Mid-term outcomes of right subaxillary approach versus median sternotomy incision for ventricular septal defect with patent ductus arteriosus.

BACKGROUND: This study aimed to evaluate and compare two surgical approaches to repair ventricular septal defect (VSD) with patent ductus arteriosus (PDA) and to explore the patients' health-related quality of life (HRQoL). METHODS: We conducted a retrospective study of all patients who had surgical repair of VSD and PDA between 2013 and 2015 using the right subaxillary approach (group A) or the median sternotomy incision (group B). The outcomes of both techniques were compared. Paediatric QoL Inventory 4.0 scale was applied to assess patients' HRQoL in the 6th postoperative year. Multiple linear regression analysis was performed to explore factors associated with higher HRQoL scores. RESULTS: A total of 128 patients were included (group A, n = 70 and group B, n = 58). Patients in group A were older and heavier than patients in group B. In group B, the diameters of VSD and PDA were larger and the pulmonary artery pressures were higher than those in group A (p < 0.001). No mortality occurred on a mean follow-up of 8.3 ± 1.2 years. Patients in group A had higher HRQoL scores than those in group B in terms of emotional and social functioning dimensions. The right subaxillary approach (OR: 3.56; 95% CI 1.65-5.46), higher parents' education level (OR: 1.62; 95% CI 0.65-2.31), and better family economic status (OR: 1.48; 95% CI 0.79-2.45) were associated with higher HRQoL scores. CONCLUSIONS: Younger and smaller patients receiving median sternotomy incisions due to large defects and pulmonary hypertension had lower HRQoL scores. The right subaxillary approach, higher parents' education level, and better family economic status were associated with higher HRQoL scores.

Do obese patients benefit from isolated aortic valve replacement through a partial upper sternotomy?

OBJECTIVE: Controversial opinions exist for aortic valve replacement (AVR) through partial upper sternotomy in obese patients. Moreover, this study sought to investigate the potential clinical advantage of partial upper sternotomy aortic valve replacement (mini-AVR) over conventional full sternotomy aortic valve replacement (con-AVR) in obese patients. METHODS: This was a retrospective and observational study. From January 2015 to December 2020, a total of 184 obese [body mass index (BMI) ≥ 30 kg  m2] patients undergoing isolated primary AVR were included: 98 patients underwent conventional full sternotomy, and 86 patients underwent partial upper sternotomy. Propensity score (PS) matching was applied to eliminate the bassline imbalances in the mini-AVR and the con-AVR groups. RESULTS: After one-to-one propensity score matching, two groups of 60 patients were obtained. No in-hospital death occurred in the two groups. In addition, cardiopulmonary bypass time and total operative time were similar across the 2 groups, but the aortic cross-clamp time was significantly shorter in the con-AVR group (P = .0.022). The amount of mediastinal drainage at 48 h after surgery (P = 0.018) and postoperative blood transfusions (P = 0.014) were significantly lower in the mini-AVR group. There was no difference in ventilation time (P = .0.145), but a shorter intensive care unit stay time (P = 0.021) in the mini-AVR group. CONCLUSION: This study demonstrates that aortic valve replacement through a mini-AVR in obese patients is a safe and effective procedure. It outperformed con-AVR in terms of blood loss, blood product transfusion, and ICU stay.

METTL14-mediated epitranscriptome modification of MN1 mRNA promote tumorigenicity and all-trans-retinoic acid resistance in osteosarcoma.

