RESUMO
Haploid males of hymenopteran species produce gametes through an abortive meiosis I followed by meiosis II that can either be symmetric or asymmetric in different species. Thus, one spermatocyte could give rise to two spermatids with either equal or unequal amounts of cytoplasm. It is currently unknown what molecular features accompany these postmeiotic sperm cells especially in species with asymmetric meiosis II such as bees. Here we present testis single-cell RNA sequencing datasets from the honeybee (Apis mellifera) drones of 3 and 14 days after emergence (3d and 14d). We show that, while 3d testes exhibit active, ongoing spermatogenesis, 14d testes only have late-stage spermatids. We identify a postmeiotic bifurcation in the transcriptional roadmap during spermatogenesis, with cells progressing toward the annotated spermatids (SPT) and small spermatids (sSPT), respectively. Despite an overall similarity in their transcriptomic profiles, sSPTs express the fewest genes and the least RNA content among all the sperm cell types. Intriguingly, sSPTs exhibit a relatively high expression level for Hymenoptera-restricted genes and a high mutation load, suggesting that the special meiosis II during spermatogenesis in the honeybee is accompanied by phylogenetically young gene activities.
Assuntos
Sêmen , Espermatogênese , Abelhas/genética , Masculino , Animais , Espermatogênese/genética , Espermátides/metabolismo , Testículo , Espermatócitos/metabolismo , Meiose/genéticaRESUMO
The HIV-1 envelope is a heavily glycosylated class 1 trimeric fusion protein responsible for viral entry into CD4+ immune cells. Developing neutralizing antibodies against the specific envelope glycans is an alternative method for antiviral therapies. This work presents the first-ever development and characterization of artificial neutralizing antibodies using molecular imprinting technology to recognize and bind to the envelope protein of HIV-1. The prepared envelope glycan-imprinted nanoparticles (GINPs) can successfully prevent HIV-1 from infecting target cells by shielding the glycans on the envelope protein. In vitro experiments showed that GINPs have strong affinity toward HIV-1 (Kd = 36.7 ± 2.2 nM) and possess high anti-interference and specificity. GINPs demonstrate broad inhibition activity against both tier 1 and tier 2 HIV-1 strains with a pM-level IC50 and exhibit a significant inhibitory effect on long-term viral replication by more than 95%. The strategy provides a promising method for the inhibition and therapy of HIV-1 infection.
Assuntos
Infecções por HIV , HIV-1 , Humanos , Anticorpos Neutralizantes , Anticorpos Anti-HIV/metabolismo , Glicosilação , Infecções por HIV/tratamento farmacológico , Polissacarídeos/metabolismoRESUMO
Enhancing the antitumor immune response and targeting ability of oncolytic viruses will improve the effect of tumor immunotherapy. Through infecting neural stem cells (NSCs) with a capsid dual-modified oncolytic adenovirus (CRAd), we obtained and characterized the "oncolytic extracellular vesicles" (CRAdEV) with improved targeted infection and tumor killing activity compared with CRAd. Both ex vivo and in vivo studies revealed that CRAdEV activated innate immune cells and importantly enhanced the immunomodulatory effect compared to CRAd. We found that CRAdEV effectively increased the number of DCs and activated CD4+ and CD8+ T cells, significantly increased the number and activation of B cells, and produced higher levels of tumor-specific antibodies, thus eliciting enhanced antitumor activity compared with CRAd in a B16 xenograft immunocompetent mice model. This study provides a novel approach to oncolytic adenovirus modification and demonstrates the potential of "oncolytic extracellular vesicles" in antitumor immunotherapy.
Assuntos
Adenoviridae , Vesículas Extracelulares , Terapia Viral Oncolítica , Vírus Oncolíticos , Animais , Camundongos , Adenoviridae/genética , Terapia Viral Oncolítica/métodos , Humanos , Linhagem Celular Tumoral , Imunoterapia , Células-Tronco Neurais/imunologia , Imunomodulação/efeitos dos fármacos , Melanoma Experimental/imunologia , Melanoma Experimental/terapia , Melanoma Experimental/patologia , Linfócitos T CD8-Positivos/imunologiaRESUMO
The solar-driven overall water splitting (2H2Oâ2H2 + O2) is considered as one of the most promising strategies for reducing carbon emissions and meeting energy demands. However, due to the sluggish performance and high H2 cost, there is still a big gap for the current photocatalytic systems to meet the requirements for practical sustainable H2 production. Economic feasibility can be attained through simultaneously generating products of greater value than O2, such as hydrogen peroxide (H2O2, 2H2OâH2 + H2O2). Compared with overall water splitting, this approach is more kinetically feasible and generates more high-value products of H2 and H2O2. In several years, there has been an increasing surge in exploring the possibility and substantial progress has been achieved. In this review, a concise overview of the importance and underlying principles of PIWS is first provided. Next, the reported typical photocatalysts for PIWS are discussed, including commonly used semiconductors and cocatalysts, essential design features of these photocatalysts, and connections between their structures and activities, as well as the selected approaches for enhancing their stability. Then, the techniques used to quantify H2O2 and the operando characterization techniques that can be employed to gain a thorough understanding of the reaction mechanisms are summarized. Finally, the current existing challenges and the direction needing improvement are presented. This review aims to provide a thorough summary of the most recent research developments in PIWS and sets the stage for future advancements and discoveries in this emerging area.
