New role of sertraline against Toxoplasma gondii-induced depression-like behaviours in mice.

Toxoplasma gondii (T. gondii) is a neurotropic protozoan parasite, which can cause mental and behavioural disorders. The present study aimed to elucidate the effects and underlying molecular mechanisms of sertraline (SERT) on T. gondii-induced depression-like behaviours. In the present study, a mouse model and a microglial cell line (BV2 cells) model were established by infecting with the T. gondii RH strain. In in vivo and in vitro experiments, the underlying molecular mechanisms of SERT in inhibiting depression-like behaviours and cellular perturbations caused by T. gondii infection were investigated in the mouse brain and BV2 cells. The administration of SERT significantly ameliorated depression-like behaviours in T. gondii-infected mice. Furthermore, SERT inhibited T. gondii proliferation. Treatment with SERT significantly inhibited the activation of microglia and decreased levels of pro-inflammatory cytokines such as tumour necrosis factor-alpha, and interferon-gamma, by down-regulating tumour necrosis factor receptor 1/nuclear factor-kappa B signalling pathway, thereby ameliorating the depression-like behaviours induced by T. gondii infection. Our study provides insight into the underlying molecular mechanisms of the newly discovered role of SERT against T. gondii-induced depression-like behaviours.

Usnic acid suppresses cervical cancer cell proliferation by inhibiting PD-L1 expression and enhancing T-lymphocyte tumor-killing activity.

The programmed cell death 1 (PD-1)/programmed death ligand 1 (PD-L1) pathway is abnormally expressed in cervical cancer cells. Moreover, PD-1/PD-L1 blockade reduces the apoptosis and exhaustion of T cells and inhibits the development of malignant tumors. Usnic acid is a dibenzofuran compound originating from Usnea diffracta Vain and has anti-inflammatory, antifungal, and anticancer activities. However, the molecular mechanism of its antitumor effects has not been fully elucidated. In this work, we first observed that usnic acid decreased the expression of PD-L1 in HeLa cells and enhanced the cytotoxicity of co-cultured T cells toward tumor cells. Usnic acid inhibited PD-L1 protein synthesis by reducing STAT3 and RAS pathways cooperatively. It was subsequently shown that usnic acid induced MiT/TFE nuclear translocation through the suppression of mTOR signaling pathways, and promoted the biogenesis of lysosomes and the translocation of PD-L1 to the lysosomes for proteolysis. Furthermore, usnic acid inhibited cell proliferation, angiogenesis, migration, and invasion, respectively, by downregulating PD-L1, thereby inhibiting tumor growth. Taken together, our results show that usnic acid is an effective inhibitor of PD-L1 and our study provide novel insights into the mechanism of its anticancer targeted therapy.

Panaxadiol inhibits programmed cell death-ligand 1 expression and tumour proliferation via hypoxia-inducible factor (HIF)-1α and STAT3 in human colon cancer cells.

Panaxadiol is a triterpenoid sapogenin monomeric compound found in the roots of Panax ginseng and has a variety of biological activities such as neuroprotective and anti-tumour functions. However, the mechanisms how panaxadiol exerts the anticancer effects remain unknown. The current study aimed to investigate the potential activity of panaxadiol on programmed cell death-ligand 1 (PD-L1) expression and tumour proliferation in human colon cancer cells and to identify the underlying mechanism. Results showed that panaxadiol showed little cytotoxicity as assessed by a cytotoxicity assay and significantly inhibited PD-L1 expression at the protein and mRNA level in a dose-dependent manner. Furthermore, panaxadiol supressed the hypoxia-induced synthesis of hypoxia-inducible factor (HIF)-1α via the phosphoinositide 3-kinase (PI3K) and mitogen-activated protein kinase (MAPK) pathways without affecting HIF-1α degradation. Simultaneously, panaxadiol inhibited STAT3 activation through the JAK1, JAK2, and Src pathways. Moreover, pre-treatment with panaxadiol enhanced the activity of cytotoxic T lymphocytes (CTL) and regained their capacity of tumour cell killing in a T cell and tumour cell co-culture system. Immunoprecipitation showed that panaxadiol inhibited PD-L1 expression by blocking the interaction between HIF-1α and STAT3. The inhibitory effect of panaxadiol on tumour proliferation was further demonstrated by colony formation and EdU labelling assays. The anti-proliferative effect of panaxadiol was also proved by a xenograft assay in vivo. Taken together, the current work highlights the anti-tumour effect of panaxadiol, providing insights into development of cancer therapeutic through PD-L1 inhibition.

