Objective To observe the effect of glucocorticoid on osteoblast-like cell apoptosis and the cell cycle. Methods Osteoblast-like cells OS-732 at the density of 1?105/mL were cultured in vitro with dexamethasone (DMX) at the concentrations of 10-7,10-8 and 10-9 mmol/L. The apoptosis of OS-732 was detected by flow cytometry after the culturing for 24 and 48 hours,and then DNA content was detected and the cell cycle was analysed. Results At the time point of the 24th hour,DNA content at G2/M phase of OS-732 cell cycle was increased obviously in DMX1(10-7 mmol/L)group (P

Objective:A study was conducted to investigate the effect of Prunella vulgaris sulfated polysaccharide (PVSP) on the expression of angiogenic growth factors (bFGF, VEGF, and IL-8) and angiogenesis in hepatocellular carcinoma. Methods:ELISA as-say was used to observe the effects of PVSP and the negative control drug Lentinan (LNT, non-sulfate radical drug) on secretions of the angiogenic growth factors, namely, bFGF, VEGF, and IL-8, in HepG2 cells in vitro. In an in vivo experiment, the microvessel density in hepatocellular carcinoma tissue sections treated with PVSP and LNT was calculated, analyzed, and compared with the microvessel den-sity in the phosphate-buffered saline (PBS) control. Results:Compared with the PBS control group, PVSP at 200μg/mL inhibited bF-GF secretion (P<0.01), whereas LNT failed to affect bFGF secretion. Neither PVSP nor LNT affected the secretions of VEGF and IL-8. In vivo results showed that PVSP at 200 mg/kg reduced the microvessel density in tumor tissue sections (P=0.03), whereas LNT at 10 mg/kg failed to affect microvessel density. Conclusion:Inhibition of bFGF secretion is a probable mechanism underlying the preven-tive effect of PVSP on hepatocellular carcinoma angiogenesis.

BACKGROUND: Bone marrow-derived mononuclear cells (BM-MNCs) hold the potential of differentiating into osteoclasts. Polygonatum sibiricum polysaccharide (PSP) may inhibit the differentiation of BM-MNCs into osteoclasts and it is expected to become a new drug for the treatment of osteoporosis. OBJECTIVE: To investigate the effect of PSP on the differentiation of mouse BM-MNCs into osteoclasts induced by receptor activator of nuclear factor kappa-B ligand (RANKL) and bone resorption in vivo. METHODS: Mouse bone marrow-derived macrophages cultured in vitro, the effect of macrophage colony stimulating factor and PSP (5, 10, 20, 40, 80,160, 320, 640, 1280, 2560 mg/L) on the proliferation of mouse BM-MNCs was detected by cell counting kit-8 assay to determine the PSP concentration range; the mouse BMMs were cultured and induced in DMEM medium containing macrophage colony stimulating factor, RANKL and 5, 10, 20, 40, 80,160, 320, 640 mg/L PSP, respectively; those cultured without PSP served as control group. The morphological changes of cells were observed under an inverted microscope.; the number of osteoclasts was detected by tartrate-resistant acid phosphatase staining; the mRNA expression levels of osteoclast-related genes including tartrate-resistant acid phosphatase, matrix metalloproteinase-9, cathepsin K, and nuclear factor of activated T cells c1 were evaluated by quantitative real-time PCR. A mouse model of calvarial osteolysis induced by lipopolysaccharide was established to receive PSP intervention, and then micro CT scanning, three-dimensional reconstruction and relevants software were used for quantitative analysis of bone volume/volume percentage, trabecular number, trabecular bone spacing and thickness. The number of osteoclasts was identified by tartrate-resistant acid phosphatase staining and quantitative analysis of bone resorption area was conducted. RESULTS AND CONCLUSION: Compared with the control group, the concentration of PSP below 640 mg/L showed no significant effect on the proliferation of BMMs (P > 0.05). Different concentrations of PSP (40-640 mg/L) significantly reduced the number of osteoclasts, osteoclast differentiation and maturation, and the mRNA expression levels of tartrate-resistant acid phosphatase, matrix metalloproteinase-9, cathepsin K, and nuclear factor of activated T cells c1 TRAP, MMP-9, CtsK and NFATc1 (P < 0.05). Compared with lipopolysaccharide, PSP could effectively alleviate the lipopolysaccharide-induced calvarial osteolysis, and the bone volume/volume percentage, trabecular number, and trabecular bone spacing were significantly decreased (P < 0.05); additionally, the number of osteoclasts and the area of bone resorption were decreased significantly (P < 0.01). To conclude, PSP can inhibit the differentiation and maturation of mouse BMMs to osteoclasts and alleviate lipopolysaccharide-induced calvarial osteolysis.

