Objective To evaluate the effect of tissue-engineered nerve graft combined with tetramethylpyrazine in the repair of ischiadic nerve defect in rats.Methods Fifty-five adult SD rats were allocated into groups A (n =15),B (n =15),C (n =15) and D (n =10),according to the random number table.All experimental sciatic nerves were at the right hind side.After the rats were anesthetized with 10 g/L pentobarbital through abdominal injection,a 1.5 cm nerve sciatic nerve was excised at the point 0.5 cm away from the lower margin of the piriformis.Four grafts were used to bridge the nerve defect,including tissue-engineered nerve transplanted neural stem cells (NSCs) that were cultured in medium containing tetramethylpyrazine for 1 week (group A),tissue-engineered nerve transplanted NSCs that were cultured in tetramethylpyrazine-free medium for 1 week (group B),acellular nerve allograft (group C),and nerve autograft (group D).After operation,group A was administered tetramethylpyrazine for 12 weeks in the operative site.For other groups,the same volume of normal saline was used instead of tetramethylpyrazine.Repair results were detected with the sciatic function index (SFI),nerve electrophysiological evaluation,fluorescent microscopy,horseradish peroxidas (HRP) retrograde tracer and nerve fiber regeneration study.Results At postoperative 2 weeks,no differences were found in SFI and electrophysiological evaluation among the groups,there was an intensive fluorescence in groups A and B and no fluorescence in group C,and large nestin-positive cells were observed in groups A and B.At postoperative 12 weeks,SFI was-17.52 ± 2.41 in group A,-25.74 ± 2.85 in group B,-36.12 ± 3.41 in group C and-15.87 ± 2.26 in group D;electrophysiological evaluation showed nerve conduction velocity of (9.43 ± 0.40) m/s in group A,(7.76 ± 0.31) m/s in group B,(5.87 ± 0.67) m/s in group C and (10.16 ± 0.39) m/s in group D;fluorescence microscope and histological staining showed NSCs could survive in vivo,migrate and differentiate into neurons and gliocytes;HRP-positive neurons were 885.40 ± 19.91 in group A,684.57 ± 38.37 in group B,390.33 ± 43.41 in group C and 941.67 ± 32.54 in group D;regenerated nerve fibers,often myelinated were large in group A.All the measures in group A were better compared to groups B and C (P ＜ 0.05),but didn't differ from those in group D (P ＞0.05).Conclusion Tissue-engineered nerve transplanted NSCs cultured in medium including tetramethylpyrazine can effectively repair rat sciatic nerve defect,and promote peripheral nerve regeneration and lower limb motor function recovery.

BACKGROUND:Acellular nerve scaffolds have the three-dimensional structure of natural nerves and low immunogenicity,but their effect on long nerve defects is still not ideal.Therefore,it is necessary to construct tissueengineered nerve using acellular nerve and seed cells in order to improve the therapeutic effect.OBJECTIVE:To systemically review the efficacy of combination of acellular nerve grafts (ANGs) and mesenchymal stem cells (MSCs) or Schwann cells (SCs) transplantation in the treatment of sciatic nerve defects in a rat model.METHODS:Randomized controlled trials (RCTs) about the effects of combination of ANGs and MSCs or SCs transplantation for sciatic nerve defects in rats were searched in PubMed,The Cochrane Library,EMbase,CNKI,WanFang and VIP from inception to July 2016.Three reviewers independently screened literature according to the inclusion and exclusion criteria,extracted data,and assessed the risk of bias of included studies.Then,a Meta-analysis was performed using Review Manger5.3 software.RESULTS AND CONCLUSION:A total of 10 RCTs involving 252 rats were included.The results of meta-analysis showed that:compared with the control group (simple acellular nerve scaffold group),the sciatic functional index (SFI) of the combined group (combination of ANGs and MSCs or SCs transplantation) were superior at 2 weeks [SMD=2.73,95％ CI (1.92,3.45),P ＜ 0.000 01],4 weeks [SMD=4.57,95％ CI (3.43,5.70),P ＜ 0.000 01],6 weeks [SMD=1.62,95％CI (0.18,3.06),P=-0.03],8 weeks [SMD=4.90,95％ CI (2.96,6.84),P ＜ 0.000 01] after surgery.The nerve conduction velocity [SMD=1.39,95％ CI (0.99,1.78),P ＜ 0.000 01),latency period (MD=-0.98,95％ CI (-1.19,-0.76),P ＜ 0.000 01],and amplitude [SMD=1.23,95％ CI (0.62,1.85),P ＜ 0.000 1] were superior at 12 weeks after surgery.The myelin sheath thickness was superior at 8 weeks [MD=0.14,95％ CI (0.07,0.21),P ＜ 0.000 1],12 weeks [SMD=1.85,95％ CI (1.63,2.08),P ＜ 0.000 01] and the number of myelinated nerve fibers were superior at 12 weeks [SMD=3.59,95％CI (2.63,4.55),P ＜ 0.000 01] after surgery.The gastrocnemius wet weight was superior at 8 weeks after surgery [SMD=4.22,95％ CI (2.40,6.03),P ＜ 0.000 01].Current evidence indicates that the combination of ANGs and MSCs or SCs can promote the regeneration and functional recovery of the peripheral nerve.Due to the limited quality of the included studies,the above conclusion should be verified by conducting high-quality and large-scale RCTs.

