Erythropoietin administration is associated with improved T-cell properties in patients with myelodysplastic syndromes.

The immune system is impaired in myelodysplastic syndromes (MDS) and plays a role in the pathogenesis of the disease. Here we show effects of recombinant human erythropoietin (rHuEPO) on T cell (CD4+, CD8+ and CD4+CD25+) number and function in MDS patients. Healthy (20 subjects), MDS patients without rHuEPO treatment ('MDS', 13), and MDS patients treated with rHuEPO ('MDS+EPO', 17) were examined. CD4+ and CD8+ T cell numbers were reduced and increased respectively in MDS compared to healthy subjects. EPO treatment normalized these levels. CD4+CD25+ cell numbers, lower in MDS, were normalized in MDS+EPO. In vitro activation of CD4+ and CD8+ cells with phytohemagglutinin as measured by CD69 expression, demonstrated a 7.2 fold increase in CD4+ activation vs 13.6 fold for MDS and MDS+EPO respectively (p=0.004); and 10.2 fold (MDS) vs 18.6 fold (MDS+EPO, p<0.003) for CD8+ T cells. Expression of the co-stimulatory marker CD28, decreased in CD4+ and CD8+ T cells in MDS, was normalized in MDS+EPO CD4+ T cells. Subgroup analysis of milder disease (WHO RA and RARS) and more advanced disease revealed no difference in CD4+ and CD8+ T cell numbers. However, the activation of these cells in the RA/RARS subgroup was impaired in EPO-untreated and enhanced in EPO-treated MDS patients. Our data suggest that EPO treatment improves immune abnormalities in MDS and may depend on disease severity.

Erythropoietin treatment is associated with an augmented immune response to the influenza vaccine in hematologic patients.

The objective of this study was to examine whether treatment with recombinant human erythropoietin (rHuEPO), previously found to be associated with a positive effect on cell-mediated immunity and humoral immunity (hepatitis B vaccine), is associated with an improved response to the seasonal influenza (flu) vaccine. Three groups received flu vaccine: healthy controls, hematologic patients not treated with rHuEPO ("No EPO" group), and hematologic patients receiving rHuEPO for their anemia ("EPO" group). Anti-flu Ab titer was measured (complement fixation test) from blood samples drawn before and approximately 3-4 weeks, 7-8 weeks and 4 months after vaccination. Nineteen healthy subjects were compared with 17 No EPO and 17 EPO patients. Mean ages were 59.5, 61.3, and 73.1 years, respectively (EPO patients were older; p = 0.005). In the healthy group, the percentage of those sustaining only a partial (twofold) response, a strong (fourfold or greater) response, and an overall response (combined partial and strong responses) were 31.6%, 57.9%, and 89.5%, respectively. In the No EPO group, values were 35.3%, 17.6%, and 52.9%, respectively. EPO group results were similar to those of the healthy controls: 23.5%, 58.8%, and 82.4% (p = 0.016, EPO vs. No EPO). In conclusion, hematologic patients (NoEPO group) respond poorly to the flu vaccine, compared with healthy subjects, and rHuEPO treatment is associated with an improved immune response to the flu vaccine in hematologic patients, with titers similar to those of healthy subjects.

Non-erythroid effects of erythropoietin: are neutrophils a target?

We have previously shown that erythropoietin (EPO) potentiates the immune response. Analysis of various possible cellular mediators was performed on EPO-injected mice and transgenic mice overexpressing human EPO (tg6). Here we present our studies on neutrophils, peritoneal (casein induced), and from the peripheral blood, spleen and bone marrow. Neutrophil counts were elevated in peripheral blood and spleens of the tg6 mice, yet, no other EPO-associated effects were detected in the count and function of the different neutrophil populations. Hence, neutrophils are probably not mediators of the EPO immunological effects, although their counts may be affected by extreme EPO levels.

Non-erythroid activities of erythropoietin: Functional effects on murine dendritic cells.

Erythropoietin (EPO) is the main hormone that promotes proliferation and differentiation of erythroid progenitor cells via binding to its surface receptor (EPO-R). Recent studies suggest that this hormone may affect also other cell types, besides the red blood cell lineage. We have previously demonstrated that the immune system is a target of EPO; however, the direct target cells of EPO, as well as the molecular mechanisms underlying its role as an immunomodulator, are unknown. Here we present evidence for functional effects of EPO on dendritic cells (DCs), which are known to initiate the immune response. In-vivo experiments in EPO-injected mice and in transgenic mice over-expressing human EPO showed an increased splenic DC population with a higher cell surface expression of CD80 and CD86. Further analysis based on mouse models, showed that DCs derived in-vitro from bone marrow (BM-DCs) express EPO-R mRNA. In-vitro stimulation of these DCs with recombinant human EPO enhanced viability, upregulated CD80, CD86 and MHC class II and augmented the secretion of IL-12. Biochemical analysis of EPO mediated signaling in the BM-DCs showed activation of the AKT, MAPK and NF-kappaB pathways. EPO stimulation of the BM-DCs led to Tyr-phosphorylation of STAT3. The inability to detect EPO mediated activation of STAT5 in the BM-DCs, suggests that in DCs, STAT3 may play a more important role than STAT5 in EPO-R signaling. Taken together, our data support the premise that DCs are direct targets of EPO, thereby providing an insight to the immunomodulatory functions of EPO.

