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Objective:To observe any regulatory effect of a pulsed electromagnetic field (PEMF) on A2A adenosine receptors (A2ARs) in the nucleus pulposus of rats with intervertebral disc degeneration (IDD), and to explore any combination with the A2ARs′ agonist-mediating ROS/PI3K/Akt signaling pathway.Methods:Fifty Sprague-Dawley rats were randomly divided into a control group, an intervertebral disc degeneration group (the model group), an A2AR agonist CGS-21680 treatment group (the agonist group), a PEMF group and a PEMF combined with CGS-21680 treatment group (the observation group). IDD was modeled in all except the rats in the control group. 100μL of CGS-21680 (100μg/kg) was injected into the L 5-6 intervertebral discs of the agonist group, while the PEMF group was given 30 minutes of PEMF intervention daily for 14 days at 1.5mT and 75Hz with a pulse width of 150μs. The observation group was injected with CGS-21680 and then given the same PEMF intervention. Primary nucleus pulposus cells from each group (50ng/mL) were cultured and the expressions of 8-OHDG, SOD, MDA and ROS were detected by immunohistochemistry, immunofluorescence or with an ELISA kit. The A2AR, PI3K, AKT and p-AKT protein levels were detected using western blotting. Results:The nucleus pulposus cells and the annulus fibrosus were obviously wrinkled, necrotic and broken in the model group but the annulus fibrosus was intact and the nucleus pulposus was almost normal in the observation group. Compared with the model group, the levels of SOD and A2AR, PI3K, p-AKT and AKT protein were higher in the agonist, PEMF and observation groups, while the expressions of MDA, ROS and 8-OHDG were weaker. The ROS level in the observation group was significantly lower than that in the agonist and PEMF groups, and the phosphorylation level of p-AKT in the observation group was significantly higher than in the agonist and PEMF groups. The average levels of SOD, A2AR, PI3K, p-AKT and AKT protein in the nucleus pulposus cells of the agonist, PEMF and observation groups were significantly higher than the IL-1β group′s average, while the average levels of MDA, ROS and 8-OHDG were significantly lower. The ROS levels in the observation group were significantly lower than in the agonist and PEMF groups, while the A2AR protein content and p-AKT phosphorylation in the observation group were significantly greater. The average Bax levels in the nucleus pulposus cells of the agonist, PEMF and observation groups were significantly lower than that in the IL-1β group, and the expression of Bcl-2 was significantly increased. There was significantly less apoptosis observed in the observation group than in the agonist and PEMF groups, while the expression of Bcl-2 was significantly higher.Conclusions:PEMF plays an anti-oxidative stress role by up-regulating A2AR activity and reducing ROS generation. Treatment with PEMF and A2AR agonist could further activate the phosphorylation of PI3K/Akt, down-regulate Bax and up-regulate Bcl-2, thus inhibiting the apoptosis of nucleus pulposus cells and alleviating the malignant progression of IDD.
