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1.
Chinese Journal of Biologicals ; (12): 551-558, 2023.
Article in Chinese | WPRIM | ID: wpr-996369

ABSTRACT

@#Objective To analyze the topology of IFN-induced transmembrane(IFITM) protein in porcine peripheral blood lymphocytes(PBMCs) and detect the change of IFITM mRNA transcription in PBMCs after porcine reproductive and respiratory syndrome virus(PRRSV) infection in vitro.Methods PRRSV,porcine circovirus 2(PCV2) and Japanese encephalitis virus(JEV) negative anticoagulant blood of piglets were collected aseptically and isolated for PBMCs.Porcine IFITM CDS sequence was amplified by PCR,sequenced and analyzed for topology.PBMCs were infected with PRRSV in vitro.Cell samples were collected at 12,24,36 and 48 h after infection,detected for PRRSV infection by RT-PCR,and detected for mRNA transcription level changes of IFITM1,IFITM2 and IFITM3 by RT-PCR.Results The porcine PBMCs were successfully isolated and the full-length sequence of IFITM CDS derived from PBMCs was cloned.The porcine IFITM protein might have two topological structures.PBMCs inoculated with PRRSV for 24 h produced obvious cytopathic effect.PRRSV was replicated in PBMCs.The transcription levels of IFITM1,IFITM2 and IFITM3 mRNA in PBMCs were significantly up-regulated at the early stage of PRRSV infection,and reached the peak at 12h after infection,and then gradually decreased;The transcription level of IFITM1 mRNA increased at 36 h after virus infection and then declined rapidly.Conclusion PRRSV infection in vitro significantly up-regulated the transcription level of IFITM mRNA in PBMCs,indicating that IFITM was involved in the antiviral immune response of PBMCs.This study provided a reference for revealing the natural immune response against PRRSV in vivo.

2.
Afr. J. Gastroenterol. Hepatol ; 6(1): 1-18, 2023. figures, tables
Article in English | AIM | ID: biblio-1512671

ABSTRACT

Aims: the current research aimed to investigate LncRNA-MIAT in patients with nonHodgkin lymphoma (NHL) and to assess its correlation with clinicopathological features and treatment protocols of NHLs among Egyptian patients with Occult hepatitis C virus (HCV) infection (OCI). Patients & Methods: This study was conducted on 20 patients with NHL and 30 healthy subjects as the control group. All subjects were screened for HCV-RNA in both plasma and PBMCs. RT-PCR determined lncRNA-MIAT. Results: lncRNA-MIAT relative expression level was upregulated in NHL groups (2.73±0.86) compared to controls (1.06±0.07), P ˂0.001*. Among NHL, patients with OCI (3.2±0.63) had significantly higher levels of lncRNA-MIAT compared to HCV (2.6±1.08) and non-HCV (2.4±0.4), P ˂0.001*. Additionally, the relative expression levels of lncRNA-MIAT were significantly positively correlated with laboratory and clinicopathological features of NHL. Interestingly, concerning the treatment of DLBCLNHL, there were significantly higher levels of lncRNA-MIAT in no treatment subgroup (n=10, 3.31±0.95) compared to successfully treated subgroups [CHOP (n=7, 1.58±0.34) and R-CHOP (n=3, 11.16±0.21), P ˂0.001* Conclusions: lncRNA-MIAT level was upregulated in NHL patients, particularly patients with OCI. Thus, circulatory lncRNA-MIAT may serve as a promising non-invasive diagnostic marker for NHL associated with OCI


Subject(s)
Humans , Male , Female , Lymphoma, Non-Hodgkin , RNA, Long Noncoding , Myocardial Infarction
3.
Journal of Xi'an Jiaotong University(Medical Sciences) ; (6): 373-382, 2022.
Article in Chinese | WPRIM | ID: wpr-1011564

