Search Results (35)

The retinal pigment epithelium (RPE) is strongly involved in the pathogenesis of several retinal diseases, such as age-related macular degeneration (AMD). RPE models addressing specific pathological pathways are of high importance for understanding cellular pathomechanisms and pre-clinical screening of potential new therapeutics. The goal of this study is to establish standard operation protocols for single-eye porcine RPE preparation for AMD-relevant models of oxidative stress (RPE-Ox) and inflammation (RPE-Inf). Porcine primary RPE were prepared from one eye and seeded into one well of 12-well plates or, for polar differentiation, in transwell inserts. Different coatings (Poly-ᴅ-Lysine and laminin) and serum content of media (10%, 5%, and 1%) were tested to determine optimal culture parameters. For RPE-Ox, cells were treated with NaIO₃, CoCl₂, or erastin; cell viability (thiazolyl blue tetrazolium bromide, MTT), and gene expression (RT-qPCR) were determined. For RPE-Inf, cells were treated with lipopolysaccharide (LPS), polyinosinic/polycytidylic acid (Poly I:C), or tumor necrosis factor alpha (TNF-α); cell viability (MTT), cytokine secretion (ELISA), and gene expression (RT-qPCR) were determined. For transwell plates in RPE-Inf, cell viability (MTT), polar cytokine secretion (ELISA), gene expression (RT-qPCR), and transepithelial electrical resistance (TEER) for barrier assessment were conducted. For RPE-Ox, effective LD₅₀ could be achieved by using 24 h stimulation with 25 µm erastin, seven days after preparation in 5% serum cultures, without coating. For gene expression assessment, the use of Poly-ᴅ-Lysine is recommended. For RPE-Inf, three days of LPS stimulation (1 µg/mL) showed effective cytokine activation with 5% serum on uncoated 12-well plates. Transwell plates are not recommended for cytokine secretion assessment. It can be used for cell barrier assays in which LPS also showed effective cell barrier decrease and gene expression assays. Two specific best practice protocols for the use of porcine single-eye cultures in AMD research concerning oxidative stress and inflammation with optimized parameters were established and are provided. Full article

The increasing integration of physical and nanotechnological treatments in agriculture has unlocked new possibilities for enhancing seed performance and the functional properties of seedlings. This study aimed to determine the effect of the coupled use of pulsed electric field (PEF) and the soaking (coating) of sunflower seeds in metal nanoparticles (AgNP and CuNP) on their germination capacity and on the stem and root length, content of pigments (chlorophyll a, chlorophyll b, carotenoids), color profile, and antioxidative properties (FRAP, polyphenols, TPC, ABTS, and DPPH) of sunflower seedlings. The study results enable the drawing of explicit conclusions that the higher PEF energy applied (5.5 kJ kg⁻¹) and seed treatment with nanoparticle solutions, in most cases, diminished the germination capacity of sunflower seeds (from 3.50 to 44.11%) compared to the control samples. A decreased seedling stem length was determined at both PEF energy levels tested, i.e., 1 kJ kg⁻¹ and 5.5 kJ kg⁻¹, with the values obtained being 11.86% to 46.14% lower compared to the respective control samples. The root length of the seedlings decreased as well, i.e., by 7.34 to 41%. The content of chlorophyll a (chl a) increased in the seedlings from all experimental variants compared to the control, whereas that of chlorophyll b (chl b) decreased by 3.24 to 7.86% in the control variant with PEF and CuNP. The FRAP value, total content of polyphenols, and TPC ranged from 10.20 to 12.95 (mg TE g⁻¹ DM), from 42.23 to 49.19 (mg GAE g⁻¹ DM), and from 20.20 to 23.90 (mg GAE g⁻¹ DM), respectively, and showed an upward trend compared to the control samples. The results of this study indicate that further research is needed to understand how the analyzed treatments affect seedling growth and demonstrate reduced germination capacity and enhanced antioxidant activity due to the synergistic effect of a high PEF and nanoparticle solutions. Full article

