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Objective To explore the role and efficacy of VEGF and HGF gene adenovirus vector in promoting angiogenesis in ischemic tissue.Methods 84 Kunming mice were randomly divided into sham group,control group,VEGF group,HGF group and VEGF+HGF group,and the left lower limb ischemia model was established.The blood supply of ischemic tissue was observed by rheometer,and the expression levels of VEGF and HGF in each group were detected by Western Blot and ELISA.Immunohistochemical staining was used to detect angiogenesis(CD31,SMA)in ischemic tissues.Safety was assessed by side effects during treatment in mice.Results After the successful modeling,the blood flow velocity of the left lower limb in each group decreased significantly.On the 7th day after operation,the blood flow of the left lower limb in each group was significantly better than that on the 0th day after operation(P<0.05),and the blood flow of the left lower limb in Ad-VEGF-HGF group was significantly better than that in other groups(P<0.05).On the 28th day after operation,the blood flow of the left lower limb in Ad-VEGF-HGF group gradually stabilized,the blood flow in Ad-VEGF-HGF group was significantly better than that in other groups,and both VEGF group and HGF group were significantly better than the control group(P<0.05).On the 7th,14th,and 28th days following surgery,HGF and VEGF protein levels in the Ad-HGF,Ad-VEGF,and Ad-VEGF-HGF groups were substantially greater than those in the control group(P<0.05).The expression level in the Ad-VEGF-HGF group peaked on the 14th day(all P<0.001)and subsequently declined to preoperative levels on the 28th day after operation.Conclusion Ad-VEGF-HGF gene injection can effectively boost VEGF and HGF protein expression and rapidly reach the relative peak level,encour-aging angiogenesis after lower limb ischemia,increasing blood flow,and improving lower limb circulation.
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Objective:To construct a column-line diagram diagnostic model based on serum and joint fluid inflammatory markers for the diagnosis of periprosthetic joint infections (PJI) after joint arthroplasty and to validate its predictive ability.Methods:The clinical data of 181 patients diagnosed with PJI or aseptic loosening in the Department of Orthopedics of the First Affiliated Hospital of Chongqing Medical University from January 2015 to June 2020 were retrospectively collected as a modeling group. The best indicators for diagnosing PJI were screened by lasso regression, single-factor and multifactor analysis. By comprehensively considering the weights and intrinsic connections of the indicators, a column-line diagram diagnostic model was constructed and used to develop a clinical decision support system (CDSS). Prospectively, the clinical data of patients diagnosed with PJI or aseptic loosening in the Department of Orthopedics of the First Hospital of Chongqing Medical University from July 2020 to December 2022 were collected as a validation group, and the diagnostic performance of the column-line diagram model was externally validated by methods such as receiver operating characteristic curve (ROC).Results:There were 85 cases of PJI in the 181 cases modeling group and 23 cases of PJI in the 49 cases validation group. Among the 27 potential factors analyzed by lasso regression analysis, body mass index (BMI), blood tests including platelet (PLT), absolute lymphocyte value, interferon γ (IFN-γ), ESR, IL-6, C-reactive protein, D-dimer, and joint fluid tests including C-reactive protein, IL-1, IL-4, IL-6, percentage of multinucleated neutrophils (PMN%), and CD64 may be potential indicators for the diagnosis of PJI. Univariate found significant differences between hematologic tests including sedimentation, C-reactive protein, IL-6, D-dimer and joint fluid tests including C-reactive protein, joint fluid CD64 index, C-reactive protein, IL-1, IL-4, IL-6, PMN%( P<0.05). Further multifactorial regression analysis screened serum IL-6, D-dimer, joint fluid CD64 index, C-reactive protein, IL-1, IL-4, IL-6, and percentage of multinucleated neutrophils, and based on that, the column-line graph model and CDSS system were constructed. The area under the ROC in the validation group was 0.978, and the AUC in the internal validation was 0.995; the C-index of the calibration curve was 99.50%, and the C-index of the internal validation was 99.53%, suggesting that the column-line diagram model has a good predictive ability. Conclusions:The column-line diagram for diagnosing PJI based on multiple diagnostic indicators showed good diagnostic performance. The CDSS system constructed by column-line diagrams could assist clinicians in diagnosing PJI and making reasonable strategies in time.
