Review: Molecular mechanism of microglia stimulated glioblastoma invasion

Glioblastoma multiforme is one of the deadliest human cancers and is characterized by a high degree of microglia and macrophage infiltration. The role of these glioma infiltrating macrophages (GIMs) in disease progression has been the subject of recent investigation. While initially thought to reflect an immune response to the tumor, the balance of evidence clearly suggests GIMs can have potent tumor-tropic functions and assist in glioma cell growth and infiltration into normal brain. In this review, we focus on the evidence for GIMs aiding mediating glioblastoma motility and invasion. We survey the literature for molecular pathways that are involved in paracrine interaction between glioma cells and GIMs and assess which of these might serve as attractive targets for therapeutic intervention.     

SARI suppresses colitis‐associated cancer development by maintaining MCP‐1‐mediated tumour‐associated macrophage recruitment

SARI (suppressor of AP‐1, regulated by IFN) impaired tumour growth by promoting apoptosis and inhibiting cell proliferation and tumour angiogenesis in various cancers. However, the role of SARI in regulating tumour‐associated inflammation microenvironment is still elusive. In our study, the colitis‐dependent and ‐independent primary model were established in SARI deficiency mice and immuno‐reconstructive mice to investigate the functional role of SARI in regulating tumour‐associated inflammation microenvironment and primary colon cancer formation. The results have shown that SARI deficiency promotes colitis‐associated cancer (CAC) development only in the presence of colon inflammation. SARI inhibited tumour‐associated macrophages (TAM) infiltration in colon tissues, and SARI deficiency in bone marrow cells has no observed role in the promotion of intestinal tumorigenesis. Mechanism investigations indicated that SARI down‐regulates p‐STAT1 and STAT1 expression in colon cancer cells, following inhibition of MCP‐1/CCR2 axis activation during CAC development. Inverse correlations between SARI expression and macrophage infiltration, MCP‐1 expression and p‐STAT1 expression were also demonstrated in colon malignant tissues. Collectively, our results prove the inhibition role of SARI in colon cancer formation through regulating TAM infiltration.     

Oesophageal squamous cell carcinoma–associated IL‐33 rewires macrophage polarization towards M2 via activating ornithine decarboxylase

BackgroundThe tumour microenvironment primarily constitutes macrophages in the form of an immunosuppressive M2 phenotype, which promotes tumour growth. Thus, the development of methodologies to rewire M2‐like tumour‐associated macrophages (TAMs) into the M1 phenotype, which inhibits tumour growth, might be a critical advancement in cancer immunotherapy research.MethodsThe expressions of IL‐33 and indicators related to macrophage polarization in oesophageal squamous cell carcinoma (ESCC) tissues and peripheral blood mononuclear cell (PBMC)–derived macrophages were determined. Inhibition of ornithine decarboxylase (ODC) with small interfering RNA was used to analyse the phenotype of macrophage polarization and polyamine secretory signals. CCK‐8, wound‐healing and Transwell assays were used to detect the proliferation and migration of ECA109 cells in vitro. The tumour xenograft assay in nude mice was used to examine the role of IL‐33 in ESCC development in vivo.ResultsThis study showed the substantially elevated IL‐33 expression in ESCC tissues compared with the normal tissues. Additionally, enhanced infiltration of M2‐like macrophages into the ESCC tumour tissue was also observed. We observed a strong correlation between the IL‐33 levels and the infiltration of M2‐like macrophages in ESCC tumours locally. Mechanistically, IL‐33 induces M2‐like macrophage polarization by activating ODC, a key enzyme that catalyses the synthesis of polyamines. Inhibition of ODC suppressed M2‐like macrophage polarization. Finally, in vivo, we confirmed that IL‐33 promotes tumour progression.ConclusionsThis study revealed an oncogenic role of IL‐33 by actively inducing M2‐like macrophage differentiation; thus, contributing to the formation of an immunosuppressive ESCC tumour microenvironment. Thus, IL‐33 could act as a novel target for cancer immunotherapies.     

