The knockdown of lncRNA DLGAP1-AS2 suppresses osteosarcoma progression by inhibiting aerobic glycolysis via the miR-451a/HK2 axis.

Osteosarcoma (OS) is one of the most aggressive bone tumors worldwide. Emerging documents have shown that long noncoding RNAs (lncRNAs) elicit crucial regulatory functions in the process of tumorigenesis. LncRNA DLGAP1-AS2 is recognized as a regulator in several types of cancers, but its biological functions and molecular mechanisms in OS remain to be elucidated. RT-qPCR and In situ hybridization (ISH) were used to evaluate DLGAP1-AS2 expression in OS samples. Western blotting was used for the measurement of the protein levels of hexokinase 2 (HK2) and epithelial-mesenchymal transition (EMT)-related markers. The proliferation of OS cells was determined using a CCK-8 assay and EdU assay. TUNEL assay and flow cytometry were performed to assess OS cell apoptosis. Glucose metabolism in vitro assays were used. The binding relations among miR-451a, HK2, and DLGAP1-AS2 were validated by luciferase reporter assay. The cellular distribution of DLGAP1-AS2 in OS cells was determined by FISH and subcellular fractionation assays. Mouse xenograft models were established to perform the experiments in vivo. We found that DLGAP1-AS2 expression was upregulated in OS tissues and cells. Downregulation of DLGAP1-AS2 expression suppressed the malignancy of OS cells by restraining cell proliferation, the EMT process, invasiveness, migration, and aerobic glycolysis and accelerating apoptotic behaviors. Of note, silenced DLGAP1-AS2 restrained tumor growth and metastasis in vivo. However, DLGAP1-AS2 overexpression accelerated the progression of OS. We further found that DLGAP1-AS2 upregulation was induced by hypoxia and low glucose. Additionally, DLGAP1-AS2 bound to miR-451a to upregulate HK2 expression. Rescue assays revealed that the DLGAP1-AS2/miR-451a/HK2 axis contributed to OS cell malignancy by promoting aerobic glucose metabolism. Overall, these findings revealed a new regulatory pathway where DLGAP1-AS2 upregulated HK2 expression by sponging miR-451a to accelerate OS development.

Is There an Association Between Bone Microarchitecture and Fracture in Patients who were Treated for High-grade Osteosarcoma? A Controlled Study at Long-term Follow-up Using High-resolution Peripheral Quantitative CT.

BACKGROUND: Neoadjuvant chemotherapy in patients with primary osteosarcoma improves survival rates, but it also causes side effects in various organs including bone. Low bone mineral density (BMD) can occur owing partly to chemotherapy or limited mobility. This can cause a higher risk of fractures compared with those who do not receive such treatment. Changes in BMD alone cannot explain the propensity of fractures. Studying microarchitectural changes of bone might help to understand the effect. QUESTIONS/PURPOSES: (1) Do patients who were treated for osteosarcoma (more than 20 years previously) have low BMD? (2) Do these patients experience more fractures than controls who do not have osteosarcoma? (3) What differences in bone microarchitecture are present between patients treated for high-grade osteosarcoma and individuals who have never had osteosarcoma? METHODS: We contacted 48 patients who were treated for osteosarcoma and who participated in an earlier study. These patients underwent multimodal treatment including chemotherapy more than 20 years ago. Of the original patient group, 60% (29 of 48) were missing, leaving 40% (19 of 48) available for inclusion in this study; all 19 agreed to participate. There were nine men and 10 women with a mean age of 46 ± 4 years and a mean time from surgery to examination of 28 ± 3 years. BMD was measured by dual-energy x-ray absorptiometry, and any fracture history was assessed using a questionnaire. Additionally, high-resolution peripheral quantitative CT was performed to compare the groups in terms of microarchitectural changes, such as cortical and trabecular area, cortical and trabecular thickness, cortical porosity, and endocortical perimeter. Participants in the control group were selected from a cohort consisting of a population-based random sample of 499 healthy adult women and men. Osteoporosis or low BMD was not an exclusion criterion for entering this study; however, the patients in the control group were selected based on a normal BMD (that is, T score > -1.0 at both the spine and hip). Also, the participants were matched based on age and sex. Differences between patients and controls were assessed using the Wilcoxon rank sum test for continuous variables and a chi-square test for categorical variables. A multiple regression analysis was performed. Model assumptions were checked using histograms and quantile-quantile plots of residuals. RESULTS: Twelve of 19 patients who were treated for osteosarcoma had either osteopenia (eight patients) or osteoporosis (four patients). More patients with osteosarcoma reported sustaining fractures (11 of 19 patients) than did control patients (2 of 19 controls; p < 0.001). Among all microarchitectural parameters, only the endocortical perimeter was increased in patients compared with the control group (75 ± 15 mm versus 62 ± 18 mm; p = 0.04); we found no differences between the groups in terms of cortical and trabecular area, cortical and trabecular thickness, or cortical porosity. CONCLUSION: Although patients who were treated for osteosarcoma had osteopenic or osteoporotic BMD and a higher proportion of patients experienced fractures than did patients in the control group, we could not confirm differences in microarchitectural parameters using high-resolution peripheral quantitative CT. Therefore, it seems that bone geometry and microstructural parameters are not likely the cause of the increased proportion of fractures observed in our patients who were treated for osteosarcoma. Until we learn more about the bone changes associated with chemotherapy in patients with osteosarcoma, we recommend that patients undergo regular BMD testing, and we recommend that physicians consider osteoporosis treatment in patients with low BMD. These data might provide the impetus for future multicenter prospective studies examining the association between chemotherapy and bone microarchitecture. LEVEL OF EVIDENCE: Level III, therapeutic study.

