Objective To clone and express bradyzoite antigen 1(BAG1) gene of T. gondii,and analyze the immunoreactivity of the recombinant product. Methods The differentiation of T. gondii RH strain tachyzoites into bradyzoites was induced in vitro,and the coding sequence of BAG1 was amplified from bradyzoites by RT-PCR. The PCR product was analyzed by sequencing. The BAG1 coding sequence was further subcloned into the plasmid pET32a(+). The plasmid pET32a(+)-BAG1 was then transformed into BL21(DE3) to express after IPTG induction. The expression product was purified with Ni-NTA agarose and the purified BAG1 was further analyzed by Western blotting and ELISA. Results BAG1 cDNA was amplified from bradyzoites. After IPTG induction,BAG1 was expressed in a fusional form in E. coli. Western blotting showed that the purified recombinant protein could be specifically recognized by sera from mice chronically infected by T. gondii B36 strain. ELISA showed that the positive rate of T. gondii IgG antibodies of 350 human sera detected by the recombinant BAG1(17.4%) was higher than by recombinant SAG1 (12.6%)(P

OBJECTIVE:To provide reference for rational use of antibiotics in terminal stage patients with malignant tumor. METHODS:The inpatients with malignant tumor who died in the department of medical oncology of our hospital from Mar. 2013 to Mar. 2016 were analyzed retrospectively in respects of general situation,infection situation and antibiotics use. RESULTS:Among 149 inpatients,a total of 129 infection cases were found in 113 patients(75.8%). The most frequent sites of infection were lung(65.9%),followed by digestive tract/abdomen(13.2%),and skin/wound(6.2%). 100 patients (67.1%)received antibiot-ics,61.1% of which were empiric treatment. β-lactam/β-lactam lactamase inhibitor(41.8%),fluoroquinolone(21.7%)and cepha-losporin(16.9%)were the top 3 frequently prescribed antibiotics. The effective rate of antibiotics treatment was as low as 15.8%, which was believed to be linked with the survival duration since infection occurred (P<0.001),the Karnofsky performance scale (KPS)score when infection occurred(P<0.001)and the duration of antibiotics treatment(P=0.025). CONCLUSIONS:Terminal stage patients with malignant tumor are vulnerable to infections,especially to pulmonary infection. The empirical broad-spectrum an-tibiotics are widely used in terminal patients with malignant tumor,but the effectiveness rate of antibiotic treatment is in low level. For those terminal stage patients with malignant tumor and with KPS score<60 points,when futile antibiotics treatment last for more than 7 days,timely termination of antibiotics treatment is a better choice.

Objective To clarify the intravenous thrombolysis utilization of acute ischemic stroke patients in Huashan hospital,and to analyze the factors affecting thrombolytic therapy compliance.Methods The data from a prospective cohort were analyzed.Consecutive acute stroke and transient ischemic attack patients from Huashan Hospital emergency room were recruited in 2014.Eligible ischemic stroke patients were divided into two groups according to intravenous thrombolysis or not.Results Totally 220 patients from emergency room were assessed in 2014.Among eligible patients,43 patients refused intravenous thrombolysis,whereas 59 patients chose this therapy.After multiple analysis,age,baseline NIHSS score,limb weakness,hemiplegic paralysis,facial paralysis or speech symptoms were significantly different between the two groups (U =936.000,P =0.024;U =284.500,P ＜ 0.01;x2 =8.824,P =0.003;x2 =7.732,P=0.005;x2 =5.169,P=0.038;x2 =5.040,P=0.025).Patients with NIHSS score ＜7 tended to refuse thrombolysis therapy in the receiver operating characteristic curve analysis (sensitivity 0.93,specificity 0.71).From 2008 to 2014,244 cases were analyzed in the thrombolysis database.Compared with patients with higher baseline NIHSS score,intracranial hemorrhage rate (2.6％ vs 19.4％;x2 =12.466,P ＜0.01),7-day mortality rate (1.3％ vs 16.9％;x2 =12.308,P ＜0.01) and 3-month mortality rate (3.8％ vs 21.1％;x2 =11.993,P ＜0.01) were lower in patients whose baseline NIHSS score ＜ 7 (minor group).A higher rate of excellent outcome (3-month modified Rankin Scale score ≤ 1)was observed in minor group (78.2％ vs 38.0％;x2 =34.403,P ＜ 0.01).Conclusions Intravenous thrombolysis was performed in 54.6％ of eligible ischemic stroke patients.Age,baseline NIHSS score,limb weakness,hemiplegic paralysis,facial paralysis or speech symptoms were associated with patients' decision of thrombolysis.The effectiveness and safety of intravenous thrombolysis were promising for patients with mild stroke.

