SummaryAging and obesity induce ectopic adipocyte accumulation in bone marrow cavities. This process is thought to impair osteogenic and hematopoietic regeneration. Here we specify the cellular identities of the adipogenic and osteogenic lineages of the bone. While aging impairs the osteogenic lineage, high-fat diet feeding activates expansion of the adipogenic lineage, an effect that is significantly enhanced in aged animals. We further describe a mesenchymal sub-population with stem cell-like characteristics that gives rise to both lineages and, at the same time, acts as a principal component of the hematopoietic niche by promoting competitive repopulation following lethal irradiation. Conversely, bone-resident cells committed to the adipocytic lineage inhibit hematopoiesis and bone healing, potentially by producing excessive amounts of Dipeptidyl peptidase-4, a protease that is a target of diabetes therapies. These studies delineate the molecular identity of the bone-resident adipocytic lineage, and they establish its involvement in age-dependent dysfunction of bone and hematopoietic regeneration.

Proteasome-catalyzed peptide splicing (PCPS) represents an additional activity of mammalian 20S proteasomes recently identified in connection with antigen presentation. We show here that PCPS is not restricted to mammalians but that it is also a feature of yeast 20S proteasomes catalyzed by all three active site ␤ subunits. No major differences in splicing efficiency exist between human 20S standard-and immuno-proteasome or yeast 20S proteasome. Using H 2 18 O to monitor the splicing reaction we also demonstrate that PCPS occurs via direct transpeptidation that slightly favors the generation of peptides spliced in cis over peptides spliced in trans. The 20S proteasome with its proteolytically active site ␤-subunits (␤1, ␤2, and ␤5) is a N-terminal nucleophilic hydrolase, widely conserved during evolution from yeast to mammals. It is the central proteolytic machinery of the ubiquitin proteasome system and the catalytic core of the 26S proteasome that is built by the association of 19S regulator complexes with the 20S proteasome. As part of the 26S proteasome the 20S core degrades poly-ubiquitylated proteins to peptides of 3 to 20 residues in length (1). A small percentage of these peptides is transported to the endoplasmic reticulum, bound by major histocompatibility complex (MHC) 1 class I molecules, and presented at the cell surface to CD8ϩ cytotoxic T lymphocyte for immune recognition. This antigen presentation pathway is usually restricted to the proteasome-dependent processing of self-and viral-proteins (2). Antigen presentation is generally increased after IFN-␥ stimuli because it induces, among others, the synthesis of alternative catalytic subunits (␤1i, ␤2i, and ␤5i) and the concomitant formation of immunoproteasomes (i-proteasomes) (2).All active ␤ subunits carry an N-terminal threonine residue as reactive nucleophile. Therefore, their distinct cleavage preferences are determined by the structural features of the substrate binding pockets. In particular, the nonprimed substrate binding site of the active site ␤ subunits binds the residues of the peptide substrate that are located at the N-terminal side of the cleaved residue. The residues of the peptide located C-terminally of the cleavage site are bound by the primed substrate binding site. The binding to both substrate binding sites of the active site ␤ subunit provides the stability and the orientation of the substrate, which is mandatory to carry out the proteolytic cleavage (3).Peptides can be produced by proteasomes during the degradation of proteins or polypeptides by conventional peptide bond hydrolysis or by proteasome-catalyzed peptide splicing (PCPS). The latter has been demonstrated in vivo so far only for four MHC class I-restricted epitopes (4 -8), leading to the assumption that PCPS is most likely a rare event that lacks any wider functional importance (9). PCPS was suggested to occur in a direct transpeptidation reaction, in either cis or trans, by linking two proteasomal cleavage products (PCPs) derived either from the same or from two ...

