Objective：With the regular mixed lymphocytes culture （MLC） to detect the allograft rejection, the reactivity of the activated lymphocytes （primed lymphocytes） of a recipient shows sometimes increase and sometimes...Objective：With the regular mixed lymphocytes culture （MLC） to detect the allograft rejection, the reactivity of the activated lymphocytes （primed lymphocytes） of a recipient shows sometimes increase and sometimes decrease against the antigens from the donor, which is inconsistent with the clinical results. In order to establish a convenient method for testing the specificity of the activated lymphocytes in vitro, so as to know the rejection occurred or not by testing the existence of the specific activated lymphocytes against donor＇s HLA antigens in the recipient＇s peripheral blood. Methods： Anti-IL-2 neutralizing monoclonal antibody （anti-IL-2 N-mAb） and immunosuppressors were introduced in this test system in the presence of specific stimulators and activated lymphocytes. Results ： When the activated lymphocytes were chosen from the one-way MLC 4 d to undergo re-stimulation by specific stimulators, the activity of activated lymphocytes in the treatment group was suppressed significantly compared with that in the control group. The result of this test method is consistent with the biopsy in the clinical diagnosis of rejection. Conclusion：h suggests that the activated lymphocytes can be inactivated by specific antigens in certain conditions. This can be a useful tool to define the specificity of the activated lymphocytes.展开更多
Objective To investigate the role of TNF receptor-associated factor 2 (TRAF-2) and TRAF6 in CD40-induced nuclear factor-κB (NF-κB) signaling pathway and whether CD40 signaling requires TRAF2. Methods Human B cell li...Objective To investigate the role of TNF receptor-associated factor 2 (TRAF-2) and TRAF6 in CD40-induced nuclear factor-κB (NF-κB) signaling pathway and whether CD40 signaling requires TRAF2. Methods Human B cell lines were transfected with plasmids expressing wild type TRAF2 or dominant negative TRAF2,TRAF2-shRNA,or TRAF6-shRNA. The activation of NF-κB was detected by Western blot,kinase assay,transfactor enzyme-linked immunosorbent assay (ELISA),and fluorescence resonance energy transfer (FRET). Analysis of the role of TRAF-2 and TRAF-6 in CD40-mediated NF-κB activity was examined following stimulation with recombinant CD154. Results TRAF2 induced activity of IκB-kinases (IKKα,IKKi/ε),phosphorylation of IκBα,as well as nuclear translocation and phosphorylation of p65/RelA. In contrast,TRAF6 strongly induced NF-κB activation and nuclear translocation of p65 as well as p50 and c-Rel. Engagement of CD154-induced nuclear translocation of p65 was inhibited by a TRAF6-shRNA,but conversely was enhanced by a TRAF2-shRNA. Examination of direct interactions between CD40 and TRAFs by FRET documented that both TRAF2 and TRAF6 directly interacted with CD40. However,the two TRAFs competed for CD40 binding. Conclusions These results indicate that TRAF2 can signal in human B cells,but it is not essential for CD40-mediated NF-κB activation. Moreover,TRAF2 can compete with TRAF6 for CD40 binding,and thereby limit the capacity of CD40 engagement to induce NF-κB activation.展开更多
Objective This study was performed to investigate the effect of high mobility group box-1 protein(HMGB1)on immune function of human T lymphocytes in vitro and explore its potential role in cell-mediated immune dysfunc...Objective This study was performed to investigate the effect of high mobility group box-1 protein(HMGB1)on immune function of human T lymphocytes in vitro and explore its potential role in cell-mediated immune dysfunction.Methods Fresh blood was obtained from healthy adult volunteers and peripheral blood mononuclear cells(PBMCs)were isolated,then rhHMGB1 was added to PBMCs.Four-color flow cytometric(FCM)analysis was used for the measurement of intracellular cytokine including interleukin IL-4 and interferon IFN-?ELISA kits were employed for the determination of IL-2 and sIL-2R protein levels in cell culture supernatants.Results(1)Different stimulating time and dosage of rhHMGB1 did not alter the number of IFN-αpositive cells(Th1).rhHMGB1 stimulation provoked a dose-dependent and time-dependent increase in Th2 subset and decrease in ratio of Th1 to Th2.(2)Compared with the untreated cells,when the cells were coincubated with rhHMGB1(10-100ng/ml)for 12 hrs,protein release of IL-2 and sIL-2R were significantly up-regulated.At 48 hrs,in contrast,protein production was relatively lower in cells after exposure to 100-1000 ng/ml rhHMGB1.Conclusions These findings demonstrated that HMGB1 has a dual influence on immune functions of human T lymphocytes.展开更多
The objective of this study was to isolate and identify the intracellular bioactive peptides from mouse lymphocytes before and after lipopolysaccharide(LPS)stimulation,to explore novel peptides and to research the bio...The objective of this study was to isolate and identify the intracellular bioactive peptides from mouse lymphocytes before and after lipopolysaccharide(LPS)stimulation,to explore novel peptides and to research the bioactive function.Mouse spleen lymphocytes were isolated and cultured with LPS stimulation(experimental group)or not(control group)to collect intracellular peptides.Totally 385 peptides were analyzed by nanoliter liquid phase-Q Exactive quadrupole ultra-high resolution orbitrap mass spectrometer(Nano LC-Q Exactive Plus)and identifi ed by PEAKS X software.After compared with peptides reported,131 novel peptides were discovered,which then were predicted bioactivity by Peptide Ranker and 6 peptides with high bioactivity were predicted function by BIOPEP-UMW database.Prediction data showed that they may have dipeptidyl peptidase IV(DPP-IV)inhibitory activity.Finally,two peptides showed better potent inhibition were verifi ed with competitive and noncompetitive modes.展开更多
