BACKGROUND:It is presumed that urinary trypsin inhibitor could have protective effects on local and systemic tissues and could inhibit osteoclast proliferation and activation under long-term chronic inflammation conditions and in ischemic and anoxic environment which was induced by prosthetic wear. OBJECTIVE:To investigate the inhibitory effect of ulinastatin on receptor activator for nuclear factor-κb ligand-induced differentiation, proliferation and osteoclastogenesis of RAW264.7 cells and its effects on matrix metal oproteinase-2, matrix metal oproteinase-9 expression level and activity. METHODS:Mouse monocyte/macrophage cellline RAW264.7 was treated with different concentrations of urinary trypsin inhibitor (0, 500, 5 000 U/mL) for 24, 48 and 72 hours. Experiments were divided into four groups:the blank group (RAW264.7 cells), receptor activator for nuclear factor-κb ligand-induced group (0 U/mL ulinastatin), 500 U/mL ulinastatin group and 5 000 U/mL ulinastatin group. RESULTS AND CONCLUSION:(1) MTT results indicated that there was no significant difference on the proliferation of RAW264.7 cells treated with urinary trypsin inhibitor at 0-5 000 U/mL (P>0.05) (2) Tartrate-resistant acid phosphatase staining results revealed that compared with receptor activator for nuclear factor-κb ligand-induced group, the number of tartrate-resistant acid phosphatase-positive cells was significantly less in the ulinastatin group (P<0.05), showing a time-dose dependent manner. (3) Immunohistochemisical results found that compared with receptor activator for nuclear factor-κb ligand-induced group, the percentage of matrix metal oproteinase-9-positive cells was apparently lower in the ulinastatin group. (4) Western blot assay results demonstrated that matrix metal oproteinase-9 expression was low in the RAW264.7 cells alone. At 48 hours after addition of receptor activator for nuclear factor-κb ligand, matrix metal oproteinase-9 protein expression was large. At 72 hours after culture in the 5 000 U/mL ulinastatin group, matrix metal oproteinase-9 protein expression was evidently reduced. (5) Gelatin zymography results showed that compared with the receptor activator for nuclear factor-κb ligand-induced group, matrix metal oproteinase-9 expression was significantly lower in the 5 000 U/mL ulinastatin group (P<0.05). Results suggested that urinary trypsin inhibitor inhibited receptor activator for nuclear factor-κb ligand-induced osteoclastogenesis and diminished matrix metal oproteinase-9 expression and activity.

BACKGROUND:Previous studies have indicated that ulinastatin can inhibit RANKL-induced osteoclastogenesis on RAW264.7 cells and also lower matrix metal oproteinase-9 expression and activity. However, it remains be unclear whether ulinastatin has the intervention effect on polymethyl methacrylate (PMMA)-induced MC3T3-E1 mouse preosteoblast apoptosis or not. ＜br> OBJECTIVE:To explore the intervention role of ulinastatin on the PMMA-induced MC3T3-E1 mouse preosteoblast apoptosis and its effects on type I col agen, osteocalcin, matrix metal oproteinase-2 mRNA expression. ＜br> METHODS:MC3T3-E1 mouse preosteoblasts at passages 6 and 7 were divided into four groups:blank group (only cultured MC3T3-E1 mouse preosteoblast), PMMA-induced group (MC3T3-E1 mouse preosteoblast+1 g/L PMMA bone cement suspension), low dose ulinastatin group (MC3T3-E1 mouse preosteoblast+1 g/L PMMA bone cement suspension+500 U/mL ulinastatin) and high dose ulinastatin group (MC3T3-E1 mouse preosteoblast+1 g/L PMMA bone cement suspension+5 000 U/mL ulinastatin). MTT method was adopted to detect the proliferation activity of proliferative activity of MC3T3-E1 mouse preosteoblast;alizarin red staining method was used to observe mineralization nodules of MC3T3-E1 mouse preosteoblast among different groups;the change of apoptosis rate for MC3T3-E1 cells was detected by flow cytometry analysis;semi-quantitative RT-PCR was taken to analyze type I col agen, osteocalcin, matrix metal oproteinase-2 mRNA expression level in MC3T3-E1 mouse preosteoblasts among different groups. ＜br> RESULTS AND CONCLUSION:Compared with the blank group, PMMA significantly inhibited the proliferation activity of MC3T3-E1 mouse preosteoblast (P＜0.05), and however significantly promoted cells apoptosis (P＜0.05). After addition of different concentrations of ulinastatin (500, 5 000 U/mL), the proliferation activity of MC3T3-E1 mouse preosteoblasts significantly raised (P＜0.05), and cells apoptosis rate significantly decreased (P＜0.05), showing the dose and time-dependent relation. Type I col agen and osteocalcin mRNA expression levels both significantly decreased after co-culture in PMMA group compared with the blank group (P＜0.05), matrix metal oproteinase-2 mRNA expression level, however, significantly increased (P＜0.05). After intervention with 5000 U/mL ulinastatin, type I col agen and osteocalcin mRNA expression levels both significantly increased, while matrix metal oproteinase-2 mRNA expression level significantly decreased (P＜0.05). PMMA group showed no obvious mineralization nodules. Yet, mineralization nodules were formed in the blank group, high and low dose ulinastatin groups. These results indicate that ulinastatin could have the inhibitory effect on the PMMA-induced MC3T3-E1 mouse preosteoblast apoptosis, and it could promote type I col agen and osteocalcin mRNA expression and yet suppress matrix metal oproteinase-2 mRNA expression.

