Exposure to Polypropylene Microplastics Causes Cardiomyocyte Apoptosis Through Oxidative Stress and Activation of the MAPK-Nrf2 Signaling Pathway.

Microplastics are a growing concern as pollutants that impact both public health and the environment. However, the toxic effects of polypropylene microplastics (PP-MPs) are not well understood. This study aimed to investigate the effects of PP-MPs on cardiotoxicity and its underlying mechanisms. The cardiotoxicity of exposure to different amounts of PP-MPs were investigated in both ICR mice and H9C2 cells. Our results demonstrated that sub-chronic exposure to 5 and 50 mg/L PP-MPs led to myocardial structural damage, apoptosis, and fibrosis in mice cardiomyocytes. Flow cytometry analysis revealed that PP-MPs could decrease mitochondrial membrane potential and induce apoptosis in H9C2 cells. Western blotting revealed decreased expression of Bcl-2, poly(ADP-ribose) polymerase (PARP) and caspase 3 and increased expression of Bax, cleaved-PARP, and cleaved-caspase 3 in PP-MPs-treated cardiac tissue and H9C2 cells. These results confirmed the apoptotic effects induced by PP-MPs. Moreover, PP-MPs treatment triggered oxidative stress, as evidenced by the increased levels of malondialdehyde; reduction in glutathione peroxidase, superoxide dismutase, and catalase activities in mice cardiac tissues; and increased reactive oxygen species levels in H9C2 cells. Finally, western blotting demonstrated that exposure to PP-MPs significantly reduced the expression levels of Nrf2 and p-ERK proteins associated with MAPK-Nrf2 pathway in both cardiac tissue and H9C2 cells. Overall, our findings indicate that PP-MPs can induce cardiomyocyte apoptosis through MAPK-Nrf2 signaling pathway, which is triggered by oxidative stress. This study provides a foundation for determining the effects of PP-MPs on cardiotoxicity and their underlying mechanisms.

Design, synthesis, and biological evaluation of triazole-pyrimidine-methylbenzonitrile derivatives as dual A2A/A2B adenosine receptor antagonists.

A series of novel dual A2A/A2B AR antagonists based on the triazole-pyrimidine-methylbenzonitrile core were designed and synthesised. The A2A AR antagonist cAMP functional assay results were encouraging for most target compounds containing quinoline or its open-ring bioisosteres. In addition, compound 7i displayed better inhibitory activity on A2B AR (IC50 14.12 nM) and higher potency in IL-2 production than AB928. Moreover, molecular docking studies were carried out to explain the rationality of molecular design and the activity of compound 7i. Further studies on 7f and 7i revealed good liver microsomes stabilities and acceptable in vivo PK profiles. This study provides insight into the future development of dual A2A/A2B AR antagonists for cancer immunotherapy.

Bifunctional enzyme ATIC promotes propagation of hepatocellular carcinoma by regulating AMPK-mTOR-S6 K1 signaling.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the cancer types with poor prognosis. To effectively treat HCC, new molecular targets and therapeutic approaches must be identified. 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase/inosine monophosphate (IMP) cyclohydrolase (ATIC), a bifunctional protein enzyme, catalyzes the last two steps of the de novo purine biosynthetic pathway. Whether ATIC contributes to cancer development remains unclear. METHODS: ATIC mRNA levels in different types of human HCC samples or normal tissues were determined from Gene Expression across Normal and Tumor tissue (GENT) database. The expression level of ATIC in human HCC samples or cell lines were examined by RT-PCR and western blot. Overall survival and disease-free survival of HCC patients in the ATIC low and ATIC high groups were determined by Kaplan-Meier analysis. Effects of ATIC knockdown by lentivirus infection were evaluated on cell-proliferation, cell-apoptosis, colony formation and migration. The mechanisms involved in HCC cells growth, apoptosis and migration were analyzed by western blot and Compound C (C-C) rescue assays. RESULTS: Here, we first demonstrated that expression of ATIC is aberrantly up-regulated in HCC tissues and high level of ATIC is correlated with poor survival in HCC patients. Knockdown of ATIC expression resulted in a dramatic decrease in proliferation, colony formation and migration of HCC cells. We also identified ATIC as a novel regulator of adenosine monophosphate-activated protein kinase (AMPK) and its downstream signaling mammalian target of rapamycin (mTOR). ATIC suppresses AMPK activation, thus activates mTOR-S6 K1-S6 signaling and supports growth and motility activity of HCC cells. CONCLUSION: Taken together, our results indicate that ATIC acts as an oncogenic gene that promotes survival, proliferation and migration by targeting AMPK-mTOR-S6 K1 signaling.

