Objective To study the apoptosis of human colorectal cancer HT-29 cells induced by Aralia elata Seem leaf total saponin (ETS) and its effects on the expression of relevant proteins. Methods MTT assay was used to detect the proliferation of human colorectal cancer HT-29 cells cultivated with different concentrations (6.25, 12.5, 25, 50, 100, 200 mg/kg) of ETS. Hoechst33258 staining and laser confocal imaging were used to detect the apoptotic cells. Morphological changes were observed. The expressions of Bcl-2 and Bax were detected by immuno-histochemistry. Results ETS could induce apoptosis of HT-29 cells and apoptosis was in a dose-dependent manner in a certain range. ETS could decrease the expression of Bcl-2 and increase the expression of Bax in HT-29 cells (P<0.05, P<0.01), showing a significant dose-effect relationship. Conclusion ETS can induce the apoptosis of HT-29 cells, and the mechanism may be related to reducing the expression of Bcl-2 and increasing the expression of Bax.

BACKGROUND: To apply mouse anti-human cTnI monoclonal antibody as the drug vector in the treatment and diagnosis of myocardial injury, it is important to degrade the immunity of murine antibody and overcome human anti-mouse reaction. Humanization has been applied as an attempt to resolve this problem.OBJECTIVE: To clone murine anti-cTnI Fab fragment and analyse the nucleotide and deduced amino acid sequences.DESIGN: Single sample study.SETTING: An institute of cardiovascular disease under a medical university-affiliated hospitalMATERIALS: The study was conducted in the Institute of Cardiovascular Diseases, First Affiliated Hospital of Nanjing Medical University from January 2003 to May 2004. The hybridoma cell line JS200202 which secrets the anti-cTnI monoclonal antibody was provided by Institute of Cardiovascular Disease, First Affiliated Hospital of Nanjing Medical University.METHODS: IgG heavy chain primers and κ light chain primers of amplified mouse were designed. Total RNA was extracted from hybridoma cells which secrete cTnI. Reverse transcription polymerase chain reaction(RT-PCR) was amplified. Cloning and subsequent sequence analysis of the Fab fragment was performed. The deduced amino acid sequence was compared and analysed with previously published sequences.MAIN OUTCOME MEASURES: Heavy chain Fd segment and κ light chain gene sequence and its subgroups.RESULTS: A band of approximate 700 and 800 base pairs were amplified using IgG heavy chain primers and κ light chain primers respectively. Sequence analysis indicated that the deduced amino acid sequences were in consistent with the characterization of the amino acid in the murine IgGl Fab fragment(GenBank accession NO AY484430, AY484431; Protein Bank accession NO AAR83243, AAR83244).CONCLUSION: A complete murine anti-cTnI Fab fragment was obtained in this study, which may provide basis for the production of the chimeric anti-cTnI antibody.

Objective To study the surgical treatment strategy for heamophilic arthropathy and musculoskeletal appara?tus. Methods A total of 120 heamophilic patients underwent 166 primary operations from January 1996 to June 2015 in Pe?king Union Medical College Hospital, with the average age of 29.7±12 years (from 6 to 61 years). Hemophilic type A accounted for 109 patients and hemophilic type B accounted for 11. Eighty?seven patients presented with bleeds within the joints, with 63 cases for knee involved, 29 cases for hip, 11 cases for ankle. Fifteen patients presented with intramuscular bleeds, 21 patients with heamophilic pseudotumors. Strategy of clotting factor replacement therapy was designed according to the different level of opera?tion procedure. The clinical manifestation, operative strategy, clinical outcome and complications were retrospectively recorded. Results 41 cases (34.2%) of patients underwent surgeries for more than one location. Totally, 166 procedures were performed for 120 patients. There were 103 procedures of joint arthroplasty (62.0%, 103/166), 21 procedures of pseudotumor resection (12.7%, 21/166), 15 procedures of tendon lengthening, 12 procedures of ankle arthrodesis, 13 procedures for knee flexion contracture. There were 30 cases of postoperative complications, with the rate of 18.1% (30/166). The coagulation related complication was 4.8%in this group (8/166). The surgical procedure related complication was 7.2%(12/166). All the preoperative symptoms were relieved during the follow?up. Conclusion Surgical treatment was effective for the heamophilic arthropathy and lesion of mus?culoskeletal apparatus, under the reasonable clotting factor replacement therapy. The incidence of perioperative complication in heamophilic patients is higher than that in non?hemophilic patients. Closely inspection was inevitable for perioperative treat?ment in hemophilic patients.

