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S-Nitrosylation in Organs of Mice Exposed to Low or High Doses of γ-Rays: The Modulating Effect of Iodine Contrast Agent at a Low Radiation Dose.

Nicolas F, Wu C, Bukhari S, de Toledo SM, Li H, Shibata M, Azzam EI - Proteomes (2015)

Bottom Line: The covalent addition of nitric oxide (NO(•)) onto cysteine thiols, or S-nitrosylation, modulates the activity of key signaling proteins.To gain insight into the biochemical changes induced by low-dose ionizing radiation, we determined global S-nitrosylation by the "biotin switch" assay coupled with mass spectrometry analyses in organs of C57BL/6J mice exposed to acute 0.1 Gy of (137)Cs γ-rays.The major results were as follows: (i) iopamidol alone had significant effects on S-nitrosylation in brain, lung and liver; (ii) relative to the control, exposure to 0.1 Gy without iopamidol resulted in statistically-significant SNO changes in proteins that differ in molecular weight in liver, lung, brain and blood plasma; (iii) iopamidol enhanced the decrease in S-nitrosylation induced by 0.1 Gy in brain; (iv) whereas a decrease in S-nitrosylation occurred at 0.1 Gy for proteins of ~50 kDa in brain and for proteins of ~37 kDa in liver, an increase was detected at 4 Gy in both organs; (v) mass spectrometry analyses of nitrosylated proteins in brain revealed differential modulation of SNO proteins (e.g., sodium/potassium-transporting ATPase subunit beta-1; beta tubulins; ADP-ribosylation factor 5) by low- and high-dose irradiation; and (vi) ingenuity pathway analysis identified major signaling networks to be modulated, in particular the neuronal nitric oxide synthase signaling pathway was differentially modulated by low- and high-dose γ-irradiation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Health Informatics, Rutgers School of Health Related Professions, Newark, NJ 07107, USA; nicolafa@shrp.rutgers.edu.

ABSTRACT

The covalent addition of nitric oxide (NO(•)) onto cysteine thiols, or S-nitrosylation, modulates the activity of key signaling proteins. The dysregulation of normal S-nitrosylation contributes to degenerative conditions and to cancer. To gain insight into the biochemical changes induced by low-dose ionizing radiation, we determined global S-nitrosylation by the "biotin switch" assay coupled with mass spectrometry analyses in organs of C57BL/6J mice exposed to acute 0.1 Gy of (137)Cs γ-rays. The dose of radiation was delivered to the whole body in the presence or absence of iopamidol, an iodinated contrast agent used during radiological examinations. To investigate whether similar or distinct nitrosylation patterns are induced following high-dose irradiation, mice were exposed in parallel to acute 4 Gy of (137)Cs γ rays. Analysis of modulated S-nitrosothiols (SNO-proteins) in freshly-harvested organs of animals sacrificed 13 days after irradiation revealed radiation dose- and contrast agent-dependent changes. The major results were as follows: (i) iopamidol alone had significant effects on S-nitrosylation in brain, lung and liver; (ii) relative to the control, exposure to 0.1 Gy without iopamidol resulted in statistically-significant SNO changes in proteins that differ in molecular weight in liver, lung, brain and blood plasma; (iii) iopamidol enhanced the decrease in S-nitrosylation induced by 0.1 Gy in brain; (iv) whereas a decrease in S-nitrosylation occurred at 0.1 Gy for proteins of ~50 kDa in brain and for proteins of ~37 kDa in liver, an increase was detected at 4 Gy in both organs; (v) mass spectrometry analyses of nitrosylated proteins in brain revealed differential modulation of SNO proteins (e.g., sodium/potassium-transporting ATPase subunit beta-1; beta tubulins; ADP-ribosylation factor 5) by low- and high-dose irradiation; and (vi) ingenuity pathway analysis identified major signaling networks to be modulated, in particular the neuronal nitric oxide synthase signaling pathway was differentially modulated by low- and high-dose γ-irradiation.

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Modulation of S-nitrosylation, in vivo, by ionizing radiation and/or radiocontrast agent. Western blot analyses, following the biotin switch assay, of nitrosylated proteins from plasma (A), brain (B), liver (C), and lung (D) of C57 of C57Bl/6J mice exposed 13 days earlier to 137Cs γ-rays in the presence or absence of iopamidol. Proteins from mouse organs were freshly extracted and subjected to the biotin switch assay. The biotinylated proteins were detected with anti-biotin antibody. Protein aliquots (15 µg), before enrichment, were used as the input standard, and the expression level of GAPDH was used as the loading control. In the case of plasma from circulating blood, staining of the membrane with Ponceau S Red (not shown) indicated equal loading.
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Figure 1: Modulation of S-nitrosylation, in vivo, by ionizing radiation and/or radiocontrast agent. Western blot analyses, following the biotin switch assay, of nitrosylated proteins from plasma (A), brain (B), liver (C), and lung (D) of C57 of C57Bl/6J mice exposed 13 days earlier to 137Cs γ-rays in the presence or absence of iopamidol. Proteins from mouse organs were freshly extracted and subjected to the biotin switch assay. The biotinylated proteins were detected with anti-biotin antibody. Protein aliquots (15 µg), before enrichment, were used as the input standard, and the expression level of GAPDH was used as the loading control. In the case of plasma from circulating blood, staining of the membrane with Ponceau S Red (not shown) indicated equal loading.

