OBJECTIVETo analyze the expression of foreign gene in the filial generation of the transgenic plants on the base of the original transgenic tomatoes seeds carrying the gene encoding saliva-binding region (SBR) in PAc of Streptococcus mutans (S. mutans) gained.

METHODSThe tomatoes total DNA was extracted by CTAB methods, and the filial generation transgenic tomatoes carrying the gene encoding SBR in PAc of S. mutans were selected by PCR. The tomatoes total RNA was extracted by trizol and the transcription of the foreign gene was analyzed by RT-PCR. Protein was extracted from fruit tissue and the content of the total protein was determined by Bradford's methods G250. The expression of foreign protein was analyzed by Western blot and the lever of the foreign protein was analyzed by ELISA.

RESULTSThe fragment encoding SBR in S. mutans PAc gene integrated in the tomato genomic DNA and was expressed. The foreign protein lever was up to 1.2% of the total soluble protein in tomato fruit tissue.

CONCLUSIONThe foreign protein gene in the filial generation of the transgenic plants could express the foreign protein.

Objective To investigate the clinical efficacy and safety of salmeterol xinafoate/ fluticasone propionate in the treatment of chronic obstructive pulmonary disease(COPD).Methods A total of 70 patients with COPD were randomly divided into control group and treatment group,35 cases in each group.Patients in the two groups were all given regular treatment.Patients in the control group were given inhalation treatment with 18 μg tiotropium bromide powder inhalation,qd.Patients in the treatment group were given 50 μg salmeterol fluticasone propionate powder,qd.Patients in the two groups were treated for three cycles (28 days per cycle).The clinical efficacy,pre-and post-treatment of forced vital capacity (FVC),one second forced expiratory volume (FEV1),peak expiratory flow rate value(PEF),FEV1 / FVC,the predicted percentage of FEV1 and PEF value,St.George Hospital Respiratory Questionnaire (SGRQ),multi factor grading system (BODE:body mass index,airflow obstruction,dyspnea and exercise capacity) index score and the incidence of adverse reaction were compared between the two groups.Results After treatment,the total clinical effective rates in the treatment group and the control group were 91.43％ (32/35 cases) and 77.14％ (27/35 cases),respectively,with statistically significant difference (P＜0.05).After treatment,the FEV1 in the treatment group and the control group were(1.86 ± 0.19)L and (1.42 ± 0.17) L,respectively.The FEV1/FVC were (42.93 ± 4.25) ％ and (39.73 ± 3.68) ％,respectively.The PEF were (3.69 ±0.37)L and(3.11 ±0.35)L,respectively.The FEV1/Predicted value were(48.57 ±4.97)％ and (44.47 ±4.51)％,respectively.The PEF/ Predicted value were (48.92 ±5.05)％ and (41.53 ± 4.55)％,respectively.The SGBQ score were (44.97 ±4.96) and (42.01 ±4.67) points,respectively.The BODE score were (5.98 ±0.59) and (5.67 ± 0.59) points.There were statistically significant differences in all the items above between the two groups(P ＜ 0.05).The adverse drug reactions in the treatment group were headache (1 case),tremor (1 case) and palpitation (1 case),while headache(2 cases),tremor(1 case) and hoarseness(2 cases) were found in the control group.The rates of adverse drug reaction in the treatment group and the control group were 8.57％ (3/35 cases) and 14.29％ (5/35 cases) respectively,and no statistical significance was found (P ＞ 0.05).Conclusion Salmeterol xinafoate/fluticasone propionate was safe and effective in the clinical treatment of COPD.