Objective To evaluate the relationship between the endogenous protection induced by autophagy against acute lung injury and inflammatory responses in the septic mice.Methods A total of 130 pathogen-free male C57BL/6 mice,aged 6 weeks,weighing 20-25 g,were randomly divided into 5 groups (n =26 each) using a random number table:sham operation group (group S),sepsis group (group Sep),autophagy inducer rapamycin group (group Rap),autophagy inhibitor 3-methyladenine (3-MA) group (group 3-MA),and lysosome inhibitor bafilomycin group (group Baf).Sepsis was produced by cecal ligation and puncture in chloral hydrate-anesthetized mice.Rapamycin 10 mg/kg,3-MA 15 mg/kg,and bafilomycin 1 mg/kg were injected intraperitoneally at 1 h after operation in Rap,3-MA and Baf groups,respectively.Twenty mice in each group were selected and observed for the survival at 7 days after operation,and the 7-day survival rate was calculated.Six mice in each group were selected,and arterial blood samples were obtained at 24 h after operation,the mice were then sacrificed,and lung tissues were obtained for determination of the levels of tumor necrosis factor-alpha (TNF-α),high-mobility group box 1 (HMGB1),interleukin-6 (IL-6),IL-10 and monocyte chemotactic protein-1 (MCP-1) in serum and lung tissues by enzyme-linked immunosorbent assay.The expression of autophagy-related proteins microtubule-associated protein 1 light chain 3 Ⅱ (LC3 Ⅱ),Beclin-1 and lysosomal-associated membrane protein 2 (LAMP2),and Ras-related protein 7 (Rab7) in lung tissues was detected by Western blot.Results Compared with group S,the 7-day survival rate was significantly decreased,and the levels of TNF-α,HMGB1,IL-6,IL-10 and MCP-1 in serum and lung tissues and LC3 Ⅱ,Beclin-1,LAMP2 and Rab7 in lung tissues were significantly increased in group Sep (P＜0.05).Compared with group Sep,the 7-day survival rate and levels of IL-10,LAMP2 and Rab7 in serum and lung tissues were significantly increased in group Rap and decreased in 3-MA and Baf groups,and the levels of TNF-α,HMGB1,IL-6 and MCP1 in serum and lung tissues were significantly decreased in group Rap and increased in 3-MA and Baf groups,and the levels of LC3 Ⅱ and Beclin-1 in lung tissues were significantly increased in Rap and Baf groups and decreased in group 3-MA (P＜0.05).Conclusion Autophagy-induced endogenous protection against acute lung injury is related to inhibition of inflammatory responses in the septic mice.

