Nitric oxide-dependent expansion of erythroid progenitors in a murine model of chronic psychological stress.

Anaemia occurs frequently in patients with heart failure and its current treatment lacks clear targets. Emerging evidence suggests that erythroid progenitor cell expansion is an integral part of physiological response to anaemia associated with chronic stress. Understanding the underlying mechanism may provide a novel approach to anaemia management. In this study, we aimed to examine a role for nitric oxide (NO) in the regulation of bone marrow erythroid progenitor response to chronic stress. For this purpose, adult male mice were subjected to 2 h daily restraint stress for 7 or 14 consecutive days. The role of NO was assessed by subcutaneous injection with NG-nitro-L-arginine methyl ester, 30 min prior to each restraint. Chronic exposure to stress resulted in significantly increased number of bone marrow erythroid progenitors, and blockade of NO biosynthesis prior to daily stress completely prevented stress-induced erythroid progenitor cell expansion. Furthermore, chronic stress exposure led to altered expression of neural, endothelial and inducible nitric oxide synthases (NOS) in the bone marrow, both on mRNA and protein level. Decreased expression of neural and endothelial NOS, as well as reduced expression of NF-kappaB/p65 in bone marrow nuclear cell fraction, was accompanied by elevated bone marrow expression of inducible NOS in chronically stressed animals. This is the first study to demonstrate a role for NO in adaptive response of erythroid progenitors to chronic stress. Targeting NO production may be beneficial to improve bone marrow dysfunction and reduced erythroid progenitor cell expansion in chronic heart failure patients.

Proliferation and differentiation markers of colorectal adenocarcinomaand their correlation with clinicopathological factors.

BACKGROUND/AIM: The purpose of this study was to investigate proliferation and differentiation markers in colorectal adenocarcinoma and their correlation with clinicopathological factors. MATERIALS AND METHODS: Samples were collected from 38 patients with colorectal adenocarcinoma and 10 healthy controls. E-cadherin, carcinoembryonic antigen (mCEA), cyclin B1, vascular endothelial growth factor (VEGF), and erythropoietin (EPO) receptor (EPOR) were examined by immunohistochemistry; VEGF and EPO were examined by real-time PCR. RESULTS: The tumor samples were mostly characterized by large dimension (pT3), moderate level of differentiation (G2), negative lymph node status (N0), and no metastasis. Cyclin B1 and VEGF gene and protein expressions were significantly higher in tumor tissues than in control tissues; E-cadherin expression was significantly decreased in tumor samples and in positive correlation with mCEA. EPO was almost undetectable in tumor tissues of colorectal adenocarcinoma. Significant positive correlation was detected between tumor size and cyclin B1, tumor grade, and lymph node status. CONCLUSION: Decreased expression of EPO, high levels of VEGF and cyclin B1 expression, predominant moderate tumor differentiation, absence of metastasis, and negative lymph node status may suggest low level of aggressiveness, better prognosis, and longer patient survival.

Macrophage migration inhibitory factor is an endogenous regulator of stress-induced extramedullary erythropoiesis.

Macrophage migration inhibitory factor is a well-known proinflammatory cytokine that is released during systemic stress response. Although MIF can affect erythrocyte production, the role of this cytokine in stress-induced erythropoiesis is completely unknown. To extend our previous findings showing that chronic psychological stress stimulates extramedullary erythropoiesis, here we examined whether MIF is involved in the control of stress-induced erythropoietic response. Adult male C57BL/6 wild-type (WT) and MIF-KO (knock-out) mice were subjected to 2-h daily restraint stress for either 7 or 14 consecutive days. The number of erythroid progenitors and CD71/Ter119 profile of erythroid precursors were analyzed in the bone marrow and spleen. Additionally, MIF protein expression was assessed in WT mice. Our results demonstrated that chronic restraint stress enhanced the number of both erythroid progenitors and precursors in the spleen. Stress-induced increase in the number of splenic late erythroid progenitors as well as in the percentage of CD71(+)Ter119(+)-double-positive precursors was significantly more pronounced in MIF-KO mice compared to WT animals. Furthermore, repeatedly stressed WT animals demonstrated an augmented MIF expression in the spleen. Unlike the spleen, the bone marrow of chronically stressed WT mice exhibited less prominent changes in erythropoietic stress response and no significant alteration in MIF expression. In addition, MIF deficiency did not influence the bone marrow erythropoiesis in stressed animals. These findings suggest that MIF regulates extramedullary erythropoiesis by inhibiting an overexpansion of splenic immature erythroid cells during chronic stress and indicate a novel role for this cytokine under chronic stress conditions.

