[Efficacy and mechanism of Xuefu Zhuyu Decoction on model rats of coronary heart disease with heart blood stasis syndrome based on metabolomics].

This study investigated the effects of Xuefu Zhuyu Decoction on myocardial metabolites in a rat model of coronary heart disease with heart blood stasis syndrome and explored the therapeutic mechanism of blood circulation-promoting and blood stasis-removing therapy. SD rats were randomly divided into a sham operation group, a model group, a Xuefu Zhuyu Decoction group(14.04 g·kg~(-1)), and a trimetazidine group(5.4 mg·kg~(-1)). The sham operation group underwent thread insertion without ligation, while the other groups underwent coronary artery left anterior descending branch ligation to induce a model of coronary heart disease with heart blood stasis syndrome. Three days after modeling, drug intervention was performed, and samples were taken after 14 days of intervention. General conditions were observed, and electrocardiogram and cardiac ultrasound indices were measured. Hematoxylin-eosin(HE) staining and Masson staining were used to observe tissue pathological morphology. The enzyme linked immunosorbent assay(ELISA) was used to measure the levels of triglyceride(TG) and total cholesterol(TC) in the serum. Ultra high performance liquid chromatography-quantitative exactive-mass spectrometry(UHPLC-QE-MS) technology was used to screen differential metabolites in myocardial tissue and conduct metabolic pathway enrichment analysis. The results showed that Xuefu Zhuyu Decoction significantly improved the general condition of the model rats, reduced heart rate and ST segment elevation in the electrocardiogram, increased left ventricular ejection fraction(LVEF) and left ventricular fractional shortening(LVFS), and decreased left ventricular internal diameter in diastole(LVIDd) and left ventricular internal diameter in systole(LVIDs). HE staining and Masson staining showed that Xuefu Zhuyu Decoction effectively alleviated myocardial tissue structural disorders, inflammatory cell infiltration, and collagen fiber deposition in the model rats. ELISA results showed that Xuefu Zhuyu Decoction effectively regulated serum TG and TC levels in the model rats. There were significant differences in the metabolic phenotypes of myocardial samples in each group. Fourteen differential metabolites were identified in the Xuefu Zhuyu Decoction group, involving five metabolic pathways, including arginine and proline metabolism, glycerophospholipid metabolism, aminoacyl-tRNA biosynthesis, ether lipid metabolism, and alanine, aspartate, and glutamate metabolism. Xuefu Zhuyu Decoction improved cardiac function and myocardial structural damage in the rat model of coronary heart disease with heart blood stasis syndrome, and its biological mechanism involved the regulation of lipid metabolism, choline metabolism, amino acid metabolism, energy metabolism, and protein synthesis pathways.

A conformation-based intra-molecular initiation factor identified in the flavivirus RNA-dependent RNA polymerase.

The flaviviruses pose serious threats to human health. Being a natural fusion of a methyltransferase (MTase) and an RNA-dependent RNA polymerase (RdRP), NS5 is the most conserved flavivirus protein and an important antiviral target. Previously reported NS5 structures represented by those from the Japanese encephalitis virus (JEV) and Dengue virus serotype 3 (DENV3) exhibit two apparently different global conformations, defining two sets of intra-molecular MTase-RdRP interactions. However, whether these NS5 conformations are conserved in flaviviruses and their specific functions remain elusive. Here we report two forms of DENV serotype 2 (DENV2) NS5 crystal structures representing two conformational states with defined analogies to the JEV-mode and DENV3-mode conformations, respectively, demonstrating the conservation of both conformation modes and providing clues for how different conformational states may be interconnected. Data from in vitro polymerase assays further demonstrate that perturbing the JEV-mode but not the DENV3-mode intra-molecular interactions inhibits catalysis only at initiation, while the cell-based virological analysis suggests that both modes of interactions are important for virus proliferation. Our work highlights the role of MTase as a unique intra-molecular initiation factor specifically only through the JEV-mode conformation, providing an example of conformation-based crosstalk between naturally fused protein functional modules.

