scholarly journals Decreased Tim-3 Expression Level of Peripheral Blood Natural Killer Cells in Severe Aplastic Anemia

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 4767-4767
Author(s):  
Rong Fu ◽  
Xin Yuan ◽  
Chunyan Liu ◽  
Hui Liu ◽  
Yihao Wang ◽  
...  
Keyword(s):  
Aplastic Anemia ◽  
Nk Cells ◽  
Peripheral Blood ◽  
Nk Cell ◽  
Conflicts Of Interest ◽  
Early Stage ◽  
Bone Marrow Failure ◽  
Severe Aplastic Anemia ◽  
Killer Cells ◽  
Before And After

Abstract Severe aplastic anemia (SAA) is a life threatening disease characterized by severe pancytopenia and bone marrow failure. T-cell immunoglobulin- and mucin domain-containing (Tim)-3 has been initially used to identify dysfunctional T cells, but recent studies have demonstrated that Tim-3 is widely detected on nature killer cells (NK cell) and may serves as a marker for activation and maturation of NK cells. In our study, Tim-3, expressed on peripheral blood of NK cells in SAA patients, was quantitatively analyzed by Flow cytometry before and after immunosuppressive therapy (IST). Results showed that the expression of Tim-3 in patients before IST [(61.11±15.46)%] was significantly lower than that in patients after IST[(74.30±12.63)%, P<0.05] and normal controls[(70.39±12.73)%, p<0.05]. However, no difference was observed between patients before and after IST. Therefore, we concluded that low expression of Tim-3 in NK cells may play a crucial role in early stage of SAA. Disclosures No relevant conflicts of interest to declare.

Aplastic Anemia in Thailand: Incidence and Treatment Outcome from a Prospective Nationwide Population-Based Study

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 8-9
Author(s):  
Lalita Norasetthada ◽  
Somchai Wongkhantee ◽  
Jindaratn Chaipokam ◽  
Kanyaporn Charoenprasert ◽  
Suporn Chuncharunee ◽  
...  
Keyword(s):  
Aplastic Anemia ◽  
Anabolic Steroid ◽  
Real World ◽  
Conflicts Of Interest ◽  
Bone Marrow Failure ◽  
Population Based ◽  
Annual Incidence ◽  
Severe Aplastic Anemia ◽  
Treatment Modalities ◽  
Population Based Study

Background: Incidence of Aplastic Anemia (AA) in Asia tends to be higher than in western countries, but contemporary real-world incidence and outcomes of AA in Asia remain limited. This study aimed to explore the incidence across the country regions and to evaluate the patient outcomes according to age, the severity of disease, and treatment modalities. Method: This is a prospective multicenter nationwide population-based observational study of patients with AA aged over 15 years old, diagnosed between August 1st, 2014 to July 31st, 2016, with a longitudinal follow-up period over 2 years, from 30 medical centers. Patients with suspected hypocellular MDS and congenital bone marrow failure syndrome were excluded. Results: During the study period of 2 years, there were 348 newly diagnosed patients with aplastic anemia, giving the annual incidence of 4.6 per million inhabitants. There was a higher annual incidence of severe (SAA) and very severe aplastic anemia (VSAA) (3.8 per million) than non-severe aplastic anemia (NSAA) (0.8 per million). The incidence was greater among older patients with a peak incidence in patients aged 60-89 years old. (Figure 1) There was a high variation in the geographic incidences across country regions, ranging from 2.6 to 6.6 per million per year. (Figure 2) The 2-year overall survival (OS) for NSAA, SAA, and VSAA were 65.5%, 49.3%, and 20.1%, respectively (P &lt; 0.001). Patients aged older than 60 years had the worst OS (42.6% as compared with 47.7% for the age 41-60 years and 64.5% for the age 15-40 years, P = 0.002). Among patients with SAA and VSAA (n = 280), the overall response rate (ORR) among patients treated with rabbit anti-thymocyte globulin and/or cyclosporin A (rATG±CsA) was significantly superior than those treated with CsA-based therapy and those treated with anabolic steroid (44.4% vs. 36.4% and 31.2%, respectively, P &lt; 0.001). Among evaluable patients, ORR after the 1st treatment with rATG±CsA at 3, 6, 12 and 24 months were 23.9%, 43.8%, 68.4% and 89.2%, respectively. The 2-year OS among SAA/VSAA patients treated with rATG±CsA, CsA-based therapy, and anabolic steroid were 54.8%, 54.5%, and 37.6% (P = 0.037), respectively (Figure 3). From multivariate analysis, age &gt; 60 years (HR 1.63, 95%CI, 1.14-2.33, P = 0.007), VSAA (HR 2.24, 95% CI, 1.45-3.46, P &lt; 0.001) and not receiving immunosuppressive therapy or anabolic steroid (HR 4.96, 95%CI, 2.88-8.54, P &lt;0.001), were independently associated with inferior OS among patients with SAA/VSAA. Conclusion: The incidence rate of AA in Thailand from this contemporary nationwide population-based study is high, especially in the elderly. Patients treated with rATG±CsA had superior survival than those receiving anabolic steroid. The real-world outcome of patients with SAA/VSAA, especially in those aged over 60 years, is substantially poor. Figure 1 Disclosures No relevant conflicts of interest to declare.

