Objective To constructan eukaryotic expression recombinant plasmid named pEGFP-N2-PRNP .Methods Total RNA was extracted from alzheimer (AD) disease peripheral blood ,and the PRNP gene was amplified by reverse transcription-poly-merase chain reaction(RT-PCR) .By using gene recombination technique ,human PRNP cDNA was inserted into retroviral vector pEGFP-N2 .The recombinant plasmid was identified by a pair of specified primers containing the restriction sites of Xho Ⅰ and EcoRⅠ .Results The PRNP gene could be obtained by RT-PCR ,the recombinant plasmid was identified by restriction endonucle-ase analysis ,PCR and sequence analysis ,and the expression vector pEGFP-N2-PRNP ,which could be stably expressed in SH-SY5Y cells .Conclusion The recombinant plasmid pEGFP-N2-PRNP is constructed successfully ,Which offers a basic for the further re-search on PRNP biological fuction .

OBJECTIVE: To observe the effects of Kangfengshi Granules (KFSG) on expressions of the mRNAs of osteoprotegerin (OPG), receptor activator of nuclear factor-kappaB ligand (RANKL) and macrophage colony stimulating factor (M-CSF) in bone tissues of rats with collagen-induced arthritis. METHODS: Forty SD rats were randomly divided into four groups: normal control group, untreated group, cyclosporine A (CsA)-treated group and KFSG-treated group. Except the rats in the normal control group, all the other rats received subcutaneous injection of collagen II to establish collagen-induced arthritis (CIA) models. Then the rats in each group were fed normal saline or corresponding drugs for four weeks. Total RNA was extracted from carpal and digital bones. The expressions of OPG, RANKL and M-CSF mRNAs were examined by real-time PCR. RESULTS: The total incidence of arthritis induced by collagen II in the rats was approximately 90%. The expression levels of RANKL and M-CSF mRNAs and the RANKL mRNA/OPG mRNA ratio in the untreated group, KFSG-treated group and CsA-treated group were all significantly higher than those in the normal control group, while the expression levels of OPG mRNA in those three groups were significantly lower than that in the normal control group. The expression level of OPG mRNA in the KFSG-treated group was obviously higher while the expression level of M-CSF mRNA and the RANKL mRNA/OPG mRNA ratio in the same group were both lower as compared with those in the untreated group. CONCLUSION: The molecular mechanism of effects of KFSG on bone erosion and destruction induced by rheumatoid arthritis is closely correlated with up-regulating the expression of OPG mRNA, down-regulating the expression of M-CSF mRNA and RANKL mRNA/OPG mRNA ratio.

BACKGROUND:Natural colagen is considered to have low immunogenicity and good biocompatibility relatively. OBJECTIVE:To evaluate the immunogenicity of animal-based colagenin vitro. METHODS:Type I colagen was extracted from bovine tendon after immunogenicity removal. The colagen purity was detected by high performance liquid chromatography and residual DNA was measured quantitatively by fluorescence staining. Fifty BALB/c mice were randomly divided into five groups: subcutaneous injection of normal saline solution (negative control), bovine colagen (positive control), 33.4, 66.8, 133.4 mg/kg of bovine tendon colagen, respectively, once a day. After 12 days of continuously subcutaneous injection, lymphocyte proliferation, and cel classification and NK cel kiling function of mice were detected; after 3 weeks of continuous injection, the spleen, liver, spleen and lung tissue of mice were taken for histological examination. RESULTS AND CONCLUSION:Compared with the standard type I colagen, the purity of purified type I bovine colagen reached more than 99%, but the residual DNA was below 1 mg/L which was far less than the residue level of conventional cel-free DNA in the matrix (dry weight: 50-100 μg/g). After 12 days of continuous injection, there were no changes in lymphocyte proliferation, NK cel kiling function and the proportion of lymphocyte subsets. After 3 weeks of injection, the spleen and lymph sheath of mice around the smal artery became thickened in the 66.8 and 133.6 mg/kg bovine tendon colagen groups, which could cause accidental liver injury and lung injury, but the splenic corpuscle germinal center area had no change. These findings indicate that continuously subcutaneous injection of animal-based colagen can cause the lower lymphocyte immune response to the spleen of BALB/c mice, which may cause accidental liver and lung injuries.