Pharmacological inhibition of MyD88 suppresses inflammation in tubular epithelial cells and prevents diabetic nephropathy in experimental mice.

Emerging evidence shows that chronic inflammation mediated by toll-like receptors (TLRs) contributes to diabetic nephropathy. Myeloid differentiation primary-response protein-88 (MyD88) is an essential adapter protein of all TLRs except TLR3 in innate immunity. It is unclear whether MyD88 could be a therapeutic target for diabetic nephropathy. Here, we used a new small-molecule MyD88 inhibitor, LM8, to examine the pharmacological inhibition of MyD88 in protecting kidneys from inflammatory injury in diabetes. We showed that MyD88 was significantly activated in the kidney of STZ-induced type 1 diabetic mice in tubular epithelial cells as well as in high glucose-treated rat tubular epithelial cells NRK-52E. In cultured tubular epithelial cells, we show that LM8 (2.5-10 µM) or MyD88 siRNA attenuated high-concentration glucose-induced inflammatory and fibrogenic responses through inhibition of MyD88-TLR4 interaction and downstream NF-κB activation. Treatment with LM8 (5, 10 mg/kg, i.g.) significantly reduced renal inflammation and fibrosis and preserved renal function in both type 1 and type 2 diabetic mice. These renoprotective effects were associated with reduced MyD88-TLR4 complex formation, suppressed NF-κB signaling, and prevention of inflammatory factor expression. Collectively, our results show that hyperglycemia activates MyD88 signaling cascade to induce renal inflammation, fibrosis, and dysfunction. Pharmacological inhibition of MyD88 may be a therapeutic approach to mitigate diabetic nephropathy and the inhibitor LM8 could be a potential candidate for such therapy.

SARS-CoV-2 replicon for high-throughput antiviral screening.

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) virus, which is highly pathogenic and classified as a biosafety level 3 (BSL-3) agent, has greatly threatened global health and efficacious antivirals are urgently needed. The high requirement of facilities to manipulate the live virus has limited the development of antiviral study. Here, we constructed a reporter replicon of SARS-CoV-2, which can be handled in a BSL-2 laboratory. The Renilla luciferase activity effectively reflected the transcription and replication levels of the replicon genome. We identified the suitability of the replicon in antiviral screening using the known inhibitors, and thus established the replicon-based high-throughput screening (HTS) assay for SARS-CoV-2. The application of the HTS assay was further validated using a few hit natural compounds, which were screened out in a SARS-CoV-2 induced cytopathic-effect-based HTS assay in our previous study. This replicon-based HTS assay will be a safe platform for SARS-CoV-2 antiviral screening in a BSL-2 laboratory without the live virus.

UBE2S exerts oncogenic activities in urinary bladder cancer by ubiquitinating TSC1.

Ubiquitin-conjugating enzyme E2S (UBE2S), an important E2 enzyme in the process of ubiquitination, has exhibited oncogenic activities in various malignant tumors. However, it remains unknown whether UBE2S plays a role in urinary bladder cancer (UBC) development. In the current study, our data confirmed UBE2S upregulation in UBC. In vitro and in vivo experiments demonstrated that UBE2S knockdown resulted in attenuated proliferation and enhanced apoptosis, which was inverse to the phenotypes with UBE2S overexpression. Gain and loss of function assays confirmed that UBE2S exerts oncogenic activities in UBC by mediating the activation of the mammalian target of rapamycin complex 1 (mTORC1) pathway. Furthermore, we discovered that this UBE2S-modulated carcinogenic mechanism was in the consequence of directly targeting tuberous sclerosis 1 (TSC1), which is the upstream inhibitor of mTOR signaling for ubiquitous degradation. Taken together, this study demonstrated that UBE2S is a carcinogen in UBC and promotes UBC progression by ubiquitously degrading TSC1. This consequently mediates the activation of the mTOR pathway, suggesting a potential therapeutic regimen for UBC by targeting the newly identified UBE2S/TSC1/mTOR axis.