scholarly journals Battle of Thermopylae: 300 Spartans (natural killer cells plus obinutuzumab) versus the immortal warriors (chronic lymphocytic leukemia cells) of Xerxes’ army

2019 ◽  
Vol 5 (10) ◽  
pp. FSO425
Author(s):  
Ricardo García-Muñoz ◽  
María-Josefa Nájera ◽  
Jesús Feliu ◽  
Judith Antón-Remírez ◽  
Enrique Ramalle-Gómara ◽  
...  
Keyword(s):  
Chronic Lymphocytic Leukemia ◽  
Nk Cells ◽  
Natural Killer ◽  
Peripheral Blood ◽  
Nk Cell ◽  
Killer Cell ◽  
Lymphocytic Leukemia ◽  
Leukemic Cells ◽  
Lymphokine Activated Killer Cells ◽  
Killer Cells

Aim: To analyze the effects of subcutaneous or intravenous rituximab + lymphokine-activated killer cells, obinutuzumab or ibrutinib on natural killer (NK) cell levels in chronic lymphocytic leukemia and follicular lymphoma patients. Patients & methods: The distribution of peripheral blood NK cells of 31 patients was analyzed by flow cytometry. Results: We detected a decrease of NK cells in peripheral blood below normal range after obinutuzumab treatment. During maintenance treatment with subcutaneous rituximab, an NK cell reduction was less pronounced than after intravenous rituximab treatment, despite lymphokine-activated killer cell infusions. Conclusion: After one dose of obinutuzumab, each NK cell in peripheral blood destroys 25 leukemic cells.

Differential Genomic and Proteomic Signatures In Cord Blood (CB) Vs Peripheral Blood (PB) CD56+Dim NK Cells: Over Expression of CD34 In CB Vs PB CD56+Dim NK Cells

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2781-2781
Author(s):  
Nancy S Day ◽  
Evan Shereck ◽  
Janet Ayello ◽  
Catherine McGuinn ◽  
Prakash Satwani ◽  
...  
Keyword(s):  
Cell Death ◽  
Programmed Cell Death ◽  
Cord Blood ◽  
Nk Cells ◽  
Peripheral Blood ◽  
Nk Cell ◽  
Conflicts Of Interest ◽  
Hematopoietic Stem ◽  
Over Expression ◽  
Apoptotic Genes

