The Experts below are selected from a list of 309 Experts worldwide ranked by ideXlab platform
Raz Krizevski - One of the best experts on this subject based on the ideXlab platform.
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benzaldehyde is a precursor of phenylpropylamino alkaloids as revealed by targeted metabolic profiling and comparative biochemical analyses in ephedra spp
Phytochemistry, 2012Co-Authors: Raz Krizevski, Asaf Levy, Einat Bar, Or Shalit, Jillian M Hagel, Korey Kilpatrick, Frederic MarsolaisAbstract:Ephedrine and pseudoephedrine are phenylpropylamino alkaloids widely used in modern medicine. Some Ephedra species such as E. sinica Stapf (Ephedraceae), a widely used Chinese medicinal plant (Chinese name: Ma Huang), accumulate ephedrine alkaloids as active constituents. Other Ephedra species, such as E. foeminea Forssk. (syn. E. campylopoda C.A. Mey) lack ephedrine alkaloids and their postulated metabolic precursors 1-phenylpropane-1,2-dione and (S)-cathinone. Solid-phase microextraction analysis of freshly picked young E. sinica and E. foeminea stems revealed the presence of increased benzaldehyde levels in E. foeminea, whereas 1-phenylpropane-1,2-dione was detected only in E. sinica. Soluble protein preparations from E. sinica and E. foeminea stems catalyzed the conversion of benzaldehyde and pyruvate to (R)-phenylacetylcarbinol, (S)-phenylacetylcarbinol, (R)-2-hydroxypropiophenone (S)-2-hydroxypropiophenone and 1-phenylpropane-1,2-dione. The activity, termed benzaldehyde carboxyligase (BCL) required the presence of magnesium and thiamine pyrophosphate and was 40 times higher in E. sinica as compared to E. foeminea. The distribution patterns of BCL activity in E. sinica tissues correlates well with the distribution pattern of the ephedrine alkaloids. (S)-Cathinone reductase enzymatic activities generating (1R,2S)-Norephedrine and (1S,1R)-Norephedrine were significantly higher in E. sinica relative to the levels displayed by E. foeminea. Surprisingly, (1R,2S)-Norephedrine N-methyltransferase activity which is a downstream enzyme in ephedrine biosynthesis was significantly higher in E. foeminea than in E. sinica. Our studies further support that benzaldehyde is the metabolic precursor to phenylpropylamino alkaloids in E. sinica.
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naturally occurring Norephedrine oxazolidine derivatives in khat catha edulis
Planta Medica, 2012Co-Authors: Raz Krizevski, Yaron Sitrit, Efraim Lewinsohn, Nativ Dudai, Asaf Levy, Shimon BenshabatAbstract:Khat ( Catha edulis Forsk.) is a perennial shrub whose young leaves are chewed for their psychostimulating and anorectic properties. The main active principles of khat are believed to be the phenylpropylamino alkaloids, primarily (−)-cathinone [( S )- α -aminopropiophenone], (+)-cathine [(1 S )(2 S )-norpseudoephedrine], and (−)-Norephedrine [(1 R )(2 S )-Norephedrine]. GC-MS analyses of young leaf extracts indicated the presence of two oxazolidine derivatives, 2,4-dimethyl-5-phenyloxazolidine and 4-methyl-2-( trans -1-pentenyl)-5-phenyloxazolidine. To ascertain the chemical identity of these compounds, we synthesized the putative compounds by condensation of Norephedrine and acetaldehyde or trans -2-hexenal, respectively. Spectroscopic analyses (GC-MS, NMR) of the structures of these synthetic compounds showed them to have identical retention indexes and mass spectra characteristic to 2,4-dimethyl-5-phenyloxazolidine and 4-methyl-2-( trans -1-pentenyl)-5-phenyloxazolidine. Marked differences in the ratios between each of these two Norephedrine oxazolidine derivatives and total phenylpropylamino alkaloids were found among thirteen different khat accessions further indicating polymorphism in alkaloid ratios and content in C. edulis .
