P65 Protein Research Articles

Ferritin nanoparticles significantly enhance the immune response to the African swine fever virus p34 protein.

African swine fever (ASF) is a highly contagious disease, and the core-shell protein p34 is an important antigen that can induce immune responses. The use of ferritin nanoparticles for the orderly and repetitive display of antigens on the particle surface can improve the immunogenicity of subunit vaccines. Here, we used the SpyCatcher/Spytag system to conjugate ferritin nanoparticles with the p34 protein (F-p34). The N-terminus of ferritin was conjugated to a truncated SpyCatcher (SC-ferritin). The antigen of ASFV p34 was linked to SpyTag (p34-ST). SC-Ferritin and p34-ST were expressed in E. coli and purified via CaptoTM Core 700 and Ni-NTA columns, respectively. Based on the isopeptide bonds formed between SpyCatcher and SpyTag, p34 linked to SpyTag was readily surface-displayed on SC-ferritin via in vitro covalent conjugation (F-p34). F-p34 can be taken up by bone marrow dendritic cells (BMDCs) and effectively stimulate BMDC maturation. Compared with the monomeric p34 protein, in vivo studies confirmed that the recombinant F-p34 nanoparticle can induce more robust T-cell responses and stronger specific IgG antibody responses against ASFV. Moreover, F-p34 can increase serum cytokines, which is also significantly greater than that of the p34 protein. These results indicate that the recombinant nanoparticles can induce not only humoral immune responses but also cellular immune responses. In addition, there were no significant pathological changes in the heart, liver, spleen, lung or kidney tissue sections of the mice immunized with F-p34, demonstrating that the recombinant nanoparticles exhibit favorable histocompatibility or safety. These results demonstrate that we successfully designed a recombinant plasmid and obtained a recombinant protein through the prokaryotic expression system of E. coli. The covalently coupled recombinant F-p34 nanoparticles significantly increased the antigenicity of p34 and contributed to research on African swine fever subunit vaccines.

Gastroprotective effect of fucoidan from Sargassum siliquastrum against ethanol-induced gastric mucosal injury.

The ethanol-induced BALB/c mice and human gastric epithelial cell (Ges-1 cell) models were used to investigate the Sargassum siliquastrum fucoidan (SFuc) gastroprotective capability. The injury score and histopathological sections of the stomach were used to evaluate the gastroprotective capability. The western blotting and RT-PCR methods determined the signaling mechanism of mice's gastric injury. SFuc is fucoidan with a molecular weight of 300.7 and 25.1kDa. The injury score and ulcer index of the SFuc-200 group decreased by 3.85 and 2.06 folds in contrast with the Model group, respectively. The findings indicated that SFuc reduced oxidative stress and inflammatory factor expression in the gastric mucosa by downregulating the levels of associated genes within the TLR-4, MyD88, and MAPK/NF-κB signaling pathways. Meanwhile, the SFuc-200 group promoted the expressions of EGF and PGE 2 by 1.53 and 1.52 folds, respectively. Together with the expression inhibition of p38, ERK, JNK, and NF-κB proteins in gastric tissue to help for differentiation of gastric cells. In addition, SFuc significantly reduced apoptosis occurrence in mice and Ges-1 cells. Our study provides potential mechanism clues of the SFuc's resistance to ethanol-induced gastric mucosal damage, suggesting its potential functional food for gastric protection.

Integrative pan-cancer genomic analysis highlights mitochondrial protein p32 as a potential therapeutic target in Myc-driven tumorigenesis.

Tumor metabolic reprogramming, particularly involving mitochondrial metabolism, is a hallmark of malignancy. The mitochondrial protein p32 (C1QBP) has emerged as a critical regulator in various cancers, frequently associated with poor patient prognosis. However, the role of p32 across different cancer types remains largely unexplored. Our bioinformatics analysis demonstrates that p32 is significantly overexpressed in several malignancies and is closely involved in multiple oncogenic pathways related to tumor progression and metabolic reprogramming. Moreover, p32 expression positively correlates with genomic heterogeneity and drug sensitivity. We identified a strong association between p32 and c-Myc in both normal and cancerous tissues. We confirmed that p32 is a direct transcriptional target of c-Myc, which upregulates p32 by binding to its promoter. Functional experiments established that p32 is crucial for MYC-driven tumorigenesis, with its knockdown or knockout inhibiting tumor proliferation and extending survival. Targeting p32 may inhibit MYC-driven tumorigenesis, highlighting its potential as a therapeutic target in MYC-driven cancers.

