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Yang Liu,1,* Wei Huang,2,* Chuanfeng Xiong,2 Yuxian Huang,2 Benny J Chen,2,3 Luigi Racioppi,2,4 Nelson Chao,2,3 Tuan Vo-Dinh1,3,5–61Department of Biomedical Engineering, Duke University, Durham, NC 27708, USA; 2Department of Medicine, Duke University Medical Center, Durham, NC 27710, USA; 3Duke Cancer Institute, Duke University Medical Center, Durham, NC 27710, USA; 4Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico II, Naples 80131, Italy; 5Fitzpatrick Institute for Photonics, Duke University, Durham, NC 27708, USA; 6Department of Chemistry, Duke University, Durham, NC 27708, USA*These authors contributed equally to this workAim: To quantitatively and sensitively investigate the biodistribution of immune cells after systemic administration.Methods: Immune cells were loaded with plasmonic gold nanostars (GNS) tracking probes. Inductively coupled plasma mass spectrometry (ICP-MS) was used for quantitative gold mass measurement and two-photon photoluminescence (TPL) was used for high-resolution sensitive optical imaging.Results: GNS nanoparticles were loaded successfully into immune cells without negative effect on cellular vitality. Liver and spleen were identified to be the major organs for macrophage cells uptake after systematic administration. A small amount of macrophage cells were detected in the tumor site in our murine lymphoma animal model.Conclusion: GNS has great potential as a biocompatible marker for quantitative tracking and high-resolution imaging of immune cells at the cellular level.Keywords: GNS, ICP-MS, two-photon microscopy, immune cells, biodistribution &nbsp

Bone remodeling, a physiological process characterized by bone formation by osteoblasts (OBs) and resorption of preexisting bone matrix by osteoclasts (OCs), is vital for the maintenance of healthy bone tissue in adult humans. Imbalances in this vital process result in pathological conditions including osteoporosis. Owing to its initial asymptomatic nature, osteoporosis is often detected only after the patient has sustained significant bone loss or a fracture. Hence, anabolic therapeutics that stimulate bone accrual is in high clinical demand. Here we identify Ca2þ/calmodulin (CaM)‐dependent protein kinase kinase 2 (CaMKK2) as a potential target for such therapeutics because its inhibition enhances OB differentiation and bone growth and suppresses OC differentiation. Mice null for CaMKK2 possess higher trabecular bone mass in their long bones, along with significantly more OBs and fewer multinuclear OCs. In vitro, although Camkk2/ mesenchymal stem cells (MSCs) yield significantly higher numbers of OBs, bone marrow cells from Camkk2/ mice produce fewer multinuclear OCs. Acute inhibition of CaMKK2 by its selective, cell‐permeable pharmacological inhibitor STO‐609 also results in increased OB and diminished OC formation. Further, we find phospho‐protein kinase A (PKA) and Ser133 phosphorylated form of cyclic adenosine monophosphate (cAMP) response element binding protein (pCREB) to be markedly elevated in OB progenitors deficient in CaMKK2. On the other hand, genetic ablation of CaMKK2 or its pharmacological inhibition in OC progenitors results in reduced pCREB as well as significantly reduced levels of its transcriptional target, nuclear factor of activated T cells, cytoplasmic (NFATc1). Moreover, in vivo administration of STO‐609 results in increased OBs and diminished OCs, conferring significant protection from ovariectomy (OVX)‐induced osteoporosis in adult mice. Overall, our findings reveal a novel function for CaMKK2 in bone remodeling and highlight the potential for its therapeutic inhibition as a valuable bone anabolic strategy that also inhibits OC differentiation in the treatment of osteoporosis. © 2013 American Society for Bone and Mineral Research.

Calcium/calmodulin-dependent kinase kinase 2 (CaMKK2) plays a key role in regulating food intake and energy expenditure at least in part by its actions in hypothalamic neurons. Previously, we showed that loss of CaMKK2 protected mice from high-fat diet (HFD)-induced obesity and glucose intolerance. However, although pair feeding HFD to WT mice to match food consumption of CAMKK2-null mice slowed weight gain, it failed to protect from glucose intolerance. Here we show that relative to WT mice, HFD-fed CaMKK2-null mice are protected from inflammation in adipose and remain glucose-tolerant. Moreover, loss of CaMKK2 also protected mice from endotoxin shock and fulminant hepatitis. We explored the expression of CaMKK2 in immune cells and found it to be restricted to those of the monocyte/macrophage lineage. CaMKK2-null macrophages exhibited a remarkable deficiency to spread, phagocytose bacteria, and synthesize cytokines in response to the Toll-like receptor 4 (TLR4) agonist lipopolysaccharide (LPS). Mechanistically, loss of CaMKK2 uncoupled the TLR4 cascade from activation of protein tyrosine kinase 2 (PYK2; also known as PTK2B). Our findings uncover an important function for CaMKK2 in mediating mechanisms that control the amplitude of macrophage inflammatory responses to excess nutrients or pathogen derivatives.