Doxycycline Inhibits IL-17-Stimulated MMP-9 Expression by Downregulating ERK1/2 Activation: Implications in Myogenic Differentiation.

Interleukin 17 (IL-17) is a cytokine with pleiotropic effects associated with several inflammatory diseases. Although elevated levels of IL-17 have been described in inflammatory myopathies, its role in muscle remodeling and regeneration is still unknown. Excessive extracellular matrix degradation in skeletal muscle is an important pathological consequence of many diseases involving muscle wasting. In this study, the role of IL-17 on the expression of matrix metalloproteinase- (MMP-) 9 in myoblast cells was investigated. The expression of MMP-9 after IL-17 treatment was analyzed in mouse myoblasts C2C12 cell line. The increase in MMP-9 production by IL-17 was concomitant with its capacity to inhibit myogenic differentiation of C2C12 cells. Doxycycline (Doxy) treatment protected the myogenic capacity of myoblasts from IL-17 inhibition and, moreover, increased myotubes hypertrophy. Doxy blocked the capacity of IL-17 to stimulate MMP-9 production by regulating IL-17-induced ERK1/2 MAPK activation. Our results imply that MMP-9 mediates IL-17's capacity to inhibit myoblast differentiation during inflammatory diseases and indicate that Doxy can modulate myoblast response to inflammatory induction by IL-17.

Bradykinin stimulation of nitric oxide production is not sufficient for gamma-globin induction.

INTRODUCTION: Hydroxycarbamide, used in therapy of hemoglobinopathies, enhances nitric oxide (NO) production both in primary human umbilical vein endothelial cells (HUVECs) and human bone marrow endothelial cell line (TrHBMEC). Moreover, NO increases gamma-globin and fetal hemoglobin levels in human erythroid progenitors. OBJECTIVE: In order to find out whether simple physiologic stimulation of NO production by components of hematopoietic microenvironment can increase gamma-globin gene expression, the effects of NO-inducer bradykinin were examined in endothelial cells. METHODS: The study was performed in co-cultures of human erythroid progenitors, TrHBMEC and HUVECs by ozone-based chemiluminescent determination of NO and real-time quantitative RT-PCR. RESULTS: In accordance with previous reports, the endogenous factor bradykinin increased endothelial cell production of NO in a dose- and time-dependent manner (0.1-0.6 microM up to 30 minutes).This induction of NO in HUVECs and TrHBMEC by bradykinin was blocked by competitive inhibitors of NO synthase (NOS), demonstrating NOS-dependence. It has been shown that bradykinin significantly reduced endothelial NOS (eNOS) mRNA level and eNOS/beta-actin ratio in HUVEC (by twofold). In addition, bradykinin failed to increase gamma-globin mRNA expression in erythroid progenitors only, as well as in co-culture studies of erythroid progenitors with TrHBMEC and HUVEC after 24 hours of treatment. Furthermore, bradykinin did not induce gamma/beta globin ratio in erythroid progenitors in co-cultures with HUVEC. CONCLUSION: Bradykinin mediated eNOS activation leads to short time and low NO production in endothelial cells, insufficient to induce gamma-globin gene expression. These results emphasized the significance of elevated and extended NO production in augmentation of gamma-globin gene expression.

Transcriptional profiling of erythroid progenitors from G-CSF mobilized and nonmobilized peripheral blood.

