AIM： To investigate the effect of a vaccine with recombinant adenovirus interleukin-12 （AdVIL-12） transduced dendritic cells （DCs） against colon cancer in mice. METHODS： DCs and AdVIL-12 were incubated together ...AIM： To investigate the effect of a vaccine with recombinant adenovirus interleukin-12 （AdVIL-12） transduced dendritic cells （DCs） against colon cancer in mice. METHODS： DCs and AdVIL-12 were incubated together at different time intervals and at different doses. Supernatant was collected and tested for IL-12 by enzyme-linked immunosorbent assay （ELISA）. In order to determine whether tumor cell lysate-pulsed （TP） AdVIL-12/DCs enhance therapeutic potential in the established tumor model, CT26 colon tumor cells were implanted subcutaneously （s.c.） in the midflank of naive BALB/c mice. Tumor-bearing mice were injected with a vaccination of CT26 TP AdVIL-12/DCs on d 3 and 10. As a protective colon tumor model, naive BALB/c mice were immunized s.c. in their abdomens with CT26 TP AdVIL-12/DCs twice at seven day intervals. After the immunization on d 7, the mice were challenged with a lethal dose of CT26 tumor cells and survival times were evaluated. Subsequently, cytotoxic T lymphocyte （CTL） activity and interferon gamma （IFNy） secretion was evaluated in the immunized mice, and assayed CTL ex vivo. RESULTS： Murine DCs were retrovirally transduced with AdVIL-12 efficiency, and the AdVIL-12 transduced DCs secreted a high level of IL-12 （AdVIL-12/DCs, 615.27 ± 42.3 pg/mL vs DCs, 46.32 ± 7.29 pg/mL, P 〈 0.05）. Vaccination with CT26 TP AdVIL-12/DCs could enhance anti-tumor immunity against CT26 colon tumor in murine therapeutic models （tumor volume on d 19：CT26 TP AdVIL-12/DCs 107 ± 42 mm^3 vs CT26 TP DCs 383± 65 mm^3, P 〈 0.05） and protective models. Moreover, the CT26 TP AdVIL-12/DC vaccination enhances tumor-specific CTL activity, producing high levels of IFN7 in immunized mice. Ex vivo primed T cells with AdVIL-12/DCs were able to induce more effective CTL activity than in primed T cells with CT26 TP/DCs （E：T = 100：1, 69.49% ± 6.11% specific lysis vs 37.44% ＋ 4.32% specific lysis, P 〈 0.05）.CONCLUSION： Vaccination with recombinant AdVIL-12 transduced DC pulsed tumor cell lysate enhance antitumor immunity specific to colon cancer in mice.展开更多
Crohn’s disease (CD) and ulcerative colitis (UC) are chronic inflammatory disorders of the gastrointestinal tract that share clinical and pathological characteristics. The most accredited hypothesis is that both CD a...Crohn’s disease (CD) and ulcerative colitis (UC) are chronic inflammatory disorders of the gastrointestinal tract that share clinical and pathological characteristics. The most accredited hypothesis is that both CD and UC result from a deregulated mucosal immune response to normal constituents of the gut microflora. Evidence, however, indicates that the main pathological processes in these two diseases are distinct. In CD, the tissue- damaging inflammatory reaction is driven by activated type 1 helper T-cell (Th1), whereas a humoral response predominates in UC. Consistently, a marked accumulation of macrophages making interleukin (IL)-12, the major Th1-inducing factor, is seen in CD but not in UC mucosa. Preliminary studies also indicate that administration of a monoclonal antibody blocking the IL-12/p40 subunit can be useful to induce and maintain clinical remission in CD patients. Notably, the recently described IL-23 shares the p40 subunit with IL-12, raising the possibility that the clinical benefit of the anti-IL-12/p40 antibody in CD may also be due to the neutralization of IL-23 activity. This review summarizes the current information on the expression and functional role of IL-12 and IL- 12-associated signaling pathways both in patients with CD and experimental models of colitis, thus emphasizing major differences between IL-12 and IL-23 activity on the development of intestinal inflammation.展开更多
