Browsing by Author "Kittiphong Paiboonsukwong"
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Publication Metadata only Abnormal red blood cell morphological changes in thalassaemia associated with iron overload and oxidative stress(2019-08-01) Pornthip Chaichompoo; Ariz Qillah; Pornpan Sirankapracha; Jirada Kaewchuchuen; Poramate Rimthong; Kittiphong Paiboonsukwong; Suthat Fucharoen; Saovaros Svasti; Suchin Worawichawong; Faculty of Medicine, Ramathibodi Hospital, Mahidol University; Mahidol University© Author(s) (or their employer(s)) 2019. No commercial re-use. See rights and permissions. Published by BMJ. Aims Iron overload is a major factor contributing to the overall pathology of thalassaemia, which is primarily mediated by ineffective erythropoiesis and shorter mature red blood cell (RBC) survival. Iron accumulation in RBCs generates reactive oxygen species (ROS) that cause cellular damage such as lipid peroxidation and RBC membrane deformation. Abnormal RBCs in patients with thalassaemia are commonly known as microcytic hypochromic anaemia with poikilocytosis. However, iron and ROS accumulation in RBCs as related to RBC morphological changes in patients with thalassaemia has not been reported. Methods Twenty-one patients with thalassaemia, including HbH, HbH with Hb Constant Spring and β-thalassaemia/HbE (splenectomy and non-splenectomy) genotypes, and five normal subjects were recruited. RBC morphology was analysed by light and scanning electron microscopy. Systemic and RBC iron status and oxidative stress were examined. Results Decreased normocytes were observed in the samples of patients with thalassaemia, with RBC morphological abnormality being related to the type of disease (α-thalassaemia or β-thalassaemia) and splenic status. Target cells and crenated cells were mainly found in splenectomised patients with β-thalassaemia/HbE, while target cells and teardrop cells were found in non-splenectomised patients. Patients with thalassaemia had high levels of serum ferritin, red cell ferritin and ROS in RBCs compared with normal subjects (p<0.05). Negative correlations between the amount of normocytes and serum ferritin (r s =-0.518, p=0.011), red cell ferritin (r s =-0.467, p=0.025) or ROS in RBCs (r s =-0.672, p<0.001) were observed. Conclusions Iron overload and its consequent intracellular oxidative stress in RBCs were associated with reduce normocytes in patients with thalassaemia.Publication Metadata only Decreased nitrite reductase activity of deoxyhemoglobin correlates with platelet activation in hemoglobin E/ß-thalassemia subjects(2018-09-01) Attaphon Chamchoi; Sirada Srihirun; Kittiphong Paiboonsukwong; Thanaporn Sriwantana; Piyadon Sathavorasmith; Kovit Pattanapanyasat; Rhoda Elison Hirsch; Alan N. Schechter; Nathawut Sibmooh; Mahidol University; Faculty of Medicine, Siriraj Hospital, Mahidol University; National Institute of Diabetes and Digestive and Kidney Diseases; Albert Einstein College of Medicine of Yeshiva University© Copyright: This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication. Nitric oxide (NO) can be generated from nitrite by reductase activity of deoxygenated hemoglobin (deoxyHb) apparently to facilitate tissue perfusion under hypoxic condition. Although hemoglobin E (HbE) solutions have been shown to exhibit decreased rate of nitrite reduction to NO, this observation has never been reported in erythrocytes from subjects with hemoglobin E/ß-thalassemia (HbE/ß-thal). In this study, we investigated the nitrite reductase activity of deoxyHb dialysates from 58 non-splenectomized and 23 splenectomized HbE/ß-thal subjects compared to 47 age- and sex-matched normal subjects, and examined its correlation with platelet activity. Iron-nitrosyl-hemoglobin (HbNO) was measured by tri-iodide reductive chemiluminescence as a marker of NO generation. HbNO produced from the reaction of nitrite with deoxyHb dialysate from both non-splenectomized and splenectomized HbE/ß-thal subjects was lower than that of normal (AA) hemoglobin subjects. P-selectin expression, a marker of platelet activation, at baseline and in reactivity to stimulation by adenosine diphosphate (ADP), were higher in HbE/ß-thal subjects than normal subjects. HbNO formation from the reactions of nitrite and deoxyHb inversely