Molecular Mechanism Analysis of the Effect of Hederagenin Combined with L-OHP on Chemosensitivity of AGS/L-OHP Based on Network Pharmacology.

AIMS AND OBJECTIVES: This study aimed to evaluate the pharmacological mechanism of Hederagenin (HD) combined with oxaliplatin (L-OHP) in treating gastric cancer (GC) through network pharmacology combined with experimental verification. MATERIAL AND METHODS: Network pharmacology methods were used to screen potential targets for HD, L-OHP, and GC-related targets from public databases, and the intersection of the three gene sets was taken. Cross genes were analyzed through protein-protein interaction (PPI) networks to predict core targets, and related pathways were predicted through GO and KEGG enrichment analysis. The experimental results were verified by the in vitro experiments. HD was applied on AGS/L-OHP cells, and then cellular chemosensitivity and the expressions of P-gp, Survivin, Bcl-2, p-Akt, and p-PI3K genes were detected. Wound assay and Transwell Chamber assay were employed to detect the effect of HD on AGS/L-OHP cells. Nude mice xenograft models transfected using AGS/L-OHP cells were also treated with HD in order to verify the results. The size and weight of the tumor, as well as the expressions of P-gp, Survivin, Bcl-2, p- Akt and p-PI3K genes, were also measured. RESULTS: KEGG analysis showed that the anti-gastric cancer effect of HD was mediated mainly by PI3K-Akt signaling pathways. The PI3K-Akt signaling pathway containing more enriched genes may play a greater role in anti-gastric cancer. It was observed that for AGS/L-OHP cells jointly treated with HD and L-OHP, their activity, migration and invasion were significantly lower than those treated only using HD or L-OHP group. Moreover, expressions of p-Akt, p- PI3K, Bcl-2, P-gp, and Survivin for the HD+L-OHP group decreased significantly. Results of the in vivo experiments showed that the sizes and weights of tumors in the HD+L-OHP group were the lowest compared to the HD group and L-OHP group. CONCLUSION: Our findings suggest that HD may reduce the resistance of AGS/L-OHP cells to LOHP by regulating the PI3K/Akt signaling pathway.

Urokinase plasminogen activator surface receptor restricts HIV-1 replication by blocking virion release from the cell membrane.

The urokinase-type plasminogen activator (uPA) system consists of the proteinase uPA, its receptor (PLAUR/uPAR). Under physiological conditions, uPA and PLAUR are predominantly expressed by blood cells, including neutrophils, monocytes, and macrophages, and play important roles in cell activation, adhesion, migration, and extravasation. Here, we report that PLAUR, which is highly expressed in macrophages and dendritic cells (DCs) but hardly expressed in CD4+ T cells, inhibits the release of HIV-1 progeny virions from the cell membrane. Silencing PLAUR markedly enhanced the transmission of HIV-1 in macrophages and DCs. We further demonstrated that PLAUR is localized at the cell membrane to block the release of HIV-1 virions. Interestingly, we found that uPA compromises the PLAUR-mediated inhibition to slightly enhance HIV-1 production in primary macrophages and DCs. In the absence of PLAUR, this enhanced effect induced by uPA is abrogated. In conclusion, PLAUR is a new anti-HIV-1 protein produced in both macrophages and DCs where it inhibits HIV-1 transmission. This discovery may provide a novel therapeutic target for combating HIV.

Membrane Protein OTOF Is a Type I Interferon-Induced Entry Inhibitor of HIV-1 in Macrophages.

In humans, HIV-1 infection induces innate immune responses mediated mainly by type I interferon (IFN). Type I IFN restricts HIV-1 replication by upregulating the expression of IFN-stimulated genes with diverse anti-HIV properties. In this study, we report that the cell membrane protein otoferlin (OTOF) acts as a type I IFN-induced effector, inhibiting HIV-1 entry in myeloid lineage macrophages and dendritic cells (DCs). OTOF is significantly induced by type I IFN in macrophages and DCs but not in CD4+ T lymphocytes. Silencing OTOF abrogates the IFN-mediated suppression of HIV-1 infection in macrophages and DCs. Moreover, OTOF overexpression exhibits anti-HIV activity in macrophages and CD4+ T cells. Further evidence reveals that OTOF inhibits HIV-1 entry into target cells at the cell membrane. Collectively, OTOF is a downstream molecule induced by type I IFN to inhibit HIV-1 entry in macrophages; it is a new potential agent for the treatment of HIV infection. IMPORTANCE In patients with HIV-1 infection, the virus is recognized by innate immune sensors that trigger the production of type I interferons (IFNs), which are well-known cytokines that exert broad antiviral effects by inducing the expression of antiviral genes. By comparing the gene expression profiles of untreated patients and healthy donors, we systematically identified OTOF as a new antiviral gene induced by IFN-α in primary macrophages and dendritic cells (DCs). Additionally, silencing OTOF alleviates IFN-α-induced resistance to HIV-1 infection in both myeloid cell lineage macrophages and DCs. In contrast, OTOF overexpression potently restricts HIV-1 transmission in macrophages. We further explored the molecular mechanism through which OTOF inhibits the HIV-1 virion across the cell membrane. Overall, OTOF is a newly identified type I IFN-induced antiviral factor that inhibits the transmembrane activity of HIV-1 in myeloid cells.

