Browsing by Author "Rachanee Udomsangpetch"

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    Characterization of inhibitory anti-duffy binding protein II immunity: Approach to plasmodium vivax vaccine development in Thailand
    (2012) Patchanee Chootong; Tasanee Panichakul; Chongrak Permmongkol; Samantha J. Barnes; Rachanee Udomsangpetch; John H. Adams; P. Chootong; Department of Clinical Microbiology and Applied Technology, Faculty of Medical Technology, Mahidol University, Bangkok, Thailand; email: pchooton@gmail.com
    Plasmodium vivax Duffy binding protein region II (DBPII) is an important vaccine candidate for antibody-mediated immunity against vivax malaria. A significant challenge for vaccine development of DBPII is its highly polymorphic nature that alters sensitivity to neutralizing antibody responses. Here, we aim to characterize naturally-acquired neutralizing antibodies against DBPII in individual Thai residents to give insight into P. vivax vaccine development in Thailand. Anti-DBPII IgG significantly increased in acute vivax infections compared to uninfected residents and naive controls. Antibody titers and functional anti-DBPII inhibition varied widely and there was no association between titer and inhibition activity. Most high titer plasmas had only a moderate to no functional inhibitory effect on DBP binding to erythrocytes, indicating the protective immunity against DBPII binding is strain specific. Only 5 of 54 samples were highly inhibitory against DBP erythrocyte-binding function. Previously identified target epitopes of inhibitory anti-DBPPII IgG (H1, H2 and H3) were localized to the dimer interface that forms the DARC binding pocket. Amino acid polymorphisms (monomorphic or dimorphic) in H1 and H3 protective epitopes change sensitivity of immune inhibition by alteration of neutralizing antibody recognition. The present study indicates Thai variant H1.T1 (R308S), H3.T1 (D384G) and H3.T3 (K386N) are the most important variants for a DBPII candidate vaccine needed to protect P. vivax in Thai residents. © 2012 Chootong et al.
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    In vitro production of functional immune cells derived from human hematopoietic stem cells
    (Leibniz Research Centre for Working Environment and Human Factors, 2015) Witchuda Payuhakrit; Tasanee Panichakul; Natthawut Charoenphon; Panus Chalermsaenyakorn; Adithep Jaovisidha; Chokdee Wongborisuth; Rachanee Udomsangpetch
    Hematopoietic stem cells (HSC) from cord blood are potentially high sources for transplantation due to their low immunogenicity and the presence of the multipotent cells. These cells are capable of differentiating to produce various lineages of blood cells under specific conditions. We have enriched highly purified CD34+ cells from cord blood, determined in vitro growth of the cells in culture systems in the absence (condition A) or presence of GM-CSF and G-CSF (condition B), and determined the profile of immune cells during the period of cultivation by using flow cytometry. PhytohemagglutininA (PHA) was used as a mitogen to stimulate T lymphocytes derived from hematopoietic stem cells. GM-CSF and G-CSF prolonged the survival of the growing cells and also maintained expansion of cells in blastic stage. By day 12 of cultivation, when cell numbers peaked, various types of immune cells had appeared (CD14+ cells, CD40+HLA-DR+ cells, CD3+CD56+ cells, CD19+ cells, CD3+CD4+ cells, CD3+CD8+cells and CD3-CD56+). A significantly higher percentage of monocytes (p = 0.002) were observed under culture with GM-CSF, G-CSF when compared with culture without GM-CSF, G-CSF. In addition, T lymphocytes derived from HSC responded to 50 _g/ml of PHA. This is the first report showing the complete differentiation and proliferation of immune cells derived from CD34+ HSC under in vitro culture conditions. Lymphocytes, monocytes, dendritic cells and polymorph nuclear cells derived from HSC in vitro are unique, and thus may benefit various studies such as innate immunity and pathophysiology of immune disorders. � 2015, Leibniz Research Centre for Working Environment and Human Factors. All right reserved.