Avicularin Reduces the Expression of Mediators of Inflammation and Oxidative Stress in Bradykinin-Treated MG-63 Human Osteoblastic Osteosarcoma Cells.

BACKGROUND Avicularin is a plant-derived flavonoid used in traditional Chinese medicine to treat conditions that include ankle fracture. Bradykinin stimulated MG-63 human osteoblastic osteosarcoma cells has previously been studied in an in vitro model. This study aimed to investigate the effects of avicularin on bradykinin-treated MG-63 human osteoblastic osteosarcoma cells in vitro. MATERIAL AND METHODS MG-63 cells were treated with increasing concentrations of avicularin for 48 hours, followed by 1 µM of bradykinin for 24 h. The MTT assay was used to measure cell viability. Quantitative real-time polymerase chain reaction (qRT-PCR) and enzyme-linked immunosorbent assay (ELISA) were used to measure the expression of inflammatory mediators, interleukin-1ß (IL-1ß), IL-6, and tumor necrosis factor-alpha (TNF-alpha) mRNA and protein, respectively. The expression of oxidative stress factors, malondialdehyde (MDA), superoxide dismutase (SOD), and catalase were measured. Western blot and qRT-PCR were performed to determine p38, p65, inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2) protein levels and mRNA expression, respectively. RESULTS Avicularin had no cytotoxic effect on MG-63 cells. Avicularin significantly upregulated the expression levels IL-1ß, IL-6, and TNF-alpha in the bradykinin treated group in a dose-dependent manner. Avicularin reduced the level of MDA and the activity of SOD and catalase in the bradykinin treated MG-63 cells, reduced p-p38, p-p65, iNOS, and COX-2 expression, and decreased the p-p38/p38 ratio and the p-p65/p65 ratio in bradykinin treated MG-63 cells in a dose-dependent manner. CONCLUSIONS Avicularin reduced the expression of inflammatory cytokines and the levels of markers of oxidative stress in MG-63 human osteoblastic osteosarcoma cells in vitro.

[The role of interleukin-11 in osteosarcoma]. / Die Rolle von Interleukin-11 beim Osteosarkom.