ABSTRACT

【Objective】 Through bioinformatics methods to analyze the differences in the gene expression profiles of peripheral blood mononuclear cells (PBMCs) between middle-aged and elderly women and normal people, so as to explore the diagnosis and treatment targets of OA. 【Methods】 We downloaded the GSE48556 data set from GEO databases. We utilized the R language to screen out the differentially expressed genes (DEGs) between OA and NC. By gene set enrichment analysis (GSEA), we obtained the target gene subset. The GO and KEGG pathways of the target gene subset were analyzed by DAVID. We applied STRING and Cytoscape software to construct PPI network. The module analysis was performed by the Mcode and centiscape plug-in, and the key genes were screened out by Cytohubba. 【Results】 By GSEA analysis and P.adjust 0.2, a total of 292 target genes were screened, consisting of 81 upregulated genes and 211 downregulated genes. The GO enrichment analysis of all target genes mainly focused on the biological functions, such as “regulation of NIK/NF-κB”, “monocytes”, “proliferation”, “regulation of apoptosis signaling pathway”, “TNF-mediated signaling pathway”, “regulation of Wnt signaling pathway”, “regulation of MAP kinase activity”, and “regulation of autophagy”. KEGG was mainly enriched in four pathways: cytotoxicity of natural killer cell mediation, TNF signaling pathway, MAPK signaling pathway, and apoptosis. We employed PPI network and related plug-ins to screen out eight core genes highly related to OA inflammation and apoptosis, namely, MAPK1, IL10, PTGS2, IL18, GSK3B, NFKBIA, TNFRSF1A, and EGR1. 【Conclusion】 Bioinformatics analysis revealed that the differences in PBMCs gene expressions between OA and NC were concentrated in the biological events of apoptosis and inflammation, making blood expression profile an effective breakthrough for monitoring OA target markers.

4.
China Occupational Medicine ; (6): 248-2022.
Article in Chinese | WPRIM | ID: wpr-965158

ABSTRACT

@#Abstract: Objective To screen and validate the differentially expressed microRNAs (miRNAs) in peripheral blood Methods - mononuclearcells(PBMCs)ofpatientswithsilicosis. Forty eightpatientswithoccupationalsilicosisatstageⅠ(case group)and45healthycontrols(controlgroup)wereselectedasresearchsubjectsbyrandomnumbertablemethod.PBMCswere-separatedbyFicollPaquegradientcentrifugationfromperipheralblood.Threepeoplefromeachgroupwererandomlyselected for miRNAs transcriptome sequencing. R Studio software was used to screen differentially expressed miRNAs, and FunRich software was used to predict the upstream transcription factors related to the differentially expressed miRNAs in PBMCs. The - Results differentially expressed miRNAs were verified by real time quantitative polymerase chain reaction. A total of 124 - - differentiallyexpressedmiRNAswerescreened,amongthem,97miRNAswereupregulatedand27miRNAswere down regulated.- The 67 targetgenes predicted by differentialmiRNAs were mainly involved in intracellularprocesses and nucleic acid binding transcriptionfactoractivities,includingcelladhesionmolecules,ratsarcoma,osteoclastdifferentiationandotherpathways.The-------------topfivemiRNAsdifferentiallyupregulatedwerehsamiR1373p,hsamiR500b3p,hsamiR190a5p,hsamiR1413pand------------hsamiR223p.ThetopfivedifferentiallydownregulatedmiRNAswerehsamiR2025p,hsamiR548ai,hsamiR55873p,------hsamiR5705pandhsamiR103983p.ThechangeofthesemiRNAswereconsistentwiththeresultspredictedaccordingto the miRNAs transcriptome sequencing. The transcription factors including specificity protein 1, early growth response 1, zinc Conclusion finger protein 161, etc., were obtained according to the differentially expressed miRNAs. The differentially

5.
Braz. j. med. biol. res ; 54(8): e10850, 2021. tab, graf
Article in English | LILACS | ID: biblio-1249328

ABSTRACT

The conversion of adenosine to inosine is catalyzed by adenosine deaminase (ADA) (EC 3.5.4.4), which has two isoforms in humans (ADA1 and ADA2) and belongs to the zinc-dependent hydrolase family. ADA modulates lymphocyte function and differentiation, and regulates inflammatory and immune responses. This study investigated ADA activity in lymphocyte-rich peripheral blood mononuclear cells (PBMCs) in the absence of disease. The viability of lymphocyte-rich PBMCs isolated from humans and kept in 0.9% saline solution at 4-8°C was analyzed over 20 h. The incubation time and biochemical properties of the enzyme, such as its Michaelis-Menten constant (Km) and maximum velocity (Vmax), were characterized through the liberation of ammonia from the adenosine substrate. Additionally, the presence of ADA protein on the lymphocyte surface was determined by flow cytometry using an anti-CD26 monoclonal human antibody, and the PBMCs showed long-term viability after 20 h. The ADA enzymatic activity was linear from 15 to 120 min of incubation, from 2.5 to 12.5 µg of protein, and pH 6.0 to 7.4. The Km and Vmax values were 0.103±0.051 mM and 0.025±0.001 nmol NH3·mg-1·s-1, respectively. Zinc and erythro-9-(2-hydroxy-3-nonyl) adenine (EHNA) inhibited enzymatic activity, and substrate preference was given to adenosine over 2′-deoxyadenosine and guanosine. The present study provides the biochemical characterization of ADA in human lymphocyte-rich PBMCs, and indicates the appropriate conditions for enzyme activity quantification.


