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Spectrochimica Acta. Part A, Molecular... Jun 2024This work aimed to set inline Raman spectroscopy models to monitor biochemically (viable cell density, cell viability, glucose, lactate, glutamine, glutamate, and...
This work aimed to set inline Raman spectroscopy models to monitor biochemically (viable cell density, cell viability, glucose, lactate, glutamine, glutamate, and ammonium) all upstream stages of a virus-like particle-making process. Linear (Partial least squares, PLS; Principal components regression, PCR) and nonlinear (Artificial neural networks, ANN; supported vector machine, SVM) modeling approaches were assessed. The nonlinear models, ANN and SVM, were the more suitable models with the lowest absolute errors. The mean absolute error of the best models within the assessed parameter ranges for viable cell density (0.01-8.83 × 10 cells/mL), cell viability (1.3-100.0 %), glucose (5.22-10.93 g/L), lactate (18.6-152.7 mg/L), glutamine (158-1761 mg/L), glutamate (807.6-2159.7 mg/L), and ammonium (62.8-117.8 mg/L) were 1.55 ± 1.37 × 10 cells/mL (ANN), 5.01 ± 4.93 % (ANN), 0.27 ± 0.22 g/L (SVM), 4.7 ± 2.6 mg/L (SVM), 51 ± 49 mg/L (ANN), 57 ± 39 mg/L (SVM) and 2.0 ± 1.8 mg/L (ANN), respectively. The errors achieved, and best-fitted models were like those for the same bioprocess using offline data and others, which utilized inline spectra for mammalian cell lines as a host.
PubMed: 38880076
DOI: 10.1016/j.saa.2024.124638 -
Veterinary Medicine and Science Jul 2024This study presents the case of non-purulent encephalomyelitis associated with astrovirus infection in a sheep from Eastern Anatolia, Türkiye.
BACKGROUND
This study presents the case of non-purulent encephalomyelitis associated with astrovirus infection in a sheep from Eastern Anatolia, Türkiye.
METHODS
A necropsy was performed on a sheep showing nervous signs. Afterwards, brain tissue samples were taken and examined with histopathological, immunohistochemical and molecular techniques.
RESULTS
Neuropathologic changes included neuronal degeneration, diffuse gliosis, multifocal perivascular cuffing, neuronophagy and neuronal necrosis in the cerebrum, the cerebellum and the cervical spinal cord. Aerobic and anaerobic bacterial culture, selective culture for Listeria monocytogenes, and PCR analysis for rabies virus, tick-borne encephalitis virus, Türkiye encephalitis virus, small ruminant lentiviruses and border disease virus were negative. However, the presence of astrovirus RNA in cerebral, cerebellar and spinal cord samples was demonstrated by a pan-astrovirus RT-PCR. Immunohistochemical examinations revealed astrovirus antigens within the neuronal cytoplasm. High-throughput sequencing techniques identified the causative agent as a member of the genotype species Mamastrovirus 13 but representing a distinct genetic lineage with similarity to ovine astrovirus 1 in the open-reading frames (ORF)1ab region and muskox astrovirus in the ORF2 region.
CONCLUSION
This report provides evidence that astroviruses are potentially encephalitis-causing pathogens in ovine populations in Türkiye, featuring an astrovirus strain distinct from those previously identified in sheep.
Topics: Animals; Sheep; Astroviridae Infections; Sheep Diseases; High-Throughput Nucleotide Sequencing; Encephalomyelitis; Sheep, Domestic; Astroviridae; Mamastrovirus; Phylogeny
PubMed: 38879881
DOI: 10.1002/vms3.1499 -
Biochemical and Biophysical Research... Jun 2024This study introduces an innovative brain-targeted drug delivery system, RVG-Exo/CBD, utilizing rabies virus glycoprotein (RVG)-engineered exosomes for encapsulating...
This study introduces an innovative brain-targeted drug delivery system, RVG-Exo/CBD, utilizing rabies virus glycoprotein (RVG)-engineered exosomes for encapsulating cannabidiol (CBD). The novel delivery system was meticulously characterized, confirming the maintenance of exosomal integrity, size, and successful drug encapsulation with a high drug loading rate of 83.0 %. Evaluation of the RVG-Exo/CBD's brain-targeting capability demonstrated superior distribution and retention in brain tissue compared to unmodified exosomes, primarily validated through in vivo fluorescence imaging. The efficacy of this delivery system was assessed using a behavioral sensitization model in mice, where RVG-Exo/CBD notably suppressed methamphetamine-induced hyperactivity more effectively than CBD alone, indicating a reduction in effective dose and enhanced bioavailability. Overall, the RVG-Exo/CBD system emerges as a promising strategy for enhancing the therapeutic efficacy and safety of CBD, particularly for neurological applications, highlighting its potential for addressing the limitations associated with traditional CBD administration in clinical settings.
