Osteoporosis is a global public health problem affecting more than 200 million people worldwide. We previously showed that treatment with α-1 antitrypsin (AAT), a multifunctional protein with antiinflammatory properties, mitigated bone loss in an ovariectomized mouse model. However, the underlying mechanisms of the protective effect of AAT on bone tissue are largely unknown. In this study, we investigated the effect of AAT on osteoclast formation and function in vitro. Our results showed that AAT dose-dependently inhibited the formation of receptor activator of nuclear factor κB ligand (RANKL)-induced osteoclasts derived from mouse bone marrow macrophage/monocyte (BMM) lineage cells and the RAW 264.7 murine macrophage cell line. To elucidate the possible mechanisms underlying this inhibition, we tested the effect of AAT on the gene expression of cell surface molecules, transcription factors and cytokines associated with osteoclast formation. We showed that AAT inhibited macrophage colony-stimulating factor (M-CSF)-induced cell surface RANK expression in osteoclast precursor cells. In addition, AAT inhibited RANKL-induced TNF-α production, cell surface CD9 expression and dendritic cell-specific transmembrane protein (DC-STAMP) gene expression. Importantly, AAT treatment significantly inhibited osteoclast-associated mineral resorption. Together, these results uncover new mechanisms for the protective effects of AAT and strongly support the notion that AAT has therapeutic potential for the treatment of osteoporosis.

To be a successful pathogen, S. aureus has to adapt its metabolism to the typically oxygen- and glucose-limited environment of the host. Under fermenting conditions and in the presence of glucose, S. aureus uses glycolysis to generate ATP via substrate level phosphorylation and mainly lactic acid fermentation to maintain the redox balance by re-oxidation of NADH equivalents. However, it is less clear how S. aureus proceeds under anoxic conditions and glucose limitation, likely representing the bona-fide situation in the host. Using a combination of proteomic, transcriptional and metabolomic analyses, we show that in the absence of an abundant glycolysis substrate the available carbon source pyruvate is converted to acetyl-CoA (AcCoA) in a pyruvate formate-lyase (PflB)-dependent reaction to produce ATP and acetate. This process critically depends on de-repression of the catabolite control protein A (CcpA), leading to upregulation of pflB transcription. Under these conditions, ethanol production is repressed to prevent wasteful consumption of AcCoA. In addition, our global and quantitative characterization of the metabolic switch prioritizing acetate over lactate fermentation when glucose is absent illustrates examples of carbon source-dependent control of colonization and pathogenicity factors. Importance: Under infection conditions, S. aureus needs to ensure survival when energy production via oxidative phosphorylation is not possible, e.g. either due to the lack of terminal electron acceptors or by the inactivation of components of the respiratory chain. Under these conditions, S. aureus can switch to mixed acid fermentation to sustain ATP production by substrate-level phosphorylation. The drop in the cellular NAD+/NADH ratio is sensed by the repressor Rex, resulting in de-repression of fermentation genes. Here we show that expression of fermentation pathways is further controlled by CcpA in response to the availability of glucose to ensure optimal resource utilization under growth limiting conditions. We provide evidence for carbon source-dependent control of colonization and virulence factors. These findings add another level to the regulatory network controlling mixed acid fermentation in S. aureus and provide additional evidence for the lifestyle-modulating effect of carbon sources available in S. aureus.

Furan-2,5-dicarboxylic acid (FDCA) is a bio-based platform chemical with the potential to replace terephthalic acid in the production of polymers. A critical step for enzymatic and whole-cell production of FDCA from 5-(hydroxymethyl)­furfural (HMF) is the transformation of 5-(hydroxymethyl)­furoic acid (HMFA) into 5-formylfuroic acid (FFA). Here, we establish periplasmic pyrroloquinoline quinone (PQQ)-dependent alcohol dehydrogenases (ADHs) as biocatalytic tools for the oxidation of HMFA, HMF, and 5-formylfurfural (FFF). Further, we identify several amino acid residues including the “lid loop” of the substrate channel as promising targets for future engineering steps toward a fully periplasmic oxidation pathway to FDCA.