The aim of this study was to investigate the renal function in 149 patients receiving myeloablative allogeneic hematopoietic stem cell transplantation (allo-HSCT) from June 2005 to June 2010 in our hospital, and analyze the risk factors resulting in kidney insufficiency and experience in diagnose and therapy. The creatinine clearance (CrCL) and serial creatinine level were evaluated before and after allo-HSCT within 100 days and 1 year. Non-radiation conditioning regimens were used for any patients. The acute kidney insufficiency (AKI) was defined as at least a 1.5-fold rise in serum creatinine level after allo-HSCT within the first 100 days. The chronic kidney insufficiency (CKI) was defined as the creatinine clearance < basal level within 3 months to 1 year after allo-HSCT. The results showed that the kidney insufficiency was found in 41 patients, in which the incidence of AKI was 32/149 (21.5%). CsA, amphotericin B (P = 0.025) and ES (P = 0.022) were defined as risk factors for AKI. The incidence of CKI was 18/138 (13%). cGVHD (P = 0.013) and TA-TMA (P = 0.012) were associated with the development of CKI. The 2-year survival was lower in patients with kidney dysfunction than that in patients without kidney dysfunction (39% vs 74.1%, P < 0.001). The main factors resulting in kidney insufficiency were defined as infection (52%), GVHD (20%), TA-TMA (12%) and tumor relapse (12%). It is concluded that kidney insufficiency is an important complication of allo-HSCT. Careful monitoring kidney function, minimizing the use of amphotericin B, prophylaxis and effective treatment of fungal infection, GVHD and TA-TMA may be effective preventive measures to decrease the incidence of kidney insufficiency.
Acute Kidney Injury ; etiology ; Adolescent ; Adult ; Female ; Hematopoietic Stem Cell Transplantation ; adverse effects ; Humans ; Male ; Middle Aged ; Renal Insufficiency ; etiology ; Retrospective Studies ; Risk Factors ; Transplantation, Homologous ; Young Adult

Background The transplantation of limbal epithelial cells cultivated on amniotic membrane is a newly developed treatment for limbal stem cell deficiency. The purpose of our study was to investigate the biological characteristics of limbal epithelial cells and evaluate the effect of transplantation of cultivated human limbal epithelial cells on ocular surface reconstruction in limbal stem cell deficiency rat model. Methods Human limbal cells were isolated and cultivated in vitro. Cytokertins 3, 12, and 19 (K3, K12 and K19) and p63 were detected by immunofluorescent staining or RT-PCR. BrdU labelling test was used to identify the slow cycling cells in the cultures. Limbal stem cell deficiency was established in rat cornea by alkali burn. Two weeks after injury, the rats received transplants of human limbal stem cells cultivated on amniotic membrane carrier. The therapeutic effect was evaluated by slit lamp observation, Hemotoxin and Eosin (HE) staining and immunofluorescent staining.Results On day 7 in primary culture, p63 and K19 were strongly expressed by most cells but only a few cells expressed K3. On days 14 and 21, p63 and K19 were still expressed by a majority of cells, but the expressive intensity of p63 decreased in a number of cells, while the proportion of K3 positive cells increased slightly and some cells coexpressed p63 and K3. RT-PCR showed that gene expression of both p63 and K12 were positive in cultivated limbal cells, but in mature superficial epithelial cells, only K12 was detected. BrdU labelling test showed that most cells were labelled with BrdU after 7 days' labelling and BrdU label retaining cells were observed after chasing for 21 days with BrdU free medium. For in vivo test, slit lamp observation, HE staining and immunofluorescent staining showed that the rats receiving transplant of human limbal stem cells cultivated on amniotic membrane grew reconstructed corneas with intact epithelium, improved transparency and slight or no neovascularization. A majority of epithelial cells of the reconstructed cornea were positive to antihuman nuclear antibody and cells expressing K3 were found mainly in superfacial epithelium.Conclusions Limbal stem cells can be cultivated in vitro: the cells are characterized by high proliferation and slow cycling and identified as p63/K19 positive and K3/K12 negative. During culture, some stem cells can proliferate and differentiate into mature cornea epithelial cells. Amniotic membrane is a suitable carrier for limbal stem cells. Transplantation of human limbal stem cells cultivated on amniotic membrane can functionally reconstruct rat cornea with limbal stem cell deficiency.

