OBJECTIVESTo induce autologous bone marrow derived mesenchymal stem cell (aMSC) into chondrocyte, and to confirm the effects of 3 dimensional (3D) dynamic inducing in vitro and their long-term animal model repairing in vivo.

METHODSaMSC were separated from rabbits bone marrow aspirates, then respectively experienced 3D dynamic inducing in alginate drops in modified rotating wall bioreactor culture or in two dimensional (2D) inducing (culture flask) for 10 d. The induced cells were harvest and then mixed with fibrin sealant (FS) to repair rabbit knee femoral trochlea cartilage defects model. After 8, 12, 24, 48 weeks animals were euthanized. Gross appearance, histological appearances were examined.

RESULTSFlask culture groups showed a little chondrocyte differentiation, 3D inducing group showed obviously chondrocyte differentiation, improved collagen II and proteoglycan production. For 3D inducing ones in vivo, the cartilage defects were smoothly repaired by white translucent hard tissue with obvious hyaline-like cartilage histological appearance after 8, 12 weeks, and the defects boundary were hard to be identified with hyaline like cartilage with sustained histological appearance and score after 24, 48 weeks. For 2D ones in vivo, the cartilage defects were smoothly repaired after 8 weeks by hyaline like cartilage which showed accelerated degeneration after 24 weeks and lose cartilage performance completely after 48 weeks.

CONCLUSIONS3D dynamic inducing may assist aMSC on differentiating into chondrocyte, improve its long-term in vivo repairing effects, and enlighten its further applications in tissue engineering cartilage.

Animals ; Bone Marrow Cells ; cytology ; Cartilage, Articular ; injuries ; physiopathology ; surgery ; Cell Culture Techniques ; Cell Differentiation ; Cells, Cultured ; Chondrocytes ; cytology ; Chondrogenesis ; Disease Models, Animal ; Mesenchymal Stem Cell Transplantation ; Mesenchymal Stromal Cells ; cytology ; Rabbits ; Tissue Engineering ; methods ; Transplantation, Autologous ; Wound Healing

OBJECTIVETo study osteoblastic phenotype expression of New Zealand rabbit skin fibroblasts transfected with mouse core binding factor a1/osteoblast specific transplanting factor-2 gene (Cbfa1/Osf2).

METHODSCbfa1/Osf2 gene, engineered into eukaryotic expression vector pSG5, was introduced into New Zealand rabbit skin fibroblasts with catholyte liposomes-Lipofectamine 2000. Meanwhile, those transfected pSG5 and un-transfected were set the control groups. The expression of Cbfa1 gene, osteocalcin (OCN) gene, alkaline phosphatase (ALP) gene and pre-peptide 2 alpha gene of collagen type I were detected by RT-PCR assay. Cbfa1 protein was detected by Western-Blot assay, in-cell ALP activity by p-nitrophenyl phosphate (PNPP) assay and OCN content in the supernatant by radio-immunity method. The ossification nodules was detected by Alizarin-Red staining and scanning electron microscope.

RESULTSCbfa 1mRNA and Cbfa1 protein were expressed in New Zealand rabbit skin fibroblasts transfected with pSG5-Cbfa1/Osf2 from the first day to the fifth day, but they were not detected in the control groups. In the pSG5-Cbfa1/Osf2 transfected group, the expression of ALP gene and OCN gene were respectively induced from the third day and the forth day, pre-peptide 2 alpha gene of collagen type I was enhanced from the third day. From the sixth day, ALP activity greatly increased, OCN strongly secreted, and they were maintained at a high level for about 4 weeks, and the difference was significant compared with the control group (P < 0.05). On the forty-second day, ossification nodules were found on the surface of pSG5-Cbfa1/Osf2 gene transfected cells.

CONCLUSIONSNew Zealand rabbit skin fibroblasts transfected with pSG5-Cbfa1/Osf2 can express osteogenesis-related genes and proteins, and form ossification nodules on their surface.

Alkaline Phosphatase ; biosynthesis ; genetics ; Animals ; Cell Adhesion Molecules ; biosynthesis ; genetics ; Cells, Cultured ; Core Binding Factor Alpha 1 Subunit ; biosynthesis ; genetics ; Fibroblasts ; cytology ; physiology ; Gene Expression ; Genetic Vectors ; Mice ; Osteocalcin ; biosynthesis ; genetics ; Osteogenesis ; genetics ; physiology ; Rabbits ; Reverse Transcriptase Polymerase Chain Reaction ; Transfection