Convallatoxin protects against dextran sulfate sodium-induced experimental colitis in mice by inhibiting NF-κB signaling through activation of PPARγ.

Convallatoxin (CNT) is a cardiac glycoside isolated from Adonis amurensis Regel et Radde and has both anti-inflammatory and anti-proliferative properties. In the present study, the anti-inflammatory mechanisms of action of CNT was investigated in vitro and in vivo. Stimulation of mouse macrophages with lipopolysaccharide induced secretion of proinflammatory cytokines via suppression of peroxisome proliferator-activated receptor gamma (PPARγ) and activation of nuclear factor-κB (NF-κB), two transcription factors implicated in many inflammatory diseases. Notably, the effects of lipopolysaccharide were reversed by concomitant treatment of macrophages with CNT. Knockdown of PPARγ by siRNA inhibited the effect of convallatoxin on NF-κB activation. Because these transcription factors play a role in the development of ulcerative colitis in humans, the mice with experimental colitis induced by dextran sodium sulfate (DSS) was employed. Indeed, concomitant treatment with CNT ameliorated DSS-induced colitis symptoms, tissue damage, inflammatory cell infiltration, and proinflammatory cytokine production in the colon, and also reversed the activation of NF-κB and suppression of PPARγ. Collectively, these data indicate that CNT ameliorates colitic inflammation via activation of PPARγ and suppression of NF-κB, and suggest that CNT may be a promising treatment for inflammatory bowel disease (IBD).

Cellular immunopathogenesis in primary Toxoplasma gondii infection during pregnancy.

Congenital toxoplasmosis is caused by the vertical transmission of infection from mother to foetus through the placenta when a pregnant woman is infected with Toxoplasma gondii (T. gondii). Congenital infection can have serious consequences, such as intrauterine abortion, foetal death and severe neurological, ocular or other organ damage in the foetus. In this review, we focus on recent publications investigating vertical transmission of T. gondii infection, cellular immunopathogenesis and protective immunity in primary toxoplasmosis during pregnancy.

Chelidonine inhibits TNF-α-induced inflammation by suppressing the NF-κB pathways in HCT116 cells.

Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) is a complex that regulates several hundreds of genes, including those involved in immunity and inflammation, survival, proliferation, and the negative feedback of NF-κB signaling. Chelidonine, a major bioactive, isoquinoline alkaloid ingredient in Chelidonium majus, exhibits antiinflammatory pharmacological properties. However, its antiinflammatory molecular mechanisms remain unclear. In this work, we explored the effect of chelidonine on TNF-induced NF-κB activation in HCT116 cells. We found chelidonine inhibited the phosphorylation and degradation of the inhibitor of NF-κB alpha and nuclear translocation of RELA. Furthermore, by inhibiting the activation of NF-κB, chelidonine downregulated target genes involved in inflammation, proliferation, and apoptosis. Chelidonine also inhibited mitogen-activated protein kinase pathway activation by blocking c-Jun N-terminal kinase and p38 phosphorylation. These results suggest that chelidonine may be a potential therapeutic agent against inflammatory diseases in which inhibition of NF-κB activity plays an important role.

Amorfrutin A inhibits TNF-α induced JAK/STAT signaling, cell survival and proliferation of human cancer cells.

