Objective To discuss the diagnosis, treatment and prognosis of stage Ⅲ osteosarcoma. Methods A retrospective analysis of 23 patients with stage Ⅲ osteosarcoma of extremities from December 1989 to December 2003 was studied. There were 9 females and 14 males, aging from 16 to 31 years with a mean of 22.4 years. 15 patients presented with lung metastases, 7 with bone metastases (including 5 of jumping metastases and 2 of osteosarcomas) and 1 with lung and bone metastases simultaneously. Patients received chemotherapy followed by resection of primary and metastatic lesions and additional chemotherapy. Results After preoperative chemotherapy, lung metastases disappeared in 1 patient, whereas in 1 with lung and bone metastases simultaneously, the lesion remained surgically unresectable because of new metastases after removal of the primary lesion. In 2 patients with osteosarcoma, primary lesion could only be removed, lung metastasis appeared in 2 of 5 patients with jumping metastases respectively after the removal of primary lesion and jumping metastases. 16 patients with lung metastases received thoracotomy and resection of the lung metastatic lesions, and 6 of them received a second thoracotomy because of a second lung metastasis inclunding 3 cases with extra-pulmonary metastases without any additional treatment. The tumor necrosis rate was not found obviously different between primary lesions and metastatic lesions. Of the 23 patients who achieved a mean 74.6 months follow-up (range, 5-168 months), 9 remained continuously free of disease, 4 relapsed with new metastases, and 10 died of tumors. The outcome of the Cox model proportional hazard regression showed the relation of the number of the metastases and the prognosis were significant(P

AIM: To study whether caspase-3,8 is activated during azurin-induced apoptosis in U2OS cells. METHODS: AnnexinV /PI method was used to detect apoptosis. The changes of procaspase-3 were analyzed by Western blot, the changes of caspase-3 mRNA were detected by semi-quantitative RT-PCR, and caspase-3 relative activity was determined by colorimetric assay. RESULTS: After U2OS cells were treated with 0, 25, 50, 100, 200, 500 mg/L azurin for 24 h, respectively, the level of procaspase-3 protein decreased and the level of caspase-3 mRNA increased as azurin concentration increased. When the cells were treated with 100 mg/L azurin for 6, 12, 24, 48 h , respectively, the caspase-3 activity began to rise from 6 h,reached the peak at 24 h,and was still higher than the control group at 48 h ( P

AIM: To down-regulate expression of c-myc through antisense therapy and to investigate its effect on the sensitivity of osteosarcoma MG-63 cells to cisplatin-induced apoptosis. METHODS: The recombinant adenovirus (Ad-Asc-myc) encoding antisense c-myc fragment was constructed and transfected into osteosarcoma MG-63 cells in vitro in order to down-regulate the expression of c-myc, and the change in the sensitivity to cisplatin-induced apoptosis was observed. MTT, Western blot, RT-PCR, flow cytometry (FCM) and electron microscope were used to evaluate tumor cell proliferation in vitro, genes expression related to apoptosis regulation and effects on the sensitivity of osteosarcoma MG-63 cells to cisplatin-induced apoptosis. RESULTS: Ad-Asc-myc down-regulated the expression of c-myc protein after transfected MG-63 cells for 48 h, combined with the treatment of 2.0 mg/L cisplatin for 2 h inhibited tumor cell proliferation in vitro by 38.0%. RT-PCR revealed that Ad-Asc-myc down-regulated the expression of Bcl-2 and up-regulated the expression of Bax. No appreciable change was observed in the expression of E_2F-1. FCM showed that Ad-Asc-myc induced apoptosis in intransfected cells, and rendered it more sensitive to cisplatin. CONCLUSION: Antisense c-myc is able per se to induce apoptosis and sensitize osteosarcoma cells to cisplatin-induced apoptosis.

