Prokaryote-derived phosphorylated Tau epitope vaccine is immunogenic and non-T-cell activated in the mice model.

Accumulation of phosphorylated Tau protein is a prominent pathological hallmark of Alzheimer's disease (AD). However, current vaccines targeting phosphorylation sites are primarily modified using chemical reactions, which exhibit low efficiency in terms of linking to the vaccine carrier. Despite the identification of over 2000 phosphorylation sites on approximately 20% of E. coli proteins through proteomic studies, it remains unclear whether recombinant Tau proteins expressed in bacteria undergo direct phosphorylation. Additionally, limited information is available regarding the immunogenicity and safety profiles of prokaryotic-derived pTau epitope vaccines. Our study discovered that the prokaryotic system can induce phosphorylation on four residues (T181, T205, S262, and S396) of the full-length Tau protein. Based on this finding, we developed a prokaryotic-modified phosphorylated Tau protein vaccine and immunized wild-type mice, resulting in enhanced immunogenicity and a favorable safety profile.

Hyperphosphorylated tau mediates neuronal death by inducing necroptosis and inflammation in Alzheimer's disease.

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.

ZEB2/TWIST1/PRMT5/NuRD Multicomplex Contributes to the Epigenetic Regulation of EMT and Metastasis in Colorectal Carcinoma.

(1) Background: The EMT plays a crucial role in tumor metastasis, which is the major cause for colorectal carcinoma-related mortality. However, the underlying regulators and mechanisms of EMT in CRC metastasis are still poorly understood; (2) Methods: The transcriptional regulators of EMT in CRC and their functions were examined using RT2212PCR, Western blotting, and luciferase reporter assay. The components of ZEB2/TWIST1 complex and their mutual interactions were identified via affinity purification, mass spectrometry, co-immunoprecipitation, and pull-down experiments. The functional mechanisms of ZEB2/TWIST1/PRMT5/NuRD axis were determined by chromatin immunoprecipitation and luciferase reporter assay. The contribution of ZEB2/TWIST1/PRMT5/NuRD complex in the CRC metastasis was investigated using wound healing, transwell assay, and in vivo xenograft mouse model; (3) Results: We found that ZEB2 and TWIST1 were both significantly upregulated in CRC tissues and EMT of CRC cells. ZEB2 could recruit TWIST1 to the E-cadherin promoter and synergistically repressed its transcription. In addition, ZEB2 physically interacted with TWIST1, PRMT5, and the nucleosome remodeling and deacetylase (NuRD) complex to form a novel repressive multicomplex, leading to epigenetic silencing of E-cadherin in CRC cells. Notably, the combined inhibition of ZEB2 and TWIST1 and epigenetic inhibition markedly reduced CRC metastasis in mice; (4) Conclusions: We revealed for the first time that ZEB2 could recruit TWIST1, PRMT5, and NuRD to form a repressive multicomplex and epigenetically suppresses the transcription of E-cadherin, thereby inducing the EMT process and metastasis in CRC. Our results also confirmed the therapeutic potential of epigenetic inhibitors in CRC.

Discovery of thiosemicarbazone derivatives as effective New Delhi metallo-ß-lactamase-1 (NDM-1) inhibitors against NDM-1 producing clinical isolates.

New Delhi metallo-ß-lactamase-1 (NDM-1) is capable of hydrolyzing nearly all ß-lactam antibiotics, posing an emerging threat to public health. There are currently less effective treatment options for treating NDM-1 positive "superbug", and no promising NDM-1 inhibitors were used in clinical practice. In this study, structure-activity relationship based on thiosemicarbazone derivatives was systematically characterized and their potential activities combined with meropenem (MEM) were evaluated. Compounds 19bg and 19bh exhibited excellent activity against 10 NDM-positive isolate clinical isolates in reversing MEM resistance. Further studies demonstrated compounds 19bg and 19bh were uncompetitive NDM-1 inhibitors with Ki = 0.63 and 0.44 µmol/L, respectively. Molecular docking speculated that compounds 19bg and 19bh were most likely to bind in the allosteric pocket which would affect the catalytic effect of NDM-1 on the substrate meropenem. Toxicity evaluation experiment showed that no hemolysis activities even at concentrations of 1000 mg/mL against red blood cells. In vivo experimental results showed combination of MEM and compound 19bh was markedly effective in treating infections caused by NDM-1 positive strain and prolonging the survival time of sepsis mice. Our finding showed that compound 19bh might be a promising lead in developing new inhibitor to treat NDM-1 producing superbug.

