Vol 27, No 4 (2024)
- Year: 2024
- Articles: 14
- URL: https://rjraap.com/1386-2073/issue/view/10052
Chemistry
Bridging Interregional Cooperation in Bioprospection
497-498
In Memoriam - Professor Mahabir Gupta 1942-2020
499-501
Design and Diversity Analysis of Chemical Libraries in Drug Discovery
Abstract
Chemical libraries and compound data sets are among the main inputs to start the drug discovery process at universities, research institutes, and the pharmaceutical industry. The approach used in the design of compound libraries, the chemical information they possess, and the representation of structures, play a fundamental role in the development of studies: chemoinformatics, food informatics, in silico pharmacokinetics, computational toxicology, bioinformatics, and molecular modeling to generate computational hits that will continue the optimization process of drug candidates. The prospects for growth in drug discovery and development processes in chemical, biotechnological, and pharmaceutical companies began a few years ago by integrating computational tools with artificial intelligence methodologies. It is anticipated that it will increase the number of drugs approved by regulatory agencies shortly.
502-515
Bioactivity of Raphanus Species against Agricultural Phytopathogens and its Role in Soil Remediation: A Review
Abstract
Phytopathogens and weeds represent around 20-40% of global agricultural productivity losses. Synthetic pesticide products are the most used to combat these pests, but it reiterates that their use has caused tremendous pressure on ecosystems' self-cleansing capacity and resistance development by pathogens to synthetic fungicides. In the last decades, researchers have demonstrated the vast biological properties of plants against pathogens and diseases. Raphanus species (Brassicaceae) possesses antimicrobial, antioxidant, anti-inflammatory, anticancer, hepatoprotective, antidiabetic, insecticidal, nematicidal, allelopathic, and phytoremediators properties. These are due to the presence of structurally diverse bioactive compounds, such as flavonoids and glucosinolates. In this review, we have provided an update on the biological properties of two Raphanus species (R. sativus and R. raphanistrum), detailing the type of natural product (extract or isolated compound), the bioassays displayed, and the results obtained for the main bioactivities of this genus cited in the literature during the last 30 years. Moreover, preliminary studies on phytopathogenic activities performed in our laboratory have also been depicted. We conclude that Raphanus species could be a source of natural bioactive molecules to treat phytopathogens and weeds that affect crops and remediate contaminated soils.
516-544
Understanding the Artemia Salina (Brine Shrimp) Test: Pharmacological Significance and Global Impact
Abstract
Background:The microplate benchtop brine shrimp test (BST) has been widely used for screening and bio-guided isolation of many active compounds, including natural products. Although the interpretation given to the results appears dissimilar, our findings suggest a correlation between positive results with a specific mechanism of action.
Objective:This study aimed to evaluate drugs belonging to fifteen pharmacological categories having diverse mechanisms of action and carry out a bibliometric analysis of over 700 citations related to microwell BST.
Methods:Test compounds were evaluated in a serial dilution on the microwell BST using healthy nauplii of Artemia salina and after 24 hrs of exposition, the number of alive and dead nauplii was determined, and the LC50 was estimated. A metric study regarding the citations of the BST miniaturized method, sorted by type of documents cited, contributing country, and interpretation of results was conducted on 706 selected citations found in Google Scholar.
Results:Out of 206 drugs tested belonging to fifteen pharmacological categories, twenty-six showed LC50 valuep <000 µM, most of them belonging to the category of antineoplastic drugs; compounds with different therapeutical uses were found to be cytotoxic as well. A bibliometric analysis showed 706 documents citing the miniaturized BST; 78% of them belonged to academic laboratories from developing countries located on all continents, 63% interpreted their results as cytotoxic activity and 35% indicated general toxicity assessment.
Conclusion:BST is a simple, affordable, benchtop assay, capable of detecting cytotoxic drugs with specific mechanisms of action, such as protein synthesis inhibition, antimitotic, DNA binding, topoisomerase I inhibitors, and caspases cascade interfering drugs. The microwell BST is a technique that is used worldwide for the bio-guided isolation of cytotoxic compounds from different sources.
545-554
Yield and Chemical Characteristics of Salvia hispanica L. (Chia) Oil from Native Seeds from Four Provenances of Guatemala
Abstract
Introduction:Chia (Salvia hispanica L.) is a plant from the Lamiaceae family that has been used as ancestral food, medicine, and oil, with culinary, artistic, and religious purposes by most of the Mesoamerican civilizations. Native from Mesoamerica, introduced into South America, Australia, and Europe, it is presently consumed as a nutritional and functional food.
Objective:This research aims to characterize ancient native cultivars from four provenances in Guatemala to recommend their direct consumption by the population as well as to establish its trade.
Methods:Seed samples were collected in four places where they have been cultivated for several generations. The oil was obtained by expression and analyzed chemically by gas chromatography following standard qualitative and quantitative methods.