Hepatitis C virus (HCV) chronic infection is a worldwide health problem, and numerous efforts have been invested to develop novel vaccines. An efficient vaccine requires broad immune response induction against viral p...Hepatitis C virus (HCV) chronic infection is a worldwide health problem, and numerous efforts have been invested to develop novel vaccines. An efficient vaccine requires broad immune response induction against viral proteins. To achieve this goal, we constructed a DNA vaccine expressing nonstructural 3 (NS3) gene (pcDNA3.1-HCV-NS3) and assessed the immune response in C57BL/6 mice. In this study, the NS3 gene was amplified with a nested-reverse transcriptase-polymerase chain reaction (RT-PCR) method using sera of HCV-infected patients with genotype 1a. The resulting NS3 gene was subcloned into a pcDNA3.1 eukaryotic expression vector, and gene expression was detected by western blot. The resultant DNA vaccine was co-administered with interleukin-12 (IL-12) as an adjuvant to female C57BL/6 mice. After the final immunizations, lymphocyte proliferation, cytotoxicity, and cytokine levels were assessed to measure immune responses. Our data suggest that co-administration of HCV NS3 DNA vaccine with IL-12 induces production of significant levels of both IL-4 and interferon (IFN)-γ (p<0.05). Cytotoxicity and lymphocyte proliferation responses of vaccinated mice were significantly increased compared to control (p<0.05). Collectively, our results demonstrated that co-administration of HCV NS3 and IL-12 displayed strong immunogenicity in a murine model.展开更多
AIM: To investigate the effect of moxibustion on intestinal flora and release of interleukin-12 (IL-12) and tumor necrosis factor-α (TNF-α) from the colon in rat with ulcerative colitis (UC). METHODS: A rat model of...AIM: To investigate the effect of moxibustion on intestinal flora and release of interleukin-12 (IL-12) and tumor necrosis factor-α (TNF-α) from the colon in rat with ulcerative colitis (UC). METHODS: A rat model of UC was established by local stimulation of the intestine with supernatant from colonic contents harvested from human UC patients. A total of 40 male Sprague-Dawley rats were randomly divided into the following groups: normal (sham), model (UC), herb-partition moxibustion (HPM-treated), and positive control sulfasalazine (SA-treated). Rats treated with HPM received HPM at acupuncture points ST25 and RN6, once a day for 15 min, for a total of 8 d. Rats in the SA group were perfused with SA twice a day for 8 d. The colonic histopathology was observed by hematoxylin-eosin. The levels of intestinal flora, including Bifidobacterium, Lactobacillus, Escherichia coli (E. coli), and Bacteroides fragilis (B. fragilis), were tested by real-time quantitative polymerase chain reaction to detect bacterial 16S rRNA/DNA in order to determine DNA copy numbers of each specific species. Immunohistochemical assays were used to observe the expression of TNF-α and IL-12 in the rat colons. RESULTS: HPM treatment inhibited immunopathology in colonic tissues of UC rats; the general morphological score and the immunopathological score were significantly decreased in the HPM and SA groups compared with the model group [3.5 (2.0-4.0), 3.0 (1.5-3.5) vs 6.0 (5.5-7.0), P < 0.05 for the general morphological score, and 3.00 (2.00-3.50), 3.00 (2.50-3.50) vs 5.00 (4.50-5.50), P < 0.01 for the immunopathological score]. As measured by DNA copy number, we found that Bifidobacterium and Lactobacillus, which are associated with a healthy colon, were significantly higher in the HPM and SA groups than in the model group (1.395 ± 1.339, 1.461 ± 1.152 vs 0.045 ± 0.036, P < 0.01 for Bifidobacterium, and 0.395 ± 0.325, 0.851 ± 0.651 vs 0.0015 ± 0.0014, P < 0.01 for Lactobacillus). On the other hand, E. coli and B. fragilis, which are associated with an inflamed colon, were significantly lower in