Comparative Proteomic Analysis of Popcorn Genotypes Identifies Differentially Accumulated Proteins Associated with Resistance Pathways to Southern Leaf Blight Disease

Southern leaf blight (SLB), caused by Bipolaris maydis, poses a significant threat to maize and popcorn production. To understand the molecular mechanisms underlying SLB resistance, we conducted a high-throughput proteomic analysis comparing SLB-resistant (L66) and SLB-susceptible (L51) popcorn genotypes at four and ten days after inoculation (DAI). A total of 717 proteins were identified, with 151 differentially accumulated proteins (DAPs) between the genotypes. Eighteen DAPs exhibited the same regulatory pattern in both the SLB-resistant and SLB-susceptible genotypes at four (R4/S4) and ten (R10/S10) DAI. The protein-protein interaction (PPI) network of differentially accumulated proteins (DAPs) linked to SLB resistance and susceptibility enriched specific metabolic pathways in the SLB response, including photosynthesis, ribosome, ascorbate and aldarate metabolism, glutathione metabolism, and carbon metabolism. Proteins such as photosystem II 11 kD protein (B4FRJ4, PSB27-1), which was up-regulated at both time points (R4/S4 and R10/S10), and 60S acidic ribosomal protein P0 (A0A1D6LEZ7, RPP0B), which was unique to the resistant genotype at both time points (R4 and R10), highlighted the importance of maintaining photosynthetic efficiency and protein synthesis during pathogen attack. Additionally, dehydroascorbate reductase like-3 (B4F817, DHAR3) was consistently up-regulated at both time points in resistant genotypes, emphasizing its role in redox balance and ROS detoxification. In contrast, glyceraldehyde-3-phosphate dehydrogenase (K7UGF5, GAPC2), a glycolytic enzyme, was unique to the susceptible genotype, suggesting its involvement in managing energy metabolism under stress conditions. Our findings suggest that resistance to SLB in popcorn involves a combination of enhanced photosynthetic repair, redox homeostasis, and ribosomal protein activity, providing new potential molecular targets, such as DHAR3 and RPP0B, for genetic improvement in SLB resistance. These results offer valuable insights into breeding programs aimed at developing SLB-resistant popcorn varieties.

Klotho supplementation decreases blood pressure and albuminuria in mice with lupus nephritis.

Klotho deficiency is prevalent in various chronic kidney diseases. Although klotho is known to bind transforming growth factor β (TGFβ) receptor 1 to antagonize renal fibrosis, TGFβ also maintains regulatory T cells with inducing forkhead box protein P3 (FOXP3). Female New Zealand Black/White F1 (NZBWF1) mice were divided into two groups (n=10 for each): one group was treated with daily subcutaneous injection of klotho protein (30μg/kg/day) for 8 weeks, and the other only received vehicle. Klotho supplementation suppressed blood pressure, 8-epi-prostaglandin F2α excretion, albuminuria, and renal angiotensin II levels (p<0.05 for all) without affecting the glomerular filtration rate (GFR) in NZBWF1 mice. Klotho protein supplementation reduced the number of cluster of differentiation (CD)4+FOXP3+ T cells (p<0.05) without altering the anti-DNA antibody levels. Klotho supplementation augmented glomerular cellularity, but decreased glomerular crescent formation and interstitial fibrosis in NZBWF1 mice (p<0.05). Klotho protein supplementation inactivated renal renin-angiotensin system, ameliorating blood pressure and albuminuria in NZBWF1 mice. Klotho supplementation hampered regulatory T cells without altering autoantibodies, exerting dual effects on glomerular pathology in NZBWF1 mice without changes in GFR.

First report of Gulupa baciliform virus A associated with passion fruit in China.

