Publications by authors named "Vahid Younesi"

33 Publications

Prevalence of Anti-SARS-CoV-2 Specific Antibodies in Health-Care Workers Compared to General Population during an Early Phase of the Pandemic, Tehran-Iran.

Iran J Immunol 2021 03;18(1):82-92

Department of Immunology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran,Iran.

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) rapidly transmits in general population, mainly between health-care workers (HCWs) who are in close contact with patients.

Objective: To study the seropositivity of HCWs as a high-risk group compared to general population.

Methods: 72 samples were obtained from HCWs working in Masih Daneshvari hospital as one of the main COVID-19 admission centers in Tehran, during April 4 to 6, 2020. Also we collected 2021 blood samples from general population. The SARS-CoV-2 specific IgM, and IgG antibodies in the collected serum specimens were measured by commercial ELISA kits.

Results: Based on the clinical manifestations, 25.0%, 47.2%, and 27.8% of HCWs were categorized as symptomatic with typical symptoms, symptomatic with atypical symptoms, and asymptomatic, respectively. Symptomatic individuals with typical and atypical symptoms were 63.2% and 36.8% positive in RT-PCR test, respectively. Anti-SARS-CoV-2 IgM and IgG antibodies were detected in 15.3% and 27.8% of HCWs samples, respectively. Antibody testing in the general population indicated that SARS-CoV-2 specific IgM and IgG were found in (162/2021) 8%, and (290/2021) 14.4%, respectively. The frequency of positive cases of IgM and IgG were significantly increased in HCWs compared to general population (p= 0.028 for IgM and p= 0.002 for IgG).

Conclusion: The frequency of SARS-CoV-2 specific antibodies in HCWs was higher than general population indicating a higher viral transmission via close exposure with COVID-19 patients.
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http://dx.doi.org/10.22034/iji.2021.88168.1851DOI Listing
March 2021

Nanocurcumin improves regulatory T-cell frequency and function in patients with multiple sclerosis.

J Neuroimmunol 2019 02 17;327:15-21. Epub 2019 Jan 17.

Drug Applied Research Center, Tabriz University of Medical Sciences, Tabriz, Iran; Department of Immunology, School of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran. Electronic address:

Background: Multiple sclerosis is a chronic incapacitating disease of the central nervous system, it has been reported that the disturbance in the development and function of Treg subpopulations is associated with the disability status in the RRMS. Accordingly, in the current study, the objective was to specify nanocurcumin effects on Treg cells frequency, and function in patients with RRMS.

Methods And Materials: 50 patients with RRMS were enrolled in this study in which 25 were treated for at least six months with nanocurcumin capsules while the other half received placebo capsules as the control group. The blood sample was collected prior to the administration of nanocurcumin and placebo capsules and following six months. At baseline and after a six-month treatment, the frequency of Treg lymphocytes, the expression of transcription factor related to these cells and the secretion levels of cytokines were assessed by flowcytometry, real-time PCR and ELISA, respectively.

Results: A significant reduction was observed in the proportion of peripheral Treg cell frequency, and the levels of TGF-β, IL-10 and FoxP3 expression in patients with RRMS. Our data revealed that the frequency of Treg cells (p = .0027), the expression of FoxP3 (p = .0005), TGF-β (p = .0005), and IL-10 (p = .0002) and the secretion levels of the TGF-β (p = .033), and IL-10 (p = .029) in cultured PBMCs are increased in nanocurcumin-treated group compared to placebo group.

Conclusion: The results of the current work indicated that nanocurcumin is capable of restoring the frequency and function of Treg cells in MS patients.
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http://dx.doi.org/10.1016/j.jneuroim.2019.01.007DOI Listing
February 2019

Chitosan (CMD)-mediated co-delivery of SN38 and Snail-specific siRNA as a useful anticancer approach against prostate cancer.

Pharmacol Rep 2018 Jun 20;70(3):418-425. Epub 2017 Nov 20.

Drug Applied Research Center, Tabriz University of Medical Sciences, Tabriz, Iran; Department of Immunology, Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran. Electronic address:

Background: Prostate cancer is known as the most common malignancy in men. Chitosan has generated great interest as a useful biopolymer for the encapsulation of small interfering RNA (siRNA). Due to cationic nature, chitosan is able to efficiently encapsulate siRNA molecules and form nanoparticles. Furthermore, the biocompatible and biodegradable attributes of chitosan have paved the way for its potential application in the in vivo delivery of therapeutic siRNAs. In this study, we aimed to design chitosan/CMD nanoparticles for the efficient encapsulation of the anti-cancer drugs SN38 and Snail-specific siRNA.

Methods: Physicochemical characteristics, growth inhibitory properties, and anti-migratory capacities of the dual delivery of SN38-Snail siRNA CMD-chitosan nanoparticles were investigated in prostate cancer cells.

Results: Our findings provided evidence for the suggestion that, ChNP-CMD-SN38-siRNA treated cells, the mRNA level of snail decreased from 1.00 to 0.30 (±0.14) and 0.09 (±0.04) after 24h and 48h, respectively. Additionally, the fold induction of E-cadherin and Claudin-1 increased from 1.00 to now 3.12 (±0.62), 3.02 (±0.28) after 24h and 5.6 (±0.91), 4.42 (±0.51) after 48h, respectively. Also, co-delivery of SN38 and Snail-specific siRNA by an appropriate nanocerrier (chitosan nanoparticles) could reduce the viability, proliferation, and migration of PC-3 cells.

Conclusions: In conclusion, ChNPs encapsulating SN38 and Snail-specific siRNA may represent huge potential as an effective anti-cancer drug delivery system for the treatment of prostate cancer.
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http://dx.doi.org/10.1016/j.pharep.2017.11.005DOI Listing
June 2018

Correction to: Peripheral Th17/Treg imbalance in elderly patients with ischemic stroke.

Neurol Sci 2018 Apr;39(4):655

Drug Applied Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.

In the original article the terms RORγt and GF-β were misspelled throughout the text. They should read RORγt and TGF-β instead. We apologize for the inconvenience. The original article has been corrected.
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http://dx.doi.org/10.1007/s10072-018-3265-xDOI Listing
April 2018

Peripheral Th17/Treg imbalance in elderly patients with ischemic stroke.