Microbial HSP70 peptide epitope 407-426 as adjuvant in tumor-derived autophagosome vaccine therapy of mouse lung cancer.

Tumor-derived autophagome (DRibble) is an effective therapeutic cancer vaccine inducing T cell recognition and death of tumor cells in mice. However, the potential for improved anti-tumor response still remains. Our previous study demonstrated that two repeats of a mycobacterial HSP70407-426 (M2) peptide acted as adjuvant in improving anti-tumor efficacy of human umbilical vein endothelial cell (HUVEC) vaccine. Here, a DRibble vaccine conjugated with M2 (DRibble-M2) was designed as a novel vaccine to enhance anti-tumor activity. Compared with DRibble alone, DRibble-M2 vaccination more significantly inhibited the growth of mouse Lewis lung cancer both in a subcutaneous tumor model and in a lung metastasis model. Higher expression of antigen-specific CTL was induced by DRibble-M2. DRibble-M2 induced higher CD83 and CD86 expression in DC2.4 and also improved the internalization of DRibble antigen into DC2.4. Our data indicated that DRibble-M2 is a potential vaccine for clinical cancer therapy.

Morusin, a novel inhibitor of ACLY, induces mitochondrial apoptosis in hepatocellular carcinoma cells through ROS-mediated mitophagy.

OBJECTIVE: Morusin (Mor), a prenylated flavonoid isolated from the root bark of Morus alba L., exhibits potent anti-tumour effects; however, the molecular target of Mor is still not entirely clear. This study aimed to elucidate the mechanism of Mor against hepatocellular carcinoma (HCC) and identify potential molecular targets. METHODS: Mitochondrial function was assessed by measuring the mitochondrial membrane potential, mitochondrial ultrastructure, oxygen consumption, and ATP levels. Mor-induced mitophagy was confirmed using western blotting, immunofluorescence, and fluorescent probes. Transcriptomics, flow cytometry, western blotting, qRT-PCR and biochemical assays were used to reveal the molecular mechanisms and targets of Mor against HCC. We further validated the interaction between Mor and the target proteins using molecular docking and biolayer interferometry (BLI). The inhibitory effect of Mor in vivo was evaluated using a Hep3B murine xenograft model. RESULTS: Mor significantly reduced the ATP citrate lyase (ACLY) expression and inhibited ACLY activity in HCC cells. BLI analysis demonstrated a direct interaction between Mor and the ACLY active domain. Mor-induced ACLY inhibition led to ROS accumulation in HCC cells, which caused mitochondrial damage, triggered PINK1/Parkin-mediated mitophagy, and ultimately induced mitochondrial apoptosis. We further verified that ROS is crucial in the apoptotic action of Mor through experiments regarding an ROS scavenger. Mor also significantly inhibited tumour xenograft growth in vivo. In addition, analysis of human liver cancer clinical samples revealed elevated ACLY levels positively correlated with histologic grade. CONCLUSION: Collectively, our findings highlight Mor as a potent bioactive inhibitor of ACLY and a promising candidate for HCC therapy.

Tiliroside disrupted iron homeostasis and induced ferroptosis via directly targeting calpain-2 in pancreatic cancer cells.