CONTEXT: Amorfrutin A is a natural product isolated from the fruits of Amorpha fruticosa L. and has been shown to exhibit multiple bioeffector functions. In the present study, we investigated whether amorfrutin A exerts anticancer effects by inhibiting STAT3 activation in cervical cancer cells. OBJECTIVE: To investigate the effectiveness of amorfrutin A as a treatment of cancer, and determine the underlying pharmacological mechanism of action. MATERIALS AND METHODS: HeLa, SK-Hep1, MDA-MB-231 and HCT116 cells were used in this study. Major assays were luciferase reporter assay, MTT, Western blot analysis, immunofluorescence assay, reverse transcription-PCR (RT-PCR), flow cytometric analysis, EdU labeling and immunofluorescence, xenografted assay. RESULTS: Amorfrutin A significantly inhibited tumor necrosis factor-α (TNF-α)-induced phosphorylation and nuclear translocation of STAT3 in human cervical carcinoma cells. Amorfrutin A also inhibited activation of the upstream kinases Janus-activated kinase 1 (JAK1), JAK2 and Src signaling pathways. Furthermore, amorfrutin A increased the expression of p53, p21, p27, induced cell cycle arrest in the G1 phase as well as decreased levels of various oncogene protein products. In vivo studies further confirmed the inhibitory effect of amorfrutin A on the expression of STAT3 proteins, leading to a decrease growth of HeLa cells in a xenograft tumor model. DISCUSSION AND CONCLUSIONS: The results indicated that amorfrutin A is a potent inhibitor of STAT3 and provide new perspectives into the mechanism of its anticancer activity.

Artemisinin inhibits inflammatory response via regulating NF-κB and MAPK signaling pathways.

Artemisinin, isolated from the Chinese plant Artemisia annua, has been used for many years to treat different forms of malarial parasites. In this study, we explored the anti-inflammatory activity of artemisinin and the underlying mechanism of this action. We demonstrated that the anti-inflammatory effects of artemisinin in TPA-induced skin inflammation in mice. Then the artemisinin significantly inhibited the expression of NF-κB reporter gene induced by TNF-α in a dose-dependent manner. Artemisinin also inhibited TNF-α induced phosphorylation and degradation of IκBα, p65 nuclear translocation. Artemisinin also has an impact on upstream signaling of IKK through the inhibition of expression of adaptor proteins, TNF receptor-associated factor 2 (TRAF2) and receptor interacting protein 1 (RIP1). Furthermore, pretreatment of cells with artemisinin prevented the TNF-α-induced expression of NF-κB target genes, such as anti-apoptosis (c-IAP1, Bcl-2, and FLIP), proliferation (COX-2, cyclinD1), invasion (MMP-9), angiogenesis (VEGF), and major inflammatory cytokines (TNF-α, iNOS, and MCP1). We also proved that artemisinin potentiated TNF-α-induced apoptosis. Moreover, artemisinin significantly impaired the ROS production and phosphorylation of p38 and ERK, but did not affect the phosphorylation of JNK. Taken together, artemisinin may be a potentially useful therapeutic agent for inflammatory-related diseases.

Dihydromyricetin suppresses TNF-α-induced NF-κB activation and target gene expression.

Nuclear factor-kappa B (NF-κB) has been reported to play a pivotal role in many physiological processes including inflammation, apoptosis, and angiogenesis. We discovered a potent natural NF-κB inhibitor, dihydromyricetin, from the traditional herb Ampelopsis grossedentata, which has a long history of use in food and medicine. In this study, we demonstrated the effect of dihydromyricetin on NF-κB activation in TNF-α-induced HeLa cells. Dihydromyricetin was found to markedly inhibit the phosphorylation and degradation of the inhibitor of NF-κB alpha (IκBα), and subsequent nuclear translocation of p65. Dihydromyricetin also has an impact on upstream signaling of IKK through the inhibition of expression of adaptor proteins, TNF receptor-associated factor 2 (TRAF2), and receptor-interacting protein 1 (RIP1). Furthermore, the current results reveal that dihydromyricetin led to the downregulation of target genes involved in inflammation, proliferation, as well as potentiation of TNF-α-induced apoptosis through suppressing the activation of NF-κB. In conclusion, our data indicate that dihydromyricetin may be a potentially useful therapeutic agent for inflammatory diseases.

Protective effect of arctiin against Toxoplasma gondii HSP70-induced allergic acute liver injury by disrupting the TLR4-mediated activation of cytosolic phospholipase A2 and platelet-activating factor.

