VEGF-C/VEGFR-3/iNOS Signaling in Osteosarcoma MG63 Cells Mediates Stimulatory Effects on Human Umbilical Vein Endothelial Cell Proliferation.

Objective To assess the effects of vascular endothelial growth factor-C (VEGF-C)/vascular endothelial growth factor receptor-3 (VEGFR-3) signaling on nitric oxide (NO) production and inducible nitric oxide synthase (iNOS) expression in human osteosarcoma MG63 cells and the subsequent impact on the proliferation of human umbilical vein endothelial cells (HUVECs). MethodsMG63 cells were treated with VEGF-C alone (VEGF-C group), VEGF-C + iNOS inhibitor aminoguanidine (AG; AG group), and VEGF-C + VEGFR-3 inhibitor MAZ51 (MAZ51 group); untreated MG63 cells were used as controls. NO production was evaluated by a colorimetric method involving nitrate reductase. Meanwhile, mRNA and protein levels of iNOS were examined by reverse transcription polymerase chain reaction (RT-PCR) and Western blot. To explore the effect of VEGF-C/VEGFR-3/iNOS signaling of MG63 cells on proliferation of HUVECs, we set up six groups: HUVECs, HUVECs+MG63, HUVECs+VEGF-C, HUVECs+MG63+VEGF-C, HUVECs+MG63+VEGF-C+AG, and HUVECs+MG63+VEGF-C+MAZ51 groups. The proliferation of HUVEC cells was assessed by Cell Counting Kit-8 (CCK-8), 5-ethynyl-2'-deoxyuridine (EdU) incorporation assay, and proliferating cell nuclear antigen (PCNA) expression quantitation. ResultsVEGF-C treatment enhanced iNOS expression at both gene and protein levels (mRNA: LSD-t=4.152, P<0.01; protein: LSD-t=3.486, P<0.01) and increased NO release of MG63 cells (LSD-t=3.774, P<0.01); treatment with either AG or MAZ51 decreased these effects (mRNA: LSD-t=9.183, P<0.001; LSD-t=8.639, P<0.001; protein: LSD-t=5.170, P<0.001; LSD-t=7.255, P<0.001; NO production:LSD-t=10.326, P<0.001; LSD-t=10.540, P<0.001). Interestingly, co-incubation of HUVECs with MG63 cells and/or VEGF-C significantly promoted HUVEC proliferation (EdU: LSD-t=5.374, P<0.001; LSD-t=2.984, P<0.05; LSD-t=8.526, P<0.001; PCNA: LSD-t=9.267, P<0.001; LSD-t=5.515, P<0.001; LSD-t=14.873, P<0.001).The proliferation effects of HUVEC induced by MG63 cells and VEGF-C attenuated by the treatment of AG (EdU: LSD-t=10.770, P<0.001; PCNA: LSD-t=19.940, P<0.001) or MAZ51 (EdU: LSD-t=6.950, P<0.001; PCNA: LSD-t=14.001, P<0.001). ConclusionIn human osteosarcoma MG63 cells, activation of VEGFR-3 by VEGF-C promotes iNOS expression and NO production, which subsequently induces HUVEC proliferation.

[Study on the effect of hydraulic delivery vertebroplasty on vertebral morphology and bone cement diffusion].

OBJECTIVE: To evaluate the effect of different bone cement injection methods during percutaneous vertebroplasty(PVP) on vertebral morphology and cement diffusion. METHODS: The clinical data of 52 patients with single-segment osteoporotic vertebral compression fracture treated from January 2016 to December 2017 were retrospectively analyzed. The patients were divided into hydraumatic group (28 cases) and pusher group (24 cases) according to bone cement injection method during PVP. By comparing visual analogue scale(VAS), height of anterior vertebral body, compression ratio, kyphosis angle before and after operation and analyzing filling ratio of bone cement in the first 1/3, median line and back 1/3 of the vertebral body in lateral X-rays and the conditions of bone cement diffusion in AP X-rays were to evaluate the effect of different bone cement injection methods on vertebral morphology and cement diffusion. RESULTS: Postoperative VAS was obviously improved in all patients and hydraumatic group was better than pusher group(P<0.05). Postoperative height of anterior vertebral body, compression ratio and kyphosis angle obviously restored in all patients while there was no significant difference between two groups(P>0.05). There was no significant difference in filling ratio of bone cement in the first 1/3 and median line of the vertebral body by lateral X-ray films between two groups(P>0.05), but in the back 1/3 of the vertebral body filling ratio of bone cement in hydraumatic group was better than in pusher group(P<0.05). The distribution of bone cement from AP X-ray films were more significant in hydraumatic group(P<0.05). CONCLUSIONS: Hydraulic delivery vertebroplasty (HDVP) has better clinical efficacy and it can guarantee sufficient distribution of bone cement into the fractured vertebra and preferably restore the morphology of vertebral body, which is worthy of clinical application.