CTNNBL1 restricts HIV-1 replication by suppressing viral DNA integration into the cell genome.

Retroviral integration is mediated by a unique enzymatic process shared by all retroviruses and retrotransposons. During integration, double-stranded linear viral DNA is inserted into the host genome in a process catalyzed by viral-encoded integrase (IN). However, host cell defenses against HIV-1 integration are not clear. This study identifies ß-catenin-like protein 1 (CTNNBL1) as a potent inhibitor of HIV-1 integration via association with viral-encoded integrase (IN) and its cofactor, lens epithelium-derived growth factor/p75. CTNNBL1 overexpression blocks HIV-1 integration and inhibits viral replication, whereas CTNNBL1 depletion significantly upregulates HIV-1 integration into the genome of various target cells. Further, CTNNBL1 expression is downregulated in CD4+ T cells by activation, and CTNNBL1 depletion also facilitates HIV-1 integration in resting CD4+ T cells. Thus, host cells may employ CTNNBL1 to inhibit HIV-1 integration into the genome. This finding suggests a strategy for the treatment of HIV infections.

Neuropilin-1, a myeloid cell-specific protein, is an inhibitor of HIV-1 infectivity.

Myeloid lineage cells such as macrophages and dendritic cells (DCs), targeted by HIV-1, are important vehicles for virus dissemination through the body as well as viral reservoirs. Compared to activated lymphocytes, myeloid cells are collectively more resistant to HIV-1 infection. Here we report that NRP-1, encoding transmembrane protein neuropilin-1, is highly expressed in macrophages and DCs but not CD4+ T cells, serving as an anti-HIV factor to inhibit the infectivity of HIV-1 progeny virions. Silencing NRP-1 enhanced the transmission of HIV-1 in macrophages and DCs significantly and increased the infectivity of the virions produced by these cells. We further demonstrated that NRP-1 was packaged into the progeny virions to inhibit their ability to attach to target cells, thus reducing the infectivity of the virions. These data indicate that NRP-1 is a newly identified antiviral protein highly produced in both macrophages and DCs that inhibit HIV-1 infectivity; thus, NRP-1 may be a novel therapeutic strategy for the treatment of HIV-1 infection.

Differently PEGylated Polymer Nanoparticles for Pancreatic Cancer Delivery: Using a Novel Near-Infrared Emissive and Biodegradable Polymer as the Fluorescence Tracer.

In this study, a chemically synthetic polymer, benzo[1,2-b:4,5-b']difuran(BDF)-based donor-acceptor copolymer PBDFDTBO, was individually coated by amphiphilic poly(ethylene oxide)-block-poly(ε-caprolactone) (PEO-PCL) and 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-methoxy(polyethylene glycol) (DSPE-PEG or PEG-DSPE), to form stably fluorescent nanoparticles in the near-infrared (NIR) window. The physicochemical properties of the synthesized nanoparticles were characterized and compared, including their size, surface charge, and morphology. In addition, in vitro studies were also performed using two pancreatic cancer cell lines, assessing the cell viability of the PBDFDTBO-included PEGylated nanoparticles formulations. Moreover, in vivo studies were also conducted, using subcutaneous murine cancer models to investigate the polymeric nanoparticles' circulation time, tumor accumulation, and preferred organ biodistribution. The overall results demonstrated that even with the same PEGylated surface, the hydrophobic composition anchored on the encapsulated PBDFDTBO core strongly affected the biodistribution and tumor accumulation of the nanoparticles, to a degree possibly determined by the hydrophobic interactions between the hydrophobic segment of amphiphilic polymers (DSPE or PCL moiety) and the enwrapped PBDFDTBO. Both PEGylated nanoparticles were compared to obtain an optimized coating strategy for a desired biological feature in pancreatic cancer delivery.

