Objective To observe the induction of tolerogenic dendritic cells (DCs) by curcumin (Cur) and its mechanism.Methods After immature DCs from bone marrow cells of Wistar rats were treated with different concentrations of Cur (0,10,20 and 30?mol/L) respectively,and then the DCs were tested by flow cytometry for the surface molecules expression.After the immature DCs were treated by 30?mol/L Cur with or without stimulation of LPS,endocytosis of DCs to dextran was tested by flow cytometry.The production of IL-12 in DC culture supernatant was determined by ELISA.The levels of NF-?B p65 and RelB translocation to the nucleus were investigated by Western- blot.The activity of NF-?B was detected by NF-?B-binding ELISA and luciferase reporter gene analy- sis.The ability of DCs to stimulate the proliferation of T cells from Lewis rats were analyzed by mixed leukocyte reactions (MLR).Results Cur suppressed LPS-indueed cell-surface expression of costimu- latory molecules (CD80,CD86 and CD40) in a dose-dependent manner.When Cur was used at a con- centration of 30?mol/L,there was no marked difference in the surface molecules expression of LPS- inducing DCs as compared with immature DCs.After DCs were induced by LPS (LPS group),the positive rate of FITC-Dextran uptake was (36.6?7.2)%,and the secretory amounts of IL-12 were (415.9?42.7) pg/ml.In DCs of LPS group,the intranuclear RelB and p65 were highly expressed and their DNA binding activity was 0.65?0.08 and 0.74?0.07 respectively.The luciferase activity of reporter gene in LPS group DCs was remarkably increased to 435% as compared with that in the controls.DCs in LPS group showed strong capacity to stimulate T cells proliferation.When DCs were treated with 30?mol/L Cur followed by induction with LPS (Cur+LPS group),the positive rate of Dextran uptake was (78.6?14.2)% and remarkably higher than in LPS group (P

OBJECTIVE To investigate the effect of overexpression of vascular cell adhesion molecule-1(VCAM-1)on the migration in vitro of the murine mesenchymal stem cells(MSCs)and its possible mechanism. METHODS The migration ability of normal mouse MSC (C3) ,empty vector-transfected MSC(C3+N) and VCAM-1 transfected MSC(C3+VCAM-1)was assessed by Transwell culture system in vitro after incubation for 8 and 12 h,respectively. The fetal bovine serum (FBS) was used as the chemotactic agent to induce MSC migration. The transmigrated cells were detected with methylosaniliam chloride(crystal violet)as well as DAPI staining.Furthermore,the specific chemical inhibitors of mitogen-activation protein kinase (MAPK) pathway ( SB203580,PD98059 and JNK inhibitorⅡ)were added to the Transwell system for 12 h and the alteration of the MSC migration ability was evaluated. RESULTS After incubation with FBS for 8 and 12 h,the absolute migrated cell number(7467 ± 485 and 8795 ± 255)and migration rate〔(14.9 ± 1.0)% and(17.6 ± 0.5)%〕of MSC in C3+VCAM-1 group were significantly increased compared with C3 group〔2731±562 and 4779±224, (5.5 ± 1.1)%and(9.6 ± 0.4)%〕and C3+N group〔2539 ± 321 and 5645 ± 1080,(5.1 ± 0.6)%and(11.3 ± 1.1)%〕(P<0.05,P<0.01),but there was no significant difference between C3 and C3+N groups. Moreover,the MSC migration ability of C3+VCAM-1 group was partially suppressed by addition of JNK inhibitorⅡ. The transmigrated cell number(4843 ± 167)and migration rate〔(9.7 ± 0.3)%〕were decreased compared with those of C3+VCAM-1 group without JNK inhibitorⅡ(P<0.01). SB203580 and PD98059,as specific chemical inhibitors of MAPK pathway,had no effect on MSC migration. CONCLUSION VCAM-1 can enhance mouse MSC migration in vitro and th4e mechanism may be related to JNK/MAPK pathway activation.

