Objective To observe the effect of silencing the expression of ADM using RNA interfering technique on the expression ofβ-catenin and terminal differentiation of osteosarcoma cells.Methods After the intervention of 0 h,48 h and 72 h by ADM siRNA,we observed the change of distribution ofβ-catenin in F5M2 using immunocytochemistry staining.The expression levels of total and phosphorylatedβ-catenin and GSK3βwere detected by Western blot after the intervention by ADM siRNA.The F5 M2 cells treated with ADM siRNA were subjected to HE staining,alkaline phosphatase (ALP)assay and immunocytochemistry staining to investigate the biological effects of ADM siRNA on the morphology and terminal differentiation of F5M2 cells.Results After the intervention of 0 h,48 h and 72 h by ADM siRNA,the distribution ofβ-catenin was transferred from the cytoplasm to the nucleus.ADM siRNA downregulated the expression level of P-β-catenin and upregulated P-GSK3βdetected by Western blot.HE staining revealed that the configuration of F5M2 had undergone restorational changes similar to those of normal cells after ADM siRNA treatment.ALP assay and immunocytochemistry staining showed that the expression of the earlier and later molecular biomarkers of terminal differentiation,including ALP and osteocalcin were strongly positive.Conclusion Silencing the expression of ADM can activateβ-catenin to transfer to the nucleus from the cytoplasm,and induce osteosarcoma cells to make terminal differentiation through activatingβ-catenin signaling pathway.

Bilirubin is a metabolic end product degraded under the action of heme oxygenase.It is an endogenous antioxidant and has the function of anti-atherosclerosis, clearing free radicals and protecting the tissues and organs of the body.This article reviews the metabolism and biological characteristics of bilirubin, and the correlations between bilirubin and vascular risk factors, as well as between bilirubin and ischemic stroke.

Objective To observe the clinical effect and safety of percutaneous kyphoplasty (PKP) on osteoporosis vertebral compres-sion fractures (OVCF) in elderly. Methods From January, 2014 to May, 2015, 88 patients with OVCF treated with PKP were retrospective studied. Visual Analogue Scale (VAS) and Oswestry Dysfunction Index (ODI) were used to evaluate the pain and quality of life, and the ky-photic angle (Cobb's angle) and anterior vertebral body height were recorded before operation, one week after operation and the last fol-low-up. The cement leakage and its effect on nerve function were recorded. Results Bone cement leakage was found in eleven patients, in which two in pedicle, three in venous leakage, three in intervertebral disc and three in vertebral posterior. Nerve root symptoms appeared in one patient, who was treated with decompression. The scores of VAS and ODI, and the Cobb's angel were significantly lower (t>29.518, P<0.001), and the vertebral body height was significantly higher (t>35.173, P<0.001) one week after operation and the last follow-up than be-fore operation. Conclusion PKP is safe and effective for OVCF in elderly. It can efficiently reduce the pain, correct kyphotic deformity and improve the quality of life.

Objective To establish the animal model of cervical pain induced by formalin injection and explore the effect of cervical pain on cervical vertigo.Methods Formalin of 40 mL/L was injected near the nerve root of left side of neck to establish the model of cervical pain in rabbits.A total of 30 rabbits were divided randomly into two groups.Formalin-induced pain group(15 rabbits): indexes related to the vertebral artery were observed respectively before 2 mL of formalin injection and after it at 5,10 and 30 minutes.Lidocaine-blocking pain group(15 rabbits): pain induced by formalin was blocked by lidocaine of 10 g/L after 15-minute drug administration and the same indexes were observed at the same time and position of the neck.Results In formalin-induced pain group,vertebral artery systolic velocity,diastolic velocity and pulsatility indexes were decreased whereas resistance index was increased(P

Objective To evaluate the value of MRI diffusion tensor imaging (DTI) on diagnosis of traumatic brain injury(TBI). Methods Twenty two patients with TBI 1 to 7 days post-injury and 14 healthy controls were studied with DTI and conventional MRI. The fractional anisotropy (FA) was quantified from different regions of interest (ROI) including the genu of corpus callosum (CC), the splenium of CC, the genu of internal capsule (IC) and the posterior limb of IC bilaterally. The FA value of the same ROI was compared between TBI group and control group, and FA value of the lesion side was compared with the mirror healthy side in TBI group. Correlations between the FA and Glasgow coma scale (GCS) in TBI patients were analyzed with Pearson linear correlation. Results Compared with control group, the FA value decreased significantly in each ROI (P<0.01). Compared with the healthy side, FA value was also significantly lower in lesion side in TBI group (P<0.01). The FA value in all the sites were not correlated with GCS in TBI group (P>0.01).Conclusion DTI is sensitive for detecting the acute traumatic injury of white matter and evaluate the degree of injury. It offers the possibility to diagnose TBI earlier and accurately.