BACKGROUND: Osteosarcoma (OS) is the most common primary malignant bone tumor in adolescents. The molecular mechanism behind OS progression and metastasis remains poorly understood, which limits the effectiveness of current therapies. RNA N6-methyladenosine (m6A) modification plays a critical role in influencing RNA fate. However, the biological significance of m6A modification and its potential regulatory mechanisms in the development of OS remain unclear. METHODS: Liquid chromatography-tandem mass spectrometry (LC-MS/MS), dot blotting, and colorimetric ELISA were used to detect m6A levels. Western blotting, quantitative real-time PCR (RT-qPCR) and immunohistochemistry (IHC) were used to investigate METTL14 expression levels. Methylated RNA immunoprecipitation sequencing (MeRIP-seq) and transcriptomic RNA sequencing (RNA-seq) were used to screen the target genes of METTL14. RNA pull-down and RNA immunoprecipitation (RIP) assays were conducted to explore the specific binding of target genes and relevant m6A "readers". RNA stability and polysome analysis assays were used to detect the half-lives and translation efficiencies of the downstream genes of METTL14. IHC and clinical data were applied to explore the clinical correlations of METTL14 and its downstream target genes with the prognosis of OS. FINDINGS: We observed the abundance of m6A modifications in OS and revealed that METTL14 plays an oncogenic role in facilitating OS progression. MeRIP-seq and RNA-seq revealed that MN1 is a downstream gene of METTL14. MN1 contributes to tumor progression and all-trans-retinoic acid (ATRA) chemotherapy resistance in OS. Mechanistically, MN1 is methylated by METTL14, specifically in the coding sequence (CDS) regions, and this modification is recognized by the specific m6A reader insulin-like growth factor 2 mRNA binding protein 2 (IGF2BP2) to prevent MN1 mRNA degradation and promote it translation efficiency. IHC showed that MN1 expression was positively correlated with METTL14 and IGF2BP2 expression in OS tissues. The METTL14-IGF2BP2-MN1 panel demonstrated more promising prognostic value for OS patients than any of these molecules individually. INTERPRETATION: Our study revealed that METTL14 contributes to OS progression and ATRA resistance as an m6A RNA methylase by regulating the stability and translation efficiency of MN1 and thus provides both an underlying biomarker panel for prognosis prediction in OS patients. FUNDING: This work was supported by the National Natural Science Foundation of China (Grants 81972510 and 81772864).

Extensive repair of acute type A aortic dissection through a partial upper sternotomy and using complete stent-graft replacement of the arch.

BACKGROUND: Partial upper sternotomy (mini-ER) can be used in some adult cardiac surgeries but is seldom performed in the treatment of acute type A aortic dissection (AAAD). This study aimed to assess the feasibility and short-term outcomes of complete stent-graft replacement of the arch with triple-branched stent graft for AAAD through a mini-ER. METHODS: From 2015 to 2018, 254 patients with AAAD underwent complete stent-graft replacement of the arch with a triple-branched stent graft. Replacement was performed with conventional full sternotomy (con-ER) in 142 patients and with mini-ER in the other 112 patients. Using propensity score matching, the clinical data were compared between 100 patients in the mini-ER group and 100 patients in the con-ER group. RESULTS: After propensity score matching, there were no significant between-group differences in aortic cross-clamp time, cardiopulmonary bypass time, or total operative time. The amount of mediastinal drainage and number of red blood cell units were significantly lower in the mini-ER group compared with the con-ER group (P < .001). The intubation time was significantly shorter in the mini-ER group (P < .001). The treatment costs were also lower in the mini-ER group (P < .001). There were no significant between-group differences in 30-day mortality (9% vs 8%; P > .99) or postoperative complications. CONCLUSIONS: This study shows that extensive repair of AAAD through a mini-ER is feasible. It was superior to con-ER in terms of blood loss, postoperative ventilation time, and treatment costs.

A Modified Approach with Caval Transection for Supracardiac Total Anomalous Pulmonary Venous Connection: Comparison Between Conventional and Sutureless Surgery in 173 Patients.