BACKGROUND: Tiliroside (TIL) is a flavonoid compound that exists in a variety of edible plants. These dietary plants are widely used as food and medicine to treat various diseases. However, the effect of TIL on pancreatic cancer (PC) and its underlying mechanisms are unclear. PURPOSE: This study aims to reveal the anti-PC effect of TIL and clarify its mechanism. METHODS: The inhibitory effects of TIL on PC growth were studied both in vitro and in vivo. Flow cytometry, transmission electron microscopy, immunofluorescence, biochemical analyses, RT-qPCR, genetic ablation, and western blotting were employed to evaluate ferroptosis, autophagy, and iron regulation. Additionally, RNA sequencing (RNA-seq), biomolecular layer interferometry (BLI), and molecular simulation analysis were combined to identify TIL molecular targets. The clinicopathological significance of Calpain-2 (CAPN2) was determined through immunohistochemistry (IHC) on a PC tissue microarray. RESULTS: Herein, we showed that TIL was an effective anti-PC drug. CAPN2 was involved in the TIL - induced elevation of the labile iron pool (LIP) in PC cells. TIL directly bound to and inhibited CAPN2 activity, resulting in AKT deactivation and decreased expression of glucose transporters (GLUT1 and GLUT3) in PC cells. Consequently, TIL impaired ATP and NADPH generation, inducing autophagy and ROS production. The accumulation of TIL-induced ROS combined with LIP iron causes the Fenton reaction, leading to lipid peroxidation. Meanwhile, TIL-induced reduction of free iron ions promoted autophagic degradation of ferritin to regulate cellular iron homeostasis, which further exacerbated the death of PC cells by ferroptosis. As an extension of these in vitro findings, our murine xenograft study showed that TIL inhibited the growth of PANC-1 cells. Additionally, we showed that CAPN2 expression levels were related to clinical prognoses in PC patients. CONCLUSION: We identify TIL as a potent bioactive inhibitor of CAPN2 and an anti-PC candidate of natural origin. These findings also highlight CAPN2 as a potential target for PC treatment.

Potential Effects of Orally Ingesting Polyethylene Terephthalate Microplastics on the Mouse Heart.

Polyethylene terephthalate microplastics (PET MPs) are widespread in natural environment, and can enter organisms and accumulate in the body, but its toxicity has not been well studied. Therefore, in order to investigate the toxic effects of PET microplastics on mammals, this study investigated the toxic effects of PET MPs on ICR mice and H9C2 cells by different treatment groups. The results indicated the cardiac tissue of mice in the PET-H (50 µg/mL) group showed significant capillary congestion, myocardial fiber breakage, and even significant fibrosis compared to the PET-C (control) group (P < 0.01). Results of the TUNEL assay demonstrated significant apoptosis in myocardial tissue in the PET-H and PET-M (5 µg/mL) groups (P < 0.01). Meanwhile, Western blotting showed increased expression of the apoptosis-related protein Bax and decreased expression of PARP, caspase-3, and Bcl-2 proteins in both myocardial tissues and H9C2 cells. In addition, flow cytometry confirmed that PET MPs decreased the mitochondrial membrane potential and apoptosis in H9C2 cells; however, this trend was reversed by N-acetylcysteamine application. Moreover, PET MP treatment induced the accumulation of reactive oxygen species (ROS) in H9C2 cells, while the MDA level in the myocardial tissue was elevated, and the activities of catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GSH-Px) were decreased (P < 0.01), indicating a change in the redox environment. In conclusion, PET MPs promoted cardiomyocyte apoptosis by inducing oxidative stress and activating mitochondria-mediated apoptotic processes, ultimately leading to myocardial fibrosis. This study provides ideas for the prevention of PET MP toxicity and promotes thinking about enhancing plastic pollution control.

Enhanced antitumor efficacy by combination treatment with a human umbilical vein endothelial cell vaccine and a tumor cell lysate-based vaccine.

