Objective:To synthesize analogues of 6-[4-(4-substituted acyl piperazinyl)phenyl]-4, 5-dihydro-3(2H)pyri-dazinones and 6-[4-(4-substituted acyl piperazinyl)phenyl]-5-methyl-4,5-dihydro-3(2H)pyridazinones in search for more potent and selective antithrombotic drugs. Methods:Many reactions such as Friedel-Crafts reaction,hydrolysis,cyclation, acyla-tion,substitution were used to synthesize the title compounds. Born method was applied for preliminary pharmacological test in vitro. Results : Twelve title compounds were synthesized, in which 10 compounds were firstly reported. Their structures were identified by element analysis and 1H-NMR. Conclusion:Results of preliminary pharmacological test show that all synthesized compounds are effective against platelet aggregation induced by ADP in vitro. The activity of compound (9) is the most potent. The activity of compound (6), (7), (10), (11), (12) are also better than that of CCI-17810.

BACKGROUND: It has been demonstrated that electromagnetic field (EMF) can adjust proliferation and differentiation of bone marrow mesenchymal stem cells, but the specific mechanism is not clear. OBJECTIVE: To investigate the effects of EMF-activated ERK1/2 pathway on proliferation and osteogenic differentiation of rat bone marrow mesenchymal stem cells.METHODS: The 3rd passage of rat bone marrow mesenchymal stem cells were received EMF treatment (15 Hz, 1 mT, sine wave), 20 μmol/L PD98059 + EMF treatment, or only PD98059 treatment. Simultaneously, a normal control group was established. Western blotting was applied to detect the activation of ERK signal pathway after EMF exposure. MTT assay was used to determine the activation of proliferation of cells. And alkaline phosphatase (ALP) activity in cells was detected by an ALP kit. RESULTS AND CONCLUSION: The ERK1/2 phosphorylation, proliferation and ALP activity of rat bone marrow mesenchymal stem cells were remarkably increased after exposure to EMF (P < 0.01). PD98059 could effectively block the increasing of ERK1/2 phosphorylation and cell proliferation (P < 0.01), but elevate ALP activity in a certain level (P < 0.01). EMF stimulation can fast activate ERK1/2 signal pathway and then promote the proliferation of rat bone marrow mesenchymal stem cells, however, ERK1/2 signal pathway activation has a less effect on osteogenic differentiation of bone marrow mesenchymal stem cells.

0.05),there were more apoptosis-starting cells found in normal skin(1738.33?348.89/mm2)than in keloid(891.67?395.00/mm2)(t=-5.565,P=0.000).The number of positive stained fibroblast cells in dermis significantly increased in keloid group(911.67?323.61/mm2),compared with that in normal skin group(220.00?80.00/mm2)(t=7.188,P=0.000).This result that the protein expression level of Bid was higher in keloid(0.46?0.08)than that in normal skin tissue(0.02?0.01)(t=18.905,P=0.000)was proven by western blotting test.Conclusions The number of apoptosis-starting cells in epidermis of normal skin is more than that in keloid.Bid protein in dermis of keloid is over expressed.

Objective To investigate the expression of nestin in keloid and hypertrophic scarring.Methods Samples were obtained from patients with normal skin(n=8),flat scars(n=7),keloids(n=8) and hypertrophic scars(n=8),respectively.Immunohistochemistry staining was used to measure the nestin expression in above samples.Nestin positive cells were counted to evaluate the expression levels.Results ①Nestin expression was detected in basal cells,prickle cells,sweat glands,hair follicles,sebaceous glands,and vascular endothelial cells in 8 samples of normal skin,and in fibroblasts in 7 of them.Nestin was also positively expressed in abovementioned cells or skin appendages in samples of flat scars,keloids and hypertrophic scars.②Nestin positive cell counts(epidermic cells,fibroblasts,and vascular endothelial cells) in keloids and hypertrophic scars were significantly higher than those in normal skin and flat scars(P0.05).Conclusions The increased expression of nestin in epidermic cells,fibroblasts,and vascular endothelial cells in keloid and hypertrophic scarring supports a possible role of nestin as fibrosis inducing factors in keloids and hypertrophic scars.