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Prosthetic joint infection (PJI) is a catastrophic complication after joint replacement.It has become the primary cause of revision surgery after knee arthroplasty and the third cause of revision after hip replacement surgery.The risk factors involve the patient (intrinsic factor) and the environment (extrinsic factor),which can be further divided into uncontrollable factors such as the history of surgical infection and other factors related to age and sex of tumor,as well as the risk factors caused by patients' own characteristics and associated diseases,such as body mass index,smoking,diabetes,rheumatoid arthritis and medicine use.The authors summarize the patients' own characteristics,associated diseases and medication to comprehensively evaluate and understand how to reduce the risk of PJI in the perioperative period,so as to provide some reference for clinical treatment.
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Objective To evaluate the construction of expression vector for fusion protein of cell-penetrating pep-tide CCL (PEP-CCL).Methods CCL6-PEP-6XHis was inserted into plasmid pABP,pABP-CCL6-PEP plasmid was extracted and then transfected into HEK293 cells,CCL6-PEP-6XHis was expressed and purified by chromatog-raphy and detected with Western Blot.Results PEP-CCL express vector was successfully constructed and purified. PCR product of CCL6-PEP-6XHis Tag was ligated with T vector,recombinant was transferred into the host cells, then host cells were cultured,plasmid was extracted and sequenced,the sequence was identical to targeted gene. CCL6-PEP-6XHis was successfully inserted into the eukaryotic expression vector pABP,plasmid was extracted and digested,electrophoresis results revealed that a fragment with 430bp was digested by Hind Ⅲ+XbaⅠ,which was identical to the expected value.Western Blot revealed that CCL6-PEP fusion protein could be recognized by His monoclonal antibody.Conclusion PEP-CCL express vector can be constructed and expressed in eukaryotic cells.
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Objective To investigate the evaluative meaning of the traditional Chinese medicine (TCM)sweat detector and temperature detector to atheriosclerosis obliterans (ASO).Methods 68 limb plantar of ASO patients and 20 left plantar of healthy subjects were tested by TCM sweat detector and temperature detector to observe the differences of sweating rate and temperature between the two groups.Results ①Sweating rate and temperature of ASO patients' limb plantar showed different graphic morphology from that of healthy control group.The sweating rate of patients was worse than that of healthy subjects in responding to external stimuli.②ASO patients'limb plantar baseline values of the sweating rate and temperature were lower than that of healthy subjects,and the result was statistical significant (P＜0.01).Conclusion Data obtained from TCM sweat detector and temperature detector can be used to distinguish ASO patients from healthy people.
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Objective To investigate the in vivo transduction capability of fusion protein PEP-1-EGFP with mice.Methods Two prokaryotic expression plasmids pET15b-EGFP and pET15b-PEP-1-EGFP were constructed and transformed into E.coli BL21(DE3) to express EGFP and fusion protein PEP-1-EGFP,respectively.The expressed EGFP and PEP-1-EGFP were purified with Ni2+-resin affinity chromatography.Five hundred micrograms of EGFP and PEP-1-EGFP fusion protein were injected into mouse through caudal vein,respectively,the mice were euthanized and perfused with PBS 2 hours after administration.Then,the heart,brain,liver,spleen and kidney were removed and sectioned with a cryostat at 7 ?m for visualization with a inverted fluorescent microscope.ResultsThe brain,heart,liver,spleen and kidney injected with PEP-1-EGFP showed bright and homogenous green fluorescence whereas that with EGFP showed no green fluorescence at all.Conclusion The successful expression and purification of PEP-1-EGFP fusion protein and its efficient transduction into mice in vivo provide a basis for the research on transmembrane delivery of macromolecule drugs mediated by the cell-penetrating peptide,PEP-1.
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Objective To discuss the curative effects of replacement of artificial femoral secured with cement in old aged patients with femoral neck fractures. Methods 131 cases (101 fresh and 30 obsolete; 23 type Ⅱ, 62 type Ⅲ, 46 type IV by Garden typing) were replaced with artificial femoral head secured with cement by a Gibson approach, and were injected with medicine to avoid venous thrombus. Results All cases were followed up for 0.5～8 years. The excellent and good rate was 96.4%. Conclusions The replacement of artificial femoral head secured with cement is suitable for old aged patients, because it is convenient and the patients can recover fast after the operation.