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Objective To compare the proportion of CD3~+T cell and CD4~+CD25~(high) regulatory T cell (Treg cell) and the production of inferon γ ( IFN-γ) and interleukin-12 (IL-12) in peripheral blood mononuclear cells (PBMC) between patients with non-small cell lung cancer (NSCLC) and healthy people, and to analyze the changes of CD3~+T cell, CD4~+CD25~(high)Treg cell, IFN-γ and IL-12 of NSCLC patients before and after chemotherapy, so as to determine immune function changes of NSCLC patients caused by chemotherapy. Methods Twenty NSCLC patients and 20 healthy volunteers according to the including criteria were selected. Three mL of blood was drawn from NSCLC patients before chemotherapy (0 d), on the 3~(rd) day (3 d) and 7~(th) day (7 d) after chemotherapy. PBMC cells were separated from the blood samples. The proportions of CD4~+CD25~(high)Treg cell and CD3~+T cell (%) in PBMC were tested by FACS, and the IFN-γ and IL-12 (pg/mL) in the supernatants were also detected. Results The proportions of CD3~+T cell in NSCLC patients on 0, 3 and 7 d were (55.15±20.11)%,(57.73±14.08)% and (62.79±7.80)%,respectively, and there was no statistical difference between any two of these results. The proportion of CD4~+CD25~(high)Treg cell in healthy volunteers was (2.14±0.85)%, while that of NSCLC patients was (2.76±0.53)% on 0 d with statistical difference compared to the healthy volunteers (P<0.05). The CD4~+CD25~(high)Treg cell proportion (%) of NSCLC patients on 3 d and 7 d were (2.54±0.57)% and (2.72±0.29)%, respectively, which were both significantly lower than that of 0 d. On 3 d it was even much lower than that on 7 d (P<0.05). IFN-γ and IL-12 of NSCLC patients on 0 d were (34.36±4.38) pg/mL and (33.24±4.36) pg/mL, and no statistical difference was observed when compared with (34.36±4.38) pg/mL and (33.24±4.36) pg/mL in the healthy volunteers. On 3 d and 7 d, IFN-γ of NSCLC patients were (40.42±5.66) pg/mL and (39.27±6.07) pg/mL, respectively, and both were higher than that on 0 d (P<0.05); IL-12 of NSCLC patients were (35.51±5.03) pg/mL and (38.62±6.44) pg/mL, also both were higher than that on 0 d (P<0.05). Conclusions This study suggests that chemotherapy can improve immune functions of NSCLC patients, and may reinforce the anti-tumor immune response.
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objective To investigate the tissue localization of CD4+T cells producing IL-17,namely Th17 cells.in patients with systemic lupus erythematosus (SLE),as well as its relationship with the activity of lupus.Methods By using H&E staining.double-label immunofluorescence.immunohistochemistry and confocal microscopy.the localization of Th17 cells was carried out in peripheral blood mononuclear cells (PBMCs).affected tissue of skin and lung obtained from 4 patients with active SLE and 2 normal human controls.Flow cytometry.reverse transcription PCR.ELISA were used to detect the proportion of Th17 cells in PBMCs,the mRNA expression of interleukin-17(IL-17)A and IL-17 F,and serum level of interleukin 17,respectively,in 50 consecutive adult patients with SLE and 15 normal human controls.Results Th17 cells were detected in PBMCs of patients with active SLE.and the fuorescence intensity of IL-17 was significantly higher in patients with active SLE than in normal human controls(127.6±20.5 vs 40.6±11.1,P＜0.001).Infiltrates of Th17 cells were noted in both skin and lung tissues of patients with active SLE.but not in those of normal human controls.The proportion of Th17 cells in PBMCs was increased in patients with active SLE.and the proportion positively correlated with SLE disease activity index(SLEDAI) (r=0.725,P＜0.01).Further more.a significant increase was observed in the mRNA expression of IL-17 A and IL-17 F and serum level of IL-17 in patients with active SLE compared with normal human controls.The amount of Th17 cells was positively correlated with the development of vasculitis.and it experienced a decrease with the remission of SLE.Conclusions A proliferation of Th17 cells is noted in patients with active SLE.which seems to closely correlated with the activity of SLE and may take part in the development of vasculitis in SLE.
