Idiopathic Macrocytosis in Smokers

Transcription

1 ORIGINAL ARTICLE Ningen Dock 27 : , 2012 Idiopathic Macrocytosis in Smokers Masuhiro Takahashi 1), Akie Yamahira 1), Takayoshi Uchiyama 1), Minami Iwabuchi 1), Naoya Sato 1), Jun Takizawa 2), Hirohito Sone 2), Miwako Narita 1) Abstract Objective: Smoking has been reported to be one of the risk factors of myelodysplastic syndrome (MDS). In order to clarify the process of progression to MDS in a certain group of smokers, it is important to identify the initial hematological abnormalities associated with the early stage of MDS in smokers. Methods: Relationships of hematological parameters such as WBC count, RBC count, Hb, Ht, MCH and MCV with level of smoking were investigated by univariate and multivariate analyses in 234 male workers at a single workplace after obtaining informed consent. Results: Univariate analysis demonstrated that correlations of smoking-related factors (number of cigarettes per day, duration of smoking [years] and smoking index [defined as number of cigarettes per day x duration of smoking]) with WBC count, Ht, MCH and MCV were significantly positive. Multivariate analysis using age, smoking-related factors, alcohol intake and BMI as explanatory valuables showed that age, alcohol intake and BMI were not confounding variables for smoking-related factors and revealed a remarkable significant positive correlation between each smoking-related factor and MCV. Conclusion: Smoking-associated macrocytosis, which was demonstrated in the present study, was not due to deficiency of vitamin B 12 or folic acid because of no pancytopenia observed but was considered to be a qualitative abnormality of erythrocytes. Taken together with the findings of previous studies concerning smoking-associated functional abnormalities of erythrocytes and leukocytes and the high incidence of MDS in smokers, our results indicate that idiopathic macrocytosis in smokers could be an initial sign of progression to MDS, in which macrocytosis and multi-lineage abnormalities are characteristic findings. Keyword smoking, leukocytosis, macrocytosis, myelodysplastic syndrome Smoking is believed to be associated with elevation of white blood cell (WBC) counts and hematocrit. Besides, smoking has been demonstrated to accelerate arteriosclerosis, which causes obstructive disorders of blood vessels 1). Myelodysplastic syndrome (MDS) is a hematopoietic stem cell disorder characterized by dysplasia in one or more cell lineages and increased risk of developing acute myeloid leukemia (AML) 2). MDS covers hematological disorders ranging from the state close to aplastic anemia to that next to acute myelogenous leukemia. MDS is classified into 5 categories (refractory anemia (RA), RA with ring sideroblasts (RARS), RA with excess of blasts (RAEB), chronic myelomonocytic leukemia (CMML) and RAEB in transformation (RAEB in T)) in the FAB classification 3) and into 7 categories (refractory cytopenia with unilineage dysplasia (RCUD), RARS, RA with multilineage dysplasia (RCMD), RAEB, MDS associated with isolated del( 5 q), MDS unclassified (MDS-U), RC of childhood) in the 2008 revision of the WHO classification 4). As for the etiological role of smoking in myelodysplastic syndrome (MDS), several studies have found a significant association between ever smoking and occurrence of MDS 5-10), and the risk of MDS was demonstrated to increase with duration of smoking 7). On the other hand Antilla et al reported that most elderly patients with idiopathic macrocytosis who did not fulfill the FAB criteria for MDS had at least none of the following abnormalities of hematological stem cell differentiation: aberrant DNA-methylation, abnormal peripheral erythroid progenitor cell colony formation, and chromosomal abnormalities. They suggested that idiopathic macrocytosis might represent early phases of stem cell disorders such as MDS 11 ). When Dohy et al analyzed the clinical and hematological features of secondary MDS developing subsequent to chemotherapy and radiotherapy for hematological and nonhematological disorders, they found that some patients exhibited macrocytosis before the onset of severe 1) School of Health Sciences, Faculty of Medicine, Niigata University 2) Division of Hematology, Endocrinology and Metabolism, Department of internal Medicine Faculty of Medicine, Niigata University Contact : 2-746, Asahimachi, Chuo-ku, Niigata Japan Tel & Fax : matak@clg.niigata-u.ac.jp 19 ( 689 )