Toxoplasma gondii (T. gondii)-derived heat shock protein 70 (T.g.HSP70) is a toxic protein that downregulates host defense responses against T. gondii infection. T.g.HSP70 was proven to induce fatal anaphylaxis in T. gondii infected mice through cytosolic phospholipase A2 (cPLA2) activated-platelet-activating factor (PAF) production via Toll-like receptor 4 (TLR4)-mediated signaling. In this study, we investigated the effect of arctiin (ARC; a major lignan compound of Fructus arctii) on allergic liver injury using T.g.HSP70-stimulated murine liver cell line (NCTC 1469) and a mouse model of T. gondii infection. Localized surface plasmon resonance, ELISA, western blotting, co-immunoprecipitation, and immunofluorescence were used to investigate the underlying mechanisms of action of ARC on T. gondii-induced allergic acute liver injury. The results showed that ARC suppressed the T.g.HSP70-induced allergic liver injury in a dose-dependent manner. ARC could directly bind to T.g.HSP70 or TLR4, interfering with the interaction between these two factors, and inhibiting activation of the TLR4/mitogen-activated protein kinase/nuclear factor-kappa B signaling, thereby inhibiting the overproduction of cPLA2, PAF, and interferon-γ. This result suggested that ARC ameliorates T.g.HSP70-induced allergic acute liver injury by disrupting the TLR4-mediated activation of inflammatory mediators, providing a theoretical basis for ARC therapy to improve T.g.HSP70-induced allergic liver injury.

A novel mechanism of resveratrol alleviates Toxoplasma gondii infection-induced pulmonary inflammation via inhibiting inflammasome activation.

BACKGROUND: Infection by Toxoplasma gondii can lead to severe pneumonia, with current treatments being highly inadequate. The NLRP3 inflammasome is one member of the NOD-like receptor family with a pyrin domain, which is crucial in the innate immune defense against T. gondii. Research has shown that resveratrol (RSV) prevents lung damage caused by this infection by inhibiting the T. gondii-derived heat shock protein 70/TLR4/NF-κB pathway, thus reducing the macrophage-driven inflammatory response. However, it should be mentioned that the participation of NLRP3 inflammasome in the immune response to the lung injuries caused by T. gondii infections is not entirely clear. PURPOSE: This study aims to clarify how RSV ameliorates lung damage triggered by Toxoplasma gondii infection, with a particular focus on the pathway involving TLR4, NF-κB, and the NLRP3 inflammasome. METHODS: Both in vitro and in vivo models of infection were developed by employing the RH strain of T. gondii in BALB/c mice and RAW 264.7 macrophage cell lines. The action mechanism of RSV was explored using techniques such as molecular docking, surface plasmon resonance, ELISA, Western blot, co-immunoprecipitation, and immunofluorescence staining. RESULTS: Findings indicate that the suppression of TLR4 or NF-κB impacts the levels of proteins associated with the NLRP3 inflammasome pathway. Additionally, a significant affinity for binding between RSV and NLRP3 was observed. Treatment with RSV led to a marked reduction in the activation and formation of the NLRP3 inflammasome within lung tissues and RAW 264.7 cells, alongside a decrease in IL-1ß concentrations in the bronchoalveolar lavage fluid. These outcomes align with those seen when using the NLRP3 inhibitor CY-09. Moreover, the application of CY-09 prior to RSV negated the latter's anti-inflammatory properties. CONCLUSION: Considering insights from previous research alongside the outcomes of the current investigation, it appears that the TLR4/NF-κB/NLRP3 signaling pathway emerges as a promising target for immunomodulation to alleviate lung injury from T. gondii infection. The evidence gathered in this study lays the groundwork for the continued exploration and potential future clinical deployment of RSV as a therapeutic agent with anti-Toxoplasma properties and the capability to modulate the inflammatory response.

Resveratrol inhibits Toxoplasma gondii-induced lung injury, inflammatory cascade and evidences of its mechanism of action.