Erythropoietin effects on dendritic cells: potential mediators in its function as an immunomodulator?

OBJECTIVE: Modulatory effects of erythropoietin (EPO) on the cellular and humoral compartments of the immune system have been described; however, the mechanism of action by which EPO affects the lymphocyte number and functions has yet to be elucidated. Because no EPO receptors (EPO-R) could be detected on lymphocytes, we searched for cells that might express the EPO-R and thereby mediate these immunomodulatory effects. We thus focused on dendritic cells (DCs), the most potent antigen-presenting and T-cell-priming cells, as possible mediators of the immunomodulatory actions of EPO. MATERIALS AND METHODS: We examined the in vitro effects of EPO on human DCs. Expression of EPO-R, expression of costimulatory molecules, antigen uptake, secretion of cytokines, and DC maturation were investigated. RESULTS: We demonstrate that the EPO-R is expressed in human DCs and that EPO directly affects their phenotype and function. When applied in vitro, EPO increased the percentage of peripheral blood DCs and monocyte-derived DCs (MoDCs) expressing the costimulatory molecules CD80 and CD86. EPO treatment of MoDCs was also associated with an increase in surface expression of CD80 and CD86 as well as that of HLA-DR. EPO enhanced MoDC function, as manifested in increased antigen uptake and secretion of interleukin 12. When applied to immature MoDCs, EPO in itself induced their maturation. CONCLUSION: Our finding that DCs are directly affected by EPO renders them as potential candidates that mediate the immunomodulatory actions of EPO.

Erythropoietin enhances immune responses in mice.

Erythropoietin (Epo) is the main erythropoietic hormone. Recombinant human Epo (rHuEpo) is thus used in clinical practice for the treatment of anemia. Accumulating data reveals that Epo exerts pleiotropic activities. We have previously shown an anti-neoplastic activity of Epo in murine multiple myeloma (MM) models, and in MM patients. Our findings that this anti-neoplastic effect operates via CD8+ T lymphocytes led us to hypothesize that Epo possesses a wider range of immunomodulatory functions. Here we demonstrate the effect of Epo on B lymphocyte responses, focusing on three experimental models: (i) tumor-bearing mice, (5T2 MM mouse); (ii) antigen-injected healthy mice; and (iii) antigen-injected transgenic mice (tg6), overexpressing human Epo. In the MM model, despite bone marrow dysfunction, Epo-treated mice retained higher levels of endogenous polyclonal immunoglobulins, compared to their untreated controls. In both Epo-treated wild type and tg6 mice, Epo effect was manifested in the higher levels of splenocyte proliferative response induced in vitro by lipopolysaccharide. Furthermore, these mice had increased in vivo production of anti-dinitrophenyl (DNP) antibodies following immunization with DNP-keyhole limpet hemocyanin. Epo-treated mice showed an enhanced immune response also to the clinically relevant hepatitis B surface antigen. These findings suggest a potential novel use of rHuEpo as an immunomodulator.

Erythropoietin treatment in advanced multiple myeloma is associated with improved immunological functions: could it be beneficial in early disease?

Erythropoietin (Epo) is the main growth regulator of red blood cells, and recombinant human erythropoietin (rHuEpo) is thus used in clinical practice for the treatment of anaemia, primarily in kidney disease and cancer. rHuEpo treatment was found to be associated with prolonged survival of multiple myeloma (MM) patients. This clinical observation was then supported by studies on murine myeloma models. It thus appeared that rHuEpo had an anti-myeloma effect, causally related to an immunomodulatory function of rHuEpo. The present study investigated whether rHuEpo-treated MM patients acquire improved immunological functions. Treatment with rHuEpo, prescribed for anaemia that occurs in advanced disease, was associated with effects on a variety of immunological parameters and functions. This was expressed in an actual normalisation of the CD4:CD8 cell ratio, enhanced T cell phytohaemagglutinin-mediated activation and proliferation potential, T cell expression of the costimulatory CD28 and inhibitory CTLA-4 molecules, as well as reduced interleukin-6 serum values to normal levels. Furthermore, it was demonstrated that immunological abnormalities manifest in patients even in the early stages of MM. Our findings thus suggest that rHuEpo treatment might be effective in the early stages of MM, before anaemia develops. It is expected that this would boost the immune system, consequently achieving an anti-myeloma function; affecting disease progression and improving the prognosis.

Erythropoietin in clinical practice: current use, effect on survival, and future directions.

Recombinant human erythropoietin has become an essential part of the management of anemic patients with end-stage renal disease. It is also used to treat the anemia associated with cancer and other diseases, and it improves quality of life. In recent years, studies in animals and humans have focused on the use of rHuEPO for other indications. It has been found to play a role in both cardioprotection and neuroprotection. It has effects on the immune system, and can cause regression in hematologic diseases such as multiple myeloma. It may also improve the response of solid tumors to chemotherapy and radiation therapy. On the other hand, concerns have been raised following two studies of patients with solid tumors in whom those treated with rHuEPO had diminished survival. Criticism of the design of these studies makes it clear that large, well-designed, randomized trials must be performed to determine the role of rHuEPO in the treatment of cancer, and more generally to clarify the full clinical benefits of the drug, while minimizing the harm.