Angiogenesis inhibitors combined with other anticancer drugs have been shown to inhibit tumor growth in animal models and some of them were recently used in clinical trials. In the present study, a whole hepatocellular carcinoma cell lysate based vaccine with diphtheria toxin (DT) and two tandem repeats of microbial HSP70 peptide epitope 407-426 (2 mHSP70407-426, M2) as adjuvant, which was called HDM, was combined with a whole human umbilical vein endothelial cell (HUVEC) vaccine to develop a combination treatment regimen. This combination treatment regimen was named HUVEC-HDM, which was supposed to enhance its antitumor efficiency. HUVEC-HDM was administrated subcutaneously in both prophylactic and therapeutic procedures. Compared to either single vaccine, HUVEC-HDM induced a more significant inhibition on the growth and metastasis of H22 hepatocellular carcinoma in mice and prolonged the survival of tumor-bearing mice. Besides, HUVEC-HDM immunization elicited strong humoral and cellular immune responses targeting tumor cell as well as tumor angiogenesis, which could be responsible for the enhanced antitumor effect. Moreover, histochemistry analysis showed that HUVEC-HDM induced large areas of continuous necrosis within tumors, correlating well with the extent of tumor inhibition. These results not only highlight the superiority of the combined HUVEC-HDM treatment regimen, but also support the translation of such approaches into the clinic for the treatment of patients with hepatocellular carcinoma.

Improved efficacy of therapeutic vaccination with viable human umbilical vein endothelial cells against murine melanoma by introduction of OK432 as adjuvant.

Vaccination with xenogeneic or syngeneic endothelial cells targeting tumor angiogenesis is effective for inhibiting tumor growth. OK432, an effective adjuvant, was mixed with viable human umbilical vein endothelial cells (HUVECs) to prepare a novel HUVECs-OK432 vaccine, which could have an improved therapeutic efficacy. In this study, HUVECs-OK432 was administrated in mice by subcutaneous injection in a therapeutic procedure. The results showed that a stronger HUVEC-specific Abs and cytotoxic T lymphocyte immune response were elicited, which resulted in significant inhibition on the growth of B16F10 melanoma and remarkably prolonged survival of B16F10 melanoma-bearing mice compared with HUVECs. Besides, parallel results were obtained in vitro showing a stronger inhibition of HUVEC proliferation by immune sera of HUVECs-OK432 than that of HUVECs. Moreover, histochemistry and immunohistochemistry analysis showed that HUVECs-OK432 induced large areas of continuous necrosis within tumors and significantly reduced the vessel density, correlating well with the extent of tumor inhibition. Our present results suggest that OK432 could be employed as an effective adjuvant for HUVEC vaccines and therefore should be useful for adjuvant immunotherapy of cancer.

Tiliroside targets TBK1 to induce ferroptosis and sensitize hepatocellular carcinoma to sorafenib.

BACKGROUND: Combination therapy with other antineoplastic agent is a favorable approach for targeting the molecules involved in sorafenib resistance. PURPOSE: In the present study, we determined whether tiliroside, a natural flavonoid glycoside isolated from oriental paperbush flower, could improve the sensitivity of hepatocellular carcinoma (HCC) cells to sorafenib. Furthermore, we investigated the mechanisms and identified the potential drug targets of tiliroside. METHODS: Synergy was performed using CalcuSyn. Transcriptomic studies were adopted to investigate whether tiliroside could induce ferroptosis and inhibit the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway in HCC cells. Ferroptosis was analyzed using western blotting, flow cytometry, and transmission electron microscopy. Immunofluorescence, co-immunoprecipitation, and Nrf2 knockdown or overexpression were performed to confirm the involvement of Nrf2 in tiliroside-induced ferroptosis. Additionally, molecular docking and biolayer interferometry-based measurements were used to confirm the direct target of tiliroside. Finally, subcutaneous xenograft and orthotopic xenograft tumors in nude mice were used to assess the effects of tiliroside in vivo. RESULTS: Tiliroside significantly enhanced the anti-HCC activity of sorafenib without any discernible side effects. Moreover, the combination of tiliroside and sorafenib induced synergistic effects against HCC in vitro. The inhibitory effects of tiliroside on HCC were antagonized by N-acetylcysteine and the ferroptosis inhibitor liproxstatin-1. Studies on the mechanism of action revealed that tiliroside could directly bind to TANK-binding kinase 1 (TBK1) and inhibit its enzymatic activity. Inhibition of TBK1 by tiliroside decreased the phosphorylation of serine 349 on sequestosome-1 (p62) and the affinity of p62 for kelch like ECH-associated protein 1 (Keap1) and promoted Keap1-mediated Nrf2 ubiquitination and degradation. The downstream target proteins of Nrf2, including glutathione peroxidase 4, ferritin heavy chain 1, and glucose-6-phosphate dehydrogenase, demonstrated similar results to that of Nrf2 protein, inducing ferroptosis in tiliroside-treated HCC cells. We extended these findings in vivo and found that tiliroside inhibited the growth of HepG2 tumors in both subcutaneous xenograft and orthotopic xenograft tumor models of HCC. CONCLUSION: Our findings imply that tiliroside is a potent TBK1 inhibitor and a candidate natural anti-cancer product that could function as a sensitizer of sorafenib in HCC treatment by targeting TBK1 to induce ferroptosis.

