Voltage gated inter-cation selective ion channels from graphene nanopores

With the ability to selectively control ionic flux, biological protein ion channels perform a fundamental role in many physiological processes. For practical applications that require the functionality of a biological ion channel, graphene provides a promising solid-state alternative, due to its atomic thinness and mechanical strength. Here, we demonstrate that nanopores introduced into graphene membranes, as large as 50 nm in diameter, exhibit inter-cation selectivity with a ~20x preference for K+ over divalent cations and can be modulated by an applied gate voltage. Liquid atomic force microscopy of the graphene devices reveals surface nanobubbles near the pore to be responsible for the observed selective behavior. Molecular dynamics simulations indicate that translocation of ions across the pore likely occurs via a thin water layer at the edge of the pore and the nanobubble. Our results demonstrate a significant improvement in the inter-cation selectivity displayed by a solid-state nanopore device and by utilizing the pores in a de-wetted state, offers an approach to fabricating selective graphene membranes that does not rely on the fabrication of sub-nm pores

Adipocyte-specific protein tyrosine phosphatase 1B deletion increases lipogenesis, adipocyte cell size and is a minor regulator of glucose homeostasis

Peer reviewedPublisher PD

Protein kinase activity of phosphoinositide 3-kinase regulates cytokine-dependent cell survival

Extent: 14 p.The dual specificity protein/lipid kinase, phosphoinositide 3-kinase (PI3K), promotes growth factor-mediated cell survival and is frequently deregulated in cancer. However, in contrast to canonical lipid-kinase functions, the role of PI3K protein kinase activity in regulating cell survival is unknown. We have employed a novel approach to purify and pharmacologically profile protein kinases from primary human acute myeloid leukemia (AML) cells that phosphorylate serine residues in the cytoplasmic portion of cytokine receptors to promote hemopoietic cell survival. We have isolated a kinase activity that is able to directly phosphorylate Ser585 in the cytoplasmic domain of the interleukin 3 (IL-3) and granulocyte macrophage colony stimulating factor (GM-CSF) receptors and shown it to be PI3K. Physiological concentrations of cytokine in the picomolar range were sufficient for activating the protein kinase activity of PI3K leading to Ser585 phosphorylation and hemopoietic cell survival but did not activate PI3K lipid kinase signaling or promote proliferation. Blockade of PI3K lipid signaling by expression of the pleckstrin homology of Akt1 had no significant impact on the ability of picomolar concentrations of cytokine to promote hemopoietic cell survival. Furthermore, inducible expression of a mutant form of PI3K that is defective in lipid kinase activity but retains protein kinase activity was able to promote Ser585 phosphorylation and hemopoietic cell survival in the absence of cytokine. Blockade of p110α by RNA interference or multiple independent PI3K inhibitors not only blocked Ser585 phosphorylation in cytokine-dependent cells and primary human AML blasts, but also resulted in a block in survival signaling and cell death. Our findings demonstrate a new role for the protein kinase activity of PI3K in phosphorylating the cytoplasmic tail of the GM-CSF and IL-3 receptors to selectively regulate cell survival highlighting the importance of targeting such pathways in cancer.Daniel Thomas, Jason A. Powell, Benjamin D. Green, Emma F. Barry, Yuefang Ma, Joanna Woodcock, Stephen Fitter, Andrew C.W. Zannettino, Stuart M. Pitson, Timothy P. Hughes, Angel F. Lopez, Peter R. Shepherd, Andrew H. Wei, Paul G. Ekert and Mark A. Guthridg

Inducible liver-specific knockdown of protein tyrosine phosphatase 1B improves glucose and lipid homeostasis in adult mice.

