Immune-mediated Bone Marrow Failure Syndromes Research Articles

Severe aplastic anemia after COVID-19 mRNA vaccination: Causality or coincidence?

The development of various autoimmune diseases has been reported after COVID-19 infections or vaccinations. However, no method for assessing the relationships between vaccines and the development of autoimmune diseases has been established. Aplastic anemia (AA) is an immune-mediated bone marrow failure syndrome. We report a case of severe AA that arose after the administration of a COVID-19 vaccine (the Pfizer-BioNTech mRNA vaccine), which was treated with allogeneic hematopoietic stem cell transplantation (HSCT). In this patient, antibodies against the SARS-CoV-2 spike protein were detected both before and after the HSCT. After the patient's hematopoietic stem cells were replaced through HSCT, his AA improved despite the presence of anti-SARS-CoV-2 antibodies. In this case, antibodies derived from the COVID-19 vaccine may not have been directly involved in the development of AA. This case suggests that the measurement of vaccine antibody titers before and after allogeneic HSCT may provide clues to the pathogenesis of vaccine-related autoimmune diseases. Although causality was not proven in this case, further evaluations are warranted to assess the associations between vaccines and AA.

Inhibition of 15-PGDH Protects Mice from Immune-Mediated Bone Marrow Failure.

Aplastic anemia (AA) is a human immune-mediated bone marrow failure syndrome that is treated by stem cell transplantation for patients who have a matched related donor and by immunosuppressive therapy (IST) for those who do not. Responses to IST are variable, with patients still at risk for prolonged neutropenia, transfusion dependence, immune suppression, and severe opportunistic infections. Therefore, additional therapies are needed to accelerate hematologic recovery in patients receiving front-line IST. We have shown that inhibiting 15-hydroxyprostaglandin dehydrogenase (15-PGDH) with the small molecule SW033291 (PGDHi) increases bone marrow (BM) prostaglandin E2 levels, expands hematopoietic stem cell (HSC) numbers, and accelerates hematologic reconstitution following murine BM transplantation. We now report that in a murine model of immune-mediated BM failure, PGDHi therapy mitigated cytopenias, increased BM HSC and progenitor cell numbers, and significantly extended survival compared with vehicle-treated mice. PGDHi protection was not immune-mediated, as serum IFN-γ levels and BM CD8+ T lymphocyte frequencies were not impacted. Moreover, dual administration of PGDHi plus low-dose IST enhanced total white blood cell, neutrophil, and platelet recovery, achieving responses similar to those seen with maximal-dose IST with lower toxicity. Taken together, these data demonstrate that PGDHi can complement IST to accelerate hematologic recovery and reduce morbidity in severe AA.

Research progress of abnormal immunologic mechanism in acquired aplastic anemia

Aplastic anemia (AA) is considered as an immune-mediated bone marrow failure syndrome, characterized by peripheral pancytopenia and destruction of hematopoietic stem/progenitor cells. Acquired aplastic anemia (aAA) is more common in clinic. Abnormal immunity is one of the major factors mediating the pathogenesis of aAA. This review discusses the current understanding of the immunobiology in aAA, so as to clarify the immune mechanism of this disease, and to provide a theoretical basis for the clinical treatment of aAA. Key words: Anemia, aplastic; Immunity; Immunosuppressive agent; Children; Acquired aplastic anemia

Eltrombopag maintains human hematopoietic stem and progenitor cells under inflammatory conditions mediated by IFN-γ

AbstractThe proinflammatory cytokine interferon-γ (IFN-γ) has been implicated in human hematopoietic stem and progenitor cell (HSPC) depletion in immune-mediated bone marrow failure syndromes. We show that IFN-γ specifically prevents full engagement of thrombopoietin (TPO), a primary positive regulator of HSPC survival, to its receptor (c-MPL) via steric occlusion of the low-affinity binding site, contributing to perturbation of TPO-induced signaling pathways and decreased survival of human HSPCs. Eltrombopag, a synthetic small molecule mimetic of TPO that interacts with c-MPL at a position distinct from the extracellular binding site of TPO, bypasses this inhibition, providing an explanation for its clinical activity in bone marrow failure, despite already elevated endogenous TPO levels. Thus, IFN-γ–mediated perturbation of TPO:c-MPL complex formation and the resulting inhibition of a critical pathway of growth factor cell signaling may represent a general mechanism by which IFN-γ impairs the function of human HSPCs. This understanding could have broad therapeutic implications for various disorders of chronic inflammation.