Age-related macular degeneration (AMD) is the main cause of blindness in Western nations. AMD models addressing specific pathological pathways are desired. Through this study, a best-practice protocol for polarized porcine single-eye retinal pigment epithelium (RPE) preparation for AMD-relevant models of RPE barrier and polarity is established. Single-eye porcine primary RPE cells (from one eye for one well) were prepared in 12-well plates including Transwell inserts. Different coatings (laminin (Lam), Poly-ᴅ-Lysine (PDL), fibronectin (Fn) and collagens) and varying serum contents (1%, 5% and 10%) were investigated to determine optimal culture parameters for this model. Success rates of cultures, cell number (trypan-blue exclusion assay), morphology/morphometry (light and fluorescence microscopy), protein secretion/expression (ELISA, Western blot), gene expression (qPCR), transepithelial electric resistance (TEER) and polar location of bestrophin 1 (BEST1) by cryosectioning (IHC-Fr) were assessed. Cells seeded on Lam exhibited the highest level of epithelial cells and confluence properties. Fn resulted in the highest cell number growth. Lam and Fn exhibited the highest culture success rates. TEER values and vascular endothelial growth factor secretion were highest when Lam was used. For the first time, polar (Transwell) porcine single-eye RPE morphometry parameters were determined. RPE on Lam showed bigger cells with a higher variety of cell shapes. CIV displayed the lowest claudin 19 expression. The highest basolateral expression of BEST1 was achieved with Lam coating. The higher the serum, the better the cell number increase and confluence success. A reduction in serum on Lam showed positive results for RPE morphology, while morphometry remained stable. A five percent serum on Lam showed the highest culture success rate and best barrier properties. RPE65 expression was reduced by using 10% serum. Altogether, the most suitable coating of Transwell inserts was Lam, and a reduction in serum to 5% is recommended, as well as a cultivation time of 28 days. A protocol for the use of polar porcine single-eye cultures with validated parameters was established and is provided herein. Full article

Plant fungal diseases remain a major threat to global agricultural production, necessitating eco-friendly and sustainable strategies. Conventional chemical fungicides often lead to the development of resistant pathogen strains and cause environmental contamination. Therefore, the development of biocontrol agents is particularly important. In this study, we identified Burkholderia stagnalis YJ-2 from the rhizosphere soil of Woodsia ilvensis as a promising biocontrol strain using 16S rRNA and whole-genome sequencing. This strain demonstrated broad-spectrum antifungal activity against plant fungal pathogens, with its bioactive extracts maintaining high stability across a temperature range of 25–100 °C and pH range of 2–12. We used in vitro assays to further show that the metabolites of B. stagnalis YJ-2 disrupted the hyphal morphology of Valsa mali, resulting in swelling, reduced branching, and increased pigmentation. Fluorescence labeling confirmed that B. stagnalis YJ-2 stably colonized the roots and stems of tomato and wheat plants. Furthermore, various formulations of microbial agents based on B. stagnalis YJ-2 were evaluated for their efficacy against plant pathogens. The seed-coating formulation notably protected tomato seedlings from Alternaria solani infection without affecting germination (p > 0.1), while the wettable powder exhibited significant control effects on early blight in tomatoes, with the preventive treatment showing better efficacy than the therapeutic treatment. Additionally, the B. stagnalis YJ-2 bone glue agent showed a substantial inhibitory effect on apple tree canker. Whole-genome analysis of B. stagnalis YJ-2 revealed a 7,705,355 bp genome (67.68% GC content) with 6858 coding genes and 20 secondary metabolite clusters, including three clusters (YJ-2_GM002015-YJ-2_GM002048, YJ-2_GM0020090-YJ-2_GM002133, and YJ-2_GM06534-YJ-2_GM006569) that are related to the antifungal activity of YJ-2 and are homologous to the biosynthetic gene clusters of known secondary metabolites, such as icosalide, ornibactin, and sinapigladioside. We further knocked out core biosynthetic genes of two secondary metabolic gene clusters and found that only the YJ-2_GM006534-YJ-2_GM006569 gene cluster had a corresponding function in two potential antifungal gene clusters. In contrast to the wild-type strain YJ-2, only deletion of the YJ-2_GM006563 gene reduced the antifungal activity of B. stagnalis YJ-2 by 8.79%. These findings highlight the biocontrol potential of B. stagnalis YJ-2, supporting a theoretical foundation for its development as a biocontrol agent against plant fungal diseases and thereby promoting sustainable agricultural disease management. Full article