correlated with baseline platelet P-selectin expression, HbE levels, and tricuspid regurgitant velocity (TRV). Nitrite plus deoxygenated erythrocytes from HbE/ß-thal subjects had a lower ability to inhibit ADP-induced P-selectin expression on platelets than erythrocytes from normal subjects. We conclude that deoxyHb in erythrocytes from HbE/ß-thal subjects has a decreased ability to reduce nitrite to NO, which is correlated with increased platelet activity in these individuals.Publication Metadata only Deferiprone increases endothelial nitric oxide synthase phosphorylation and nitric oxide production(2018-01-01) Thanaporn Sriwantana; Pornpun Vivithanaporn; Kittiphong Paiboonsukwong; Krit Rattanawonsakul; Sirada Srihirun; Nathawut Sibmooh; Mahidol University© 2018, Canadian Science Publishing. All rights reserved. Iron chelation can improve endothelial function. However, effect on endothelial function of deferiprone has not been reported. We hypothesized deferiprone could promote nitric oxide (NO) production in endothelial cells. We studied effects of deferiprone on blood nitrite and blood pressure after single oral dose (25 mg/kg) in healthy subjects and hemoglobin E/β-thalassemia patients. Further, effects of deferiprone on NO production and endothelial NO synthase (eNOS) phosphorylation in primary human pulmonary artery endothelial cells (HPAEC) were investigated in vitro. Blood nitrite levels were higher in patients with deferiprone therapy than those without deferiprone (P = 0.023, n = 16 each). Deferiprone increased nitrite in plasma and whole blood of healthy subjects (P = 0.002 and 0.044) and thalassemia patients (P = 0.003 and 0.046) at time 180 min (n = 20 each). Asymptomatic reduction in diastolic blood pressure (P = 0.005) and increase in heart rate (P = 0.009) were observed in healthy subjects, but not in thalassemia patients. In HPAEC, deferiprone increased cellular nitrite and phospho-eNOS (Ser1177) (P = 0.012 and 0.035, n = 6) without alteration in total eNOS protein and mRNA. We conclude that deferiprone can induce NO production by enhancing eNOS phosphorylation in endothelial cells.Publication Metadata only Derivation of the human induced pluripotent stem cell line MUi017-A from a patient with homozygous Hemoglobin Constant Spring(2017-04-01) Wasinee Wongkummool; Warun Maneepitasut; Thongperm Munkongdee; Pirut Tong-ngam; Amornrat Tangprasittipap; Saovaros Svasti; Narisorn Kitiyanant; Kittiphong Paiboonsukwong; Suthat Fucharoen; Alisa Tubsuwan; Mahidol University© 2017 The Authors Hemoglobin Constant Spring (HbCS, HBA2: c.427T > C) is a common nondeletional α-thalassemia resulting from a nucleotide substitution at the termination codon of the HBA2 gene. Homozygosity for HbCS is characterized with mild anemia, jaundice, and splenomegaly. In this study, the human induced pluripotent stem cell line MUi017-A was successfully generated from peripheral blood CD34 + hematopoietic progenitors of a 52 year old female with homozygous HbCS. The MUi017-A cell line exhibited embryonic stem cell characteristics with consistent expression of specific pluripotency markers and the capability of differentiating into the three germ layers. The cell line may be used for the disease modeling.Publication Metadata only Development of DNA controls for detection of β-thalassemia mutations commonly found in Asian(2020-01-01) Thongperm Munkongdee; Tiwaporn Nualkaew; Nattrika Buasuwan; Nurmeeha Hinna; Kittiphong Paiboonsukwong; Orapan Sripichai; Saovaros Svasti; Pranee Winichagoon; Suthat Fucharoen; Natee Jearawiriyapaisarn; Mahidol University; Faculty of Medicine, Siriraj Hospital, Mahidol University; National Institutes of Health (NIH)© 2020 John Wiley & Sons Ltd Introduction: Several DNA-based approaches including a reverse dot-blot hybridization (RDB) have been established for detection of β-thalassemia genotypes to provide accurate genetic counseling and prenatal diagnosis for prevention and control of severe β-thalassemia. However, one of major concerns of these techniques is a risk of misdiagnosis due to a lack of DNA controls. Here, we constructed positive DNA controls for β-thalassemia genotyping in order to ensure that all steps in the analysis are performed properly. Methods: Four recombinant β-globin plasmids, including a normal