Objective To contrast the roles of Asparagus officinalis polysaccharide on erythrocyte of S180 mice played in immunological reaction of lymphocytes.To study the effect of Asparagus officinalis polysaccharide on the er...Objective To contrast the roles of Asparagus officinalis polysaccharide on erythrocyte of S180 mice played in immunological reaction of lymphocytes.To study the effect of Asparagus officinalis polysaccharide on the erythrocyte function of S180 mice.Methods Suspensions of lymphocytes(1×106/mL)and autologous plasma were respectively separated from anticoaguted whole blood of healthy mice with the lymphocyte separation medium.The erythrocytes(1×108/mL)were separated from whole blood of Asparagus officinalis polysaccharide mice.Using the autologous plasm as reactive medium,the role of erythrocytes in regulating the immunological reaction of lymphocytes was appraised.The expression of CD25 on lymphocytes was detected using flow cytometry.Results The immunogical regulating ability of erythrocyte in mice with control groups is much lower than that of normal groups,and the immunogical regulating ability of erythrocyte in mice with Asparagus officinalis polysaccharide groups is much higher than that of control groups.Conclusions According to the effects of erythrocyte CD35 on the immuno-response of lymphocyte and the different of the expression of CD25 on lymphocytes,we prove that Asparagus officinalis polysaccharide can improve the erythrocyte function of S180 mice.展开更多
Neuroleukin(NLK),a protein(Mr 56000),acts on both the nervous and the im-mune systems and its expression is regulated at mRNA levels.One hundred fifty Sprague-Daw-ley(SD)male rats were divided into 5 groups:control gr...Neuroleukin(NLK),a protein(Mr 56000),acts on both the nervous and the im-mune systems and its expression is regulated at mRNA levels.One hundred fifty Sprague-Daw-ley(SD)male rats were divided into 5 groups:control group(N)and groups of cold-exposurefor 1,2,3 and 4 weeks(ClW,C2W,C3W and C4W).With RNA dot blot experiments,changes of NLK mRNA levels in both neuroendocrine system(pituitary and hypothalamus)andimmune system(splenic lymphocytes,LC)of SD male rats during cold acclimation presentedsimilar responses.C/N ratios of scan gray intensity level in RNA dots are presented as follows:there was a rise in C/N ratio to 1.63～11.31 in CIW,a fall to 0.63 in C2W and a rise again to1.42～21.09 in C3W.It might thus be regarded as a general response to cold surroundings.Itwas reported that SDS-PAGE autoradiogram of synthetic proteins in LC and secretive proteinsin supernatant cultured in vitro showed that NLK content in LC was not parallel to its mRNAlevel because NLK was extremely meager over the 1-week cold exposure whereas it increasedobviously over the 3-and 4-week cold exposure.This is probably a major cause of why im-munoglobulin synthesis in cold-acclimated animals was greatly enlarged.展开更多
Glioma-infiltrating lymphocytes(GIL)were isolated from 9 surgical biopsy specimens of pri-mary brain gliomas using mechanical and enzymatic digestion and discontinuous density gradientcentfifugation.Durng culture in t...Glioma-infiltrating lymphocytes(GIL)were isolated from 9 surgical biopsy specimens of pri-mary brain gliomas using mechanical and enzymatic digestion and discontinuous density gradientcentfifugation.Durng culture in the presence of interleukin-2(IL-2)for a period of four weeks,GIL were expanded by 48.4-fold on the avea-age,even up to 118-fold.GIL activated by IL-2 hadspcific cytolytic activity against autologous glioma cells.Analysis of cell surface phenotypes offreshly isolated GIL showed that CD3^+ cells were 71.0±11.9％,CD4^+ cells 34.2±6.1％ and CD8^+cells 37.0±7.6％.Ability of IL-2-activated GIL to secrete γ-interferon(γ-IFN)was significantlyhigher than that of freshly isolated GIL and autologous peripheral blood lymphocytes(PBL).Theresults suggest that GIL have many advantages as an adoptive immunotherapy for patients withgliomas and as a new type of antitumor immune effector.展开更多
The dose-response curves obtained by premature condensed chromosome (PCC)and conventional cellular genetic methods can be represented by two linear equations. The ratiO of the slopes, Kpcc/KM,, is about 28. In compari...The dose-response curves obtained by premature condensed chromosome (PCC)and conventional cellular genetic methods can be represented by two linear equations. The ratiO of the slopes, Kpcc/KM,, is about 28. In comparison to the conventional method, the PCC method has many advantages; e,g. it is faster, simpler, more sensitive and accurate. Its significance in the study of radiation damage is also discussed.展开更多
Studied features of cellular and humoral factors of immune system and activation markers of lymphocytes at breast cancer patients.Analyzed biophysical parameters of lymphocyte superficial membranes of periphery blood ...Studied features of cellular and humoral factors of immune system and activation markers of lymphocytes at breast cancer patients.Analyzed biophysical parameters of lymphocyte superficial membranes of periphery blood of breast cancer patients.Features of immune system reaction were:expressed Т - cellular immunodeficiency,deficiency of CD4+Т-helpers/inducers against raised expression of СD8+Т-lymphocyte,suppression of immunoregulatory index,imbalance of immunity humoral link,activity of expression CD16 + and CD56 + on natural killers and rising of expression CD38 + and CD95 + on lymphocytes.Studying of biophysical parameters of lymphocytes superficial membranes has allowed to reveal statistically significant changes characterized by intensifying of electric field and rising ofmicroviscosity,obviously leading to disappearance or weakening of intercellular interactions.展开更多
