The clinical research on using the anterior half of the peroneus longus tendon as an autograft source / 中华骨科杂志

Objective To evaluate availability and outcomes of using anterior half of the peroneus longus tendon (AHPLT) in knee ligament reconstruction as an autograft source.Methods From July 2007 to January 2008,100 patients with knee ligament injuries were recruited in this study.There were 33 males and 67 females aging from 16 to 62 years (mean,32.3 years).49 cases had undergone medial patellofemoral ligament reconstruction,19 cases multiligament reconstruction,18 cases double-bundle posterior cruciate ligament (PCL) reconstruction and 14 cases double-bundle anterior cruciate ligament (ACL) reconstruction.AHPLT was used as sole (49 cases) or part (51 cases) of reconstruction materials.One-incision or two-incision striping techniques were adopted to harvest AHPLT.Ligaments were fixed with screws.Post-operative assessments included Kujala knee score,Lysholm knee score,Marx knee score,International Knee Documentation Committee (IKDC) subjective evaluation form and objective evaluation grade,the Foot and Ankle Disability Index (FADI) and the American Orthopedic Foot and Ankle Society (AOFAS) scale.Results 92 cases were followed up for more than 24 months.Postoperative Kujala score,IKDC subjective score,Lysholm score and Marx score were improved significantly in all four groups of patients.According to IKDC objective evaluation grade,the number of patients reaching Grade A (normal) or Grade B (near-normal) in multiligament,PCL and ACL reconstruction were 17,15 and 12,with an excellent rate of 89.5％ (17/19),93.7％ (15/16) and 100％ (12/12),respectively.Preoperative and postoperative AOFAS scores were 97.4±2.0 and 97.2±1.6,respectively,while the FADI scores preoperatively and postoperatively were 96.8±2.2 and 96.9±2.5,respectively.These results had no statistical significance.No signs of peroneal nerve injury or peroneus longus tendon rupture was found.Conclusion It is acceptable to use AHPLT as an autograft due to its feasibility to harvest,good clinical outcome,and low rate of donor site morbidity at a minimum of two-year follow-up.

Expression of human retinol-binding protein 4 in insect baculovirus system and preparation of its polyclonal antibody / 生物工程学报

To prepare recombinant human retinol binding protein 4 (RBP4) by using the baculovirus expression system and to detect its immunogenicity, the fusion DNA fragment of secretory signal peptide SS64 and human RBP4 gene was subcloned into a baculovirus transfer vector pFastBac-dual(pFBd), and the corresponding recombinant transfer plasmid was transformed into E. coli strain DH10bac, after transposition recombinant shuttle bacmid was screened out. The logarithmic phase Sf9 cells were transfected with the recombinant bacmid and then the recombinant baculovirus containing hRBP4 expression box were generated. After amplification of recombinant baculovirus, the recombinant baculovirus seeds were obtained. To express human RBP4, logarithmic phase Sf9 cells were infected with the virus seeds and SDS-PAGE and Western blotting were used to detect and identify the expression. Finally, to prepare a batch of RBP4 protein, logarithmic phase Sf9 cells in suspension culture were infected with recombinant baculovirus seeds and the supernatant was harvested after 120 hours post-infection for purification. Finally for preparation of polyclonal antibody and evaluation of immunogenicity, the recombinant hRBP4 from insect cells and from E. coli were immunized rabbits. Restriction enzyme digestion and sequencing confirmed that the recombinant baculovirus transfer plasmid was constructed correctly, and subsequently recombinant RBP4-bacmid was generated successfully. SDS-PAGE and Western blotting analysis suggested that human RBP4 protein was highly expressed in Sf9 cells with the molecular weight of approximately 23 kDa. The recombinant RBP4 protein could be secreted into the medium efficiently, and the expression level was calculated amount of 100 mg/L. Finally the rabbit antiserum was harvested after recombinant RBP4 immunization, therein the titer of antiserum against baculovirus recombinant RBP4 is 1:100 000 whereas the titer of antiserum against E. coli recombinant RBP4 is only 1:10 000. Overall, human RBP4 was high efficiently expressed successfully with good antigenicity in baculovirus system, and high affinity antiserum was obtained. A solid foundation was laid for the next step of the preparation of human serum RBP4 detection kit.

Effects of IL-12 coexpression level on antigen expression and immune responses induced by HBsAg DNA vaccination / 中华微生物学和免疫学杂志

Objective To investigate the effects of IL-12 coexpression level on antigen expression and immune responses induced by HBsAg DNA vaccination. Methods DNA vaccine plasmid pHBV carrying codon-optimized preS2-S gene of reference sequence CHN-HBV07-C in China was constructed. Three DNA vaccine plasmids pHBV-12i, pHBV-12l and pHBV-12h were also constructed by subcloning three different IL-12 expression cassettes with various expression strengths to plasmid pHBV, respectively. Expression levels of IL-12 and HBsAg in vaccine plasmid-tranfected 293T cells were measured by quantitative ELISA. DNA vaccines were administered intramuscularly to BALB/c mice and HBsAg-specific cellular immune responses were determined by IFN-γ ELISPOT. HBsAg-specific antibodies were tested by Chemiluminescence Quantitative Immunoassay. Results The HBsAg expression level in 293T cells was 70 ng/ml when transfected by plasmid pHBV without IL-12 expression cassette, and the HBsAg level was 18 ng/ml when transfected by plasmid pHBV-12l carrying low-level IL-12 expression cassette, whereas the HBsAg level was only 6 ng/ml when transfected by plasmid pHBV-12h carrying high-level IL-12 expression cassette.Results of DNA vaccination revealed that HBsAg-specific humoral and cellular immune responses were significantly decreased in mice administering vaccine pHBV-12h carrying high-level IL-12 expression cassette. Although HBsAg-specific antibody responses in mice inoculated with pHBV-12l were also decreased when compared with those in pHBV-vaccinated mice without IL-12 expression, the HBsAg-specific cellular immune responses were significantly increased. Conclusion High-level coexpression of IL-12 may suppress the expression of HBsAg, Whereas modest coexpression of IL-12 significantly enhanced the HBsAg-specific T cell responses induced by DNA vaccination. Therefore, it is so important to balance the expression between adjuvant and antigen to enhance the immune response.