The color of the walnut seed coat is a critical determinant of its market value; however, research into the mechanisms responsible for seed coat color formation is yet to be determined. Using two walnut clones with contrasting pale-yellow and light purple seed coats, we characterized pigmentation, particularly anthocyanin content, using spectrophotometry. We then conducted integrated transcriptomic and metabolomic analyses to identify the molecular mechanisms and pathways underlying their formation. The anthocyanin content in the light purple seed coat clone was significantly greater than that in the clone with a white seed coat. The results of comparative metabolomics indicated that four anthocyanins (delphinidin, cyanidin-3-(caffeoylglucoside), pelargonidin-3-(6″-caffeoylglucoside), and delphinidin-3-O-sophoroside) were significantly more abundant in the light purple seed coat clone. These anthocyanins were the key pigments responsible for the light purple coloration of the walnut seed coat. Furthermore, comparative transcriptomics revealed that structural genes in the anthocyanin biosynthesis pathway (e.g., phenylalanine ammonia-lyase, 4-coumarate-CoA ligase, chalcone isomerase, and bronze-1) were significantly upregulated in the purple seed coat clone. Coexpression network analysis revealed that several transcription factors (e.g., ARF, bHLH, and MYB-related) were significantly correlated with the upregulation of these structural genes and the accumulation of four key anthocyanins. These transcription factors may serve as critical regulators influencing seed coat color formation. In conclusion, these findings establish a strong theoretical foundation for walnut breeding aimed at developing diverse seed coat colors. Full article

As an agri-food by-product, the rice bran of pigmented rice, encompassing varieties such as red, black, and purple rice, has garnered increasing attention due to its richness in terms of bioactive compounds. Being mainly composed of the pericarp, aleuron, seed coat, and germ, the brown outer layer of the rice kernel offers potential health benefits and has applications in skincare. Human skin serves as the primary barrier against external threats, including pathogens, pollutants, and ultraviolet (UV) radiation. Notably, UV radiation accelerates the aging process and contributes to various skin issues. Recent trends suggest a heightened interest in incorporating pigmented rice into skincare regimens, motivated by its potential to mitigate oxidative stress, inflammation, and pigmentation, which are pivotal factors in skin aging and photodamage. With increasing consumer demand for natural and sustainable ingredients, pigmented rice has emerged as a promising candidate within the skincare and personal care sectors, effectively bridging the gap between nutrition and dermatological health. This review examines the applications of pigmented rice in skincare, with a particular focus on its bioactive components and potential mechanisms of action that contribute to skin health. The unique chemical composition of pigmented rice, which includes compounds such as anthocyanins, flavonoids, phenolic acids, and vitamin E, underlies its antioxidant, anti-inflammatory, and skin-protective properties. Despite the increasing recognition of its benefits, a comprehensive understanding of the underlying mechanisms remains limited, underscoring the necessity for further research to exploit the potential of pigmented rice in skincare applications fully. Full article

Morus nigra L., or mulberry, is a susceptible fleshy fruit due to its high respiratory rate and low storage stability, which shortens its shelf life and makes it difficult to commercialize in natura. Edible coatings, thin membranes produced directly on the desired surface, could improve food preservation, among other properties. Annatto (Bixa orellana L.) seeds are natural pigments with high antioxidant activity. This work aimed to develop a pectin-based edible coating with annatto extract to increase the shelf life of fruits, using mulberries as a study model. The mulberries were randomly separated into five groups: without coating, coated with different extract concentrations (0%, 5%, and 10%), and a layer-by-layer treatment consisting of a pectin layer under a 10% extract layer. The samples were evaluated for the following parameters: titratable acidity, maturity index, mass loss, pH, soluble solids, moisture contents, and bioactive compounds. The coated group with 10% annatto extract had the best result for the maturity index (25.52), while the group with 5% showed better mass loss and moisture (37.28% and 83.66%, respectively). Herein, it was demonstrated that pectin-based edible coatings with annatto extract delay the maturation and senescence of mulberries, preserving the bioactive compounds and increasing shelf life. Full article