Passion fruit (Passiflora edulis) is a commercially important crop known for its nutritional value, high antioxidant content, and use in beverages and desserts. Gulupa baciliform virus A (GBVA), tentatively named Badnavirus in the family Caulimoviridae, is a cryptic circular double-stranded DNA (dsDNA, ≈6,951 bps) virus recently reported in Colombia with asymptomatic infection of passion fruit (Sepúlveda et al. 2022). In July 2024, viral-like symptoms, including vein clearing, mottling, leaf distortion, and yellowing, were observed on passion fruit leaves in Pingtang, Guizhou, China (Supplementary figure S1), with a 30% incidence rate, and 30 samples were collected. Total RNA was extracted from these samples using TRIzol reagent (Invitrogen, CA, USA). The intact RNA from the two pools (each consisting of 14 samples) was subjected to library construction using TruseqTM Small RNA sample prep Kit (Illumina) followed by Hiseq sequencing (Hiseq2000 Truseq SBS Kit v3-HS, Illumina platform) to complete the sRNA sequencing. Approximately 12,923,899 and 10,877,466 raw while 10,135,256 and 4,184,706 clean reads were obtained for the samples in pool A and pool B, respectively. Next, the de novo assembly was carried out using metaSPAdes (Nurk et al., 2017) which resulted in 4,770 virus related trimmed contigs with the maximum length of 883 bp. BLASTn searches of sequence contigs against the NCBI viral genome database confirmed the presence of telosma mosaic virus (TEMV, Hanoi isolate, accession No. NC_009742, >96.0% nucleotide identity), East Asian passiflora virus (EAPV, Taiwan isolate, accession No. KP114137, >97.5% nucleotide identity), and papaya leaf curl Guangdong virus (PaLCuGdV, Fujian isolate, accession No. FJ495184, >98.0% nucleotide identity). Interestingly, more than 70 sequence contigs revealed >98.7% nucleotide identity with the GBVA Colombia isolate (accession No. MW393828). Whereas, the overall similarity of newly identified GBVA Guizhou isolate was calculated to be more than 99% with corresponding sequences in NCBI database. Moreover, out of 4,770 trimmed contigs, the mapping of 241 contigs to the reference genome sequence (accession No. MW393828) using Geneious R9 further confirmed the presence of GBVA. To validate the Hiseq sequencing results, total RNA was extracted from the same samples used for high throughput sequence (HTS) analysis, and reverse-transcription polymerase chain reaction (RT-PCR) was performed using specific primers (GBVA-P2-F1: 5´-ATGAATTGGAAATCAACYGTAG-3´/GBVA-P2-R1: 5´-TTCTTCTCGAGGCGTCTCTT-3´) targeting the p2 protein region of the reference genome (accession No. MW393828) (Sepúlveda et al. 2022). Amplicon of expected size of 408bp was subjected to Sanger sequencing, and the resulted sequence (GBVA, Guizhou isolate) was submitted to GenBank (accession No. PQ240853). Multiple sequence alignment was executed based on the p2 protein sequence of GBVA in MEGA X with the ClustalW program. Subsequently, the phylogenetic tree was constructed employing the maximum likelihood method (Supplementary figure S1) (Kumar et al. 2018). Phylogenetic analysis showed that the newly identified isolate has highest similarity with the corresponding isolates from Colombia, indicating the close origin of Colombian and Chinese isolates. To our knowledge, this is the first report of GBVA associated with passion fruit in China, which indicates that further research into it's exact symptom expression may be warranted for global passion fruit production.

Executioner caspases degrade essential mediators of pathogen-host interactions to inhibit growth of intracellular Listeria monocytogenes

Cell death mediated by executioner caspases is essential during organ development and for organismal homeostasis. The mechanistic role of activated executioner caspases in antibacterial defense during infections with intracellular bacteria, such as Listeria monocytogenes, remains elusive. Cell death upon intracellular bacterial infections is considered altruistic to deprive the pathogens of their protective niche. To establish infections in a human host, Listeria monocytogenes deploy virulence mediators, including membranolytic listeriolysin O (LLO) and the invasion associated protein p60 (Iap), allowing phagosomal escape, intracellular replication and cell-to-cell spread. Here, by means of chemical and genetical modifications, we show that the executioner caspases-3 and -7 efficiently inhibit growth of intracellular Listeria monocytogenes in host cells. Comprehensive proteomics revealed multiple caspase-3 substrates in the Listeria secretome, including LLO, Iap and various other proteins crucially involved in pathogen-host interactions. Listeria secreting caspase-uncleavable LLO or Iap gained significant growth advantage in epithelial cells. With that, we uncovered an underappreciated defense barrier and a non-canonical role of executioner caspases to degrade virulence mediators, thus impairing intracellular Listeria growth.

Concurrent inhibition of p300/CBP and FLT3 enhances cytotoxicity and overcomes resistance in acute myeloid leukemia.