Objective To investigate the expression of microRNA (miRNA)-3620 in the plasma of patients with anti-tuberculosis drug-induced hepatotoxicity (ATDH).Methods A total of 35 patients with ATDH and 35 non-ATDH paired individuals were included in this study.Plasma miRNA-3620 levels were detected using real-time Polymerase chain reaction.Comparison between two groups was done with t test.Receiver operation characteristic (ROC) curve analysis was performed to determine the diagnostic value of miRNA-3620 in ATDH.Results The relative expression of plasma miRNA-3620 of patients with ATDH and non-ATDH were 1.65±1.43 and 0.71±0.45, respectively, with significantly statistical difference (t=3.703, P<0.01).The cut off value of miRNA-3620 expression was 1.15 and the area under ROC curve were 0.71(95% CI: 0.43-1.45).Based on this cutoff value, the sensitivity and specificity of miRNA-3620 in diagnosing ATDH were 60.0% and 82.9%, respectively;the positive predictive value was 77.8% and the negative predictive value was 67.4%.Twenty-one ATDH cases and 29 non-ATDH cases was correctly diagnosed, with the accuracy of 71.4%.Conclusion The expression of miRNA-3620 in plasma is significantly increased in ATDH patients.

Objective To evaluate the level of plasma endothelial cell protein C receptor (EPCR) and thrombomodulin (TM)in the perioperative period of artificial joint replacement.Methods 119 patients (male 32,female 87,age distribution 57.75 ±12.04) who underwent total knee replacement and total hip arthroplasty from March to June 2015 in Department of Orthopedics of Beijing Jishuitan Hospital were selected.The levels of plasma endothelial cell EPCR and TM were compared be fore and after operation for 1 and 3 days.Results ①The concentration of plasma EPCR on the first day after surgery was reduced by 40.34％ compared with preoperative,and on the third day after surgery reduced by 49.45％ compared with pre operative,and on the third day after surgery was reduced by 15.26％ compared with the first day after surgery.The differences were all statistically significant (F=5.63,P＜0.05).②The concentration of plasma TM on the first day after surgery was reduced by 12.77％ compared with preoperative,and on the third day after surgery reduced by 40.53％ compared with preoperative,and on the third day after surgery was reduced by 31.83％ compared with the first day after surgery.The differences were all statistically significant (F=7.87,P＜0.05).Conclusion The concentration of EPCR and TM were pro gressively reduced within 3 days after arthroplasty.

BACKGROUNDProteins or peptides can be directly transferred into cells when covalently linked to protein transduction domains (PTDs). TAT is one of the most widely studied PTDs. The effect of fusion protein TAT and heme oxygenase-1 (HO-1) on liver sinusoidal endothelial cells (SECs) apoptosis during cold storage is unknown. The present study aimed to determine whether fusion protein TAT-HO-1 would transduce efficiently into liver during cold storage, and, if so, to determine whether TAT-HO-1 would attenuate SECs apoptosis during preservation injury in rat.

METHODSLivers of Sprague-Dawley rats were harvested and randomly assigned to group 1 (HTK solution) and group 2 (HTK solution containing TAT-HO-1 fusion protein) according to the type of the preservation solution. The transduction efficiency of TAT-HO-1 was examined and the impairment of SECs was assessed during the period of cold storage followed by 1 hour of reperfusion.

