Project description:Skeletal fragility is associated with type 2 diabetes mellitus (T2D), but the underlying mechanism is not well understood. Here, in a mouse model for youth-onset T2D, we show that both trabecular and cortical bone mass are reduced due to diminished osteoblast activity. Stable isotope tracing in vivo with 13C-glucose demonstrates that both glycolysis and glucose fueling of the TCA cycle are impaired in diabetic bones. Similarly, Seahorse assays show suppression of both glycolysis and oxidative phosphorylation by diabetes in bone marrow mesenchymal cells as a whole, whereas single-cell RNA sequencing reveals distinct modes of metabolic dysregulation among the subpopulations. Metformin not only promotes glycolysis and osteoblast differentiation in vitro, but also improves bone mass in diabetic mice. Finally, osteoblast-specific overexpression of either Hif1a, a general inducer of glycolysis, or Pfkfb3 which stimulates a specific step in glycolysis, averts bone loss in T2D mice. The study identifies osteoblast-intrinsic defects in glucose metabolism as an underlying cause of diabetic osteopenia, which may be targeted therapeutically.
Project description:Skeletal fragility is associated with type 2 diabetes mellitus (T2D), but the underlying mechanism is not well understood. Here, in a mouse model for youth-onset T2D, we show that both trabecular and cortical bone mass are reduced due to diminished osteoblast activity. Stable isotope tracing in vivo with 13 C-glucose demonstrates that both glycolysis and glucose fueling of the TCA cycle are impaired in diabetic bones. Similarly, Seahorse assays show suppression of both glycolysis and oxidative phosphorylation by diabetes in bone marrow mesenchymal cells as a whole, whereas single-cell RNA sequencing reveals distinct modes of metabolic dysregulation among the subpopulations. Metformin not only promotes glycolysis and osteoblast differentiation in vitro, but also improves bone mass in diabetic mice. Finally, targeted overexpression of Hif1a or Pfkfb3 in osteoblasts of T2D mice averts bone loss. The study identifies osteoblast-intrinsic defects in glucose metabolism as an underlying cause of diabetic osteopenia, which may be targeted therapeutically.
Project description:Skeletal fragility is associated with type 2 diabetes mellitus (T2D), but the underlying mechanism is not well understood. Here, in a mouse model for youth-onset T2D, we show that both trabecular and cortical bone mass is reduced due to diminished osteoblast activity. Stable isotope tracing in vivo with 13C-glucose demonstrates that both glycolysis and glucose fueling of the TCA cycle are impaired in diabetic bones. Similarly, Seahorse assays show suppression of both glycolysis and oxidative phosphorylation by diabetes in bone marrow mesenchymal cells as a whole, whereas single-cell RNA sequencing reveals distinct modes of metabolic dysregulation among the subpopulations. Metformin not only promotes glycolysis and osteoblast differentiation in vitro, but also improves bone mass in diabetic mice. Finally, osteoblast-specific overexpression of either Hif1a, a general inducer of glycolysis, or Pfkfb3 which stimulates a specific step in glycolysis, averts bone loss in T2D mice. The study identifies osteoblast-intrinsic defects in glucose metabolism as an underlying cause of diabetic osteopenia, which may be targeted therapeutically.
Project description:Neonates often generate incomplete immunity against intracellular pathogens, although the mechanism of this defect is poorly understood. An important question is whether the impaired development of memory CD8+ T cells in neonates is due to an immature priming environment or lymphocyte-intrinsic defects. Here we show that neonatal and adult CD8+ T cells adopted different fates when responding to equal amounts of stimulation in the same host. While adult CD8+ T cells differentiated into a heterogeneous pool of effector and memory cells, neonatal CD8+ T cells preferentially gave rise to short-lived effector cells and exhibited a distinct gene expression profile. Surprisingly, impaired neonatal memory formation was not due to a lack of responsiveness, but instead because neonatal CD8+ T cells expanded more rapidly than adult cells and quickly became terminally differentiated. Collectively, these findings demonstrate that neonatal CD8+ T cells exhibit an imbalance in effector and memory CD8+ T cell differentiation, which impairs the formation of memory CD8+ T cells in early life mRNA profiles of effector CD8+ T cells from neonatal and adult mice
Project description:Global expression profile of human osteoblast treated with melphalen, compared to human osteoblast cells treated with diluent control
Project description:Protein disulfide isomerase (PDI) is an oxidoreductase responsible for the formation, reduction and isomerization of disulfide bonds of nascent proteins in endoplasmic reticulum (ER). So far, the role of PDI in bone biology has never been characterized using genetically-modified animal models. In this study we generated osteoblast- specific PDI-deficient mice by crossing PDI-floxed (PDIfl/fl) mice with Osx-Cre mice. Compared with their littermate control PDIfl/fl mice, homozygous osteoblast-knockout mice (Osx-Cre/PDIfl/fl) were embryonically lethal, but heterozygous knockout mice (Osx-Cre/PDIfl/wt) displayed significantly pronounced growth retardation and reduced bone length. Besides, the decreases in bone density, osteoblast and osteoclast numbers, collagen fiber content and bone formation rate were observed in Osx-Cre/PDIfl/wt mice. Osteoblast precursors isolated from PDIfl/fl mice were infected with Cre recombinant adenovirus to produce PDI-deficient osteoblasts, followed by induction of differentiation. Osteoblasts deficient of PDI had decreased alkaline phosphatase activity, mineralizing capacity, and differentiation. Quantitative protein mass spectrometry analysis and immunoblotting showed that PDI deficiency markedly decreased the expression of the α-subunits of collagen prolyl 4-hydroxylase (C-P4H), including P4HA1, P4HA2 and P4HA3. These results demonstrate that PDI plays an essential role in osteoblast differentiation and bone formation and is required for the expression of the α-subunit of C-P4H in osteoblasts.
Project description:Global expression profile of human osteoblast treated with recombinant TGF-beta compared to human osteoblast treated with growth media alone