Objective To construct the third-generation self-inactivating lentiviral vectors including BMP-2,BMP-4,BMP-6,BMP-7,BMP-9 and Wnt3a,then integrate them into the genome of mouse embryonic osteoblast cell,MC3T3-E1,and to explore the capability of osteogenic differentiation of individual bone morphogenetic control factor and the effective approaches to further improve the capability of osteogenic differentiation.Methods The plasmid vectors of gene expression including BMP-2,BMP-4,BMP-6,BMP-7,BMP-9 and Wnt3a were constructed,which were identified through enzyme cutting and further confirmed through sequencing.After packing pELNS-BMP-2,BMP-4,BMP-6,BMP-7,BMP-9 and Wnt3a,mouse embryonic osteoblast cell,MC3T3-E 1 was transfected,and the transfection efficiency was confirmed by GFP fluorescence imaging.The expression level of Runx2 mRNA and the transfection efficiency of individual bone morphogenetic control factor was detected by Real time PCR.Eight groups of MC3T3-E1 were dual-gene co-transfected,and the transfection efficiency was verified by GFP fluorescence imaging.ELISA was adopted to detect the expression level of BGP and ALP in MC3T3-E1culture supernatants;Real time PCR was adopted to detect the expression level of Runx2 mRNA;Western blot was adopted to detect the expression level of protein of BMP-2,BMP-4,BMP-6,BMP-7,BMP-9 and Wnt3a.Thus,the effectiveness of osteogenic differentiation of dual-gene co-transfection were evaluated.Results The recombination of lentiviruses,pELNS-BMP-2,pELNS-BMP-4,pELNS-BMP-6,pELNS-BMP-7,pELNS-BMP-9 and pELNS-Wnt3a were successfully constructed.MC3T3-E1 was successfully transfected.The expression levels of Runx2 mRNA were:BMP-2 ＞ BMP-4 ＞ BMP-9 ＞ BMP-7 ＞ Wnt3a ＞ BMP-6.Successful transfection of the dual-gene co-transfection of eight groups of MC3T3-E1 were verified by GFP fluorescence imaging.The expression level of Runx2 mRNA,the expression of BGP and ALP showed BMP-2 and BMP-7 co-transfection group was the most efficient in osteogenesis transfection.Western blot revealed thatthe protein expression of BMP-2,BMP-4,BMP-6,BMP-7,BMP-9 and Wnt3a increased after cotransfection of MC3T3-E1 by BMP-2 and BMP-7.Conclusion The third-generation lentiviral vector,pELNS can lead BMP-2,BMP-4,BMP-6,BMP-7,BMP-9 and Wnt3a into MC3T3-E1,mouse embryonic osteoblast cell,and stabilize its expression.Individual bone morphogenetic control factors can promote MC3T3-E1's differentiation to osteoblasts.The dual-gene co-transfection of BMP-2 and BMP-7 can effectively promote osteoblast conversion,which provides significant theoretical basis and technical support for remodeling of tissue engineering bone.

This paper discussed the roots of American ethical review system, the development of twists and turns, and puts forward the enlightenment to our country, the ethical review system mainly include:perfecting the ethical review at the top of specification design, improve specification status;improve the system of education and training, to strengthen the construction of information communication channels, establish standard operating proce-dures, improve the regulatory system;improve the system of continuous review, formulate specific implementation specification.

Objective To evaluate effects of chronic arsenic exposure and arsenic exposure time on oxidative DNA lesions in humans. Methods A cross-sectional study was conducted in 108 subjects exposed to high concentrations of arsenic in drinking water and 75 control subjects. A cohort study was conducted in 64 subjects exposed to high levels of arsenic in drinking water for 7 or 9 years. Urinary 8-oxo-7,8-dihydredeoxygnanine(8-OHdG) levels were analyzed by the enzyme-linked immunosorbent assay kit(ELISA). Urinary arsenic concentration was detected with hydride generation atomic absorption spectroscopy. Results In the cross-sectional study, the median of urinary arsenic concentration was 484.17 mg/kg Cr for the arsenic-exposed group, and 13.80 mg/kg Cr for the control group, and the difference between the two groups was statistically significant (t=32.57, P<0.01). The median of urinary 8-OHdG levels was 16.60 and 21.88 mg/kg Cr for arsenic-exposed children and adults respectively, much higher than control children(10.50 mg/kg Cr) and adults (9.11 mg/kg Cr), and the difference was statistically significant (t=5.049, 6913, all P<0.01). Urinary 8-OHdG levels were signifieandy lower for children than adults in the exposed group(t=-1.997, P<0.05). In the cohort study, the median of urinary arsenic concentration was 461.3 mg/kg Cr for the 7-year-exposed subjects and 422.90 mg/kg Cr for the 9-year-expesed subjects, and no significant difference was observed(t=-0.250, P 0.05). The median of urinary 8- OHdG levels for 9-year-exposed children and adults were 23.46 and 24.30 mg/kg Cr respectively, significantly increased compared with those of 7-year-exposed(14.29 and 18.38 mg/kg Cr), and the difference had statical signhqcanees (t= -2.949,-3.055, all P<0.01). Conclusions Chronic arsenic exposure can lead to oxidative DNA lesions in humans. The arsenic-induced DNA lesions may aggravate with the exposure time in a certain period.

