ABSTRACT
Chronic idiopathic neutropenia (CIN) is a granulocytic disorder associated with presence of activated, myelosuppressive T-lymphocytes. In the present study we have evaluated constituents of humoral immunity in CIN patients (n=48) compared to healthy controls (n=52). CIN patients displayed lower serum IgG levels due to a reduction in IgG1, IgG3, IgG4 but not IgG2, lower IgA and increased IgM levels compared to controls. The proportion of CD19+ cells did not differ between patients and controls; however the proportion of the naïve IgD+/CD27- B-cells was increased and the proportion of class-switched memory IgD-/CD27+ B-cells was decreased in the patients. The percentage of CD40+ B-cells did not differ between patients and controls and no aberrations in the CD40-meadiated signal transduction pathway or in CD40-gene polymorphisms were identified. These data provide further evidence that immune disturbances are associated with the pathophysiology of CIN and point out for the first time the implication of the B-cell system.
Subject(s)
B-Lymphocyte Subsets/immunology , B-Lymphocytes/immunology , Immunoglobulins/immunology , Neutropenia/immunology , Adolescent , Adult , Aged , CD40 Antigens/immunology , CD40 Ligand/immunology , Case-Control Studies , Chronic Disease , Female , Humans , Immunoglobulin Class Switching , Immunoglobulin D/immunology , Immunoglobulin G/immunology , Immunoglobulin M/immunology , Male , Middle Aged , Signal Transduction , Tumor Necrosis Factor Receptor Superfamily, Member 7/immunology , Young AdultABSTRACT
Chronic idiopathic neutropenia (CIN) is a bone marrow (BM) disorder characterized by presence of activated T-lymphocytes in peripheral blood (PB) and BM. We investigated the pattern of T-cell responses in CIN by analyzing the T-cell receptor ß-chain variable (Vß) gene repertoire. Compared to controls, CIN patients displayed different patterns of Vß gene usage in PB CD3(+), CD4(+) and CD8(+) cells. The frequency of Vß skewing and the number of expanded Vß families per subject were higher in patients compared to controls in all cell subpopulations. Skewing was more profound within the CD8(+) cells. The number of Vß expansions per patient was higher in BM compared to PB. The majority of patients displayed a skewed oligoclonal/monoclonal pattern within the PB and/or BM CD8(+) cells and a polyclonal profile within the CD4(+) cells. We concluded that aberrant T-cell expansions are invariably detected in CIN patients and may have a role in the disease pathogenesis.
Subject(s)
Gene Rearrangement, beta-Chain T-Cell Antigen Receptor , Neutropenia/genetics , Neutropenia/immunology , Receptors, Antigen, T-Cell, alpha-beta/genetics , Receptors, Antigen, T-Cell, alpha-beta/immunology , Receptors, Antigen, T-Cell, gamma-delta/genetics , Receptors, Antigen, T-Cell, gamma-delta/immunology , T-Lymphocytes/immunology , Adult , Aged , Aged, 80 and over , Bone Marrow/immunology , CD3 Complex/immunology , CD4-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/immunology , Chronic Disease , Female , Flow Cytometry , Humans , Male , Middle AgedABSTRACT
OBJECTIVE: Impaired granulopoiesis in chronic idiopathic neutropenia (CIN) has been associated with an inflammatory bone marrow (BM) microenvironment consisting of pro-inflammatory and pro-apoptotic mediators, such as tumor necrosis factor (TNF)-alpha, transforming growth factor (TGF)-beta1, and Fas-Ligand (Fas-L). In this study, we evaluated the frequency of TNF-alpha, TGF-beta1 and Fas-L gene polymorphisms in CIN patients and explored their role in excessive cytokine production and their association with CIN development. METHODS: The TNF-alpha-308G/A, TGF-beta1 -509C/T, +869T/C, +915G/C, and Fas-L -844T/C polymorphisms were studied in 57 CIN patients, and 100 healthy controls from Crete, a well-defined area with genetically homogeneous population, using a polymerase chain reaction-based restriction fragment length polymorphism assay. RESULTS: The mutant genotype C/T or T/T of TGF-beta1 -509C/T polymorphism was more common in CIN patients than in controls (P = 0.033). Compared to wild-type genotype, the TT genotype was associated with increased risk for CIN development (OR: 5.7; 95% CI: 1.18-27.26; P = 0.033). Compared to controls, patients with CT and TT genotypes displayed increased TGF-beta1 levels in serum (P < 0.0001 and P = 0.0002, respectively) and BM (P < 0.0001 and P = 0.0002, respectively). No significant difference was found between patients and controls in the frequency of TNF-alpha-308G/A, TGF-beta1 +869T/C and +915G/C and Fas-L -844T/C polymorphisms. CONCLUSIONS: The TGF-beta1 -509C/T polymorphism is associated with increased risk for CIN and contributes to the pathophysiology of the disorder by inducing TGF-beta1 overproduction. This is the first study providing evidence that genetic factors may predispose to CIN and may have a role in the pathophysiology of the disorder.
