl.) I.M. Johnst. Leaf Extracts: A Detailed Insight into Phytochemical Analysis and Antioxidant and Enzyme Inhibition Assays.

BACKGROUND: A previously unstudied medicinal plant, Leucophyllum frutescens (Berland.) I.M. Johnst. (Scrophulariaceae) was investigated to evaluate its potential in preventing and treating neurodegenerative diseases, including Alzheimer's disease. METHODS: Methanolic leaf extract (MELE) and its fractions (HELE, CHLE, and BULE) were evaluated for their polyphenolic content and antioxidant activity by five different methods, including in vitro enzyme inhibition assays, which are clinically linked to neurodegenerative diseases. The potentially active n-butanol fraction (BULE) was further evaluated for its neuroprotective effects using an albino rat animal model and phytoconstituents profiling using Liquid chromatography with tandem mass spectrometry (LC-MS/MS), and in silico molecular docking by Maestro® Schrödinger. RESULTS: The n-butanol fraction (BULE) in the hydroalcoholic leaf extract exhibited the highest total phenolic content (230.435 ± 1.575 mg gallic acid equivalent gm-1± SD). The chloroform leaf extract exhibited the highest total flavonoid content (293.343 ± 3.756 mg quercetin equivalent gm-1± SD) as well as the highest antioxidant content, which was equivalent to Trolox, with five assay methods. Similarly, the chloroform and n-butanol fractions from the hydroalcoholic leaf extract significantly inhibited human acetylcholinesterase and butyrylcholinesterase with their IC50 values of 12.14 ± 0.85 and 129.73 ± 1.14 µg∙mL-1, respectively. The in vivo study revealed that BULE exhibited a significant neuroprotective effect at doses of 200 and 400 mg/kg/day in an aluminum chloride-induced neurodegenerative albino rat model. The LC-MS/MS analysis of BULE tentatively confirmed the presence of biologically active secondary metabolites, such as theobromine, propyl gallate, quercetin-3-O-glucoside, myricetin-3-acetylrhamnoside, isoquercitrin-6'-O-malonate, diosmetin-7-O-glucuronide-3'-O-pentose, pinoresinol diglucoside, asarinin, eridictoyl, epigallocatechin, methyl gallate derivative, and eudesmin. The results from the computational molecular docking of the identified secondary metabolites revealed that diosmetin-7-O-glucuronide-3'-O-pentose had the highest binding affinity to human butyrylcholinesterase, while isoquercetin-6'-O-malonate had the highest to human acetylcholinesterase, and pinoresinol diglucoside to human salivary alpha-amylase. CONCLUSIONS: The present study concluded a need for further exploration into this medicinal plant, including the isolation of the bioactive compounds responsible for its neuroprotective effects.

Pigment Produced by Glycine-Stimulated Macrophomina Phaseolina Is a (-)-Botryodiplodin Reaction Product and the Basis for an In-Culture Assay for (-)-Botryodiplodin Production.

An isolate of Macrophomina phaseolina from muskmelons (Cucumis melo)was reported by Dunlap and Bruton to produce red pigment(s) in melons and in culture in the presence of added glycine, alanine, leucine, or asparagine in the medium, but not with some other amino acids and nitrogen-containing compounds. We explored the generality and mechanism of this pigment production response using pathogenic M. phaseolina isolates from soybean plants expressing symptoms of charcoal rot disease. A survey of 42 M. phaseolina isolates growing on Czapek-Dox agar medium supplemented with glycine confirmed pigment production by 71% of isolates at the optimal glycine concentration (10 g/L). Studies in this laboratory have demonstrated that some pathogenic isolates of M. phaseolina produce the mycotoxin (-)-botryodiplodin, which has been reported to react with amino acids, proteins, and other amines to produce red pigments. Time course studies showed a significant positive correlation between pigment and (-)-botryodiplodin production by selected M. phaseolina isolates with maximum production at seven to eight days. Pigments produced in agar culture medium supplemented with glycine, beta-alanine, or other amines exhibited similar UV-vis adsorption spectra as did pigments produced by (±)-botryodiplodin reacting in the same agar medium. In a separate study of 39 M. phaseolina isolates, red pigment production (OD520) on 10 g/L glycine-supplemented Czapek-Dox agar medium correlated significantly with (-)-botryodiplodin production (LC/MS analysis of culture filtrates) in parallel cultures on un-supplemented medium. These results support pigment production on glycine-supplemented agar medium as a simple and inexpensive in-culture method for detecting (-)-botryodiplodin production by M. phaseolina isolates.

