Objective To develop a duplex loop-mediated isothermal amplification(LAMP) assay for rapid detection of Clostridium difficile toxin A(TcdA) and B(TcdB), so as to provide technical support for the rapid detection and toxin typing of Clostridium difficile(CD) infection.Methods According to the conserved sequences of TcdA and TcdB genes, two sets of LAMP primers were designed respectively, and duplex LAMP amplification was performed in the same system. By optimizing the reaction system and reaction conditions, a rapid LAMP detection method for TcdA and TcdB was developed. The specificity and sensitivity of this method were verified, and the melting curves of the LAMP amplified products were analyzed. The 18 stool samples from patients with diarrhea were detected by using the developed method.Results The duplex LAMP reaction conditions for good amplification of both TcdA and TcdB were as follows: Mg~(2+) concentration 6. 8 mmol/L, dNTPs concentration 1. 2 mmol/L, and amplification temperature 64 ℃. The amplification products were preliminarily detected by using SYBR Green I staining, and the target gene(bacteria) showed a bright green fluorescence, while the six other common intestinal pathogens(Escherichia coli, Enterococcus faecium, Enterococcus faecalis, Clostridium perfringens, Bacteroides fragilis,and Clostridium botulinum) exhibited no fluorescence. The sensitivity of the duplex LAMP system for detection of TcdA and TcdB was 10~0 and 10~1 plasmids/reaction respectively, and the exact target toxins contained in CD strains were determined by analyzing the characteristic peaks of the melting curves. Eight of 18 stool samples from patients with diarrhea were positive in preliminary detection, and the melting curve analysis showed that all of them contained TcdA and TcdB.Conclusion A duplex LAMP detection method for TcdA and TcdB has been developed with good sensitivity and specificity, which can be used for the rapid detection of CD infection and can realize the simultaneous rapid detection of TcdA and TcdB.

Objective To observe the effect of fluid resuscitation on hemodynamies and oxygen in severe trauma patients. Methods Parameters of hemodynamic and oxygen metabolism in different fluid volume resuscita-tion in 24 severe trauma patients who were injured on abdominal were measured. Results By fluid resuscitation, as the systolic blood pressure was raised from 80～90 mm Hg(1 mm Hg=0.133 kPa) to 100～120 mm Hg,cardiac in-dex rose from (2.0±0.5 ) L/(min·m2) to (3.2±0.6) L/(min·m2) (P<0.05), systemic vascular resistance index rose from (1857.6±750.2) dyn·s/(cm5·m2) to (3741.5±862.1) dyn·s,/(cm5·m2) significantly (P<0.05). Meanwhile, oxygen delivery index rose from (301.1±74.1) ml/(min·m2) to (554.1±80.0) mL/(min·m2) (P<0.05), and oxygen consumption index also rose (99.7±51.4) mL/( min·m2) to (147.2± 60.1) mL/(min·m2) remarkably (P<0.05), the rate of oxygen expenditure decreased from (33.1±9.1) % to (26.6±8.0) % significantly (P<0.05). Conclusion Large volume resuscitation maybe benefit to the improve-ment of hemodynamics and oxygen metabolism in the emergency resuscitation.

Objective:To analyze data of 5604 cases who received health examination in our hospital ,and understand morbidities of overweight and obesity in these people and their relationship with blood glucose level . Methods :Morning venous blood was taken from the 5604 subjects undergoing health examination to measure blood glucose lev‐el ,and their body weight and height were measured at the same time ,then body mass index (BMI) was calculated and analyzed .Results:Among the 5604 subjects , prevalence rates of overweight and obesity were 42.1% and 13. 3% respectively ,and they were 50% and 16. 4% in men ,and 25. 4% and 6. 8% in women respectively .Among different age groups (except ≥71 years group) , prevalence rate of overweight in men was significantly higher than that of women , P<0.01 all;except ≥61 years subjects , prevalence rate of obesity in men was significantly higher than that of women ,P<0.01 .In overweight group ,risk relative (RR) of impaired fasting blood glucose and dia‐betes mellitus to those of subjects with normal body weight was 2.32 (95% CI 1.91~2.82) and 2.92 (95% CI 2.31~3.70) respectively ;in obesity group , RR of impaired fasting blood glucose and diabetes mellitus to those of subjects with normal body weight was 3.2 (95% CI 2.57~3.99) and 5.17 (95% CI 4.53~5.90) respectively , P=0.001 all . Conclusion:Overweight and obesity have become a common problem among our customer groups .Compared with those with normal body weight ,overweight and obesity greatly increase risk ratio of impaired fasting blood glucose .

Micro RNA (miRNA)were small RNAs encoded by the non-coding RNA genes,which functioned through degrada-tion of mRNA or inhibition of mRNA translation.The miRNA was involved in numerous biological processes,including cell proliferation,apoptosis,development and differentiation.And ectopic expression of miRNA could result in diseases,such as neoplastic hematologic disorder and solid tumor.miR-1 06b-25 cluster contained miR-1 06b,miR-93 and miR-25.These miR-NAs were correlated with the development of tumor.Therefore, we reviewed recent articles on the relationship of miR-1 06b-25 cluster with tumor.

