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Deorphanization and characterization of the ectopically expressed olfactory receptor OR51B5 in myelogenous leukemia cells

View Article: PubMed Central - PubMed

ABSTRACT

The ectopic expression of olfactory receptors (ORs) in the human body has been of major interest in the past decade. Several studies have reported the expression of ORs not only in healthy tissues such as heart, sperm or skin cells, but also in cancerous tissues of the liver, prostate or intestine. In the present study, we detected the expression of OR51B5 in the chronic myelogenous leukemia (CML) cell line K562 and in white blood cell samples of clinically diagnosed acute myelogenous leukemia (AML) patients by reverse transcription-PCR and immunocytochemical staining. The known OR51B5 ligand isononyl alcohol increased the levels of intracellular Ca2+ in both AML patient blood cells and K562 cells. With calcium imaging experiments, we characterized in greater detail the OR51B5-mediated signaling pathway. Here, we observed an involvement of adenylate cyclase and the downstream L-type and T-type calcium channels. In addition, the activation of OR51B5 leads to an inhibition of cell proliferation in K562 cells. In western blot experiments, we found that incubation with isononyl alcohol led to a reduction in p38-MAPK (mitogen-activated protein kinase) phosphorylation that might be responsible for the decreased cell proliferation. In the present study, we characterized the OR51B5-mediated signaling pathway downstream of the activation with isononyl alcohol, which leads to reduced proliferation and therefore provide a novel pharmacological target for CML and AML, the latter of which remains difficult to treat.

No MeSH data available.


Related in: MedlinePlus

Calcium imaging experiments in the native blood samples of clinically diagnosed AML patients. (a) Native white blood cells from AML patients showed an increase in their intracellular Ca2+ levels during isononyl alcohol application. (b) Representative calcium imaging with the application of 100 μM isononyl alcohol. (c) SQ-22536 abolished the isononyl alcohol-induced increase in intracellular Ca2+. (d) In absence of extracellular Ca2+ the isononyl alcohol-induced increase in intracellular Ca2+ was abolished. (e) Isononyl alcohol (100 μM) led to a significant increase in intracellular Ca2+ in ~35% of all AML cells. SQ-22536 and the extracellular Ca2+ chelator EGTA reduced the number of responding cells.
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fig4: Calcium imaging experiments in the native blood samples of clinically diagnosed AML patients. (a) Native white blood cells from AML patients showed an increase in their intracellular Ca2+ levels during isononyl alcohol application. (b) Representative calcium imaging with the application of 100 μM isononyl alcohol. (c) SQ-22536 abolished the isononyl alcohol-induced increase in intracellular Ca2+. (d) In absence of extracellular Ca2+ the isononyl alcohol-induced increase in intracellular Ca2+ was abolished. (e) Isononyl alcohol (100 μM) led to a significant increase in intracellular Ca2+ in ~35% of all AML cells. SQ-22536 and the extracellular Ca2+ chelator EGTA reduced the number of responding cells.

Mentions: To investigate whether this signaling pathway is also activated in native, undifferentiated myeloid cells, we isolated white blood cells from clinically diagnosed AML patients due to their relatively high levels of blast cells in the peripheral bloodstream. During the treatment with isononyl alcohol, the AML cells showed a rapid increase in their amount of intracellular Ca2+ with similar kinetics as those of K562 cells (Figures 4a and b). In total, 100 μM isononyl alcohol could increase the cytosolic Ca2+ level of ~34% of all AML cells. To investigate whether the same signaling pathway as in K562 cells is involved, we used the same inhibitors that we used on K562 cells. Both the AC inhibitor SQ-22536 and depletion of extracellular Ca2+ significantly reduced the intracellular Ca2+ levels, thus validating the involvement of an AC-mediated signaling pathway downstream of the OR51B5 activation in AML patient blood cells (Figures 4c and d). An overview of the calcium imaging results obtained in AML cells is shown in Figure 4e.


