Stress is thought to be immunosuppressive but paradoxically exacerbates inflammatory and autoimmune diseases (K.Viswanathan et al., 2005). Many aspects of the stress-induced immunomodulation remain controversial especially in terms of the age-related aspects of the problem. Growing body in known to be particularly vulnerable to the stress exposure but the details of the stress-mediated immunosuppression are mainly investigated at the level of the central lymphoid organs which is described as an accidental thymic involutation, while the involvement of the peripheral immune organs remains underestimated (Q.Li et al., 2005; R.G.Tseng et al., 2005).
The links among the stress-associated increased level of plasma glucocorticoids, catecholamines and immunity were examined in a number of studies. As peripheral blood is generally the only "window" available through which the human immune response can be studied, it is difficult to assess the mechanisms by which neuroendocrine responses affect either the inductive or effector phases of immunity, as both generally occur in tissues and not in the blood, hence immunohistochemical methods of the tissue-specific changes evaluation are invaluable in getting information regarding the stress-associated immunomodulation at the level of the peripheral lymphoid organs (J.K. Kiecolt-Glaser et al., 1995; D.A. Padgett et al., 2003; J.Diao et al., 2006; M.E. Truckenmiller et al., 2006).
The objective of the present investigation was to reveal the stress-induced immunomodulatory changes in the growing body evaluated at the level of the secondary lymphoid organs (lymph nodes).
Prepubertal Sprague-Dawley rats aged 21 and 30 days corresponding to the weaning and infant periods accordingly, were exposed to the severe chronic (restraint) stress (R.Kvetnansky et al., 1970) with 7 daily 5-hour sessions. Each age group of the experimental animals contained 8 rats with another 8 rats serving as an age-matched control, with total number of the animals equal to 32 species.
After the last session of stress the animals were sacrificed, their thymus, spleen and inguinal lymph nodes were sampled, weighed, fixed in formalin and embedded in paraffin. Histological sections of the lymph nodes were stained with hematoxylin-eosin for routine histological examination and immunohistochemically proceessed for CD8 (T-suppressor/cytotoxic lymphocytes) and CD20 (B-lymphocytes) markers using streptavidin-biotin-peroxidase method with subsequent quantitative evaluation of the volume density and the numeric density of the immunopositive cells using NIKON image analyzer with Image Pro Plus 4.5 software.
It was demonstrated that chronic stress induced prominent immunosuppressive changes in the lymph nodes of the prepubertal rats of both age groups. They included considerable reduction of both T- and B-zones in the lymph nodes with increased number of apoptotic cells mainly in the В-zones. The diameter of the primary and secondary lymphoid nodules and the number of the secondary lymphoid nodules was reduced in the senior age group of the experimental animals while the width and the volume density of the paracortical zone decreased in both experimental groups.
The immunohistochemical staining for the СD8 demonstrated that the immunoreactive cells were concentrated in the paracortical zone of the lymph nodes and were scanty in the mantle zone of the lymphoid nodules and in the medullary cords. After stress exposure the number of immunopositive cells in the paracortical zone decreased while single immunoreactive cells were present in the medullary cords and in the cortex of the lymph nodes.
The immunohistochemical staining for CD20 exhibited accumulation of the immunopositive cells in the lymphatic follicles being less densely distributed in the medullary cords of the lymph nodes.
Image analysis demonstrated that the volume density of the CD8+ immunoreactive cells was significantly reduced in the weaning and infant (p<0,01) age groups of the experimental animals compared to the age-matched control rats, while the volume density of the CD20+ immunoreactive cells was significantly reduced in the weaning (p<0,001) and infant (p<0,05) age groups accordingly with a different level of significance.
All these changes were accompanied by a significant reduction of the body (p<0,05), thymus (p<0,001 and p<0,01) and spleen (p<0,01 and p<0,05) mass in weaning and infant experimental animals accordingly.
The results of the investigation revealed considerable immunosuppressive changes in the lymphoid organs of the growing rats demonstrating prominent immunomodulation in the T-zones of the inguinal lymph nodes of both age groups of experimental animals and more severe changes in the B-zones of the lymph nodes of the junior animals of the present study. These findings allow to develop age-related strategies for the prophylaxis of the stress-associated immunosuppressive changes in the growing body.
The work is submitted to the IV Scientific International Conference "Basic research", Italy (Neapolitan Riviera), October, 11-18, 2008, came to the editorial office on 21.08.2008.