BACKGROUND:Transplantation of exogenous stem cells for functional cardiac celldeath or apoptosis easily induced complications after transplantation. Therefore, cardiac progenitor cells of heart itself have been ideal seed cells. OBJECTIVE:To establish stable celllines of cardiac progenitor cells from mouse heart, and to provide ideal cellmodels for studying proliferation and differentiation factors affecting cardiac progenitor cells during adult myocardial cells were damaged. METHODS:(1) Myocardiocytes were isolated from embryonic 15.5 days mice. (2) The cultured myocardiocytes were immortalized using retrovirus SSR69. Immortalized monoclonal myocardial cells were obtained using antibiotic selection and infinite dilution. (3) Monoclonal cellline with the property of cardiac progenitor cells was screened out. RESULTS AND CONCLUSION:(1) Myocardiocytes were successful y isolated and cultured. Partial cells showed obvious beating. (2) Myocardiocytes infected with retrovirus SSR69 were cloned and got 76 clones, then were named as CP15-#, (3) Screening the first 20 clones, the reasonable clones with the characteristics of cardiac progenitor cells were obtained according to myocardial cellmarker genes. The results suggested that immortalized cardiac progenitor cells were established mediated by reversible SV40 T antigen.

OBJECTIVETo compare the efficacy between elongated needle therapy and regular needle therapy at Tiantu (CV 22) on the basis of xingnao kaiqiao (activiting brain and regaining consciousness) acupuncture therapy so as to explore the effective therapeutic method in treatment of dysphagia induced by bulbar palsy.

METHODSSeventy one cases of dysphagia induced by bulbar palsy were randomized into two groups. The xingnao kaiqiao acupuncture therapy was applied at Shuigou (GV 26), Neiguan (PC 6), Sanyinjiao (SP 6) and the others in the two groups. In the elongated needle therapy group, on the basis of xingnao kaiqiao acupuncture therapy, the elongated needle was used to puncture Tiantu (CV 22). In the regular needle therapy, the regular acupuncture technique was used at Tiantu (CV 22). In both groups, the treatment was given once a day in a week except Sunday and lasted for 4 weeks totally. Before and after treatment, the swallowing condition and the standardized swallowing assessment (SSA) were observed in the patients and the efficacy was compared between the two groups.

RESULTSThe total effective rate was 97.2% (35/36) in the elongated needle therapy group, which was better than 77.1% (27/35) in the regular needle therapy group (P<0.05). After treatment, SSA score was reduced significantly as compared with that before treatment in the two groups (both P<0.05). SSA score in the elongated needle therapy group was reduced much more apparently as compared with that in the regular needle therapy group after treatment (P<0.05).

CONCLUSIONOn the basis of the xingnao kaiqiao acupuncture therapy, the elongated needle therapy at Tiantu (CV 22) achieves the superior effect on bulbar palsy-induced dysphagia as compared with the regular acu- puncture at Tiantu (CV 22).

Acupuncture Points ; Acupuncture Therapy ; Aged ; Bulbar Palsy, Progressive ; complications ; Deglutition Disorders ; etiology ; therapy ; Female ; Humans ; Male ; Middle Aged ; Treatment Outcome