BACKGROUND: Toxoplasma gondii is an opportunistic protozoan that can infect host to cause toxoplasmosis. We have previously reported that resveratrol (RSV) has protective effects against liver damage in T. gondii infected mice. However, the effect of RSV on lung injury caused by T. gondii infection and its mechanism of action remain unclear. PURPOSE: In this work, we studied the protective effects of RSV on lung injury caused by T. gondii infection and explored the underlying mechanism. METHODS: Molecular docking and localized surface plasmon resonance assay were used to detect the molecular interactions between RSV and target proteins. In vitro, the anti-T. gondii effects and potential anti-inflammatory mechanisms of RSV were investigated by quantitative competitive-PCR, RT-PCR, ELISA, Western blotting and immunofluorescence using RAW 264.7 cells infected with tachyzoites of T. gondii RH strain. In vivo, the effects of RSV on lung injury caused by T. gondii infection were assessed by observing pathological changes and the expression of inflammatory factors of lung. RESULTS: RSV inhibited T. gondii loads and T. gondii-derived heat shock protein 70 (T.g.HSP70) expression in RAW 264.7 cells and lung tissues. Moreover, RSV interacts with T.g.HSP70 and toll-like receptor 4 (TLR4), respectively, and interferes with the interaction between T.g.HSP70 and TLR4. It also inhibited the overproduction of inducible nitric oxide synthase, TNF-α and high mobility group protein 1 (HMGB1) by down-regulating TLR4/nuclear factor kappa B (NF-κB) signaling pathway, which is consistent with the effect of TLR4 inhibitor CLI-095. In vivo, RSV improved the pathological lung damage produced by T. gondii infection, as well as decreased the number of inflammatory cells in bronchoalveolar lavage fluid and the release of HMGB1 and TNF-α. CONCLUSION: These findings indicate that RSV can inhibit the proliferation of T. gondii and T.g.HSP70 expression both in vitro and in vivo. RSV can inhibit excessive inflammatory response by intervening T.g.HSP70 and HMGB1 mediated TLR4/NF-κB signaling pathway activation, thereby ameliorating lung injury caused by T. gondii infection. The present study provides new data that may be useful for the development of RSV as a new agent for the treatment of lung damage caused by T. gondii infection.

Coixol ameliorates Toxoplasma gondii infection-induced lung injury by interfering with T. gondii HSP70/TLR4/NF-κB signaling pathway.

Toxoplasma gondii (T. gondii) is an obligate intracellular protozoan parasite that causes pulmonary toxoplasmosis, although its pathogenesis is incompletely understood. There is no cure for toxoplasmosis. Coixol, a plant polyphenol extracted from coix seeds, has a variety of biological activities. However, the effects of coixol on T. gondii infection have not been clarified. In this study, we infected a mouse macrophage cell line (RAW 264.7) and BALB/c mice with the T. gondii RH strain to establish infection models in vitro and in vivo, respectively, to explore protective effects and potential mechanisms of coixol on lung injury caused by T. gondii infection. Anti-T. gondii effects and underlying anti-inflammatory mechanisms of coixol were investigated by real-time quantitative PCR, molecular docking, localized surface plasmon resonance, co-immunoprecipitation, enzyme-linked immunosorbent assay, western blotting, and immunofluorescence microscopy. The results show that coixol inhibits T. gondii loads and T. gondii-derived heat shock protein 70 (T.g.HSP70) expression. Moreover, coixol reduced inflammatory cell recruitment and infiltration, and ameliorated pathological lung injury induced by T. gondii infection. Coixol can directly bind T.g.HSP70 or Toll-like receptor 4 (TLR4) to disrupt their interaction. Coixol prevented overexpression of inducible nitric oxide synthase, tumor necrosis factor-α, and high mobility group box 1 by inhibiting activation of the TLR4/nuclear factor (NF)-κB signaling pathway, consistent with effects of the TLR4 inhibitor CLI-095. These results indicate that coixol improves T. gondii infection-induced lung injury by interfering with T.g.HSP70-mediated TLR4/NF-κB signaling. Altogether, these findings suggest that coixol is a promising effective lead compound for the treatment of toxoplasmosis.

Protective effect of ginsenoside Rh2 against Toxoplasma gondii infection-induced neuronal injury through binding TgCDPK1 and NLRP3 to inhibit microglial NLRP3 inflammasome signaling pathway.

