Search Results (25)

The aim of this study was to explore the effects of different lactic acid bacteria additives (Lactiplantibacillus plantarum or Lentilactobacillus buchneri) on the fermentation quality, chemical composition, in vitro digestibility, bacterial community structure, and predictive function of S. perfoliatum silage feed. Fresh S. perfoliatum was wilted overnight, then its moisture content was adjusted between 65 and 70%. The experiment was performed in three groups as follows: (1) the control group (CK group), which lacked a Lactobacillus preparation; (2) the Lactiplantibacillus plantarum (L. plantarum) group (LP group), which was inoculated with L. plantarum at 5 × 10⁶ cfu/g FW; and (3) the Lentilactobacillus buchneri (L. buchneri) group (LB group), which was inoculated with L. buchneri at 5 × 10⁶ cfu/g FW. The results showed that L. plantarum significantly reduced pH and increased lactic acid (LA) content in S. perfoliatum silage compared with the control. L. buchneri, on the other hand, excelled in reducing ammonia nitrogen (NH₃-N) content and significantly increased acetic acid (AA) content. At 60 days of fermentation, the CP content was significantly higher (p < 0.05) in the LP and LB groups than in the CK group (19.29 vs. 15.53 and 15.87). At 60 days of fermentation, the ivCPD was significantly higher (p < 0.05) in the LB group than in the CK and LP groups (57.80 vs. 54.77 and 55.77). The 60-day silage process completely altered the bacterial community of S. perfoliatum silage. In the fresh samples, the dominant genera were Weissella_A and Pantoea_A. Weissella_A and Pantoea_A were gradually replaced by Lentilactobacillus and Lactiplantibacillus after S. perfoliatum ensiling. After 45 days of fermentation, L. buchneri became the dominant strain in CK, LP and LB groups. Inoculation with L. plantarum altered the succession of the bacterial community from 7 to 15 days of fermentation of S. perfoliatum. In contrast, inoculation with L. buchneri affected the succession of the bacterial community from 30 to 60 days of S. perfoliatum fermentation. In S. perfoliatum silage aged 7 to 60 days, the amino acid metabolic pathway in the LB group remained upregulated. The experimental results revealed that inoculation with L. buchneri had a stronger effect on S. perfoliatum silage than inoculation with L. plantarum. Thus, L. buchneri should be selected as an additive for S. perfoliatum silage fermentation in practical production. Full article

Heme is a chemical compound crucial for various biological processes and industrial applications. However, the microbial production of heme is often limited by its intracellular accumulation and associated toxicity. To address this, we employed a two-step approach involving in vivo cell cultivation for the production of a heme precursor (coproporphyrin III or coproheme) followed by its in vitro conversion(s) to heme. For the first step, we engineered Escherichia coli strains by implementing the coproporphyrin-dependent (CPD) pathway for bacterial cell cultivation, extracellularly producing up to 251 mg/L coproporphyrin III and 85 mg/L coproheme, respectively. For the second step, we cloned the hemH and hemQ genes for expression in E. coli, and the expressed gene products, i.e., coproheme decarboxylase (ChdC/HemH) and heme synthase (HemQ), were purified. Using the purified enzymes with modulated reaction conditions, we achieved up to a 77.2% yield to convert coproporphyrin III to coproheme and a 45.8% yield to convert coproheme to heme. This in vitro approach not only bypassed the intracellular toxicity constraint associated with in vivo cell cultivation but also enabled precise reaction control, leading to a higher efficiency and yield for heme (and coproheme) production. By applying novel strategies in strain engineering and bioprocessing to overcome inherent bioprocess challenges, this study paves the way for industrial biotechnology for the sustainable, efficient, and even large-scale bio-based production of heme. Full article

The breadth and volume of research exploring the complexities of childhood trauma in Early Childhood Education and Care (ECEC) environments has grown significantly in recent years, yet, little is known about this trajectory in an Irish context. Consequently, the purpose of the current study was twofold: (i) consult with influential figures in the wider ECEC domain to access expert knowledge on the provision of trauma awareness education in Ireland and to (ii) gain invaluable insights into the trauma awareness knowledge, training, and experiences of ECEC professionals working in Irish ECEC settings. Utilising a qualitative methodological approach, eight semi-structured interviews with ECEC sector representatives (i.e., IV Group) and six focus groups with managers (n = 8), room leaders (n = 8), and practitioners (n = 8) (i.e., FG Group) were conducted as explorative processes to investigate the presence of trauma awareness in ECEC, if any, and the factors supporting/hindering this discourse from the perspectives of diverse stakeholders and professionals in Ireland. Results revealed that the provision of trauma awareness is challenged by the dearth of relevant educational opportunities during both Initial Practitioner Education (IPE) and Continuous Professional Development (CPD) pathways, with an absence of neuro-informed practice among the FG Group. Contrastingly, while sector representatives in the IV Group demonstrated an awareness of childhood trauma, these participants identified the need for (i) government investment, (ii) training reform, and (iii) professional practice considerations. Based on the cumulative findings, this research recommends that policymakers in Ireland prioritise the provision of funded and/or incentivised professional development opportunities for the ECEC profession to enhance the knowledge and skills associated with trauma-informed practice. Full article