AIM:To differentiate bone marrow mesenchymal stem cells(BMSCs) into functional insulin-producing cells to produce sufficient pancreatic islet cells for transplantation.METHODS:Recombinant adenovirus vectors carrying PDX1 and NKX6.1 genes were constructed and the bone marrow mesenchymal stem cells were infected by the recombinant adenovirus combined with several cytokines for differentiation.The expressions of PDX1,NKX6.1 and insulin and C-peptide in the differentiated bone marrow mesenchymal stem cells were detected by RT-PCR and Western blotting.After the differentiated bone marrow mesenchymal stem cells were transplanted into subrenal capsule of diabetic mice,cell morphology of the grafts as well as their secretion of insulin and C-peptide were detected.Besides,regulating capacities of grafts on the blood glucose level of the diabetic mice were also detected.RESULTS:BMSCs induced by recombinant adenovirus(pAdxsi-CMV-PDX1/CMV-NKX6.1) and several cytokines showed positive dithizone staining and the expressions of ?-cell related molecule such as insulin and glucose transporter 2 were detected by RT-PCR,which showed a sustaining and stable expression.The similar results were showed by Western blotting,immunohistochemical staining and indirect immunofluorescence.The insulin secretions in the cells stimulated with glucose at concentrations of 5.5 and 25 mmol/L in the experimental group were(1 240.4?109.3) mU/L and(3 539.8?245.1) mU/L,respectively,and were significantly higher than those in control group.Moreover,transplantation of the cells to STZ mice in treatment group made serum glucose recover to normal level.CONCLUSION:PDX1 and NKX6.1 gene-modified bone marrow mesenchymal stem cells differentiate into insulin-producing cells in vitro.When these cells transplanted into STZ induced diabetic mice,their serum glucose could return to the normal level and they could live well.Thus this is a promising method for diabetes treatment.

AIM: To construct a recombinant plasmid vector containing human pancreatic duodenal homeobox1(Pdx1) and neurogenic differentiation 1(NeuroD1) genes,and to detect its effective expression in eukaryocytes and the ability to induce differentiation of hepatocytes.METHODS: Using human embryo pancreas mRNA as template,Pdx1 and NeuroD1 genes were amplified by RT-PCR and cloned into the two different multiple cloning sites(MCSA and MCSB) of plasmid pIRES.The recombinant plasmid pI/Pdx1/NeuroD1 was transfected into L02 cells.The expression of Pdx1 and NeuroD1 in transfected cells was detected by immunocytochemistry,IFA,RT-PCR and Western blotting,respectively.RESULTS: The length and sequence of cloned segments were correct.Pdx1 and NeuroD1 were expressed in eukaryocytes.Furthermore,the hepatic cells were induced to express glucose transporter 2(GLUT2) and eukaryocyte transcription factor Nkx6.1,which were functionally correlated to ? cells.CONCLUSION: pI/Pdx1/NeuroD1 plasmid is successfully constructed and expressed in human eukaryocytes,with which the cells express the eukaryocyte transcription factor and GLUT2,indicating the transfected cells functionally correlates to ? cells.The results suggest that Pdx1 and NeuroD1 genes can induce the differentiation the cells from hepatic cells to pancreatic endocrine secretion cells.

Objective:To investigate the prevalence and related risk factors of diabetic retinopathy (DR) among registered residents of type 2 diabetes mellitus in Jiading Industrial Zone Community Health Service Center,Shanghai.Methods:In this cross-sectional study,diabetic retinopathy was screened among registered residents with type 2 diabetes in Jiading Industrial Zone Community Health Service Center,Shanghai.The prevalence and risk factors of DR were evaluated by combining questionnaire and clinical examination.Results:Among 776 subjects who were chosen,695 people completed the study,and the overall response rate was 89.56％.26 cases were excluded because the refractive medium was opaque and the retina could not be seen clearly.DR was detected in 91 patients (13.60％).The mean duration of diabetes in patients with DR and without DR was (9.63±6.44) years and (7.02±5.32) years (OR=1.073,95％ CI 1.036-1.111,P＜0.001),the fasting blood glucose was (8.66±2.66) mmol/L and (7.52±2.15) mmol/L (OR=1.219,95％ CI 1.119-1.328,P＜0.001),glycated hemoglobin was (7.68±1.55)％ and (6.99±1.21)％ (OR=1.495,95％ CI 1.275-1.754,P＜0.001).There was also statistically significant difference in the use of insulin between DR patients and non-DR patients (OR=4.202,95 ％ CI 2.504-7.051,P＜0.001).Conclusions:The prevalence of DR among type 2 diabetes patients is 13.6％ in Jiading Industrial Zone,Shanghai.The longer the duration of diabetes is,the higher the fasting blood glucose is,the higher the glycosylated hemoglobin is,the greater the risk of DR will be.