FMS-like tyrosine kinase-3 (FLT3), a class 3 receptor tyrosine kinase, can be activated by mutations of internal tandem duplication (FLT3-ITD) or point mutations in the tyrosine kinase domain (FLT3-TKD), leading to constitutive activation of downstream signaling cascades, including the JAK/STAT5, PI3K/AKT/mTOR and RAS/MAPK pathways, which promote the progression of leukemic cells. Despite the initial promise of FLT3 inhibitors, the discouraging outcomes in the treatment of FLT3-ITD-positive acute myeloid leukemia (AML) promote the pursuit of more potent and enduring therapeutic approaches. The histone acetyltransferase complex comprising the E1A binding protein P300 and its paralog CREB-binding protein (p300/CBP) is a promising therapeutic target, but the development of effective p300/CBP inhibitors faces challenges due to inherent resistance and low efficacy, often exacerbated by the absence of reliable clinical biomarkers for patient stratification. In this study we investigated the role of p300/CBP in FLT3-ITD AML and evaluated the therapeutic potential of targeting p300/CBP alone or in combination with FLT3 inhibitors. We showed that high expression of p300 was significantly associated with poor prognosis in AML patients and positively correlated with FLT3 expression. We unveiled that the p300/CBP inhibitors A485 or CCS1477 dose-dependently downregulated FLT3 transcription via abrogation of histone acetylation in FLT3-ITD AML cells; in contrast, the FLT3 inhibitor quizartinib reduced the level of H3K27Ac. Concurrent inhibition of p300/CBP and FLT3 enhanced the suppression of FLT3 signaling and H3K27 acetylation, concomitantly reducing the phosphorylation of STAT5, AKT, ERK and the expression of c-Myc, thereby leading to synergistic antileukemic effects both in vitro and in vivo. Moreover, we found that p300/CBP-associated transcripts were highly expressed in quizartinib-resistant AML cells with FLT3-TKD mutation. Targeting p300/CBP with A485 or CCS1477 retained the efficacy of quizartinib, suggesting marked synergy when combined with p300/CBP inhibitors in quizartinib-resistant AML models, as well as primary FLT3-ITD+ AML samples. These results demonstrate a potential therapeutic strategy of combining p300/CBP and FLT3 inhibitors to treat FLT3-ITD and FLT3-TKD AML.

Combination effects of Pistachio hull and carfilzomib on NF-κB p65, MDR1, MRP1, and Caspase3 gene expression in breast cancer cell line

ObjectiveThis study aimed to investigate the synergistic effects of the chemotherapy drug Carfilzomib (CFZ) and Pistachio hull extract on the SK-BR3 breast cancer cell line.MethodsIn this experimental study, we evaluated the effect of Pistachio hull extract and CFZ as standalone treatments on cell viability using the MTT assay at 24- and 48-hours post-treatment. Following this, we conducted combination therapy analyses to assess the potential synergistic relationship between Pistachio hull extract and CFZ after 24- and 48-hours of treatment on both the SK-BR3 breast cancer cell line and the MCF10A normal cell line. We utilized real-time PCR to measure the expression levels of MDR1, MRP1, NF-κB p65, and Caspase3 genes. Additionally, the NF-κB p65 transcription factor was evaluated using ELISA after 24- and 48-hours.ResultsThe MTT assay revealed IC50 values of 2.014 mg/mL and 1.031 mg/mL in the SK-BR3 cell line, and 3.265 mg/mL and 2.994 mg/mL in the MCF10A cell line at 24- and 48-hours post-treatment with Pistachio hull extract. CFZ concentrations of 0.181 × 10− 3 mg/mL and 0.0057 × 10− 3 mg/mL in the SK-BR3 cell line, as well as 5.54 × 10− 3 mg/mL and 2.51 × 10− 3 mg/mL in the MCF10A cell line, inhibited growth by up to 50%. The analysis of combination therapy indicated a synergistic effect between the two treatments after both 24- and 48-hours of exposure. Real-time PCR results demonstrated significant alterations in the expression of MDR1, MRP1, NF-κB p65, and Caspase3 genes, along with changes in NF-κB p65 protein levels in both cell lines following treatment with Pistachio hull extract, CFZ, or their combination compared to the control group (p < 0.05).ConclusionThe findings highlight the effectiveness of CFZ as a proteasome inhibitor when used in conjunction with Pistachio hull extract in breast cancer cell lines. Therefore, both CFZ and Pistachio hull extract, whether administered alone or in combination, represent promising molecular targets for breast cancer treatment.

Borrelia burgdorferi tolerates alteration to P66 porin function in a murine infectivity model.