Ultraviolet (UV) radiation plays a crucial role in the development of melanoma and non-melanoma skin cancers. The types of UV radiation are differentiated by wavelength: UVA (315 to 400 nm), UVB (280 to 320 nm), and UVC (100 to 280 nm). UV radiation can cause direct DNA damage in the forms of cyclobutane pyrimidine dimers (CPDs) and 6-4 photoproducts (6-4PPs). In addition, UV radiation can also cause DNA damage indirectly through photosensitization reactions caused by reactive oxygen species (ROS), which manifest as 8-hydroxy-2′-deoxyguanine (8-OHdG). Both direct and indirect DNA damage can lead to mutations in genes that promote the development of skin cancers. The development of melanoma is largely influenced by the signaling of the melanocortin one receptor (MC1R), which plays an essential role in the synthesis of melanin in the skin. UV-induced mutations in the BRAF and NRAS genes are also significant risk factors in melanoma development. UV radiation plays a significant role in basal cell carcinoma (BCC) development by causing mutations in the Hedgehog (Hh) pathway, which dysregulates cell proliferation and survival. UV radiation can also induce the development of squamous cell carcinoma via mutations in the TP53 gene and upregulation of MMPs in the stroma layer of the skin. Full article

Meningioma, a primary brain tumor, is commonly encountered and accounts for 39% of overall CNS tumors. Despite significant progress in clinical research, conventional surgical and clinical interventions remain the primary treatment options for meningioma. Several proteomics and transcriptomics studies have identified potential markers and altered biological pathways; however, comprehensive exploration and data integration can help to achieve an in-depth understanding of the altered pathobiology. This study applied integrated meta-analysis strategies to proteomic and transcriptomic datasets comprising 48 tissue samples, identifying around 1832 common genes/proteins to explore the underlying mechanism in high-grade meningioma tumorigenesis. The in silico pathway analysis indicated the roles of extracellular matrix organization (EMO) and integrin binding cascades in regulating the apoptosis, angiogenesis, and proliferation responsible for the pathobiology. Subsequently, the expression of pathway components was validated in an independent cohort of 32 fresh frozen tissue samples using multiple reaction monitoring (MRM), confirming their expression in high-grade meningioma. Furthermore, proteome-level changes in EMO and integrin cell surface interactions were investigated in a high-grade meningioma (IOMM-Lee) cell line by inhibiting integrin-linked kinase (ILK). Inhibition of ILK by administrating Cpd22 demonstrated an anti-proliferative effect, inducing apoptosis and downregulating proteins associated with proliferation and metastasis, which provides mechanistic insight into the disease pathophysiology. Full article

The final three steps of heme biogenesis exhibit notable differences between di- and mono-derm bacteria. The former employs the protoporphyrin-dependent (PPD) pathway, while the latter utilizes the more recently uncovered coproporphyrin-dependent (CPD) pathway. In order to devise a rapid screen for potential inhibitors that differentiate the two pathways, the genes associated with the protoporphyrin pathway in an Escherichia coli YFP strain were replaced with those for the CPD pathway from Staphylococcus aureus (SA) through a sliding modular gene replacement recombineering strategy to generate the E. coli strain Sa-CPD-YFP. Potential inhibitors that differentially target the pathways were identified by screening compound libraries against the YFP-producing Sa-CPD-YFP strain in comparison to a CFP-producing E. coli strain. Using a mixed strain assay, inhibitors targeting either the CPD or PPD heme pathways were identified through a decrease in one fluorescent signal but not the other. An initial screen identified both azole and prodigiosin-derived compounds that were shown to specifically target the CPD pathway and which led to the accumulation of coproheme, indicating that the main target of inhibition would appear to be the coproheme decarboxylase (ChdC) enzyme. In silico modeling highlighted that these inhibitors are able to bind within the active site of ChdC. Full article