RESUMO
Assembly of the adenovirus capsid protein hexon depends on the assistance of the molecular chaperone L4-100K. However, the chaperone mechanisms remain unclear. In this study, we found that L4-100K was involved in the hexon translation process and could prevent hexon degradation by the proteasome in cotransfected human cells. Two nonadjacent domains, 84-133 and 656-697, at the N-terminal and C-terminal regions of human adenovirus type 5 L4-100K, respectively, were found to be crucial and cooperatively responsible for hexon trimer expression and assembly. These two chaperone-related domains were conserved in the sequence of L4-100K and in the function of hexon assembly across different adenovirus serotypes. Different degrees of cross-activity of hexon trimerization with different serotypes were detected in subgroups B, C, and D, which were proven to be controlled by the interaction between the C-terminal chaperone-related domain of L4-100K and hypervariable regions (HVR) of hexon. Additionally, HVR-chimeric hexon mutants were successfully assembled with the assistance of the 1-697 mutant. Structural analysis of 656-697 by nuclear magnetic resonance and structural prediction of L4-100K using Robetta showed that the two conserved domains are mainly composed of α-helices and are located on the surface of the highly folded core region. Our research provides a more complete understanding of hexon assembly and guidance for the development of hexon-chimeric adenovirus vectors that will be safer, smarter, and more efficient. IMPORTANCE Adenovirus vectors have been widely used in clinical trials of vaccines and gene therapy, although some deficiencies remain. Chimeric modification of the hexon was expected to improve the potency of preexisting immune evasion and targeting, but in many cases, viral packaging is prevented by the inability of the chimeric hexon to assemble correctly. So far, few studies have examined the mechanisms of hexon trimer assembly. Here, we show how the chaperone protein L4-100K contributes to the assembly of the adenovirus capsid protein hexon, and these data will provide a guide for novel adenovirus vector design and development, as we desired.
Assuntos
Adenovírus Humanos , Chaperonas Moleculares , Proteínas não Estruturais Virais , Humanos , Adenovírus Humanos/genética , Adenovírus Humanos/metabolismo , Capsídeo/metabolismo , Proteínas do Capsídeo/metabolismo , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo , Proteínas não Estruturais Virais/genética , Proteínas não Estruturais Virais/metabolismoRESUMO
Solar energy is an ideal clean and inexhaustible energy source. Solar-driven formic acid (FA) dehydrogenation is one of the promising strategies to address safety and cost issues related to the storage, transport, and distribution of hydrogen energy. For FA dehydrogenation, the O-H and C-H cleavages are the key steps, and developing a photocatalyst with the ability to break these two bonds is critical. In this work, both density functional theory (DFT) calculation and experimental results confirmed the positive synergistic effect between brookite and rutile TiO2 for O-H and C-H cleavage in HCOOH. Further, brookite TiO2 is beneficial to the generation of the â¢OH radical and significantly promotes C-H cleavage in formate. Under optimized conditions, the H2 production efficiency of FA dehydrogenation can reach up to 30.4 µmol·mg-1·h-1, which is the highest value compared with similar reported TiO2-based systems and over 1.7 times the reported highest value of Au0.75Pd0.25/TiO2 photocatalysts. More importantly, after more than 42 days (>500 h) of irradiation, the system still demonstrated high H2 production activity, indicating the potential for practical application. This work provides a valuable strategy to improve both the efficiency and stability of photocatalytic FA dehydrogenation under mild conditions.
