Predictable Risk Factors of Spontaneous Venous Thromboembolism in Patients Undergoing Spine Surgery.

OBJECTIVE: The purpose of this study was to conduct a meta-analysis to identify the risk factors for formation of venous thromboembolism (VTE) in patients after spine surgery. METHODS: This study retrieved potential academic articles on the related factors for VTE formation in patients after spine surgery from MEDLINE, PubMed, EMBASE, and the Cochrane Library. The reference articles for the identified studies were carefully reviewed to ensure that all available documents were represented in the study. RESULTS: A total of 21 articles (20 retrospective studies and 1 prospective study) involving 2,870,105 patients were identified in the analysis, including 7829 patients who presented with VTE after spine surgery; the incidence of VTE was 0.273%. Our meta-analysis showed that compared with patients who did not have VTE after spine surgery, there was significantly more blood loss (weighted mean difference [WMD], 93.295; 95% confidence interval [CI], 60.521-126.069; P < 0.001), higher age (WMD, 6.011; 95% CI, 3.647-9.376; P < 0.001), thoracolumbar surgery (odds ratio [OR], 0.233; 95% CI, 0.198-0.274; P < 0.001), and longer duration of surgery (WMD, 45.672; 95% CI, 10.433 to -80.911; P = 0.011) among the patients with VTE. Patients with a history of hypertension (OR, 1.785; 95% CI, 1.516-2.103; P < 0.001), diabetes (OR, 1.535; 95% CI, 1.286-1.832; P < 0.001), and preoperative walking disability (OR, 4.882; 95% CI, 2.044-11.663; P < 0.001) showed a significantly higher rate of VTE after spine surgery. However, no significant differences were found in gender (P = 0.289), fusion surgery (P = 0.979), body mass index (P = 0.157), history of heart disease (P = 0.397), and level of D-dimer (P = 0.220). CONCLUSIONS: A higher rate of postoperative VTE is closely associated with the elderly, longer duration of surgery, thoracolumbar surgery, greater blood loss, and patients with a history of hypertension, preoperative walking disability, or diabetes after spinal surgery; these risk factors should be guarded against.

[Relationship between Golgi apparatus and cell migration direction in vivo and in vitro].

OBJECTIVE: To explore the relationship between Golgi apparatus and the direction of tumor cell migration in vivo and in vitro. METHODS: Cell migration assays were conducted with rat C6 glioma cells, human U251 and SNB19 glioma cells respectively. Then immunofluorescence was used to detect the position of Golgi apparatus in migrating cells. The percentage of cells with Golgi apparatus facing towards wound edge was calculated. Cell pseudopodium was stained with TRITC-phalloidin and the relationship between Golgi apparatus and pseudopodium detected. Immunohistochemistry was used to reveal the Golgi apparatus in tumor tissue samples. And the percentage of cells with Golgi apparatus facing opposite to the necrotic zones was calculated. RESULTS: In cells located at wound edge, the Golgi apparatus was found facing towards the wound in the vast majority of cells (C6 83% ± 6%, U251 80% ± 7%, SNB19 82% ± 6%). In U251 and SNB19 cells, the golgi apparatus was located in the same direction with cellular pseudopodium. Immunohistochemical staining showed that in cells located around the necrotic zone, the Golgi apparatus faced opposite to the necrotic zones in most cells (rat tissue samples 80% ± 7%, human tissue samples 82% ± 6%). CONCLUSIONS: The Golgi apparatus is closely correlated with cell migration and it may be considered as a direction indicator of cell migration. And it provides an important index for the study of tumor cell invasion both in vivo and in vitro.

Rac1+ cells distributed in accordance with CD 133+ cells in glioblastomas and the elevated invasiveness of CD 133+ glioma cells with higher Rac1 activity.