Neurol Sci 2018 Apr 20;39(4):647-654. Epub 2018 Jan 20.

Drug Applied Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.

CD4CD25 regulatory T (Treg) cells and Th17 cells play important roles in peripheral immunity. Immune responses are main elements in the pathogenesis of ischemic stroke (IS). The contribution of Th17 cells in IS patients has not been proved, and whether the balance of Treg/Th17 cells is changed in IS patients remains unidentified. In the present study, we studied Th17 and Treg cell frequency, cytokine secretion, expression of transcription factors, and microRNAs related to Th17 and Treg cells differentiation, which is compared between IS patients and control group. Thirty patients with IS and 30 individuals as control group were enrolled in this study. The frequency of Th17 and Treg lymphocytes, the expression of transcription factors and microRNAs related to these cells, and the serum levels of associated cytokines were assessed by flow cytometry, real-time PCR, and ELISA, respectively. A significant reduction in proportion of peripheral Treg cell frequency and the levels of TGF-β and FOXP3 expression were observed in patients with IS compared with controls, while the proportions of Th17 were increased dramatically, and these effects were along with increases in the levels of IL-17A and RORγt expression in IS patients. The levels of mir-326 and mir-106b-25 expression were increased in patients with IS. These studies suggest that the increase in proportion of Th17 cells and decrease in Treg cells might contribute to the pathogenesis of IS. Manipulating the balance between Tregs and Th17 cells might be helpful for the treatment of IS.
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http://dx.doi.org/10.1007/s10072-018-3250-4DOI Listing
April 2018

Nanocurcumin restores aberrant miRNA expression profile in multiple sclerosis, randomized, double-blind, placebo-controlled trial.

J Cell Physiol 2018 07 19;233(7):5222-5230. Epub 2018 Jan 19.

Drug Applied Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.

In the current study, we aimed to identify nanocurcumin effects on microRNAs (miRNAs) in the peripheral blood of patients with relapsing-remitting multiple sclerosis (RRMS). We intended to investigate the expression pattern of these miRNAs in experimental settings in vivo. The expression levels of the selected 27 miRNAs known to be involved in the regulation of immune responses were analyzed in 50 RRMS patients and 35 healthy controls. The miRNA expression profiles were investigated by quantitative PCR (qPCR) at baseline and after 6 months of nanocurcumin therapy. Our data revealed that the expression of a number of microRNAs including miR-16, miR-17-92, miR-27, miR-29b, miR-126, miR-128, miR-132, miR-155, miR-326, miR-550, miR-15a, miR-19b, miR-106b, miR-320a, miR-363, miR-31, miR-150, and miR-340 is regulated by nanocurcumin. The results of the current work indicate that nanocurcumin is able to restore the expression pattern of dysregulated miRNAs in MS patients. We discovered that some miRNAs are deregulated in untreated patients compared with healthy controls and nanocurcumin-treated patients. This is a new finding that might represent the potential contribution of these miRNAs to MS pathogenesis. Taken together, these data provide novel insights into miRNA-dependent regulation of the function of B and T cells in MS disease and enrich our understanding of the effects mediated by a therapeutic approach that targets B and T cells.
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http://dx.doi.org/10.1002/jcp.26301DOI Listing
July 2018

Regulatory T cells improve pregnancy rate in RIF patients after additional IVIG treatment.

Syst Biol Reprod Med 2017 Dec 3;63(6):350-359. Epub 2017 Nov 3.

b Drug Applied Research Center , Tabriz University of Medical Sciences , Tabriz , Iran.

RIF (repeated implantation failure) women with immunologic basis and cellular abnormalities may benefit from intravenous immunoglobulins (IVIG) as an immunomodulator based on different studies. In this study, we evaluated the effect of IVIG on the frequency and function of Th17 and Treg cells, as two important subgroups of CD T cells in implantation and pregnancy rates. Seventy-two RIF patients with preconception Th1⁄Th2 ratio and natural killer (NK) cells frequency and activity elevation were selected and divided into two groups; 40 out of 72 received IVIG, aspirin, and heparin (anoxaparin) and constituted the treatment group and 32 patients received aspirin and heparin (anoxaparin) and no IVIG and were the control group. Th17, Treg frequency, transcription factors, cytokine gene expression, and cytokine secretion were evaluated by flow cytometry, real-time PCR, and ELISA, respectively. Post-treatment evaluation of the IVIG grouprevealed a significant increase in Treg associated parameters such as Treg frequency (p = 0.0186), Foxp3 (p = 0.0004), and cytokine mRNA levels (IL-10, p = 0.0058 and TGF- β, p = 0.0038), however, in the case of Th17, a significant difference was only observed in a reduction in the RORγt mRNA level (p = 0.0218). In conclusion, IVIG therapy may be a good choice in the treatment of implantation failure in RIF women especially with an immunologic basis, and may improve the implantation and pregnancy rate by affecting immunoregulatory mechanisms such as Tregs.

Abbreviations: RIF: repeated implantation failure; IVIG: intravenous immunoglobulin; Th17: T helper 17; Treg: T regulatory; NK cells: natural killer cells; PCR: polymerase chain reaction; ELISA: enzyme-linked immunosorbent assay; RORγt: RAR-related orphan receptor gamma; Foxp3: forkhead box protein P3; IL-17: interleukin-17; LMWH: low-molecular weight heparin; dNK: decidual NK cells.
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http://dx.doi.org/10.1080/19396368.2017.1390007DOI Listing
December 2017

Intravenous immunoglobulin (IVIG) treatment modulates peripheral blood Th17 and regulatory T cells in recurrent miscarriage patients: Non randomized, open-label clinical trial.

Immunol Lett 2017 12 10;192:12-19. Epub 2017 Oct 10.

Drug Applied Research Center, Tabriz University of Medical Sciences, Tabriz, Iran; Department of Immunology, School of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran. Electronic address:

Background: Th17 cells and Treg cells have been proposed as new risk factors for recurrent miscarriage (RM). In this study, we investigated the effect of Intravenous immunoglobulin G (IVIG) on the levels and function of Th17 and Treg cells and pregnancy outcome in women with RM.