PubMed: 38878760
DOI: 10.1016/j.bbrc.2024.150260 -
Frontiers in Immunology 2024Rabies virus (RABV) causes a fatal neurological disease, consisting of unsegmented negative-strand RNA, which encodes five structural proteins (3'-N-P-M-G-L-5')....
Rabies virus (RABV) causes a fatal neurological disease, consisting of unsegmented negative-strand RNA, which encodes five structural proteins (3'-N-P-M-G-L-5'). Apolipoprotein D (ApoD), a lipocalin, is upregulated in the nervous system after injury or pathological changes. Few studies have focused on the role of ApoD during virus infection so far. This study demonstrated that ApoD is upregulated in the mouse brain () and C8-D1A cells () after RABV infection. By upregulating ApoD expression in C8-D1A cells, we found that ApoD facilitated RABV replication. Additionally, Co-immunoprecipitation demonstrated that ApoD interacted with RABV glycoprotein (G protein). The interaction could promote RABV replication by upregulating the cholesterol level. These findings revealed a novel role of ApoD in promoting RABV replication and provided a potential therapeutic target for rabies.
Topics: Animals; Female; Humans; Male; Mice; Apolipoproteins D; Brain; Cell Line; Cholesterol; HEK293 Cells; Rabies; Rabies virus; Up-Regulation; Virus Replication
PubMed: 38868762
DOI: 10.3389/fimmu.2024.1392804 -
Experimental Neurology Jun 2024Cerebral ischemic stroke is a serious disease with high mortality and disability rates. However, few neuroprotective drugs have been used for ischemic stroke in the...
Cerebral ischemic stroke is a serious disease with high mortality and disability rates. However, few neuroprotective drugs have been used for ischemic stroke in the clinic. Two main reasons may be responsible for this failure: difficulty in penetrating the blood-brain barrier (BBB) and easily inactivated in the blood circulation. Ferroptosis, a lipid oxidation-related cell death, plays significant roles in cerebral ischemia-reperfusion injury. We utilized RVG29, a peptide derived from Rabies virus glycoprotein, to obtain BBB-targeted lipid nanoparticles (T-LNPs) in order to investigate whether T-LNPs improved the neuroprotective effects of Ferrostatin-1 (Fer1, an inhibitor of ferroptosis) against cerebral ischemic damage. T-LNPs significantly increased BBB penetration following oxygen/glucose deprivation exposure in an in vitro BBB model and enhanced the fluorescence distribution in brain tissues at 6 h post-administration in a cerebral ischemic murine model. Moreover, T-LNPs encapsulated Fer1 (T-LNPs-Fer1) significantly enhanced the inhibitory effects of Fer1 on ferroptosis by maintaining the homeostasis of NADPH oxidase 4 (NOX4) and glutathione peroxidase 4 (GPX4) signals in neuronal cells after cerebral ischemia. T-LNPs-Fer1 significantly suppressed oxidative stress [heme oxygenase-1 expression and malondialdehyde (the product of lipid ROS reaction)] in neurons and alleviated ischemia-induced neuronal cell death, compared to Fer1 alone without encapsulation. Furthermore, T-LNPs-Fer1 significantly reduced cerebral infarction and improved behavior functions compared to Fer1-treated cerebral ischemic mice after 45-min ischemia/24-h reperfusion. These findings showed that the T-LNPs helped Fer1 penetrate the BBB and improved the neuroprotection of Fer1 against cerebral ischemic damage in experimental stroke, providing a feasible translational strategy for the development of clinical drugs for the treatment of ischemic stroke.
PubMed: 38857748
DOI: 10.1016/j.expneurol.2024.114849 -
Frontiers in Cellular and Infection... 2024The rabies virus enters the nervous system by interacting with several molecular targets on host cells to modify behavior and trigger receptor-mediated endocytosis of...
