Bioequivalence decision for nanoparticular iron complex drugs for parenteral administration based on their disposition.

Although parenteral iron products have been established to medicinal use decades before, their structure and pharmacokinetic properties are not fully characterized yet. With its' second reflection paper on intravenous iron-based nano-colloidal products (EMA/CHMP/SWP/620008/2012) the European Medicine Agency provided an extensive catalogue of methods for quality, non-clinical and pharmacokinetic studies for the comparison of nano-sized iron products to an originator (EMA, 2015). For iron distribution studies, the reflection paper assumed the use of rodents.

Effects of sorafenib and cisplatin on preneoplastic foci of altered hepatocytes in fetal turkey liver.

Foci of altered hepatocytes (FAH) were induced in fetal turkey liver (FTL) by diethyl nitrosamine. FAH in FTL were resistant to iron overload similar to FAH in humans and rodents. The mitotic index was significantly higher in FAH (6.

Differences in tissue distribution of iron from various clinically used intravenous iron complexes in fetal avian heart and liver.

Nanomedicines are more complex than most pharmacologically active substances or medicines and have been considered as non-biological complex drugs. For nanomedicines pivotal pharmacokinetic properties cannot be assessed by plasma concentration data from standard bioequivalence studies. Using intravenous iron complexes (IICs) as model we show that fetal avian tissues can be used to study time dependent tissue concentrations in heart and liver.

Development of the mammalian axial skeleton requires signaling through the Gα(i) subfamily of heterotrimeric G proteins.

129/SvEv mice with a loss-of-function mutation in the heterotrimeric G protein α-subunit gene Gnai3 have fusions of ribs and lumbar vertebrae, indicating a requirement for Gα(i) (the "inhibitory" class of α-subunits) in somite derivatives. Mice with mutations of Gnai1 or Gnai2 have neither defect, but loss of both Gnai3 and one of the other two genes increases the number and severity of rib fusions without affecting the lumbar fusions. No myotome defects are observed in Gnai3/Gnai1 double-mutant embryos, and crosses with a conditional allele of Gnai2 indicate that Gα(i) is specifically required in cartilage precursors.

Comparative toxicity and cell-tissue distribution study on nanoparticular iron complexes using avian embryos and HepG2-cells.

In this study the toxicity and intracellular availability of iron from iron dextran (FeD), iron sucrose (FeS), and iron gluconate (FeG) was compared in organs of avian (turkey) embryos and in isolated cells (HepG2) in cell culture. Iron uptake was more pronounced in embryonic liver than in renal tissue. Cellular iron uptake in liver and kidney was more or less similar for the different compounds.

Differential regulation of lipopolysaccharide and Gram-positive bacteria induced cytokine and chemokine production in macrophages by Galpha(i) proteins.

Heterotrimeric G(i) proteins play a role in signalling activated by lipopolysaccharide (LPS), Staphylococcus aureus (SA) and group B streptococci (GBS), leading to production of inflammatory mediators. We hypothesized that genetic deletion of G(i) proteins would alter cytokine and chemokine production induced by LPS, SA and GBS stimulation. LPS-induced, heat-killed SA-induced and heat-killed GBS-induced cytokine and chemokine production in peritoneal macrophages from wild-type (WT), Galpha(i2) (-/-) or Galpha(i1/3) (-/-) mice were investigated.

Galphai2-mediated signaling events in the endothelium are involved in controlling leukocyte extravasation.

The trafficking of leukocytes from the blood to sites of inflammation is the cumulative result of receptor-ligand-mediated signaling events associated with the leukocytes themselves as well as with the underlying vascular endothelium. Our data show that Galpha(i) signaling pathways in the vascular endothelium regulate a critical step required for leukocyte diapedesis. In vivo studies using knockout mice demonstrated that a signaling event in a non-lymphohematopoietic compartment of the lung prevented the recruitment of proinflammatory leukocytes.

An obligatory requirement for the heterotrimeric G protein Gi3 in the antiautophagic action of insulin in the liver.

Heterotrimeric G proteins of the G(i) class have been implicated in signaling pathways regulating growth and metabolism under physiological and pathophysiological conditions. Knockout mice carrying inactivating mutations in both of the widely expressed Galpha(i) class genes, Galpha(i2) and Galpha(i3), demonstrate shared as well as gene-specific functions. The presence of a single active allele of Galpha(i3) is sufficient for embryonic development, whereas at least one allele of Galpha(i2) is required for extrauterine life.

Differential regulation of lipopolysaccharide and Gram-positive bacteria induced cytokine and chemokine production in splenocytes by Galphai proteins.

Heterotrimeric Gi proteins play a role in lipopolysaccharide (LPS) and Staphylococcus aureus (SA) activated signaling leading to inflammatory mediator production. We hypothesized that genetic deletion of Gi proteins would alter cytokine and chemokine production induced by LPS and SA. LPS- and heat killed SA-induced cytokine and chemokine production in splenocytes from wild type (WT), Galpha(i2) (-/-) or Galpha(i1/3) (-/-) mice were investigated.