The members of Microbacterium isolated from different environments are known to form peptidoglycan. In this study, we compared the biofilm-forming abilities of Microbacterium sp. PAMC22086 (PAMC22086), which was isolated from the soil in the South Shetland Islands and Microbacterium sp. PAMC21962 (PAMC21962), which was isolated from algae in the South Shetland Islands. The analysis of average nucleotide identity and phylogeny of PAMC22086 revealed a 97% similarity to Microbacterium oxydans VIU2A, while PAMC21962 showed a 99.1% similarity to Microbacterium hominis SGAir0570. For the comparative genomic analysis of PAMC22086 and PAMC21962, the genes related to biofilm formation were identified using EggNOG and KEGG pathway databases. The genes possessed by both PAMC22086 and PAMC21962 are cpdA, phnB, rhlC, and glgC, which regulate virulence, biofilm formation, and multicellular structure. Among the genes indirectly involved in biofilm formation, unlike PAMC21962, PAMC22086 possessed csrA, glgC, and glgB, which are responsible for attachment and glycogen biosynthesis. Additionally, in PAMC22086, additional functional genes rsmA, which is involved in mobility and polysaccharide production, and dksA, GTPase, and oxyR, which play roles in cell cycle and stress response, were identified. In addition, the biofilm-forming ability of the two isolates was examined in vivo using the standard crystal violet staining technique, and morphological differences in the biofilm were investigated. It is evident from the different distribution of biofilm-associated genes between the two strains that the bacteria can survive in different niches by employing distinct strategies. Both strains exhibit distinct morphologies. PAMC22086 forms a biofilm that attaches to the side, while PAMC21962 indicates growth starting from the center. The biofilm formation-related genes in Microbacterium are not well understood. However, it has been observed that Microbacterium species form biofilm regardless of the number of genes they possess. Through comparison between different Microbacterium species, it was revealed that specific core genes are involved in cell adhesion, which plays a crucial role in biofilm formation. This study provides a comprehensive profile of the Microbacterium genus’s genomic features and a preliminary understanding of biofilm in this genus, laying the foundation for further research. Full article

Cyclobutane pyrimidine dimers (CPDs) are ultraviolet radiation (UV)-induced carcinogenic DNA photoproducts that lead to UV signature mutations in melanoma. Previously, we discovered that, in addition to their incident formation (iCPDs), UV exposure induces melanin chemiexcitation (MeCh), where UV generates peroxynitrite (ONOO⁻), which oxidizes melanin into melanin-carbonyls (MCs) in their excited triplet state. Chronic MeCh and energy transfer by MCs to DNA generates CPDs for several hours after UV exposure ends (dark CPD, dCPDs). We hypothesized that MeCh and the resulting dCPDs can be inhibited using MeCh inhibitors, and MC and ONOO⁻ scavengers. Here, we investigated the efficacy of Acetyl Zingerone (AZ), a plant-based phenolic alkanone, and its chemical analogs in inhibiting iCPDs and dCPDs in skin fibroblasts, keratinocytes, and isogenic pigmented and albino melanocytes. While AZ and its methoxy analog, 3-(4-Methoxy-benzyl)-Pentane-2,4-dione (MBPD) completely inhibited the dCPDs, MBPD also inhibited ~50% of iCPDs. This suggests the inhibition of ~80% of total CPDs at any time point post UV exposure by MBPD, which is markedly significant. MBPD downregulated melanin synthesis, which is indispensable for dCPD generation, but this did not occur with AZ. Meanwhile, AZ and MBPD both upregulated the expression of nucleotide excision repair (NER) pathways genes including Xpa, Xpc, and Mitf. AZ and its analogs were non-toxic to the skin cells and did not act as photosensitizers. We propose that AZ and MBPD represent “next-generation skin care additives” that are safe and effective for use not only in sunscreens but also in other specialized clinical applications owing to their extremely high efficacy in blocking both iCPDs and dCPDs. Full article