AIM: To study the relationship between respiratory syncytial virus (RSV) infection and apoptosis, between RSV infection and expressions of FasL, Fas, Bcl-2 and Bax. METHODS: Apoptotic cells were examined by flow cytometry and transmission electron microscope. Immunohistochemical analysis was used to detect the expressions of apoptosis-associated gene FasL, Fas, Bcl-2 and Bax in A549 cells during RSV infection. RESULTS: Apoptotic index increased at 72 h and 120 h postinfection. Apoptotic cells were detected by transmission electron microscope. High-expressions of FasL, Fas and Bax genes and low-expression of Bcl-2 gene were detected by immunohistochemical staining. CONCLUSION: Apoptosis in A549 cells was induced by RSV infection. This apoptosis may be induced by up-regulating the expression of FasL, Fas, Bax genes and down-regulating the expression of Bcl-2 gene.

Objective To investigate technique and clinical effect of total en bloc spondylectomy for thoracic and lumbar chondrosarcoma.Methods From January 2010 to March 2012,6 patients with thoracic or lumbar chondrosarcoma underwent total en bloc spondylectomy.There were 4 males and 2 females,aged from 25 to 54 years (average,38 years).The tumor ranged from T3 to L3; 1 located in T3 and T4,1 in T7,1in T11,1 in L1,1 in L2 and 1 in L3.According to Tomita surgical classification system,there was 1 case of type 2,1 case of type 4,3 cases of type 5 and 1 case of type 6.One patient underwent tumor resection through single posterior approach,while the other 5 patients underwent anterior dissection and posterior resection of tumor.All spines were reconstructed by posterior fixation with pedicle screws and anterior interbody fusion with titanium mesh cages or artificial vertebrae.Results The average amount of blood loss was 3200 ml (range,2100 to 6300 ml).The duration of operation ranged from 3.5 to 12 hours (average,5.5hours).Two patients obtained wide resection,3 obtained marginal resection,and 1 had intralesional margin.The complications included 2 cases of cerebrospinal leak,1 case of pleural effusion and 1 case of pulmonary infection.There was no wound infection and death during peroperative period.All patients were followed up for 6 to 32 months (average,19 months).The neurological function improved from preoperative Frankel C to postoperative Frankel E in 2 cases.All patients obtained bone union 6 to 12 months (average,8 months) after operation.At final follow-up,all patients could walk without aid,and there was no recurrence.Conclusion The total en bloc spondylectomy is an effective method for thoracolumbar chondrosarcoma,which could provide a satisfied tumor control and neurological function improvement.

Objective To investigate technique and indications of proximal femoral autograft for pelvic reconstruction after tumor resection.Methods Between October 2006 and May 2011,5 patients with primary malignant pelvic tumor underwent tumor resection,pelvic reconstruction by proximal femoral autograft,and hip reconstruction by tumor prosthesis.There were 3 males and 2 females,aged from 19 to 55years (average,30.6 years).There were 3 cases of chondrosarcoma and 2 cases of primitive neuroectodermal tumor (PNET).The tumor involved zone Ⅰ and zone Ⅱ in 3 cases,and zone Ⅱ and zone Ⅲ in other 2cases.All 5 patients were followed up,and complications,MSTS (Musculoskeletal Tumour Society) score and prognosis were recorded.Results At final follow-up,one patient died of pulmonary metastasis,one patient survived with local recurrence,and other 3 patients survived without recurrence.The complications consisted of local recurrence,prosthesis loosening,nonunion,infection,and sciatic nerve palsy.One patient underwent revision surgery for prosthesis loosening 26 months after the initial operation.One patient underwent amputation for local recurrence 6 months after the initial operation.The average MSTS score was 19.2.Conclusion Proximal femoral autograft is an effective method for pelvic reconstruction after tumor resection,which can be used for defect in zone Ⅱ and zone Ⅲ,as well as in zone Ⅰ and zone Ⅱ.However,the method has a high rate of complications.The short-term result is similar to that of pelvic prosthesis,while the long-term result needs further observation.