Short Direct Repeats in the 3' Untranslated Region Are Involved in Subgenomic Flaviviral RNA Production.

Mosquito-borne flaviviruses consist of a positive-sense genome RNA flanked by the untranslated regions (UTRs). There is a panel of highly complex RNA structures in the UTRs with critical functions. For instance, Xrn1-resistant RNAs (xrRNAs) halt Xrn1 digestion, leading to the production of subgenomic flaviviral RNA (sfRNA). Conserved short direct repeats (DRs), also known as conserved sequences (CS) and repeated conserved sequences (RCS), have been identified as being among the RNA elements locating downstream of xrRNAs, but their biological function remains unknown. In this study, we revealed that the specific DRs are involved in the production of specific sfRNAs in both mammalian and mosquito cells. Biochemical assays and structural remodeling demonstrate that the base pairings in the stem of these DRs control sfRNA formation by maintaining the binding affinity of the corresponding xrRNAs to Xrn1. On the basis of these findings, we propose that DRs functions like a bracket holding the Xrn1-xrRNA complex for sfRNA formation.IMPORTANCE Flaviviruses include many important human pathogens. The production of subgenomic flaviviral RNAs (sfRNAs) is important for viral pathogenicity as a common feature of flaviviruses. sfRNAs are formed through the incomplete degradation of viral genomic RNA by the cytoplasmic 5'-3' exoribonuclease Xrn1 halted at the Xrn1-resistant RNA (xrRNA) structures within the 3'-UTR. The 3'-UTRs of the flavivirus genome also contain distinct short direct repeats (DRs), such as RCS3, CS3, RCS2, and CS2. However, the biological functions of these ancient primary DR sequences remain largely unknown. Here, we found that DR sequences are involved in sfRNA formation and viral virulence and provide novel targets for the rational design of live attenuated flavivirus vaccine.

Infectious Chikungunya Virus (CHIKV) with a Complete Capsid Deletion: a New Approach for a CHIKV Vaccine.

Chikungunya virus (CHIKV) is a mosquito-borne alphavirus that causes epidemics of debilitating disease worldwide. Currently, there are no licensed vaccines or antivirals available against CHIKV infection. In this study, we generated a novel live attenuated vaccine (LAV) candidate for CHIKV with a complete deficiency of capsid (ΔC-CHIKV). It could propagate in BHK-21 cells, and had antigenic properties similar to those of native CHIKV. Vaccination of either immunocompromised IFNAR-/- mice or immunocompetent C57BL/6 mice with a single dose of ΔC-CHIKV conferred complete protection upon challenge with wild-type (WT) CHIKV. Taken together, this vaccine candidate appeared to be safe and efficacious, representing a novel strategy for CHIKV vaccine design.IMPORTANCE Currently, there is no licensed vaccine against CHIKV infection. An ideal CHIKV vaccine should generate an optimal balance between efficacy and safety. Live attenuated vaccines that can elicit strong immune responses often involve a trade-off of reduced safety. Here, a novel live attenuated vaccine candidate for CHIKV lacking the entire capsid gene, ΔC-CHIKV, was developed. It was demonstrated to be genetically stable, highly attenuated, immunogenic, and able to confer complete protection against lethal CHIKV challenge after a single dose of immunization. Such an infectious vaccine candidate devoid of capsid provides a novel strategy for the development of a live attenuated CHIKV vaccine.

[Comparison between Ye Tianshi and Wu Jutong in treatment of summer-heat disease].

