Aged ; Albuminuria ; drug therapy ; urine ; Diabetes Mellitus, Type 2 ; drug therapy ; urine ; Diabetic Nephropathies ; drug therapy ; urine ; Drugs, Chinese Herbal ; therapeutic use ; Female ; Flavonoids ; Humans ; Male ; Middle Aged ; Phytotherapy

Objective To construct vectors based on adeno-associated virus type 2 (rAAV2) carrying bone morphogenetic protein-7 (BMP-7) and observe their expression in adipose-derived adult stem cells, which can be served as a new gene therapy method and cell source for bone tissue engineering. Methods The coding sequence (1. 3 kb) of BMP-7 was amplified by PCR from the pcDNAl. 1 ( + ) plasmid containing the human BMP-7 cDNA. After purified, the gene fragments were cloned into a plasmid pUC18 and termed plasmid pUC18-hBMP7. The recombinant pUC18-hBMP7 was digested and further ligated to the pSNAV by T4DNA ligase and termed plasmid pSNAV-hBMP7. BHK-21 cells were transfected with the purified pSNAV-BMP7 plasmid according to a standard calcium phosphate precipitation method. The cells were isolated and the integrity of hBMP7 gene was determined by polymerase chain reaction (PCR) using the above PCR primers. To package the virus, stably transfected BHK-21 cells were subsequently infected with recombinant herpes simplex virus type 1 (rHSV-1). The collected cells were processed by chloroform treatment, PEG8000/NaCl precipitation and chloroform extraction for purification. The tiler was determined using quantitative DNA dot blots and the purity was examined by sodium dodecyl sulphatepolyacrylamide gel electrophoresis. Following infection with rAAV2-BMP7 at multiplicities of infection of 1?105 vector genomes per cell and subsequent culture, adipose-derived adult stem cells were assessed qualitatively for BMP7 production. Results The recombinant plasmid pSNAV-hBMP7 was identified by PCR and digested with restriction enzyme. Transfection showed an efficiency of 90 % in ADAS cells. BMP-7 expression in ADAS cells was identified by Western blot. Conclusions The hBMP-7 recombinant adeno-associated virus vectors can be successfully constructed in vitro and rAAV2-hBMP7 can infect ADAS cells.

Relevant literatures on the relationship betw een fluorosis and oxidative stress were reviewed.Based on most of the original papers published in recent years,we can see the increased free radicals and oxidative stress may occur in certain stage of fluoride intoxication,but confirmation of the causality between oxidative stress and fluoride-induced damages still remains much work to do.

BACKGROUND:Skin-derived mesenchymal stem cels may reflect the onset of psoriasis.
OBJECTIVE:To analyze the biological characteristics of skin-derived mesenchymal stem cels in psoriasis patients.
METHODS:Skin-derived mesenchymal stem cels from 30 patients with psoriasis and 20 healthy controls were isolated and cultured by trypsin. Flow cytometry was used to detect the celular immune phenotypes CD34, CD44, CD29, CD45, CD90, CD105, CD73 and HLA-DR. The mesenchymal stem cels were induced by the corresponding cartilage, osteogenic and osteogenic inducing agents, to identify the multi-directional differentiation ability. The cel proliferation curve was plotted at passage 3, and the levels of transforming growth factor-β1 and epidermal growth factor in culture supernatant were detected by ELISA assay.
RESULTS AND CONCLUSION: Under an inverted phase contrast microscope, primary skin-derived mesenchymal stem cels isolated from patients with psoriasis and normal controls both exhibited heterogeneity. In the two groups, CD29, CD90, CD44, CD73 and CD105 were highly expressed, and CD45, CD34, and HLA-DR were lowly expressed. Under certain conditions, skin-derived mesenchymal stem cels were induced to differentiate into adipocytes, osteoblasts or chondrocytes. Proliferation of skin-derived mesenchymal stem cels in the psoriasis group was significantly faster than that in control group, but the final number of cels in the two groups tended to be consistent. The levels of transforming growth factor-β1 and epidermal growth factor in the psoriatic skinhad no correlation with the severity of the disease (P > 0.05). Compared with the control group, the epidermal growth factor level in the cel supernatant was significantly higher in the psoriasis group (P < 0.01), while the level of transforming growth factor-β1 was significantly lower (P < 0.01). These results showed that there is heterogeneity in the morphology of skin-derived mesenchymal stem cels from psoriasis patients, and the biological activity of mesenchymal stem cels is abnormal.