Objective To investigate the quantities of bone marrow CD5+ B lymphocytes in the patients with autoimmune hemocytopenia and the relationship between quantities of CD5+ B lymphocytes and clinical or laboratorial parame...Objective To investigate the quantities of bone marrow CD5+ B lymphocytes in the patients with autoimmune hemocytopenia and the relationship between quantities of CD5+ B lymphocytes and clinical or laboratorial parameters. Methods Quantities of CD5+ B lymphocytes in the bone marrow of 14 patients with autoimmune hemolytic anemia (AIHA) or Evans syndrome, 22 immunorelated pancytopenia (IRP) patients, and 10 normal controls were assayed by flow cytometry. The correlation between their clinical or laboratorial parameters and CD5+ B lymphocytes was analyzed. Results The quantity of CD5+ B lymphocytes of AIHA/Evans syndrome (34.64%±19.81%) or IRP patients (35.81%±16.83%) was significantly higher than that of normal controls (12.00%±1.97%, P<0.05). However, there was no significant difference between AIHA/Evans syndrome and IRP patients (P>0.05). In all hemocytopenic patients, the quantity of bone marrow CD5+ B lymphocytes showed significantly negative correlation with serum complement C3 level (r=-0.416, P<0.05). In the patients with AIHA/Evans syndrome, the quantity of bone marrow CD5+ B lymphocytes showed significantly positive correlation with serum indirect bilirubin level (r=1.00, P<0.05). In Evans syndrome patients, the quantity of CD5+ B lymphocytes in bone marrow showed significantly positive correlation with platelet-associated immunoglobulin G (r=0.761, P<0.05) and platelet-associated immunoglobulin M (r=0.925, P<0.05). The quantity of CD5+ B lymphocytes in bone marrow of all hemocytopenic patients showed significantly negative correlation with treatment response (tau-b=-0.289, P<0.05), but had no correlation with colony forming unit-erythroid (r=-0.205, P>0.05) or colony forming unit-granulocyte-macrophage colonies (r=-0.214, P>0.05). Conclusions The quantity of bone marrow CD5+ B lymphocytes in the patients with autoimmune hemocytopenia significantly increases and is correlated with disease severity and clinical response, which suggest that CD5+ B lymphocytes might play an important role in the pathogenesis of autoimmune hemocytopenia.展开更多
Background: In clinical studies, the findings on sulfur mustard(SM) toxicity for CD3+CD4+ and CD3+CD8+ T lymphocyte subsets are contradictory. In animal experiments, the effect of SM on the T cell number and prolifera...Background: In clinical studies, the findings on sulfur mustard(SM) toxicity for CD3+CD4+ and CD3+CD8+ T lymphocyte subsets are contradictory. In animal experiments, the effect of SM on the T cell number and proliferation is incompatible and is even the opposite of the results in human studies. In this study, we observed the dynamic changes of T lymphocytes in the first week in a high-dose SM-induced model.Methods: Mice were exposed to SM by subcutaneous injection(20 mg/kg) and were sacrificed 4 h, 24 h, 72 h and 168 h later. Spleen T lymphocyte proliferation was evaluated by 3H-Td R. Flow cytometric analysis was used to observe the percentage of CD3+CD4+ and CD3+CD8+ T lymphocyte subsets. The IL-1e assayed using the Luminex method. DNA damage in bone marrow ceβ, IL-6, IL-10 and TNF-lls was observed with α levels in plasma werthe single cell gel electrophoresis technique(SCGE).Results: SM continuously inhibited the proliferation of lymphocytes for 7 days, and there was a significant rebound of Con A-induced T lymphocyte proliferation only at 24 h. The percentage of CD3+CD4+ and CD3+CD8+ lymphocytes was upregulated, which was accompanied by increased IL-1β and TNF-creased in the PG group at 4 h. The peak of lymphocytic apoptα and decreased IL-10. The IL-6 level was gradually deosis and DNA damage occurred at 24 h and 72 h, respectively. Conclusion: Our results show that SM significantly inhibited T lymphocyte proliferation as well as induced CD3+CD4+ and CD3+CD8+ upregulation. SM intoxication also significantly increased the levels of pro-inflammatory cytokines(IL-1β, IL-6 and TNF-α) and inhibited the level of anti-inflammatory cytokine IL-10. Our results may partly be due to the significant SM induced significant apoptosis and necrosis of lymphocytes as well as DNA damage of bone marrow cells. The results provided a favorable evaluation of SM immune toxicity in an animal model.展开更多
Tumor-infiltrating lymphocytes (TIL)isolated from metastatic lymph nodes in patients with nonoperable advanced gastric cancer were induced to become LAK-like cytotoxic activrty of TIL after in vitro culture with rlL-2...Tumor-infiltrating lymphocytes (TIL)isolated from metastatic lymph nodes in patients with nonoperable advanced gastric cancer were induced to become LAK-like cytotoxic activrty of TIL after in vitro culture with rlL-2.Twenty-three patients with advanced gastric cancer were treated by intravenously transfer of autologous TIL combined with rlL-2. The tumor forus disappeared (complete remission, CR) in 3 patients (13. 0%) and significantly decreased (partial remission, PR) in 5 patients (21. 7%). Fifteen patients did not respond to the treatment. The amount of soluable IL-2 receptor in serum was significantly decreased after treatment, the cytotoxicity of NK cells and OT test were significantly increased. No significant difference in CD4/CD8 was found between before and after treatment. No serious side effect was obseved in the treatment.展开更多
Objective: To analyze the effect of lymphocytes on proliferation of keratinocytes in patients with psoriasis. Methods: Lymphocytes in lesion and peripheral blood were isolated and amplified, then cultured together wit...Objective: To analyze the effect of lymphocytes on proliferation of keratinocytes in patients with psoriasis. Methods: Lymphocytes in lesion and peripheral blood were isolated and amplified, then cultured together with normal keratinocytes. By MTT method, the living cells were quantified in the mixed culture. Results: Compared with normal controls, lymphocytes from lesion and peripheral blood of psoriasis both promote the proliferation of keratinocytes (P<0. 01 and P<0. 05 respectively). The concentrations of IL-2 and IFN-7 in the mixture of lesion lymphocytes and keratinocytes were significantly higher than that of controls. Tripterygium glycosides inhibited this promotion. Conclusion: Lymphocytes in patients with psoriasis (mainly Th1 cell) play an important role in proliferation of keratinocytes. This psoriasis cell model is useful for studies on signal transduction in psoriasis.展开更多
