Objective To explore a method of preparing the acellular tendons of human with chemical approaches.Methods From April 2011 to June 2012,several treatments were performed on human tendons using 1.00％ tri(n-butyl) Phosphate (TnBP) combinded with Triton X-100 at the concentration of 0,0.25％,0.50％,1.00％ respectively.Specimens were examined using histological observation,scanning electron microscopy,biomechanical testing and hydroxyproline quantitation.The results were then compared with fresh tendons of control group,so as to value the characteristics of decellularized human tendon scaffold.Results Acellular human tendons had glossy surface,intact aponeurotic membrane and satisfactory flexibility.A small number of disrupted cells remained in the 1.00％ TnBP + 0％ Triton X-100 treated tissue,while other three experimental groups successfully eliminate all cells.Intact and regular collagen architecture was retained in 1.00％ TnBP +0.25％ Triton X-100 treated tissue.1.00％ TnBP + 0.50％ Triton X-100 and 1.00％ TnBP + 1.00％ Triton X-100 treated tissue were nearly identical to 1.00％ TnBP +0.25％ Triton X-100 treated tissue,but the interval of collagen was slightly wider than the control group,the maximum load (385.22 ± 80.32N,398.22 ± 127.20N),ultimate tensile strength(46.69 ± 16.30Mpa,46.20 ±5.52Mpa) and hydroxyproline content(0.282663 ± 0.0110109 μg/mg,0.279355 ± 0.0102129 μg/mg) were statistically lower (P ＜ 0.05) compared with those of the control group,maximum load(533.28 ± 135.77N),ultimate tensile strength (65.56 ± 14.40Mpa) and hydroxyproline content (0.292882 ± 0.0100988 μg/mg) respectively.Conclusion The decellularization treatment with 1.00％ TnBP + 0.25％ Triton X-100 could be optimized for preparing acellular human tendons.

The cancer stem cells (CSCs) from human osteosarcoma by serum-free three-dimensional culture combined with anticancer drugs were isolated and identified. The primary cells derived from human osteosarcoma were digested by trypsin to prepare a single-cell suspension, and mixed homogeneously into 1.2% alginate gel. Single-cell alginate gel was cultured with serum-free DMEM/F12 medium. Epirubicin (0.8 mug/mL) was added to the medium to enrich CSCs. After cultured conventionally for 7 to 10 days, most of cells suspended in alginate gel were killed by epirubicin. But few cells survived and some single-cell cloning spheres formed. Immunofluorescent staining for Oct3/4 and Nanog was implemented to find cells with properties of self-renewal and multi-potential differentiation. Cells from cloning spheres were transplanted into BALB/c mice to detect the tumorigenicity in vivo. The results showed that some cells positive for Oct3/4 (TRITC) and Nanog (TRITC) were found in single-cell cloning spheres, and most of positive cells were concentrated in the core of sphere. Cells from spheres could form osteosarcoma in the body of mice. It was concluded that cells from single-cell cloning spheres had the properties of the expression of parts of stem cell genes (Oct3/4 and Nanog), resisting anti-cancer drugs, and tumorigenicity in vivo. To sum up, it is believed that cells obtained from osteosarcoma by serum-free three-dimensional culture combined with anticancer drugs are cancer stem cells.

Objective To evaluate the role of early cleavage embryo in combination with embryo growth rate and morphology scoring in embryo selection in in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) cycles. Methods Six hundred and ten IVF/ICSI cycles were randomly assigned to group A(269 cycles) and group B(341 cycles). In group A, transferred embryos were chosen according to embryo growth rate and morphology scoring by 72 h(D3) after fertilization, while early cleavage embryo was added to the selecting system in group B. The pregnancy rate and implantation rate were compared between two groups, and the clinic outcomes were compared between transfers with early cleavage embryos and without early cleavage embryos in group B. Results The pregnancy rate and implantation rate in group B were significantly higher than those in group A (P < 0.05). Transfers with early cleavage embryos also achieved much higher pregnancy rate and implantation rate in group B (P < 0.01). Conclusion Compared with embryo growth rate and morphology scoring, early cleavage embryo in combination with embryo growth rate and morphology scoring can improve the clinical outcomes in IVF/ICSI cycles.