Subject(s)
Neutropenia/genetics , Polymorphism, Single Nucleotide , Transforming Growth Factor beta1/genetics , Adolescent , Adult , Case-Control Studies , Chronic Disease , Fas Ligand Protein/genetics , Female , Genetic Predisposition to Disease , Genotype , Greece/epidemiology , Humans , Male , Middle Aged , Neutropenia/epidemiology , Neutropenia/physiopathology , Polymerase Chain Reaction , Polymorphism, Restriction Fragment Length , Risk , Transforming Growth Factor beta1/biosynthesis , Transforming Growth Factor beta1/blood , Tumor Necrosis Factor-alpha/genetics , Up-Regulation , Young AdultABSTRACT
BACKGROUND AND AIM: T-lymphocyte migration is implicated in the pathogenesis of Crohn's disease (CD) and ulcerative colitis (UC). CXC chemokines MIG, IP-10, and I-TAC act by binding to CXCR3 receptor on T-lymphocytes. We investigated the role of these chemokines and their receptor in patients with UC, CD, and normal controls (NC). METHODS: Chemokine expression and serum levels were examined in colonic biopsies from patients and NC using reverse transcription-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay. HT-29 and Caco2 colonic epithelial cells were studied following in vitro stimulation with proinflammatory (Th1) and Th2-derived cytokines. CXCR3 receptor expression was assessed in CD3+ peripheral blood lymphocytes (PBL) from patients and NC and in stimulated Jurkat leukaemia cells, using RT-PCR and flow cytometry. RESULTS: Full size CXCR3 mRNA (FS) expression was found in CD3+ PBL from controls and UC, but not from CD patients. In contrast, CD3+ PBL from CD patients showed a marked mRNA expression of the spliced variant CXCR3 (TV). This finding explains the high expression of CXCR3 on CD3+ PBL from CD patients in flow cytometry. Increased chemokine expression and production was found in colonic biopsies and serum from CD compared to UC patients and controls. Stimulation of epithelial cells with proinflammatory cytokines significantly induced chemokine production. The addition of Th2 cytokines had an inhibitory effect. Stimulation of Jurkat cells with cytokines and supernatant conditioned media from epithelial cells induced CXCR3TV expression. CONCLUSIONS: These data demonstrate that PBL from CD patients express a spliced variant of the CXCR3 receptor and suggest a role for the colonic epithelial cells in T-lymphocyte migration in intestinal inflammation.