The efficacy of primary sutureless repair for supracardiac total anomalous pulmonary venous connection (TAPVC) needs to be confirmed. This study aimed to compare the long-term outcomes between the conventional surgery and the sutureless technique with a modified approach in superior TAPVC. Between January 2008 and December 2018, 173 patients with supracardiac TAPVC underwent surgery either with the conventional procedure (n = 130) or the sutureless repair (n = 43). Multivariate analysis and competing-risk analysis were used to identify risk factors for early death and postoperative pulmonary venous obstruction (PVO), respectively. Among 173 patients who underwent repair of supracardiac TAPVC, 46 (28%) had preoperative PVO, and 22 (12.7%) had postoperative PVO. The sutureless group had a lower postoperative PVO rate compared with the conventional group (p = 0.027). The risk factors for death were age ≤ 28 days [odds ratio (OR), 11.56; 95% confidence interval (CI) 1.33-100.47, p = 0.015], weight ≤ 3 kg (OR 9.57; 95% CI 1.58-58.09, p = 0.009), emergency operation (OR 19.24; 95% CI 3.18-116.35, p = 0.002), cardiopulmonary bypass time (OR 2.16; 95% CI 1.36-3.43, p = 0.003), cross-clamp time (OR 1.73; 95% CI 1.20-2.50, p = 0.022), and duration of ventilation (OR 1.11; 95% CI 1.02-1.21, p = 0.027). Age ≤ 28 days [Hazard Ratio (HR) 1.92; 95% CI 1.92-11.02, p < 0.001] and preoperative PVO (HR 41.70; 95% CI 8.15-213.5, p < 0.001) were associated with postoperative PVO. The sutureless repair is a reliable technique for supracardiac TAPVC. Age ≤ 28 days is associated with 30-day mortality and postoperative PVO.

A Novel Method to Treat Progressive Desmoid Tumors Involving Neurovascular Bundles: A Retrospective Cohort Study.

BACKGROUND: More effective therapies are needed to treat progressive desmoid tumors when active surveillance and systemic therapy fail. OBJECTIVE: To assess the efficacy and safety of sandwich isolation surgery on the local control of progressive desmoid tumors involving neurovascular bundles. METHODS: A total of 27 patients with progressive desmoid tumors at extremities involving neurovascular bundles who received surgery at our hospital between August 2014 and August 2018 were identified. A total of 13 patients received sandwich isolation surgery, in which R2 resection was performed in neurovasculature-involving regions, and a biomaterial patch was used to envelop involved neurovascular structures and isolate residual tumors. In non-neurovasculature-involving regions, wide resection was performed without isolation. A total of 14 patients received traditional surgery, which included tumor resection without isolation procedure. RESULTS: In sandwich isolation group, tumor progressions and local recurrences occurred in 3 patients outside the isolated neurovasculature-involving regions. However, no progressions or recurrences occurred in any patients in the isolated neurovasculature-involving regions where R2 resection was performed. Sandwich isolation surgery group and traditional surgery group shared similar baseline clinical characteristics. The estimated 3-yr event-free survival rate was 76.9% after sandwich isolation surgery, and 32.7% after traditional surgery (P = .025). Patients who received sandwich isolation surgery were less likely to have local recurrence (hazard ratio: 0.257, P = .040). No complications were noted except intermittent mild pain in operative regions (2 cases). CONCLUSION: Sandwich isolation surgery is effective and safe for local control of desmoid tumors involving neurovascular bundles.

Discontinuous polyostotic fibrous dysplasia with multiple systemic disorders and unique genetic mutations: A case report.

BACKGROUND: Polyostotic fibrous dysplasia (PFD) is an uncommon developmental bone disease in which normal bone and marrow are replaced by pseudotumoral tissue. The etiology of PFD is unclear, but it is generally thought to be caused by sporadic, post-zygotic mutations in the GNAS gene. Herein, we report the case of a young female with bone pain and lesions consistent with PFD, unique physical findings, and gene mutations. CASE SUMMARY: A 27-year-old female presented with unbearable bone pain in her left foot for 4 years. Multiple bone lesions were detected by radiographic examinations, and a diagnosis of PFD was made after a biopsy of her left calcaneus with symptoms including pre-axial polydactyly on her left hand and severe ophthalmological problems such as high myopia, vitreous opacity, and choroidal atrophy. Her serum cortisol level was high, consistent with Cushing syndrome. Due to consanguineous marriage of her grandparents, boosted whole exome screening was performed to identify gene mutations. The results revealed mutations in HSPG2 and RIMS1, which may be contributing factors to her unique findings. CONCLUSION: The unique findings in this patient with PFD may be related to mutations in the HSPG2 and RIMS1 genes.