OBJECTIVE: To investigate the diagnostic significance of basophil activation test (BAT) in anaphylaxis to non-ionic contrast media through testing the content of CD63, mast cell-carboxypeptidase A3 (MC-CPA3), and terminal complement complex SC5b-9 of the individuals by testing their levels in the normal immune group and the anaphylaxis groups to β-lactam drugs and non -ionic contrast media. METHODS: The CD63 expression of basophilic granulocyte in blood was detected by flow cytometry. The levels of MC-CPA3 in blood serum and SC5b-9 in blood plasma were detected by ELISA. RESULTS: The CD63 expression of basophilic granulocyte in blood, the levels of MC-CPA3 and SC5b-9 of anaphylaxis to non-ionic contrast media and β-lactam drugs were significantly higher than that in normal immune group (P < 0.05). CONCLUSION There is activation of basophilic granulocytes, mast cells and complement system in anaphylaxis to non-ionic contrast media. BAT can be used to diagnose the anaphylaxis to non-ionic contrast media.
Anaphylaxis/diagnosis* ; Basophils/cytology* ; Carboxypeptidases A/metabolism* ; Complement Membrane Attack Complex/metabolism* ; Contrast Media ; Flow Cytometry ; Granulocytes/cytology* ; Humans ; Mast Cells/cytology* ; Tetraspanin 30/metabolism*

OBJECTIVETo detect the character of surgical treatment of massive soft tissue sarcoma in the shoulder girdle and analyze the impact factor to the result.

METHODSSeven patients with massive soft tissue sarcoma in the shoulder girdle were treated in our department between 2005 and 2009. There were 4 males and 3 females. All the patients were referred to our hospital after local recurrence post-operatively. The mean age was 43.8 years old (range 14 - 75). The maximum diameter of the tumor varied from 10 to 16 centimeters. All the patients were performed surgery, wide margin in 4 cases and marginal margin in 3 cases. Five were performed tumor resection and reconstruction with latissimus dorsi muscle flap transfer and skin graft. One was reconstructed with advanced skin flap and skin graft. The other one was treated with skin graft. The diagnosis included 3 malignant fibrous histiocytomas, 1 low grade myxoid fibrosarcoma, 1 Primitive neuroectodermal tumor, 1 rhabdomyosarcoma, 1 dermatofibrosarcomas protuberans. The MSTS score system was used to evaluate the shoulder function.

RESULTSSeven patients were followed up with long time. The mean follow up was 29 months (range 10 to 46 months). Two patients suffered local recurrence and one died of pulmonary metastasis 6 months after the second surgery for local recurrence. One patient suffered pulmonary metastasis. The last four patients were disease-free at the end of follow-up. The function of shoulder girdle was satisfactory. The mean MSTS score was 28.

CONCLUSIONSSoft tissue sarcomas in the shoulder girdle are easy to be misdiagnosed and mistreated. Wide surgical margin was the key impact factor to the local recurrence of soft tissue sarcoma in the shoulder girdle. The surgical margin and invasion of the tumor are the key factor to the prognosis. The soft tissue defect after surgery is often reconstructed by muscle flap transfer or skin flap transfer. The latissimus dorsi muscle flap transfer is often used.

Adolescent ; Adult ; Aged ; Female ; Humans ; Male ; Middle Aged ; Prognosis ; Retrospective Studies ; Sarcoma ; diagnosis ; surgery ; Shoulder ; pathology ; Soft Tissue Neoplasms ; diagnosis ; surgery ; Treatment Outcome ; Young Adult

BACKGROUND: Our previous findings have shown that poly(lactic acid)/hydroxyapatite (PLA/HA) composite biomaterials prepared by the supercritical carbon dioxide method have good physiochemical properties and biocompatibility. However, the composite materials only have osteoconductivity but no osteoinductivity. OBJECTIVE: To prepare icariin-PLA/HA (IC-PLA/HA) composite biomaterials with good osteoconduction and osteoinduction. METHODS: The supercritical carbon dioxide method was used to prepare IC-PLA/HA composite biomaterials containing 10-4, 10-5, 10-6mmol/g IC, named as IC-PLA/HA(1 000), IC-PLA/HA(100) and IC-PLA/HA(10). PLA/HA composite material served as controls. Biomechanical properties, porosity, sustained release characteristics were detected, and scanning electron microscope observation was performed, in order to screen out the optimal IC content in the composite materials. RESULTS AND CONCLUSION: (1) Compressive strength and elastic modulus of IC-PLA/HA(1 000), IC-PLA/HA(100) and IC-PLA/HA(10) showed no difference from those of PLA/HA. (2) The porosity of all the composite materials was over 75%, and there was still no difference among groups. (3) The IC release from IC-PLA/HA was faster within the first 3 days, and then reduced gradually. However, after 7 days, the IC release plateaued, and the IC release amount from the IC-PLA/HA(100) was close to 10-7mol/L that had been confirmed to be an effective and safe concentration in the previous experiments. (4) Under the scanning electron microscope, HA and PLA were mixed homogeneously and IC was difficult to be identified. The pore size of the IC-PLA/HA(100) ranged from 50 μm to 150 μm. Overall, the IC-PLA/HA composite biomaterials have good mechanical and sustained-release properties.