RESULTSTAT-HO-1 can transduce efficiently into liver during cold storage. A significantly lower apoptotic index of SECs was observed in group 2, at 6, 12 and 18 hours of cold storage after 1 hour reperfusion, when compared with group 1. TAT-HO-1 reduced HA and ET levels in liver at each time point. Both Bcl-2 and Bax protein were expressed in hepatocytes and SECs at the periphery of the sinusoidal space. Moreover, higher Bcl-2 expression and lower Bax expression were observed in group 2.

CONCLUSIONSTAT-HO-1 can transduce efficiently into rat livers and shows a protective effect on SECs by attenuating apoptosis during cold ischemia/reperfusion injury. Protein transduction will be a novel therapeutic strategy to reduce the risk of preservation injury in liver transplantation.

Animals ; Apoptosis ; drug effects ; Endothelial Cells ; cytology ; drug effects ; Heme Oxygenase-1 ; genetics ; Immunohistochemistry ; In Situ Nick-End Labeling ; In Vitro Techniques ; Liver ; cytology ; Male ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Radioimmunoassay ; Rats ; Rats, Sprague-Dawley ; Recombinant Fusion Proteins ; genetics ; metabolism ; pharmacology ; bcl-2-Associated X Protein ; metabolism ; tat Gene Products, Human Immunodeficiency Virus ; genetics

Objectives To evaluate the efficacy of rituximab combined with chemotherapy in the treatment of diffuse large B-cell lymphoma (DLBCL) and the relationship of clinical prognosis with the International Prognostic Index (IPI) by the using rituximab in autologous peripheral stem cell transplantation (APBSCT) for the patients of DLBCL. Methods 21 patients with DLBCL, 11 patients of them were at IPI low risk, and 3 patients were IPI at low intermediate risk, 3 patients were at IPI high intermediate risk, 4patients IPI high risk. Rituximab combined with CHOP regimen (cyclophosphamide, adriamycin, vincfistine and prednisone) was given for 4～8 courses. 5 patients received APBSCT. The mobilizing regimen was rituximab combined with cyclophosphamide(CTX) and etoposide(VP16). The conditioning regimen were CBV(CTX combined with VP16 and carmustine). Results In 21 patients, the complete response rate was 61.9 %,with overall response rate 90.5 %. 2-year progression free survival was (69.74±10.43)%. 2-year overall survival was (84.44:1:8.35) %. The complete response rate was 92.9 % and overall response rate was 100 % in the patients IPI≤2. The overall response rate was 71.4 % in the patients with IPI≥3. The complete response rate was higher in the patients with IPI≤ 2 (P＜0.01). The amount of mononuclear cells (M NC) in harvest were 7.34 (4.6～8.53)×108/kg. The CD+34 cells in harvest were 8.82 (2.1～10.34)×1O6/kg. The mean time of neutrephil recovering to 0.5×109/L after APBSCT was +9 day. The mean time of platelet recovering to 20×109/L after APBSCT was +12 day. The major adverse reaction were infusion related response (14.3 %) and hematological toxieities. Conclusion The efficacy of rituximab combined with chemotherapy in the treatment of DLBCL is effective, The complete response rate was higher in the patients with IPI≤2 than in the patients with IPI≥3.Using rituximab in mobilizing regimen, all patients had harvested enough CD+34 cells. Rituximab given at +1day did not affect the hematopoiesis reconstruction.

OBJECTIVETo evaluate causes and clinical management of postoperative pain after total knee arthroplasty (TKA).

METHODSFrom January 2004 to June 2009, 41 patients (44 knees) with knee pain post TKA were treated. There were 9 male and 32 female patients aging from 51 to 84 years with a mean of 63.5 years. The diagnosis followed to Brown diagnostic system. One case of extraarticular pain was complex regional pain syndrome type 1 (CRPS-1) and underwent conservative treatment, the remaining 5 cases were treated by surgery. Three cases of joint instability, 1 case of patellar baja, 2 cases of soft tissue impingement caused by overhang of the prosthesis, 1 case of popliteal tendon impact underwent conservative treatment, the other 27 cases underwent surgical intervention. The patients were followed up and the Knee Society Score (KSS) knee score, pain visual analog scale (VAS) score were recorded.