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Objective To observe whether peripheral blood mononuclear cells (PBMCs) from patients with active systemic lupus erythematosus (SLE) can induce the adhesion of T lymphocytes to endothelial cells. Methods Sera and PBMCs were obtained from patients with active SLE and normal human controls. PBMCs were cultivated and culture supematants were harvested. Human umbilical vein endothelial cells (HUVECs) were cultured in vitro with or without the presence of the sera or culture super-natants of PBMCs. Some cells were pretreated with the antibody to IL-17 before the treatment with the sera or supematants. After another 48-hour culture, RT-PCR and real-time PCR were used to detect the mRNA expressions of adhesion molecules, including intercellular adhesion moleeule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) and E-cadherin in HUVECs, wound healing assay to estimate the motility of HUVECs. Additionally, T lymphocytes were added to HUVECs 48 hours after stimulation with the sera or supematants, the adhering of T lymphocytes to HUVECs was observed by microscopy. Results After stimulation with supematants of PBMCs from patients with active SLE, the mRNA expressions of ICAM-1, VCAM-1 and E-cadherin were significantly increased in HUVEC, while the increase could be inhibited by the antibody to IL-17. The elevation of adhesion molecule expression subsequently promoted the motility of HUVEC, mediated the adhesion of T lymphocytes to HUVEC, and the antibody to IL-17 could suppress the adhesion of T lymphocytes and motility of HUVEC. Conclusion The culture supematants of PBMCs from patients with active SLE can induce the expression of vascular cell adherin molecules and promote the adherin of T lymphocytes, which may in turn mediate the development of lupus vasculitis.
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Objective: To evaluate the efficacy of 3LL/GM-CSF tumor vaccine combined with pacilitaxel chemotherapy in treatment of mice bearing transplanted Lewis lung carcinoma. Methods: The tumor vaccine 3LL/GM-CSF was prepared by infecting Lewis lung carcinoma cell line 3LL with adenovirus encoding GM-GSF. Mice model of Lewis lung carcinoma was established by subcutaneous injection of 2?104 3LL cells into C57BL/6(H-2b)mice. The sensitivity of Lewis lung carcinoma cell line-3LL to the treatment of pacilitaxel was detected in vivo and in vitro. The mice tumor model was first treated with pacilitaxel chemotherapy and then with 3LL/GM-CSF, or first with 3LL/GM-CSF and then with pacilitaxel. Tumor growth and the long-term survival of mice were observed after treatment. The immune memory and cytotoxicity against target cells were studied in the mice. Results: Pacilitaxel at 100 nmol/L killed 32.10% 3LL cells after 24 hour in vitro; but pacilitaxel at 5-25 mg/kg only had a poor effect on growth of 3LL cells in vivo. The tumors rebated in 70% of mice treated with pacilitaxel chemotherapy and 3LL/GM-CSF vaccination successively, and the survival of these mice was obviously longer than that of pure pacilitaxel chemotherapy group (70.0 days vs 27.5 days). The killing rate of 3LL/GM-CSF after pacilitaxel chemotherapy was 41.35% on day 3. Meanwhile, the survival mice could resist the re-attack of 3LL cells (2?104). We also noticed that first treatment with 3LL/GM-CSF and then pacilitaxel chemotherapy had no effect on tumors. Conclusion: Application of tumor vaccine shortly after pacilitaxel chemotherapy can induce specific immune responses and prolong the survival of experimental mice, which provide a basis for future clinical practice.