The rabies virus enters the nervous system by interacting with several molecular targets on host cells to modify behavior and trigger receptor-mediated endocytosis of the virion by poorly understood mechanisms. The rabies virus glycoprotein (RVG) interacts with the muscle acetylcholine receptor and the neuronal α4β2 subtype of the nicotinic acetylcholine receptor (nAChR) family by the putative neurotoxin-like motif. Given that the neurotoxin-like motif is highly homologous to the α7 nAChR subtype selective snake toxin α-bungarotoxin (αBTX), other nAChR subtypes are likely involved. The purpose of this study is to determine the activity of the RVG neurotoxin-like motif on nAChR subtypes that are expressed in brain regions involved in rabid animal behavior. nAChRs were expressed in oocytes, and two-electrode voltage clamp electrophysiology was used to collect concentration-response data to measure the functional effects. The RVG peptide preferentially and completely inhibits α7 nAChR ACh-induced currents by a competitive antagonist mechanism. Tested heteromeric nAChRs are also inhibited, but to a lesser extent than the α7 subtype. Residues of the RVG peptide with high sequence homology to αBTX and other neurotoxins were substituted with alanine. Altered RVG neurotoxin-like peptides showed that residues phenylalanine 192, arginine 196, and arginine 199 are important determinants of RVG peptide apparent potency on α7 nAChRs, while serine 195 is not. The evaluation of the rabies ectodomain reaffirmed the observations made with the RVG peptide, illustrating a significant inhibitory impact on α7 nAChR with potency in the nanomolar range. In a mammalian cell culture model of neurons, we confirm that the RVG peptide binds preferentially to cells expressing the α7 nAChR. Defining the activity of the RVG peptide on nAChRs expands our understanding of basic mechanisms in host-pathogen interactions that result in neurological disorders.
Topics: alpha7 Nicotinic Acetylcholine Receptor; Animals; Rabies virus; Humans; Xenopus laevis; Glycoproteins; Oocytes; Viral Proteins; Viral Envelope Proteins; Host-Pathogen Interactions; Protein Binding; Rabies; Acetylcholine; Neurotoxins
PubMed: 38836054
DOI: 10.3389/fcimb.2024.1394713 -
Vaccine Jun 2024SYN023 is an anti-rabies monoclonal antibody mixture administered as part of post-exposure prophylaxis regimens. The rabies virus neutralizing antibody (RVNA)...
A phase 2b, Randomized, double blinded comparison of the safety and efficacy of the monoclonal antibody mixture SYN023 and human rabies immune globulin in patients exposed to rabies.
BACKGROUND
SYN023 is an anti-rabies monoclonal antibody mixture administered as part of post-exposure prophylaxis regimens. The rabies virus neutralizing antibody (RVNA) concentration generally accepted as an adequate immune response to vaccination is ≥ 0.5 IU/mL.
METHODS
Within 54 h of potential rabies exposure, 448 patients in two risk substrata of WHO Category III exposure were randomized to receive either 0.3 mg/kg SYN023 or 0.133 mL/kg human rabies immunoglobulin (HRIG) injected in and around the wound site(s) plus a course of rabies vaccination. Patients were followed for safety and absence of rabies for ≥ 365 days.
RESULTS
GMT RVNA was higher with SYN023 throughout the 2-week post-treatment period. In the primary analysis group (n = 368), 99.4 % of SYN023 recipients versus 4.5 % of HRIG recipients had protective RVNA levels on Day 4. On Day 8, 98.1 % SYN023 versus 12.2 % HRIG recipients were protected. The SYN023:HRIG ratio of geometric mean titer of RVNA (RVNA GMTs) on Day 8 (19.42) exceeded the 10 % superiority margin (P < 0.0001) indicating higher Day 8 RVNA with SYN023. On Day 99, the SYN023:HRIG RVNA GMT ratio (0.66) was below the non-inferiority margin of 20 % (P = 0.9485) suggesting some moderation of vaccine immune response by SYN023 relative to HRIG. The ratio of percent SYN023:HRIG recipients achieving RVNA ≥ 0.5 IU/mL on Day 99 (0.98) met the non-inferiority margin of 20 % (P = 0.013) indicating anti-rabies immune response with SYN023 was non-inferior to HRIG despite this effect. There were no probable/confirmed rabies cases in any patient. Study regimens were well tolerated.
CONCLUSIONS
SYN023 provided higher RVNA than HRIG soon after rabies exposure. By Day 99 post-treatment, GM RVNA with SYN023 was lower than HRIG, however, the percent of SYN023 recipients with a protective response was not inferior at this time point. No rabies cases were reported in the study. The SYN023 safety profile was acceptable.
CLINICALTRIALS
gov ID: NCT03961555.
PubMed: 38834432
DOI: 10.1016/j.vaccine.2024.05.066 -
Scientific Reports May 2024Rabies virus (RABV) causes fatal neurological disease. Pre-exposure prophylaxis (PrEP) and post-exposure prophylaxis (PEP) using inactivated-virus vaccines are the most...