Cyclosporin A inhibits adipogenic differentiation and regulates immunomodulatory functions of murine mesenchymal stem cells

Aplastic anemia (AA) is generally considered as an immune-mediated bone marrow failure syndrome. Several studies show that bone marrow mesenchymal stem cells (BM-MSCs), as key cellular components of the bone marrow microenvironment, are also involved in the pathogenic mechanism of AA. Cyclosporin A (CsA) is a classic immunosuppressive drug for AA, and it specifically inhibits mammalian T cells by preventing activation of transcription factors involved in cytokine gene expression. However, little is known about the effect of CsA on the BM-MSCs. In this study, murine BM-MSCs were stimulated in the presence of CsA. Further, we found that CsA could inhibit murine BM-MSC proliferation and promote BM-MSC apoptosis, what's more CsA could inhibit adipogenic differentiation. Our study also showed that CsA could inhibit interleukin-6 expression in BM-MSCs, while promoting programmed death-ligand 2 expression. In conclusion, our results proposed that CsA may exert an effect on regulating the bone marrow environment by influencing BM-MSCs, which have a beneficial effect on treating AA.

Persistent elevated bone marrow plasma levels of thrombopoietin in patients with aplastic anemia

Acquired aplastic anemia (AA) is an immune-mediated bone marrow (BM) failure syndrome. Plasma thrombopoietin (TPO) levels are elevated in AA patients with reduced blood counts. However, whether elevated TPO can recover to normal level in AA patients in complete remission (CR) after efficient immunosuppressive therapy (IST) is unknown. We measured TPO levels of BM plasma in a large cohort of AA patients with focusing on patients in CR after IST. Our data showed that BM plasma TPO levels were still high in patients who had reached normal blood counts levels for a long time. We speculate that downregulated expression of MPL (TPO receptor) in HSPC (hematopoietic stem and progenitor cells) may be related to the upregulated TPO in AA.

Increased Frequency of Paroxysmal Nocturmal Hemoglobinuria Type Cells in Patients with Chronic Idiopathic Neutropenia Correlates with the Severity of the Disease and the Presence of Skewed T-Cell Expansions

Introduction: Chronic idiopathic neutropenia (CIN) of adults is an acquired disorder of granulopoiesis characterized by an unexplained, prolonged reduction in the number of peripheral blood (PB) neutrophils. We have previously shown that CIN pathophysiology is related to T-cell activation and increased apoptosis of granulocytic progenitor cells. Notably, in our cohort of patients we exclude cases with antineutrophil antibody activity, i.e. autoimmune neutropenia cases.Aim of the study: Preliminary data have shown the presence of minor populations of paroxysmal nocturmal hemoglobinuria (PNH) type cells in the PB of CIN patients. The present study aims to investigate further the presence of PNH type cells in the PB of a large cohort of patients fulfilling the diagnostic criteria of CIN and probe the possible association with the severity of the disease and the skewed T-cell profile.Patients - Methods: We have studied 91 adults with CIN and 55 hematologically healthy subjects, age- and sex-matched with the patients. We have used flow cytometry for the detection of PNH type cells according to the International Clinical Cytometry Society guidelines and for the identification of T-cell expansions based on the analysis of T-cell receptor beta variable (TRBV) gene repertoire.Results: CIN patients displayed increased proportion of PB PNH type FLAER-/CD24- neutrophils (0.05% ± 0.18%) and FLAER-/CD14- monocytes (1.33% ± 1.38%) compared to healthy individuals (0.005% ± 0.01% and 0.64% ± 0.66%, respectively; P =0.0044 and P =0.011, respectively). Similarly, the patients displayed increased proportion of both, type II CD235+/CD59dim and type III CD235+/CD59- (0.059% ± 0.29% and 0.065% ± 0.28%, respectively) PB PNH red blood cells, compared to healthy controls (0.01% ± 0.03% and 0.01% ± 0.03%, respectively; P<0.0001 and P< 0.0001, respectively). Interestingly, an inverse correlation was found between the proportion of PB PNH type neutrophils and the number of absolutely neutrophil counts in CIN patients (r=-0.2525, P =0.0157) whereas no correlation was found between the proportion of PNH red blood cells or monocytes and absolutely neutrophil counts. In accordance with our previously reported data, increased frequency of skewed T-cell expansions were identified in CIN patients (71.4%) compared to healthy subjects (29.9%; P<0.0001). PNH type neutrophils (FLAER-/CD24-) were identified at a higher frequency in patients with skewed TRBV repertoire (63.08%) compared to those with normal TRBV distribution (34.61%) (P = 0.0194) suggesting a possible pathogenetic/pathophysiologic association between T-cell activation and emergence of PNH neutrophils in CIN.Conclusion: CIN patients display minor populations of PNH type PB neutrophils, monocytes and red blood cells. The frequency of PNH type neutrophils is associated with the severity of neutropenia and it is higher among patients with skewed TRBV repertoire. These data support further the hypothesis that CIN displays common pathophysiologic features with immune-mediated bone marrow failure syndromes and should be included in this spectrum of disease entities. DisclosuresPapadaki:Alexion Pharmaceuticals: Research Funding.