Objective To explore a new method of functional reconstruction of hand digits and joints with free transfer of foot tissues so as to increase the success rate of the operation.Methods After micro-anatomic study of the plantar and dorsal metatarsal arteries,retrograde and free grafts of foot tissues pedicled with plantar metatarsal arteries were designed and applied in transplantation to treat 76 cases of hand digital or joint defects.The surgeries included 58 cases of transfer of the second toe,four cases of transfer of composite tissues of the second toe, eight cases of transfer of proximal interphalangeal joint,and six cases of nail flap transfer.Results The mi- cro-anatomic study found that the first plantar metatarsal artery was anatomically constant and the diameter of its branch to the second toe was larger than that of the first dorsal metatarsal artery.Flaps survived in 75 of the 76 patients(98.7%),with fine appearance and significantly improved function.One patient who had received free transfer of the second toe to reconstruct the thumb function had to undergo a second repair with infraclavicula skin tube because of refractory arteriospasm of anastomosed vessels.Conclusion Transfer with free retrograde grafts of foot tissues pedicled with plantar metatarsal artery to reconstruct hand functions can effectively improve the success rate of the operation,because it is free of the shortcomings of great anatomic variation of blood vessels and time-consuming and complex procedures in conventional transfer.

OBJECTIVETo explore the anti-tumor immunity in vitro induced by prostate cancer cell vaccine transfected with recombinant adenovirus encoding 4-1BBL in mice.

METHODSThe replication-deficient adenovirus AdEasy-1 system was used to construct recombinant adenovirus Ad-m4-1BBL and Ad-eGFP. The prostate cancer cell RM-1 of mice was transfected with Ad-m4-1BBL and Ad-eGFP, and treated with mitomycin (MMC) to produce TCV, TCV-Ad-eGFP and TCV-Ad-m4-1BBL, followed by co-culture with syngeneic murine spleen cells. Then the cytotoxic activity of the lymphocytes against RM-1 cells was analyzed with CCK-8 solution, and IL-2 and INF-gamma were detected by ELISA.

RESULTSThe 4-1BBL protein was highly expressed in the TCV-Ad-m4-1BBL of the 4-1BBL-transfected mice. TCV-Ad-m4-1BBL significantly increased the expressions of IL-2 ([180.24 +/- 2.22] pg/ml) and INF-gamma ([1512.46 +/- 23.64] pg/ml) as compared with TCV and TCV-Ad-eGFP (P < 0.05), and induced higher RM-1 cell specific cytotoxicity ([34.24 +/- 2.64]%) than the latter two ([9.82 +/- 1.48]%) and ([14.65 +/- 3. 21]%), (P < 0.05). But none of them exhibited significant cytotoxicity against hepatocellular carcinoma Hepal-6.

CONCLUSIONThe m4-1BBL-expressing prostate cancer cell vaccine can effectively induce anti-tumor immune responses.

4-1BB Ligand ; genetics ; immunology ; Animals ; Cancer Vaccines ; genetics ; immunology ; Cell Line, Tumor ; Coculture Techniques ; Cytotoxicity, Immunologic ; genetics ; Female ; Interleukin-2 ; metabolism ; Male ; Mice ; Mice, Inbred C57BL ; Prostatic Neoplasms ; Transfection

Objective:To compare the clinical and prognostic characteristics of ovarian endometrioid carcinoma (OEC) patients with synchronous endometrial lesions and patients with pure OEC.Methods:A retrospective review of the medical records of patients received initial treatment and a postoperative pathological diagnosis of OEC at Peking University People′s Hospital between August 1998 and December 2017 were performed. According to the inclusion criteria, a total of 56 patients with OEC were included in the study, including 13 patients concurrent with simultaneous endometrial lesions (Group A) and 43 patients with pure OEC (Group B).Results:Patients with synchronous endometrial lesions accounted for 23% (13/56). Mean age of Group A at diagnosis was (44.9±8.3) years old, 2/13 of patients were postmenopausal, and no one had a history of hypertension, the first symptom of 5/13 people was irregular vaginal bleeding. Mean age of Group B patients at diagnosis was (52.7±10.2) years old, 53% (23/43) of patients were postmenopausal, and 28% (12/43) patients had the history of hypertension, the first symptom of 4 (9%, 4/43) people was irregular vaginal bleeding. The differences of age, menopause status, history of hypertension and initial symptoms between the two groups were statistically significant (all P<0.05). There were no significant differences in fertility history, dysmenorrhea history, age of menarche, history of endometriosis, preoperative and postoperative CA 125 level, International Federation of Gynecology and Obstetrics (FIGO) stage, tumor grade, metastatic site and platinum-based chemotherapy drug resistance between the two groups (all P>0.05). The overall 5-year survival rate of OEC patients was 91.6%, and the overall 5-year progression-free survival rate was 76.6%. Among them, the 5-year survival rate of the OEC concurrent with simultaneous endometrial lesions group was 80.2%, and the pure OEC group was 93.4%; the 5-year progression-free survival rate of the OEC concurrent with simultaneous endometrial lesions group was 74.1%, and the 5-year progression-free survival rate of the pure OEC group was 77.3%. There were no significant differences between the two groups (all P>0.05). Multivariate analysis showed that the independent factors for the prognosis of OEC patients were FIGO stage ( P=0.006) and residual lesion size ( P=0.020). Conclusions:OEC patients have a high proportion of simultaneous endometrial lesions. OEC with simultaneous endometrial lesions are younger than patients with pure OEC. Synchronous endometrial lesions do not affect the prognosis of patients with OEC.