Abstract Abstract 2781 Background. Umbilical cord blood (UCB) is a viable alternative source of allogeneic hematopoietic stem cells for the treatment of both malignant and non-malignant disease (Cairo et al BBMT 2008). UCB transplantation (UCBT) is known to be associated with decrease severe acute graft-versus-host disease (GvHD) compared to unrelated bone marrow (BM) and peripheral blood (PB) transplantation; however, it is associated with delayed hematopoietic and immune reconstitution (Szabolcs/Cairo et al Seminars in Hematology 2010). NK cells play important roles in both innate and adaptive immunity and are characterized as a CD56+ cell population. NK cell recovery is prompt by 2 months after hematopoietic stem cell transplantation (HSCT), while T-cell (after at least 9 mo HSCT) and B-cell (after 3 to 4 mo HSCT) reconstitutions are gradual and delayed. CD56+dim cells are primarily cytotoxic and make up 90% of PB NK populations (Shereck/Cairo PBC 2007). We previously demonstrated the ability to ex-vivo expand CB MNC into various phenotypes of CD56+dim and CD56+bright NK cells (totally 60%) and NKT cells (40%) with profound in vitro and in vivo cytotoxicity against hematological malignancies (Ayello/Cairo BBMT 2006 & Exp. Hematology 2009). Proteomic studies from our group demonstrated differential protein expression including ↑NKG2A, ↓IP3R type 3, ↓MAPKAPK5, and ↑NOTCH 2 in CB vs PB CD56+dim (Shereck/Cairo, ASH 2007; Shereck/Day/Cairo, ASBMT 2009). Objective. In these studies, we sought to determine the similarity or differences in genetic signatures in CB vs APB CD56+dim NK cells. Methods. CB MNCs were isolated on a ficoll gradient and NK CD56+16+dim cells isolated using a 2-step magnetic activated cell separation (MACS) process via a standard kit (Miltenyi Biotec). Enrichment was at least 94%. Isolated RNA from CB and PB CD56+dim cells were subjected to microarray studies (Affymetrix, U133A_2) as we have previously described (Jiang/Cairo et al J Immunol 2004). Data were analyzed by Agilent GeneSpring and Ingenuity pathway analyses. Welch test were used to perform statistical analysis and fold change of < 1.5 and values of p<0.05 were considered to be significant. Two-color ECL Plex fluorescence Western blotting (WB) was preformed to validate the proteomic data. Protein samples were separated using SDS-PAGE followed by transblotting. WB membranes were then incubated with target and control (GAPDH) primary antibodies. After rinse and wash, the membranes were further incubated with CY5 and CY3 conjugated secondary antibodies. The membranes were scanned with TYPHOON by green (532 laser and 580 filter) and red (633 laser and 670 filter) setting for CY3 and CY5, respectively, and then observed and quantified using ImageQuant. Results. CB vs PB CD56+dim cells significantly altered expressed 796 genes, in which 486 genes were over expressed, at the genomic level including: pro-apoptotic genes: CASP10 (3.1F), TNFSF11 (4.7F), CDC2 (3.0F), BCL2L1 (4.3F), NOTCH2 (1.5F); and cell development: PBX1 (7.6F), IL1RN (5.1F), CD24 (5.3F), CD34 (3.5F), CD55 (2.1F), CCL13 (2.2F). Conversely, there was significant under expression of NF1 (5.1F), MAP2K3 (1.7F), PIK3CD (2.1F), BAX (2.9F), and JUN (2.2F). Our WB results indicate that NOTCH2 (2.4F) and PBX1 (2.2F) proteins are increased in CB vs PB CD56+dim NK cells, consistent with our proteomic results. Conclusion. These results suggest that CB vs PB CD56+dim NK are more prone to undergo programmed cell death (apoptosis) secondary to over expression of numerous pro-apoptotic genes, and may be earlier in development (pro-NK) with over expression of the CD34 gene. Furthermore, decrease CB vs PB NK cytotoxicity maybe in part secondary to increase programmed cell death in particularly increase NOTCH2 at the genomic and proteomic levels. (The first two authors contribute equally.) Disclosures: No relevant conflicts of interest to declare.

scholarly journals The Effect of Pyruvate Kinase M2: Inhibitor(shikonin) on the Function of Mdc in Severe Aplastic Anemia Mouse Model

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 5021-5021
Author(s):  
Mengying Zheng ◽  
Chunyan Liu ◽  
Yuanyuan Shao ◽  
Rong Fu ◽  
Huaquan Wang ◽  
...  
Keyword(s):  
Mouse Model ◽  
Aplastic Anemia ◽  
Pyruvate Kinase ◽  
Conflicts Of Interest ◽  
Bone Marrow Failure ◽  
Autoimmune Response ◽  
Severe Aplastic Anemia ◽  
Pyruvate Kinase M2 ◽  
Cd8 Cells ◽  
Cytotoxic Molecules