PURPOSE: The purpose of this study was to examine the gene expression profile of granulocyte colony stimulating factor (G-CSF)-mobilized peripheral blood (mPB)-derived progenitors, used in transplantation. METHODS: We correlated gene expression patterns of highly enriched steady-state peripheral blood (PB)- and mPB-derived CD71+ cells by microarray and ingenuity pathway analyses, to identify the transcriptional program during in vitro erythroid differentiation. RESULTS: The gene expression was more than doubled in mPB-derived (4180 genes) compared to PB-derived erythroid progenitors (1667 genes) while PB-and mPB-derived erythroid progenitors shared 1534 common genes. Comparative analysis of transcript levels showed differential expression of 54 genes between cultured erythroid progenitors of PB and mPB origin, where we identified common 13 downregulated and 30 upregulated genes. The most significant genes in mPB-derived erythroid progenitors were P4HB, DDIA3, ARPC2 and ATP5G3. Regarding G-CSF stimulation the G-CSF receptor CSF2RB (1.1-fold) was linked via STAT3 to erythroid-specific ALAS2 (2.9-fold) and GATA2 (1.3-fold) factors, all upregulated in mPB-derived erythroid progenitors, coupled to common upregulated NUDC gene involved in the proliferation of erythroid cells. CONCLUSION: This report provides an extensive transcriptional profile of cultured erythroid progenitors and leads to a better understanding of diversity among the progenitor sources.

Chronic psychological stress activates BMP4-dependent extramedullary erythropoiesis.

Psychological stress affects different physiological processes including haematopoiesis. However, erythropoietic effects of chronic psychological stress remain largely unknown. The adult spleen contains a distinct microenvironment favourable for rapid expansion of erythroid progenitors in response to stressful stimuli, and emerging evidence suggests that inappropriate activation of stress erythropoiesis may predispose to leukaemic transformation. We used a mouse model to study the influence of chronic psychological stress on erythropoiesis in the spleen and to investigate potential mediators of observed effects. Adult mice were subjected to 2 hrs daily restraint stress for 7 or 14 consecutive days. Our results showed that chronic exposure to restraint stress decreased the concentration of haemoglobin in the blood, elevated circulating levels of erythropoietin and corticosterone, and resulted in markedly increased number of erythroid progenitors and precursors in the spleen. Western blot analysis revealed significantly decreased expression of both erythropoietin receptor and glucocorticoid receptor in the spleen of restrained mice. Furthermore, chronic stress enhanced the expression of stem cell factor receptor in the red pulp. Moreover, chronically stressed animals exhibited significantly increased expression of bone morphogenetic protein 4 (BMP4) in the red pulp as well as substantially enhanced mRNA expression levels of its receptors in the spleen. These findings demonstrate for the first time that chronic psychological stress activates BMP4-dependent extramedullary erythropoiesis and leads to the prolonged activation of stress erythropoiesis pathways. Prolonged activation of these pathways along with an excessive production of immature erythroid cells may predispose chronically stressed subjects to a higher risk of leukaemic transformation.

Characteristics of human adipose mesenchymal stem cells isolated from healthy and cancer affected people and their interactions with human breast cancer cell line MCF-7 in vitro.

Adipose tissue is an attractive source of mesenchymal stem/stromal cells (MSCs) with potential applications in reconstructive plastic surgery and regenerative medicine. The aim of this study was to characterise human adipose tissue MSCs (ASCs) derived from healthy individuals and cancer patients and to compare their interactions with tumour cells. ASCs were isolated from adipose tissue of healthy donors, breast cancer-adjacent adipose tissue of breast cancer patients and tumour-adjacent adipose tissue of non-breast cancer patients. Their proliferation, differentiation, immunophenotype and gene expression were assessed and effects on the proliferation of human breast cancer cell line MCF-7 compared. ASCs from all sources exhibited similar morphology, proliferative and differentiation potential, showing the characteristic pattern of mesenchymal surface markers expression (CD90, CD105, CD44H, CD73) and the lack of HLA-DR and hematopoietic markers (CD11a, CD33, CD45, Glycophorin-CD235a), but uneven expression of CD34. ASCs also shared a common positive gene expression of HLA-DR, HLA-A, IL-6, TGF-ß and HIF-1, but were negative for HLA-G, while the expression levels of Cox-2 and IDO-1 varied. All ASCs significantly stimulated the proliferation of MCF-7 tumour cells in direct mixed co-cultures and transwell system, although their conditioned media displayed antiproliferative activity. Data obtained showed that ASCs with similar characteristics are easily isolated from various donors and sites of origin, although ASCs could both suppress and favour tumour cells growth, emphasising the importance of cellular context within the microenvironment and pointing to the significance of safety studies to exclude any potential clinical risk of their application in regenerative medicine.