UVB radiation is known to trigger the block of DNA replication and transcription by forming cyclobutane pyrimidine dimer (CPD), which results in severe skin damage. CPD photolyase, a kind of DNA repair enzyme, can efficiently repair CPDs that are absent in humans and mice. Although exogenous CPD photolyases have beneficial effects on skin diseases, the mechanisms of CPD photolyases on the skin remain unknown. Here, this study prepared CPD photolyase nanoliposomes (CPDNL) from Antarctic Chlamydomonas sp. ICE-L, which thrives in harsh, high-UVB conditions, and evaluated their protective mechanisms against UVB-induced damage in mice. CPDNL were optimized using response surface methodology, characterized by a mean particle size of 105.5 nm, with an encapsulation efficiency of 63.3%. Topical application of CPDNL prevented UVB-induced erythema, epidermal thickness, and wrinkles in mice. CPDNL mitigated UVB-induced DNA damage by significantly decreasing the CPD concentration. CPDNL exhibited antioxidant properties as they reduced the production of reactive oxygen species (ROS) and malondialdehyde. Through activation of the NF-κB pathway, CPDNL reduced the expression of pro-inflammatory cytokines including IL-6, TNF-α, and COX-2. Furthermore, CPDNL suppressed the MAPK signaling activation by downregulating the mRNA and protein expression of ERK, JNK, and p38 as well as AP-1. The MMP-1 and MMP-2 expressions were also remarkably decreased, which inhibited the collagen degradation. Therefore, we concluded that CPDNL exerted DNA repair, antioxidant, anti-inflammation, and anti-wrinkle properties as well as collagen protection via regulation of the NF-κB/MAPK/MMP signaling pathways in UVB-induced mice, demonstrating that Antarctic CPD photolyases have the potential for skincare products against UVB and photoaging. Full article

Brassinosteroid (BR) signaling is very important in plant developmental processes. Its various components interact to form a signaling cascade. These components are widely studied in Arabidopsis; however, very little information is available on tomatoes. Brassinosteroid Insensitive 2 (BIN2), the downstream suppressor of BR signaling, plays a critical role in BR signal pathway, while FRIGIDA as a key suppressor of Flowering Locus C with overexpression could cause early flowering; however, how the BR signaling regulates FRIGIDA homologous protein to adjust flowering time is still unknown. This study identified 12 FRIGIDA-LIKE proteins with a conserved FRIGIDA domain in tomatoes. Yeast two-hybrid and BiFC confirmed that SlBIN2 interacts with 4 SlFRLs, which are sub-cellularly localized in the nucleus. Tissue-specific expression of SlFRLs was observed highly in young roots and flowers. Biological results revealed that SlFRLs interact with SlBIN2 to regulate early flowering. Further, the mRNA level of SlBIN2 also increased in SlFRL-overexpressed lines. The relative expression of SlCPD increased upon SlFRL silencing, while SlDWF and SlBIN2 were decreased, both of which are important for BR signaling. Our research firstly provides molecular evidence that BRs regulate tomato flowering through the interaction between SlFRLs and SlBIN2. This study will promote the understanding of the specific pathway essential for floral regulation. Full article

In this study, effects of smoking on colour vision with the Farnsworth–Munsell 100 Hue test (FM100h) and achromatic (A), red-green (RG), and blue-yellow (BY) contrast sensitivity functions were evaluated. In total, 50 non-smoker controls and 25 smokers, divided into two groups (group 1, less than 10 cigarettes per day, with 15 patients, and group 2, >10 cigarettes per day, with 10 patients) took part in the experiments. Best-corrected visual acuity (BCVA), FM100h, and A, RG, and BY contrast sensitivity functions were measured. Total and partial RG and BY error scores (TES and PTES) and colour axis index (CA) were used in the analysis. No differences between smoker and non-smoker groups were found in BCVA, CA and A and BY contrast sensitivity, but TES and PTES values and RG contrast sensitivity at 1 cpd were statistically different. Differences between smoker groups were not significant. Error scores in smokers were positively correlated with the number of cigarettes smoked per day, and in BY also with age. Tobacco caused discrimination losses in both chromatic mechanisms but affected the red-green pathway more than the blue-yellow, and therefore, a partial RG score of FM100h test seems to be a good predictor of smoker colour deficiencies. Full article

Coproheme decarboxylase (ChdC) is an important enzyme in the coproporphyrin-dependent pathway (CPD) of Gram-positive bacteria that decarboxylates coproheme on two propionates at position 2 and position 4 sequentially to generate heme b by using H₂O₂ as an oxidant. This work focused on the ChdC from Geobacillus stearothermophilus (GsChdC) to elucidate the mechanism of its sequential two-step decarboxylation of coproheme. The models of GsChdC in a complex with substrate and reaction intermediate were built to investigate the reorienting mechanism of harderoheme. Targeted molecular dynamics simulations on these models validated that harderoheme is able to rotate in the active site of GsChdC with a 19.06-kcal·mol⁻¹ energy barrier after the first step of decarboxylation to bring the propionate at position 4 in proximity of Tyr145 to continue the second decarboxylation step. The harderoheme rotation mechanism is confirmed to be much easier than the release–rebinding mechanism. In the active site of GsChdC, Trp157 and Trp198 comprise a “gate” construction to regulate the clockwise rotation of the harderoheme. Lys149 plays a critical role in the rotation mechanism, which not only keeps the Trp157–Trp198 “gate” from being closed but also guides the propionate at position 4 through the gap between Trp157 and Trp198 through a salt bridge interaction. Full article

