Table 1 shows the demographic data of the sixteen RA patients and a healthy individual and their blood test results of commonly used RA diagnostic markers indicating their disease activities.
Based on the DASCRP index of 16 RA patients, 3 of these patients were classified as high disease activity, 8 as moderate activity, 3 as remission, and the remaining two as unknown status due to incomplete data collection.
Out of the seventeen flasks, 15 EBV-transformed LCLs except 4E and 14A were generated one week after infection in which visible cell clusters were seen from the flasks. When visualized under microscope, the LCLs showed typical rosette morphology with various sizes as indicated by the red arrows Figure 1. The success rate of the LCLs generation was We then determined the growth rate of each generated category of LCLs using a nonlytic and real-time luminescence-based cell proliferation assay.
Table 2 shows the calculated doubling time and growth rate of respective LCLs using an online calculator [ 15 ]. The expression level was then presented in percentage from the total number of analysed lymphocytes which were 20, cells.
Following the EBV transformation, T-cell populations reduced to This clearly indicates the success of B-cell immortalization by EBV which has taken over the T-cell population. In RA patients, elevated levels of CDexpressing B-cells have been previously reported to positively correlate with the disease activity, and they could potentially serve as a therapeutic target [ 10 , 18 ].
From all LCLs, while B-cells coexpressing CD23 and CD27 were 5. A statistical analysis was performed to test the correlation between the disease activity and surface marker expressions; however, none of them showed significance. Similarly, the disease activity did not correlate with the growth rate of corresponding LCLs. From the doubling time of each category of LCLs, mean values for both fast- and slow-growing LCLs were determined and subjected to statistical analysis.
However, this is not statistically significant Table 4. Further studies with larger sample size are required to validate these findings. There is evidence showing that B-cell immortalization might rely on the cellular CD23 expression. The study showed that CDnegative B-cells could be infected by EBV but could not be immortalized unless supplementing with specific growth factors [ 19 ]. The former B-cell subpopulation actively proliferated while the latter subpopulation ceased to proliferate.
Further studies with a larger sample size are warranted to investigate in depth on the implication of the surface marker expression on the EBV maintenance and cellular growth. The authors declare that there are no conflicts of interest regarding the publication of this article. We would like to thank Dr. Supplementary Materials. This is an open access article distributed under the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Article of the Year Award: Outstanding research contributions of , as selected by our Chief Editors. Read the winning articles. Journal overview. Special Issues. Academic Editor: Finn S. Received 30 Nov Revised 20 Feb Under normal conditions, the HIF1A protein level is low because of proteasome-dependent degradation. At high oxygen levels, a few prolines e. Our findings show that HIF1A is stabilized in EBV-transformed B-cells and, given that mRNA levels are not changed, it seems likely that the stabilization is due to the inhibition of hydroxylation, as we have demonstrated here.
PDK1 phosphorylates the mitochondrial pyruvate dehydrogenase complex responsible for oxidative decarboxylation of pyruvate. The level of GLUT1, a glucose transporter responsible for glucose uptake, is high in thyroid [19] and colorectal cancers [20].
PGK1, a phosphoglycerate kinase, is secreted by fibrosarcoma cells at levels several-fold higher than for normal fibroblasts [21]. Hexokinase HK phosphorylates glucose into glucosephosphate [22] , which is the first step of the glycolytic pathway. At high oxygen concentration, glucose is metabolized to CO 2 and the cells produce ATP about 36 molecules per molecule of glucose and precursors for amino acid synthesis, whereas only minimal amounts of lactate are generated from pyruvate reviewed in [23].
At low oxygen concentrations, the cells convert most of the pyruvate to lactate and much less ATP is generated 2—4 molecules per 1 molecule of glucose , in a process referred to as anaerobic glycolysis.
However, the rate of ATP production is higher and overall metabolism is more efficient during glycolysis reviewed in [24]. It has been shown that tumor cells and rapidly proliferating mitogen-activated T-cells reviewed in [10] , [25] prefer glycolysis even under normoxic conditions; this phenomenon is usually referred to as the Warburg effect. As a result, the cells can produce more NADH reduced form of the nicotinamide adenine dinucleotide , intermediates for the synthesis of fatty acids, nonessential amino acids and sugars for nucleotides ; this is believed to facilitate cellular protein synthesis.
Thus, only 1. Consistent with the high rate of anaerobic glycolysis in rapidly proliferating cells, the activities of hexokinase, 6-phosphofructokinase PFK and PKLR were increased 10—fold relative to their resting state [25]. Many enzymes involved in glycolysis are the products of HIF1A-responsive genes. Our findings indicate that under normoxic conditions, aerobic glycolysis is active in LCLs and in freshly EBV-infected B-cells, but not in mitogen-activated B-cells.
It has been shown that activation of AKT signaling also results in the induction of aerobic glycolysis [24] , [26] , [27] , and it was proposed that the AKT pathway could be activated by LMP1 upon CD40 mimicking, discussed in [28]. Recently, it was shown that the AKT pathway is also activated in EBV-transformed B-cells where it counteracts apoptosis and favors cell survival [29].
LCLs produce much higher levels of ROS than mitogen-activated B-cells without any apparent detrimental effects on the cells. Importantly, our findings indicate that EBV-transformed lymphoblastoid cells that have never been exposed to hypoxia exhibit the Warburg effect, a phenomenon that is characteristic of tumor cells.
The transformed cells use both the aerobic and nonaerobic glycolytic pathways for energy production, even in the presence of abundant oxygen Figure 7. This results in conversion of pyruvate to lactate, i. Browse Subject Areas? Click through the PLOS taxonomy to find articles in your field. Abstract Background Epstein-Barr virus EBV encodes six nuclear transformation-associated proteins that induce extensive changes in cellular gene expression and signaling and induce B-cell transformation.
Conclusions Our data suggest that activation of the aerobic glycolytic pathway, corresponding to the Warburg effect, occurs in EBV-transformed lymphoblastoid cells, in contrast to mitogen-activated B-cells. Introduction Epstein-Barr virus EBV is a ubiquitous human gamma herpesvirus that causes lymphoproliferative disease in immunosuppressed patients and is associated with Burkitt's lymphoma, Hodgkin's lymphoma, other B- and T-cell lymphomas, nasopharyngeal carcinoma NPC and some gastric carcinomas [1].
Immunostaining and imaging Immunostaining and digital image capture were performed as described elsewhere. Download: PPT. Figure 1. Science and Engineering. Health Sciences. Description Epstein-Barr virus EBV belongs to the herpesvirus family and it is the main agent that cause human mononucleosis.
How does it work. Establishment of cell lines from normal adult human blood leukocytes by exposure to Epstein-Barr virus and neutralization by human sera with Epstein-Barr virus antibody. Proc Soc Exp Biol Med. The oncogenicity of Epstein-Barr virus. J Infect Dis. Epstein-Barr virus: transformation of lymphocytes separated by size or exposed to bromodeoxyuridine and light.
Epstein-Barr virus DNA is amplified in transformed lymphocytes. J Virol. J Mol Biol. Two small RNAs encoded by Epstein-Barr virus and complexed with protein are precipitated by antibodies from patients with systemic lupus erythematosus. Sci Am.
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