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Objective:To explore the effect of combining pulsed electromagnetic field (PEMF) stimulation with A 2A adenosine receptor agonist CGS-21680 on apoptosis and inflammation of nucleus pulposus cells in cases of intervertebral disc degeneration. Methods:Forty-eight Sprague-Dawley rats were randomly divided into a sham operation group (Sham group), an intervertebral disc degenerative disease group (Model group), an A 2A adenosine receptor agonist CGS-21680 group (Agonist group), and a group in which PEMF was combined with CGS-21680 (Observation group). The rat model of intervertebral disc degeneration (IDD) was established in all other groups than the sham operation group. The rats in the Agonist group were injected with 100μL of CGS-21680 (100μg/kg) at the L 5-6 intervertebral disc. The Observation group was injected with CGS-21680 similarly but then received 14 conse-cutive days of PEMF stimulation (30min/time). The Sham and Model groups were injected with the same amount of normal saline solution. Eight weeks later, HE staining was used to evaluate the pathological changes in the intervertebral disc tissues. The expression of type II collagen was determined by immunohistochemistry. The content and mRNA expression of inflammatory factors interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) were determined using enzyme-linked immunosorbent assays and reverse transcription-polymerase chain reactions (RT-PCRs). The protein and mRNA levels of A 2A, NLRP3 and caspase-3 were determined by western blotting and RT-PCR. Results:The degeneration in the nucleus pulposus and annulus fibrosus in the Model group was significant, while significantly less shrinkage, necrosis and fibrous annulus rupture was observed in the Observation group. Compared with Model and Agonist groups, the positive expression of Col Ⅱ in the nucleus pulposus, A 2AR protein levels and relative expression of its mRNA had all increased significantly in the Observation group, while the average levels and mRNA expression of pro-inflammatory cytokines IL-6 and TNF-α had decreased significantly. The average protein level and mRNA expression of NLRP3 and caspase-3 in the intervertebral disc tissues of the Observation group were significantly lower than in the Model and Agonist groups. Conclusions:Combining pulsed electromagnetic field stimulation with A 2A adenosine receptor agonist CGS-21680 can inhibit the apoptosis of nucleus pulposus cells, alleviate disease response and delay IDD by up-regulating the activity of A 2A receptors and down-regulating the expressions of pro-inflammatory factors IL-6, TNF-α, NLRP3 and caspase-3 in nucleus pulposus cells.
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Dengue virus (DENV) is the most widely transmitted arbovirus in the world. Due to the lack of diagnostic technology to quickly identify the virus serotypes in patients, severe dengue hemorrhagic fever cases caused by repeated infections remain high. To realize the rapid differential diagnosis of different serotypes of DENV infection by immunological methods, in this study, four DENV serotype NS1 proteins were expressed and purified in mammalian cells. Monoclonal antibodies (MAbs) against NS1 protein were obtained by hybridoma technology after immunizing BALB/c mice. Enzyme-linked immunosorbent assay, indirect immunofluorescence assay, dot blotting, and Western blotting were used to confirm the reactivity of MAbs to viral native NS1 and recombinant NS1 protein. These MAbs include not only the universal antibodies that recognize all DENV 1-4 serotype NS1, but also serotype-specific antibodies against DENV-1, DENV-2 and DENV-4. Double antibody sandwich ELISA was established based on these antibodies, which can be used to achieve rapid differential diagnosis of serotypes of DENV infection. Preparation of DENV serotype-specific MAbs and establishment of an ELISA technology for identifying DENV serotypes has laid the foundation for the rapid diagnosis of DENV clinical infection.
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Animals , Humans , Mice , Antibodies, Monoclonal , Antibodies, Viral/metabolism , Dengue/diagnosis , Dengue Virus/immunology , Enzyme-Linked Immunosorbent Assay , Mice, Inbred BALB C , Sensitivity and Specificity , Serogroup , Viral Nonstructural Proteins/immunologyABSTRACT
Objective:To observe electromagnetic field irradiation′s ability to inhibit heterotopic ossification (HO) and explore the mechanisms involved.Methods:A total of 72 male Sprague-Dawley rats were randomly divided into a control group, a model group and an observation group, each of 24. The rats in the model and observation groups were made to model HO by clamping and cutting the Achilles tendon, while among those in the control group the Achilles tendon was only exposed without clamping and cutting. The rats in the observation group were exposed to a 1mT electromagnetic field alternating sinusoidally at 50Hz (SEMF) twice a day for 2h each time. At 3 days, 14 days, 1 month and 3 months after the operation, Achilles tendon tissue was resected from 6 rats in each group for gross and histological examination and immunohistochemical staining. The protein content and gene expression were detected using western blotting or reverse transcription polymerase chain reaction. X-ray examination was performed at 1 and 3 months after the operation to observe any HO.Results:No significant changes were observed in the control group after the operation. Compared with the model group, HO in the observation group was significantly inhibited. The average expression of the MMP-9 gene in the model and observation groups was significantly higher than in the control group, and it increased with time. The average tissue metalloproteinase-1 (TIMP-1) gene expression was significantly lower, and it decreased with time. Compared with the model group, the average expression of MMP-9 in the observation group was significantly less, while that of TIMP-1 was significantly greater. Compared with the model group, there was also a significant increase in the expression of decapentaplegic homolog 7 (Smad7) protein and a significant decrease in the expression of phosporylated Smad3 protein at each time point.Conclusion:Applying 50Hz, 1mT SEMF has a definite effect in inhibiting HO, at least in rats. It may be related to promoting the expression of Smad7 and inhibiting the phosphorylation of Smad3, thus inhibiting the TGF-β/Smad signal transduction pathway and promoting the restoration of a dynamic balance of MMP-9 with TIMP-1.