BACKGROUND: Recent studies have suggested that cancer stem cells are one of the major causes for tumor recurrence due to their resistance to radiotherapy and chemotherapy. Although the highly invasive nature of glioblastoma (GBM) cells is also implicated in the failure of current therapies, it is not clear how glioma stem cells (GSCs) are involved in invasiveness. Rac1 activity is necessary for inducing reorganization of actin cytoskeleton and cell movement. In this study, we aimed to investigate the distribution characteristics of CD133+ cells and Rac1+ cells in GBM as well as Rac1 activity in CD133+ GBM cells, and analyze the migration and invasion potential of these cells. METHODS: A series of 21 patients with GBM were admitted consecutively and received tumor resection in Tianjin Medical University General Hospital during the first half of the year 2011. Tissue specimens were collected both from the peripheral and the central parts for each tumor under magnetic resonance imaging (MRI) navigation guidance. Immunohistochemical staining was used to detect the CD133+ cells and Rac1+ cells distribution in GBM specimens. Double-labeling immunofluorescence was further used to analyze CD133 and Rac1 co-expression and the relationship between CD133+ cells distribution and Rac1 expression. Serum-free medium culture and magnetic sorting were used to isolate CD133+ cells from U87 cell line. Rac1 activation assay was conducted to assess the activation of Rac1 in CD133+ and CD133 - U87 cells. The migration and invasive ability of CD133+ and CD133 - U87 cells were determined by cell migration and invasion assays in vitro. Student's t-test and one-way analysis of variance (ANOVA) test were used to determine statistical significance in this study. RESULTS: In the central parts of GBMs, CD133+ cells were found to cluster around necrosis and occasionally cluster around the vessels under the microscope by immunohistological staining. In the peripheral parts of the tumors, CD133+ cells were lined up along the basement membrane of the vessels and myelinated nerve fibers. Rac1 expression was high and diffused in the central parts of the GBMs, and the Rac1+ cells were distributed basically in accordance with CD133+ cells both in the central and peripheral parts of GBMs. In double-labeling immunofluorescence, Rac1 was expressed in (83.14 ± 4.23)% of CD133+ cells, and CD133 and Rac1 co-expressed cells were located around the vessels in GBMs. Significantly higher amounts of Rac1-GTP were expressed in the CD133+ cells (0.378 ± 0.007), compared to CD133- cells (0.195 ± 0.004) (t = 27.81; P < 0.05). CD133+ cells had stronger ability to migrate (74.34 ± 2.40 vs. 38.72 ± 2.60, t = 42.71, P < 0.005) and invade (52.00 ± 2.28 vs. 31.26 ± 1.82, t = 30.76, P < 0.005), compared to their counterpart CD133- cells in transwell cell migration/invasion assay. CONCLUSIONS: These data suggest that CD133+ GBM cells highly express Rac1 and have greater potential to migrate and invade through activated Rac1-GTP. The accordance of distribution between Rac1+ cells and CD133+ cells in GBMs implies that Rac1 might be an inhibited target to prevent invasion and migration and to avoid malignant glioma recurrence.

[Glioma stem cells enhanced angiogenesis and its relationship with microvessel].

OBJECTIVES: To dynamically observe how glioma stem cells promote the tumor formation and angiogenesis, and to study the correlation between the distribution of glioma stem cells and microvessels within different growth stages of subcutaneous tumor. METHODS: Stem cell medium culture and magnetic activated cell sorting were carried out to obtain CD133+ cells from C6 rat glioma cell line. Sprague Dawley (SD) rat ears model were established to observe glioma stem cells promoting blood vessel formation. Subcutaneous glioma model of C6 and immunohistochemical staining of hypoxia inducible factor-1α (HIF-1α) and CD133 were used to investigate the relationship between distribution of glioma stem cells and microvessels. Expressions of CD133 protein in each stage of the subcutaneous tumor were detected by Western blot. RESULTS: Isolation and identification of glioma stem cells deprived from C6 glioma cell line successfully, the establishment of ears model showed real-time dynamic observation of CD133+ cells involved in angiogenesis and tumor formation. SD rat model of subcutaneous glioma showed the initial of tumor formation, CD133+ cells scattered. With tumor growth, CD133+ cells began to tend to capillaries, in late distributed clusters in perivascular. Meanwhile as tumor growth, CD133 protein expression was gradually increased: the values of Western blot analysis of CD133 expression on 6, 9, 12, 15, 20 d were 0.208±0.004, 0.282±0.003, 0.360±0.004, 0.564±0.135, 0.756±0.007, the differences were significant between different groups (F=2601.681, P<0.01). At a high magnification, the CD133 scores with immunohistochemical staining on 6, 9, 12, 15 d were 0.8±0.4, 2.4±0.5, 4.0 ± 0.7, 6.0±0.7; HIF-1α scores were 0.8±0.4, 2.8±0.8, 5.0±0.7, 6.8±0.4. By Spearman rank correlation analysis found that the relationship between CD133 and HIF-1α expression was positively correlated (r=0.921, P<0.01). CONCLUSIONS: Glioma stem cells promote angiogenesis more than non-stem cells; HIF-1α and its downstream gene product might mediate the distribution of glioma stem cells around the perivascular.

