Search Results (801)

Daphnis nerii cypovirus-23 (DnCPV-23) is a new type of cypovirus that has a lethal effect on many species of Sphingidae pests. DnCPV-23 can replicate in Spodoptera frugiperda Sf9 cells, but the replication characteristics of the virus in this cell line are still unclear. To determine the replication characteristics of DnCPV-23 in Sf9 cells, uninfected Sf9 cells and Sf9 cells at 24 and 72 h after DnCPV-23 infection were collected for transcriptome analysis. Compared to uninfected Sf9 cells, a total of 188 and 595 differentially expressed genes (DEGs) were identified in Sf9 cells collected at 24 hpi and 72 h, respectively. KEGG analyses revealed that 139 common DEGs in two treatment groups were related to nutrition and energy metabolism-related processes, cell membrane integrity and function-related pathways, detoxification-related pathways, growth and development-related pathways, and so on. We speculated that these cellular processes might be manipulated by viruses to promote replication. This study provides an important basis for further in-depth research on the mechanism of interaction between viruses and hosts. It provides additional basic information for the future exploitation of DnCPV-23 as a biological insecticide. Full article

Background/Objectives: Myelodysplastic syndrome (MDS) is a heterogeneous clonal hematopoietic disorder characterized by ineffective hematopoiesis and leukemic transformation risk. Current therapies show limited efficacy, with ~50% of patients failing hypomethylating agents. This review aims to synthesize recent discoveries through an integrated network model and examine translation into precision therapeutic approaches. Methods: We reviewed breakthrough discoveries from the past three years, analyzing single-cell multi-omics technologies, epitranscriptomics, stem cell architecture analysis, and precision medicine approaches. We examined cell-type-specific splicing aberrations, distinct stem cell architectures, epitranscriptomic modifications, and microenvironmental alterations in MDS pathogenesis. Results: Four interconnected mechanisms drive MDS: genetic alterations (splicing factor mutations), aberrant stem cell architecture (CMP-pattern vs. GMP-pattern), epitranscriptomic dysregulation involving pseudouridine-modified tRNA-derived fragments, and microenvironmental changes. Splicing aberrations show cell-type specificity, with SF3B1 mutations preferentially affecting erythroid lineages. Stem cell architectures predict therapeutic responses, with CMP-pattern MDS achieving superior venetoclax response rates (>70%) versus GMP-pattern MDS (<30%). Epitranscriptomic alterations provide independent prognostic information, while microenvironmental changes mediate treatment resistance. Conclusions: These advances represent a paradigm shift toward personalized MDS medicine, moving from single-biomarker to comprehensive molecular profiling guiding multi-target strategies. While challenges remain in standardizing molecular profiling and developing clinical decision algorithms, this systems-level understanding provides a foundation for precision oncology implementation and overcoming current therapeutic limitations. Full article

Background/Objectives: Curcumin is a promising therapy for glioblastoma but is limited by poor water solubility, rapid metabolism, and low blood–brain barrier penetration. This study aimed to evaluate curcumin and six curcumin derivatives with improved activity against a glioblastoma cell line and favorable absorption, distribution, metabolism, excretion, and toxicity (ADMET) properties. Methods: Twenty-one curcumin derivatives were assessed and subjected to in vitro MTT cytotoxicity assays in SF268 glioblastoma and Vero cells. On the basis of the cytotoxicity results, six derivatives with the most favorable characteristics were selected for additional mechanistic studies, which included microtubule depolymerization, mitochondrial membrane potential (ΔΨm), and BAX activation assays. ADMET properties were determined in silico. Results: Compounds 2–4, 6, and 11 demonstrated better activity (IC₅₀: 0.59–3.97 µg/mL and SI: 3–20) than curcumin (IC₅₀: 6.3 µg/mL; SI: 2.5). Lead derivatives destabilized microtubules, induced ΔΨm collapse, and activated BAX. In silico ADMET prediction analysis revealed that compounds 4 and 6 were the most promising for oral administration from a biopharmaceutical and pharmacokinetic point of view. Conclusions: Strategic modifications were made to one or both hydroxyl groups of the aromatic rings of curcumin to increase its physicochemical stability and activity against glioblastoma cell line SF268. Compound 4, bearing fully protected aromatic domains, was identified as a prime candidate for in vivo validation and formulation development. Full article

