The Role of Exercise Stress and Catecholamines in

Immunity

Erika VielmasFebruary 23, 2009Biol 520

Background Information

Catecholamines (CAs)

Catecholamines Known as epinephrine and norepinephrine

(adrenaline/noradrenaline). Water soluble hormones that circulate in the

blood. Produced from chromaffin cells;

neuroendocrine cells found in the adrenal medulla.

β-adrenergic receptors: G-coupled protein receptors that are targets for CAs.

http://en.wikipedia.org/wiki/Adrenergic_receptorBenschop et al 1994

Catecholamine Synthesis

OH CH2-CH-COO-

+NH3

TyrosineOH

OH

OH OH

OH OH

CH2-CH2

+NH3

CH-CH2

+NH3

OH

Dopamine

Norepinephrine

CH-CH2

OH

H2N+-CH3

Epinephrine

Nelson & Cox 2005

Dopamine β-Hydroxylase

PhenylethanolamineN-methyltransferase

Tyrosine hydroxylaseAromatic amino acid decarboxylase

The Hypothalamic–Pituitary–Adrenal (HPA) Axis

PVN of the hypothalamus releases Cortico releasing hormone (CRH).

CRH stimulates the anterior pituitary gland to release adrenocorticotropin (ACTH) hormone into circulation.

The ACTH induces the adrenal glands to synthesize and secrete glucocorticoids and catecholamines.

Webster-Marketon et al 2008www.ott.zynet.co.uk/.../images/encephf1.gif

PVN

A.P

Sympathetic Nervous System

The SNS is activated by the secretion of acetylcholine from the pre ganglionic sympathetic fibers in the adrenal medulla.

SNS activation induces secretion of epinephrine into blood circulation.

Norepinephrine is released from the nerve terminals in the vicinity of immune cells.

Webster-Marketon et al 2008users.rcn.com/.../BiologyPages/P/PNS.html

CAs can be Synthesized by Immune Cells

For many years, CAs were believed to be synthesized only by neurons and endocrine cells.

Recent studies suggest that lymphocytes and mature CD3 T- cells can synthesize and secrete CAs.

Cells need high levels of norepinephrine to stimulate the production of more CAs.

Leposavic et al 2008

Lymphocyte Differentiation

Mammalian stem cells differentiate into several kinds of blood cells within the bone marrow by a process called haematopoiesis.

All lymphocytes originate from a common lymphoid progenitor before differentiating into their distinct lymphocyte types.

The formation of lymphocytes is known as lymphopoiesis.

http://en.wikipedia.org/wiki/Lymphocyte

www.textbookofbacteriology.net

Natural Killer Cells (NK)

Natural Killer Cells are large granular lymphoid-like cells important in innate immunity.

NK cells are produced in the bone marrow. These cells can detect virus-infected cells without having any

antigen specificity . (“natural killer”). To control their cytotoxic activity, NK cells possess two types

of surface receptors: activating receptors and inhibitory receptors.

Inhibitory receptors recognize MHC I molecules and activating receptors recognize interferons and cytokines (which may be caused by stress); interferon γ (INF-γ) and interleukin-2 (IL-2) .

80% of NK cells express CD8.

http://jem.rupress.org/cgi/content/abstract/190/7/1005Calcagni et al 2006Webster-Marketon et al 2008

Two Functionally Distinct Cells for NK Cells : CD56dim & CD56bright

CD56dim constitutes 90% of peripheral NK cells and these are more cytotoxic than the CD56bright . These cells express higher levels of CD16 than their subset.

CD16 is a low-affinity immunoglobulin G (IgG) Fc receptor that is expressed on natural killer (NK) cells.

CD56bright cells have the capacity to produce an abundant amount of cytokines.

Timmons et al 2006

T-lymphocytes

T lymphocytes Migrate to and mature in the thymus. Enter circulation and peripheral lymphoid

organs (spleen & lymph nodes) Patrol for invading pathogens and/or tumor

cells. Involved in adaptive immunity differentiate

further after exposure to an antigen.

http://en.wikipedia.org/wiki/Lymphocyte

T-lymphocytes

Have a special surface receptor that recognize specific antigens. (Express CD3, and CD8).

CD8 cells differentiate into cytotoxic T cells or can be expressed by NK cells.

CD3 complex is the complex of α:β or γ:δ T-cell receptor chains.

Assist in the recognition and rejection of foreign tissues.