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    Plasmodium vivax inhibits erythroid cell growth through altered phosphorylation of the cytoskeletal protein ezrin
    (BioMed Central Ltd., 2015) Tasanee Panichakul; Saranyoo Ponnikorn; Sittiruk Roytrakul; Atchara Paemanee; Suthathip Kittisenachai; Suradej Hongeng; Rachanee Udomsangpetch
    Background: The underlying causes of severe malarial anaemia are multifactorial. In previously reports, Plasmodium vivax was found to be able to directly inhibited erythroid cell proliferation and differentiation. The molecular mechanisms underlying the suppression of erythropoiesis by P. vivax are remarkably complex and remain unclear. In this study, a phosphoproteomic approach was performed to dissect the molecular mechanism of phosphoprotein regulation, which is involved in the inhibitory effect of parasites on erythroid cell development. Methods: This study describes the first comparative phosphoproteome analysis of growing erythroid cells (gECs), derived from human haematopoietic stem cells, exposed to lysates of infected erythrocytes (IE)/uninfected erythrocytes (UE) for 24, 48 and 72 h. This study utilized IMAC phosphoprotein isolation directly coupled with LC MS/MS analysis. Results: Lysed IE significantly inhibited gEC growth at 48 and 72 h and cell division resulting in the accumulation of cells in G0 phase. The relative levels of forty four phosphoproteins were determined from gECs exposed to IE/UE for 24-72 h and compared with the media control using the label-free quantitation technique. Interestingly, the levels of three phosphoproteins: ezrin, alpha actinin-1, and Rho kinase were significantly (p_<_0.05) altered. These proteins display interactions and are involved in the regulation of the cellular cytoskeleton. Particularly affected was ezrin (phosphorylated at Thr567), which is normally localized to gEC cell extension peripheral processes. Following exposure to IE, for 48-72 h, the ezrin signal intensity was weak or absent. This result suggests that phospho-ezrin is important for actin cytoskeleton regulation during erythroid cell growth and division. Conclusions: These findings suggest that parasite proteins are able to inhibit erythroid cell growth by down-regulation of ezrin phosphorylation, leading to ineffective erythropoiesis ultimately resulting in severe malarial anaemia. A better understanding of the mechanisms of ineffective erythropoiesis may be beneficial in the development of therapeutic strategies to prevent severe malarial anaemia. � 2015 Panichakul et al.; licensee BioMed Central.
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    Suppression of erythroid development in vitro by Plasmodium vivax
    (2012) Tasanee Panichakul; Witchuda Payuhakrit; Panyu Panburana; Chokdee Wongborisuth; Suradej Hongeng; Rachanee Udomsangpetch; T. Panichakul; Faculty of Science and Technology, Suan Dusit Rajabhat University, Bangplat, Bangkok, 10700, 204/3 Sirindhorn Rd., Thailand; email: tasanee_p@yahoo.com
    Background: Severe anaemia due to dyserythropoiesis has been documented in patients infected with Plasmodium vivax, however the mechanism responsible for anaemia in vivax malaria is poorly understood. In order to better understand the role of P. vivax infection in anaemia the inhibition of erythropoiesis using haematopoietic stem cells was investigated. Methods: Haematopoietic stem cells/CD34 + cells, isolated from normal human cord blood were used to generate growing erythroid cells. Exposure of CD34 + cells and growing erythroid cells to P. vivax parasites either from intact or lysed infected erythrocytes (IE) was examined for the effect on inhibition of cell development compared with untreated controls. Results: Both lysed and intact infected erythrocytes significantly inhibited erythroid growth. The reduction of erythroid growth did not differ significantly between exposure to intact and lysed IE and the mean growth relative to unexposed controls was 59.4 ± 5.2 for lysed IE and 57 ± 8.5% for intact IE. Interestingly, CD34 + cells/erythroid progenitor cells were susceptible to the inhibitory effect of P. vivax on cell expansion. Exposure to P. vivax also inhibited erythroid development, as determined by the reduced expression of glycophorin A (28.1%) and CD 71 (43.9%). Moreover, vivax parasites perturbed the division of erythroid cells, as measured by the Cytokinesis Block Proliferation Index, which was reduced to 1.35 ± 0.05 (P-value²0.01) from a value of 2.08 ± 0.07 in controls. Neither TNF-a nor IFN-g was detected in the culture medium of erythroid cells treated with P. vivax, indicating that impaired erythropoiesis was independent of these cytokines. Conclusions: This study shows for the first time that P. vivax parasites inhibit erythroid development leading to ineffective erythropoiesis and highlights the potential of P. vivax to cause severe anaemia. © 2012 Panichakul et al.; licensee BioMed Central Ltd.