The role of dietary fibre and short-chain fatty acids (SCFA) in obesity development is controversially discussed. Here, we investigated how various types of dietary fibre and different SCFA ratios affect metabolic syndrome-related disorders. Male mice (B6) were fed high-fat diets supplemented with dietary fibres (either cellulose, inulin or guar gum) or different Ac:Pr ratios (high acetate (HAc) or propionate (HPr)) for 30 weeks. Body-fat gain and insulin resistance were greatly reduced by inulin, but not by guar gum, and completely prevented by SCFA supplementation. Only inulin and HAc increased body temperature, possibly by the induction of beige/browning markers in WAT. In addition, inulin and SCFA lowered hepatic triglycerides and improved insulin sensitivity. Both, inulin and HAc reduced hepatic fatty acid uptake, while only inulin enhanced mitochondrial capacity and only HAc suppressed lipogenesis in liver. Interestingly, HPr was accompanied by the induction of Nrg4 in BAT. Fermentable fibre supplementation increased the abundance of bifidobacteria; B. animalis was particularly stimulated by inulin and B. pseudolongum by guar gum. We conclude that in contrast to guar gum, inulin and SCFA prevent the onset of diet-induced weight gain and hepatic steatosis by different mechanisms on liver and adipose tissue metabolism.

Objective Aging is accompanied by loss of brown adipocytes and a decline in their thermogenic potential, which may exacerbate the development of adiposity and other metabolic disorders. Presently, only limited evidence exists describing the molecular alterations leading to impaired brown adipogenesis with aging and the contribution of these processes to changes of systemic energy metabolism. Methods Samples of young and aged murine brown and white adipose tissue were used to compare age-related changes of brown adipogenic gene expression and thermogenesis-related lipid mobilization. To identify potential markers of brown adipose tissue aging, non-targeted proteomic and metabolomic as well as targeted lipid analyses were conducted on young and aged tissue samples. Subsequently, the effects of several candidate lipid classes on brown adipocyte function were examined. Results Corroborating previous reports of reduced expression of uncoupling protein-1, we observe impaired signaling required for lipid mobilization in aged brown fat after adrenergic stimulation. Omics analyses additionally confirm the age-related impairment of lipid homeostasis and reveal the accumulation of specific lipid classes, including certain sphingolipids, ceramides, and dolichols in aged brown fat. While ceramides as well as enzymes of dolichol metabolism inhibit brown adipogenesis, inhibition of sphingosine 1-phosphate receptor 2 induces brown adipocyte differentiation. Conclusions Our functional analyses show that changes in specific lipid species, as observed during aging, may contribute to reduced thermogenic potential. They thus uncover potential biomarkers of aging as well as molecular mechanisms that could contribute to the degradation of brown adipocytes, thereby providing potential treatment strategies of age-related metabolic conditions.

Aged tissues usually show a decreased regenerative capacity accompanied by a decline in functionality. During aging pancreatic islets also undergo several morphological and metabolic changes. Besides proliferative and regenerative limitations, endocrine cells lose their secretory capacity, contributing to a decline in functional islet mass and a deregulated glucose homeostasis. This is linked to several features of aging, such as induction of cellular senescence or the formation of modified proteins, such as advanced glycation end products (AGEs) - the latter mainly examined in relation to hyperglycemia and in disease models. However, age-related changes of endocrine islets under normoglycemic and non-pathologic conditions are poorly investigated. Therefore, a characterization of pancreatic tissue sections as wells as plasma samples of wild-type mice (C57BL/6J) at various age groups (2.5, 5, 10, 15, 21 months) was performed. Our findings reveal that mice at older age are able to secret sufficient amounts of insulin to maintain normoglycemia. During aging the pancreatic islet area increased and the islet size doubled in 21 months old mice when compared to 2.5 months old mice, whereas the islet number was unchanged. This was accompanied by an age-dependent decrease in Ki-67 levels and pancreatic duodenal homeobox-1 (PDX-1), indicating a decline in proliferative and regenerative capacity of pancreatic islets with advancing age. In contrast, the number of p16Ink4a-positive nuclei within the islets was elevated starting from 10 months of age. Interestingly, AGEs accumulated exclusively in the islet blood vessels of old mice associated with increased amounts of inflammatory markers, such as the inducible nitric oxide synthase (iNOS) and 3-nitrotyrosine (3-NT). In summary, the age-related increase in islet size and area was associated with the induction of senescence, accompanied by an accumulation of non-enzymatically modified proteins in the islet vascular system.