the HPM and SA groups than in the model group (0.244 ± 0.107, 0.628 ± 0.257 vs 1.691 ± 0.683, P < 0.01 for E. coli, and 0.351 ± 0.181, 0.416 ± 0.329 vs 1.285 ± 1.039, P < 0.01 for B. fragilis). The expression of TNF-α and IL-12 was decreased after HPM and SA treatment as compared to UC model alone (4970.81 ± 959.78, 6635.45 ± 1135.16 vs 12333.81 ± 680.79, P < 0.01 for TNF-α, and 5528.75 ± 1245.72, 7477.38 ± 1259.16 vs 12550.29 ± 1973.30, P < 0.01 for IL-12). CONCLUSION: HPM treatment can regulate intestinal flora and inhibit the expression of TNF-α and IL-12 in the colon tissues of UC rats, indicating that HPM can improve colonic immune response.展开更多
Objective. To investigate the anti- tumor effects of human single chain interleukin- 12 (hscIL- 12). Method. pcDNA/ hscIL- 12 recombinant was transfected into human hepatic carcinoma cells (7721 cells) by lipofectin m...Objective. To investigate the anti- tumor effects of human single chain interleukin- 12 (hscIL- 12). Method. pcDNA/ hscIL- 12 recombinant was transfected into human hepatic carcinoma cells (7721 cells) by lipofectin method. The 7721/hscIL- 12 cells which secrete hscIL- 12 stably, were obtained via G418 selection, and in vitro the influence of hscIL- 12 gene transduction on the growth of tumor cells was evaluated by cell cycle analysis. In vivo, genetically engineered 7721 cells (7721/hscIL- 12, 7721/pcDNA) and parental cells were implanted into BALB/c nude mice,respectively. 7721/pcDNA and 7721/hscIL- 12 groups were divided into two sub- groups on day 8: one was administered with hPBL twice, 6 days at interval; the other was given equal volume of PBS. Mice were sacrificed on day 26, and spleens and tumors were taken out for histologic assay. Results. hscIL- 12 produced stably by 7721/hscIL- 12 cells had bioactivity, and it was proved by Western blot, immunocytochemistry, and in situ hybridization. In vitro, compared with 7721 and 7721/pcDNA, the 7721/hscIL- 12 grew much more slowly. FACS assay showed apparent G1 arrest of 7721/hscIL- 12 cells. In animal experiment, on day 8 after inoculation, the tumors of 7721 and 7721/pcDNA group were up to 5～ 7mm,while those of 7721/hscIL- 12 group were 2～ 4mm.When treated with hPBL, the tumor of 7721/hscIL- 12 group disappeared completely. Histologically, the tumors from 7721/hscIL- 12 without hPBL treatment had numerous lymphocyte infiltration, the tumor cells displayed depression looking, atrophy, focal necrosis and apoptosis , whereas the tumors of 7721 and 7721/pcDNA groups grew thrivingly. Conclusion. hscIL- 12 transduced 7721 cells could induced significant antitumor immune response which resulted in tumor regression totally when the hPBL was inoculated, and also hscIL- 12 has certain effects on mice immune system. These findings suggest that hscIL- 12 and hscIL- 12 gene therapy might have promising prospects in clinical application.展开更多
Objective: To investigate the effect of interleukin-12 (IL-12) and interleukin-18 (IL-18)DNA immunization on immune response induced by HIV-1 DNA vaccine and to explore new strategies for therapeutic HIV DNA vaccine. ...Objective: To investigate the effect of interleukin-12 (IL-12) and interleukin-18 (IL-18)DNA immunization on immune response induced by HIV-1 DNA vaccine and to explore new strategies for therapeutic HIV DNA vaccine. Methods: The recombinant expression vector pCI-neoGAG was constructed by inserting HIV Gag gene into the eukaryotic expression vector pCI-neo. Balb/c mice were immunized with pCI-neoGAG alone or co-immunized with the DNA encoding for IL-12 or IL-18.Anti-HIV antibody and IFN-γ were tested by ELISA,and splenocytes were isolated for detecting antigen-specific lymphoproliferative responses and specific CTL response by MTT assay and LDH assay respectively. Results: The anti-HIV antibody titers of mice co-immunized with pCI-neoGAG and the DNA encoding for IL-12 or IL-18 were lower than