TNIP1‐mediated TNF‐α/NF‐κB signalling cascade sustains glioma cell proliferation

As a malignant tumour of the central nervous system, glioma exhibits high incidence and poor prognosis. Although TNIP1 and the TNF‐α/NF‐κB axis play key roles in immune diseases and inflammatory responses, their relationship and role in glioma remain unknown. Here, we revealed high levels of TNIP1 and TNF‐α/NF‐κB in glioma tissue. Glioma cell proliferation was activated with TNF‐α treatment and showed extreme sensitivity to the TNF receptor antagonist. Furthermore, loss of TNIP1 disbanded the A20 complex responsible for IκB degradation and NF‐κB nucleus translocation, and consequently erased TNFα‐induced glioma cell proliferation. Thus, our investigation uncovered a vital function of the TNIP1‐mediated TNF‐α/NF‐κB axis in glioma cell proliferation and provides novel insight into glioma pathology and diagnosis.     

MicroRNA‐155‐3p promotes glioma progression and temozolomide resistance by targeting Six1

The prognosis of glioma is generally poor and is the cause of primary malignancy in the brain. The role of microRNAs has been implicated in tumour inhibition or activation. In several cancers, the Six1 signalling pathway has been found to be aberrant and also relates to the formation of tumours. We analysed the database for expression profiles and clinical specimens of various grades of glioma to assess microRNA‐155‐3p (miR‐155‐3p) expression. The role of miR‐155‐3p in glioblastoma, cell cycle, proliferation, apoptosis and resistance to temozolomide was assessed in vitro through flow cytometry and cell proliferation assays. Bioinformatics analyses, and assays using luciferase reporter, and immunoblotting revealed that miR‐155‐3p targets Six1 and that the relationship between glioma and healthy brain tissues was significantly inverse. In rescue experiments, overexpressed Six1 revoked the changes in cell cycle distribution, proliferation and resistance to temozolomide estimated by apoptosis induced by overexpressed miR‐155‐3p. MiR‐155‐3p inhibition reduced glioma cell growth and proliferation in the brain of a mouse model and increased the survival of mice with gliomas. Thus, miR‐155‐3p modulates Six1 expression and facilitates the progression of glioblastoma and resistance to temozolomide and may act as a novel diagnostic biomarker and a target for glioma treatment.     

Immunohistochemical analysis of glioma-infiltrating leucocytes after peripheral therapeutic immunization with interferon-γ-transfected glioma cells

We have shown previously that rejection of preinduced rat brain tumours is possible following therapeutic immunizations with interferon-γ (IFNγ)-transfected glioma cells (N32-IFNγ). In the present study we have used the same model to evaluate whether quantitative differences in tumour-infiltrating lymphocytes can be detected between animals receiving therapeutic immunizations with either IFNγ-transfected glioma cells, wild-type glioma cells or no treatment. Since leucocyte transpedesis into the tumour can be anticipated to depend on the state of vascularization, we have mapped the development of microvessels in the tumour in parallel with the leucocyte infiltration. Our results show that microvessels start to form at day 7 and then gradually increase in number and size, indicating the establishment of an extensive vascularization by day 24. Leucocyte infiltration displays a biphasic pattern after tumour grafting. We have therefore studied the infiltration kinetics after an early immunization (1 day after intracerebral isografting) and compared the effects with those of a late immunization (10 days after intracerebral isografting) with N32-IFNγ or wild-type N32. Our results show (1) an early infiltration of granulocytes 3 days after isografting; (2) a T-cell-receptor-positive (TCR⁺) T-cell infiltration starting on day 10; (3) a macrophage infiltration starting on day 13; (4) a CD8⁺ cell infiltration starting on day 13. The proportions of TCR⁺ T cells, CD8⁺ cells and natural killer cells differs significantly between animals immunized with N32-IFNγ and those receiving wild-type N32, when analysed 14 days after immunization at day 10. This difference can only be detected when animals are immunized at later stages of tumour growth. We propose that this could depend on an early-immunization-independent leucocyte infiltration during tumour establishment. This has to be considered when evaluating studies of leucocyte infiltration in experimental tumours. Received: 7 October 1999 / Accepted: 10 December 1999     