Background: Age-related macular degeneration (AMD) is the leading cause of severe vision loss in industrialized nations. AMD models based on standardized optimized cell culture models are warranted. The aim of this study is to implement a standard operation protocol for the preparation of porcine retinal pigment epithelium (RPE) from pig eyes with cells from one eye designated for one culture, generating a genetic homology within one culture and genetic heterogeneity between cultures, thereby implementing a relevant in vitro model for AMD investigations. In addition, the use of pigs’ eyes from slaughterhouse waste material is an active measure to reduce animal experimentation in ophthalmological research. Methods: Primary porcine RPE cells were prepared, and cells from one eye were seeded into one well of a twelve-well polystyrene plate. They were cultured for 7, 14, and 28 days. Different post-mortem times (2, 4, and 6 h), coatings (collagen I, IV, Poly-d-Lysine, fibronectin, and laminin), and serum content of media (1%, 5%, and 10%) were tested. The success rate for confluence and survival was determined. At the different time points, cell number (trypan blue exclusion assay), confluence and morphology (microscope imaging), VEGF content of supernatants, and gene and protein expression, as well as tight junctions (fluorescence imaging), were examined. In addition, a baseline for RPE morphometry using CellProfiler software was established. Results: A post-mortem time of 4 to 5 h is most suitable. A coating with Poly-d-Lysine is recommended due to high success rates, the fastest confluence, the highest expression of RPE65, and suitable morphologic properties. The results for confluence, protein expression, and morphology showed that a reduction of 10% to 5% FBS is acceptable. Conclusion: A basic best practice protocol for the preparation of porcine single-eye cultures with optimized parameters was established and is provided. Full article

B-box (BBX) proteins, a subfamily of zinc-finger transcription factors, are involved in various environmental signaling pathways. In this study, we conducted a comprehensive analysis of BBX family members in Brassica crops. The 482 BBX proteins were divided into five groups based on gene structure, conserved domains, and phylogenetic analysis. An analysis of nonsynonymous substitutions and (Ka)/synonymous substitutions (Ks) revealed that most BBX genes have undergone purifying selection during evolution. An analysis of transcriptome data from rapeseed (Brassica napus) organs suggested that BnaBBX3d might be involved in the development of floral tissue-specific RNA-seq expression. We identified numerous light-responsive elements in the promoter regions of BnaBBX genes, which were suggestive of participation in light signaling pathways. Transcriptomic analysis under shade treatment revealed 77 BnaBBX genes with significant changes in expression before and after shading treatment. Of these, BnaBBX22e showed distinct expression patterns in yellow- vs. black-seeded materials in response to shading. UPLC-HESI-MS/MS analysis revealed that shading influences the accumulation of 54 metabolites, with light response BnaBBX22f expression correlating with the accumulation of the flavonoid metabolites M46 and M51. Additionally, BnaBBX22e and BnaBBX22f interact with BnaA10.HY5. These results suggest that BnaBBXs might function in light-induced pigment accumulation. Overall, our findings elucidate the characteristics of BBX proteins in six Brassica species and reveal a possible connection between light and seed coat color, laying the foundation for further exploring the roles of BnaBBX genes in seed development. Full article