Objective To evaluate the midterm outcomes of autologous femoral head grafting in total hip arthroplasty for developmental dysplasia of the hip.Methods From October 2001 to December 2011,36 patients (36 hips) with developmental dysplasia of the hip were treated by total hip arthroplasty with autologous femoral head grafting,of which 34 cases were followed up for at least 30 months.There were 6 males and 28 females.16 patients were involved in the left hip and 18 in right hip.The average age at the operation was 51 years (range,28-68 years).According to Crowe classification for developmental dysplasia of the hip,12 were type Ⅱ,16 type Ⅲ,and 6 type Ⅳ.Cementless prostheses were used for all,but one patient,who received a hybrid prosthesis.Clinical outcomes were evaluated according to Harris score and complications.Components migration,periprosthetic bone changes,the polyethylene wear rate were measured radiologically.Components position and bone healing were assessed in the anteriorposterior and lateral X-ray of the hips.Results Thirty-four patients (34 hips) were followed up for an average duration of 64.1 months (range,31-153 months).The average Harris score was 35.4 (range,23-56) preoperatively,and was 89 (range,82-95) at the latest follow-up.Twenty-eight were classified as excellent,and 6 were good,and the excellent and good rate was 100％.10 cases presented with bone absorption 2-8 months postoperatively.Four patients suffered from dislocation after surgery.All the cases were treated successfully with close reduction,except one with redislocation.Close reduction was performed for this case,and no dislocation occurred again.One patient had periprosthetic fracture due to a fall and open reduction and plate fixation was performed.The fracture was healed at 1 year follow-up,with good position of the implants.Another patient had hypertensive intracerebral hemorrhage 1 year after the surgery,who was treated by conservative treatment and had hemiplegia and apraxia of the surgical hip.Conclusion Autologous femoral head grafting for the treatment of developmental dysplasia of the hip has a satisfactory clinical and radiographic outcomes at an average of five-year follow-up.Bone graft absorption is a natural process of structural bone grafting.Joint dislocation is the most common complication of THA in developmental dysplasia of the hip.

Objective To evaluate the role of autophagy in hydrogen-induced reduction of lung injury in septic mice.Methods Sixty pathogen-free healthy male ICR mice,aged 6 weeks,weighing 20-25 g,were divided into 5 groups (n =12 each) using a random number table:sham operation group (group Sh),sepsis group (group Sep),sepsis plus hydrogen group (group Sep+H2),sepsis plus autophagy inhibitor 3-methyladenine (3-MA) group (group Sep+3-MA) and sepsis plus 3-MA plus hydrogen group (group Sep+3-MA+H2).Sepsis was produced by cecal ligation and puncture.At 1 h before operation,3-MA 10 mg/kg was intraperitoneally injected.The mice inhaled 2％ H2 for 1 h starting from 1 and 6 h after operation.Blood samples were collected from the common carotid artery at 24 h after operation for measurement of arterial oxygen partial pressure,and the oxygenation index (OI) was calculated.Pulmonary specimens were obtained for examination of the pathological changes which were scored.Pulnonary mitochondria were isolated for determination of mitochondrial membrane potential (MMP) and ATP content using fluorescence spectrophotometry and a bioluminescence assay,respectively,and the respiratory control rate (RCR) was calculated.The expression of autophagy-related protein microtubule-associated protein 1 light chain 3 (LC3) was determined by Western blot,and the ratio of LC3-Ⅱ to LC3-Ⅰ expression (LC3-Ⅱ/LC3-Ⅰ ratio) was calculated.Results Compared with group Sh,the pathological scores were significantly increased,the OI and contents of mitochondrial RCR,MMP and ATP were decreased,and the LC3-Ⅱ/LC3-Ⅰ ratio was increased in Sep and Sep+H2 groups (P＜0.05).Compared with group Sep,the pathological scores were significantly decreased,the OI and contents of mitochondrial RCR,MMP and ATP were increased,and the LC3-Ⅱ/LC3-Ⅰ ratio was increased in group Sep+H2,and the pathological scores were significantly increased,the OI and contents of mitochondrial RCR,MMP and ATP were decreased,and the LC3-Ⅱ/LC3-Ⅰ ratio was decreased in group Sep+3-MA (P＜0.05),and no significant change was found in each parameter mentioned above in group Sep+3-MA+H2 (P＞0.05).Compared with group Sep+H2,the pathological scores were significantly increased,the OI and contents of mitochondrial RCR,MMP and ATP were decreased,and the LC3-Ⅱ/LC3-Ⅰ ratio was decreased in group Sep+3-MA+H2 (P＜0.05).Conclusion The mechanism by which hydrogen ameliorates lung injury is related to enhanced level of autophagy in septic mice.