Diversity and ethnomycological importance of mushrooms from Western Himalayas, Kashmir.

BACKGROUND: Wild edible mushrooms (WEM) are economically significant and used in traditional medicines worldwide. The region of Jammu and Kashmir (Western Himalayas) is enriched with the diversity of edible mushrooms, collected by the rural people for food and income generation. This is the first detailed study on diversity and ethno-medicinal uses of mushrooms from the State of Jammu and Kashmir. METHODS: Consecutive surveys were conducted to record ethnomycological diversity and socio-economic importance of wild edible mushrooms value chain in rural areas of Azad Jammu and Kashmir during 2015-2019. Ethnomycological data were collected with a semi-structured questionnaire having a set of questions on indigenous mycological knowledge and collection and retailing of wild edible mushrooms. A total of 923 informants from the study area provided the results identifying the gender, type of mushroom species, medicinal uses, and marketing of mushrooms. Diversity of mushrooms was studied by using quadrat and transect methods. Principal component analysis (PCA) and detrended correspondence analysis (DCA) were also applied to the dataset to analyse the relationship between species distribution, the underlying environmental factors, and habitat types. PCA identified the major species-specific to the sites and put them close to the sites of distribution. RESULTS: A total of 131 mushroom species were collected and identified during 2015-2019 from the study area. Ninety-seven species of mushrooms were reported new to the State of Azad Jammu and Kashmir. The dominant mushroom family was Russulaceae with 23 species followed by Agaricaceae, 16 species. Major mushroom species identified and grouped by the PCA were Coprinus comatus, Lactarius sanguifluus, Amanita fulva, Armillaria gallica, Lycoperdon perlatum, Lycoperdon pyriforme, and Russula creminicolor. Sparassis crispa, Pleurotus sp, and Laetiporus sulphureus were recorded most edible and medicinally significant fungi. Morels were also expensive and medicinally important among all harvested macro-fungal species. These were reported to use against common ailments and various health problems. CONCLUSIONS: Collection and retailing of WEM contribute to improving the socio-economic status, providing alternative employment and food security to rural people of the area. These mushrooms are used as a source of food and traditional medicines among the rural informants and could be used as a potential source of antibacterial and anticancer drugs in the future.

Dendrimeric alkylated polyethylenimine nano-carriers with acid-cleavable outer cationic shells mediate improved transfection efficiency without increasing toxicity.

PURPOSE: Improved polycation-based non-viral DNA vectors were sought by preparing dendrimers with polyethylenimine cores surrounded by various shells incorporating structural features intended to facilitate steps in transfection mechanisms. Dendrimeric vectors were designed with (a) an outer oligocation shell, intended to facilitate DNA release inside cells, (b) a hydrophobic C-16 alkyl shell, and (c) a polycationic core, the latter two intended to combine lipid-depletion and osmotic burst endosome escape mechanisms, respectively, and were (d) attached through an a acid-cleavable linker reported to hydrolyze at endosomal pH values. METHODS: Vectors and DNA complexes were characterized by dynamic and static light scattering. Flow cytometry was used to quantitate transfection activity and cytotoxicity in CHO-K1 cells. RESULTS: About 5-fold increased transfection activity was obtained for a vector constructed with an outer shell of oligocations attached through acid-cleavable linkers, relative to a control dendrimer with an acid-stable linker. The most effective oligocation component of outer shells tested was spermine. Neither modification was associated with increased cytotoxicity. This vector design did not permit an evaluation of the benefit of combining endosome release mechanisms. CONCLUSION: Using acid-cleavable linkers to attach an outer shell of short, highly-charged oligocations to a PEI-based dendrimeric vector substantially increased transfection efficiency without increasing cytotoxicity.

Megalin-targeted enhanced transfection efficiency in cultured human HK-2 renal tubular proximal cells using aminoglycoside-carboxyalkyl- polyethylenimine -containing nanoplexes.

