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1.
Angew Chem Int Ed Engl ; 62(52): e202306019, 2023 Dec 21.
Article in English | MEDLINE | ID: mdl-37610759

ABSTRACT

In this review the applications of isotopically labeled compounds are discussed and put into the context of their future impact in the life sciences. Especially discussing their use in the pharma and crop science industries to follow their fate in the environment, in vivo or in complex matrices to understand the potential harm of new chemical structures and to increase the safety of human society.


Subject(s)
Biological Science Disciplines , Humans , Research
2.
Toxicol Lett ; 330: 65-70, 2020 May 04.
Article in English | MEDLINE | ID: mdl-32380121

ABSTRACT

Metiram is a polymeric plant protection fungicidal product. Since farm workers can potentially be exposed to the used solo-formulation, polyram, its dermal penetration is important for the assessment of its safety. Previous dermal penetration studies indicated a low penetration of metiram (≤ 1% of the applied dose), when applied in polyram or in the almost identical technical concentrate, metiram TK. Here, we present an in vitro human skin absorption study conducted according to OECD guideline 428. In this study, synthesized polymeric 14C-radiolabelled metiram in polymeric "unlabeled" solo-formulation, polyram, was used. Single doses of radioactive metiram were applied to human skin preparations in vitro (4 donors per dose group, 2 replicates each) for 8 h, under semi-occluded conditions in Franz-like diffusion cells, using a flow-through diffusion system. Under these test conditions, dermal absorption of the low dose, which represents spray dilution concentrations, was 0.34 ± 0.48% of applied dose; and dermal absorption of the high dose (formulation concentrate) was negligible (<0.0015 % of the applied dose). These results confirm the low dermal absorption of polymeric metiram and indicate that slight differences in applied formulations have minimal impact on its penetration properties.

5.
Stem Cells Dev ; 17(2): 221-32, 2008 Apr.
Article in English | MEDLINE | ID: mdl-18447638

ABSTRACT

Recently, it has been shown that human unrestricted somatic stem cells (USSCs) from umbilical cord blood represent pluripotent, neonatal, nonhematopoietic stem cells with the potential to differentiate into the neural lineage. However, molecular and functional characterization of the neural phenotype and evaluation of the degree of maturity of the resulting cells are still lacking. In this study, we addressed the question of neuronal differentiation and maturation induced by a defined composition of growth and differentiation factors (XXL medium). We demonstrated the expression of different neuronal markers and their enrichment in USSC cultures during XXL medium incubation. Furthermore, we showed enrichment of USSCs expressing tyrosine hydroxylase (TH), an enzyme specific for dopaminergic neurons and other catecholamine-producing neurons, accompanied by induction of Nurr1, a factor regulating dopaminergic neurogenesis. The functionality of USSCs has been analyzed by patch-clamp recordings and high-performance liquid chromatography (HPLC). Voltage-gated sodium-channels could be identified in laminin-predifferentiated USSCs. In addition, HPLC analysis revealed synthesis and release of the neurotransmitter dopamine by USSC-derived cells, thus correlating well with the detection of TH transcripts and protein. This study provides novel insight into the potential of unrestricted somatic stem cells from human umbilical cord blood to acquire a neuronal phenotype and function.


Subject(s)
Cell Differentiation , Dopamine/metabolism , Fetal Blood/cytology , Neurons/physiology , Stem Cells/physiology , Animals , Cell Cycle/drug effects , Cells, Cultured , Culture Media/pharmacology , Humans , Neurons/metabolism , Phenotype , Rats , Sodium Channels/metabolism , Sodium Channels/physiology , Stem Cells/metabolism
6.
Cell Physiol Biochem ; 20(5): 397-404, 2007.
Article in English | MEDLINE | ID: mdl-17762167

ABSTRACT

We were interested whether PKC alpha, delta, epsilon or zeta is the isoform actually employed in the activation of hypertonicity-induced cation channels (HICCs) in primary cultures of rat hepatocytes. Quantitative SDS-page and Western-blot experiments revealed that PKC alpha, delta and epsilon were stimulated by Indolactam V (as a DAG substitute for activation of c and nPKCs) but that only PKC delta and epsilon did respond to hypertonic stress. Furthermore, chelation of intracellular Ca(++) by BAPTA-AM did not alter HICC activation in cable-analysis experiments whereas Indolactam V as well as V8 (an Indolactam derivative specific for PKC delta and epsilon) activated HICC currents under isotonic conditions. Finally, by use of Rottlerin (as an inhibitor exhibiting a slight preference for PKC delta over epsilon) PKC epsilon could be identified as the most likely isoform responsible for the activation of the HICC.


