An improved assay for quantitation of murine macrophage-mediated antibody-dependent cellular cytotoxicity (MMADCC) by hemoglobin-enzyme release assay (Hb-ERA) has been developed The method is based on the coloration measurement by spectrophotometer,because hemoglobin has peroxidase activity capable of catalyzing the oxidation of o-phenylenediamine and producing color reaction. This method was applied to demonstrate variation of MMADCC activity with varying effector:target ratio,incubation time,antibody concentration and macrophage in different stage of activation. The method provides advantages of(l)elimination of the need for expensive and hazardous radioactive materials,(2)relative ease and rapidity, (3)sensitiyity and reproducibility.

Total knee arthroplasty (TKA) identified as an effective treatment for ultimate knee joint disease can effectively relieve pain, correct deformity, improve knee function and enhance the quality of life of patients. Patient satisfaction has been increasingly considered as an important factor in evaluating the success of primary TKA. Anterior knee pain that usually appears in the region of the anterior knee is a recognized complaint for primary TKA and has a strong impact on the improvement of knee function and patient satisfaction of primary TKA. Accordingly, the relief of anterior knee pain has become one of the primary goals of primary TKA. At present, soft tissue lesions around the patellar caused by patellar maltracking and the elevation of internal pressure in subchondral bone because of the high contact stress of patellofemoral joint are both considered as the mechanism of anterior knee pain. For the past few years,on increasing number of studies have focused on the prevention of anterior knee pain following primary TKA. However, none of the past treatment such as patellar resurfacing, patellar denervation without patellar resurfacing or a mobile-bearing prosthesis has a good and affirmative effect on it. The prevention and treatment of anterior knee pain following primary TKA still is a difficult solved problem. To address this problem, we need further researches about the cause of anterior knee pain, knee joint prosthesis and biomechanics of patellofemoral joint, as well as lots of randomized controlled trials.
Arthralgia ; etiology ; prevention & control ; Arthroplasty, Replacement, Knee ; adverse effects ; Humans ; Knee Joint ; surgery ; Randomized Controlled Trials as Topic

Objective To explore the clinical efficacy of 1251 stent implantation in treatment of malignant tracheal stenosis with the help of ventilation catheter under local anesthesia.Methods Totally 180 consecutive patients with malignant tracheal stenosis underwent 125I airway stent implantation were analyzed retrospectively.The anhelation grade,oxygen saturation and respiratory rate before and after operation were recorded and analyzed,125I stent situation,clinical symptoms and survival situation were followed up after operation.Results The success rate of 125I stent placement was 100％ (180/180),and the stent type was tubular stent in 132 cases,Y-shaped stent in 34 cases,and L-shaped stent in 14 cases.Dyspnea was significantly relived in all patients.Oxygen saturation and respiratory rate improved from (80.60±3.87)％ and (29.36± 3.20)times/min before operation to (94.31±3.40)％ and (19.29±2.19)times/min after operation (t=-30.52,35.09,both P＜0.01).Patients were followed up 3-13 mouth after operation,and stent restenosis were occurred in 6 patients.The lifetime was 49-401 days and the average lifetime was (182±94) days.The 60-day survival rate was 0.99 and the 180-day survival rate was 0.65.Conclusion 125I stent implantation in treatment of malignant tracheal stenosis with the help of ventilation catheter under local anesthesia is a safe and effective method.

HIV-1 envelope glycoprotein gp41,which is a hopeful target for HIV-1 fusion inhibitors,plays a critical role in the fusion of viral and cellular membranes.In order to build up the screening assay of HIV-1 fusion inhibitors targeting gp41,HIV-1 gp41 5-helix and 6-helix were expressed in prokaryotic cells.Gp41 5-helix and 6-helix recombined plasmids were constructed by using PCR,enzyme digestion and ligation taking the clade B HIV-1 genome as a template.The plasmid was transferred into E.coli BL21(DE3)and then induced by IPTG.The expressed protein was purified by affinity chromatography after denaturation and renaturation.The SDS-PAGE analysis was used during expression and purification.Native-PAGE was used to identify the interaction between gp41 5-helix and T-20.The result will be helpful to build up the screening assay of HIV-1 fusion inhibitors targeting gp41.