Subject(s)
Colitis, Ulcerative/immunology , Colon/immunology , Crohn Disease/immunology , Epithelial Cells/immunology , Intestinal Mucosa/immunology , Receptors, CXCR3/metabolism , T-Lymphocytes/immunology , Adult , CD3 Complex/analysis , Caco-2 Cells , Case-Control Studies , Chemokines/metabolism , Chemotaxis , Colitis, Ulcerative/genetics , Colitis, Ulcerative/pathology , Colon/pathology , Colonoscopy , Crohn Disease/genetics , Crohn Disease/pathology , Cytokines/metabolism , Epithelial Cells/pathology , Female , Flow Cytometry , HT29 Cells , Humans , Inflammation Mediators/metabolism , Intestinal Mucosa/pathology , Jurkat Cells , Male , Protein Isoforms , RNA, Messenger/metabolism , Receptors, CXCR3/geneticsABSTRACT
BACKGROUND: Chronic idiopathic neutropenia (CIN) is a bone marrow (BM) failure syndrome characterized by accelerated apoptosis of myeloid progenitor cells because of a local imbalance between pro-inflammatory and anti-inflammatory cytokines. In this study, we investigated the interplay among transforming growth factor-beta1 (TGF-beta1), interleukin-10 (IL-10), and soluble flt-3 ligand (sFL) within the BM of CIN patients and probed the role of these cytokines in the pathophysiology of CIN. DESIGN: We used long-term BM cultures (LTBMC) to evaluate TGF-beta1, IL-10, and sFL levels in CIN patients (n = 70) and healthy subjects (n = 35). Cytokine levels in LTBMC supernatants were correlated with the number of circulating neutrophils and the proportion of BM CD34+/CD33+ myeloid progenitor cells. RESULTS: CIN patients had increased TGF-beta1 and sFL levels in LTBMCs compared with controls and individual cytokine values were found to be correlated inversely with the number of neutrophils and the proportion of CD34+/CD33+ cells. Patients displayed low supernatant IL-10 levels compared with controls and cytokine values were found to be correlated positively with the number of neutrophils and the proportion of CD34+/CD33+ cells. The levels of TGF-beta1 were found to be inversely correlated with IL-10 and positively with sFL values in LTBMC, supernatants suggesting a possible interplay among these cytokines in CIN BM. Neutralization of TGF-beta1 in LTBMCs increased IL-10 levels significantly in patients but not in controls, while neutralization had no effect on sFL levels. CONCLUSION: Excessive production of TGF-beta1 within the BM microenvironment of CIN patients results in downregulation of IL-10 and reduction of myeloid progenitor cells. Overexpression of sFL probably represents a compensatory mechanism to the low myeloid progenitor cells.
Subject(s)
Bone Marrow Cells/metabolism , Interleukin-10/biosynthesis , Neutropenia/blood , Transforming Growth Factor beta1/metabolism , Adolescent , Adult , Aged , Antigens, CD/biosynthesis , Antigens, CD34/biosynthesis , Antigens, Differentiation, Myelomonocytic/biosynthesis , Cytokines/metabolism , Female , Humans , Interleukin-10/metabolism , Male , Membrane Proteins/metabolism , Middle Aged , Neutropenia/metabolism , Sialic Acid Binding Ig-like Lectin 3ABSTRACT
OBJECTIVE: The aim of this study is to probe the mechanisms underlying anemia in patients with chronic idiopathic neutropenia (CIN) by evaluating parameters of bone marrow (BM) erythropoiesis. PATIENTS AND METHODS: Ten CIN patients fulfilling the criteria of anemia of chronic disease, 27 nonanemic CIN patients, and 30 healthy volunteers were enrolled in the study. Reserves and survival characteristics of BM erythroid cells were evaluated using flow cytometry and clonogenic assays. Serum erythropoietin (EPO) was measured with ELISA. Expression of EPO receptors (EPORs) on BM erythroid cells was evaluated by flow cytometry and reverse-transcription polymerase chain reaction. RESULTS: CIN patients display defective erythropoiesis in addition to previously reported impaired granulopoiesis. Patients have low number of CD34(+)/CD71(+) progenitor and CD36(-)/ Glycophorin A(+) (GlycoA(+)) precursor BM cells, and increased proportion of apoptotic cells within the CD34(+)/CD71(+) and CD36(+)/GlycoA(+) compartments. Burst-forming units erythroid (BFU-Es) in BM mononuclear or purified CD34(+) cells were significantly reduced in the patients. Patient BFU-Es increased significantly following in vitro treatment with tumor necrosis factor-alpha (TNF-alpha) and/or interferon gamma (IFN-gamma)-neutralizing antibodies. Local TNF-alpha and IFN-gamma production was higher in anemic than in nonanemic patients. EPO production was appropriate in the patients, but EPOR expression was significantly reduced in patient GlycoA(+) cells, especially in anemic patients. CONCLUSION: Impaired BM erythropoiesis in CIN patients is probably the result of increased local production of TNF-alpha and IFN-gamma that induce apoptosis, cell growth inhibition, and downregulation of EPOR expression on erythroid cells. We suggest that anemia in CIN patients displays overlapping pathophysiologic features with anemia of chronic disease.