The aim of this paper is to report the synthesis of the mPEG-PCL-g-PEI copolymers as small interfering RNA (siRNA) delivery vector, and exploration of the siRNA delivery potential of mPEG-PCL-g-PEI in vitro. The diblock copolymers mPEG-PCL-OH was prepared through the ring-opening polymerization. Then, the hydroxyl terminal (-OH) of mPEG-PCL-OH was chemically converted into the carboxy (-COOH) and N-hydroxysuccinimide (NHS) in turn to prepare mPEG-PCL-NHS. The branched PEI was reacted with mPEG-PCL-NHS to synthesize the ternary copolymers mPEG-PCL-g-PEI. The structure of mPEG-PCL-g-PEI copolymers was characterized with Fourier transform infrared spectroscopy (FTIR), nuclear magnetic resonance (NMR) and gel permeation chromatography (GPC). The mPEG-PCL-g-PEI/siRNA nanoparticles were prepared by complex coacervation, and the nanoparticles size and zeta potential were determined, separately. The cytotoxicities of mPEG-PCL-g-PEI/siRNA nanoparticles and PEI/siRNA nanoparticles were compared through cells MTT assays in vitro. The inhibition efficiencies of firefly luciferase gene expression by mPEG-PCL-g-PEI/ siRNA nanoparticle at various N/P ratios were investigated through cell transfection in vitro. The experimental results suggested that the ternary (mPEG5k-PCL(1.2k))1.4-g-PEI(10k) copolymers were successfully synthesized. (mPEG(5k)-PCL(1.2k))1.4-g-PEI(10k) could condense siRNA into nanoparticles (50-200 nm) with positive zeta potential. MTT assay results showed that the cytotoxicity of (mPEG(5k)-PCL(1.2k))1.4-g-PEI(10k)/siRNA nanoparticles was significantly lower than that of PEI(10k)/siRNA nanoparticles (P < 0.05). The expression of firefly luciferase gene could be significantly down-regulated at a range of N/P ratio from 50 to 150 (P < 0.01), and maximally inhibited at the N/P ratio of 125. The mPEG-PCL-g-PEI polymers could delivery siRNA into cells to inhibit the expression of target gene with very low cytotoxicity, which suggested that mPEG-PCL-g-PEI could serve as a new type of siRNA delivery vector.
Cell Line, Tumor ; Cell Survival ; Drug Carriers ; Genes, Reporter ; Genetic Vectors ; Humans ; Luciferases ; metabolism ; Molecular Weight ; Nanoparticles ; Particle Size ; Polyesters ; chemistry ; Polyethylene Glycols ; chemistry ; Polyethyleneimine ; chemistry ; Polymers ; chemistry ; RNA, Small Interfering ; administration & dosage ; genetics ; metabolism ; Transfection

BACKGROUNDThe optimal revascularization strategy in patients with heart failure with preserved ejection fraction (HFPEF) remains unclear. The aim of the present study was to compare the effects of percutaneous coronary intervention (PCI) and coronary artery bypass grafting (CABG) in patients with HFPEF.

METHODSFrom July 2003 through September 2005, a total of 920 patients with coronary artery disease (CAD) and HFPEF (ejection fraction ≥ 50%) underwent PCI (n = 350) or CABG (n = 570). We compared the groups with respect to the primary outcome of mortality, and the secondary outcomes of main adverse cardiac and cerebral vascular events (MACCE), including death, myocardial infarction, stroke and repeat revascularization, at a median follow-up of 543 days.

RESULTSIn-hospital mortality was significantly lower in the PCI group than in the CABG group (0.3% vs. 2.5%, adjusted P = 0.016). During follow-up, there was no significant difference in the two groups with regard to mortality rates (2.3% vs. 3.5%, adjusted P = 0.423). Patients receiving PCI had higher MACCE rates as compared with patients receiving CABG (13.4% vs. 4.0%, adjusted P < 0.001), mainly due to higher rate of repeat revascularization (adjusted P < 0.001). Independent predictors of mortality were age, New York Heart Association (NYHA) class and chronic total occlusion.

CONCLUSIONAmong patients with CAD and HFPEF, PCI was shown to be as good as CABG with respect to the mortality rate, although there was a higher rate of repeat revascularization in patients undergoing PCI.

Aged ; Angioplasty, Balloon, Coronary ; mortality ; Coronary Artery Bypass ; mortality ; Female ; Heart Failure ; physiopathology ; therapy ; Hospital Mortality ; Humans ; Male ; Middle Aged ; Stents

BACKGROUNDIn patients with chronic total occlusion (CTO) and multivessel coronary artery disease, the comparison of surgical and the percutaneous revascularization strategies has rarely been conducted. The aim of this study was to compare long term clinical outcomes of drug eluting stent (DES) implantation with coronary artery bypass surgery (CABG) in the patients with CTO and multivessel disease.