Self-Assembled Virus-Like Particle Vaccines via Fluorophilic Interactions Enable Infection Mimicry and Immune Protection.

Influenza epidemics persistently threaten global health. Vaccines based on virus-like particles (VLPs), which resemble the native conformation of viruses, have emerged as vaccine candidates. However, the production of VLPs via genetic engineering remains constrained by challenges such as low yields, high costs, and being time consuming. In this study, a novel VLP platform is developed that could mimic infection and confer influenza protection through fluorination-driven self-assembly. The VLPs closely mimick the key steps in viral infection including dendritic cell (DC) attachment and pH-responsive endo-lysosomal escape, which enhances DC maturation and antigen cross-presentation. It is also observed that the VLPs migrate from the injection site to the draining lymph nodes efficiently. Immunization with VLPs triggers both Th1 and Th2 cellular responses, thereby inducing an improved CD8+ T cell response along with strong antigen-specific antibody responses. In several infected mouse models, VLP vaccines ameliorate weight loss, lung virus titers, pulmonary pathologies, and confer full protection against H1N1, H6N2, H9N2, and mixed influenza viruses. Therefore, the results support the potential of VLPs as an effective influenza vaccine with improved immune potency against infection. A methodology to generate VLPs based on fluorophilic interactions, which can be a general approach for development of pathogenic VLPs, is reported.

Vaccination with the repeat ß-hCG C-terminal peptide carried by heat shock protein-65 (HSP65) for inducing antitumor effects.

The ß-subunit of human chorionic gonadotropin (ß-hCG) is ectopically expressed in various types of cancer and has been utilized as an antigenic target in anti-cancer vaccines. In view of the low immunogenicity of this self-peptide, we designed a method based on the isocaudamer technique to generate 14 tandem repeats of the 10-residue sequence X of ß-hCG (109-118). These tandemly repeated copies were then combined with ß-hCG C-terminal 37 peptides (CTP37) and finally fused to mycobacterial heat-shock protein 65 (HSP65) to construct a fusion protein HSP65-X14-ßhCGCTP37 as an immunogen. In this study, BALB/c female mice were immunized via subcutaneous injection of the designed protein. Humoral immune and cellular immune responses were effectively elicited. A high titer of anti-ß-hCG antibody was detected in immunized mice sera by enzyme-linked immunosorbent assay and verified by Western blot analysis. The fusion protein, HSP65-X14-ß-hCGCTP37, effectively inhibited the growth of Ehrlich ascites carcinoma in mice. These results suggest that HSP65-X14-ßhCGCTP37 may be an effective tumor vaccine, and the use of multiple tandem repeats of a certain epitope is an effective method to overcome the low immunogenicity of self-peptide antigens.