BACKGROUND: Ipsilateral femoral shaft and neck fracture is a rare clinical injury, and its treatment has attracted more and more attention from scholars. At present, there are many internal fixation methods. OBJECTIVE: To retrospectively analyze efficacy of three internal fixation methods in the treatment of ipsilateral femoral shaft and neck fracture, and its indications. METHODS: Thirty-three cases of ipsilateral femoral neck and shaft fracture treated by three different internal fixation implants between January 2015 and September 2018 were retrospectively studied. Patients were divided into three groups according to different internal fixation: intramedullary nail only group (group A; n=11), retrograde nail with cannulated screw group (group B; n=10) and plate with cannulated screw group (group C; n=12). The general clinical data, the intraoperative situations, the postoperative follow-up conditions and complications were compared among the three groups. RESULTS AND CONCLUSION: (1) There was no significant difference between the three groups with regard to age, gender, cause of injury, AO fracture classification, Garden classification and average time from injury to surgery (P > 0.05). (2) There were statistically significant differences among the three groups in incision length (P < 0.05); no significant difference in incision length was found between the groups A and B (P > 0.05); the incision length was smaller in groups A and B than in group C (P < 0.05). There were no significant differences among the three groups in operation time and intraoperative blood loss (P > 0.05). (3) There were no significant differences among the three groups in time to bony union in both femoral shaft and neck, complication rate, and excellent and good rate of Harris hip score (P > 0.05). (4) Femoral neck fracture delayed healing occurred in one case of the group A; re-fracture occurred in one case of the group B; and the fracture end was located under the femoral tuberosity; screw loosening occurred in one case of the group C. (5) It is indicated that all three internal fixation methods can achieve good therapeutic effects. The operator can choose the skilled internal fixation method according to the specific conditions of the patient.

Objective: To explore the relationship between adolescents physical fitness and depressive symptoms, and to provide reference for the early prevention and intervention of depressive symptoms and improvement of physical fitness in Chinese adolescents. Methods: From September to December 2021, a total of 8 102 adolescents were selected by random cluster sampling method in Shanghai, Urumqi, Changsha and Kunming. The Center for Epidemiologic Studies Depression Scale(CES-D) was used to investigate the depressive symptoms, and completed grip strength, standing long jump, 50 m running, modified sitting forward flexion, 20 s repeated traverse, 30 s sit ups, 20 m round trip running (20 m SRT) test. χ 2 test, Goodman Kruskal Gamma and Logistic regression analysis were used to analyze the relationship between physical fitness index(PFI) and depressive symptoms. Results: The overall detection rate of depressive symptoms in adolescents with high level PFI was 23.4%, and the detection rate of low level adolescents was 26.3%, with a statistically significant difference ( χ 2=6.73, P =0.01). There was a significant positive correlation between PFI and depressive symptoms in the high school group ( G=0.09, P <0.05) and the boy group ( G=0.12, P < 0.05 ), and there was no significant association between PFI and depressive symptoms in the junior high school group and the girl group ( P >0.05). After adjusting for gender and age in the Logistic regression model, compared with those with high PFI, the risk of depressive symptoms in those with low PFI was 1.18 times (95% CI =1.05-1.33). Conclusion There is a correlation between physical fitness and depressive symptoms in adolescents. Adolescents with low PFI are at higher risk of developing depressive symptoms than those with high PFI.