METHODSFrom a prospective registry of 6000 patients in our institution, we included patients with CTO and multivessel coronary artery disease who underwent either CABG (n = 679) or DES (n = 267) treatment. Their propensity risk score was used for adjusting baseline differences.

RESULTSAt a median follow-up of three years, propensity score adjusted Cox regression analysis showed that the rate of major adverse cardiac cerebrovascular events (MACCE) was lower in CABG group (12.7% vs. 24.3%, hazard ratio (HR) 1.969, 95%CI 1.219 - 3.179, P = 0.006) mainly due to lower rate of target vessel revascularization in CABG group than in DES group (3.1% vs. 17.2%, HR 16.14, 95%CI 5.739 - 45.391, P < 0.001). The incidence of cardiac death or myocardial infarction (composite end point) was not significantly different between these two groups. On multivariate analysis, the significant predictors of MACCE were only the type of revascularization. Age, left ventricular ejection fraction (LVEF), and complete revascularization were identified as significant predictors of composite end points.

CONCLUSIONSOur study shows that in patients with CTO and multivessel coronary disease, DES can offer comparable long term outcomes in cardiac death and myocardial infraction free survival in comparison with CABG. However, there is an increased rate of MACCE which results from more repeat revascularizations. Obtaining a complete revascularization is crucial for decreasing adverse cardiac events.

Angioplasty, Balloon, Coronary ; methods ; Chronic Disease ; Coronary Angiography ; Coronary Artery Bypass ; methods ; Coronary Artery Disease ; surgery ; therapy ; Coronary Occlusion ; surgery ; therapy ; Drug-Eluting Stents ; Humans ; Prospective Studies

OBJECTIVEThe aim of this study was to observe the association between the occurrence of esophageal cancer lesions and esophageal mucosa fold (white ridges), and further identify where is the initial origin of esophageal cancer lesions in the esophagus mucosa.

METHODSThis was a cohort study which recruited 551 subjects underwent endoscopic examination in a high risk area of esophageal cancer in Linxian, Henan Province in 1987. 339 subjects with esophageal white ridges, and with red area or erosion lesion at the surface of the white ridges, was studied as exposure group. Other 212 subjects whose esophagus had no white ridges and pathological diagnosis was negative, was studied as control group. The endpoint was occurrence of pathologically confirmed esophageal cancer. After a 15-year follow-up, the results were compared between two groups.

RESULTSAmong the 551 subjects, there were 339 cases with esophageal mucosal white ridges in the exposure group. During the period of 15 year follow-up, the incidence of esophageal cancer was 11.8% (9/76) in 76 case with simple mucosal white ridges, 33.5% (88/263) in 263 subjects with white ridges and red area, or erosions on the surface of white ridge. While only 8.0% of subjects (17/212) developed esophageal cancer after the 15-year follow up in the control group. There was a significant difference between the two groups (P < 0.001).

CONCLUSIONEsophageal mucosal white ridge, especially white ridge with red area or erosions is closely associated with subsequent esophageal cancer occurrence in the esophageal cancer high risk area in China. It is suggested that esophageal mucosa with white ridge may be the initial origin of esophageal cancer. Further investigations focused on this spot are required.

Adult ; Aged ; Carcinoma, Squamous Cell ; epidemiology ; pathology ; China ; epidemiology ; Cohort Studies ; Esophageal Neoplasms ; epidemiology ; pathology ; Esophagoscopy ; Esophagus ; pathology ; Female ; Follow-Up Studies ; Humans ; Hyperplasia ; pathology ; Incidence ; Male ; Middle Aged ; Mouth Mucosa ; pathology ; Precancerous Conditions ; epidemiology ; pathology ; Prospective Studies

OBJECTIVETo summarize the diagnostic and therapeutic experiences of acute mesenteric venous thrombosis (MVT).

METHODSThe clinical data of 27 cases of acute MVT treated between 1983 and July 2007 were analyzed retrospectively.

RESULTSThe courses of disease were from 1 to 14 days (mean, 6.1 days). Eighteen cases (66.7%) had the history of portal hypertension, deep vein thrombosis, acute MVT or other hypercoagulability. The diagnostic sensitivity of ultrasonography, CT, angiography and serum D-Dimer level were 70.6% (12/17), 75.0% (6/8), 100% (6/6), 100% (6/6), respectively. Bowel necrosis occurred in all the 16 cases with bloody ascites. The thrombolytic and anticoagulation therapy are effective in 36.4% of cases (4/11). Twenty-two cases received operation, and resection of necrotic bowel was performed in all and thrombectomy in 3 cases. The main postoperative complications included 3 cases of deep vein thrombosis, 1 acute cardiac infarction, 3 short bowel syndrome. MVT recurred in 4 cases within a week after operation. Eight patients died within a month after confirmed with acute MVT, in which 7 patients died after operation. Anticoagulation medication was implemented in all the 19 survived patients. Fifteen patients were followed-up for 1-120 months (mean, 39.2 months), 7 of them continued the anticoagulation therapy during this period.