The dosage-efficacy/toxicity relationship of the 50% alcohol extracts of Polygonum multiflorum was comparatively investigated on either normal or CCl4-induced chronic liver injury rats, by determining the general condition, serum biochemical indices and liver histopathology, coupled with the factor analysis. The dosages were 10 and 20 g raw materials per kg body weight. Compared with the normal control group, the normal high dose group showed significant increases of the serum alanine transaminase (ALT), total bilirubin (TBIL), high mobility group box 1 (HMGB-1) and interleukin-1β (IL-1β) (P < 0.05 or P < 0.01), as well the frequent incidences of inflammatory cell infiltration, hepatic sinus enlargement and fiber stripes formation in histopathological sections. Compared with the model control group, the model low dose group showed significant declines of serum ALT, aspartate transaminase (AST) and total bile acid (TBA) (P < 0.05), as well the alleviation of vacuoles of hepatocytes, but no amelioration of the inflammatory cell infiltration and fibrous tissue hyperplasia; moreover, the model high dose group showed significant degeneration declines of serum HMGB-1, tumor necrosis factor-α (TNF-α) and IL-1β (P < 0.05, P < 0.01), as well the evident alleviation of vacuoles degeneration of hepatocytes, inflammatory cells infiltration and fibrosis degree. The factor analysis showed that the low dosage treatment had almost neither injuring effect on the normal rats nor protective effect on the model rats; while the high dosage treatment showed observable injuring effect on the normal rats, expressed by the significant increases of the factor-1 (HMGB-1, TNF-α and IL-1β as the main contributors) and factor-2 (TBIL, ALT and TBA as the main contributors) relative to the normal control group. The liver protective effect of the high dosage treatment could be observed with the significant reduction of the factor-1, indicating the effective alleviation of the expression of inflammatory cytokines. In conclusion, it could illustrated the phenomenon of symptom-based prescription theory of Polygonum multiflorum on rat livers: the high dosage of the herb had either an injuring effect on normal rats, or a therapeutic effect on the rats with chronic liver injury.
Alanine Transaminase ; blood ; Animals ; Aspartate Aminotransferases ; blood ; Bile Acids and Salts ; metabolism ; Bilirubin ; blood ; Chemical and Drug Induced Liver Injury ; drug therapy ; Drugs, Chinese Herbal ; pharmacology ; Fallopia multiflora ; chemistry ; HMGB1 Protein ; metabolism ; Hepatocytes ; drug effects ; Interleukin-1beta ; metabolism ; Liver ; drug effects ; pathology ; Plant Extracts ; pharmacology ; Rats ; Tumor Necrosis Factor-alpha ; metabolism

OBJECTIVETo investigate the mechanism of rosiglitazone (RSG, the activator of peroxisome proliferators activated receptor lambda) for inhibiting endothelin-1 (ET-1)-induced neonatal rat cardiac myocyte hypertrophy and the role of protein kinase C (PKC) and c-fos.

METHODSIn vitro cultured neonatal rat cardiac myocytes were treated with ET-1, phorbol ester (PMA, the PKC activator), ET-1+RSG, ET-1+chelerythrine (che, the PKC inhibitor), PMA+RSG, or without treatment (control), respectively. The effects of RSG on the protein content, (3)H-leucine incorporation, PKC activity and C-fos protein expression were observed in the cardiac myocytes stimulated with ET-1 or PMA.

RESULTSAfter two days of culture, the intracellular protein content in ET-1 group and PMA group were increased by 15% (339-/+15 microg/ml) and 13% (329-/+14 microg/ml) as compared with the control cells (290-/+13 microg/ml), respectively (P<0.01). Compared with the ET-1 group, cells treated with ET-1+10(-8) mol/L RSG, ET-1+10(-7) mol/L RSG, and ET-1+che showed decreased intracellular protein content by 10% (303-/+14 microg/ml, P<0.05), 12% (292-/+11 microg/ml, P<0.05), and 13% (291-/+12 microg/ml, P<0.01), respectively. The intracellular protein content in PMA+10(-7) mol/LRSG group was decreased by 10% (P<0.05) in comparison with the PMA group. RSG inhibited protein synthesis enhancement and increased (3)H-leucine incorporation induced by ET-1 and PMA, and antagonized the effects of ET-1 and PMA in promoting PKC activity and c-fos protein expression in the myocytes.