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Objective@#To explore the expression of the A2A adenosine receptor and the inflammatory cytokines interleukin-1 beta (IL-1β) and tumor necrosis factor alpha (TNF-α) in human degenerative nucleus pulposus (NP) cells after they have been treated with a pulsed electromagnetic field (PEMF).@*Methods@#Human degenerative NP cells were cultured in vitro and treated using an 0.8mT PEMF with a pulse frequency of 50Hz. The pulse width was 150μs and the exposure time was 30min, repeated 5 times at 12 hour intervals. The expression of the A2A adenosine receptor in NP cells was determined using western blotting and reverse transcription polymerase chain reactions. The expression of the inflammatory cytokines IL-1β and TNF-α were detected using enzyme-linked immunosorbent assays (ELISA). The human degenerative NP cells were also treated with an antagonist and agonist of the A2A adenosine receptor, and the expression of IL-1β and TNF-α were also determined using ELISA.@*Results@#After the PEMF treatment the expression of the A2A adenosine receptor increased significantly, while the expression of IL-1β and TNF-α decreased significantly. However, the A2A adenosine receptor antagonist reversed the inhibitory effect of the PEMF on the expression of IL-1β and TNF-α, while the agonist played an opposite role.@*Conclusion@#A PEMF can significantly inhibit the expression of IL-1β and TNF-α in human degenerative NP cells, which could be related to up-regulation of the expression of the A2A adenosine receptor in those cells.
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Objective To systematically evaluate the effects of metformin on serum vitamin B12,folic acid and homocysteine (Hcy) levels in the patients with type 2 diabetes mellitus (T2DM).Methods The databases of Cochrane Library,EMbase,PubMed,CBM,CNKI,VIP and Wanfang were retrieved by computer to collect the randomized controlled trials (RCTs) on the association between metformin and serum vitamin B12 level in T2DM patients.Two researchers independently screened the literatures,assessed the risk of bias of included RCTs and extracted the data.The RevMan5.3 software was used for conducting the meta analysis.Results A total of 5 RCTs were included.The Meta-analysis results showed that serum vitamin B12 level in the metformin group was significantly decreased compared with the placebo group (MD=-55.86,95 ％ CI(-86.89,-24.84),P =0.000 4);the subgroup analysis showed that serum vitamin B12 level was negatively correlated with the duration of diabetes,metformin use time and dose.The serum Hcy level in the metformin group was increased compared with the placebo group (MD=2.44,95 ％ CI(1.41,3.46),P＜0.01);the serum folic acid level had no statistical differences between the two groups [MD=-2.39,95 ％CI (-4.93,0.15),P=0.06];the incidence rate of adverse reactions in the metformin group was higher than that in the placebo group[RR=2.0,95 ％ CI(1.32,3.03),P =0.001].Conclusion Metformin treatment may lead to decrease of serum vitamin B12 level and increase of Hcy level while has no obvious influence on the folic acid level in T2DM patients,but has higher incidence rate of adverse reactions.
