Prevention of cyclophosphamide-induced immune suppression by polysaccharides from Apocynum venetum flowers via enhancing immune response, reducing oxidative stress, and regulating gut microbiota in mice.

Introduction: Emerging proof suggests that Apocynum venetum flowers polysaccharide (AVFP) has immunomodulatory effects in vitro. However, the action mechanism of AVFA is still unclear in vivo. The purpose of this study is to probe into the potential mechanism of AVFA in immunosuppressed mice by investigating organ index, cytokine levels, anti-oxidative stress capacity, transcriptomics, and gut microbiota. Methods: Immunocompromised mice induced by cyclophosphamide (CTX) were divided into six groups. The enzyme-labeled method, hematoxylin and eosin, transcriptomics, and high-throughput sequencing were used to detect the regulatory effects of AVFP on immunocompromised mice and the function of AVFP on the concentration of short-chain fatty acids (SCFAs) by high-performance liquid chromatography (HPLC) analysis. The Spearman correlation analysis was used to analyze the correlation between the intestinal microbiota and biochemical indexes. Results: The experimental results illustrated that AVFP has protective effects against CTX-induced immunosuppression in mice by prominently increasing the organ index and levels of anti-inflammatory factors in serum in addition to enhancing the antioxidant capacity of the liver. Meanwhile, it could also signally decrease the level of pro-inflammatory cytokines in serum, the activity of transaminase in serum, and the content of free radicals in the liver, and alleviate the spleen tissue damage induced by CTX. Transcriptomics results discovered that AVFP could play a role in immune regulation by participating in the NF-κB signaling pathway and regulating the immune-related genes Bcl3, Hp, Lbp, Cebpd, Gstp2, and Lcn2. Gut microbiota results illustrated that AVFP could increase the abundance of beneficial bacteria, reduce the abundance of harmful bacteria, and regulate the metabolic function of intestinal microorganisms while dramatically improving the content of SCFAs, modulating immune responses, and improving the host metabolism. The Spearman analysis further evaluated the association between intestinal microbiota and immune-related indicators. Conclusion: These findings demonstrated that AVFP could enhance the immune effects of the immunosuppressed mice and improve the body's ability to resist oxidative stress.

Late Positivity Correlates with Subjective Reports: Evidence from the Low-frequency and High-frequency Reporting Tasks.

Identifying the neural correlates of consciousness (NCCs) is an important way to understand the fundamental nature of consciousness. By recording event-related potentials (ERPs) using EEG, researchers have found three potential electrophysiological NCCs: early positive correlate of consciousness (enhanced P1), visual awareness negativity (VAN), and late positivity (LP). However, LP may reflect post-perceptual processing associated with subjective reports rather than consciousness per se. The present experiment investigated the relationship between LP and subjective reports. We adopted two subjective reporting tasks that differed in the requirement for subjective reports. In the low-frequency reporting task, participants needed to report whether they saw the target picture in 25% of trials, whereas in the high-frequency reporting task, participants needed to report whether they saw the target picture in each trial. Behavioral results showed that the hit rates were lower and false alarm rates were higher on reporting trials in low-frequency reporting tasks than on reporting trials in high-frequency reporting tasks. Unexpectedly, VAN was larger on reporting trials in the low-frequency reporting task than on reporting trials in the high-frequency reporting task. Importantly, our ERP results showed that LP was larger on reporting trials in the high-frequency reporting task than on reporting trials in the low-frequency reporting task. Thus, our findings indicated that when the frequency of reports was increased, the task relevance of the stimuli increased, which led to larger LP amplitudes. These findings suggest that LP correlates with subjective reports.

Carbonic anhydrase IX inhibitor S4 triggers release of DAMPs related to immunogenic cell death in glioma cells via endoplasmic reticulum stress pathway.

BACKGROUND: Immunogenic cell death (ICD), which releases danger-associated molecular patterns (DAMP) that induce potent anticancer immune response, has emerged as a key component of therapy-induced anti-tumor immunity. The aim of this work was to analyze whether the carbonic anhydrase IX inhibitor S4 can elicit ICD in glioma cells. METHODS: The effects of S4 on glioma cell growth were evaluated using the CCK-8, clonogenic and sphere assays. Glioma cell apoptosis was determined by flow cytometry. Surface-exposed calreticulin (CRT) was inspected by confocal imaging. The supernatants of S4-treated cells were concentrated for the determination of HMGB1and HSP70/90 expression by immunoblotting. RNA-seq was performed to compare gene expression profiles between S4-treated and control cells. Pharmacological inhibition of apoptosis, autophagy, necroptosis and endoplasmic reticulum (ER) stress was achieved by inhibitors. In vivo effects of S4 were evaluated in glioma xenografts. Immunohistochemistry (IHC) was performed to stain Ki67 and CRT. RESULTS: S4 significantly decreased the viability of glioma cells and induced apoptosis and autophagy. Moreover, S4 triggered CRT exposure and the release of HMGB1 and HSP70/90. Inhibition of either apoptosis or autophagy significantly reversed S4-induced release of DAMP molecules. RNA-seq analysis indicated that the ER stress pathway was deregulated upon exposure to S4. Both PERK-eIF2α and IRE1α- XBP1 axes were activated in S4-treated cells. Furthermore, pharmacological inhibition of PERK significantly suppressed S4-triggered ICD markers and autophagy. In glioma xenografts, S4 significantly reduced tumor growth. CONCLUSIONS: Altogether, these findings suggest S4 as a novel ICD inducer in glioma and might have implications for S4-based immunotherapy. Video Abstract.