that of mice immunized with pCI-neoGAG alone(P<0.01). In contrast, the IFN-γ level of mice co-immunized with pCI-neoGAG and the DNA encoding for IL-12 or IL-18 was higher than that of mice immunized with pCI-neoGAG alone (P<0.01).Furthermore, compared with mice injected with pCI-neoGAG alone, the specific CTL cytotoxity activity and antigen-specific lymphoproliferative responses of mice immunized with pCI-neoGAG and the DNA encoding for IL-12 or IL-18 were significantly enhanced respectively (P<0.01). Conclusion: The DNA encoding for IL-12 or IL-18 together with HIV DNA vaccine may enhance specific Th-1 responses and cellular immune response elicited in mice. Hence, the DNA encoding for IL-12 or IL-18 are promising immune adjuvants for HIV-1 DNA vaccine.展开更多
Background Our previous study showed that the combined Chinese herbs containing scutellaria baicalensis georgi and gardenia jasminoids ellis inhibited atherosclerosis. In this study, we sought to determine if baicalin...Background Our previous study showed that the combined Chinese herbs containing scutellaria baicalensis georgi and gardenia jasminoids ellis inhibited atherosclerosis. In this study, we sought to determine if baicalin and geniposide could inhibit atherosclerosis through Wntl and dickkopf-related protein-1 （DKK1）. Methods The wild-type and ApoE-/- mice were treated with baicalin, geniposide, and baicalin plus geniposide daily by gavage for 12 weeks. Blood lipid levels were measured with an automatic biochemistry analyzer. Aortic atherosclerotic lesion areas were analyzed with Image-ProPlus software. The mRNA and protein expression of DKK1, Wntt and nuclear factor-r,B （NF-κB） were measured with RT-PCR and Westem Blot. Serum levels of interleukin-12 （IL-12） were quantified with ELISA. Results The baicalin or geniposide monotherapy as well as combination therapy inhibited the development of atherosclerotic lesions, increased Wntl and decreased DKKI expression and elevated the ratio of Wntl/DKK1 compared with high-lipid diet group. However, only baicalin or geniposide monotherapy decreased NF-κB expression. Moreover, baicalin and geniposide monoor combination therapy lowered IL-12 levels. Geniposide reduced both serum total cholesterol and low density lipoprotein levels, while baicalin either alone or in combination with geniposide did not affect serum lipid levels. In human, umbilical vein endothelial ceils stimulated by oxidized low density lipoprotein, baicalin and geniposide also increased Wntl and decreased DKK1 expression and elevated the ratio of Wntl/DKK1. Condusions Baicalin and geniposide exert inflammation-regulatory effects and may prevent atherosclerotic lesions through enhancing Wntl and inhibit- ing DKK1 expression.展开更多
Objective To determine the in vitro expression of interleukin-12 (IL-12) and its effect on signal transducers and activators of transcription (STAT) signaling molecules in peripheral blood mononuclear cells (PBMCs) in...Objective To determine the in vitro expression of interleukin-12 (IL-12) and its effect on signal transducers and activators of transcription (STAT) signaling molecules in peripheral blood mononuclear cells (PBMCs) in patients with systemic lupus erythematosus (SLE).Methods Peripheral blood mononuclear cells in 39 patients with definite systemic lupus erythematosus and 11 healthy volunteers were collected. Expression of IL-12 P40mRNA in PBMCs was determined with reverse transcription-polymerase chain reaction (RT-PCR). Quantity of IL-12 protein supernatant was measured by enzyme-linked immunosorbent assay (ELISA). The levels of phosphorylated STAT3 and STAT4 signaling molecules in PBMCs were detected by immunoblot. Results Levels of IL-12 protein and mRNA expression in patients with active or inactive SLE were significantly higher than those in controls. Phytohemagglutinin (PHA) may promote the expression of IL-12. IL-12 alone induced the phosphorylation of STAT3 and STAT4 in PBMCs from patients with SLE, especially in active SLE. However it had no obvious effect on normal PBMCs. Phosphorylated STAT3 and STAT4 might be observed in normal PBMCs treated with IL-12 plus PHA.Conclusion IL-12 is produced aberrantly in patients with SLE. IL-12 might exert its biological role in SLE via the aberrantly phosphorylated STAT3 and STAT4 signaling molecules.展开更多