Objective To investigate mutation rate and types of KRAS in colorectal carcinoma , and to analyze the relationship between KRAS mutation and clinicopathological parameters in patients with colorectal carcinoma ( CRC) .Methods Scorpions Amplification Refractory Mutation System ( ARMS) fluorescence quantitative PCR was performed to detect the mutations in codons 12 and 13 of KRAS and to correlate between clinicopathological characteristics and the presence of various KRAS mutations of colorectal carcinoma .Results KRAS mutations were identified in 518 patients ( 42.92%) , including G12D ( 197 cases, 16.32%) , G12V ( 125 cases, 10.36%),G12C(40 cases, 3.31%),G12S(29 cases, 2.40%),G12A(21cases, 1.74%),G12R(7 cases, 0.58%),13D(117 cases, 9.69%).Female patients had a higher KRAS mutation rate than male (46.21%, 238/515 vs.40.46%, 280/692, P<0.05 ) .KRAS mutation was significantly higher in right colon cancer (46.45%,131/282)and in rectal cancer(44.50%,255/573)than that in left colon cancer(37.50%,132/352) (P<0.05,P<0.05).Conclusions There are many types of KRAS mutations in colorectal cancer, and many mutation types exist simultaneously .The detection rate of KRAS mutation is higher in female CRC patients than in the males.The detection rate of KRAS mutation is significantly higher in right colon cancer and in rectal cancer than that in left colon cancer.

The clinical value of transvaginal ultrasound in clinical surgical treatment of cesarean scar pregnancy (CSP) was evaluated.The clinical data of 104 patients with CSP admitted at Tongji Hospital from 2013 to 2016 were collected and analyzed retrospectively,including the patients' age,gestational age,the size of gestational sac or uneven mass,the uterine scar thickness,β human chorionic gonadotropin (hCG) levels and so on.Of these 104 cases,30 cases were subjected to laparotomy,29 cases to laparoscopy,27 cases to hysteroscopy,16 cases to ultrasound-guided uterine curettage,and 2 cases to conservative treatment.The transvaginal ultrasound showed that uterine scar thickness and gestational sac or uneven mass size had significant difference (P＜0.05) among different surgical methods by comparatively analyzing the patients' data.It was suggested that transvaginal ultrasound may provide the valuable reference for choosing clinical surgical procedures for CSP.

Objective To explore the predictive value of baseline CD4⁺ cell counts and CD4/CD8 ratio in immune reconstitution among HIV/AIDS patients receiving highly effective antiretroviral therapy (HAART). Methods A total of 90 HIV/AIDS patients who had received over 24 months of HAART regimen and had viral road <50 copies/ml were enrolled in this study. The patients were subgrouped into immunological non-responders (INR_s) (<200 cells/μl), immunological inadequate responders (IIR_s) (200~500 cells/μl) and immunological responders (IR_s) (>500 cells/μl) according to the CD4 cell counts after two-year HAART regimen. The characteristics and dynamic CD4⁺ cell counts among the three groups were compared, and the factors of immune reconstitution were analyzed. Results The CD4⁺ cell counts in the immunological responders (IR_s) were significantly higher than those in the immunological non-responders (INR_s) and immunological inadequate responders (IIR_s) (P<0.001). The analysis of area under the ROC curve reveled that the number of CD4⁺ cells reached the highest predictive value at 1 year of HAART. Conclusion Our findings suggested that early initiation of HAART could reduce the failure of immune reconstitution. The combination of baseline CD4⁺ cell counts and baseline CD4/CD8 ratio may serve as a valid predictor of immune reconstitution prognosis after HAART.

Objective To establish animal models with anterior transposition of the ulnar nerve and evaluate the safety of anterior transposition of the ulnar nerve at molecular level.Methods Location of the ulnar nerve of elbow in 5 rats were found similar to human being by anatomy.Twenty healthy adult SD rats,weighting about 250 g,were performed the anterior transposition of the ulnar nerve in the right forelimbs and the left forelimbs was considered as control group.The bilateral flexor carpi ulnaris muscles were weighed and the slice of cervical spinal cord(C_6-T_1)level were prepared 1 month after the operation.Nissl staining,NADPH-d histochemical staining,IB4 staining and ChAT-immunohistochemical staining were employed to observe the spinal cord(C_6-T_1)level at molecular level;electron microscope was used to observe the ultrastructure of ChAT-positive neurons.Statistical analysis was paired T test.Results The flexor carpi ulnaris muscles in the model group(92.3±9.13mg)and control group(93.2±7.29 mg)were not significantly different(P>0.05).After anterior transposition of the ulnar nerve in rats,no significant differences in cell number and morphology in the cervical spinal cord(C_6-T_1)were found between the model group and the control group(P>0.05).No changes between the 2 groups were noted in the fine structure of anterior horn motor neurons and the expression of nenrotransmitters(P>0.05).Conclusion Anterior transposition of the ulnar nerve can be safely done in the animal models(rats).