Osteosarcoma is an often highly malignant mesenchymal tumor. By definition, osteosarcoma cells are able to form osteoid, which can mature into tumor bone. Osteosarcoma metastasizes preferentially to the lung. In Europe, the incidence is between 2 and 5 new diagnoses per 1,000,000 people per year. The underlying mechanisms for osteosarcoma formation are not well understood. However, previous radiotherapy or exposition to nuclear radiation increase the risk of osteosarcoma. Patients are usually treated with a neoadjuvant chemotherapy, followed by complete surgical resection of the tumor and post-surgical chemotherapy, which leads to a five-year survival rate of approximately 70% for all stages. Scientific publications in recent years have shown that expression of the cell surface protein interleukin-11 receptor (IL-11R) correlates with a worse prognosis for patients. The IL-11R is activated by its ligand, the cytokine IL-11. IL-11 activates several intracellular signaling cascades within its target cells and is known to be an important regulator of bone homeostasis. Patients with dysfunctional IL-11 signaling display different forms of craniosynostosis. IL-11 induces proliferation of osteosarcoma cell lines in vitro, and the IL-11 signaling cascade was further used to reduce tumor growth and lung metastasis in preclinical mouse models of primary intratibial osteosarcoma. This article gives a comprehensive overview of the frequency, classification, and etiology of osteosarcoma and describes the basic biology of the cytokine IL-11. Furthermore, it summarizes current knowledge about the functional role of IL-11 in osteosarcoma and lists possible therapeutic opportunities.

Molecular mechanisms underlying the inhibitory effects of bovine lactoferrin on osteosarcoma.

Osteosarcoma (OS) is one the most common primary malignancies of the bone in children and young adults with high metastasis. The use of non-toxic naturally derived compounds is one of present strategies in OS therapy to reduce secondary effects and chemo-resistance. Lactoferrin (LF), a transferrin protein derived from milk, currently appears to be an anticancer agent. However, its suppressive effects on OS have not been fully investigated. Therefore, we aimed to examine the molecular mechanism underlying the inhibitory effects of bovine LF (bLF) on OS. OS cell lines (NOS1, U2OS, MG63, and 143B) and an osteoblastic (ST2) were treated with bLF. Effects of bLF on OS-cell proliferation and migration were examined by proliferation and wound-healing assays. Expression levels of low-density-lipoprotein-receptor-related protein 1 (LRP1) and cytokines including interleukin-1 beta (IL-1ß), IL-6, and receptor-activator of nuclear factor kappa-Β ligand (RANKL) were measured using western blotting. Osteoclast formation was examined by co-culture of 143B, ST2, and bone marrow cells. We found that bLF down-regulated IL-1ß, IL-6, and RANKL expression and suppressed phosphorylation of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) p65 in 143B cells; bLF also drastically suppressed 143B-activated RANKL production in ST2 cells. This may have contributed to the reduction in the number of differentiated osteoclasts. Taken together, these data reveal that bLF down-regulates NF-κB to attenuate proliferation, migration, and bone resorption in OS and the OS-microenvironment. This study provides new findings and the precise underlying mechanisms of the inhibitory effects of bLF on OS. bLF can be a possible therapeutic agent for OS patients.

Singe nucleotide polymorphisms in osteosarcoma: Pathogenic effect and prognostic significance.

PURPOSE: Osteosarcoma (OS) is a common malignant bone tumor in children and adolescents. Pathogenesis and prognosis of OS can be associated with several environmental and genetic factors. Single nucleotide polymorphisms (SNPs) are crucial genetic changes that can be involved in clinical and therapeutic outcomes of OS. The aim of this review is to present a synopsis of the role of SNPs in pathogenesis and prognosis of OS tumor cells as well as their potential as therapeutic targets to improve the outcomes of patients. METHOD: The content used in this paper has been obtained by an electronic databases search of English language (1998-2018) articles using the terms "Single nucleotide polymorphisms", "Osteosarcoma", "Pathogenesis", "Prognosis", and "Clinical Outcomes". DISCUSSION: SNPs can affect a number of biological processes such as proliferation, apoptosis, adhesion, invasion, and drug resistance of OS tumor cells, playing a key role in pathogenesis, prognosis, and clinical outcomes after chemotherapy in this disease. CONCLUSION: Considering the importance of SNPs in OS pathophysiology, these genetic changes may be used as potential pathogenic and prognostic biomarkers for OS. It is hoped that targeting these changes using new therapeutic approaches leads to the effective treatment of this debilitating tumor. However, better understanding of OS biology and further clinical trials are needed to achieve this goal.