ABSTRACT:Objective To study the expressions of adrenomedullin (ADM)andβ-catenin in human osteosarcoma of different differentiation grading and their clinical significance.Methods We chose 33 cases of human osteosarcoma with different pathological grading (14 cases of high differentiation grading and 1 9 cases of low grading)and 10 cases of human osteochondroma to detect the expressions of ADM protein,total (T)β-catenin and phosphorylated (P)β-catenin using immunohistochemical method. Results The expression of ADM was detected in the cytoplasm.In osteosarcomas of higher and lower differentiation grading,the positive rate of ADM was 57.1% (8/14)and 100% (1 9/1 9),respectively,with significant differences (P =0.003<0.05).The expression of T-β-catenin was observed in both the cytoplasm and the nucleus in higher differentiation grading and in only cytoplasm in lower grading.In osteosarcomas of higher and lower differentiation grading,the positive rate of T-β-catenin was 85.7%(12/14)and 94.7% (18/1 9),with no significant difference (P =0.561>0.05).The expression of P-β-catenin was detected in the cytoplasm.In osteosarcomas of higher and lower differentiation grading,the positive rate of P-β-catenin was 35.7% (5/14)and 78.9% (1 5/1 9),with a significant difference (P =0.029 <0.05).There was a significant correlation between ADM and P-β-catenin (r =0.424,P =0.025<0.05),but no significant correlation between ADM and T-β-catenin (P = 0.078 > 0.05 )or between T-β-catenin and P-β-catenin (P = 0.052 > 0.05 ).Conclusion The expressions of ADM and P-β-catenin proteins and the different distributions of expression of T-β-catenin protein in cytoplasm and/or nucleus are important indicators for judging the pathological grading of osteo-sarcoma.

Objective To study the anti-tumor activity of quercetin in NB4 leukemia cells and the roles of PI3K/Akt,bcl-2,and Bax on the quercetin-induced apoptosis,and to investigate the potential underlying mechanism.Methods MTT assay was used to monitor cell proliferation,Hoechst 33258 fluorescent staining and flow cytometry were employed to detect apoptosis in NB4 cells.Western blot was used to detect the expression changes of related proteins in quercetin treated NB4 cells.Confocal laser microscopy was used to test the distributional variation of Akt between cytoplasm and nucleus.Results Quercetin significantly inhibited the NB4 cell proliferation in a dose-and time-dependent manner (20-160 μ mol/L).In addition,treated by 20,40 80 μmol/L quercetin,the rates of apoptosis were (9.25±0.11) ％,(20.83±2.10) ％and (41.43±2.90) ％,there were statistical difference compared with blank cells (t were 4.14,6.56 and 7.02,all P ＜ 0.05).This was concentration dependent and accompanied by morphological changes characteristic of apoptosis.Further,quercetin induced a G~M arrest,which might account for its cytotoxic effects.Quercetin decreased PI3k/Akt expression and caused an inhibition of the anti-apoptotic protein bcl-2,while increasing the expression of Bax.Quercetin had no effects on total Akt,but it promoted Akt translocation from cell nucleus to the cytoplasm (F =15.12,P ＜ 0.05).Conclusion Quercetin induces the leukemia NB4 cell apoptosis by affecting multiple signal pathways and plays a strong anti-leukemia effect.In addition,our results suggest that PI3K/Akt pathway could be a novel target for the leukemia chemotherapy.