Telangiectatic osteosarcoma: Outcome analyses and a diagnostic model for differentiation from aneurysmal bone cyst.

BACKGROUND AND PURPOSE: Telangiectatic osteosarcoma (TOS), a rare variant of osteosarcoma, may be easily misdiagnosed as aneurysmal bone cyst (ABC). The aims of this study were to investigate the diagnostic and prognostic factors of TOS by reviewing our experience with TOS and to develop a diagnostic model that may distinguish TOS from ABC. MATERIALS AND METHODS: We identified 51 cases of TOS treated at the First Affiliated Hospital of Sun Yat-Sen University from March 2001 to January 2016 and reviewed their records, imaging information and pathological studies. A diagnostic model was developed to differentiate TOS and ABC by Bayes discriminant analysis and was evaluated. The log-rank test was used to analyze the prognostic factors of TOS and to compare the outcome differences between TOS and other high-grade osteosarcoma subtypes. RESULTS: The multi-disciplinary diagnostic method employed that combined clinical, imaging, and pathological studies enhanced the diagnostic accuracy. Age 18 years or younger and pathologic fracture were more common among the TOS patients than among the ABC patients (P = .004 and .005, respectively). The average white blood cell (WBC), platelet, lactate dehydrogenase (LDH), and alkaline phosphatase (ALP) values of the TOS patients were higher than those of the ABC patients (P = .002, .003, .007, and .007, respectively). Our diagnostic model, including the aforementioned factors, accurately predicted 62% and 78% of the TOS patients in the training and validation sets, respectively. The 5-year estimates of event-free survival and overall survival of the TOS patients were 52.5 ± 9.4% and 54.9 ± 8.8%, respectively, which were similar to those of patients with other osteosarcoma subtypes (P = .950 and .615, respectively). Tumor volume and the LDH level were predictive prognostic factors (P = .040 and .044) but not the presence of pathologic fracture or misdiagnosis (P = .424 and .632, all respectively). CONCLUSIONS: The multi-disciplinary diagnostic method and diagnostic model based on predictive factors, i.e., age, the presence of pathologic fracture, and platelet, LDH, ALP and WBC levels, aided the differentiation of TOS and ABC. Smaller tumors and normal LDH levels were associated with better outcomes.

LDOC1 regulates Wnt5a expression and osteosarcoma cell metastasis and is correlated with the survival of osteosarcoma patients.

Osteosarcomas are common bone malignancies in children and adolescents. LDOC1 (leucine zipper, down-regulated in cancer 1), a tumor suppressor, is down-regulated in many cancers. In this study, we investigated the role of LDOC1 in tumor metastasis and its prognostic significance in osteosarcomas. We established osteosarcoma cells stably expressing LDOC1, driven by an HIV-based lentiviral system. We investigated the impact of LDOC1 on migration and invasion abilities in these cells using a transwell assay. LDOC1-associated changes in expression of metastasis-promoting genes were analyzed with a quantitative real-time polymerase chain reaction primer array. A xenograft tumor model (n = 7 mice/group) was used to assess the effect of LDOC1 on osteosarcoma metastasis in vivo. The overall survival and disease-free survival of osteosarcoma patients (n = 74) were analyzed retrospectively based on immunohistochemical analysis of LDOC1 levels in tumors and Kaplan-Meier analysis. LDOC1-expressing osteosarcoma cells displayed decreased migration and invasion in vitro. The quantitative real-time polymerase chain reaction primer array data showed that increased LDOC1 expression up-regulated many metastasis-suppressor genes. In the xenograft model, micro-computed tomography imaging data indicated that increased LDOC1 expression is associated with weaker lung metastasis ability. The Wnt5a signaling pathway promotes osteosarcoma metastasis; LDOC1 expression decreased Wnt5a levels in osteosarcoma cells. Kaplan-Meier analysis showed that higher LDOC1 expression was associated with improved osteosarcoma patient overall survival and disease free survival (p = 0.022). Our data show that LDOC1 is a tumor suppressor in osteosarcoma, and that it regulates metastasis of osteosarcoma cells. Furthermore, LDOC1 might be a valuable prognostic marker in osteosarcomas.