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Objective To evaluate the effects of posterior decompression,subtotal vertebrectomy,intervertebral fusion and internal fixation through unilateral pedicle in the treatment of severe throracolumar fractures.Methods A retrospective study was conducted to evaluate the outcomes of 11 patients with severe thmracolumar fracture who had been treated with posterior decompression,subtotal vertebrectomy,intervertebral fusion and internal fixation through unilateral pedicle at our department from March 2009 to March 2012.They were 10 males and one female,aged from 20 to 42 years (mean,31.1 years).According to AO classification,there were 2 cases of type B1,3 cases of type B2 and 6 cases of type C2.The Gaines scores for anterior column stability were more than 6 in all.All the patients had neurological disorders.The fracture reduction,fusion of bone grafting and recovery of neurological function were followed up.Results Their operation time ranged from 200 to 300 minutes,averaging 243.2 minutes;the amount of bleeding ranged from 800 to 1,600 mL,averaging 1,023.3 mL.No deterioration of neurological symptoms or no new neurological symptoms were observed postoperatively.Cerebrospinal fluid leakage was found in 3 patients who healed spontaneously following extubation,compressive dressing and elevation of the bed end.All the patients were followed up for 12 to 30 months (average,18.3 months).A loosened screw cap was found in one patient after 6 months who had no discomfort and received no treatment.X-ray films showed satisfactory fracture reduction and fine bone grafting locations.No spinal canal stenosis was found by CT scans.It took 3 to 6 months (mean,4.3 months) for the bone grafts to get fused.No height loss of the injured vertebrae was no found at the last follow-ups.No functional recovery was found in the patients with complete spinal cord injury whose Frankle grade remained A.The spinal function recovered from Frankel grade B to grade D in one patient with incomplete spinal injury.Of the 8 patients with medullary cone injury,7 recovered Frankle grade E and one Frankle grade D.The symptoms were relieved in one patient with simple cauda equine injury.Conclusion Posterior decompression,subtotal vertebrectomy,intervertebral fusion and internal fixation through unilateral pedicle is an effective alternative for treatment of severe throracolumar fractures,because it can lead to satisfactory fracture reduction,thorough decompression of the spinal canal and good reconstruction of spinal stability.
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Objective To explore the correlation between cardiovascular risk factors and characteristics of coronary artery le‐sions ,the degree of stenosis in patients with coronary artery disease(CAD) .Methods Totally 156 patients who under went coro‐nary angiography(CAG) in our hospital were divided into CAD group(n=98) and non‐CAD group(n=58) which the patients with coronary stenosis 0 .05) .The number of coronary artery lesion branch were increased with these risk factors incresed ,such as smoking ,SBP ,LDL‐C ,FBG ,no significant difference with age ,male ,TCH ,TG .The degree of stenosis were increased with SBP ,FBG ,no significant difference with age ,male ,smoking ,TCH ,TG and LDL‐C .Conclusion The smoking ,males ,FBP and SBP are the prominent risk factors of CAD in Qinghai area ,prosmote the occurrence and development of CAD together .
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Objective To investigate the treatment effect of aescuven forte on the postoperative complications of breast cancer.Methods One hundred and twenty patients with breast cancer radical surgery were randomly divided into control group(n =60) and treatment group(n =60).Patients in control group were given postoperative routine treatment,while in treatment group were administrated aescuven forte pills at 300 mg orally,2 times/day for 4 weeks beside the conventional treatment.Results (1)After the 1st,2nd weeks therapy,the flap congestion disappear rate in the treatment group were 80.0％ (48/60) and 93.3％ (56/60),better than that in the control group 60.0％ (36/60) and 71.6％ (43/60),the difference was statistically significant (P ＜ 0.05).However this trend was not seen in third weeks treatment (P ＞ 0.05).(2) The total efficiency of remission of upper limb edema was 93％ (56/60),higher than that in control group 77％ (46/60),the difference was statistically significant(x2 =5.17,P ＜ 0.05).(3) Visual analogue scale(VAS) pain score in treatment group were decreased form (8.87 ±0.74) in before treatment to (3.21 ±0.92) at after treatment.And the VAS score in control was from (8.91 ±0.85) down to (4.87 ± 1.34),the difference was statistically significant (P ＜ 0.05).Moreover VAS score in treatment group was lower than that in control group (P ＜ 0.05).(4) There was no adverse effect of the medication process.Conclusion Aescuven forte showed a ability to reduce congestion disappear time of breast cancer and shorten the recovery time of upper limb swelling and pain and other symptoms.