Subject(s)
Humans , Adenosine Deaminase , Dipeptidyl Peptidase 4 , Leukocytes, Mononuclear , Adenine , Lymphocytes
6.
Chinese Journal of Experimental Traditional Medical Formulae ; (24): 1-8, 2019.
Article in Chinese | WPRIM | ID: wpr-801722

ABSTRACT

Objective:To observe the clinical efficacy of modified Erchentang on CXC chemokine ligand receptors (CXCR1/2)and their ligands CXCL8,macrophage inflammatory protein -2(MIP-2) in patients of chronic obstructive pulmonary disease(AECOPD)at acute exacerbation stage,and assess the effect and mechanism of modified Erchentang on anti-inflammatory in patients of AECOPD. Method:This study was a multicenter, randomized single blind, controlled trial. The authors selected 200 cases in conformity to the standards of AECOPD. The AECOPD patients were randomly divided into modified Erchentang group and control group. In addition to the western medicine, modified Erchentang was also given to the modified Erchentang group, and Jizhitangjiang was given to the control group for 14 days. Each group was observed for the alleviation of the symptoms. Euzyme-linked immunosorbent assay (ELISA) was used to determine the levels of CXCL8 and MIP-2 in the patients' plasma of all groups before and after treatment. Western blot were used to detect the levels of CXCR1, CXCR2 and CXCL8 protein in peripheral blood mononuclear cells(PBMCs). Immunocytochemistry (ICC) method was used to detect the expressions of CXCL8, CXCR1 and CXCR2 protein in PBMCs. Result:The level of CXCL8 in plasma, and the expressions of CXCR1, CXCR2 and CXCL8 mRNA and protein in the modified Erchentang group were decreased significantly than those in the control group(PPConclusion:Modified Erchentang has an anti-inflammatory effect on AECOPD. Its mechanism may be related to the down-regulation of the expressions of CXCL8, CXCR1 and CXCR2, the reduction of synthesis and release of CXCL8 and MIP-2, the inhibition of the chemotaxis and activity of inflammatory cells, and the prevention of inflammation progress.

7.
Journal of Practical Stomatology ; (6): 812-815, 2017.
Article in Chinese | WPRIM | ID: wpr-697431

ABSTRACT

Objective:To investigate the expression of miR-132 in peripheral blood mononuclear ceils(PBMCs) and plasma of patients with oral lichen planus (OLP) and the relationship of miR-132 with clinical forms of OLP.Methods:40 OLP patients (24 nonerosive OLP,16 erosive OLP) and 40 healthy controls were included.The relative expression of miR132 in peripheral blood mononuclear cells (PBMCs) and plasma was examined by quantitative real-time PC R.Results:The expression of miR-132 in both PBMCs and plasma of OLP patients was significantly higher than those in healthy controls(P < 0.01).miR-132 expression of PBMCs and plasma in erosive OLP group was significantly higher than that in non-erosive OLP group and healthy control(P < 0.01).However,the expression of miR-132 in PBMCs and plasma showed no significant difference between non-erosive OLP group and healthy control (P >0.05).Conclusion:miR-132 might be involved in the abnormal immune response of OLP patients,and miR-132 can be utilized as an assistant biomarker for the evaluation of the severity of OLP.

