LOX-catalyzed collagen stabilization is a proximal cause for intrinsic resistance to chemotherapy.

The potential of altering the tumor ECM to improve drug response remains fairly unexplored. To identify targets for modification of the ECM aiming to improve drug response and overcome resistance, we analyzed expression data sets from pre-treatment patient cohorts. Cross-evaluation identified a subset of chemoresistant tumors characterized by increased expression of collagens and collagen-stabilizing enzymes. We demonstrate that strong collagen expression and stabilization sets off a vicious circle of self-propagating hypoxia, malignant signaling, and aberrant angiogenesis that can be broken by an appropriate auxiliary intervention: Interfering with collagen stabilization by inhibition of lysyl oxidases significantly enhanced response to chemotherapy in various tumor models, even in metastatic disease. Inhibition of collagen stabilization by itself can reduce or enhance tumor growth depending on the tumor type. The mechanistical basis for this behavior is the dependence of the individual tumor on nutritional supply on one hand and on high tissue stiffness for FAK signaling on the other.

Amino Acid Catabolism in Staphylococcus aureus and the Function of Carbon Catabolite Repression.

Staphylococcus aureus must rapidly adapt to a variety of carbon and nitrogen sources during invasion of a host. Within a staphylococcal abscess, preferred carbon sources such as glucose are limiting, suggesting that S. aureus survives through the catabolism of secondary carbon sources. S. aureus encodes pathways to catabolize multiple amino acids, including those that generate pyruvate, 2-oxoglutarate, and oxaloacetate. To assess amino acid catabolism, S. aureus JE2 and mutants were grown in complete defined medium containing 18 amino acids but lacking glucose (CDM). A mutation in the gudB gene, coding for glutamate dehydrogenase, which generates 2-oxoglutarate from glutamate, significantly reduced growth in CDM, suggesting that glutamate and those amino acids generating glutamate, particularly proline, serve as the major carbon source in this medium. Nuclear magnetic resonance (NMR) studies confirmed this supposition. Furthermore, a mutation in the ackA gene, coding for acetate kinase, also abrogated growth of JE2 in CDM, suggesting that ATP production from pyruvate-producing amino acids is also critical for growth. In addition, although a functional respiratory chain was absolutely required for growth, the oxygen consumption rate and intracellular ATP concentration were significantly lower during growth in CDM than during growth in glucose-containing media. Finally, transcriptional analyses demonstrated that expression levels of genes coding for the enzymes that synthesize glutamate from proline, arginine, and histidine are repressed by CcpA and carbon catabolite repression. These data show that pathways important for glutamate catabolism or ATP generation via Pta/AckA are important for growth in niches where glucose is not abundant, such as abscesses within skin and soft tissue infections.IMPORTANCES. aureus is a significant cause of both morbidity and mortality worldwide. This bacterium causes infections in a wide variety of organ systems, the most common being skin and soft tissue. Within a staphylococcal abscess, levels of glucose, a preferred carbon source, are limited due to the host immune response. Therefore, S. aureus must utilize other available carbon sources such as amino acids or peptides to proliferate. Our results show that glutamate and amino acids that serve as substrates for glutamate synthesis, particularly proline, function as major carbon sources during growth, whereas other amino acids that generate pyruvate are important for ATP synthesis via substrate-level phosphorylation in the Pta-AckA pathway. Our data support a model whereby certain amino acid catabolic pathways, and acquisition of those particular amino acids, are crucial for growth in niches where glucose is not abundant.

FcγRIIB on liver sinusoidal endothelial cells is essential for antibody-induced GPVI ectodomain shedding in mice.

The activating platelet collagen receptor glycoprotein VI (GPVI) is a promising antithrombotic target because of its central role in arterial thrombosis and its minor relevance for normal hemostasis. The receptor can be specifically targeted by antibodies and irreversibly downregulated in circulating platelets in vivo, resulting in long-term antithrombotic protection in mice. This GPVI immunodepletion predominantly occurs through ectodomain shedding, which is accompanied by a transient drop in peripheral platelet counts. Mechanistic studies on this targeted GPVI loss have been hampered because it cannot be reproduced in isolated platelets in vitro. Here we show that both the transient thrombocytopenia and GPVI ectodomain shedding depend on the Fc portion of the anti-GPVI antibody and its interaction with the inhibitory Fcγ receptor (FcγR)IIB. In wild-type, but not Fcgr2b(-/-) mice, anti-GPVI-opsonized platelets became transiently trapped in the liver followed by the appearance of the soluble GPVI ectodomain in the plasma. Depletion of Kupffer cells neither affected anti-GPVI-induced platelet accumulation nor GPVI shedding, demonstrating that the other major FcγRIIB-expressing cell type, liver sinusoidal endothelial cells, is required for both processes to occur. These results reveal a novel and unexpected function of hepatic FcγRIIB in the targeted downregulation of GPVI in vivo.