BACKGROUND: Toxoplasma gondii (T. gondii) is a neurotropic obligate intracellular parasite that can activate microglial and promote neuronal apoptosis, leading to central nervous system diseases. The NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome signaling complex plays a key role in inducing neuroinflammation. Our previous studies have found that ginsenoside Rh2 (GRh2) inhibits T. gondii infection-induced microglial activation and neuroinflammation by downregulating the Toll-like receptor 4/nuclear factor-kappa B signaling pathway. However, whether GRh2 reduces T. gondii infection-induced neuronal injury through actions on microglial NLRP3 inflammasome signaling has not yet been clarified. METHODS: In this study, we employed T. gondii RH strain to establish in vitro and in vivo infection models in BV2 microglia cell line and BALB/c mice. Molecular docking, localized surface plasmon resonance assay, quantitative competitive-PCR, ELISA, western blotting, flow cytometric analysis, and immunofluorescence were performed. RESULTS: Our results showed that GRh2 alleviated neuropathological damage and neuronal apoptosis in cortical tissue of T. gondii-infected mice. GRh2 and CY-09 (an inhibitor of NLRP3) exhibited potent anti-T. gondii effects through binding T. gondii calcium-dependent protein kinase 1 (TgCDPK1). GRh2 decreased Iba-1 (a specific microglial marker) and NLRP3 inflammasome signaling pathway-related protein expression by binding NLRP3. Co-culture of microglia/primary cortical neurons revealed that T. gondii-induced microglial activation caused neuronal apoptosis, but GRh2 reduced this effect, consistent with the effects of CY-09. CONCLUSION: Taken together, our results show that GRh2 has a protective effect against T. gondii infection-induced neuronal injury by binding TgCDPK1 and NLRP3 to inhibit NLRP3 inflammasome signaling pathway in microglia.

Ginsenoside Rh2 reduces depression in offspring of mice with maternal toxoplasma infection during pregnancy by inhibiting microglial activation via the HMGB1/TLR4/NF-κB signaling pathway.

BACKGROUND: Maternal Toxoplasma gondii (T. gondii) infection during pregnancy has been associated with various mental illnesses in the offspring. Ginsenoside Rh2 (GRh2) is a major bioactive compound obtained from ginseng that has an anti-T. gondii effect and attenuates microglial activation through toll-like receptor 4 (TLR4)/nuclear factor-kappa B (NF-κB) signaling pathway. GRh2 also alleviated tumor-associated or lipopolysaccharide-induced depression. However, the effects and potential mechanisms of GRh2 on depression-like behavior in mouse offspring caused by maternal T. gondii infection during pregnancy have not been investigated. METHODS: We examined GRh2 effects on the depression-like behavior in mouse offspring, caused by maternal T. gondii infection during pregnancy, by measuring depression-like behaviors and assaying parameters at the neuronal and molecular level. RESULTS: We showed that GRh2 significantly improved behavioral measures: sucrose consumption, forced swim time and tail suspended immobility time of their offspring. These corresponded with increased tissue concentrations of 5-hydroxytryptamine and dopamine, and attenuated indoleamine 2,3-dioxygenase or enhanced tyrosine hydroxylase expression in the prefrontal cortex. GRh2 ameliorated neuronal damage in the prefrontal cortex. Molecular docking results revealed that GRh2 binds strongly to both TLR4 and high mobility group box 1 (HMGB1). CONCLUSION: This study demonstrated that GRh2 ameliorated the depression-like behavior in mouse offspring of maternal T. gondii infection during pregnancy by attenuating the excessive activation of microglia and neuroinflammation through the HMGB1/TLR4/NF-κB signaling pathway. It suggests that GRh2 could be considered a potential therapy in preventing and treating psychiatric disorders in the offspring mice of mothers with prenatal exposure to T. gondii infection.

Formononetin represses cervical tumorigenesis by interfering with the activation of PD-L1 through MYC and STAT3 downregulation.

A. membranaceus is a traditional Chinese medicine that regulates blood sugar levels, suppresses inflammation, protects the liver, and enhances immunity. In addition, A. membranaceus is also widely used in diet therapy and is a well-known health tonic. Formononetin is a natural product isolated from A. membranaceus that has multiple biological functions, including anti-cancer activity. However, the mechanism by which formononetin inhibits tumor growth is not fully understood. In this present study, we demonstrated that formononetin suppresses PD-L1 protein synthesis via reduction of MYC and STAT3 protein expression. Furthermore, formononetin markedly reduced the expression of MYC protein via the RAS/ERK signaling pathway and inhibited STAT3 activation through JAK1/STAT3 pathway. Co-immunoprecipitation experiments illustrated that formononetin suppresses protein expression of PD-L1 by interfering with the interaction between MYC and STAT3. Meanwhile, formononetin promoted PD-L1 protein degradation via TFEB and TFE3-mediated lysosome biogenesis. T cell killing assay revealed that formononetin could enhance the activity of cytotoxic T lymphocytes (CTLs) and restore ability to kill tumor cells in a co-culture system of T cells and tumor cells. In addition, formononetin inhibited cell proliferation, tube formation, cell migration, and promoted tumor cell apoptosis by suppressing PD-L1. Finally, the inhibitory effect of formononetin on tumor growth was confirmed in a murine xenograft model. The present study revealed the anti-tumor potential of formononetin, and the findings should support further research and development of anti-cancer drugs for cervical cancer.