Isotemporal substitution model is a powerful tool to explore the real association between physical behavior and health outcomes, which has the potential of the application in large-scale cohort study. This paper systematically introduces the principle of isotemporal substitution model and its implementation method in specific analysis to provide analytical ideas for the epidemiological research related to physical behavior in China. The baseline data of Regional Ethic Cohort Study in Northwest China conducted in Shaanxi province were used to analyze the relationship between physical behavior and cardiovascular disease with single-factor model, partition model and isotemporal substitution model. The advantages and disadvantages of different models were compared, and the advantages of isotemporal substitution model in quantifying physical activity health risk were introduced. Isotemporal substitution model could qualify physical behavior and health outcomes, which has wide application value in epidemiological research.
Humans ; Cohort Studies ; Epidemiologic Studies ; Cardiovascular Diseases ; China/epidemiology*

Objective To investigate the distribution characteristics and risk factors of intracranial and extracranial aterial lesions in Chinese patients with ischemic stroke . Methods In this multi‐center study ,2 310 continuously inpatients with ischemic stroke diagnosed in 20 stroke screening and prevention project base hospitals from June 2015 to M ay 2016 were enrolled . Carotid ultrasonography and transcranial color‐coded sonography or transcranial Doppler were performed in all patients to confirm the presence of cerebral artery stenosis or occlusion . According to the distribution of lesions ,the subjects were divided into 2 groups :the simple intracranial artery stenosis group and the simple extracranial artery stenosis group . T he difference of risk factors between the two groups was compared . Results Of the 2 310 patients with ischemic stroke ,1 516 ( 65 .6% ) had simple intracranial artery stenosis and 794 ( 34 .4% ) had simple extracranial artery stenosis . T he incidence of anterior circulation artery stenosis was higher in the group of intracranial artery stenosis than that in the extracranial artery stenosis group ( 68 .1% vs 48 .7% , P ＜0 .001) . Posterior circulation artery stenosis and combined anterior with posterior circulation artery stenosis were more common in patients with extracranial artery stenosis group than those in intracranial artery stenosis group ( 36 .4% vs 22 .1% ,14 .9% vs 9 .8% ;all P ＜0 .001) . Univariate analysis of risk factors for stroke showed that patients with intracranial arterial stenosis had a higher prevelence of hypertension , diabetes ,obesity ,and family history of stroke ,and their systolic blood pressure ,diastolic blood pressure , body mass index ( BM I) ,fasting blood‐glucose ,glycosylated hemoglobin ,triacylglycerol ,total cholesterol , and low‐density lipoprotein cholesterol were significantly higher than those in the extracranial arterial stenosis group ( all P ＜ 0 .05 ) . T he proportion of elderly ( ≥ 65 years old ) ,male and smokers in the extracranial arterial stenosis group was significantly higher than that in the intracranial arterial stenosis group ( all P ＜0 .05) . Multivariate logistic regression analysis showed that elderly ( ≥65 years old) ,male , and smoking history were independent risk factors for extracranial arterial stenosis ( OR＝ 2 .012 ,1 .637 , 1 .325 ,respectively ;all P ＜0 .05) . While hypertension ,diabetes ,less physical activity ,and high BM I levels were independent risk factors for simple intracranial arterial disease ( OR ＝ 1 .301 ,1 .252 ,1 .248 ,1 .030 , respectively ;all P ＜0 .05) . Conclusions There are significant differences in the distribution characteristics and risk factors of intracranial and extracranial aterial lesions in patients with ischemic stroke in China .