The susceptibility was compared between murine bone marr ow hemopoietic cells and splenic lymphocytes to four major factors of cryopreservation process: toxicity of dimethyl sulfoxide (DMSO), cooling rate,Tris-NH4Cl t...The susceptibility was compared between murine bone marr ow hemopoietic cells and splenic lymphocytes to four major factors of cryopreservation process: toxicity of dimethyl sulfoxide (DMSO), cooling rate,Tris-NH4Cl treatment, and dilution after thawing. When the concentration of DMSO was over 20%, the proliferative function or viability of both kinds of cells decreased markedly. Injury to hemopoietic stem cells and lymphoeytes was found more severe in rapid cooling than in slow cooling, and the intensity of iujury seemed alike under the same fast cooling condition. Pretreatment of bone marrow cells with Tris-NH4Cl did not significantly accentuate the injury of frozen-thawed hemopoietic stem cells. Dilution of the frozen-thawed cells in ice bath or at a fast rate was harmful to both cells. No significant differential susceptibilities of the two kinds of cells to each factor were found. These results indicate that selective destruction or inactivation of lymphocytes by cold treatment among bone marrow cells may be impossible.展开更多
A series of artificial membranes called small unilaminar vesicles (SUVs) comprising different molar ratios of phosphatidylcholine (PC)/cholesterol (Ch) were constructed and exogenous PC and Ch were transferred into hu...A series of artificial membranes called small unilaminar vesicles (SUVs) comprising different molar ratios of phosphatidylcholine (PC)/cholesterol (Ch) were constructed and exogenous PC and Ch were transferred into human peripheral blood lymphocytes through incubating the cells with SUVs in order to study the effects of the changed constituents of membrane lipids on the expression of CD 71 in lymphocytes, the binding characteristics of CD 71 molecules with its ligand and its internalization. It was found that after the treatment of SUVs, the Ch content in the plasmic membrane was increased with a decrease of PC/Ch ratio. After incubation with ConA for 48 h,the percentage of CD 71-positive lymphocytes increased with the increase of the availab1e binding sites for CD 71 to its ligand, but the affinity of CD 71 with its ligand was lowered- When Ch synthesis was blocked with HMG-CoAreductase inhibitor (Lovastatin), the number of CD71 molecules was obviously reducce but the afinity of CD 71with its ligand elevated- It is believed that that the construction of the lipid environment can directly affect the properties of CD 71 molecules. Less fluid phase of the lipid environment might be suitable to the move;nent of CD 71 in the membrane but minimize the tendency for CD 71 to maintain a con formation of high affinity state.展开更多
Objective The purpose of our study is to observe the voltage-gated potassium channel Kvl.3 expressed on CD4＋CD28~ T cells from the peripheral blood of acute coronary syndrome （ACS） patients by the patch clamp techn...Objective The purpose of our study is to observe the voltage-gated potassium channel Kvl.3 expressed on CD4＋CD28~ T cells from the peripheral blood of acute coronary syndrome （ACS） patients by the patch clamp technique. Methods Kvl.3 potassium channels expression from 17 patients with ACS and 11 healthy age-match controls was detected in single cell（CD4＋CD28null T cells and CD4＋CD28＋T cells） by fluorescence mieroscopy and patch clamp. Results The percentage of CD4＋CD28mullT cells was higher in the ACS patients [（6.97±2.05）%] than that in the controls [（1.38±0.84）%, P〈0.05]. The concentration of hsCRP was directly correlated with the number of the CD4~CD28nul~ T cells in the ACS patients （r=0.52, P〈0,05）. The conductance （6.89±1.17ns vs 3.36±0.66ns）, dens （1.95±0.80 μm2 vs 1.13±0.57 μm2） and numbers （574.5±97.6 n/cell vs. 280.3±55.3 n/cell） of the Kv1.3 channels on the CIM＋CD28null T cells were significantly higher than those on the CD4＋CD28＋ T cells （all P〈0.01） in ACS patients, but were similar on CD4＋CD28＋T betweenACS patients and controls. Conclusion The CD4＋CD28nullT cells and the numbers of Kvl.3 channels on the CD4＋CD28nullT cells from patients with ACS are significantly upregulated and might contribute to the pathogenesis of ACS （d Geriatr Cardio12010; 7：40-46）.展开更多
To put forward a method of automatic recognition of the micronuclei on computerized image in bi-nucleated lymphocytes. Methods: The procedure was as the follows: ① The nuclei and cytoplasm images were automatically e...To put forward a method of automatic recognition of the micronuclei on computerized image in bi-nucleated lymphocytes. Methods: The procedure was as the follows: ① The nuclei and cytoplasm images were automatically extracted from the background in order that the nuclei and cytoplasm images could be preprocessed individually.② Noise in two images was reduced, the edges in the nuclei image smoothed and then the big-point noise in the cytoplasm image taken off. ③ The nuclei was identified and the single cytoplasm acquired individually form nuclei and cytoplasm image. ④ The two preprocessed images were put together to identify the bi-nuclei and micronuclei form the overlapped map. Results: Many new models for image processing have been established. Preliminary measurement showed that the accuracy of recognition reached a high percentage of 85%. Conclusion: It is of great value to put the automatic recognition system into use.展开更多