BACKGROUND:Pauwels Ⅲ femoral neck fracture is a typical unstable fracture characterized by high vertical shear force and high incidence of postoperative complications.At present,there are many fixation methods for Pauwels Ⅲ fracture,and there is no clear conclusion as to which internal fixation is the best fixation method for Pauwels Ⅲ femoral neck fracture in young adults. OBJECTIVE:To compare the clinical effect of three inverted triangle cannulated screws assisted transverse lag screws and femoral neck system in fixing Pauwels Ⅲ femoral neck fractures of young adults. METHODS:From May 2021 to December 2022,21 young and middle-aged patients with Pauwels Ⅲ femoral neck fracture were treated with three inverted triangle cannulated screws assisted by transverse lag screws and femoral neck system in Affiliated Hospital of Southwest Medical University.Of them,9 patients were treated with three inverted triangle cannulated screws and one transverse lag screws perpendicular to the fracture line as the 3+1 cannulated screw group and 12 patients were treated with femoral neck system as the femoral neck system group.The two fixation methods were compared in terms of operation time,intraoperative blood loss,total incision length,intraoperative fluoroscopy times,fracture healing time,and limb function. RESULTS AND CONCLUSION:(1)All patients were followed up.Patients in the 3+1 cannulated screw group were followed up for 10-25 months,with a mean of(17.44±4.30)months.The patients in the femoral neck system group were followed up for 8-24 months,with a mean of(15.58±4.68)months.(2)The intraoperative fluoroscopy times and Harris score at 3 months postoperatively in the femoral neck system group were better than those in the 3+1 cannulated screw group,and the difference was statistically significant(P<0.05).The intraoperative blood loss,total incision length,and femoral neck shortening distance in the 3+1 cannulated screw group were better than those in the femoral neck system group,and the differences were statistically significant(P<0.05).There was no significant difference in operation time,fracture healing time,and Harris score at the last follow-up between the two groups(P>0.05).(3)It is indicated that three inverted triangle cannulated screws assisted transverse lag screw and femoral neck system can achieve good clinical effects in the treatment of young and middle-aged Pauwels Ⅲ femoral neck fracture.The femoral neck system has fewer intraoperative fluoroscopy times and better early overall stability,while the 3+1 cannulated screw is more minimally invasive,easier to operate.Both fixation methods are worthy of clinical application and promotion,and can be selected according to the actual clinical situation.

Objective To observe biomechanical characteristics and advantages of the self-developed anatomical locking plate of sternoclavicular joint by comparison with the radial distal oblique T-shaped locking plate and sternoclavicular hook plate. Methods Nine embalming and moistening adult corpses were selected, including 6 males and 3 females. Model of complete dislocation of the sternoclavicular joint caused by complete removal of the sternoclavicular joint specimen. The bilateral sternoclavicular joints of 9 specimen models were randomly numbered, matched and divided into Group A(experimental group, sternoclavicular joint anatomical locking plate), Group B(control group 1, the radial distal oblique T-shaped locking plate) and Group C(control group 2, sternoclavicular hook plate), with 6 sternoclavicular joints in each group. All specimens were placed with steel plates on both sides and fixed on the universal mechanical test machine. Three biomechanical experiments were carried out, including loading of distal clavicle, torsion of distal clavicle and anti-pull out of sternum handle screw. Results Distal clavicle loading test: the load-displacement of specimens in three groups showed a linear relationship. The compressive deformation resistance in experimental group was stronger than that in two control groups. Distal clavicle torsion test: the relationship between torque and torsion angle was linear. The torsional deformation resistance in experimental group was stronger than that in two control groups. Anti-pullout test of sternum handle screw: there was a significant difference in the maximum anti-pullout force of sternum handle screw among the three groups (P＜0.05). The anti-pullout performance of the sternum handle screw in the experimental group was better than that in the two control groups. Conclusions The self-developed sternoclavicular joint anatomical locking plate is superior to the oblique T-shaped locking plate of distal radius and the plate of sternoclavicular hook in terms of anti-compression, anti-torsion and anti-pullout of sternoclavicular screw, so as to provide an ideal internal fixation device for the treatment of fracture and dislocation of sternoclavicular joint.

Biological sample pretreatment is an important step in biological sample analysis. Due to the diversity of biological matrices, the analysis of target substances in these samples presents significant challenges to sample processing. To meet these emerging demands on biopharmaceutical analysis, this paper summarizes several new techniques of on-line biological sample processing: solid phase extraction, solid phase micro-extraction, column switching, limited intake filler, molecularly imprinted solid phase extraction, tubular column, and micro-dialysis. We describe new developments, principles, and characteristics of these techniques, and the application of liquid chromatography-mass spectrometry (LC-MS) in biopharmaceutical analysis with these new techniques in on-line biological sample processing.