The CRISPR/Cas9 system is a powerful genome-editing tool that is applied in baculovirus engineering. In this study, we present the first report of the AcMNPV genome deletions for bioproduction purposes, using a dual single-guide RNA (sgRNA) CRISPR/Cas9 approach. We used this method to remove nonessential genes for the budded virus and boost recombinant protein yields when applied as BEVS. We show that the co-delivery of two distinct ribonucleoprotein (RNP) complexes, each assembled with a sgRNA and Cas9, into Sf9 insect cells efficiently generated deletions of fragments containing tandem genes in the genome. To evaluate the potential of this method, we assessed the expression of two model proteins, eGFP and HRPc, in insect cells and larvae. The gene deletions had diverse effects on protein expression: some significantly enhanced it while others reduced production. These results indicate that, although the targeted genes are nonessential, their removal can differentially affect recombinant protein yields depending on the host. Notably, HRPC expression increased up to 3.1-fold in Spodoptera frugiperda larvae. These findings validate an effective strategy for developing minimized baculovirus genomes and demonstrate that dual-guide CRISPR/Cas9 editing is a rapid and precise tool for baculovirus genome engineering. Full article

The extensive diversity of volatile organic compounds, along with their minor structural variations, presents significant challenges in the development of chemosensory-based biosensors. Previously, we generated sensor cells expressing insect odorant receptors (ORs) in Sf21 cells, demonstrating their potential as cell-based odorant sensor elements. However, it remains unclear whether the selectivity of cells expressing ORs in vitro for diverse compounds aligns with the receptor’s in vivo performance, aside from the response to target compounds. To address this, we assessed the ligand responses of sensor cells expressing ORs from Drosophila melanogaster using a high-throughput calcium imaging system. Our results demonstrate that in vitro receptor responses exhibit ligand selectivity comparable to in vivo conditions across different chemical categories. Broadly tuned OR-expressing sensor cells (Or13a, Or47a, and Or98a) displayed differential affinities, whereas the narrowly tuned Or56a-expressing sensor cells selectively responded to geosmin. Moreover, cell responses varied with subtle differences in chemical structure, including carbon chain length and functional group positioning. These findings provide valuable insights into insect OR–ligand interactions in vitro, demonstrating that receptor selectivity in sensor cells closely mirrors in vivo conditions. In addition to this consistency, our results highlight the subtle ligand differentiation capabilities of sensor cells enabling fluorescence-based visualization of receptor–ligand interactions. Full article

Background: Treatment of metastatic cancer remains a challenge, because cancer cells acquire resistance even to the most contemporary therapies. This study analyzed the role of the phosphoprotein Stathmin 1 (STMN1) in regulating cancer cell growth and metastatic potential. Methods: Public datasets with metastatic castration-resistant prostate cancer (mCRPC) and breast cancer (BC) were analyzed to determine the interrelationship between STMN1, hepatocyte growth factor (HGF) and MET proto-oncogene (MET) expression, overall survival, and response to chemotherapy. Site-directed mutagenesis, cell cycle analysis, proliferation, and migration and invasion assays determined the impact of STMN1 phosphorylation on proliferation and metastatic potential. Results: Increased STMN1 associates with HGF and MET gene expression in mCRPC, and taxane chemotherapy further increases HGF expression. STMN1 and HGF are highest, and overall survival is poorest in mCRPC in the liver compared to other sites, implying the metastatic site influences their expression levels and potentially the pattern of metastatic spread. Increased STMN1 and MET also predict taxane responsiveness in BC patients. Analysis of STMN1 serine (S)16, 25, 38, and 63 determined that total (t) STMN1 and STMN1 S16 phosphorylation (pSTMN1^S16) are co-regulated by HGF/MET during cell cycle progression, pSTMN1^S16 alone can promote cell proliferation, and pSTMN1^S16 shortens the cell cycle similar to HGF treatment, while STMN1^S16 dephosphorylation lengthens the cell cycle to arrest cell growth in G2/M, similar to HGF plus the MET inhibitor AMG337. Importantly, STMN1^S16 does not promote metastasis. Conclusions: Selectively inhibiting STMN1^S16 phosphorylation may provide an alternative strategy for inhibiting MET-mediated cell growth to eliminate metastatic cancer cells and inhibit further metastasis. Full article