Objective To track the effects of hyperbaric oxygen (HBO) therapy on neonatal rats with hypoxic-ischernic brain damage (HIBD).MethodsA total of 126 healthy,seven-day-old SD rats were used to model the HIBD and then randomly divided into seven groups of 18:a sham group,a hypoxic-ischemic (HI) group,a 6 h HBO group,a 24 h HBO group,a 48 h HBO group,a 72 h HBO group and a 1 week HBO group.One hour of HBO treatment at 2 atmospheres absolute pressure was administered once daily for a week to the rats in the latter 5groups,starting at 6,24,48,72 hours and 1 week post the HIBD modeling,while no HBO was administered to the sham and HI groups.The effects were assessed in terms of histological changes ( the neuron density and the percentage of neuron apoptosis in the CA1 region of the hippocampus) and nitric oxide (NO),malondialdehyde (MDA) and superoxygen dismustase (SOD) concentrations after 15 days.ResultsIn the 6 h HBO,24 h HBO,48 h HBO and 72 h HBO groups,average neuron density in the CA1 region was significantly higher than in the HI group.But in the 1 week HBO group the average density was not significantly different than in the HI group.In the 6 h HBO,24 h HBO,48 h HBO and 72 h HBO groups the average percentage of neuron cell apoptosis in the CA1 area of the hippocampus was significantly lower than in the HI group,but the 1 week HBO group again showed no significant difference.There were significant differences in average NO,MDA or SOD levels among the 6 h HBO,24 h HBO,48 h HBO,72 h HBO and HI groups,but the 1 week HBO group showed no significantly higher average levels than the HI group.ConclusionsThe optimal therapeutic window for using HBO to protect against HIBD is within the first week.The best effect can be obtained in the first 6 hours,but after 1 week HBO no longer has a significant effect.The earlier the HBO is administered,the better the effect obtained.

Objective To construct the recombinant plasmid containing human filaggrin gene,purify and identify the immunoreactivity of the recombinant protein,and establish the indirect ELISA to detect AFA for diagnosis of RA.Methods The constructed plasmids were transformed into E. Coli Rosettagami(DE3).This fusion protein was purified by NAT chromatography.ELISA coated with the fusion protein Was established to detect the AFA in serum of patients,which included 114 cases of RA,56 cases of SLE,32 cases of OA and 40 cases of normal controls. The correlation between the results of AFA and anti-CCP in RA group were compared. Results 321 bp fragment of filaggrin gene was amplified and the recombinant expression vector pET-28a( + )-filaggrin was constructed. The sequence of filaggrin gene was the same as the sequence reported in the literatures. The Rosetta-gami (DE3) strains of E. Coli with recombinant vector showed high level of filaggrin protein after induction. The SDS-PAGE showed that the plasmid expressed the filaggrin fusion protein with molecule weight of 14 000 Da. The expression protein could be purified by Ni-NAT with activity. The absorbance value of AFA in RA group was 0.473 ±0. 248 while they were 0. 160 0. 088, 0. 121±0. 070, 0.050 0. ±018 in SLE, OA and normal groups respectively. There were significant differences of absorbance values of AFA between RA and SLE, OA, control group (t = 12.004, 14. 464, 18.078, P<0. 01, respectively). The positivities of anti-filaggrin in RA, SLE and OA were 48.2%, 5.4% and 3. 1% respectively. The positivities of AFA were significantly different between RA, OA and normal control groups (x~2 = 67. 088, P < 0. 01). There was positive correlation of results between AFA and anti-CCP antibody (r = 0.42, P < 0. 05 ) . The consistency rate of results between AFA and anti-CCP was 70. 1%. Anti-CCP was negative in 10 out of 114 patients with AFA positive. AFA can be used to diagnose RA with sensitivity of 48. 2% , specificity of 96.9% , positive predictive value of 93. 2% and negative predictive values of 67. 9% . Conclusions The purified human filaggrin fusion protein is successfully purified. The indirect ELISA method based on the recombinant protein shows good sensitivity and specificity. Joint detection with AFA and anti-CCP can improve the positive rate of detection.