Materials And Methods: 94 pregnant women with RM were enrolled in this study. Blood was drawn at the time of positive pregnancy. On the same day, IVIG 400mg/kg was administered intravenously for 44 patients. 50 other RM patients were included as no IVIG interfering control group. Following the first administration, IVIG was given every 4 weeks through 32 weeks of gestation. Peripheral blood was drawn after the last administration (32 weeks after pregnancy).

Results: IVIG down-regulated Th17 cells population and function and up-regulated Treg cells population and function were significant in the treated group. Pregnancy outcome in IVIG treated subjects was successful in 38 out of 44 RM women (86.3%). However, pregnancy outcome was successful in 21 out of 50 untreated RM women (42%).

Conclusion: Administration of IVIG in RM women with cellular immune cells abnormalities during pregnancy influences Th17/Treg ratio in peripheral blood and enhances Treg and decreases Th17 responses.
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http://dx.doi.org/10.1016/j.imlet.2017.10.003DOI Listing
December 2017

Effect of Intravenous immunoglobulin on Th1 and Th2 lymphocytes and improvement of pregnancy outcome in recurrent pregnancy loss (RPL).

Biomed Pharmacother 2017 Aug 12;92:1095-1102. Epub 2017 Jun 12.

Drug Applied Research Center, Tabriz University of Medical Sciences, Tabriz, Iran; Department of Immunology, Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran. Electronic address:

Background: Women with elevated natural killer (NK) cell frequency and function during pregnancy, suffer from recurrent pregnancy loss (RPL). In the present study, the possible effect of intravenous immunoglobulin (IVIG) administration on Th1 and Th2 cell frequency, cytokine secretion, and expression of transcription factors is compared between RPL patients and control group.

Materials And Methods: Totally, 44 women with a history of RPL (32 women as treated group and 12 as control group) were enrolled in the study. The frequency of Th1 and Th2 lymphocytes, the expression of transcription factors related to these cells and the serum levels of associated cytokines were assessed by flowcytometry, real-time PCR and ELISA, respectively. All, assessments were performed both before and after treatment with IVIG.

Results: A significant reduction in Th1 lymphocyte frequency, transcription factor expression and cytokine levels were observed in IVIG-treated group, while all the above parameters indicated a significant increase for Th2 lymphocytes. Th1/Th2 ratio decreased significantly (p value<0.0001) at the end of treatment and 28 out of 32 (87.5%) women in IVIG-treated group had live birth in comparison with 5 out of 12 (41.6%) in untreated group.

Conclusion: IVIG administration proves to be an efficient therapeutic strategy which is able to enhance the success rate of pregnancy through a shift in Th2 responses. Furthermore, IVIG presents efficacy for the treatment of reproduction failures especially in subjects with immune cell abnormalities and increased NK cell level and function.
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http://dx.doi.org/10.1016/j.biopha.2017.06.001DOI Listing
August 2017

Co-delivery of insulin-like growth factor 1 receptor specific siRNA and doxorubicin using chitosan-based nanoparticles enhanced anticancer efficacy in A549 lung cancer cell line.

Artif Cells Nanomed Biotechnol 2018 Mar 31;46(2):293-302. Epub 2017 Mar 31.

h Liver and Gastrointestinal Diseases Research Center , Tabriz University of Medical Sciences , Tabriz , Iran.

Here, we investigated the effects of dual delivery of IGF-1R siRNA and doxorubicin by chitosan nanoparticles on viability of A549 lung cancer cells line by utilization of MTT and qRT-PCR. Furthermore apoptosis and migration of treated cells were assessed by Annexin-PI and wound healing assays, respectively. The chitosan nanoparticles had about 176 nm size with zeta potential and polydispersive index about 11 mV and 0.3, respectively. The IGF-1R siRNA had synergistic effect on DOX-induced cytotoxicity and apoptosis in tumour cells. In addition, siRNA/DOX-loaded chitosan nanoparticles could significantly decrease migration and expressions of mmp9, VEGF and STAT3 in A549 cells.
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http://dx.doi.org/10.1080/21691401.2017.1307212DOI Listing
March 2018

Selection of single chain antibody fragments binding to the extracellular domain of 4-1BB receptor by phage display technology.

Tumour Biol 2017 Mar;39(3):1010428317695924

3 Hybridoma Laboratory, Department of Immunology, Pasteur Institute of Iran, Tehran, Iran.

The 4-1BB is a surface glycoprotein that pertains to the tumor necrosis factor-receptor family. There is compelling evidence suggesting important roles for 4-1BB in the immune response, including cell activation and proliferation and also cytokine induction. Because of encouraging results of different agonistic monoclonal antibodies against 4-1BB in the treatment of cancer, infectious, and autoimmune diseases, 4-1BB has been suggested as an attractive target for immunotherapy. In this study, single chain variable fragment phage display libraries, Tomlinson I+J, were screened against specific synthetic oligopeptides (peptides I and II) designed from 4-1BB extracellular domain. Five rounds of panning led to selection of four 4-1BB specific single chain variable fragments (PI.12, PI.42, PII.16, and PII.29) which showed specific reaction to relevant peptides in phage enzyme-linked immunosorbent assay. The selected clones were successfully expressed in Escherichia coli Rosetta-gami 2, and their expression was confirmed by western blot analysis. Enzyme-linked immunosorbent assay experiments indicated that these antibodies were able to specifically recognize 4-1BB without any cross-reactivity with other antigens. Flow cytometry analysis demonstrated an acceptable specific binding of the single chain variable fragments to 4-1BB expressed on CCRF-CEM cells, while no binding was observed with an irrelevant antibody. Anti-4-1BB single chain variable fragments enhanced surface CD69 expression and interleukin-2 production in stimulated CCRF-CEM cells which confirmed the agonistic effect of the selected single chain variable fragments. The data from this study have provided a rationale for further experiments involving the biological functions of anti-4-1BB single chain variable fragments in future studies.
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http://dx.doi.org/10.1177/1010428317695924DOI Listing
March 2017

Antiproliferative and Apoptotic Effects of Novel Anti-ROR1 Single-Chain Antibodies in Hematological Malignancies.