Highly Active Anti-Retroviral Therapy (HAART) has effectively inhibited the prevalence of HIV-1 and reduced the death rate caused by AIDS. In recent years,the emergence of resistance-conferring RT gene mutations in HIV-1 strains has become the major reason for HAART failure. The detection of drug resistance is important for the HAART regimen choice and novel drug development. A novel assay for the detection of HIV-1 RT drug resistance mutations was developed. HIV-1 drug resistance and wild strains in B subtypes were investigated using Two-Step Mutagenically-Separated PCR (MS-PCR),and point mutations including M41L,K70R,K103N,Y181C,T215F were detected. A longer mutant type primer was designed,using microplates hybridization and ELISA technique to detect several point mutations within a mixed mutant-wild type population. The results indicate that the Two-Step MS-PCR is as sensitive and specific as that in the traditional MS-PCR and MS-PCR combined with ELISA can give a good P/N quotient with better sensitivity,low cost,relatively less time consumption and high-throughput screening. It will be used in clinic usage for the detection of HIV-1 drug resistance mutations as well as other point mutations.

Objective:To investigate the clinical features of direct and indirect carotid cavernous fistulas (CCFs).Methods:Patients with CCF treated in the Affiliated Hospital of Xuzhou Medical University from January 2010 to August 2020 were enrolled retrospectively. Relevant clinical data were collected, including the main clinical manifestations, neuroimaging features, and treatment methods. The clinical features of direct and indirect CCFs were compared.Results:A total of 31 patients were enrolled in the study, 29 (93.5%) had ocular symptoms, of which conjunctival hyperemia and edema ( n=24, 77.4%), exophthalmos ( n=19, 61.3%) and orbital murmur ( n=18, 58.1%) were most common. There were 23 patients (74.2%) in direct CCF group and 8 (25.8%) in indirect CCF group. The former had more history of head trauma (78.2% vs. 12.5%; P=0.002), more flow volume (high-flow CCFs: 100% vs. 37.5%; P<0.001) and more likely to cause orbital murmur (69.6% vs. 25.0%; P=0.043). Endovascular embolization was safe and effective. The common methods of endovascular embolization were EVAL glue combined with coil embolization ( n=18, 66.7%) and detachable balloon embolization alone ( n=6, 22.2%). Conclusion:Ocular manifestations are most prominent in patients with CCFs. Direct CCF is more common, usually with a history of head trauma, and the clinical and imaging features are more typical. Interventional embolization is the preferred treatment option for patients with CCF.

To explore the plasticity of human amniotic mesenchymal cells(hAMCs) into cardiomyocyte-like cells,hAMCs were isolated from human amnion with collagenase digestion.Phenotype of the isolated cells was analyzed by flow cytometry(FCM).hAMCs were treated with 5-azacytidine and basic fibroblast growth factor(bFGF) to investigate their ability of differentiation into cardiomyocytes.The induced differentiated cells were evaluated by immunofluorescence for desmin and ?-actin expression and by RT-PCR for Nkx2.5,GATA-4 and alpha-myosin heavy chain(?-MHC) mRNA expression.The results showed that,after primary culture,hAMCs could reach a confluence of 80% with swirl like growth at 6 days.The cells proliferated rapidly after passages with a 100% confluence at 3~4 d.hAMCs were positive expression of CD44 and vimentin,but negative for CK19.After induced differentiation at 8~10d,the differentiated cells have close-up arranged with long spindle-shape.At 2 weeks and 4 weeks,induced cells expressed ?-actinin and cardiac-specific transcription factor Nkx2.5.Expressions of GATA-4 and desmin can be detected but ?-MHC can not in the hAMCs both before and after the induction.In conclusion,hAMCs have the ability of differentiation into cardiomyocyte-like cells,which means that hAMCs can be regarded as candidate cells for cellular cardiomyoplasty(CCM).