Objective Congestive heart failure （CHF） is the final common pathway of various heart diseases.Calcineurin,a calcium/calmodulindependent phosphatase consisting of a catalytic subanit A （CnA） and a regulatory calci...Objective Congestive heart failure （CHF） is the final common pathway of various heart diseases.Calcineurin,a calcium/calmodulindependent phosphatase consisting of a catalytic subanit A （CnA） and a regulatory calcium-binding subunit B （CnB）,is activated in heart failure.This study aimed to investigate the relationship between mRNA level of calcineurin in circulating T-lymphocyte and that in myocardium in patients with CHF. Methods A total of 38 patients with CHF （aged from 29 to 62 years） were included in this study.The mRNA levels of alpha-and beta-isoform of CnA in left ventricular anterior papillary muscle and peripheral lymphocytes were determined by semi-quantitative reverse transcription polymerase chain reaction.Pearson linear correlation analysis was performed,and difference was considered statistically significant at a P value 〈0.05. Results Calcineurin mRNA levels in lymphocytes were positively correlated with those in myocardium （for CnA-alpha mRNA,r=0.820;for CnA-beta mRNA,r=0.875;both P〈0.01）.CnA-beta mRNA levels in both circulating lymphocytes and myocardium increased significantly with increasing NYHA class （r=0.877 for peripheral blood and r=0.805 for cardiac muscle;both P〈0.01）. Conclusions The mRNA level of CnA-beta in circulating lymphocytes is positively correlated with that in myocardium and is a promising marker for the severity of cardiac dysfunction in patients with CHF.展开更多
文摘Objective：With the regular mixed lymphocytes culture （MLC） to detect the allograft rejection, the reactivity of the activated lymphocytes （primed lymphocytes） of a recipient shows sometimes increase and sometimes decrease against the antigens from the donor, which is inconsistent with the clinical results. In order to establish a convenient method for testing the specificity of the activated lymphocytes in vitro, so as to know the rejection occurred or not by testing the existence of the specific activated lymphocytes against donor＇s HLA antigens in the recipient＇s peripheral blood. Methods： Anti-IL-2 neutralizing monoclonal antibody （anti-IL-2 N-mAb） and immunosuppressors were introduced in this test system in the presence of specific stimulators and activated lymphocytes. Results ： When the activated lymphocytes were chosen from the one-way MLC 4 d to undergo re-stimulation by specific stimulators, the activity of activated lymphocytes in the treatment group was suppressed significantly compared with that in the control group. The result of this test method is consistent with the biopsy in the clinical diagnosis of rejection. Conclusion：h suggests that the activated lymphocytes can be inactivated by specific antigens in certain conditions. This can be a useful tool to define the specificity of the activated lymphocytes.
基金Supported by Key Projects of the National Science & Technology Pillar Program in the Eleventh Five-year Plan Period (2008-BAI59B02)
文摘Objective To investigate the role of TNF receptor-associated factor 2 (TRAF-2) and TRAF6 in CD40-induced nuclear factor-κB (NF-κB) signaling pathway and whether CD40 signaling requires TRAF2. Methods Human B cell lines were transfected with plasmids expressing wild type TRAF2 or dominant negative TRAF2,TRAF2-shRNA,or TRAF6-shRNA. The activation of NF-κB was detected by Western blot,kinase assay,transfactor enzyme-linked immunosorbent assay (ELISA),and fluorescence resonance energy transfer (FRET). Analysis of the role of TRAF-2 and TRAF-6 in CD40-mediated NF-κB activity was examined following stimulation with recombinant CD154. Results TRAF2 induced activity of IκB-kinases (IKKα,IKKi/ε),phosphorylation of IκBα,as well as nuclear translocation and phosphorylation of p65/RelA. In contrast,TRAF6 strongly induced NF-κB activation and nuclear translocation of p65 as well as p50 and c-Rel. Engagement of CD154-induced nuclear translocation of p65 was inhibited by a TRAF6-shRNA,but conversely was enhanced by a TRAF2-shRNA. Examination of direct interactions between CD40 and TRAFs by FRET documented that both TRAF2 and TRAF6 directly interacted with CD40. However,the two TRAFs competed for CD40 binding. Conclusions These results indicate that TRAF2 can signal in human B cells,but it is not essential for CD40-mediated NF-κB activation. Moreover,TRAF2 can compete with TRAF6 for CD40 binding,and thereby limit the capacity of CD40 engagement to induce NF-κB activation.
基金This study was supported by the National Natural Science Foundation of China(No.30672178)National Basic Research Program of China(No.2005CB522602)the National Natural Science Outstanding Youth Foundation of China(No.30125020).
文摘Objective This study was performed to investigate the effect of high mobility group box-1 protein(HMGB1)on immune function of human T lymphocytes in vitro and explore its potential role in cell-mediated immune dysfunction.Methods Fresh blood was obtained from healthy adult volunteers and peripheral blood mononuclear cells(PBMCs)were isolated,then rhHMGB1 was added to PBMCs.Four-color flow cytometric(FCM)analysis was used for the measurement of intracellular cytokine including interleukin IL-4 and interferon IFN-?ELISA kits were employed for the determination of IL-2 and sIL-2R protein levels in cell culture supernatants.Results(1)Different stimulating time and dosage of rhHMGB1 did not alter the number of IFN-αpositive cells(Th1).rhHMGB1 stimulation provoked a dose-dependent and time-dependent increase in Th2 subset and decrease in ratio of Th1 to Th2.(2)Compared with the untreated cells,when the cells were coincubated with rhHMGB1(10-100ng/ml)for 12 hrs,protein release of IL-2 and sIL-2R were significantly up-regulated.At 48 hrs,in contrast,protein production was relatively lower in cells after exposure to 100-1000 ng/ml rhHMGB1.Conclusions These findings demonstrated that HMGB1 has a dual influence on immune functions of human T lymphocytes.
