Cytokine profiles during delivery affect cord blood hematopoietic stem and progenitors cells.

The study was aimed to determine the correlations between serum levels of cytokines (GM-CSF, IL-4, IL-10 and TNF) in maternal (MB) and cord blood (CB) and some features of cord blood hematopoietic stem and progenitor cells (CB HSPCs). Study material was MB and concomitant CB samples collected from 98 volunteers at the moment of delivery. The IL-4, IL-10, TNF and GM-CSF concentrations in serum and in supernatants from PMA-stimulated mononuclear cells isolated from both blood types were measured using BD Cytometric Bead Array Flex Set System. CB HSPCs (CD34(+)CD45(low)) proportion was also estimated by flow cytometry. The most relevant results concerned the tendency to down regulation of CB HSPCs number with an increase of IL-4, IL-10 and GM-CSF levels, only the TNF concentration seems to have no influence on HSPCs pole size. The strongest positive correlations were found between CD34(+)CD45(low) HSPCs number and IL-10 and GM-CSF in MB serum and GM-CSF and TNF from CB supernatants. The strongest negative correlations were found between CD34(+)CD45(low) HSPCs number and IL-4 and GM-CSF in CB serum and IL-10 in MB supernatants. Interestingly, we observed 'opposite correlation' between serum and supernatant from CB and MB. We concluded that elevated serum levels of IL-4, IL-10 and GM-CSF in CB are indicative of enhanced differentiation of HSPCs and characterize a normal perinatal development. Elevated levels of cytokines seem to stimulate differentiation of HSPCs what is advantageous for neonates during perinatal period.

Ex vivo expansion of CD4(+)CD25(+) T regulatory cells for immunosuppressive therapy.

Immunosuppressants are powerful drugs, capable of triggering severe adverse effects. Hence, there is tremendous interest in replacing them with less-toxic agents. Adoptive therapy with CD25(+)CD4(+) T regulatory cells (Tregs) holds promise as an alternative to immunosuppressants. Tregs have been described as the most potent immunosuppressive cells in the human body. In a number of experimental models, they have been found to quench autoimmune diseases, maintain allogeneic transplants, and prevent allergic diseases. A major stumbling block in their clinical application is related to Treg phenotype and the very limited number of these cells in the periphery, not exceeding 1-5% of total CD4(+) T cells. Recent progress in multicolor flow cytometry and cell sorting as well as cellular immunology has found ways of overcoming these obstacles, and has opened the doors to the clinical application of Tregs. In the review, we describe Treg sorting and expansion techniques that have been developed in recent years. In the experience of our laboratory, as well as some published reports, Treg adoptive therapy is a promising tool in immunosuppressive therapy, and should be considered for clinical trials.

From bench to bedside and back: the SENIEUR Protocol and the efficacy of influenza vaccination in the elderly.

Prophylaxis with vaccines is important in geriatrics as, apart from specific protection, it reduces incidence of potentially fatal infectious complications and exacerbations of existing medical conditions. Post-vaccination protection depends on immunity and therefore markers of immune status could be used to predict efficacy of vaccination. From the practical point of view, the determination of simple and robust methods for assessing immunity, which could enable practitioners to distinguish patients at risk of vaccination unresponsiveness, is desirable. Additional care, necessary to avoid possible complications, could be then targeted to such patients. Here, we discuss correlations between immune parameters and the clinical status of elderly people challenged with anti-influenza vaccination and ways to classify immune status, based on results obtained between 1999 and 2004 in the elderly immunized annually against influenza. These studies document a correlation between health status assessed according to the SENIEUR Protocol and response to the vaccination. Patients classified as SENIEUR-compatible responded to the vaccine, while non-SENIEURs did not. Factors most strongly associated with unresponsiveness were, perhaps unexpectedly, circulatory and psychiatric diseases, although both humoral and cellular immune responsiveness did correlate with levels of proinflammatory cytokines in the serum. Patients whose humoral responses improved during subsequent immunizations were characterized by well-preserved natural killer (NK)-mediated cytotoxicity. In contrast to the first-immunization responders, the second- immunization responders were characterized by elevated levels of proinflammatory cytokines and Cytomegalovirus seropositivity, which placed them in the non-SENIEUR group, although they were not yet frail and still lived independently. From this series, we conclude that detailed clinical data together with some simple markers such as IL 6 levels seem sufficient to provide clinicians with presumptive information on the condition of the immune system and to allow for initial prediction of vaccination efficiency. Those patients predicted to respond poorly could be considered for alternative treatment.