Protective antitumor immunity induced by tumor cell lysates conjugated with diphtheria toxin and adjuvant epitope in mouse breast tumor models.

Cancer cell vaccine-based immunotherapy has received increasing interest in many clinical trials involving patients with breast cancer. Combining with appropriate adjuvants can enhance the weak immunogenic properties of tumor cell lysates (TCL). In this study, diphtheria toxin (DT) and two tandem repeats of mycobacterial heat shock protein 70 (mHSP70) fragment 407-426 (M2) were conjugated to TCL with glutaraldehyde, and the constructed cancer cell vaccine was named DT-TCL-M2. Subcutaneous injection of DT-TCL-M2 in mice effectively elicited tumor-specific polyclonal immune responses, including humoral and cellular immune responses. High levels of antibodies against TCL were detected in the serum of immunized mice with ELISA and verified with Western blot analyses. The splenocytes from immunized mice showed potent cytotoxicity on Ehrlich ascites carcinoma cells. Moreover, the protective antitumor immunity induced by DT-TCL-M2 inhibited tumor growth in a mouse breast tumor model. DT-TCL-M2 also attenuated tumor-induced angiogenesis and slowed tumor growth in a mouse intradermal tumor model. These findings demonstrate that TCL conjugated with appropriate adjuvants induced effective antitumor immunity in vivo. Improvements in potency could further make cancer cell vaccines a useful and safe method for preventing cancer recurrence after resection.

Chrysin induces autophagy-dependent ferroptosis to increase chemosensitivity to gemcitabine by targeting CBR1 in pancreatic cancer cells.

Recent studies have verified that inducing reactive oxygen species (ROS) is one of the gemcitabine anti-tumor mechanisms of action. Human carbonyl reductase 1 (CBR1) plays an important role in protecting cells against oxidative damage. However, it is unclear whether CBR1 is involved in pancreatic cancer (PC) progression and resistance to gemcitabine. Based on the GEPIA database, we analyzed tumor tissue samples from PC patients using immunohistochemistry (IHC) and revealed that CBR1 was highly expressed in PC tissues and that this was significantly correlated with the clinicopathological features of PC. Genetic inhibition of CBR1 suppressed PC cell proliferation by regulating ROS generation. Furthermore, gemcitabine upregulated CBR1 expression, which could limit the anti-tumor activity of gemcitabine, and attenuation of CBR1 enhanced gemcitabine sensitivity in vitro and in vivo. Additionally, we report that chrysin directly binds to CBR1, which inhibited its enzymatic activity both at the molecular and cellular levels. Inhibition of CBR1 by chrysin increased cellular ROS levels and led to ROS-dependent autophagy, which resulted in the degradation of ferritin heavy polypeptide 1 (FTH1) and an increase in the intracellular free iron level that participates in ferroptosis in PC cells. Finally, our results showed that chrysin enhanced PC sensitivity to gemcitabine by inducing ferroptotic death in vitro and in vivo. Collectively, these findings indicate that CBR1 is a potential therapeutic target for PC treatment. In addition, we elucidated a novel mechanism underlying the anti-tumor effects of chrysin.

Low-dose docetaxel enhances the anti-tumour efficacy of a human umbilical vein endothelial cell vaccine.