Subject(s)
Bone Marrow Cells/metabolism , Down-Regulation , Erythroid Cells/metabolism , Erythropoiesis , Neutropenia/metabolism , Receptors, Erythropoietin/biosynthesis , Adult , Aged , Antigens, CD/metabolism , Apoptosis , Bone Marrow Cells/pathology , Cell Proliferation , Chronic Disease , Down-Regulation/genetics , Erythroid Cells/pathology , Erythropoiesis/genetics , Female , Glycophorins/metabolism , Humans , Interferon-gamma/biosynthesis , Male , Middle Aged , Neutropenia/genetics , Neutropenia/pathology , Receptors, Erythropoietin/genetics , Tumor Necrosis Factor-alpha/metabolismABSTRACT
Granulopoiesis abnormalities have been described in association with thyroid disorders (TD). However, data regarding systematic evaluation of adult neutropenia and concurrent or prior TD are scarce. To investigate the frequency of TD among patients presenting with neutropenia, and the immunophenotypic and immunologic profile of neutropenic patients with concomitant thyroidopathy. Two hundred eighteen consecutive neutropenic patients were prospectively evaluated in our outpatient Hematology Clinic, with a detailed laboratory screen, including thyroid function tests, antineutrophil antibodies, blood lymphocytes immunophenotyping, and detection of T-cell clonality by PCR. Among 218 patients with neutropenia, 95 (43.6%) had TD, 65 chronic immunologic neutropenia, 20 clonal proliferation of T-large granular lymphocytes (T-LGL), 5 autoimmune disorders, and 33 other diagnoses. TD-patients had an increased frequency of recurrent infections compared with other patients (Pâ=â0.045). The following correlations were found: negative correlation between FT3 and absolute neutrophil count (ANC) (r²â=â-0.274, Pâ=â0.007), negative correlation between TPO-Abs/TG-Abs and C4 (r²â=â-0.16, Pâ=â0.045; r²â=â-0.266, Pâ=â0.001), and CD4⺠counts were inversely correlated to T4 and positively to TSH (r²â=â-0.274, Pâ=â0.024; r²â=â0.16, Pâ=â0.045). In addition, TD-patients had significantly higher percentages of CD4⺠lymphocytes (Pâ=â0.003). Among TD-patients, 23.4% had Hashimoto thyroiditis (HT), 4.1%, Graves disease (GD), 8.2% nontoxic multinodular goiter (NTMG), 5% subclinical hypothyroidism, and 2.8% had undergone total thyroidectomy associated with nodules (TTM). Thirteen TD-patients displayed T-LGL. Patients with autoimmune thyroidopathy had an increased frequency of concomitant autoimmune manifestations (Pâ=â0.03). Significant differences between the different thyroidopathies included: HT-patients had higher percentages of B-lymphocytes, while the opposite was evident for the TTM-subgroup (Pâ=â0.009, 0.02); GD-patients showed an increase of the proportion of NK cells and a decrease in the percentage of TCRγδ+ lymphocytes (Pâ=â0.001, 0.045); and NTMG-patients had significantly higher ANC (Pâ=â0.004) compared to other thyroidopathies. Antineutrophil antibodies were found in 37.2% of TD-patients tested. Anti-TPO titers were significantly higher in patients with positive antineutrophil antibodies (Pâ=â0.04). The frequency of TD among neutropenic patients may be higher than previously reported. The existence of antineutrophil antibodies, as well as the different distribution of lymphocyte subsets among patients with different TD, suggests both humoral and cellular mechanisms in the pathophysiology of thyroid disease-associated neutropenia.