Results:Variations in oil yield and some of the characteristic parameters of the phytochemical analysis were obtained. In general, the profile was similar to most of the reported data in the literature, with the saturated fatty acids (8.54-9.25%) relatively lower than the references (7.95-11.45%) but a higher concentration of unsaturated fatty oils, particularly of omega-3 (64.68-68.62%).
Conclusion:The oil from native cultivars contains high quantities of omega-3, which might help pregnant women during gestation and to control other conditions such as metabolic syndrome, particularly in low- and middle-income populations where these seeds are consumed regularly. The suggestion is made to encourage the cultivation and use of these ancestral seeds with the possibility of commercialization abroad with an appellation of origin label.
555-561
In silico Molecular Docking Analysis of Three Molecules Isolated from Litsea guatemalensis Mez on Anti-inflammatory Receptors
Abstract
Background:The Litsea genus has four native species from Mesoamerica. Litsea guatemalensis Mez. is a native tree, traditionally used as a condiment and herbal medicine in the region. It has demonstrated antimicrobial, aromatic, anti-inflammatory and antioxidant activity. Bioactive fractionation attributed the anti-inflammatory and anti-hyperalgesic activities to pinocembrin, scopoletin, and 5,7,3´4´-tetrahydroxy-isoflavone. In silico analysis, these molecules were analyzed on receptors involved in the anti-inflammatory process to determine which pathways they interact.
Objective:To analyze and evaluate 5,7,3',4'-tetrahydroxyisoflavone, pinocembrin, and scopoletin using the in silico analysis against selected receptors involved in the inflammatory pathway.
Method:Known receptors involved in the anti-inflammatory process found as protein-ligand complex in the Protein Data Bank (PDB) were used as references for each receptor and compared with the molecules of interest. The GOLD-ChemScore function, provided by the software, was used to rank the complexes and visually inspect the overlap between the reference ligand and the poses of the studied metabolites.
Results:53 proteins were evaluated, each one in five conformations minimized by molecular dynamics. The scores obtained for dihydroorotate dehydrogenase were greater than 80 for the three molecules of interest, scores for cyclooxygenase 1 and glucocorticoid receptor were greater than 50, and identified residues with interaction in binding sites overlap with the reference ligands in these receptors.
Conclusion:The three molecules involved in the anti-inflammatory process of L. guatemalensis show in silico high affinity to the enzyme dihydroorotate dehydrogenase, glucocorticoid receptors and cyclooxygenase-1.
562-572
One-step Bio-guided Isolation of Secondary Metabolites from the Endophytic Fungus Penicillium crustosum Using High-resolution Semi-preparative HPLC
Abstract
Background:An endophytic fungal strain Penicillium crustosum was isolated from the seagrass Posidonia oceanica and investigated to identify its antimicrobial constituents and characterize its metabolome composition. The ethyl acetate extract of this fungus exhibited antimicrobial activity against methicillin-resistant Staphylococcus aureus (MRSA) as well as an anti-quorum sensing effect against Pseudomonas aeruginosa.
Methods:The crude extract was profiled by UHPLC-HRMS/MS, and the dereplication was assisted by feature-based molecular networking. As a result, more than twenty compounds were annotated in this fungus. To rapidly identify the active compounds, the enriched extract was fractionated by semipreparative HPLC-UV applying a chromatographic gradient transfer and dry load sample introduction to maximise resolution. The collected fractions were profiled by 1H-NMR and UHPLC-HRMS.
Results:The use of molecular networking-assisted UHPLC-HRMS/MS dereplication allowed preliminary identification of over 20 compounds present in the ethyl acetate extract of P. crustosum. The chromatographic approach significantly accelerated the isolation of the majority of compounds present in the active extract. The one-step fractionation allowed the isolation and identification of eight compounds (1-8).
Conclusion:This study led to the unambiguous identification of eight known secondary metabolites as well as the determination of their antibacterial properties.
573-583
HILIC UPLC/ QTof MS Method Development for the Quantification of AGEs Inhibitors - Trouble Shooting Protocol
Abstract
Objective:The paper reports an attempt to develop and validate a HILIC UPLC/ QTof MS method for quantifying N-ε-carboxymethyl-L-lysine (CML) in vitro, testing N-ε- carboxy[D2]methyl-L-lysine (d2-CML), and N-ε-carboxy[4,4,5,5-D4]methyl-L-lysine (d4-CML) as internal standards.
Method:During the method development, several challenging questions occurred that hindered the successful completion of the method. The study emphasizes the impact of issues, generally overlooked in the development of similar analytical protocols. For instance, the use of glassware and plasticware was critical for the accurate quantification of CML. Moreover, the origin of atypical variation in the response of the deuterated internal standards, though widely used in other experimental procedures, was investigated.