Miscorrelation of Functional Outcome and Sociooccupational Status of Childhood, Adolescent, and Young Adult Generation With Bone and Soft Tissue Sarcoma Patients.

Few studies have examined the relationship between functional outcome and sociooccupational or psychological status in adolescent and young adults (AYA) generation and childhood sarcoma patients. We retrospectively analyzed clinical (prognostic and functional) and sociooccupational outcomes in 50 patients; 22 children aged under 14 years and 28 AYAs generation (15 to 29 y). There were 35 cases of bone sarcomas and 15 of soft tissue sarcomas. Limb-sparing surgery was performed in 30 of 37 extremity cases. The most prevalent problems among patients were as follows: limited activities; drop-out or delayed studies among high school and college students; limitation in job searching; and changes in social relationships. These problems were unaffected by limb-sparing. Regression analysis between functional and sociooccupational disability showed that the correlation coefficient was significant (P=0.005) in all limb-salvaged patients, but there was no significant correlation among osteosarcoma patients (P=0.07). These findings suggest that quality of life is a multidimensional measure: it depends on physical status, spiritual health, and social well-being of both patients and family members. To overcome the disadvantages of this type of disease, it is essential to provide comprehensive care at the earliest convenience using multidimensional approaches.

microRNA-211 regulates cell proliferation, apoptosis and migration/invasion in human osteosarcoma via targeting EZRIN.

BACKGROUND: In recent years, microRNA-211 (miR211) has been considered as a tumor suppressor in multiple malignancies. However, the function of miR211 in human osteosarcoma has not been explored intensively so far. In this study, the relationship between miR211 and EZRIN was analyzed in human osteosarcoma. METHODS: The expression levels of miR211 and EZRIN were measured in both human osteosarcoma cells and tissues. The direct regulatory relationship between miR211 and EZRIN was evaluated using dual-luciferase assay. The effect of miR211 and EZRIN overexpression on cell proliferation, migration/invasion, and apoptosis was detected. RESULTS: The expression of miR211 was obviously lower in osteosarcoma tissues than paracancerous tissues. EZRIN was identified as the direct target of miR211, and up-regulation of miR211 increased the percentage of cell apoptosis, and suppressed cell proliferation as well as cell migration/invasion via directly regulating EZRIN. CONCLUSIONS: Our study indicated that miR211 has an important role in the development and progress of osteosarcoma, and it might become a novel target in the diagnosis and treatment of human osteosarcoma.

Identifying bone sarcoma survivors facing psychosocial challenges. A study of trajectories following treatment.

OBJECTIVE: Bone sarcoma survivors face a number of physical and psychosocial challenges in relation to the late effects they experience following treatment. The present study aimed to identify and explore the different trajectories that bone sarcoma survivors might navigate during follow-up. METHODS: In-depth and semi-structured interviews were conducted, and an inductive thematic analysis was performed. RESULTS: When they were interviewed three to ten years after the primary diagnosis, the eighteen bone cancer survivors were found to be in three different rehabilitation phases that followed fairly distinct trajectories, namely, back to normal, a new normal and still struggling. Only three participants felt that they had returned to a life that was quite similar to the one they had lived prior to having cancer. Fifteen participants considered their lives and their bodies to be significantly altered. CONCLUSION: Sarcoma survivors who undergo life-changing treatment and return to very different lives than they had before should be identified by healthcare professionals and guided through this demanding phase to better cope with their new living conditions. Information on and tailored guidance related to psychosocial challenges may be of particular importance. Active focus on reorientation, as well as possibilities for growth, seems to be important.

Use of Vascularized Fibular Free Flap in the Reconstruction of the Femur in Pediatric and Adolescent Bone Sarcomas: Complications and Functional Outcome.