文摘AIM： To investigate the effect of a vaccine with recombinant adenovirus interleukin-12 （AdVIL-12） transduced dendritic cells （DCs） against colon cancer in mice. METHODS： DCs and AdVIL-12 were incubated together at different time intervals and at different doses. Supernatant was collected and tested for IL-12 by enzyme-linked immunosorbent assay （ELISA）. In order to determine whether tumor cell lysate-pulsed （TP） AdVIL-12/DCs enhance therapeutic potential in the established tumor model, CT26 colon tumor cells were implanted subcutaneously （s.c.） in the midflank of naive BALB/c mice. Tumor-bearing mice were injected with a vaccination of CT26 TP AdVIL-12/DCs on d 3 and 10. As a protective colon tumor model, naive BALB/c mice were immunized s.c. in their abdomens with CT26 TP AdVIL-12/DCs twice at seven day intervals. After the immunization on d 7, the mice were challenged with a lethal dose of CT26 tumor cells and survival times were evaluated. Subsequently, cytotoxic T lymphocyte （CTL） activity and interferon gamma （IFNy） secretion was evaluated in the immunized mice, and assayed CTL ex vivo. RESULTS： Murine DCs were retrovirally transduced with AdVIL-12 efficiency, and the AdVIL-12 transduced DCs secreted a high level of IL-12 （AdVIL-12/DCs, 615.27 ± 42.3 pg/mL vs DCs, 46.32 ± 7.29 pg/mL, P 〈 0.05）. Vaccination with CT26 TP AdVIL-12/DCs could enhance anti-tumor immunity against CT26 colon tumor in murine therapeutic models （tumor volume on d 19：CT26 TP AdVIL-12/DCs 107 ± 42 mm^3 vs CT26 TP DCs 383± 65 mm^3, P 〈 0.05） and protective models. Moreover, the CT26 TP AdVIL-12/DC vaccination enhances tumor-specific CTL activity, producing high levels of IFN7 in immunized mice. Ex vivo primed T cells with AdVIL-12/DCs were able to induce more effective CTL activity than in primed T cells with CT26 TP/DCs （E：T = 100：1, 69.49% ± 6.11% specific lysis vs 37.44% ＋ 4.32% specific lysis, P 〈 0.05）.CONCLUSION： Vaccination with recombinant AdVIL-12 transduced DC pulsed tumor cell lysate enhance antitumor immunity specific to colon cancer in mice.
文摘Crohn’s disease (CD) and ulcerative colitis (UC) are chronic inflammatory disorders of the gastrointestinal tract that share clinical and pathological characteristics. The most accredited hypothesis is that both CD and UC result from a deregulated mucosal immune response to normal constituents of the gut microflora. Evidence, however, indicates that the main pathological processes in these two diseases are distinct. In CD, the tissue- damaging inflammatory reaction is driven by activated type 1 helper T-cell (Th1), whereas a humoral response predominates in UC. Consistently, a marked accumulation of macrophages making interleukin (IL)-12, the major Th1-inducing factor, is seen in CD but not in UC mucosa. Preliminary studies also indicate that administration of a monoclonal antibody blocking the IL-12/p40 subunit can be useful to induce and maintain clinical remission in CD patients. Notably, the recently described IL-23 shares the p40 subunit with IL-12, raising the possibility that the clinical benefit of the anti-IL-12/p40 antibody in CD may also be due to the neutralization of IL-23 activity. This review summarizes the current information on the expression and functional role of IL-12 and IL- 12-associated signaling pathways both in patients with CD and experimental models of colitis, thus emphasizing major differences between IL-12 and IL-23 activity on the development of intestinal inflammation.