WITHDRAWN: BRCA1 protein expression and TOP2A gene aberration and protein expression in four molecular subtypes of breast cancer.

Ahead of Print article withdrawn by publisher.

Mesenchymal stem cells isolated from peripheral blood and umbilical cord Wharton's jelly.

INTRODUCTION: Mesenchymal stem cells (MSCs) are a promising tool for regenerative medicine, but due to the heterogeneity of their populations, different sources and isolation techniques, the characteristics defining MSCs are inconsistent. OBJECTIVE: The aim of this study was to compare the characteristics of MSCs derived from two different human tissues: peripheral blood (PB-MSCs) and umbilical cord Wharton's Jelly (UC-MSCs). METHODS: The PB-MSC and UC-MSC were isolated by adherence to plastic after gradient-density separation or an explant culture method, respectively, and compared regarding their morphology, clonogenic efficiency, proliferating rates, immunophenotype and differentiation potential. RESULTS: MSCs derived from both sources exhibit similar morphology, proliferation capacity and multilineage (osteogenic, chondrogenic, adipogenic and myogenic) differentiation potential. Differences were observed in the clonogenic capacity and the immunophenotype, since UC-MSCs showed higher CFU-F (colony-forming units-fibroblastic) cloning efficiency, as well as higher embryonic markers (Na-nog, Sox2, SSEA4) expression. When additional surface antigens were analyzed by flow cytometry (CD44, CD90, CD105, CD33, CD34, CD45, CD11b, CD235a) or immunofluorescent labeling (vimentin, STRO-1 and alpha-smooth muscle actin), most appeared to have similar epitope profiles irrespective of MSC source. CONCLUSION: The results obtained demonstrated that both MSCs represent good alternative sources of adult MSCs that could be used in cell therapy applications.

The potential of interleukin-17 to mediate hematopoietic response.

It has long been known that T cells have the potential to modulate hematopoietic response in different ways. More recently, the importance of interleukin (IL)-17-secreting Th17 cells in T-cell-mediated regulation of hematopoiesis was indicated by the line of evidence that IL-17 links T-cell function and hematopoiesis through stimulation of granulopoiesis and neutrophil trafficking. Furthermore, our data demonstrated that IL-17 also affects other cells of hematopoietic system, such as erythroid progenitors, as well as mesenchymal stem cells. In order to better understand the regulatory role of IL-17 in hematopoiesis, molecular mechanisms underlying the effects of IL-17 on hematopoietic and mesenchymal stem cells were also studied.

IL-17 and FGF signaling involved in mouse mesenchymal stem cell proliferation.

The mouse is a suitable experimental model to study the biology of mesenchymal stem cells (MSCs), as well as to be used in biocompatibility studies and tissue engineering models. However, the isolation and purification of murine MSCs is far more challenging than their counterparts from other species. In this study, we isolated, expanded and characterized mouse MSCs from bone marrow (BM-MSCs). Additionally, we analyzed the effects of two regulatory molecules, interleukin 17 (IL-17) and basic fibroblast growth factor (bFGF), on BM-MSCs growth and elucidated the signaling pathways involved. The results revealed that IL-17 increased the frequency of colony-forming units fibroblast (CFU-F) as well as the BM-MSCs proliferation in a dose-dependent manner, while bFGF supplementation had no significant effect on CFU-F frequency but induced an increase in cell proliferation. Their combined usage did not produce additive effects on BM-MSCs proliferation and even induced reduction in the number of CFU-F. Also, the involvement of both p38 and extracellular signal-regulated kinase (ERK) mitogen-activated protein kinases (MAPKs) signaling in proliferative activity of IL-17 and bFGF on murine BM-MSCs and, moreover, the increased co-activation of a common signaling molecule, p38 MAPK, were demonstrated. Together, the data presented highlighted the role of IL-17 and bFGF in murine BM-MSCs proliferation and pointed to the complexity and specificity of the signaling networks leading to MSCs proliferation in response to different regulatory molecules.