Objective To evaluate the value of human epididymis secretory protein 4(HE4)and CAl25 in the diagnosis of ovariall malignancy.Methods HF4 and CA125 in the serum specimens of malignant ovarian tumor group(30 cases),benign ovarian diseases(110 cases;45 benign ovarian tumor,57endometriotic diseases and 8 pelvic inflammation were included) and healthy women group( 137 cases)were assayed double blindly . The levels and the diagnosis efficiency of the HE4 and CA125 were analyzed.Results (1) The median levels of HE4 and CA125 were significantly higher in malignant ovarian tumor group (244 pmoi/L and 601 kU/L respectively) than those of the benign ovarian diseases group( 32 pmol/L and 22 kU/L respectively)and healthy women group (32 pmoi/L and 11 kU/L respectively) (P =0. 000-0. 029). The median levels of CA125 were also higher in endometriotic diseases and pelvic inflammation groups(53 and 41 kU/L respectively) than those of benign ovarian tumor group and healthy women group (12 and 11 kU/L respectively;P = 0. 000-0. 031 ). (2) The positive rate of HE4 was lower than that of CA12s in malignant ovarian tumor group ( P = 0. 036 ). HE4 was negative in benign diseases and healthy women groups. But the positive rates of CA125 were 56. 1% and 5/8 respectively in endometriotic diseases and pelvic inflammation groups and there were significant differences compared with HE4( P =0. 000). (3)The HE4 assay had advantage over the CA125 assay in receiver operating characteristic-area under the curve (ROC-AUC) and sensitivity with a specificity of 100% when ovarian malignancy was compared with controls having benign diseases and healthy women, benign tumor or benign diseases groups respectively. The CA125 assay had advantage over the HE4 assay in ROC-AUC and sensitivity with the same specificity when ovarian cancers were compared with controls having healthy women group. (4) Combined assay of HE4 and CA125was better than CA125 alone when ovarian malignancy was compared with controls having any group. (5)Combined assay was better than HE4 alone in ROC-AUC and sensitivity with the same specificity when ovarian cancers were compared with controls having benign diseases and healthy women or healthy women groups. And combined assay was lower in the ROC-AUC and the sensitivity with specificity of 100% than HE4 when ovarian cancers were compared with controls having benign tumors or benign diseases groups respectively. (6) The diagnosis efficiency of the HE4 assay at the level 86 pmol/L determined in ROC curve with controls having benign diseases and healthy women group and at the 95% reference level 50 pmol/L of healthy women or 150 pmol/L recommended by the kit respectively was compared. The sensitivity of 50 pmol/L was 73% higher than 150 pmol/L and 86 pmoi/L, while the specificity and positive predictive value were lower ( P = 0. 002, P = 0. 000 ). The specificity, accuracy and positive predictive value of HE4 assay at the set point of 150 pmol/L and 86 pmol/L were 100%, 96% and 96%. The set point of 86 pmol/L had advantage over 150 pmol/L at the sensitivity of diagnosis, 70% and 63% respectively. But the positive predictive value was 95% lower than 150 pmol/L, being 100%. There was no significant difference( P =0. 883, P = 0. 883 ). Conclusions The specificity of HF4 assay is higher than CA125 assay in the diagnosis of ovarian cancer and HE4 combined with CA125 assay can improve the diagnoses. The set point of 150 pmol/L is advantageous for the accurate diagnosis, while the set point of 86 pmol/L is advantageous for the screening of malignant ovarian cancer.

OBJECTIVETo develop a tight tetracycline-controlled HCV-C double transgenic mouse model.

METHODSBy crossbreeding of ApoE-rtTA-tTS transgenic mice with TRE-HCV-C transgenic mice, the double transgenic mice were produced in the F1 generation. The presence of HCV-C and tTS gene in the F1 generation was confirmed by PCR, followed by further identification and quantification of the transgene using Southern blot hybridization. The expression of HCV-C in the liver of the mouse model was detected immunohistochemically.