Subject(s)
Neutropenia/complications , Thyroid Diseases/complications , Thyroid Diseases/epidemiology , Adolescent , Adult , Aged , Aged, 80 and over , Ambulatory Care , Female , Humans , Male , Middle Aged , Neutropenia/blood , Neutropenia/immunology , Prospective Studies , Thyroid Diseases/blood , Thyroid Diseases/immunology , Young AdultABSTRACT
Eltrombopag is a nonpeptidyl thrombopoietin receptor agonist. We evaluated the ex vivo effect of eltrombopag on megakaryopoiesis of patients with lower risk myelodysplastic syndromes (MDSs). At a concentration of 0.1µg/mL, eltrombopag resulted in a significant increase in the number of megakaryocytic colonies in MDS patients and healthy controls compared to baseline. This dose of eltrombopag did not exert any significant change in the proliferation rate or the survival characteristics of patient CD34(+) cells that might clinically imply an unfavorable effect on patients' outcome. These encouraging preclinical data support the rationale of using eltrombopag in the clinic for alleviation of thrombocytopenia in lower risk MDS patients.
Subject(s)
Benzoates/therapeutic use , Cell Proliferation/drug effects , Hydrazines/therapeutic use , Megakaryocyte Progenitor Cells/drug effects , Myelodysplastic Syndromes/drug therapy , Pyrazoles/therapeutic use , Receptors, Thrombopoietin/agonists , Thrombopoiesis/drug effects , Aged , Aged, 80 and over , Antigens, CD34/metabolism , Apoptosis/drug effects , Blotting, Western , Bone Marrow Cells/drug effects , Bone Marrow Cells/pathology , Female , Humans , Male , Megakaryocyte Progenitor Cells/pathology , Middle Aged , Myelodysplastic Syndromes/pathology , Risk Factors , Tumor Cells, CulturedABSTRACT
Alterations of the ten-eleven translocation-2 (TET2) gene have been recently identified in patients with myeloid malignancies using molecular, comparative genomic hybridization and single nucleotide polymorphism array techniques. We have performed TET2 fluorescence in situ hybridization analysis in a cohort of patients with myeloid disorders including myeloid malignancies and chronic idiopathic neutropenia, aiming to determine the usefulness of the technique in the identification of TET2 gene alterations. A TET2 deletion was found in one patient with chronic myelomonocytic leukemia suggesting that fluorescence in situ hybridization may have a role in identification of TET2 deletions, at least in this group of patients.
Subject(s)
DNA-Binding Proteins/genetics , Gene Deletion , In Situ Hybridization, Fluorescence , Leukemia, Myelomonocytic, Chronic/genetics , Myelodysplastic Syndromes/genetics , Myeloproliferative Disorders/genetics , Neutropenia/genetics , Proto-Oncogene Proteins/genetics , Aged , Aged, 80 and over , Chronic Disease , Dioxygenases , Female , Humans , Leukemia, Myelomonocytic, Chronic/pathology , Male , Middle Aged , Myelodysplastic Syndromes/pathology , Myeloproliferative Disorders/pathology , Neutropenia/pathologyABSTRACT
Chronic idiopathic neutropenia (CIN) is a granulopoiesis disorder associated with an inhibitory bone marrow (BM) microenvironment consisting of activated T-lymphocytes and pro-inflammatory mediators. In this study, we investigated the possible involvement of BM mesenchymal stem cells (MSCs) in the pathophysiology of CIN by assessing the frequency and function of BM MSCs in terms of the proliferative/clonogenic characteristics, the differentiation capacity, the potential to produce pro-inflammatory cytokines, and the ability to suppress T-cell proliferation. The frequency, differentiation capacity toward adipocytes, chondrocytes, or osteoblasts, and immunosuppressive potential to inhibit mitogen-induced T-cell proliferation did not differ significantly between patient (n = 14) and normal (n = 21) MSCs. Tumor necrosis factor-α, interleukin-1ß, and interleukin-6 levels in MSC supernatants did not differ significantly between patients and controls; however, transforming growth factor (TGF)-ß1 levels were significantly elevated in patients, particularly in those displaying the -509C/T TGF-ß1 polymorphism. Patient MSCs displayed defective proliferative/clonogenic potential, which could not be attributed to altered cellular survival characteristics or to increased TGF-ß1 production as TGF-ß1 neutralization did not restore the impaired colony formation by patient MSCs. We conclude that although BM MSCs do not exert a significant role in the immune deregulation associated with CIN, they contribute to the inhibitory microenvironment by overproducing TGF-ß1, at least in patients displaying the -509C/T polymorphism.