SKIP is required for TGF-ß1-induced epithelial mesenchymal transition and migration in transformed keratinocytes.

Transforming growth factor-ß1 (TGF-ß1) potently induces the epithelial-mesenchymal transition (EMT) during tumoral progression. Although Sky-interacting protein (SKIP) regulates TGF-ß1-induced Smad activation, its role in the induction of cell malignance remains uncertain. We found that TGF-ß1 increases SKIP expression in PDV cells. In cells stably transfected with SKIP antisense, AS-S, Smad3 activation decreased, along with an inhibition of TGF-ß1-induced EMT, and the cells were sensitized to the TGF-ß1-dependent inhibition of proliferation. Also, AS-S cells showed a weaker migration and invasion response. Moreover, TGF-ß1-induced urokinase-type plasminogen activator expression was inhibited, concomitantly with a TGF-ß1-independent increment of the plasminogen-activator inhibitor-1 expression. Thus, these results suggest that SKIP is required for EMT and invasiveness induced by TGF-ß1 in transformed cells.

Syphacia obvelata modifies mitogen-activated protein kinases and nitric oxide synthases expression in murine bone marrow cells.

Syphacia obvelata is a rodent nematode parasite with high prevalence in laboratory mice. In our previous work we have demonstrated that this gut-dwelling helminth induces significant hematopoietic changes, characterized by increased myelopoiesis and erythropoiesis in infected animals, and accompanied with altered reactivity of bone marrow hematopoietic progenitors to interleukin (IL)-17. In this study we extended these investigations by demonstrating that naturally acquired S. obvelata infection induces significant alterations in murine bone marrow cells manifested at the molecular level. Namely, S. obvelata infection induced sustained phosphorylation of the members of three major groups of distinctly regulated mitogen-activated protein kinases (MAPKs), the p38, the c-Jun amino-terminal kinase (JNK) and the extracellular signal-regulated kinase (ERK), as well as enhanced expression of mRNA for the inducible nitric oxide synthase (iNOS) in the bone marrow cells of infected animals. Furthermore, the infection interfered with the IL-17-mediated effects in bone marrow cells, since in normal mice IL-17 significantly enhanced phosphorylation of p38 MAPK and upregulated the expression of iNOS and the constitutive, endothelial (e)NOS mRNA, while in S. obvelata-infected animals IL-17 did not influence the MAPKs activation, but markedly down-regulated the expression of both NOS isoforms. The data obtained demonstrating that S. obvelata is able to manipulate signal transduction pathways in the hosts' bone marrow cells, pointed to the multiple layers of immunomodulatory ability of this pinworm parasite and highlighted the importance of working under pinworm-free conditions when using experimental murine models for immunohematopoietic investigations.

Low O2 concentrations enhance the positive effect of IL-17 on the maintenance of erythroid progenitors during co-culture of CD34+ and mesenchymal stem cells.

Co-culture of haematopoietic cells with a stromal cell layer does not mimic the physiological, micro-environmental niche, whose major feature is a low oxygen (O2) concentration. Thus, in order to study the effects of IL-17 in a context which better approximates the physiological state, we investigated its effects on cell expansion, colony-forming ability, and the phenotypical profile of normal, human blood CD34+ cells co-cultured for five days with MSC layers at various O2 concentrations (20%, 12.5% and 3% O2. We demonstrated that IL-17 enhances CD34+ and total CFC production during the five days of MSC/CD34+ co-culture. This effect depends upon the O2 concentration, reaching its maximum at 3% O2, and is more pronounced on erythroid progenitors (BFU-E). In addition, the stimulation of IL-6 production by IL-17 in MSC cultures and co-cultures is enhanced by low O2 concentration. The expression of some differentiation markers (CD34, CD13 and CD41) on haematopoietic cells in co-cultures also depends upon the oxygen concentration. Our results strengthen the concept that physiological levels of O2 (mistakenly called hypoxia), should be considered as an important environmental factor that significantly influences cytokine activity.

p38 MAPK signaling mediates IL-17-induced nitric oxide synthase expression in bone marrow cells.