SLAS Discov 2017 04 31;22(4):408-417. Epub 2017 Jan 31.

2 Drug Applied Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.

Receptor tyrosine kinase-like orphan receptor (ROR) proteins are a conserved family of tyrosine kinase receptors that function in developmental processes including cell survival, differentiation, cell migration, cell communication, cell polarity, proliferation, metabolism, and angiogenesis. ROR1 has recently been shown to be expressed in various types of cancer cells but not normal cells. Pharmacokinetics and pharmacodynamics of single-chain Fragment variable (scFv) antibodies provide potential therapeutic advantages over whole antibody molecules. In the present study, scFvs against a specific peptide from the extracellular domain of ROR1 were selected using phage display technology. The selected scFvs were further characterized using polyclonal and monoclonal phage enzyme-linked immunosorbent assay (ELISA), soluble monoclonal ELISA, colony PCR, and sequencing. Antiproliferative and apoptotic effects of selected scFv antibodies were also evaluated in lymphoma and myeloma cancer cell lines using MTT and annexin V/PI assays. The results of ELISA indicated specific reactions of the isolated scFvs against the ROR1 peptide. Colony PCR confirmed the presence of full-length V and Vκ inserts. The percentages of cell growth after 24 h of treatment of cells with individual scFv revealed that the scFv significantly inhibited the growth of the RPMI8226 and chronic lymphocytic leukemia (CLL) cells in comparison with the untreated cells ( p < 0.05). Interestingly, 24-h treatment with specific scFv induced apoptosis cell death in the RPMI8226 and CLL cells. Taken together, our results demonstrate that targeting of ROR1 using peptide-specific scFv can be an effective immunotherapy strategy in hematological malignancies.
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http://dx.doi.org/10.1177/2472555216689659DOI Listing
April 2017

Isolation and characterization of anti ROR1 single chain fragment variable antibodies using phage display technique.

Hum Antibodies 2017 ;25(1-2):57-63

Drug Applied Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.

Receptor tyrosine kinase-like orphan receptor (ROR1) belongs to one of the families of receptor tyrosine kinases (RTKs). RTKs are involved in the various physiologic cellular functions including proliferation, migration, survival, signaling and differentiation. Several RTKs are deregulated in various cancers implying the targeting potential of these molecules in cancer therapy. ROR1 has recently been shown to be expressed in various types of cancer cells but not in normal adult cells. Hence a molecular inhibitor of extracellular domain of ROR1 that inhibits ROR1-cell surface interaction is of great therapeutic importance. In an attempt to develop molecular inhibitors of ROR1, we screened single chain variable fragment (scFv) phage display libraries, Tomlinson I + J, against one specific synthetic oligopeptide from extracellular domain of ROR1 and selected scFvs were characterized using various immunological techniques. Several ROR1 specific scFvs were selected following five rounds of panning procedure. The scFvs showed specific binding to ROR1 using immunological techniques. Our results demonstrate successful isolation and characterization of specific ROR1 scFvs that may have great therapeutic potential in cancer immunotherapy.
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http://dx.doi.org/10.3233/HAB-170310DOI Listing
March 2017

Isolation and characterization of anti c-met single chain fragment variable (scFv) antibodies.

J Immunotoxicol 2017 12 16;14(1):23-30. Epub 2017 Jan 16.

g Liver and Gastrointestinal Diseases Research Center , Tabriz University of Medical Sciences , Tabriz , Iran.

The receptor tyrosine kinase (RTK) Met is the cell surface receptor for hepatocyte growth factor (HGF) involved in invasive growth programs during embryogenesis and tumorgenesis. There is compelling evidence suggesting important roles for c-Met in colorectal cancer proliferation, migration, invasion, angiogenesis, and survival. Hence, a molecular inhibitor of an extracellular domain of c-Met receptor that blocks c-Met-cell surface interactions could be of great thera-peutic importance. In an attempt to develop molecular inhibitors of c-Met, single chain variable fragment (scFv) phage display libraries Tomlinson I + J against a specific synthetic oligopeptide from the extracellular domain of c-Met receptor were screened; selected scFv were then characterized using various immune techniques. Three c-Met specific scFv (ES1, ES2, and ES3) were selected following five rounds of panning procedures. The scFv showed specific binding to c-Met receptor, and significantly inhibited proliferation responses of a human colorectal carcinoma cell line (HCT-116). Moreover, anti- apoptotic effects of selected scFv antibodies on the HCT-116 cell line were also evaluated using Annexin V/PI assays. The results demonstrated rates of apoptotic cell death of 46.0, 25.5, and 37.8% among these cells were induced by use of ES1, ES2, and ES3, respectively. The results demonstrated ability to successfully isolate/char-acterize specific c-Met scFv that could ultimately have a great therapeutic potential in immuno-therapies against (colorectal) cancers.
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http://dx.doi.org/10.1080/1547691X.2016.1251512DOI Listing
December 2017

Epigenetic modifications and epigenetic based medication implementations of autoimmune diseases.

Biomed Pharmacother 2017 Mar 10;87:596-608. Epub 2017 Jan 10.

Drug Applied Research Center, Tabriz University of Medical Sciences, Tabriz, Iran; Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran; Department of Immunology, School of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran. Electronic address:

Recent genome-wide association studies have documented a number of genetic variants to explain mechanisms underlying autoimmune diseases. However, the precise etiology of autoimmune diseases remains largely unknown. Epigenetic mechanisms like alterations in the post-translational modification of histones and DNA methylation may potentially cause a breakdown of immune tolerance and the perpetuation of autoreactive responses. Recently, several studies both in experimental models and clinical settings proposed that the epigenome may hold the key to a better understanding of autoimmunity initiation and perpetuation. More specifically, data support the impact of epigenetic changes in autoimmune diseases, in some cases based on mechanistical observations. Epigenetic therapy already being employed in hematopoietic malignancies may also be associated with beneficial effects in autoimmune diseases. In this review, we will discuss on what we know and expect about the treatment of autoimmune disease based on epigenetic aberrations.
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http://dx.doi.org/10.1016/j.biopha.2016.12.072DOI Listing
March 2017

Chitosan nanoparticles as a dual drug/siRNA delivery system for treatment of colorectal cancer.