Objective To investigate the effects and mechanisms of 1,25-Dihydroxyvitamin D3 [1,25(OH)2 D3] on hyperoxia-induced lung injury of neonatal rats.Methods Neonatal Sprague-Dawley rats were randomly divided into air group,hyperoxia group and 1,25(OH)2D3 group within 12 hours after birth,eight in each group.Rats in air group were exposed to air,while those in hyperoxia and 1,25 (OH) 2 D3 group were exposed to hyperoxia (≥85 ％ oxygen concentration).Rats in 1,25(OH)2D3 group were injected with 1,25(OH)2D3 0.5 μg/(kg · d) intraperitoneally once a dayfor seven days,meanwhile the rats in the other two groups received 0.9 ％ saline in the same way.All rats were sacrificed on day 7.Lung tissue sections were HE stained in order to assess lung histological changes and lung radical alveolar count (RAC).The levels of tumor necrosis factor-α(TNF-α),interleukin(IL)-1β and IL-6 mRNA were measured by real time-polymerase chain reaction.Analysis of variance and LSD test were applied for statistics.Results Compared with the air group,the weight of rats in the hyperoxia group was significantly lower on day 7 [(8.48±1.34) g vs (12.51±0.47) g,t=8.05,P＜0.05],while the weight of rats in 1,25(OH)2D3 group [(10.29±1.00) g] was higher than that in the hyperoxia group (t=3.61,P＜0.05).Lung tissue structure was normal in the air group.In the hyperoxia group,inflammatory exudation was observed in pulmonary interstitial,the alveolar size was uneven,and the RAC was lower than that in the air group (5.6±0.1 vs 6.8±0.2,t=21.45,P＜0.05).The RAC in 1,25(OH)2D3 group (6.2±0.1) was significantly increased compared with that in the hyperoxia group (t=11.76,P＜0.05),but still lower than that in the air group (t=9.69,P＜0.05).The expressions of TNF-α,IL-1β and IL-6 mRNA in hyperoxia group (0.0348±0.0006,0.0269±0.0003 and 0.0368 ± 0.0006) were higher than those in the air group (0.0111±0.0007,0.0040±0.0003 and 0.0162 ±0.0007,t=56.54,111.12 and 49.26,P＜0.05,respectively).The expressions of TNF-α,IL-1β and IL-6 mRNA in the 1,25 (OH)2D3 group (0.0203±0.0009,0.0141±0.0004 and 0.0251±0.0009) were lower than those in the hyperoxia group (t=34.44,61.93 and 27.99,P＜0.05,respectively),but higher than those in the air group (t=22.10,49.19 and 21.27,P＜0.05,respectively).Conclusions 1,25(OH)2D3 could attenuate hyperoxia-induced lung injury by inhibiting the expression of inflammatory cytokines.

Objective To evaluate the role of calcium channel in the mechanism of the generation and maintenance of bursting firing of substantia nigra pars compacta (SNc) dopaminergic neurons in rats.Methods Using the patch clamp technique,we observed the firing pattern switching features after adding 10 μmol/L N-methyl-D-aspartic acid (NMDA),compared the changes of whole-calcium current and L-type calcium current with or without NMDA,and analyzed the correlation between the generation of burst firing and L-type calcium channel activation.Results After NMDA treatment,the firing pattern of SNc dopaminergic neurons changed to burst firing,which was compromised by a charastistic high plateau potential and series of action potential on it.The current density of L-type calcium current increased significantly after adding NMDA,which,from (2.86 ±0.26) pA/pF (n =28),significantly increased to (3.75 ± 0.18) pA/pF (n =34 ; t =7.52,P =0.002 8).The high plateau potential was almost abolished with the application of verapamil,a specific antagonist of L-type calcium channel.Consiusion NMDA could induce the firing pattern changed to burst firing in SNc dopaminergic neurons,while L-type calcium channel contributes to the process of generation and maintenance of burst firing.