Severe aplastic anemia (SAA) is a rare disease characterized by severe pancytopenia and bone marrow failure. Over-activated myeloid dendritic cells (mDCs) play an important role in the pathogenesis of SAA. In recent years, the role of pyruvate kinase M2 (PKM2) in DC function and autoimmune response has been gradually recognized. In this study, an immune attack-mediated AA mouse model was constructed by total body irradiation and lymphocyte infusion. The AA model mice showed pancytopenia, decreased ratio of CD4+/CD8+ cells, increased expression of cytotoxic molecules perforin and granzyme in CD8+ cells, increased levels of co-stimulatory molecules CD80 and CD86 in DCs and inadequate Treg number. In-vitro animal experiments confirmed the activation of PKM2 in mDCs, which promoted glycolytic metabolism. High levels of PKM2 in mice contributed to the poor survival rate. Additionally, intervention treatment with shikonin or cyclosporin A in the AA mouse model reduced the expression not only of co-stimulatory molecules CD80 and CD86 in mDCs but also of cytotoxic molecules in CD8+ cells. In conclusion, we confirmed the activation of PKM2 in mDCs in AA mouse models. PKM2 is involved in mDC activation and proliferation, which might contribute to activating the downstream cytotoxic T cells (CTLs). PKM2 is a possible novel target in SAA treatment. Disclosures No relevant conflicts of interest to declare.

T/NK Lymphocytosis in CML Ph+ Patients During Dasatinib Therapy.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3279-3279
Author(s):  
Antonella Russo Rossi ◽  
Alessandra Ricco ◽  
Paola Carluccio ◽  
Mario Delia ◽  
Manuela Leo ◽  
...  
Keyword(s):  
Nk Cells ◽  
Peripheral Blood ◽  
Nk Cell ◽  
Conflicts Of Interest ◽  
Cytotoxic T Cell ◽  
Molecular Response ◽  
Imatinib Treatment ◽  
Long Term Outcome ◽  
Significant Difference ◽  
The Impact

Abstract Abstract 3279 Poster Board III-1 Background. Some authors reported that Natural Killer (NK) cells from CML patients are defective in NK cell activity and NK cell number decrease as the disease progresses to the advanced phase and probably the abnormal BCR/ABL gene causes abnormal NK cell differentiation.TK inhibitors reduce BCR/ABL transcription and could restore NK cell numbers and/or function.Moreover Dasatinib,by the blockade of SRC Kinases,could affect the development of NK cells as well as T-lymphocytes.Our aim was to verify the impact of Dasatinib treatment on T CD8+ and NK cells modulation. Methods. We evaluated 24 patients with CML resistant/intollerant to Imatinib and treated with Dasatinib at a starting dose of 70 mg/BID or 100 mg/QD.Blood count were monitored; lymphocytosis has been definited by an increased number of peripheral blood lymphocyte counts ≥ 3.0×10(e)9/L and by the predominance of LGLs in peripheral blood smear. Immunophenotyping was done with flow-cytometry using antibodies against the following antigens: CD2, CD3, CD4, CD5, CD7, CD8, CD16, and CD56. Results. With a median of 19 mo. of Dasatinib therapy (range 3–43), 15/24 cases (62.5%) developed peripheral blood lymphocytosis. Median onset of lymphocytosis was 3 months after the initiation of Dasatinib therapy (range 1–12) and duration was 14 months (range 6–40).Lymphocytosis was CD3+/CD8+/Cytotoxic T Cell in 9 patients (60%) and CD3-/CD16+/CD56+/NK Cell in 6 patients (40%).In all 15 patients no symptoms or signs suggestive of LGL leukemia or viral infections were documented.There was no significant difference in terms of the frequency of severe adverse events, including pleural effusion between patients with and without lymphocytosis. 11(73%) of the 15 patients who developed lymphocytosis achieved MMolR and 4/11 presented Bcr/Abl mutation at the time of imatinib treatment (F317L, E255K, F359V, E255K),whereas only 3(33%) of the 9 patients without lymphocytosis achieved MMolR and 1/3 presented Bcr/Abl mutation (F359V). Moreover molecular response was earlier in the group of patients with lymphocytosis (8vs12 mo.). Conclusions. The development of lymphocytosis in our patients seems to be associated to an improved response to dasatinib in terms of molecular response and time to response. The assessment of higher frequency lymphocytosis requires further analysis; a larger patients'cohort should be needed to explore the biological role of lymphocytosis and the impact on the long term outcome in patients treated with dasatinib. Disclosures: No relevant conflicts of interest to declare.