Objective To screen possible factors affecting prognosis of chondrosarcoma.Methods A total of 37 patients with chondrosarcoma,who had undergone surgical treatment from December 2005 to March 2008 in our hospital and had complete follow-up data and definitive pathological diagnosis,were enrolled in this retrospective study.There were 16 males and 21 females,ageing from eleven to eighty-four years (average,42.8 years).The univariate analysis of survival rate was performed with Kaplan-Meier method and tested with the Log-rank test.Chi-squared test or Fisher's exact test were used to analyze numeration data,and then the significant indexes after univariate analysis were performed multivariate analysis with COX regression model to screen the independent factors affecting prognosis.On the basis of literatures,8factors including gender,age,duration of disease,tumor site,Enneking stage,surgical approach,distant metastasis and local recurrence were analyzed.Results Statistical significance was found in Enneking stage,surgical approach and distant metastasis,which indicated that they were related to survival rate 3years after surgery.However,gender,age,duration of disease,tumor site,and local recurrence had no significant correlation with prognosis.After multivariate analysis of Enneking stage,surgical approach,distant metastasis,the results showed that Enneking stage and surgical procedure were the independent prognostic factors,while distant metastasis was not an independent prognostic factor.Conclusion Enneking stage and surgical approach are the independent prognostic factors for chondrosarcoma,which can be used to evaluate prognosis of chondrosarcoma.

Objective To explore the short?term functional outcomes of the reconstruction of the proximal humerus by re?verse shoulder arthroplasty after tumor rescetion. Methods 8 patients who underwent reverse shoulder arthroplasty after tumor resection between January 2013 and December 2014. 5 were female and 3 were male, mean aged was 38 years old (25-61). 2 chon?drosarcomas and 6 giant cell tumors. Enneking stageing of 2 cases with chondrosarcoma were stageⅠB and stageⅡB. 6 giant cell tumors were Campanacci stage 3, meanwhile 3 cases had pathological fractures. The deltoid and axillary nerve were intact in all patients by image analysis before the operation. The proximal humerus was resected according to Malawer typeⅠresection. Then reverse shoulder arthroplasty reconstruction and bone graft was performed. The follow?up was scheduled, and the patient received X?ray examination of the shoulder. The range of motion of the shoulder was measured, the Constant?Murley score and musculoskel?etal tumor society(MSTS) score was recorded. Results The mean duration of the operation was 2.7 h (2-3.5 h). The bleeding in the operation was 510 ml (300-850 ml). The mean length of humerus resection was 8 cm (6-10 cm). The allografts were used in 7 cases and reimplantation after tumor bone deactivation was used in one. The latissimus dorsi transfer were performed in 2 cases. The rotator cuff were resected 1-1.5 cm from the great and lesser tubercles. The follow?up was 13 months (3-26 months). No infec?tion, dislocation, or loosening of prosthesis was found by the last follow?up. The X?ray showed the case who received reimplanta?tion after tumor bone deactivation had achieved bone union 1 year postoperation,7 cases received allograft had still nonunion at the host?graft junction. Bone resorption were found in all cases in different extent but the prosthesis were stable. No local recur?rence of the tumor was found. At last follow?up, active abduction was 155° (100°-175° ) and active forward elevation was 150° (115°-170°) and Constant?Murley score was 76%(68%-87%). The MSTS score was 92%(87%-97%). Conclusion The func?tional outcomes of the reconstruction of the proximal humerus by reverse shoulder arthroplasty after tumor rescetion was satisfied in early period. The reverse shoulder arthroplasty can be used in younger patient, but long?term results need further study.