Vpr counteracts the restriction of LAPTM5 to promote HIV-1 infection in macrophages.

The HIV-1 accessory proteins Vif, Vpu, and Nef can promote infection by overcoming the inhibitory effects of the host cell restriction factors APOBEC3G, Tetherin, and SERINC5, respectively. However, how the HIV-1 accessory protein Vpr enhances infection in macrophages but not in CD4+ T cells remains elusive. Here, we report that Vpr counteracts lysosomal-associated transmembrane protein 5 (LAPTM5), a potent inhibitor of HIV-1 particle infectivity, to enhance HIV-1 infection in macrophages. LAPTM5 transports HIV-1 envelope glycoproteins to lysosomes for degradation, thereby inhibiting virion infectivity. Vpr counteracts the restrictive effects of LAPTM5 by triggering its degradation via DCAF1. In the absence of Vpr, the silencing of LAPTM5 precisely phenocopied the effect of Vpr on HIV-1 infection. In contrast, Vpr did not enhance HIV-1 infection in the absence of LAPTM5. Moreover, LAPTM5 was highly expressed in macrophages but not in CD4+ T lymphocytes. Re-expressing LAPTM5 reconstituted the Vpr-dependent promotion of HIV-1 infection in primary CD4+ T cells, as observed in macrophages. Herein, we demonstrate the molecular mechanism used by Vpr to overcome LAPTM5 restriction in macrophages, providing a potential strategy for anti-HIV/AIDS therapeutics.

Biosynthetic Polymalic Acid as a Delivery Nanoplatform for Translational Cancer Medicine.

Poly(ß-L-malic acid) (PMLA) is a natural polyester produced by numerous microorganisms. Regarding its biosynthetic machinery, a nonribosomal peptide synthetase (NRPS) is proposed to direct polymerization of L-malic acid in vivo. Chemically versatile and biologically compatible, PMLA can be used as an ideal carrier for several molecules, including nucleotides, proteins, chemotherapeutic drugs, and imaging agents, and can deliver multimodal theranostics through biological barriers such as the blood-brain barrier. We focus on PMLA biosynthesis in microorganisms, summarize the physicochemical and physiochemical characteristics of PMLA as a naturally derived polymeric delivery platform at nanoscale, and highlight the attachment of functional groups to enhance cancer detection and treatment.

Development and application of a comparative risk assessment method for ranking chemical hazards in food.

The aim of this study was to develop a comparative risk assessment method to prioritise the public health risks posed by chemical hazards in food. Through a literature review, and in light of expert opinions, a bottom-up, semi-quantitative scoring method was applied to screen the ranking metrics and assign a score. In addition, a metrics system and a ranking model were constructed. The fuzzy comprehensive analysis model was used to assess typical chemical hazards in a specific food, as well as to rank risks in many foods. Data were collected from the National Food Surveillance System in China, the Food Consumption of Chinese Residents Database, government reports, public websites and databases of authoritative organisations. The comparative risk assessment method was applied to case studies on ranking chemical hazards in different kinds of food. According to application testing, the method truly reflects the overall risk and ranking of chemical hazards in food.

Exosomal PD-L1: New Insights Into Tumor Immune Escape Mechanisms and Therapeutic Strategies.

As a classical immune checkpoint molecule, PD-L1 on the surface of tumor cells plays a pivotal role in tumor immunosuppression, primarily by inhibiting the antitumor activities of T cells by binding to its receptor PD-1. PD-1/PD-L1 inhibitors have demonstrated unprecedented promise in treating various human cancers with impressive efficacy. However, a significant portion of cancer patients remains less responsive. Therefore, a better understanding of PD-L1-mediated immune escape is imperative. PD-L1 can be expressed on the surface of tumor cells, but it is also found to exist in extracellular forms, such as on exosomes. Recent studies have revealed the importance of exosomal PD-L1 (ExoPD-L1). As an alternative to membrane-bound PD-L1, ExoPD-L1 produced by tumor cells also plays an important regulatory role in the antitumor immune response. We review the recent remarkable findings on the biological functions of ExoPD-L1, including the inhibition of lymphocyte activities, migration to PD-L1-negative tumor cells and immune cells, induction of both local and systemic immunosuppression, and promotion of tumor growth. We also discuss the potential implications of ExoPD-L1 as a predictor for disease progression and treatment response, sensitive methods for detection of circulating ExoPD-L1, and the novel therapeutic strategies combining the inhibition of exosome biogenesis with PD-L1 blockade in the clinic.

Membrane metalloprotease TRABD2A restricts HIV-1 progeny production in resting CD4+ T cells by degrading viral Gag polyprotein.