The effects of interleukin (IL)-17 on nitric oxide (NO) synthase (NOS) expression, as well as the participation of mitogen-activated protein kinases (MAPKs) in IL-17-mediated effects were examined in murine bone marrow cells. The results demonstrated the ability of IL-17 to upregulate the expression of mRNA for both inducible NOS and constitutive, endothelial NOS isoforms, as well as to enhance the phosphorylation of p38 MAPK. Moreover, both the NOS-inducing effect of IL-17 and the in vitro IL-17-mediated inhibition colony forming unit-erythroid (CFU-E) growth were dependent on p38 MAPK activity. The data demonstrating that the in vivo reducing effect of IL-17 on bone marrow CFU-E was prevented by co-treatment with the NOS inhibitor Nw-nitro-l-arginine methyl ester hydrochloride (L-NAME), implied that this effect is mediated through NOS activation. Besides revealing a link between the IL-17, NO, and haematopoiesis, data presented gave an insight into the mechanisms by which IL-17 exerts its modulatory effects on bone marrow cells.

Interleukin-6 (IL-6) and low O(2) concentration (1%) synergize to improve the maintenance of hematopoietic stem cells (pre-CFC).

Low O(2) concentration (1%) favors the self-renewal of hematopoietic stem cells and inhibits committed progenitors (CFC). Since IL-6 influences both stem cells and committed progenitors at 20% O(2), we studied its effects in cultures at 1% O(2). The pre-CFC activity in Lin- population of mouse bone marrow was analyzed following 10 days of serum-free culture in medium (LC1) supplemented with IL-3 with and without IL-6, at 20 and 1% O(2) and phenotypic differentiation and proliferative history monitored. The IL-6 receptor expression and initiation of VEGF-A synthesis were also investigated. At 20% O(2), the effects of IL-6 on pre-CFC were negligible but effects on CFC were apparent; conversely, at 1% O(2), the IL-6 enhances activity of pre-CFC but not of CFC. Unlike at 20% O(2), at 1% O(2) a subpopulation of cells remained Lin- in spite of extensive proliferation. However, the absolute number of Lin- cells, did not correlate with pre-CFC activity. A relative increase in VEGF transcripts at 1% O(2) in presence of IL-3 alone was enhanced by the addition of IL-6. IL-6 enhanced pre-CFC activity at 1% O(2) and this was correlated to the induction of VEGF. These data reinforce the concept that physiologically low oxygenation of bone marrow is a regulator of stem cell maintenance. Since the 20% O(2) does not exist in tissues in vivo, further studies in vitro at lower O(2) concentrations should revise our knowledge relating to cytokine effects on stem and progenitor cells.

Signaling pathways implicated in hematopoietic progenitor cell proliferation and differentiation.

The objective of this study was to investigate the signal transduction pathways associated with the clonal development of myeloid and erythroid progenitor cells. The contribution of particular signaling molecules of protein tyrosine kinases (PTKs), mitogen-activated protein (MAP) kinase, and PI-3 kinase signaling to the growth of murine bone marrow colony forming unit-granulocyte-macrophage (CFU-GM) and erythroid (burst forming unit-erythroid [BFU-E] and colony forming unit-erythroid [CFU-E]) progenitors was examined in studies performed in the presence or absence of specific signal transduction inhibitors. The results clearly pointed to different signal transducing intermediates that are involved in cell proliferation and differentiation depending on the cell lineage, as well as on the progenitors' maturity. Lineage-specific differences were obtained when chemical inhibitors specific for receptor- or nonreceptor-PTKs, as well as for the main groups of distinctly regulated MAPK cascades, were used because all of these compounds suppressed the growth of erythroid progenitors, with no major effects on myeloid progenitors. At the same time, differential involvement of MEK/extracellular signal-regulated kinase (ERK) MAPK transduction pathway was observed in the proliferation and/or differentiation of early, BFU-E, and late, CFU-E, erythroid progenitor cells. The results also demonstrated that phosphatydylinositol (PI)-3 kinase and nuclear factor kappaB (NF-kappaB) transcriptional factor were required for maintenance of both myeloid and erythroid progenitor cell function. Overall, the data obtained indicated that committed hematopoietic progenitors express a certain level of constitutive signaling activity that participates in the regulation of normal steady-state hematopoiesis and point to the importance of evaluating the impact of signal transduction inhibitors on normal bone marrow when used as potential therapeutic agents.

