Search Results (33)

PDE11A is a little-studied phosphodiesterase sub-family that breaks down cAMP/cGMP, with the PDE11A4 isoform enriched in the memory-related hippocampal formation. Age-related increases in PDE11A expression occur in human and rodent hippocampus and cause age-related cognitive decline of social memories. Interestingly, age-related increases in PDE11A4 protein ectopically accumulate in spherical clusters that group together in the brain to form linear filamentous patterns termed “PDE11A4 ghost axons”. The biophysical/physiochemical mechanisms underlying this age-related clustering are not known. Here, we determine if age-related clustering of PDE11A4 reflects liquid–liquid phase separation (LLPS; biomolecular condensation), and if PDE11A inhibitors can reverse this LLPS. We show human and mouse PDE11A4 exhibit several LLPS-promoting sequence features, including intrinsically disordered regions, non-covalent pi–pi interactions, and prion-like domains that were particularly enriched in the N-terminal regulatory region. Further, multiple bioinformatic tools predict PDE11A4 undergoes LLPS. Consistent with these predictions, aging-like PDE11A4 clusters in HT22 hippocampal neuronal cells were membraneless spherical droplets that progressively fuse over time in a concentration-dependent manner. Deletion of the N-terminal intrinsically disordered region prevented PDE11A4 LLPS despite equal protein expression between WT and mutant constructs. 1,6-hexanediol, along with tadalafil and BC11-38 that inhibit PDE11A4, reversed PDE11A4 LLPS in HT22 hippocampal neuronal cells. Interestingly, PDE11A4 inhibitors reverse PDE11A4 LLPS independently of increasing cAMP/cGMP levels via catalytic inhibition. Importantly, orally dosed tadalafil reduced PDE11A4 ghost axons in old mouse ventral hippocampus by 50%. Thus, PDE11A4 exhibits the four defining criteria of LLPS, and PDE11A inhibitors reverse this age-related phenotype both in vitro and in vivo. Full article

Background/Objectives: We generated a novel recombinant Vibrio cholerae ghost (rVCG)-based subunit vaccine incorporating the A1 subunit of cholera toxin (CTA1) and a multiepitope Chlamydia trachomatis (CT) antigen (MECA) derived from five chlamydial outer membrane proteins (rVCG-MECA). The ability of this vaccine to protect against a CT transcervical challenge was evaluated. Methods: Female C57BL/6J mice were immunized thrice at two-week intervals with rVCG-MECA or rVCG-gD2 (antigen control) via the intramuscular (IM) or intranasal (IN) route. PBS-immunized mice or mice immunized with live CT served as negative and positive controls, respectively. Results: Vaccine delivery stimulated robust humoral and cell-mediated immune effectors, characterized by local mucosal and systemic CT-specific IgG, IgG2c, and IgA antibody and IFN-γ (Th1 cytokine) responses. The elicited mucosal and systemic IgG2c and IgA antibody responses persisted for 16 weeks post-immunization. Immunization with rVCG-MECA afforded protection comparable to that provided by IN immunization with live CT EBs without any side effects, irrespective of route of vaccine delivery. Conclusions: The results underline the potential of a multiepitope vaccine as a promising resource for protecting against CT genital infection and the potential of CTA1 on the VCG platform as a mucosal and systemic adjuvant for developing CT vaccines. Full article

Background/Objectives Bacterial ghosts (BGs), non-living empty envelopes of bacteria, are produced either through genetic engineering or chemical treatment of bacteria, retaining the shape of their parent cells. BGs are considered vaccine candidates, promising delivery systems, and vaccine adjuvants. The practical use of BGs in vaccine development for humans is limited because of concerns about the preservation of viable bacteria in BGs. Methods: To increase the efficiency of Klebsiella pneumoniae BG formation and, accordingly, to ensure maximum killing of bacteria, we exploited previously designed plasmids with the lysis gene E from bacteriophage φX174 or with holin–endolysin systems of λ or L-413C phages. Previously, this kit made it possible to generate bacterial cells of Yersinia pestis with varying degrees of hydrolysis and variable protective activity. Results: In the current study, we showed that co-expression of the holin and endolysin genes from the L-413C phage elicited more rapid and efficient K. pneumoniae lysis than lysis mediated by only single gene E or the low functioning holin–endolysin system of λ phage. The introduction of alternative lysing factors into K. pneumoniae cells instead of the E protein leads to the loss of the murein skeleton. The resulting frameless cell envelops are more reminiscent of bacterial sacs or bacterial skins than BGs. Although such structures are less naive than classical bacterial ghosts, they provide effective protection against infection by a hypervirulent strain of K. pneumoniae and can be recommended as candidate vaccines. For our vaccine candidate generated using the O1:K2 hypervirulent K. pneumoniae strain, both safety and immunogenicity aspects were evaluated. Humoral and cellular immune responses were significantly increased in mice that were intraperitoneally immunized compared with subcutaneously vaccinated animals (p < 0.05). Conclusions: Therefore, this study presents novel perspectives for future research on K. pneumoniae ghost vaccines. Full article