文摘Hepatitis C virus (HCV) chronic infection is a worldwide health problem, and numerous efforts have been invested to develop novel vaccines. An efficient vaccine requires broad immune response induction against viral proteins. To achieve this goal, we constructed a DNA vaccine expressing nonstructural 3 (NS3) gene (pcDNA3.1-HCV-NS3) and assessed the immune response in C57BL/6 mice. In this study, the NS3 gene was amplified with a nested-reverse transcriptase-polymerase chain reaction (RT-PCR) method using sera of HCV-infected patients with genotype 1a. The resulting NS3 gene was subcloned into a pcDNA3.1 eukaryotic expression vector, and gene expression was detected by western blot. The resultant DNA vaccine was co-administered with interleukin-12 (IL-12) as an adjuvant to female C57BL/6 mice. After the final immunizations, lymphocyte proliferation, cytotoxicity, and cytokine levels were assessed to measure immune responses. Our data suggest that co-administration of HCV NS3 DNA vaccine with IL-12 induces production of significant levels of both IL-4 and interferon (IFN)-γ (p<0.05). Cytotoxicity and lymphocyte proliferation responses of vaccinated mice were significantly increased compared to control (p<0.05). Collectively, our results demonstrated that co-administration of HCV NS3 and IL-12 displayed strong immunogenicity in a murine model.
基金Supported by National Natural Science Foundation of China, No. 81001549National Basic Research Program of China (973 program), No. 2009CB522900+1 种基金Shanghai Health System of Outstanding Young Talent Cultivation Program, No. XYQ2011068Shanghai Rising-Star Program, No. 10QA1406100
文摘AIM: To investigate the effect of moxibustion on intestinal flora and release of interleukin-12 (IL-12) and tumor necrosis factor-α (TNF-α) from the colon in rat with ulcerative colitis (UC). METHODS: A rat model of UC was established by local stimulation of the intestine with supernatant from colonic contents harvested from human UC patients. A total of 40 male Sprague-Dawley rats were randomly divided into the following groups: normal (sham), model (UC), herb-partition moxibustion (HPM-treated), and positive control sulfasalazine (SA-treated). Rats treated with HPM received HPM at acupuncture points ST25 and RN6, once a day for 15 min, for a total of 8 d. Rats in the SA group were perfused with SA twice a day for 8 d. The colonic histopathology was observed by hematoxylin-eosin. The levels of intestinal flora, including Bifidobacterium, Lactobacillus, Escherichia coli (E. coli), and Bacteroides fragilis (B. fragilis), were tested by real-time quantitative polymerase chain reaction to detect bacterial 16S rRNA/DNA in order to determine DNA copy numbers of each specific species. Immunohistochemical assays were used to observe the expression of TNF-α and IL-12 in the rat colons. RESULTS: HPM treatment inhibited immunopathology in colonic tissues of UC rats; the general morphological score and the immunopathological score were significantly decreased in the HPM and SA groups compared with the model group [3.5 (2.0-4.0), 3.0 (1.5-3.5) vs 6.0 (5.5-7.0), P < 0.05 for the general morphological score, and 3.00 (2.00-3.50), 3.00 (2.50-3.50) vs 5.00 (4.50-5.50), P < 0.01 for the immunopathological score]. As measured by DNA copy number, we found that Bifidobacterium and Lactobacillus, which are associated with a healthy colon, were significantly higher in the HPM and SA groups than in the model group (1.395 ± 1.339, 1.461 ± 1.152 vs 0.045 ± 0.036, P < 0.01 for Bifidobacterium, and 0.395 ± 0.325, 0.851 ± 0.651 vs 0.0015 ± 0.0014, P < 0.01 for Lactobacillus). On the other hand, E. coli and B. fragilis, which are associated with an inflamed colon, were significantly lower in the HPM and SA groups than in the model group (0.244 ± 0.107, 0.628 ± 0.257 vs 1.691 ± 0.683, P < 0.01 for E. coli, and 0.351 ± 0.181, 0.416 ± 0.329 vs 1.285 ± 1.039, P < 0.01 for B. fragilis). The expression of TNF-α and IL-12 was decreased after HPM and SA treatment as compared to UC model alone (4970.81 ± 959.78, 6635.45 ± 1135.16 vs 12333.81 ± 680.79, P < 0.01 for TNF-α, and 5528.75 ± 1245.72, 7477.38 ± 1259.16 vs 12550.29 ± 1973.30, P < 0.01 for IL-12). CONCLUSION: HPM treatment can regulate intestinal flora and inhibit the expression of TNF-α and IL-12 in the colon tissues of UC rats, indicating that HPM can improve colonic immune response.