Stimulation Effects and Mechanisms of Different Adjuvants on a Norovirus P Particle-Based Active Amyloid-ß Vaccine.

BACKGROUND: Adjuvants are important components of vaccines and effectively enhance the immune response of specific antigens. However, the role of adjuvants or combinations of adjuvants in stimulating immunogenicity of the amyloid-ß (Aß) vaccine, as well as molecular mechanisms underlying such stimulation still remain unclear. A previous study of ours developed a norovirus P particle-based active Aß epitope vaccine, PP-3copy-Aß1-6-loop123, which stimulates a high titer of Aß-specific antibodies in mouse Alzheimer's disease (AD) models. OBJECTIVE: The most effective and safe adjuvant that maximizes the immunogenicity of our protein vaccine was determined. METHODS: We investigated four adjuvants (CpG, AS02, AS03, and MF59), and combinations of those, for capacity to enhance immunogenicity, and performed transcriptome analysis to explore mechanisms underlying the role of these in AD immunotherapy. RESULTS: Addition of the adjuvant, AS02, remarkably improved the immunogenicity of the PP-3copy-Aß1-6-loop123 vaccine without triggering an Aß-specific T-cell response. Combinations of adjuvants, particularly CpG +  AS02 and CpG + AS03, elicited a significantly elevated and prolonged Aß-specific antibody response. Gene ontology and Kyoto Encyclopedia of Genes and Genomes analyses indicated that a combination of two adjuvants was more effective in activating immune-related pathways, thereby enhancing the immunogenicity of PP-3copy-Aß1-6-loop123. CONCLUSION: These findings demonstrated that adjuvants can be used as enhancers in AD protein vaccination, and that a combination of CpG and AS-related adjuvants may be a very effective adjuvant candidate suitable for further clinical trials of the PP-3copy-Aß1-6-loop123 vaccine. Our studies also revealed potential mechanisms underlying the stimulation of immune response of protein vaccines by adjuvants.

The behavioural and neuropathologic sexual dimorphism and absence of MIP-3α in tau P301S mouse model of Alzheimer's disease.

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.

USP28 contributes to the proliferation and metastasis of gastric cancer.

USP28, a member of the deubiquitinating enzymes family, plays a vital role in the physiological process of cell proliferation, differentiation and apoptosis, DNA repair, immune response, and stress response. USP28 has been reported to be overexpressed in bladder cancer, colon cancer, breast carcinomas, and so on. Nevertheless, the role of USP28 in gastric cancer has not yet been investigated. In our study, we examined the USP28 expression in 87 paired samples of gastric cancer and normal gastric tissues. We found that USP28 was overexpressed in gastric cancer compared with normal gastric tissues (P < 0.01), and its overexpression was related to the degree of differentiation and metastases. Inhibiting USP28 expression in vitro suppressed the proliferation and invasion of gastric cancer cells by downregulating lysine specific demethylase 1. On the basis of our data, it can be concluded that USP28 may be a novel therapeutic target for gastric cancer.

Lysine-specific demethylase 1 activation by vitamin B2 attenuates efficacy of apatinib for proliferation and migration of gastric cancer cell MGC-803.

B vitamins play an essential role in the biosynthesis of nucleotides, replication of DNA, supply of methyl-groups, growth and repair of cells, aberrancies of which have all been implicated in carcinogenesis. Although the potential role of vitamin B in relation to the risk of cancer, including breast, and colorectal cancer, has been investigated in several observational studies, the mechanism of action is still unclear. In this study, vitamin B2 exhibited efficient activation of LSD1 by occupying the active sites where FAD stands. Interestingly, vitamin B2 significantly downregulated expression of CD86, a sensitive surrogate biomarker of LSD1 inhibition, and showed marked activation of gastric cancer cell migration and invasion. Meanwhile, vitamin B2 induced activation of LSD1 may attenuate the proliferation inhibition, and anti-migration effects of apatinib in gastric cancer cells. These findings suggested that vitamin B supplementation may interfere with the efficacy of apatinib in patients with gastric cancer.