Resting CD4+ T cells are highly resistant to the production of human immunodeficiency virus type 1 (HIV-1). However, the mechanism by which resting CD4+ T cells restrict such production in the late viral replication phase of infection has remained unclear. In this study, we found that the cell membrane metalloprotease TRAB domain-containing protein 2A (TRABD2A) inhibited this production in resting CD4+ T cells by degrading the virion structural precursor polyprotein Gag at the plasma membrane. Depletion or inhibition of metalloprotease activity by TRABD2A profoundly enhanced HIV-1 production in resting CD4+ T cells. TRABD2A expression was much higher in resting CD4+ T cells than in activated CD4+ T cells and was considerably reduced by T cell activation. Moreover, reexpressing TRABD2A reinforced the resistance of activated CD4+ T cells to the production of HIV-1 progeny. Collectively, these results elucidate the molecular mechanism employed by resting CD4+ T cells to potently restrict the assembly and production of HIV-1 progeny.

Immunosuppression of Human Adipose-Derived Stem Cells on T Cell Subsets via the Reduction of NF-kappaB Activation Mediated by PD-L1/PD-1 and Gal-9/TIM-3 Pathways.

Adipose-derived stem cells (ADSCs) are a type of multipotent mesenchymal stem cells with immunosuppressive capacities. However, the underlying mechanisms involved in the inhibitory effects of ADSCs on T cells are not completely elucidated. In this study, human peripheral blood mononuclear cells (PBMCs) stimulated with anti-CD3/CD28 antibody-coated beads were cultured with or without allogeneic ADSCs (ADSC-to-PBMC ratio, 1:5). Surface marker levels, violet-labeled cell proliferation, apoptosis, interferon-gamma (IFN-gamma) production, and nuclear factor-kappaB (NF-kappaB) phosphorylation of CD4+ and CD8+ T cells were detected using flow cytometry. It was observed that ADSCs significantly suppressed the proliferation and IFN-gamma production but enhanced apoptosis of both CD4+ and CD8+ T cells in T cell receptor (TCR)-stimulated PBMCs. The expressions of programmed death-ligand 1 (PD-L1) and galectin 9 (Gal-9) on ADSCs were significantly upregulated and induced during coculture with PBMCs. TCR-stimulated CD4+ and CD8+ T cells cultured with ADSCs had higher expression levels of programmed death-1 (PD-1) and T cell immunoglobulin and mucin-containing protein-3 (TIM-3) than those in cells cultured without ADSCs. Moreover, the suppressive effects of ADSCs on T cells in terms of proliferation and IFN-gamma production were significantly reversed in the presence of anti-PD-L1 and anti-Gal-9 antibodies. Importantly, the phosphorylation of NF-kappaB in CD4+ and CD8+ T cells cocultured with ADSCs was significantly inhibited, and this inhibition was significantly attenuated via the PD-L1 and Gal-9 blockades. In conclusion, human ADSCs perform immunoregulatory functions partially through the inhibition of NF-kappaB activation in T cells via the PD-L1/PD-1 and Gal-9/TIM-3 pathways, which provide new insights into the mechanism of human ADSC-mediated immunomodulation.

Alteration of CCR6+CD95+CD4+ naïve T cells in HIV-1 infected patients: Implication for clinical practice.

The profound deficiency of Th17 cells contributes to HIV disease progression. The mechanisms of their perturbation remain unclear. Recently, CCR6+CD95+CD4+ naïve T cells (CCR6+CD95+CD4+ TNA), identified as pre-committed Th17 precursors, were recognized as a subpopulation of CD4+ T cells with stem cell properties. Following phenotypical identification, we evaluated their level in patients during chronic HIV infection and following antiretroviral therapy (ART) using flow cytometry. The levels of CCR6+CD95+CD4+ TNA were decreased during chronic HIV infection and correlated with CD4+ T cell counts. Immunological responders harbored higher frequency of CCR6+CD95+CD4+ TNA, which was associated with CD4/CD8 T cell ratio. Immunological non-responders with lower frequency of CCR6+CD95+CD4+ TNA failed to exhibit a correlation between CCR6+CD95+CD4+ TNA and CCR6+CD95+CD4+ TCM, and displayed elevated ratio of CCR6+CD95+CD4+ TCM/TNA. The number of CCR6+CD95+CD4+ TNA was increased following early ART. These findings shed light on the importance of targeting pre-committed Th17 precursors that enhance immune reconstitution.