Deorphanization and characterization of the ectopically expressed olfactory receptor OR51B5 in myelogenous leukemia cells
Calcium imaging experiments in the native blood samples of clinically diagnosed AML patients. (a) Native white blood cells from AML patients showed an increase in their intracellular Ca2+ levels during isononyl alcohol application. (b) Representative calcium imaging with the application of 100 μM isononyl alcohol. (c) SQ-22536 abolished the isononyl alcohol-induced increase in intracellular Ca2+. (d) In absence of extracellular Ca2+ the isononyl alcohol-induced increase in intracellular Ca2+ was abolished. (e) Isononyl alcohol (100 μM) led to a significant increase in intracellular Ca2+ in ~35% of all AML cells. SQ-22536 and the extracellular Ca2+ chelator EGTA reduced the number of responding cells.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4979495&req=5

fig4: Calcium imaging experiments in the native blood samples of clinically diagnosed AML patients. (a) Native white blood cells from AML patients showed an increase in their intracellular Ca2+ levels during isononyl alcohol application. (b) Representative calcium imaging with the application of 100 μM isononyl alcohol. (c) SQ-22536 abolished the isononyl alcohol-induced increase in intracellular Ca2+. (d) In absence of extracellular Ca2+ the isononyl alcohol-induced increase in intracellular Ca2+ was abolished. (e) Isononyl alcohol (100 μM) led to a significant increase in intracellular Ca2+ in ~35% of all AML cells. SQ-22536 and the extracellular Ca2+ chelator EGTA reduced the number of responding cells.
Mentions: To investigate whether this signaling pathway is also activated in native, undifferentiated myeloid cells, we isolated white blood cells from clinically diagnosed AML patients due to their relatively high levels of blast cells in the peripheral bloodstream. During the treatment with isononyl alcohol, the AML cells showed a rapid increase in their amount of intracellular Ca2+ with similar kinetics as those of K562 cells (Figures 4a and b). In total, 100 μM isononyl alcohol could increase the cytosolic Ca2+ level of ~34% of all AML cells. To investigate whether the same signaling pathway as in K562 cells is involved, we used the same inhibitors that we used on K562 cells. Both the AC inhibitor SQ-22536 and depletion of extracellular Ca2+ significantly reduced the intracellular Ca2+ levels, thus validating the involvement of an AC-mediated signaling pathway downstream of the OR51B5 activation in AML patient blood cells (Figures 4c and d). An overview of the calcium imaging results obtained in AML cells is shown in Figure 4e.

View Article: PubMed Central - PubMed

ABSTRACT

The ectopic expression of olfactory receptors (ORs) in the human body has been of major interest in the past decade. Several studies have reported the expression of ORs not only in healthy tissues such as heart, sperm or skin cells, but also in cancerous tissues of the liver, prostate or intestine. In the present study, we detected the expression of OR51B5 in the chronic myelogenous leukemia (CML) cell line K562 and in white blood cell samples of clinically diagnosed acute myelogenous leukemia (AML) patients by reverse transcription-PCR and immunocytochemical staining. The known OR51B5 ligand isononyl alcohol increased the levels of intracellular Ca2+ in both AML patient blood cells and K562 cells. With calcium imaging experiments, we characterized in greater detail the OR51B5-mediated signaling pathway. Here, we observed an involvement of adenylate cyclase and the downstream L-type and T-type calcium channels. In addition, the activation of OR51B5 leads to an inhibition of cell proliferation in K562 cells. In western blot experiments, we found that incubation with isononyl alcohol led to a reduction in p38-MAPK (mitogen-activated protein kinase) phosphorylation that might be responsible for the decreased cell proliferation. In the present study, we characterized the OR51B5-mediated signaling pathway downstream of the activation with isononyl alcohol, which leads to reduced proliferation and therefore provide a novel pharmacological target for CML and AML, the latter of which remains difficult to treat.

No MeSH data available.


Related in: MedlinePlus