Through the traditional Chinese medicine inheritance platform system, with the help of medical records, Ye Tianshi and Wu Jutong's medication characteristics for summer heat sickness were analyzed, the laws of the two people's medication were summarized, and the similarities and differences between the two were explored to explore the relationship. As a result, it was found that both of them recognized the relationship between summer heat and wetness, and Wu Jutong believed that "wind" was also an important pathogenic factor. Both of the patients were treated with cold medicine and warm medicine. They used mostly bitter, sweet, pungent taste and lungs, spleen, stomach, and heart meridian are the main components; two are commonly used Armeniacae Semen Amarum, Talcum, Rehmanniae Radix, Ophiopogonis Radix, Pinelliae Rhizoma and other drugs, Ye Tianshi use Scrophulariae Radix, Tetrapanacis Medulla, Coicis Semen and other drugs more, Wu Jutong use Gypsum Fibrosum, Sojae Semen Praeparatum, Menthae Haplocalycis Herba and other drugs more; at the same time, a combination of two high-frequency medicines used by two people has been excavated, and a new prescription has been deduced to provide a reference for further understanding and treatment of summer diseases.

Recovery of the Zika virus through an in vitro ligation approach.

In this study, an in vitro ligation method was developed to assemble a full-length infectious cDNA clone of the Zika virus (ZIKV). Four contiguous cDNA subclones covering the complete ZIKV genome were constructed with unique BglI restriction sites at the ends of each fragment. The BglI restriction sites only allow in vitro ligation to happen between interconnecting restriction sites from adjacent cDNA fragments, resulting in an intact full-length cDNA of ZIKV. RNA transcripts derived from the full-length cDNA were infectious. The recombinant virus replicated as efficiently as the wild-type virus with similar growth kinetics and plaque morphologies in Vero and C6/36 cells. Both viruses were inhibited by NITD008 treatment. This in vitro ligation method will facilitate manipulation of the viral genome through genetic modifications of four separated subclones of ZIKV for the rapid and rational development of candidate vaccines and viral replication study.

Transmembrane Domains of NS2B Contribute to both Viral RNA Replication and Particle Formation in Japanese Encephalitis Virus.

UNLABELLED: Flavivirus nonstructural protein 2B (NS2B) is a transmembrane protein that functions as a cofactor for viral NS3 protease. The cytoplasmic region (amino acids 51 to 95) alone of NS2B is sufficient for NS3 protease activity, whereas the role of transmembrane domains (TMDs) remains obscure. Here, we demonstrate for the first time that flavivirus NS2B plays a critical role in virion assembly. Using Japanese encephalitis virus (JEV) as a model, we performed a systematic mutagenesis at the flavivirus conserved residues within the TMDs of NS2B. As expected, some mutations severely attenuated (L38A and R101A) or completely destroyed (G12L) viral RNA synthesis. Interestingly, two mutations (G37L and P112A) reduced viral RNA synthesis and blocked virion assembly. None of the mutations affected NS2B-NS3 protease activity. Because mutations G37L and P112A affected virion assembly, we selected revertant viruses for these two mutants. For mutant G37L, replacement with G37F, G37H, G37T, or G37S restored virion assembly. For mutant P112A, insertion of K at position K127 (leading to K127KK) of NS2B rescued virion assembly. A biomolecular fluorescent complementation (BiFC) analysis demonstrated that (i) mutation P112A selectively weakened NS2B-NS2A interaction and (ii) the adaptive mutation K127KK restored NS2B-NS2A interaction. Collectively, our results demonstrate that, in addition to being a cofactor for NS3 protease, flavivirus NS2B also functions in viral RNA replication, as well as virion assembly. IMPORTANCE: Many flaviviruses are important human pathogens. Understanding the molecular mechanisms of the viral infection cycle is essential for vaccine and antiviral development. In this study, we demonstrate that the TMDs of JEV NS2B participate in both viral RNA replication and virion assembly. A viral genetic study and a BiFC assay demonstrated that interaction between NS2B and NS2A may participate in modulating viral assembly in the flavivirus life cycle. Compensatory-mutation analysis confirmed that there was a correlation between viral assembly and NS2B-NS2A interaction. TMDs of NS2B may serve as novel antiviral targets to prevent flavivirus infection, and the structure determination of NS2B will help us to understand the functional mechanism of NS2B in viral RNA replication and assembly. The results have uncovered a new function of flavivirus NS2B in virion assembly, possibly through interaction with the NS2A protein.

