Standard chemotherapy of Glioblastoma multiforme (GBM) using temozolomide (TMZ) frequently fails due to acquired chemoresistance. Tumor-associated macrophages and microglia (TAMs) as major immune cell population in the tumor microenvironment are potential modulators of TMZ response. However; little is known about how TAMs participate in TMZ induced chemoresistance. Members of the metzincin superfamily such as Matrix Metalloproteases (MMPs) and A Disintegrin and Metalloprotease (ADAM) proteases are important mediators of cellular communication in the tumor microenvironment. A qPCR screening was performed to identify potential targets within the ADAM and MMP family members in GBM cells. In co-culture with macrophages ADAM8 was the only signature gene up-regulated in GBM cells induced by macrophages under TMZ treatment. The relationship between ADAM8 expression and TAM infiltration in GBM was determined in a patient cohort by qPCR; IF; and IHC staining and TCGA data analysis. Moreover; RNA-seq was carried out to identify the potential targets regulated by ADAM8. CCL2 expression levels were determined by qPCR; Western blot; IF; and ELISA. Utilizing qPCR; IF; and IHC staining; we observed a positive relationship between ADAM8 expression and TAMs infiltration level in GBM patient tissues. Furthermore; ADAM8 induced TAMs recruitment in vitro and in vivo. Mechanistically; we revealed that ADAM8 activated HB-EGF/EGFR signaling and subsequently up-regulated production of CCL2 in GBM cells in the presence of TMZ treatment; promoting TAMs recruitment; which further induced ADAM8 expression in GBM cells to mediate TMZ chemoresistance. Thus; we revealed an ADAM8 dependent positive feedback loop between TAMs and GBM cells under TMZ treatment which involves CCL2 and EGFR signaling to cause TMZ resistance in GBM.

TAF participated in the progression of various cancers, including PA via the release of soluble factors. Exosomes belonged to extracellular vesicles, which were revealed as a crucial participator in intercellular communication. However, the expression pattern and effect of TAF-derived exosomes remained largely unknown in PA. In the present study, we performed in silico analysis based on public RNA-seq datasets to generate the circRNA/miRNA regulatory network. The qRT-PCR, Western blotting, RNA pull-down, and luciferase assay were performed to investigate the effect of TAF-derived exosomes. TAF-derived exosomal circDennd1b was significantly upregulated in PA and promoted the proliferation, migration, and invasion of PA cells via sponging miR-145-5p in PA cells. In addition, miR-145-5p directly regulated One Cut homeobox 2 (ONECUT2/OC2) expression and inhibited the promoting effect of ONECUT2 on PA. We further demonstrated that ONECUT2 transcriptionally increased fibroblast growth factor receptor 3 (FGFR3) expression, which further activates the mitogen-activated protein kinases (MAPK) pathway, thus promoting PA progression. Moreover, the suppression of TAFs by ABT-263 and ONECUT2 by CSRM617 inhibited the growth of PA. In conclusion, our study illustrated that TAF-derived exosomal circDennd1b affected PA progression via regulating ONECUT2 expression, which provides a potential therapeutic strategy against aggressive PA.

Background: Pituitary adenoma (PA) bone invasion results in adverse outcomes, such as reduced rates of complete surgical resection and biochemical remission as well as increased recurrence rates, though few studies have been conducted. Methods: We collected clinical specimens of PAs for staining and statistical analysis. Evaluation of the ability of PA cells to induce monocyte–osteoclast differentiation by coculturing PA cells with RAW264.7 in vitro. An in vivo model of bone invasion was used to simulate the process of bone erosion and evaluate the effect of different interventions in alleviating bone invasion. Results: We found an overactivation of osteoclasts in bone-invasive PAs and concomitant aggregation of inflammatory factors. Furthermore, activation of PKCθ in PAs was established as a central signaling promoting PA bone invasion through the PKCθ/NF-κB/IL-1β pathway. By inhibiting PKCθ and blocking IL1β, we were able to significantly reverse bone invasion in an in vivo study. Meanwhile, we also found that celastrol, as a natural product, can obviously reduce the secretion of IL-1β as well as alleviate the progression of bone invasion. Conclusions: By activating the PKCθ/NF-κB/IL-1β pathway, pituitary tumors are able to induce monocyte–osteoclast differentiation in a paracrine manner and promote bone invasion, which can be alleviated by celastrol.

Objective: This study aimed to improve the antitumor immunocompetence of a tumor lysate-pulsed dendritic cell (DC) vaccine through differential centrifugation and provide a theoretical basis for its clinical application in glioblastoma. Methods: Peripheral blood mononuclear cells were extracted using Ficoll-Paque PLUS and induced into mature DCs in vitro with a cytokine cocktail. The modified tumor lysate was generated by differential centrifugation. The maturity markers of DCs in each group, namely the modified tumor lysate, tumor lysate, and negative and positive control groups, were assessed using flow cytometry. Furthermore, their ability to stimulate lymphocyte proliferation and in vitro antitumor effects were assessed using Cell Trace TM CFSE. IFN-γ secretion levels were measured with ELISA. Intracellular reactive oxygen species were measured using 2′,7′-dichlorofluorescein diacetate (DCFDA) staining. The results were statistically analyzed using an unpaired Student’s t-test and were considered significant at P < 0.05. Results: Compared with tumor lysate-pulsed DCs, modified tumor lysate-pulsed DCs had a higher expression of maturity markers: CD1a (7.38 ± 0.53% vs. 4.47 ± 0.75%) and CD83 (19.81 ± 4.09% vs. 9.64 ± 1.50%), were better capable of stimulating lymphocyte proliferation [proliferation index (PI): 8.54 ± 0.16 vs. 7.35 ± 0.05], secreting IFN-γ, and inducing stronger in-vitro cytotoxic T lymphocyte (CTL) cytotoxicity against glioblastoma cells. In addition, we found that the level of ROS in modified tumor lysate-pulsed DCs was lower than that in tumor lysate-pulsed DCs. Conclusion: Differential centrifugation of tumor lysates can improve the antitumor immunocompetence of DC vaccines, and reactive oxygen species may be the key to affecting DC function in the whole tumor lysate.