Backgrounds: Imaging of parotid tumors is crucial for surgery planning, but it cannot distinguish malignant from benign lesions with absolute reliability. The aim of the study was to establish a diagnostic MRI algorithm to differentiate parotid tumors. Methods: A retrospective study was conducted including all patients with parotid tumors, who underwent 3T-MRI and surgery. Morphological characteristics and normalized T2 and late postcontrast T1 signal intensities (SI) were assessed. “Ghosting sign” on late postcontrast T1 sequence was defined as indistinguishability of the tumor except for a thin peripheral enhancement. Patients were divided according to histology and imaging data were compared. A diagnostic MRI algorithm was established. Results: Thirty-six patients were included. The combination of normalized late T1 postcontrast SI, normalized T2 SI and “ghosting sign” allowed for the distinguishing of malignant from benign parotid tumors with high sensitivity (100%), specificity (93%), positive predictive value (80%), negative predictive value, (100%) and accuracy (94%). Moreover, pleomorphic adenomas often showed a homogeneous T2 signal and a complete capsule (p < 0.01), Warthin tumors protein-rich cysts and calcifications (p < 0.005 and p < 0.05), and malignant tumors an inhomogeneous contrast enhancement (p < 0.01). Conclusions: High field MRI represents a promising tool in parotid tumors, allowing for an accurate differentiation of malignant and benign lesions. Full article

Salmonella is a significant zoonotic foodborne pathogen, and the global spread of multidrug-resistant (MDR) strains poses substantial challenges, necessitating alternatives to antibiotics. Among these alternatives, vaccines protect the community against infectious diseases effectively. This review aims to summarize the efficacy of developed Salmonella vaccines evaluated in various animal hosts and highlight key transitions for future vaccine studies. A total of 3221 studies retrieved from Web of Science, Google Scholar, and PubMed/Medline databases between 1970 and 2023 were evaluated. One hundred twenty-seven qualified studies discussed the vaccine efficacy against typhoidal and nontyphoidal serovars, including live-attenuated vaccines, killed inactivated vaccines, outer membrane vesicles, outer membrane complexes, conjugate vaccines, subunit vaccines, and the reverse vaccinology approach in different animal hosts. The most efficacious vaccine antigen candidate found was recombinant heat shock protein (rHsp60) with an incomplete Freund’s adjuvant evaluated in a murine model. Overall, bacterial ghost vaccine candidates demonstrated the highest efficacy at 91.25% (95% CI = 83.69–96.67), followed by the reverse vaccinology approach at 83.46% (95% CI = 68.21–94.1) across animal hosts. More than 70% of vaccine studies showed significant production of immune responses, including humoral and cellular, against Salmonella infection. Collectively, the use of innovative methods rather than traditional approaches for the development of new effective vaccines is crucial and warrants in-depth studies. Full article