CONCLUSIONThe inhibitory effect of RSG on ET-1- or PMA-induced myocyte hypertrophy is associated with PKC-c-fos pathway.

Animals ; Animals, Newborn ; Blotting, Western ; Cell Enlargement ; drug effects ; Cells, Cultured ; Dose-Response Relationship, Drug ; Endothelin-1 ; pharmacology ; Hypoglycemic Agents ; pharmacology ; Myocytes, Cardiac ; cytology ; drug effects ; metabolism ; Protein Kinase C ; metabolism ; Proto-Oncogene Proteins c-fos ; biosynthesis ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; drug effects ; Tetradecanoylphorbol Acetate ; pharmacology ; Thiazolidinediones ; pharmacology

This study was purposed to investigate the effect of phycocyanin at different concentration on proliferation of K562 cells, to detect the changes of integrin beta1 expression and intracellular focal adhesion kinase (FAK) gene expression on the surface K562 cells treated with phycocyanin, and to explore the possible mechanism of integrin beta1 effect on phycocyanin inhibiting proliferation of K562 cells. The expression level of integrin beta1 on the surface of K562 cells was evaluated by flow cytometry (FCM); the growth of K562 cells treated with phycocyanin was measured by MTT assay; the expression level of FAK mRNA was analyzed by relatively quantitative RT-PCR after four-day culture of K562 cells with phycocyanin of 40 microg/ml, 80 microg/ml and 160 microg/ml, respectively. The results showed that integrin beta1 expression on the surface of K562 cells was significantly higher than that in bone marrow mononuclear cells (BMMNC) from normal subjects. Phycocyanin could not change the level of integrin beta1 expression. Phycocyanin could increase the expression of FAK gene on K562 cells and inhibit the proliferation of K562 cells. It is concluded that phycocyanin can inhibit the proliferation of K562 cells through enhancing the conjunction of cell stroma with integrin beta1 on K562 cell surface, up-regulating the expression level of FAK gene in K562 cells, restoring the signaling pathway of proliferation inhibition mediated by integrin beta1. The possible mechanism of phycocyanin in the proliferation inhibition of K562 cells is to increase the expression of FAK gene. The phycocyanin may be considered as a potential agent for inhibition of cancer cell proliferation.
Antineoplastic Agents, Phytogenic ; pharmacology ; Cell Proliferation ; drug effects ; Focal Adhesion Kinase 1 ; biosynthesis ; genetics ; Humans ; Integrin beta1 ; biosynthesis ; genetics ; K562 Cells ; Phycocyanin ; pharmacology ; RNA, Messenger ; biosynthesis ; genetics

OBJECTIVETo study the cleavage activity of specific deoxyribozyme to hepatitis C virus in vitro.

METHODSThree deoxyribozymes were designed to cleave at sites 157, 168, 173 in HCV 5'-noncoding region with the active region of 5'-GGCTAGCTACAACGA-3' respectively. Plasmid pCMV/T7-NCRC -Delta Luc was completely linearized with restriction endonuclease Xba I. HCV RNA5'-NCRC was transcribed in vitro from the linearized products and radiolabelled with [alpha-32P] UTP. Under the conditions of 37 degrees C, pH7.5, Mg2+ 10 mmol/L, the three deoxyribozymes were mixed with substrate RNA individually for 120 minutes and then the reactions were terminated. The cleavaged products were separated with 8% denaturated polyacrylamide gel electrophoresis and displayed by autoradiography. DRz3 was mixed with the substrate RNA at different Mg2+ concentrations. The cleavage efficiency was analyzed with a gel document action analyzing systems.

RESULTSUnder the adopted conditions the three deoxyribozymes efficiently cleaved to the target RNA in vitro and the cleavage activity of DRz3 was increased with the increase of Mg2+ concentration.

CONCLUSIONThe designed deoxyribozymes can cleave 5'-NCR mRNA of HCV efficiently in vitro and it is dose-respondent to Mg2+ concentration.

DNA, Catalytic ; genetics ; DNA, Single-Stranded ; genetics ; Genetic Therapy ; Hepacivirus ; genetics ; Hepatitis C ; therapy ; Humans ; RNA, Messenger ; genetics