A proposed classification system for guiding surgical strategy in cases of severe spinal deformity based on spinal cord function.

PURPOSE: Spinal cord function classification systems are not useful for guiding surgery in patients with severe spinal deformities. The aim of this study is to propose a classification system for determining a surgical strategy that minimizes the risk of neurological dysfunction in patients with severe spinal deformities. METHODS: The records of 89 patients with severe spinal deformities treated with vertebral column reconstruction from 2008 to 2013 were retrospectively analyzed. Based on neurophysiological monitoring, magnetic resonance imaging, and neurological symptoms patients were categorized into three groups: group A, normal spinal cord, normal evoked potentials and no neurological symptoms; group B, spinal cord abnormalities and/or abnormal evoked potentials but no neurological symptoms; group C, neurological symptoms with or without spinal cord abnormalities/abnormal evoked potentials. Outcomes and complications were compared between the groups. RESULTS: A total of 89 patients (51 male, 38 female) were included with 47 (52.8 %), 16 (18.0 %), and 26 (29.2 %) patients in groups A, B and C, respectively, and a mean follow-up 34.5 months. There were no differences in age, gender, average preoperative scoliosis, and kyphosis among three groups, but there were differences with respect to the causes of severe spinal deformity and the corrective rate of scoliosis and kyphosis. Changes in intraoperative evoked potentials were different in these three types according to this new classification, and the recovery rates of changes in the three groups were 71.1, 50.0, and 14.1 %, respectively. Postoperative spinal cord injury was positively related to intraoperative changes of evoked potentials. CONCLUSION: The classification system may be useful for guiding surgical decisions in patients with severe spinal deformities to minimize the risk of neurological complications.

Ampelopsin induces apoptosis by regulating multiple c-Myc/S-phase kinase-associated protein 2/F-box and WD repeat-containing protein 7/histone deacetylase 2 pathways in human lung adenocarcinoma cells.

Ampelopsin (AMP), a plant flavonoid, has been reported to inhibit cell growth and/or induce apoptosis in various types of tumor. The aim of the present study was to assess the apoptosis-inducing activity of AMP in A549 human lung adenocarcinoma epithelial cells and the associated underlying mechanism. A549 cells were incubated with different concentrations of AMP in culture medium. Cell growth and apoptosis were evaluated by MTT assay and Annexin V/propidium iodide double staining and flow cytometry, respectively. In addition, western blotting and reverse transcription quantitative polymerase chain reaction analysis were used to examine the time-dependent changes in protein expression. Certain changes in apoptotic protein expression were detected following exposure to AMP, including X-linked inhibitor of apoptosis protein release, reduced B-cell lymphoma 2, myeloid cell leukemia 1 and survivin expression levels, increased Bcl-2-associated X protein expression levels and cleaved-poly ADP ribose polymerase expression. The results revealed that AMP was a potent inhibitor of A549 cell proliferation. The c-Myc/S-phase kinase-associated protein 2 (Skp2) and histone deacetylase (HDAC)1/2 pathways were found to exert an important role in AMP-induced A549 cell apoptosis, as increased levels of c-Myc mRNA and reduced levels of c-Myc/Skp2 and HDAC1 and 2 proteins following AMP treatment were observed. The levels of F-box and WD repeat-containing protein 7α (Fbw7α), Fbw7ß, Fbw7γ, phosphorylated-(p-)c-Myc (Thr58) and glycogen synthase kinase 3ß (GSK3ß) proteins involved in c-Myc ubiquitin-dependent degradation were also analyzed. Following exposure to AMP, the expression levels of Fbw7α, Fbw7γ and GSK3ß were reduced and p-c-Myc (Thr58) expression levels were increased. The results suggest that AMP exerts an anticancer effect, which is associated with the degradation of c-Myc, Skp2 and HDAC1 and 2. The ability of AMP to induce apoptosis independently of Fbwα and Fbw7γ suggests a possible use in drug-resistant cancer associated with Fbw7 deficiency. Understanding the exact underlying mechanism requires further investigation of the association between c-Myc and Fbw7α/γ reversal, and analysis of whether Thr58 phosphorylation of c-Myc is dependent on GSK3ß.