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composition and stereochemistry of ephedrine alkaloids accumulation in ephedra sinica stapf
Phytochemistry, 2010Co-Authors: Raz Krizevski, Shimon Benshabat, Yaron Sitrit, Einat Bar, Or Shalit, Efraim LewinsohnAbstract:Ephedra sinica Stapf (Ephedraceae) is a widely used Chinese medicinal plant (Chinese name: Ma Huang). The main active constituents of E. sinica are the unique and taxonomically restricted adrenergic agonists phenylpropylamino alkaloids, also known as ephedrine alkaloids: (1R,2S)-Norephedrine (1S,2S)-norpseudoephedrine, (1R,2S)-ephedrine, (1S,2S)-pseudoephedrine, (1R,2S)-N-methylephedrine and (1S,2S)-N-methylpseudoephedrine. GC–MS analysis of freshly picked young E. sinica stems enabled the detection of 1-phenylpropane-1,2-dione and (S)-cathinone, the first two putative committed biosynthetic precursors to the ephedrine alkaloids. These metabolites are only present in young E. sinica stems and not in mature stems or roots. The related Ephedra foemina and Ephedra foliata also lack ephedrine alkaloids and their metabolic precursors in their aerial parts. A marked diversity in the ephedrine alkaloids content and stereochemical composition in 16 different E. sinica accessions growing under the same environmental conditions was revealed, indicating genetic control of these traits. The accessions can be classified into two groups according to the stereochemistry of the products accumulated: a group that displayed only 1R stereoisomers, and a group that displayed both 1S and 1R stereoisomers. (S)-cathinone reductase activities were detected in E. sinica stems capable of reducing (S)-cathinone to (1R,2S)-Norephedrine and (1S,2S)-norpseudoephedrine in the presence of NADH. The proportion of the diastereoisomers formed varied according to the accession tested. A (1R,2S)-Norephedrine N-methyltransferase capable of converting (1R,2S)-Norephedrine to (1R,2S)-ephedrine in the presence of S-adenosylmethionine (SAM) was also detected in E. sinica stems. Our studies further support the notion that 1-phenylpropane-1,2-dione and (S)-cathinone are biosynthetic precursors of the ephedrine alkaloids in E. sinica stems and that the activity of (S)-cathinone reductases directs and determines the stereochemical branching of the pathway. Further methylations are likely due to N-methyltransferase activities.
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quantitative stereoisomeric determination of phenylpropylamino alkaloids in khat catha edulis forsk using chiral gc ms
Israel Journal of Plant Sciences, 2008Co-Authors: Raz Krizevski, Ilana Dessow, Nativ Dudai, Efraim LewinsohnAbstract:Khat (Catha edulis Forsk., Celastraceae) is a perennial shrub that was introduced to Israel by Yemenite immigrants. Its young leaves are chewed for their psycho-stimulating properties. Young khat leaves contain the phenylpropylamino alkaloids (-)-cathinone [(S)-α-aminopropiophenone], (+)-cathine [(1S)(2S)-norpseudoephedrine], and (-)-Norephedrine [(1R)(2S)-Norephedrine] as the main active principles. A novel GC-MS analysis method for the quantitative determination of phenylpropylamino alkaloids and their putative biosynthetic precursor 1-phenylpropane-1,2- dione in khat leaves was developed. We utilized an alkaline-organic extraction, coupled with gas chromatography and a chiral permethylated beta cyclodextrin phase, to allow a full separation between the two diastereoisomers (1S)(2S)-cathine and (1R)(2 S)-Norephedrine. We found a marked diversity in the phenylpropylamino alkaloid content and composition in three different locally grown accessions and the commercial cultivar ‘Mahanaim’.
Dennis R. Feller - One of the best experts on this subject based on the ideXlab platform.