RESUMO
An oncolytic virus is a promising strategy for glioblastoma (GBM) therapy. However, there are still some challenges such as the blood-brain barrier (BBB) and preexisting immunity for targeted treatment of GBM with an oncolytic virus. In this study, two kinds of cell membrane-coated oncolytic adenoviruses (NCM-Ad and GCM-Ad) were prepared using neural stem cells (NSCs) and GBM cells as sources of membranes, respectively, and were shown to improve the targeted infectivity on GBM cells and avoid the immune clearance of preexisting neutralizing antibodies in vitro and in vivo. Specifically, NCM-Ad showed a strong ability to cross the BBB and target tumor cells in vivo. To improve the cytotoxicity to GBM, a capsid dual-modified oncolytic adenovirus (A4/k37) was also encapsulated, and NCM-A4/k37 showed outstanding tumor targeting and inhibition capacity in an orthotopic xenograft tumor model of GBM upon intravenous administration. This study provides a promising oncolytic virus-based targeted therapeutic strategy for glioma.
Assuntos
Neoplasias Encefálicas , Glioblastoma , Terapia Viral Oncolítica , Vírus Oncolíticos , Humanos , Glioblastoma/terapia , Glioblastoma/patologia , Adenoviridae/genética , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Ensaios Antitumorais Modelo de Xenoenxerto , Vírus Oncolíticos/genética , Membrana Celular/metabolismoRESUMO
With the advent of cancer immunotherapy, there is a growing interest in vaccine development as a means to activate the cellular immune system against cancer. Despite the promise of DNA vaccines in this regard, their effectiveness is hindered by poor immunogenicity, leading to modest therapeutic outcomes across various cancers. The role of Type 1 conventional dendritic cells (cDC1), capable of cross-presenting vaccine antigens to activate CD8+T cells, emerges as crucial for the antitumor function of DNA vaccines. To address the limitations of DNA vaccines, a promising approach involves targeting antigens to cDC1 through the fusion of XCL1, a ligand specific to the receptor XCR1 on the surface of cDC1. Here, female C57BL/6 mice were selected for tumor inoculation and immunotherapy. Additionally, recognizing the complexity of cancer, this study explored the use of combination therapies, particularly the combination of cDC1-targeted DNA vaccine with the chemotherapy drug Gemcitabine (Gem) and the anti-PD1 antibody in a mouse lung cancer model. The study's findings indicate that fusion antigens with XCL1 effectively enhance both the immunogenicity and antitumor effects of DNA vaccines. Moreover, the combination of the cDC1-targeted DNA vaccine with Gemcitabine and anti-PD1 antibody in the mouse lung cancer model demonstrates an improved antitumor effect, leading to the prolonged survival of mice. In conclusion, this research provides important support for the clinical investigation of cDC1-targeting DNA vaccines in combination with other therapies.
Assuntos
Vacinas Anticâncer , Neoplasias Pulmonares , Vacinas de DNA , Animais , Feminino , Camundongos , Linfócitos T CD8-Positivos , Células Dendríticas , Gencitabina , Neoplasias Pulmonares/terapia , Camundongos Endogâmicos C57BL , Vacinas de DNA/imunologia , Vacinas de DNA/uso terapêutico , Vacinas Anticâncer/imunologia , Vacinas Anticâncer/uso terapêuticoRESUMO
Viral infection generally induces polyclonal neutralizing antibody responses. However, how many lineages of antibody responses can fully represent the neutralization activities in sera has not been well studied. Using the newly designed stable HIV-1 Env trimer as hook, we isolated two distinct broadly neutralizing antibodies (bnAbs) from Chinese rhesus macaques infected with SHIV1157ipd3N4 for 5 years. One lineage of neutralizing antibodies (JT15 and JT16) targeted the V2-apex in the Env trimers, similar to the J038 lineage bnAbs identified in our previous study. The other lineage neutralizing antibody (JT18) targeted the V3 crown region in the Env, which strongly competed with human 447-52D. Each lineage antibody neutralized a different set of viruses. Interestingly, when the two neutralizing antibodies from different lineages isolated from the same macaque were combined, the mixture had a neutralization breath very similar to that from the cognate sera. Our study demonstrated that a minimum of two different neutralizing antibodies can fully recapitulate the serum neutralization breadth. This observation can have important implications in AIDS vaccine design.