Background: Rabbit hemorrhagic disease (RHD) is an acute, hemorrhagic and highly lethal infectious disease caused by rabbit hemorrhagic disease virus (RHDV), which causes huge economic losses to the rabbit breeding industry. Moreover, there is limited cross-protection between the two different serotypes of classic RHDV (GI.1) and RHDV2 (GI.2). The shortcomings of traditional inactivated vaccines have led to the development of novel subunit vaccines that can protect against both strains, and the VP60 capsid protein is the ideal antigenic protein. This study focused on developing a bivalent RHDV vaccine that can prevent infection with both GI.1 and GI.2 strains. Methodology: Baculovirus vectors containing classic RHDV and RHDV2 VP60 were co-transfected with linearized baculovirus into sf9 cells and transferred to baculovirus via homologous recombination of the VP60 gene. Infected sf9 cells were lysed, and after purification via Ni-NTA chromatography, VLPs were observed using transmission electron microscopy (TEM). In order to evaluate the immunogenicity of the chimeric RHDV VLP vaccine in rabbits, the RHDV VP60-specific antibody, IL-4, IFN-γ and neutralizing antibody titers were analyzed in serum using ELISA and HI. Results: The recombinant baculovirus system successfully expressed chimeric RHDV VLPs with a diameter of 32–40 nm. After immunization, it could produce specific antibodies, IL-4 and IFN-γ. Following the second immunization, neutralizing antibodies, determined using hemagglutination inhibition (HI) assays, were elicited. Conclusions: These data show that the chimeric RHDV VLP bivalent vaccine for immunized New Zealand rabbits can induce humoral immunity and cellular immunity in vivo, and the immunization effect of the high-dose group is similar to that of the current commercial vaccine. Full article

Background/Objectives: Feline panleukopenia, caused by FPV, is a highly contagious disease in cats. Current vaccines face challenges including complex production, high cost, and safety risks. Developing safer, more efficient alternatives is crucial. This study aimed to produce FPV virus-like particles (VLPs) using a recombinant baculovirus system expressing the VP2 gene and evaluate their immunogenicity and protective efficacy in cats. Methods: Sf9 insect cells were infected with recombinant baculovirus to express VP2 protein. The VP2 protein was purified using ultrafiltration and size-exclusion chromatography (SEC). Dynamic light scattering (DLS) and transmission electron microscopy (TEM) confirmed the assembly of VLPs. Twenty healthy cats were randomly divided into four groups; three groups received different doses (5 μg, 15 μg, and 45 μg) of FPV VLP vaccine, while the fourth group served as the control group immunized with PBS. Blood samples were collected on day 21 to measure hemagglutination inhibition (HI) and virus-neutralizing (VN) antibody responses. Cats in the 15 μg dose group were challenged with virulent FPV strain 708 on day 21, and clinical signs and white blood cell counts were monitored for 10 days. Results: Immunized cats exhibited significantly higher HI and VN antibody titers compared to controls. After challenge, vaccinated cats showed no clinical signs of disease, and their white blood cell counts remained stable. In contrast, control cats developed severe symptoms and experienced significant leukopenia. Conclusions: The FPV VLP vaccine generated in this study are highly immunogenic and provide effective protection against virulent FPV challenge, demonstrating their potential as a safer vaccine candidate for feline panleukopenia. Full article