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effects of synephrine and β phenethylamine on human α adrenoceptor subtypes
Planta Medica, 2010Co-Authors: Supriya A. Bavadekar, Ikhlas A. Khan, Brian T. Schaneberg, Dennis R. FellerAbstract:Synephrine and β-phenethylamine, two naturally occurring compounds, are structurally related to ephedrine. In this study, the effects of synephrine and β-phenethylamine on α-adrenergic receptor ( α-AR) subtypes are investigated in human embryonic kidney (HEK293) or Chinese hamster ovary (CHO) cells, and compared to that of 1R,2S-Norephedrine. The rank order of binding affinities was found to be the same for the subtypes tested ( α 1A -, α 2A -, and α 2C -AR) viz, 1R,2S-Norephedrine > β-phenethylamine > synephrine. Functional studies on the α 1A -AR subtype showed that synephrine was a partial agonist giving a maximal response at 100 μM that was equal to 55.3 % of the L-phenylephrine maximum. In contrast, neither 1R,2S-Norephedrine nor β-phenethylamine exhibited agonist activity at the highest concentration tested (300 μM). β-Phenethylamine was more potent as an antagonist than 1R,2S-Norephedrine and synephrine on the α 1A -AR subtype. Functional studies on the α 2A - and α 2C -AR subtypes indicated that synephrine and β-phenethylamine did not act as agonists. Similar to 1R,2S-Norephedrine, both of these analogs reversed the effect of medetomidine against forskolin-induced cAMP elevations at 300 μM, and the rank order of antagonist potency was: 1R,2S-Norephedrine = β-phenethylamine > synephrine; and β-phenethylamine > 1R,2S-Norephedrine > synephrine, respectively. These differences suggest that the presence of a 4-hydroxy group, as in synephrine, reduced the potency in these subtypes. In conclusion, at the α 1A -AR, synephrine acted as a partial agonist, while β-phenethylamine did not exhibit any direct agonist activity. Both, synephrine and β-phenethylamine, may act as antagonists of pre-synaptic α 2A/2C -ARs present in nerve terminals.
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effects of synephrine and beta phenethylamine on human alpha adrenoceptor subtypes
Planta Medica, 2010Co-Authors: Supriya A. Bavadekar, Ikhlas A. Khan, Brian T. Schaneberg, Dennis R. FellerAbstract:Synephrine and beta-phenethylamine, two naturally occurring compounds, are structurally related to ephedrine. In this study, the effects of synephrine and beta-phenethylamine on alpha-adrenergic receptor (alpha-AR) subtypes are investigated in human embryonic kidney (HEK293) or Chinese hamster ovary (CHO) cells, and compared to that of 1R,2S-Norephedrine. The rank order of binding affinities was found to be the same for the subtypes tested (alpha(1A)-, alpha(2A)-, and alpha(2C)-AR) viz, 1R,2S-Norephedrine > beta-phenethylamine > synephrine. Functional studies on the alpha(1A)-AR subtype showed that synephrine was a partial agonist giving a maximal response at 100 microM that was equal to 55.3 % of the L-phenylephrine maximum. In contrast, neither 1R,2S-Norephedrine nor beta-phenethylamine exhibited agonist activity at the highest concentration tested (300 microM). beta-Phenethylamine was more potent as an antagonist than 1R,2S-Norephedrine and synephrine on the alpha(1A)-AR subtype. Functional studies on the alpha(2A)- and alpha(2C)-AR subtypes indicated that synephrine and beta-phenethylamine did not act as agonists. Similar to 1R,2S-Norephedrine, both of these analogs reversed the effect of medetomidine against forskolin-induced cAMP elevations at 300 microM, and the rank order of antagonist potency was: 1R,2S-Norephedrine = beta-phenethylamine > synephrine; and beta-phenethylamine > 1R,2S-Norephedrine > synephrine, respectively. These differences suggest that the presence of a 4-hydroxy group, as in synephrine, reduced the potency in these subtypes. In conclusion, at the alpha(1A)-AR, synephrine acted as a partial agonist, while beta-phenethylamine did not exhibit any direct agonist activity. Both, synephrine and beta-phenethylamine, may act as antagonists of pre-synaptic alpha(2A/2C)-ARs present in nerve terminals.
Hans Jørgen Aarstad - One of the best experts on this subject based on the ideXlab platform.