Quantifying division of aortal smooth muscle cells in culture stimulated by 1,25(OH)2D3.

Inhibitory effect of 1alpha,25dihydroxycholecalciferol (1,25D(3)=calcitriol) in different cell type is well recognized but its promoting effect on vascular smooth muscle cells (SMCs) is poor established. Therefore, the aim of this study was to determine stimulatory effect of calcitriol on aortal SMCs proliferation in culture. We used the cell division analysis procedure based on the quantitative sequential halving of the stably incorporating fluorescent dye carboxyfluorescein diacetate succinimidyl ester (CFSE). This technique allowed the visualization of cycles of SMCs division by flow cytometry. Rat aortal SMCs were labeled with CFSE and cultured for up to 10 days with defined concentration of calcitriol in medium. Proliferative activity as the percentage of SMCs in different phases of the cell cycle using propidium iodide was determined. Apoptosis was assessed using Annexin-V/CFDA method. The results suggest that low concentrations of an active form of vitamin D-1alpha,25dihydroxycholecalciferol applied in supraphysiological concentration of 10 nmol/l is a mitogenic factor for aortal SMCs. None of the applied concentrations of calcitriol caused apoptosis. The findings well support our morphological (LM) and ultrastructural (TEM and SEM) observations.

Anti-tumor action of tumor necrosis factor against Bomirski Ab melanoma in hamsters.

INTRODUCTION: Tumor necrosis factor (TNF) is a cytokine able to exert anti-tumor activity in various models and modes of applications. However, the exact mechanism mediating the in vivo anti-tumor effect of TNF has not yet been clarified. MATERIALS AND METHODS: The effects of intratumoral injection of rat TNF into hamsters bearing Bomirski Ab amelanotic melanoma, a fast growing tumor of high metastatic potential, were tested. Subcutaneous injections of the anti-angiogenic compound TNP-470 allowed analysis of its influence on the effects of TNF administration. RESULTS: TNF application resulted in a significant inhibition of tumor growth and changes in metastasis pattern. Accelerated hemorrhagic necrosis was also observed, indicating the effect of the cytokine on tumor vessels. Moreover, the synergistic anti-tumor effect of TNF and anti-angiogenic agent TNP-470 suggested a cooperative activity of both substances on tumor vasculature. Microscopically, the effect of TNF injections was expressed by an increase in the amount of tumor cells with nuclear pyknosis and karryorrhexis. In vitro assays indicated a direct cytotoxic effect of TNF against Ab melanoma cells, most probably as an outcome of apoptosis. Intratumoral application of TNF also caused some modulation of cytokine response in melanoma-bearing hamsters as evidenced by increased levels of IL-6 in blood serum. CONCLUSIONS: This study established Bomirski Ab melanoma as a useful model for complex analysis of the anti-tumor activity of TNF.

NK cell compartment in patients with coronary heart disease.

BACKGROUND: Viral and bacterial infections have been considered as a risk factor for Coronary Heart Disease (CHD). NK cells, as a first line of defense against those infections, may play a role in CHD development. Thus, the main aim of our study was to determine NK cell compartment in patients with CHD undergoing coronary artery by-pass grafting. RESULTS: Ninety three patients with CHD were included into the study; the control group consisted of 49 healthy volunteers. As compared to controls, CHD patients had lower NK cytotoxic activity. CHD group had also a decreased absolute number and percentage of total NK cells and CD3-CD56dim cytotoxic NK subset. In addition, we observed tendency toward lower percentage of the CD3-CD56bright regulatory NK subset and CD3-CD56+IFN-gamma+ cells in CHD patients. CONCLUSION: These data indicate that CHD is associated with an impairment of NK cells compartment.