Assuntos
Anticorpos Neutralizantes , Anticorpos Anti-HIV , HIV-1 , Macaca mulatta , Síndrome de Imunodeficiência Adquirida dos Símios , Macaca mulatta/imunologia , Animais , HIV-1/imunologia , Anticorpos Anti-HIV/imunologia , Anticorpos Anti-HIV/sangue , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/sangue , Humanos , Síndrome de Imunodeficiência Adquirida dos Símios/imunologia , Síndrome de Imunodeficiência Adquirida dos Símios/sangue , Síndrome de Imunodeficiência Adquirida dos Símios/virologia , Infecções por HIV/imunologia , Infecções por HIV/virologia , Infecções por HIV/sangue , Vírus da Imunodeficiência Símia/imunologia , Produtos do Gene env do Vírus da Imunodeficiência Humana/imunologia , Testes de NeutralizaçãoRESUMO
Apolipoprotein B mRNA-editing catalytic polypeptide-like 3 family members (APOBEC3s) are host restriction factors that inhibit viral replication. Viral infectivity factor (Vif), a human immunodeficiency virus type 1 (HIV-1) accessory protein, mediates the degradation of APOBEC3s by forming the Vif-E3 complex, in which core-binding factor beta (CBFß) is an essential molecular chaperone. Here, we screened nonfunctional Vif mutants with high affinity for CBFß to inhibit HIV-1 in a dominant negative manner. We applied the yeast surface display technology to express Vif random mutant libraries, and mutants showing high CBFß affinity were screened using flow cytometry. Most of the screened Vif mutants containing random mutations of different frequencies were able to rescue APOBEC3G (A3G). In the subsequent screening, three of the mutants restricted HIV-1, recovered G-to-A hypermutation, and rescued APOBEC3s. Among them, Vif-6M showed a cross-protection effect toward APOBEC3C, APOBEC3F, and African green monkey A3G. Stable expression of Vif-6M in T lymphocytes inhibited the viral replication in newly HIV-1-infected cells and the chronically infected cell line H9/HXB2. Furthermore, the expression of Vif-6M provided a survival advantage to T lymphocytes infected with HIV-1. These results suggest that dominant negative Vif mutants acting on the Vif-CBFß target potently restrict HIV-1. IMPORTANCE Antiviral therapy cannot eliminate HIV and exhibits disadvantages such as drug resistance and toxicity. Therefore, novel strategies for inhibiting viral replication in patients with HIV are urgently needed. APOBEC3s in host cells are able to inhibit viral replication but are antagonized by HIV-1 Vif-mediated degradation. Therefore, we screened nonfunctional Vif mutants with high affinity for CBFß to compete with the wild-type Vif (wtVif) as a potential strategy to assist with HIV-1 treatment. Most screened mutants rescued the expression of A3G in the presence of wtVif, especially Vif-6M, which could protect various APOBEC3s and improve the incorporation of A3G into HIV-1 particles. Transduction of Vif-6M into T lymphocytes inhibited the replication of the newly infected virus and the chronically infected virus. These data suggest that Vif mutants targeting the Vif-CBFß interaction may be promising in the development of a new AIDS therapeutic strategy.
Assuntos
Subunidade beta de Fator de Ligação ao Core , Infecções por HIV , HIV-1 , Produtos do Gene vif do Vírus da Imunodeficiência Humana , Desaminases APOBEC/genética , Desaminases APOBEC/metabolismo , Animais , Linhagem Celular , Chlorocebus aethiops , Subunidade beta de Fator de Ligação ao Core/genética , HIV-1/genética , HIV-1/fisiologia , Interações Hospedeiro-Patógeno , Humanos , Linfócitos T/virologia , Replicação Viral , Produtos do Gene vif do Vírus da Imunodeficiência Humana/genéticaRESUMO
BACKGROUND: Progressive neuronal death is the key pathological feature of Alzheimer's disease (AD). However, the molecular mechanisms underlying the neuronal death in AD patients have not been fully elucidated. Necroptosis reportedly activates and induces neuronal death in patients with Alzheimer's disease (AD); however, the main mediators and mechanisms underlying necroptosis induction in AD remain elusive. METHODS: The function of hyperphosphorylated tau (pTau) in inducing necroptosis in neuronal cell was examined using Western blotting, RT-PCR and flow cytometry. Tau-induced inflammation was identified via RNA sequencing and transwell assay. Pharmacological methods and CRISPR-Cas9 technology were used to verify the role of necrosome proteins in pTau-stimulated neuronal death and inflammation. TauP301S model mice were treated with Nec-1 s to evaluate the role of necroptosis in tau pathology. RESULTS: Hyperphosphorylated tau could induce necroptosis in neuronal cells by promoting the formation of the RIPK1/RIPK3/MLKL necrosome. In addition, pTau significantly stimulated cell-autonomous overexpression of cytokines and chemokines via the intracellular nuclear factor kappa B (NF-κB) signaling pathway. Importantly, the RIPK1/RIPK3/MLKL axis was essential for the pTau-mediated NF-κB activation and cytokine storm. Furthermore, necroptosis stimulation, NF-κB activation, and cytokine induction have been detected in TauP301S mice and blocking necroptosis markedly ameliorated behavioral defects and excessive neuroinflammation in AD mice. CONCLUSIONS: Our study, for the first time, revealed that pTau contributes to neuronal death by inducing necroptosis and inflammation, mediated by activating the RIPK1/RIPK3/MLKL and NF-κB pathways, thereby delineating the hierarchical molecular network of neuronal necroptosis induction in AD.