sequence and three different mutant panels covering 10 common β-thalassemia mutations in Asia, were constructed by a conventional cloning method followed by sequential rounds of site-directed mutagenesis. These positive DNA controls were further validated by RDB analysis. Results: We demonstrated the applicability of established positive DNA controls for β-thalassemia genotyping in terms of accuracy and reproducibility by RDB analysis. We further combined three mutant β-globin plasmids into a single positive control, which showed positive signals for both normal and mutant probes of all tested mutations. Therefore, only two positive DNA controls, normal and combined mutant β-globin plasmids, are required for detecting 10 common β-thalassemia mutations by RDB, reducing the cost, time, and efforts in the routine diagnosis. Conclusion: The β-globin DNA controls established here provide efficient alternatives to a conventional DNA source from peripheral blood, which is more difficult to obtain. They also provide a platform for future development of β-globin plasmid controls with other mutations, which can also be suitable for other DNA-based approaches.Publication Metadata only Elevated levels of circulating monocytic myeloid derived suppressor cells in splenectomised β-thalassaemia/HbE patients(2020-01-01) Sirikwan Siriworadetkun; Rattanawan Thubthed; Chayada Thiengtavor; Kittiphong Paiboonsukwong; Archrob Khuhapinant; Suthat Fucharoen; Kovit Pattanapanyasat; Jim Vadolas; Saovaros Svasti; Pornthip Chaichompoo; Ramkhamhaeng University; Monash University; Mahidol University; Faculty of Medicine, Siriraj Hospital, Mahidol University; Hudson Institute of Medical ResearchPublication Metadata only Establishment of MUi009 – A human induced pluripotent stem cells from a 32 year old male with homozygous β°-thalassemia coinherited with heterozygous α-thalassemia 2(2017-04-01) Wasinee Wongkummool; Warun Maneepitasut; Pirut Tong-ngam; Amornrat Tangprasittipap; Thongperm Munkongdee; Chanikarn Boonchuay; Saovaros Svasti; Narisorn Kitiyanant; Kittiphong Paiboonsukwong; Suthat Fucharoen; Alisa Tubsuwan; Mahidol University© 2017 The Authors The thalassemias are a group of genetic disorders characterized by a deficiency in the synthesis of globin chains. In this study the MUi009-A human induced pluripotent stem cell line was successfully generated from peripheral blood CD34 + haematopoietic progenitors of a 32 year old male who had coinherited a homozygous β°-thalassemia mutation at codon 41/42 (-TCTT) and a heterozygous α-thalassemia 4.2 deletion. The MUi009-A cell line exhibited embryonic stem cell characteristics with consistent pluripotency marker expression and the capability of differentiating into the three germ layers. The cell line may provide a tool for drug testing and gene therapy studies.Publication Metadata only Glycated Albumin Measurement Using an Electrochemical Aptasensor for Screening and Monitoring of Diabetes Mellitus(2019-11-01) Sasinee Bunyarataphan; Tararaj Dharakul; Suthat Fucharoen; Kittiphong Paiboonsukwong; Deanpen Japrung; Mahidol University; Faculty of Medicine, Siriraj Hospital, Mahidol University; Thailand National Science and Technology Development Agency© 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim Glycated albumin (GHSA) is a medium-term glycaemic control marker of diabetes, which can be used as an alternative to or together with glycated haemoglobin (HbA1c). Currently available methods for the measurement of GHSA are limited in clinical practice because they involve slow and cumbersome processes of sample preparation, proteolytic digestion, and thermal incubation, and they suffer from limited analytical performance, and/or a lack of normalization to total albumin (HSA) levels. In this paper, we developed a simple electrochemical biosensor to measure GHSA values based on two DNA aptamers that specifically bind to GHSA and HSA. We used square wave voltammetry (SWV) to measure binding of the target proteins to their specific biotinylated aptamers, which had been immobilised on separate streptavidin (STR)-modified screen-printed carbon electrodes (SPCEs), in the presence of the redox mediator ferricyanide (Fe(CN)63−). This electrochemical aptasensing system had a detection limit of 3 ng/ml for GHSA and 0.2 μg/ml