Serum homocysteine concentrations of Chinese intellectuals and the influential factors concerned

doi:10.4236/health.2009.12014

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Vol.1, No.2, 83-87 (2009)

doi:10.4236/health.2009.12014

SciRes

Health

Serum homocysteine concentrations of Chinese

intellectuals and the influential factors concerned

Yue Hou*, Yan Hong, Wei-Qiang Chen, Dong-Lan Wang, Yi-Yong Cheng

*Institution of Hygiene and Environmental Medicine, Tianjin, China; Corresponding author: [email protected]

Received 23 July 2009; revised 6 August 2009; accepted 7 August 2009.

ABSTRACT

Objective: To observe the concentration of se-

rum homocysteine in intellectuals and the re-

lated influential factors. Methods: The concen-

trations serum homocysteine and saliva cortisol

were measured in 138 intellectuals from three

cities, Tianjin, Guangzhou and Chengdu in

China. All the subjects had senior titles of tech-

nical post, aged 40-69 years. Results: The mean

value of serum homocysteine concentration in

intellectuals was 20.6±0.8μmol/L, higher than

the reference value. With the increase of cortisol

levels the homocysteine concentrations rise (P

<0.05). The mean value of homocysteine con-

centration was highest in 40-49 years old group.

Men had higher homocysteine level than women

in this investigation. According to the mean

value of homocysteine concentration among

different cities, Tianjin was highest, Chengdu

medium, Guangzhou lowest. Conclusion: The

serum homocysteine concentration of intellec-

tuals is higher than the reference value. The

stress level, gender and resident cities might

contribute to the differences in serum homo-

cysteine concentration in Chinese intellectuals.

Keywords: Homocysteine; Psychological Stress;

Intellectual

1. INTRODUCTION

Stress induces several physiological and behavioral al-

terations that increase cardiovascular morbidity and

mortality [1-3]. It is well known that total plasma ho-

mocysteine (tHcy) is now established as a clinical risk

factor for coronary artery disease, as well as other arte-

rial and venous occlusive disease in adult populations [4].

Nonetheless, only a few studies have evaluated the rela-

tionship between stress and tHcy level. In order to ex-

amine whether any acute effect on total plasma Homo-

cysteine (Hcy) concentrations in rats would occur four

distinct acute stressors in rats, i.e., swimming, restrain,

novelty and cold exposure were used. Plasma corticos-

terone and adrenocorticotropic hormone concentrations

were also measured to demonstrate the ability of the

chosen manipulations to activate the hypothalamic-pi-

tuitary-adrenal (HPA) axis. Three of the four stressors

activated the HPA axis and only restrain increased tHcy

concentrations [5]. Stoney et al.’s study was to test if

acute psychological stress could induce elevations in

plasma Hcy concentrations. Thirty-four healthy women

participated in this study. The results indicated signifi-

cant elevations in plasma Hcy during acute psychologi-

cal stress, with a return to baseline concentrations during

recovery [6]. The complexity of the physiological re-

sponses to stress, the peculiarities of stress responses and

the intricate regulatory systems involved in Hcy metabo-

lism must be taken into account in order to clarify the

increasing effect of restrain (mainly a psychological

stressor) on total plasma Hcy in rats and to evaluate its

meaning in human pathology.

Stress is one of the biggest problems faced by intel-

lectuals today. The increasingly demanding nature of

their jobs has also increased pressure levels dramatically.

Research shows that intellectuals are now facing greater

day-to-day problems with occupational stress than most

other employees. Psychological stressors and depressive

and anxiety disorders also are associated with psycho-

somatic disease such as heart disease, hypertension and

diabetes, which have become the main factors impairing

the intellectuals’ health [7-10]. The intellectuals were

selected from different resident cities in China as sub-

jects and their serum Hcy levels and some influential

factors were observed.

2. MATERIALS AND METHODS

2.1. Subjects

One hundred and thirty-eight volunteers between the

ages of 40 and 69 years (mean age= 54.2 years) partici-

pated in the study after providing written informed con-

sent. The 138 intellectuals come from three cities, Tianjin,

Y. Hou et al. / HEALTH 1 (2009) 83-87

Openly accessible at

Table 1. The distribution of subjects by area, age and sex.

40-49(y) 50-59(y) 60-69(y)

City

Male Female Male Female Male Female

Total

Tianjin 8 8 8 8 8 8 48

Guangzhou 5 5 5 5 5 5 30

Chengdu 10 10 10 10 10 10 60

Total 46 46 46 138

Guangzhou and Chengdu, and all the subjects had senior

titles of technical post in university or science research

institute or design institute. The distribution of partici-

pants is presented in Table 1.

