Prognostic significance of the expression of HER family members in primary osteosarcoma.

The prognosis of patients with metastatic osteosarcoma is poor and has shown no significant improvement in nearly 20 years. The human epidermal growth factor (EGF) receptor (HER) family is frequently overexpressed in the majority of human carcinomas, and is involved in promoting the proliferation and survival of cancer cells. However, the role of EGFR and HER-2 expression in osteosarcoma survival remains controversial and no previous study has simultaneously investigated the association of the expression of all the four HER family members with the prognostic significance of osteosarcoma. Therefore, the present study investigated the expression levels of the complete members of the HER family in osteosarcoma specimens, as well as their associations with the clinicopathological parameters, progression-free survival (PFS) and overall survival (OS) time of patients with osteosarcoma. The expression of HER family members was detected in osteosarcoma tumor specimens from 60 patients using immunohistochemistry. The association of the expression of HER receptors in osteosarcoma with clinicopathological parameters was analyzed using χ2 test and Fishers exact test. Survival analyses were evaluated by Kaplan-Meier method and Cox proportional hazards regression model. Overall, 18 (30%), 13 (22%), 23 (38%) and 19 (32%) patients presented with high expression of EGFR, HER-2, HER-3 and HER-4, respectively, and the co-expression of 2, 3 and all 4 members of the HER family was observed. High expression of EGFR and HER-4 was associated with distant metastasis. High HER-3 expression was significantly associated with an advanced Enneking stage and distant metastasis. Multivariate analysis demonstrated that the expression of EGFR, HER-3, HER-4, EGFR/HER-3, EGFR/HER-4 and HER-3/HER-4 was an independent predictor of poor PFS and OS time in osteosarcoma patients with stage I-IIB disease. In patients with stage IIB osteosarcoma, the expression of HER-4 and EGFR/HER-4 demonstrated a more significant effect on PFS and OS time. In conclusion, therapies targeting EGFR, HER-3 and HER-4 may provide promising strategies for primary osteosarcoma.

The clinical significance of the Ezrin gene and circulating tumor cells in osteosarcoma.

PURPOSE: The aim of this study was to investigate the clinical significance of circulating tumor cells (CTCs) in the peripheral blood of an osteosarcoma and the Ezrin gene expressed in CTCs. PATIENTS AND METHODS: CTC enrichment was done with CanPatrol™ CTC enrichment technique in 41 patients with osteosarcoma. The characterization of CTCs was performed using a multiple messenger RNA in situ analysis (MRIA). The expression of the Ezrin gene in CTCs was detected by RNA probe technology. The correlations of CTC counts, cell type and the expression level of the Ezrin gene with clinical stage and metastasis of osteosarcoma were analyzed using SPSS 16.0 software. RESULTS: The CTC counts correlated significantly with Enneking stage (P<0.001). The ratio of mesenchymal CTCs correlated with the distant metastases (P<0.001). Ezrin gene expression in CTCs correlated significantly with distant metastases (χ2=152.51, P=0.000). CONCLUSION: The ratio of mesenchymal CTCs in the peripheral blood of osteosarcoma correlates with distant metastases. High expression of Ezrin gene in CTCs correlates with distant metastases.

Dual-probe electrochemical DNA biosensor based on the "Y" junction structure and restriction endonuclease assisted cyclic enzymatic amplification for detection of double-strand DNA of PML/RARα related fusion gene.

Taking advantage of "Y" junction structure and restriction endonuclease assisted cyclic enzymatic amplification, a dual-probe electrochemical DNA (DE-DNA) biosensor was designed to detect double-stranded DNA (dsDNA) of acute promyelocytic leukemia (APL) related gene. Two groups of detection probes were designed, and each group was composed of a biotinylated capture probe and an assisted probe. They were separately complementary with two strands of target dsDNA in order to prevent the reannealing of the two separate strands from target dsDNA. First, thiol functionalized capture probes (C1 and C2) were severally assembled onto two different gold electrodes, followed by hybridizing with target dsDNA (S1a-S1b) and assistant probes to form two Y-junction-structure ternary complexes. Subsequently, restriction sites on the ternary complexes were digested by Rsa I, which can release S1a, S1b and biotins from the electrode surfaces. Meanwhile, the released S1a and S1b can further hybridize with the unhybridized corresponding detection probes and then initiate another new hybridization-cleavage-separation cycle. Finally, the current signals were produced by the enzyme-catalyzed reaction of streptavidin-horse reddish peroxidase (streptavidin-HRP). The distinct difference in current signals between different sequences allowed detection of target dsDNA down to a low detection limit of 47 fM and presented excellent specificity with discriminating only a single-base mismatched dsDNA sequence. Moreover, this biosensor was also used for assay of polymerase chain reaction (PCR) samples with satisfactory results. According to the results, the power of the DE-DNA biosensor as a promising tool for the detection of APL and other diseases.