Therapeutic Effects of ALT-803, an IL-15 Superagonist, in Combination with Anti-CD20 Chimeric Antigen Receptor Modified Expanded Natural Killer CELLS Against Pediatric Burkitt Lymphoma (BL)

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3085-3085 ◽  
Author(s):  
Yaya Chu ◽  
Fangyu Lee ◽  
Janet Ayello ◽  
Brian Hang ◽  
Melanie Zhang ◽  
...  
Keyword(s):  
Natural Killer Cells ◽  
Nk Cells ◽  
Natural Killer ◽  
Peripheral Blood ◽  
Burkitt Lymphoma ◽  
Nk Cell ◽  
Target Cells ◽  
Killer Cells ◽  
Anti Cd20

Abstract Background: The outcome for children with Burkitt lymphoma (BL)has improved significantly but for patients who relapse, the prognosis is dismal due to chemo-immunotherapy resistance (Cairo et al, JCO, 2012, Cairo et al, Blood, 2007). NK cells are bone marrow-derived cytotoxic lymphocytes that play a major role in the rejection of tumors. A variety of activating and inhibitory receptors on the NK cell surface are engaged to regulate NK cell activities and to discriminate target cells from other healthy 'self' cells. However, NK therapy is limited by several factors, including small numbers of active NK cells in unmodified peripheral blood, lack of tumor targeting specificity, and multiple mechanisms of tumor escape of NK cell immunosurveillance. Our group has successfully modified expanded peripheral blood Natural Killer cells (exPBNK) with an anti-CD20 CAR to target rituximab sensitive/resistant CD20+ BL cells in vitro and in NSG mice (Chu/Cairo, et al, Can Imm Res 2015). However, the short lifespan/persistence of adoptively transferred NK cells has limited the therapeutic efficacy. ALT-803 (Altor BioScience Corporation) is a superagonist of an IL-15 variant bound to an IL-15Rα-Fc fusion with enhanced IL-15 biological activity (Zhu et al. 2009 J Immunol), longer half-life and increased potency (Han, et al. Cytokine. 2011). It is currently in several clinical trials in patients with variety of cancers such as refractory indolent non-Hodgkin's lymphoma (NCT02384954). Objective: We hypothesize that ALT-803, IL-15 superagonist complex, promotes exPBNK persistence and significantly enhances the cytotoxicity of anti-CD20 CAR exPBNK against CD20+ BL. Method: PBMCs were expanded with lethally irradiated K562-mbIL21-41BBL cells (Dean Lee et al, PLoS One, 2012). CD56+ CD3- exPBNK cells were isolated using Miltenyi NK cell isolation kit. Anti-CD20-4-1BB-CD3 ζ mRNA (CAR mRNA) was producedin vitro and nucleofected into exPBNK as we have previously described (Chu/Cairo, et al, Can Imm Res 2015). ALT-803 was provided by Altor BioScience Corporation. ExPBNK cells were cultured with 0.35ng/ml or 3.5ng/ml ALT-803. NK proliferation was monitored with MTS assays. NK receptors expression and cytotoxicity were examined by flow cytometry (Chu/Cairo, et al, ASH 2014). NK resistant BL cells Raji and Daudi were used as target cells. Results: % CD56+ CD3- PBNK cells were significantly increased compared to media alone at day 14 (mean 81.85% vs 14.91%, n=3, p<0.001) when co-cultured with the irradiated feeder cell K562-mbIL21-41BBL. The absolute NK numbers were enhanced with irradiated K562-mbIL21-41BBL cells as feeders compared to IL-2 alone after normalized to the INPUT NK cell numbers (mean 2247 fold±293.7 vs 0.516 fold±0.225, n=3, p<0.001) at day 14. Different doses of ALT-803 or IgG were added to the culture medium of purified expanded exPBNK. Proliferation assays were performed at day 3, 7,11, and 17. ALT-803 significantly promoted exPBNK proliferation and persistence compared to IgG in vitro in a dose-dependent manner (A490 reading at 3.5ng/ml dose: ALT803 vs IgG=0.3383+0.009 vs 0.0987+0.0007, P<0.0001 at d17). And ALT-803 significantly enhanced exPBNK cytotoxicity against NK resistant BL cells: Raji (ALT803 vs IgG= 49.54%+2.7% vs 5.99+0.34%, p<0.001, E:T=10:1) and Daudi (ALT803 vs IgG= 63.73%+3.09% vs 2.58+1.96%, p<0.001, E:T=10:1). It also maintained the highcytoxicity of exPBNK at d4, d10 and d18 against Raji (E:T=10:1, d4 vs d10 vs d18=62.07% vs 49.54% vs 61.47%) and against Daudi (E:T=10:1, d4 vs d10 vs d18=76.02% vs 63.73% vs 55%) by maintaining the activating receptors expression such as NKp30, NKp44, and NKp46. Further-more, we demonstrated ALT-803 significantly enhanced the cytotoxicity of anti-CD20 CAR modified exPBNK against Raji (CAR vs MOCK= 81.19%+0.35% vs 66.19+0.94%, p<0.001, E:T=10:1) and Daudi (CAR vs MOCK= 91.41%+0.45% vs 80.56+1.07%, p<0.001, E:T=10:1) compared to mock modified exPBNK. ALT-803 also significantly enhanced the cytotoxicity of anti-CD20 CAR modified exPBNK against NK resistant BL cells: Raji and Daudi compared to anti-CD20 CAR modified exPBNK maintained in medium without ALT803 (Fig.1). Conclusions: ALT-803 maintained the cytotoxicity of exPBNK and in vitro persistence and significantly enhanced anti-CD20 CAR exPBNK cytotoxicity against pediatric NK resistant BL. The in vivo effect of ALT-803 on CAR exPBNK using humanized NSG models is under investigation. Disclosures Wong: Altor BioScience Corporation: Employment, Other: stockholder of Altor Bioscience Corporation. Lee:Intrexon, Ziopharm, Cyto-Sen: Equity Ownership.

