Ovbective To evaluate the feasibility of a tissue engineered patch constructed by a new copolymer of p(3HB-co-3HH) and polyurethane(PU) as in partial aortic replacement. Methods The autologous cell-polymer scaffold were used to replace 2.0～ 2.5cm abdominal aortic segments in dogs (group TE,n=6). The patches were seeded with 1?10 7 myofibroblasts each day for 4 days,and then 1?10 7 endothelial cells were seeded onto the scaffold. After 48 hours' incubation,the cell-polymer scaffolds were implanted to replace a segment of canine abdominal. In the control group (n=4),aortic segments were replaced with acellular polymer patches. No anticoagulation agent was used. At 2nd,4th,8th,12th,24th,and 48th weeks after operation,the seeded group animals were killed. The control group animals were killed at 4th,12th,24th,and 48th weeks. Explanted patches were examined histologically with scanning electron microscopy,and biochemically. Results In the group of TE patches,the tissue-engineered patches were covered with endothelium-like tissue macroscopically,and there was no thrombus formation on any of the specimens. Histological staining showed uniform layered tissue with endothelium and laminated fibrous tissue with collagen as predominant extracellular matrix. A confluent smooth surface was observed by scanning electron microscopy. An increase in the content of collagen and elastin was observed,and at 48th weeks after operation and their contents equaled to the level of native aorta. There was no endothelium formation in the acellular control,the collagen and elastin content were also smaller than that of the TE groups. Conclusion The autologous aortic grafts with biological characteristics resembling the native aorta can be created by using TE approach.

BACKGROUND:cellmarker technology has been widely applied in many studies concerning celltransplantation. Chlormethylbenzamido-1,1-dioctadecyl-3,3,3’3’-tetramethylin-docarbocyamine (CM-DIL) and 4’,6-diamidino-2-phenylindole (DAPI) are commonly used for labeling cells. To our knowledge, there are few reports on comparing the two fluorescent dyes.
OBJECTIVE:To compare the effects of CM-DIL and DAPI on labeling bone marrow mesenchymal stem cells in vitro and in vivo.
METHODS:Isolation and expansion of bone marrow-derived mesenchymal stem cells were performed according to attachment culture. The cells were labeled by CM-DIL and DAPI, respectively. cellviability was assessed via trypan blue exclusion assay. Growth curves of bone marrow-derived mesenchymal stem cells were depicted using MTS assay. The reduction of fluorescent intensity was observed under an inverted fluorescent contrast phase microscope from passage 1 to passage 3 after labeling. Myocardial infarction was induced by left anterior artery ligation in Sprague-Dawley rats. One week later, bone marrow-derived mesenchymal stem cells labeled by CM-DIL or DAPI were injected randomly into the border area of infarct myocardium. After 3 days, transplanted celldistribution was examined under the fluorescent microscope through paraffin sections and frozen sections respectively.
RESULTS AND CONCLUSION:In vitro, bone marrow-derived mesenchymal stem cells labeled by both CM-DIL and DAPI showed decreased cellproliferation during the early period;the percentage of fluorescent-positive cells was approximately 100%in the two groups;however, the fluorescent intensity was significantly reduced from passage 1 to passage 3 in bone marrow-derived mesenchymal stem cells labeled by DAPI. In vivo, the transplanted cells were detected in a concentrated way both on the paraffin sections and frozen ones;the background color of frozen sections was lower in the CM-DIL group than in the DAPI group;false positive results of fluorescent expression could be eliminated in the CM-DIL group by using fluorescent mounting medium with the fluorescence of DNA staining. These data indicates that CM-DIL is more appropriate to in vivo tracing cells than DAPI.