RESULTS AND CONCLUSIONTwo transgenic mice were obtained, which contained ApoE-rtTA-tTS and TRE-HCV-C genes in the genome. Five founders contained HCV-C gene as confirmed by PCR and Southern blot hybridization. The tight tetracycline-controlled system may facilitate further study of HCV-C gene expression and gene therapy of hepatic cellular carcinoma.

Animals ; Apolipoproteins E ; genetics ; Blotting, Southern ; Breeding ; Crosses, Genetic ; Female ; Gene Expression Regulation, Viral ; drug effects ; Hepacivirus ; genetics ; immunology ; Hepatitis C Antigens ; genetics ; immunology ; Male ; Mice ; Mice, Transgenic ; Polymerase Chain Reaction ; Tetracycline ; pharmacology ; Trans-Activators ; genetics ; Viral Core Proteins ; genetics

Objective To explore the virulence of enterovirus 71 from infected children in neonatal mice. Methods Three strains of EV71 were isolated from the mild, severe and dead patients. Symptoms, weight and death of mice were recorded throughout 14 days. The mice were sacrificed on the first, third, fifth, seventh and ninth days post infection to gain the tissue virus load including the liver, spleen, lung, intestine, brain and muscle tissue which were used to detect the virus tilter by real-time RT-QPCR, and pathological lesions using HE staining. Results As to the severity of symptoms, no significant difference was found between the severe and mild groups (P＝0. 693), which were more serious than that of the fatal group. (P＝0. 000 ＜ 0. 05/6, P＝0. 000 ＜ 0. 05/6). The survival rate of the mice with mild, severe and fatal virus infection was 77. 2%, 81. 7% and 97. 8%, respectively, and there was a significant difference among the three groups (P＝0. 0010 ＜ 0. 05, P＝0. 001 ＜ 0. 05, P＝0. 0004 ＜ 0. 05). Lung hemorrhage of the mild group was the most serious, and there were no significant differences in pathological lesions of the brain, muscle, spleen and intestine. Virus titer in the liver and muscle was higher than the other tissues and that in mild group of different tissues tended to be higher than the other two groups. Conclusions Neonatal mice infected with the mild strain of enterovirus 71 presents heaviest symptoms, which are not consistent with the outcomes of humans. It is considered to be related to the virus gene, host and other factors.

Objective Clinical significance of using ELISA to determine ?-amyloid(A?)_(1-42) antibody levels in the sera of patients with Alzheimer's disease(AD).Methods 96 wells PVC plate was coated with A?_(1-42)peptide.Serum of AD patient was competing with mouse A?_(1-42)monoclonal antibody in this assay.The second antibody was horseradish peroxidase(HRP)conjugated goat anti-mouse IgG.Serum A?_(1-42)antibody levels were determined by ELISA.Results The sensitivity of this assay was about 1 ng/ml.The recovery rate of this test was between 96.5% and 104.7%.The residual A?_(1-42)antibody levels in human serum or horse serum after A?_(1-42)antibody was removed by absorption were less than 1 ng/ml. Serum A?_(1-42)antibody levels in 37 AD patients[(5.1?1.9)ng/ml]were remarkably lower than those in normal people[(12.6?3.3)ng/ml,P

Objective To detect the expression of matrix metallproteinases(MMPs) in aortic valve of children who suffered from rheumatic heart disease(RHD) and to explore the pathological role of MMPs in children′s rheumatic aortic valve disease.Methods RHD group composed of 18 aortic valves from children suffered from RHD.Controls were 8 children who were died accidentally without cardiovascular system diseases.Hematoxylin and eosin stain observing the histological characteristic of the 2 groups.Immunohistochemistry was used to detect expression of MMP2 and MMP9 on aortic valves in 2 groups.Results Hematoxylin and eosin stain showed:in RHD the valves′ structure were destroyed along with fibrous tissue proliferation,mucinous degeneration,collagen and fiber hyalinization,blood vessel and blood capillary proliferation,lymphocyte,plasmocyte,monocyte infiltration.Immunohistochemistry showed that MMP2 and MMP9 expression were significantly higher than those in the aortic of RHD(68.85?13.08,64.35?9.59) compared with control group(107.31?23.39,116.28?6.99)(t=3.92,10.18 all P