Local control of giant cell tumors of the long bone after aggressive curettage with and without bone cement.

BACKGROUND: Aggressive curettage has been well established for the treatment of giant cell tumors (GCTs) of the bone. The purpose of this study was to review our experience and evaluate the role of different implant materials in patients with GCTs of the extremities after aggressive curettage. METHODS: A total of 119 patients with GCTs of the long bone were treated at the First Affiliated Hospital of Sun Yat-Sen University between 2004 and 2009. We excluded patients presenting metastases, recurrent tumors, and soft tissue involvement and those with Jaffe pathological grade III. The remaining 65 patients were treated with aggressive curettage using a bone graft or bone cement to fill the cavity. The recurrence rates and functional scores associated with the different fillings were analyzed. RESULTS: Aggressive curettage and bone grafting was performed in 34 cases (52.3%), and aggressive curettage with bone cement was performed in 31 cases (47.7%). The overall recurrence rate after the aggressive intralesional procedures was 35.3% with bone grafting and 12.9% when bone cement was used as an adjuvant filling. The recurrence rate following aggressive curettage and bone grafting was higher than that following aggressive curettage with cement (p = 0.038). The Musculoskeletal Tumor Society (MSTS) score for bone graft patients was 91.1%, which was significantly lower than that for patients treated with bone cement (94.7%). CONCLUSIONS: The use of bone cement was associated with a significantly lower recurrence rate than bone grafting following aggressive intralesional curettage to treat benign giant cell tumors of the long bone. Better MSTS functional results were also observed in the bone cement group compared to the bone graft group.

[Proteomics research of bufalin-induced apoptosis in osteosarcoma cell lines].

OBJECTIVE: To study the apoptosis inducing effects of bufalin on various human osteosarcoma cells and the concerning molecular mechanisms. METHOD: 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. RESULT: 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. CONCLUSION: 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.

Dihydromyricetin activates AMP-activated protein kinase and P38(MAPK) exerting antitumor potential in osteosarcoma.

Numerous patients with osteosarcoma either are not sensitive to chemotherapy or develop drug resistance to current chemotherapy regimens. Therefore, it is necessary to develop several potentially useful therapeutic agents. Dihydromyricetin is the major flavonoid component derived from Ampelopsis grossedentata, which has a long history of use in food and medicine. The present study examined the antitumor activity both in vitro and in vivo without noticeable side effects and the underlying mechanism of action of dihydromyricetin in osteosarcoma cells. We found that dihydromyricetin induced increased p21 expression and G2-M cell-cycle arrest, caused DNA damage, activated ATM-CHK2-H2AX signaling pathways, and induced apoptosis in osteosarcoma cells as well as decreasing the sphere formation capability by downregulating Sox2 expression. Mechanistic analysis showed that the antitumor potential of dihydromyricetin may be due to the activation of AMPKα and p38(MAPK), as the activating AMPKα led to the inactivation of GSK3ß in osteosarcoma cells. Moreover, GSK3ß deletion or GSK3ß inhibition by LiCl treatment resulted in increased p21 expression and reduced Sox2 expression in osteosarcoma cells. Taken together, our results strongly indicate that the antitumor potential of dihydromyricetin is correlated with P38(MAPK) and the AMPKα-GSK3ß-Sox2 signaling pathway. Finally, immunohistochemical analysis indicated that some patients had a lower p-AMPK expression after chemotherapy, which supports that the combination of dihydromyricetin and chemotherapy drug will be beneficial for patients with osteosarcoma. In conclusion, our results are the first to suggest that dihydromyricetin may be a therapeutic candidate for the treatment of osteosarcoma.