ObjectiveTo investigate the clinical application value of enzyme-linked immunosorbent assay (ELISA) and time-resolved fluorescence analysis(TRFIA) and latex immune chromatography (GICA) in detecting hepatitis B serum markers.MethodsOne hundred and forty-five suspected patients of hepatitis B were detected serum markers of hepatitis B by ELISA,TRFIA and GICA method,and the results were compared and analyzed.ResultsWhen TRFIA method was as gold standard,the positive coincidence rate of ELISA and GICA method in HBsAg,HBeAb,HBcAb was 71％ (57/80),45％ (36/80),and in HBsAb,HBeAb,HBcAb was 33％( 1/3),0 (0/3),and there were significant differences between two methods (P＜0.05 ) ; the others were no significant differences (P ＞ 0.05 ).There was significant difference in the sensitivity of HBsAg between ELISA method and GICA method (P ＜ 0.05 ),but there was no significant difference in HBsAb,HBeAg,HBeAb,HBcAb(P ＞ 0.05 ).There was no significant difference in the specificity of HBsAg,HBsAb,HBsAg,HBeAb and HBcAb between ELISA method and GICA method (P ＞0.05).There was significant difference in HBsAg among three methods(P ＜0.05),but there was no significant difference between ELISA method and TRFIA method (P＞0.05),and there was significant difference between GICA method and TRFIA method (P＜0.05).There was no significant difference in HBsAb,HBeAg,HBeAb and HBcAb among three methods (P ＞ 0.05 ); there was significant difference in both HBsAb and HBeAb positive among three methods (P ＜ 0.05),and there was significant difference between ELISA method,GICA method and TRFIA method (P＜ 0.05).ConclusionTRFIA method has supreme measuring range,sensitivity and specificity supreme,but the price is higher;ELISA method is in the intermediate level of three methods and price is cheap,and it as well as TRFIA is suitable for the batch detection; GICA accuracy is low,but quick and simple,it is more suitable for the complement of the first two methods.

Objective To investigate the value of CT-guided 125I particles implantation combined with gemcitabine plus S-1 (GS) regimen in the treatment of locally advanced pancreatic cancer.Methods 42 patients with unresectable local advanced pancreatic cancer were given with CT-guided 125I seed implantation.3-4 cycles of GS regimen was given based on the tolerance of patient s body within 3-7 d after implantation of particles.Review of blood,CA199,chest X-ray,CT scan + enhanced or MRI were performed at 2nd,4th,6th,12th month after surgery.Results 2nd,4th,6th month after surgery,tumor lesions were significantly reduced,ORRs were 59.5 ％ (25/42),66.7 ％ (28/42) and 73.8 ％ (31/42),respectively,DCRs were 83.3 ％ (35/42),78.6 ％ (33/42) and 76.2 ％ (32/42),respectively.No serious adverse reactions were observed,patient could tolerate these reactions.The 6th,12th,24th month survival rates were 100 ％ (42/42),47.6 ％ (20/42) and 11.9 ％ (5/42),respectively,mPFS was 8.27 months and mOS was 12.00 months.Conclusion CT-guided 125I particles implantation combined with GS regimen is convenient,safe,high efficacy in the treatment of locally advanced pancreatic cancer.

Objective To evaluate the efficacy and safety of CT-guided 125I radioactive seed implantation combined with gemcitabine and Gio (gemcitabine and S-1, GS scheme) chemotherapy in treating advanced pancreatic carcinoma. Methods Sixty-eight patients with inoperable advanced pancreatic carcinoma were randomly divided into two groups. Patients in group A(n=38) were treated with CT-guided 125I radioactive seed implantation combined with GS chemotherapy scheme, while patients in group B (n=30) received GS chemotherapy scheme only. The short-term effect, the median progression-free survival time, the median survival time and adverse reactions of the two groups were determined , and the results were compared between the two groups. Results The objective response rate (ORR), disease control rate (DCR) and clinical benefit rate (CBR) of the group A were 57.9%, 73.7%and 84.2%respectively, while those of group B were 26.7%, 46.7% and 60.0% respectively. The differences between the two groups were statistically significant (P<0.05). In group A the median progression-free survival time and the median survival time were 8.00 months and 11.84 months respectively, which were strikingly higher than those in group B (5.63 months and 10.40 months respectively), the differences between the two groups were statistically significantly (P<0.05). No significant differences in gastrointestinal reactions, blood toxicity, liver toxicity and other adverse reactions existed between the two groups (P>0.05). Conclusion For advanced pancreatic carcinoma, CT-guided 125I radioactive seed implantation combined with GS program is a safe and effective treatment.