Rabies virus (RABV) causes fatal neurological disease. Pre-exposure prophylaxis (PrEP) and post-exposure prophylaxis (PEP) using inactivated-virus vaccines are the most effective measures to prevent rabies. In Japan, HEP-Flury, the viral strain, used as a human rabies vaccine, has historically been propagated in primary fibroblast cells derived from chicken embryos. In the present study, to reduce the cost and labor of vaccine production, we sought to adapt the original HEP-Flury (HEP) to Vero cells. HEP was repeatedly passaged in Vero cells to generate ten- (HEP-10V) and thirty-passaged (HEP-30V) strains. Both HEP-10V and HEP-30V grew significantly better than HEP in Vero cells, with virulence and antigenicity similar to HEP. Comparison of the complete genomes with HEP revealed three non-synonymous mutations in HEP-10V and four additional non-synonymous mutations in HEP-30V. Comparison among 18 recombinant HEP strains constructed by reverse genetics and vesicular stomatitis viruses pseudotyped with RABV glycoproteins indicated that the substitution P(L115H) in the phosphoprotein and G(S15R) in the glycoprotein improved viral propagation in HEP-10V, while in HEP-30V, G(V164E), G(L183P), and G(A286V) in the glycoprotein enhanced entry into Vero cells. The obtained recombinant RABV strain, rHEP-PG4 strain, with these five substitutions, is a strong candidate for production of human rabies vaccine.
Topics: Animals; Vero Cells; Chlorocebus aethiops; Rabies Vaccines; Rabies virus; Amino Acid Substitution; Humans; Rabies; Genome, Viral
PubMed: 38822013
DOI: 10.1038/s41598-024-63337-9 -
Methods in Molecular Biology (Clifton,... 2024Vectored RNA vaccines offer a variety of possibilities to engineer targeted vaccines. They are cost-effective and safe, but replication competent, activating the humoral... (Review)
Review
Vectored RNA vaccines offer a variety of possibilities to engineer targeted vaccines. They are cost-effective and safe, but replication competent, activating the humoral as well as the cellular immune system.This chapter focuses on RNA vaccines derived from negative-strand RNA viruses from the order Mononegavirales with special attention to Newcastle disease virus-based vaccines and their generation. It shall provide an overview on the advantages and disadvantages of certain vector platforms as well as their scopes of application, including an additional section on experimental COVID-19 vaccines.
Topics: Animals; Humans; COVID-19; Genetic Vectors; Newcastle disease virus; RNA Viruses; SARS-CoV-2; Viral Vaccines; mRNA Vaccines
PubMed: 38814390
DOI: 10.1007/978-1-0716-3770-8_3 -
Journal of Virology May 2024Rabies virus (RABV) is highly lethal and triggers severe neurological symptoms. The neuropathogenic mechanism remains poorly understood. Ras-related C3 botulinum toxin...
Rabies virus (RABV) is highly lethal and triggers severe neurological symptoms. The neuropathogenic mechanism remains poorly understood. Ras-related C3 botulinum toxin substrate 1 (Rac1) is a Rho-GTPase that is involved in actin remodeling and has been reported to be closely associated with neuronal dysfunction. In this study, by means of a combination of pharmacological inhibitors, small interfering RNA, and specific dominant-negatives, we characterize the crucial roles of dynamic actin and the regulatory function of Rac1 in RABV infection, dominantly in the viral entry phase. The data show that the RABV phosphoprotein interacts with Rac1. RABV phosphoprotein suppress Rac1 activity and impedes downstream Pak1-Limk1-Cofilin1 signaling, leading to the disruption of F-actin-based structure formation. In early viral infection, the EGFR-Rac1-signaling pathway undergoes a biphasic change, which is first upregulated and subsequently downregulated, corresponding to the RABV entry-induced remodeling pattern of F-actin. Taken together, our findings demonstrate for the first time the role played by the Rac1 signaling pathway in RABV infection and may provide a clue for an explanation for the etiology of rabies neurological pathogenesis.IMPORTANCEThough neuronal dysfunction is predominant in fatal rabies, the detailed mechanism by which rabies virus (RABV) infection causes neurological symptoms remains in question. The actin cytoskeleton is involved in numerous viruses infection and plays a crucial role in maintaining neurological function. The cytoskeletal disruption is closely associated with abnormal nervous symptoms and induces neurogenic diseases. In this study, we show that RABV infection led to the rearrangement of the cytoskeleton as well as the biphasic kinetics of the Rac1 signal transduction. These results help elucidate the mechanism that causes the aberrant neuronal processes by RABV infection and may shed light on therapeutic development aimed at ameliorating neurological disorders.
PubMed: 38809020
DOI: 10.1128/jvi.00606-24