Our previous studies have indicated that human umbilical vein endothelial cell (HUVEC) vaccination appears to be a potentially promising anti-angiogenesis therapy, but the modest therapeutic anti-tumour efficiency limits its clinical use. This highlights the importance of identifying more potent therapeutic HUVEC vaccine strategies for clinical testing. In the present study, the immune-modulating doses of docetaxel (DOC) was combined with 1 × 106 viable HUVECs as a means to enhance the therapeutic anti-tumour efficiency of the HUVEC vaccine. Our results demonstrated that 5 mg/kg DOC administrated prior to HUVEC vaccine could most effectively assist HUVEC vaccine to display a remarkable suppression of tumour growth and metastasis as wells as a prolongation of survival time in a therapeutic procedure. CD31 immunohistochemical analysis of the excised tumours confirmed a significant reduction in vessel density after treatment with the HUVEC vaccine with 5 mg/kg DOC. Additionally, an increased HUVEC-specific antibody level, activated CTLs and an elevated IFN-γ level in cultured splenocytes were revealed after treatment with HUVEC vaccine with 5 mg/kg DOC. Finally, 5 mg/kg DOC coupled with the HUVEC vaccine led to induction of significant increases in CD8+T cells and decrease in Tregs in the tumour microenvironment. Taken together, all the results verified that 5 mg/kg DOC could assist HUVEC vaccine to elicit strong HUVEC specific humoral and cellular responses, which could facilitate the HUVEC vaccine-mediated inhibition of cancer growth and metastasis. These findings provide the immunological rationale for the combined use of immune-modulating doses of DOC and HUVEC vaccines in patients with cancer.

TLR2 deficiency attenuated chronic intermittent hypoxia-induced neurocognitive deficits.

Chronic intermittent hypoxia (CIH) is the main symptom of obstructive sleep apnea syndrome (OSAS) and causes neural damage and cognitive deficits via neuroinflammation. Toll-like receptors (TLRs), especially TLR2, play an important role in neuroinflammation. However, the mechanisms by which TLR2 participates in CIH-induced cognitive deficits remain unclear. In this study, wild-type (WT) and TLR2 knock out (KO) mice were exposed to CIH for 8 weeks, and their social novelty discrimination, spatial learning and memory were severely compromised. Additionally, seriously damaged neurons and abnormally activated glia were observed in the CA1 and dentate gyrus (DG) areas of the hippocampus. Mechanistically, knocking out the TLR2 gene significantly alleviated these pathological changes and improved the behavioral performance. Together, these findings demonstrate that the TLR2-MyD88 signaling pathway might play an important role in CIH-induced cognitive deficits.

YAP Inhibition by Nuciferine via AMPK-Mediated Downregulation of HMGCR Sensitizes Pancreatic Cancer Cells to Gemcitabine.

Nuciferine, a major aporphine alkaloid constituent of lotus leaves, is a raw material for obesity treatment. Extensive studies have revealed that obesity is associated with pancreatic cancer (PC). However, it has not been clarified whether nuciferine could be used in PC treatment or prevention. Here, we show that nuciferine could enhance the sensitivity of PC cells to gemcitabine in both cultured cells and the xenograft mouse model. The mechanism study demonstrated that nuciferine induced YAP Ser127 phosphorylation [pYAP(Ser127)] through AMPK-mediated 3-hydroxy-3-methyl-glutaryl-coA reductase (HMGCR) downregulation. Remarkably, wild-type YAP overexpression or YAP Ser127 mutant could resist to nuciferine and no longer sensitize PC cells to gemcitabine. Knockdown of AMPK attenuated pYAP(Ser127) induced by nuciferine. Moreover, knockdown of AMPK reversed nuciferine-mediated HMGCR downregulation. Notably, HMGCR inhibiting could restrain YAP by phosphorylation Ser 127, and therefore enhance the efficiency of gemcitabine in PC cells. In line with this consistent, overexpression of HMGCR reduced growth inhibition caused by nuciferine and/or gemcitabine treatment in PC cells. In summary, these results provide an effective supplementary agent and suggest a therapeutic strategy to reduce gemcitabine resistance in PC.

Ailanthone Induces Cell Cycle Arrest and Apoptosis in Melanoma B16 and A375 Cells.

