As an effective method of studying soluble protein-protein interactions, yeast display system is now widely used for affinity maturation of single-chain antibodies.Due to the strong homology recombination machinery of yeast and the high-throughput nature of FACS detection, a rapid scan for interaction between antigen-antibody pairs could be easily achieved.Based on this system, a novel and reliable method for determining conformational epitopes was developed.Different fragments of macrophage migration inhibitory factor(MIF) and several point mutations of MIF were displayed on yeast cell surface using homologous recombination technology.Three MIF monoclonal antibodies, 10C3, 2A12 and 4E10, were screened for their binding affinity to each displayed peptide.Utilizing this technology, the key amino acids of MIF that bind to the MIF monoclonal antibodies were easily identified.

Tumor necrosis factor alpha(TNF-?) is a multi-functional cytokine that plays a significant role in many autoimmune diseases. The key biological functional domain of mouse TNF-? was determined by identifying the binding site of mouse TNF-? neutralizing antibody 9C6. Using yeast surface display technology, it was determined that 9C6 can recognize the linear amino acid fragment 29 ～40 of mature mouse TNF-? protein. Through mutagenesis experiments of this TNF-? region, the critical amino acids necessary for 9C6 binding were identified. Finally, wild-type mouse TNF-? and mutant variants that loses binding ability to 9C6 were expressed in Escherichia coli, purified, and used in a L929 cell killing assay. The assay results proved that the key amino acids for 9C6 binding were consistent with mouse TNF-? functional domain.

ObjectiveTo explore the influence of gender,age and types of peer acceptance on the ability of theory of mind among rural parent-absent children.Methods 312 rural parent-absent children participated in the study (including 162 boys,150 girls; 102 members of 8 year-old,119 members of 9 year-old,and 91 members of 10 year-old).Their ability of theory of mind and types of peer acceptance were examined by the 2nd-order False-beliefs Tasks and Peer nomination.The date were analyzed by Multivariate analysis of variance (ANOVA),LSD checkout and Simple effect analysis.ResultsThere were statistically significant effects on ages in theory of mind,types of peer acceptance ( F=4.57,P=0.011 ; F =3.82,P =0.005 ) and the interaction among gender,ages and types of peer acceptance(F=3.20,P =0.042; F =2.50,P =0.043 ).The least significant different(LSD) assay of the ability of theory of mind age and types of peer acceptance shows:there were statistically significant lies in the divergence of age 8 and age 9 group,and also in age 9 and age 10 group,(I-J =-0.51,P =0.016;I-J =- 0.90,P =0.000 ).In the same way,there were also statistically significant lies in the divergence of the ability of theory of mind performance-popular and rejected ( I-J =1.05,P =0.001 ) ; popular and neglected ( I-J =0.84,P =0.003 ) ; popular and controversial( I-J =1.03,P =0.002) ; rejected and average ( I-J =- 0.66,P =0.02 ) ; controversial and average ( I-J =- 0.64,P=0.034).Results of simple effect shows:there were statistically significant lies in the gender divergence of the 9 age group (F =4.98,P =0.026),the age divergence of girls (F =7.50,P =0.001 ),the gender divergence of controversial groups (F=5.76,P =0.017 ),the peer acceptance divergence of boys(F=4.97,P =0.001),and also the peer acceptance divergence of girls(F =2.54,P =0.04).Conclusion There is substantial connection between the ability of theory of mind of rural parent-absent children and ages and types of peer acceptance.

BACKGROUND:Cerebral ischemia often occurs in underlying pathological conditions, such as hypertension,
hyperlipidemia and diabetes. Therefore, it is of great significance to construct a cerebral ischemia rat model with hyperlipidemia and to study the effect of basic pathological changes on the cerebral ischemia.
OBJECTIVE:To observe the brain tissue pathological changes of rat models with coexistence of hyperlipidemia and cerebral ischemia, and the effect of hyperlipidmia on cerebral ischemia.
METHODS:The rats were fed with high-fat diet to establish the hyperlipidmia models, and then focal cerebral
ischemia models were prepared with suture method. At 3 and 7 days after modeling, the 2,3,5-triphenyltetrazolium chloride staining was used to observe the volume of brain tissue ischemia, and hematoxylin-eosin staining was
performed to observe pathological change of the margin of the brain tissue ischemia zone.
RESULTS AND CONCLUSION:2,3,5-Triphenyltetrazolium chloride staining staining results showed that the volume of cerebral ischemia was significantly reduced in the hyperlipidemia+cerebral ischemia 7 day group. Hematoxylin-eosin staining results showed there was typical ischemic changes in al the cerebral ischemia models, and the number of microglial cel s after cerebral ischemia for 7 days was significantly smal er than that after cerebral ischemia for 3 days, and the changes were more obvious in the hyperlipidemia+7-day cerebral ischemia group when compared with the hyperlipidemia+3-day cerebral ischemia group. Ultrastructure showed there were neuronal and glial nuclear membrane shrinkage in al the cerebral ischemia models, mitochondria cristae was disappeared completely, endothelial cel mitochondria was decreased, most of the synaptic vesicles of nerve synapse were dissolved;the damages above were improved after ischemia for 7 days, especial y
hyperlipidemia+cerebral ischemia for 7 days, the neuronal degeneration and necrosis were reduced, the
mitochondrial damage was repaired, the number of mitochondrial cristae was increased significantly, and the synaptic vesicles of nerve synapse were recovered significantly. The results indicate that hyperlipidemia can promote the recovery of cerebral ischemic injury,
probably because the hyperlipidemia factors can activate the protection mechanism.

