Development of a bionanodevice for detecting stress levels

S Nomura1,3

, S Handri1, and H Honda

2,3

1 Top Runner Incubation Center for Academia-Industry Fusion, Nagaoka University of

Technology, 1603-1 Kamitomioka, Nagaoka 940-2188, Japan 2 Department of Bioengineering, Nagaoka University of Technology, 1603-1

Kamitomioka, Nagaoka 940-2188, Japan

Abstract. Recent advances in molecular analysis techniques have enabled scientists to assess

the tiny amounts of biochemical substances secreted in our bodies. This has revealed that the

levels of various secretory hormones and immune substances vary sensitively with the mental

state of a person. Such hormones and immune substances exhibit transient increases with

various psychological stressors. They thus have the potential to be used as a novel biometric

for monitoring stress. Biomarkers that occur in saliva can be monitored non-invasively and are

thus potentially useful as practical indicators of mental stress. Stress biomarkers are considered

to be released into the blood stream or other secretory fluids by physiological stress reactions.

Stress biomarkers are expected to be detectable in sweat and other humoral fluids that are

exuded from the skin surface. Based on this, we have developed a bionanodevice for detecting

stress by capturing stress biomarkers on the skin surface in a non-invasive manner. A prototype

bionanodevice is described in which a motor protein is introduced for molecular handling.

1. Introduction

Mental health problems represent an acute global problem. They are not just problems of individuals

but they also represent a social problem. The International Labor Organization reported various effects

of mental health problems on society; for example, it estimated the financial cost of mental health

problems in the European Union to be about 3 to 4 % of the GNP [1]. However, it is generally difficult

for individuals to monitor their own mental stress levels making it hard to manage mental stress

unaided. It is thus important to devise a practical methodology for evaluating, managing, and

controlling mental stress.

Recent advances in molecular analysis techniques allow scientists to monitor tiny amounts of

substances contained in human secretory fluids. The levels of some hormones and immune substances

in the body have been found to vary in response to a person’s mental state. For example, salivary

immunoglobulin A (IgA) transiently increases in response to short-term psychological stressors such as

mental arithmetic tasks, stroop tasks, and academic presentations [2]. Therefore, the levels of such

substances are expected to be useful as novel biometrics for stress. Several such potential stress

biomarkers have been reported in behavioural medicine and psychophysiological studies [3].

Moreover, since these substances are secreted into the blood stream or external/internal secretory

fluids through activation of physiological stress reactions, they are expected to be determinable by

analyzing sweat and other humoral fluids that are exuded from the skin. This gave us the idea of

3 Corresponding authors. E-mail: nomura@kjs.nagaokaut.ac.jp, hhonda@vos.nagaokaut.ac.jp

International Symposium on Global Multidisciplinary Engineering 2011 (S-GME2011) IOP PublishingIOP Conf. Series: Materials Science and Engineering 21 (2011) 012029 doi:10.1088/1757-899X/21/1/012029

Published under licence by IOP Publishing Ltd 1

developing a bionanodevice for detecting stress that obtains tiny amounts of stress biomarkers from

the skin surface in a non-invasive manner.

In this paper, we briefly review previous stress biomarker studies; in particular, we consider the

results of our past studies on potential stress biomarkers. We also describe a prototype bionanodevice

that employs a motor protein for molecular handling.

2. Stress biomarkers: background and example

2.1. Physiological stress reaction pathways

A wide range of secretory substances have been reported to change their levels in response to certain

stressors. These substances are considered as potential biomarkers for stress. Seven biomarkers are

currently used as stress biomarkers; they include hormones, immune substances, proteins, and

enzymes and are the following: immunoglobulin A (IgA), cortisol, chromogranin A (CgA), alpha

amylase, dehydroepiandrosterone (DHEA), dehydroepiandrosterone sulphate (DHEA-S), and

testosterone (TE) [2].

The reaction mechanisms of these substances in response to mental stress are considered in relation

to the human physiological stress reaction pathways. The two internal stress reaction pathways are the

hypothalamus-pituitary-adrenal (HPA) and sympatho-adrenal-medullary (SAM) systems [4] (Figure

1). The above seven biomarkers are considered to be released into the blood or other secretory fluids

and thus reflect the activation of one of the two reaction pathways: IgA, CgA, and amylase are

considered to reflect SAM activation, while cortisol, DHEA, and DHEA-S are considered to reflect

HPA activation.

