Applied Biochemistry and Biotechnology最新文献

Osteoporosis (OP) is a genetic disorder characterized by an imbalance between osteoblast-mediated bone formation and osteoclast-induced bone resorption. However, the underlying gene-related mechanisms of its pathogenesis remain to be fully elucidated. Aberrantly expressed neuralized E3 ubiquitin-protein ligase 3 (NEURL3), which is related to osteoclastic differentiation, was identified through the analysis of the microarray profile GSE176265. Bone marrow-derived macrophages (BMMs) were isolated from the femurs and tibias of C57BL/6 J mice and treated with 30 ng/mL macrophage-colony-stimulating factor (M-CSF) and 100 ng/mL receptor activator of nuclear factor-kappa B ligand (RANKL) to induce osteoclastic differentiation, thereby mimicking OP in vitro. To model OP in vivo, ovariectomy (OVX)-induced bone loss was performed in mice. High expression levels of NEURL3 were confirmed in clinical samples, OP model cells, and OP model mice using quantitative real-time polymerase chain reaction (qRT-PCR). The impact of NEURL3 on osteoclastic differentiation was assessed by evaluating cell viability and the expression levels of osteoclastogenesis-related marker genes. Additionally, bone loss in mice was quantified using micro-computed tomography before and after NEURL3 inhibition. Mechanistically, the effects of NEURL3 on osteogenic differentiation were investigated by determining the protein levels of osteogenic markers via Western blotting. NEURL3 was markedly overexpressed in serum samples collected from patients with OP, OVX-induced OP mouse models, and induced osteoclasts. Inhibition of NEURL3 leads to a 20% decrease in BMM survival rate and a reduction in the number of tartrate-resistant acid phosphatase (TRAP) positive cells, which is a characteristic of mature osteoclasts. Furthermore, the expression levels of osteoclastogenesis-related marker genes were reduced by 50%. In vivo studies revealed that suppressing NEURL3 resulted in a 38% improvement in trabecular bone volume (BV/TV) and a 28% increase in bone mineral density (BMD) in the OVX-induced OP mice. Mechanistically, NEURL3 promoted osteoclast differentiation by increasing the ubiquitination levels of BMP7. Inhibition of BMP7 reversed the effects of NEURL3 on osteoclast differentiation in BMMs. Suppression of NEURL3 inhibits osteoclast differentiation of BMMs in vitro and alleviates bone loss in vivo. The underlying mechanism may involve NEURL3-induced ubiquitination of BMP7. Collectively, the downregulation of NEURL3 represents a promising therapeutic strategy for suppressing osteoclast differentiation and treating OP.

The tree bean (Parkia timoriana (DC). Merr) is an underutilized legume and is abundantly found in Southeast Asia. It is valued for its nutritious pods and cultivated for food and timber. Despite of the presence of several nutrients, the regulatory networks involved in secondary metabolite biosynthesis in the tree bean remain largely unexplored. Recent studies have highlighted that consumption of its pods provides numerous health benefits, including antioxidant, α-glucosidase inhibitory, antibacterial, antidiabetic, and insecticidal activities. To elucidate the biosynthesis of specific metabolites in this plant, a comparative metabolite and transcriptomic analysis of the leaf and root tissues of P. timoriana was carried out. The study revealed that P. timoriana leaf and root tissues contain varying levels of phenolics, flavonoids, and terpenoids. ¹H nuclear magnetic resonance (¹H NMR) analysis identified 16 significant metabolites in the leaf and root tissues, including sugars, amino acids, and organic acids. L-dihydroxyphenylalanine (L-DOPA), an amino acid derivative and precursor to dopamine, was detected for the first time in the seeds. Additionally, the presence of pinitol in P. timoriana was also confirmed. De novo RNA-sequence analysis identified differentially expressed genes (DEGs) in both the tissues. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis identified pathways associated with shikimate pathway, such as phenylpropanoid and flavonoid biosynthesis. MapMan pathway analysis revealed a high number of transcripts related to phenylalanine, tryptophan, tyrosine, and condensed tannin biosynthesis. The research conducted identified secondary metabolites in P. timoriana, and their probable biosynthetic pathway which can be used for medicinal and nutritional purposes.