The technique conditions of decolonization of fermentation broth were successively optimized using single factor assay and orthogonal layout method in Cordyceps jiangxiensis. The optimal condition of decolorization was investigated to be 3g/100mL active carbon, 5 min absorption time, pH5 of fermented broth and 25℃absorption temperature. Under the optimal condition, the maximum decolorization rate of fermented broth reached 89. 6% , simultaneously 10. 7% consuming rate of exopolysaccahride was minimum. Subsequently, the extract condition of exopolysaccharide of C. jiangxiensis was further optimized by orthogonal layout design. The maximum exopolysaccharide production was 0. 38 g/L under the optimal condition, i. e. firstly fermented filtrate decolorized and deproteined was concentrated to 1/7 of its total volume, secondly concentration broth was mixed with four times its volume of absolute ethanol and stirred vigorously, lastly precipitation of exopolysaccharide proceeded at 4℃for 16 hrs and the exopolysaccharide collected by centrifugal ion and dryness.

Objective To discuss whether preoperative extracorporeal shock wave lithotripsy (ESWL) could improve the efficacy of percutaneous nephrolithotomy (PCNL) for complicated renal calculi. Methods 160 cases of complicated renal calculi patients were divided into observation group (80) and control group (80) at random. Take conventional PCNL treatment for control group, and ESWL treatment one day before conventional PCNL treatment for observation group. Record the operation time, intraoperative blood loss, postoperative calculi clearance rate, complications, treatment costs, hospital stays of two groups of patients, then compare the curative effects between the two groups. Results Results are very different in the two groups, and the therapeutic effect of observation group is much better than control group (P < 0.05). Conclusions For complicated renal calculi, compared with conventional PCNL, in reducing the residual stone rate shorting the operation time, reducing the operation number, complications, treatment costs and the length of hospital stay, PCNL preoperative ESWL have obvious advantages. It is a kind of both economic and safe and effective treatment method.

The pol gene of HIV-1 encodes mainly three enzymes: reverse transcriptase (RT), protease (PR) and integrase (IN). Currently, FDA approved drugs targeting RT and PR are available and administered in various combinations, while no anti-IN drug was approved. HIV-1 integrase is an essential enzyme for the viral replication and an interesting target for the design of new pharmaceuticals for multi-drug therapy of AIDS. The 288 amino acids of IN (32kDa) recognizes specific sequences in the long terminal repeats (LTRs) of the retroviral DNA. The IN protein catalyzes the 3′-processing step and the 5′-strand transfer step reaction in vivo, which was called integration and this reaction could be analysed by ELISA Assay in vitro. It has been reported that F185K and C280S mutations of HIV-1 integrase would improve the enzyme solubility, and the catalytical activity of the enzyme was the same as that of the wild-type enzyme in vitro. In order to build the platform of screening inhibitor against integrase of HIV-1 virus, the IN enzyme was expressed and the function of integrase protein was assayed. The cDNA of clade B HIV-1 genome was used as a template, overlapped PCR was used to construct site mutagenesis of F185K/C280S and NdeI/Xho I restriction sites were brought in. The PCR product was cloned into the prokaryotic vector pET-28a(+) to form a recombined plasmid, transferred into the host cell E.coli(BL21 DE3). The recombined clones were identified by PCR and Nde I/Xho I digestion .The positive plasmid was sequenced, and the successfully recombined plasmid in the host cell was induced by IPTG. The expressed IN protein was puriied sy the Co+ affinity chromatography column and SDS-PAGE was used to analyze the molecular weight and specificity. In addition, ELISA assay was used to analyze the function of the recombined IN protein. The recombinant protein was soluble, and expressed highly and stably in E.coli. The molecular weight of the expression product was identical to the expectation. The IN protein was proved to be functional in 3′ processing and 5′strand transfer by ELISA. It will be helpful to build the platform of screening inhibitors against HIV-1 integrase.