Genomics ; methods ; Humans ; Microbiology

Background Stickler syndrome is a genetic connective tissue disorder that affects the ocular,skeletal,orofacial and auditory systems.To determine the gene mutation loci can offer a basis for genetic diagnosis and management of Stickler syndrome.Objective The aim of this study was to research the clinical characteristics of a pedigree with Stickler syndrome and identify the disease-causing gene mutation.Methods This study was approved by Ethic Committee of Peking Union Medical College Hospital.The clinical study and pedigree analysis were performed in one family with Stickler syndrome type Ⅰ (STL Ⅰ).Nine family members were examined with informed consent.The entire coding regions of COL2A1 gene with flanking intronic regions were amplified by PCR and directly sequenced.The detected sequence change was confirmed to be mutationloci by examining whether they existed in normal control individuals.Mutant proteins were predicted with online software.Results There were 4 generations and 11 members in this family,and 2 members died,including 1 patient.Three patients were found in 9living families.Inheritance of this family complicd with an autosomal dominant inheritance mode.All affected individuals showed the consistent phenotypes with STL Ⅰ,including high myopia,membranous vitreous anomaly and surface central flat,short nose,palatoschisis,etc.Mutation screening of COL2A1 gene revealed that the first base of intron 12 was deleted(IVS12+1G del).Nucleotide sequence analysis showed that this mutation led to the functional abnormal of this gene by forming termination cordon in advance.This mutation occurred in all affected individuals,however,no mutation was observed in any unaffected member or 100 normal unrelated individuals.Conclusions This study identifies a novel splice-site mutation(IVS12+ 1G del)in COL2A1 gene in a Chinese STL Ⅰ pedigree.This is the first report on a mutation in a Chinese STL Ⅰ family.

ObjectiveTo observe the effect of Tongxinfang on coronary collateral formation and stent-restenosis in patients with coronary artery disease after stenting.Methods61 patients with coronary artery disease were randomly divided into the treatment group (n=30), and control group (n=31). Patients of the treatment group were taking Chinese medicine Tongxinfang for six months, but cases of the control group only received basic treatment. Coronary collateral formation, stent-restenosis and ejection fraction (EF) of patients of two groups were assessed through aniograms and echocardiography before treatment and after six months.ResultsCoronary collateral formation and EF of patients of the treatment group improved significantly compared with cases of the control group (P<0.01 or P<0.05), but the minimal lumen diameter and percent of diamter stenosis were not significantly different between two groups (P>0.05).ConclusionTongxinfang can improve coronary collateral formation and EF.

ObjectiveTo observe the effect of Yiqihuoxue recipe on left ventricular remodeling after acute myocardial infarction (AMI).Methods40 patients with AMI were randomly divided into treatment group and control group with 20 cases in each group and received Yiqihuoxue recipe or fasinopril respectively for 6 months. Changes of clinical symptoms, the level of plasma angiotensin Ⅱ(ATⅡ), aldosterone (ALD), endothelin (ET) and echocardiographical indexes: left ventricular end-diastolic volume index (LVEDVI), left ventricular end-systolic volume index (LVESVI), ejection fraction (EF), left ventricular mass index (LvmassI), wall motion index (WMI), wall thick (WT), E/A were assessed before discharge, and in the end of 3rd and 6th month after AMI.ResultsClinical symptoms of patients of treatment group improved significantly (P<0.01). EF and E/A of all patients in two groups increased (P<0.01 or P<0.05), LvmassI and WT reduced (P<0.05), but there were no significantly differences between two groups in LVEDVI, LVESVI, EF, LvmassI, WMI, E/A and WT(P>0.05). The level of plasma ET decreased in treatment group (P<0.05), the level of plasma ATⅡ and ALD of control group decreased more than that of treatment group (P<0.05).ConclusionYiqihuoxue recipe can significantly improve clinical symptoms, cardic function, and left ventricular remodeling, showing a better clinical efficacy.

