Pigment Epithelium-Derived Factor Blocks Tumor Extravasation by Suppressing Amoeboid Morphology and Mesenchymal Proteolysis

Metastatic melanoma cells are highly adaptable to their in vivo microenvironment and can switch between protease-dependent mesenchymal and protease-independent amoeboid invasion to facilitate metastasis. Such adaptability can be visualized in vitro, when cells are cultured in conditions that recapitulate three-dimensional microenvironments. Using thick collagen layers in cell culture and in vivo extravasation assays, we found that pigment epithelium-derived factor (PEDF) suppressed lung extravasation of aggressive melanoma by coordinated regulation of cell shape and proteolysis. In cells grown on a thick collagen bed, PEDF overexpression and exogenous PEDF blocked the rapidly invasive, rounded morphology, and promoted an elongated, mesenchymal-like phenotype associated with reduced invasion. These changes in cell shape depended on decreased RhoA and increased Rac1 activation and were mediated by the up-regulation of Rac1-GEF, DOCK3 and down-regulation of Rac1-GAP, ARHGAP22. Surprisingly, we found that PEDF overexpression also blocked the trafficking of membrane-tethered, MT1-MMP to the cell surface through RhoA inhibition and Rac1 activation. In vivo, knockdown of Rac1 and DOCK3 or overexpression of MT1-MMP was sufficient to reverse the inhibitory effect of PEDF on extravasation. Using functional studies, we demonstrated that PEDF suppressed the rounded morphology and MT1-MMP surface localization through its antiangiongenic, 34-mer epitope and the recently identified PEDF receptor candidate, PNPLA2. Our findings unveil the coordinated regulation of cell shape and proteolysis and identify an unknown mechanism for PEDF''s antimetastatic activity.     

The Chemokine CCL2 Increases Prostate Tumor Growth and Bone Metastasis through Macrophage and Osteoclast Recruitment

CC chemokine ligand 2 (CCL2, also known as monocyte chemoattractant protein-1) has been demonstrated to recruit monocytes to tumor sites. Monocytes are capable of being differentiated into tumor-associated macrophages (TAMs) and osteoclasts (OCs). TAMs have been shown to promote tumor growth in several cancer types. Osteoclasts have also been known to play an important role in cancer bone metastasis. To investigate the effects of CCL2 on tumorigenesis and its potential effects on bone metastasis of human prostate cancer, CCL2 was overexpressed into a luciferase-tagged human prostate cancer cell line PC-3. In vitro, the conditioned medium of CCL2 overexpressing PC-3luc cells (PC-3^lucCCL2) was a potent chemoattractant for mouse monocytes in comparison to a conditioned medium from PC-3^lucMock. In addition, CCL2 overexpression increased the growth of transplanted xenografts and increased the accumulation of macrophages in vivo. In a tumor dissemination model, PC-3^lucCCL2 enhanced the growth of bone metastasis, which was associated with more functional OCs. Neutralizing antibodies targeting both human and mouse CCL2 inhibited the growth of PC-3^luc, which was accompanied by a decrease in macrophage recruitment to the tumor. These findings suggest that CCL2 increases tumor growth and bone metastasis through recruitment of macrophages and OCs to the tumor site.     

肿瘤微环境与前列腺癌骨转移研究进展

前列腺癌（prostate cancer, PCa）转移中大约80%是发生骨转移,是造成患者死亡的主要原因。前列腺癌骨转移主要呈现成骨和破骨混合性损伤。然而,诱发肿瘤细胞转移到骨以及引起混合样病变的机制仍不十分清楚。宿主微环境与前列腺癌细胞间的相互作用是其中一个重要原因。本文主要讨论宿主微环境如何影响前列腺癌骨转移各个阶段的发生及其中的机制,探讨多种细胞因子、分泌蛋白、微环境中的免疫细胞、成骨细胞、破骨细胞等分泌的因子在前列腺癌骨转移中的重要作用。     

Induction of Tumor Necrosis Factor in Mice by Recombinant Human Macrophage Colony-stimulating Factor

The effect of recombinant human macrophage colony-stimulating factor (rhM-CSF) on endogenous production of tumor necrosis factor (TNF) was investigated in mice. The intravenous injection of lipopolysaccharide (LPS) after the administration of rhM-CSF via the same route induced the production of endogenous cytotoxic activity in serum as assessed by using TNF-sensitive miirine L929 cells. The intravenous injection of LPS alone or rhM-CSF alone did not induce cytotoxic activity. The priming effect of rhM-CSF was transient and the optimal duration between injections of primer and trigger was 3 h, while the optimal duration between trigger injection and serum sampling was 1 h after LPS injection. Moreover, preinjection of rhM-CSF enhanced the priming effect of recombinant mouse interferon-γ. No triggering effect of rhM-CSF was observed. The cytotoxic activity in the serum was completely neutralized by anti-mouse TNF-α polyclonal antibody. These results indicate that rhM-CSF can be used as a priming agent for endogenous production of TNF in vivo , and raise the possibility of using rhM-CSF in cancer immunotherapy.     