2 anemia and leukemia, which indicates that idiopathic macrocytosis could be an initial sign of hematological neoplasms including MDS 12). These findings imply that some of idiopathic macrocytosis could progress to MDS and further to acute myelogenous leukemia. We investigated the effects of smoking on WBC counts, red blood cell (RBC) counts and RBC indexes (MCH and MCV) in 234 male workers at a single workplace. WBC counts increased in smokers according to the number of cigarettes smoked and the duration of smoking. Although RBC count was not related with smoking in the present study, hemoglobin (Hb) and hematocrit (Ht) were positively correlated with level of smoking, which resulted in high in smokers. The idiopathic macrocytosis seen in the smokers might be a sign of transition to MDS since idiopathic macrocytosis is frequently an initial feature of it. Materials and Methods Ninety-eight male non-smokers and 136 male smokers (total 234 males) at a single workplace (oil refining company) were enrolled in the present study after obtaining informed consent. The 98 non-smokers included 26 past-smokers, who had quit smoking before a year from the beginning of the study, and three past-smokers, who had quit smoking between 3 months and 1 year before the beginning of the study. The 29 past-smokers were assigned to a non-smoker group, which prevented an imbalance in the number of the persons in the smoker and non-smoker groups. In addition, if there were a difference in a particular parameter between a smoker group and a non-smoker group including past-smokers, the difference between smokers and true non-smokers would presumably be more definite. The 234 males were analyzed after confirming that they had no definite disorders through the following methods: responses to a questionnaire, interview and physical examination by a doctor, urinalysis, hematological examination, biochemical examination, chest and esophagus/stomach radiography, and electrocardiograms. The age range was years old (yo) (mean: 45.3 yo, median: 46 yo) for nonsmokers and yo (mean: 42.9 yo, median: 43.5 yo) for smokers. The number of cigarettes smoked in a day by smokers was 5-60 (mean: 21.8, median: 20), the duration of smoking was 2-40 years (mean: 22.5 years, median: 22 years) and the smoking index, which was defined as number of cigarettes a day x duration of smoking, was (mean: 505, median: 445). WBC counts, RBC counts, Hb, Ht, MCH, MCV and WBC differentials were measured using an automated blood cell analyzer (Sysmex, Kobe, Japan). The significances of differences in WBC counts, RBC counts, Hb, Ht, MCH, and MCV between smokers and non-smokers were evaluated using the Student s t-test. Correlations between smoking-related factors and WBC counts, RBC counts, Hb, Ht, MCH, or MCV were evaluated by making scatter diagrams. The correlation coefficient (r) and significance level (p) was calculated using Spearman s nonparametric rank statistic by means of PRISM software (GraphPad, La Jolla, CA, USA). p<0.05 was defined as significant. Multiple regression analyses using WBC count, RBC count, Hb, Ht, MCH, or MCV as response variables and age, smoking-related factors, alcohol intake and BMI as explanatory variables were performed using Mac Multiple Regression Analysis (Esumi, Tokyo, Japan). Results WBC counts in smokers (6,890 ± 1,820/ μ L: mean ± SD) were significantly increased compared with those in non-smokers (5,440 ± 970/ μ L) (p<0,001). There were no differences in RBC counts between smokers (4.88 x10 6 ± 