Effects of rotating shift work on biomarkers of metabolic syndrome and inflammation

Introduction

The major function of the circadian system is the internal cycling of physiological and metabolic events [1]. In fact, many physiological processes display day–night rhythms. Besides, feeding behaviour, lipid and carbohydrate metabolism and blood pressure are subject to daily variation.

Modern society has evolved to rely increasingly on 24-h operations in many diverse settings and as many as 20% of workers in industrialized nations are shift workers [2, 3].

The circadian rhythm and environmental conditions can become desynchronized in rotating shift workers whose night activity is out of phase with many coupled rhythms owing to desynchronization of the normal phase relationships between biological rhythms within the circadian system [4]. In addition, because individual biological rhythms re-entrain to a time shift at different rates, each time the work schedule rotates, for a period of time after the time shift, the circadian system will be in a desynchronized state [5].

Immediate disturbances associated with shift work are symptoms such as sleep disturbances and fatigue. These symptoms are often short term or related to specific phases of the work schedule. However, the symptoms sometimes reflect a more serious chronic disease process [6].

In this regard, it has been reported that the effects of rotating shift work schedule on the cardiovascular and metabolic system show that rotating shift work may be directly responsible for increased body fatness, higher blood pressure levels and some features of the metabolic syndrome (MS) [7–10]. Furthermore, in several reports rotating shift workers had a significantly higher risk of death due to ischaemic heart disease [11–14].

Interestingly, the association between shift work and coronary heart disease was thoroughly discussed in both longitudinal and cross-sectional studies by Knutsson [15] postulating three major disease pathways encompassing collision between the circadian rhythm and myocardial performance, changes in behaviour and disturbed sociotemporal rhythmicity leading to distress reactions.

Recent epidemiological studies have reported an association between inflammation and thrombogenesis as possible causes for cardiovascular events in patients suffering from the MS [16]. Additionally, several prospective studies have shown a positive association between elevated leucocyte count and risk of coronary heart disease suggesting that leucocyte count is associated with the cluster of metabolic and haemodynamic disorders typical of the MS. In fact, it was postulated that increased leucocyte count may be another component of this syndrome [17].

In the light of the above comments, the present study sought to explore the effect of rotating shift work schedule on leucocyte count and its relation with risk factors of MS.

Subjects and methods

Healthy male subjects recruited from a factory in Buenos Aires metropolitan area who underwent the annual health examination during the 2005 period were included in a cross-sectional population-based study.

A total of 1351 men of self-reported European ancestry, aged 34.4 ± 8.4 years were included in this study, in which 877 day workers were compared with 474 rotating shift workers.

The job schedule type was divided into rotating shift work and daytime work. Rotating shift work was based on two shifts and clockwise rotation in 28 days (four work days, three rest days, two work nights, three rest days, four work nights and three rest days, two work days, three rest days, four work days; see Fig. 1). Day and night work periods started at 06:00 am and 6:00 pm, respectively, and the workers were engaged in an alternating daytime work and night-time work based on 12 labour hours. Regarding day workers they work 8 h/day, 5 days a week starting between 7:00 and 8:00 am.

None of the subjects interchanged the job schedule; all the rotating shift workers were engaged in this schedule at baseline.

Medical history was investigated by using a self-administered questionnaire. In addition, the contents were confirmed by individual interviews conducted by occupational doctors.

The participants were asked to fast for at least 8 h. After a 5 minutes’ rest in a quiet room, systolic (SABP) and diastolic (DABP) arterial blood pressure were measured on the right arm with a standard mercury sphygmomanometer at sitting position. The health examinations included anthropometric measurements, a questionnaire on health-related behaviours and biochemical determinations. Information about the years of work, either shift work or day work duration and past medical history were also included. Height and weight were measured at every examination. Subjects were measured in light clothing and without shoes.

Body mass index (BMI) was calculated as kg m⁻² and was used as the index for relative weight. Additionally, waist circumference was assessed in the standing position, midway between the highest point of the iliac crest and the lowest point of the costal margin in the mid-axillary line, by trained staff. Hip circumference was measured at the level of the femoral greater trochanter by the same observer. Those with a BMI of 30 or more were classified as obese.

As for health-related behaviours, the questions about alcohol intake included items about the type of alcoholic beverage and the usual amount consumed daily. Weekly alcohol intake was calculated and then converted to daily alcohol consumption (grams of ethanol per day). Questions about current smoking habits were asked to as the number of cigarettes smoked per day.