CAs, can inhibit lymphocyte proliferation and cytotoxicity.

http://en.wikipedia.org/wiki/Lymphocyte

Stress Induced by Exercise

Exhaustive Exercise Vigorous physical activity triggers

the response of CAs during exhaustive exercise. Know to induce a shift in peripheral blood NK

cells. Associated with significant shifts in peripheral

immune cell counts. Redistribution of lymphocytes among different

compartments of the immune system.

Imrich et al 2004, Zhang et al 2006cache.boston.com/.../swim_06_30/swim13.jpg

Webster-Marketon et al 2008

The HPA axis and the SNS are activated which induces a release of glucocorticoids and catecholamines which can modulate different aspects in the immune system.

Hypothesis

Exercise-induced stress: Increases catecholamine levels Decreases natural killer cell cytotoxicity Decreases T-lymphocyte activity.

Overview

We will be looking at Plasma concentrations of CAs prior and after vigorous exercise.

NK cell and T-lymphocyte cell counts and activity prior and after vigorous exercise.

Show the connection of CAs, stress (from exercise), and immune cells.

Experimental Results

Suzui et al 2004

The training group consisted of 15 women in pre-season training for 1 month training for 5h/day.The control group consisted of 7 women instructed to avoid vigorous exercise.Sample of blood from pre-training, during (10th day), end (1day prior to end training, and post-training ( 1 week after).

NK Cell Count Increase Throughout Training

Concentrations of the CAs and Cytolytic Activity of the NKs

Suzui et al 2004

Catecholamine levels were not affected. There was no significant difference during the 1 month training process.

Nk cell cytolytic activity during and after training.

CAs levels and Cell Counts for Immune Cells

Imrich et al 2004

The training group consisted of 10 healthy males.6 weeks of endurance training: 1h run, 4 times a weekFollowed by 5 days of bed rest.

Flow cytometry

Method for quantitating components or structural features of cells & measures fluorescence per cell or particle.

Scattered and emitted light from cells and particles are converted to electrical pulses by optical detectors. Parallel light waveforms is picked up by confocal lenses focused at the intersection point of cells and the light source.

After the different signals or pulses are amplified they are processed by an Analog to Digital Converter (ADC) which in turn allows for events to be plotted on a graphical scale(One Parameter, Two parameter Histograms).

http://biology.berkeley.edu/crl/flow_cytometry_basic.html

CD56dim & CD56bright Cell Distribution

Timmons et al 2006

Dot plot: CD56 and CD3A total of 10,000 events were collected in thelymphocyte gate based on forward- vs. side-scatter characteristics.N=11 female adolescents age 14Exercise: Vigorous CyclingRest: prior to exercisePost: 60 min. after exerciseRecovery: 60 min. rest period after exercise.

NK Cell Count and CA Concentrations

Zhang et al 2006

Epinephrine* P< 0.001

(pg.

ml-1

)

Lancaster et al 2004

Effects of Exercise on T-Lymphocytes

Subjects consisted of 7 malesExercise: Cycling til’ exhaustionPre-exercise60 minPost-exercise1h post-exerciseTrial 1: 6 day Intense Training Period work out everyday.Trial 2: 2 week training, 3-4 sessions per weekTrial 3: Same as first trial

NK cells in Lung Tissue

NK cells have different activity in different organs.

Kanemi and his team investigated the effect of restraint stress on lung lymphocytes including NK cells.

C57BL/6 female mice were exposed to 2 h restraint stress.

Kanemi et al 2004

Kanemi et al 2004

Plasma concentrations of Epinephrine and Norepinephrine

N=6Samples measure by HPLCAfter the mice were stressed, the CAs levels immediately increased but the value for Norepinephrine was not shown to be significant despite the increase.

Con= controlRS0= 0 hrRS1= 1 hrRS2= 2 hr

Kanemi et al 2004

Y-axis is the number of cells. Levels of lung NK cells decreased significantly as well as NK cells in the Plasma. Initial decrease occurred during stress and lasted throughout the recovery phase.

Immunohistochemical analysis of the lung tissue: a) control mice b) stressed mice.c) NK cells decreased compared to control.

Stress on pulmonary NK cells

Conclusion

Summary

Supports Hypothesis

Partial support of Hypothesis

Refutes Hypothesis

Suzui et al 2004

Lancaster et al 2004

Imrich et al 2004

Timmons et al 2006

Kanemi et al 2006

Zhang et al 2006

NK decrease in cytotoxicity, CAs not affected

CAs increased but Immune cells not affectedIncreased CAs and decrease NK cell

CA’s increased in girls,no change in boys; Nk increased in girls, not in boys

CAs increased and NK cell increase

(cytotoxicity not mentioned)

Increase in CAs, Increase in T-cell (CT not

mentioned)

Take Home MessageWomen Exercise decreased cytolytic activity despite an

overall increase in NK cells and T-cells. CAs are not affected.