Objective To investigate and analyze the mineral and bone disorder (MBD) in the patients with chronic kidney disease (CKD),reveal the change of related indexes of CKD-MBD.Methods A cross-sectional study was carried out in the First Affiliated Hospital of Harbin Medical University.From October 2011 to May 2014,1318 inpatients and hemodialysis outpatients were enrolled.Parameters related to MBD,including serum phosphorus (P),total calcium (t-Ca),intact parathyroid hormone (iPTH) and alkaline phosphatase (AKP) were analyzed.Last,it was analyzed with multiple regression analysis to related factors of the secondary hyperparathyroidism (SHPT) in patients with CKD.Results Serum calcium,phosphorus and iPTH had no obvious abnormalities at the early stages of CKD [GFR ＞ 60 ml· min-1· (1.73 m2)-1],and relatively stable before GFR ＞ 30 ml· min-1· (1.73m2)-1.After entering the CKD4 stage,serum phosphorus,iPTH increased sharply and serum calcium decreased obviously along with the decreased glomerular filtration rate (GFR).Serum P,t-Ca and iPTH levels were statistically significant in CKD 1 to 5D patients,respectively,serum P:(1.13±0.20) mmol/L,(1.14±0.22) mmol/L,(1.26±0.23) mmol/L,(1.48±0.34) mmol/L,(2.05±0.61) mmol/L and (2.08±0.58)mmol/L;serum t-Ca (mmol/L) (2.35±0.13) mmol/L,(2.35±0.12) mmol/L,(2.35±0.15) mmol/L,(2.26± 0.18) mmol/L,(2.07±0.29) mmol/L and (2.31±0.26) mmol/L;iPTH:57.8(45.6,91.8) ng/L,54.1(37.8,74.6) ng/L,71.6(45.8,102.2) ng/L,131.1(81.7,205.1) ng/L,277.5(173.6,395.3) ng/L and 354.9 (194.4,720.3) ng/L;The stepwise logistic regression analysis showed:hypocalcemia (OR=3.32,P ＜ 0.01) and decreased GFR (OR=5.28,P ＜ 0.01) were independent risk factors of iPTH elevation at stage CKD3～ 5.Conclusions From the beginning of the CKD3 stage,serum t-Ca,P,iPTH level began to be relatively abnormal as renal function declined.Hyperphosphatemia,SHPT has not been improved significantly in CKD5D stage patients even with hemodialysis.The regulation of hemodialysis on serum calcium showed overcorrecting phenomenon.

AIM :To investigate the association of primary open-glaucoma (POAG), intraocular pressure (IOP) and systemic blood pressure in a rural population aged 50 years old or above in Shaanxi Province, China.METHODS: In the population-based, cross-sectional study, 1 775 (83.53%) residents, aged 50 years old or above, from 3 counties of Shaanxi Province, China, undertook an interview with a standard questionnaire and a detailed eye examination, including logarithm of the minimum angle of resolution visual acuity, slit-lamp biomicroscopy, applanation tonometry and dilated fundus examination. Two blood pressure reading in the sitting position were taken. Gonioscopy was performed if a narrower peripheral anterior chamber (less than one fourth of the corneal thickness) was indicated. Automated visual field testing was performed by participants assessed to have suspicion of glaucomatous disc damage or if IOP was 22mmHg or higher.RESULTS: IOP significantly correlated with systemic blood pressure, and both IOP and systolic blood pressure increased significantly with increasing age. No association between POAG and hypertension was found. The frequency of POAG increased significantly with lower diastolic perfusion pressure.CONCLUSION: Data in our study are accordance with those reported in other population-based studies, and confirm that lower diastolic perfusion pressure is a significant risk factor for primary open angle glaucoma.

AIM: To assess the prevalence and risk factors for age-related macular degeneration (AMD) in a rural population in Shaanxi Province of China.METHODS: A total of 2 835 (81.00%) people aged 40 years old or more, from Fuping county, Jingbian county and Yang county of Shaanxi Province, China, underwent a comprehensive interview and a relative eye examination. The present of AMD was classified into neovascular AMD (NV) and pure geographic atrophy (GA) by using direct ophthalmoscopy for fundus examination according to International Classification System.RESULTS: The prevalence (95% CI) of AMD was 3.00% (2.42, 3.71) in this population, of which NV accounted for 1.45% (1.05, 1.98) and 1.55% (1.14, 2.10)for GA. The prevalence of AMD increased significantly with increasing age (P ＜0.001). AMD was present in 0.47% of participants aged 40 to 49 years, rising to 11.90% of participants older than 80 years, of which the corresponding data increased from 0.28% to 4.76% for NV and from 0.19% to 7.14% for GA. No significant difference was found in the prevalence of NA and GA between genders in this population. With multiple logistic analyses, apart from advancing age, only smoking was found to have a strong association with any type of AMD.CONCLUSION: The prevalence of AMD in the rural population of Shaanxi Province of China is lower than that reported from other population-based studies in different provinces of China, less than that reported in whites, more than that reported in blacks. Except increasing age, smoking is also a significant well-known risk factor for AMD.