[Role of Rac1 in the SDF-1-induced migration and invasion of human glioma cell line U251].

OBJECTIVE: To explore the role of Rac1 in the SDF-1-induced migration and invasion of glioma cells with a specific Rac1 inhibitor. METHODS: Human glioma cell lines U251 treated with SDF-1 or/and specific Rac1 inhibitor were used. The migration and invasion capacities of cells in 2D cell migration/3D invasion assay were assessed. Western blot was employed to detect the levels of Rac1 and GAPDH in cell lysates and the Rac1 activity measured by Rac1 activation assays. Immunofluorescence was used to identify the expression and intracellular location of Rac1 in U251 cells. RESULTS: SDF-1 significantly increased the migration and invasion capacities of U251 cells (P < 0.05). The stimulation of SDF-1 boosted the activity of Rac1 versus the unstimulated cells (P < 0.05). And Rac1 was recruited to protruding edge in SDF-1-stimulated cells. Inhibition of Rac1 with specific Rac1 inhibitor decreased the migration and invasion capacities of SDF-1-induced U251 cells (P < 0.05). In comparison with the SDF-1 treated group, the activity of Rac1 significantly decreased (P < 0.05) and the recruitment of Rac1 to protruding edge significantly decreased in the NSC23766 pre-treated group. CONCLUSIONS: This study provides novel evidence that Rac1 modulates the SDF-1-induced migration and invasion of glioma cells. It suggests that the inhibition of Rac1 activation may be a new therapeutic target for glioma.

Enhanced invasion in vitro and the distribution patterns in vivo of CD133+ glioma stem cells.

BACKGROUND: Recent studies have suggested that cancer stem cells cause tumor recurrence based on their resistance to radiotherapy and chemotherapy. Although the highly invasive nature of glioblastoma cells is also implicated in the failure of current therapies, it is not clear whether cancer stem cells are involved in invasiveness. This study aimed to assess invasive ability of glioma stem cells (GSCs) derived from C6 glioma cell line and the distribution patterns of GSCs in Sprague-Dawley (SD) rat brain tumor. METHODS: Serum-free medium culture and magnetic isolation were used to gain purely CD133(+) GSCs. The invasive ability of CD133(+) and CD133(-) C6 cells were determined using matrigel invasion assay. Immunohistochemical staining for stem cell markers and luxol fast blue staining for white matter tracts were performed to show the distribution patterns of GSCs in brain tumor of rats and the relationship among GSCs, vessels, and white matter tracts. The results of matrigel invasion assay were estimated using the Student's t test and the analysis of Western blotting was performed using the one-way analysis of variance (ANOVA) test. RESULTS: CD133(+) GSCs (number: 85.3 ± 4.0) were significantly more invasive in vitro than matched CD133(-) cells (number: 25.9 ± 3.1) (t = 14.5, P < 0.005). GSCs invaded into the brain diffusely and located in perivascular niche of tumor-brain interface or resided within perivascular niche next to white fiber tracts. The polarity of glioma cells containing GSCs was parallel to the white matter tracts. CONCLUSIONS: Our data suggest that CD133(+) GSCs exhibit more aggressive invasion in vitro and GSCs in vivo probably disseminate along the long axis of blood vessels and transit through the white matter tracts. The therapies targeting GSCs invasion combined with traditional glioblastoma multiforme therapeutic paradigms might be a new approach for avoiding malignant glioma recurrence.