Relationship between antigen presenting cells and pathogenesis of aplastic anemia

Aplastic anemia(AA) is defined as an immune-mediated bone marrow failure syndrome, characterized by pancytopenia and hypocellular bone marrow. At present, the pathogenesis of AA is concerned chiefly with abnormal hematopoietic stem cells, hematopoietic microenvironment abnormality and disorder of immune system. Antigen presenting cell (APC) is the main initiator of adaptive immunity. It plays a role in processing and presenting antigens to T lymphocyte, and leading to an immune response or immune tolerance. Activation of APC may lead to the imbalance and dysfunction of T lymphocyte subsets, such as the polarization of helper T lymphocyte (Th)1, activation of CD8+ T cells and secretion of negative regulators of hematopoiesis. All of these compose a cytokine network to destruct stem/progenitor cells as well as hematopoietic stem cells, mesenchymal stem cells and angioblasts/endothelial progenitor cells. Thus, this article reviews literatures on the relationship between APC including dendritic cells (DC), monocytes, B lymphocytes and the pathogenesis of AA. Key words: Anemia, aplastic; Antigen-presenting cells; Receptor, pattern recognition

Decreased expression of vitamin D receptor may contribute to the hyperimmune status of patients with acquired aplastic anemia.

Acquired aplastic anemia (AA) is an immune-mediated bone marrow failure syndrome. 1α,25-Dihydroxyvitamin D3 [1,25(OH)2 D3 ], the biologically active metabolite of vitamin D, is a critical modulator of immune response via binding with vitamin D receptor (VDR). Previous studies have established that 1,25(OH)2 D3 and VDR were involved in the pathogenesis of some autoimmune diseases. In this study, we evaluated the involvement of 1,25(OH)2 D3 and VDR on T-cell responses in AA. Plasma 25(OH)D3 levels were comparable between patients with AA and healthy controls. Surprisingly, VDR mRNA was significantly lower in untreated patients with AA than in healthy controls. Subsequent invitro experiments revealed that 1,25(OH)2 D3 treatment suppressed the proliferation of lymphocytes and inhibited the secretion of interferon-γ, tumor necrosis factor-α, and interleukin-17A, meanwhile promoting the production of transforming growth factor-β1 in patients with AA. Moreover, 1,25(OH)2 D3 inhibited the differentiation of type 1 and Th17 cells but induced the differentiation of type 2 and regulatory T cells. Interestingly, VDR mRNA was elevated in healthy controls after 1,25(OH)2 D3 treatment, but not in patients with AA. In conclusion, decreased expression of VDR might contribute to the hyperimmune status of AA and appropriate vitamin D supplementation could partly correct the immune dysfunction by strengthening signal transduction through VDR in patients with AA.