Objective To compare the effectiveness of medicated γintrauterine device (IUD) and medicated genefix IUD inserted immediately after abortion. Methods A multicenter clinical trail was performed for the study from Mar. 2012 to Jan. 2013. Totally 840 women who volunteered to participate were randomly allocated to γ-group (medicated γ IUD) or genefix-group (medicated genefix IUD) immediately after abortion. While 464 abortion women who had not used IUD or steroids contraceptive methods were chosen as control group. The effectiveness of the IUD were followed up for 1 year. All women were required to record the number of vaginal bleeding days and blood volume of vaginal bleeding within 3 months after abortion. Results At the 12th month, the expulsion was the most common reason for termination. The expulsion rates of genefix-group and γ-group were 2.48/100 women years and 3.12/100 women years, respectively (P>0.05). For the expulsion reasons, IUD moving down could account for more than seventy percent. The removal rate for IUD usage of two IUD groups were almost equal (3.91/100 women years verus 4.35/100 women years), the differences were not statistically significant (P>0.05). At the 90th day after abortion, comparing with control group, the bleeding and (or) spotting days of genefix-group and γ-group extended by 3.9 and 2.6 days respectively, the differences had statistical significance between the three groups (P<0.05). Among the bleeding and (or) spotting days, spotting days prolonged significantly. At the 12th month, spotting days of genefix-group andγ-group were (9.2±5.9) days and (8.5±4.6) days respectively, more than (5.2 ± 4.0) days of control group. The differences had statistical significance between IUD with control group (P<0.05), and had not between the two kinds of IUD (P>0.05). Conclusion The insertion of medicated genefix IUD and medicated γ IUD immediately after abortion is safe, feasible, has slight side effects and could be effective contraception.

OBJECTIVETo explore the surgical method and its clinical effects of minimally invasive osteosynthesis on the treatment of complex ankle fractures.

METHODSFrom January 2007 to December 2011, 53 patients with complex ankle fractures were treated with minimally invasive osteosynthesis. There were 31 males and 22 females, with an average age of 38.2 years old (ranged, 18 to 65). According to the system of Lauge-Hansen, 32 fractures were supination external rotation injury (grade WV), 13 fractures were pronation external rotation (grade III or IV), 5 fractures were pronation abduction (grade III); and 3 fractures can not be classified due to serious comminution fracture of fibula. According to the system of Denis-Weber, there were 4 cases with type A, 34 cases with type B and 15 cases with type C. Seven cases were open fractures. The duration from injuries to operation ranged from 2 hours to 14 days with an average of 5 days. The sequence of reduction and fixation of ankle fractures was firstly posterior malleolus, then medial malleolus and lateral malleolus, and inferior tibiofibular syndesmosis lastly. The fractures of posterior malleolus were reduced and fixed through anterior ankle approaches; the fractures of medial and lateral malleolus were percutaneously fixed with bolts or blade plate or tensile force band; and inferior tibiofibular syndesmosis were firmly fixed if necessary. Baird-Jackson scoring system was used to evaluate clinical effects.

RESULTSForty-eight patients were followed up from 10 to 36 months with an average of 13 months. The fractures got healing with an average time of 12 weeks (ranged, 10 to 18). According to the Baird-Jackson scoring system, the mean score of ankle function was 94.7 +/- 4.2, and 28 cases obtained excellent results, 15 good, 3 fair and 2 poor. One case experienced superficial infections and was cured by changing dressings, 2 cases experienced fixed syndesmosis screw breakage.

CONCLUSIONThe surgical method of minimally invasive osteosynthesis can ensure the anatomical join restoration, protect the blood supply of fracture end, rebuild the function of ankle joint, obtain satisfactory clinical results in treating complex ankle fractures.

Adolescent ; Adult ; Aged ; Ankle Fractures ; Ankle Joint ; physiopathology ; surgery ; Female ; Fracture Fixation, Internal ; methods ; Humans ; Male ; Middle Aged ; Minimally Invasive Surgical Procedures ; methods

Objective:To prepare and preliminarily identify the monoclonal antibodies(mAbs) specifically against 3D protein of Enterovirus 71(EV71),using bioinformatics to predict the epitopes of 3D,with HBc protein as a carrier.Methods: Artificial screening of 3D protein epitope sequences by bioinformatic method,inserted into the major immunodominant region(MIR) area of Hepatitis B virus core protein(HBc),to construct the recombinant protein.BALB/c mice were immunized with the recombinant virus like particles(VLPs),to prepare the mAbs against 3D protein of EV71.Affinity chromatography technology was used to purify the mAb.The indirect ELISA,ELISPOT,immunofluorescence and immunohistochemistry staining methods were used to identify the characteristic of the mAb.Results: We displayed 3D(aa34-43),3D(aa61-76) and 3D(aa151-164) epitopes by constructing fusion protein using HBc VLPs as a vector,after hybridization,one positive hybridoma cell line(3E1) was selected by ELISA.The isotype of 3E1 was IgG2a.The results of immunofluorescence and immunohistochemistry staining assay showed that the mAb 3E1 could specifically recognize EV71.Conclusion: The prepared mAb 3E1 can specifically recognizes the EV71,which laid the foundation for the detection of virus and further study on 3D protein,and verified the bioinformatics technology combined with HBc carrier displaying peptides could prepare mAb quickly and efficiently.