[Correlation between the distribution of CD133-positive cells and the proliferation of microvessels in glioblastoma multiforme].

OBJECTIVE: To investigate the expression of CD133 and CD34 in different parts of glioblastoma and its margin and explore the invasive path of glioma stem cells within the tumor and surrounding tissue. METHODS: The surgical specimens were collected from the core of mass, junctional zones between tumor and peritumoral edematous areas and edematous areas in 52 patients with glioblastoma. Immunohistochemical cell staining and Western blot were employed to evaluate the expression of CD133 in different specimens while immunohistochemistry was used to detect the CD34-microvessel postforming. A correlation analysis was performed between them. RESULTS: The expression of CD133 was not detected in the control groups while the scores were 7.3 ± 1.4, 5.2 ± 1.1, 2.7 ± 1.0 in junctional zones, tumor cores and edematous areas with immunohistochemistry and 0.79 ± 0.03, 0.38 ± 0.01, 0.20 ± 0.04 with Western blot respectively. There were significant differences between different groups (P < 0.05). Under light microscope, the CD133-positive cells frequently forming perivascular pseudorosettes were dense in junctional zones and mostly clustered near the microvessels in tumor cores and scattered in edematous areas. At a high magnification (200×), the CD34-MVD (/HP) values were 31.3 ± 4.0, 21.8 ± 2.6, 15.3 ± 2.4, 4.7 ± 1.5 respectively in junctional zones, tumor cores, edematous areas and control tissues. Significant differences were also found in these groups (P < 0.05). The expression level of CD133-positive cell was positively correlated with the distribution of CD34-microvessels (r = 0.948, P < 0.05). CONCLUSION: Glioma stem cells tend to assemble in the junctional zones where the microvessels are enriched. There is probably an intimate nourishing relationship with the microvessels. The distribution of glioma stem cells may be related with the invasiveness within glioblastoma.

Expression and Distribution Characteristics of Human Ortholog of Mammalian Enabled (hMena) in Glioma.

OBJECTIVE: To investigate the utility of hMena, a family of enabled/vasodilator-stimulated phosphoprotein (Ena/VASP), we sought to characterize the expression profile and distribution characteristics of hMena in a large panel of glioma samples and determine whether hMena expression levels might correlate with the pathological grade of glioma. METHODS: Sixty-five specimens of glioma with different pathological grades and five control brain tissues were collected. In 6 of the 21 glioblastoma patients, multi-specimens were obtained respectively from the main tumor mass, the junction zone between the tumor and the normal tissue, and adjacent brain tissue 1.5 cm away from the tumor boundary under assistance of neuronavigation system during the operation. Immunohistochemistry was used to detect the expression and distribution characteristics of hMena. hMena expression was analyzed by Western blot in 20 specimens. RESULTS: The hMena expression was negative in control brain tissue but positive in different grades of glioma. The expression rate of hMena was positively correlated with the increasing grade of the World Health Orgnization (WHO) classification (r(s)=0.682, P=0.000). hMena was located in cytoplasm. Positive cells only distributed around the vessels within the tumor mass in low grade glioma, while in high grade glioma, these cells were able to be detected not only in the tumor but also in the boundary zone and adjacent brain parenchyma. In the tumor mass, hMena expressed highly and diffusedly. In the junction zone, hMena positive cells formed radiolitic pattern around the vessels. In adjacent brain parenchyma, single positive cell was scattered. hMena expression was markedly elevated in Grade III and IV glioma compared with Grade II and I. CONCLUSION: Our data suggested that the expression of hMena is closely related to malignant grade of glioma. hMena can label the migrating cells, and indicate the migrating path of glioma cells from the tumor to adjacent tissue along with the vascular basement membranes and tracts of white matter.