BACKGROUND: There still was not any report about inducing stem cells into matured cells to form products in vitro.OBJECTIVE: To induce CD34+ cells of umbilical cord blood to differentiate into mature megakaryocytes, and to investigate the mechanism of production of platelets.DESIGN, TIME AND SETTING: This cytology in vitro study was conducted at the Central Laboratory of Xiangya Hospital and Xiangya Third Hospital from 2004 to 2006. MATERIALS: Umbilical cord was collected from healthy full-term pregnant puerperants at the Xiangya Hospital.METHODS: The CD34+ cells were isolated from umbilical cord blood by magnetic activated cell sorting (MACS) and then cultured in 24-well culture plate at 5x107/L in StemPro-34 serum-free medium, supplemented with L-glutamine, saturated human transferrin, CaCl2, insulin, deionized bovine serum albumin and recombinant human thrombopoietin at 37℃, under 0.05 volume fraction CO2 saturated humidity to be differentiated into megakaryocytes for 14-21 days. Cell medium was absorbed, and centrifuged to obtain supernatant. Samples were centrifuged again, and then supernatant was removed. The remaining was platelet-like particles in cell culture plate. Platelet was isolated from normal platelet-rich plasma.MAIN OUTCOME MEASURES: The following parameters were measured: morphological changes in cultured cells and platelet-like particles in supematant; results of immunohistochemistry; observation results under a microscope; platelet aggregation; CD41 expression.RESULTS: At day 10, silk-like substances were found in megakaryocyte culture medium, with the presence of platelet-sized particles. The production of platelet-sized particles reached a peal at day 16. Cultured cells were strongly positively for platelet-specific antigen GP Ⅱb Ⅲa. Under the optical microscope, mature megakaryocytes were detected, with the presence of some immature megakaryocytes, and platelet-sized particles were found surrounding megakaryocytes. Under the electron microscope, a majority of mature megakaryocytes and a few apoptotic megakaryocytes were detected, and platelet-sized particles in the supernatant had the same size and structure with the platelet in the platelet-rich plasma. Some platelet surfaces were smooth or irregular. Platelet-sized particles in the supematant aggregated in response to thrombin as platelets in normal platelet-rich plasma. Flow cytometry demonstrated that the cultured platelets had the same high expression rate of CD41 as the platelets from platelet rich plasma.CONCLUSION: Umbilical cord blood CD34+ cells can be induced to differentiate into pudfied and mature megakaryocytes and platelets in vitro.

Drug metabolism studies, including in vivo and in vitro metabolism studies, are significant in the design of candidate compounds and screening of lead compounds at drug discovery/development stages. Compared with in vivo metabolism studies, in vitro metabolism studies have the advantages of rapidity, simplicity, without consumption of large amounts of samples and animals. Moreover, it is convenient for researchers to observe the selective interaction between compound and target. Therefore, in vitro metabolism studies are appropriate for high throughput screening of compounds which are lack of metabolism information and have been widely used during drug discovery stages. This article briefly introduced the application of in vitro drug metabolism studies based on the metabolic stability, reaction phenotyping and metabolic drug-drug interactions, aiming to raise valuable evaluation strategies for innovative drug discovery in China.

Objective To identify the electrophysiological charateristics and cause of ridge gap related reentry after MI ablation in atrial ifbrillation patients. Methods Activation and entrainment mapping was performed in 82 redo cases for OAT recurrence in whom MI was ablated during the index produre. Once ridge gap related reentry was conifrmed, detailed mapping was performed in MI and ridge region. In addition, in 36 cases undergoing MI ablation and fulfilling criterion for bidirectional block, differential pacing was repeated at the ridge to identify a ridge gap. Results Out of 82 redo cases for OAT recurrence in whom MI was ablated during the index produre, 7 (8.5%) was found to be ridge gap related reentry. TCL was (247.9±19.2) ms, and the left atrial endocardial activation time was (145.4±17.7) ms, accounting for (58.5±3.2)%of TCL. However, wide double potential was recorded along the previous ablated MI line where PPI was (34.3±6.6) ms longer than TCL, while PPI was signiifcantly shorter at the ridge[PPI-TCL (11.4±3.9) ms, P<0.001]. Tachycardia was terminated at the ridge in 6 cases and at the corresponding site in coronary sinus in 1 case. No recurrence was found during follow-up for (11.1±4.5) months. In addition, in 36 patients undergoing MI ablation in whom criterion of bi-directional block was fuliflled, conduction gap located at the ridge was found in 5 (13.9%) cases. Conclusions MI ridge gap related reentry is a distinctive OAT, in which the ridge was used as the critical isthmus, whereas the previous ablated MI line is not part of the reentry. MI pseudo-block due to the ridge gap may lead to this type of recurrent tachycardia.