Borrelia burgdorferi exists in a complex enzootic life cycle requiring differential gene regulation. P66, a porin and adhesin, is upregulated and essential during mammalian infection, but is not produced or required within the tick vector. We sought to determine whether the porin function of P66 is essential for infection. Vancomycin treatment of B. burgdorferi cultures was used to screen for P66 porin function and found to generate spontaneous mutations in p66 (bb0603). Three novel, spontaneous, missense P66 mutants (G175V, T176M, and G584R) were re-created by site-directed mutagenesis in an infectious strain background and tested for infectivity in mice by ID50 experiments. Two of the three mutants retained infectivity comparable to the isogenic control, suggesting that B. burgdorferi can tolerate alteration to P66 porin function during infection. The third mutant exhibited highly attenuated infectivity and produced low levels of P66 protein. Interestingly, four isolates that were recovered for p66 sequencing from mouse tissues revealed novel secondary point mutations in genomic p66. However, these secondary mutations did not rescue P66 porin function. New structural modeling of P66 is presented and consistent with these experimental results. This is the first work to assess the contribution of P66 porin function to B. burgdorferi pathogenesis.

HPV E6/E7-Induced Acetylation of a Peptide Encoded by a Long Non-Coding RNA Inhibits Ferroptosis to Promote the Malignancy of Cervical Cancer.

Although a fraction of functional peptides concealed within long non-coding RNAs (lncRNAs) is identified, it remains unclear whether lncRNA-encoded peptides are involved in the malignancy of cervical cancer (CC). Here, a 92-amino acid peptide is discovered, which is named TUBORF, encoded by lncRNA TUBA3FP and highly expressed in CC tissues. TUBORF inhibits ferroptosis to promote the malignant proliferation of CC cells. Mechanistically, human papillomavirus (HPV) oncogenes E6 and E7 upregulate TUBORF through CREB-binding protein (CBP)/E1A-binding protein p300 (p300)-mediated histone H3 lysine 27 acetylation (H3K27ac) of lncTUBA3FP enhancer. Furthermore, E6 and E7 elevate and recruit acetyltransferase establishment of sister chromatid cohesion N-acetyltransferase 1 (ESCO1) to bind to and acetylate TUBORF, which facilitates the degradation of immunity-related GTPase Q (IRGQ) via a ubiquitin-proteasome pathway, resulting in the inhibition of ferroptosis and promotion of the malignant proliferation of CC cells. Importantly, silencing ESCO1 or TURORF amplifies anticancer effects by paclitaxel both in CC cells and in vivo. These novel findings reveal oncopeptide TUBORF and its acetyltransferase ESCO1 as important regulators of ferroptosis and tumorigenesis during cervical cancer pathogenesis and establish the scientific basis for targeting these molecules for treating CC.

Two Novel Peptides from Buthotus saulcyi Scorpion Venom: Proteomic Analysis and Approaches

Background and Objectives: Venomous scorpions play a crucial role in medicine and public health. Buthotus saulcyi scorpion is known as one of the most populous species in East Asia and Iran, while its venom proteome has still not been fully determined. background: In the present study, we report for the first time a proteomic profile of Buthotus saulcyi scorpion venom with the aim of looking ahead and determining the structural and functional characteristics of these compounds for use as medical tools. Aims: In the current research, the proteomic profile of Buthotus saulcyi scorpion venom to determine the structural and functional characteristics of its compounds used for treatment will be examined for the first time. objective: In the present study, we report for the first time a proteomic profile of Buthotus saulcyi scorpion venom with the aim of looking ahead and determining the structural and functional characteristics of these compounds for use as medical tools. Method:: 2D-PAGE, HPLC, SDS-PAGE, sequencing, and MALDI-TOF MS techniques were used to investigate the properties of these peptides. Result: The 2D-PAGE analysis of crude toxin from B. saulcyi revealed a minimum of 96 protein spots, with isoelectric points ranging from 4 to 9 and molecular weights spanning from 3.6 to 205 kDa. Following this, HPLC was used to isolate 14 fractions of crude toxins, and the protein content of these fractions was measured. SDS-PAGE analysis identified 7 protein bands within the B. saulcyi crude toxin fractions, with molecular weights ranging from 13 to 217 kDa. Further examination of fraction 7 through amino acid sequencing resulted in the identification of two protein bands labeled peptide 3 and peptide 4. Ultimately, these protein bands were extracted, and their molecular mass and amino acid sequences were analyzed using MALDI-TOF MS. result: 2D-PAGE data from the B. saulcyi crude toxin showed at least 96 protein spots in isoelectric point between 4 and 9 and a molecular weight between 3.6 and 205 kDa. Then 14 fractions of crude toxins were isolated using HPLC and their protein content was estimated. SDS-PAGE analysis of fractions of B. saulcyi crude toxin showed 7 protein bands with a molecular weight between 13 and 217 kDa. Subsequently, the amino acid sequencing of Fraction 7 led us to two protein bands designated as Peptide 3 and Peptide 4 peptides. Finally, these protein bands were removed, and molecular mass and amino acid sequence analysis was performed using MALDI-TOF MS. Conclusion:: According to our results, the alignment of P3 and P4 protein sequences revealed the highest similarity to chrysophsin 2 and pheromone-bound protein 2, respectively.