CONCLUSIONSThe determination of serum D-Dimer level and such adjuvant examinations as ultrasonography, CT and angiography are important diagnostic means for acute MVT. Anticoagulation and thrombolysis should be considered firstly if there is no active bleeding and bowel necrosis. We recommend laparotomy when bowel necrosis is suspected.

Adult ; Aged ; Female ; Follow-Up Studies ; Humans ; Male ; Mesenteric Veins ; Middle Aged ; Retrospective Studies ; Venous Thrombosis ; diagnosis ; therapy

AIM:To explore whether there is synergistic effect of recombinant human endostatin ( rh-Endo ) and paclitaxel (Pac) in the time window of vascular normalization and the role of magnetic resonance imaging (MRI) in early assessment of chemotherapy by observing the response of human triple -negative breast cancer ( TNBC) to Pac after vascular normalization in nude mice .METHODS:The human TNBC MDA-MB-231 cells were planted in the subcutaneous region of right lower abdomen of BALB/c-nu female nude mice .These nude mice were randomly divided into 4 groups (n=7).rh-Endo was given for 17 consecutive days in rh-Endo group and rh-Endo+Pac group.Pac was given on the 6th and 12th days in Pac group and rh-Endo+Pac group.The dosage of both drugs was 10 mg· kg-1· d-1(ip).On the day before the treatment and the 5th, 11th and 17th days after treatment, all the transplanted tumors were examined by MRI . All the mice were killed by cervical dislocation and their transplanted tumors were taken down for examinations after the last MRI on the 17th day.The changes of pathology, immunohistochemisty, microvessel density (MVD) and Ki67 expression were measured.RESULTS:On the 17th day, the volume of transplanted tumor in rh-Endo+Pac group was smaller than that in model group and rh-Endo group ( P<0.05 ) , and no difference between rh-Endo+Pac group and Pac group was found.On the 17th day, the tumor inhibitory rates in rh-Endo group, Pac group and rh-Endo+Pac group were 14.61%, 39.08%and 54.79%, respectively.The slow diffusion coefficient in Pac group was increased compared with model group , while it was decreased compared with rh-Endo+Pac group (P<0.05).No distant metastatic lesion in the tumor-bearing mice was observed .The necrotic rates in rh-Endo+Pac group and Pac group were higher than those in model group and rh-Endo group.The MVD in model group was higher than that in the other 3 groups.The MVD in rh-Endo+Pac group was decreased compared with Pac group and rh-Endo group .The Ki67 level in rh-Endo+Pac group was decreased compared with rh-Endo group , and no difference between rh-Endo+Pac group and Pac group was detected .CONCLUSION:In the time window of vascular normalization , the combination of Pac and rh-Endo has a significant antitumor effect on TNBC , but this study did not observe a significant synergistic effect of the 2 drugs.The change of slow diffusion coefficient can predict the therapeutic effect in advance .

OBJECTIVETo investigate the transfection efficiency and the optimal conditions of delivering latent membrane protein-1 (LMP-1) gene to dendritic cells (DCs) by ultrasound exposure combined with contrast agent.

METHODSHuman DCs were cultured in vivo and transfected with the recombinant plasmid pEGFP-C3-LMP1 under varying conditions including ultrasound intensities, exposure time and microbubble contrast agent concentration. The transfection efficiency was assessed by fluorescent microscopy and flow cytometry, and the cell viability by trypan blue exclusion test.

RESULTSAn exposure time of 60 s at MI 1.0 with a microbubble contrast agent concentration of 20% resulted in the optimal effect of delivering the recombinant plasmid pEGFP-C3-LMP1 into the DCs, with a transfection efficiency of (14.37∓2.12)%. Over 90% of the transfected cells were viable after the transfection.

CONCLUSIONMicrobubble contrast agent combined with ultrasound exposure can enhance the delivery of recombinant plasmid pEGFP-C3-LMP1 into the DCs.

Adaptor Proteins, Signal Transducing ; genetics ; Cells, Cultured ; Contrast Media ; administration & dosage ; pharmacology ; Cytoskeletal Proteins ; genetics ; Dendritic Cells ; drug effects ; metabolism ; Humans ; LIM Domain Proteins ; genetics ; Microbubbles ; Plasmids ; Transfection ; Ultrasonics

Biopsy ; methods ; standards ; Chronic Disease ; Humans ; Liver ; pathology ; Liver Diseases ; diagnosis ; pathology ; Quality Control