8.
Chinese Journal of Immunology ; (12): 1195-1199, 2016.
Article in Chinese | WPRIM | ID: wpr-495087

ABSTRACT

Objective:To study the expression of CXCL 8 in the serum and CXCL8 mRNA in the peripheral blood mononuclear cells(PBMCs) of the children with Mycoplasma pneumoniae pneumonia (MPP) and its clinical significance.Methods: Forty-eight children(severe cases 12,light cases 36) with MPP were recruited from October 2013 to March 2015 in the Maternal and Child Health-Care Hospital of Huainan.The concentration of the CXCL8 in serum and the level of CXCL8 mRNA in the PBMCs were measured by enzyme linked immunosorbent assay ( ELISA) and polymerase chain reaction ( PCR).Taking GAPDH as the internal reference ,the ratio of lgcDNA/lgGAPDH was regarded as the extreme level of CXCL 8 mRNA.Results: The serum level of CXCL8 and expression of CXCL8 mRNA in PBMCs in the children with MPP were ( 298.917 ±51.860 ) pg/ml and ( 1.848 ±0.525 ) lgcDNA/lgGAPDH.Compared with the normal control ,there were significant differences between the two groups ( P0.05).However, the expression of CXCL8 mRNA in peripheral blood of the children with severe illness was significantly higher than those in light cases (P<0.05).Intravenous infusion of Erythromycin was provided in the acute phase for seven to ten days ,so that the children′s condition could be significantly controlled , and the symptoms of pulmonary inflammation were also relieved .Followed by the use of sequential therapy of Azithromycin for about two to three weeks ,the children′s condition were gradually from acute stage to recovery stage .At this time,the CXCL8 and its mRNA levels in peripheral blood of the sick children were all significantly decreased comparing with those in the acute stage(P<0.05).Conclusion: The expression of CXCL8 and its mRNA were increased in the peripheral blood of the sick children with Mycoplasma pneumonia ,and also correlated with the severity of the disease .CXCL8 can participate in the pathogenesis of Mycoplasma pneumonia ,and has a certain cue effect on the severity and prognosis of the disease .Azithromycin can reduce the content of CXCL8 in serum of the sick children via the pathway of inhibiting the proliferation of Mycoplasma pneumoniae ,and down regulate the expression of mRNA ,so that the immune injury mediated by Mycoplasma pneumoniae may be gradually inhibited .

9.
Chinese Journal of Immunology ; (12): 1424-1427,1436, 2016.
Article in Chinese | WPRIM | ID: wpr-605660

ABSTRACT

Objective:To observe the changes of gene expression in peripheral blood mononuclear cells( PBMCs) of benign and malignant breast tumor based on gene expression profiling. Methods: Datasets of gene expression profiling were downloaded from the GEO database,including PBMCs profilings of benign breast tumor,breast cancer and healthy controls. GEO2R tool was used to analyze the data to identify the differentially expressed genes (DEGs). Function of DEGs were annotated by DAVID. Protein interaction analysis and hub gene select were then performed using STRING database. Results:563 and 237 DEGs respectively were identified. DEGs in breast cancer involved in biological process of leukocyte activation,angiogenesis and leukocyte transendothelial migration. The hub genes are IL8,RHOB,ITGB1. Conclusion:The data suggests that gene expression patterns of these two profilings are different at a certain degree. PBMCs maybe a better noninvasive material for biomarker detection of benign and malignant breast tumor.

10.
Br Biotechnol J ; 2014 Dec; 4(12): 1263-1271
Article in English | IMSEAR | ID: sea-162548

ABSTRACT

Low frequency electromagnetic fields (LF-EMFs) which can be generated from homes and workplaces appliances can lead to alteration in oncogenes causing cancer diseases such as leukemia, nervous system tumors, lymphoma and breast cancer. To investigate the effect of LF-EMFs on c-myc oncogene expression level, primary cell culture of human peripheral blood mononuclear cells (PBMCs) were exposed to AC (50 Hz) electromagnetic flux density: 0.37mT, 0.82mT, 1.22mT, 1.68mT, 2.1mT, 2.47mT, 2.85mT, 3.33mT, 3.72mT, 3.92mT, 4.32mT and 4.67mT using exposure unit. After four days of exposure, when initial changes in cell viability, morphology and count between exposed and unexposed cells were noted, total RNA was extracted to evaluate the expression level of c-myc oncogene by quantitative real time PCR (qRT-PCR). The results showed that c-myc oncogene expression level began to increase gradually from value 1.69% at 0.82mT reaching to the maximum expression level value 4.65% at 4.67mT. This result refers to an increasing in LF-EMFs exposure offset by an increasing in c-myc oncogene expression level, affecting different cellular functions. This study indicates that LF-EMF is environmental pollution effects on the expression level of oncogenes which increase the risk of human cancer diseases.