Background/Objectives: Mycobacterium tuberculosis is the causative agent of tuberculosis and the leading cause of death from a single infection with the microorganism. Tuberculosis remains globally one of the major diseases leading to high mortality rates, with serious implications for public health and economic development. Therefore, tuberculosis prevention and control is crucial for global health and socio-economic stability. The development of effective preventive vaccines remains an urgent task in the fight against tuberculosis. Methods: The Mycobacterium tuberculosis antigen PPE18 was fused to Zera, and Bacmid was extracted and transfected into Sf9, which was purified and characterized for the formation of nanoparticle protein bodies. BALB/c mice and calves were immunized, and the immunogenicity of the nanoparticle vaccine was assessed by serum antibodies and splenic lymphocytes. Results: Zera-71CA-mCherry can be expressed in Sf9 cells, forming 0.5–1.2 μm protein bodies. Excising the mCherry sequence, Zera-71CA/Zera-PPE18 candidate nanoparticle-immunized mice were able to elicit serum antibody levels and the proliferation of splenic lymphocytes, and immunized calves were determined to have high levels of serum antibody levels, and IFN-γ and TNF-α levels. Conclusions: The results indicated that Zera-71CA/Zera-PPE18 recombinant nanoparticles had good immunogenicity as a subunit vaccine in both BALB/c mice and calves and are potential candidates for further development as effective subunit vaccines. Full article

Background: Human Immunodeficiency Virus (HIV) infections remain a source of cardiopulmonary complications among people receiving antiretroviral therapy. Still to this day, pulmonary hypertension (PH) severely affects the prognosis in this patient population. The persistent expression of HIV proteins, even during viral suppression, has been implicated in vascular dysfunction; however, little is known about the specific effects of these proteins on the pulmonary vasculature. This study investigates the impact of Nef variants derived from HIV-positive pulmonary hypertensive and normotensive donors on pulmonary vascular cells in vitro. Methods: We utilized well-characterized Nef molecular constructs to examine their effects on cell adhesion molecule gene expression (ICAM1, VCAM1, and SELE), pro-apoptotic gene expression (BAX, BAK), and vasoconstrictive endothelin-1 (EDN1) gene expression in endothelial nitric oxide synthase (eNOS) nitric oxide and the production and secretion of pro-inflammatory cytokines over 24, 48, and 72 h post-transfections with Nef variants. Results: HIV Nef variants SF2, NA7, and PH-associated Fr17 and 3236 induced a significant increase in adhesion molecule gene expression of ICAM1, VCAM1, and SELE. Pulmonary normotensive Nef 1138 decreased ICAM1 gene expression, but had increased VCAM1. PH Nef ItVR showed a consistent decrease in ICAM1 and no changes in SELE and VCAM1 expression. Further gene expression analyses of pro-apoptotic genes BAX and BAK demonstrated that Nef NA7, SF2, normotensive Nef 1138, and PH Nef Fr8, Fr9, Fr17, and 3236 variants significantly increased gene expression for apoptosis. Normotensive Nef 1138, as well as PH Nef Fr9 and ItVR, all displayed a statistically significant decrease in BAX expression. The expression of EDN1 had a statistically significant increase in samples treated with Nef NA7, SF2, normotensive Nef 2044 and PH Nef 3236, Fr17, and Fr8. Notably, PH-associated Nef variants sustained pro-inflammatory cytokine production, including IL-2, IL-4, and TNFα, while anti-inflammatory cytokine levels remained insufficient. Furthermore, eNOS was transiently upregulated by all Nef variants except for normotensive Nef 2044. Conclusions: The distinct effects of Nef variants on pulmonary vascular cell biology highlight the complex interplay between Nef, host factors, and vascular pathogenesis according to the variants. Full article