Mentions: Deregulation of S-nitrosylation has been associated with degenerative diseases [36]. Further, recent findings have suggested that exposures to moderate and even low doses of ionizing radiation can induce degenerative conditions [37], including neurodegeneration and neuroinflammation [38]. To characterize the dependence of nitrosative stress on radiation dose, we investigated global S-nitrosylation by the ‘biotin switch’ assay in whole organs of irradiated mice. To this end, changes in the levels of SNO proteins with varying molecular weights in liver, lung, brain, as well as plasma were examined 13 days after exposure of 17-week-old C57BL/6J male mice to acute doses of 0, 0.1 or 4 Gy of 137Cs γ-rays. To gain understanding of the modulating effect of contrast agents used in radiodiagnostic procedures, mouse exposure to 0 or 0.1 Gy was carried out in the presence or absence of iopamidol, an iodinated agent that was delivered intravenously immediately prior to irradiation. The biotin switch assay results in Figure 1 describe representative radiation dose-dependent changes in the levels of nitrosylated proteins extracted from freshly-harvested organs of three individual mice. The Western blot analyses in the various panels show that relative to the control, exposure to 0.1 Gy resulted in prominent changes in the levels of SNO proteins in plasma, brain and liver. Interestingly, the decreases observed at 0.1 Gy in brain and liver are contrasted with increases in SNO proteins with a similar molecular weight (40–60 kDa) in brain and liver of mice exposed to 4 Gy. In plasma, decreases in similar magnitude to those observed at 0.1 Gy were detected at 4 Gy for proteins of the molecular weight range shown in (A) of Figure 1. In lung, comparable increases were detected at 0.1 and 4 Gy. The full range of changes in SNO proteins of different molecular weights is shown in Supplementary Figure S1.


S-Nitrosylation in Organs of Mice Exposed to Low or High Doses of γ-Rays: The Modulating Effect of Iodine Contrast Agent at a Low Radiation Dose.

Nicolas F, Wu C, Bukhari S, de Toledo SM, Li H, Shibata M, Azzam EI - Proteomes (2015)

Modulation of S-nitrosylation, in vivo, by ionizing radiation and/or radiocontrast agent. Western blot analyses, following the biotin switch assay, of nitrosylated proteins from plasma (A), brain (B), liver (C), and lung (D) of C57 of C57Bl/6J mice exposed 13 days earlier to 137Cs γ-rays in the presence or absence of iopamidol. Proteins from mouse organs were freshly extracted and subjected to the biotin switch assay. The biotinylated proteins were detected with anti-biotin antibody. Protein aliquots (15 µg), before enrichment, were used as the input standard, and the expression level of GAPDH was used as the loading control. In the case of plasma from circulating blood, staining of the membrane with Ponceau S Red (not shown) indicated equal loading.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4548934&req=5

Figure 1: Modulation of S-nitrosylation, in vivo, by ionizing radiation and/or radiocontrast agent. Western blot analyses, following the biotin switch assay, of nitrosylated proteins from plasma (A), brain (B), liver (C), and lung (D) of C57 of C57Bl/6J mice exposed 13 days earlier to 137Cs γ-rays in the presence or absence of iopamidol. Proteins from mouse organs were freshly extracted and subjected to the biotin switch assay. The biotinylated proteins were detected with anti-biotin antibody. Protein aliquots (15 µg), before enrichment, were used as the input standard, and the expression level of GAPDH was used as the loading control. In the case of plasma from circulating blood, staining of the membrane with Ponceau S Red (not shown) indicated equal loading.
Mentions: Deregulation of S-nitrosylation has been associated with degenerative diseases [36]. Further, recent findings have suggested that exposures to moderate and even low doses of ionizing radiation can induce degenerative conditions [37], including neurodegeneration and neuroinflammation [38]. To characterize the dependence of nitrosative stress on radiation dose, we investigated global S-nitrosylation by the ‘biotin switch’ assay in whole organs of irradiated mice. To this end, changes in the levels of SNO proteins with varying molecular weights in liver, lung, brain, as well as plasma were examined 13 days after exposure of 17-week-old C57BL/6J male mice to acute doses of 0, 0.1 or 4 Gy of 137Cs γ-rays. To gain understanding of the modulating effect of contrast agents used in radiodiagnostic procedures, mouse exposure to 0 or 0.1 Gy was carried out in the presence or absence of iopamidol, an iodinated agent that was delivered intravenously immediately prior to irradiation. The biotin switch assay results in Figure 1 describe representative radiation dose-dependent changes in the levels of nitrosylated proteins extracted from freshly-harvested organs of three individual mice. The Western blot analyses in the various panels show that relative to the control, exposure to 0.1 Gy resulted in prominent changes in the levels of SNO proteins in plasma, brain and liver. Interestingly, the decreases observed at 0.1 Gy in brain and liver are contrasted with increases in SNO proteins with a similar molecular weight (40–60 kDa) in brain and liver of mice exposed to 4 Gy. In plasma, decreases in similar magnitude to those observed at 0.1 Gy were detected at 4 Gy for proteins of the molecular weight range shown in (A) of Figure 1. In lung, comparable increases were detected at 0.1 and 4 Gy. The full range of changes in SNO proteins of different molecular weights is shown in Supplementary Figure S1.