for HSA. The results exhibited high selectivity for GHSA over other molecules present in the blood. The developed sensor was able to measure the amount of GHSA in plasma samples. A statistically significant difference was observed in the elevated plasma GHSA levels in diabetic versus non-diabetic patients. Moreover, the trends in these GHSA levels were consistent with those obtained using the HbA1c test. The sensing system reported herein could be applied as a point-of-care-testing (POCT) device for measurements of clinically relevant GHSA values.Publication Metadata only Graphene based aptasensor for glycated albumin in diabetes mellitus diagnosis and monitoring(2016-08-15) Chayachon Apiwat; Patraporn Luksirikul; Pacharapon Kankla; Prapasiri Pongprayoon; Kiatnida Treerattrakoon; Kittiphong Paiboonsukwong; Suthat Fucharoen; Tararaj Dharakul; Deanpen Japrung; Thailand National Science and Technology Development Agency; Kasetsart University; Mahidol University© 2016 Elsevier B.V. We selected and modified DNA aptamers specifically bound glycated human serum albumin (GHSA), which is an intermediate marker for diabetes mellitus. Our aptamer truncation study indicated that the hairpin-loop structure with 23 nucleotides length containing triple G-C hairpins and 15-nucleotide loop, plays an important role in GHSA binding. Fluorescent quenching graphene oxide (GO) and Cy5-labeled G8 aptamer were used in this study to develop simple and sensitive graphene based aptasensor for GHSA detection. The limit of detection (LOD) of our aptasensor was 50 μg/mL, which was lower than other existing methods. In addition, with the nuclease resistance system, our GHSA detection platform could also be used in clinical samples. Importantly, our approach could significantly reveal the higher levels of GHSA concentrations in diabetes than normal serums. These indicate that our aptasensor has a potential for diagnosis and monitoring of diabetes mellitus.Publication Metadata only Hemoglobin-bound platelets correlate with the increased platelet activity in hemoglobin E/β-thalassemia(2020-01-01) Attaphon Chamchoi; Sirada Srihirun; Kittiphong Paiboonsukwong; Thanaporn Sriwantana; Peerawich Kongkaew; Suthat Fucharoen; Kovit Pattanapanyasat; Nathawut Sibmooh; Mahidol University; Faculty of Medicine, Siriraj Hospital, Mahidol University© 2020 John Wiley & Sons Ltd Introduction: An increase in platelet activity is a contributing factor to vascular complications in hemoglobin E/β-thalassemia (HbE/β-thal). Plasma-free hemoglobin (Hb) increases in HbE/β-thal patients and correlates with platelet activation, but the levels of Hb-bound platelets have never been reported. In this study, we aimed to investigate the levels of Hb-bound platelets and its association with platelet activity in HbE/β-thal patients. Methods: Hb-bound platelets were measured by flow cytometry in 22 healthy subjects and 26 HbE/β-thal patients (16 nonsplenectomized and 10 splenectomized HbE/β-thal patients). Plasma Hb was measured by the chemiluminescence method based on the consumption of nitric oxide (NO) by Hb. Expression of P-selectin and activated glycoprotein (aGP) IIb/IIIa on platelets was measured by flow cytometry as a marker of platelet activity. Results: Both nonsplenectomized and splenectomized HbE/β-thal patients had higher levels of Hb-bound platelets and plasma Hb than healthy subjects. In vitro incubation of dialyzed Hb from patients with platelets of healthy subjects caused an increase in Hb-bound platelets, which was partially inhibited by anti-GPIbα antibody. Plasma Hb positively correlated with Hb-bound platelets. Platelet P-selectin expression at baseline and in response to adenosine diphosphate (ADP, 1 µM) stimulation was higher in nonsplenectomized and splenectomized HbE/β-thal patients than healthy subjects. The ADP-induced aGPIIb/IIIa expression on platelets was also higher in HbE/β-thal patients than healthy subjects. Hb-bound platelets correlated with baseline P-selectin expression and ADP-induced P-selectin expression. Conclusion: HbE/β-thal patients have increased Hb-bound platelets, which is associated with increased baseline platelet activation and reactivity.Publication Metadata only High-level induction of fetal haemoglobin by pomalidomide in β-thalassaemia/HbE