Immunol Lett 2017 01 1;181:79-86. Epub 2016 Dec 1.

Drug Applied Research Center, Tabriz University of Medical Sciences, Tabriz, Iran; Department of Immunology, School of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran. Electronic address:

Nanoparticles are widely used to deliver anticancer drugs and inhibit tumor growth without systemic toxicity. Recently, chitosan has received much attention as a functional biopolymer for encapsulation of small interfering RNA (siRNA). Because of cationic nature, chitosan efficiently encapsulate siRNA and forming nanoparticles. Moreover, biocompatible and biodegradable properties represent chitosan as potential candidate for in vivo siRNA delivery. In the present study we designed carboxymethyl dextran (CMD) chitosan nanoparticles (ChNPs) to encapsulate snail siRNA and anticancer drug doxorubicin (DOX). The effect of ChNPs-drug/siRNA on cell growth and Epithelial mesenchymal transition (EMT) gene expression of HCT-116 cell lines were investigated. Furthermore the efficacy of dual agent nanoparticle to induce apoptosis and inhibit migration of colorectal cancer cells were assessed using Annexin-V and wound healing assays, respectively. The results demonstrated that treatment with dual agent nanoparticles led to significant changes of EMT genes (i.e down regulation of MMP-9 and Vimentin and up regulation of E-cadherin), apoptosis cell death and migration inhibition in HCT-116 cells. In conclusion, ChNPs encapsulating DOX and snail siRNA can be considered as an effective anti-cancer drugs delivery system for the treatment of colorectal cancer.
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http://dx.doi.org/10.1016/j.imlet.2016.11.013DOI Listing
January 2017

Developing and characterization of single chain variable fragment (scFv) antibody against frizzled 7 (Fzd7) receptor.

Bioengineered 2017 Sep 16;8(5):501-510. Epub 2016 Nov 16.

b Drug Applied Research Center, Tabriz University of Medical Sciences , Tabriz , Iran.

ABSTACT Wnt/β-catenin signaling pathway through Frizzled receptors has been shown to play a key role in both normal development and tumorigenesis. Overexpression of Wnt pathway genes, such as Fzd7 in several malignancies is well-documented. Therefore, targeting of Fzd7 and its ligand inhibits cancer cells proliferation metastasis. In the present study we isolated single chain variable fragments (scFvs) against Fzd7 receptor using phage display method. Semi-synthetic human naive antibody libraries (Tomlinson I + J) was employed in panning procedure to isolate specific scFv against specific peptide from extracellular domain of Fzd7 receptor. The reactivity and growth inhibition effects of the selected antibodies was evaluated using enzyme-linked immunosorbent assay (ELISA), MTT and annexin V assays, respectively. Seven scFvs reactive to Fzd7 were selected following 4 rounds of panning. The results showed that the selected scFvs inhibits cell growth through apoptosis cell death in a triple negative breast cancer cells, MDA-MB-231. Given that Fzd7 and Wnt pathway plays a critical role in tumor progression, selected blocking scFvs represent significant potential for immunotherapy of breast cancer cells.
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http://dx.doi.org/10.1080/21655979.2016.1255383DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5639855PMC
September 2017

Antiproliferative and apoptotic effects of a specific anti-insulin-like growth factor I receptor single chain antibody on breast cancer cells.

Tumour Biol 2016 Nov 17;37(11):14841-14850. Epub 2016 Sep 17.

Drug Applied Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.

Insulin-like growth factor I receptor (IGF-IR) is expressed on breast cancer cells and involves in metastasis, survival, and proliferation. Currently, application of IGF-IR-targeting monoclonal antibodies (mAbs), alone or in combination with other drugs, is a promising strategy for breast cancer therapy. Single-chain fragment variable (scFv) antibodies have been introduced as appropriate tools for tumor-targeting purposes because of their advantages over whole antibodies. In the present study, we employed a naïve phage library and isolated scFvs against a specific epitope from extracellular domain of IGF-IR by panning process. The selected scFvs were further characterized using polyclonal and monoclonal phage ELISA, soluble monoclonal ELISA, and colony PCR and sequencing. Antiproliferative and apoptotic effects of selected scFv antibodies on breast cancer cell lines were also evaluated by MTT and Annexin V/PI assays. The results of ELISA indicated specific reactions of the isolated scFvs against the IGF-IR peptide, and analyses of PCR product and sequencing confirmed the presence of full length V and Vκ inserts. Treatment of MCF7 and SKBR3 cells with anti-IGF-IR scFv led to a significant growth inhibition. The results also showed that scFv treatment significantly augmented trastuzumab growth inhibitory effects on SKBR3 cells. The percentage of the apoptotic MCF7 and SKBR3 cells after 24-h treatment with scFv was 39 and 30.70 %, respectively. Twenty-four-hour treatment with scFv in combination with trastuzumab resulted in 44.75 % apoptosis of SKBR3 cells. Taken together, our results demonstrate that the targeting of IGF-IR by scFv can be an effective strategy in the treatment of breast cancer and provide further evidence for effectiveness of dual targeting of HER2 and IGF-IR in breast cancer therapy.
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http://dx.doi.org/10.1007/s13277-016-5323-4DOI Listing
November 2016

T Helper 1 (Th1), Th2, and Th17 Responses to Leishmania major Lipophosphoglycan 3.

Immunol Invest 2016 Oct 9;45(7):692-702. Epub 2016 Sep 9.

b Drug Applied Research Center , Tabriz University of Medical Sciences , Tabriz , Iran.

