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NAD(P)H: quinone oxidoreductase 1 deficiency conjoint with marginal vitamin C deficiency causes cigarette smoke induced myelodysplastic syndromes.

Das A, Dey N, Ghosh A, Das T, Chatterjee IB - PLoS ONE (2011)

Bottom Line: Apoptosis precedes MDS but disappears later with marked decrease in the p53 protein.However, after the onset of MDS vitamin C becomes ineffective.Our results suggest that human smokers having NQO1 deficiency combined with marginal vitamin C deficiency are likely to be at high risk for developing MDS and that intake of a moderately large dose of vitamin C would prevent MDS.

View Article: PubMed Central - PubMed

Affiliation: Department of Biotechnology and Dr. B. C. Guha Centre for Genetic Engineering and Biotechnology, Calcutta University College of Science, Kolkata, West Bengal, India.

ABSTRACT

Background: The etiology of myelodysplastic syndromes (MDS) is largely unknown. Exposure to cigarette smoke (CS) is reported to be associated with MDS risk. There is inconsistent evidence that deficiency of NAD(P)H-quinone: oxidoreductase 1 (NQO1) increases the risk of MDS. Earlier we had shown that CS induces toxicity only in marginal vitamin C-deficient guinea pigs but not in vitamin C-sufficient ones. We therefore considered that NQO1 deficiency along with marginal vitamin C deficiency might produce MDS in CS-exposed guinea pigs.

Methodology and principal findings: Here we show that CS exposure for 21 days produces MDS in guinea pigs having deficiency of NQO1 (fed 3 mg dicoumarol/day) conjoint with marginal vitamin C deficiency (fed 0.5 mg vitamin C/day). As evidenced by morphology, histology and cytogenetics, MDS produced in the guinea pigs falls in the category of refractory cytopenia with unilineage dysplasia (RCUD): refractory anemia; refractory thrombocytopenia that is associated with ring sideroblasts, micromegakaryocytes, myeloid hyperplasia and aneuploidy. MDS is accompanied by increased CD34(+) cells and oxidative stress as shown by the formation of protein carbonyls and 8-oxodeoxyguanosine. Apoptosis precedes MDS but disappears later with marked decrease in the p53 protein. MDS produced in the guinea pigs are irreversible. MDS and all the aforesaid pathophysiological events do not occur in vitamin C-sufficient guinea pigs. However, after the onset of MDS vitamin C becomes ineffective.

Conclusions and significance: CS exposure causes MDS in guinea pigs having deficiency of NQO1 conjoint with marginal vitamin C deficiency. The syndromes are not produced in singular deficiency of NQO1 or marginal vitamin C deficiency. Our results suggest that human smokers having NQO1 deficiency combined with marginal vitamin C deficiency are likely to be at high risk for developing MDS and that intake of a moderately large dose of vitamin C would prevent MDS.

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Identification of p-BQ-protein adducts and detection of oxidative stress in bone marrow cells of guinea pigs.(Panel A) p-BQ protein adducts in the bone marrow cells of CS-exposed guinea pigs on day 21 in vivo. (Panel B) p-BQ protein adducts in CS-exposed guinea pigs at different time period in vivo. (Panel C) p-BQ protein adducts formed in marrow cells in vitro after incubation with p-BQ and AECS (aqueous extract of cigarette smoke), respectively; M, marker (cropped). (Panel D) ROS production in MDS guinea pigs at different time periods, as evidenced by flow cytometry. The X-axis represents the intensity of dichlorodihydrofluorescein diacetate (H2DCFDA). (Panel E) Protein oxidation as evidenced by oxyblot indicating formation of protein carbonyl. (Panel F) DNA oxidation as evidenced by the formation of 8-oxodG; upper row: green fluorescence indicates formation of 8-oxodG; lower row: stained with DAPI; (magnification 200×). (Panel G) Quantitative evaluation of 8-oxodG; * indicates significant difference from 0 and 7 days. Vit C means vitamin C.
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pone-0020590-g004: Identification of p-BQ-protein adducts and detection of oxidative stress in bone marrow cells of guinea pigs.(Panel A) p-BQ protein adducts in the bone marrow cells of CS-exposed guinea pigs on day 21 in vivo. (Panel B) p-BQ protein adducts in CS-exposed guinea pigs at different time period in vivo. (Panel C) p-BQ protein adducts formed in marrow cells in vitro after incubation with p-BQ and AECS (aqueous extract of cigarette smoke), respectively; M, marker (cropped). (Panel D) ROS production in MDS guinea pigs at different time periods, as evidenced by flow cytometry. The X-axis represents the intensity of dichlorodihydrofluorescein diacetate (H2DCFDA). (Panel E) Protein oxidation as evidenced by oxyblot indicating formation of protein carbonyl. (Panel F) DNA oxidation as evidenced by the formation of 8-oxodG; upper row: green fluorescence indicates formation of 8-oxodG; lower row: stained with DAPI; (magnification 200×). (Panel G) Quantitative evaluation of 8-oxodG; * indicates significant difference from 0 and 7 days. Vit C means vitamin C.