Subject(s)
Hepatocytes/metabolism , Ion Channels/metabolism , Protein Kinase C-epsilon/metabolism , Acetophenones/pharmacology , Animals , Benzopyrans/pharmacology , Cells, Cultured , Enzyme Activation , Hepatocytes/drug effects , Indoles/pharmacology , Isoenzymes/metabolism , Lactams/pharmacology , Rats
7.
Proc Natl Acad Sci U S A ; 103(28): 10606-11, 2006 Jul 11.
Article in English | MEDLINE | ID: mdl-16809424

ABSTRACT

Protein phosphatases have very recently emerged as important targets for chemical biology and medicinal chemistry research, and new phosphatase inhibitor classes are in high demand. The underlying frameworks of natural products represent the evolutionarily selected fractions of chemical space explored by nature so far and meet the criteria of relevance to nature and biological prevalidation most crucial to inhibitor development. We refer to synthesis efforts and compound collection development based on these criteria as biology-oriented synthesis. For the discovery of phosphatase inhibitor classes by means of this approach, four natural product-derived or -inspired medium-sized compound collections were synthesized and investigated for inhibition of the tyrosine phosphatases VE-PTP, Shp-2, PTP1B, MptpA, and MptpB and the dual-specificity phosphatases Cdc25A and VHR. The screen yielded four unprecedented and selective phosphatase inhibitor classes for four phosphatases with high hit rates. For VE-PTP and MptpB the first inhibitors were discovered. These results demonstrate that biology-oriented synthesis is an efficient approach to the discovery of new compound classes for medicinal chemistry and chemical biology research that opens up new opportunities for the study of phosphatases, which may lead to the development of new drug candidates.


Subject(s)
Biological Products/classification , Biological Products/chemical synthesis , Enzyme Inhibitors/classification , Enzyme Inhibitors/chemical synthesis , Phosphoprotein Phosphatases/antagonists & inhibitors , Phosphoprotein Phosphatases/chemistry , Biological Products/chemistry , Drug Design , Enzyme Inhibitors/chemistry , Humans
8.
J Med Chem ; 48(4): 1179-87, 2005 Feb 24.
Article in English | MEDLINE | ID: mdl-15715484

ABSTRACT

Multidrug resistance (MDR) is a major limiting factor in the development and application of drug candidates. MDR caused by MRP-1 is known to be modulated by the nonsteroidal antiinflammatory drug indomethacin. We have synthesized and biologically evaluated a library of indomethacin analogues. The indomethacin-derived compound library was synthesized employing the Fischer-indole synthesis as the key transformation and making use of a "resin-capture-release" strategy. Sixty representative members of the library were evaluated in a cell biological cytotoxicity assay employing the MRP-1 expressing human glioblastoma cell line T98G as a model system. Nine of the 60 tested derivatives increased the doxorubicin-mediated cytotoxicity at a comparable or higher level than indomethacin itself. Analysis of these derivatives revealed an interesting structure-function relationship. Most remarkably, two substances increased the toxicity, when doxorubicin was used at clinically relevant low concentrations, at a higher degree than indomethacin.


Subject(s)
Anti-Inflammatory Agents, Non-Steroidal/chemical synthesis , Drug Resistance, Multiple , Indomethacin/chemical synthesis , Multidrug Resistance-Associated Proteins/physiology , Anti-Inflammatory Agents, Non-Steroidal/chemistry , Anti-Inflammatory Agents, Non-Steroidal/pharmacology , Cell Line, Tumor , Combinatorial Chemistry Techniques , Doxorubicin/pharmacology , Drug Resistance, Neoplasm , Drug Synergism , Humans , Indomethacin/chemistry , Indomethacin/pharmacology , Reverse Transcriptase Polymerase Chain Reaction , Structure-Activity Relationship
9.
J Exp Med ; 200(2): 123-35, 2004 Jul 19.
Article in English | MEDLINE | ID: mdl-15263023