AIMS/HYPOTHESIS Protein tyrosine phosphatase 1B (PTP1B) is a key negative regulator of insulin signalling. Hepatic PTP1B deficiency, using the Alb-Cre promoter to drive Ptp1b deletion from birth in mice, improves glucose homeostasis, insulin sensitivity and lipid metabolism. The aim of this study was to investigate the therapeutic potential of decreasing liver PTP1B levels in obese and insulin-resistant adult mice. METHODS Inducible Ptp1b liver-specific knockout mice were generated using SA-Cre-ER(T2) mice crossed with Ptp1b floxed (Ptp1b(fl/fl)) mice. Mice were fed a high-fat diet (HFD) for 12 weeks to induce obesity and insulin resistance. Tamoxifen was administered in the HFD to induce liver-specific deletion of Ptp1b (SA-Ptp1b(-/-) mice). Body weight, glucose homeostasis, lipid homeostasis, serum adipokines, insulin signalling and endoplasmic reticulum (ER) stress were examined. RESULTS Despite no significant change in body weight relative to HFD-fed Ptp1b(fl/fl) control mice, HFD-fed SA-Ptp1b(-/-) mice exhibited a reversal of glucose intolerance as determined by improved glucose and pyruvate tolerance tests, decreased fed and fasting blood glucose and insulin levels, lower HOMA of insulin resistance, circulating leptin, serum and liver triacylglycerols, serum NEFA and decreased HFD-induced ER stress. This was associated with decreased glycogen synthase, eukaryotic translation initiation factor-2α kinase 3, eukaryotic initiation factor 2α and c-Jun NH2-terminal kinase 2 phosphorylation, and decreased expression of Pepck. CONCLUSIONS/INTERPRETATION Inducible liver-specific PTP1B knockdown reverses glucose intolerance and improves lipid homeostasis in HFD-fed obese and insulin-resistant adult mice. This suggests that knockdown of liver PTP1B in individuals who are already obese/insulin resistant may have relatively rapid, beneficial therapeutic effects

Synthesis and Characterization of Lanthanum Phosphate Nanoparticles as Carriers for ²²³Ra and ²²⁵Ra for Targeted Alpha Therapy

Introduction: Targeted alpha therapy (TAT) has the potential for killing micro-metastases with minimum collateral damage to surrounding healthy tissue. In vivo generator radionuclides, such as223Ra, 225Ra, and 225Ac, are of special interest for radiotherapeutic applications as they emit multiple α-particles during their decay. Utilizing appropriate carriers capable of retaining both the parent radioisotope as well as daughter products is important for the effective delivery of the radioisotope to the tumor site while mitigating global in vivo radiotoxicity. In this work, LaPO4 core and core + 2 shells nanoparticles (NPs) (NPs with 2 layers of cold LaPO4 deposited on the core surfaces) were synthesized containing either 223Ra or225Ra/225Ac, and the retention of the parents and daughters within the NPs in vitro was investigated. Methods: Core LaPO4 NPs were synthesized in aqueous solution by reacting 1 equivalent of La(NO3)3, along with few microcuries of either 223Ra or 225Ra/225Ac, with 1 equivalent of sodium tripolyphosphate (TPP) under moderate heating and purified by membrane dialysis. Core-shell NPs were also synthesized with one (core + 1 shell) and two (core + 2 shells) cold LaPO4 layers deposited onto the radioactive cores. The NPs were then characterized by transmission electron microscopy (TEM) and powder x-ray diffraction (XRD). Identification and quantification of radioactive parents and daughters released from the NPs in vitro were investigated using gamma-ray spectroscopy. Results: XRD and TEM analysis revealed that the NPs crystallized in the rhabdophane phase with mean diameters of 3.4 and 6.3nm for core and core + 2 shells, respectively. The core LaPO4 NPs retained up to 88% of 223Ra over 35days. However, in the core + 2 shells NPs, the retention of 223Ra and its daughter, 211Pb, was improved to \u3e 99.9% over 27days. Additionally, the retention of 225Ra/225Ac parents was \u3e 99.98% and ~80% for the 221Fr and 213Bi daughters over 35days for the core + 2 shells NPs. Conclusions: The in vitro retention of both parents and daughters results suggests that LaPO4 NPs are potentially effective carriers of radium isotopes