Expression of b-FGF and endostatin and their clinical significance in human osteosarcoma.

OBJECTIVE: To investigate the expression and the clinical significance of basic fibroblast growth factor (b-FGF) and endostatin in osteosarcoma. METHODS: From January 2003 to December 2005, expression of b-FGF, endostatin and CD34 were detected in 30 osteosarcoma and 30 osteochondroma tissue specimens by the immunohistochemical Elivision method. All data were post-processed with SPSS 13.0 software and prepared for investigation and analysis of these expressions and the relationships between the parameters. RESULTS: (i) The rates of expression of b-FGF, endostatin and CD34 protein in osteosarcoma were 76.7%, 93.3%, and 96.7%, respectively, and in osteochondroma 43.3%, 40.0% and 16.7%, respectively. Each of the three expressions showed obvious differences between the osteosarcoma and the osteochondroma group. (ii) In the osteosarcoma group, expression of endostatin was positively correlated with that of CD34 (P < 0.05, γs = 0.528), and expression of endostatin in poorly differentiated osteosarcoma was much greater than that in highly differentiated osteosarcoma (P= 0.004). Expression of endostatin correlated with osteosarcoma metastasis (P= 0.036). (iii) There was no correlation between b-FGF and endostatin expression rates (P= 0.182) in the osteosarcoma group. CONCLUSION: Angiogenesis is the basis of tumor metastasis, as well as being an important factor in tumor growth. Expression of endostatin could be adopted as a parameter for the diagnosis of postoperative metastases and for assessing prognosis, and could act as an adjuvant indicator in the grading of osteosarcoma.

[Operative strategy of atlantoaxial instability].

OBJECTIVE: To evaluate the operative strategy and therapeutic outcomes of the atlantoaxial instability. METHODS: Clinical data of 29 patients with atlantoaxial instability were retrospectively analyzed. There were 27 males and 2 females. The mean age was 33 years old with a range from 18 to 54 years. There were fracture of anterior arch of atlas accompanied with ligamentum transversum rupture in 5 cases, odontiod fracture in 7 cases, Hangman fracture in 6 cases, dysplasia of atlas and axis in 10 cases, ankylosing spondylitis in 1 case. The clinical and imaging manifestation of atlantoaxial instability were found in all patients. The symptoms and physical signs of superior cervical spinal cord disease or cervical spinal injury were found in 18 cases. The patients were treated with simple modified Magerl method (7 cases), cannutated screw fixation(6 cases), resection of C2,3 disc throuth the anterior approach and fusion with Zephir titanium plate (4 cases), percutaneous pedicle screw fixation of C2 (2 cases), release and reduction through anterior oropharynx (LRAO) combined with modified Magerl method (4 cases), LRAO and atlas lateral mass screw and plate fixation through posterior approach (3 cases), cervical occipital fusion through C2 pedicle (3 cases). RESULTS: All patients were followed up with an average time of 17.2 months ranging from 11 to 38 months. All patients obtained anatomical reduction and bone healing. Using Odom standard to evaluation for 18 cases with spinal injury before operation, the results were excellent in 9 cases,good in 7,fair in 2. No injury of vertebral artery, nerve root, spinal cord, infection of incisional wound, breaking or loosening of internal fixatir were found in the study. CONCLUSION: Identifying the causes of atlantoaxial instability, rational plan of operation can get satisfactory clinical results.