Development of a stable Japanese encephalitis virus replicon cell line for antiviral screening.

Japanese encephalitis virus (JEV), an important pathogen in Eastern and Southern Asia and the Pacific, has spread to Australia and other territories in recent years. Although the vaccine for JEV has been used in some countries, development of efficient antiviral drugs is still an urgent requirement. Replicon systems have been widely used in the research of viral replication and antiviral screening for West Nile virus (WNV), yellow fever virus (YFV) and dengue virus (DENV). Here, a novel JEV replicon harboring the Rluc and Pac gene (JEV-Pac-Rluc-Rep) was constructed. Furthermore, we established a BHK-21 cell line harboring JEV-Pac-Rluc-Rep (BHK-21/PAC/Rluc cell line) through continuous puromycin selection. Characterization of cell line stability showed that the replicon RNA could persistently replicate in this cell line for at least up to 10 rounds of passage. Using a known flavivirus inhibitor, the JEV replicon cell line was validated for antiviral screening. The JEV replicon cell line will be a valuable tool for both compound screening and viral replication studies.

Induction of Heme Oxygenase-1 by Sodium 9-Hydroxyltanshinone IIA Sulfonate Derivative Contributes to Inhibit LPS-Mediated Inflammatory Response in Macrophages.

BACKGROUND/AIM: Sodium 9-acetoxyltanshinone IIA sulfonate (ZY-1A4), a novel compound derived from sodium 9-hydroxyltanshinone IIA sulfonate, was synthesized with potential biological activities. This study aimed to explore the effects of ZY-1A4 on lipopolysaccharide (LPS)-triggered inflammatory response and the underlying mechanisms. METHODS: Activation of RAW264.7 macrophages was induced by LPS. The effects of ZY-1A4 on inducible nitric oxide synthase (iNOS) expression, nitric oxide (NO) generation, nuclear factor-κB (NF-κB) activation, heme oxygenase-1 (HO-1) expression, and nuclear factor-erythroid 2 related factor 2 (Nrf2) pathway were evaluated to elucidate its underlying mechanisms on inflammatory responses. RESULTS: ZY-1A4 concentration-dependently reduced iNOS expression and NO production, and inhibited c-Jun-N-terminal kinase 1/2 (JNK1/2) phosphorylation and NF-κB activation in LPS-stimulated macrophages. In addition, ZY-1A4 concentration- and time-dependently induced HO-1 expression associated with degradation of Kelch-like ECH-associated protein 1 (Keap1) and nuclear translocation of Nrf2, while the effect of ZY-1A4 was abolished by a phosphoinositide 3-kinase (PI3K) inhibitor LY294002. Intriguingly, pharmacological inactivation of HO-1 with zinc protoporphyrin IX reversed anti-inflammatory effect of ZY- 1A4, but the anti-inflammatory effect of ZY-1A4 was largely mimicked by HO-1 by-products carbon monoxide and bilirubin. Furthermore, the inhibitory effect of ZY-1A4 on LPS-induced iNOS expression and NO release was abolished by HO-1 siRNA or LY294002. CONCLUSION: Our results demonstrated that ZY-1A4 suppressed LPS-induced iNOS expression and NO generation via modulation of NF-κB activation and HO-1 expression. This new finding might shed light to the prevention and therapy of cardiovascular diseases.

Identification of fangchinoline as a broad-spectrum enterovirus inhibitor through reporter virus based high-content screening.