Mentions: CS contains substantial amount of p-BSQ, [15], [17] a major long-lived radical. We had shown before that p-BSQ is oxidized to p-BQ by transition metal containing proteins [14]. The amount of p-BQ calculated to be formed in the lungs from the inhaled cigarette smoke is 22.5 µg/day (Supporting Information Text S1). Using antibody against p-BQ and immunoblotting, here we show that bone marrow proteins of marginal vitamin C-deficient CS-exposed guinea pigs contain p-BQ-protein adducts (Figure 4, A and B). Also, incubation of bone marrow cells in vitro with p-BQ or aqueous extract of cigarette smoke (AECS) produced p-BQ-protein adducts (Figure 4C). This would indicate that probably one mechanism of CS-induced MDS is p-BQ-induced modification of bone marrow proteins. The formation of p-BQ-protein adduct is prevented by vitamin C (15 mg/day) (Figure 4A). p-BQ is also a redox cycling agent [42]. The reactive oxygen species (ROS) generated by redox cycling lead to the formation of protein carbonyls as well as 8-oxo-7, 8-dihydroguanosine (8-oxodG) that are associated with carcinogenesis [42]. As depicted in Figure 4D, bone marrow cells of DC-treated vitamin C-deficient guinea pig show increase in ROS at day 14 of CS-exposure, which is decreased after continuation of exposure for 21 days. The oxidative stress is further demonstrated by measuring protein carbonyl, an evidence of protein oxidation, (Figure 4E) and 8-oxodG, an evidence of DNA oxidation (Figure 4, F and G). No ROS was produced in the guinea pigs fed 15 mg vitamin C (Figure 4D), indicating that moderately high dose of vitamin C prevents CS-induced oxidative stress.


NAD(P)H: quinone oxidoreductase 1 deficiency conjoint with marginal vitamin C deficiency causes cigarette smoke induced myelodysplastic syndromes.

Das A, Dey N, Ghosh A, Das T, Chatterjee IB - PLoS ONE (2011)

Identification of p-BQ-protein adducts and detection of oxidative stress in bone marrow cells of guinea pigs.(Panel A) p-BQ protein adducts in the bone marrow cells of CS-exposed guinea pigs on day 21 in vivo. (Panel B) p-BQ protein adducts in CS-exposed guinea pigs at different time period in vivo. (Panel C) p-BQ protein adducts formed in marrow cells in vitro after incubation with p-BQ and AECS (aqueous extract of cigarette smoke), respectively; M, marker (cropped). (Panel D) ROS production in MDS guinea pigs at different time periods, as evidenced by flow cytometry. The X-axis represents the intensity of dichlorodihydrofluorescein diacetate (H2DCFDA). (Panel E) Protein oxidation as evidenced by oxyblot indicating formation of protein carbonyl. (Panel F) DNA oxidation as evidenced by the formation of 8-oxodG; upper row: green fluorescence indicates formation of 8-oxodG; lower row: stained with DAPI; (magnification 200×). (Panel G) Quantitative evaluation of 8-oxodG; * indicates significant difference from 0 and 7 days. Vit C means vitamin C.
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Related In: Results  -  Collection