AID recruits the RNA exosome to degrade HIV-1 nascent transcripts through interaction with the Tat-P-TEFb-TAR RNP complex.

Activation-induced cytidine deaminase (AID), a member of the APOBEC family that induces antibody diversification, has been shown to inhibit the replication of hepatitis B virus, Kaposi's sarcoma-associated herpesvirus, and retro-transposons. However, whether AID can inhibit human immunodeficiency virus 1 (HIV-1) replication remains unclear. Here, we report that AID impairs the synthesis of HIV-1 components by interacting with the complex of Tat. This interaction recruits the RNA exosome to degrade the nascent HIV-1 transcript. AID also targets the HIV-1-integrated genome via the Tat-P-TEFb-TAR complex. Thus, we propose a novel function for AID as an adaptor protein that represses viral transcription. Our findings provide insights into developing anti-HIV therapeutics and understanding how host cells restrict integrated virus replication.

The Per-1 Short Isoform Inhibits de novo HIV-1 Transcription in Resting CD4+ T-cells.

BACKGROUND: Understanding of the restriction of HIV-1 transcription in resting CD4+ Tcells is critical to find a cure for AIDS. Although many negative factors causing HIV-1 transcription blockage in resting CD4+ T-cells have been found, there are still unknown mechanisms to explore. OBJECTIVE: To explore the mechanism for the suppression of de novo HIV-1 transcription in resting CD4+ T-cells. METHODS: In this study, a short isoform of Per-1 expression plasmid was transfected into 293T cells with or without Tat's presence to identify Per-1 as a negative regulator for HIV-1 transcription. Silencing of Per-1 was conducted in resting CD4+ T-cells or monocyte-derived macrophages (MDMs) to evaluate the antiviral activity of Per-1. Additionally, we analyzed the correlation between Per-1 expression and viral loads in vivo, and silenced Per-1 by siRNA technology to investigate the potential anti-HIV-1 roles of Per-1 in vivo in untreated HIV-1-infected individuals. RESULTS: We found that short isoform Per-1 can restrict HIV-1 replication and Tat ameliorates this inhibitory effect. Silencing of Per-1 could upregulate HIV-1 transcription both in resting CD4+ Tcells and MDMs. Moreover, Per-1 expression is inversely correlated with viral loads in Rapid progressors (RPs) in vivo. CONCLUSION: These data together suggest that Per-1 is a novel negative regulator of HIV-1 transcription. This restrictive activity of Per-1 to HIV-1 replication may contribute to HIV-1 latency in resting CD4+ T-cells.

TGF-ß triggers HBV cccDNA degradation through AID-dependent deamination.

The covalently closed circular DNA (cccDNA) of hepatitis B virus (HBV) is a viral center molecule for HBV infection and persistence. However, the cellular restriction factors of HBV cccDNA are not well understood. Here, we show that TGF-ß can induce nuclear viral cccDNA degradation and hypermutation via activation-induced cytidine deaminase (AID) deamination activity in hepatocytes. This suppression by TGF-ß is abrogated when AID or the activity of uracil-DNA glycosylase (UNG) is absent, which indicates that AID deamination and the UNG-mediated excision of uracil act in concert to degrade viral cccDNA. Moreover, the HBV core protein promotes the interaction between AID and viral cccDNA. Overall, our results indicate a novel molecular mechanism that allows cytokine TGF-ß to restrict viral nuclear cccDNA in innate immunity, thereby suggesting a novel method for potentially eliminating cccDNA.

TGF-ß suppression of HBV RNA through AID-dependent recruitment of an RNA exosome complex.

Transforming growth factor (TGF)-ß inhibits hepatitis B virus (HBV) replication although the intracellular effectors involved are not determined. Here, we report that reduction of HBV transcripts by TGF-ß is dependent on AID expression, which significantly decreases both HBV transcripts and viral DNA, resulting in inhibition of viral replication. Immunoprecipitation reveals that AID physically associates with viral P protein that binds to specific virus RNA sequence called epsilon. AID also binds to an RNA degradation complex (RNA exosome proteins), indicating that AID, RNA exosome, and P protein form an RNP complex. Suppression of HBV transcripts by TGF-ß was abrogated by depletion of either AID or RNA exosome components, suggesting that AID and the RNA exosome involve in TGF-ß mediated suppression of HBV RNA. Moreover, AID-mediated HBV reduction does not occur when P protein is disrupted or when viral transcription is inhibited. These results suggest that induced expression of AID by TGF-ß causes recruitment of the RNA exosome to viral RNP complex and the RNA exosome degrades HBV RNA in a transcription-coupled manner.