Objective To approach the risk factors of bronchopulmonary dysplasia(BPD)with very low birth weight infants whose gestational age less than 32 weeks,thus it could provide a basis direction for prevention.Methods To summarize 70 neonates with BPD in October 2012 to October 2014 in our hospital, and randomly select 70 very low birth weight infants didn't have oxygen requirement of gestation age less than 32 weeks as control group.The perinatal risk factors,oxygen therapy and use caffeine were analyzed by using the statistical analysis of Chi-square test and logistic regression,so the risk factors and prevention direc-tion of BPD could be provided.Results Male gender were more prone to BPD(P ﹦0.000).Gestational age (P ﹦0.000)and birth weight(P ﹦0.002)were statistical lower in infants with BPD compared with no BPD.Family history of asthma,fetal distress,amniotic fluid turbidity,pulmonary hemorrhage,respiratory dis-tress syndrome,use of pulmonary surfactant,patent ductus arteriosus,intrauterine infection,ventilator time,na-sal continuous positive airway pressure /nasal intermittent positive pressure ventilation time,duration of oxy-gen therapy and use of caffeine were statistical significances (P 〈0.05,respectively).Logistic regression analysis demonstrated that gender(OR ﹦3.574,P ﹦0.003),gestational age≥28 weeks(OR ﹦58.665,P ﹦0.002), birth weight 〈1 250 g(OR ﹦36.453,P ﹦0.012)and ventilator time(OR ﹦703.696,P ﹦0.000)were independent risk factors for BPD infants.Using of caffeine(OR ﹦0.025,P ﹦0.010)and nasal continuous positive airway pressure/nasal intermittent positive pressure ventilation(OR ﹦0.004,P ﹦0.002)were protective factors for BPD infants. Conclusion The incidence of BPD could be reduced by strengthening prenatal care,minimizing intrauterine in-fection and preterm delivery,reducing the patent ductus arteriosus by strict fluid management,using mechanical ventilation rationally,choosing a non-invasive mechanical ventilation and caffeine.

Objective To investigate the clinic signification of newborn hearing screening combined with deafness susceptibility genes screening. Methods 1 440 newborns(3 ~ 5 days after birth) were screened for 8 hot spot hearing loss associated mutations from GJB2, mt12S rRNA and SLC26A4. At the same time, all infants received hearing screening. Those who failed to pass two-step test were referred to further audiological assessment. Results The carrier rate of commonmutations was 1.46% for GJB2 c.235delC, 0.35% for GJB2 c.299-300delAT, 0.42% for mt12S rRNA c.1555A > G, 0.42% for SLC26A4 c.IVS7-2A > G and 0.14% for SLC26A4 c.2168A > G. The total carrier rate was 2.78%. 10 infants were diagnosed as hearing loss in the hearing screening and follow-up audiology assessment (6.94‰) and 5 were diagnosed as severe hearing loss (3.47‰). 32 hearing loss associated mutation carriers passed the hearing screening. Conclusions Genetic screening of newborn hearing screening can be helpful to find out neonates with late-onset and progressive hearing impairment, which were significant for early intervention, regular follow-up and reduction of deafness.

Objective To explore the curative effects of unilateral Orthofix external fixation in the treatment of tibial defects.Methods From June 2012 to February 2016,we treated 19 patients with tibial defects using unilateral Orthofix external fixation.They were 14 men and 5 women,aged from 26 to 79 years (mean,43.0 years).The right tibia was involved in 12 cases and the left tibia in 7.The soft tissue defects averaged 6.4 cm × 3.7 cm.The mean bone defect before bone transport was 8.9 cm in length.The anteroposterior and lateral X-ray films of the affected tibia were taken regularly after surgery.Edwards tibial fracture scores were used to evaluate the functional recovery and complications of the affected limb at final follow-ups.Results All the patients were followed up for 10 to 40 months (average,18.5 months).Bone union time ranged from 4 to 16 months (average,9.0 months).Lengths of bone transport ranged from 6 to 10 cm (average,7.8 cm).All the bone defects were reconstructed.According to the Edwards scoring at the final follow-ups,10 cases were rated as excellent,6 as good and 3 as poor.Complications were observed in 4 cases,giving a complication rate of 21.1％.Conclusion Unilateral Orthofix external fixation and bone transport can lead to preferable curative effects in the management of tibial defects.

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This study was conducted to find perfect temperature sintered bone as carrier of bone morphogenetic protein (BMP). The different temperature active sintered bones, which were made up of calcine bone and bone morphogenetic protein, were implanted into the defects of rabbit radius. Compared with the sintered bone of 600 degrees C, the sintered bone of 900 degrees C and 1200 degrees C could induce more pieces of bone formation and be replaced by new bone. There were more pieces of new bone formation in sintered bone of 900 degrees C and 1200 degrees C than those in sintered bone of 600 degrees C (P<0.05). There was no difference between the sintered bone of 900 degrees C and 1 200 degrees C (P>0.05). In comparison with the sintered bone of 600 degrees C and 1200 degrees C, the sintered bone of 900 degrees C may be the choicest carrier of bone morphogenetic protein.
Animals ; Bone Morphogenetic Proteins ; chemistry ; Bone and Bones ; chemistry ; Drug Carriers ; chemistry ; Female ; Hot Temperature ; Male ; Rabbits ; Temperature

