BACKGROUND:Construction of tissue engineering adult cardiac myocytes has been a new research hotspot in cardiovascular fields.
OBJECTIVE:To explore a simple, fast method for the separation of adult rat cardiomyocytes, and preliminarily explore the construction methods of tissue engineering adult cardiac myocytes.
METHODS:Segmented enzymatic digestion method was used to isolate cardiac myocytes from adult rats. Subsequently, cardiac myocytes were transfected with adenovirus and liposome-mediated red fluorescent protein gene. Construction efficiency of tissue engineering cells was qualified using inverted fluorescence microscopy and flow cytometry. Final y, cardiac myocytes were transfected with adenovirus-mediated hypoxia-inducible factor-1a, and the expression of hypoxia-inducible factor-1a was examined by western blot analysis.
RESULTS AND CONCLUSION:A lot of cardiac myocytes were col ected using the segmented enzyme digestion method. Flow cytometric analysis showed that the survival of adult cardiomyocytes was (87.03±0.70)%. Compared with liposomal transfection (transfection efficiency was 0), adenovirus infection efficiency was (70.31± 1.39)%, and the cells expressed red fluorescence under fluorescence microscope. After 4 days of adenovirus infection, transfected cells expressed hypoxia-inducible factor-1a protein. These results showed that segmented enzyme digestion is a fast way to isolate adult cardiac myocytes, and recombinant adenovirus vector is a good vector to transfect cardiac myocytes.

Objective To explore the effect of psychotherapy in patients with methadone maintenance treatment.Methods 60 patients accepted drug and 69 patients accepted drug with psychotherapy treatment were evaluated with Self-Rating Anxiety Scale(SAS) and Self-rating Depression Scale(SDS) in the beginning of psychotherapy therapy and 3 months later.All of them were assessed for heroin use at 3 months after the end of a 3-month treatment period.Results SAS、SDS did not show any significant differences between the two groups in the beginning(P ＞0.05).After 3 months of psychotherapy,the differences in SDS and SAS between the two groups showed statistically significant(t =- 2.858,- 2.465,all P ＜0.05).After 2 months of psychotherapy,the number of relapsing again patients of group psychotherapy treatment was significantly lower than control group(x2 =6.906,5.054,5.100,4.247,all P ＜0.05).Conclusion The mental healthy status of patients with heroin dependence is poor.Psychological treatment plays an important role in short-term prevent hernoin addicts from relapse again.

Objective The experiment aims at probing the best condition of the isolation and purification of rat islets.Methods The islets were isolated from rat pancreas by hepatopancreatic duct perfusing with collagenase and purified with Ficoll 400 discontinuous density gradient centrifugation.Then the purified islets were subjected to histological staining,electron microscopy and radioimmunoassay for identification of specificity and viability.Results The histological staining revealed that the viability and the purity of the purified islets were above 95％and 85％respectively.Electron microscopy showed that the purified islets were morphologically intact with clear membrane and plenty of secreting granules.Radioimmunoassay demonstrated the secreted insulin concerntration between low-glucose groups and high-glucose groups varied significantly,which verified the good function of the islets.Conclusion Hepatopancreatic duct perfusing with collagenase is a good method for digestion.There aye many factors that influence the quantity and quality of the acquired islets,such as the completed expansion of pancreas,the concentration and viability of collagenase and the digested time,etc.

Objective The experiment aims at probing the best condition of the isolation and purification of rat islets. Methods The islets were isolated from rat pancreas by hepatopancreatic duct perfusing with collagenase and purified with Ficoll 400 discontinuous density gradient centrifugation.Then the purified islets were subjected to histological staining,electron microscopy and radioimmunoassay for identification of specificity and viability. Results The histological staining revealed that the viability and the purity of the purified islets were above 95% and 85% respectively.Electron microscopy showed that the purified islets were morphologically intact with clear membrane and plenty of secreting granules.Radioimmunoassay demonstrated the secreted insulin concerntration between low-glucose groups and high-glucose groups varied significantly,which verified the good function of the islets.Conclusion Hepatopancreatic duct perfusing with collagenase is a good method for digestion.There are many factors that influence the quantity and quality of the acquired islets,such as the completed expansion of pancreas,the concentration and viability of collagenase and the digested time,etc.

To study the clinicopathological and immunohistochemical characteristics of omental and mesenteric stromal tumor, 8 cases of stromal tumor were investigated, and followed up. The results showed that these tumors were solitary nodule, most were large. Histologically, cytoplasmic eosinophilia and perinuclear vacuolization were often seen in omental and mesenteric stromal tumor of spindle cell type. 3 cases showed epithelioid cytologic features. An immunohistochemical study showed that all the patients were C kit positive (100%),7 were CD34 positive, whereas 6 were SMA positive, positive expression of p53 and PCNA was found in 2 cases of malignant stromal tumor, Lower expression of PCNA was observed in 1 case of benign one.The results suggested that omental and mesenteric stromal tumors have distinct clinicopathological features, the expression of C kit and CD34 plays an important role in its diagnosis.

Objective To observe the pulmonary X-ray changes of malignant malaria.Methods The chest routine X-ray examination in 86 cases with malaria in company with respiratory tract symptoms such as fever was performed,the pulmonary changes were obsenved.Results Of 86 cases, 58 cases were abnormal on chest filmes,including lung framework marked in 33 cases,miliary,small or large patchy shadow in 24 cases and multiple round opaque mass lesion in one case.Conclusion The pulmonary X-ray changes of malignant malaria are varied,the common characteristic is that most of them are situated at middle and lower portion of the lungs.This may attribute to blood dynamic abnormality resulting from malaria protozoan and the pathological changes of involved tissues.