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RESEARCH ARTICLE Open Access
2016Co-Authors: Hans Jørgen Aarstad, Anne Christine Johannessen, Olav Karsten Vintermyr, Øystein Bruserud, Bjørn Tore GjertsenAbstract:Distinct single cell signal transduction signatures in leukocyte subsets stimulated with khat extract, amphetamine-like cathinone, cathine or Norephedrine Treatment with Norephedrine resulted in significantly increased T-lymphocyte proliferation, whereas khat-extract Bredholt et al. BMC Pharmacology and Toxicology 2013, 14:3
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Distinct single cell signal transduction signatures in leukocyte subsets stimulated with khat extract, amphetamine-like cathinone, cathine or Norephedrine
BMC Pharmacology and Toxicology, 2013Co-Authors: Therese Bredholt, Elisabeth Ersvær, Bjarte Skoe Erikstein, André Sulen, Håkon Reikvam, Hans Jørgen Aarstad, Anne Christine Johannessen, Olav Karsten Vintermyr, Øystein Bruserud, Bjørn Tore GjertsenAbstract:Background Amphetamine and amphetamine derivatives are suggested to induce an immunosuppressive effect. However, knowledge of how amphetamines modulate intracellular signaling pathways in cells of the immune system is limited. We have studied phosphorylation of signal transduction proteins (Akt, CREB, ERK1/2, NF-κB, c-Cbl, STAT1/3/5/6) and stress sensors (p38 MAPK, p53) in human leukocyte subsets following in vitro treatment with the natural amphetamine cathinone, the cathinone derivatives cathine and Norephedrine, in comparison with a defined extract of the psychostimulating herb khat ( Catha edulis Forsk.). Intracellular protein modifications in single cells were studied using immunostaining and flow cytometry, cell viability was determined by Annexin V-FITC/Propidium Iodide staining, and T-lymphocyte proliferation was measured by ^3H-thymidine incorporation. Results Cathinone, cathine and Norephedrine generally reduced post-translational modifications of intracellular signal transducers in T-lymphocytes, B-lymphocytes, natural killer cells and monocytes, most prominently affecting c-Cbl (pTyr700), ERK1/2 (p-Thr202/p-Tyr204), p38 MAPK (p-Thr180/p-Tyr182) and p53 (both total p53 protein and p-Ser15). In contrast, the botanical khat-extract induced protein phosphorylation of STAT1 (p-Tyr701), STAT6 (p-Tyr641), c-Cbl (pTyr700), ERK1/2 (p-Thr202/p-Tyr204), NF-κB (p-Ser529), Akt (p-Ser473), p38 MAPK (p-Thr180/p-Tyr182), p53 (Ser15) as well as total p53 protein. Cathinone, cathine and Norephedrine resulted in unique signaling profiles, with B-lymphocytes and natural killer cells more responsive compared to T-lymphocytes and monocytes. Treatment with Norephedrine resulted in significantly increased T-lymphocyte proliferation, whereas khat-extract reduced proliferation and induced cell death. Conclusions Single-cell signal transduction analyses of leukocytes distinctively discriminated between stimulation with cathinone and the structurally similar derivatives cathine and Norephedrine. Cathinone, cathine and Norephedrine reduced phosphorylation of c-Cbl, ERK1/2, p38 MAPK and p53(Ser15), and Norephedrine induced T-lymphocyte proliferation. Khat-extract induced protein phosphorylation of signal transducers, p38 MAPK and p53, followed by reduced cell proliferation and cell death. This study suggests that protein modification-specific single-cell analysis of immune cells could unravel pharmacologic effects of amphetamines and amphetamine-like agents, and further could represent a valuable tool in elucidation of mechanism(s) of action of complex botanical extracts.