Blood sample was drawn from the subjects between 6:00 and 9:00 am, after at least, 8 h or overnight fast in all the cases regardless of the work schedule; all the subjects were in a supine resting position for at least 30 min. Serum insulin, total cholesterol, HDL and LDL cholesterol, triglycerides, plasma glucose, leucocyte count, uric acid and erythrocyte sedimentation rate were measured by standard clinical laboratory techniques. Leukocyte count was measured automatically in a Sysmex XE 2100 equipment (Roche Diagnostics, Buenos Aires, Argentina) (normal range 6000–10000 cells μL⁻¹).

Homeostasis Model Assessment (HOMA) was used to evaluate an insulin resistance index and was calculated as fasting serum insulin (μU mL⁻¹) × fasting plasma glucose (mmol L⁻¹)/22.5.

Metabolic syndrome was defined as the presence of three of the five components (elevated levels of blood pressure, serum triglycerides, waist circumference, fasting glucose level and low HDL cholesterol) according to the National Cholesterol Education Program Adult Panel III guidelines [18]. Specifically, elevated blood pressure was defined as SABP ≥130 mmHg and/or DABP 85 mmHg or receipt of anti-hypertensive medications. Low HDL cholesterol was defined as <40 mg dL⁻¹, high serum triglycerides were defined as ≥150 mg dL⁻¹, elevated total cholesterol was defined as ≥200 mg dL⁻¹ and elevated plasma glucose as ≥110 mg dL⁻¹. We included the measure of insulin resistance as it has been shown to provide incremental information regarding atherosclerotic cardiovascular disease beyond current National Cholesterol Education Program Adult Treatment Panel III MS [19]. Estimation of insulin resistance was defined as HOMA index ≥2.4 [20]. Finally, abdominal obesity was defined as a waist circumference ≥102 cm.

Determining the 10-year risk for developing coronary heart disease outcomes (myocardial infarction and coronary death) was carried out using Framingham risk scoring. The risk factors included in the Framingham calculation are age, total cholesterol, HDL cholesterol, systolic blood pressure, treatment for hypertension and cigarette smoking (Third Report of the Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults (Adult Treatment Panel III) Executive Summary. National Cholesterol Education Program of the National Institutes of Health (NIH) National Heart, Lung, and Blood Institute).

All the investigations performed in this study were conducted in accordance with the guidelines of The Declaration of Helsinki. Written consent from individuals had been granted, in accordance with the procedures approved by the Ethical Committee of our institution.

Results

Clinical features, anthropometric variables and laboratory findings of the participants according to the job schedule are shown in Table 1. Rotating shift workers had elevated BMI, waist-hip ratio, DABP, fasting insulin, HOMA index, triglycerides and uric acid.

Additionally, the simultaneous presence of three or more metabolic risk factors (abdominal obesity, hypertension, high triglycerides and insulin resistance) was significantly more common in rotating shift workers (17.2%) in comparison with day workers (10.7%, P < 0.005). The OR for MS in rotating shift workers when compared with day workers was 1.51 (95% CI 1.01–2.25, P < 0.04), independently of age and physical activity.

Besides, leucocyte count was higher in shift workers when compared with day workers (Table 1). By logistic regression analysis we observed that amongst age, physical activity, BMI, waist-hip ratio, DABP, leucocyte count, fasting serum insulin, HOMA, triglycerides and uric acid only age (estimate ± SE: 0.92 ± 0.26, P < 0.0004), waist-hip ratio (2.54 ± 1.04, P < 0.014) and uric acid (0.7 ± 0.30, P < 0.02) are independently associated with rotating shift work.

We then decided to explore further the potential role of leucocyte count as a biomarker of inflammation in this population. Taking the population as a whole, leucocyte count significantly increased with cumulative risk factors for the MS defined according to the National Cholesterol Education Program Adult Panel III guidelines [18] (Spearman R = 0.19, P < 0.0001). Furthermore, by logistic regression analysis, leucocyte count was a risk factor of MS independently of age, educational status, smoking and physical activity (OR per cell: 1.0002, 95% CI 1.0001–1.0003, P < 0.003).

Although leucocyte count was higher in subjects with MS (P < 0.0001), rotating shift work induced an additive effect on leucocyte count (P < 0.04) (Fig. 2). Besides, in multiple regression analysis, amongst age, education status, rotating shift work, smoking, physical activity, alcohol consumption, BMI, waist, HOMA index, total and HDL cholesterol, triglycerides and hypertension, only rotating shift work, smoking, HOMA index, triglycerides and hypertension appeared to be correlated with leucocyte count (Table 2).