Assuntos
Doença de Alzheimer , Necroptose , Doença de Alzheimer/genética , Animais , Apoptose/genética , Inflamação/patologia , Camundongos , NF-kappa B/metabolismo , Necrose/patologia , Proteínas Quinases/genética , Proteína Serina-Treonina Quinases de Interação com Receptores/genética , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismoRESUMO
HIV remains a health challenge worldwide, partly because of the continued development of resistance to drugs. Therefore, it is urgent to find new HIV inhibitors and targets. Apolipoprotein B mRNA-editing catalytic polypeptide-like 3 family members (APOBEC3) are important host restriction factors that inhibit HIV-1 replication by their cytidine deaminase activity. HIV-1 viral infectivity factor (Vif) promotes proteasomal degradation of APOBEC3 proteins by recruiting the E3 ubiquitin ligase complex, in which core-binding factor ß (CBFß) is a necessary molecular chaperone. Interrupting the interaction between Vif and CBFß can release APOBEC3 proteins to inhibit HIV-1 replication and may be useful for developing new drug targets for HIV-1. In this study, we identified a potent small molecule inhibitor CBFß/Vif-3 (CV-3) of HIV-1 replication by employing structure-based virtual screening using the crystal structure of Vif and CBFß (PDB: 4N9F) and validated CV-3's antiviral activity. We found that CV-3 specifically inhibited HIV-1 replication (IC50 = 8.16 µm; 50% cytotoxic concentration >100 µm) in nonpermissive lymphocytes. Furthermore, CV-3 treatment rescued APOBEC3 family members (human APOBEC3G (hA3G), hA3C, and hA3F) in the presence of Vif and enabled hA3G packaging into HIV-1 virions, which resulted in Gly-to-Ala hypermutations in viral genomes. Finally, we used FRET to demonstrate that CV-3 inhibited the interaction between Vif and CBFß by simultaneously forming hydrogen bonds with residues Gln-67, Ile-102, and Arg-131 of CBFß. These findings demonstrate that CV-3 can effectively inhibit HIV-1 by blocking the interaction between Vif and CBFß and that this interaction can serve as a new target for developing HIV-1 inhibitors.
Assuntos
Desaminases APOBEC/metabolismo , Fármacos Anti-HIV/química , Fármacos Anti-HIV/farmacologia , Subunidade beta de Fator de Ligação ao Core/metabolismo , HIV-1/efeitos dos fármacos , Produtos do Gene vif do Vírus da Imunodeficiência Humana/metabolismo , Linhagem Celular , Infecções por HIV/tratamento farmacológico , Infecções por HIV/metabolismo , HIV-1/fisiologia , Interações Hospedeiro-Patógeno/efeitos dos fármacos , Humanos , Simulação de Acoplamento Molecular , Mapas de Interação de Proteínas/efeitos dos fármacos , Proteólise/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Replicação Viral/efeitos dos fármacosRESUMO
SAM and HD domain-containing protein 1 (SAMHD1) is a host factor that restricts reverse transcription of lentiviruses such as HIV in myeloid cells and resting T cells through its dNTP triphosphohydrolase (dNTPase) activity. Lentiviruses counteract this restriction by expressing the accessory protein Vpx or Vpr, which targets SAMHD1 for proteasomal degradation. SAMHD1 is conserved among mammals, and the feline and bovine SAMHD1 proteins (fSAM and bSAM) restrict lentiviruses by reducing cellular dNTP concentrations. However, the functional regions of fSAM and bSAM that are required for their biological functions are not well-characterized. Here, to establish alternative models to investigate SAMHD1 in vivo, we studied the restriction profile of fSAM and bSAM against different primate lentiviruses. We found that both fSAM and bSAM strongly restrict primate lentiviruses and that Vpx induces the proteasomal degradation of both fSAM and bSAM. Further investigation identified one and five amino acid sites in the C-terminal domain (CTD) of fSAM and bSAM, respectively, that are required for Vpx-mediated degradation. We also found that the CTD of bSAM is directly involved in mediating bSAM's antiviral activity by regulating dNTPase activity, whereas the CTD of fSAM is not. Our results suggest that the CTDs of fSAM and bSAM have important roles in their antiviral functions. These findings advance our understanding of the mechanism of fSAM- and bSAM-mediated viral restriction and might inform strategies for improving HIV animal models.