Non-viral vectors are of interest as therapeutic gene delivery agents in gene therapy, because they are simple to prepare, easy to modify and have definable safety profiles compared to viral vectors. The potential of gene therapy in the treatment of renal diseases is limited by a lack of effective kidney-targeted gene delivery systems. Aminoglycoside antibiotics gentamicin and neomycin were connected by amide linkages to carboxyl groups on carboxyalkylated-PEI25 (25kDa PEI) or carboxyalkylated-PEI10 (10kDa PEI). Aminoglycoside-carboxyalkylated-PEI conjugates were characterized with respect to size, surface charge density, DNA condensation ability, and buffering capacity. Polyplexes prepared by electrostatic interaction between aminoglycoside-carboxyalkylated-PEIs and enhanced green fluorescent protein-expressing (EGFP) plasmid DNA had appropriate nano-scale size (143-173nm). Their targeting potential was investigated in cultured HK-2 immortalized human cortex/proximal tubule kidney epithelial cells, which expresses megalin, a scavenger receptor that mediates endocytosis of a diverse group of ligands, including aminoglycoside antibiotics. Aminoglycoside-carboxyalkylated-PEIs significantly increased EGFP gene transfection efficiency in HK-2 cells by ∼13-fold for aminoglycoside-carboxyalkylated-PEI25 and ∼7-fold increase for aminoglycoside-carboxyalkylated-PEI10 relative to the corresponding PEIs without aminoglycosides. The transfection efficiency of polyplexes was dependent on the weight ratio of aminoglycoside-containing ligand in the carrier. In the presence of a range of concentrations of human serum albumin, which competes for megalin binding, aminoglycoside-carboxyalkylated-PEI-mediated transfection was reduced to background levels. These results suggest that aminoglycoside-carboxyalkylated-PEI polyplexes can target megalin-expressing kidney-derived cells in vitro resulting in improved transfection efficiency with low cytotoxicity.

Efficient megalin targeted delivery to renal proximal tubular cells mediated by modified-polymyxin B-polyethylenimine based nano-gene-carriers.

Non-viral vectors have attracted great interest, as they are simple to prepare, easy to modify and relatively safe, compared to viral vectors. Kidney-targeted gene delivery systems depict a promising technology to improve drug efficacy in renal diseases treatments. In order to develop a novel kidney-targeted gene delivery system, we synthesized polyamine-PEI conjugates using polymyxin B as ligand and investigated their potential targeting efficiency. After grafting either PEI25 kDa or PEI10 kDa with polymyxin B through amidation reaction, the modified-polymyxin-PEI/DNA-nanoplexes were produced via electrostatic attraction between the cationic polymers and EGFP plasmid. The properties of modified polymers including size, surface charge density, DNA condensation ability, buffering capacity and cytotoxicity were evaluated. Results revealed that the average size of -modified-polymyxin- PEI25kDa was about 143-180nm and modified-polymyxin-PEI10kDa 115-194nm. The zeta potentials were in the range of 16.4±1.87 to 23.43±1.25mV and 11.3±1.4 to 19.3±2.1mV for conjugates based on PEI25 and PEI10 respectively. The AFM images revealed that the complexes were spherical and nano-sized at C/P=4. The buffering capacity of both PEI 10 and 25kDa increased as the percentage of polymyxin B grafting increased. In vitro study demonstrated that modified-polymyxin-PEI conjugates could remarkably improve the gene transfection efficiency to kidney cells. The transfection efficiency of the polyplexes was dependent on the weight ratio of ligand in the formulation (~12 and 8 fold increase for PEI25 and PEI10kDa, respectively) and was significantly higher than that of unmodified PEIs/DNA nanoparticles. These results suggest that modified-polymyxin-PEI /DNA nanoparticles can effectively target megalin-expressing kidney cells and show improved transfection efficiency and low cytotoxicity in In vitro and In vivo studies. Animal studies confirmed the in vivo study. Thus, these conjugates can be considered as a safe and efficient non-viral therapeutic therapy vector for kidney diseases.

Heterocyclic amine-modified polyethylenimine as gene carriers for transfection of mammalian cells.