Iridoids and active ones in patrinia: A review.

Iridoid is a special class of monoterpenoids, whose basic skeleton is the acetal derivative of antinodilaldehyde with a bicyclic H-5/H-9ß, ß-cisfused cyclopentan pyran ring. They were often existed in Valerianaceae, Rubiaceae, Scrophulariaceae and Labiaceae family, and has various biological activities, such as anti-inflammatory, hypoglycemic, neuroprotection, and soon. In this review, iridoids from Patrinia (Valerianaceae family), and the active ones as well as their mechanisms in recent 20 years were summarized. Up to now, a total of 115 iridoids had been identified in Patrinia, among which 48 had extensive biological activities mainly presented in anti-inflammatory, anti-tumor and neuroprotective. And the mechanisms involved in MAPK, NF-κB and JNK signal pathways. The summary for iridoids and their activities will provide the evidence to exploit the iridoids in Patrinia.

UM-164, a Dual Inhibitor of c-Src and p38 MAPK, Suppresses Proliferation of Glioma by Reducing YAP Activity.

UM-164 is a dual inhibitor of c-Src and p38 MAPK, and has been a lead compound for targeting triple-negative breast cancer. UM-164 shows stronger binding to the active sites of Src compared with the conventional Src inhibitor Dasatinib. While Dasatinib has displayed some inhibitory effects on glioma growth in clinical trials, whether UM-164 can suppress glioma growth has not been reported. Here we show that UM-164 suppressed the proliferation, migration and spheroid formation of glioma cells, and induced cell cycle arrest in the G1 phase. Moreover, UM-164 triggered YAP translocation to the cytoplasm and reduced the activity of YAP, as evidenced by a luciferase assay. Accordingly, UM-164 markedly decreased the expression levels of YAP target genes CYR61 and AXL. Importantly, ectopic expression of wild-type YAP or YAP-5SA (YAP constitutively active mutant) could rescue the anti-proliferative effect induced by UM-164. Intriguingly, p38 MAPK appears to play a greater role than Src in UM-164-mediated inhibition of YAP activity. Furthermore, the in vitro anti-glioma effect mediated by UM-164 was confirmed in a xenograft glioma model. Together, these findings reveal a mechanism by which UM-164 suppresses the malignant phenotypes of glioma cells and might provide a rationale for UM-164-based anti-glioma clinical trials.

Circular RNA circ_0001588 sponges miR-211-5p to facilitate the progression of glioblastoma via up-regulating YY1 expression.

BACKGROUND: As the most common and detrimental brain tumor with high invasiveness and poor prognosis, glioblastoma (GBM) has severely threatened people's health globally. Therefore, it is of great importance and necessary to identify the molecular mechanisms involved in tumorigenesis and development, thus contributing to potential therapeutic targets and strategies. METHODS: The level of circ_0001588 was detected in 68 pairs of GBM tissues and adjacent normal tissues and human glioma cell lines via a real-time quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). Then, the effect of circ_0001588 on the proliferation, migration and invasion of glioma cells was evaluated. In addition, potential downstream targets of circ_0001588 were forecasted by circBANK and Starbase. Their interaction was confirmed by introducing luciferase reporter assays. Moreover, sh-circ_0001588 transfected U251 cells were used to form tumors in vivo. Finally, the functional mechanism of circ_0001588 was identified by qRT-PCR, western blotting, xenograft and immunohistochemistry (IHC) assays. RESULTS: The expression of circ_0001588 is markedly up-regulated in GBM tissues and human gliomas cells. Additionally, increased expression of circ_0001588 is positively relevant with poor survival in GBM patients. The down-regulation of circ_0001588 distinctly inhibits the proliferation, migration and invasion of GBM in vitro, as well as tumor growth in vivo. Moreover, knockdown of circ_0001588 reduces the tumor volume and weight, enhances the relative IHC staining index of E-cadherin and decreases the relative IHC staining index of Ki-67, Yin Yang 1 (YY1) and vinmentin in vivo. Mechanistically, circ_0001588 locates in the cytoplasm, which is directly bound with miR-211-5p. Furthermore, circ_0001588 can positively regulate YY1 via sponging miR-211-5p. Moreover, circ_0001588 accelerates the proliferation, migration and invasion of GBM by modulating miR-211-5p/YY1 signaling. CONCLUSIONS: These results illustrate a new circ_0001588/miR-211-5p/YY1 regulatory signaling axis in GBM.

Downregulation of miR-485-3p promotes glioblastoma cell proliferation and migration via targeting RNF135.