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distinct single cell signal transduction signatures in leukocyte subsets stimulated with khat extract amphetamine like cathinone cathine or Norephedrine
BMC Clinical Pharmacology, 2013Co-Authors: Therese Bredholt, Elisabeth Ersvær, Bjarte Skoe Erikstein, André Sulen, Håkon Reikvam, Hans Jørgen Aarstad, Anne Christine JohannessenAbstract:Amphetamine and amphetamine derivatives are suggested to induce an immunosuppressive effect. However, knowledge of how amphetamines modulate intracellular signaling pathways in cells of the immune system is limited. We have studied phosphorylation of signal transduction proteins (Akt, CREB, ERK1/2, NF-κB, c-Cbl, STAT1/3/5/6) and stress sensors (p38 MAPK, p53) in human leukocyte subsets following in vitro treatment with the natural amphetamine cathinone, the cathinone derivatives cathine and Norephedrine, in comparison with a defined extract of the psychostimulating herb khat (Catha edulis Forsk.). Intracellular protein modifications in single cells were studied using immunostaining and flow cytometry, cell viability was determined by Annexin V-FITC/Propidium Iodide staining, and T-lymphocyte proliferation was measured by 3H-thymidine incorporation. Cathinone, cathine and Norephedrine generally reduced post-translational modifications of intracellular signal transducers in T-lymphocytes, B-lymphocytes, natural killer cells and monocytes, most prominently affecting c-Cbl (pTyr700), ERK1/2 (p-Thr202/p-Tyr204), p38 MAPK (p-Thr180/p-Tyr182) and p53 (both total p53 protein and p-Ser15). In contrast, the botanical khat-extract induced protein phosphorylation of STAT1 (p-Tyr701), STAT6 (p-Tyr641), c-Cbl (pTyr700), ERK1/2 (p-Thr202/p-Tyr204), NF-κB (p-Ser529), Akt (p-Ser473), p38 MAPK (p-Thr180/p-Tyr182), p53 (Ser15) as well as total p53 protein. Cathinone, cathine and Norephedrine resulted in unique signaling profiles, with B-lymphocytes and natural killer cells more responsive compared to T-lymphocytes and monocytes. Treatment with Norephedrine resulted in significantly increased T-lymphocyte proliferation, whereas khat-extract reduced proliferation and induced cell death. Single-cell signal transduction analyses of leukocytes distinctively discriminated between stimulation with cathinone and the structurally similar derivatives cathine and Norephedrine. Cathinone, cathine and Norephedrine reduced phosphorylation of c-Cbl, ERK1/2, p38 MAPK and p53(Ser15), and Norephedrine induced T-lymphocyte proliferation. Khat-extract induced protein phosphorylation of signal transducers, p38 MAPK and p53, followed by reduced cell proliferation and cell death. This study suggests that protein modification-specific single-cell analysis of immune cells could unravel pharmacologic effects of amphetamines and amphetamine-like agents, and further could represent a valuable tool in elucidation of mechanism(s) of action of complex botanical extracts.
Bjørn Tore Gjertsen - One of the best experts on this subject based on the ideXlab platform.
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RESEARCH ARTICLE Open Access
2016Co-Authors: Hans Jørgen Aarstad, Anne Christine Johannessen, Olav Karsten Vintermyr, Øystein Bruserud, Bjørn Tore GjertsenAbstract:Distinct single cell signal transduction signatures in leukocyte subsets stimulated with khat extract, amphetamine-like cathinone, cathine or Norephedrine Treatment with Norephedrine resulted in significantly increased T-lymphocyte proliferation, whereas khat-extract Bredholt et al. BMC Pharmacology and Toxicology 2013, 14:3
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Distinct single cell signal transduction signatures in leukocyte subsets stimulated with khat extract, amphetamine-like cathinone, cathine or Norephedrine
BMC Pharmacology and Toxicology, 2013Co-Authors: Therese Bredholt, Elisabeth Ersvær, Bjarte Skoe Erikstein, André Sulen, Håkon Reikvam, Hans Jørgen Aarstad, Anne Christine Johannessen, Olav Karsten Vintermyr, Øystein Bruserud, Bjørn Tore GjertsenAbstract:Background Amphetamine and amphetamine derivatives are suggested to induce an immunosuppressive effect. However, knowledge of how amphetamines modulate intracellular signaling pathways in cells of the immune system is limited. We have studied phosphorylation of signal transduction proteins (Akt, CREB, ERK1/2, NF-κB, c-Cbl, STAT1/3/5/6) and stress sensors (p38 MAPK, p53) in human leukocyte subsets following in vitro treatment with the natural amphetamine cathinone, the cathinone derivatives cathine and Norephedrine, in comparison with a defined extract of the psychostimulating herb khat ( Catha edulis Forsk.). Intracellular protein modifications in single cells were studied using immunostaining and flow cytometry, cell viability was determined by Annexin V-FITC/Propidium Iodide staining, and T-lymphocyte proliferation was measured by ^3H-thymidine incorporation. Results Cathinone, cathine and Norephedrine generally reduced post-translational modifications of intracellular signal transducers in T-lymphocytes, B-lymphocytes, natural killer cells and monocytes, most prominently affecting c-Cbl (pTyr700), ERK1/2 (p-Thr202/p-Tyr204), p38 MAPK (p-Thr180/p-Tyr182) and p53 (both total p53 protein and p-Ser15). In contrast, the botanical khat-extract induced protein phosphorylation of STAT1 (p-Tyr701), STAT6 (p-Tyr641), c-Cbl (pTyr700), ERK1/2 (p-Thr202/p-Tyr204), NF-κB (p-Ser529), Akt (p-Ser473), p38 MAPK (p-Thr180/p-Tyr182), p53 (Ser15) as well as total p53 protein. Cathinone, cathine and Norephedrine resulted in unique signaling profiles, with B-lymphocytes and natural killer cells more responsive compared to T-lymphocytes and monocytes. Treatment with Norephedrine resulted in significantly increased T-lymphocyte proliferation, whereas khat-extract reduced proliferation and induced cell death. Conclusions Single-cell signal transduction analyses of leukocytes distinctively discriminated between stimulation with cathinone and the structurally similar derivatives cathine and Norephedrine. Cathinone, cathine and Norephedrine reduced phosphorylation of c-Cbl, ERK1/2, p38 MAPK and p53(Ser15), and Norephedrine induced T-lymphocyte proliferation. Khat-extract induced protein phosphorylation of signal transducers, p38 MAPK and p53, followed by reduced cell proliferation and cell death. This study suggests that protein modification-specific single-cell analysis of immune cells could unravel pharmacologic effects of amphetamines and amphetamine-like agents, and further could represent a valuable tool in elucidation of mechanism(s) of action of complex botanical extracts.
Anne Christine Johannessen - One of the best experts on this subject based on the ideXlab platform.
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RESEARCH ARTICLE Open Access
2016Co-Authors: Hans Jørgen Aarstad, Anne Christine Johannessen, Olav Karsten Vintermyr, Øystein Bruserud, Bjørn Tore GjertsenAbstract:Distinct single cell signal transduction signatures in leukocyte subsets stimulated with khat extract, amphetamine-like cathinone, cathine or Norephedrine Treatment with Norephedrine resulted in significantly increased T-lymphocyte proliferation, whereas khat-extract Bredholt et al. BMC Pharmacology and Toxicology 2013, 14:3
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Distinct single cell signal transduction signatures in leukocyte subsets stimulated with khat extract, amphetamine-like cathinone, cathine or Norephedrine
BMC Pharmacology and Toxicology, 2013Co-Authors: Therese Bredholt, Elisabeth Ersvær, Bjarte Skoe Erikstein, André Sulen, Håkon Reikvam, Hans Jørgen Aarstad, Anne Christine Johannessen, Olav Karsten Vintermyr, Øystein Bruserud, Bjørn Tore GjertsenAbstract:Background Amphetamine and amphetamine derivatives are suggested to induce an immunosuppressive effect. However, knowledge of how amphetamines modulate intracellular signaling pathways in cells of the immune system is limited. We have studied phosphorylation of signal transduction proteins (Akt, CREB, ERK1/2, NF-κB, c-Cbl, STAT1/3/5/6) and stress sensors (p38 MAPK, p53) in human leukocyte subsets following in vitro treatment with the natural amphetamine cathinone, the cathinone derivatives cathine and Norephedrine, in comparison with a defined extract of the psychostimulating herb khat ( Catha edulis Forsk.). Intracellular protein modifications in single cells were studied using immunostaining and flow cytometry, cell viability was determined by Annexin V-FITC/Propidium Iodide staining, and T-lymphocyte proliferation was measured by ^3H-thymidine incorporation. Results Cathinone, cathine and Norephedrine generally reduced post-translational modifications of intracellular signal transducers in T-lymphocytes, B-lymphocytes, natural killer cells and monocytes, most prominently affecting c-Cbl (pTyr700), ERK1/2 (p-Thr202/p-Tyr204), p38 MAPK (p-Thr180/p-Tyr182) and p53 (both total p53 protein and p-Ser15). In contrast, the botanical khat-extract induced protein phosphorylation of STAT1 (p-Tyr701), STAT6 (p-Tyr641), c-Cbl (pTyr700), ERK1/2 (p-Thr202/p-Tyr204), NF-κB (p-Ser529), Akt (p-Ser473), p38 MAPK (p-Thr180/p-Tyr182), p53 (Ser15) as well as total p53 protein. Cathinone, cathine and Norephedrine resulted in unique signaling profiles, with B-lymphocytes and natural killer cells more responsive compared to T-lymphocytes and monocytes. Treatment with Norephedrine resulted in significantly increased T-lymphocyte proliferation, whereas khat-extract reduced proliferation and induced cell death. Conclusions Single-cell signal transduction analyses of leukocytes distinctively discriminated between stimulation with cathinone and the structurally similar derivatives cathine and Norephedrine. Cathinone, cathine and Norephedrine reduced phosphorylation of c-Cbl, ERK1/2, p38 MAPK and p53(Ser15), and Norephedrine induced T-lymphocyte proliferation. Khat-extract induced protein phosphorylation of signal transducers, p38 MAPK and p53, followed by reduced cell proliferation and cell death. This study suggests that protein modification-specific single-cell analysis of immune cells could unravel pharmacologic effects of amphetamines and amphetamine-like agents, and further could represent a valuable tool in elucidation of mechanism(s) of action of complex botanical extracts.