Objective To analyze the clinical effect of minimally invasive transforaminal lumbar interbody fusion (MI-TLIF) in combination with unilateral pedicle fixation on lumbar degenerative diseases in the elderly.Methods From June 2011 and June 2013,51 elderly patients with lumbar degenerative diseases who underwent MI-TLIF in combination with unilateral pedicle fixation,including 39 males and 12 females,aged from 60 to 72 years (average 65 years old),were included in this study.All the 51 cases had mono segmental lesion,including 11 cases with the lesion in L3.4,22 cases with the lesion in L4.5,and 18 cases with the lesion in L5-S1.The operation time,blood loss,postoperative drainage and complications were recorded.Postoperative outpatient follow-ups were conducted once every 2-3 months for all cases,and JOA scoring criteria (29 points) of lower back pain were adopted to evaluate the clinical effects.Thin-layer helical CT scanning reconstruction was applied to evaluate the fusion rate of bone grafting.X-ray film of lumbar vertebra AP & LAT was employed to evaluate the intervertebral space height,the internal fixation and the position of cage.Results The operation time ranged between 50-90 min with a mean value of 65 min; blood loss ranged between 80-180 ml (average 110 ml); and postoperative drainage volume ranged between 20-70 ml (average 40 ml).Healing phase I was achieved in 49 cases,while post-operative local cutaneous necrosis around the incision site occurred in 2 case and the wound healed well after wound dressing.Dural laceration without nerve root injury occurred in 2 cases.The follow up in 51 patients ranged between 18 24 months (average 20 months).In terms of JOA scoring,the preoperative score was (9.7± 1.6),the post operative score was (21.4 ± 1.4) 7 d after operation and the score was (25.4 ± 1.2) in the final follow up.During the final follow-up,35 cases were evaluated as excellent,11 cases as good and 5 cases as moderate,the excellent and good rate was 90.2％,and the fusion rate of bone grafting was 88.2％.The intervertebral space height ranged from (9.2±1.5) mm before operation to (11.2±1.3) rnm 7 d after operation,to (11.0±1.2) mm in the final follow-up.There were significant difference in intervertebral space height before and after operation (both P＜0.05),and no difference was found between 7d after operation and the final follow-up (P＞0.05).Looseness,fracture or cage displacement were not observed.Conclusions The clinical effect of MI-TLIF in combination with unilateral pedicle fixation on lumbar degenerative diseases in the elderly is satisfied.It has the advantages of high fusion rate,less trauma,quicker recoverer,higher cost effectiveness and convenience.However,the indications for operation should be restricted and the long-term curative effect remains to be studied in further clinical studies.

Objective To investigate feasible immunotherapy strategy using tumor specific cell against osteosarcoma,and to analyze the therapeutic effect of adoptive cellular infusion therapy on osteosarcoma.Methods Decitabine (DAC) was employed as a hypomethylating agent for the treatment in osteosarcoma cell lines HOS and U2OS.After treatment,the expression of cancer-testis antigen (CTA) was evaluated by PCR and Western Blot.In animal studies,human osteosarcoma cell line HOS,which was transfected by luciferase and HLA-A0201 in previous,was inoculated into immune deficient NOD-SCID mice to establish osteosarcoma xenografts.Ex-vivo expanded CTA specific homo CD8+T-ells were labeled with DiR and injected into the mice via the tail vein.In vivo imaging system was utilized to detect the distribution of administrated CD8+ T-cells.In addition,the progression of tumor xenografts was monitored.Moreover,mouse K7M2 osteosarcoma cell line was used to establish animal models in immune competent BALB/c mice.Immune competent models were utilized to evaluate the effectiveness of hypomethylating treatment in regarding to spontaneous immune attack against tumors.Flow cytometry was used to analyze the proportion of intratumoral lymphocytes and the status of these effector antitumor immune cells,and to reveal the effect of hypomethylating treatment in facilitating lymphocyte infiltration and activation.Results The expression of all the evaluated cancer/testis antigens were elevated in HOS and U2OS osteosarcoma cell lines after hypomethylating treatment with DAC.The proliferation of in vitro cultured osteosarcoma cells can be significantly suppressed after at least 5 d treatment with DAC.Besides,DAC alone controlled osteosarcoma cell proliferation.In immune deficient mouse models,hypomethylating pre-treatment resulted in successful T-cell homing to tumor sites.Moreover,the combination treatment with DAC and CTA specific T-cell adoptive transfer significantly suppressed tumor proliferation.In immune competent mouse models,hypomethylating treatment with DAC improved autologous T-cell infiltration into the tumor,and strengthened the activity of intratumoral CD8+ T-cells,elevated the secretion of IFN-gamma,granzyme B and perforin by CD8+ T-cells.Conclusion Hypomethylating treatment is able to suppress osteosarcoma cell proliferation,improve the expression of CTA in osteosarcoma cells,and consequently provide optimal environment for CTA specific T-cell adoptive therapy.