We read with special interest the article of Busiello and colleagues in Blood. [1][1] They described atypical features of familial hemophagocytic lymphohistiocytosis (FHLH) in a patient/family presenting 2 different perforin gene alterations: an already known homozygous A91V and a novel heterozygous

Bone remodeling, a physiological process characterized by bone formation by osteoblasts (OBs) and resorption of preexisting bone matrix by osteoclasts (OCs), is vital for the maintenance of healthy bone tissue in adult humans. Imbalances in this vital process result in pathological conditions including osteoporosis. Owing to its initial asymptomatic nature, osteoporosis is often detected only after the patient has sustained significant bone loss or a fracture. Hence, anabolic therapeutics that stimulate bone accrual is in high clinical demand. Here we identify Ca²⁺/calmodulin (CaM)-dependent protein kinase kinase 2 (CaMKK2) as a potential target for such therapeutics because its inhibition enhances OB differentiation and bone growth and suppresses OC differentiation. Mice null for CaMKK2 possess higher trabecular bone mass in their long bones, along with significantly more OBs and fewer multinuclear OCs. In vitro, although Camkk2⁻/⁻ mesenchymal stem cells (MSCs) yield significantly higher numbers of OBs, bone marrow cells from Camkk2⁻/⁻ mice produce fewer multinuclear OCs. Acute inhibition of CaMKK2 by its selective, cell-permeable pharmacological inhibitor STO-609 also results in increased OB and diminished OC formation. Further, we find phospho-protein kinase A (PKA) and Ser¹³³ phosphorylated form of cyclic adenosine monophosphate (cAMP) response element binding protein (pCREB) to be markedly elevated in OB progenitors deficient in CaMKK2. On the other hand, genetic ablation of CaMKK2 or its pharmacological inhibition in OC progenitors results in reduced pCREB as well as significantly reduced levels of its transcriptional target, nuclear factor of activated T cells, cytoplasmic (NFATc1). Moreover, in vivo administration of STO-609 results in increased OBs and diminished OCs, conferring significant protection from ovariectomy (OVX)-induced osteoporosis in adult mice. Overall, our findings reveal a novel function for CaMKK2 in bone remodeling and highlight the potential for its therapeutic inhibition as a valuable bone anabolic strategy that also inhibits OC differentiation in the treatment of osteoporosis.

Bone remodeling, a physiological process characterized by bone formation by osteoblasts (OBs) and resorption of preexisting bone matrix by osteoclasts (OCs), is vital for the maintenance of healthy bone tissue in adult humans. Imbalances in this vital process result in pathological conditions including osteoporosis. Owing to its initial asymptomatic nature, osteoporosis is often detected only after the patient has sustained significant bone loss or a fracture. Hence, anabolic therapeutics that stimulate bone accrual is in high clinical demand. Here we identify Ca²⁺/calmodulin (CaM)-dependent protein kinase kinase 2 (CaMKK2) as a potential target for such therapeutics because its inhibition enhances OB differentiation and bone growth and suppresses OC differentiation. Mice null for CaMKK2 possess higher trabecular bone mass in their long bones, along with significantly more OBs and fewer multinuclear OCs. In vitro, although Camkk2⁻/⁻ mesenchymal stem cells (MSCs) yield significantly higher numbers of OBs, bone marrow cells from Camkk2⁻/⁻ mice produce fewer multinuclear OCs. Acute inhibition of CaMKK2 by its selective, cell-permeable pharmacological inhibitor STO-609 also results in increased OB and diminished OC formation. Further, we find phospho-protein kinase A (PKA) and Ser¹³³ phosphorylated form of cyclic adenosine monophosphate (cAMP) response element binding protein (pCREB) to be markedly elevated in OB progenitors deficient in CaMKK2. On the other hand, genetic ablation of CaMKK2 or its pharmacological inhibition in OC progenitors results in reduced pCREB as well as significantly reduced levels of its transcriptional target, nuclear factor of activated T cells, cytoplasmic (NFATc1). Moreover, in vivo administration of STO-609 results in increased OBs and diminished OCs, conferring significant protection from ovariectomy (OVX)-induced osteoporosis in adult mice. Overall, our findings reveal a novel function for CaMKK2 in bone remodeling and highlight the potential for its therapeutic inhibition as a valuable bone anabolic strategy that also inhibits OC differentiation in the treatment of osteoporosis.

Transplantation to a DiGeorge syndrome affected patient is reported