Transcriptional response of human umbilical vein endothelial cell to H9N2 influenza virus infection.

Endothelial cells are believed to play an important role in response to virus infection. Here, we used a microarray technology to study the gene expression profile in human umbilical vein endothelial cells at 24h postinfection with H9N2 viruses or inactivated H9N2 viral particles. The results showed that H9N2 virus infection induced an abundance of differential expressed genes, exhibiting a transcriptional signature of viral infection. High levels of chemokine gene expressions were detected following treatment. Surprisingly, the most significantly up-regulated genes were mainly interferon-stimulated genes (ISGs), although there was no change in interferon gene expression and interferon protein level. We also found that viral particles were more potent than viruses in inducing ISGs expression. These results suggest that induction of expression of ISGs is mainly dependent on the interaction between viral particles and endothelial cells. Our data offer further insight into the interaction between endothelial cells and H9N2 influenza viruses.

Expression of hypoxia-inducible factor 1α mRNA in hearts and lungs of broiler chickens with ascites syndrome induced by excess salt in drinking water.

Hypoxia-inducible factor 1 (HIF-1) is a ubiquitously expressed heterodimeric transcription factor that mediates adaptive responses to hypoxia in all nucleated cells of metazoan organisms. Hypoxia-inducible factor 1α is involved in the pathogenesis of pulmonary hypertension in humans and animals, but whether HIF-1α is associated with the development of pulmonary hypertension syndrome (also known as ascites syndrome, AS) in broiler chickens has not been determined. In the present paper we addressed this issue by measuring the expression of HIF-1α mRNA in hearts and lungs of broiler chickens with AS induced by excess salt in drinking water. We conducted 2 experiments. The first experiment was used to observe the effects of excess salt on AS incidence. The results indicated that total incidence (20%) of AS in excess salt group (receiving 0.3% NaCl in drinking water) was much higher compared with the control group (receiving tap water) over a 43-d time course (P < 0.05). In the second experiment, we determined mean pulmonary arterial pressure (mPAP), ascites heart index (AHI), and expression of HIF-1α mRNA in lungs and hearts of broiler chickens after the excess salt treatment. Our results showed that excess salt induced pulmonary hypertension (indicated by higher mPAP) and right ventricular hypertrophy (greater ascites heart index) in broiler chickens. Meanwhile, the expression levels of HIF-1α mRNA in lungs and hearts were significantly increased at different time points in the excess salt group compared with the control group. Linear correlation analysis showed that the expression of HIF-1α mRNA in lungs was significantly positively correlated with mPAP (correlation coefficient = 0.79, P < 0.001), demonstrating that expression of HIF-1α mRNA was gradually increased in the excess salt group with the increase of pulmonary arterial pressure. In addition, the ascitic chickens showed significantly higher transcriptional levels of HIF-1α in hearts and lungs, compared with the age-matched healthy chickens, respectively. Our findings hinted that HIF-1α might be associated with the development of AS induced by excess salt in drinking water in broiler chickens.

[Effects of extracts from different parts of Folium perillae (L.) Britt. on Hemorrheological parameters in rats].

In order to provide experimental data for the development and application of drug in clinic, we determined the effects of extracts from different parts of folium perillae (L. ) Britt. on hemorheological parameters, extracted from leaves (folium perillae), seeds (fructus perillae) and peduncles (caulis perillae). The results showed that all extracts from different parts of folium perillae (L. ) Britt. can significantly reduce the whole blood viscosity at low shear rate (10 s(-1)), erythrocyte aggregation index, erythrocyte electrophoresis index (P<0.05), and the whole blood reductive viscosity at low shear rate (10 s(-1)) (P<0.01). Extracts from folium perillae and caulis perillae can significantly decrease erythrocyte deformation index (P<0.05), whereas extracts from fructus perillae can not. Extracts from fructus perillae and caulis perillae can significantly decrease plasma viscosity at low shear rate(10 s(-1)), but extracts of folium perillae can not. Aspirin can only decrease the whole blood reductive viscosity at low shear rate and plasma viscosity (P<0.05). All extracts from different parts of folium perillae (L. ) Britt. had no significant effects on hematocrit, erythrocyte rigidity index, fibrinogen concentration , the whole blood viscosity and the whole blood reductive viscosity at middle and high shear rate (60 s(-1),120 s(-1)).