Proteomics research of bufalin-induced apoptosis in osteosarcoma cell lines / 中国中药杂志

<p><b>OBJECTIVE</b>To study the apoptosis inducing effects of bufalin on various human osteosarcoma cells and the concerning molecular mechanisms.</p><p><b>METHOD</b>MTT assay was used to detect the growth inhibition rates of osteosarcoma cells U-20S, U-20S/MTX300, SaOS-2, IOR/OS9 treated with bufalin in different concentrations and times. The apoptosis of cells was observed flow cytometry 48 h following bufalin treatment. The proteomic techniques were used to separate and compare the treated and control groups 48 h after bufalin-incubation. Then, the proteomic results were validated by western blot.</p><p><b>RESULT</b>Bufalin inhibited the growth of human osteosarcoma cells U20S, U20S/MTX300 (methotrexate resistant cells), SAOS2, IOR/OS9 in a dose- and time-dependent manner. The 72 h IC50 were (37.43 +/- 4.1), (32.24 +/- 5.3) nmol x L(-1) in U20S,U20S/MTX300 cells,respectivly. Flow cytometry showed that the apoptosis cells were increased following bufalin treatment. The protein expression profile showed 24 differentiated expression proteins. Among these proteins, the level of an anti-apoptotic protein, heat shock protein 27 (Hsp27) decreased significantly and the result was then validated by western blot. Ectopic expression of Hsp27 could reduce the bufalin-induced apoptosis remarkably in U20S and U20S/MTX300 cells.</p><p><b>CONCLUSION</b>Bufalin could inhibit the cell growth and induce apoptosis on human osteosarcoma cells. The effect of bufalin may be related to the joint intervention with multiple protein targets. Among them, downregulation of Hsp27 plays a critical role in the bufalin-induced apoptosis in human osteosarcoma cells.</p>

Histone deacetylases inihibitor PCI-24781 induces apoptosis in methotrexate resistant cell line U2-OS/MTX300 in vitro / 中国药学杂志

OBJECTIVE: To study the antitumor activity and the related mechanism of histone deacetylase inihibitor PCI-24781 on methotrexate resistant osteosarcoma cell line, U2-OS/MTX300. METHODS: U-20S/MTX300 cells were treated with PCI-24781. Cell viability was evaluated by MTT and colony formation assay. Apoptosis was evaluated by flow cytometry and fluorescent staining. Western Blot was used to detect the expression of apoptosis-related proteins and the acetylated level of histone 3 and 4. RESULTS: PCI-24781 inhibited the viability of U-20S/MTX300 in a concentration-dependent manner. The IC50 value of PCI-24781 on U-20S/ MTX300 at 48 h after administration was (0.55±0.03) μmol · L -1, which was similar to its IC50 value in the MTX sensitive cells (P>0.05). 0.5 μmol · L-1 PCI-24781 treatment could inhibit the colony formation obviously, and the colony inhibit rate was (61±7)%. Typical apoptotie bodies were observed at 48 h after PCI-24781 treatment and the apoptosis rate was (29±4)%. PCI-24781 could induce up-regulation of cleaved-PARP and P53. The acetylated level of histone 3 and 4 were also significantly increased after PCI-24781 treatment. CONCLUSION: Histone deacetylases inihibitor PCI-24781 could inhibit the growth and induce apoptosis in methotrexate resistant osteosarcoma cells. Up-regulation of acetylated histone proteins and P53 are involved in this process.