Dose-Dependent Effect of a New Biotin Compound in Hippocampal Remyelination in Rats.

Demyelination is commonly observed in neurodegenerative disorders, including multiple sclerosis (MS). Biotin supplementation is known to stabilize MS progression. To reduce the effective dose of biotin, we synthesized a new and superior form of biotin, a complex of magnesium ionically bound to biotin (MgB) and compared its dose-dependent effect with biotin alone after inducing demyelination using lysolecithin (LPC) in rats. Myelination was assessed using luxol fast blue staining and immunostaining against MBP protein, revealing that the most significant remyelination occurred in the MgB groups. Additionally, both biotin and MgB-treated animals showed dose-dependent improvements in spatial memory. Moreover, we detected a decrease in inflammatory proteins in both treatment groups, which was more prominent in high-dose MgB-treated animals and correlated with decreased expression of NF-κB p65, OP, and MMP-9 proteins. Further analysis of biotin-related proteins demonstrated that both biotin and, notably, MgB reversed the demyelination-dependent reduction of these proteins. Furthermore, biotin, particularly MgB, improved neuronal transmission proteins, Synapsin-1, PSD-93, and PSD-95. Additionally, both treatment groups exhibited increased BDNF, GAP43, and ICAM levels, with significant increments observed in high-dose MgB-treated animals. Increased GFAP, indicative of reactive gliosis, was observed in LPC-treated animals, and this effect was notably reversed by high-dose MgB treatment. The current data emphasize the dose-dependent beneficial effect on the remyelination process. Furthermore, the combination of biotin with Mg resulted in a more potent effect compared to biotin by itself. The strong influence of MgB encourages proof-of-concept studies using MgB in patients with MS.

Mechanism of dexmedetomidine in brain injury of infant rats via the IRE1α/NF-κB/CHOP pathway

Objective We investigated the mechanism of Dexmedetomidine (Dex) in infant rats with brain injury. Methods The infant rats underwent brain injury modelling. The motor function, spatial learning and memory abilities in rats, and the hippocampal CA1 region Nissl body level and apoptosis were evaluated by behavioural tests and histological stainings. Levels of the hippocampal CA1 region p-IRE1α, nuclear/cytoplasmic p65, CHOP, Bax and Bcl-2 proteins were determined by Western blot. Results Propofol anaesthesia caused brain injury in infant rats. Dex increased the hippocampal CA1 region Nissl body level, abated cell apoptosis, reduced p-IRE1α, ATF6, p-PERK/PERK and CHOP levels, decreased the Bax protein level, elevated the Bcl-2 protein level, and alleviated brain injury in infant rats. After ERS induction and the NF-κB pathway inhibition, the hippocampal CA1 region nuclear/cytoplasmic p65 ratio, CHOP level, and apoptosis were reduced in infant rats with brain injury treated with Dex, while the learning and memory abilities of rats were enhanced. Conclusion Dex reduced the hippocampal CA1 region cell apoptosis and enhanced learning and memory abilities by inhibiting the ERS-mediated IRE1α/NF-κB/CHOP pathway, thereby alleviating brain injury in infant rats.

Genomic, Evolutionary, and Pathogenic Characterization of a New Polerovirus in Traditional Chinese Medicine Viola philippica.

Viola philippica, a medicinal herbaceous plant documented in the Chinese Pharmacopoeia, is a promising candidate for research into plant-derived pharmaceuticals. However, the study of newly emerging viruses that threaten the cultivation of V. philippica remains limited. In this study, V. philippica plants exhibiting symptoms such as leaf yellowing, mottled leaves, and vein chlorosis were collected and subjected to RNA sequencing to identify potential viral pathogens. A novel polerovirus, named Viola Philippica Polerovirus (VPPV), was identified in V. philippica. VPPV possesses a linear, positive-sense, single-stranded RNA genome consisting of 5535 nucleotides (nt) and encodes seven highly overlapping open reading frames (ORFs). Two potential recombination events were identified within ORF2, ORF3a, and ORF3, providing insights into the genetic diversity and evolution history of this novel polerovirus. An infectious cDNA clone of VPPV was successfully constructed and shown to infect Nicotiana benthamiana. Using a PVX-based heterologous expression system, the VPPV P0 protein was shown to trigger a systemic hypersensitive response (HR)-like reaction in N. benthamiana, indicating that P0 functions as the main pathogenicity determinant. These findings contributed to the detection and understanding of pathogenic mechanisms and control strategies for VPPV in V. philippica.