0.34 x10 6 / μ L) and non-smok- Table 1. Comparison of Smoking-related and Hematological Parameters between Non-smokers and Smokers parameter statistics non-smokers smokers number of subjects age (years old) number of cigarettes smoked per day duration of smoking (years) smoking index* alcohol (g) drunk per day BMI WBC count (/ μ L) RBC count (x10 6 / μ L) Hb (g/dl) Ht % MCH pg MCV (fl) Student's t-test range mean ± SD 45.3 ± ± 8.1 p=0.019 median range mean ± SD ± 8.9 median 0 20 range mean ± SD ± 8.2 median 0 22 range mean ± SD ± 301 median range mean ± SD 31.6 ± ± 20.0 p=0.040 median range mean ± SD 23.8 ± ± 3.2 p=0.531 median range mean ± SD 5440 ± ± 1820 p<0.001 median range mean ± SD 4.89 ± ± 0.34 p=0.838 median range mean ± SD 15 ± ± 0.9 p=0.001 median range mean ± SD 45.7 ± ± 2.6 p<0.001 median range mean ± SD 30.6 ± ± 1.6 p<0.001 median range mean ± SD 93.4 ± ± 4.3 p<0.001 median *number of cigarettes per day x duration of smoking (years) 20 ( 690 )

3 Takahashi, et al. : Idiopathic Macrocytosis in Smokers ers (4.89x10 6 ± 0.37x10 6 / μ L) in the present study. Hb in smokers (15.4 ± 0.9 g/dl) was significantly increased compared with that in non-smokers (15.0 ± 0.9 g/dl) (p=0,001) and Ht in smokers (47.4±2.6 %) was also significantly increased compared with nonsmokers (45.7 ± 2.6 %) (p<0.001). MCH in smokers (31.9 ± 1.6 pg) was significantly increased compared with non-smokers ( ± 1. 7 pg) (p< 0, 001 ) and MCV in smokers (97.6 ± 4.3 g/dl) was also significantly increased compared with non-smokers ( ± 4. 7 g/dl) (p< 0, 001 ) (Table 1 ). Regarding BMI, there was no difference between non-smokers (23.8 ± 3.8) and smokers (23.5 ± 3.2). The amount of alcohol drunk per day was not statistically different between non-smokers (32.68 ± 1.94 g, converted into pure alcohol) and smokers ( ± g) (p= ). Correlations between individual smoking parameters and WBC count were significant: r=0.474 (p<0.0001) for number of cigarettes per day, r=0.441 (p< ) for duration of smoking (years), and r=0.481 (p<0.0001) for smoking index (Fig. 1). Correlations between number of cigarettes per day, duration of smoking or smoking index and RBC count were not significant, though there was a tendency towards a negative correlation (Fig. 2 ). Correlations of smoking parameters with Hb were r=0.204 (p=0.0029) for number of cigarettes per day, r=0.179 (p= ) for duration of smoking, and r= (p= ) for smoking index (Fig. 3 ). Although correlation coefficients between Ht and smoking parameters were not high ( ) correlations were significant for all of them: r=0.245 (p=0.0004) for number of cigarettes per day, r=0.232 (p=0.0008) for duration of smoking, and r=0.229 (p=0.0010) for smoking index (Fig. 4). Correlations of all smoking parameters with MCH were significant with r=0.348 (p<0.0001) for number of cigarettes per day, r=0.384 (p< ) for duration of smoking, and r= (p<0.0001) for smoking index (Fig. 5). Correlations of all smoking parameters with MCV were remarkably significant with r=0.406 (p<0.0001) for number of cigarettes per day, r=0.457 (p<0.0001) for duration of smoking, and r=0.461 (p<0.0001) for smoking index (Fig. 6). WBC differentials in eight smokers, whose degree of leukocytosis was 10, 725 ± 819 / μ L (mean ± SD, n=8), were 2.4 ± 1.5% stab neutrophils, 45.9 ± 4.6% segmented neutrophils, 44.1 ± 2.9% lymphocytes, 4.9 ± 1.7% monocytes, 2.6 ± 2.3% eosinophils and 0.3 ± 0.5% basophils. There was no increase in specific Fig. 1. Scatter Diagrams between Smoking-related Factors and WBC Fig. 2. Scatter Diagrams between Smoking-related Factors and RBC 21 ( 691 )