· AIM: To assess the prevalence and related risk factors for primary angle-closure glaucoma in a rural population for 40 years of age or older in Shaanxi Province.· METHODS: By using a stratified, cluster-based, random sampling technique, 8 500 persons of all ages were selected randomly, including 3 500 those aged 40 years or more, from the North, the South and the Middle of Shaanxi Province in Western China from July to December in 2003. All participants had an interview with a standard questionnaire and those questions related to glaucoma of previous diagnosis and treatment, family history and outbreak history. Then a detailed and relative eye examination was performed, including logarithm of the minimum angle of resolution visual acuity, external eye examination with slit-lamp biomicroscopy and fundus examination. The intraocular pressure was measured with Perkins applanation tonometry to those persons aged 50 years old or more and those with suspected increased IOP. A further examination was performed to those persons with suspicious glaucoma, including repeated tonometric examination, gonioscopy, dark room test, automated visual field testing, et al.·RESULTS: 6 815 of the eligible 8 500 persons of all ages were interviewed and examined from July to December in 2003, a response rate of 80.18%, including 2835 of the eligible 3 500 persons of 40 years old or more with a response rate of 81.00%. According to Van Herick method, the percentage of shallow peripheral anterior chamber was 13.6%, of which 10.4% was in grade 2, 3.0% in grade 1, and 0.2% in grade 0. In this rural population for 40 years of age or older 31 participants were found to have primary angle-closure glaucoma, with the prevalence of 1.09%. With multiple logistic analyses, the prevalence increased significantly with age (P=0.008), whereas no significant difference (Pgender=0.180, Peducation=0.199) was found in the rate with gender and education, although women and illiteracy seemed to be at increased risk of the disease (OR: 1.77, 95%CI,0.77-4.10; OR: 1.71, 95%CT, 0.76-3.87). Of 31 participants, 21 persons (67.74%) had not been previously diagnosed and treated. Of 10 participants previously diagnosed, 6 (60%) had been received peripheral iridectomy; nevertheless, no any other treatments were performed. 48.39% participants suffered from visual impairment in various degrees, of which the percentage of blindness in either eye accounted for 29.03%. Of those with primary angle-closure glaucoma, 22 persons (70.97%) were classified as having chronic from of the disease.· CONCLUSION: In this rural population in Shaanxi Province of China, the rate of occludable angles is 13.6%, more common than that reported previously in other Chinese populations. The prevalence of primary angle-closure glaucoma is close to that from other Asian populations, increasing with age. A majority of glaucoma was undiagnosed and untreated previously.

· AIM: To assess the prevalence and related risk factors of primary open angle glaucoma (POAG) in a rural population in Shaanxi Province.· METHODS: By using a stratified, duster-based, random sampling technique, 8 500 persons of all ages were selected randomly, from the North, the South and the Middle of Shaanxi Province from July to December in 2003. All participants had an interview with a standard questionnaire and those questions related to glaucoma of previous diagnosis and treatment, family history and outbreak history. Then a detailed and relative eye examination was performed, including logarithm of the minimum angle of resolution visual acuity, external eye examination with slit-lamp biomicroscopy and fundus examination. The intraocular pressure was measured with Perkins applanation tonometry to those persons aged 50years old or more and those with suspected increased IOP. A further examination was performed to those persons with suspicious glaucoma, including repeated tonometric examination, gonioscopy, dark room test, automated visual field testing, et al.· RESULTS: 6 815 of the eligible 8 500 persons of all ages were interviewed and examined from July to December in 2003, a response rate of 80.18%. 9 participants were found to have POAG, with the prevalence of 0.13%, age ranging from 38 to 80 years (mean year:62.0). In those aged 30 years old or more, the prevalence of the disease was 0.23%, 0.28% and 0.39% for those more than 40 and 50 years old, respectively. The rate of suspicious POAG was 0.18% (12 cases) with age ranging from 35 to 77 years (mean year: 54.7). With multiple logistic analyses, the prevalence increased significantly with age (P=0.023). Other than increasing age, myopia was also a strong risk factor for POAG. Of 9participants with POAG, only 2 cases (22.22%) had been previously diagnosed. No one with POAG was received any treatment previously. 66.67% (6 cases) participants with POAG suffered from visual impairment in various degrees secondary to POAG. The percentage of blindness in either eye was 33.33%.· CONCLUSION: The prevalence of POAG is close to that from other Chinese populations, increasing with age. A majority of glaucoma was undiagnosed and untreated previously.