erythroid progenitor cells(2020-01-01) Pinyaphat Khamphikham; Tiwaporn Nualkaew; Phitchapa Pongpaksupasin; Woratree Kaewsakulthong; Duantida Songdej; Kittiphong Paiboonsukwong; James D. Engel; Suradej Hongeng; Suthat Fucharoen; Orapan Sripichai; Natee Jearawiriyapaisarn; University of Michigan, Ann Arbor; Faculty of Medicine, Ramathibodi Hospital, Mahidol University; Mahidol University; Thammasat University; Faculty of Medicine, Siriraj Hospital, Mahidol University; National Institutes of HealthPublication Metadata only Impact of the detection of ζ-globin chains and hemoglobin Bart's using immunochromatographic strip tests for α0-thalassemia (-SEA) differential diagnosis(2019-01-01) Supansa Pata; Witida Laopajon; Matawee Pongpaiboon; Weeraya Thongkum; Nattapong Polpong; Thongperm Munkongdee; Kittiphong Paiboonsukwong; Suthat Fucharoen; Chatchai Tayapiwatana; Watchara Kasinrerk; Mahidol University; Chiang Mai University© 2019 Pata et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. α0-Thalassemia is an inherited hematological disorder caused by the deletion of α-globin genes. The Southeast Asian deletion (-SEA) is the most common type of α0-thalassemia observed in Southeast Asian countries. Regarding WHO health policy, an effective α0-thalassemia screening strategy is needed to control new severe α-thalassemia cases. In this study, a monoclonal antibody panel was used to develop immunochromatographic (IC) strip tests for detecting the Hb Bart's and ζ-globin chain. Among 195 samples, all α0-thalassemia traits (78 α0-thalassemia (-SEA) and 4 α0-thalassemia (-Thai)) had low MCV or MCH values. The sensitivity, specificity, PPV and NPV of the IC strip tests for ζ-globin and Hb Bart's for screening α0-thalassemia (-SEA) within the low MCV or MCH samples were 100%, 65.2%, 90.7%, 100% and 96.2%, 47.8%, 86.6%, 78.6%, respectively. All 4 α0-thalassemia (-Thai) traits were negative for ζ-globin chains but positive for Hb Bart's using the IC strip tests. These results led to a α0-thalassemia screening being proposed in which blood samples are first evaluated by MCV, MCH and Hb typing. Samples with high MCV and MCH values are excluded for the presence of the α0-thalassemia gene. Samples with low MCV or MCH values are assayed using the developed IC strip tests, where only samples testing positive are further assayed for α0-thalassemia by PCR. Patients with Hb H, EA Bart's or EF Bart's diseases do not need to use this IC strip assay. Thus, in this study, a simple and cost effective α0-thalassemia point of care test was developed.Publication Metadata only Increased ferritin levels in non-transfusion-dependent β°-thalassaemia/HbE are associated with reduced CXCR2 expression and neutrophil migration(2019-01-01) Chayada Thiengtavor; Sirikwan Siriworadetkun; Kittiphong Paiboonsukwong; Suthat Fucharoen; Kovit Pattanapanyasat; Jim Vadolas; Saovaros Svasti; Pornthip Chaichompoo; Ramkhamhaeng University; Monash University; Mahidol University; Faculty of Medicine, Siriraj Hospital, Mahidol University; Hudson Institute of Medical Research© 2019 British Society for Haematology and John Wiley & Sons Ltd Severe bacterial infection is a major complication causing morbidity and mortality in β-thalassaemia/HbE patients. Innate immunity constitutes the first line of defence against bacterial infection. This study aimed to comprehensively investigate the innate immune phenotype and function related to factors predisposing to infection in non-transfusion-dependent (NTD) β°-thalassaemia/HbE patients. Twenty-six patients and 17 healthy subjects were recruited to determine complement activity (C3, C4, mannose-binding lectin and CH50) and surface receptor expression including markers of phagocytosis (CD11b, CD16 and C3bR), inflammation (C5aR) and migration (CD11b, CXCR1 and CXCR2) on neutrophils and monocytes. In addition, phagocytosis and oxidative burst activity of neutrophils and monocytes against Escherichia coli and neutrophil migration were examined. Decreased C3 and surface expression of CD11b and C3bR on neutrophils were found in patients. However, phagocytosis of neutrophils in patients was still in the normal range. Interestingly, patients displayed a significant reduction of surface expression of CXCR2 [1705 ± 217 mean fluorescent intensity (MFI)] on