Hydroxyurea generates nitric oxide in human erythroid cells: Mechanisms for c-globin gene activation

Tzu Fang Lou; Manisha Singh; Ashley Mackie; Wei Li; Betty S. Pace

doi:10.3181/0811-RM-339

Hydroxyurea generates nitric oxide in human erythroid cells: Mechanisms for c-globin gene activation

Tzu Fang Lou, Manisha Singh, Ashley Mackie, Wei Li, Betty S. Pace

Research output: Contribution to journal › Article › peer-review

48 Scopus citations

Abstract

Hydroxyurea (HU) induces fetal hemoglobin synthesis through activation of cyclic guanine monophosphate (cGMP) signaling. Studies in sickle cell patients demonstrated increased circulating nitric oxide (NO) levels after oral HU treatment. However, the direct measurement of NO in erythroid cells and its role in fetal hemoglobin induction have not been defined. Therefore, we quantified the level of nitrate and nitrite (NOx) generated by HU in human erythroid progenitors in the presence of three nitric oxide synthase inhibitors (NOS), including N^G-monomethyl-L-arginine (L-NMMA). In addition, cGMP levels were measured in the presence or absence of the pathway inhibitor 1H-(1,2,4)oxadiazolo(4,3-a)quinoxalin-1-one, which blocks soluble guanylyl cyclase formation. HU treatment increased NOx levels and γ-globin transcription in K562 and primary erythroid cells, which was augmented when HU was combined with L-NMMA. Pretreatment with the cGMP pathway inhibitor reversed γ-gene activation by HU. These data demonstrate the direct stimulation of cellular NO and cGMP signaling in erythroid progenitors by HU as a possible mechanism for γ-globin gene activation.

Original language	English (US)
Pages (from-to)	1374-1382
Number of pages	9
Journal	Experimental Biology and Medicine
Volume	234
Issue number	11
DOIs	https://doi.org/10.3181/0811-RM-339
State	Published - Nov 2009
Externally published	Yes

Keywords

Fetal hemoglobin
Hydroxyurea
Nitric oxide
Sickle-cell disease
cGMP

ASJC Scopus subject areas

Biochemistry, Genetics and Molecular Biology(all)

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title = "Hydroxyurea generates nitric oxide in human erythroid cells: Mechanisms for c-globin gene activation",

abstract = "Hydroxyurea (HU) induces fetal hemoglobin synthesis through activation of cyclic guanine monophosphate (cGMP) signaling. Studies in sickle cell patients demonstrated increased circulating nitric oxide (NO) levels after oral HU treatment. However, the direct measurement of NO in erythroid cells and its role in fetal hemoglobin induction have not been defined. Therefore, we quantified the level of nitrate and nitrite (NOx) generated by HU in human erythroid progenitors in the presence of three nitric oxide synthase inhibitors (NOS), including NG-monomethyl-L-arginine (L-NMMA). In addition, cGMP levels were measured in the presence or absence of the pathway inhibitor 1H-(1,2,4)oxadiazolo(4,3-a)quinoxalin-1-one, which blocks soluble guanylyl cyclase formation. HU treatment increased NOx levels and γ-globin transcription in K562 and primary erythroid cells, which was augmented when HU was combined with L-NMMA. Pretreatment with the cGMP pathway inhibitor reversed γ-gene activation by HU. These data demonstrate the direct stimulation of cellular NO and cGMP signaling in erythroid progenitors by HU as a possible mechanism for γ-globin gene activation.",

keywords = "Fetal hemoglobin, Hydroxyurea, Nitric oxide, Sickle-cell disease, cGMP",

author = "Lou, {Tzu Fang} and Manisha Singh and Ashley Mackie and Wei Li and Pace, {Betty S.}",

note = "Funding Information: This research was supported by grant HL69234 from the National Heart, Lung, and Blood Institute to Betty Pace, MD and state funding of the Center for Applied Biology at the University of Texas at Dallas. ",

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AU - Singh, Manisha

AU - Mackie, Ashley

AU - Li, Wei

AU - Pace, Betty S.

N1 - Funding Information: This research was supported by grant HL69234 from the National Heart, Lung, and Blood Institute to Betty Pace, MD and state funding of the Center for Applied Biology at the University of Texas at Dallas.

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N2 - Hydroxyurea (HU) induces fetal hemoglobin synthesis through activation of cyclic guanine monophosphate (cGMP) signaling. Studies in sickle cell patients demonstrated increased circulating nitric oxide (NO) levels after oral HU treatment. However, the direct measurement of NO in erythroid cells and its role in fetal hemoglobin induction have not been defined. Therefore, we quantified the level of nitrate and nitrite (NOx) generated by HU in human erythroid progenitors in the presence of three nitric oxide synthase inhibitors (NOS), including NG-monomethyl-L-arginine (L-NMMA). In addition, cGMP levels were measured in the presence or absence of the pathway inhibitor 1H-(1,2,4)oxadiazolo(4,3-a)quinoxalin-1-one, which blocks soluble guanylyl cyclase formation. HU treatment increased NOx levels and γ-globin transcription in K562 and primary erythroid cells, which was augmented when HU was combined with L-NMMA. Pretreatment with the cGMP pathway inhibitor reversed γ-gene activation by HU. These data demonstrate the direct stimulation of cellular NO and cGMP signaling in erythroid progenitors by HU as a possible mechanism for γ-globin gene activation.

AB - Hydroxyurea (HU) induces fetal hemoglobin synthesis through activation of cyclic guanine monophosphate (cGMP) signaling. Studies in sickle cell patients demonstrated increased circulating nitric oxide (NO) levels after oral HU treatment. However, the direct measurement of NO in erythroid cells and its role in fetal hemoglobin induction have not been defined. Therefore, we quantified the level of nitrate and nitrite (NOx) generated by HU in human erythroid progenitors in the presence of three nitric oxide synthase inhibitors (NOS), including NG-monomethyl-L-arginine (L-NMMA). In addition, cGMP levels were measured in the presence or absence of the pathway inhibitor 1H-(1,2,4)oxadiazolo(4,3-a)quinoxalin-1-one, which blocks soluble guanylyl cyclase formation. HU treatment increased NOx levels and γ-globin transcription in K562 and primary erythroid cells, which was augmented when HU was combined with L-NMMA. Pretreatment with the cGMP pathway inhibitor reversed γ-gene activation by HU. These data demonstrate the direct stimulation of cellular NO and cGMP signaling in erythroid progenitors by HU as a possible mechanism for γ-globin gene activation.

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Hydroxyurea generates nitric oxide in human erythroid cells: Mechanisms for c-globin gene activation

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