Electrochemical immunosensor for detection of topoisomerase based on graphene-gold nanocomposites.

A facile electrochemical immunosensor based on graphene-three dimensional nanostructure gold nanocomposites (G-3D Au) using simple and rapid one-step electrochemical co-reduction technique was developed for sensitive detection of topoisomerase. The resultant G-3D Au nanocomposites were characterized by scanning electron microscopy, cyclic voltammetry and electrochemical impedance spectroscopy, and then were used as a substrate for construction of the "sandwich-type" immunosensor. Amperometric current-time curve was employed to monitor the immunoreaction on the protein modified electrode. The proposed method could respond to topoisomerase with a linear calibration range from 0.5 ng mL(-1) to 50 ng mL(-1) with a detection limit of 10 pg mL(-1). This new biosensor exhibited a fast amperometric response, high sensitivity and selectivity, and was successfully used in determining the topoisomerase which was added in human serum with a relative standard deviation (n=5)<5%. The immunosensor served as a significant step toward the practical application of the immunosensor in clinical diagnosis and prognosis monitor.

Overexpressing neuroglobin improves functional recovery by inhibiting neuronal apoptosis after spinal cord injury.

The current study was performed to evaluate the mechanisms and therapeutic effects of overexpressing neuroglobin (Ngb) on spinal cord injury (SCI). Adeno-associated virus (AAV) was injected in the T12 section 7 days before SCI. Animals were randomly divided into four groups: a sham group, a vehicle group, an AAV-EGFP group and an AAV-Ngb group. Recovery of hind limb locomotor function was determined during the 3-week post operation period by the Basso, Beattie and Bresnahan locomotor rating scale. At 24 h after SCI and at the end of the study, the segments of spinal cord, centered with the lesion site were harvested for histopathological analysis. Immunofluorescence was performed using antibodies to recognize neuN in the lesion sections. At 24 h after SCI, the spinal cord tissue samples were removed to analyze tissue concentrations of superoxide dismutase (SOD) and malondialdehyde (MDA). Apoptotic cells were assessed using a terminal deoxynucleotidyl transferase, dUTP nick end labeling (TUNEL) kit. The expression of bcl-2, bax, cytochrome c, and cleaved caspase-3, were determined by Western blot assay and immunostaining analysis. The results showed that animals overexpressing Ngb had significantly greater recovery of locomotor function, less neuronal loss and fewer apoptotic cells. In addition, overexpressing Ngb significantly increased bcl-2 expression and SOD level, decreased bax expression, attenuated the release of cytochrome c from mitochondria to the cytosol fraction, and reduced the activity of caspase-3 and MDA level after SCI. These findings suggest, that overexpressing Ngb can significantly improve the recovery of locomotor function. This neuroprotective effect may be associated with the inhibition of neural apoptosis via the mitochondrial pathway.

A sandwich-type DNA biosensor based on electrochemical co-reduction synthesis of graphene-three dimensional nanostructure gold nanocomposite films.

A novel electrochemical DNA biosensor based on graphene-three dimensional nanostructure gold nanocomposite modified glassy carbon electrode (G-3D Au/GCE) was fabricated for detection of survivin gene which was correlated with osteosarcoma. The G-3D Au film was prepared with one-step electrochemical coreduction with graphite oxide and HAuCl4 at cathodic potentials. The active surface area of G-3D Au/GCE was 2.629cm(2), which was about 3.8 times compared to that of a Au-coated GCE under the same experimental conditions, and 8.8 times compared to a planar gold electrode with a similar geometric area. The resultant nanocomposites with high conductivity, electrocatalysis and biocompatibility were characterized by scanning electron microscopy (SEM), cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). A "sandwich-type" detection strategy was employed in this electrochemical DNA biosensor and the response of this DNA biosensor was measured by CV and amperometric current-time curve detection. Under optimum conditions, there was a good linear relationship between the current signal and the logarithmic function of complementary DNA concentration in a range of 50-5000fM with a detection limit of 3.4fM. This new biosensor exhibited a fast amperometric response, high sensitivity and selectivity and has been used in a polymerase chain reaction assay of real-life sample with a satisfactory result.