Objective:To measure the safe range from the foramen rotundum to the siphon of internal carotid artery using three-dimensional reconstruction technique, and to provide guidance for operation through pterygopalatine fossa.Methods:The skulls of 121 volunteers were scanned to get the final results with thin-section computed tomographic images.The position of the siphon of internal carotid artery (point A)and foramen rotundum (pointB)were ascertained.Three-dimensional reconstruction technique was used to build a coordinate system paralleled to the frankfort horizontal plane and the nasal septum plane.The coordinate system took point A as the coordinate origin.Point C and point D were the projections of point A in two planes parallel with frankfort horizontal plane and nasal septum plane which included point B.The distances of AC,AB,and BC were measured. The angles of the line went through A and B to the three planes were also measured.Results:The distance of AC was measured as 13.22 (3.79)mm (range,8.33 - 105.67 mm;95%CI:8.55 - 21.39 mm).The angle to the sagittal plane was measured as 33.54 (9.23)° (range,5.38- 66.58°;95%CI:30.88 - 34.20°). The angle to the coronal plane was measured as 53.17 (10.48)°(range,5.60-75.02°;95%CI:51.29-55.06°).The angle to the horizontal plane was measured as 9.43 (12.91 ) mm (range,- 28.44 - 82.22;95% CI:7.11 - 11.76 ). Conclusion:The safety distance from foramen rotundum to the siphon of internal carotid artery in the operation through pterygopalatine fossa (PPF) under nasoendoscope is obtained by thin-section computed tomographic images.

Objective: To explore the feasibility and clinical effect of single bundle anatomic intrathecal reconstruction of proximal injury of anterior cruciate ligament (ACL) using dissociate peroneus longus tendon combined with semitendinosus tendon. Methods: From January 2015 to September 2016, a total of 24 patients with proximal injury of ACL, confirmed by arthroscopy, were admitted to the sports medicine department of our hospital. The ACL was completely ruptured from the proximal footprint and the tibial side residual remained intact. There were 19 males and 5 females; 16 cases on the right side and 8 cases on the left side. The mean age was 27.88±7.13 years old; The interval between injury and surgery was 14.83±9.09 d; The dissociate peroneus longus tendon and semitendinosus tendonfrom the injured extremity were folded in half, then braided and trimmed into ACL graft for use. The ACL remnant was preserved and the graft was pulled through the stump for single bundle anatomic intrathecal reconstruction. The ACL graft was fixed with Endobutton on the femoral side and interference screw on the tibial side. The results of Lachman test, Lysholm scores, Tegner scores and International Knee Documentation Committee (IKDC) subjective scores of knee before operation and at final visit were recorded to evaluate the stability and function of the knee. Visual analogue score (VAS) was used to record the changes of pain at the site where the peroneal longus tendon was harvested. The American Orthopaedic Foot and Ankle Society (AOFAS) Ankle-Hindfoot Scores were recorded before injury and at the final visit to assess the effect of ankle function after peroneal longus tendon resection. Results: The diameter of the ACL graft made of dissociate peroneus longus tendon and semitendinosus tendon is 8.88±0.30 mm. All patients were followed up at the outpatient clinic, with an average of 34.38±5.40 months, and no serious complications such as rerupture and joint infection were found. There were 16 grade B, 6 grade C and 2 grade D for preoperative Lachman test, none with hard end point. At last vist, there were 23 cases of grade A and 1 grade B, all with hard end points for Lachman test. At the preoperative and final visit, the Lysholm scores of the knee joint were 35.20±11.92 and 94.29±2.92 (t=23.850, P=0.000). Tegner scores were 3.46±0.93 and 8.04±1.00 (t=16.653, P=0.000). The subjective IKDC scores of knee joint were 47.63±13.06 and 91.71±3.75 (t=15.972, P=0.000). At the final visit, all the indicators of the knee were improved compared with those before surgery, and the difference was statistically significant.The AOFAS scores before injury and at the final visit were 98.83 (2.78) and 98.17 (4.01), respectively, with no statistically significant difference (t=1.850, P=0.076) . The VAS scores for the harvest of theperoneus longus tendon were 4.50±1.41, 0.54±0.65 and 0.29±0.55 immediately and 6 months after the operation and at the last visit.There was statistically significant difference between the scores immediately and 6 months after the operation (t=14.900, P=0.001).There was statistically significant differences between the scores at 6 months after the operation and the final visit (t=2.770, P=0.011). Conclusion Single bundle anatomic intrathecal reconstruction of proximal injury of ACL using dissociate peroneus longus tendon and semitendinosus tendon is feasible with good clinical effect.