Bone Marrow Immunophenotyping By Flow Cytometry in Refractory Cytopenia of Childhood

Refractory cytopenia of childhood (RCC) is the most common type of childhood myelodysplastic syndrome (MDS). Because the majority of patients with RCC have a normal karyotype and a hypocellular bone marrow, differentiating RCC from the immune-mediated bone marrow failure syndrome (very) severe aplastic anemia ((v)SAA) can be challenging. The histopathological differentiation between RCC and (v)SAA is mainly based on the presence of patchy erythropoiesis with defective maturation and/or the presence of micromegakaryocytes in RCC, and the absence of erythropoiesis and megakaryopoiesis in (v)SAA. Flow cytometric immunophenotyping of bone marrow has been shown to be a valuable additional diagnostic tool in differentiating MDS from non-clonal cytopenias in adults, but in childhood MDS, only a limited number of flow cytometric immunophenotyping studies have been reported.Here, we performed the first comprehensive flow cytometric analysis of myeloid and lymphoid progenitor cells, maturing granulocytes, monocytes and erythrocytes in bone marrow aspirates obtained from a large prospective cohort of 81 RCC patients, collected by the European Working Group of MDS in Childhood (EWOG-MDS). RCC was diagnosed according to WHO criteria and confirmed by central review of bone marrow morphology and histology. Bone marrow aspirates obtained from healthy adult individuals (n=9) and pediatric patients with (v)SAA (n=17) or advanced MDS (n=7) were used as controls. We employed a 7-tube, 6-color flow cytometry protocol, and data analysis was performed largely in line with European LeukemiaNet recommendations for flow cytometry in MDS.RCC patients had a strongly reduced myeloid compartment compared to healthy controls, but not as severe as (v)SAA patients. In the majority of RCC patients, immature myeloid and/or lymphoid cells were reduced in numbers, but still detectable, while in the vast majority of (v)SAA patients, myeloid blast cells and CD34+B-cell precursors were absent: both cell types were absent in 27 of 81 RCC patients (33%) and in 15 of 17 (v)SAA patients (88%). Furthermore, the number of flow cytometric abnormalities was significantly higher in RCC patients than in healthy controls and in pediatric patients with (v)SAA, but lower than in advanced MDS. The most frequently occurring flow cytometric abnormalities in RCC were the heterogeneous expression of CD71 and CD36 on immature erythroid cells. Two or more flow cytometric abnormalities were detected in 49 of 81 RCC patients (60%), and in 2 of 17 (v)SAA patients (12%). If a diagnosis of RCC was considered if myeloid blast cells and/or CD34+ B-cell precursors were present, or if two or more immunophenotypic abnormalities were detected, 61 of 81 RCC patients (84%) could be correctly classified, using histopathology as gold standard, whereas the specificity of this combination, using (v)SAA as a control group, was 76% (13 of 17 (v)SAA patients). No significant associations were detected between the presence of flow cytometric abnormalities (defined as two or more abnormalities) in RCC patients and age or sex, the presence of HLA-DR15, bone marrow cellularity, transfusion dependency at diagnosis, the presence of a PNH clone, or skewing of the T-cell receptor Vβ chain. Of interest, although only 5 patients with monosomy 7 were included in the present study, all of them displayed at least 2 flow cytometric abnormalities. RCC with monosomy 7 confers a high risk of progression to AML, but histopathologically, no differences can be detected between RCC cases with or without monosomy 7.In conclusion, our results indicate that, although flow cytometric abnormalities in RCC patients are present at a relatively low frequency, flow cytometric immunophenotyping might be a relevant addition to histopathology and cytogenetic analysis in the diagnostic work-up of RCC. DisclosuresNo relevant conflicts of interest to declare.

Increased bone marrow (BM) plasma level of soluble CD30 and correlations with BM plasma level of interferon (IFN)-γ, CD4/CD8 T-cell ratio and disease severity in aplastic anemia.