neutrophils, leading to impaired neutrophil migration (9·2 ± 7·7%) when compared to neutrophils from healthy subjects (2261 ± 627 MFI and 27·8 ± 9% respectively). Moreover, surface expression of CXCR2 on neutrophils was associated with splenectomy status, serum ferritin and haemoglobin levels. Therefore, impaired neutrophil migration could contribute to the increased susceptibility to infection seen in NTD β°-thalassaemia/HbE patients.Publication Metadata only Inhaled nebulized sodium nitrite decreases pulmonary artery pressure in β-thalassemia patients with pulmonary hypertension(2018-06-01) Teerapat Yingchoncharoen; Teewin Rakyhao; Suporn Chuncharunee; Piyamitr Sritara; Pavit Pienvichit; Kittiphong Paiboonsukwong; Piyadon Sathavorasmith; Kanjana Sirirat; Thanaporn Sriwantana; Sirada Srihirun; Nathawut Sibmooh; Faculty of Medicine, Ramathibodi Hospital, Mahidol University; Mahidol University© 2017 Elsevier Inc. Pulmonary hypertension is a life-threatening complication in β-thalassemia. Inhaled sodium nitrite has vasodilatory effect on pulmonary vasculature. However, its effect on pulmonary artery pressure (PAP) in β-thalassemia subjects with pulmonary hypertension has never been reported. In this study, we investigated the change in PAP during inhalation of sodium nitrite in 5 β-thalassemia patients. We demonstrated that sodium nitrite administered by nebulization rapidly decreased PAP as measured by echocardiography and right heart catheterization. The effect of nitrite was short as PAP returned to baseline at end of inhalation. Our findings support acute pulmonary vasodilation effect of nitrite in β-thalassemia with pulmonary hypertension.Publication Metadata only Microparticles from β-thalassaemia/HbE patients induce endothelial cell dysfunction(2018-12-01) Wasinee Kheansaard; Kunwadee Phongpao; Kittiphong Paiboonsukwong; Kovit Pattanapanyasat; Pornthip Chaichompoo; Saovaros Svasti; Mahidol University; Faculty of Medicine, Siriraj Hospital, Mahidol University© 2018, The Author(s). Thromboembolic complication occurs frequently in β-thalassaemia/HbE patients, particularly in splenectomised patients. Endothelial cells play an important role in thrombosis. There is strong evidence of endothelial cell activation and dysfunction in β-thalassaemia. Microparticles (MPs) are associated with thrombosis and endothelial cell dysfunction in many diseases including β-thalassaemia. However, the effect of thalassaemic-MPs on endothelial cells mediating thrombus formation has not been elucidated. In this study, the effects of circulating MPs from β-thalassaemia/HbE patients on endothelial cell functions were investigated. The results showed that MPs directly induce tissue factor, interleukin (IL)-6, IL-8, intracellular adhesion molecule-1, vascular cell adhesion molecule-1 and E-selectin expression in human umbilical vein endothelial cells (HUVECs). Notably, the levels of these endothelial cell activation markers were significantly increased in HUVECs treated with MPs obtained from splenectomised β-thalassaemia/HbE patients when compared to MPs from non-splenectomised patients or normal subjects. The increased endothelial cell activation ultimately lead to increased monocyte-endothelial cell adhesion. THP-1 and HUVECs adhesion induced by MPs from normal subjects, non-splenectomised and splenectomised patients increased to 2.0 ± 0.4, 2.3 ± 0.4 and 3.8 ± 0.4 fold, respectively when compared to untreated cells. This finding suggests that MPs play an important role on thrombosis and vascular dysfunction in β-thalassaemia/HbE disease, especially in splenectomised cases.Publication Metadata only miR-144 regulates oxidative stress tolerance of thalassemic erythroid cell via targeting NRF2(2019-09-01) Kanitta Srinoun; Nuankanya Sathirapongsasuti; Kittiphong Paiboonsukwong; Somporn Sretrirutchai; Malai Wongchanchailert; Suthat Fucharoen; Faculty of Medicine, Prince of Songkia University; Faculty of Medicine, Ramathibodi Hospital, Mahidol University; Mahidol University; Prince of Songkla University© 2019, Springer-Verlag GmbH Germany, part of Springer Nature. Thalassemia has a high prevalence in Thailand. Oxidative damage to erythroid cells is known to be one of the major etiologies in thalassemia pathophysiology. Oxidative stress status of thalassemia is