Men Immune cells were not affected by exercise but CAs

levels increased significantlyAdolescents Increase in CAs for girls. Increase in cytolytic activity. Incease in CAs for boys. No significant change in

cytolytic activityMice Stress increased CAs. Decrease in NK cells.

Conclusion

According to the data, my hypothesis was partially supported.

CAs increase in certain instances as a result of stress from exercise.

Cytotoxic activity is not directly mediated by CAs increase.

Not enough studies done to show the differences among men, women, adolescents, and rodent models.

Future Experiments

Use proper controls. Men: Trained vs. Untrained Women: Trained vs. Untrained Boys: Trained vs. Untrained Girls: Trained vs. Untrained Run a hormone analysis of testosterone, estrogen, and

CAs among adolescents and adults after exercise. In vitro experiments to check cytotoxic activity of NK

cells and T-cells: Using NK-sensitive and T-sensitive K562 target cell

lines and label with non-radioactive europium.

References

Benschop, R.J., Nijkamp, F.P., Ballieux, R.E., Heijnen, C. (1994) The effects of B-adrenoreceptor Stimulation on Adhesion of Human Natural Killer Cells to Cultured Endothelium. Br. Journal of Pharmacology 113, 1311-1316

Calcagni, E., Elenkov, I. (2006) Stress System Activity, Innate and T Helper Cytokines, and Susceptibility to Immune-Related Diseases. Annual New York Academy of Sciences 1069, 62-76

Imrich, R., Tibenska, E., Koska, J., Ksinantova, L., Kvetnansky, R., Bergendiova-Sedlackova, K., Blazicek, P., Vigas, M. (2004) Repeated Stress-Induced Stimulation of Catecholamines Response is not Followed by Altered Immune Cell Redistribution. Anneals New York Academy of Sciences 1018, 266-272

Kanemi, O., Zhang, X., Sakamoto, Y., Ebina, M,. Nagatomi, R.(2005) Acute Stress Reduces Intraparenchymal Lung Natural Killer Cells Via beta-adrenergic Stimulation. Clinical and Experimental Immunology, 139, 25–34

Lancaster G.I., Halson, S. L., Khan, Q., Drysdale, P., Wallace, F., Jeukendrup, A.E., Drayson, M.T., Gleeson, M. (2004) Effects of Acute Exhaustive Exercise and Chronic Exercise Training on Type 1 and Type 2 T Lymphocytes. Exercise Immunology Rev 10, 91-106

Leposavic, G., Pilipovic, I., Radojevic, K., Pesic, V., Perisic, M., Kosec, D. (2008) Catecholamines as Immunomodulators: A role for Adrenoceptor-mediated Mechanisms in Fine Tuning of T-cell Development. Autonomic Neuroscience:Basic and Clinical 144, 1-12

Nelson, D.L., Cox, M.M. (2005)Lehninger Principles of Biochemistry. W.H. Freeman and Company. Suzui, M., Kawai, T., Kimura, H., Takeda, K., Yagita, H., Okumura, K., Shek, P.N., Shephard, R.J. (2004) Natural Killer Cell

Lytic Activity and CD56dim and CD56bright Cell Distributions During and After Intensive Training. Journal of Applied Physiology 96: 2167–2173, 2004

Timmons, B.W., Tarnopolsky, M.A., Bar-Or, O.( 2006) Sex-based effects on the Distribution of NK Cell Subsets in Response to Exercise and Carbohydrate Intake in Adolescents. Journal of Applied Physiology 100, 1513–1519

Webster-Marketon, J.I., Glaser, R. (2008) Stress Hormones and Immune Function. Cellular Immunology 252, 16-26 Zhang, X., Matsuo, K., Farmawati, A., Higashi, K. O., Nagata, K., Nagatomi, R. (2006) Exhaustive Exercise Induces

Differential Changes in Serum Granulysin and Circulating Number of Natural Killer Cells. Journal of Experimental Medicine 210, 117-124

http://en.wikipedia.org/wiki/Adrenergic_receptor http://en.wikipedia.org/wiki/Lymphocyte http:// www.textbookofbacteriology.net http://jem.rupress.org/cgi/content/abstract/190/7/1005 http://www.ott.zynet.co.uk/.../images/encephf1.gif http://users.rcn.com/.../BiologyPages/P/PNS.html http://cache.boston.com/.../swim_06_30/swim13.jpg

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