[Tuftsin--new analogues and properties]. / Tuftsyna--nowe analogi i wlasciwosci.

Tuftsin, a natural tetrapeptide of sequence TKPR, occuring in the blood of humans and other mammals, capable of stimulating certain white blood cells (monocytes, macrophages, and neutrophils), was isolated at Tufts University in 1970 by Najjar and Nishioka. Tuftsin is a compound with a wide spectrum of biological activities, notable enhances phagocytosis, immune response, bactericidal, tumoricidal and antifungal activities. This article concerns new analogues and properties of tuftsin.

Interleukin 6 polymorphism corresponds to the number of severely stenosed coronary arteries.

IL6 gene promoter polymorphisms may influence the outcome of cardiovascular diseases. The aim of our study was to find out whether the -174G>C polymorphism, as well as the IL6 secretory profile, may be linked to the number of severely (> or = 75%) occluded coronary arteries in patients with advanced coronary heart disease (CHD). Three hundred and twenty patients awaiting elective coronary artery bypass grafting were enrolled into the study. Blood was taken the day before surgery. The PCR-RFLP method was used for IL6 gene polymorphism analysis. Spontaneous IL6 release was measured by bioassay in supernatants of whole blood cell cultures (WBCC) incubated for 24 h and 48 h. We found that significantly more patients with triple vessel disease were found within the -174GG group as compared to the -174GC and CC genotype carriers. The highest IL6 serum levels were found in the -174GG and the lowest in the -174CC genotype patients. Spontaneous in vitro IL6 secretion appeared to be significantly higher at all time points in the -174GG as compared to the CC and GC genotype carriers. The serum concentration of IL6 and the spontaneous IL6 secretion were directly related to the number of obstructed coronary vessels. Our results emphasize the role of IL6 as an important, non-classical risk factor predicting the number of severely affected coronary vessels.

The atypical pattern of cell death in B16F10 melanoma cells treated with TNP-470.

TNP-470 is an acknowledged anti-angiogenic factor, and was studied clinically as an anti-cancer drug. We previously reported on an additional property of this molecule: the intracellular generation of reactive oxygen species in B16F10 melanoma cells. We showed that a massive generation of ROS occurred in the first few hours after treatment with TNP-470 and that this event was critical to subsequent cell death. In this study, we analyzed the process of cell death and noticed an atypical pattern of death markers. Some of these, such as DNA fragmentation or condensation of chromatin, were characteristic for programmed cell death, while others (the lack of phosphatidylserine flip-flop but permeability to propidium iodide, the maintenance of adhesion to the substratum, no change in mitochondrial transmembrane potential, no effect of the panspecific caspase inhibitor) rather suggested a necrotic outcome. We concluded that TNP-470 induced at least some pathways of programmed cell death. However, increasing damage to critical cell functions appears to cause a rapid switch into the necrotic mode. Our data is similar to that in other reports describing the action of ROS-generating agents. We hypothesize that this rapid programmed cell death/necrosis switch is a common scenario following free radical stress.

Serum and urinary cytokine homeostasis and renal tubular function in children with type 1 diabetes mellitus.