文摘The objective of this study was to isolate and identify the intracellular bioactive peptides from mouse lymphocytes before and after lipopolysaccharide(LPS)stimulation,to explore novel peptides and to research the bioactive function.Mouse spleen lymphocytes were isolated and cultured with LPS stimulation(experimental group)or not(control group)to collect intracellular peptides.Totally 385 peptides were analyzed by nanoliter liquid phase-Q Exactive quadrupole ultra-high resolution orbitrap mass spectrometer(Nano LC-Q Exactive Plus)and identifi ed by PEAKS X software.After compared with peptides reported,131 novel peptides were discovered,which then were predicted bioactivity by Peptide Ranker and 6 peptides with high bioactivity were predicted function by BIOPEP-UMW database.Prediction data showed that they may have dipeptidyl peptidase IV(DPP-IV)inhibitory activity.Finally,two peptides showed better potent inhibition were verifi ed with competitive and noncompetitive modes.
文摘Objective To contrast the roles of Asparagus officinalis polysaccharide on erythrocyte of S180 mice played in immunological reaction of lymphocytes.To study the effect of Asparagus officinalis polysaccharide on the erythrocyte function of S180 mice.Methods Suspensions of lymphocytes(1×106/mL)and autologous plasma were respectively separated from anticoaguted whole blood of healthy mice with the lymphocyte separation medium.The erythrocytes(1×108/mL)were separated from whole blood of Asparagus officinalis polysaccharide mice.Using the autologous plasm as reactive medium,the role of erythrocytes in regulating the immunological reaction of lymphocytes was appraised.The expression of CD25 on lymphocytes was detected using flow cytometry.Results The immunogical regulating ability of erythrocyte in mice with control groups is much lower than that of normal groups,and the immunogical regulating ability of erythrocyte in mice with Asparagus officinalis polysaccharide groups is much higher than that of control groups.Conclusions According to the effects of erythrocyte CD35 on the immuno-response of lymphocyte and the different of the expression of CD25 on lymphocytes,we prove that Asparagus officinalis polysaccharide can improve the erythrocyte function of S180 mice.
文摘Neuroleukin(NLK),a protein(Mr 56000),acts on both the nervous and the im-mune systems and its expression is regulated at mRNA levels.One hundred fifty Sprague-Daw-ley(SD)male rats were divided into 5 groups:control group(N)and groups of cold-exposurefor 1,2,3 and 4 weeks(ClW,C2W,C3W and C4W).With RNA dot blot experiments,changes of NLK mRNA levels in both neuroendocrine system(pituitary and hypothalamus)andimmune system(splenic lymphocytes,LC)of SD male rats during cold acclimation presentedsimilar responses.C/N ratios of scan gray intensity level in RNA dots are presented as follows:there was a rise in C/N ratio to 1.63～11.31 in CIW,a fall to 0.63 in C2W and a rise again to1.42～21.09 in C3W.It might thus be regarded as a general response to cold surroundings.Itwas reported that SDS-PAGE autoradiogram of synthetic proteins in LC and secretive proteinsin supernatant cultured in vitro showed that NLK content in LC was not parallel to its mRNAlevel because NLK was extremely meager over the 1-week cold exposure whereas it increasedobviously over the 3-and 4-week cold exposure.This is probably a major cause of why im-munoglobulin synthesis in cold-acclimated animals was greatly enlarged.
文摘Glioma-infiltrating lymphocytes(GIL)were isolated from 9 surgical biopsy specimens of pri-mary brain gliomas using mechanical and enzymatic digestion and discontinuous density gradientcentfifugation.Durng culture in the presence of interleukin-2(IL-2)for a period of four weeks,GIL were expanded by 48.4-fold on the avea-age,even up to 118-fold.GIL activated by IL-2 hadspcific cytolytic activity against autologous glioma cells.Analysis of cell surface phenotypes offreshly isolated GIL showed that CD3^+ cells were 71.0±11.9％,CD4^+ cells 34.2±6.1％ and CD8^+cells 37.0±7.6％.Ability of IL-2-activated GIL to secrete γ-interferon(γ-IFN)was significantlyhigher than that of freshly isolated GIL and autologous peripheral blood lymphocytes(PBL).Theresults suggest that GIL have many advantages as an adoptive immunotherapy for patients withgliomas and as a new type of antitumor immune effector.
文摘The dose-response curves obtained by premature condensed chromosome (PCC)and conventional cellular genetic methods can be represented by two linear equations. The ratiO of the slopes, Kpcc/KM,, is about 28. In comparison to the conventional method, the PCC method has many advantages; e,g. it is faster, simpler, more sensitive and accurate. Its significance in the study of radiation damage is also discussed.
文摘Studied features of cellular and humoral factors of immune system and activation markers of lymphocytes at breast cancer patients.Analyzed biophysical parameters of lymphocyte superficial membranes of periphery blood of breast cancer patients.Features of immune system reaction were:expressed Т - cellular immunodeficiency,deficiency of CD4+Т-helpers/inducers against raised expression of СD8+Т-lymphocyte,suppression of immunoregulatory index,imbalance of immunity humoral link,activity of expression CD16 + and CD56 + on natural killers and rising of expression CD38 + and CD95 + on lymphocytes.Studying of biophysical parameters of lymphocytes superficial membranes has allowed to reveal statistically significant changes characterized by intensifying of electric field and rising ofmicroviscosity,obviously leading to disappearance or weakening of intercellular interactions.