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Objective To investigate the significance of 25 hydroxyvitamin D3 (25 (OH)D3) in patients with type 2 diabetes mellitus by comparing the differences of serum 25-hydroxyvitamin D levels between type 2 diabetes mellitus patients and healthy people.Methods One hundred and forty-seven patients with type 2 diabetes mellitus were selected as the T2DM group,and 135 healthy people were selected as the normal control (NC) group.The level of serum 25 (OH) D3 was detected by ELISA.Clinical data were recorded and biochemical indexes were also detected.Statistical analysis was performed.Results (1) Serum 25 (OH) D3 level in T2DM group was low,but there was no significant difference between two groups((23.10 ± 1.59) μg/L vs.(23.11 ± 1.65) μg/L; t =0.064,P =0.949),as well as between men and women in the 2 populations (T2DM men vs.women(23.22 ± 1.54) μg/L vs.(22.95 ± 1.65) μg/L; t =0.209,P ＞0.05; NC men vs women(23.03 ± 1.73) μg/L vs.(23.18 ± 1.59) μg/L; t =0.436,P ＞0.05) ; (2)The prevalence of vitamin D insufficient and deficiency in T2DM group was high,but it was not statistically significant between two groups (98.0％ vs.94.8 ％ and 2.0％ vs.5.2％ ; x2 =2.034,P =0.154) ; (3) Serum 25 (OH) D3 in T2DM group has a significant negative correlation with triglycerides (r =-0.218,P ＜ 0.01).Conclusion (1) Vitamin D level was low in both the healthy and T2DM populations in the southern region of China,the prevalence of vitamin D insufficient and deficiency was up to 90％.(2)Triglyceride is the only independent risk factor to T2DM population.
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Since the study of the mechanism of breast cancer occurrence and development deepens, breast cancer stem cells are receiving more and more attention. Studies have shown that a group of breast cancer stem cells were undifferentiated, with self-renewal and multi-differentiation potential. These cells have a resistance to chemotherapy, radiotherapy, hypoxic, high tumorigenic, high invasion and metastasis. In breast cancer's recurrence,development, and even metastasis, they play an extremely important role. In-depth study of breast cancer stem cell related signal transduction pathways and the regulation of microenvironment are meaningful for clinical targeted treatment of breast cancer. Therefore, we summarized the latest development on breast cancer stem cells in the treatment of breast cancer.
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BACKGROUND: It has been proved that electromagnetic field can adjust and control proliferation and differentiation of bone marrow mesenchymal stem cells v/a cyclic adenosine monophosphate (cAMP)-protein kinase A (PKA) signal transduction system. However, there are few relevant reports about Ca2+ as the second messenger in application. OBJECTIVE: To study the effects of verapamil on the proliferation and differentiation of bone marrow rnesenchymal stem cells stimulated by electromagnetic fields and to conclude influx changes of Ca2+.DESIGN, TIME AND SETTING: Electrostimulative cytological observation in vitro, which was performed in Laboratory of Orthopedic Surgery, Tongji Hospital between April and June 2005.MATERIALS: Six 4-5-week SD rats of clean grade were selected in this study. Verapami| was provided by Sigma Company, USA, and Helmholtz coil-magnetic field producer was made in Department of Electric Machine, Navy Engineering University.METHODS: The bone marrow mesenchymal stem cells were isolated and cultured in vitro with adherence method and digested with trypsin. The fourth-passage cells were harvested, adjusted to 1 × 107 L-1 in density, and divided into A, B, C and D groups in 96-well plate with 200 μ I/well. Cells in the normal control group were not performed with any agent. On the second day of inoculation, cells in the magnetic field (EMF) group were cultured in Helmholtz-coil magnetic field (0.8 mT, 50 Hz) in 0.05% CO2 saturated humidity incubator at 37 ℃, 30 minutes for each, 12 hours for interval, six time in total. Cells in the verapamil group were cultured with 20 μ mol/L verapamil, and cells in the combination group were cultured with 20 μ mol/L verapamil and magnetic stimulation.MAIN OUTCOME MEASURES: Proliferative activity was tested with MTT method, content of alkaline phosphate differentiated to osteoblasts was measured, and cells were stained with modified Gomori Ca-Co staining. RESULTS: Proliferative activity was significantly increased in the EMF group as compared with that in the normal control group after 3-day magnetic stimulation (P < 0.01), but verapamil could inhibit promotive effect on proliferation. Content of alkaline phosphate in the normal control group was similar to that in the EMF group, while those two contents were significantly higher than those in the verapamil group and the combination group (P < 0.01); furthermore, content of alkaline phosphate in the combination group was significant higher than that in the EMF group (P < 0.01). Qualitative analysis of alkaline phosphate showed a coincident result as mentioned above.CONCLUSION: EMF of 50 Hz frequency and 0.8 mT intensity can change intracellular free calcium ion concentration of bone marrow mesenchymal stem cells, and the change play a key role in the cellular proliferation and play a partial role in the differentiation of bone marrow mesenchymal stem cells into osteoblasta.