BACKGROUND: The pathological characteristics of pulmonary interstitial fibrosis are the proliferation of a large number of fibroblasts and the increasing deposition of matrix collagen that takes the place of normal lung structure. Fluvastatin can inhibit the proliferation of fibroblasts and many other cells.OBJECTIVE: To investigate the effects of fluvastatin in inhibiting the proliferation of rat lung fibroblasts cultured in vitro and its influence on bleomycin-induced pulmonary fibrosis and ventilation function.DESIGN: A randomized controlled trial.SETTING: Department of Respiratory Diseases, Xijing Hospital, Fourth Military Medical University of Chinese PLA; Teaching and Research Section of Pathology, Department of Basic Medicine, Fourth Military Medical University of Chinese PLA; Research Institute ofOrthopedics, Xijing Hospital,Fourth Military Medical University of Chinese PLA.PARTICIPANTS: The study was conducted in the laboratory of Department of Respiratory Diseases, Xijing Hospital of Fourth Military Medical University of Chinese PLA from January to December 2001. Thirty-one healthy adult male Sprague-Dawley(SD) rats of grade Ⅰ were selected in this study.INTERVENTIONS: The fibroblasts derived from the lung normal of one rat were cultured in vitro in media containing fluvastatin. The effect of fluvastatin on the growth curve and the effect of its different concentrations(0, 1 × 10-7,1 ×10-6, 1 ×10-5, 1 ×10-4, 1 ×10 3and 1 ×10-2 mol/L, fluvastatin of 0 mol/L was taken as the blank control group) in inhibiting the cultured cells were observed with MTT colorimetry. The effect of fluvastatin on the division index of the fibroblasts was analyzed by direct cell counting Hydroxyproline colorimetry was used to detect the influence of fluvastatin on the collagen secretion in the media. The other 30 SD rats were divided into six groups: normal control group, bleomycin-induced group and fluvastatin-treated groups(TH 1,TE1, TH15 and TL15 groups) named according to the date of giving fluvastatin,i. e. the 1st day and the 15th day, after the rats were given bleomycin A5. All the rats were killed 28 days later. The number of fibroblasts, the thickness of alveolar wall and the area of mesenchyma in lung tissue were measured by HE staining. The extracellular matrix and collagen in lung tissue were observed by Masson and sirius red staining, and hydroxyproline in lung tissue homogenates was measured.MAIN OUTCOME MEASURES: Fibroblast growth curve and division index of rat lung, hydroxyproline in the media and lung tissue homogenates,number of fibroblasts and the thickness of alveolar wall, the area of mesenchyma, extracellular matrix and collagen contents in lung tissue.RESULTS: Fluvastatin could inhibit the proliferation of the rat lung fibroblasts cultured in vitro(t=4.20 to 17.52, P < 0.01), and its inhibitory effect was increased with the increased dose of fluvastatin, which showed a dose-dependent effect. The 1 × 10-4 mol/L fluvastatin could completely inhibit the proliferation of the cultured cells, and the A490 value from the 2nd day on the fibroblasts by MTT colorimetry was not insignificantly different from those on the 1st day( P > 0.05) . The division index of the fibroblasts and secretion of collagen were obviously decreased by fluvastatin( t = 8. 037,P <0.01; t =3.99 to 10. 84, P <0.05 or P <0.01). In vivo, the number of fibroblasts, the thickness of lung alveolar wall, the area of mesenchyma and the content of hydroxyproline in lung tissue were significantly higher in bleomycin group than in control group( t =4. 62 to 11.93, P < 0. 01), while those in the fluvastatin-treated groups were lower than those in bleomycin group in different degrees( t = 2.69 to 7.65, P < 0.05 to 0.01 ) . The distribution of extracellular matrix and types Ⅰ and Ⅲ collagen in lung tissue were obviously increased in bleomycin group as compared with that in control group, but decreased in different degrees in fluvastatin-treated groups.CONCLUSION: Fluvastatin can significantly inhibit the proliferation of rat lung fibroblasts in vitro, suggesting that it may be an effective drug for pulmonary fibrosis. Treatment at earlier stage is more effective than at advanced stage.

Objective To study the expression of thymic stromal lymphopoietin(TSLP) and the activation of DCs in OVA-induced murine asthma model, and investigate the effects and underlying mecha-nisms of TSLP on lung inflammation. Methods Thirty BALB/c mice were randomly divided into control group, OVA group and TSLP neutralizing antibody treated group. The asthma model was evaluated by airway responsiveness and histological analysis of lung tissues ; The levels of TSLP mRNA in lungs were determined by quantitative real-time PCR; The expression of TSLP in lungs were determined by immunohistochemistry and Western blot; The expression of CD40, CD80, CD86 in BALF was detected by FACS. Results Both the histological analysis of lung tissues and the airway responsiveness were all consistent with the characteris-tic of murine asthma model. The expression of TSLP and TSLP mRNA in the OVA group was significantly in-creased compared with blank group. The expression of CD40, CD80, CD86 in BALF from OVA group was increased significantly compared with the control group. Furthermore, treating mice with TSLP neutralizing antibody reduced the expression of CD40, CD80, CD86 on dendritic cells, and IL-4, IL-5, IL-13 in the OVA group. Conclusion Our study indicate that TSLP was highly expressed in the bronchial epithelia of murine asthma model, via upregulation of CD40, CD80, CD86, induce DCs to active CD4~+ T cells and pro-duce type 2 responses, so that aggravating the lung inflammation of asthma. Blocking TSLP is capable of in-hibiting the production of Th2 cytokines, thus presents a promising strategy for the treatment of asthma.