Bottom Line: The covalent addition of nitric oxide (NO(•)) onto cysteine thiols, or S-nitrosylation, modulates the activity of key signaling proteins.To gain insight into the biochemical changes induced by low-dose ionizing radiation, we determined global S-nitrosylation by the "biotin switch" assay coupled with mass spectrometry analyses in organs of C57BL/6J mice exposed to acute 0.1 Gy of (137)Cs γ-rays.The major results were as follows: (i) iopamidol alone had significant effects on S-nitrosylation in brain, lung and liver; (ii) relative to the control, exposure to 0.1 Gy without iopamidol resulted in statistically-significant SNO changes in proteins that differ in molecular weight in liver, lung, brain and blood plasma; (iii) iopamidol enhanced the decrease in S-nitrosylation induced by 0.1 Gy in brain; (iv) whereas a decrease in S-nitrosylation occurred at 0.1 Gy for proteins of ~50 kDa in brain and for proteins of ~37 kDa in liver, an increase was detected at 4 Gy in both organs; (v) mass spectrometry analyses of nitrosylated proteins in brain revealed differential modulation of SNO proteins (e.g., sodium/potassium-transporting ATPase subunit beta-1; beta tubulins; ADP-ribosylation factor 5) by low- and high-dose irradiation; and (vi) ingenuity pathway analysis identified major signaling networks to be modulated, in particular the neuronal nitric oxide synthase signaling pathway was differentially modulated by low- and high-dose γ-irradiation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Health Informatics, Rutgers School of Health Related Professions, Newark, NJ 07107, USA; nicolafa@shrp.rutgers.edu.

ABSTRACT

The covalent addition of nitric oxide (NO(•)) onto cysteine thiols, or S-nitrosylation, modulates the activity of key signaling proteins. The dysregulation of normal S-nitrosylation contributes to degenerative conditions and to cancer. To gain insight into the biochemical changes induced by low-dose ionizing radiation, we determined global S-nitrosylation by the "biotin switch" assay coupled with mass spectrometry analyses in organs of C57BL/6J mice exposed to acute 0.1 Gy of (137)Cs γ-rays. The dose of radiation was delivered to the whole body in the presence or absence of iopamidol, an iodinated contrast agent used during radiological examinations. To investigate whether similar or distinct nitrosylation patterns are induced following high-dose irradiation, mice were exposed in parallel to acute 4 Gy of (137)Cs γ rays. Analysis of modulated S-nitrosothiols (SNO-proteins) in freshly-harvested organs of animals sacrificed 13 days after irradiation revealed radiation dose- and contrast agent-dependent changes. The major results were as follows: (i) iopamidol alone had significant effects on S-nitrosylation in brain, lung and liver; (ii) relative to the control, exposure to 0.1 Gy without iopamidol resulted in statistically-significant SNO changes in proteins that differ in molecular weight in liver, lung, brain and blood plasma; (iii) iopamidol enhanced the decrease in S-nitrosylation induced by 0.1 Gy in brain; (iv) whereas a decrease in S-nitrosylation occurred at 0.1 Gy for proteins of ~50 kDa in brain and for proteins of ~37 kDa in liver, an increase was detected at 4 Gy in both organs; (v) mass spectrometry analyses of nitrosylated proteins in brain revealed differential modulation of SNO proteins (e.g., sodium/potassium-transporting ATPase subunit beta-1; beta tubulins; ADP-ribosylation factor 5) by low- and high-dose irradiation; and (vi) ingenuity pathway analysis identified major signaling networks to be modulated, in particular the neuronal nitric oxide synthase signaling pathway was differentially modulated by low- and high-dose γ-irradiation.

No MeSH data available.


Related in: MedlinePlus