Zidovudine (AZT), the first drug approved by the US Food and Drug Administration for the treatment of human immunodeficiency virus (HIV) infection, is metabolized in the host cells to 5'-AZT triphosphate (AZT-TP) which inhibits HIV reverse transcriptase. As the pharmacokinetics of AZT and its phosphorylated metabolites in human peripheral blood mononuclear cells (hPBMCs) is limited, the aim of this study was to determine the pharmacokinetic parameters of AZT and its phosphorylated metabolites in hPBMCs from 12 healthy Chinese male subjects after a single oral dose of 600 mg of AZT. Blood samples were collected prior to drug administration, then at 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 5, 6, 8 and 10 h after drug administration. Mononuclear cells collected by Ficoll-Hypaque density gradient centrifugation were used for determination of AZT and metabolites [AZT monophosphate (AZT-MP), AZT diphosphate (AZT-DP) and AZT-TP] and the plasma was used to evaluate the pharmacokinetics of AZT. Plasma concentration of AZT peaked within 0.583 h and intracellular concentrations of AZT, AZT-MP, AZT-DP and AZT-TP peaked within 1.083, 1.500, 1.417 and 1.583 h, respectively. AZT in plasma was eliminated rapidly with t 1/2 of 2.022 h, and AZT-MP, AZT-DP and AZT-TP were eliminated with t 1/2 of 13.428, 8.285 and 4.240 h, respectively. The plasma concentration of the phosphorylated metabolites was not quantifiable.

Objective: To isolate a bacteriophage against pan-drug resistant Klebsiella pneumoniae in a burn patient, and to study its biological characteristics, genomic information, and effects on bacterial biofilm. Methods: (1) In 2018, pan-drug resistant Klebsiella pneumoniae UA168 (hereinafter referred to as the host bacteria) solution isolated from the blood of a burn patient in Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine (hereinafter referred to as Ruijin Hospital) was used to isolate and purify the bacteriophage against pan-drug resistant Klebsiella pneumoniae from the sewage of Ruijin Hospital with sewage co-culture method, drip plate method, and double-agar plate method. The bacteriophage was named as phage KP168 and the plaque morphology was observed. (2) The phage KP168 solution was taken for cesium chloride density gradient centrifugation and dialysis, and then the morphology of phage KP168 was observed through transmission electron microscope after phosphotungstic acid negative staining. (3) The phage KP168 solution was taken to determine the lytic ability of the phage KP168 against 20 strains of pan-drug resistant Klebsiella pneumoniae isolated from the burned patients′ blood in Ruijin Hospital by the drip plate method, and then the lysis rate was calculated. (4) The phage KP168 solution at a initial titer of 9.3×10¹¹ plaque-forming unit (PFU)/mL (400 μL per tube) and the host bacteria solution at a concentration of 1×10⁹ colony-forming unit (CFU)/mL (4 mL per tube) were conventionally shaking cultured together for 4 hours at multiplicity of infection (MOI) of 10.000, 1.000, 0.100, 0.010, or 0.001, respectively (1 tube per MOI). The titer of phage KP168 was measured by the double-agar plate method (the measurement method was the same below) to select the optimal MOI. The experiment was repeated three times. (5) The host bacteria solution at a concentration of 1×10⁹ CFU/mL (4 mL per tube) and the phage KP168 solution at an adjusted titer of 5×10⁷ PFU/mL (400 μL per tube) were mixed at the MOI of 0.005. The plaques were counted 0 (immediately), 1, 2, 3, 4, 5, 15, and 30 minutes (1 tube at each time point) after mixing by the double-agar plate method (the counting method was the same below), and the percentage of adsorbed phages was calculated to screen for the optimal adsorption time. The experiment was repeated three times. (6) The host bacteria solution at a concentration of 1×10⁹ CFU/mL (300 μL per tube) and the phage KP168 solution at a titer of 5×10⁸ PFU/mL (60 μL per tube) were mixed at MOI of 0.005 and conventionally shaking cultured after standing for the optimal adsorption time. The phage KP168 titer was measured 0 (immediately), 10, 20, 30, 40, 50, 60, 70, 80, 90, and 100 minutes after culture, and a one-step growth curve was drawn. The experiment was repeated three times. (7) The phage KP168 solution at a titer of 2.5×10¹⁰ PFU/mL was