11.
Indian J Med Microbiol ; 2014 April-June ; 32 (2): 164-168
Article in English | IMSEAR | ID: sea-156884

ABSTRACT

The conventional method of transfection of suspension cells by chemical has proven to be very difficult. We present a new transfection protocol, wherein, low‑speed centrifugation of cell culture plates immediately after adding the lipid: DNA complex significantly enhances the transfection efficiency. Peripheral blood mononuclear cells (PBMCs) were transfected with BLOCK‑iT™ Fluorescent Oligo (scrambled siRNA) and lipofectamine complex using conventional and low‑speed centrifugation modified transfection protocols. The efficiency of transfection was determined using flowcytometer and cell viability was checked using MTT assay. Incorporation of low‑speed centrifugation significantly enhances the transfection efficiency of BLOCK‑iT™ in the suspension culture of PBMCs as compared to conventional transfection method (99.8% vs 28.3%; P < 0.0001), even at a low concentration of 40 picomoles without affecting the cell viability. Centrifugation enhanced transfection (CET) technique is simple, time‑saving and novel application without compromising the cell viability in the context of recently popular RNA interference in suspension cultures of PBMCs. This undemanding modification might be applicable to a wide variety of cell lines and solve crucial problem of researchers working with RNA interference in suspension cultures.

12.
Chinese Journal of Immunology ; (12): 946-949, 2014.
Article in Chinese | WPRIM | ID: wpr-452575

ABSTRACT

Objective:To study the HBV infection in peripheral blood mononuclear cells in mediating the role of mother -to-child transmission of hepatitis B virus.Methods: The peripheral blood mononuclear cells ( PBMCs ) in maternal and cord blood mononuclear cells ( CBMCs ) in newborns were conventionally isolated by Ficoll-Hypaque medium.The loads of HBV-DNA in peripheral blood of maternal and cord blood of newborns were both detected by PCR .Results:The clinical data showed that the positive detection rates of HBV-DNA in serum and PBMCs of pregnant women with HBeAg (+) were 100.00%( 25/25 ) and 72.00%( 18/25),and the positive detection rates of HBV-DNA in the neonatal umbilical cord blood serum and CBMCs were 60.00%(15/25) and 44.00%(11/25),respectively.There were significantly difference between HBeAg (+) and HBeAg(-) in the pregnant women (P<0.05 ).The positive detection rates of HBV-DNA in neonatal umbilical cord blood serum and CBMCs were higher in the group with high HBV loads (more than 106copies/ml) in PBMCs than those of low HBV loading group (102-103copies/ml).The significantly difference was explored between the two groups.Conclusion: Mononuclear cells can not only be infected by HBV , but also play a critical role in the intrauterine vertical transmission of HBV via the pathway transmitted from PBMCs in pregnant women to CBMCs in newborns.

13.
Article in English | IMSEAR | ID: sea-163813

ABSTRACT

Highly active anti-retroviral therapy (HAART) is the current HIV/AIDS treatment modality. Despite the fact that HAART is very effective in suppressing HIV-1 replication and reducing the mortality of HIV/AIDS patients, it has become increasingly clear that HAART does not offer an ultimate cure to HIV/AIDS. The high cost of the HAART regimen has impeded its delivery to over 90% of the HIV/AIDS population in the world. This reality has urgently called for the need to develop inexpensive alternative anti-HIV/AIDS therapy. This need has further manifested by recent clinical trial failures in anti-HIV-1 vaccines and microbicides. . In the current study, we characterized a panel of extracts of traditional medicinal plants for their activities against HIV-1 replication. The aim of the present study was to evaluate the invitro anti- HIV activity of Eclipta alba plant extracts. Extracts were prepared from dried fruits in n-hexane, ethyl acetate and n butanol. Peripheral Blood Mononuclear Cells (PBMCs) isolated from healthy donors by ficoll-hypaque density gradient centrifugation method. A toxicity study was performed on all crude extracts by MTT assay using PBMCs isolated from whole blood. HIV-1 RT inhibition activity of the all solvent extracts of Eclipta alba was determined by a RetrsoSys HIV-1 RT activity kit (Innovagen, Sweden). The aerial parts of Eclipta alba extracts are shows anti-HIV-1 activity and this plant has great potential for developing useful drugs.