Assuntos
HIV/genética , Lentivirus/genética , Transcrição Reversa/genética , Proteína 1 com Domínio SAM e Domínio HD/genética , Animais , Gatos , Bovinos , Células HEK293 , HIV/patogenicidade , Interações Hospedeiro-Patógeno/genética , Humanos , Lentivirus/patogenicidade , Células Mieloides/virologia , Domínios Proteicos/genética , Proteína 1 com Domínio SAM e Domínio HD/química , Linfócitos T/virologia , Replicação Viral/genéticaRESUMO
BACKGROUND: Tau hyper-phosphorylation has been considered a major contributor to neurodegeneration in Alzheimer's disease (AD) and related tauopathies, and has gained prominence in therapeutic development for AD. To elucidate the pathogenic mechanisms underlying AD and evaluate therapeutic approaches targeting tau, numerous transgenic mouse models that recapitulate critical AD-like pathology have been developed. Tau P301S transgenic mice is one of the most widely used mouse models in AD research. Extensive studies have demonstrated that sex significantly influences AD pathology, behavioral status, and therapeutic outcomes, suggesting that studies using mouse models of AD must consider sex- and age-related differences in neuropathology, behavior, and plasma content. METHOD: We systematically investigated differences in tau P301S transgenic mice (PS19 line) and wildtype littermates of different sex behavioral performance, tau neuropathology, and biomarkers in plasma and brain. RESULTS: Male P301S transgenic mice exhibited significant changes in weight loss, survival rate, clasping, kyphosis, composite phenotype assessment, nest building performance, tau phosphorylation at Ser202/Thr205, and astrocyte activation compared to that of wild-type littermates. In contrast, female P301S transgenic mice were only sensitive in the Morris water maze and open field test. In addition, we characterized the absence of macrophage-inflammatory protein (MIP-3α) and the upregulation of interferon (IFN)-γ, interleukin (IL)-5, and IL-6 in the plasma of P301S transgenic mice, which can be served as potential plasma biomarkers in P301S Tg mice. Male P301S transgenic mice expressed more monokine induced by IFN-γ (MIG), tumor necrosis factor-α (TNF-α), IL-10, and IL-13 than those of female P301S mice. CONCLUSION: Our findings highlight sexual dimorphism in the behavior, neuropathology, and plasma proteins in tau P301S transgenic AD mice, indicating that the use of male P301S transgenic mice may be more suitable for assessing anti-phosphorylated tau therapeutic strategies for AD and related tauopathies, and the MIP-3α may be a new potential plasma biomarker.
Assuntos
Doença de Alzheimer , Quimiocina CCL20/sangue , Modelos Animais de Doenças , Caracteres Sexuais , Proteínas tau/genética , Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Animais , Biomarcadores/sangue , Feminino , Humanos , Masculino , Camundongos , Camundongos TransgênicosRESUMO
BST2 is a single-pass type II transmembrane (TM) protein, which has a cytoplasmic domain, a transmembrane domain, and an extracellular domain, each domain is important for biologic function of BST2. BST2 is a host restriction factor that can effectively inhibit retrovirus release. Rhesus monkeys are considered as relevant natural animal models for studying AIDS in humans. In order to recognize rhesus BST2 (RhBST2) protein and detect its function accurately, we prepared a polyclonal antibody (pAb) especially for RhBST2. Meanwhile, we constructed RhBST2 proteins with the addition of an HA-tag at the N-terminus (RhBST2-NHA) or inside of the ectodomain (RhBST2-IHA) to compare the recognition ability of rabbit anti-RhBST2 pAb and anti-HA mAb. The results showed that the anti-HA mAb and rabbit anti-RhBST2 pAb had the same ability to identify RhBST2. RhBST2 demonstrated antiviral activity and the ability to activate NF-κB. Moreover, the N-glycosylation states, cell surface level and intracellular localization of RhBST2 were detected. However, HA tags relatively changed part of the biological function of RhBST2. These results show that the RhBST2 polyclonal antibody is more suitable for analyzing the properties and functions of RhBST2, and the natural domain of RhBST2 is very important for its function.