Branched polyethylenimine (PEI) is extensively used as a polycationic non-viral vector for gene delivery. Polyplexes formed with PEI are believed to be released from endocytotic vesicles by the osmotic burst mechanism in the rate-limiting step in transfection. Increasing the buffering capacity of PEI derivatives in the endosomal pH range of 4.5-7.5 should enhance transfection efficiency. In this study, PEI was derivatized by covalently attaching heterocyclic amine moieties (piperazine, pyridine and imidazole rings with pKa values from 5.23 to 6.04) through amide bonds. PEI derivatives with 50% of the primary amines on PEI exhibited increased buffering capacity, increased transfection activity and decreased cytotoxicity in murine neuroblastoma (Neuro-2a) cells. The relative effectiveness in enhancing transfection efficiency was piperazine>pyridine>histidine, but each type of amine was the most effective under a particular set of conditions. Modified vectors could significantly improve transfection efficiency in murine mesenchymal stem cells. PEI25 derivatized at 50% with histidine administered as polyplexes in the tail veins of mice resulted in remarkably enhanced luciferase gene expression in the expected organ distribution and much lower toxicity than underivatized PEI25.

From rationally designed polymeric and peptidic systems to sophisticated gene delivery nano-vectors.

Lack of safe, efficient and controllable methods for delivering therapeutic genes appears to be the most important factor preventing human gene therapy. Safety issues encountered with viral vectors have prompted substantial attention to in vivo investigations with non-viral vectors throughout the past decade. However, developing non-viral vectors with effectiveness comparable to viral ones has been a challenge. The strategy of designing multifunctional synthetic carriers targeting several extracellular and intracellular barriers in the gene transfer pathway has emerged as a promising approach to improving the efficacy of gene delivery systems. This review will explain how sophisticated synthetic vectors can be created by combining conventional polycationic vectors such as polyethylenimine and basic amino acid peptides with additional polymers and peptides that are designed to overcome potential barriers to the gene delivery process.

Molecular weight-dependent genetic information transfer with disulfide-linked polyethylenimine-based nonviral vectors.

One strategy for improving gene vector properties of polyethylenimine is to facilitate individual transfection mechanism steps. This study investigates (i) improving transfection efficiency by attaching peptide nuclear localization signals (nuclear localization signals: SV40 large T antigen nuclear localization signal or C-terminus of histone H1) to polyethylenimine (10 kDa) and (ii) using disulfide linkages, which are expected to be stable during polyplex formation, but cleaved inside cells giving improved gene release. Nuclear localization signal-containing polyplexes exhibited low cytotoxicity, whereas transfection efficiency with high molecular weight plasmid DNA increased up to 3.6 times that of underivatized polyethylenimine in Neuro2A cells at higher molar ratio of polyethylenimine-nitrogen to DNA-phosphate (N/P) ratios. However, with luciferase-specific low molecular weight small interfering RNA in Neuro2A/EGFPLuc cells, nuclear localization signal-containing polyplexes with disulfide linkages caused substantial cytotoxicity at N/P ratios >15 and no consistent significant reduction in luciferase expression. Possible explanations for molecular weight-dependent differences in genetic information transfer by polyplexes containing disulfide-linked nuclear localization signals are discussed.

Arginine-rich hydrophobic polyethylenimine: potent agent with simple components for nucleic acid delivery.

Conjugation of various arginine-rich peptide sequences to vectors based on 10 kDa polyethylenimine (PEI) and its hydrophobic derivative (hexanoate-PEI) was investigated as a strategy for improving pDNA and siRNA transfection activities. Six different arginine-histidine (RH) sequences and two arginine-serine (RS) sequences with a range of R/H ratios were designed and coupled to PEI and hexanoate-PEI. All arginine-rich peptide derivatives of PEI significantly enhanced luciferase gene expression compared to PEI 10 kDa alone. Hexanoate-PEI derivatives exhibited higher transfection activity than underivatized PEI vectors. Improved transfection activity may have resulted at least in part from use of higher vector/DNA ratios made possible by reduced cytotoxicity of vectors, and to use of vectors with higher molecular weights. Vectors that were the most efficient in pDNA delivery and transfection were also the most effective in siRNA delivery and protein expression knock down.