Objective: To observe the changes of endothelial-dependent vasodilatation and cyclophilin A (CyPA) expression at different phases of atherosclerosis in experimental rats. Methods: A total of 30 male Wistar rats were divided into 4 groups: Control group, the rats received normal diet, and 3 atherosclerosis groups, the rats received high cholesterol diet for different period of time and a single dose intraperitoneal injection of Vitamin D3 at 600,000 IU/kg, as Atherosclerosis at 8 weeks (AS 8W) group, AS 12W group and AS 15W group, n=8 in each AS group. The rats were sacriifced at the same time to isolate aorta abdominalis. The expression of CyPA at the wall of aorta abdominalis was detected by HE staining and immunohistochemistry. In addition, the aorta abdominalises was cut into 5 mm rings to observe its response to acethylcholine in exvivo organ bath. Results: With prolonged modeling time, the rats at different groups presented normal vessel structure, endothelial cell damage, vessel smooth muscle cell proliferation, atherosclerosis plaque formation and calciifed plaque formation for differentpathological characteristics. The endothelial-dependent vasodilatation was decreased and the maximum vasodilatation percentage in Control group, AS 8W group, AS 12W group and AS 15W group were at (93.46 ± 2.80) %, (82.58 ± 3.25) %, (61.19 ± 3.72)% and (41.28 ± 2.68)% respectively,P<0.05 between each group. The CyPA expression in endothelial cell and vessel smooth muscle cell were increased accordingly in 4 groups by OD value as (0.25 ± 0.06), (0.34 ± 0.09), (0.53 ± 0.09) and (0.68 ± 0.13) respectively,P<0.05 between each group. Conclusion: The CyPA expression increased and the endothelial-dependent vasodilatation decreased with the progress of atherosclerosis accordingly in experimental rats. The expression level of CyPA is related to atherosclerosis degree and it is one of the initial factors for atherosclerosis in rats.

Objective To design and manufacture a hypobaric cabin simulator for experimental animals.Methods The simulator was composed of a fully transparent working chamber,an electrical chamber,a pressure control component and a program control component,which had its body made of polymethyl methacrylate plastics,the hatch made of metal plate and support by reinforced metal bar.The air inside chamber was exhausted by the diaphragm vacuum pump,the air inflow was controlled by proportional valve,and the inner pressure and the speed of rise and decline were exactly controlled by the balance of inflow and exhaust.Results The new type of hypobaric cabin simulator had the characteristics of accurate pressure control,low fluctuation range and controllable up and down time,low noise as well as adaptable air exchange rate.Conclusion The new type of hypobaric cabin simulator matches the national standard for experimental animals,and can be used in making the animal model of high altitude diseases.

BACKGROUND: At present, bone morphogenetic protein (BMP) is the only cytokine characterized as the ability of osteoblast, and it can promote marrow mesenchymal stem cells (MSCs) differentiating into chondroblast and osteoblast.OBJECTIVE: To investigate the differentiation into osteoblast from MSCs of rabbits transfected by human bone morphogenetic protein-7 (hBMP-7) gene with genetic engineering technique. DESIGN: Single sample observation. SETTING: Department of Orthopaedics, the First Hospital Affiliated to Sun Yat-sen University. MATERIALS: pcDNA1.1/AMP-hBMP7 plasmid.METHODS: The experiment was completed at the Surgery Laboratory of the First Hospital Affiliated to Sun Yat-sen University from May 2004 to March 2005. MSCs of rabbits were cultured with whole bone marrow technique, transfected with pcDNA1.1/AMP-hBMP7 and pcDNA1.1/AMP plasmids respectively in vitro, and left the blank controls. Transcription and expression of transfected genes were detected so as to observe form and growth of cells. Component of calcium content in cytoplasm was measured and osteogenic phenotype was identified with alkaline phosphatase (ALP) staining, calcium salts staining and transmission electron microscopy (TEM).MAIN OUTCOME MEASURES: ① Form and growth of MSCs of rabbits; ② evaluation of expressive product; ③ results of ALP staining (Ca-Co technique); ④ results of chinalizarin staining; ⑤ results of TEM; ⑥ results of scanning electron m icroscopy (SEM); ⑦ assay of osteocalcin.RESULTS: hBMP-7 gene was contained in MSCs after transfection and expressed the relevant mRNA. Cellular form changed a little after expression of amid genes, but growth curve did not changed as compared with that in non-transfected group. Expression of osteocalcin was increased as compared with that in non-transfected group, and ALP staining and calcium salts staining of transfected cells were positive.CONCLUSION: Transfection of hBMP-7 gene can promote MSCs of rabbits differentiating into osteoblast.