Idiopathic aplastic anemia (AA) is an immune-mediated bone marrow failure syndrome. Immune abnormalities such as decreased lymphocyte counts, inverted CD4/CD8 T-cell ratio and increased IFN-γ-producing T cells have been found in AA. CD30, a surface protein belonging to the tumor necrosis factor receptor family and releasing from cell surface as a soluble form (sCD30) after activation, marks a subset of activated T cells secreting IFN-γ when exposed to allogeneic antigens. Our study found elevated BM plasma levels of sCD30 in patients with SAA, which were closely correlated with disease severity, including absolute lymphocyte count (ALC) and absolute netrophil count (ANC). We also noted that sCD30 levels were positively correlated with plasma IFN-γ levels and CD4/CD8 T-cell ratio in patients with SAA. In order to explain these phenomena, we stimulated T cells with alloantigen in vitro and found that CD30+ T cells were the major source of IFN-γ, and induced CD30+ T cells from patients with SAA produced significantly more IFN-γ than that from healthy individuals. In addition, increased proportion of CD8+ T cells in AA showed enhanced allogeneic response by the fact that they expressed more CD30 during allogeneic stimulation. sCD30 levels decreased in patients responded to immunosuppressive therapy. In conclusion, elevated BM plasma levels of sCD30 reflected the enhanced CD30+ T cell-mediated immune response in SAA. CD30 as a molecular marker that transiently expresses on IFN-γ-producing T cells, may participate in mediating bone marrow failure in AA, which also can facilitate our understanding of AA pathogenesis to identify new therapeutic targets.

Role of CD8(+)T cells and their secreted cytokines in the pathogenesis of aplastic anemia

Aplastic anemia(AA) is mostly considered as an immune-mediated bone marrow failure syndrome, characterized by pancytopenia and bone marrow hypoplasia. The pathogenesis of AA is complicated, until now it is not fully understood. Further study on the pathological mechanism will be helpful for the diagnosis and treatment of AA. CD8(+) T cells and their secreted cytokines play important roles in the abnormal immunity during the process of AA. Thus, this review focuses on the role of CD8(+) T cells and their secreted cytokines in the pathogenesis of AA.

Mesenchymal Stem Cells in Immune-Mediated Bone Marrow Failure Syndromes

Immune-mediated bone marrow failure syndromes (BMFS) are characterized by ineffective marrow haemopoiesis and subsequent peripheral cytopenias. Ineffective haemopoiesis is the result of a complex marrow deregulation including genetic, epigenetic, and immune-mediated alterations in haemopoietic stem/progenitor cells, as well as abnormal haemopoietic-to-stromal cell interactions, with abnormal release of haemopoietic growth factors, chemokines, and inhibitors. Mesenchymal stem/stromal cells (MSCs) and their progeny (i.e., osteoblasts, adipocytes, and reticular cells) are considered as key cellular components of the bone marrow haemopoietic niche. MSCs may interfere with haemopoietic as well as immune regulation. Evidence suggests that bone marrow MSCs may be involved in immune-mediated BMFS underlying pathophysiology, harboring either native abnormalities and/or secondary defects, caused by exposure to activated marrow components. This review summarizes previous as well as more recent information related to the biologic/functional characteristics of bone marrow MSCs in myelodysplastic syndromes, acquired aplastic anemia, and chronic idiopathic neutropenia.

Differential Gene Expression Profile Associated with the Abnormality of Bone Marrow Mesenchymal Stem Cells in Aplastic Anemia