Subject(s)
Bone Marrow/immunology , Mesenchymal Stem Cells/immunology , Neutropenia/immunology , Neutropenia/metabolism , Transforming Growth Factor beta1/immunology , Adolescent , Adult , Aged , Bone Marrow/metabolism , Cell Differentiation , Cell Proliferation , Cells, Cultured , Cytokines/biosynthesis , Female , Humans , Lymphocyte Activation/immunology , Male , Mesenchymal Stem Cells/cytology , Mesenchymal Stem Cells/metabolism , Middle Aged , Polymorphism, Restriction Fragment Length , Polymorphism, Single Nucleotide , T-Lymphocytes/metabolism , T-Lymphocytes/pathology , Transforming Growth Factor beta1/biosynthesisABSTRACT
CD40 is a member of the TNFR family and upon interaction with its cognate ligand (CD40L), induces diverse biologic responses related to cell survival/growth. As altered CD40/CD40L interactions have been associated with neutropenia, we investigated the role of CD40/CD40L on human granulopoiesis using immunomagnetically sorted CD34(+), CD34(-)/CD33(+), and CD34(-)/CD33(-)/CD15(+) BM cells, which represent sequential stages of the granulocytic development, the KG-1 cells that constantly express CD34 and CD33, and LTBMCs that mimic the BM microenvironment. CD40 and CD40L were minimally expressed on CD34(+), CD34(-)/CD33(+), and CD34(-)/CD33(-)/CD15(+) cells, but CD40 was substantially induced in the presence of TNF-α. Cross-linking of CD40 in the above cell populations resulted in induction of apoptosis that was enhanced further in the presence of FasL. CD40 activation in primary as wells as in KG-1 cells resulted in Fas up-regulation, providing a mechanism for the CD40-mediated apoptosis. Addition of CD40L in clonogenic assays resulted in a significant decrease in the colony-forming capacity of BMMCs from patients with chronic neutropenia, presumably expressing high levels of CD40 in the progenitor cells, and this effect was reversed upon CD40 blockade. CD40 was constitutively expressed on LTBMC stromal cells and upon activation, resulted in an increase in G-CSF and GM-CSF production. These data show that CD40/CD40L interactions may promote granulopoiesis under steady-state conditions by inducing the stromal release of granulopoiesis-supporting cytokines, whereas under inflammatory conditions, they may affect the granulocytic progenitor/precursor cell survival by accelerating the Fas-mediated apoptosis.