文摘Objective To investigate the quantities of bone marrow CD5+ B lymphocytes in the patients with autoimmune hemocytopenia and the relationship between quantities of CD5+ B lymphocytes and clinical or laboratorial parameters. Methods Quantities of CD5+ B lymphocytes in the bone marrow of 14 patients with autoimmune hemolytic anemia (AIHA) or Evans syndrome, 22 immunorelated pancytopenia (IRP) patients, and 10 normal controls were assayed by flow cytometry. The correlation between their clinical or laboratorial parameters and CD5+ B lymphocytes was analyzed. Results The quantity of CD5+ B lymphocytes of AIHA/Evans syndrome (34.64%±19.81%) or IRP patients (35.81%±16.83%) was significantly higher than that of normal controls (12.00%±1.97%, P<0.05). However, there was no significant difference between AIHA/Evans syndrome and IRP patients (P>0.05). In all hemocytopenic patients, the quantity of bone marrow CD5+ B lymphocytes showed significantly negative correlation with serum complement C3 level (r=-0.416, P<0.05). In the patients with AIHA/Evans syndrome, the quantity of bone marrow CD5+ B lymphocytes showed significantly positive correlation with serum indirect bilirubin level (r=1.00, P<0.05). In Evans syndrome patients, the quantity of CD5+ B lymphocytes in bone marrow showed significantly positive correlation with platelet-associated immunoglobulin G (r=0.761, P<0.05) and platelet-associated immunoglobulin M (r=0.925, P<0.05). The quantity of CD5+ B lymphocytes in bone marrow of all hemocytopenic patients showed significantly negative correlation with treatment response (tau-b=-0.289, P<0.05), but had no correlation with colony forming unit-erythroid (r=-0.205, P>0.05) or colony forming unit-granulocyte-macrophage colonies (r=-0.214, P>0.05). Conclusions The quantity of bone marrow CD5+ B lymphocytes in the patients with autoimmune hemocytopenia significantly increases and is correlated with disease severity and clinical response, which suggest that CD5+ B lymphocytes might play an important role in the pathogenesis of autoimmune hemocytopenia.
基金supported by grants from the military medical science foundation projects (08G142)Chinese scientific and technological major special project (2009ZXJ09002-012, 2013ZX09J13103-01B and 2014ZX09J14103-03A)state key laboratory of toxicology and medical countermeasures
文摘Background: In clinical studies, the findings on sulfur mustard(SM) toxicity for CD3+CD4+ and CD3+CD8+ T lymphocyte subsets are contradictory. In animal experiments, the effect of SM on the T cell number and proliferation is incompatible and is even the opposite of the results in human studies. In this study, we observed the dynamic changes of T lymphocytes in the first week in a high-dose SM-induced model.Methods: Mice were exposed to SM by subcutaneous injection(20 mg/kg) and were sacrificed 4 h, 24 h, 72 h and 168 h later. Spleen T lymphocyte proliferation was evaluated by 3H-Td R. Flow cytometric analysis was used to observe the percentage of CD3+CD4+ and CD3+CD8+ T lymphocyte subsets. The IL-1e assayed using the Luminex method. DNA damage in bone marrow ceβ, IL-6, IL-10 and TNF-lls was observed with α levels in plasma werthe single cell gel electrophoresis technique(SCGE).Results: SM continuously inhibited the proliferation of lymphocytes for 7 days, and there was a significant rebound of Con A-induced T lymphocyte proliferation only at 24 h. The percentage of CD3+CD4+ and CD3+CD8+ lymphocytes was upregulated, which was accompanied by increased IL-1β and TNF-creased in the PG group at 4 h. The peak of lymphocytic apoptα and decreased IL-10. The IL-6 level was gradually deosis and DNA damage occurred at 24 h and 72 h, respectively. Conclusion: Our results show that SM significantly inhibited T lymphocyte proliferation as well as induced CD3+CD4+ and CD3+CD8+ upregulation. SM intoxication also significantly increased the levels of pro-inflammatory cytokines(IL-1β, IL-6 and TNF-α) and inhibited the level of anti-inflammatory cytokine IL-10. Our results may partly be due to the significant SM induced significant apoptosis and necrosis of lymphocytes as well as DNA damage of bone marrow cells. The results provided a favorable evaluation of SM immune toxicity in an animal model.
文摘Tumor-infiltrating lymphocytes (TIL)isolated from metastatic lymph nodes in patients with nonoperable advanced gastric cancer were induced to become LAK-like cytotoxic activrty of TIL after in vitro culture with rlL-2.Twenty-three patients with advanced gastric cancer were treated by intravenously transfer of autologous TIL combined with rlL-2. The tumor forus disappeared (complete remission, CR) in 3 patients (13. 0%) and significantly decreased (partial remission, PR) in 5 patients (21. 7%). Fifteen patients did not respond to the treatment. The amount of soluable IL-2 receptor in serum was significantly decreased after treatment, the cytotoxicity of NK cells and OT test were significantly increased. No significant difference in CD4/CD8 was found between before and after treatment. No serious side effect was obseved in the treatment.
基金Supported by the National Natural Science Foundation of China (No. 39970684)
文摘Objective: To analyze the effect of lymphocytes on proliferation of keratinocytes in patients with psoriasis. Methods: Lymphocytes in lesion and peripheral blood were isolated and amplified, then cultured together with normal keratinocytes. By MTT method, the living cells were quantified in the mixed culture. Results: Compared with normal controls, lymphocytes from lesion and peripheral blood of psoriasis both promote the proliferation of keratinocytes (P<0. 01 and P<0. 05 respectively). The concentrations of IL-2 and IFN-7 in the mixture of lesion lymphocytes and keratinocytes were significantly higher than that of controls. Tripterygium glycosides inhibited this promotion. Conclusion: Lymphocytes in patients with psoriasis (mainly Th1 cell) play an important role in proliferation of keratinocytes. This psoriasis cell model is useful for studies on signal transduction in psoriasis.