Objective To explore the application effect of the adult education theory in the nursing round. Methods 340 trainee nurses, who were trained at our surgical department between June 2008 and December 2009, were randomly divided into the experimental group(178 cases)and the control group(162cases). The adult education theory was adopted in the experimental group, whereas the traditional education method was used in the control group, a questionnaire was carried out after the nursing visit. Results The experimental group was more satisfied with the round visit than the control group. Conclusions The application of the adult education theory in the round visit is beneficial not only to the students' selection of the topics of nursing visit, but also to the improvement of the students' understanding and communicating capacity, the students' studying interests and the students' creative thinking, finally the degree of satisfaction to the round visit is increased by the adoption of this theory.

Objective To observe bone formation of compound of autologous periosteum cells plus calcium alginate gelatin in animal body and investigate the best mixing ratio of the two components.Methods A total of 96 healthy adult New Zealand rabbits were randomly divided into six groups (Groups A,B,C,D,E and control group).The excised bilateral periostea from rabbits in each experimental group were all made into 0.4 ml cell suspensions.The compounds were prepared by blending 0.4 ml calcium alginate gelatin with 1,1/4,1/16,1/64 and 1/256 of the periosteum cell suspension respectively,and then were applied to autologous right radial defect area.Gross observation,X-ray examination and histological study were carried out at 2,4,8,12 weeks postoperatively.Levels of serum alkaline phosphatase and serum calcium were determined as well.Results The compounds containing periosteum cell suspension with cell counts of 1/4 or 1/16 and calcium alginate gelatin reached the best osteogenesis.Conclusion Compound of autologous periosteum cell suspension-calcium alginate gelatin induces obvious bone formation and is worthy of clinical application,for it has advantages of satisfactory bone defect repair and easy operation.

Objective To explore the effect of NPY on activation of primary microglia and the production of in?terleukin-1β. Methods Rat primary cortical microglia was cultured and divided into control group, LPS group, NPY+LPS group, NPY group and BIBP3226+NPY+LPS group. Microglia in control group were incubated with serum-free me?dium for 6 h;microglia in LPS group were incubated with serum-free medium plus LPS for 6 h;microglia in NPY+LPS group were incubated with serum-free medium plus NPY and LPS for 6 h; microglia cells in NPY group were incubat?ed in serum-free medium plus NPY for 6 h; microglia cells in BIBP3226+NPY+LPS group were incubated in se?rum-free medium including BIBP3226 、NPY and LPS for 6 h. After 6 h , Primary cultured microglia were stained us?ing IBA-1 antibody and examined under the fluorescence microscope. The protein levels of IL-1βin the culture media and the mRNA expression levels of IL-1βin the microglia of different groups were detected using the methods of Elisa and RT-PCR. Results After 6 h, the contents of IL-1 βin the culture media and the mRNA expression levels of IL-1βin the cells of LPS group increased remarkably compared with control group (P<0.05) and the microglia were activat? ed. Compared with LPS group, the contents of IL-1 βin the culture media. the mRNA expression levels of IL-1β and the activity of microglia in LPS+NPY group were significantly decreased .Compared with LPS+NPY group, the contents of IL-1βin the culture media. the mRNA expression levels of IL-1β and the activity of microglia in BIBP3226+NPY+LPS group were increased (P<0.05). There were no significant differences in the contents of IL-1βin the culture media. the mRNA expression levels of IL-1βand the activity of microglia between BIBP3226+NPY+LPS group and LPS group or between NPY group and the control group. Conclusion NPY can inhibit the biological activity of microglia and IL-1βproduction through NPY Y1 receptorin the microglia.