Rat Prostate Tumor Cells Progress in the Bone Microenvironment to a Highly Aggressive Phenotype

Prostate cancer generally metastasizes to bone, and most patients have tumor cells in their bone marrow already at diagnosis. Tumor cells at the metastatic site may therefore progress in parallel with those in the primary tumor. Androgen deprivation therapy is often the first-line treatment for clinically detectable prostate cancer bone metastases. Although the treatment is effective, most metastases progress to a castration-resistant and lethal state. To examine metastatic progression in the bone microenvironment, we implanted androgen-sensitive, androgen receptor–positive, and relatively slow-growing Dunning G (G) rat prostate tumor cells into the tibial bone marrow of fully immune-competent Copenhagen rats. We show that tumor establishment in the bone marrow was reduced compared with the prostate, and whereas androgen deprivation did not affect tumor establishment or growth in the bone, this was markedly reduced in the prostate. Moreover, we found that, with time, G tumor cells in the bone microenvironment progress to a more aggressive phenotype with increased growth rate, reduced androgen sensitivity, and increased metastatic capacity. Tumor cells in the bone marrow encounter lower androgen levels and a higher degree of hypoxia than at the primary site, which may cause high selective pressures and eventually contribute to the development of a new and highly aggressive tumor cell phenotype. It is therefore important to specifically study progression in bone metastases. This tumor model could be used to increase our understanding of how tumor cells adapt in the bone microenvironment and may subsequently improve therapy strategies for prostate metastases in bone.Abbreviations: PC, prostate cancer; DTCs, disseminated tumor cells; BrdU, bromodeoxyuridine; DHT, dihydrotestosterone; T, testosterone; FBS, fetal bovine serum; AR, androgen receptor; CSC, cancer stem cells     

Tumor Cells Modulate Macrophage Phenotype in a Novel In Vitro Co-Culture Model of the NSCLC Tumor Microenvironment

IntroductionMacrophage phenotype in the tumor microenvironment correlates with prognosis in NSCLC. Immunosuppressive macrophages promote tumor progression, whereas proinflammatory macrophages may drive an antitumor immune response. How individual NSCLCs affect macrophage phenotype is a major knowledge gap.MethodsTo systematically study the impact of lung cancer cells on macrophage phenotypes, we developed an in vitro co-culture model that consisted of molecularly and clinically annotated patient-derived NSCLC lines, human cancer-associated fibroblasts, and murine macrophages. Induced macrophage phenotype was studied through quantitative real-time polymerase chain reaction and validated in vivo using NSCLC xenografts through quantitative immunohistochemistry and clinically with The Cancer Genome Atlas (TCGA)–“matched” patient tumors.ResultsA total of 72 NSCLC cell lines were studied. The most frequent highly induced macrophage-related gene was Arginase-1, reflecting an immunosuppressive M2-like phenotype. This was independent of multiple clinicopathologic factors, which also did not affect M2:M1 ratios in matched TCGA samples. In vivo, xenograft tumors established from high Arginase-1–inducing lines (Arg^hi) had a significantly elevated density of Arg1+ macrophages. Matched TCGA clinical samples to Arg^hi NSCLC lines had a significantly higher ratio of M2:M1 macrophages (p = 0.0361).ConclusionsIn our in vitro co-culture model, a large panel of patient-derived NSCLC lines most frequently induced high-expression Arginase-1 in co-cultured mouse macrophages, independent of major clinicopathologic and oncogenotype-related factors. Arg^hi cluster-matched TCGA tumors contained a higher ratio of M2:M1 macrophages. Thus, this in vitro model reproducibly characterizes how individual NSCLC modulates macrophage phenotype, correlates with macrophage polarization in clinical samples, and can serve as an accessible platform for further investigation of macrophage-specific therapeutic strategies.     