Stress biomarker studies investigate secretory hormones and immune substances that accurately

reflect activation of the HPA or SAM system so that they can be used as objective indicators of mental

stress levels. Below, two important biomarkers in this field, IgA and cortisol, are described.

Figure 1. Stress reaction pathway and a method for detecting stress levels using a

bionanodevice.

Pituitary

ANS Center

Cerebral Cortex

Adrenal cortex

Adrenal Medulla

Hypothalamus

DHEA Cortisol

N, NE

IgA Amylase

Stressor

SA

M s

ystemH

PA

System

Bio-nanodevicetoward stress detection

Specimen collectionOn-site analysis

International Symposium on Global Multidisciplinary Engineering 2011 (S-GME2011) IOP PublishingIOP Conf. Series: Materials Science and Engineering 21 (2011) 012029 doi:10.1088/1757-899X/21/1/012029

2

2.2. Possible stress biomarkers: immunoglobulin A (IgA) and cortisol

Immunoglobulin A (IgA) is one of the most important substances in the human immune system. IgA

functions non-specifically and thus plays a very important role in our health (e.g., inhibiting

pathogenic viruses from entering epithelial cells). Thus, salivary IgA is frequently referred to as our

first line of defense against influenza or other respiratory tract infections. Behavioral medicine studies

conducted in the 1970s revealed that salivary IgA levels vary in response to various psychological

factors, including desirable and undesirable daily events, daily hassles, negative and positive moods,

academic stress, stressful short-term cognitive tasks, and relaxation [5]. We observed temporarily

elevated IgA levels in university students who were studying for their final examinations [6] (Figure

2).

(a)

(b)

Figure 2. (a) Profile of mood states (POMS; a psychological scale) and (b) IgA

concentrations of undergraduate university students before (day I) and after (day

II) their final examinations (from [5]). Within-subjects design (n = 20) was

employed. POMS consist of six mood factors: tensionanxiety (TA),

depressiondejection (D), angerhostility (AH), vigor (V), fatigue (F), and

confusion (C)

0

5

10

15

20

25

30

T-A D A-H V F C

Day I

Day II

Sco

res

of

PO

MS

***

**

*****

**

*** p<0.001

** p<0.01

0

50

100

150

200

250

Day I Day II

IgA

Co

nce

ntr

atio

n [μ

g/m

L] ***

*** p <0.001

International Symposium on Global Multidisciplinary Engineering 2011 (S-GME2011) IOP PublishingIOP Conf. Series: Materials Science and Engineering 21 (2011) 012029 doi:10.1088/1757-899X/21/1/012029

3

Cortisol is the most potent glucocorticoid produced. It is secreted by the adrenal cortex. It plays an

important role in that it maintains adequate blood glucose levels. Cortisol is considered to be released

into the blood stream by activation of the HPA system, so it is a potential stress biomarker [4]. A

transient increase in salivary cortisol has been observed in response to short-term stressors such as

mental arithmetic, stroop tasks, and oral presentations [4,7]. Moreover, the level of cortisol released

immediately after waking has recently been reported to reflect chronic stress levels [8]. This was

confirmed by an experiment we conducted that demonstrated that performing mild exercise prior to

going to bed alleviates hypersecretion of cortisol after awakening (Figure 3).

Although there must be many potential mediators that affect the secretion of these stress biomarkers,

(e.g., sex, age, personality), this method promises to provide a completely new methodology for

objectively estimating stress levels.

Figure 3. Cortisol awakening response: the salivary cortisol

concentration immediately before going to bed (tn), immediately

after waking (t0), 30 min after waking (t15), and 45 min after

waking (t45). Within-subject design (n = 8) was employed. The

“Exercise” group performed mild exercise about an hour before

going to bed.

3. Bionanodevice for detecting human stress — a developing idea

The ultimate goal of the above-mentioned stress biomarker studies is on-site assessment of human

stress. As mentioned above, stress biomarkers can be released into the blood or saliva via activation of

the two physiological stress reaction pathways (i.e., HPA and SAM). Stress biomarkers are also

expected to be determinable by analyzing sweat and other humoral fluids that are exuded from the skin.

Therefore, an on-site stress detection device needs to be compact, integrated, sensitive, and non-

invasive.