Lipopolysaccharide- and gram-positive bacteria-induced cellular inflammatory responses: role of heterotrimeric Galpha(i) proteins.

Heterotrimeric G(i) proteins may play a role in lipopolysaccharide (LPS)-activated signaling through Toll-like receptor 4 (TLR4), leading to inflammatory mediator production. Although LPS is a TLR4 ligand, the gram-positive bacterium Staphylococcus aureus (SA) is a TLR2 ligand, and group B streptococci (GBS) are neither TLR2 nor TLR4 ligands but are MyD88 dependent. We hypothesized that genetic deletion of G(i) proteins would alter mediator production induced by LPS and gram-positive bacterial stimulation.

Macrophages induce the inflammatory response in the pulmonary Arthus reaction through G alpha i2 activation that controls C5aR and Fc receptor cooperation.

Complement and FcgammaR effector pathways are central triggers of immune inflammation; however, the exact mechanisms for their cooperation with effector cells and their nature remain elusive. In this study we show that in the lung Arthus reaction, the initial contact between immune complexes and alveolar macrophages (AM) results in plasma complement-independent C5a production that causes decreased levels of inhibitory FcgammaRIIB, increased levels of activating FcgammaRIII, and highly induced FcgammaR-mediated TNF-alpha and CXCR2 ligand production. Blockade of C5aR completely reversed such changes.

TCR-mediated hyper-responsiveness of autoimmune Galphai2(-/-) mice is an intrinsic naïve CD4(+) T cell disorder selective for the Galphai2 subunit.

Heterotrimeric Gi signaling regulates immune homeostasis, since autoimmunity occurs upon disruption of this pathway. However, the role of the lymphocyte-expressed Galphai subunits (Galphai2 and 3) on T cell activation and cytokine production is poorly understood. To examine this role, we studied T lymphocytes from mice deficient in the Galphai2 or Galphai3 subunits.

Noninvasive quantification of bowel inflammation through positron emission tomography imaging of 2-deoxy-2-[18F]fluoro-D-glucose-labeled white blood cells.

Treatment of inflammatory bowel disease (IBD) generally relies on long-term use of anti-inflammatory and immunosuppressive agents. The adverse effects of those drugs make it important to prescribe the minimal regimen that is effective. An objective method for noninvasively quantifying severity of bowel inflammation would thus be valuable in guiding inflammatory bowel disease therapy.

Functional G-protein heterotrimers are associated with vesicles of putative glutamatergic terminals: implications for regulation of transmitter uptake.

Changes in the vesicular transmitter content modulate synaptic strength and may contribute to synaptic plasticity. Several transporters mediating transmitter uptake into small synaptic vesicles (SSVs) have been identified but their regulation is largely unknown. Here we show by quantitative immunoelectron microscopy that the heterotrimeric G-protein subunits Galphao(2), Galpha(q/11), Gbeta(2), and Ggamma(7) are associated with vesicle-containing areas in terminals of cerebellar parallel fibers.

B cell developmental requirement for the G alpha i2 gene.

Null mutation of the Galphai2 trimeric G protein results in a discrete and profound mucosal disorder, including inflammatory bowel disease (IBD), attenuation of IL-10 expression, and immune function polarized to Th1 activity. Genetic and adoptive transfer experiments have established a role for B cells and IL-10 in mucosal immunologic homeostasis and IBD resistance. In this study, we addressed the hypothesis that Galphai2 is required for the development of IL-10-producing B cells.

Roles of G beta gamma in membrane recruitment and activation of p110 gamma/p101 phosphoinositide 3-kinase gamma.

Receptor-regulated class I phosphoinositide 3-kinases (PI3K) phosphorylate the membrane lipid phosphatidylinositol (PtdIns)-4,5-P2 to PtdIns-3,4,5-P3. This, in turn, recruits and activates cytosolic effectors with PtdIns-3,4,5-P3-binding pleckstrin homology (PH) domains, thereby controlling important cellular functions such as proliferation, survival, or chemotaxis. The class IB p110 gamma/p101 PI3K gamma is activated by G beta gamma on stimulation of G protein-coupled receptors.

Identification and characterization of the autophosphorylation sites of phosphoinositide 3-kinase isoforms beta and gamma.

Class I phosphoinositide 3-kinases (PI3Ks) are bifunctional enzymes possessing lipid kinase activity and the capacity to phosphorylate their catalytic and/or regulatory subunits. In this study, in vitro autophosphorylation of the G protein-sensitive p85-coupled class I(A) PI3K beta and p101-coupled class I(B) PI3K gamma was examined. Autophosphorylation sites of both PI3K isoforms were mapped to C-terminal serine residues of the catalytic p110 subunit (i.