OBJECTIVETo compare components in volatile oils of nutmeg and prepared nutmeg.

METHODVolatile oil from nutmeg and prepared nutmeg were extracted by vapor distillation. The chemical components in two kinds of volatile oils were determined and indentified by GC-MS.

RESULTThe change in quantity and quality of components in volatile oils were observed after processing. 13 new components occurred and 4 components disappeared in volatile oils after processing. The contents of methyleugenol and methylisoeugenol that are active ingredients were increased. The contents of myristicin and safrol that are toxic ingredients in volatile oils were decreased.

CONCLUSIONThe processing method of nutmeg by soaking with water and roasting with bran is scientific.

Anisoles ; analysis ; Benzyl Compounds ; analysis ; Dioxolanes ; analysis ; Eugenol ; analogs & derivatives ; analysis ; Gas Chromatography-Mass Spectrometry ; Hot Temperature ; Myristica fragrans ; chemistry ; Oils, Volatile ; chemistry ; isolation & purification ; Plants, Medicinal ; chemistry ; Pyrogallol ; analogs & derivatives ; analysis ; Safrole ; analysis ; Technology, Pharmaceutical ; methods

Tanshinones are the bioactive components of the Chinese medicinal herb Salvia miltiorrhiza, while its biosynthetic pathway remains to be characterized. Rapid identification and characterization of the genes correlated to tanshinones biosynthesis is very important. As one of the intermediates of tanshinones biosynthesis, the ferruginol content is relative low in both root and engineered bacteria. It is urgent to construct an efficient system for conversion of miltiradiene to ferruginol to obtain large amount of ferruginol as the substrates for further identifying other downstream genes involved in tanshinones biosynthesis. In this study, we constructed the whole-cell yeast biocatalysts co-expressing miltiradiene oxidase CYP76AH1 and cytochrome P450 reductases (SmCPR1) from Salvia miltiorrhiza, and then characterized it with RT-PCR. After permeabilization, the yeast whole-cell could catalyze turnover of miltiradiene to ferruginol efficiently through single-step biotransformation with a conversion efficiency up to 69.9%. The yeast whole-cell biocatalyst described here not only provide an efficient platform for producing ferruginol in recombinant yeast but also an alternative strategy for identifying other CYP genes involved in tanshinones biosynthesis.

AIM: To explore a new compute tomography (CT) for the localization of intraocular foreign bodies (IOFBs).METHODS: After CT ocular horizontal and ocular axial scan,the foreign bodies on the eyeball wall or retinal surface in 26eyes were localized by means of the combination of CT ocular axial scan and meridian plane reconstruction (new method),ocular horizontal scan and ocular axial scan (conventional method Ⅰ) as well as ocular horizontal scan right angle coordination (conventional method Ⅱ) separately. According to the criteria of indirect ophthalmoscope localization and direct observation during operation, the relative accuracies of corresponding points of the foreign bodies on sclera surface along meridian and latitude were measured.RESULTS: In the 26 cases, the mean relative accuracies of corresponding points of foreign bodies on sclera surface along meridian and latitude were 1.53mm、1.64mm (new method), 1.37mm、1.64mm (conventional method Ⅰ) and 2.02mm、2.55mm (conventional method Ⅱ) respectively.There was no statistical difference between the new method and the conventional method Ⅰ, whereas there was statistical difference between the new methods and the conventional method Ⅱ (along meridian: P＜0.05; along latitude:P＜0.01).CONCLUSION: Compared with the conventional methods,the new method is simpler, more visible and more potential in the clinic.