About the problem of colinearity amongst the independent factors we observed that the tolerance was higher than 0.80 for all the independent predictor variables except for total cholesterol and triglycerides that was higher than 0.70.

Discussion

In this study we observed that rotating shift workers had most of the risk factors of the MS, including elevated BMI, waist-hip ratio, DABP, fasting insulin, HOMA index and triglycerides. Additionally, uric acid and leucocyte count were higher in rotating shift workers when compared with day workers. Besides, rotating shift workers showed a more sedentary life style compared with day workers. This may be a consequence of their particular desynchronized life style.

It is well known that biological processes are coordinated for optional functioning of the organism by the circadian system. Rotating shift work disrupts the synchronous relationship between the body's internal clock and the environment, causing deleterious effects not only on hormonal levels but also on the immune system and activity-rest cycles. Moreover, shift work schedules have been reported to have harmful effects on biomarkers of cardiovascular disease.

Apart from performance decrements during work, rotating shift work is widely felt to give rise to widespread health concerns. This scenario of metabolic and clinical disturbances provides an explanation for the increased risk of cardiovascular events reported in this population [11, 12, 21].

In this context, it is noteworthy that, in our findings, leucocyte count was significantly increased in rotating shift workers independently of other leucocyte-increasing well-known factors such as smoking and HOMA index. Furthermore, in multiple regression analysis, only rotating shift work, smoking, HOMA index, triglycerides and hypertension appeared to be correlated with leucocyte count.

It should be noted that we may exclude the possibility that the colinearity between the predictor variables influenced the effects, as the tolerance was higher than 0.80 for almost all the independent predictor variables.

This finding, although novel in the population mentioned above, is consistent with previous observations. For instance, it was reported that increased leucocyte count, a biological marker of systemic inflammation, is associated with components of MS [17, 22]. Besides, inflammation and thrombogenesis have been suggested as possible causes for cardiovascular events in patients suffering from MS [16].

Although the relation between rotating shift work and health hazards is not a matter of debate, the mechanisms involved are still under study. However, it is reasonable to believe that the main cause of the observed clinical and metabolic disorders is the desynchronization imposed by the rotating shift work schedule and the resulting conflict with the biological rhythms. This observation was experimentally supported by a recent study that reported the development of MS in mice with altered circadian timing brought about by a mutation in a gene that functions in the biological clock [23].

Furthermore, the circadian pacemakers, which time the approximately 24-h rhythms in sleep and wakefulness, neuroendocrine, thermoregulatory and other body functions, resynchronize only slowly after an abrupt phase shift in environmental time cues [24]. In this regard, rotating night shifts are particularly disruptive for sleep, wakefulness, eating patterns and social activities.

Likewise, a recent human study suggested a supportive influence of regular nocturnal sleep on immune responses to experimental infection [25]. In fact, it was stated that immune response may be modified, and therapeutically manipulated, by circadian effector signals [26].

Finally, circadian control depends on oscillating transcription factors, master switches synchronized by stimuli such as light and feeding. Here we showed that rotating shift work affected many metabolic pathways along with a higher leucocyte count in comparison with day workers. Therefore, circadian changes in the metabolism and secretion patterns of endocrine and immunological factors may well be partly responsible for the changes observed in the inflammatory response in rotating shift workers.

Our observations were based on a cross-sectional study and all the samples were drawn from the subjects at the same time during the morning regardless of the work schedule. It may introduce a selection bias amongst the groups considering the possibility of biological circadian variation of different blood parameters.

In conclusion, a common denominator in most forms of rotating shift work is sleep deprivation, which not only leads to a loss in cognitive and physical performance, but, as others have previously reported [7–11] and we show here, has serious metabolic consequences with an increase in cardiovascular morbidity and mortality.

Because the MS is an important cause of morbidity and mortality in developed countries, and is also becoming increasingly prevalent in the developing world [27], we reinforce that special attention should be given to rotating shift workers in order to prevent cardiovascular disease. In addition, health problems in rotating shift workers are of economic and industrial importance, as they represent approximately 20% of the work force worldwide. It is therefore important to find out ways to lessen the consequences of the circadian disruption and related harmful health effects not only by pharmacological intervention but also through the implementation of chronobiotic measures to ameliorate the effects of the phase shifts.

Conflict of interest statement

No conflict of interest was declared.