Leishmania major is the main causal agent of cutaneous leishmaniasis (CL) that remains a serious public health concern in many tropical and subtropical countries. A long-lasting protective vaccine against leishmaniasis remains as a medical unmet need. Lipophosphoglycan 3 (LPG3) is one of the class II LPG genes from HSP90 family involved in the host immune responses. The aim of the present study is to investigate the capability of recombinant LPG3 (rLPG3) to induce Th1, Th2, Th17 responses. The results showed that rLPG3 in moderate and high concentrations significantly induced expression of Th1 lineage-specific transcription factor (T-bet) and cytokine (IFN-γ)(P < 0.05). Moreover, the Th1-stimulating effect of rLPG3 was confirmed by significant induction of IFN-γ secretion from treated T cells (P < 0.01). However, no significant effect of rLPG3 on Th2 and Th17 lineage cells was observed even in high concentration. Our findings demonstrate that rLPG3 induces Th1, but not Th2 and Th17, lineage responses. Further studies are needed to investigate adjuvant properties of rLPG3 for leishmania therapy.
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http://dx.doi.org/10.1080/08820139.2016.1208217DOI Listing
October 2016

Design and construction of immune phage antibody library against Tetanus neurotoxin: Production of single chain antibody fragments.

Hum Antibodies 2015 Dec;23(3-4):73-9

Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.

Background: Tetanus neurotoxin (TeNT) is composed of a light (LC) and heavy chain (HC) polypeptides, released by anaerobic bacterium Clostridium tetani and can cause fatal life-threatening infectious disease. Toxin HC and LC modules represents receptor binding and zinc metalloprotease activity, respectively. The passive administration of animal-derived antibodies against tetanus toxin has been considered as the mainstay therapy for years. However, this treatment is associated with several adverse effects due to the presence of anti-isotype antibodies.

Objective: In the present study, we have produced the fully human single chain antibody fragments (HuScFv) from two human antibody phage display libraries.

Material And Methods: Twenty-four different HuscFvs were isolated from two anti TeNT immune libraries. Our produced human ScFv (HuScFv) were converted to IgG platform and analyzed regarding their specific reactivity to TeNT.

Results: All of the selected scFvs have the same VL but different VH. Three HuscFvs from the first library (TTX15, 51, 75) and two HuscFvs from the second library (TTX16, 20) were chosen to convert to IgG1 using pOptiVEC and pcDNA3.3 systems. Production of IgG1 from transfected DG44 and binding capacity of them to tetanus toxin and toxoid were measured by ELISA. ELISA results showed no detectable production of TTX16 and TTX20 IgG1. Although, TTX51 and TTX75 were converted and produced as IgG1, no reactivity to tetanus toxin and toxoid was observed. However, TTX15 was successfully produced as whole IgG1 platform with reactivity to both tetanus toxin and toxoid. The latter would be an appropriate replacement for conventional polyclonal antibodies if would meet the further characterization including specificity determination, affinity measurement and toxin neutralizing assays.

Conclusion: Our results demonstrated production of functional IgG1 derived from TTX15 scFv and might be an appropriate replacement for polyclonal Tetabulin but it needs further characterization.
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http://dx.doi.org/10.3233/HAB-150287DOI Listing
December 2015

Evaluation of EBV transformation of human memory B-cells isolated by FACS and MACS techniques.

J Immunotoxicol 2016 07 4;13(4):490-7. Epub 2016 Apr 4.

a Immunology Research Center, Tabriz University of Medical Sciences , Tabriz , Iran ;

Several studies have been performed to develop effective neutralizing monoclonal antibodies. The Epstein-Barr virus (EBV) can efficiently immortalize B-cells to establish lymphoblastoid cell lines (LCL) and so it has been used extensively for transformation of B-cells to produce and secrete immunoglobulin. The present study addressed the effect of TLR7/8 agonist (R848), feeder cells layer and fluorescence-activated cell sorting (FACS) and magnetic-activated cell sorting (MACS) cell separation methods on the transformation efficiency of antibody-producing memory B-cells. For these studies, the antigen used for analyses of antibody formation was the tetanus neurotoxin (TeNT) derived from Clostridium tetani. The results here showed that employing an HFFF.PI6 feeder cell layer, R848 agonist and FACS-mediated purification of memory B-cells led to increased transformation efficiency. Altogether, the effects of the R848 and the feeder cells provided an efficient method for EBV transformation of human B-cells. Moreover, there was an advantage in using FACS sorting of B-cells over the MACS method in the context of EBV transformation and immortalization of precursors of antigen-specific B-cells.
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http://dx.doi.org/10.3109/1547691X.2015.1132288DOI Listing
July 2016

Recombinant Leishmania major lipophosphoglycan 3 activates human T-lymphocytes via TLR2-independent pathway.

J Immunotoxicol 2016 16;13(2):263-9. Epub 2015 Jul 16.

b Immunology Research Center , and.

Leishmaniasis is one of the most common infectious diseases transmitted by an obligate intracellular genus Leishmania. As there is no efficient vaccination strategy for leishmaniasis, new immunostimulatory components may enhance protective immune responses against this parasite. Lipophosphoglycan 3 (LPG3) is an essential protein required for LPG assembling. In this study, the ability of recombinant LPG3 (rLPG) and its fragments to activate isolated healthy human T-cells and cytokine secretion was evaluated in vitro. The results showed that rLPG3 and its N-terminal fragment (rNT-LPG3) enhanced expression of CD69 on the surface of T-cells and promoted differentiation of CD4(+) T-lymphocytes toward a T-helper 1 (T(H)1) phenotype, in part, through up-regulation of interferon (IFN)-γ expression in a TLR2-independent manner. These results indicated the protective effects of LPG3 (particularly NT-LPG3 fragment) as a potent immunostimulatory component of leishmania in vaccination against leishmaniasis. Further investigations in in vivo assays are clearly warranted.
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http://dx.doi.org/10.3109/1547691X.2015.1066906DOI Listing
September 2016

Comparative human and mouse antibody responses against tetanus toxin at clonal level.

J Immunotoxicol 2016 20;13(2):243-8. Epub 2015 May 20.

a Department of Immunology , School of Public Health, Tehran University of Medical Sciences , Tehran , Iran .