Erianin regulates programmed cell death ligand 1 expression and enhances cytotoxic T lymphocyte activity.

ETHNOPHARMACOLOGICAL RELEVANCE: Dendrobium chrysotoxum Lindl is a cultivation of Dendrobium which belongs to the family of Orchidaceae. D. chrysotoxum Lindl is a traditional Chinese medicine with a wide range of clinical applications including tonic, astringent, analgesic and anti-inflammatory properties as early as the 28th century B.C. Erianin is a representative index component for the quality control of the D. chrysotoxum Lindl, which is included in the Pharmacopoeia of the People's Republic of China (2020 version). AIM OF THE STUDY: To clarify the anti-tumour mechanisms of erianin in vitro and in vivo. MATERIALS AND METHODS: We detected the anti-tumour activity of erianin using in vitro HeLa cell models and in vivo cervical cancer xenograft models. We performed MTT, western blot, RT-PCR, homology modeling, flow cytometry, and immunoprecipitation assays to study the proteins, genes, and pathways related to erianin's anti-tumour activity. LysoTracker Red staining was performed to detect lysosome function. Transwell, wound healing, tube formation, colony formation and EdU labelling assays were performed to determine cell proliferation, migration and invasion abilities, respectively. Cytotoxic T lymphocytes ability was confirmed using HeLa/T-cell co-culture model. RESULTS: Experimental data demonstrated that erianin inhibited PD-L1 expression and induced the lysosomal degradation of PD-L1. Erianin suppressed HIF-1α synthesis through mTOR/p70S6K/4EBP1 pathway, and inhibited RAS/Raf/MEK/MAPK-ERK pathway. Immunoprecipitation experiments demonstrated that erianin reduced the interaction between RAS and HIF-1α. Experiments using a co-cultivation system of T cells and HeLa cells confirmed that erianin restored cytotoxic T lymphocytes ability to kill tumour cells. Erianin inhibited PD-L1-mediated angiogenesis, proliferation, invasion and migration. The anti-proliferative effects of erianin were supported using in vivo xenotransplantation experiments. CONCLUSIONS: Collectively, these results revealed previously unknown properties of erianin and provided a new basis for improving the efficacy of immunotherapy against cervical cancer and other malignant tumours through PD-L1.

Resveratrol modulates Toxoplasma gondii infection induced liver injury by intervening in the HMGB1/TLR4/NF-κB signaling pathway.

Toxoplasma gondii (T. gondii) is an obligate intracellular parasite that can cause liver diseases in the host, including hepatitis and hepatomegaly. High mobility group box 1 (HMGB1) is the main inflammatory mediator causing cell injury or necrosis. HMGB1 binds to toll like receptor 4 (TLR4), then activates the nuclear factor-κB (NF-κB) signaling pathway, which promotes the release of inflammatory factors. Our previous studies showed that HMGB1 mediated TLR4/NF-κB signaling pathway plays an important role in liver injury induced by T. gondii infection. Resveratrol (RSV) is a small polyphenol, which has anti-inflammatory, anti-cancer, anti-T. gondii effect. However, the effect of RSV on liver injury caused by T. gondii infection is unclear. This study used the RH strain tachyzoites of T. gondii to infect murine liver line, NCTC-1469 cells to establish an in vitro model and acute infection of mice for the in vivo model to explore the protective effect of RSV on liver injury induced by T. gondii infection. The results showed that RSV inhibited the proliferation of T. gondii in the liver, reduced the alanine aminotransferase/aspartate aminotransferase levels and pathological liver damage. Additionally, RSV inhibited the production of tumor necrosis factor-α, inducible nitric oxide synthase and HMGB1 by interfering with the TLR4/NF-κB signaling pathway. These results indicate that RSV can protect liver injury caused by T. gondii infection by intervening in the HMGB1/TLR4/NF-κB signaling pathway. This study will provide a theoretical basis for RSV treatment of T. gondii infection induced liver injury.