Aplastic anemia (AA) is generally considered as an immune-mediated bone marrow failure syndrome with defective hematopoietic stem cells (HSCs) and marrow microenvironment. Previous studies have demonstrated the defective HSCs and aberrant T cellular-immunity in AA using a microarray approach. However, little is known about the overall specialty of bone marrow mesenchymal stem cells (BM-MSCs). In the present study, we comprehensively compared the biological features and gene expression profile of BM-MSCs between AA patients and healthy volunteers. In comparison with healthy controls, BM-MSCs from AA patients showed aberrant morphology, decreased proliferation and clonogenic potential and increased apoptosis. BM-MSCs from AA patients were susceptible to be induced to differentiate into adipocytes but more difficult to differentiate into osteoblasts. Consistent with abnormal biological features, a large number of genes implicated in cell cycle, cell division, proliferation, chemotaxis and hematopoietic cell lineage showed markedly decreased expression in BM-MSCs from AA patients. Conversely, more related genes with apoptosis, adipogenesis and immune response showed increased expression in BM-MSCs from AA patients. The gene expression profile of BM-MSCs further confirmed the abnormal biological properties and provided significant evidence for the possible mechanism of the destruction of the bone marrow microenvironment in AA.

Genetic Associations in Acquired Immune-Mediated Bone Marrow Failure Syndromes: Insights in Aplastic Anemia and Chronic Idiopathic Neutropenia

Increasing interest on the field of autoimmune diseases has unveiled a plethora of genetic factors that predispose to these diseases. However, in immune-mediated bone marrow failure syndromes, such as acquired aplastic anemia and chronic idiopathic neutropenia, in which the pathophysiology results from a myelosuppressive bone marrow microenvironment mainly due to the presence of activated T lymphocytes, leading to the accelerated apoptotic death of the hematopoietic stem and progenitor cells, such genetic associations have been very limited. Various alleles and haplotypes of human leucocyte antigen (HLA) molecules have been implicated in the predisposition of developing the above diseases, as well as polymorphisms of inhibitory cytokines such as interferon-γ, tumor necrosis factor-α, and transforming growth factor-β1 along with polymorphisms on molecules of the immune system including the T-bet transcription factor and signal transducers and activators of transcription. In some cases, specific polymorphisms have been implicated in the outcome of treatment on those patients.

Autoimmune Disease Genetics

Autoimmune Disease Genetics

Immunosuppressive therapies in the management of acquired immune-mediated marrow failures

Immunosuppression is a key treatment strategy for patients suffering from aplastic anemia or related immune-mediated bone marrow failure syndromes. Several attempts have been performed to improve the standard immunosuppression regimen of horse antithymocyte globulin (h-ATG) and cyclosporine A (CyA). The addition of a third immunosuppression agent to h-ATG + CyA did not result in any improvement. Antilymphocyte agents other than h-ATG have been investigated. A rabbit-ATG preparation, which was known to be more immunosuppressive than h-ATG, resulted in markedly inferior outcome in a large randomized study from the National Institutes of Health. These data seem to be confirmed in additional experiences with rabbit-ATG from other groups. Cyclophosphamide and alemtuzumab have been proven to be biologically active in small studies, but available data suggest inferior outcomes when compared with h-ATG. All these alternative agents result in a more pronounced lymphocyte depletion, suggesting that the actual mechanisms of action of immunosuppressive therapy in aplastic anemia are not fully understood. Immunosuppression by h-ATG and CyA remains the standard of care for aplastic anemia patients lacking a low-risk transplant procedure, resulting in a 60-70% response rate. Rabbit-ATG, cyclophosphamide and alemtuzumab demonstrated a biological activity, but resulted in inferior outcome as compared with h-ATG; thus, they are not recommended as front-line therapy of aplastic anemia.

Abnormal telomere shortening of peripheral blood mononuclear cells and granulocytes in patients with chronic idiopathic neutropenia