Objective:To explore the possibility of using peritoneal alternatively activated M2 macrophages to prevent rejection after islet allotransplantation in a murine model.Methods:Peritoneal monocytes from C57BL/6 mice were induced and modulated to M2 and M0 macrophages in vitro,then the phenotype of macrophage was assessed by flow cytometry.C57BL/6 mice were induced diabetic by streptozotocin (STZ) injection and transplanted with islets isolated from BALB/c mice under the left kidney capsule.The recipients were randomly divided to 3 groups (n=8).A total of 2.5× 106 M2 macrophages were injected intravenously at 0,3,7 d after transplantation in islet+M2 group;2.5×106 M0 macrophages were injected intravenously at 0,3,7 d after transplantation in islet+M0 group;the mice in islet+PBS group were injected with PBS.Blood glucose was monitored after transplantation.On day 10 after transplantation,2 recipients in each group were randomly selected and sacrificed,and the left kidneys were resected for pathological examination.Results:Achievement of euglycemia was significantly prolonged after islet transplantation in the islet+M2 group than that in the other two groups (P＜0.01).The median survival time of islet allografts in the islet+PBS group,the islet+M0 group,and the islet+M2 group were 6.5 (4-10),7.5 (4-10),and 24(＞ 15) d,respectively.Pathological examination also showed that the grafts in islet+M2 group remained an intact structure with positive insulin stain and no apparent lymphocytes infiltration,while the graft was rejected in other 2 groups with negative insulin stain and massive lymphocytes infiltration.Conclusion:Peritoneal alternatively activated M2 macrophages can prevent rejection after islet allotransplantation,prolong the survival time of islet allografts and enhance the tolerance of the recipient to blood glucose in mice.

In this paper, three spoilage organisms were separated from five transmutative soy milks, and all the three spoilage bacteria could survive condition of both 1?105Pa,30min and 300mg/kg Nisin. Morpha character, physiological and biochemical characteristics, and a phylogenetic analysis based on 16S rDNA gene sequences reveal that these three strains are Bacillus licheniformis, Bacillus pumilus and Brevibacillus borstelensis respectively. GenBank accessions for these three strains are EF439666-EF439668。

Objective Through detecting CD4+CD25+ T regulatory cells(Treg)in the peripheral blood in children suffering autoimmune diseases and normal controls to learn about the changes of Tregs during the diseases and to acquire some references for clinical diagnosis and treatment.Methods The data were reviewed for CD4+CD25+ Treg cells of the 93 children diagnosed as pediatric autoimmune disease in Children′s Hospital Affiliated to Capital Institute of Pediatrics from Nov.2007 to Jun.2008.Thirty-five normal children in the contempora-neous physical examination were selected as the control group.The percentage of CD4+CD25+ Treg cells and CD4+ T cells to the total T cells were determined by flow cytometric method.Data of the JRA group(22 cases),SLE group(12 cases) and HSP group(12 cases),which were the first three according to the number of cases,were respectively compared with the controls.Independent-samples t test was performed for a statistic analysis with SPSS 11.5 software.Results 1.The percentages of CD4+CD25+ Treg cells to the total T cells and CD4+ T cells in the autoimmune diseases children[(6.14?3.21)% and(21.85?11.68)%,respectively] were both higher than those in the control group[(3.68?1.02)% and(12.83?3.61)%,respectively Pa

Objective：This study was designed to investigate the apoptosis of HL 60 cells induced by the peptide from hemocyte of horseshoe crab (horseshoe crab hemocyte peptide). Methods： Cytotoxicity and cell viability were assayed by MTT method. Morphologic assessment of apoptosis was performed with fluorescence microscope; the apoptosis of the cells was confirmed by flow cytometry; cell cycle was analyzed by flow cytometry, changes of cell membrane were observed by scanning electron microscope. Results： HL 60 cells treated with horseshoe crab hemocyte peptide were showed apparently cytotoxicity with IC50 24 μ g/ml. The morphologic changes including reduction in volume, nuclear chromatin condensation, fluorescence strength were observed with fluorescence microscope. Treated with horseshoe crab hemocyte peptide 50- 100 μ g/ml for 6 h, apoptostic HL 60 cells were predominant. However the HL 60 cells were shown necrotic and apoptostic cells were reduced when treatment with horseshoe crab hemocyte peptide of 200 μ g/ml. SubG1 peak was detected by flow cytometry. Cell cycle analysis showed Phase G1 cells decreased and Phase G2 cells increased with horseshoe crab hemocyte peptide 50 μ g/ml treating for 0- 12 h. Ratio of apoptosis increase depended on concentrations of horseshoe crab hemocyte peptide in 25- 100 μ g/ml but it reduced in 200 μ g/ml, the HL 60 cells were shown necrotic mainly. Scanning electron microscope showed membrane of HL 60 cells was damaged by horseshoe crab hemocyte peptide treatment,cell membrane showed some holes. Conclusion： Peptide from hemocytes of horseshoe crab can induce apoptosis in HL 60 cell,it was early event,and may correlate with membrane damage. The HL 60 cell in Phase G1 seemed to be sensitive to the peptide.