Trypanosoma brucei causes African trypanosomiasis in humans. Infection with T. brucei elicits a potent pro-inflammatory immune response within infected human hosts, and this response is thought to at least be partially due to Toll-like receptor (TLR) activation. In response to stimulation by lipopolysaccharide and other pathogen antigens, TLR4 translocates to lipid rafts, which induces the expression of pro-inflammatory genes. However, cholesterol efflux is acknowledged as anti-inflammatory due to promoting lipid raft disruption. In this study, we wanted to assess the impact of T. brucei “ghosts”, which are non-viable T. brucei essentially devoid of intracellular contents, in stimulating macrophage TLR4 translocation to lipid rafts, and whether promoting cholesterol efflux in macrophages incubated with T. brucei ghosts attenuates TLR4-target gene expression. When cultured macrophages were exposed to T. brucei ghosts, we observed an increase in lipid raft TLR4 protein content, which suggests certain surface molecules of T. brucei serve as ligands for TLR4. However, pretreating macrophages with cholesterol acceptors before T. brucei ghost exposure decreased lipid raft TLR4 protein content and the expression of pro-inflammatory TLR4-target genes. Taken together, these results imply that macrophage cholesterol efflux weakens pro-inflammatory responses which occur from T. brucei infection via increasing macrophage lipid raft disruption. Full article

Chlamydia abortus (C. abortus) is an important zoonotic pathogen that seriously endangers the development of animal husbandry. Vaccination is the most effective approach to preventing C. abortus infection. We previously reported a recombinant Escherichia coli ghost (rECG)-based C. abortus vaccine that demonstrated outstanding protective efficacy. In this study, we further attempted to fuse the cholera toxin B subunit (CTB), a widely studied potent mucosal immune adjuvant, with macrophage infectivity potentiator (MIP), a candidate antigen of C. abortus, on the surface of the rECG and explore its protective effect against C. abortus infection. The MIP fusion protein was highly expressed in the rECGs, and the CTB-modified rECGs significantly induced the activation of mouse bone marrow-derived dendritic cells in vitro. Intranasal immunization with rECGs induced a Th1-biased cellular immune response. Compared to the rECGs without CTB, the CTB-modified rECGs induced higher concentrations of IgA in the serum and vaginal wash solution. Moreover, in a mouse infection model, the CTB-modified rECGs significantly improved the clearance efficiency of C. abortus and reduced the pathological damage to the uterus. This study demonstrates that incorporating CTB into rECGs significantly enhances the immunogenic potential of the rECG vaccine and can significantly enhance its protective efficacy against a C. abortus challenge. Full article

Bacterial ghosts (BGs) are hollow bacterial cell envelopes with intact cellular structures, presenting as promising candidates for various biotechnological and biomedical applications. However, the yield and productivity of BGs have encountered limitations, hindering their large-scale preparation and multi-faceted applications of BGs. Further optimization of BGs is needed for the commercial application of BG technology. In this study, we screened out the most effective lysis protein ID52-E-W4A among 13 mutants based on phage ID52 lysis protein E and optimized the liquid culture medium for preparing Escherichia coli Nissle 1917 (EcN). The results revealed a significantly higher lysis rate of ID52-E-W4A compared to that of ID52-E in the 2xYT medium. Furthermore, EcN BGs were cultivated in a fermenter, achieving an initial OD₆₀₀ as high as 6.0 after optimization, indicating enhanced BG production. Moreover, the yield of ID52-E-W4A-induced BGs reached 67.0%, contrasting with only a 3.1% yield from φX174-E-induced BGs. The extended applicability of the lysis protein ID52-E-W4A was demonstrated through the preparation of Salmonella pullorum ghosts and Salmonella choleraesuis ghosts. Knocking out the molecular chaperone gene slyD and dnaJ revealed that ID52-mediated BGs could still undergo lysis. Conversely, overexpression of integral membrane enzyme gene mraY resulted in the loss of lysis activity for ID52-E, suggesting that the lysis protein ID52-E may no longer rely on SlyD or DnaJ to function, with MraY potentially being the target of ID52-E. This study introduces a novel approach utilizing ID52-E-W4A for recombinant expression, accelerating the BG formation and thereby enhancing BG yield and productivity. Full article

Lactobacillus helveticus 34.9 was isolated from a sample of Romanian home-made fermented milk, producing both surface layer proteins and a class III bacteriocin. The present study aimed to investigate the biological and functional role of the S-layer in correlation with its probiotic properties. The presence of S-layer proteins resulted in various degrees of co-aggregation of L. helveticus 34.9 with pathogens and with other lactic acid bacteria, but the removal of these proteins reduced the co-aggregation with all the tested strains. Moreover, the S-layer proved to be involved in cell wall hydrophobicity and cellular protection during freeze-drying. In the simulated passage through the gastrointestinal tract, S-layer depleted cells exhibited increased vulnerability, with greater viability loss in low pH and pepsin treatment compared to control cells. Subsequently, in the small intestine simulation, these cells lost all viability, underscoring the vital role of extracellular proteins for cell protection. The morphological effects of these treatments were observed by scanning electron microscopy. Severe structural damage was noticed when the S-layer was absent, including loss of cell shape and integrity as well as many ghost cells emptied of their content. Finally, the elimination of surface proteins reduced the interaction between L. helveticus 34.9 and mammalian cells. Full article