AIM: To explore the relationships between tumor necrosis factor-alpha (TNFalpha), interleukin-6 (IL-6) and urinary N-acetyl-beta-D-glucosaminidase (NAG) and the function of renal proximal tubules in children with type 1 diabetes mellitus (DM1). METHODS: Fifty-six children with DM1 and 35 healthy controls were analyzed. We measured NAG (A and B isoforms) in urine as well as serum TNFalpha and urinary IL-6. RESULTS: The children with DM1 with microalbuminuria (group A) had significantly higher urinary IL-6 and serum TNFa than the children without microalbuminuria (group B). The diabetic patients with no sign of nephropathy showed significantly higher TNFalpha and NAG and its A and B isoforms in urine compared to the healthy group. Additionally, groups A and B both showed a positive significant correlation between serum TNFalpha and urinary isoform B. CONCLUSIONS: From our pilot results it appears that TNFalpha might be a sensitive marker of damage to the renal proximal tubules occurring prior to microalbuminuria. Conversely, the increase in NAG and its isoform B activity in patients with no clinical sign of diabetic nephropathy may indicate the onset of microalbuminuria.

The biological activity of new tuftsin derivatives--induction of phagocytosis.

Phagocytosis plays a crucial role in a host defense against invading microorganisms. This process can be induced by many phagocytosis stimulating factors. One of them is an endogenous tetrapeptide - tuftsin that occurs in the blood of mammals including human beings. Tuftsin is capable of potentiating granulocyte and macrophage functions such as: phagocytosis, motility, and chemotaxis as well as bactericidal and tumoricidal activity. The other particle able to induce phagocytosis is muramyl dipeptide (MDP), the smallest synthetic glycopeptide of bacterial origin that possesses an immunogenic activity. MDP is known to affect most functions of macrophages. Phagocytosis stimulating properties of a new group of tuftsin and MDP analogues (one tuftsin analogue and four conjugates of tuftsin/retro tuftsin and muramyl dipeptide or nor-muramyl dipeptide) were tested. The results of the study show that all of the examined conjugates are able to generate oxidative burst. The most promising analogues proved to be kd6 and kd7.

Lower percentage of CD8(high+)CD152(+) but not CD8(high+)CD28(+) T lymphocytes in the elderly may be reverted by interleukin 2 in vitro.

An expression of the surface co-stimulatory molecules-the CD152 and the CD28 has been compared between young and old individuals on the CD8(high+) lymphocytes. Sixty five elderly healthy (65-96 years old) and 31 young (19-40 years old) volunteers were examined. An expression of CD152 and CD28 surface antigens was analyzed by flow cytometry ex vivo and on whole blood cell cultures lymphocytes stimulated with interleukin 2 (IL2). The elderly population was characterized by a lower percentage of the CD8(high+) lymphocytes than the young population. The percentages of CD28(+) lymphocytes as well as those of CD8(high+)CD28(+) subpopulation were lower in the old group compared to the young group. The surface expression of CD152 antigen was similar to that of CD28 with a lower percentage of the CD152(+) lymphocytes and CD8(high+)CD152(+) cells in the old group. Stimulation of lymphocytes in vitro with IL2 resulted in an increase of the CD8(high+)CD152(+) cells in the elderly, while it had no effect on lymphocytes of the young group. Our results indicate that lymphocytes of the elderly population are characterized by a lower expression of the surface CD28 and CD152 molecules. An age-related decrease of an expression of the co-stimulatory molecules CD28 and CD152 on the surface of lymphocytes, found in our study, may be compatible with a hypothesis of a 'remodelling' of immune response in the healthy elderly.

Synthesis and cytotoxic activity of conjugates of muramyl and normuramyl dipeptides with batracylin derivatives.

The synthesis of MDP (muramyl dipeptide) or nor-MDP (normuramyl dipeptide) conjugates modified at the peptide part with batracylin (BAT) or batracylin derivatives is described. Batracylin was synthesized by our modified method (Scheme 3). The synthesis of BAT via this modified route now appears to be feasible on a multigram scale. Preliminary screening data obtained at the National Cancer Institute (NCI, Bethesda, MD) have revealed that the conjugates did not expose any cytotoxic activity even at 10(-4)-10(-8) M or microg/mL. During tests performed at Medical University of Gdansk, Poland, two analogues 11c and 11e reduced the proliferation of Ab melanoma cells in vitro compared with batracylin alone (Table 2, Figure 1).