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pone-0020590-g004: Identification of p-BQ-protein adducts and detection of oxidative stress in bone marrow cells of guinea pigs.(Panel A) p-BQ protein adducts in the bone marrow cells of CS-exposed guinea pigs on day 21 in vivo. (Panel B) p-BQ protein adducts in CS-exposed guinea pigs at different time period in vivo. (Panel C) p-BQ protein adducts formed in marrow cells in vitro after incubation with p-BQ and AECS (aqueous extract of cigarette smoke), respectively; M, marker (cropped). (Panel D) ROS production in MDS guinea pigs at different time periods, as evidenced by flow cytometry. The X-axis represents the intensity of dichlorodihydrofluorescein diacetate (H2DCFDA). (Panel E) Protein oxidation as evidenced by oxyblot indicating formation of protein carbonyl. (Panel F) DNA oxidation as evidenced by the formation of 8-oxodG; upper row: green fluorescence indicates formation of 8-oxodG; lower row: stained with DAPI; (magnification 200×). (Panel G) Quantitative evaluation of 8-oxodG; * indicates significant difference from 0 and 7 days. Vit C means vitamin C.
Mentions: CS contains substantial amount of p-BSQ, [15], [17] a major long-lived radical. We had shown before that p-BSQ is oxidized to p-BQ by transition metal containing proteins [14]. The amount of p-BQ calculated to be formed in the lungs from the inhaled cigarette smoke is 22.5 µg/day (Supporting Information Text S1). Using antibody against p-BQ and immunoblotting, here we show that bone marrow proteins of marginal vitamin C-deficient CS-exposed guinea pigs contain p-BQ-protein adducts (Figure 4, A and B). Also, incubation of bone marrow cells in vitro with p-BQ or aqueous extract of cigarette smoke (AECS) produced p-BQ-protein adducts (Figure 4C). This would indicate that probably one mechanism of CS-induced MDS is p-BQ-induced modification of bone marrow proteins. The formation of p-BQ-protein adduct is prevented by vitamin C (15 mg/day) (Figure 4A). p-BQ is also a redox cycling agent [42]. The reactive oxygen species (ROS) generated by redox cycling lead to the formation of protein carbonyls as well as 8-oxo-7, 8-dihydroguanosine (8-oxodG) that are associated with carcinogenesis [42]. As depicted in Figure 4D, bone marrow cells of DC-treated vitamin C-deficient guinea pig show increase in ROS at day 14 of CS-exposure, which is decreased after continuation of exposure for 21 days. The oxidative stress is further demonstrated by measuring protein carbonyl, an evidence of protein oxidation, (Figure 4E) and 8-oxodG, an evidence of DNA oxidation (Figure 4, F and G). No ROS was produced in the guinea pigs fed 15 mg vitamin C (Figure 4D), indicating that moderately high dose of vitamin C prevents CS-induced oxidative stress.

Bottom Line: Apoptosis precedes MDS but disappears later with marked decrease in the p53 protein.However, after the onset of MDS vitamin C becomes ineffective.Our results suggest that human smokers having NQO1 deficiency combined with marginal vitamin C deficiency are likely to be at high risk for developing MDS and that intake of a moderately large dose of vitamin C would prevent MDS.

View Article: PubMed Central - PubMed

Affiliation: Department of Biotechnology and Dr. B. C. Guha Centre for Genetic Engineering and Biotechnology, Calcutta University College of Science, Kolkata, West Bengal, India.

ABSTRACT

Background: The etiology of myelodysplastic syndromes (MDS) is largely unknown. Exposure to cigarette smoke (CS) is reported to be associated with MDS risk. There is inconsistent evidence that deficiency of NAD(P)H-quinone: oxidoreductase 1 (NQO1) increases the risk of MDS. Earlier we had shown that CS induces toxicity only in marginal vitamin C-deficient guinea pigs but not in vitamin C-sufficient ones. We therefore considered that NQO1 deficiency along with marginal vitamin C deficiency might produce MDS in CS-exposed guinea pigs.

Methodology and principal findings: Here we show that CS exposure for 21 days produces MDS in guinea pigs having deficiency of NQO1 (fed 3 mg dicoumarol/day) conjoint with marginal vitamin C deficiency (fed 0.5 mg vitamin C/day). As evidenced by morphology, histology and cytogenetics, MDS produced in the guinea pigs falls in the category of refractory cytopenia with unilineage dysplasia (RCUD): refractory anemia; refractory thrombocytopenia that is associated with ring sideroblasts, micromegakaryocytes, myeloid hyperplasia and aneuploidy. MDS is accompanied by increased CD34(+) cells and oxidative stress as shown by the formation of protein carbonyls and 8-oxodeoxyguanosine. Apoptosis precedes MDS but disappears later with marked decrease in the p53 protein. MDS produced in the guinea pigs are irreversible. MDS and all the aforesaid pathophysiological events do not occur in vitamin C-sufficient guinea pigs. However, after the onset of MDS vitamin C becomes ineffective.

Conclusions and significance: CS exposure causes MDS in guinea pigs having deficiency of NQO1 conjoint with marginal vitamin C deficiency. The syndromes are not produced in singular deficiency of NQO1 or marginal vitamin C deficiency. Our results suggest that human smokers having NQO1 deficiency combined with marginal vitamin C deficiency are likely to be at high risk for developing MDS and that intake of a moderately large dose of vitamin C would prevent MDS.

Show MeSH
Related in: MedlinePlus