文摘Objective. To investigate the anti- tumor effects of human single chain interleukin- 12 (hscIL- 12). Method. pcDNA/ hscIL- 12 recombinant was transfected into human hepatic carcinoma cells (7721 cells) by lipofectin method. The 7721/hscIL- 12 cells which secrete hscIL- 12 stably, were obtained via G418 selection, and in vitro the influence of hscIL- 12 gene transduction on the growth of tumor cells was evaluated by cell cycle analysis. In vivo, genetically engineered 7721 cells (7721/hscIL- 12, 7721/pcDNA) and parental cells were implanted into BALB/c nude mice,respectively. 7721/pcDNA and 7721/hscIL- 12 groups were divided into two sub- groups on day 8: one was administered with hPBL twice, 6 days at interval; the other was given equal volume of PBS. Mice were sacrificed on day 26, and spleens and tumors were taken out for histologic assay. Results. hscIL- 12 produced stably by 7721/hscIL- 12 cells had bioactivity, and it was proved by Western blot, immunocytochemistry, and in situ hybridization. In vitro, compared with 7721 and 7721/pcDNA, the 7721/hscIL- 12 grew much more slowly. FACS assay showed apparent G1 arrest of 7721/hscIL- 12 cells. In animal experiment, on day 8 after inoculation, the tumors of 7721 and 7721/pcDNA group were up to 5～ 7mm,while those of 7721/hscIL- 12 group were 2～ 4mm.When treated with hPBL, the tumor of 7721/hscIL- 12 group disappeared completely. Histologically, the tumors from 7721/hscIL- 12 without hPBL treatment had numerous lymphocyte infiltration, the tumor cells displayed depression looking, atrophy, focal necrosis and apoptosis , whereas the tumors of 7721 and 7721/pcDNA groups grew thrivingly. Conclusion. hscIL- 12 transduced 7721 cells could induced significant antitumor immune response which resulted in tumor regression totally when the hPBL was inoculated, and also hscIL- 12 has certain effects on mice immune system. These findings suggest that hscIL- 12 and hscIL- 12 gene therapy might have promising prospects in clinical application.
文摘Objective: To investigate the effect of interleukin-12 (IL-12) and interleukin-18 (IL-18)DNA immunization on immune response induced by HIV-1 DNA vaccine and to explore new strategies for therapeutic HIV DNA vaccine. Methods: The recombinant expression vector pCI-neoGAG was constructed by inserting HIV Gag gene into the eukaryotic expression vector pCI-neo. Balb/c mice were immunized with pCI-neoGAG alone or co-immunized with the DNA encoding for IL-12 or IL-18.Anti-HIV antibody and IFN-γ were tested by ELISA,and splenocytes were isolated for detecting antigen-specific lymphoproliferative responses and specific CTL response by MTT assay and LDH assay respectively. Results: The anti-HIV antibody titers of mice co-immunized with pCI-neoGAG and the DNA encoding for IL-12 or IL-18 were lower than that of mice immunized with pCI-neoGAG alone(P<0.01). In contrast, the IFN-γ level of mice co-immunized with pCI-neoGAG and the DNA encoding for IL-12 or IL-18 was higher than that of mice immunized with pCI-neoGAG alone (P<0.01).Furthermore, compared with mice injected with pCI-neoGAG alone, the specific CTL cytotoxity activity and antigen-specific lymphoproliferative responses of mice immunized with pCI-neoGAG and the DNA encoding for IL-12 or IL-18 were significantly enhanced respectively (P<0.01). Conclusion: The DNA encoding for IL-12 or IL-18 together with HIV DNA vaccine may enhance specific Th-1 responses and cellular immune response elicited in mice. Hence, the DNA encoding for IL-12 or IL-18 are promising immune adjuvants for HIV-1 DNA vaccine.