Decreased TIM-3 Expression Level of Peripheral Blood Natural Killer Cells in Severe Aplastic Anemia

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 5078-5078
Author(s):  
Rong Fu ◽  
Tian Zhang ◽  
Xin Yuan ◽  
Zonghong Shao
Keyword(s):  
T Cells ◽  
Aplastic Anemia ◽  
Nk Cells ◽  
Mrna Expression ◽  
Natural Killer ◽  
Peripheral Blood ◽  
Statistical Significance ◽  
Severe Aplastic Anemia ◽  
Hematopoietic Stem ◽  
Normal Controls

Abstract Severe aplastic anemia (SAA) is a life threatening disease characterized by severe pancytopenia and bone marrow failure. Natural killer (NK) cells are large granular lymphocytes which are one important component of the innate immune system and play a core role in regulation of adaptive immunity. T cell immunoglobulin mucin-3 (TIM-3), a member of the TIM family, appears to play an important negative regulatory role in T cells initial. Now, TIM-3 is widely detected on NK cells, and may contribute as a marker for activation and maturation of NK cells. Our previous studies have confirmed that the decrease of total NK cells, and CD56bright, CD56dim NK cell subsets and the higher expressions of NKp46 and perforin on NK cells may cause the over-function of T lymphocytes and thus lead to hematopoiesis failure in SAA. But, the expression of TIM-3 on NK cells in patients with SAA was still unknown. In this study, we investigated the expression of TIM-3 and its mRNA on NK cells in peripheral blood of untreated and recovered SAA patients by flow cytometry and Real-time PCR. Results showed that the expression of TIM-3 on peripheral blood NK cells in SAA patients before IST was (63.57±12.14) %, which was significantly decreased than that in normal controls (85.62±9.03) % ( p<0.01). The expression of TIM-3 on CD56dim NK cells was (66.41±11.74) % and (83.83±1.59) % separately in SAA patients before IST and normal controls. TIM-3 expressed in SAA patients before IST was lower than that in normal controls ( p<0.01). We also measured TIM-3 expressed on the surface of CD56bright NK cells, but the result showed that there was no statistical difference between SAA patients before IST (61.11±24.99%) and normal controls (62.64±12.06%) (p>0.05). More interesting, the expression of TIM-3 on NK cells was (75.88±12.83) % in SAA patients after IST, which was significantly increased than that in SAA patients before IST, and was no difference with normal controls. As well, we found TIM-3 expression on both CD56dim NK cells and CD56bright NK cells in SAA patients after IST also has a rising trend compared with SAA patients before IST. However, these differences had no statistical significance. Further, we detected TIM-3 mRNA expression in NK cells isolated from peripheral blood. TIM-3 mRNA expression in peripheral blood NK cells from newly diagnosis SAA patients, recovering SAA patients and normal controls was evaluated, respectively. The relative TIM-3 mRNA expression was significantly increased in NK cells in SAA patients after IST compared with SAA patients before IST and normal controls, and this difference had statistical significance. Meanwhile, the relative TIM-3 mRNA expression was lower in NK cells in SAA patients before IST compared with normal controls. However, the difference had no statistical significance. In SAA patients, the expression of TIM-3 on NK cells was positively correlated with the level of WBC (r=0.685, p=0.000), proportion of neutrophil (r=0.825, p=0.000), and proportion of reticulocyte in peripheral blood (r=0.465, p=0.029). And it was negatively correlated with the proportion of lymphocyte in peripheral blood (r=-0.802, p=0.000). According to our series studies, we hypothesize that the lower numbers and dysfunction of NK cells induce the failure of suppressing the over function of DC cells and T cells, that lead damage to healthy hematopoietic stem cells and the onset of SAA. Disclosures No relevant conflicts of interest to declare.