MicroRNAs (miRNAs) are small non-coding RNAs that serve pivotal roles in human diseases. Several miRNAs, such as miR-485-3p, have been identified as potential biomarkers for predicting overall survival of patients with glioblastoma (GBM). However, the underlying mechanism of miRNAs in promoting GBM progression remains unknown. In the present study, decreased miR-485-3p expression was detected in tumor tissues from patients with GBM. Using western blot analysis, reverse transcription-quantitative PCR and dual luciferase reporter assay, ring finger protein 135 (RNF135) was confirmed as a target gene of miR-485-3p in GBM cells. Through silencing of RNF135, miR-485-3p inactivated the mitogen-activated protein kinase/ERK1/2 pathway in GBM cells. Moreover, functional assays demonstrated that miR-485-3p inhibited GBM cell proliferation and migration whilst overexpression of RNF135 reversed this effect. Additionally, a negative correlation between miR-485-3p and RNF135 mRNA expression was observed in tissues from patients with glioblastoma. In conclusion, the present results demonstrated that miR-485-3p functioned as a tumor suppressor which suggested that miR-485-3p might have a role in GBM progression.

MicroRNA-374a Governs Aggressive Cell Behaviors of Glioma by Targeting Prokineticin 2.

MicroRNA-374a has been abnormally expressed in several cancer types; however, its role in glioma remains unclear. Therefore, we aimed to investigate whether microR-374a participated in the progression of glioma. Expression of microR-374a in glioma cell lines and normal cell line was measured by quantitative real-time polymerase chain reaction. Luciferase reporter assay and Western blot were used to detect the targets of microR-374a. In vitro functional experiments were conducted to investigate the biological role of microR-374a. Low expression of microR-374a was found in glioma cell lines. Prokineticin 2 was identified as a direct target of microR-374a in glioma. Investigations on the mechanisms related to glioma progression showed that microR-374a inhibited glioma cell proliferation, cell cycle progression, and cell invasion through targeting Prokineticin 2. Taken together, these results revealed that microR-374a functions as tumor suppressor by targeting Prokineticin 2, suggesting it might be a novel therapeutic target for glioma.

Bufalin Induces Apoptosis and Improves the Sensitivity of Human Glioma Stem-Like Cells to Temozolamide.

Glioma is the most common malignant tumor of the central nervous system, and it is characterized by high relapse and fatality rates and poor prognosis. Bufalin is one of the main ingredients of Chan-su, a traditional Chinese medicine (TCM) extracted from toad venom. Previous studies revealed that bufalin exerted inhibitory effects on a variety of tumor cells. To demonstrate the inhibitory effect of bufalin on glioma cells and glioma stem-like cells (GSCs) and discuss the underlying mechanism, the proliferation of glioma cells was detected by MTT and colony formation assays following treatment with bufalin. In addition, we investigated whether bufalin inhibits or kills GSCs using flow cytometry, Western blotting, and reverse transcription polymerase chain reaction analysis (RT-PCR). Finally, we investigated whether bufalin could improve the therapeutic effect of temozolomide (TMZ) and discussed the underlying mechanism. Taken together, our data demonstrated that bufalin inhibits glioma cell growth and proliferation, inhibits GSC proliferation, and kills GSCs. Bufalin was found to induce the apoptosis of GSCs by upregulating the expression of the apoptotic proteins cleaved caspase 3 and poly(ADP-ribose) polymerase (PARP) and by downregulating the expression of human telomerase reverse transcriptase, which is a marker of telomerase activity. Bufalin also improved the inhibitory effect of TMZ on GSCs by activating the mitochondrial apoptotic pathway. These results suggest that bufalin damages GSCs, induces apoptosis, and enhances the sensitivity of GSCs to TMZ.

S-nitrosocaptopril interrupts adhesion of cancer cells to vascular endothelium by suppressing cell adhesion molecules via inhibition of the NF-кB and JAK/STAT signal pathways in endothelial cells.

Inflammatory cytokines can induce the expression of cell adhesion molecules (CAMs) in endothelial cells. The induction may play an important role in attracting circulating tumor cells (CTCs) to endothelial cells. S-nitrosocaptopril (CapNO) is known to produce vasorelaxation and interfere the hetero-adhesion of CTCs to vascular endothelium via down-regulating the expression of CAMs. To elucidate the mechanisms underlying the inhibition of CapNO on CAMs, in this study, we examined the relationship between cytokines and CAMs expression and investigated the effects of CapNO on cytokine-induced NF-кB and JAK/STAT signal pathways. The activation of CAMs by cytokines was dependent on concentrations and reaction time of cytokines, and the combination of cytokines could produce a strong synergistic effect. IL-1ß induced the expression of CAMs on endothelial cells by activating NF-кB and JAK/STAT pathways. CapNO inhibited IL-1ß-stimulated NF-кB pathway by down-regulating IKK-α and inducing IкB-α directly. CapNO also inhibited JAK/STAT pathway by inhibiting JAK2 and STAT3 expressions. These effects bring about down-regulating CAMs expression on endothelial cells. These results suggest that CapNO may interrupt adhesion of cancer cells to endothelium by suppressing CAMs via inhibiting the NF-кB and JAK/STAT pathways in endothelial cells.