RESULTSForty-one cases were followed up for 1 to 6 years. At the last time of follow-up, the 5 cases received surgical treatment to extra-articular pain showed VAS score as 2.5 ± 0.2, KSS clinical and functional score as 92.8 ± 2.6 and 89.0 ± 3.4. There was significantly difference compared with preoperative (P < 0.05). One case of CRPS-1 performed conservative treatment, the therapy was effective. In the infected 12 cases of intra-articular pain, 1 case received amputation, 3 cases received antibiotic bone cement insert, 8 cases received two stage revision. All infections were cured, and VAS score was 3.8 ± 0.2, KSS clinical score was 88.3 ± 4.6, function score was 85.0 ± 4.6 postoperatively, with significantly difference compared with preoperative (P < 0.05). In the 8 cases received conservative treatment in non-infected group, at the last time of follow-up, VAS score was 4.5 ± 0.4, KSS clinical and functional score was 85.4 ± 4.2 and 84.2 ± 2.3, with significantly difference compared with preoperative (P < 0.05). Fifteen cases underwent surgical treatment, at the last time of follow-up, VAS score was 3.4 ± 0.1, KSS clinical and functional score was 86.6 ± 5.4 and 87.1 ± 2.4, with significantly difference compared with preoperative (P < 0.05).

CONCLUSIONSPatients with knee pain post TKA need systematic assessment to identify the causes. Appropriate treatment due to the positive diagnosis generally lead to satisfactory results, surgical intervention with indefinite causes is strictly prohibited.

Aged ; Aged, 80 and over ; Arthroplasty, Replacement, Knee ; adverse effects ; Female ; Follow-Up Studies ; Humans ; Male ; Middle Aged ; Pain, Postoperative ; etiology ; therapy ; Retrospective Studies

OBJECTIVETo investigate serum level and gene polymorphisms of matrix metalloproteinase 9 (MMP-9), and platelet glycoprotein VI (GPVI) in patients with acute coronary syndrome (ACS).

METHODSIn a prospective study of 179 patients with documented ACS and 164 controls, we measured baseline serum MMP-9 levels using ELISA and determined the MMP-9/C-1562T and MMP-9/G5564A genotypes using PCR-restriction fragment length polymorphism. Fib serum level was measured by Clauss assay. We also analyzed the Fib/Bbeta-148C/T and GPVI/T13254C polymorphisms.

RESULTSSerum levels of MMP-9 and Fib in ACS patients were significantly higher than in controls (P < 0.001), and serum level of Fib in the acute myocardial infarction group was higher than in patients with unstable angina (P < 0.05). No significant difference between ACS patients and controls was found in frequencies of MMP-9/C-1562T, MMP-9/G5564A, Fib/Bbeta-148C/T, and GPVI/T13254C genotypes and alleles (P > 0.05). The T allele of the Fib/Bbeta-148T polymorphism was associated with increased plasma Fib level (P < 0.05). There was a strong positive correlation between serum level of MMP-9 and Fib (r = 0.289, P < 0.01).

CONCLUSIONSerum levels of MMP-9 and Fib were independent risk factors of ACS. There was an obvious relationship between the Bbeta-148C/T mutation and high Fib level. No significant difference between controls and ACS patients was found in the frequencies of MMP-9 C-1562T and G5564A, Fib Bbeta-148C/T and GPVI T13254C genotypes and alleles (P > 0.05).

Acute Coronary Syndrome ; genetics ; Adult ; Aged ; Case-Control Studies ; Female ; Humans ; Male ; Matrix Metalloproteinase 9 ; blood ; genetics ; Middle Aged ; Platelet Membrane Glycoproteins ; genetics ; Polymorphism, Single Nucleotide

OBJECTIVETo prove if it is possible for using the shattering extraction with solvent to extract ingredients of traditional Chinese medicine.

METHODThe shattering extraction with solvent, the refluxing extraction and the ultrasonic extraction were used to extract paeoniflorin from Radix Paeoniae rubra, and to extract baicalein from Radix Scutellariae, and to extract chlorogenic acid from Flos lonicerae japonicae respectively, using ingredient content and extract yield as the measuring indexes.

RESULTThe content of each every ingredient obviously higher by using shattering extraction with solvent than using refluxing extraction or the ultrasonic extraction.

CONCLUSIONThe shattering extraction with solvent is a high efficiency, simple and quick extraction. It may be used to extract the ingredient of three kinds of traditional Chinese medicine.

Benzoates ; isolation & purification ; Bridged-Ring Compounds ; isolation & purification ; Chemical Fractionation ; methods ; Chlorogenic Acid ; isolation & purification ; Drugs, Chinese Herbal ; chemistry ; Flavanones ; isolation & purification ; Glucosides ; isolation & purification ; Monoterpenes ; Solvents ; chemistry ; Time Factors