BACKGROUND: In vitro construction of tissue engineered cardiac muscle becomes a hot spot in recent years, and the selection and design of scaffold is the key link. However, there is lack of ideal cardiac tissue engineering scaffold material. OBJECTIVE: To evaluate the novel biodegradable polyurethane in vitro, and to discuss the feasibility of polyurethane as cardiac tissue engineeru scaffold. METHODS: A new type polyurethane (PV-Lys) was synthesized using diphenylmethane-4,4'-diisocyanate as hard segment and lysine as expand chain. The tensile and suture strength were tested in vitro respectively, hydrolytic degradation was carded out in phosphate buffer saline of pH 7.4 at 37 ℃, and cytotoxicity was evaluated by MTT measurement and morphological observation. RESULTS AND CONCLUSION: The tensile strength of the polyurethane was up to (8.1±0.1) MPa, and the suture strength was (12.2+0.8) N. The average value of the mass loss of PV-Lys was (13.1+0.3)% at 8 weeks of in vitro hydrolytic degradation. MTT assay results showed that the cytotoxic grade of the novel PV-Lys was 0-1. Cell morphology observation showed that the L929 cells were spindle-shaped or tdangular with good stretch. This PV-Lys scaffold is with favorable mechanical property, cytocompatibility, biodegradable property, which meets the requirements of tissue engineering application.

ObjectiveTo evaluate the feasibility of constructing tissue engineering cardiac patch with photooxidationfixed acellular bovine pericardium.MethodsFresh bovine pericardia were treated by dye-mediated photooxidation after decellularization.Some of them were seeded with bone marrow stromal cells(MSCs) isolated from male SD rats to construct cardiac patches.Myocardial infarction(MI) model was made in female SD rats by left anterior descending coronary ligation(LAD).One week later, the confirmed MI rats were divided into three groups randomly, group MI (n = 15)without any treatment; group P (n = 18) with photooxidated pericardia implantation ; group P + C (n = 18) with seeded pericardia implantation.A sham group (n = 10) was also performed with opening and closing chest twice only.The heart were explanted at 2 or 4 weeks after implantation, and examined histologically and immunohistochemically.The heart function was evaluated by echocardiography at 4 weeks before excising the rats.ResultsThere were no cells or cell debris remained in bovine pericardium tissue.The fiber structure became condensed after photooxidation.The seeded cells formed a continuous layer on the surface of the tissue.The pericardial degradation level and newly formed microvessel density were larger in group P + C than in group P after 2 [ (13.7 ±5.2)个/mm2 vs (7.1 ±3.1)个/mm2, P＜0.05]and4 [(22.6 ±4.9)个/mrn2 vs (14.1 ±5.3)个/mm2, P＜0.05]weeks.Four weeks after transplantation, cardiac echocardiography showed left ventricular ejection fraction(LVEF) was lower in group MI (44.8 ± 4.4) ％ and group P (48.4 ± 5.0) ％ compared with group P + C (49.3 ± 4.8) ％, left ventricular fractional shorterning(LVFS) was lower in group MI (18.0 ± 2.2) ％ and group P (19.8 ± 2.5) ％ compared with group P + C (20.4 ±2.5) ％, the difference between P + C and MI was significant.ConclusionTransplantation of the tissue engineered bovine pericardial patches with dye-mediated photooxidation can improve heart function in MI rats.This kind of patches demonstrates a promising prospect in the future.