Assuntos
Anticorpos/imunologia , Antígenos CD , Antivirais , Síndrome da Imunodeficiência Adquirida , Animais , Antígenos CD/biossíntese , Antígenos CD/química , Antígenos CD/imunologia , Antivirais/química , Antivirais/imunologia , Antivirais/metabolismo , Proteínas Ligadas por GPI/biossíntese , Proteínas Ligadas por GPI/química , Proteínas Ligadas por GPI/imunologia , Células HEK293 , HIV-1/imunologia , Humanos , Macaca mulatta , Domínios Proteicos , CoelhosRESUMO
An integrase-defective SIV (idSIV) vaccine delivered by a DNA prime and viral particle boost approach can suppress viral loads (VLs) during the acute infection stage after intravenous SIVmac239 challenge. This study investigated how idSIV DNA and viral particle immunization alone contributed to the suppression of VLs in Chinese rhesus macaques after SIV challenge. Two macaques were immunized with idSIV DNA five times and two macaques were immunized with idSIV viral particles three times. Cellular and humoral immune responses were measured in the vaccinated macaques after immunization. The VLs and CD4+ T cell counts were monitored for 28 weeks after the intravenous SIVmac239 challenge. The SIV-specific T cell responses were only detected in the DNA-vaccinated macaques. However, binding and neutralizing antibodies against autologous and heterologous viruses were moderately better in macaques immunized with viral particles than in macaques immunized with DNA. After the challenge, the mean peak viremia in the DNA group was 2.3 logs lower than that in the control group, while they were similar between the viral particle immunization and control groups. Similar CD4+ T cell counts were observed among all groups. These results suggest that idSIV DNA immunization alone reduces VLs during acute infection after SIV challenge in macaques and may serve as a key component in combination with other immunogens as prophylactic vaccines.
Assuntos
Provírus/imunologia , Vacinas contra a SAIDS/imunologia , Vacinas de DNA/imunologia , Viremia/prevenção & controle , Animais , Anticorpos Neutralizantes , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Linfócitos T CD4-Positivos , Modelos Animais de Doenças , Imunidade Humoral , Imunização , Macaca mulatta , Provírus/genética , Vacinas contra a SAIDS/genética , Vacinas contra a SAIDS/uso terapêutico , Vírus da Imunodeficiência Símia/genética , Vacinação , Vacinas de DNA/uso terapêutico , Carga ViralRESUMO
Castration-resistant progression of prostate cancer is a major cause of prostate cancer mortality, and increased expression and activity of the full-length and the splice variants of androgen receptor (AR) have been indicated to drive castration resistance. Consequently, there is an urgent need to develop agents that can target both the full-length and the splice variants of AR for more effective treatment of prostate cancer. In the present study, we showed that raddeanin A (RA), an oleanane-type triterpenoid saponin, suppresses the transcriptional activities of both the full-length and the splice variants of AR. This is attributable to their decreased expression as a result of RA induction of proteasome-mediated degradation and inhibition of the transcription of the AR gene. We further showed the potential of using RA to enhance the growth inhibitory efficacy of docetaxel, the first-line chemotherapy for prostate cancer. This study identifies RA as a new agent to target both the full-length and the splice variants of AR and provides a rationale for further developing RA for prostate cancer treatment.
Assuntos
Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Neoplasias da Próstata/genética , Splicing de RNA , Receptores Androgênicos/genética , Saponinas/uso terapêutico , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Docetaxel/uso terapêutico , Regulação para Baixo/efeitos dos fármacos , Sinergismo Farmacológico , Humanos , Masculino , Células PC-3 , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/patologia , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/patologia , Isoformas de Proteínas/genéticaRESUMO
Functional E-cadherin loss, a hallmark of epithelial-mesenchymal transition (EMT), is important for metastasis. However, the mechanism of Snail2 in hepatocellular carcinoma (HCC) EMT and metastasis remains unclear. Here, we showed that Snail2 was upregulated in primary HCC, and significantly increased during transforming growth factor-ß-induced liver cell EMT. Snail2-overexpressing and knockdown cell lines have been established to determine its function in EMT in HCC. H3K9 methylation was upregulated and H3K4 and H3K56 acetylation were downregulated at the E-cadherin promoter in Snail2-overexpressing cancer cells. Furthermore, Snail2 interacted with G9a and histone deacetylases (HDACs) to form a complex to suppress E-cadherin transcription. Snail2 overexpression enhanced migration and invasion in HCC cells, whereas G9a and HDAC inhibition significantly reversed this effect. Moreover, Snail2 overexpression in cancer cells increased tumor metastasis and shortened survival time in mice, whereas G9a and HDAC inhibitors extended survival. Our study not only reveals a critical mechanism underlying the epigenetic regulation of EMT but also suggests novel treatment strategies for HCC.