Regulatory Mutations in GATA2 Associated with Aplastic Anemia

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 3488-3488 ◽  
Author(s):  
Danielle M. Townsley ◽  
Amy Hsu ◽  
Bogdan Dumitriu ◽  
Steven M. Holland ◽  
Neal S. Young
Keyword(s):  
Bone Marrow ◽  
Aplastic Anemia ◽  
Immunosuppressive Therapy ◽  
Nk Cell ◽  
Conflicts Of Interest ◽  
Bone Marrow Failure ◽  
Early Adulthood ◽  
Mycobacterial Disease ◽  
Regulatory Regions ◽  
Exon 2

Abstract Abstract 3488 Germline heterozygous mutations in GATA2 have been reported to cause familial myelodysplasia–acute myeloid leukemia (MDS/AML), monocytopenia and mycobacterial infection (monoMAC syndrome), dendritic cell, myeloid and NK cell lymphopenia (DCML), and Emberger syndrome (lymphedema and MDS). GATA2 is a zinc finger transcription factor that plays a crucial role in regulating growth of hematopoietic progenitors. In some pedigrees, patients or family members have manifestations of bone marrow failure. We hypothesized that patients with aplastic anemia (AA) may harbor mutations in GATA2. The coding regions and regulatory regions of GATA2 were sequenced in 99 patients with confirmed AA. Sequences from 100 normal individuals as well as published human genomes from unaffected individuals (dbSNP build 135 and 1000 Genomes Project) were used as controls. Genetic variants were confirmed in hematopoietic and somatic tissues. We identified 4 heterozygous mutations in regulatory regions of GATA2 in 5 patients. In two patients, a mutation at nucleotide 59T>G in exon 1 of isoform 2 was identified; both had severe AA in early adulthood refractory to immunosuppressive therapy. We noted this 59T>G mutation in two unrelated individuals with severe disseminated mycobacterial disease. We identified a mutation at nucleotide 20G>A in exon 2 of isoform 1, in a 3 year-old male with hepatitis-associated severe AA whose disease was refractory to multiple rounds of immunosuppressive therapy. Another mutation was present in 38G>A in exon 2 of isoform 1 in a 32 year old male with moderate AA and paroxysmal nocturnal hemoglobinuria (PNH). We also identified the exon 2 38G>A mutation in a patient with disseminated mycobacterial disease where reduced transcription of the mutant 38G>A allele was noted on RT-PCR. Finally, an intron 5, c.512+573 G>A variant was identified in an 18 year old male with severe AA who progressed after immunosuppressive therapy to MDS/AML. This variant, which causes a disruption of the FLI1 binding site, has also been found to be pathogenic in monoMAC syndrome. In summary, a subset of patients with AA were found to have mutations in GATA2 suggesting a role for the gene in the pathogenesis of bone marrow failure. It also may identify patients at higher risk of infectious complications, those who may have less advantageous responses to immune suppression, and command earlier bone marrow transplantation. Disclosures: No relevant conflicts of interest to declare.