BACKGROUND: Most reports on skeletal reconstruction using vascularized fibular free flap include patients with varying age groups and anatomic locations. This study has limited the inclusion criteria to pediatric and adolescent patients diagnosed with bone sarcoma of the femoral shaft. METHODS: Forty-one patients, diagnosed with a malignant bone tumor of the femoral shaft (21 Ewing's sarcomas and 20 osteosarcomas), were locally treated by joint sparing wide resection and reconstruction using a vascularized fibular free flap. All clinical and radiographic data were reviewed for graft healing and hypertrophy as well as oncologic and functional outcome. RESULTS: The mean follow-up period was 48.7 months (12-104 months). The mean age at presentation was 10.3 years (5-17 years). The average length of the resected femoral shaft was 19.2 cm (15-24 cm) and the average length of the harvested fibula was 17.4 cm (15-21 cm). The mean time to union was 4.8 months (1-6 months) and the mean hypertrophy index was 78% (15.5-184%). Complications included 12 fractures (33.3%), 5 non-unions (13.8%), and 5 failures of graft hypertrophy (13.8%). At the latest clinical evaluation, the mean MSTS score was 81% (56-100%) and the mean limb length inequality was 4.75 cm (3-11 cm). CONCLUSION: Despite the high functional demand and deleterious effect of chemotherapy on bone healing, reconstruction of the femur by vascularized fibular free flap in pediatric bone sarcomas can lead to a good functional outcome. Complications, such as fracture and non-union, can be successfully treated by revision of fixation and autologous iliac crest grafting. LEVEL OF EVIDENCE: IV.

Inositol polyphosphate-4-phosphatase type II and rucaparib treatment inhibit the growth of osteosarcoma cells dependent on phosphoinositide 3-kinase/protein kinase B pathway.

Osteosarcoma (OS) is an aggressive malignant tumor of bone, which often occurs in children and adolescents. Currently, the effective method for the treatment of OS is still limited. The study aimed to investigate the synergistic antitumor effect of inositol polyphosphate-4-phosphatase, type-II (INPP4B) and rucaparib on OS cells. The expression levels of INPP4B in OS tissues and OS cell lines were examined by quantitative real-time polymerase chain reaction and Western blot analysis. SaOS2 and U2OS cells were then transfected with overexpression vector of INPP4B or were treated with different concentrations of rucaparib, and cell viability, cell cycle, and apoptosis were determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and flow cytometry. Western blot assay uncovered the combined effects of INPP4B and rucaparib on cell cycle, apoptosis and phosphoinositide 3-kinase/protein kinase B (PI3K/AKT) signal pathway. Further, the tumor formation was examined in vivo. Results showed that INPP4B was low expressed in OS tissues and in OS cell lines. INPP4B overexpression significantly decreased cell viability and induced apoptosis in SaOS2 and U2OS cells. Additionally, rucaparib remarkably reduced cell viability in a dose-dependent and time-dependent manner. Meanwhile, rucaparib suppressed cell cycle progression in the S phase and promoted apoptosis in a dose-dependent manner. Further, combination of INPP4B overexpression and rucaparib declined Myc, cyclin E1 and cyclin D1 expressions, enhanced Bad, Bax, and cleaved-caspase-3 expressions, and blocked PI3K/AKT signal pathway in SaOS2 and U2OS cells. Finally, combination of INPP4B overexpression and rucaparib inhibited tumor formation in vivo. The study demonstrated that INPP4B and rucaparib exhibited synergistic antitumor effect by regulating PI3K/AKT pathway in OS cells.

miR-149 promotes human osteocarcinoma progression via targeting bone morphogenetic protein 9 (BMP9).

OBJECTIVES: To investigate the roles of miR-149 in the progression of human osteosarcoma (OS). RESULTS: miR-149 level was upregulated in tissues from OS patients more than in normal subjects. Cell proliferation and apoptosis assays revealed that miR-149 increased cell proliferation and inhibited cell apoptosis in OS cell line (MG63). An increase of Bcl-2 gene expression and a decrease of cleaved-caspase-3, and cleaved-PARP expression were observed in MG63 cells with transfection of miR-149. Additionally, bone morphogenetic protein 9 (BMP9) was identified as a target of miR-149 in MG63 cells, and BMP9 expression was negatively correlated with miR149 level in OS clinical samples. Co-overexpression of BMP9 with miR-149 in MG63 cells prohibited miR-149-mediated promotive effects on OS progression. Importantly, overexpression of miR-149 conferred chemoresistance in MG63 cells. CONCLUSIONS: miR-149 promotes OS progression via targeting BMP9.