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The aim of the current study was to determine whether tumor-specific T cells can be primed in and obtained from sponge implants loaded with tumor associated peptides. Naive C57BL/6 mice as well as C57BL/6 mice previously primed with FBL-3 tumor cells were implanted with small polyurethane sponges containing FBL-3 gag peptides CCLCLTVFL(gPr80 gag 85-93)or RSPTNLAKV(Pr65 gag p30 131-139). Both FBL-3 gag peptides were shown could bind to H-2 Db molecules. Ten days later, cells that had accumulated in the sponges were harvested, stimulated in vitro with the immunizing peptide, and tested for cytolytic activity against FBL-3 tumor and FBL-3 gag peptides. The results demonstrated that peptide-specific CD8+ CTL could be elicited and obtained from the sponge implants of both naive and immune mice. The FBL-3 gag p85-93 peptide induced CTL could specifically lyse syngeneic targets pulsed with the FBL-3 gag p85-93 peptide as well as FBL-3 tumor. However, the FBL-3 gag p131-139 peptide induced CTL lysed only the FBL-3 gag p131- 139 peptide pulsed syngeneic targets but not the FBL- 3 tumor. Tumor-specific T cells obtained from peptide-loaded sponge implants could be induced to grow to large numbers in vitro by periodic restimulation with the immunizing peptide plus syngeneic APC and low concentrations of IL- 2. Adoptive transfer of the resultant expanded FBL-3 gag p85-93 peptide-induced CTL into mice with disseminated FBL-3 could mediate effective anti-tumor therapy. Thus,in vivo immunization with peptide-loaded sponges provides a potentially useful technique for procuring primed peptide- specific T cells for tumor therapy.
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Objective: To study the anti-tumor efficacy induced by antibodized tumor epitope PDTRP gene immunization. Methods: Three copies of tumor associate gene PDTRP from MUCI tandem repeats were designed and mimicked the conformation of MUCI by Insight Ⅱ . The ?lneo-PDTRP plasmid was further constructed, in which the PDTRP target gene was inserted into CDR3 of the ?1 -neo vector. The specific humoral and cellular immune responses towards to PDTRP were detected after intraspleen immunized Balb/c mice with "ylneo-PDTRP. And the immune protection assay was also done to observe whether the mice immunized with ?lneo-PDTRP could prolong the survival after tumor challenge. Results: The conformation of three copies of PDTRP mimicked the conformation of MUCI tandem repeats. The expression of ?lneo-PDTRP could be detected after in vitro transfect. The specific antibody against PDTRP epitope could be induced and increase to a higher titer after intraspleen injection with a ?lneo-PDTRP plasmid. And the specific proliferation and cytotox-ic function of lymphocyte were also increased. There is a significant survival from mice immunized with ?lneo-PDTRP a-gainst the 4T1-PDTRP tumor challenge. Conclusions: Gene immunization with ?lneo-PDTRP could elicit both humoral and cellular tumor specific immune response and had the protective effect.
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Objective: To investigate whether the plasmid ?1neo-hgp100 could be expressed and presented in vitro and could protect the immunized mice from B16F10 challenge in vivo. Methods: ?1neo-hgp100 plasmid was constructed in which the DNA sequence encoding hgp100 CTL epitope inserted into CDR3 of ?1-neo vector. The expression of anti-bodized antigen and IFN-? in supernatant was measured by ELISA respectively after transfection J558L with ?1neo-hgp100 and further co-culture of J588L transfacted with ?1 neo-hgp100 and pmel TCR transgenic T cell. After introspleenic inoculation of ?1neo-hgp100, the protective efficacy of the gene vaccine was observed by means of measuring the tumor area every two days. Results: ?1neo-hgp100 could be expressed and presented in vitro, the immunogenecity of CTL epitope of hgp100 was strong enough and could activate gp100 specific T cell, the mice immunized with the gene vaccine could resist the tumor challenging in vivo. The mean survival time was prolonged to 36 days, compared to control group (P
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Objective:To investigate the effects of IP10(IFN-? inducible protein 10,CXCL10) on anti-tumor immune response and to explore its mechanisms involved in.Methods:A mammary carcinoma cell line 4T1 was transfected with pcDNA3-IP10(IP10-4T1) by electrophoration and positive clones were screened in the presence of G418.Growth kinetics of IP10-4T1 cells was observed in vitro and in vivo.Survival rate among the animals was determined by daily assessment.Proliferation activity of lymphocytes was analyzed with()~3H-TdR incorporation.The phenotypes of lymphocytes isolated from tumors by ficoll density gradient centrifugation were assayed by flow cytometry.Results:The growth rate of IP10-4T1 cells was similar with that of parental 4T1 cells and neo-vector transfected 4T1 cells in vitro.Growth of the tumors formed by IP10-4T1 cells was inhibited in vivo.Compared to those of controls,the size and the weight of the tumors formed by IP10-4T1 cells decreased significantly 35 days post tumor transplantation(P