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<p><b>OBJECTIVE</b>To investigate the relationship between six-transmembrane epithelial antigen of the prostate (STEAP) expression and the histologic grading of prostatic carcinoma (PCa).</p><p><b>METHODS</b>Different prostatic tissues and non-prostatic tumors, 131 cases of PCa, 164 cases of benign prostate hyperplasia (BPH), and 56 cases of non-prostatic malignancies, were analyzed for the expression of STEAP by using STEAP monoclonal antibody and SP immunohistochemical staining. The positive area unit (PU) was introduced to describe the intensity of STEAP expression.</p><p><b>RESULTS</b>The prostatic tissues of all but 3 cases of PCa and 5 cases of BPH were stained positively, while all of the non-prostatic tumors were stained negatively. There was a significant negative correlation between the STEAP expression and the histologic grading of PCa.</p><p><b>CONCLUSION</b>STEAP can be a prognostic marker of PCa and a potential therapeutic target in PCa.</p>
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Humans , Male , Antigens, Neoplasm , Genetics , Immunohistochemistry , Oxidoreductases , Prostatic Neoplasms , Diagnosis , Allergy and Immunology , PathologyABSTRACT
<p><b>OBJECTIVE</b>To evaluate the clinical efficacy of intra-articular injection with 2% chitosan as a treatment for irreducible anterior disc displacement of the TMJ.</p><p><b>METHODS</b>Total 30 patients with irreducible anterior disc displacement were divided into two groups according to quasi-randomizaion. The patients in test group received intra-articular injection with 1.0 ml of 2% chitosan into upper cavities of the suffered joints, in control group with 12.5 mg of prednisolone. The patients were followed up at the 1st day and 14th day after injection and the maximal mouth opening was measured.</p><p><b>RESULTS</b>The maximal mouth opening at 14th day was (36.73 +/- 4.69) mm in test group, with 11.73 mm increase from baseline; and (28.53 +/- 5.81) mm in control group, with 3.86 mm increase, respectively. The increase of maximal mouth opening in test group was significantly higher than that of control group (P < 0.05).</p><p><b>CONCLUSION</b>Chitosan is an effective biomaterial in curing irreducible anterior disc displacement of TMJs.</p>
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Adolescent , Adult , Female , Humans , Male , Biocompatible Materials , Chitin , Chitosan , Follow-Up Studies , Injections, Intra-Articular , Prednisolone , Range of Motion, Articular , Temporomandibular Joint Disc , Wounds and Injuries , Pathology , Temporomandibular Joint Disorders , Drug Therapy , Pathology , Treatment OutcomeABSTRACT
Objective To study the cotransfection mB7-1 and mCD1D gene into pancreatic cancer cells of rats and to observe its anti-tmor responses.Methods Recombinant retroviral vectors expressing mB7-1and mCD1D gene were packaged into GP2-293 cell lines and transfected.The expressions of mB7-1 and mCD1D were detected with PCR and Western blot.The positive cells of mB7-1 and mCD1D were used to induce the anti-tumor immunity in vitro.Results Anti-tumor immunity was induced after B7-1 and CD1D positive cells were coinoculated in syngeneic mice.Furthermore,the growth of tumor was inhibited.Conclusions Cotransfection of B7-1 and CD1D could induce anti-tumor effect.This study provide a foundation for the application of B7-1 and CD1D gene therapy in tumor.