Antibiotic anisomycin induces cell cycle arrest and apoptosis through inhibiting mitochondrial biogenesis in osteosarcoma.

The anti-cancer activities of antibiotic anisomycin have been demonstrated in kidney, colon and ovarian cancers whereas its underlying mechanisms are not well elucidated. In this work, we investigated whether anisomycin is effective in sensitizes osteosarcoma cell response to chemotherapy. We show that anisomycin inhibits proliferation via inducing osteosarcoma cell arrest at G2/M phase, accompanied by the increased levels of mitotic marker cyclin B and the decreased levels of Rb and E2F-1. Anisomycin also induces apoptosis in a caspase-dependent manner in osteosarcoma cells. Importantly, anisomycin is less effective in normal control NIH3T3 cells compared to osteosarcoma cells. In addition, anisomycin inhibits osteosarcoma growth in xenograft mouse model and enhances the inhibitory effects of doxorubicin in osteosarcoma in vitro and in vivo. Mechanistically, anisomycin targets mitochondrial biogenesis in osteosarcoma as shown by the decreased mitochondrial membrane potential, suppressed mitochondrial respiration via decreasing complex I activity, reduced ATP production. Furthermore, mitochondrial biogenesis stimulator acetyl-L-Carnitine (ALCAR) significantly rescues the inhibitory effects of anisomycin in osteosarcoma cells. Our work demonstrates that anisomycin is active against osteosarcoma cells and the molecular mechanism of its action is the inhibition of mitochondrial biogenesis.

LncRNA SNHG15 contributes to proliferation, invasion and autophagy in osteosarcoma cells by sponging miR-141.

BACKGROUND: LncRNA small nucleolar RNA host gene 15 (SNHG15) was reported to play an oncogenic role in tumors. However, the role of SNHG15 and its molecular mechanism in osteosarcoma (OS) cells are largely unknown. METHODS: qRT-PCR was performed to evaluate the expression levels of SNHG15 and miR-141 in OS tissues and cells. Cell transfection with different siRNAs, miRNAs or pcDNAs into U2OS and MG63 cells were carried out by Lipofectamine 2000. The effects of SNHG15 and miR-141 on OS cell proliferation, invasion and the levels of autophagy-related proteins were analyzed by MTT assay, Transwell invasion/migration assay and western blot, respectively. Luciferase reporter assay was used to confirm whether SNHG15 could directly interact with miR-141. RESULTS: We found that up-regulation of SNHG15 was inversely correlated with miR-141 expression in OS tissues. SNHG15 knockdown and miR-141 overexpression significantly suppressed cell proliferation, invasion, migration and autophagy while SNHG15 overexpression and miR-141 repression exhibited the opposite effects on OS cells. Besides, SNHG15 could directly interact with miR-141 and regulate its expression. Furthermore, miR-141 suppressing significantly overturned the inhibition on proliferation, invasion, migration and autophagy mediated by SNHG15 knockdown while miR-141 overexpression remarkably attenuated SNHG15 overexpression-induced proliferation, invasion, migration and autophagy in OS cells. CONCLUSION: Our data showed that SNHG15 contributes to proliferation, invasion, migration and autophagy in OS by negatively regulating miR-141, providing a new potential target and prognostic biomarker for the treatment of OS.

Inherited germline ATRX mutation in two brothers with ATR-X syndrome and osteosarcoma.