Objective To discuss the diagnosis and treatment of the benign tumors of pancreas,especially the surgical options and the prevention and cure of the postoperative complications.Methods Clinical data of 23 patients with benign lesions of the pancreas were retrospectively analyzed in our hospital from Jan.2004 to Dec.2009.All of the patients underwent surgical operation.Five patients were treated with pancreatic enucleation,8 patients were treated with distal pancreatectomy with splenectomy,1 patient was treated with central pancreatectomy,5 patients were treated with pancreaticoduodenectomy,4 patients were treated with pancreatic cyst- jejunum R-Y anastomosis.Eighteen patients were treated with somatostatin analogue.Results The pathological diagnosis included pancreatic cyst in 8,islet cell tumor in 4,mucous cystadenomas in 6,serous cystadenomas in 3,solid-pseudopapillary tumor in 2.Seven of 10 cases having pancreatic leak occoured secondary abdominal infection.One patient died of multiple organ dysfunction syndrome after pancreaticoduodenectomy.Conclusion There is no specific symptoms and serologic laboratory examinations for pancreatic benign tumors.CT and ERCP is very helpful for diagnosis and treatment.Surgical option is according to the location of the lesion and the judgement of the tumor whether being benign or malignant.Pancreaticoduodenectomy,central pancreatectomy,distal pancreatectomy with or without splenectomy and pancreatic enucleation is the reasonable surgical option.Pancreatic leakage is the main post-operation complication.Ligation the pancreatic duct reliably,treating the pancreatic raw surface appropriately,external drainage of the pancreatic fluid can reduce the rate of pancreatic leakage.

BACKGROUND: Recent studies have reported that more and more methods were used to prevent and cure tendon adhesion following tendon rupture by repairing tendinous sheath. Especially, amnion membrane is commonly used to effective prevent and cure adhesion and promote healing of biomembrane; however, the effect on tendon adhesion needs to be further studied. OBJECTIVE: To verify the efficacy of amnion membrane preserved in Honghua injection on preventing and curing tendon adhesion following transplanting into foot flexor tendon. METHODS: Bilateral foot flexor tendons of 32 healthy mature chickens were cut off. By anastomosis, amnion membrane preserved in Honghua injection was transplanted into left foot flexor tendon, considering as experimental group. Right foot flexor tendons were randomly divided into two groups: blank control group, anastomosis was performed alone; positive control group, amnion membrane not preserved in any injections was transplanted. At 4 weeks after fixation by plaster cast, sliding function of tendon was detected using biomechanics, and local samples were obtained for histopathological observation. RESULTS AND CONCLUSION: In the experimental group, broken end of left tendon was well healed; fiber tissues were formed surrounding tendon; tissue adhesion was not observed surrounding tendon. Proliferative quantity and adhesion of fiber tissues, as well as content of hydroxyproline in the experimental group were significantly less than in the blank control and positive control groups (P < 0.05, P < 0.01); total inflexion angle of articulationes digitorum pedis and slipping distance of flexor digitorum profundus tendon in the experimental group were significantly greater than in the blank control and positive control groups (P < 0.05, P < 0.01). The results indicated that amnion membrane preserved in Honghua injection might prevent tendon adhesion and effectively promote tendon healing.

BACKGROUND: It has been proved that electromagnetic field can adjust and control proliferation and differentiation of bone marrow mesenchymal stem cells v/a cyclic adenosine monophosphate (cAMP)-protein kinase A (PKA) signal transduction system. However, there are few relevant reports about Ca2+ as the second messenger in application. OBJECTIVE: To study the effects of verapamil on the proliferation and differentiation of bone marrow rnesenchymal stem cells stimulated by electromagnetic fields and to conclude influx changes of Ca2+.DESIGN, TIME AND SETTING: Electrostimulative cytological observation in vitro, which was performed in Laboratory of Orthopedic Surgery, Tongji Hospital between April and June 2005.MATERIALS: Six 4-5-week SD rats of clean grade were selected in this study. Verapami| was provided by Sigma Company, USA, and Helmholtz coil-magnetic field producer was made in Department of Electric Machine, Navy Engineering University.METHODS: The bone marrow mesenchymal stem cells were isolated and cultured in vitro with adherence method and digested with trypsin. The fourth-passage cells were harvested, adjusted to 1 × 107 L-1 in density, and divided into A, B, C and D groups in 96-well plate with 200 μ I/well. Cells in the normal control group were not performed with any agent. On the second day of inoculation, cells in the magnetic field (EMF) group were cultured in Helmholtz-coil magnetic field (0.8 mT, 50 Hz) in 0.05% CO2 saturated humidity incubator at 37 ℃, 30 minutes for each, 12 hours for interval, six time in total. Cells in the verapamil group were cultured with 20 μ mol/L verapamil, and cells in the combination group were cultured with 20 μ mol/L verapamil and magnetic stimulation.MAIN OUTCOME MEASURES: Proliferative activity was tested with MTT method, content of alkaline phosphate differentiated to osteoblasts was measured, and cells were stained with modified Gomori Ca-Co staining. RESULTS: Proliferative activity was significantly increased in the EMF group as compared with that in the normal control group after 3-day magnetic stimulation (P < 0.01), but verapamil could inhibit promotive effect on proliferation. Content of alkaline phosphate in the normal control group was similar to that in the EMF group, while those two contents were significantly higher than those in the verapamil group and the combination group (P < 0.01); furthermore, content of alkaline phosphate in the combination group was significant higher than that in the EMF group (P < 0.01). Qualitative analysis of alkaline phosphate showed a coincident result as mentioned above.CONCLUSION: EMF of 50 Hz frequency and 0.8 mT intensity can change intracellular free calcium ion concentration of bone marrow mesenchymal stem cells, and the change play a key role in the cellular proliferation and play a partial role in the differentiation of bone marrow mesenchymal stem cells into osteoblasta.