Corticotropin-releasing hormone affects cytokine production in human HaCaT keratinocytes.

CRH cutaneous expression is significantly enhanced after exposure to various stimuli (Physiol Rev 2000, 80;979-1020). We evaluated the effect of CRH on cytokine production in HaCaT keratinocytes, a cell line shown to express CRH receptors coupled to cAMP activation and calcium-dependent transmission pathways. It is demonstrated for the first time that exogenously added CRH stimulates production of IL-6 and IL-11. It also inhibits production of IL-1beta and does not affect TNF-alpha production. Our results indicate that CRH function(s) during cutaneous stress may be mediated by differential effects on cytokine production.

Differences in kinetics of donor lymphoid cells in response to G-CSF administration may affect the incidence and severity of acute GvHD in respective HLA-identical sibling recipients.

Granulocyte-colony stimulating factor (G-CSF), in addition to myeloid and stem cells, mobilizes a large number of lymphoid cells. We examined which lymphoid populations were mobilized in 21 consecutive donors of peripheral blood stem cells (PBSC) and whether the differences in mobilization could affect the incidence of acute and chronic GvHD in respective HLA-identical recipients. G-CSF administration induced significant increases of donor B (CD3-CD19+) lymphocytes and slight increases of T (CD3+) and cytotoxic (CD16+CD56+) NK cells. The number of extrathymic cells (CD3+ cells with NK markers, or CD7+) remained unchanged except for an increase of CD3+CD57+CD8+ cells. Donors of patients without subsequent grade II-IV acute GvHD compared to donors of patients who developed significant acute GvHD were found to have in peripheral blood stable numbers of CD3+CD4+ cells producing IL2, with a concomitant increased number of CD3+CD4low+CD25+ T regulatory cells and decreased NK-mediated cytotoxicity, together with a higher number of suppressive extrathymic CD57+CD3+ cells in the blood and G-PBMC grafts. Increasing numbers of activated T and NK cells in the blood were associated with the development of chronic GvHD. We suggest that differences in steady-state levels and kinetics of G-CSF induced mobilization of donor lymphoid cells may in addition to other well-known factors affect the incidence of GvHD in HLA-identical recipients. However, owing to the small number of donor-recipient pairs studied, our results must be verified in a larger group of patients.

The effect of angiogenesis inhibitor TNP-470 on the blood vessels of the lungs, kidneys and livers of treated hamsters.

The growth of solid tumours and their metastases is dependent on the development of new blood vessels (angiogenesis). Therefore angiogenesis inhibitors are potential antitumour drugs. In our previous studies it was found that the angiogenesis inhibitor TNP-470 given to transplantable melanoma-bearing hamsters can decrease the rate of the tumour growth, although the survival time of the animals treated was not significantly affected. It was found finally that TNP-470 given in the vicinity of the growing tumour can cause complete remission of the melanoma in hamsters treated in this way. To check what side-effects could be evoked by such treatment, an examination of the morphology of the blood vessels of the lungs, kidneys and livers of the treated animals was carried out. It was found that the angiogenesis inhibitor applied did not cause any changes which could be observed by light and electron microscopes in the structure of the examined blood vessels of the treated animals.

Ultrastructure of the ventromedial hypothalamic nucleus in fasted and refed young and old rats.