Decreased TIM-3 expression of peripheral blood natural killer cells in patients with severe aplastic anemia

2017 ◽  
Vol 318 ◽  
pp. 17-22 ◽  
Author(s):  
Tian Zhang ◽  
Xin Yuan ◽  
Chunyan Liu ◽  
Yi Li ◽  
Hui Liu ◽  
...  
Keyword(s):  
Natural Killer Cells ◽  
Aplastic Anemia ◽  
Natural Killer ◽  
Peripheral Blood ◽  
Severe Aplastic Anemia ◽  
Killer Cells

NK Cell Subsets Isolated From Human Thymus Differ From Peripheral Blood NK Cells in Their Cytotoxic and Cytokine Secreting Capability

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 1053-1053
Author(s):  
Stephanie Krieg ◽  
Wolfgang Koehler ◽  
Gordon Heidkamp ◽  
Julia Schneider ◽  
Franziska Ganss ◽  
...  
Keyword(s):  
Nk Cells ◽  
Peripheral Blood ◽  
Nk Cell ◽  
Conflicts Of Interest ◽  
Immune Therapy ◽  
Expression Patterns ◽  
Killer Cell ◽  
Lymphoid Tissues ◽  
Special Focus ◽  
Cell Subpopulations

Abstract Abstract 1053 Until recently, studies on human NK cells have been mainly focused on NK cells from peripheral blood. Little is known about the functional role of human NK cell subpopulations in different lymphoid organs. We therefore aimed to characterize the phenotypical and functional profile of human NK cell subsets from different organs with a special focus on early forms of thymic NK cells in comparison to mature peripheral blood NK cells from adults and children. Human NK cells can be classified according to the expression of CD56 and CD16 into immunoregulatory, cytokine producing CD56highCD16dim and mature cytotoxic CD56dimCD16high NK cells. It is widely accepted that NK cell development takes place in secondary lymphoid tissues leading to the maturation from CD56highCD16dim into CD56dimCD16high NK cells. Additionally, it has been reported that thymic CD34+ cells can differentiate into NK cells under treatment with IL-2 in vitro. Here we report for the first time on a different composition of human NK cell subpopulations in the thymus compared to peripheral blood. NK cells isolated from the thymus did not only express lower levels of CD56 but also showed a different ratio of CD16high/CD16dim NK cells compared to peripheral blood NK cells. Extensive phenotypical analysis revealed significant alterations in expression patterns of killer-cell immunoglobulin-like receptors, natural cytotoxicity receptors and other maturation or differentiation markers on thymic NK cells. Functional analyses showed that the whole NK cell population from thymus as well as from peripheral blood of young children produced only low amounts of IFN-γ compared to adult peripheral blood NK cells. Furthermore, functional assays of sorted NK cell subsets revealed important differences of CD16high and CD16dim thymic NK cells. Upon stimulation with IL-2 +/− K562 tumor cells, thymic CD16 high NK cells gain cytokine producing capacity, while CD16dim develop a tumor killing capacity comparable to naive peripheral blood NK cells. In summary, this comparative study of NK cell subsets provides important information on the development and function of NK cells that may be of great value for the optimization of cellular immune therapy. Disclosures: No relevant conflicts of interest to declare.

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