Chronic idiopathic neutropenia is characterized by immune-mediated suppression of neutrophil production. Because patients with immune-mediated bone marrow failure syndromes display age-inappropriate telomere shortening in leukocytes, we investigated telomere lengths in peripheral blood mononuclear cells and granulocytes of patients with chronic idiopathic neutropenia. We studied 37 patients with chronic idiopathic neutropenia and 68 age- and sex-matched healthy controls. Relative telomere length and telomerase reverse transcriptase expression were assessed by a quantitative real time polymerase chain reaction. Telomerase activity was determined by a polymerase chain reaction-based immunoassay. The mean relative telomere values of peripheral blood mononuclear cells and granulocytes were significantly lower in patients compared to controls, and significantly lower than expected on the basis of the age-adjusted healthy control distribution. The difference in the relative telomere lengths between patients and controls in both peripheral blood mononuclear cells and granulocytes was prominent in those under the age of 50 years. Contrary to the peripheral blood mononuclear cells, in which an inverse correlation was observed between relative telomere values and age, no significant correlation was noted between granulocyte telomere values and patient age. A significant correlation was observed between individual relative telomere values and absolute neutrophil counts. There was no difference in expression of telomerase reverse transcriptase in peripheral blood mononuclear cells between patients and controls but telomerase activity was identified at a significantly higher frequency in controls than in patients. No correlation was found between telomerase activity or telomerase reverse transcriptase expression and relative telomere lengths of peripheral blood mononuclear cells. Patients with chronic idiopathic neutropenia display age-inappropriate telomere shortening of peripheral blood cells and low telomerase activity in peripheral blood mononuclear cells. A compensatory increased proliferation of bone marrow hematopoietic progenitor cells in association with lymphocyte replicative exhaustion probably account for these abnormalities.

Experimental bone marrow failure in mice ameliorated by OCH via tippling the balance of released cytokines from Th1 to Th2

Background: OCH was reported to stimulate natural killer T (NKT) cells to produce predominantly T helper type 2 (Th2) cytokines. The present study was attempted to evaluate potential protection of OCH on acquired bone marrow failure syndromes (BMFS) in mice model.Methods: BMFS in mice model was established by exposure to sublethal irradiation followed by infusion with 5 × 106 B6 lymph nodes cells. Mice were injected intraperitoneally (I.P.) with either OCH or α-galactosylceramide (α-GC) at a dose of 100 µg kg−1 twice a week for two weeks after post-irradiation. The control mice were I.P. with vehicle alone (10% dimethyl sulfoxide in phosphate-buffered saline). Meanwhile, anti-interleukin-4 (anti-IL-4) monoclonal antibody (mAb) (500 µg/animal) was given I.P. 2 hours prior to vehicle or OCH administration. Both interferon-γ (IFN-γ) and IL-4 levels in the serum were measured by enzyme-linked immunosorbent assay. Colony-forming unit-granulocyte-macrophage (CFU-GM) by bone marrow (BM) mononuclear cells was counted. The percentage of NKT cell in BM cells and intracellular cytokines were determined by flow cytometry.Results: The treatment of OCH in vivo decreased the IFN-γ/IL-4 ratio in the serum, and increased the transformation of NKT cells into NKT2 cells. The treatment of OCH in vitro increased the colonies of CFU-GM.Conclusion: Our data suggests that OCH ameliorated immune-mediated BMFS in CByB6F1 mice via activation of NKT cells, and shifting the balance from Th1 to Th2.

Interleukin‐27 enhances the production of tumour necrosis factor‐α and interferon‐γ by bone marrow T lymphocytes in aplastic anaemia

Aplastic anaemia (AA) is considered as an immune-mediated bone marrow failure syndrome. The mechanism is involved with a variety of immune molecules including interferon-γ (IFN-γ), tumour necrosis factor-α (TNF-α) and interleukins (ILs). IL-27 is a novel member of the IL-12 family, which mediates T cell response and enhances the production of IFN-γ. However, little is known about the role of IL-27 in the development of AA. This study investigated the role of IL-27 and its receptor IL-27R in the pathogenesis of AA. Both the mRNA expression of IL-27/IL-27R subunits in the bone marrow mononuclear cells (BMMNCs) and the levels of IL-27 in the marrow plasma in AA were found to be higher than in controls. Increased IL-27 correlated with the disease severity of AA. Subsequently, we stimulated marrow T lymphocytes with recombinant human (rh)IL-27 and found that rhIL-27 enhanced the production of TNF-α and IFN-γ in both CD4(+) and CD8(+) T lymphocytes from AA patients. We also detected increased TNF-α and IFN-γ in the supernatants of BMMNCs from AA patients after IL-27 stimulation. In conclusion, our data suggest that elevated IL-27 and IL-27-induced TNF-α and IFN-γ overproduction might be involved in the pathogenesis of AA.