4 Fig. 3. Scatter Diagrams between Smoking-related Factors and Hb Fig. 4. Scatter Diagrams between Smoking-related Factors and Ht Fig. 5. Scatter Diagrams between Smoking-related Factors and MCH WBC subpopulations in smokers with leukocytosis, which suggested that the predominant WBC subpopulations, segmented neutrophils and lymphocytes, were mainly elevated by smoking (Table 2). Multiple regression analyses using WBC count, RBC count, Hb, Ht, MCH, and MCV as response variables and age, smoking-related factors (number of cigarettes per day, duration of smoking or smoking index), alcohol intake and BMI as explanatory variables were performed. For the response variable of WBC count, the only significant explanatory variable was smokingrelated factors. For the response variable of RBC count, age and alcohol intake were significant explanatory factors, with negative standardised partial regression coefficients. For the response variable of Hb, there was a significant negative standardized partial regression coefficient with age and a significant positive standardised partial regression coefficient with smoking-related 22 ( 692 )

5 Takahashi, et al. : Idiopathic Macrocytosis in Smokers Fig. 6. Scatter Diagrams between Smoking-related Factors and MCV Nnumber of cigarettes smoked in a day, duration of smoking (years) and smoking index, which was defined as number of cigarettes per day x duration Table 2. WBC Differentials in Eight Smokers with Leukocytosis smoker (No) age (years old) number of cigarettes smoked per day duration of smoking (years) smoking index* WBC counts (/ μ L) stab neutrophils segmented neutrophils lympho-cytes monocytes eosinophils basophils mean ± SD 47 ± ± ± ± ± ± ± ± ± ± ± 0.5 *number of cigarettes per day x duration of smoking (years) factors (number of cigarettes per day and duration of smoking). For the response variable of Ht, there was a significant negative standardised partial regression coefficient with age and a significant positive standardised partial regression coefficients with all smoking-related factors (number of cigarettes per day, duration of smoking and smoking index). For the response variables of MCH and MCV, there were positive standardised partial regression coefficients with all smoking-related factors and alcohol intake and significance was remarkable. Regarding levels of significance for the response variable MCV, the p-value for all smoking-related factors was but that for alcohol intake was These data show that the correlation of smoking-related factors with MCV was more significant than that of alcohol intake, which was supported by higher values of standardised partial regression coefficient and F-values in smoking-related factors compared with alcohol intake. In all multiple regression analyses in the present study, the VIF (variance inflation factor) was below 1.1 for each of the 4 explanatory variables, which showed that there was no correlation among age, each of smoking-related parameters, alcohol intake and BMI (Table 3). These findings show that age, alcohol intake and BMI are not confounding variables for smokingrelated factors (number of cigarettes per day, duration of smoking or smoking index). Discussion We investigated a relationship between smoking and hematological findings in male workers at a single workplace. More than sixty percent of them had a smoking habit at the time of the present study. We did not have a separate category for past history of smoking, so workers who had smoked before but were not smoking at the time of the study were included in the non-smoker group. Despite this, WBC counts, Hb, Ht, MCH and MCV were significantly correlated with the level of smoking. Although it is clear from previous studies that WBC counts are elevated by smoking 13-14), the mechanism of smoking-associated leukocytosis has not been fully elucidated. It has been speculated that leukocytosis is at least partly due to chronic bronchitis caused by smoking 15), but no subjects with chronic bronchitis were enrolled in the present study. In this regard, catecholamine, which levels are significantly higher in the plasma of smokers than that of non-smokers 16), is presumed to release leukocytes from the marginal pool to the circulating pool and give rise to peripheral leukocytosis. However, cotinine, a metabolite of nicotine, has been reported to suppress the growth of hematopoietic progenitor cells at a concentration range similar to its serum levels in smokers 17). In addition to generating leukocytosis, smoking has been reported to induce a 23 ( 693 )