Selective Gamma-Secretase Inhibition by CHF5074 Attenuates Inflammation and Neovascularization in a Murine Model of Choroidal Neovascularization

Purpose Chronic inflammation plays an important role in the pathogenesis of choroidal neovascularization (CNV). This study aimed to investigate the effect of the CHF5074, a γ-secretase inhibitor, on angiogenesis in a laser-induced CNV model and elucidate its possible molecular mechanism. Methods Male C57/BL6J mice aged between 6 to 8 weeks were employed to set up a laser-induced model of CNV. Then, CHF5074 was injected intraperitoneally on the day after laser modeling, as well as on the second, third, and fourth days. Immunofluorescence staining was used to evaluate the retinal and choroidal complex. The markers used were CD31 for neovascularization and IBA1 for microglia staining in ocular tissue slices. Fundus fluorescein angiography on days 3d, 7d, and 14d analyzed neovascularization and leakage areas. Inflammatory indicators were examined by Western blot (WB) and enzyme-linked immunosorbent assay (ELISA). High-throughput whole-tissue sequencing of retinal choroids identified relevant cell pathways. Key regulatory factors modulated by CHF5074 were identified via WB. Co-culture of BV2 cells and human umbilical vein endothelial cells (HUVEC) were used to explore the function of CHF5074 on the inhibition of tube formation. Results CHF5074 significantly decreased CD31 expression in the choroid on 3d, 7d, and 14d post-laser modeling (p < 0.05) and decreased both neovascularization and leakage areas (p < 0.05). Additionally, CHF5074 significantly lowered TNF-α, IL-10, and IL-1β expression levels in the choroid (p < 0.05), as demonstrated by WB analysis and ELISA. High-throughput whole-tissue sequencing identified P38-MAPK, JNK, and Wnt signaling pathways associated with neovascularization. CHF5074 decreased P38 protein phosphorylation (p < 0.05) as confirmed by WB analysis. CHF5074 inhibited the tube formation of HUVECs co-cultured with LPS and ATP-treated BV2 cells. Conclusion CHF5074 significantly suppresses angiogenesis in laser-induced choroidal neovascularization models, suggesting its potential as a novel agent for preventing and treating CNV.

Differential roles of human CD4+ and CD8+ regulatory T cells in controlling self-reactive immune responses

Here we analyzed the relative contributions of CD4+ regulatory T cells expressing Forkhead box protein P3 (FOXP3) and CD8+ regulatory T cells expressing killer cell immunoglobulin-like receptors to the control of autoreactive T and B lymphocytes in human tonsil-derived immune organoids. FOXP3 and GZMB respectively encode proteins FOXP3 and granzyme B, which are critical to the suppressive functions of CD4+ and CD8+ regulatory T cells. Using CRISPR–Cas9 gene editing, we were able to achieve a reduction of ~90–95% in the expression of these genes. FOXP3 knockout in tonsil T cells led to production of antibodies against a variety of autoantigens and increased the affinity of influenza-specific antibodies. By contrast, GZMB knockout resulted in an increase in follicular helper T cells, consistent with the ablation of CD8+ regulatory T cells observed in mouse models, and a marked expansion of autoreactive CD8+ and CD4+ T cells. These findings highlight the distinct yet complementary roles of CD8+ and CD4+ regulatory T cells in regulating cellular and humoral responses to prevent autoimmunity.

Mycoplasma pneumoniae MPN606 induces inflammation by activating MAPK and NF-κB signaling pathways.

Mycoplasma pneumoniae (M. pneumoniae) is one of the major pathogens causing community-acquired pneumonia (CAP), and its pathogenic mechanism is not fully understood. Inflammatory response is the most basic and common pathological phenomenon of CAP, but the specific mechanism needs further investigation. In this study, the inflammatory action of M. pneumoniae MPN606 protein was confirmed and its molecular mechanism was tentatively investigated. Compared with the control group treated with PBS, stimulation of RAW264.7 cells with rMPN606 can promote the release of NO, increase the expression level of TNF-α and IL⁃6 cytokines, and up-regulate the mRNA transcription levels of iNOS, IL-6 and TNF-α in RAW264.7 cells. In addition, rMPN606 also significantly induced the expression of iNOS protein in RAW264.7 cells, resulting in increased phosphorylation levels of p65, p38 and ERK proteins. The results of cellular immunofluorescence showed that NF-κB was transferred from cytoplasm to nucleus of RAW264.7 cells after stimulation with rMPN606, and nuclear translocation of NF-κB was significantly enhanced. These results indicate that Mycoplasma pneumoniae MPN606 induces M1-type activation of macrophages and secretes pro-inflammatory factors by activating NF-κB and MAPK pathways.