Objective To investigate the primary angle-closure glaucoma iris weeks after cutting holes cut the width of the near future, long-term effect relationship, selected after close to long-term efficacy of the forecasting methods to improve iridectomy success rate. Methods 90 cases (116 eyes) with primary angle-closure glaucoma were randomly divided into three groups:A group(the width of the iris-week cut-hole cut 1.5mm) ,30 cases 42 eyes;B group(removal of the width of the iris weeks Ceccon 2 mm) ,30 cases 40 eyes;C group(the width of the iris-week cut-lole cut 3 mm) ,30 cases 34 eyes,after over 1 month,they were followed up to observe the intraocular pressure,anterior chamber angle width of the depth, depending on the features, the iris in recent weeks, as well as cutting holes,long-term concurrency disease. Results A:recent surgery success rate of 85.79% ,long-term surgical success rate of 76. 19% ;B group of recent surgery success rate of 87.5% ,long-term surgical success rate of 82.5% ;C group of recent surgery success rate of94.11% ,long-term surgical the success rate of 88.23%, X2 test the difference was significant(P＜ 0. 05) ;3 postoperative IOP decreased in varying degrees compared with the preoperative differences were significant(P＜0.01) ;peripheral angle after deep Random mean :A group of ≥ 1/3CT,compared the difference between before and after surgery were significantly( P ＜ 0.01 ) ;B group and C group ＞ 1/2CT,2 groups were compared before and after surgery the difference was highly significant( P ＜ 0.05 ). Conclusion Iridectomy was the treatment of primary angle-closure glaucoma safe and effective method of cutting holes. The width of the iris weeks to 2～3 mmwas appropriate, beneficial to control the iris weeks before the adhesion and angle-cut hole shallow narrowing.

Objective To study the effect of down-regulation of E-cadherin on the invasive ability of tumor cells. Methods Human pancreatic carcinoma cell lines JHP-1, PANC-1, and MIA PaCa-2 were selected. The immunocytochemistry, Western blot and invasive-MTT assay were used to observe the expression and the contents of E-cadherin in the tumor cells. Results Immunocytochemistry showed that E-cadherin expression was on cell membrane. The expressions of E-cadherin were preserved in JHP-1, reduced in PANC-1, and showed negative in MIA PaCa-2. Western blot analysis showed that the protein content was the highest in JHP-1 and lowest in MIA PaCa-2. According to invasive MTT assay, the invasive ability of MIA PaCa-2 was the strongest, followed by PANC-1 and JHP-1 (P

Objective To study the values of apparent diffusion coefficient (ADC) changing with time and space in cerebral infarction,and to provide the evidence in defining the infarction stages.Methods 117 work-ups in 98 patients with cerebral infarction (12 hyperacute,43 acute,29 subacute,10 steady,and 23 chronic infarctions) were imaged with both conventional MRI and diffusion weighted imaging.The average ADC values,the relative ADC (rADC) values,and the ADC values or rADC values from the center to the periphery of the lesion were calculated.Results The average ADC values and the rADC values of hyperacute and acute infarction lesion depressed obviously.rADC values in hyperacute and acute stage was minimized,and progressively increased as time passed and appeared as "pseudonormal" values in approximately 8 to 14 days.Thereafter,rADC values became greater than normal in chronic stage.There was positive correlation between rADC values and time[ (174 ±3.47)% vs (58±6.75)%,t =2.03,P ＜0.05 ].The ADC values and the rADC values in hyperacute and acute lesions had gradient signs that these lesions increased from the center to the periphery.The ADC values and the rADC values in subacute lesions had adverse gradient signs that these lesions decreased from the center to the periphery.Conclusion The ADC values of infarction lesions have evolution rules with time and space.The evolution rules with time and those in space can be helpful to provide the evidence in guiding the treatment or judging the prognosis in infarction.