Malignant melanoma is the most lethal type of skin cancer. Previous studies have shown that ailanthone has potent antitumor activity in a variety of cell lines. However, the anti-tumor effect of ailanthone on malignant melanoma remains unclear. To investigate the anti-tumor mechanisms of ailanthone in human melanoma B16 and mouse melanoma A375 cells, the cell counting kit-8 assay, colony formation assay, DNA content analysis, Hoechst 33258, and Annexin V-FITC/PI staining were used to assess cell proliferation, cell cycle distribution, and cell apoptosis, respectively. Western blotting was performed to evaluate the expression of cell cycle- and apoptosis-related proteins and regulatory molecules. The results showed that ailanthone significantly inhibited melanoma B16 and A375 cell proliferation as well as remarkably induced cell cycle arrest at the G0-G1 phase in B16 cells and the G2-M phase in A375 cells in a dose-dependent manner. Further investigation revealed that ailanthone promoted the expression of p21 and suppressed the expression of cyclin E in B16 cells or cyclin B in A375 cells through the PI3K-Akt signaling pathway. In addition, ailanthone induced B16 and A375 cell apoptosis via a caspase-dependent mechanism. Further studies showed that ailanthone remarkably downregulated Bcl-2 and upregulated Apaf-1 and Bax, and subsequently increased mitochondrial membrane permeabilization and released cytochrome c from the mitochondria in B16 cells and A375 cells. Taken together, ailanthone induces cell cycle arrest via the PI3K-Akt signaling pathway as well as cell apoptosis via the mitochondria-mediated apoptotic signaling pathway. Ailanthone may be potentially utilized as an anti-tumor agent in the management of malignant melanoma.

Combination of Human Umbilical Vein Endothelial Cell Vaccine and Docetaxel Generates Synergistic Anti-Breast Cancer Effects.

Background: A human umbilical vein endothelial cell (HUVEC) vaccine is a promising anti-angiogenesis therapy, but the modest therapeutic antitumor efficacy restricts its clinical use. Preclinical evidence supports the combination of antiangiogenic agents and chemotherapy for cancer treatment. Materials and Methods: In the present study, docetaxel (DOC) was combined with HUVEC vaccine to develop a HUVEC-DOC treatment regime. This study was designed to investigate the synergistic anti-breast cancer effects and mechanisms of the HUVEC-DOC treatment. Results: Compared with either agent monotherapy, HUVEC-DOC treatment exhibited more favorable anti-EMT-6 breast cancer effects in vivo. CD31 immunohistochemical analysis of the excised tumors showed notable decreases in vessel density after HUVEC-DOC administration, while T cells isolated from mice immunized with HUVEC-DOC showed increased cytotoxicity against HUVECs. Furthermore, the quantity of interferon gamma released from HUVEC-DOC-administered mice was significantly higher than the other three groups, and enhanced CD8+ T cell infiltration was observed more frequently in tumors excised from HUVEC-DOC-treated mice. Finally, the percentage of regulatory T cells was significantly decreased after HUVEC-DOC immunization. Conclusions: All the data verified that combining DOC with a HUVEC vaccine could generate synergistic anti-breast cancer activity, which might have the potential for combination treatment of human breast cancer.

Cannabidiol Induces Cell Cycle Arrest and Cell Apoptosis in Human Gastric Cancer SGC-7901 Cells.

The main chemical component of cannabis, cannabidiol (CBD), has been shown to have antitumor properties. The present study examined the in vitro effects of CBD on human gastric cancer SGC-7901 cells. We found that CBD significantly inhibited the proliferation and colony formation of SGC-7901 cells. Further investigation showed that CBD significantly upregulated ataxia telangiectasia-mutated gene (ATM) and p53 protein expression and downregulated p21 protein expression in SGC-7901 cells, which subsequently inhibited the levels of CDK2 and cyclin E, thereby resulting in cell cycle arrest at the G0-G1 phase. In addition, CBD significantly increased Bax expression levels, decreased Bcl-2 expression levels and mitochondrial membrane potential, and then upregulated the levels of cleaved caspase-3 and cleaved caspase-9, thereby inducing apoptosis in SGC-7901 cells. Finally, we found that intracellular reactive oxygen species (ROS) increased after CBD treatment. These results indicated that CBD could induce G0-G1 phase cell cycle arrest and apoptosis by increasing ROS production, leading to the inhibition of SGC-7901 cell proliferation, thereby suggesting that CBD may have therapeutic effects on gastric cancer.