2.2. Blood and Salivary Collection

The fasting blood samples were collected into chilled

tubes, and immediately immersed in ice. The serums

were separated by centrifugation within 30-min and

stored at -20℃ until analyses. The volunteers collected

the saliva samples in the morning. Firstly they gargled

three times, after 15 minutes they chaw on citric acid

slips for a few seconds to stimulate salivation (not use

gum or any other food-type product) and collected the

salivary with straw. The samples stored at -20℃ within

three hours and centrifuged before analysis [11,12].

2.3. Laboratory Analysis

Total Hcy was measured in serum by the Total-Homo-

cysteine Enzymatic Assay method [13]. The principle of

the assay is as follows. In reaction I, rHCYase specifi-

cally converts Hcy to α-ketobutyrate, ammonia, and H2S.

In reaction II, the H2S combines with DBPDA to form

3,7-bis (dibutyl amino) phenothiazine–5-ium chlorides,

which is highly fluorescent. For the reduction reaction,

flat-bottomed 96-well cell culture cluster plates with

low-evaporationlids were used (cat. no. 0720089; Corn-

ing). Twenty microliters of serum samples and 170 uL of

assay buffer [40mmol/L sodium phosphate buffer (pH

8.4), containing 2 mL/L Triton X- 100 and 0.25 mmol/L

DTT] were added to the wells. Two wells were used for

each sample. Reduction reaction was carried out at 37

°C for 30 min. It is necessary to break the disulfide

linkages in plasma proteins and to reduce the low mo-

lecular weight disulfides Hcy and homocysteine-cysteine

mixed disulfide in this reaction. For each sample, we

added 30 uL of rHCYase (0.05 g/L), the equivalent to 0.1

U, in 40mmol/L potassium phosphate buffer containing

20μmol/L pyridoxal phosphate. One unit of enzyme is

defined as the amount that catalyzes 1 μmol of H2S per

minute from Hcy. We added 30 uL of enzyme buffer to

the other well to serve as background. The enzymatic

reaction was carried out at room temperature for 5 min.

The lid was put on the dish to prevent possible loss of

H2S. The enzymatic reaction was stopped by the addition

of 30 uL of chromophore reagent (20 mmol/L DBPDA

in 3 mol/L H2SO4). The chromogenic reaction was car-

ried out at room temperature for 10 min. The resulting

fluorescence was measured at an excitation wavelength

of 665 nm and an emission wavelength of 690 nm in

Hcy special fluorescence spectral-photometer (JD Bio-

tech Co., Ltd.).

Cortisol level was measured with a RIA kit, a product

of Isotope Institute, Chinese Atomic Energy Science

Academy [14].

2.4. Statistical Analysis

Continuous variables were summarized as the mean ±

SE. One-way ANOVA were used to compare group

means. Statistical significance was defined as P < 0.05.

Subjects with missing data were dropped from the

analysis.

3. RESULTS

129 saliva samples were collected. The Hcy concentra-

tions and the cortisol levels of all subjects were com-

pared (see Figure 1). A significant relation between Hcy

concentrations and the cortisol levels was observed

(r=0.21, P<0.05). With the increase of cortisol levels the

Hcy concentrations rise.

The average level of salivary cortisol was 10.3μg/L. The

subjects was divided into two groups, low cortisol group

(LCor, <10.3μg/L) and high cortisol group (HCor, ≥10.3

μg/L). As expected the HCor had signify cantly higher se-

rum concentrations of Hcy than the LCor (see Table 2).

02040

Cortisol(ng/ml)

Hcy(uM/L)

Figure 1. The correlation plots of serum Hcy and cortisol

concentrations.

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Y. Hou et al. / HEALTH 1 (2009) 83-87

http://www.scirp.org/journal/HEALTH/

Openly accessible at

Table 2. Saliva cortisol and serum homocysteine concentra-

tions in LCor and HCor groups (



Group n Cortisol（μg/L） Hcy（μmol/L）

LCor 77 5.5±0.3 18.4±0.8

HCor 52 18.1±1.3 22.0±1.7

Total 129 10.3±0.7 20.3±0.9

P<0.05

Table 3. Comparison of serum homocysteine concentration by

age(

s).

Age（y） n Hcy（μmol/L）

40-49 46 23.2±1.6 a

50-59 46 19.0±1.4 b

60-69 46 19.5±1.2

Total 138 20.6±0.8

Values with different superscripts are significantly different (P<0.05)

Table 4. Differences of serum homocysteine concentrations

between male and female(

s).