A motor protein-powered device is a possible means for achieving this challenging goal. Motor

proteins perform nanoscale mechanical movements by using chemical energy generated by hydrolysis

of ATP. Various nanoscale platforms using motor proteins have been proposed [9]. We are currently

focusing on a molecular transport system that harnesses the actinmyosin interaction (Figure 4). The

-1.50

-1.00

-0.50

0.00

0.50

1.00

1.50

tn t0 t30 t45

Exercise

Control

Co

rtis

ol

Co

nce

ntr

atio

n(z

-sco

re) *

**

* p<.05, ** p<.01 vs tn

International Symposium on Global Multidisciplinary Engineering 2011 (S-GME2011) IOP PublishingIOP Conf. Series: Materials Science and Engineering 21 (2011) 012029 doi:10.1088/1757-899X/21/1/012029

4

actinmyosin interaction typically occurs in muscles; it is used to generate the extremely powerful

contractions of muscles. The platform shown in Figure 4 harnesses this motor protein interaction.

Since the actin filaments are attached to myosin, which is fixed to the substrate, the actin filaments can

act as a shuttle for transporting an attached cargo (antibodies of the target stress biomarker, in our

case). The spatial configuration of the substrate on which myosin or another motor protein are fixed is

critical for controlling the molecular transport system and thus for constructing an on-chip device.

Tracks are formed in poly(methylmethacrylate) or another substrate. These tracks constrain the actin

filaments while allowing them to move smoothly within the tracks [10].

Although these concepts are currently being demonstrated merely as proof of principle, they

promise to result in molecular handling technology for achieving on-site assessment of stress levels.

Figure 4. Actinmyosin platform for molecular transport system.

4. Conclusion

This paper reviewed previous stress biomarker studies including our experimental results and it

describes the concepts behind a bionanodevice for on-site assessment of stress levels. As mentioned

above, the problem of mental stress is currently a global crisis. Moreover, the difficulty of individuals

monitoring and managing their stress levels unaided makes mental stress more serious. Future studies

will contribute stress management research through the development of a means for objectively

measuring stress levels.

Acknowledgement

This study was financially supported by Program for Developing the Supporting System for Global

Multidisciplinary Engineering Establishment, Ministry of Education, Culture, Sports, Science and

Technology, Japan.

References

[1] Gabriel, P. & Liimatainen, M.R. (2000). Mental health in the workplace. International Labor

Organization. Retrieved August 1, 2009, from http://ilo.org

[2] Valdimarsdottir, H.B. & Stone, A.A. (1997). Psychosocial factors and secretory Immunoglobulin

A. Critical Reviews in Oral Biology & Medicine, 8(4), 461-474.

[3] Ader, R., Felten, L. & Cohen, N. (Eds.) (2001) Psychoneruoimmunology. 3rd ed. Academic Press.

International Symposium on Global Multidisciplinary Engineering 2011 (S-GME2011) IOP PublishingIOP Conf. Series: Materials Science and Engineering 21 (2011) 012029 doi:10.1088/1757-899X/21/1/012029

5

[4] Kirschbaum, C & Hellhammer, D.H. (1994). Salivary cortisol in psychoneuroendocrine research:

recent developments and applications. Psychoneuroendocrinology, 19(1), 313-333.

[5] Bosch, J.A., Ring, C., de Geus, E.J., Veerman, E.C. & Amerongen, A.V. (2002) Stress and

secretory immunity, International Review of Neurobiology, 52(1), 213-253.

[6] Nomura, S., Ohira, H. & Kamei, T. (2009) Effect of the Relief from Chronic Stress during

Graduation Examination on Salivary Biomarkers, Transactions of Japan Society of Kansei

Engineering, 8(3), 481-488.

[7] Dickerson, S.S. & Kemeny, M.E. (2004). Acute stressors and cortisol responses: A theoretical

integration and synthesis of laboratory research. Psychological Bulletin, 130(3), 335-391.

[8] Wessa, M., Rohleder, N., Kirschbaum, C. & Flor, H. (2009) Altered cortisol awakening response

in posttraumatic stress disorder. Psychoneuroendocrinology, 31(2), 209-215.

[9] van den Heuvel, M. & Dekker, C. (2007). Motor proteins at work for Nanotechnology. Science,

317, 333-336.

[10] Suzuki, H., Yamada, A., Oiwa, K., Nakayama, H. & Mashiko, S. (1997) Control of actin moving

trajectory by patterned Poly(methylmethacrylate) tracks. Biophysiological Journal, 72, 1997-

2001.

International Symposium on Global Multidisciplinary Engineering 2011 (S-GME2011) IOP PublishingIOP Conf. Series: Materials Science and Engineering 21 (2011) 012029 doi:10.1088/1757-899X/21/1/012029

6