基金This study was funded by grants from the Natural Science Foundation of Hubei Province in China （2012FFB02508）.
文摘Background Our previous study showed that the combined Chinese herbs containing scutellaria baicalensis georgi and gardenia jasminoids ellis inhibited atherosclerosis. In this study, we sought to determine if baicalin and geniposide could inhibit atherosclerosis through Wntl and dickkopf-related protein-1 （DKK1）. Methods The wild-type and ApoE-/- mice were treated with baicalin, geniposide, and baicalin plus geniposide daily by gavage for 12 weeks. Blood lipid levels were measured with an automatic biochemistry analyzer. Aortic atherosclerotic lesion areas were analyzed with Image-ProPlus software. The mRNA and protein expression of DKK1, Wntt and nuclear factor-r,B （NF-κB） were measured with RT-PCR and Westem Blot. Serum levels of interleukin-12 （IL-12） were quantified with ELISA. Results The baicalin or geniposide monotherapy as well as combination therapy inhibited the development of atherosclerotic lesions, increased Wntl and decreased DKKI expression and elevated the ratio of Wntl/DKK1 compared with high-lipid diet group. However, only baicalin or geniposide monotherapy decreased NF-κB expression. Moreover, baicalin and geniposide monoor combination therapy lowered IL-12 levels. Geniposide reduced both serum total cholesterol and low density lipoprotein levels, while baicalin either alone or in combination with geniposide did not affect serum lipid levels. In human, umbilical vein endothelial ceils stimulated by oxidized low density lipoprotein, baicalin and geniposide also increased Wntl and decreased DKK1 expression and elevated the ratio of Wntl/DKK1. Condusions Baicalin and geniposide exert inflammation-regulatory effects and may prevent atherosclerotic lesions through enhancing Wntl and inhibit- ing DKK1 expression.
文摘Objective To determine the in vitro expression of interleukin-12 (IL-12) and its effect on signal transducers and activators of transcription (STAT) signaling molecules in peripheral blood mononuclear cells (PBMCs) in patients with systemic lupus erythematosus (SLE).Methods Peripheral blood mononuclear cells in 39 patients with definite systemic lupus erythematosus and 11 healthy volunteers were collected. Expression of IL-12 P40mRNA in PBMCs was determined with reverse transcription-polymerase chain reaction (RT-PCR). Quantity of IL-12 protein supernatant was measured by enzyme-linked immunosorbent assay (ELISA). The levels of phosphorylated STAT3 and STAT4 signaling molecules in PBMCs were detected by immunoblot. Results Levels of IL-12 protein and mRNA expression in patients with active or inactive SLE were significantly higher than those in controls. Phytohemagglutinin (PHA) may promote the expression of IL-12. IL-12 alone induced the phosphorylation of STAT3 and STAT4 in PBMCs from patients with SLE, especially in active SLE. However it had no obvious effect on normal PBMCs. Phosphorylated STAT3 and STAT4 might be observed in normal PBMCs treated with IL-12 plus PHA.Conclusion IL-12 is produced aberrantly in patients with SLE. IL-12 might exert its biological role in SLE via the aberrantly phosphorylated STAT3 and STAT4 signaling molecules.