Assuntos
Caderinas/metabolismo , Carcinoma Hepatocelular/metabolismo , Epigênese Genética , Transição Epitelial-Mesenquimal/genética , Antígenos de Histocompatibilidade/metabolismo , Histona Desacetilases/metabolismo , Histona-Lisina N-Metiltransferase/metabolismo , Neoplasias Hepáticas/metabolismo , Fatores de Transcrição da Família Snail/metabolismo , Acetilação , Animais , Azepinas/uso terapêutico , Carcinoma Hepatocelular/secundário , Movimento Celular , Progressão da Doença , Regulação para Baixo , Feminino , Inibidores de Histona Desacetilases/uso terapêutico , Humanos , Ácidos Hidroxâmicos/uso terapêutico , Fígado/efeitos dos fármacos , Fígado/metabolismo , Neoplasias Hepáticas/patologia , Metilação , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Invasividade Neoplásica , Metástase Neoplásica , Quinazolinas/uso terapêutico , Transcrição Gênica , Fator de Crescimento Transformador beta/farmacologiaRESUMO
BACKGROUND: Active immunotherapy targeting amyloid-ß (Aß) is a promising treatment for Alzheimer's disease (AD). Numerous preclinical studies and clinical trials demonstrated that a safe and effective AD vaccine should induce high titers of anti-Aß antibodies while avoiding the activation of T cells specific to Aß. RESULTS: An untagged Aß1-6 chimeric protein vaccine against AD based on norovirus (NoV) P particle was expressed in Escherichia coli and obtained by sequential chromatography. Analysis of protein characteristics showed that the untagged Aß1-6 chimeric protein expressed in soluble form exhibited the highest particle homogeneity, with highest purity and minimal host cell protein (HCP) and residual DNA content. Importantly, the untagged Aß1-6 chimeric soluble protein could induce the strongest Aß-specific humoral immune responses without activation of harmful Aß-specific T cells in mice. CONCLUSIONS: The untagged Aß1-6 chimeric protein vaccine is safe and highly immunogenic. Further research will determine the efficacy in cognitive improvement and disease progression delay.
Assuntos
Vacinas contra Alzheimer/imunologia , Peptídeos beta-Amiloides/imunologia , Anticorpos/imunologia , Vetores Genéticos , Norovirus , Proteínas Recombinantes de Fusão , Doença de Alzheimer/terapia , Vacinas contra Alzheimer/administração & dosagem , Peptídeos beta-Amiloides/química , Peptídeos beta-Amiloides/isolamento & purificação , Animais , Especificidade de Anticorpos/imunologia , Modelos Animais de Doenças , Imunização , Camundongos , Norovirus/imunologia , Linfócitos T/imunologia , Linfócitos T/metabolismoRESUMO
The Norovirus (NoV) P domain, with three surface loops for foreign antigen insertion, has been demonstrated as an excellent platform for antigen presentation and novel vaccine development. The P domain alone can self-assemble into a P dimer, 12-mer small particle or 24-mer P particle, and vaccines based on those particles may elicit different levels of immunogenicity. Currently, P particles are generally produced in soluble expression systems in Escherichia coli, mainly in the 24-mer form. However, the low yield of the soluble protein has hindered further clinical applications of P particle-based protein vaccines. In this study, we inserted the Alzheimer's disease (AD) immunogen Aß1-6 into the three loops of the P particle to generate an AD protein vaccine. To increase the yield of this chimeric protein, we tested the generation of proteins in a soluble expression system and an inclusion body expression system separately in E. coli. The result showed that the inclusion body expression system could greatly enhance the product yield of the chimeric protein compared with the soluble expression system. The refolded protein from the inclusion bodies was mainly in the 12-mer form, while the protein generated from the soluble supernatant was mainly in the 24-mer form. Moreover, the immunogenicity of soluble proteins was significantly stronger than that of the refolded proteins. Thus, comparisons between the two expression methods suggested that the soluble expression system generated chimeric P particles with better immunogenicity, while inclusion body expression system yielded more P particle proteins.