Tetanus is a highly fatal disease caused by tetanus neurotoxin (TeNT) and remains a major threat to human and animal health, despite preventive strategies. TeNT is composed of heavy and light chain linked by a disulfide bond. The antibody response to TeNT is polyclonal and directed to multiple epitopes within both the light and heavy chains, leading to toxin neutralization. This study was undertaken to localize and compare neutralizing epitopes recognized by human and mouse TeNT-specific antibodies at a clonal level. In the present study, 22 murine hybridoma clones and 50 human lymphoblastoid cell lines secreting monoclonal antibodies (mAb) were generated against TeNT. The specificity of these mAb was determined using different recombinant fragments of tetanus toxin. Moreover, this study investigated the in vitro toxin neutralizing activity of these mAb by a ganglioside GT1b assay. The results showed that tetanus toxoid immunization in humans and BALB/c mice induced a vigorous humoral immune response against different fragments of TeNT, particularly the carboxyl-terminal fragment of the heavy chain (known as fragment C). The fragment C-specific human and mouse mAb could largely neutralize TeNT. However, while all fragment C-specific human mAb reacted with the carboxyl-terminal part of this fragment (H(CC)), the majority of the mouse mAb failed to recognize this region. These results suggested that fragment C is the major target for the TeNT neutralizing antibodies, although different epitopes seem to be targeted by human and mouse antibodies.
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http://dx.doi.org/10.3109/1547691X.2015.1046572DOI Listing
September 2016

Immunotherapeutic approaches for cancer therapy: An updated review.

Artif Cells Nanomed Biotechnol 2016 May 24;44(3):769-79. Epub 2015 Mar 24.

a Immunology Research Center, Tabriz University of Medical Sciences , Tabriz , Iran.

In spite of specific immune effector mechanisms raised against tumor cells, there are mechanisms employed by the tumor cells to keep them away from immune recognition and elimination; some of these mechanisms have been identified, while others are still poorly understood. Manipulation or augmentation of specific antitumor immune responses are now the preferred approaches for treatment of malignancies, and traditional therapeutic approaches are being replaced by the use of agents which potentiate immune effector mechanisms, broadly called "immunotherapy". Cancer immunotherapy is generally classified into two main classes including active and passive methods. Interventions to augment the immune system of the patient, for example, vaccination or adjuvant therapy, actively promote antitumor effector mechanisms to improve cancer elimination. On the other hand, administration of specific monoclonal antibodies (mAbs) against different tumor antigens and adoptive transfer of genetically-modified specific T cells are currently the most rapidly developing approaches for cancer targeted therapy. In this review, we will discuss the different modalities for active and passive immunotherapy for cancer.
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http://dx.doi.org/10.3109/21691401.2015.1019669DOI Listing
May 2016

Down-regulation of TLR2, 3, 9 and Signaling Mediators, MyD88 and TRIF, Gene Transcript Levels in Patients with Kawasaki Disease Treated with IVIG.

Iran J Allergy Asthma Immunol 2015 Apr;14(2):188-97

Division of Allergy and Clinical Immunology, Department of Pediatrics, Shiraz University of Medical Sciences, Shiraz, Iran

Kawasaki disease (KD) is an acute febrile systemic vasculitis of childhood characterized by elevated levels of inflammatory mediators at the acute stage. High-dose intravenous immunoglobulin (IVIG) is well accepted as a conventional therapy for KD. The aim of the present study was to determine the expression level of Toll like receptors (TLRs) and their corresponding signaling mediators in PBMCs of IVIG-treated KD patients. TLR2, 3, 9 and signaling mediators, MyD88 and TRIF transcript levels were determined in PBMCs from 31 KD patients, before (acute phase), 2 weeks later (sub-acute phase) and 6 weeks later (convalescent phase) of IVIG therapy using real time PCR. The mean age of the patients was 3.6 years and 65% of subjects were male and 35% were female. 20 age-matched irrelevant febrile patients and 20 healthy subjects were included as control groups. Elevated levels of TLR2, MyD88, and TRIF gene transcripts were observed in the PBMCs at acute phase of untreated KD patients in compression with normal subjects. IVIG therapy resulted in significant decrease in TLR2, 3 and 9 (60-90%) as well as MyD88 and TRIF (60-70%) transcripts following 2 and 6 weeks. With Regard to significant up-regulation of MyD88 and TRIF at the acute phase of KD, our findings suggest TLR signaling pathway potential in KD pathogenesis and may also support the assumption of an infectious background in KD. Down-regulation of TLR members and corresponding mediators in IVIG treated patient suggest general TLR pathway suppression as a novel anti-inflammatory mechanism of IVIG.
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April 2015

Induction of anti-proliferative and apoptotic effects by anti-IL-25 receptor single chain antibodies in breast cancer cells.

Int Immunopharmacol 2014 Dec;23(2):624-32

Overexpression of IL-25 receptor (IL-25R) on the surface of malignant, but not normal, breast epithelial cells contributes to tumorigenic potential of breast cancer cells. IL-25/IL-25R binding results in specific apoptosis in breast cancer cells. In this study, the inhibitory effects of anti-IL-25R scFv antibodies on three breast cancer cells were evaluated. Panning process was performed on a phage library to isolate scFvs against two specific epitopes of IL25R. The reactivities of scFvs were assessed using phage ELISA. Cell binding capacity of the selected scFvs on breast cancer cells was analyzed using flow cytometry. Anti-proliferative and apoptotic effects of scFvs were evaluated by MTT, Annexin V/PI and quantitative caspase 3 activity assays. Two scFvs with frequencies of 47% and 68% were selected against peptides I and II, respectively. Phage ELISA demonstrated the specific reactions of the scFvs against the corresponding peptides. The scFvs against peptides I and II bound to 14% and 21% of MDA-231, 28% and 32% of MCF7, and 21% and 30% of SKBR3 cells, respectively. Treatment of the breast cancer cells with IL-25 and two anti-IL-25R scFvs resulted in significant inhibition of the cell growth in these cell compared to that in IL-25R negative Raji cells. The results also demonstrated 66% and 70% apoptosis cell death and 15.5 and 17 fold increases in caspase- 3 activity in SKBR3 cells induced by scFvs I and II, respectively. Our findings suggest the targeting of IL-25R by scFvs as a new effective strategy in breast cancer immunotherapy.
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http://dx.doi.org/10.1016/j.intimp.2014.10.015DOI Listing
December 2014

mRNA expression of pattern recognition receptors and their signaling mediators in healthy and diseased gingival tissues.

J Indian Soc Periodontol 2014 Mar;18(2):150-4

Shiraz Institute for Cancer Research, Shiraz University of Medical Sciences, Shiraz, Iran ; Department of Immunology, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran.