We report a family in which two brothers had an undiagnosed genetic disorder comprised of dysmorphic features, microcephaly, severe intellectual disability (non-verbal), mild anemia, and cryptorchidism. Both developed osteosarcoma. Trio exome sequencing (using blood samples from the younger brother and both parents) was performed and a nonsense NM_000489.4:c.7156C>T (p.Arg2386*) mutation in the ATRX gene was identified in the proband (hemizygous) and in the mother's peripheral blood DNA (heterozygous). The mother is healthy, does not exhibit any clinical manifestations of ATR-X syndrome and there was no family history of cancer. The same hemizygous pathogenic variant was confirmed in the affected older brother's skin tissue by subsequent Sanger sequencing. Chromosomal microarray studies of both brothers' osteosarcomas revealed complex copy number alterations consistent with the clinical diagnosis of osteosarcoma. Recently, somatic mutations in the ATRX gene have been observed as recurrent alterations in both osteosarcoma and brain tumors. However, it is unclear if there is any association between osteosarcoma and germline ATRX mutations, specifically in patients with constitutional ATR-X syndrome. This is the first report of osteosarcoma diagnosed in two males with ATR-X syndrome, suggesting a potential increased risk for cancer in patients with this disorder.

Perinatal factors associated with clinical presentation of osteosarcoma in children and adolescents.

BACKGROUND: Osteosarcoma typically develops during puberty with tumors arising at sites of rapid bone growth, suggesting a role for growth-regulating pathways in tumor etiology. Birthweight is one measure of perinatal growth that has been investigated as an osteosarcoma risk factor. Whether birthweight affects clinical features of osteosarcoma remains unexplored. METHOD: Six hundred seventy patients with osteosarcoma, aged 0-19 years, were identified through the California Cancer Registry. We analyzed birth certificate data from the California Department of Public Health vital statistics unit for these patients and 2,860 controls, matched by sex, birth-year, and race/ethnicity. We examined the impact of birthweight on the risk, timing, and clinical presentation of pediatric osteosarcoma including tumor location, size, extension, differentiation, presence of metastasis, and age at onset. Regression models were adjusted for race, sex, gestational age, socioeconomic status, and tumor site. RESULTS: Higher birthweight was associated with more advanced tumor stage (P = 0.017), a trend toward greater tumor extension into surrounding tissues (P = 0.083), and with occurrence of tumors in sites other than the long bones of the arms/legs (P = 9.7 × 10-3 ). Higher birthweight was also associated with an increased likelihood of metastases present at diagnosis (P = 0.047), with each 200 g increase in birthweight associated with a 1.11-fold increase in the odds of having metastatic disease (95% confidence interval: 1.01-1.22). CONCLUSIONS: The association between higher birthweight and more aggressive osteosarcoma, frequently occurring at sites other than the long bones, suggests that growth pathways active during gestation may play an important role in future osteosarcoma progression, especially at anatomic sites with diminished rates of osteoblastic proliferation.

ViscumTT induces apoptosis and alters IAP expression in osteosarcoma in vitro and has synergistic action when combined with different chemotherapeutic drugs.

BACKGROUND: Osteosarcoma is the most common bone tumor and is associated with a poor prognosis. Conventional therapies, surgery and chemotherapy, are still the standard but soon reach their limits. New therapeutic approaches are therefore needed. Conventional aqueous mistletoe extracts from the European mistletoe (Viscum album L.) are used in complementary cancer treatment. These commercial extracts are water-based and do not include water-insoluble compounds such as triterpenic acids. However, both hydrophilic and hydrophobic triterpenic acids possess anti-cancer properties. In this study, a whole mistletoe extract viscumTT re-created by combining an aqueous extract (viscum) and a triterpene extract (TT) was tested for its anti-cancer potential in osteosarcoma. METHODS: Two osteosarcoma cell lines were treated with three different mistletoe extracts viscum, TT and viscumTT to compare their apoptotic potential. For this purpose, annexin/PI staining and caspase-3, -8 and -9 activity were investigated by flow cytometry. To determine the mechanism of action, alterations in expression of inhibitors of apoptosis (IAPs) were detected by western blot. Apoptosis induction by co-treatment of viscum, TT and viscumTT with doxorubicin, etoposide and ifosfamide was examined by flow cytometry. RESULTS: In vitro as well as ex vivo, the whole mistletoe extract viscumTT led to strong inhibition of proliferation and synergistic apoptosis induction in osteosarcoma cells. In the investigations of mechanism of action, inhibitors of apoptosis such as XIAP, BIRC5 and CLSPN showed a clear down-regulation after viscumTT treatment. In addition, co-treatment with doxorubicin, etoposide and ifosfamide further enhanced apoptosis induction, also synergistically. CONCLUSION: ViscumTT treatment results in synergistic apoptosis induction in osteosarcoma cells in vitro and ex vivo. Additionally, conventional standard chemotherapeutic drugs such as doxorubicin, etoposide and ifosfamide were able to dramatically enhance apoptosis induction. These results promise a high potential of viscumTT as an additional adjuvant therapy approach for osteosarcoma.