14.
Indian J Med Microbiol ; 2010 Jul-Sept; 28(3): 233-237
Article in English | IMSEAR | ID: sea-143704

ABSTRACT

Purpose: To investigate the difference between the abilities of Helicobacter pylori and Escherichia coli to induce expression of TNF-α in human peripheral blood mononuclear cells (PBMC). Materials and Methods: H pylori was isolated from gastric biopsy specimens. The mononuclear cells were isolated from human blood, cultured, and treated with either intact or sonicated E coli or H pylori, and mRNA expression for TNF-α was detected using semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). Results: TNF-α mRNA expression levels were significantly higher in PBMCs stimulated with E coli compared to those stimulated with H pylori at the same number and identical conditions (P < .001). The results also suggest that sonicated bacteria were significantly (P < .001) less stimulatory for PBMCs than intact bacteria for both E coli and H pylori. Conclusions: The ability of different H pylori strains isolated from biopsy samples to stimulate TNF-α from PBMCs was significantly lower than that of E coli. Sonicated bacteria, as compared to intact bacteria, was a very poor inducer of TNF-α mRNA expression, suggesting that the conformation of lipopolysaccharides (LPS) on the outer leaflet of the outer membrane is not totally conserved in sonicated bacteria.

15.
Academic Journal of Xi&#39 ; an Jiaotong University;(4): 206-212, 2008.
Article in Chinese | WPRIM | ID: wpr-844827

ABSTRACT

Objective: To evaluate the methylation status at CpG site -55 in the interferon-gamma (IFN-γ) gene promoter and its effect on IFN-γ expression in chronic hepatitis B. Method: The authors recruited 30 patients with HBeAg-positive chronic hepatitis B (CHB), 30 HBeAg-negative CHB patients, and 30 healthy blood donors. Pyrosequencing was used to determine the methylation status at CpG site -55 in the IFN-γ gene promoter following bisulfite treatment of DNA in peripheral blood, mononuclear cells (PBMCs). The expression of IFN-γ was analyzed by real-time RT-PCR and ELISA. HBV DNA in PBMCs was detected by nested PCR. Results: The methylation level at CpG site -55 in the IFN-γ gene promoter was significantly increased, resulting in subsequent down-regulation of the expression of this cytokine in CHB. The methylation level at CpG site -55 was significantly higher in HBeAg-positive patients than in HBeAg-negative ones (P < 0.01) and was also significantly higher in PBMCs from HBV DNA-positive patients than from HBV DNA-negative ones (P < 0.01); the methylation level at CpG site -55 was positively correlated with the amount of HBV DNA in serum (P < 0.01). Conclusion: IFN-γ gene expression appears to be regulated by methylation of the IFN-γ gene promoter in CHB; the methylation level at CpG site -55 is associated with HBV infection.

16.
Virologica Sinica ; (6): 330-338, 2008.
Article in Chinese | WPRIM | ID: wpr-407041

ABSTRACT

The ELISPOT assay is increasingly used for assessing cellular immune responses in clinical trials of HIV-1 or cancer vaccines. However, to date, data from clinical trials do not consistently show that immune responses are correlated with clinical endpoints. This is due in part to the lack of assay standardization and validation across laboratories and therefore, a quality control panel is required to establish competency and comparability amongst different laboratories. In this study peripheral blood mononuclear cells (PBMCs) from healthy individuals were screened and frozen in liquid nitrogen. The recovery and viability of the PBMCs and the frequencies of interferon (IFN)-γ-secreting cells after CEF peptide pool stimulation were detected after various intervals in seven different laboratories. The recovery and viability did not differ significantly after different intervals. Although the frequencies of IFN (interferon)-γ-secreting cells among thawed PBMCs (peripheral blood mononuclear cells) fluctuated after CEF peptide pool stimulation at different intervals, they were not significantly decreased compared with those among fresh PBMCs. However, the viabilities, recoveries and frequencies of IFN-γ-secreting cells differed significantly among the seven laboratories. Our results indicate that cryopreserved PBMCs could be used as a quality control panel for ELISPOT. However, the procedures for ELISPOT need to be standardized amongst different laboratories.

