IN VIVO PROTEIN SYNTHESIS DETERMINATIONS IN HUMAN IMMUNE CELLS

Intact immune responses are essential for defeating severe infections in individual patients. Insufficient function of the immune system contributes to a poor prognosis in these patients, in particular the ICU patients. Nevertheless, the immune system function is not easily monitored and evaluated. The ongoing metabolic activity of immune competent cells is reflected by their in vivo protein synthesis rate. The aim of this thesis was to apply in vivo protein synthesis measurement in cells of the immune system, in order to quantitatively characterise the state of their activation. Such measurement may add information on immune cells activation and serve as a tool for evaluation of immune competence in severely ill patients. The in vivo fractional protein synthesis rate (FSR) was determined in the circulating peripheral blood cells: T lymphocytes, mononuclear cells and whole population of leukocytes, as well as in the stationary, unfractioned cells of palatine tonsils. The FSR in the isolated T lymphocytes of healthy volunteers was 13.6 ± 0.9 %/24 h and was not affected by a 6-h cortisol infusion, either immediately after the end of the infusion or 18 h later. In contrast, a combined stress hormone infusion (cortisol, epinephrine, glucagon), as a human model of surgical trauma, given for 6 h to healthy volunteers decreased the in vivo protein synthesis rate in T lymphocytes by 34% from 13.0 ± 1.0 %/24 h to 8.6 ± 2.1 %/24 h. A more accentuated decrease by 53% was observed in the total mononuclear cells, from 13.3 ± 1.2 %/24 h to 6.3 ± 2.0 %/24 h. Following an endotoxin injection, a human model of the initial phase of sepsis, different patterns of the in vivo fractional protein synthesis rates were observed in the circulating blood cells of healthy volunteers. The isolated T lymphocytes responded with a 60% decrease of the protein synthesis rate from 9.4 ± 1.2 %/24 h to 3.8 ± 2.4 %/24 h, whereas the whole population of leukocytes showed an increase by 43% from 3.2 ± 1.2 %/24 h to 4.4 ± 1.1 %/24 h. A comparison of the in vivo fractional protein synthesis rate between the circulating and stationary immune cells of healthy subjects revealed that unfractioned tonsillar cells had a protein synthesis rate of 22.8 ± 5.7 %/24 h. This rate was higher compared with T lymphocytes, mononuclear cells and leukocytes separated from peripheral blood in these subjects, having in vivo protein synthesis rates of 10.7 ± 3.4 %/24 h, 10.8 ± 2.8 %/24 h and 3.2 ± %/24 h, respectively. Alterations in the protein synthesis rates were also observed during the early phase of the critical illness. In a pilot group of intensive care unit patients with a general systemic inflammatory activation, a distinct polarization of the protein synthesis responses was observed. The in vivo protein synthesis rate in the mononuclear cells and in the whole population of leukocytes was high, 21.6 ± 7.4 %/24 h and 8.9 ± 4.4 %/24 h, respectively, while that in T lymphocytes (12.5 ± 5.5 %/24 h) and tonsillar cells (27.9 ± 11.4 %/24 h) was not different from what was observed in healthy subjects. In summary, uniform and characteristic changes of the in vivo rate of protein synthesis in response to exogenous stimuli in healthy volunteers and during the early phase of systemic inflammation in critically ill patients were described in the individual populations of immune cells. The in vivo protein synthesis determination in the immune cells reflects the state of activation of these cells. This measurement may be used as a tool to obtain additional information on the immune competence in studies concerning function of the human immune system. LIST OF PUBLICATIONS This thesis is based on the following papers, which will be referred to in the text by their Roman numerals I-V. I. In vivo protein synthesis of circulating human T lymphocytes does not respond to a cortisol challenge within 24 hours. Januszkiewicz A, Essen P, McNurlan MA, Ringden O, Wernerman J, Garlick PJ. Acta Anaesthesiol Scand, 2000; 44: 202-209. II. Stress hormone infusion decreases protein synthesis of circulating human T lymphocytes. Januszkiewicz A, Essen P, McNurlan MA, Ringden O, Garlick PJ, Wernerman J. Metabolism, 2001; 11: 1308-1314. III. Response of in vivo protein synthesis in T lymphocytes and leukocytes to an endotoxin challenge in healthy volunteers. Januszkiewicz A, Lore K, Essen P, Andersson B, McNurlan MA, Garlick PJ, Ringden O, Andersson J, Wernerman J. Clin Exp Immunol, 2002; 130: 263-270. IV. Determination of in vivo protein synthesis in human palatine tonsil. Januszkiewicz A, Klaude M, Lore K, Andersson J, Ringden O, Rooyackers O, Wernerman J. Clin Sci, 2005; 108: 179-184. V. In vivo protein synthesis in immune cells of ICU patients – a pilot study. Januszkiewicz A, Klaude M, Lore K, Andersson J, Ringden O, Rooyackers O, Wernerman J. Manuscript. Reprints of the above articles were made with the kind permission of the publishers: Blackwell Publishing (paper I and III), Elsevier Science (paper II) and Portland Press (paper IV).