A Comparison of Vascularized Free Fibular Flaps and Nonvascularized Fibular Grafts for Reconstruction of Long Bone Defects after Tumor Resection.

Background The use of the fibula autograft has been a reliable method in the reconstruction of long bone defects after tumor resection. The objective of this study was to report the outcomes fibular grafting in terms of graft union, functional score, complications, and oncologic outcome. Methods A retrospective review of patients with fibular grafting after tumor resection was done from January 1, 1993 to December 31, 2013. The primary outcome was graft union and the revised musculoskeletal tumor society scoring system (MSTS score). The secondary outcomes were oncologic outcomes, complications, and the factors associated with graft union. Results A total of 52 patients with a mean follow-up of 42 months (SD, 33; range, 12-132 months) were included. The overall union for all fibular grafts was 37 of 52. The use of vascularized free fibula flaps had a higher union rate compared with nonvascularized fibula grafts. The use of a vascularized free fibular flap was four times likely to unite (95% CI 1.1-12.8, p = 0.039) compared with nonvascularized fibular grafts. The mean MSTS score in 36 patients was 82.5 (SD, 12.9) at 35 months from surgery (SD, 30). A total of 39 complications were present in 29 patients. On final follow-up, 45 of 52 patients were alive, six patients had died of disease and one died of other causes. Conclusion A higher union rate was achieved using vascularized free fibular flaps compared with nonvascularized fibular grafts for long bone reconstruction after tumor resection. There was no difference in terms of MSTS score between the two types of grafts.

Osteosarcopenia: A new geriatric syndrome.

BACKGROUND: Longevity, the increase in the ageing population and a lifestyle of minimal physical activity come with a hefty price. Consequently, two diseases are increasingly becoming a concern for the welfare of patients and the health industry: osteoporosis and sarcopenia. These conditions are usually interrelated through several mechanisms and metabolic pathways, and comprise a syndrome called osteosarcopenia. OBJECTIVE: As patients with osteosarcopenia represent an important subset of frail individuals at higher risk of institutionalisation, falls and fractures, the aim of this review is to further familiarise general practitioners with osteosarcopenia as a new geriatric syndrome that requires early diagnosis and effective therapeutic interventions. DISCUSSION: The most important aspects of osteosarcopenia are discussed here. These include pathogenesis, prevalence, diagnostic criteria, management and follow-up. Finally, the role of multidisciplinary clinics for the care of patients with osteosarcopenia is discussed in brief.

Daphnetin inhibits invasion and migration of LM8 murine osteosarcoma cells by decreasing RhoA and Cdc42 expression.

Daphnetin, 7,8-dihydroxycoumarin, present in main constituents of Daphne odora var. marginatai, has multiple pharmacological activities including anti-proliferative effects in cancer cells. In this study, using a Transwell system, we showed that daphnetin inhibited invasion and migration of highly metastatic murine osteosarcoma LM8 cells in a dose-dependent manner. Following treatment by daphnetin, cells that penetrated the Transwell membrane were rounder than non-treated cells. Immunofluorescence analysis revealed that daphnetin decreased the numbers of intracellular stress fibers and filopodia. Moreover, daphnetin treatment dramatically decreased the expression levels of RhoA and Cdc42. In summary, the dihydroxycoumarin derivative daphnetin inhibits the invasion and migration of LM8 cells, and therefore represents a promising agent for use against metastatic cancer.