Hand, foot, and mouth disease (HFMD) is a common pediatric illness mainly caused by enteroviruses, which are important human pathogens. Currently, there are no available antiviral agents for the therapy of enterovirus infection. In this study, an excellent high-content antiviral screening system utilizing the EV-A71-eGFP reporter virus was developed. Using this screening system, we screened a drug library containing 1042 natural compounds to identify potential EV-A71 inhibitors. Fangchinoline (FAN), a bis-benzylisoquinoline alkaloid, exhibits potential inhibitory effects against various enteroviruses that cause HFMD, such as EV-A71, CV-A10, CV-B3 and CV-A16. Further investigations revealed that FAN targets the early stage of the enterovirus life cycle. Through the selection of FAN-resistant EV-A71 viruses, we demonstrated that the VP1 protein could be a potential target of FAN, as two mutations in VP1 (E145G and V258I) resulted in viral resistance to FAN. Our research suggests that FAN is an efficient inhibitor of EV-A71 and has the potential to be a broad-spectrum antiviral drug against human enteroviruses.

[Study on HPLC-fingerprint-based identification of dao-di herb and non dao-di herb of scutellariae radix].

OBJECTIVE: To compare the discrepancies between chemical constituents in Dao-di herb and non Dao-di herb of Huangqin (the root of Scutellaria baicalensis), study the impact of habitat and growth pattern (including cultivated and wild Huangqin) on chemical substances of Huangqin, and then provide evidence for the identification of Dao-di herb and quality evaluation of Huangqin. METHOD: The chemical constituents in Huangqin collected from different habitats and under different growth patterns, were analyzed using HPLC fingerprint. The fingerprints obtained were then evaluated by hierarchical clustering analysis, principal component analysis and components peak area pattern. RESULT: The fingerprints' chemical profiles of Dao-di herb and non Dao-di Huangqin had significant disparity. The fingerprints of modem Dao-di herb Huangqin samples originated from Chengde (Hebei Province) were significantly different from those from other habitats, though the fingerprints of the non Dao-di Huangqin collected from Chifeng (Inner Mongolia) and Chengde had high similarity to each other. The chemical characteristics of Huangqin samples collected from the habitats recorded in ancient herbals, such as Qingyang (Gansu Province), Yan'an (Shaanxi Province), Linyi (Shangdong Province), Changzhi and Jinzhong (Shanxi Province) were similar. The fingerprints of modern non Dao-di samples collected from Dingxi and Longnan (Gansu Province) and Shangluo (Shaanxi Province) had high similarity. In addition, the content of acteoside in wild Huangqin was higher than that in cultivated Huangqin. CONCLUSION: Dao-di herb and non Dao-di herb of Huangqin could be distinguished using the developed HPLC fingerprints. The results obtained may provide evidence for the quality control and pharmcodynamical research of Dao-di herb and non Dao-di Huangqin.

Metric networks for enhanced perception of non-local semantic information.

Introduction: Metric learning, as a fundamental research direction in the field of computer vision, has played a crucial role in image matching. Traditional metric learning methods aim at constructing two-branch siamese neural networks to address the challenge of image matching, but they often overlook to cross-source and cross-view scenarios. Methods: In this article, a multi-branch metric learning model is proposed to address these limitations. The main contributions of this work are as follows: Firstly, we design a multi-branch siamese network model that enhances measurement reliability through information compensation among data points. Secondly, we construct a non-local information perception and fusion model, which accurately distinguishes positive and negative samples by fusing information at different scales. Thirdly, we enhance the model by integrating semantic information and establish an information consistency mapping between multiple branches, thereby improving the robustness in cross-source and cross-view scenarios. Results: Experimental tests which demonstrate the effectiveness of the proposed method are carried out under various conditions, including homologous, heterogeneous, multi-view, and crossview scenarios. Compared to the state-of-the-art comparison algorithms, our proposed algorithm achieves an improvement of ~1, 2, 1, and 1% in terms of similarity measurement Recall@10, respectively, under these four conditions. Discussion: In addition, our work provides an idea for improving the crossscene application ability of UAV positioning and navigation algorithm.