Critical role of heat shock protein 27 in bufalin-induced apoptosis in human osteosarcomas: a proteomic-based research.

Bufalin is the primary component of the traditional Chinese herb "Chan Su". Evidence suggests that this compound possesses potent anti-tumor activities, although the exact molecular mechanism(s) is unknown. Our previous study showed that bufalin inhibited growth of human osteosarcoma cell lines U2OS and U2OS/MTX300 in culture. Therefore, this study aims to further clarify the in vitro and in vivo anti-osteosarcoma effects of bufalin and its molecular mechanism of action. We found bufalin inhibited both methotrexate (MTX) sensitive and resistant human osteosarcoma cell growth and induced G2/M arrest and apoptosis. Using a comparative proteomics approach, 24 differentially expressed proteins following bufalin treatment were identified. In particular, the level of an anti-apoptotic protein, heat shock protein 27 (Hsp27), decreased remarkably. The down-regulation of Hsp27 and alterations of its partner signaling molecules (the decrease in p-Akt, nuclear NF-κB p65, and co-immunoprecipitated cytochrome c/Hsp27) were validated. Hsp27 over-expression protected against bufalin-induced apoptosis, reversed the dephosphorylation of Akt and preserved the level of nuclear NF-κB p65 and co-immunoprecipitated Hsp27/cytochrome c. Moreover, bufalin inhibited MTX-resistant osteosarcoma xenograft growth, and a down-regulation of Hsp27 in vivo was observed. Taken together, bufalin exerted potent anti-osteosarcoma effects in vitro and in vivo, even in MTX resistant osteosarcoma cells. The down-regulation of Hsp27 played a critical role in bufalin-induced apoptosis in osteosarcoma cells. Bufalin may have merit to be a potential chemotherapeutic agent for osteosarcoma, particularly in MTX-resistant groups.

Glycogen synthase kinase-3ß, NF-κB signaling, and tumorigenesis of human osteosarcoma.

BACKGROUND: Glycogen synthase kinase-3ß (GSK-3ß), a serine/threonine protein kinase, may function as a tumor suppressor or an oncogene, depending on the tumor type. We sought to determine the biological function of GSK-3ß in osteosarcoma, a rare pediatric cancer for which the identification of new therapeutic targets is urgent. METHODS: We used cell viability assays, colony formation assays, and apoptosis assays to analyze the effects of altered GSK-3ß expression in U2OS, MG63, SAOS2, U2OS/MTX300, and ZOS osteosarcoma cell lines. Nude mice (n = 5-8 mice per group) were injected with U2OS/MTX300, and ZOS cells to assess the role of GSK-3ß in osteosarcoma growth in vivo and to evaluate the effects of inhibitors and/or anticancer drugs on tumor growth. We used an antibody array, polymerase chain reaction, western blotting, and a luciferase reporter assay to establish the effect of GSK-3ß inhibition on the nuclear factor-κB (NF-κB) pathway. Immunochemistry was performed on primary tumor specimens from osteosarcoma patients (n = 74) to determine the relationship of GSK-3ß activity with overall survival. RESULTS: Osteosarcoma cells with low levels of inactive p-Ser9-GSK-3ß formed colonies in vitro and tumors in vivo more readily than cells with higher levels and cells in which GSK-3ß had been silenced formed fewer colonies and smaller tumors than parental cells. Silencing or pharmacological inhibition of GSK-3ß resulted in apoptosis of osteosarcoma cells. Inhibition of GSK-3ß resulted in inhibition of the NF-κB pathway and reduction of NF-κB-mediated transcription. Combination treatments with GSK-3ß inhibitors, NF-κB inhibitors, and chemotherapy drugs increased the effectiveness of chemotherapy drugs in vitro and in vivo. Patients whose osteosarcoma specimens had hyperactive GSK-3ß, and nuclear NF-κB had a shorter median overall survival time (49.2 months) compared with patients whose tumors had inactive GSK-3ß and NF-κB (109.2 months). CONCLUSION: GSK-3ß activity may promote osteosarcoma tumor growth, and therapeutic targeting of the GSK-3ß and/or NF-κB pathways may be an effective way to enhance the therapeutic activity of anticancer drugs against osteosarcoma.