17.
Journal of Pharmaceutical Analysis ; (6): 206-212, 2008.
Article in Chinese | WPRIM | ID: wpr-621875

ABSTRACT

Objective To evaluate the methylation status at CpG site -55 in the interferon-gamma (IFN-7) gene promoter and its effect on IFN-7 expression in chronic hepatitis B. Method The authors recruited 30 patients with UBeAg-positive chronic hepatitis B (CHB), 30 HBeAg-negative CHB patients, and 30 healthy blood donors. Pyrosequeneing was used to determine the methylation status at CpG site -55 in the IFN-γ gene promoter following bisulfite treatment of DNA in peripheral blood mononuclear cells (PBMCs). The expression of IFN-γ was analyzed by real-time RT-PCR and ELISA. HBV DNA in PBMCs was detected by nested PCR. Results The methylation level at CpG site -55 in the IFN-γ gene promoter was significantly increased, resulting in subsequent down-regulation of the expression of this cytoldne in CHB. The methylation level at CpG site -55 was significantly higher in HBeAg-positive patients than in HBeAg-negative ones (P<0.01) and was also significantly higher in PBMCs from HBV DNA-positive patients than from HBV DNA-negative ones (P<0.01) ; the methylation level at CpG site -55 was positively correlated with the amount of HBV DNA in serum (P<0.01). Oonclusion IFN-γ gene expression appears to be regulated by methylation of the IFN-γ gene promoter in CHB; the methylation level at CpG site -55 is associated with HBV infection.

18.
China Journal of Traditional Chinese Medicine and Pharmacy ; (12)2005.
Article in Chinese | WPRIM | ID: wpr-564313

ABSTRACT

Objective:With proteomic approach,to find the differentially expressed proteins spots in the peripheral blood mononuclear cells(PBMCs) of healthy people and intracerebral hemorrhage patients with liver yang forming wind syndrome(LYFWS) treated by Zhenganxifeng decoction(ZGXFD) to investigate the possible proteins of LYFWS and the cure mechanism of ZGXFD.Methods: 10 intracerebral hemorrhage patients with LYFWS were treated by ZGXFD for 7-10 days.10 healthy people were selected taken as healthy controls.10ml venous blood was drawed from every subjects and PBMCs were isolated from blood by using lymphocytes separation medium.The total proteins was extracted from PBMCs.The total proteins from either intracerebral hemorrhage patients or healthy controls were prepared by means of immobilized pH gradient based on two-dimensional gel electrophoresis.After Coomassie brilliant blue G250 staining,gel-image analysis was performed by PDQuest.The differentially expressed proteins spots were identified by matrix-assisted laser desorption ionization-time-of-flight mass spectrometry(MALD I-TOF-MS).Results: Gel-image analysis revealed that there were 22 proteins spots expressed differentially.20 proteins spots were expressed differentially in the intracerebral hemorrhage patients with LYFWS as compared with healthy control,of which 8 proteins were down-regulated,7 proteins were up-regulated and 5 proteins were lost.After treated by ZGXFD,4 proteins were down-regulated,5 proteins were up-regulated and 2 proteins were lost,9 proteins weren’t expressed di erentially as compared with healthy control.8 of 20 di erentially expressed proteins spots in the intracerebral hemorrhage patients with LYFWS identi ed by MALDI-TOF-MS as compared with healthy control.These proteins are related to cell metabolism,signal transduction and so forth,of which 6 proteins were down-regulated,1 proteins were up-regulated and 1 proteins were lost.After treated by ZGXFD,2 proteins were down-regulated,1 proteins were lost,5 proteins weren’t expressed di erentially as compared with healthy control.Conclusion: 2-DE pro les of intracerebral hemorrhage patients with LYFWS in PBMCs had been established.The proteins of intracerebral hemorrhage patients with LYFWS had been identi ed,which were related to cell metabolism,signal transduction and so forth.This research presented that ZGXFD could adjust multiple di erentially expressed proteins of intracerebral hemorrhage patients with LYFWS.It was also indicated that the cure mechanism of ZGXFD was to regulate multitarget proteins.

19.
Journal of Chinese Physician ; (12)2001.
Article in Chinese | WPRIM | ID: wpr-524779

ABSTRACT

Objective To explore the expression of interleukin 13(IL-13) mRNA in peripheral blood mononuclear cells (PBMCs) and its association with serum C-reaction protein(CRP) in patients with ulcerative colitis(UC). Methods IL-13 mRNA expression in PBMCs was detected by RT-PCR in patients with UC(21 cases of active UC and 10 cases of quiescent UC)and 20 healthy subjects. At the same time, the serum level of CRP was detected by ELISA. Results The expression level of IL-13 mRNA in PBMCs of patients with active UC was significantly lower than that in PBMCs of patients with quiescent UC and healthy subjects (6.45?1.23 vs 14.72?2.12 or 15.17?2.38,P0.05). The expression level of IL-13 mRNA in PBMCs of active UC patients was negative correlation with serum level of CRP (r=-0.589,P

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