AIM: To study the survival, transfer and distribution of bone marrow CD34+/CD45+ cells from transgenic GFP mouse after transplanted into the completed transversional spinal cord rat model. METHODS: The bone marrow cells isolated from transgenic GFP mice were cultured in vitro. The cultured cells were identified by anti-CD34 and anti-CD45 monoclonal antibodies, and were transferred into the end of transection spinal cord. Paraformaldehyde was infused into the left ventricle of the rat model at the 24 h, 48 h, 1 week, 2 weeks, 4 weeks and 8 weeks after cell transplantation. Through sank and frozen, the spinal cord was sectioned at 10 ?m thickness. The green fluorescence positive cells were observed under the fluorescence microscope. CD34+/CD45+ cells were identified by immunohistochemistry staining. RESULTS: Green fluorescence positive cells were found at the head and the end of the completed transection part of spinal cord. Most of the green fluorescence positive cells were distributed in the gray substance of spinal cord. CD34+/CD45+ cells were found by immunohistochemistry staining. CONCLUSION: CD34+/CD45+ cells survived in spinal cord of SD rat, and migrated to the head of the transection part. The distance of migration was extended by the time.

2,4-Dichlorophenoxyacetic Acid ; analogs & derivatives ; toxicity ; Animals ; Epididymis ; growth & development ; metabolism ; Female ; HSP70 Heat-Shock Proteins ; biosynthesis ; genetics ; Herbicides ; toxicity ; Male ; Rats ; Rats, Wistar ; Spermatozoa ; drug effects ; growth & development ; metabolism

Objective: To establish a method for isolation and purification of fetal membrane derived adherent cells (FMDACs) , and investigate their biological characteristics. Method: FMDACs were isolated with trypsin inducing and cultured in vitro. FMDACs were induced to differentiate into osteoblasts and adipocytes. FACS and immunocytochemistry technique were used to examine the cell surface antigen. The genetic stability was verified by karyotype analysis. Results: FMDACs were successfully isolated and expanded in vitro. They had strong proliferative ability. FMDACs were positive for CD44 and CD29, but negative for CD34, CD14 and CD45. FMDACs were differentiated into osteoblasts and adipocytes after inducement. The karyotype was stable in the sixth-passaged FMDACs and the tumorigenicity was not found. Conclusion; FMDACs have the possibility of multipotent stem cells, which have strong capacities of self-renewal and multidirectional differentiation. The genetic background of FMDACs is stable. FMDACs may be used as a kind of novel seed cells for tissue engineering.

This study was designed to evaluate the effects of drilling through the growth plate and using adipose-derived stem cells (ADSCs) and bone morphogenetic protein-2 (BMP-2) to treat femoral head epiphyseal ischemic necrosis,which can be done in juvenile rabbits.Passage-four bromodeoxyuridine (BrdU)-labeled ADSCs were cultured,assayed with MTT to determine their viability and stained with alizarin red dye to determine their osteogenic ability.Two-month-old,healthy male rabbits (1.2 to 1.4 kg,n=45) underwent ischemic induction and were randomly divided into five groups (group A:animal model control;group B:drilling;group C:drilling & ADSCs;group D:drilling & BMP-2;and group E:drilling & ADSCs & BMP-2).Magnetic resonance imaging (MRI),X-ray imaging,hematoxylin and eosin staining and BrdU immunofluorescence detection were applied 4,6 and 10 weeks after treatment.Approximately 90％ of the ADSCs were labeled with BrdU and showed good viability and osteogenic ability.Similar results were observed in the rabbits in groups C and E at weeks 6 and 10.The animals of groups C and E demonstrated normal hip structure and improved femoral epiphyseal quotients and trabecular areas compared with those of the groups A and B (P＜0.01).Group D demonstrated improved femoral epiphyseal quotients and trabecular areas compared with those of groups A and B (P＜0.05).In summary,drilling through the growth plate combined with ADSC and BMP-2 treatments induced new bone formation and protected the femoral head epiphysis from collapsing in a juvenile rabbit model of femoral head epiphyseal ischemic necrosis.