Background: Gingivitis and periodontitis are initiated by inflammation caused by microorganisms. Pathogen-associated molecular patterns (PAMPs) from these microorganisms are recognized through various toll-like receptors (TLRs) and NOD-like receptors (NLRs). In this study, we have chosen five TLRs and two NLRs as representatives taking part in the recognition and inflammation process, along with a few of their signaling mediators including CD14, MYD88, and TRIF to compare their mRNA expression levels between healthy and diseased gingival tissues. This will provide deeper insight into the mechanisms underlying gingivitis and periodontitis. Understanding the mechanisms involved in the onset and progression of the periodontal diseases could greatly help in establishing effective ways for prevention and treatment of these diseases besides decreasing the risk factor for relevant systemic disorders.

Materials And Methods: Gingival tissue samples for mRNA extraction and cDNA synthesis were taken from patients with gingivitis and periodontitis and from healthy control subjects. Messenger RNA expression of all genes was assessed using real-time polymerase chain reaction (PCR).

Results: Among the genes studied in different groups, only MYD88 mRNA expression was significantly higher in the periodontitis group compared to that of the controls. The expression level of this molecule was also significantly higher in patients with severe periodontitis compared to other patients and also compared to healthy individuals. In different tissues, positive significant correlations were observed between the mRNA expression levels of some genes.

Conclusions: Elevated mRNA levels of MYD88 in periodontitis might have a key role in the pathogenesis of this disease. Therefore, MYD88 may be a useful target for the therapy of this disease.
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http://dx.doi.org/10.4103/0972-124X.131309DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4033878PMC
March 2014

Deregulation of HER2 downstream signaling in breast cancer cells by a cocktail of anti-HER2 scFvs.

Oncol Res 2013 ;20(8):333-40

Shiraz University of Medical Sciences, Shiraz, Iran.

Human epidermal growth factor receptor 2 (HER2) is overexpressed in 30% of patients with breast cancer. HER2 targeting is the mainstay of targeted therapy for the treatment of invasive breast cancers. Due to biological and therapeutic advantages, single chain fragment variable (scFv) antibodies have emerged as promising alternative therapeutics. In this study, we assessed the capability of three scFvs against HER2 extracellular domains (II, III, IV) in deregulation of some key signaling mediators that have important roles in growth, survival, angiogenesis, and cell migration of breast tumor cells. Downregulation of activated Akt (p-Akt), increase of p27 protein levels, and downregulation of HER1, HER2, HER3 and epidermal growth factor (EGF), CXCR3, CXCL10, and MMP2 were observed following treatment of breast cancer cells (SKBR3 cell line) with the scFvs and their combination. Our results suggest that the combination of the three scFvs could be considered as an effective cocktail on HER2 tumorgenic signaling pathways that leads to tumor growth suppression and death.
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http://dx.doi.org/10.3727/096504013X13657689382734DOI Listing
September 2013

Hyperthermia increases natural killer cell cytotoxicity against SW-872 liposarcoma cell line.

Iran J Immunol 2013 Jun;10(2):93-102

Department of Immunology, Shiraz University of Medical Sciences, Shiraz, Iran, e-mail:

Background: Although there is convincing data in support of the effectiveness of hyperthermia in tumor therapy, the molecular mechanisms underlying the clinical effects of hyperthermia are still poorly understood.

Objective: To investigate natural killer (NK) cell cytotoxicity against heat-treated SW-872 and HeLa tumor cell lines.

Methods: NKG2D ligands and HLA class I transcription were examined using quantitative real-time PCR in treated tumor cell lines at 0, 2, 4, 6 and 12 h following thermal treatment at 39C and 42C for 1 h. The expression of MICA/B, ULBP1 and ULBP2 were also determined by flow cytometry. NK92-MI cytotoxic activity against heat-treated target cell lines was assessed by LDH release as well as annexin-V and 7-AAD assays.

Results: Our results showed that heat treatment at 39C improved the cytolytic activity of NK cells against SW-872 cells without increasing NKG2D ligand concentration or decreasing HLA class I levels.

Conclusion: The observed increase in the cytotoxicity of NK cells against SW-872 cells after hyperthermia does not coincide with changes in MICA/B, ULBP1 and ULBP2 ligands of NKG2, however, the expression of other ligands in target cells may have made the cells susceptible to the cytotoxic effect of NK cells.
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http://dx.doi.org/IJIv10i2A4DOI Listing
June 2013

Assessment of the effect of TLR7/8, TLR9 agonists and CD40 ligand on the transformation efficiency of Epstein-Barr virus in human B lymphocytes by limiting dilution assay.

Cytotechnology 2014 Jan 13;66(1):95-105. Epub 2013 Feb 13.

Department of Immunology, School of Public Health, Tehran University of Medical Sciences, 14155, Tehran, Iran.

Infection of human B cells with Epstein-Barr virus (EBV) induces polyclonal activation in almost all infected cells, but a small proportion of infected cells are transformed to immortalized lymphoblastoid cell lines. Since B cells are activated also by CD40 ligand (CD40L) and Toll-like receptor (TLR) agonists via a similar signaling pathway, it is likely that costimulation through these molecules could result in synergistic enhancement of the transformation efficiency of EBV. In this study, the stimulatory effect of TLR7/8 (R848), TLR9 (CpG) agonists and/or CD40L on transformation efficiency of EBV in normal human B cells was assessed using the limiting dilution assay. Costimulation of peripheral blood mononuclear cells (PBMCs) with CpG and R848, but not CD40L, increased significantly the frequency of EBV transformed B cells (p < 0.001). Neither synergistic nor additive effects were observed between TLR agonists and CD40L and also TLR7/8 and TLR9 agonists. Costimulation with R848, CpG and CD40L enhanced the proliferative response of B cells infected with EBV. This effect was more evident when enriched B cells were employed, compared to PBMCs. The promoting effect of TLR agonists stimulation, implies that EBV may take advantage of the genes induced by the TLR stimulation pathway for viral latency and oncogenesis.
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http://dx.doi.org/10.1007/s10616-013-9542-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3886530PMC
January 2014
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