Iron deficiency remains a public health challenge globally. Prebiotics have the potential to improve iron bioavailability by modulating intestinal bacterial population, increasing SCFA production, and stimulating expression of brush border membrane (BBM) iron transport proteins among iron-deficient populations. This study intended to investigate the potential effects of soluble extracts from the cotyledon and seed coat of three pea (Pisum sativum) varieties (CDC Striker, CDC Dakota, and CDC Meadow) on the expression of BBM iron-related proteins (DCYTB and DMT1) and populations of beneficial intestinal bacteria in vivo using the Gallus gallus model by oral gavage (one day old chicks) with 1 mL of 50 mg/mL pea soluble extract solutions. The seed coat treatment groups increased the relative abundance of Bifidobacterium compared to the cotyledon treatment groups, with CDC Dakota seed coat (dark brown pigmented) recording the highest relative abundance of Bifidobacterium. In contrast, CDC Striker Cotyledon (dark-green-pigmented) significantly increased the relative abundance of Lactobacillus (p < 0.05). Subsequently, the two dark-pigmented treatment groups (CDC Striker Cotyledon and CDC Dakota seed coats) recorded the highest expression of DCYTB. Our study suggests that soluble extracts from the pea seed coat and dark-pigmented pea cotyledon may improve iron bioavailability by affecting intestinal bacterial populations. Full article

Legume–rhizobia symbiosis is the most important plant–microbe interaction in sustainable agriculture due to its ability to provide much needed N in cropping systems. This interaction is mediated by the mutual recognition of signaling molecules from the two partners, namely legumes and rhizobia. In legumes, these molecules are in the form of flavonoids and anthocyanins, which are responsible for the pigmentation of plant organs, such as seeds, flowers, fruits, and even leaves. Seed-coat pigmentation in legumes is a dominant factor influencing gene expression relating to N₂ fixation and may be responsible for the different N₂-fixing abilities observed among legume genotypes under field conditions in African soils. Common bean, cowpea, Kersting’s groundnut, and Bambara groundnut landraces with black seed-coat color are reported to release higher concentrations of nod-gene-inducing flavonoids and anthocyanins compared with the Red and Cream landraces. Black seed-coat pigmentation is considered a biomarker for enhanced nodulation and N₂ fixation in legumes. Cowpea, Bambara groundnut, and Kersting’s bean with differing seed-coat colors are known to attract different soil rhizobia based on PCR-RFLP analysis of bacterial DNA. Even when seeds of the same legume with diverse seed-coat colors were planted together in one hole, the nodulating bradyrhizobia clustered differently in the PCR-RFLP dendrogram. Kersting’s groundnut, Bambara groundnut, and cowpea with differing seed-coat colors were selectively nodulated by different bradyrhizobial species. The 16S rRNA amplicon sequencing also found significant selective influences of seed-coat pigmentation on microbial community structure in the rhizosphere of five Kersting’s groundnut landraces. Seed-coat color therefore plays a dominant role in the selection of the bacterial partner in the legume–rhizobia symbiosis. Full article

The association between pre-harvest sprouting (PHS) and seed coat color has long been recognized. Red-grained wheats generally exhibit greater PHS resistance compared to white-grained wheat, although variability in PHS resistance exists within red-grained varieties. Here, we conducted a genome-wide association study on a panel consisting of red-grained wheat varieties, aimed at uncovering genes that modulate PHS resistance and red color components of seed coat using digital image processing. Twelve loci associated with PHS traits were identified, nine of which were described for the first time. Genetic loci marked by SNPs AX-95172164 (chromosome 1B) and AX-158544327 (chromosome 7D) explained approximately 25% of germination index variance, highlighting their value for breeding PHS-resistant varieties. The most promising candidate gene for PHS resistance was TraesCS6B02G147900, encoding a protein involved in aleurone layer morphogenesis. Twenty-six SNPs were significantly associated with grain color, independently of the known Tamyb10 gene. Most of them were related to multiple color characteristics. Prioritization of genes within the revealed loci identified TraesCS1D03G0758600 and TraesCS7B03G1296800, involved in the regulation of pigment biosynthesis and in controlling pigment accumulation. In conclusion, our study identifies new loci associated with grain color and germination index, providing insights into the genetic mechanisms underlying these traits. Full article