文摘The susceptibility was compared between murine bone marr ow hemopoietic cells and splenic lymphocytes to four major factors of cryopreservation process: toxicity of dimethyl sulfoxide (DMSO), cooling rate,Tris-NH4Cl treatment, and dilution after thawing. When the concentration of DMSO was over 20%, the proliferative function or viability of both kinds of cells decreased markedly. Injury to hemopoietic stem cells and lymphoeytes was found more severe in rapid cooling than in slow cooling, and the intensity of iujury seemed alike under the same fast cooling condition. Pretreatment of bone marrow cells with Tris-NH4Cl did not significantly accentuate the injury of frozen-thawed hemopoietic stem cells. Dilution of the frozen-thawed cells in ice bath or at a fast rate was harmful to both cells. No significant differential susceptibilities of the two kinds of cells to each factor were found. These results indicate that selective destruction or inactivation of lymphocytes by cold treatment among bone marrow cells may be impossible.
文摘A series of artificial membranes called small unilaminar vesicles (SUVs) comprising different molar ratios of phosphatidylcholine (PC)/cholesterol (Ch) were constructed and exogenous PC and Ch were transferred into human peripheral blood lymphocytes through incubating the cells with SUVs in order to study the effects of the changed constituents of membrane lipids on the expression of CD 71 in lymphocytes, the binding characteristics of CD 71 molecules with its ligand and its internalization. It was found that after the treatment of SUVs, the Ch content in the plasmic membrane was increased with a decrease of PC/Ch ratio. After incubation with ConA for 48 h,the percentage of CD 71-positive lymphocytes increased with the increase of the availab1e binding sites for CD 71 to its ligand, but the affinity of CD 71 with its ligand was lowered- When Ch synthesis was blocked with HMG-CoAreductase inhibitor (Lovastatin), the number of CD71 molecules was obviously reducce but the afinity of CD 71with its ligand elevated- It is believed that that the construction of the lipid environment can directly affect the properties of CD 71 molecules. Less fluid phase of the lipid environment might be suitable to the move;nent of CD 71 in the membrane but minimize the tendency for CD 71 to maintain a con formation of high affinity state.
文摘Objective The purpose of our study is to observe the voltage-gated potassium channel Kvl.3 expressed on CD4＋CD28~ T cells from the peripheral blood of acute coronary syndrome （ACS） patients by the patch clamp technique. Methods Kvl.3 potassium channels expression from 17 patients with ACS and 11 healthy age-match controls was detected in single cell（CD4＋CD28null T cells and CD4＋CD28＋T cells） by fluorescence mieroscopy and patch clamp. Results The percentage of CD4＋CD28mullT cells was higher in the ACS patients [（6.97±2.05）%] than that in the controls [（1.38±0.84）%, P〈0.05]. The concentration of hsCRP was directly correlated with the number of the CD4~CD28nul~ T cells in the ACS patients （r=0.52, P〈0,05）. The conductance （6.89±1.17ns vs 3.36±0.66ns）, dens （1.95±0.80 μm2 vs 1.13±0.57 μm2） and numbers （574.5±97.6 n/cell vs. 280.3±55.3 n/cell） of the Kv1.3 channels on the CIM＋CD28null T cells were significantly higher than those on the CD4＋CD28＋ T cells （all P〈0.01） in ACS patients, but were similar on CD4＋CD28＋T betweenACS patients and controls. Conclusion The CD4＋CD28nullT cells and the numbers of Kvl.3 channels on the CD4＋CD28nullT cells from patients with ACS are significantly upregulated and might contribute to the pathogenesis of ACS （d Geriatr Cardio12010; 7：40-46）.
文摘To put forward a method of automatic recognition of the micronuclei on computerized image in bi-nucleated lymphocytes. Methods: The procedure was as the follows: ① The nuclei and cytoplasm images were automatically extracted from the background in order that the nuclei and cytoplasm images could be preprocessed individually.② Noise in two images was reduced, the edges in the nuclei image smoothed and then the big-point noise in the cytoplasm image taken off. ③ The nuclei was identified and the single cytoplasm acquired individually form nuclei and cytoplasm image. ④ The two preprocessed images were put together to identify the bi-nuclei and micronuclei form the overlapped map. Results: Many new models for image processing have been established. Preliminary measurement showed that the accuracy of recognition reached a high percentage of 85%. Conclusion: It is of great value to put the automatic recognition system into use.
文摘Objective Congestive heart failure （CHF） is the final common pathway of various heart diseases.Calcineurin,a calcium/calmodulindependent phosphatase consisting of a catalytic subanit A （CnA） and a regulatory calcium-binding subunit B （CnB）,is activated in heart failure.This study aimed to investigate the relationship between mRNA level of calcineurin in circulating T-lymphocyte and that in myocardium in patients with CHF. Methods A total of 38 patients with CHF （aged from 29 to 62 years） were included in this study.The mRNA levels of alpha-and beta-isoform of CnA in left ventricular anterior papillary muscle and peripheral lymphocytes were determined by semi-quantitative reverse transcription polymerase chain reaction.Pearson linear correlation analysis was performed,and difference was considered statistically significant at a P value 〈0.05. Results Calcineurin mRNA levels in lymphocytes were positively correlated with those in myocardium （for CnA-alpha mRNA,r=0.820;for CnA-beta mRNA,r=0.875;both P〈0.01）.CnA-beta mRNA levels in both circulating lymphocytes and myocardium increased significantly with increasing NYHA class （r=0.877 for peripheral blood and r=0.805 for cardiac muscle;both P〈0.01）. Conclusions The mRNA level of CnA-beta in circulating lymphocytes is positively correlated with that in myocardium and is a promising marker for the severity of cardiac dysfunction in patients with CHF.