ObjectiveTo evaluate in vivo antithrombosis property of optimized FW-Ⅱ axial blood pump and provides evidence for future clinical use.MethodsA left ventricle-pump-descending aorta bypass model was established in five healthy sheep (60-70 kg) and the circulation of these sheep was assisted by FW-Ⅱ axial blood pump for 2 weeks.In preoperative and postoperative day 1,2,3,7,10 and 14,blood was drawn from the jugular vein to examine platelet activation and leukocyte-platelet aggregation respectively quantified with Annexin V,CD41/61 and CD14-PE by flow cytometry assays.Immediately after termination of the experiment,FW-Ⅱ axial blood pumps were explanted and each part was inspected for thrombus formation.Macroscopic and histological examinations were checked on heart,brain,kidney and spleen,respectively for thrombosis.ResultsCompared with preoperative baseline,the number of platelet activation and leukocyte-platelet aggregation reached a peak at postoperative day 2,it retained a high level within 7 days,then gradually decreased,but was still higher than preoperative level at dayl4.According to rotating speed,the number of platelet activation and platelet-leukocyte aggregation were lowest at the speed of 8000 r/min Minus thrombus were found in the front and rear hub of the pump rotor,and there was no thrombus at other components (flow straighter,impeller and pump housing).There were no ischemia and infarction evidences in macroscopic and histological examination of the heart,brain,kidney and spleen.ConclusionFW-II axial blood pump can be used to assist left ventricular circulation for 2 weeks with a satisfactory antithrombosis property.The level of platelet activation and leukocyte-platelet aggregation can be reduced to a lowest level at an optimized pump rotating speed.

AIM: To study the basic mechanism of transcriptional regulation, NKX3.1 gene promoter was cloned and its promoter activities in prostate cancer cell lines and other cancer cell lines were tested. METHODS: 1.04 kb - promoter fragment of NKX3. 1 gene was obtained by PCR and cloned into pGL3 - basic and pEGFP - 1 that are promoter - less reporter vectors to examine its promoter activity driving the reporter gene transcription. The promoter activity was determined by dual -luciferase reporter assay and the expression of GFP reporter observed under fluorescence micro scope. RESULTS: The sequence of the cloned 1.04 kb promoter proved to be correct by DNA sequencing. The dual - lu ciferase reporter assay (M1/M2) showed that the promoter activity in LNCaP cell transfected with pGL3 - 1.04 kb promoter was about 1.5 - fold higher than that of pGL3 - control transfection and 50 - fold higher than that of pGL3 - basic transfec tion. To investigate the 1.04 kb - promoter activity in different tumor cell lines, the constructed pGL3 - 1.04 kb promoter and pEGFP - 1.04 kb promoter were transfected into several cell lines, respectively. The results showed that the activity of 1.04 kb promoterin LNCaP was highest among the tested cell lines. Multiple consensus sequence elements have been iden tified within the 1.04 kb fragment using TRANSFAC database. Further experiments will be done to determine their founc tions. CONCLUSION: Cloned 1.04 kb fragment upstream of NKX3.1 gene presented a strong promoter activity and its activity was highest in LNCaP cell among the tested tumor cell lines.

AIM: To study the basic mechanism of transcriptional regulation,NKX3.1 gene promoter was cloned and its promoter activities in prostate cancer cell lines and other cancer cell lines were tested.METHODS:(1.04 kb)-promoter fragment of NKX3.1 gene was obtained by PCR and cloned into pGL_3-basic and pEGFP-1 that are promoter-less reporter vectors to examine its promoter activity driving the reporter gene transcription.The promoter activity was determined by dual-luciferase reporter assay and the expression of GFP reporter observed under fluorescence microscope.RESULTS: The sequence of the cloned(1.04 kb) promoter proved to be correct by DNA sequencing.The dual-luciferase reporter assay(M1/M2) showed that the promoter activity in LNCaP cell transfected with pGL_3-(1.04 kb) promoter was about 1.5-fold higher than that of pGL_3-control transfection and 50-fold higher than that of pGL_3-basic transfection.To investigate the 1.04 kb-promoter activity in different tumor cell lines,the constructed pGL_3-(1.04 kb) promoter and pEGFP-1.04 kb promoter were transfected into several cell lines,respectively.The results showed that the activity of(1.04 kb) promoter in LNCaP was highest among the tested cell lines.Multiple consensus sequence elements have been identified within the(1.04 kb) fragment using TRANSFAC database.Further experiments will be done to determine their founctions.CONCLUSION: Cloned 1.04 kb fragment upstream of NKX3.1 gene presented a strong promoter activity and its activity was highest in LNCaP cell among the tested tumor cell lines.