potentiated by the heme, nonheme iron, and free iron resulting from imbalanced globin synthesis. In addition, levels of antioxidant proteins are reduced in α-thalassemia and β-thalassemia erythrocytes. However, the primary molecular mechanism for this phenotype remains unknown. Our study showed a high expression of miR-144 in β- and α-thalassemia. An increased miR-144 expression leads to decreased expression of nuclear factor erythroid 2-related factor 2 (NRF2) target, especially in α-thalassemia. In α-thalassemia, miR-144 and NRF2 target are associated with glutathione level and anemia severity. To study the effect of miR-144 expression, the gain-loss of miR-144 expression was performed by miR inhibitor and mimic transfection in the erythroblastic cell line. This study reveals that miR-144 expression was upregulated, whereas NRF2 expression and glutathione levels were decreased in comparison with the untreated condition after miR mimic transfection, while the reduction of miR-144 expression contributed to the increased NRF2 expression and glutathione level compared with the untreated condition after miR inhibitor transfection. Moreover, miR-144 overexpression leads to significantly increased sensitivity to oxidative stress at indicated concentrations of hydrogen peroxide (H2O2) and rescued by miR-144 inhibitor. Taken together, our findings suggest that dysregulation of miR-144 may play a role in the reduced ability of erythrocyte to deal with oxidative stress and increased RBC hemolysis susceptibility especially in thalassemia.Publication Metadata only Pharmacokinetics and pharmacodynamics of single dose of inhaled nebulized sodium nitrite in healthy and hemoglobin E/β-thalassemia subjects(2019-12-01) Kanjana Sirirat; Thanaporn Sriwantana; Jirada Kaewchuchuen; Kittiphong Paiboonsukwong; Suthat Fucharoen; Garnpimol Ritthidej; Tipparat Parakaw; Sirada Srihirun; Pornpun Vivithanaporn; Piyamitr Sritara; Nathawut Sibmooh; Chulalongkorn University; Faculty of Medicine, Ramathibodi Hospital, Mahidol University; Mahidol University© 2019 Elsevier Inc. Inhaled sodium nitrite has been reported to decrease pulmonary artery pressure in hemoglobin E/β-thalassemia (HbE/β-thal) patients with pulmonary hypertension. This study investigated the pharmacokinetics and pharmacodynamics of inhaled nebulized sodium nitrite in 10 healthy subjects and 8 HbE/β-thal patients with high estimated pulmonary artery pressure. Nitrite pharmacokinetics, fraction exhaled nitric oxide (FENO), estimated right ventricular systolic pressure (eRVSP) measured by echocardiography, and platelet activation were determined. Nebulized sodium nitrite at doses used in this study (37.5 and 75 mg for healthy subjects and 15 mg for HbE/β-thal patients) was well tolerated and did not cause changes in methemoglobin levels and systemic blood pressure. Absorption of inhaled nitrite was rapid with the absolute bioavailability of 18%. In whole blood, nitrite exhibited the dose-independent pharmacokinetics with clearance (CL) of 1.5 l/h/kg, volume of distribution (Vd) of 1.3 l/kg and half-life (t1/2) of 0.6 h. CL and Vd of nitrite was higher in red blood cells (RBC) than whole blood and plasma. HbE/β-thal patients had lower nitrite CL and longer t1/2 in RBC than healthy subjects. FENO increased immediately after inhalation. Following nitrite inhalation, eRVSP remained unchanged but platelet activation was suppressed as evidenced by inhibition of adenosine diphosphate (ADP)-induced P-selectin expression and increase in phosphorylated vasodilator-stimulated phosphoprotein (P-VASPSer239) in platelets. There were no changes in markers of oxidative and nitrosative stress after inhalation. Our results support further development of inhaled nebulized sodium nitrite for treatment of pulmonary hypertension in β-thalassemia.Item Metadata only Regulation of erythropoiesis of normal and thalassemia progenitor cells from peripheral blood(Mahidol University. Mahidol University Library and Knowledge Center, 2023) Kittiphong Paiboonsukwong; Suthat FucharoenPublication Metadata only UNC0638 induces high levels of fetal hemoglobin expression in β-thalassemia/HbE erythroid progenitor cells(2020-09-01) Tiwaporn Nualkaew; Pinyaphat Khamphikham; Phitchapa Pongpaksupasin; Woratree Kaewsakulthong; Duantida