Enzymatic oligomerization of flavonoids enables the synthesis of biomolecules with different structures and improved physicochemical and biological properties and can therefore broaden their application in industry. In this study, the influence of the key reaction parameters temperature, solvent, substrate, and enzyme concentrations on the synthesis of esculin oligomers was investigated. The reaction was optimized using response surface methodology (RSM) in order to obtain the highest products' concentration and specific products' yield (per mass of enzyme). Mass spectrometry revealed that oligomers with a degree of polymerization of up to 4 were synthesized in which dimers were the most abundant, while the NMR analysis of the esculin dimer product showed that C8-C8 link between two esculin units was formed. Maximum products' concentration was obtained at 60 °C, in 14% (v/v) methanol, 7 mg/mL of esculin, and 54.6 U/L of laccase after 7 h, while the optimal conditions for specific products' yield differed in the aspect of optimal laccase concentration which was 19 U/L for this output. Synthesized esculin oligomers exhibited higher iron chelating and cupric reducing antioxidant capacities and similar or even superior free radical scavenging activity compared to monomeric esculin. Moreover, the mixture of synthesized esculin oligomers has shown a promising potential to be used as a skin prebiotics, suggesting novel applications in skincare industries.

Microplastic pollution has emerged as a new global concern because of its ubiquitous and persistent nature. Due to the rising use of plastics and discharge of plastic waste into coastal water bodies from point and non-point sources, the occurrence of microplastics along coastal ecosystems has become very prevalent. The current study is the first of its kind to evaluate the presence of microplastics in the surface water of river estuary along the coast of Odisha. Six GPS-fixed locations were used to collect the surface water samples from the Budhabalanga river estuary in Chandipur, Odisha, India. The samples were then subjected to further investigation to determine the types of microplastics present. The average microplastic abundance, according to our findings, ranged from 9.33 ± 2.11 items L^-1 to 28.50 ± 2.77 items L^-1. Microplastics come in a variety of colours and shapes, but the most prevalent kind is fibre-shaped and black in colour. The pollution load index of the sampling area was calculated to be 4.25 which is categorized under ecological risk level I. FE-SEM images clearly showed the topology of microplastics and ATR-FTIR analysis confirmed the presence of polyethylene, polypropylene, polyvinyl chloride (PVC), nylon, polycarbonate (PC), ethylene vinyl acetate (EVA) and polystyrene (PS) at sampling stations. Our investigation provides useful information that helps to reduce the ecological risk in habitats connected with contaminated sites, including both aquatic and terrestrial habitats.

Bean common mosaic virus (BCMV) is a severe plant pathogen of common bean (Phaseolus vulgaris L.), that causes huge yield losses across the globe. The virus has a wide host range and varied modes of transmission, due to which its management is challenging. Pathogen-derived resistance, which entails inserting virus-derived gene sequences into transgenic plants, is extremely effective in overcoming plant viruses. However, owing to ethical and biosecurity concerns, transgenic crops have not been widely accepted. Exogenous application of double-stranded RNA (dsRNA) is a new and intriguing method for inducing resistance against plant viruses. In this study, the efficacy of exogenous application of dsRNAs synthesized from BCMV helper component proteinase (HC-Pro) and coat protein (CP) genes were assessed in three plants: tobacco (Nicotiana tabacum), common bean (Phaseolus vulgaris L.), and cowpea (Vigna unguiculata), and both dsRNAs elicited a resistance response. dsRNA targeting the HC-Pro gene of BCMV was found more effective in inducing RNAi-based resistance than dsRNA targeting the CP gene of BCMV with a more pronounced effect in cowpea than tobacco and common bean. We showed the stability and transport of both dsHC-Pro and dsCP in inoculated to non-inoculated young leaves. We also showed the ability of mesoporous silica nanoparticles (MSP) conjugated with dsHC-Pro to provide prolonged stability and broader resistance against BCMV in common bean, extending protection from 12 dpi up to 20 dpi, compared to naked dsHC-Pro. Our results suggest that dsRNA produced from HC-Pro and CP genes of BCMV can induce RNAi-based resistance against BCMV infection.

