Cellular diversity in the brain revealed: The mammalian brain has an extraordinarily large number of cells. Although there are quite a few different cell types, many cells in any one category tend to look alike. Zeisel et al. analyzed the transcriptomes of mouse brain cells to reveal more than meets the eye. Interneurons of similar type were found in dissimilar regions of the brain. Oligodendrocytes that seemed to be all of one class were differentiated by their molecular signatures into a half-dozen classes. Microglia associated with blood vessels were distinguished from look-alike perivascular macrophages. Thus, the complex microanatomy of the brain can be revealed by the RNAs expressed in its cells. Science , this issue p. 1138

Tumors are composed of multiple cell types besides the tumor cells themselves, including innate immune cells such as macrophages. Tumor-associated macrophages (TAMs) are a heterogeneous population of myeloid cells present in the tumor microenvironment (TME). Here, they contribute to immunosuppression, enabling the establishment and persistence of solid tumors as well as metastatic dissemination. We have found that the pattern recognition scavenger receptor MARCO defines a subtype of suppressive TAMs and is linked to clinical outcome. An anti-MARCO monoclonal antibody was developed, which induces anti-tumor activity in breast and colon carcinoma, as well as in melanoma models through reprogramming TAM populations to a pro-inflammatory phenotype and increasing tumor immunogenicity. This anti-tumor activity is dependent on the inhibitory Fc-receptor, FcγRIIB, and also enhances the efficacy of checkpoint therapy. These results demonstrate that immunotherapies using antibodies designed to modify myeloid cells of the TME represent a promising mode of cancer treatment.

Previous studies have shown that the adaptor protein Shb is involved in receptor tyrosine kinase signaling. In this study, we demonstrate that Shb is phosphorylated in an Src-dependent manner upon vascular endothelial growth factor (VEGF) stimulation using porcine aortic endothelial cells expressing the human VEGF receptor 2 (VEGFR-2) (KDR). In co-immunoprecipitation experiments, we could detect an interaction between Shb and the VEGFR-2 in human telomerase-immortalized microvascular endothelial cells. Furthermore, in a glutathione S-transferase pull-down assay, the Src homology 2 domain of Shb was shown to interact with phosphorylated tyrosine 1175 in the C-terminal tail of VEGFR-2. VEGF-induced Shb phosphorylation was lost in porcine aortic endothelial cells expressing a chimeric murine VEGFR-2 (Flk-1) with a mutation at the corresponding position. Shb expression was specifically decreased by 80%, in a transient manner, by using the short interfering RNA technique. Reduced Shb expression led to a loss of stimulation of phosphatidylinositol 3-kinase, phosphorylation of focal adhesion kinase at tyrosine 576, the generation of focal adhesions, and stress fiber formation in response to VEGF. Furthermore, we show that VEGF-induced migration is inhibited in Shb short interfering RNA-treated cells. Our data demonstrate that Shb is important for VEGF signaling in endothelial cells. This is achieved by Shb binding to tyrosine 1175 in the VEGFR-2, which regulates VEGF-induced formation of focal adhesions and cell migration, of which the latter occurs in a phosphatidylinositol 3-kinase-dependent manner.

During infection, chemokines sequestered on endothelium induce recruitment of circulating leukocytes into the tissue where they chemotax along chemokine gradients toward the afflicted site. The aim of this in vivo study was to determine whether a chemokine gradient was formed intravascularly and influenced intraluminal neutrophil crawling and transmigration. A chemokine gradient was induced by placing a macrophage inflammatory protein-2 (MIP-2)–containing (CXCL2) gel on the cremaster muscle of anesthetized wild-type mice or heparanase-overexpressing transgenic mice (hpa-tg) with truncated heparan sulfate (HS) side chains. Neutrophil-endothelial interactions were visualized by intravital microscopy and chemokine gradients detected by confocal microscopy. Localized extravascular chemokine release (MIP-2 gel) induced directed neutrophil crawling along a chemotactic gradient immobilized on the endothelium and accelerated their recruitment into the target tissue compared with homogeneous extravascular chemokine concentration (MIP-2 superfusion). Endothelial chemokine sequestration occurred exclusively in venules and was HS-dependent, and neutrophils in hpa-tg mice exhibited random crawling. Despite similar numbers of adherent neutrophils in hpa-tg and wild-type mice, the altered crawling in hpa-tg mice was translated into decreased number of emigrated neutrophils and ultimately decreased the ability to clear bacterial infections. In conclusion, an intravascular chemokine gradient sequestered by endothelial HS effectively directs crawling leukocytes toward transmigration loci close to the infection site.