Efficient delivery of a DNA plasmid into antigen-presenting cells (APCs) is a potential strategy to enhance the immune responses of DNA vaccines. The bacterial ghost (BG) is a potent DNA vaccine delivery system that targets APCs. In the present work, we describe a new strategy of using E. coli BGs as carriers for an Ii-linked Hepatitis C Virus (HCV) NS3 DNA vaccine that improved both the transgene expression level and the antigen-presentation level in APCs. BGs were prepared from DH5α cells, characterized via electron microscopy and loaded with the DNA vaccine. The high transfection efficiency mediated using BGs was first evaluated in vitro, and then, the immune protective effect of the BG-Ii-NS3 vaccine was determined in vivo. It was found that the antibody titer in the sera of BG-Ii-NS3-challenged mice was higher than that of Ii-NS3-treated mice, indicating that the BGs enhanced the humoral immune activity of Ii-NS3. The cellular immune protective effect of the BG-Ii-NS3 vaccine was determined using long-term HCV NS3 expression in a mouse model in which luciferase was used as a reporter for HCV NS3 expression. Our results showed that the luciferase activity in BG-Ii-NS3-treated mice was significantly reduced compared with that in Ii-NS3-treated mice. The CTL assay results demonstrated that BG-Ii-NS3 induced a greater NS3-specific T-cell response than did Ii-NS3. In summary, our study demonstrated that BGs enhanced both the humoral and cellular immune response to the Ii-NS3 DNA vaccine and improved its immune protection against HCV infection. Full article

Epipogium roseum, commonly known as one of the ghost orchids due to its rarity and almost transparent color, is a non-photosynthetic and fully mycoheterotrophic plant. Given its special nutritional strategies and evolutionary significance, the mitogenome was first characterized, and three plastomes sampled from Asia were assembled. The plastomes were found to be the smallest among Orchidaceae, with lengths ranging from 18,339 to 19,047 bp, and exhibited high sequence variety. For the mitogenome, a total of 414,552 bp in length, comprising 26 circular chromosomes, were identified. A total of 54 genes, including 38 protein-coding genes, 13 tRNA genes, and 3 rRNA genes, were annotated. Multiple repeat sequences spanning a length of 203,423 bp (45.47%) were discovered. Intriguingly, six plastid regions via intracellular gene transfer and four plastid regions via horizontal gene transfer to the mitogenome were observed. The phylogenomics, incorporating 90 plastomes and 56 mitogenomes, consistently revealed the sister relationship of Epipogium and Gastrodia, with a bootstrap percentage of 100%. These findings shed light on the organelle evolution of Orchidaceae and non-photosynthetic plants. Full article

Ghost crabs, as a species of the Ocypode within the subfamily Ocypodinae, are distributed in the upper intertidal zone worldwide and are ecologically remarkable. They play an important role in the energy circulation in the intertidal zone and are used as an ecological indicator to predict the impacts of environmental change or anthropogenic activities on the marine ecosystem. In this study, we provide the first evidence for the distribution of O. sinensis in Jeju Island and the southern coastal area on the Korean Peninsula. We generated a high-fidelity mitochondrial genome (mitogenome) for the species. The mitogenome was assembled into a circular chromosome of 15,589 bp, including 13 protein-coding genes, two ribosomal RNA genes, and twenty-two transfer RNA genes. High genetic variation compared with closely related species enabled the precise reconstruction of phylogenetic relationships and an estimation of the divergence times among the Ocypode species. The phylogenetic inference indicated that O. sinensis forms a monophyletic clade with O. cordimanus and diverged from ancestral species approximately 20.41 million years ago. Full article