Stimulating factors in cartilage tissue engineering / 中国组织工程研究

Stimulating factors have been employed to induce,accelerate,and/or enhance cartilage formation.For instance,stimulating factors and other additives may be added to culture media in vitro or incorporated into scaffolds for in vivo delivery to control cellular differentiation and tissue formation.At present,many growth factors and other dissoluble factors such as hyaluronic acid,chondroitin sulfate,and insulin have been used alone or synergistically in cartilage tissue engineering.The application efficacy depends on cell type and culture condition.In addition,gene therapy has emerged as another method of local delivery,where cells can be engineered to over-express bioactive molecules.An additional approach is the introduction of mechanical signals through loading regimes such as hydrostatic or dynamic compression or through the use of bioreactors.Current bioreactors used in cartilage tissue engineering include flat-sheet bioreactor,rotating wall vessel bioreactor and concentric cylinder bioreactor.Bioreactor can improve nutrition transmittability,provide hydrodynamic environment,induce shear stress,and promote production of cartilage specific matrix protein.

Characteristics of tissue engineered cartilage materials / 中国组织工程研究

Scaffolds provide a three-dimensional environment that is desirable for inducing and promoting the production of cartilaginous tissue.Ideally the scaffold should:① have directed and controlled degradation;② promote cell viability,differentiation,and extracellular matrix production;③ adhere and integrate with the surrounding native cartilage;④ span and assume the size of the defect;⑤ provide mechanical integrity depending on the defect location;⑥ have no toxicity,no immunogenicity or inflammatory induction;⑦ allow for the diffusion of nutrients and waste products.To date,a wide range of natural and synthetic materials have been investigated as a scaffold for cartilage repair.Based on the morphology and structure,these materials can be divided into hydrogel,sponge,fiber mesh and so on.Natural polymers that have been explored as bioactive scaffolds for cartilage engineering include:alginate,agarose,fibrin,hyaluronic acid,collagen,gelatin,chitosan,chondroitin sulfate,and cellulose.Synthetic polymers currently explored for cartilage repair include poly(?-hydroxy esters),polyethylene glycol,poly(NiPAAm),poly(propylene fumarates),and polyurethanes.

Clinical evaluation of intraoperative block of abdomial aorta in sacral and pelvic tumors surgical operation / 中国癌症杂志

Purpose:To evaluate intraoperative block of abdomial aorta(BAA) in surgery of sacral and pelvic tumors as a useful adjuvant technique.Methods:Of 164 cases of pelvic tumor who underwent resections and hemisections, the procedures in 94 cases were non block of abdominal aorta(NBAA);in 109 cases of sacral tumors various extents of acrectomy was done,53 cases had NBAA,56 cases had BAA.Results:In the pelvic tumor group, the size of tumors for BAA were on the average bigger than NBAA by 0.8 cm, the operation was shortened by 2 h, the dose of hemorrhage decreased by 2 200 ml. The complications of operation were reduced by 6.3%, death rate reduced by 4.6%. In the sacral tumor group, the size of tumors for BAA were on the average bigger than NBAA by 0.7 cm, the operation was shortened by 1 h and 40 min, the dose of hemorrhage decrease by 1 600 ml. The complications of operations were reduced by 23%, death rate reduced by 7.5%.Conclusions:The adoption of BAA adjuvant technique can reduce the intraoperative hemorrhage,the death rate, and the complications of operation, while also shortening the surgical operating time, and is a valuable clinical technique. [

The enhancement effect of caffeine in cisplatin-induced apoptosis of osteosarcoma cells / 解放军医学杂志

Objective To explore the enhancement effect of caffeine in cisplatin-induced apoptosis of osteosarcoma cell line OS-U2. Methods The osteosarcoma cells were incubated with different concentrations of cisplatin (0.2, 2, 5, 10 and 20?g/L), caffeine (0.2, 2.0mmol/L) and caffeine + cisplatin for 24, 48 and 72 hours. The proliferation of OS-U2 cells was determined by MTT assay, the apoptotic levels were determined by flow cytometry (FCM), and the morphologic changes of apoptotic cells and the positive rates of apoptosis were determined by fluorescence microscopy. Results The proliferation of OS-U2 cells was inhibited, and the apoptotic level was increased when incubated with caffeine and/or cisplatin. There were dose- and time-effect relationships between the lethal effect of cisplatin on osteosarcoma cells and caffeine. Conclusion There is a remarkable enhancement effect of caffeine on cisplatin-induced apoptosis of osteosarcoma cell line OS-U2. It seems that the apoptosis-inducing activity of caffeine may enhance the lethal effect of cisplatin on osteosarcoma cells.