6 Table 3. Multiple Regression Analysis using WBC, RBC, Hb, Ht, MCH or MCV as Response Variable and Age Smoking-related Factors Alcohol Intake and BMI as Explanatory Variables response variable WBC RBC Hb Ht MCV MCH WBC RBC Hb Ht MCV MCH WBC RBC Hb Ht MCV MCH explanatory variable and partial regression standardised partial VIF (variance F-value p-value evaluation standard error constant term coefficient regression coefficient inflation factor) age no.cigar/day ** alcohol (g)/day BMI constant term ** age ** no. cigar/day alcohol (g)/day * BMI constant term ** age ** no. cigar/day * alcohol (g)/day BMI constant term ** age * no. cigar/day ** alcohol (g)/day BMI constant term ** age no. cigar/day ** alcohol (g)/day ** BMI constant term ** age no. cigar/day ** alcohol (g)/day ** BMI constant term ** age years of smoking ** alcohol (g)/day BMI constant term ** age ** years of smoking alcohol (g)/day * BMI constant term ** age ** years of smoking ** alcohol (g)/day BMI constant term ** age ** years of smoking ** alcohol (g)/day BMI constant term ** age years of smoking ** alcohol (g)/day ** BMI constant term ** age years of smoking ** alcohol (g)/day ** BMI constant term ** age index (no. cigar x years) ** alcohol (g)/day BMI constant term ** age * index (no. cigar x years) alcohol (g)/day * BMI constant term ** age ** index (no. cigar x years) alcohol (g)/day BMI constant term ** age ** index (no. cigar x years) ** alcohol (g)/day BMI constant term ** age index (no. cigar x years) ** alcohol (g)/day ** BMI constant term ** age index (no. cigar x years) ** alcohol (g)/day ** BMI constant term ** p<0.05: significant (*), p<0.01: remarkably significant (**) 24 ( 694 )

7 Takahashi, et al. : Idiopathic Macrocytosis in Smokers reduction in leukocyte deformability 18) and an increase in leukocyte aggregation 19), and enhance adhesion of circulating leukocytes to the endothelium in a hamster model 20). These effects of smoking, together with leukocytosis, were thought to be associated with the causative mechanism of smoking-induced arteriosclerosis and veno-occlusive disease. Further, Nobel et al have reported that leukocyte chemotaxis was depressed in smoking subjects when compared to non-smokers. Although RBC counts tended to be decreased by smoking in the present study, Hb and Ht were significantly increased, which resulted in a remarkable increase in MCH and MCV due to smoking. The smoking-induced increase in Ht is thought to be due to enhanced erythropoiesis caused by excessive secretion of erythropoietin from kidney interstitial cells, which have perceived smoking-associated oxygen deficiency, as well as a decrease in circulating plasma volume 21). If this theory is correct, as elevated erythropoietin would stimulate marrow erythropoiesis, there should be a greater increase in RBC counts than in Hb or Ht unless there is deficiency of vitamin B 12 or folic acid. Further an increase in RBC counts due to excessive erythropoietin secretion would result in a decrease in MCH and MCV due to an iron deficient state resulting from erythroid hyperplasia. However, RBC counts actually tend to decrease with a remarkable increase in MCH and MCV in smokers. Although previous studies have observed an increase in Hb and Ht in smokers, RBC counts have not been included in analyses on a relationship between smoking and hematological abnormalities. Smokingassociated RBC abnormalities may consist of both quantitative abnormalities caused by elevated erythropoietin with a decreased plasma volume and qualitative abnormalities like an increase in MCH and MCV (macrocytosis) without an increase in the number of RBC. Smoking has been reported to impair erythrocyte filterability (deformability), which seems to be a sign of qualitative abnormalities induced by smoking. Qualitative abnormalities in