Genome-wide association studies (GWAS) have identified and reproduced thousands of diseases associated loci, but many of them are not directly interpretable due to the strong linkage disequilibrium among variants. Transcriptome-wide association studies (TWAS) incorporated expression quantitative trait loci (eQTL) cohorts as a reference panel to detect associations with the phenotype at the gene level and have been gaining popularity in recent years. For nicotine addiction, several important susceptible genetic variants were identified by GWAS, but TWAS that detected genes associated with nicotine addiction and unveiled the underlying molecular mechanism were still lacking. In this study, we used eQTL data from the Genotype-Tissue Expression (GTEx) consortium as a reference panel to conduct tissue-specific TWAS on cigarettes per day (CPD) over thirteen brain tissues in two large cohorts: UK Biobank (UKBB; number of participants (N) = 142,202) and the GWAS & Sequencing Consortium of Alcohol and Nicotine use (GSCAN; N = 143,210), then meta-analyzing the results across tissues while considering the heterogeneity across tissues. We identified three major clusters of genes with different meta-patterns across tissues consistent in both cohorts, including homogenous genes associated with CPD in all brain tissues; partially homogeneous genes associated with CPD in cortex, cerebellum, and hippocampus tissues; and, lastly, the tissue-specific genes associated with CPD in only a few specific brain tissues. Downstream enrichment analyses on each gene cluster identified unique biological pathways associated with CPD and provided important biological insights into the regulatory mechanism of nicotine dependence in the brain. Full article

Ultraviolet (UV) irradiation of the skin is related to the development of skin cancer. UVB also causes DNA damage in the form of cyclobutane pyrimidine dimers (CPDs), which can result in stable mutations. Toll-like receptor 4 (TLR4), a component of innate immunity, plays a key role in cancer. Previous studies from our laboratory have observed that TLR4 deficiency resulted in the repair of UVB-induced DNA damage, inhibition of UVB-induced immune suppression, and carcinogenesis. In this study, we determined the efficacy of TLR4 antagonist TAK-242 in regulation of UVB-induced DNA damage, inflammation, and tumor development. Our results indicate that TAK-242 treatment increased the expression of xeroderma pigmentosum group A (XPA) mRNA, resulting in the repair of UVB-induced CPDs in skin of SKH-1 mice. Treatment with TAK-242 also inhibited the activation of NLR family pyrin domain containing 3 (NLRP3) in UVB-exposed skin of SKH-1 mice. Cutaneous carcinogenesis was significantly reduced in mice treated with TAK-242 in comparison to vehicle-treated mice. The proinflammatory cytokines IL-1β, IL-6, and TNF-α were also found to be significantly greater in vehicle-treated mice than TAK-242-treated mice. Finally, treatment with TAK-242 augmented anti-tumor immune responses in mice. Our data provide further evidence that activation of the TLR4 pathway promotes the development of UV-induced non-melanoma skin cancer mediated at least in part on its negative effects on DNA damage. Moreover, treatment with the TLR4 inhibitor TAK-242 may be effective for prevention of skin cancer. Full article

Brassinosteroids (BRs) play key roles in diverse plant growth processes through a complex signaling pathway. Components orchestrating the BR signaling pathway include receptors such as kinases, transcription factors, protein kinases and phosphatases. The proper functioning of the receptor kinase BRI1 and the transcription factors BES1/BZR1 depends on their dephosphorylation by type 2A protein phosphatases (PP2A). In this work, we report that an additional phosphatase family, type one protein phosphatases (PP1), contributes to the regulation of the BR signaling pathway. Co-immunoprecipitation and BiFC experiments performed in Arabidopsis plants overexpressing durum wheat TdPP1 showed that TdPP1 interacts with dephosphorylated BES1, but not with the BRI1 receptor. Higher levels of dephosphorylated, active BES1 were observed in these transgenic lines upon BR treatment, indicating that TdPP1 modifies the BR signaling pathway by activating BES1. Moreover, ectopic expression of durum wheat TdPP1 lead to an enhanced growth of primary roots in comparison to wild-type plants in presence of BR. This phenotype corroborates with a down-regulation of the BR-regulated genes CPD and DWF4. These data suggest a role of PP1 in fine-tuning BR-driven responses, most likely via the control of the phosphorylation status of BES1. Full article