Songdej; Kittiphong Paiboonsukwong; Orapan Sripichai; James Douglas Engel; Suradej Hongeng; Suthat Fucharoen; Natee Jearawiriyapaisarn; University of Michigan, Ann Arbor; Faculty of Medicine, Ramathibodi Hospital, Mahidol University; Mahidol University; Thammasat University; Faculty of Medicine, Siriraj Hospital, Mahidol University; National Institutes of Health (NIH)© 2020, Springer-Verlag GmbH Germany, part of Springer Nature. Increased expression of fetal hemoglobin (HbF) improves the clinical severity of β-thalassemia patients. EHMT1/2 histone methyltransferases are epigenetic modifying enzymes that are responsible for catalyzing addition of the repressive histone mark H3K9me2 at silenced genes, including the γ-globin genes. UNC0638, a chemical inhibitor of EHMT1/2, has been shown to induce HbF expression in human erythroid progenitor cell cultures. Here, we report the HbF-inducing activity of UNC0638 in erythroid progenitor cells from β-thalassemia/HbE patients. UNC0638 treatment led to significant increases in γ-globin mRNA, HbF expression, and HbF-containing cells in the absence of significant cytotoxicity. Moreover, UNC0638 showed additive effects on HbF induction in combination with the immunomodulatory drug pomalidomide and the DNMT1 inhibitor decitabine. These studies provide a scientific proof of concept that a small molecule targeting EHMT1/2 epigenetic enzymes, used alone or in combination with pomalidomide or decitabine, is a potential therapeutic approach for HbF induction. Further development of structural analogs of UNC0638 with similar biological effects but improved pharmacokinetic properties may lead to promising therapies and possible clinical application for the treatment of β-thalassemia.Publication Metadata only Update in Laboratory Diagnosis of Thalassemia(2020-05-27) Thongperm Munkongdee; Ping Chen; Pranee Winichagoon; Suthat Fucharoen; Kittiphong Paiboonsukwong; Mahidol University; Guangxi Medical University© Copyright © 2020 Munkongdee, Chen, Winichagoon, Fucharoen and Paiboonsukwong. Alpha- and β-thalassemias and abnormal hemoglobin (Hb) are common in tropical countries. These abnormal globin genes in different combinations lead to many thalassemic diseases including three severe thalassemia diseases, i.e., homozygous β-thalassemia, β-thalassemia/Hb E, and Hb Bart’s hydrops fetalis. Laboratory diagnosis of thalassemia requires a number of tests including red blood cell indices and Hb and DNA analyses. Thalassemic red blood cell analysis with an automated hematology analyzer is a primary screening for thalassemia since microcytosis and decreased Hb content of red blood cells are hallmarks of all thalassemic red blood cells. However, these two red blood cell indices cannot discriminate between thalassemia trait and iron deficiency or between α- and β-thalassemic conditions. Today, Hb analysis may be carried out by either automatic high-performance liquid chromatography (HPLC) or capillary zone electrophoresis (CE) system. These two systems give both qualitative and quantitative analysis of Hb components and help to do thalassemia prenatal and postnatal diagnoses within a short period. Both systems have a good correlation, but the interpretation under the CE system should be done with caution because Hb A2 is clearly separated from Hb E. In case of α-thalassemia gene interaction, it can affect the amount of Hb A2/E. Thalassemia genotypes can be characterized by the intensities between alpha-/beta-globin chains or alpha-/beta-mRNA ratios. However, those are presumptive diagnoses. Only DNA analysis can be made for specific thalassemia mutation diagnosis. Various molecular techniques have been used for point mutation detection in β-thalassemia and large-deletion detection in α-thalassemia. All of these techniques have some advantages and disadvantages. Recently, screening for both α- and β-thalassemia genes by next-generation sequencing (NGS) has been introduced. This technique gives an accurate diagnosis of thalassemia that may be misdiagnosed by other conventional techniques. The major limitation for using NGS in the screening of thalassemia is its cost which is still expensive. All service labs highly recommend to select the technique(s) they are most familiar and most economic one for their routine use.