Dioscin elicits anti‐tumour immunity by inhibiting macrophage M2 polarization via JNK and STAT3 pathways in lung cancer

Tumour‐associated macrophage (TAM) is an important component in tumour microenvironment. Generally, TAM exhibits the function of M2‐like macrophage, which was closely related to angiogenesis and tumour progression. Dioscin, a natural steroidal saponin, has shown its powerful anti‐tumour activity recently. However, the mechanism of dioscin involved in immune regulation is still obscure. Here, we observed dioscin induced macrophage M2‐to‐M1 phenotype transition in vitro and inhibited IL‐10 secretion. Meanwhile, the phagocytosis of macrophages was enhanced. In subcutaneous lung tumour models, dioscin inhibited the augmentation of M2 macrophage populations. Furthermore, dioscin down‐regulated STAT3 and JNK signalling pathways in macrophages in vitro. In BMDMs, activating JNK and inhibiting STAT3 induce macrophages to M1 polarization while inhibiting JNK and activating STAT3 to M2 polarization. Additionally, condition mediums from dioscin‐pre‐treated macrophages inhibited the migration of 3LL cells and the tube‐formation capacity of HUVECs. What's more, dioscin‐mediated macrophage polarization inhibited the in vivo metastasis of 3LL cells. In conclusion, dioscin may act as a new anti‐tumour agent by inhibiting TAMs via JNK and STAT3 pathways in lung cancer.     

Bioinformatics analysis of the prognostic value of NEK8 and its effects on immune cell infiltration in glioma

Glioma is the most common malignancy of the nervous system with high rates of recurrence and mortality, even after surgery. The 5-year survival rate is only about 5%. NEK8 is involved in multiple biological processes in a variety of cancers; however, its role in glioma is still not clear. In the current study, we evaluated the prognostic value of NEK8, as well as its role in the pathogenesis of glioma. Using a bioinformatics approach and RNA-seq data from public databases, we found that NEK8 expression is elevated in glioma tissues; we further verified this result by RT-PCR, Western blotting and immunochemistry using clinical samples. Functional enrichment analyses of genes with correlated expression indicated that elevated NEK8 expression is associated with increased immune cell infiltration in glioma and may affect the tumour microenvironment via the regulation of DNA damage/repair. Survival analyses revealed that high levels of NEK8 are associated with a poorer prognosis; higher WHO grade, IDH status, 1p/19q codeletion, age and NEK8 were identified as an independent prognostic factor. These findings support the crucial role of NEK8 in the progression of glioma via effects on immune cell infiltration and suggest that it is a new prognostic biomarker.     

TGFbeta is responsible for skin tumour infiltration by macrophages enabling the tumours to escape immune destruction

Infiltration of skin tumours by macrophages is an important step in tumour progression, although the mechanisms of macrophage recruitment to the tumour mass and the subsequent effects on tumour growth are poorly understood. Transfecting a murine regressing skin tumour with the gene for transforming growth factor (TGF)beta enabled the tumours to grow progressively in vivo thus allowing us to study the role of this cytokine in tumour growth. Flow cytometry was used to show that TGFbeta-mediated tumour progression was accompanied by an increase in tumour-associated macrophages (TAM) and a decrease in tumour-infiltrating dendritic cells (DCs). TAM in TGFbeta-secreting tumours expressed lower levels of major histocompatibility complex II and CD86 compared to DC in control tumours and had a high phagocytic capacity as measured by uptake of latex beads in vivo. Indeed, TGFbeta was directly responsible not only for the enhanced macrophage phagocytosis but also altering the ratio of antigen-presenting cells to favour macrophages over DC. Our results demonstrate that TGFbeta recruitment and retention of macrophages at the tumour site enable effective tumour evasion of the host immune system and reinforces the need to target TGFbeta in human cancer immunotherapy trials.     