Result:A narrative description of the systematic approach used to address the various drawbacks during the analytical method development and validation is presented.
Conclusion:Reporting those findings can be considered beneficial while bringing an insightful notion about critical factors and potential interferences. Therefore, some conclusion and ideas can be drawn from these trouble-shooting questions, which might help other researchers to develop more reliable bioanalytical methods, or to raise their awareness of stumbling blocks along the way.
584-598
Identification and Validation of Synapse-related Hub Genes after Spinal Cord Injury by Bioinformatics Analysis
Abstract
Background:Spinal cord injury (SCI) is a neurological disease with high morbidity and mortality. Previous studies have shown that abnormally expressed synapse-related genes are closely related to the occurrence and development of SCI. However, little is known about the interaction of these aberrantly expressed genes and the molecular mechanisms that play a role in the injury response. Therefore, deeply exploring the correlation between synapse-related genes and functional recovery after spinal cord injury and the molecular regulation mechanism is of great significance.
Methods:First, we selected the function GSE45006 dataset to construct three clinically meaningful gene modules by hierarchical clustering analysis in 4 normal samples and 20 SCI samples. Subsequently, we performed functional and pathway enrichment analyses of key modules.
Results:The results showed that related module genes were significantly enriched in synaptic structures and functions, such as the regulation of synaptic membranes and membrane potential. A protein-protein interaction network (PPI) was constructed to identify 10 hub genes of SCI, and the results showed that Snap25, Cplx1, Stxbp1, Syt1, Rims1, Rab3a, Syn2, Syn1, Cask, Lin7b were most associated with SCI. Finally, these hub genes were further verified by quantitative real-time fluorescence polymerase chain reaction (qRT-PCR) in the spinal cord tissues of the blank group and SCI rats, and it was found that the expression of these hub genes was significantly decreased in the spinal cord injury compared with the blank group (P ≤ 0.05).
Conclusion:These results suggest that the structure and function of synapses play an important role after spinal cord injury. Our study helps to understand the underlying pathogenesis of SCI patients further and identify new targets for SCI treatment.
599-610
Structural Maintenance of Chromosome Protein 4 Promotes the Progression of Cardia Adenocarcinoma via Regulation of the Wnt/β-catenin Signaling Pathway
Abstract
Background:Structural maintenance of chromosome protein 4 (SMC4) is crucial for chromosome assembly and separation, but its role and mechanism in cardia adenocarcinoma (CA) are unknown.
Methods:SMC4 expression levels were initially detected by protein profiling in 20 pairs of CA tumor tissues and adjacent normal tissues. The level of SMC4 expression in CA cells was then evaluated using a western blot analysis. Cell proliferation was evaluated by CCK-8 and clone formation tests. Scratch and transwell tests were used to investigate cell migration as well as invasion, while through the flow cytometry, we examined the cell apoptosis and progression of the cell cycle. The regulatory effects of the epithelial-mesenchymal transition (EMT) and the Wnt/β- catenin pathway were investigated using western blot. A tumorigenesis experiment was used to investigate the influence of SMC4 on tumor development in nude mice.
Results:This study showed overexpression of SMC4 in CA tissues and cells. Knockdown of SMC4 can significantly inhibit the proliferation, migration and invasion, stimulate cell apoptosis, induce cell cycle arrest in the G0/G1 phase of CA cells, and inhibit tumor growth in vivo. In addition, down-regulation of SMC4 resulted in decreased expression of Bcl-2, Cyclin D1, CDK4, CDK6, β-catenin, phosphorylated GSK-3β, N-cadherin, and Vimentin, with an increased level of proteins, i.e., Bax, cleaved-caspase3, and E-cadherin. When SMC4 was overexpressed, these effects were reversed.
Conclusion:SMC4 can facilitate the biological progression of CA, suggesting that SMC4 could be a potential therapeutic target for the disease.
611-620
New Tiaoxin Recipe Alleviates Energy Metabolism Disorders in an APPswe/PS1DE9 Mouse Model of Alzheimers Disease
Abstract
Background:Alzheimer's disease (AD) is a typical neurodegenerative disease with a complex etiology. Until now, there has been no effective treatment available for AD; however, improving energy dysmetabolism, the key pathological event in the early stage of AD, can effectively delay the progression of AD.
Objective:This paper aims to investigate the therapeutic effect and potential mechanism of the new Tiaoxin recipe on early AD.