LWJ-M30, a conjugate of DM1 and B6, for the targeted therapy of colorectal cancer with improved therapeutic effects.

Colorectal cancer (CRC) is one of the most prevalent cancers worldwide as well as a significant cause of mortality. The conventional treatment could cause serious side effects and induce drug resistance, recurrence and metastasis of cancers. Hence, specific targeting of cancer cells without affecting the normal tissues is currently an urgent necessity in cancer therapy. The emerging of peptide-drug conjugates (PDC) is regarded as a promising approach to address malignant tumors. LWJ-M30, a conjugate of DM1 and B6 peptide, targeted transferrin receptors (TfRs) on the surface of the CRC cells, showing a powerful anti-cancer effect. LWJ-M30 significantly inhibited the HCT116 cells proliferation and migration in vitro. LWJ-M30 also dramatically decreased the level of polymeric tubulin, while the disruption of microtubules caused the cell cycle to be arrested in the G2/M phase. LWJ-M30 induced the HCT116 cells apoptosis both in vivo and in vitro. The results in vivo demonstrated that LWJ-M30 could inhibit the HCT116 growth without affecting the mouse body weight. Taking these results together, our data indicated that LWJ-M30 could improve the therapeutic effects of DM1 while reducing the systemic toxicity in normal tissues.

Mechanisms of Xuefu Zhuyu Decoction in the treatment of coronary heart disease based on integrated metabolomics and network pharmacology approach.

Coronary heart disease (CHD) has become the leading cause of mortality, morbidity, and disability worldwide. Though the therapeutic effect of Xuefu Zhuyu Decoction (XFZY) on CHD has been demonstrated in China, the active ingredients and molecular mechanisms of XFZY have not been elucidated. The purpose of the current study is to explore the molecular mechanisms of XFZY in the treatment of CHD via network pharmacology, metabolomics, and experimental validation. First, we established a CHD rat model by permanently ligating the left anterior descending coronary artery (LAD), and evaluated the therapeutic effect of XFZY by hemorheology and histopathology. Second, network pharmacology was employed to screen the active ingredients and potential targets of XFZY for the treatment of CHD. Metabolomic was applied to identify the molecules present in the serum after XFZY treatment. Third, the results of network pharmacology and metabolomics were further analyzed by Cytoscape to elucidate the core ingredients and pathways. Finally, the obtained key pathways were verified by transmission electron microscopy and immunofluorescence assay. The results showed that XFZY was effective in the treatment of CHD in the rat model, and the highest dose exerted the best effect. Network pharmacology analysis revealed 215 active ingredients and 129 key targets associated with XFZY treatment of CHD. These targets were enriched in pathways of cancer, lipid and atherosclerosis, fluid shear stress and atherosclerosis, proteoglycans in cancer, chemical carcinogenesis - receptor activation, HIF-1 signaling, et al. Serum metabolomic identified 1081 metabolites involved in the therapeutic effect of XFZY on CHD. These metabolites were enriched in taurine and hypotaurine metabolism, histidine metabolism, retrograde endocannabinoid signaling pathways, et al. Cytoscape analysis combining the data from serum metabolomic and network pharmacology revealed that energy metabolism as the core pathway for XFZY treatment of CHD. Electron microscope observation identified changes in the level of autophagy in the mitochondrial structure of cardiomyocytes. Immunofluorescence assay showed that the expression levels of autophagy-related proteins LC3-B and P62/SQSTM1 were consistent with the levels of autophagy observed in mitochondria. In conclusion, our findings suggest that the possible mechanisms of XFZY in the treatment of CHD are reducing the level of autophagy, improving energy metabolism, and maintaining mitochondrial homeostasis in cardiomyocytes. Our study also shows that the combined strategies of network pharmacology, metabolomics, and experimental validation may provide a powerful approach for TCM pharmacology study.