Endoclita signifer is a prominent wood-boring insect species in eucalyptus plantations in Guangxi, China, causing significant ecological and economic damage. A novel approach to controlling the challenging wood-boring pest involves disrupting the olfactory communication between insects and the volatile compounds emitted by plants. To identify the olfactory proteins contributing to host selection based on 11 GC-EAD-active volatiles from eucalyptus leaves and to discover the highly expressed olfactory proteins, we conducted a study on the antennal transcriptomes of adult E. signifer and screened key olfactory proteins in the antennae. We identified a total of 69 olfactory proteins. When compared to the larval transcriptomes, the antennal transcriptome of adult E. signifer revealed the presence of 17 new odorant-binding proteins (OBPs), including 2 pheromone-binding proteins (PBPs), 7 previously unreported chemosensory proteins (CSPs), 17 new odorant receptors (ORs), 4 new gustatory receptors (GRs), 11 novel ionotropic receptors (IRs), and 2 sensory neuron membrane proteins (SNMPs). Through the phylogenetic tree of OBPs and ORs, we identified EsigPBP2 and EsigPBP3 as two of the three PBPs, designated EsigOR13 as EsigOrco, and recognized EsigOR10 and EsigOR22 as the newly discovered EsigPRs in E. signifer. In the adult antennae, the expression levels of EsigGOBP14, EsigGOBP13, EsigOBP14, EsigOBP17, EsigCSP14, and EsigOR16 were notably high, indicating that these proteins could be pivotal in binding to plant volatiles. Full article

AD-2 (20(R)-dammarane-3β, 12β, 20, 25-tetrol, 25-OH-PPD) was structurally modified to introduce additional amino groups, which can better exert its anti-tumor effects in MCF-7, A549, LoVo, HCT-116, HT -29, and U-87 cell lines. We investigated the cellular activity of 15 different AD-2 amino acid derivatives on HepG2 cells and the possible mechanism of action of the superior derivative 6b. An MTT assay was used to detect the cytotoxicity of the derivatives. Western blotting was used to study the signaling pathways. Flow cytometry was used to detect cell apoptosis and ghost pen peptide staining was used to identify the changes in the cytoskeleton. The AD-2 amino acid derivatives have a better cytotoxic effect on the HepG2 cells than AD-2, which may be achieved by promoting the apoptosis of HepG2 cells and influencing the cytoskeleton. The derivative 6b shows obvious anti-HepG2 cells activity through affecting the expression of apoptotic proteins such as MDM2, P-p53, Bcl-2, Bax, Caspase 3, Cleaved Caspase 3, Caspase 8, and NSD2. According to the above findings, the amino acid derivatives of AD-2 may be developed as HepG2 cytotoxic therapeutic drugs. Full article

Haemophilus influenzae is a Gram-negative bacterium characterized as a small, nonmotile, facultative anaerobic coccobacillus. It is a common cause of a variety of invasive and non-invasive infections. Among six serotypes (a–f), H. influenzae type b (Hib) is the most familiar and predominant mostly in children and immunocompromised individuals. Following Hib vaccination, infections due to other serotypes have increased in number, and currently, there is no suitable effective vaccine to induce cross-strain protective antibody responses. The current study was aimed to validate the capability of two 20-mer highly conserved synthetic tbp1 (transferrin-binding protein 1) peptide-based vaccine candidates (tbp1-E₁ and tbp1-E₂) predicted using in silico approaches to induce immune responses against H. influenzae strains. Cytokine induction ability, immune simulations, and molecular dynamics (MD) simulations were performed to confirm the candidacy of epitopic docked complexes. Synthetic peptide vaccine formulations in combination with two different adjuvants, BGs (Bacterial Ghosts) and CFA/IFA (complete/incomplete Freund’s adjuvant), were used in BALB/c mouse groups in three booster shots at two-week intervals. An indirect ELISA was performed to determine endpoint antibody titers using the Student’s t-distribution method. The results revealed that the synergistic use of both peptides in combination with BG adjuvants produced better results. Significant differences in absorbance values were observed in comparison to the rest of the peptide–adjuvant combinations. The findings of this study indicate that these tbp1 peptide-based vaccine candidates may present a preliminary set of peptides for the development of an effective cross-strain vaccine against H. influenzae in the future due to their highly conserved nature. Full article