This study investigates the genetic determinants of seed coat color and pattern variations in cowpea (Vigna unguiculata), employing a genome-wide association approach. Analyzing a mapping panel of 296 cowpea varieties with 110,000 single nucleotide polymorphisms (SNPs), we focused on eight unique coat patterns: (1) Red and (2) Cream seed; (3) White and (4) Brown/Tan seed coat; (5) Pink, (6) Black, (7) Browneye and (8) Red/Brown Holstein. Across six GWAS models (GLM, SRM, MLM, MLMM, FarmCPU from GAPIT3, and TASSEL5), 13 significant SNP markers were identified and led to the discovery of 23 candidate genes. Among these, four specific genes may play a direct role in determining seed coat pigment. These findings lay a foundational basis for future breeding programs aimed at creating cowpea varieties aligned with consumer preferences and market requirements. Full article

The loss of anthocyanin pigments is one of the most common evolutionary transitions in petal color, yet the genetic basis for these changes in flax remains largely unknown. In this study, we used crossing studies, a bulk segregant analysis, genome-wide association studies, a phylogenetic analysis, and transgenic testing to identify genes responsible for the transition from blue to white petals in flax. This study found no correspondence between the petal color and seed color, refuting the conclusion that a locus controlling the seed coat color is associated with the petal color, as reported in previous studies. The locus controlling the petal color was mapped using a BSA-seq analysis based on the F₂ population. However, no significantly associated genomic regions were detected. Our genome-wide association study identified a highly significant QTL (BP4.1) on chromosome 4 associated with flax petal color in the natural population. The combination of a local Manhattan plot and an LD heat map identified LuMYB314, an R2R3-MYB transcription factor, as a potential gene responsible for the natural variations in petal color in flax. The overexpression of LuMYB314 in both Arabidopsis thaliana and Nicotiana tabacum resulted in anthocyanin deposition, indicating that LuMYB314 is a credible candidate gene for controlling the petal color in flax. Additionally, our study highlights the limitations of the BSA-seq method in low-linkage genomic regions, while also demonstrating the powerful detection capabilities of GWAS based on high-density genomic variation mapping. This study enhances our genetic insight into petal color variations and has potential breeding value for engineering LuMYB314 to develop colored petals, bast fibers, and seeds for multifunctional use in flax. Full article

Titanium oxide (TiO₂) is the most widely used white pigment because of its brightness and very high reflective index, traits surpassed by only a few other materials; it has gained adequate ground in the fabrication of solar cells due to its wide band gap of 3.32 eV. Various natural dyes such as laali plant dye, zobo leaf dye and tomato seed dye act as sensitizers. This research intends to explore the effect of this titanium oxide on enhancing sensitivity in light harvesting by using dye-sensitized solar cell fabrication. Indium tin oxide, one of the transparent, conducting optical glasses, was chosen for the photoanode, on which the prepared titanium powder and the extracted dye were coated using the screen printing method. TiO₂ was screen printed over the TCO (ITO) or plain glass slide and annealed at 4000 °C for 3 min; then, the dyes were injected drop by drop and analysis was carried out for XRD and UV–optical. From the XRD results obtained for the laali dye, the XRD showed no prominent peaks and when improved by introducing titanium oxide, it showed the peaks as having a rutile nature which enhances light harvesting. The optical properties showed a transmittance edge at 350 nm which gradually increased as the wavelength increased with no visibility on the absorbance graph. For the tomato dye, a visible peak was observed and this increased with the addition of titanium oxide, while transmittance rose at 380 nm and fell at 550 nm, with no absorbance. The zobo dye showed no evidence of visible peaks and little change in the peak visibility with the addition of TiO₂ was observed, with the transmittance edge at 350 nm, maximum at 390 nm and constant with TiO₂ enhancement, and showing no visible absorbance properties. Laali and zobo are good transmittance materials, unlike the tomato dye which is a good absorbance material. Conclusively, TiO₂ is effective in dye-sensitized solar cell fabrication since there were visible changes within the scientific environment which further enhanced light harvesting. Full article