Methods:APP/PS1 mice were divided into a model group, a new Tiaoxin recipe group, and a donepezil group, and C57/BL mice were used for the control group. Mouse cognitive and learning abilities were tested using the Morris water maze test and a new object-recognition experiment. The 42 amino acid form of amyloid β peptide (Aβ142) content was detected by enzyme-linked immunosorbent assay, the senile plaque area was detected by thioflavin S staining, and the senescence- associated β-galactosidase (SA-β-gal)positive area was detected by chemical staining. Also, the adenosine triphosphate (ATP), nicotinamide adenine dinucleotide (NAD+), and nicotinamide adenine dinucleotide hydride (NADH) contents were detected using a biochemical method, and the cluster of differentiation 38 (CD38) and silent matingtype information regulation 2 homolog 3 (SIRT3) protein expression levels were detected by immunofluorescence and Western blot analysis.
Results:Compared with those of the control group, the learning and memory abilities of the model group were impaired; the senile plaque deposition, Aβ142 content, and SA-βgalpositive staining area were increased; the ATP concentration, NAD+ concentration, and NAD+/NADH ratio were decreased; the CD38 protein expression level was increased; and the SIRT3 protein expression level was decreased. Following intervention with the new Tiaoxin recipe, the learning and memory abilities were improved; the senile plaque deposition, Aβ142 content, and SA-βgalpositive area were reduced; the ATP concentration, NAD+ concentration, and NAD+/NADH ratio were increased; CD38 protein expression was decreased, and SIRT3 protein expression was increased.
Conclusion:This study shows that the new Tiaoxin Recipe can improve cognitive ability and reduce the Aβ1-42 content and senile plaque deposition in APP/PS1 mice, which may occur through the downregulation of CD38 protein expression, upregulation of SIRT3 protein expression, restoration of the NAD+ level, promotion of ATP synthesis, mitigation of energy metabolism disorders.
621-631
Key Mutant Genes and Biological Pathways Involved in Aspirin Resistance in the Residents of the Chinese Plateau Area
Abstract
Introduction:Aspirin is used to prevent and treat cardiovascular diseases; however, some patients develop aspirin resistance.
Aim:We aimed to explore the potential molecular mechanisms underlying aspirin resistance in people living in the Chinese plateau area.
Methods:In total, 91 participants receiving aspirin treatment from the Qinghai plateau area were divided into the aspirin resistance and aspirin sensitivity groups. Genotyping was performed using the Sequence MASSarray. Differentially mutated genes between the two groups were analyzed using MAfTools. The annotation of differentially mutated genes was conducted based on the Metascape database.
Results and Discussion:In total, 48 differential SNP and 22 differential InDel mutant genes between the aspirin resistance and aspirin sensitivity groups were screened using Fishers exact test (p < 0.05). After the χ2 test, a total of 21 SNP mutant genes, including ZFPL1 and TLR3, and 19 InDel mutant genes were found to be differentially expressed between the two groups (p < 0.05). Functional analysis revealed that these differential SNP mutations were mainly enriched in aspirin resistance pathways, such as the Wnt signaling pathway. Furthermore, these genes were related to many diseases, including various aspirin indications.
Conclusion:This study identified several genes and pathways that could be involved in arachidonic acid metabolic processes and aspirin resistance progression, which will provide a theoretical understanding of the molecular mechanism of aspirin resistance.
632-640
miR-129-5p/FGF2 Axis is Associated with Homocysteine-induced Human Umbilical Vein Endothelial Cell Injury
Abstract
Purpose:Homocysteine (Hcy)-induced endothelial cell injury is a key event in atherosclerosis pathogenesis. In this study, we aimed to explore the mechanisms underlying Hcy-induced endothelial injury by assessing the effects of Hcy on endothelial cell proliferation and the microRNA (miR)-129-5p/fibroblast growth factor 2 (FGF2) axis.
Methods:Human umbilical vein endothelial cells (HUVECs) were treated with Hcy to construct an endothelial cell injury model. Expression levels of FGF2 in Hcy-induced HUVECs were determined using quantitative real-time polymerase chain reaction and western blotting. An FGF2 overexpression lentiviral vector was constructed to upregulate FGF2 expression in HUVECs via lentivirus transduction. A cell counting kit-8 assay was used to explore the effects of FGF2 overexpression on HUVEC proliferation. An upstream regulatory miRNA was predicted, and its targetbinding relationship with FGF2 was evaluated using a dual-luciferase reporter assay.
Results:We found that FGF2 expression in HUVECs was inhibited by Hcy treatment. Lentivirus transduction led to the overexpression of FGF2 in HUVECs, which significantly reversed the effect of Hcy on endothelial cell proliferation. miR-129-5p was experimentally validated as an upstream regulator of FGF2, and its decreased levels in HUVECs led to increased FGF2 expression. In addition, HUVEC proliferation was enhanced by the knockdown of miR-129-5p, and this effect was reversed by Hcy treatment.
Conclusion:Taken together, the results of this study revealed that Hcy inhibits FGF2 expression in HUVECs, and FGF2 is regulated by upstream miR-129-5p to improve the effect of Hcy on endothelial cell proliferation.
641-648