Objective: To investigate the hygienic status of the central air conditioner ventilation system in public places in urban areas of Ma'anshan City, Anhui Province, so as to provide insights into formulation of supervision and management interventions. Methods: A total of 15 public places with the central air conditioner ventilation system were randomly sampled from main urban areas in Ma'anshan City in 2022. Sampling and detection were performed following the standard GB/T 18204.5—2013 Examination methods for public places Part 5: Central air conditioning ventilation system, including total number of bacteria and total number of fungus on the inner surface of wind pipes, total number of bacteria and fungus, particulate matter (PM10) and β-hemolytic streptococci in the air supply system, and Legionella pneumophila in the cooling water, and the detection indicators were assessed following the WS 394—2012 Guideline for hygiene of the central air conditioner ventilation system in public places. The eligible rate of samples, the detection rate of L. pneumophila were analyzed. Results: A total of 368 samples were collected from 15 public places, and the qualified rate was 50.54%, no places had all eligible measurement indicators. The qualified rates of total bacteria and fungus numbers on the inner surface of wind pipes were 52.67% and 59.33%. The qualified rates of total bacterium number, total fungus number, PM10 and β-hemolytic streptococci were 12.00%, 28.00%, 90.00% and 96.00% in the air supply system. The qualified rates of samples in administrative workplaces, hotels, bathing places and malls (supermarkets) were 32.50%, 59.24%, 61.09% and 68.92%, the qualified rates of total bacteria on the inner surface of air ducts were 8.33%, 72.46%, 66.67% and 61.90%, and the qualified rates of total fungus numbers in air supply were 0, 21.70%, 33.30% and 71.40%, respectively, with statistical significance (P<0.05). A total of 18 cooling water samples were collected, and L. pneumophila was detected in three samples (16.67%). Conclusions Poor hygiene is seen in the central air conditioning ventilation systems in public places in main urban areas of Ma'anshan City. High attention needs to be paid to contamination of bacterium, fungus and L. pneumophila, and expansion of supervision coverage and improved supervision intensity are recommended.

Objective To evaluate the effect of inhaling hydrogen (H2) on proteomics during acute lung injury in septic mice.Methods Sixty male ICR mice,aged 6 weeks,weighing 20-25 g,were divided into 4 groups (n=15 each) using a random number table:sham operation group (Sh group),sham operation plus H2 group (Sh+H2 group),sepsis group (S group) and sepsis plus H2 group (S+H2 group).Sepsis was produced by cecal ligation and puncture.The mice in Sh+H2 and S+H2 groups inhaled 2％ H2 for 1 h starting from 1 and 6 h after operation.At 24 h after operation,lungs were removed for identification of proteins by isobaric tags for relative and absolute quantification and liquid chromatography-tandem mass spectrometry analysis,and the differentially expressed proteins were screened.The differentially expressed proteins were used for KEGG pathway enrichment analysis and STRING protein-protein interaction networks analysis.Western blot was used to confirm the 4 differentially expressed proteins semaphorin 7A,transferrin,OTULIN and mitogen-activated protein kinase kinase kinase 1.Results A total of 4 472 quantifiable proteins were identified.A total of 192 proteins which were related to acute lung injury during H2 inhalationinduced reduction of sepsis were identified.The 192 proteins involved phosphatidylinositol 3-kinase/serine0threonine kinase signaling pathway,chemokine signaling pathway,hypoxia-inducible factor 1 signaling pathway,complement and coagulation cascades,peroxisome proliferator-activated receptor signaling pathway and proteins including ribosome proteins,myosin and troponin,collagen and adhesion-related proteins,coagulation-related proteins found in STRING protein-protein interaction networks.Conclusion Inhaling H2 can induce changes in the expression of 192 proteins,which may be the mechanism of lung protection in septic mice.