Subject(s)
Bone Marrow/metabolism , CD40 Antigens/metabolism , CD40 Ligand/metabolism , Granulocytes/metabolism , Hematopoietic Stem Cells/metabolism , Leukemia, Myeloid, Acute/metabolism , Neutropenia/metabolism , Apoptosis , Blotting, Western , Cells, Cultured , Colony-Forming Units Assay , Cytokines/metabolism , Flow Cytometry , Granulocyte Colony-Stimulating Factor/metabolism , Granulocyte-Macrophage Colony-Stimulating Factor/metabolism , Humans , Leukemia, Myeloid, Acute/pathology , Neutropenia/pathology , Stromal Cells/metabolismABSTRACT
OBJECTIVE: Patients with systemic lupus erythematosus (SLE) display increased apoptosis of bone marrow (BM) CD34+ hematopoietic progenitor cells. This study was undertaken to evaluate the expression of CD40 and CD40L in the BM of SLE patients, and to explore the possible involvement of these molecules in apoptosis of CD34+ cells. METHODS: The proportion and survival characteristics of CD40+ cells within the BM CD34+ fraction from SLE patients and healthy controls were evaluated by flow cytometry. The production of CD40L by BM stromal cells was assessed using long-term BM cultures, and the effect of CD40L on the survival characteristics and clonogenic potential of CD34+ cells was evaluated ex vivo by flow cytometry and clonogenic assays. RESULTS: SLE patients displayed an increased proportion of CD40+ cells within the CD34+ fraction as compared with controls. The CD34+CD40+ subpopulation contained an increased proportion of apoptotic cells compared with the CD34+CD40- fraction in patients and controls, suggesting that CD40 is involved in the apoptosis of CD34+ cells. Stimulation of patients' CD34+ cells with CD40L increased the proportion of apoptotic cells and decreased the proportion of colony-forming cells as compared with untreated cultures. The CD40L-mediated effects were amplified following treatment with recombinant Fas ligand, suggesting that the effects of these ligands are synergistic. CD40L levels were significantly increased in long-term BM culture supernatants and adherent layers of BM cells from SLE patients as compared with controls. CONCLUSION: These data reveal a novel role for the CD40/CD40L dyad in SLE by demonstrating that up-regulation and induction of CD40 on BM CD34+ cells from patients with SLE contribute to the amplification of Fas-mediated apoptosis of progenitor cells.
Subject(s)
Antigens, CD34/metabolism , Apoptosis , Bone Marrow Cells/metabolism , CD40 Antigens/metabolism , Hematopoietic Stem Cells/metabolism , Lupus Erythematosus, Systemic/metabolism , Adult , Aged , Antigens, CD34/immunology , Apoptosis/drug effects , Biomarkers/metabolism , Bone Marrow Cells/immunology , Bone Marrow Cells/pathology , CD40 Antigens/immunology , Cell Survival , Cells, Cultured , Clone Cells , Fas Ligand Protein/pharmacology , Female , Health Status , Hematopoietic Stem Cells/immunology , Hematopoietic Stem Cells/pathology , Humans , Lupus Erythematosus, Systemic/immunology , Lupus Erythematosus, Systemic/physiopathology , Male , Middle Aged , Severity of Illness Index , Stromal Cells/immunology , Stromal Cells/metabolism , Stromal Cells/pathology , Young Adult , fas Receptor/metabolismABSTRACT
Patients with chronic idiopathic neutropenia (CIN) display relatively low peripheral blood platelet counts and hypo-lobulated megakaryocytes in the bone marrow (BM). The underlying pathogenetic mechanismswere probed by studying the reserves and clonogenic potential of BM megakaryocytic progenitor cells using flow-cytometry and a collagen-based clonogenic assay for the identification of megakaryocyte colony-forming units (CFU-Meg). Thrombopoietin (TPO) and transforming growth factor-beta1 (TGFbeta1) levels were also evaluated in long-term BM culture supernatants using an enzyme-linked immunosorbent assay. CIN patients (n = 39) showed a low proportion of BM CD34(+)/CD61(+) megakaryocytic progenitor cells and low frequency of early and mixed CFU-Meg in the BM mononuclear, but not CD34(+), cell fraction, compared with healthy controls (n = 20). TPO and TGFbeta1 levels were significantly higher in patients compared with controls. TPO levels inversely correlated with platelet counts whereas TGFbeta1 values correlated inversely with CD34(+)/CD61(+) and CFU-Meg megakaryocytic progenitor cell numbers and positively with TPO levels. The addition of an anti-TGFbeta1 neutralising antibody significantly increased the numbers of CFU-Meg in CIN patients but not in controls, compared with baseline. These data suggest that increased local production of TGFbeta1 probably affects the BM megakaryocytic progenitor cell growth in CIN whereas the compensatory production of TPO finally balances the TGFbeta1-induced inhibitory effect.