Objective: To explore the sensitivity of Kit or PDGFRA mutants related to gastrointestinal stromal tumor (GIST) to Gleevec.Methods: The recombinant plasmids of KIT Del559-560, KIT Ins IPYD579, PDGFRA D842V and PDG-FRA L839P gene mutants were transiently transformed into the CHO cells by liposome methods.Western blot was used to detect the expression of the related protein and their phosphorylated forms after the cells were incubated with Gleevec for 90 min.At 72 hours after incubation with Gleevec, MTT was used to detect cell proliferation.Results: Western blot results showed that Gleevec at 0.1 μM can notably reduce phosphorylation of KIT Del559-560.Gleevec at 1μM completely blocked phosphorylation of KIT Ins IPYD579 and PDGFRA L839P, but did not affect PDGFRA D842V phosphorylation.MTT analy-sis indicated that growth of CHOPDGFRA L839P was inhibited by Gleevec at 1μM, however, CHOPDGFRA D842V was re-sistant to Gleevec at 5 μM.Conclusion: Gleevec can decrease the expression of phosphorylated protein CHOPDGFRA L839P and CHOKIT Ins IPYD579, and can remarkably inhibit the proliferation of cells containing PDGFRA L839P mutant.

ObjectiveTo study the influence of EmbryoGlue on the implantation of embryo and pregnancy outcome in vitro fertilization (IVF)-embryo transfer (ET).Methods From August 2010 to January 2011,243 infertile patients in Reproductive Medical Center of Tianjin Medical University General Hospital who underwent IVF or intracytoplasmic sperm injection (ICSI) were divided into two groups,including 129 cases used EmbryoGlue as the embryo transfer medium in experimental group and 114 cases used G-2 as embryo transfer medium in control group.Pregnancy outcome were compared between two groups.Results (1) The female age,IVF/ICSI constituent ratio,previous failure cycles and infertile factors of patients did not show statistical difference between experimental group and control group (P ＞0.05 ).(2) The implantation rate of women in experimental group increased significantly compared with the control group [ 30.4％ ( 85/280 ) vs.18.8％ ( 48/255 ),P ＜ 0.05 ] ; Clinical pregnancy rate increased significantly compared with the control group [48.8％ (63/129) vs.34.2％ (39/114),P ＜0.05] ; Multiple pregnancy rate increased significantly compared with the control group [ 34.9％ (22/63) vs.20.5％(8/39),P ＜ 0.05 ] ; Ectopic pregnancy rate decreased significantly compared with the control group [ 4.8％(3/63) vs.17.9％ ( 7/39 ),P ＜ 0.05 ].ConclusionEmbryoGlue can facilitate embryo implantation in IVF-ET and reduce the occurrence of ectopic pregnancy.

Objective To investigate the expression of nuclear factor kappaB(NF-κB) and regulated upon activation normal T cell expressed and secreted chemokine(RANTES) during the formatiom of ascending aortic aneurysm. Methods Forty Wistar rats were randomly divided into the control group(n=20) and the experimental group(n=20).The rat models were made by ligating the ascending aorta. The ascending aortas were taken after ligation for 3months. Immunohistochemistry staining was performed to detect the protein expression of NF-κB and RANTES. The expression of NF-κB and RANTES mRNA were detected by RT-PCR. Results Immunohistochemisry staining results showed NF-κB and RANTES expression significantly increased in aneurysm, while there was a little positive staining in the control group. RT-PCR results indicated that the expression levels of NF-κB and RANTES in the aneurysm were stonger than that of the control group. The expression of NF-κB and RANTES mRNA were remarkably correlated. Conclusion The expression of NF-κB and RANTES in ascendin aortic aneurysm are stronger than that in the control. NF-κB and RANTES may contribute to the pathogenesis of the ascending aortic aneurysm.

A method was developed for the determination of furocoumarin compounds(psoralen, oxypeucedanin, imperatorin, isoimperatorin) in traditional Chinese medicine samples by dispersive liquid-liquid microextraction(DLLME) coupled with high performance liquid chromatography and the relation of sample solution volume, sediment phase volume with enrichment factors was deducted. Some important parameters that influenced the extraction efficiency were optimized: 50 μL carbon tetrachloride and 300 μL acetonitrile were rapidly injected into 1.5 mL sample solution containing 2.5%(w/V) NaCl. After centrifugation at 3500 r/min for 3 min, the sedimented CCl_4 phase was pumped and recorded the volume, then dissolved with methanol and injected into the HPLC for analysis. Under the optimum conditions, a good linear relationship was obtained in the range of 0.006 6.00 mg/L of both psoralen and oxypeucedanin and 0.006-12.0 mg/L of both imperatorin and isoimperatorin. The limits of detection were 1.0-3.0 μg/L(S/N=3). The relative standard deviations were 2.3%-5.4%(n=5). The enrichment factors ranged from 12.6 to 38.5 folds. The average recoveries ranged from 97.5% to 114.8%. The four kinds of furocoumarin compounds in Radix Angelicae dahuricae and Yuanhu Zhitong tablet were determined by the proposed method with satisfactory results.