Enrichment of osteosarcoma stem cells by chemotherapy / 癌症

Osteosarcoma is the most common primary malignant bone cancer in children and adolescents. Emerging evidence has suggested that the capability of a tumor to grow is driven by a small subset of cells within a tumor, termed cancer stem cells (CSCs). Although several methods have been explored to identify or enrich CSCs in osteosarcoma, these methods sometimes seem impractical, and chemotherapy enrichment for CSCs in osteosarcoma is rarely investigated. In the present study, we found that short exposure to chemotherapy could change the morphology of osteosarcoma cells and increase sarcosphere formation in vitro, as well as increase tumor formation in vivo. Furthermore, methotrexate (MTX)-resistant U2OS/MTX300 osteosarcoma cells were larger in size and grew much more tightly than parental U2OS cells. More importantly, U2OS/MTX300 cells possessed a higher potential to generate sarcospheres in serum-free conditions compared to parental U2OS cells. Also, U2OS/MTX300 cells exhibited the side population (SP) phenotype and expressed CSC surface markers CD117 and Stro-1. Notably, U2OS/MTX300 cells showed a substantially higher tumorigenicity in nude mice relative to U2OS cells. Therefore, we conclude that chemotherapy enrichment is a feasible and practical way to enrich osteosarcoma stem cells.

Wide resection and brachytherapy management of extremity soft tissue sarcoma close to neurovascular bundle / 中华外科杂志

<p><b>OBJECTIVE</b>With the extremity soft tissue sarcoma close to neurovascular bundle, combined en bloc resection and brachytherapy or simple en bloc resection were performed to evaluate the treatment outcome of the combined en bloc resection and brachytherapy.</p><p><b>METHODS</b>Retrospectively investigation was performed for the extremity soft tissue sarcoma close to neurovascular bundle between 2000 and 2009. Inclusion criteria were primary extremity soft tissue sarcoma, MRI showed that the reaction zone involved the main neurovascular bundle, and the reaction zone closed less than 1 cm to the main neurovascular bundle. 86 cases were included in the study. There were 41 men and 45 women. The average age was 38.5 years old (Range from 15 to 73). There were malignant fibrous histiocytoma, synovial sarcoma, fibrosarcoma, liposarcoma, clear cell sarcoma, epithelioid sarcoma, leiomyosarcoma, rhabdomyosarcoma and vascular sarcoma etc. The stage were IA (8), IIA (12), IIB (10), IIC (7), III (43) and IV (6).</p><p><b>RESULTS</b>During an average follow-up of 53 months (range 24 - 102 months), the distant metastasis rate 32.56% (28/86) and the lymph node metastasis rate was 6.98% (6/86). The local recurrence rates was 13.95% (12/86). In the group of combined en bloc resection and brachytherapy with 38 cases, the local recurrence rates was 5.26% (2/38). Four cases had wound infection and six cases had wound delay healing. The MSTS functional score was 21.11 ± 1.79. In the group of simple en bloc resection with 48 cases, the local recurrence rates was 20.83% (10/48). One case had wound infection and four cases had wound delay healing. The MSTS functional score was 84.23% (26.11 ± 1.79). The local recurrence rates was significant different between.</p><p><b>CONCLUSION</b>With the extremity soft tissue sarcoma close to neurovascular bundle, combined en bloc resection and brachytherapy could decrease the local recurrence rate.</p>

Bufalin induces apoptosis in osteosarcoma U-2OS and U-2OS methotrexate 300-resistant cell lines in vitro / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To study the growth inhibition and apoptosis induction effects of bufalin on human osteosarcoma cell lines in vitro.</p><p><b>METHODS</b>U-2OS and U-2OS/methotrexate (MTX) 300-resistant cell lines were treated with bufalin. Cell viability was assessed by MTT assay. Cell-cycle status, apoptosis-inducing effects, and the expression of apoptosis-related proteins were evaluated by flow cytometry, fluorescent staining, DNA fragmentation assay, and Western blotting.</p><p><b>RESULTS</b>Bufalin inhibited cell growth in both U-2OS and U-2OS/MTX300 cells. The IC(50) values of bufalin for U-2OS and U-2OS/MTX300 cells were (8.49 ± 2.1) ng/ml and (10.19 ± 1.7) ng/ml, respectively. The induction of G(2)/M cell-cycle arrest was also seen in the bufalin-treated cells. The bufalin-induced apoptosis was confirmed by increased expression of tumor suppressor protein p53, bax and decreased expression of bcl-2.</p><p><b>CONCLUSION</b>Bufalin inhibits the growth of and induces apoptosis in both MTX-sensitive and MTX-resistant human osteosarcoma U-2OS cell lines. The apoptosis-inducing effect of bufalin is not influenced by the presence of high levels of DHFR.</p>