Nile tilapia, as an ideal model for studying sex differentiation, is a popular farmed fish worldwide with a stable XX/XY sex-determination system. In tilapia, ovarian differentiation is triggered by estradiol-17β (E2) production in undifferentiated gonads. In a previous study, we suggested that follicle-stimulating hormone (FSH) signaling might be involved in ovarian differentiation in Nile tilapia. In this study, we further investigated the role of FSH signaling in ovarian differentiation via aromatase expression, which converts testosterone to E2. Masculinization of XX fry by aromatase inhibitor or 17α-methyltestosterone leads to suppression of fshr expression. Feminization of XY fry by E2 treatment increased fshr expression from 15 days after hatching, when E2 treatment was terminated. XX tilapia developed ovaries harboring aromatase expression if fsh and fshr were double knockdowns by morpholino-oligo injections. Finally, the transcriptional activity in the upstream region of the aromatase gene (cyp19a1a) was further increased by FSH stimulation when HEK293T cells were co-transfected with foxl2 and ad4bp/sf1. Collectively, this study suggests that the role of FSH signaling is not critical in tilapia ovarian differentiation; however, FSH signaling may have a compensatory role in ovarian differentiation by increasing cyp19a1a transcription in cooperation with foxl2 and ad4bp/sf1 in Nile tilapia. Full article

(1) Background: Mopeia–Lassa reassortant ML29 virus is an investigational, reassortant virus vaccine for the prevention of Lassa fever caused by Lassa virus (LASV). (2) Methods: The vaccine virus ML29-SF was prepared in Vero cells using a serum-free culture medium under Good Manufacturing Practice. A 2-week repeat dose toxicity study was performed in guinea pigs under Good Laboratory Practice (GLP) regulations to assess the local and systemic toxicological effects. (3) Results: Following an intramuscular (IM) or subcutaneous (SC) injection of 10⁴ PFU of ML29-SF LASV vaccine at the start of the study, with a second dose 15 days later, no toxicological response attributable to the vaccine was observed. Vaccine-related effects were not observed in any in-life or post-mortem parameter evaluated, including clinical observations, injection site observations, body temperature, body weight, food consumption, ophthalmology, immunology, hematology, clinical chemistry, gross anatomical pathology, organ weights, and histopathology. An immunogenic response, as measured by the elicitation of IgG antibodies against major LASV immunogens, nucleocapsid and glycoprotein precursor, was observed in all vaccine-treated animals prior to the booster dose (Study Day 15) which endured through the end of the study (Study Day 42). There was no evidence of viral shedding in any vaccinated animal. (4) Conclusions: Overall, this single-dose vaccine was locally and systemically well tolerated even after a two-dose repeat administration, confirming the high level of safety of ML29-SF vaccination and supporting the future evaluation of this LASV vaccine, including in clinical trials. Full article

Background/Objectives: Oxidative stress plays a pivotal role in skin aging and carcinogenesis. Phytochemicals such as sulforaphane (SF, from broccoli sprouts or seeds) or curcumin (CUR, from turmeric) can be highly protective against this stress. They each induce a suite of cytoprotective and antioxidant enzymes that are coordinately transcribed via the Keap1-Nrf2-ARE pathway in mammals, such as the prototypical cytoprotective enzyme NAD(P)H dehydrogenase 1 (NQO1). Methods: Eighteen healthy human volunteers (9 males, 9 females, aged 18–69. were randomized to receive daily glucoraphanin (GR), which is converted to SF upon ingestion (450 mg; 1 mmol), CUR (1000 mg; 2.7 mmol), or both (450 mg GR + 1000 mg CUR), as oral supplements. After 8 days of a diet low in both compounds, blood and urine were collected for compliance and biomarker measurements. Randomized spots on the buttock’s skin were exposed to 2 x M.E.D. of UVB, and punch biopsies were obtained 1 and 3 days later for biomarker and histological measurement. Erythema was measured with a chromameter daily for 3 consecutive days following UVB. The process was repeated after receiving oral supplements, both with and without UVB exposure. Results: Compared to baseline, each treatment (n = 6 for each) induced NQO1 mRNA levels in skin biopsies: 3.1-fold with GR, 3.3-fold with CUR, and 3.6-fold with the combination of GR and CUR. Across all treatments (n = 18), expression of the pro-inflammatory cytokines IL-1β and TNF-α were reduced, as were IL-6, IL-17, STING, and CYR61, though less robustly. Modulation of these biomarkers persisted, but was less pronounced, in biopsies taken following UV exposure. The presence of SF and its metabolites in the skin post-treatment was confirmed by examining 6 of 12 subjects who ingested GR. Supplement effects on erythema following UV exposure were not significant, and no significant changes were measured in the same biomarkers in blood cells (PBMC), or by counting dyskeratotic keratinocytes. Supplements were well tolerated and compliance was excellent. Conclusions: Oral GR and CUR are well tolerated and have for the first time been shown to result in increased expression of cytoprotective genes and reduced expression of inflammatory cytokine genes in human skin in vivo. This mechanism-based clinical study suggests that an antioxidant, anti-inflammatory, and cytoprotective benefit from these oral supplements is delivered to the skin in humans. Full article