ABSTRACT

Here a new, intrinsically pluripotent, CD45-negative population from human cord blood, termed unrestricted somatic stem cells (USSCs) is described. This rare population grows adherently and can be expanded to 10(15) cells without losing pluripotency. In vitro USSCs showed homogeneous differentiation into osteoblasts, chondroblasts, adipocytes, and hematopoietic and neural cells including astrocytes and neurons that express neurofilament, sodium channel protein, and various neurotransmitter phenotypes. Stereotactic implantation of USSCs into intact adult rat brain revealed that human Tau-positive cells persisted for up to 3 mo and showed migratory activity and a typical neuron-like morphology. In vivo differentiation of USSCs along mesodermal and endodermal pathways was demonstrated in animal models. Bony reconstitution was observed after transplantation of USSC-loaded calcium phosphate cylinders in nude rat femurs. Chondrogenesis occurred after transplanting cell-loaded gelfoam sponges into nude mice. Transplantation of USSCs in a noninjury model, the preimmune fetal sheep, resulted in up to 5% human hematopoietic engraftment. More than 20% albumin-producing human parenchymal hepatic cells with absence of cell fusion and substantial numbers of human cardiomyocytes in both atria and ventricles of the sheep heart were detected many months after USSC transplantation. No tumor formation was observed in any of these animals.


Subject(s)
Cell Line , Fetal Blood/cytology , Placenta/blood supply , Stem Cells/cytology , Adipocytes/cytology , Albumins/metabolism , Animals , Blotting, Western , Bone and Bones/cytology , Cell Culture Techniques , Cell Differentiation , Cell Division , Cell Transplantation , Cord Blood Stem Cell Transplantation , Femur/metabolism , Flow Cytometry , Gene Expression Regulation, Developmental , Hematopoietic Stem Cells/cytology , Hippocampus/cytology , Humans , Immunophenotyping , Leukocyte Common Antigens/biosynthesis , Leukocytes, Mononuclear/metabolism , Myocardium/cytology , Myocytes, Cardiac/metabolism , Neurotransmitter Agents , Osteoblasts/metabolism , Phenotype , Polymerase Chain Reaction , Rats , Rats, Wistar , Reverse Transcriptase Polymerase Chain Reaction , Sheep , Time Factors , Umbilical Veins
10.
Brain Res ; 1018(1): 18-25, 2004 Aug 20.
Article in English | MEDLINE | ID: mdl-15262200

ABSTRACT

Understanding the structural and functional development of neurons in networks has a high impact to estimate the potentials for restorative therapies. Neurons derived from the human NT2 cell line (hNT) formed networks with a clustered neuritic architecture in vitro, whereas primary dissociated embryonic rat cortical neurons (Cx) displayed a more homogenous cell assembly. Spontaneous spikes of both cell types were recorded on microelectrode arrays within 2 weeks after seeding, but hNT showed a mostly uncorrelated firing pattern in contrast to Cx with highly synchronized bursting. hNT neurons were less sensitive to TTX (IC50 = 5.7 +/- 0.1 nM vs. IC50 = 1.1 +/- 0.2 nM), magnesium (IC50 = 1.83 +/- 0.01 mM vs. IC50 = 0.161 +/- 0.023 mM), and APV (IC50 > 100 microM vs. IC50 = 18 microM). We conclude that embryonic cortical neurons and hNT neurons have different network properties. This should be carefully considered before hNT neurons are used in therapeutic approaches, e g., central nervous system (CNS) grafting.


Subject(s)
Cell Differentiation/physiology , Nerve Net/physiology , Neurons/physiology , 2-Amino-5-phosphonovalerate/pharmacology , Action Potentials/drug effects , Action Potentials/physiology , Animals , Axons/metabolism , Axons/ultrastructure , Biomarkers , Brain Tissue Transplantation/methods , Cell Culture Techniques/methods , Cell Differentiation/drug effects , Cell Line, Tumor , Dose-Response Relationship, Drug , Fetus , Humans , Magnesium/pharmacology , Microelectrodes , Nerve Net/cytology , Nerve Net/drug effects , Nerve Tissue Proteins/metabolism , Neural Pathways/cytology , Neural Pathways/drug effects , Neural Pathways/physiology , Neurons/cytology , Neurons/drug effects , Rats , Teratocarcinoma , Tetrodotoxin/pharmacology
12.
Exp Neurol ; 184(1): 348-58, 2003 Nov.
Article in English | MEDLINE | ID: mdl-14637105