BACKGROUND:Posterior lamina resection often causes loss of spinal stability, so screw rod internal fixation technology is needed to maintain the stability of lumbar spine. Finite element analysis can be used to simulate the stress distribution of the spine and internal fixation system after spinal surgery. OBJECTIVE: To build three-dimensional finite element model of spinal L1 to L3, analyze the spinal stability and stress distribution after the total laminectomy and insertion of bilateral pedicle screw using finite element method. METHODS: L1-L3 CT data could be colected from an adult healthy male volunteer. Mimics14.01, 3-matic(V6.0) and Ansys 15.0 could be used to set up the intact lumbar spine finite element model of L1-L3 (group A), the L1-L3 finite element model after L2 total laminectomy (group B), and the finite element model of L2 total laminectomy and insertion of bilateral pedicle screw (group C). We used software to simulate flexion, extension, lateral bending and axial rotation, and three kinds of models received finite element analysis. RESULTS AND CONCLUSION: (1) Based on the maximum of Von Mises under different motion states, the maximum stress was significantly lower in group A than in group B (P< 0.05). The maximum stress was significantly lower in group B than in group C (P < 0.05). (2) Based on the total deformation under different motion states, the total deformation was significantly lower in group A than in group B (P < 0.05). The total deformation was significantly lower in group C than in groups A and B (P < 0.05). (3) After the total laminectomy, vertebral body stress increased, especialy in the lamina, pedicle and joints. The range of motion of the vertebral body increased, which influenced the stability of the vertebral body. Internal fixation could decrease range of motion. Stress concentrated on the screw. Stress on the vertebral plate and pedicle decreased. The stability of vertebral body increased. Excessive stress concentrated on screw system wil increase the risk of screw breakage.

BACKGROUND:Our previous studies have found that polygonatum sibiricum polysaccharide (PSP) promotes osteogenic differentiation of bone marrow mesenchymal stem cel s (BMSCs) by Wnt/β-catenin signaling pathway, but the molecular mechanism is unclear.OBJECTIVE:To investigate the effect of PSP promoting the osteogenic differentiation via Wnt signaling pathways in BMSCs after LRP5 silencing. METHODS:LRP5 interference vectors were constructed and then transfected into C57BL/6 mouse BMSCs cultured in vitro. The transfection efficiency of cel s was calculated under fluorescence inverted microscope and the expression of LRP5 protein was detected by western blot assay. The osteogenic potential of BMSCs after LRP5-siRNA transfection was analyzed by alkaline phosphatase staining, alizarin red staining and western blot assay. Effect of PSP on the osteogenic differentiation of LIRP5-silenced mouse BMSCs was detected by real-time PCR and dual luciferase assay. RESULTS AND CONCLUSION:Compared with the control group, the mineralization ability, the mRNA expressions of Runx2 and Osterix, and the protein expression of LRP5 were significantly decreased in the LRP5-siRNA group (P<0.05). PSP could promote LRP5-siRNA transfected mouse BMSCs differentiating into osteoblasts and significantly upregulated the expressions ofβ-catenin and Osterixin, and also induced the high expression of luciferase reporter gene (TOPFlash) containing wild type TCF binding sites (P<0.05). To conclude, LRP5 plays an important role in the process of mouse BMSCs differentiating into osteoblasts. PSP can promote the osteogenic differentiation of mouse BMSCs by activating the Wnt/β-catenin signaling pathway independent on LRP5.

Objective To investigate PSP on bone microstructures,Ca,P,OPG and RANKL of osteoporotic rat model.Methods Thirty female rats randomly divided into five groups:Sham,OVX,H-,M-,L-PSP.Sham and OVX were irrigated stomachsaline;PSP solution was gavaged to other groups.After 8-week,bone microstructures of tibial metaphyseal,Ca,P,OPG and RANKL were measured.Results Body weight,Ca,P,RANKL,Tb.Sp of OVX were significantly increased compared to Sham,OPG,BV/TV,Tb.Th,Tb.N decreased.Body weight of H-,M-PSP,Ca and Tb.Sp of PSP,P and RANKL in H-PSP were decreased compared to OVX,OPG in H-,M-PSP,BV/TV,Tb.Th,Tb.N of PSP group increased.The differences were statistically significant (P ＜ 0.05).Conclusion PSP prevents osteoporosis by improving the microstructure of trabecular bone,reducing bone turnover,increasing OPG and reducing RANKL expression.

CD44+ CD29+ and CD44- CD29+ bone marrow mesenchymal stem cells (BMSCs) were isolated from bone marrow of BALB/C male donor mouse by adherence selecting in DMEM media with low glucose and 20% fetal bovine serum. Then P5 cells were injected into the liver remnants of hepatectomined female mouse. Simultaneous assays were performed on the injected liver lobe and other liver lobe for detecting the Y-chromosome by fluorescence in situ hybridization and for defecting the albumin or cytokeratin 18 (CK18) by fluorescence immunoassays. Both Y chromosome and albumin (or CK18) positive cell could be detected at 5 days and 14 days after transplantation. At 14 days after transplantation, the liver weight in transplanted mouse was higher than that in model mouse. These results suggest that BMSCs could be induced to differentiate into hepatocytes and participate in the regeneration of liver.
Animals ; Bone Marrow Cells ; cytology ; Cell Differentiation ; physiology ; Cells, Cultured ; Female ; Hepatectomy ; Hepatocytes ; cytology ; Liver Regeneration ; physiology ; Male ; Mesenchymal Stem Cell Transplantation ; Mesenchymal Stromal Cells ; cytology ; Mice ; Mice, Inbred BALB C