Many hypothalamic nuclei are involved in the regulation of food intake and energy homeostasis. An ultrastructural investigation of the hypothalamic ventromedial nucleus (VMN), a hypothetical "satiety centre" was performed to explore the morphological basis of altered feeding behaviour of old rats in an experimental model of fasting/refeeding. Young (5 months old, n=12) and old (24 months old, n=12) male Wistar rats were fasted for 48 hours, then refed for 24 hours and sampled thereafter. Brain tissue was fixed by perfusion, histological and ultrathin sections were obtained by routine methods. Although food intake was similar in control young and old rats, during refeeding old animals consumed less chow than young ones. The EM analysis of VMN neurones of old control rats revealed, besides typical age-related residual bodies, deep indentations of the nuclear envelope and the presence of long, undulating rough endoplasmic reticulum cisternae in the cell periphery. In both young and old rats fasting for 48 hours led to the expansion of Golgi complexes and increased folds of the nuclear envelope, which is suggestive of enhanced cellular activity of the VMN neurones. These fasting-induced alterations were sustained in the VMN neurones of refed rats in both age groups. The results showed that the VMN neurones of old control rats differ at the ultrastructural level from young ones. However, starvation and subsequent refeeding cause similar alterations in the hypothalamic neurones of "satiety centre" of both young and old rats.

The effect of fasting and refeeding on the ultrastructure of the hypothalamic paraventricular nucleus in young and old rats.

In order to explore the morphological basis of the altered feeding behaviour of old rats, an ultrastructural investigation of the magnocellular neurons of the hypothalamic paraventricular nucleus (PVN) was performed. Young and old male Wistar rats, 5 and 24 months old, respectively, and with each age group comprising 12 animals, were divided into 3 groups. The rats in Group I were used as controls (normally fed), the rats of Group II were fasted for 48 hours and in Group III the rats were fasted for 48 hours and then refed for 24 hours. The brains were fixed by perfusion and histological and ultrathin sections were obtained by routine methods. Common features of the magnocellular PVN neurons of young and old rats were abundant Golgi complexes and short fragments of RER localised at the cell periphery. In contrast to young rats, the PVN neurons of old animals showed deep indentations of the nuclear envelope and age-related residual bodies. In both age groups fasting for 48 hours led to the expansion of the Golgi complexes and dilatation of RER cisternae. In contrast to those in fed rats, RER cisternae in the neurons of old fasted animals were situated between the nuclear envelope and the Golgi zone. Prolonged RER cisternae were distributed in the peripheral cytoplasm of refed old rats. Our observations suggest that at the ultrastructural level the process of ageing does not change the responsiveness of magnocellular PVN neurons to fasting-refeeding.

[Influence of erythropoietin on immunological system of patients with chronic renal failure]. / Wplyw erytropoetyny na uklad immunologiczny u chorych z przewlekla newydolnoscia nerek.