ABSTRACT

Adrenal chromaffin cells constitute one of the first cell types to have been defined as a neuroendocrine cell type. Since they produce dopamine, these cells have been proposed for the treatment of neuronal deficits in human Parkinson's disease. However, the factors involved in the development of chromaffin cells are still poorly understood. Based on recent insights from stem cell research, we decided to study the role of extracellular matrices, growth factors and neuropeptides on the neuroendocrine differentiation in a serum-free medium of PC12 cells. Employing immunohistochemistry, quantitative PCR and HPLC analysis, neuroendocrine differentiation was determined by evaluating neurite outgrowth, catecholamine biosynthesis and release as well as neuropeptide and vesicular protein mRNA expression. The combination of bFGF, NGF and PACAP could prevent the inhibition of neurite process development induced by dexamethasone in PC12 cells cultured on ECM. Whereas glucocorticoids were essential in the regulation of enzymes of catecholamine biosynthesis and metabolism, growth factors and PACAP were more efficient in inducing neuropeptide and chromogranin B expression as well as release of dopamine and 3-methoxytyramine. Therefore, in addition to glucocorticoids, chromaffin cells need a gradient of matrix, growth factors, and neuropeptides to develop the full functional phenotype of a neuroendocrine cell.


Subject(s)
Growth Substances/biosynthesis , Neuropeptides/biosynthesis , Neurosecretory Systems/physiology , Animals , Catecholamines/biosynthesis , Catecholamines/metabolism , Cell Differentiation/physiology , Cell Survival/physiology , Chromatography, High Pressure Liquid , Culture Media, Serum-Free , Dexamethasone/pharmacology , Extracellular Matrix/physiology , Immunohistochemistry , Nerve Tissue Proteins/biosynthesis , Neurites/physiology , Neuropeptides/pharmacology , Neurosecretory Systems/cytology , PC12 Cells , Pituitary Adenylate Cyclase-Activating Polypeptide , RNA, Messenger/biosynthesis , Rats , Reverse Transcriptase Polymerase Chain Reaction
13.
Chem Commun (Camb) ; (15): 1822-3, 2003 Aug 07.
Article in English | MEDLINE | ID: mdl-12931984

ABSTRACT

The Fischer indole synthesis using polymer-bound hydrazines is employed as the key step for the development of a traceless indole synthesis on a solid support.

14.
J Neurovirol ; 8(5): 411-9, 2002 Oct.
Article in English | MEDLINE | ID: mdl-12402167

ABSTRACT

The alpha chemokine receptor CXCR4 is used as the major coreceptor for the cell entry of T-cell-tropic human immunodeficiency virus-1 (HIV-1) isolates. Activation of this coreceptor by its natural ligand SDF1alpha is associated with an intracellular Ca(2+) increase. Because the HIV-1 glycoprotein 120 (gp120) is shedded from the surface of HIV-1-infected cells and is regarded as an injurious molecule in the pathogenesis of HIV-1-associated encephalopathy (HIVE), we investigated the effects of gp120 on the intracellular Ca(2+) regulation of astrocytes and neurons. After 5 days in vitro (DIV), SDF1alpha (50 nM) elicited a pertussis toxin-sensitive intracellular Ca(2+) increase due to Ca(2+) release from internal stores that was reduced by a blocking monoclonal antibody against the CXCR4 receptor in astrocytes and neurons. Parallel with the development of the SDF1alpha response, cells became sensitive to direct application of gp120 (1.25 microg/ml), which, similarly to SDF1alpha, elicited a transient intracellular Ca(2+) increase. However, short-term incubation with gp120 for 60 to 120 min induced a reduction of glutamate- or ATP-evoked intracellular Ca(2+) responses only in astrocytes and not in neurons, although functional CXCR4 receptors were expressed in both cell types. Therefore, our data strongly suggest that the CXCR4 receptor-mediated intracellular signaling pathway of gp120 differs in astrocytes and neurons.


Subject(s)
Astrocytes/metabolism , HIV Envelope Protein gp120/pharmacology , HIV-1 , Receptors, CXCR4/metabolism , Signal Transduction , AIDS Dementia Complex/physiopathology , Animals , Animals, Newborn , Astrocytes/drug effects , Calcium/metabolism , Cells, Cultured , Chemokine CXCL12 , Chemokines, CXC/pharmacology , Humans , Intracellular Membranes/metabolism , Neurons/drug effects , Neurons/metabolism , Rats , Rats, Wistar , Receptors, CXCR4/biosynthesis , Recombinant Proteins/pharmacology , Signal Transduction/drug effects , Stromal Cells/drug effects , Time Factors
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