Medicinal plants have long been recognized as a valuable source of human health due to their therapeutic potential in treating a variety of diseases. Tarenna alpestris, a plant native to the Megamalai Hills in the Western Ghats of India, has traditionally been used for numerous medicinal purposes. However, despite its extensive use in folklore, scientific validation of its therapeutic properties remains limited. This study aims to evaluate the phytochemical composition of Tarenna alpestris, assess its antioxidant properties, and explore its potential anticancer effects against HT-29 colon cancer cells. The phytochemical profile was determined using preliminary screening and gas chromatography-mass spectrometry (GC-MS). Antioxidant activity was measured through DPPH, FRAP, H₂O₂, and N₂O₂ assays. The anticancer effects were investigated using the MTT assay for cell viability, AO/EtBr staining for apoptosis detection, DAPI staining for nuclear fragmentation analysis, and flow cytometry for cell cycle analysis. The phytochemical analysis identified several bioactive compounds, including flavonoids, alkaloids, terpenoids, and phenolic acids. Sixteen phytocomponents were detected from the extract by GCMS analysis, the major compounds are 9-octadecynoic acid (21%), N-hexadecanoic acid (16%), methylene diamine, N,N'-diacetyl (15%), 1-allyl-cyclohexane-1,2-diol (11%) 2-methyl-6-methylene-octa-1,7-dien-3-ol (5%), squalene (4%), and lupeol (3%) respectively. The antioxidant assays demonstrated significant free radical scavenging activity, with IC50 values comparable to known antioxidant standards. The antioxidant enzymatic activity of Tarenna alpestris extract suggests a potent ability to neutralize reactive oxygen species and protect against oxidative damage. In vitro studies revealed that Tarenna alpestris extract significantly inhibited the proliferation of HT-29 colon cancer cells and induced apoptosis, based on concentration dependent manner. The concentration needed to inhibit 50% of cell growth, known as the IC₅₀ value, was found to be 26 ± 0.20 μg/mL. Additionally, cell cycle analysis showed G0/G1 phase arrest in treated cells. Tarenna alpestris exhibits a robust phytochemical profile with substantial antioxidant and anticancer properties. These findings support its potential as a therapeutic agent for cancer prevention and treatment. Further research, including in vivo studies, is warranted to fully elucidate its therapeutic efficacy and mechanisms of action.

Background: Solanum melongena Linnaeus (brinjal) belongs to the Solanaceae family and is also known as eggplant. It is one of the most common vegetables that is grown abundantly and consumed by a large number of people. However, it is found to be highly susceptible to harmful pests such as brinjal shoot and fruit borer, (Leucinodes orbonalis) which are responsible for causing severe damage to the plant's health and, correspondingly, its yield. Damages include shoot and leaf spoilage which leads to overall hampering of the metabolic process of the plant. This study aims to suggest that the plant, Solanum melongena L., has certain self-induced mechanisms to withstand these stress and pest attacks by secreting compounds known as "Chlorogenic Acid." Chlorogenic acid is known to be a plant-derived product and is a part of secondary metabolites. Different plant parts were examined for their diverse secondary metabolite content under laboratory conditions. The study was further proceeded by implementing chlorogenic acid exogenously, on the pest-infected plants at a concentration of 1 mg/ml in two different foliar sprays, one consisting of simple water and another 50% ethanol. Moreover, molecular analysis shows a higher expression of the genes which are pivotal for the secretion of chlorogenic acid within the plant itself. The results of this research reveal that chlorogenic acid exhibits a massive potential in controlling pest attacks against Solanum melongena L. and can be used as a potential bio-pesticide.