UNLABELLED: Recombinant human erythropoietin (epoetin) administration is a well established therapy of anaemia in maintenance hemodialysis (HD) patients. During the treatment, along with an increase in haemoglobin (Hb) level also an improvement of physical and sexual activity and cognitive functions was observed. Moreover, recent studies have shown an impact of epoetin on lipid-carbohydrate and protein metabolism, endocrinological functions and immune system [1-7]. It is still unclear whether all these changes are caused by direct epoetin activity or they are associated with the correction of anaemia and better oxygen supply and therefore an improvement of the conditions required for many metabolic function. The goal of the first study performed in our centre was to estimate the influence of epoetin alpha (Eprex) administered in the doses not affecting erythropoiesis (7-10 IU/kg/three times a week for 12 weeks) on serum levels of interleukin (IL) 2, 6 and tumor necrosis factor TNF-alpha [8, 9]. 10 HD patients (3 F, 7 M) aged from 33 to 62 years participated in that study. The level of IL-2, IL-6 and TNF-alpha was measured by means of bioassay using a highly sensitive cell line respectively CTLL, B9 and fibrosarcoma--WEHI 164 (clone 13). Cells viability was tested by colorimetric MTT assay. During the first period of observation stable Hb concentration and unchanged although significantly higher than in healthy people levels of TNF-alpha and IL-6 were noticed. Serum level of IL-2 increased significantly and in the 10 week it reached the values observed in healthy humans although after that period of time it dropped to the initial values. The aim of the following study was to estimate the influence of epoetin on IL-2, IL-10 and TNF-alpha production by whole blood cell culture [1, 2]. 10 HD patients (2 F, 8 M) aged from 35 to 53 years receiving standard doses of epoetin alpha (Eprex) for six months (in vivo experiments) and another 10 HD patients (3 F, 7 M) aged from 40 to 60 years not receiving epoetin participated in the study (in vitro experiments--cell culture were stimulated with different doses of (epoetin alpha--Eprex and epoetin beta--Recormon): 0.05; 0.1; 0.5; 1.0 IU/ml). IL 2 and TNF-alpha were measured using the bioassay mentioned above, IL 10 by ELISA immunoassay. The levels of IL10 increased in all epoetin treated patients and it was accompanied by transitory decrease of TNF-alpha. The levels of IL-2 increased in 7/10 patients under the study. Addition of epoetin in vitro to the whole blood culture of HD patients before implementation of epoetin confirmed that it is able to directly stimulate IL-2 production. The highest levels of stimulation of IL-2 secretion were observed for the physiological doses of epoetin (0.05 IU/ml). The aim of the more recent study was to examine changes in CD4+ and CD8+ T-cell subpopulations, the expression of the inhibitory molecule, CD152+ on T lymphocytes and the levels of IL-2, IL-6, IL-10, IL-12 and TNF-alpha in HD patients [10]. Additionally serum levels of C reactive protein (CRP), C3, C4 components of complement and immunoglobulin IgG, IgM and IgA were measured. 14 patients (8 F, 6 M) aged from 31 to 64 years receiving standard doses of epoetin alpha (Eprex) for twelve months (in vivo experiments) and another 4 HD patients (2 F, 2 M) aged from 43 to 57 years not receiving epoetin participated in the study (in vitro experiments--cell culture were stimulated with epoetin as in previous study). METHODS: IL-2, IL-6, and TNF-alpha were measured using bioassays described above, IL-12 and IL-10 by ELISA immunoassay. Expression of T-cell surface molecules was measured both in vivo by flow cytometry of lymphocytes sampled from peripheral blood and in vitro using whole blood cell culture stimulated with physiological as well as non-physiological doses of epoetin. The levels of C3, C4, IgG, IgM and IgA were estimated using nephelometric method. Compared with the findings before the start of epoetin therapy the CD4+/CD8+ ratio increased after 1 year of follow-up, whereas the percentage of CD152+ peripheral blood lymphocytes decreased. The increase of the CD4+/CD8+ ratio was dependent on a decrease of the percentage of CD8+ cells. The decrease of CD152+ population affected mainly CD8+152+ T cells. All these effects became apparent after 6 months of epoetin treatment. In vitro stimulation of whole blood cultures revealed that the addition of physiological concentration of epoetin decreased the percentage of CD8+152+ T cells. The pattern of the cytokines shifted towards Th1 phenotype (increase of IL-2 and IL-12) with a decreased level of proinflammatory cytokines (decrease of IL-6 and TNF-alpha). Treatment with epoetin did not alter plasma CRP, C3, C4 components of complement, immunoglobulin, as well as total count of lymphocytes. Summing up, administration of epoetin to maintenance HD patients not only treats the anaemia but also results in favourable changes in immune system. Epoetin is probably not only hemopoietic factor but also an immunomodulatory cytokine.

Analogues of muramyl dipeptide (MDP) and tuftsin limit infection and inflammation in murine model of sepsis.

Pharmacological manipulation of the balance between pro- and anti-inflammatory mediators emerges as a key aspect of a successful treatment of sepsis. A murine model of septic shock was developed and chosen conjugates (1a, 1b, 8a, 8c) and analogs (T2) of muramyl dipeptide and tuftsin were tested in this model as prospective anti-bacterial drugs or adjuvants. The phagocytic activity of monocytes/macrophages was determined (flow cytometry, bacterial clearance from vital organs). To evaluate cytokines levels (TNFalpha, IFNgamma, IL6, IL10) we used real-time PCR. The most promising immunomodulatory properties were displayed by the analogue T2 and two conjugates: 8a, 8c.