left to stand for 1 hour at 37, 40, 50, 60, or 70 ℃ (3 tubes at each time point, 1 mL per tube) for counting the plaques, and then the thermal stability curve was drawn. SM buffer at a pH values of 5.0, 6.0, 7.0, 7.4, 8.0, 9.0, or 10.0 were added to the phage KP168 solution at a titer of 3.0×10¹⁰ PFU/mL, respectively. The mixed solution was left to stand for 1 hour at 37 ℃ (3 tubes of each pH, each tube containing 100 μL phage KP168 solution and 900 μL SM buffer), and then the plaques were counted, and an acid-base stability curve was drawn. (8) The phage KP168 solution was taken for DNA extraction and sequencing after dialysis as in experiment (2). The whole genome was annotated with Prokka to obtain the coding sequence of phage KP168. Nucleotide′s BLAST function was used to proceed nucleic acid sequence alignment for finding a known phage with the highest similarity to the phage KP168 nucleic acid sequence, and Blastx function was used to translate the coding sequence into protein for its function prediction. The comparison with Antibiotic Resistance Genes Database and Virulence Factors Database was proceeded. (9) In a 96-well plate, at a MOI of 1.000, 0.100, 0.010 or 0.001 (3 wells per MOI), 20 μL phage KP168 solution at a initial titer of 5.8×10¹⁰ PFU/mL was added to 200 μL host bacteria solution at a concentration of 1.5×10⁸ CFU/mL (the same concentration below) for co-cultivation for 48 hours. After 200 μL host bacteria solution was left to stand for 48 hours, 20 μL phage KP168 solution at a titer of 1×10^6, 1×10^7, 1×10^8, 1×10^9, or 1×10¹⁰ PFU/mL (3 wells per titer) was added respectively for action for 4 hours. In both experiments, 200 μL host bacteria solution added with 20 μL SM buffer (3 wells) acted as a negative control, and 220 μL LB culture medium (3 wells) acted as a blank control. Absorbance values were measured by a microplate reader, and inhibition/destruction rates of biofilm were calculated. The experiments were both repeated three times. Results: (1) The plaques of phage KP168 successfully isolated and purified were transparent and round, and its diameter was approximately 1.5 mm. (2) The phage KP168 has a regular polyhedron structure with a diameter of about 50 nm and without a tail. (3) The phage KP168 could lyse 13 of 20 strains of Klebsiella pneumoniae from burned patients, with a lysis rate of 65.0%. (4) When MOI was 1.000, the titer was the highest after co-culturing the phage KP168 with the host bacteria for 4 hours, which was the optimal MOI. (5) After the mixing of the phage KP168 with the host bacteria for 4 minutes, the percentage of the adsorbed phage reached the highest, which was the optimal adsorption time. (6) The one-step growth curve showed that during the lysis of the host bacteria by phage KP168, the incubation period was about 10 minutes, and the lysis period was about 40 minutes. (7) With the condition of 40 ℃ or pH 7.4, the number of plaques and the activity of phage KP168 reached the highest. (8) The genome of phage KP168 was a linear double-stranded DNA with a length of 40 114 bp. There were 48 possible coding sequences. It had the highest similarity to Klebsiella phage_vB_Kp1. The most similar known proteins corresponding to the translated proteins of coding sequences contained 23 hypothetical proteins and 25 proteins with known functions. No resistance genes or virulence factor genes were found. The GeneBank accession number was KT367885. (9) After 48 hours of co-cultivation of the phage KP168 and the host bacteria at each MOI, the inhibition rates of biofilm were similar, with an average of about 45%. After the phage KP168 with a titer of 1×10⁹ PFU/mL acted on the biofilm formed by the host bacteria for 4 h, the destruction rate of biofilm was the highest, reaching an average of 42%. Conclusions In this study, a bacteriophage against pan-drug resistant Klebsiella pneumoniae from a burn patient, phage KP168, is isolated from sewage, which belongs to the tailless phage. It has a wide host spectrum, short adsorption time, and short incubation period, with certain thermal and acid-base stability. Its genomic information is clear, and it does not contain resistance genes or virulence factor genes. It also has an inhibitory effect on the formation of bacterial biofilm and a destructive effect on the formed bacterial biofilm.