Subject(s)
Bone Marrow/immunology , Megakaryocytes/immunology , Neutropenia/immunology , Thrombopoiesis/immunology , Transforming Growth Factor beta/immunology , Adult , Aged , Antibodies, Monoclonal/immunology , Cytokines/immunology , Female , Flow Cytometry , Humans , Male , Middle Aged , Transforming Growth Factor beta1ABSTRACT
Splenic volume and Helicobacter pylori (H. pylori) infection were evaluated in 67 patients with chronic idiopathic neutropenia (CIN) and 39 healthy individuals. Using ultrasound, splenomegaly was found in 61.7% of H. pylori-infected subjects compared to only 8.7% noted in the group of H. pylori-non-infected individuals (P < 0.0001). Splenomegaly was also found in 47.8% of CIN patients compared to 12.8% in the group of non-CIN subjects (P = 0.0003). However, applying the two-way ANOVA test, a statistically significant effect on splenic volume was documented for "factor H. pylori " (F1(102) = 16.800, P < 0.0001) but not for "factor CIN" (F1(102) = 3.213, P = 0.0760), suggesting that CIN-associated splenomegaly is probably due to H. pylori infection.
Subject(s)
Helicobacter Infections/complications , Neutropenia/complications , Splenomegaly/etiology , Adult , Aged , Analysis of Variance , Chronic Disease , Female , Helicobacter pylori , Humans , Male , Middle Aged , Splenomegaly/diagnostic imaging , UltrasonographyABSTRACT
The levels of serum and long-term bone marrow culture supernatant soluble flt-3 ligand (sFL) were determined in 54 patients with chronic idiopathic neutropenia (CIN) and 16 normal controls. Both serum and supernatant sFL levels were significantly increased in the patients compared with controls. Individual sFL values inversely correlated with the number of circulating neutrophils and the proportion of bone marrow CD34+ cells. Supernatant sFL values positively correlated with the levels of supernatant G-CSF. These findings suggest that the impaired myelopoiesis in CIN patients is accompanied by a compensatory mechanism attempting to increase the neutrophil production at the myeloid progenitor cell level.
Subject(s)
Bone Marrow Cells/metabolism , Membrane Proteins/blood , Neutropenia/metabolism , Adult , Aged , Bone Marrow Cells/pathology , Case-Control Studies , Cells, Cultured , Chronic Disease , Female , Granulocyte Colony-Stimulating Factor/analysis , Humans , Leukocyte Count , Male , Membrane Proteins/analysis , Membrane Proteins/metabolism , Middle Aged , Neutropenia/pathology , Neutrophils/pathology , Statistics, Nonparametric , Time FactorsABSTRACT
The prevalence of Helicobacter pylori infection was evaluated in 120 patients with chronic idiopathic neutropenia (CIN), 8 patients with monoclonal gammopathy of undetermined significance (MGUS) associated with CIN, and 74 age- and sex-matched normal volunteers, all derived from the same geographical area. The purpose of the study was to investigate the possible causal relationships of H. pylori infection with the development of MGUS in CIN patients. We found that the prevalence of H. pylori infection was elevated to 69.2% in the group of CIN patients, 100% in the group of patients with CIN-associated MGUS, and 32.4% in the group of control subjects. No statistically significant difference, however, was found in the prevalence of H. pylori infection between CIN patients with concomitant MGUS and CIN patients without MGUS, no resolution of the gammopathy after eradication of the bacterium, no significant rise in the titers of serum anti-H. pylori antibodies, and no formation of an abnormal precipitation line in immunoelectrophoresis using a saline extract of NCTC11367 H. pylori reference strain as antigen. We concluded that there is no evidence that H. pylori infection is the cause of MGUS in CIN patients.