Sex n Hcy（μmol/L）

Male 69 24.0±1.5

Female 69 16.9±0.6

P<0.05

Table 5. Serum homocysteine concentrations of subjects in

three cities(

s).

City n Hcy（μmol/L）

Tianjin 48 22.4±1.5a

Guangzhou 30 17.1±2.3b

Chengdu 60 20.5±1.0

Values with different superscripts are significantly different (P<0.05)

The mean value of serum Hcy concentration in intel-

lectuals was 20.6±0.8μmol/L. The mean value of Hcy

concentration was highest in 40-49 years old and lowest

in 50-59 years old. There was significant difference be-

tween 40-49 years group and 50-59 years group (see

Table 3).

Among the subjects the average Hcy level of men was

significant higher than women (see Table 4).

The average Hcy levels among the three cities were

compared. The average Hcy level of the volunteers in

Tianjin was highest, medium in Chengdu, lowest in

Guangzhou. But only significant difference was ob-

served between Tianjin and Guangzhou (see Table 5).

4. DISCUSSION

Hcy is a sulfur-containing amino acid generated through

the demethylation of methionine. The resulting Hcy is

either remethylated to methionine through methionine

synthase (MS), a process that utilizes folate and vitamin

B12 as cofactors, or catabolized by transsulfuration into

if excess Hcy is present, using vitamin B6 as cofactor

[15]. It is also controlled by individual genetic differ-

ences in how vitamins are utilized as cofactors in the

reactions controlling Hcy metabolism. In excess quanti-

ties, Hcy is thought to be thrombophilic and to damage

the vascular endothelium. Total plasma homocysteine

(tHcy) is now established as a independent risk factor for

coronary artery disease, as well as other arterial and ve-

nous occlusive disease in adult populations [4,16-18].

The third National Health Examination Surv

cystathionine through cystathionine-β-synthetase (CBS)

ical stress

owed the first data on Hcy concentration in a nation-

ally representative sample of Americans. The Survey

measured serum total Hcy concentrations for a nation-

ally representative samples of 3766 males and 4819 fe-

males aged ≥12 y. The normal total plasma Hcy concen-

trations of Americans was in the range of 5μmol/L to

15μmol/L and affected by age, sex and race [19]. Yan et

al. reported that Chinese average plasma Hcy level ref-

erence was (9.67±3.0) μmol/L, 95% confidence interval

was 3.79μmol/L to 15.55μmol/L. Zhang et al. investi-

gated 1226 middle- and old-age people and the average

plasma Hcy level was 9.1μmol/L [20]. The present study

reflected that the average serum Hcy level of the intel-

lectuals aged 40-69 in Tianjin, Guangzhou and Chengdu

was higher than the level other study showed.

It is now generally accepted that psycholog

a multi-dimensional and multi-level phenomenon that

is influenced by personal, situational or structural factors.

Studies of occupational stress indicate that workload and

communications are significant causes. Anticipation,

worry, helplessness and executive roles have all emerged

from laboratory studies as psychological factors induc-

ing stress. Several studies concur in finding that poor

working conditions, especially relationships with col-

leagues, overload, and poor school ethos are the major

causes of intellectual stress. There is considerable evi-

dence, mainly from self-reports, that intellectuals feel ill

as a consequence of excessive stress. However, available

absence and retirals statistics are not sufficiently specific

to support this connection [8]. Generally cortisol level

was recognized to be an index judging the degree of

stress [11,12]. Our study showed that the serum Hcy

concentration of high saliva cortisol concentration group

was significant higher than low saliva cortisol concen-

tration group. The result indicated that psychological

stress could be one of the reasons causing the increase of

serum Hcy level and the stress degree may connect with

the level of serum Hcy. Stoney et al.’s study was to test

the hypothesis that acute psychological stress induces

elevations in plasma Hcy concentrations. Thirty-four

healthy women participated in the study. The psycho-

logical stressors included standard mental arithmetic and

speech stressors and heat rate and blood samples were

also monitored. Results indicated significant elevations

in plasma Hcy during acute psychological stress, with a

Y. Hou et al. / HEALTH 1 (2009) 83-87

http://www.scirp.org/journal/HEALTH/

ical variables influ-

onstrated that men had higher

onstrated that the average serum Hcy

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intellectuals is higher than that of normal people. The

stress level, gender and resident cities might contribute

to the differences in serum homocysteine concentration

in Chinese intellectuals. The epidemiological study

showed that Hcy is not only pro-atherogenic and

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levels may cause brain injury and neuropsychiatric dis-

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very important to handle and improve the health condi-

tion of intellectuals. To verify the exact relation between

psychological stress and the difference of Hcy level,

much more study and intervention experiment should be

conducted in the future.

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