Alternative splicing (AS) is a pivotal post-transcriptional mechanism that expands the functional diversity of the proteome by enabling a single gene to generate multiple mRNA and protein isoforms. This process, which involves the differential inclusion or exclusion of exons and introns, is tightly regulated by splicing factors (SFs), such as serine/arginine-rich proteins (SRs), heterogeneous nuclear ribonucleoproteins (hnRNPs), and RNA-binding motif (RBM) proteins. These factors recognize specific sequences, including 5′ and 3′ splice sites and branch points, to ensure precise splicing. While AS is essential for normal cellular function, its dysregulation is increasingly implicated in cancer pathogenesis. Aberrant splicing can lead to the production of oncogenic isoforms that promote tumorigenesis, metastasis, and resistance to therapy. Furthermore, such abnormalities can cause the loss of tumor-suppressing activity, thereby contributing to cancer development. Importantly, abnormal AS events can generate neoantigens, which are presented on tumor cell surfaces via major histocompatibility complex (MHC) molecules, suggesting novel targets for cancer immunotherapy. Additionally, splice-switching oligonucleotides (SSOs) have shown promise as therapeutic agents because they modulate splicing patterns to restore normal gene function or induce tumor-suppressive isoforms. This review explores the mechanisms of AS dysregulation in cancer, its role in tumor progression, and its potential as a therapeutic target. We also discuss innovative technologies, such as high-throughput sequencing and computational approaches, that are revolutionizing the study of AS in cancer. Finally, we address the challenges and future prospects of targeting AS for personalized cancer therapies, emphasizing its potential in precision medicine. Full article

Genetics and mitochondrial (mt) dysfunction contribute to metabolic dysfunction-associated steatotic liver disease (MASLD). Recently, we demonstrated that the co-presence of PNPLA3, TM6SF2 and MBOAT7 polymorphisms predisposes to disease progression in MASLD patients and that their deletion triggers mt maladaptation in vitro. Here, we deepened the impact of the silencing of these genes on mt dynamism and respiration by reintroducing TM6SF2 and/or MBOAT7 wild-type proteins in deleted cells through lentiviral infection. Since hepatic mt bioenergetics is impaired in MASLD, in the attempt to identify a non-invasive signature, we then compared the enzymatic mt activity of seahorses, which was assessed in liver biopsies and peripheral blood mononuclear cells (PBMCs) of biopsy-proven MASLD patients (n = 44; Discovery cohort) stratified according to the number of the three at-risk variants (3NRV). Concerning the in vitro results, the rescue of MBOAT7 and/or TM6SF2 wild-type proteins resulted in the assembly of spaghetti-shaped mitochondria with improved oxidative phosphorylation (OXPHOS) capacity. In the Discovery cohort, the hepatic bioenergetic profile fully reflected that in PBMCs and was impaired especially in 3NRV carriers. A lowered serum respiration rate was confirmed in noninvasively assessed MASLD (n = 45; Fibroscan-MASLD cohort), while it did not change in unrelated liver disease patients (n = 45). In summary, we firstly demonstrated that mt circulating respirometry reflects that in liver and is specific in defining genetic MASLD. Full article