Inhibition of MCP-1/CCR2 pathway ameliorates the development of diabetic nephropathy

Monocyte chemoattractant protein (MCP-1) is an important mediator for macrophage recruitment in atherosclerosis and various glomerulonephritis. However, the role of MCP-1 and its receptor CCR2 in the progression of diabetic nephropathy remains unknown. Using a type 1 diabetic nephropathy model that shows noticeable glomerulosclerosis, we examined the role of MCP-1/CCR2 by propagermanium (Pro; CCR2 antagonist) treatment, and confirmed it by transfection of plasmids carrying the 7ND (a mutant of MCP-1) gene. We measured the mesangial matrix expansion, type IV collagen (Col4), transforming growth factor (TGF)-beta1 positive area, and macrophage infiltration in glomeruli after 12 weeks. Mesangial matrix expansion and macrophage infiltration were increased in diabetic mice and inhibited by Pro or 7ND-treatment. Increased glomerular expression of Col4 and TGF-beta1 in diabetic mice was also ameliorated. Thus blocking the MCP-1/CCR2 pathway ameliorated glomerulosclerosis, indicating that the MCP-1/CCR2 pathway plays a crucial role in the progression of diabetic nephropathy.     

Production of the Inaccessible Sesquiterpene (−)‐5‐Epieremophilene by Metabolically Engineered Escherichia coli

(?)‐5‐Epieremophilene, an epimer of the versatile sesquiterpene (+)‐valencene, is an inaccessible natural product catalyzed by three sesquiterpene synthases (SmSTPSs1‐3) of the Chinese medicinal herb Salvia miltiorrhiza, and its biological activity remains less explored. In this study, three metabolically engineered Escherichia coli strains were constructed for (?)‐5‐epieremophilene production with yields of 42.4–76.0 mg/L in shake‐flask culture. Introducing an additional copy of farnesyl diphosphate synthase (FDPS) gene through fusion expression of SmSTPS1‐FDPS or dividing the FDP synthetic pathway into two modules resulted in significantly improved production, and ultimately 250 mg of (?)‐5‐epieremophilene were achieved. Biological assay indicated that (?)‐5‐epieremophilene showed significant antifeedant activity against Helicoverpa armigera (EC₅₀=1.25 μg/cm²), a common pest of S. miltiorrhiza, implying its potential defensive role in the plant. The results provided an ideal material supply for studying other potential biological activities of (?)‐5‐epieremophilene, and also a strategy for manipulating terpene production in engineered E. coli using synthetic biology.     

Mathematical modelling of the use of macrophages as vehicles for drug delivery to hypoxic tumour sites

Poor drug delivery and low rates of cell proliferation are two factors associated with hypoxia that diminish the efficacy of many chemotherapeutic drugs. Since macrophages are known to migrate specifically towards, and localize within, hypoxic tumour regions, a promising resolution to these problems involves genetically engineering macrophages to perform such anti-tumour functions as inducing cell lysis and inhibiting angiogenesis. In this paper we outline a modelling approach to characterize macrophage infiltration into early avascular solid tumours, and extensions to study the interaction of these cells with macrophages already present within the tumour. We investigate the role of chemotaxis and chemokine production, and the efficacy of macrophages as vehicles for drug delivery to hypoxic tumour sites. The model is based upon a growing avascular tumour spheroid, in which volume is filled by tumour cells, macrophages and extracellular material, and tumour cell proliferation and death is regulated by nutrient diffusion. Crucially, macrophages occupy volume, and hence contribute to the volume balance and hence the size of the tumour. We also include oxygen-dependent production of macrophage chemokines, which can lead to accumulations in the hypoxic region of the tumour. We find that the macrophage chemotactic sensitivity is a key determinant of macrophage infiltration and tumour size. Although increased infiltration should be beneficial from the point of view of macrophage-based therapies, such infiltration in fact leads to increased tumour sizes. Finally, we include terms representing the induced death of tumour cells by hypoxic engineered macrophages. We demonstrate that reductions in tumour size can be achieved, but predict that a combination of therapies would be required for complete eradication. We also highlight some counter-intuitive predictions-for example, absolute and relative measures of tumour burden lead to different conclusions about prognosis. In summary, this paper illustrates how mathematical models may be used to investigate promising macrophage-based therapies.