Objective To evaluate the effect of hydrogen on mitochondrial dynamics during endotoxin-induced damage to human umbilical vein endothelial cells (HUVECs).Methods HUVECs cultured in vitro were seeded in the culture plate and divided into 4 groups using a random number table:control group (group C),hydrogen-saturated culture medium group (H group),endotoxin group (group E) and endotoxin + hydrogen-saturated culture medium group (group E+H).The cells were cultured in the plain culture medium in C and E groups.The cells were cultured in the hydrogen-saturated culture medium containing lipopolysaccharide (LPS) with the final concentration of 10 μg/ml in H and E+H groups.At 2,8 and 24 h of culture or incubation with LPS,the cell viability was detected by methyl thiazolyl tetrazolium assay,the intracellular ATP content was measured using the phosphomolybdic acid colorimetric method,and the expression of dynamin-related protein 1 (DRP1) was detected by using Western blot.The expression of DRP1 was detected by immunofluorescence at 8 h of incubation with LPS.Results Compared with group C,the cell viability and ATP content were significantly decreased,and the expression of DRP1 was up-regulated at each incubation time point in E and E +H groups (P＜0.05),and no significant change was found in the parameters mentioned above in group H (P＞0.05).Compared with group E,the cell viability and ATP content were significantly increased,and the expression of DRP1 was down-regulated at each incubation time point in group E+H (P＜0.05).Conclusion The mechanism by which hydrogen reduces endotoxin-induced damage to HUVECs is related to down-regulation of DRP1 expression and inhibition of excessive mitochondrial fission.

Objective To evaluate the long-term outcomes of total knee arthroplasty (TKA) for end-stage hemophilic arthropathy.Methods Eighteen patients (24 knees) with hemophilic arthropathy underwent TKA from June 2003 to January 2009 were retrospectively reviewed.All patients were hemophilia A with an average age of 33.7± 13.0 years (18-56tyears) old at Surgery.Pharmacokinetic tests of coagulation factors were performed after consultation of hematologist.Based on the guideline of World Federation of hemophilia,the protocol of coagulation factor replacement was adjusted according to our experience and the financial status.Normally,peak level of coagulation factor concentrations were maintained at 100％ on the day of surgery,at about 80％ in the first 3 days after surgery,at 60％ on the postoperative days of 4-6 and at 40％ on postoperative days of 7-10.The dose was then gradually tapered to 20％ or 30％.Zimmer prosthesis was used in three cases (PS prosthesis in two cases,LCCK prosthesis in one case),Centerpulse prosthesis in three cases,and the rest were from Smith & Nephew (including one case of constrained prosthesis and one case of revision prosthesis).Preoperative and last follow-up Hospital for Special Surgery (HSS) score,Knee Society Score (KSS),knee flexion contracture and complications were evaluated.Results Fourteen patients(20 knees) were followed-up,with an average duration of 124±17 months (96-145 months).Knee flexion contracture improved from 16.7°±12.2° (0°-40°) preoperative to 3.3 °±5.0° (0°-10°) at the last follow-up.The average preoperative HSS score was 42.4± 16.0 (t 0-60),whereas postoperative score was 74.8± 10.6 (59-87).The preoperative KSS clinical and functional score were 36.1 ± 10.5 (20-50) and 36.1±5.5 (25-40),which were improved to 85.8±7.1 (70-93) and 80.9±22.4 (40-100) at the last follow-up,respectively.Postoperative infection,aseptic loosening of the implant and hematoma occurred in one patient respectively.All of them recovered after a revision surgery.One patient had secondary skin ulceration due to tension blisters and recovered after anterolateral thigh myocutaneous flap transplantation.Conclusion The long-term outcomes of TKA under coagulation factor substitution for hemophilic arthropathy are promising.However,the effects are inferior to those in non-hemophilic patients,and the risk of infection,aseptic loosening of the implant and hematoma are higher.