Androgen Receptor Is a Tumor Suppressor and Proliferator in Prostate Cancer

Targeting androgens via androgen deprivation therapy (ADT) to suppress androgens/androgen receptor (AR) functions remains the standard treatment for prostate cancer. However, most tumors eventually recur in     

Dendrosomal Curcumin Suppresses Metastatic Breast Cancer in Mice by Changing M1/M2 Macrophage Balance in the Tumor Microenvironment

Curcumin, a lipid-soluble compound extracted from the plant Curcuma Longa, has been found to exertimmunomodulatory effects via macrophages. However, most studies focus on the low bioavailability issue ofcurcumin by nano and microparticles, and thus the role of macrophages in the anticancer mechanism of curcuminhas received little attention so far. We have previously shown the potential biocompatibility, biodegradabilityand anti-cancer effects of dendrosomal curcumin (DNC). In this study, twenty-seven BALB/c mice were equallydivided into control as well as 40 and 80 mg/kg groups of DNC to investigate the involvement of macrophagesin the antitumor effects of curcumin in a typical animal model of metastatic breast cancer. At the end ofintervention, the tumor volume and weight were significantly reduced in DNC groups compared to control(P<0.05). Histopathological data showed the presence of macrophages in tumor and spleen tissues. Real-timePCR results showed that DNC increased the expression of STAT4 and IL-12 genes in tumor and spleen tissues incomparison with control (P<0.05), referring to the high levels of M1 macrophages. Furthermore treatment withDNC decreased STAT3, IL-10 and arginase I gene expression (P<0.05), indicating low levels of M2 macrophage.The results confirm the role of macrophages in the protective effects of dendrosomal curcumin against metastaticbreast cancer in mice.     

Mammary Fibroblast-derived Hepatocyte Growth Factor Stimulates Growth and Morphogenesis of Mouse Mammary Tumor Cells in Primary Culture

We have recently isolated a mammary growth factor from the conditioned medium of mouse mammary stromal fibroblasts and identified it as a mouse homologue of human HGF (hepatocyte growth factor). To elucidate the role of HGF in mouse mammary tumorigenesis, we produced recombinant mouse HGF and examined its effects on primary cultures of mouse mammary tumor cells in this study. HGF at concentrations above 20 ng/ml maximally stimulated the growth of mammary tumor cells in primary monolayer culture. HGF also stimulated the three-dimensional growth and branching morphogenesis of mammary tumor cells cultured inside collagen gels. A comparison of the growth-stimulating activity of HGF with that of EGF (epidermal growth factor) and KGF (keratinocyte growth factor) revealed that HGF is the most potent growth factor among the three. Immunological studies using an antibody against mouse HGF demonstrated that 74% of the growth-stimulating activity present in the mammary fibroblast-conditioned medium was abolished by the antibody, indicating that HGF is the major growth factor produced by the fibroblasts. These observations thus suggest a role for HGF as a mammary stromal fibroblast-derived factor which stimulates growth and morphogenesis of adjacent mammary tumor cells in vivo .     

肿瘤微环境与中性粒细胞相互作用关系的研究进展

摘 要：肿瘤微环境能够通过细胞因子招募大量中性粒细胞聚集到肿瘤局部,并使其极化成为肿瘤相关中性粒细胞,而肿瘤相关中性粒细胞又能重塑肿瘤微环境的组成,对肿瘤的发生、发展中起着重要作用。肿瘤微环境与中性粒细胞存在着相互影响的作用关系,文章就肿瘤微环境和中性粒细胞的相互关系作用及其机制的研究进展进行综述。     

CD36 and Its Role in Regulating the Tumor Microenvironment

CD36 is a transmembrane glycoprotein that binds to a wide range of ligands, including fatty acids (FAs), cholesterol, thrombospondin-1 (TSP-1) and thrombospondin-2 (TSP-2), and plays an important role in lipid metabolism, immune response, and angiogenesis. Recent studies have highlighted the role of CD36 in mediating lipid uptake by tumor-associated immune cells and in promoting tumor cell progression. In cancer-associated fibroblasts (CAFs), CD36 regulates lipid uptake and matrix protein production to promote tumor proliferation. In addition, CD36 can promote tumor cell adhesion to the extracellular matrix (ECM) and induce epithelial mesenchymal transition (EMT). In terms of tumor angiogenesis, CD36 binding to TSP-1 and TSP-2 can both inhibit tumor angiogenesis and promote tumor migration and invasion. CD36 can promote tumor angiogenesis through vascular mimicry (VM). Overall, we found that CD36 exhibits diverse functions in tumors. Here, we summarize the recent research findings highlighting the novel roles of CD36 in the context of tumors.