Britannin stabilizes T cell activity and inhibits proliferation and angiogenesis by targeting PD-L1 via abrogation of the crosstalk between Myc and HIF-1α in cancer.

BACKGROUND: Programmed cell death-ligand 1 (PD-L1) is overexpressed in tumor cells, which causes tumor cells to escape T cell killing, and promotes tumor cell survival, cell proliferation, migration, invasion, and angiogenesis. Britannin is a natural product with anticancer pharmacological effects. PURPOSE: In this work, we studied the anticancer potential of britannin and explored whether britannin mediated its effect by inhibiting the expression of PD-L1 in tumor cells. METHODS: In vitro, the mechanisms underlying the inhibition of PD-L1 expression by britannin were investigated by MTT assay, homology modeling and molecular docking, RT-PCR, western blotting, co-immunoprecipitation, and immunofluorescence. The changes in tumor killing activity, cell proliferation, cell cycle, migration, invasion, and angiogenesis were analyzed by T cell killing assays, EdU labeling, colony formation, flow cytometry, wound healing, matrigel transwell invasion, and tube formation, respectively. In vivo, the antitumor activity of britannin was evaluated in the HCT116 cell xenograft model. RESULTS: Britannin reduced the expression of PD-L1 in tumor cells by inhibiting the synthesis of the PD-L1 protein but did not affect the degradation of the PD-L1 protein. Britannin also inhibited HIF-1α expression through the mTOR/P70S6K/4EBP1 pathway and Myc activation through the Ras/RAF/MEK/ERK pathway. Mechanistically, britannin inhibited the expression of PD-L1 by blocking the interaction between HIF-1α and Myc. In addition, britannin could enhance the activity of cytotoxic T lymphocytes and inhibit tumor cell proliferation and angiogenesis by inhibiting PD-L1. Finally, in vivo observations were confirmed by demonstrating the antitumor activity of britannin in a murine xenograft model. CONCLUSION: Britannin inhibits the expression of PD-L1 by blocking the interaction between HIF-1α and Myc. Moreover, britannin stabilizes T cell activity and inhibits proliferation and angiogenesis by inhibiting PD-L1 in cancer. The current work highlights the anti-tumor effect of britannin, providing insights into the development of cancer therapeutics via PD-L1 inhibition.

Ginsenoside Rh2 attenuates microglial activation against toxoplasmic encephalitis via TLR4/NF-κB signaling pathway.

BACKGROUND: Ginsenoside Rh2 (GRh2) is a characterized component in red ginseng widely used in Korea and China. GRh2 exhibits a wide range of pharmacological activities, such as anti-inflammatory, antioxidant, and anticancer properties. However, its effects on Toxoplasma gondii (T. gondii) infection have not been clarified yet. METHODS: The effect of GRh2 against T. gondii was assessed under in vitro and in vivo experiments. The BV2 cells were infected with tachyzoites of T. gondii RH strain, and the effects of GRh2 were evaluated by MTT assay, morphological observations, immunofluorescence staining, a trypan blue exclusion assay, reverse transcription PCR, and Western blot analyses. The in vivo experiment was conducted with BALB/c mice inoculated with lethal amounts of tachyzoites with or without GRh2 treatment. RESULTS AND CONCLUSION: The GRh2 treatment significantly inhibited the proliferation of T. gondii under in vitro and in vivo studies. Furthermore, GRh2 blocked the activation of microglia and specifically decreased the release of inflammatory mediators in response to T. gondii infection through TLR4/NF-κB signaling pathway. In mice, GRh2 conferred modest protection from a lethal dose of T. gondii. After the treatment, the proliferation of tachyzoites in the peritoneal cavity of infected mice markedly decreased. Moreover, GRh2 also significantly decreased the T. gondii burden in mouse brain tissues. These findings indicate that GRh2 exhibits an anti-T. gondii effect and inhibits the microglial activation through TLR4/NF-κB signaling pathway, providing the basic pharmacological basis for the development of new drugs to treat toxoplasmic encephalitis.