A PDMS/chitosan/MPMs composite film based on multi-field coupling enhancement for African swine fever virus P72 protein detection.

African swine fever (ASF) is an acute hemorrhagic disease in pigs caused by the African swine fever virus (ASFV), which has a high mortality rate and brought great damage to global pig farming industry. At present, there is no effective treatment or vaccine to combat ASFV infection, so early detection of ASFV has become particularly important. Therefore, the PDMS/chitosan/MPMs composite film was proposed to detect ASFV P72. PDMS mixed with chitosan particles were used to provide biological modification sites for the ASFV P72 antibody, which simplified the functional modification process. The specific binding of antigen and antibody leads to the mechanical deformation of the composite film, which was observedthrough optical method tochange the microsphere spacing. The polystyrene microsphere (PMs) spacing changes were acquired by simply measuring the diameters of the Debye ring diffracted by the PDMS/chitosan/PMs with a simple laser pointer. The conductive layer formed by AgNWs realize the transformation of biological signals to electrical signals. By conjugating PMs with Fe3O4 nanoparticles (MPMs) and applying a magnetic field, a magnetic-stress-electric coupling system has been constructed, which amplifies the deformation of the composite film and enhances the sensitivity of the flexible film. The experimental results showed that the flexible film has wide linear range of 10ng/mL-100μg/mL, and the optimal detection limit (LOD) is 0.059ng/mL, which was aboutone order of magnitude lower than that without magnetic field. In addition, the flexible film showed a good stability, specificity, and selectivity.

Anti-Inflammatory and Anti-Migratory Effects of Morin on Non-Small-Cell Lung Cancer Metastasis via Inhibition of NLRP3/MAPK Signaling Pathway.

Non-small-cell lung cancer (NSCLC) remains the leading cause of cancer-related deaths globally, with a persistently low five-year survival rate of only 14-17%. High rates of metastasis contribute significantly to the poor prognosis of NSCLC, in which inflammation plays an important role by enhancing tumor growth, angiogenesis, and metastasis. Targeting inflammatory pathways within cancer cells may thus represent a promising strategy for inhibiting NSCLC metastasis. This study evaluated the anti-inflammatory and anti-metastatic properties of morin, a bioactive compound derived from a Thai medicinal herb, focusing on its effects on NLRP3 inflammasome-mediated pathways in an in vitro NSCLC model. The A549 and H1299 cell lines were stimulated with lipopolysaccharide (LPS) and adenosine triphosphate (ATP) to activate the NLRP3 pathway. The inhibition effects exhibited by morin in reducing pro-inflammatory secretion in LPS- and ATP-stimulated NSCLC cells were assessed by ELISA, while wound healing and trans-well invasion assays evaluated its impact on cell migration and invasion. RT-qPCR measurement quantified the expression of inflammatory genes, and zymography and Western blotting were used to examine changes in invasive protein levels, epithelial-to-mesenchymal transition (EMT) markers, and underlying molecular mechanisms. Our findings demonstrated the significant ability of morin to decrease the production of IL-1β, IL-18, and IL-6 in a dose-dependent manner (p < 0.05), as well as suppress NSCLC cell migration and invasion. Morin downregulated invasive proteins (MMP-2, MMP-9, u-PAR, u-PA, MT1-MMP) and EMT markers (fibronectin, N-cadherin, vimentin) (p < 0.01) while also reducing the mRNA levels of NLRP3, IL-1β, IL-18, and IL-6. Mechanistic investigations revealed that morin suppressed NLRP3 inflammasome activity and inactivated MAPK pathways. Specifically, it decreased the expression of NLRP3 and ASC proteins and reduced caspase-1 activity, while reducing the phosphorylation of ERK, JNK, and p38 proteins. Collectively, these findings suggest that morin's inactivation of the NLRP3 inflammasome pathway could offer a novel therapeutic strategy for counteracting pro-tumorigenic inflammation and metastatic progression in NSCLC.