Objective:To evaluate the accuracy of lung ultrasound score (LUSS) in predicting emerging hypoxemia after tracheal extubation in the patients in postanesthesia care unit (PACU).Methods:A total of 333 patients of both sexes, aged 18-89 yr, of American Society of Anesthesiologist physical statusⅠ-Ⅲ, scheduled for elective abdominal surgery, were included in the study.Lung ultrasound examinations were performed before operation (T 0) and on admission to PACU (T 1), and the LUSS were recorded as LUSS 0 and LUSS 1.Arterial blood gas analysis was conducted at 20 min after tracheal extubation, and oxygenation index (PaO 2/FiO 2) were recorded.Patients were divided into 2 groups according to the oxygenation index: PaO 2/FiO 2<300 mmHg group (hypoxemia group), and PaO 2/FiO 2≥300 mmHg group (non-hypoxemia group). Multivariate logistic regression analysis and the receiver operating characteristic curve were used to evaluate the accuracy of LUSS 1 in predicting the emerging hypoxemia after extubation in the patients in PACU. Results:The incidence of emerging hypoxemia in PACU after extubation was 9.0%.Multivariate logistic regression analysis indicated that LUSS 1 and body mass index were independent risk factors for emerging hypoxemia after extubation in the patients in PACU.The area under the ROC curve for LUSS 1 was 0.873 ( P<0.001, 95%CI 0.812-0.935). The patients with LUSS 1<7 had a lower risk of hypoxemia after extubation (LR -=0.15, 95%CI 0.05-0.45), and the patients with LUSS 1>10 had a higher risk of hypoxemia after extubation (LR + =17.25, 95%CI 7.35-40.51). Conclusion:LUS can effectively predict the development of hypoxemia after tracheal extubation in the patients in PACU.

Kv2.1 channel currents in pancreatic beta-cells are thought to contribute to action potential repolarization and thereby modulate insulin secretion. Because of its central role in this important physiological process, Kv2.1 channel is a promising target for the treatment of type 2 diabetes. Jingzhaotoxin-XI (JZTX-XI) is a novel peptide neurotoxin isolated from the venom of the spider Chilobrachys jingzhao. Two-microelectrode voltage clamp experiments had showed that the toxin inhibited Kv2.1 potassium currents expressed in Xenopus Laevis oocytes. In order to investigate the structure-function relationship of JZTX-XI, the natural toxin and a mutant of JZTX-XI in which Arg3 was replaced by Ala, were synthesized by solid-phase chemistry method with Fmoc-protected amino acids on the PS3 automated peptide synthesizer. Reverse-phase high performance liquid chromatography (RP-HPLC) and matrix assisted laser desorption/ ionization time-of-flight mass spectrometry (MALDI-TOF/TOF MS) were used to monitor the oxidative refolding process of synthetic linear peptides to find the optimal renaturation conditions of these toxins. The experiments also proved that the relative molecular masses of refolded peptides were in accordance with their theoretical molecular masses. RP-HPLC chromatogram of co-injected native and refolded JZTX-XI was a single peak. Under the whole-cell patch-clamp mode, JZTX-XI could completely inhibit hKv2.1 and hNav1.5 channels currents expressed in HEK293T cells with IC50 values of 95.8 nmol/L and 437.1 nmol/L respectively. The mutant R3A-JZTX-XI could also inhibit hKv2.1 and hNav1.5 channel currents expressed in HEK293T cells with IC50 values of 1.22 micromol/L and 1.96 micromol/L respectively. However, the prohibitive levels of R3A-JZTX-XI on hKv2.1 and hNav1.5 channels were reduced by about 12.7 times and 4.5 times respectively, indicating that Arg3 was a key amino acid residue relative to the hKv2.1 channel activity of JZTX-XI, but it is also an amino acid residue correlated with the binding activity of JZTX-XI to hNav1.5 channel. Our findings should be helpful to develop JZTX-XI into a molecular probe and drug candidate targeting to Kv2.1 potassium channel in the pancreas.
Animals ; HEK293 Cells ; Humans ; Insulin-Secreting Cells ; metabolism ; Mutant Proteins ; genetics ; pharmacology ; NAV1.5 Voltage-Gated Sodium Channel ; metabolism ; Neurotoxins ; chemical synthesis ; genetics ; pharmacology ; Protein Refolding ; Shab Potassium Channels ; antagonists & inhibitors ; metabolism ; Sodium Channel Blockers ; pharmacology ; Spider Venoms ; genetics ; pharmacology ; Transfection