Eosinophilic granuloma of bilateral humerus with unusual diagnostic process.

A 15-year-old boy presented with a right shoulder pain and the inability to use his right upper limb after carrying a heavy object. Physical examination and local imaging revealed a benign tumor or tumor-like lesion in the upper ends of both humeri and a pathologic fracture in the right humerus. An initial biopsy was reported as fibroblastic osteosarcoma. But the CT-guided biopsy was diagnosed as eosinophilic granuloma (EG). Bone curretage and grafting confirmed diagnosis of EG. This case illustrates the importance of combining a clinical impression with imaging and pathology in bone and soft tissue tumors.

Salinomycin inhibits osteosarcoma by targeting its tumor stem cells.

Osteosarcoma is the most common primary bone tumor in children and adolescents and is typically associated with a poor prognosis. Tumor stem cells (TSCs) are presumed to drive tumor initiation and tumor relapse or metastasis. Hence, the poor prognosis of osteosarcoma likely results from a failure to target the osteosarcoma stem cells. Here, we have utilized three different methods to enrich TSCs in osteosarcoma and further evaluated whether salinomycin could selectively target TSCs in osteosarcoma. Our results indicated that sarcosphere selection, chemotherapy selection and stem cell marker OCT4 or SOX2 over-expression are all effective in the enrichment of TSCs from osteosarcoma cell lines. Further investigation found that salinomycin inhibited osteosarcoma by selectively targeting its stem cells both in vitro and in vivo without severe side effects, and the Wnt/ß-catenin signaling pathway may be involved in this inhibition of salinomycin. Taken together, we have identified that salinomycin is an effective inhibitor of osteosarcoma stem cells, supporting the use of salinomycin for elimination of osteosarcoma stem cells and implying a need for further clinical evaluation.

Enrichment of osteosarcoma stem cells by chemotherapy.

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.

Expression change of ezrin as a prognostic factor in primary osteosarcoma.

Osteosarcoma is a life-threatening malignancy that most often occurs in teenagers. Ezrin is a cytoskeleton linker protein that is actively involved in the metastatic capacity of cancer cells. In this study, we investigated the expression change of ezrin after preoperative chemotherapy and its prognostic value in patients with primary osteosarcoma. Ezrin mRNA expression level of initial biopsy specimens and resected tumor specimens after preoperative chemotherapy of 25 patients who had primary osteosarcoma with ezrin expression was detected by semiquantitative reverse transcription polymerase chain reaction (RT-PCR) to evaluate the expression change of ezrin. In addition, the prognostic factors and the relationship between the expression change of ezrin and the clinical characteristics were analyzed. Expression change of ezrin was found in 64% of all 25 patients. The expression change of ezrin had good relation with histology (P = 0.037), grade (P = 0.006), chemotherapy response (P = 0.017), and metastasis or recurrence (P = 0.041). The 2-year overall survival and event-free survival were associated with expression change of ezrin (P = 0.001 and P = 0.002, respectively) and response to preoperative chemotherapy (P = 0.001 and P = 0.012, respectively). Our findings suggest that expression change of ezrin is an independent positive prognostic factor in non-metastatic osteosarcoma.