Hematopoietic changes and altered reactivity to IL-17 in Syphacia obvelata-infected mice.

Pinworm parasites commonly infect laboratory mice with high prevalence even in well-managed animal colonies. Although often considered as irrelevant, these parasites if undetected may significantly interfere with the experimental settings and alter the interpretation of final results. There are a few reports documenting the effects of pinworms on research and the effects of pinworms on the host hematopoiesis have not yet been investigated. In this study we examined the changes within various hematopoietic cell lineages in the bone marrow, spleen, peripheral blood and peritoneal space during naturally acquired Syphacia obvelata infection in inbred CBA mice. The data obtained showed significant hematopoietic alterations, characterized by increased myelopoiesis and erythropoiesis in S. obvelata-infected animals. In order to additionally evaluate if this pinworm infection modifies hematopoietic cells' reactivity, we examined the effect of murine interleukin-17, T cell-derived cytokine implicated in the regulation of hematopoiesis and inflammation, on the growth of bone marrow progenitor cells and demonstrated that bone marrow myeloid and erythroid progenitors from S. obvelata-infected mice displayed altered sensitivity to IL-17 when compared to non-infected controls. Taken together the alterations presented pointed out that this rodent pinworm is an important environmental agent that might significantly modify the hosts' hematopoietic response, and therefore interfere with the experimental settings and alter the interpretation of the final results. However, the results obtained also contributed new data concerning the activity of IL-17 on bone marrow hematopoietic cells, supporting our previous reports that depending on physiological/pathological status of the organism IL-17 exerts differential effects on the growth of progenitor cells.

Interleukine-17-induced inhibitory effect on late stage murine erythroid bone marrow progenitors.

Recent studies have shown that the T cell-derived cytokine, interleukin-17 (IL-17), stimulates hematopoiesis, specifically granulopoiesis inducing expansion of committed and immature progenitors in bone marrow. Our previous results pointed to its role in erythropoiesis too, demonstrating significant stimulation of BFU-E and suppression of CFU-E growth in the bone marrow from normal mice. As different sensitivities of erythroid and myeloid progenitor cells to nitric oxide (NO) were found, we considered the possibility that the observed effects of IL-17 were mediated by NO. The effects of recombinant mouse IL-17, NO donor (sodium nitroprusside - SNP) and two NO synthases inhibitors (L-NAME and aminoguanidine) on erythroid progenitor cells growth, as well as the ability of IL-17 to induce nitric oxide production in murine bone marrow cells, were examined. In addition, we tested whether the inhibition of CFU-E colony formation by IL-17 could be corrected by erythropoietin (Epo), the principal regulator of erythropoiesis. We demonstrated that IL-17 can stimulate low level production of NO in murine bone marrow cells. Exogenously added NO inhibited CFU-E colony formation, whereas both L-NAME and aminoguanidine reversed the CFU-E suppression by IL-17 in a dose-dependent manner. The inhibition of CFU-E by IL-17 was also corrected by exposure to higher levels of Epo. The data obtained demonstrated that at least some of the IL-17 effects in bone marrow related to the inhibition of CFU-E, were mediated by NO generation. The fact that Epo also overcomes the inhibitory effect of IL-17 on CFU-E suggests the need for further research on their mutual relationship and co-signalling.