Metabolomic profiling of tumor-infiltrating macrophages during tumor growth.

Myeloid-derived suppressor cells (MDSCs) and tumor-associated macrophages (TAMs) are both key immunosuppressive cells that contribute to tumor growth. Metabolism and immunity of tumors depend on the tumor microenvironment (TME). However, the intracellular metabolism of MDSCs and TAMs during tumor growth remains unclear. Here, we characterized CD11b+ cells isolated from a tumor-bearing mouse model to compare intratumoral TAMs and intrasplenic MDSCs. Intratumoral CD11b+ cells and intrasplenic CD11b+ cells were isolated from tumor-bearing mice at early and late stages (14 and 28 days post-cell transplantation, respectively). The cell number of intrasplenic CD11b+ significantly increased with tumor growth. These cells included neutrophils holding segmented leukocytes or monocytes with an oval nucleus and Gr-1hi IL-4Rαhi cells without immunosuppressive function against CD8 T cells. Thus, these cells were classified as MDSC-like cells (MDSC-LCs). Intratumoral CD11b+ cells included macrophages with a round nucleus and were F4/80hi Gr-1lo IL-4Rαhi cells. Early stage intratumoral CD11b+ cells inhibited CD8 T cells via TNFα. Thus, this cell population was classified as TAMs. Metabolomic analyses of intratumoral TAMs and MDSC-LCs during tumor growth were conducted. Metabolic profiles of intratumoral TAMs showed larger changes in various metabolic pathways, e.g., glycolysis, TCA cycle, and glutamic acid pathways, during tumor growth compared with MDSL-LCs. Our findings demonstrated that intratumoral TAMs showed an immunosuppressive capacity from the early tumor stage and underwent intracellular metabolism changes during tumor growth. These results clarify the intracellular metabolism of TAMs during tumor growth and contribute to our understanding of tumor immunity.

Intrahepatic Biliary Metastasis of Colonic Adenocarcinoma: A Case Report With Immunohistochemical Analysis.

Although intrabiliary metastasis of carcinoma in the liver is unusual, intraductal and/or intraepithelial spread of cancer cells along intrahepatic bile ducts is now well recognized as hepatic metastasis. However, several clinical and laboratory findings, including images, lead us to differentially diagnose from primary intrahepatic cholangiocarcinoma. We report here on a case of colonic adenocarcinoma that metastasized to the liver with spread along with the intrahepatic bile duct of S5/6 area. The patient was a 51-year-old man and clinically diagnosed liver metastasis of sigmoid colon cancer (tub2, pMP, ly1, v0, n0), which was diagnosed and treated by sigmoidectomy 7 years ago. The right hepatic lobectomy was performed in March 2016 and histopathological examination revealed that moderately differentiated adenocarcinoma proliferated along the epithelium of intrahepatic bile ducts. Immunohistochemistry (IHC) showed that cancer cells in the intrahepatic bile ducts were positive for CK20, CDX2, CK17 and CK19, but negative for CK7, MUC-5AC, MUC-2 and CA19-9. The findings were almost the same as those of the sigmoid colon cancer removed in July 2009. We finally diagnosed the liver tumor as intrahepatic biliary metastasis of sigmoid colon cancer. Patients with liver metastasis of cancer are hard to be detected biliary invasion and spread on diagnostic image examination. Knowledge of distinctive morphological and IHC features can help to accurately diagnose this rare intrahepatic biliary metastasis of colonic cancer in routine pathological diagnostic procedures.

Combined GM-CSF treatment and M-CSF inhibition of tumor-associated macrophages induces dendritic cell-like signaling in vitro.

Macrophages demonstrate plasticity, and tumor-associated macrophages (TAM) can function as immunosuppressive cells in the tumor microenvironment. Therefore, in this study, we aimed to reprogram TAM in vitro with cytokine signal alteration. Granulocyte macrophage colony stimulating factor (GM-CSF) treatment alone did not lead to changes in the expression of M1 (including IL-1ß, TNFα and CXCL-10) or M2 (including CD36, CD206 and CCL17) molecules by TAM in vitro, although they adopted a round morphology and were less adhesive to the culture dish. When macrophage colony stimulating factor (M-CSF) signals were suppressed by siRNA against the M-CSF receptor (M-CSFR) in conjunction with GM-CSF treatment, the signal transduction pathway of TAM was altered, and the expression of STAT1, STAT5 and STAT6, which are usually expressed by dendritic cells, was increased. However, the same treatment did not alter the TAM expression pattern of M1/M2 marker molecules. With respect to the NF-κB pathway, GM-CSF and M-CSFR siRNA combination treatment significantly induced the expression of p65, which is usually not expressed by TAM, while p50 and p105 expression by TAM was not affected by the treatment. These findings indicate that our model could not redirect TAM to a monocyte-derived dendritic cell-like phenotype based on the analysis of M1/M2 marker expression, but it was able to modify cell signaling pathways toward a dendritic cell-like pattern. Therefore, the present data suggest that TAM demonstrate plasticity toward dendritic cell-like signal transduction patterns, and that the alteration of the tumor microenvironment has the potential to reverse the immunosuppressive properties of TAM.

Tumor-infiltrating myeloid-derived suppressor cells are pleiotropic-inflamed monocytes/macrophages that bear M1- and M2-type characteristics.

Here, tumor-infiltrating CD11b(+) myelomonocytoid cells in murine colon adenocarcinoma-38 and GL261 murine glioma were phenotypically characterized. Over 90% were of the CD11b(+)F4/80(+) monocyte/macrophage lineage. They also had a myeloid-derived suppressor cell (MDSC) phenotype, as they suppressed the proliferation of activated splenic CD8(+) T cells and had a CD11b(+)CD11c(+)Gr-1(low)IL-4Ralpha(+) phenotype. In addition, the cells expressed CX(3)CR1 and CCR2 simultaneously, which are the markers of an inflammatory monocyte. The MDSCs expressed CD206, CXCL10, IL-1beta, and TNF-alpha mRNAs. They also simultaneously expressed CXCL10 and CD206 proteins, which are typical, classical (M1) and alternative (M2) macrophage activation markers, respectively. Peritoneal exudate cells (PECs) strongly expressed CD36, CD206, and TGF-beta mRNA, which is characteristic of deactivated monocytes. The MDSCs also secreted TGF-beta, and in vitro culture of MDSCs and PECs with anti-TGF-beta antibody recovered their ability to secrete NO. However, as a result of secretion of proinflammatory cytokines, MDSCs could not be categorized into deactivated monocyte/macrophages. Thus, tumor-infiltrating MDSCs bear pleiotropic characteristics of M1 and M2 monocytes/macrophages. Furthermore, CD206 expression by tumor-infiltrating MDSCs appears to be regulated by an autocrine mechanism that involves TGF-beta.