Objective To investigate the relationship between rs17525495 locus polymorphism of leukotriene A4 hydrolase (LTA4H) gene and the severity of tuberculous meningitis (TBM). Methods A total of 184 TBM patients from Department of Neurology, the Second Hospital of Hebei Medical University from January 2014 to October 2016 were selected as research subjects. According to the British Medical Research Council criteria, the severity of TBM patients was divided into three stages. The single nucleotide polymorphism rs17525495 of LTA4H gene was sequenced, and the general case data, clinical manifestations and results of lumbar puncture were analyzed. Results There were 91 cases (49.5％) of CC genotypes of rs17525495 locus in LTA4H gene of 184 cases, 75 cases (40.8％) of CT genotypes and 18 cases (9.8％) of TT genotypes. The frequency of allele C was 69.8％ and T was 30.2％. Patients with different genotypes were compared for their severity, clinical manifestations and lumbar puncture results. Among CC patients, the proportion of stage Ⅰ patients(54.9％, 50/91)was higher than that of stage Ⅱ(22.0％, 20/91)and Ⅲ(23.1％, 21/91). Among TT patients, the proportion of patients with stage Ⅱ(8/18)and Ⅲ(8/18)was higher than patients with stageⅠ(2/18)(χ2=15.898,P=0.003). The incidence of headache, fever, nausea and vomiting, neck stiffness, epilepsy and disturbance of consciousness was statistically analyzed. Compared with CC and CT patients, the incidence of fever(TT:13/18,CC:42/91,CT:50/75,χ2=8.932,P=0.011)and neck stiffness(TT:12/18,CC:38/91,CT:46/75,χ2=7.993,P=0.018)was higher in TT patients. Headache, nausea and vomiting, disturbance of consciousness, and the incidence of epilepsy showed no statistically significant difference. And there was no statistically significant difference in lumbar puncture pressure, chloride, protein and glucose between different genotypes. Conclusion TBM patients with mild illness frequently prompt LTA4H gene rs17525495 locus for the CC type;while patients with severe disease prompt TT type.

Objective:To investigate the mechanism of crush syndrome (CS) induced by crush injury on myocardial cells in rats.Methods:Thirty two male Sprague Dawley (SD) rats were randomly divided into control group, CS-0 group, CS-12 group and CS-24 group with 8 rats in each group. CS model was made by self-made extruder and perfused with 4% paraformaldehyde for 0, 12 and 24 h. The morphological changes of myocardial tissue were observed by hematoxylin staining. The apoptosis of cardiomyocytes was detected by terminal dexynucleotide transferase-mediated nick end labeling (TUNEL). The levels and activities of malondialdehyde (MDA), superoxide dismutase (SOD), lactose dehydrogenase (LDH), interleukin-6 (IL-6), interleukin-1 β (IL-1 β), tumor necrosis factor-α (TNF- α) in myocardial homogenate were detected by enzyme-linked immunosorbent assay (ELISA). The expressions of Caspase-3, Bax, Bcl-2 and necrosis factor-κB (NF-κ B) were detected by Western blot.Results:Compared with the control group, the myocardial tissue of CS model group had different degrees of morphological damage; compared with the control group, the apoptosis rate, Caspase-3 and Bax protein expression levels of CS-0 group, CS-12 group and CS-24 group were significantly increased ( P<0.05), and the expression level of Bcl-2 protein was significantly decreased ( P<0.05); compared with the control group, the levels of MDA, LDH, IL-6, IL-1β, TNF-α and p65 protein phosphorylation in the myocardial homogenate of CS-0 group, CS-12 group and CS-24 group were significantly increased ( P<0.05), and SOD activity was significantly decreased ( P<0.05). Conclusions:CS may inhibit oxidative stress and induce inflammatory reaction by activating NF-κ B pathway, thus damaging myocardial cells in rats.

Objective To construct a risk prediction model of pulmonary involvement based on chest CT and clinical feature in patients with primary Sjogren's syndrome(pSS),and to explore the risk prediction value of the model.Methods A total of 360 pSS patients who had been treated at Handan Hospital of Traditional Chinese Medicine from October 2020 to August 2023 were retrospectively selected as study objects,and were then divided into a modeling group(252 patients)and a verification group(108 patients)according to a ratio of 7∶3.The patients in the modeling group were divided into a control group(201 patients)and an involvement group(51 patients)based on presence or absence of lung involvement.The data on clinical characteristics and features of chest high-resolution CT(HRCT)in the modeling group was collected.Univariate analysis was performed among the groups to determine the relevant factors affecting lung involvement in pSS patients.Binary logistic regression analysis was performed on related factors to screen independent risk factors.A prediction model was established based on the independent risk factors.A verification and value analysis of the column-line prediction model were completed through data collection of the verification group.Results Age,disease course,cough,Raynaud's phenomenon,C-reactive protein(CRP),anti-SSA antibody,and HRCT were the relevant factors affecting lung involvement in pSS patients(all P＜0.05).Further binary logistic regression analysis showed that old age,prolonged disease course,cough and abnormal HRCT imaging were independent risk factors for lung involvement in SS patients(all P＜0.05).A nomogram risk prediction model was constructed based on independent factors.The model verification results indicated that the calibration chart showed better performance in the prediction model.The AUC of the area under the receiver operating characteristic(ROC)curve was 0.993 the modeling group and 0.995 in the validation group.Conclusions The clinical characteristics and the results of chest CT are closely related with lung involvement in patients with pSS.Old age,prolonged disease course,cough,and abnormal HRCT imaging are independent risk factors affecting lung involvement in patients with pSS.The prediction model established on this basis has a higher predictive value for the occurrence of lung involvement in patients receiving after-loading radiotherapy.