Detection of serum biomakers of osteosarcoma by proteomic profiling / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To detect differentially expressed proteins in serum of patient with osteosarcoma.</p><p><b>METHODS</b>8 serum protein samples were recruited (4 cases of osteosarcoma and 4 cases of normal adults), cross-labeled with variant CyDye, followed by two-dimensional differential in-gel electrophoresis (2-D DIGE), image analysis, and identified by matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS).</p><p><b>RESULTS</b>24 protein spot-features were significantly increased, and 34 were significantly decreased in the serum from patients with osteosarcoma relative to the controls. The mass spectrometry analysis revealed 18 unique proteins that were increased, and 25 unique proteins decreased in the serum of patients with osteosarcoma. Gelsolin was down-regulated in osteosarcoma, and Western blotting also confirmed a decreased level of gelsolin in the serum of patients with osteosarcoma.</p><p><b>CONCLUSION</b>Our results indicate that gelsolin may have great potential as a biomarker of osteosarcoma and as a potential target for therapy. These preliminary data suggest that incorporation of more samples and new datasets will permit the identification of serum biomarkers for osteosarcoma.</p>

In vitro screening of 32 traditional Chinese herbal extracts against U2OS human osteosarcoma cells / 南方医科大学学报

<p><b>OBJECTIVE</b>To test 32 traditional Chinese medicinal herbs with effects against osteosarcoma in vitro.</p><p><b>METHODS</b>U(2)OS human osteosarcoma cell line was treated with the extracts of the Chinese herbs at various concentrations. The changes in cell proliferation in response to the treatment were examined by MTT assay, and the effects of these extracts against human osteosarcoma growth were compared. Morphological observation, flow cytometry and Annexin V were employed to detect the cell apoptosis after the treatments.</p><p><b>RESULTS</b>According to the results of MTT assay, several of the 32 Chinese herbs, especially Venenum Bufonis and oxgall powder, were identified to produce growth inhibition against U(2)OS cells. Further study of the aqueous extracts of Venenum Bufonis and oxgall powder demonstrated their effects in inducing U(2)OS cell apoptosis, and Venenum Bufonis showed the strongest effect. In spite of the obvious growth inhibitory effect of oxgall powder, its extract induced cell apoptosis only at high concentrations.</p><p><b>CONCLUSION</b>Several of the traditional Chinese herbs, especially Venenum Bufonis and oxgall powder, may inhibit the growth of U(2)OS cell line, and the results of this study pave the way for further identification of the effective components in the herbs that inhibit osteosarcoma growth both in vivo and in vitro.</p>

Analysis of the factors affecting the recurrence of giant cell tumor of bone / 中华外科杂志

<p><b>OBJECTIVE</b>To analyze the clinical factors affecting the recurrence of giant cell tumors (GCT) of bone.</p><p><b>METHODS</b>The complete data of 146 cases with GCT were reviewed. Thirteen clinical factors were analyzed by chi(2) analysis. And the related Campanacci's grade system and Jaffe's grade system was analyzed by Crosstabs analysis. Multipal factors were analyzed by Logistic regression analysis.</p><p><b>RESULTS</b>Nineteen of 146 cases recurred, and recurrence rate was 13.0%. Recurrence rates of curettage and enblock resection groups were 18.8% and 6.3% respectively. And recurrence rates of curettage with or without of extensive procedure were 12.9% and 38.9%. Five cases had lung metastasis, and two cases presented with malignant transformation. The metastasis rate and the rate of malignant transformation were 3.4% and 1.4% respectively. The two factors of surgery method and burst out of bone-envelope appearance were related with the recurrence. Moreover, Logistic regression revealed that the surgery method significantly affected the recurrence. And Campanacci's grade system and Jaffe's grade system were not related to each other.</p><p><b>CONCLUSIONS</b>Surgery method is the main factor affected the recurrence of GCT, and Campanacci's grade system or Jaffe's grade system has no prognostic value.</p>