Currently, there is a demand for rapid, sensitive, low-cost, portable, and visualized testing technologies for point-of-care testing (POCT). However, most traditional testing methods face challenges such as long testing times, complicated operations, and high costs, limiting their implementation in resource-limited areas and hindering the fulfillment of POCT demands. Lateral flow assay (LFA) has emerged as an ideal detection technique for POCT, particularly when utilizing gold nanoparticles (AuNPs) as labels. This approach not only enables visualization with the naked eye but also reduces the need for expensive reading instruments. The technologies reviewed in this paper encompass integrated detection technology utilizing amplification technique and LFA, integrated detection technology utilizing clustered regularly interspaced short palindromic repeats (CRISPR) system and LFA, the utilization of surface-enhanced Raman spectroscopy (SERS) in LFA detection technique, the utilization of aptamers in LFA detection technique, and the utilization of DNA barcodes in LFA detection technique. By integrating these advanced techniques, there is significant potential to overcome the limitations of LFA, including low sensitivity, poor specificity, inability to quantify, and false positives, thereby enabling broader applications in resource-constrained settings. Additionally, this article comprehensively evaluates the strengths and weaknesses of each approach, underscoring the immense developmental potential of AuNPs-based LFA in point-of-care testing (POCT).

Regular long-term exercise can benefit the body and reduce the risk of several diseases, such as cardiovascular disease, diabetes, and obesity. However, the proteomic and metabolomic changes, as well as the physiological responses associated with long-term exercise, remain incompletely understood. To investigate the effects of long-term exercise on the human body, 14 subjects with long-term exercise habits and 10 subjects without exercise habits were selected for this study. Morning urine samples were collected and analyzed for untargeted metabolomics and proteomics using liquid chromatography-mass spectrometry. A total of 404 differential metabolites and 394 differential proteins were screened in this research, and the analysis results indicated that long-term exercise may affect energy metabolism, amino acid synthesis and metabolism, nucleotide metabolism, steroid hormone biosynthesis, and the inflammatory response. These findings offer a more comprehensive understanding of the molecular effects of long-term exercise on the human body and provide a basis for future research exploring the underlying mechanisms.

Alkaliptosis, crucial in various cancers, is a specific form of cell death. This study aims to screen a prognosis-related gene in kidney renal clear cell carcinoma (KIRC) using the alkaliptosis gene set and to investigate the roles of the gene in KIRC and its association with alkaliptosis. Transcriptome and clinical information of KIRC patients were collected from the TCGA-KIRC and GSE29609 database. We detected ZACN levels in normal kidney cells and KIRC cells and assessed the ZACN and JTC801-induced alkaliptosis relationship using immunoblotting and pH measurement. In the alkaliptosis gene set (IKBKB, NFKB1, CA9, CHUK, IKBKG, and RELA), NFKB1, CHUK, and IKBKG exhibited differential expression in TCGA-KIRC. Based on these three genes, two alkaliptosis patterns were identified in TCGA-KIRC. The independent prognostic gene for KIRC, ZACN, was screened. High ZACN in KIRC patients indicated a poor prognosis. ZACN was inversely related to the infiltration of anti-cancer immune cells such as CD4 + T cells, macrophages, and neutrophils, and it regulated the immune checkpoint and gene mutations. Patients characterized by high ZACN levels exhibited a heightened drug sensitivity to ABT737_1910, 5-Fluorouracil_1073, etc. ZACN expression in KIRC cells was increased relative to normal kidney cells and was inhibited in a concentration-dependent manner by JTC801. ZACN overexpression suppressed p-p65/p65 expression, increased expression of CA9, and lowered intracellular pH. In KIRC, ZACN inhibits JTC801-induced alkaliptosis. This study sheds light on a novel mechanism involving ZACN and alkaliptosis in KIRC, offering a promising avenue for further research and potential therapeutic interventions in KIRC management.