smokers could be observed not only in erythrocytes but also in leukocytes, which are exemplified by a reduction in leukocyte deformability 18 ) and an elevation in leukocyte aggregation 19). Also qualitative RBC abnormalities due to smoking, such as a marked increase in MCH and MCV, and the significantly higher incidence of MDS in smokers, as was found by the present study, may support the hypothesis that idiopathic macrocytosis in smokers could be an initial sign of progression to MDS. Conflict of Interest The authors declare no conflict of interest. References 1) Helman N, Rubenstein LS: The effects of age, sex, and smoking on erythrocytes and leukocytes. Am J Clin Pathol 1975; 63: ) Cazzola M, Della Porta MG, Travaglino E, et al: Classification and prognostic evaluation of myelodysplastic syndromes. Semin Oncol 2011; 38: ) Bennett JM, Catovsky D, Daniel MT, et al: Proposals for the classification of the myelodysplastic syndromes. Br J Haematol 1982; 51: ) V Vardiman JW, Thiele J, Arber DA, et al: The 2008 revision of the World Health Organization (WHO) classification of myeloid neoplasms and acute leukemia: rationale and important changes. Blood 2009; 114: ) Du Y, Fryzek J, Sekeres MA, et al: Smoking and alcohol intake as risk factors for myelodysplastic syndromes (MDS). Leuk Res 2010; 34: ) Niss e C, Hagueno er JM, Grandbastien B, et al: Occupational and environmental risk factors of the myelodysplastic syndromes in the North of France. Br J Haematol 2001; 112: ) Björk J, Johansson B, Broberg K, et al: Smoking as a risk factor for myelodysplastic syndromes and acute myeloid leukemia and its relation to cytogenetic findings: a casecontrol study. Leuk Res 2009; 33: ) Strom SS, Gu Y, Gruschkus SK, et al: Risk factors of myelodysplastic syndromes: a case-control study. Leukemia 2005; 19: ) Dalamaga M, Petridou E, Cook FE, et al: Risk factors for myelodysplastic syndromes: a case-control study in Greece. Cancer Causes Control 2002; 13: ) Ido M, Nagata C, Kawakami N, et al: A case-control study of myelodysplastic syndromes among Japanese men and women. Leuk Res 1996; 20: ) Anttila P, Ihalainen J, Salo A, et al: Idiopathic macrocytic anaemia in the aged: molecular and cytogenetic findings. Br J Haematol 1995; 90: ) Dohy H, Genot JY, Imbert M, et al: Myelodysplasia and leukaemia related to chemotherapy and/or radiotherapy- -a haematological study of 13 cases. Value of macrocytosis as an early sign of bone marrow injury. Clin Lab Haematol 1980; 2: ) Taylor RG, Woodman G, Clarke SW: Plasma nicotine concentration and the white blood cell count in smokers. Thorax 1986; 41: ) Watanabe N, Fukushima M, Taniguchi A, et al: Smoking, white blood cell counts, and TNF system activity in Japanese male subjects with normal glucose tolerance. Tob Induc Dis 2011; 9: ) James AL, Knuiman MW, Divitini ML, et al: Associations between white blood cell count, lung function, respiratory illness and mortality: the Busselton Health Study. Eur Respir J 1999; 13: ) Umene-Nakano W, Yoshimura R, Yoshii C, et al: Plasma levels of metabolites of catecholamine in nicotinedependent patients treated with varenicline. Nicotine Tob Res 2012; 14: ) Takahashi M, Shigeno N, Narita M, et al: Cotinine (a metabolite of nicotine) suppresses the growth of hematopoietic progenitor cells at the concentration range equivalent to its serum levels in smokers. Stem Cells 1999; 17: ( 695 )

8 18) Drost EM, Selby C, Bridgeman MM, et al: Decreased leukocyte deformability after acute cigarette smoking in humans. Am Rev Respir Dis 1993; 148: ) Galante A, Pietroiusti A, Polucci C, et al: Influence of acute and chronic smoking on leukocyte aggregation. Clin Physiol Biochem 1993; 10: ) Lehr HA, Kress E, Menger MD, et al: Cigarette smoke elicits leukocyte adhesion to endothelium in hamsters: inhibition by CuZn-SOD. Free Radic Biol Med 1993; 14: ) Aitchison R, Russell N: Smoking--a major cause of polycythaemia. J R Soc Med 1988; 81: (Received April 26, 2012 ; Accepted September 25, 2012) 26 ( 696 )