MUC15 is an independent prognostic factor that promotes metastases of MYCN non-amplified neuroblastoma.

Background: Neuroblastoma (NB) is a cancer that arises from neural-crest-derived sympathoadrenal lineage. Less is known about the pathogenesis and molecular characteristics of MYCN non-amplified (MYCN-NA) NB. Methods: We constructed a signature model targeting mucin family according to RNA sequencing data from GSE49710 dataset, and validated the prognostic performance. We also analyzed the gene expression matrix using DESeq2 R packages to screen the most differential mucin in high-risk NB samples. We further assessed its prognostic value, particularly in MYCN-NA NB samples. Moreover, we performed functional experiments to evaluate the impact of MUC15 overexpression on the migration of MYCN-NA NB cell lines. Results: The 8-mucin signature model showed good prognostic performance in the GSE49710 dataset. Among the mucin genes, MUC15 was significantly upregulated in the high-risk NB cohort and was associated with poor prognosis, especially in MYCN-NA NB samples. Furthermore, MUC15 overexpression and exogenous MUC15 protein enhanced the migration of MYCN-NA NB cell lines. Mechanistically, MUC15 promoted the phosphorylation of focal adhesion kinase (FAK) by inhibiting the expression of MYCT1, a target of c-Myc. Conclusions: Our findings suggested a potential network in controlling NB cell metastasis. Targeting MUC15 in MYCN-NA NB patients could be a promising therapeutic strategy.

Attenuated WNV-poly(A) exerts a broad-spectrum oncolytic effect by selective virus replication and CD8+ T cell-dependent immune response.

As an emerging tumor therapy, ideal oncolytic viruses preferentially replicate in malignant cells, reverse the immunosuppressive tumor microenvironment, and eventually can be eliminated by the patient. It is of great significance for cancer treatment to discover new excellent oncolytic viruses. Here, we found that WNV live attenuated vaccine WNV-poly(A) could be developed as a novel ideal oncolytic agent against several types of cancers. Mechanistically, due to its high sensitivity to type Ι interferon (IFN-Ι), WNV-poly(A) could specifically kill tumor cells rather than normal cells. At the same time, WNV-poly(A) could activate Dendritic cells (DCs) and trigger tumor antigen specific response mediated by CD8 + T cell, which contributed to inhibit the propagation of original and distal tumor cells. Like intratumoral injection, intravenous injection with WNV-poly(A) also markedly delays Huh7 hepatic carcinoma (HCC) transplanted tumor progression. Most importantly, in addition to an array of mouse xenograft tumor models, WNV-poly(A) also has a significant inhibitory effect on many different types of patient-derived tumor tissues and HCC patient-derived xenograft (PDX) tumor models. Our studies reveal that WNV-poly(A) is a potent and excellent oncolytic agent against many types of tumors and may have a role in metastatic and recurrent tumors.

Newcastle Disease Virus (NDV)-based vaccine candidate against SARS-CoV-2 Omicron by intranasal immunization.

Despite global vaccination efforts, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to evolve and spread globally. Currently, the development of affordable vaccine against Omicron variant of concern (VOC) is necessary. Here, we assessed the safety and immunogenicity of a SARS-CoV-2 vaccine consisting of a live Newcastle disease virus vector expressing the spike (S) protein of Omicron BA.1 administrated intranasally (IN) or intramuscularly (IM) in Golden Syrian hamster model. Immunogenicity studies showed that the prime-boost regimen elicited high antibody titers and the modified S antigen (Sm-F) could induce robust antibody response in low dosage immunization through IN route. Sera of the immunized hamsters provided effective cross-neutralizing activity against different Omicron variants, the prototype and delta strains of SARS-CoV-2. Moreover, the vaccine could provide complete immunoprotection in hamsters against the Omicron BA.1 challenge by either intranasal or intramuscular immunization. Overall, our study provides an alternative nasal vaccine against the SARS-CoV-2 Omicron variants.