Objective To investigate the relative histological change of transplanted domestic expanded polytetrafluoroethylene (e-PTFE), which were treated with different methods, in order to offer the referential data for clinical application. Methods e-PTFE treated with different methods was transplanted into subcutaneous tussue of rat. The samples harvested according to time sequence were examined by using histological and histochemical methods. Dynamic change of the structure between the e-PTFE and it's surrounded tissue was investigated based on the examination. Results Cell and tissue were observed on the inside of all the e-PTFE including the control group and the experiment groups. Heavy cell infiltration on the 3rd day was the most significant in control group, and the quantity of tissue ingrowth was also the most until the 28th day. The next was trimming group. The quantity of both cell infiltration and the tissue ingrowth in high pressure steamed group and forceps squeezed group were less than that in other groups. Conclusion Cell infiltration into domestic e-PTFE is significantly achieved since 72 hrs and fibrovascular ingrowth since the 7days after implanted e-PTFE under subcutaneous tissue of the rat. Different treatment methods of e-PTFE can affect the speed of tissue ingrowth into the e-PTFE, which could be a reference for clinic application of e-PTFE.

In order to identify the differentially expressing gene of bone marrow mesenchymal stem cells (MSCs) stimulated by electromagnetic field (EMF) with osteogenesis microarray analysis, the bone marrow MSCs of SD rats were isolated and cultured in vitro. The third-passage cells were stimulated by EMFs and total RNA was extracted, purified and then used for the synthesis of cDNA and cRNA. The cRNA of stimulated group and the control group was hybridized with the rat oligo osteogenesis microarray respectively. The hybridization signals were acquired by using X-ray film after chemiluminescent detection and the data obtained were analyzed by employing the web-based completely integrated GEArray Expression Analysis Suite. RT-PCR was used to identify the target genes: Bmp1, Bmp7, Egf and Egfr. The results showed that 19 differentially expressing genes were found between the stimulated group and the control group. There were 6 up-regulated genes and 13 down-regulated genes in the stimulated group. Semi-quantitative RT-PCR confirmed that the expressions of Bmp1, Bmp7 mRNA of the stimulated group were up-regulated (P<0.05) and those of Egf, Egfr were down-regulated (P<0.05). It was suggested that the gene expression profiles of osteogenesis of the bone marrow MSCs were changed after EMF treatment. It is concluded that the genes are involved in skeletal development, bone mineral metabolism, cell growth and differentiation, cell adhesion etc.
Bone Marrow Cells/*cytology ; Cell Differentiation ; Down-Regulation ; Electromagnetic Fields ; Gene Expression Profiling ; Gene Expression Regulation ; Mesenchymal Stem Cells/*cytology ; Nucleic Acid Hybridization ; Oligonucleotide Probes/chemistry ; Osteogenesis/*genetics ; RNA, Complementary/metabolism ; Reverse Transcriptase Polymerase Chain Reaction

Objective:To investigate the changes in the expression of COX-2 in acute spinal cord injury of rats. Methods:Sixty adult SD rats were randomly divided into two groups: Trauma group (n=48) subjected to laminectomy and bounce of spinal cord; control group (n=12) only received laminectomy. The activity of COX-2 was detected using immunohistochemistry and Western blot analysis in cytoplasm of spinal cord. Results:The expression of COX-2 increased at 2-hour after injury and reached the top level at 24-hour. The expression of COX-2 dropped at 48-hour after injury and returned to basal level at 72-hour. Conclusion: The expression of COX-2 could be regarded as an index of inflammation on acute trauma spinal injury. Inhibition of COX-2 might be an new therapeutic method for spinal cord injury.