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distinct single cell signal transduction signatures in leukocyte subsets stimulated with khat extract amphetamine like cathinone cathine or Norephedrine
BMC Clinical Pharmacology, 2013Co-Authors: Therese Bredholt, Elisabeth Ersvær, Bjarte Skoe Erikstein, André Sulen, Håkon Reikvam, Hans Jørgen Aarstad, Anne Christine JohannessenAbstract:Amphetamine and amphetamine derivatives are suggested to induce an immunosuppressive effect. However, knowledge of how amphetamines modulate intracellular signaling pathways in cells of the immune system is limited. We have studied phosphorylation of signal transduction proteins (Akt, CREB, ERK1/2, NF-κB, c-Cbl, STAT1/3/5/6) and stress sensors (p38 MAPK, p53) in human leukocyte subsets following in vitro treatment with the natural amphetamine cathinone, the cathinone derivatives cathine and Norephedrine, in comparison with a defined extract of the psychostimulating herb khat (Catha edulis Forsk.). Intracellular protein modifications in single cells were studied using immunostaining and flow cytometry, cell viability was determined by Annexin V-FITC/Propidium Iodide staining, and T-lymphocyte proliferation was measured by 3H-thymidine incorporation. Cathinone, cathine and Norephedrine generally reduced post-translational modifications of intracellular signal transducers in T-lymphocytes, B-lymphocytes, natural killer cells and monocytes, most prominently affecting c-Cbl (pTyr700), ERK1/2 (p-Thr202/p-Tyr204), p38 MAPK (p-Thr180/p-Tyr182) and p53 (both total p53 protein and p-Ser15). In contrast, the botanical khat-extract induced protein phosphorylation of STAT1 (p-Tyr701), STAT6 (p-Tyr641), c-Cbl (pTyr700), ERK1/2 (p-Thr202/p-Tyr204), NF-κB (p-Ser529), Akt (p-Ser473), p38 MAPK (p-Thr180/p-Tyr182), p53 (Ser15) as well as total p53 protein. Cathinone, cathine and Norephedrine resulted in unique signaling profiles, with B-lymphocytes and natural killer cells more responsive compared to T-lymphocytes and monocytes. Treatment with Norephedrine resulted in significantly increased T-lymphocyte proliferation, whereas khat-extract reduced proliferation and induced cell death. Single-cell signal transduction analyses of leukocytes distinctively discriminated between stimulation with cathinone and the structurally similar derivatives cathine and Norephedrine. Cathinone, cathine and Norephedrine reduced phosphorylation of c-Cbl, ERK1/2, p38 MAPK and p53(Ser15), and Norephedrine induced T-lymphocyte proliferation. Khat-extract induced protein phosphorylation of signal transducers, p38 MAPK and p53, followed by reduced cell proliferation and cell death. This study suggests that protein modification-specific single-cell analysis of immune cells could unravel pharmacologic effects of amphetamines and amphetamine-like agents, and further could represent a valuable tool in elucidation of mechanism(s) of action of complex botanical extracts.