During placental development, oxygen environment is not only critical for trophoblasts migration and invasion, but also fundamental for appropriate placental perfusion. Cysteine-rich 61 (Cyr61, CCN1) was expressed in the extravillous trophoblasts (EVTs) and decreased in preeclampsia. Its regulatory properties in human first-trimester extravillous trophoblast cell line (TEV-1 cells) upon a low oxygen tension were investigated. The present study examined functional changes involved in adaptation to hypoxia of the TEV-1 cells, using cobalt chloride (CoCl(2)) as hypoxic mimic. It was found that hypoxia inhibited growth of TEV-1 cells and induced the increase of cell apoptosis (P<0.05). The Cyr61 expression in human EVTs was transcriptionally induced by CoCl(2). Inappropriate EVTs apoptosis has been implicated in the failure of trophoblasts to fully invade and modify the uterine environment and Cyr61 down-regulation, potentially leading to preeclampsia.

Objective To analyze the ultrasonographic characters in the patients with scalp arteriovenous malformations and evaluate the value of ultrasonography in the diagnosis.Methods The head soft tissue mass,bilateral superficial temporal artery and occipital artery in 9 patients with scalp arteriovenous malformations were examined using high frequency ultrasound.The caliber,systole peak velocity(Vmax),diastole end velocity(Vmin) and resistance index(RI) of the abnormal vessels were recorded.The findings were compared and analyzed with those of the cerebral angiography and ultrasonography in 40 healthy volunteers.Results These lesions displayed subcutaneous hypoecho or no echo region full of reticular or honeycombed darkspace by two dimensional ultrasound,rich signals with flickering and bright mosaic flow by color Doppler and the spectra of high velocity and low resistance by pulsed Doppler.Arterial spectral shape with high velocity and low resistance were found within the lesions and Vmax( 77.7 ? 14.3 ) cm/s,Vmin( 44.3 ? 5.7 )cm/s,RI= 0.41 ? 0.09 .Sixteen feeding arteries with spectra of high velocity and low resistance were checked out,their caliber,Vmax and Vmin were higher and RI lower significantly than those in the control (P

AIM: To investigate the effect of selective silencing of SLC7a8 on uptaking L-dopa in renal tubular epithelial cells of rat (NRK-52E). METHODS: The three siRNAs targeting SLC7a8 (siRNA-1, siRNA-2, siRNA-3) were designed and synthesized. A siRNA with nonspecific coding sequence (siRNA-con) was used for control. All siRNAs were transfected into NRK-52E cells. The siRNA-con transfected group, blank control group and gene-specific silencing SLC7a8 group were set up. The efficiency of transfection was estimated by flow cytometry. The efficiency of RNA interference was detected and screened by RT-PCR preliminarily, and was followed by Western blotting at protein level. The concentrations of L-dopa uptake into the NRK-52E cells were detected by ultraviolet spectrophotometry at different time points (6, 12, 24, 36, 48, 60, 72 and 120 min). RESULTS: The transfection efficiency was 94% detected by flow cytometry. The initial screening of RT-PCR showed that the efficiencies of RNA interference of siRNA-1 and siRNA-3 were higher, and siRNA-3 was the highest at protein level determined by Western blotting. No distinctive change was found between siRNA-con treated NRK-52E and blank control cells. The L-dopa uptake at different time points (6, 12, 24, 36, 48, 60, 72 and 120min) in siRNA-interference group was lower than that in siRNA-con transfected group and blank control group. No significant difference of L-dopa uptake between siRNA-con group and blank control group was observed. CONCLUSION: RNA interference technology selectively down-regulates SLC7a8 expression in rat renal tubular epithelial cells. The L-dopa uptake is also decreased after specifically silencing the slc7a8 expression.

Objective To define individualized internal target volume (ITV) for hepatocellular car-cinoma using four-dimensional (4D) CT, and to compare the differences in target volume definition and dose distribution among 3D, 4D and respiratory-gated plans. Methods 4DCT scanning was obtained for 12 pa-tients with hepatoceUular. Gross tumor volume (GTV), clnical target volume (CTV) and normal tissues were contoured on all 10 respiratory phases of 4DCT images. The 3D, 4D and gated treatment plans were prepared for each patient using three different planning target volumes (PTVs): 1) PTV3D was derived from a single CTV plus conventional margins;2) PTV4D was derived from ITV4D, which encompassed all 10 CTVs plus setup margins (SMs);3) PTV_(Gating) was derived from ITV_(Gating), which encompassed 3 CTVs within ga-ting-window at end-expiration plus SMs. The PTV volume and dose distribution were compared among differ-ent plans. Results The PTV3D was the largest in all 12 patients, but still missed partial target volume in 5 patients when comparing with PTV4D. Both the 4D plans and the gated plans spared more normal tissues than the 3D plans, especially the hver. Without increasing normal tissue dose, the 4D plans allowed for increas-ing the calculated dose from (50.8±2.0) Gy (3D plans) to (54.7±3.3) Gy, and the gated plans could further increase the dose to (58.0±3.9) Gy. Conclusions The 4DCT-based plans can ensure optimal tar-get coverage with less irradiation of normal tissues and allow dose escalation when compared with 3D plans.Respiratory gated radiotherapy can further reduce the target volumes to spare more surrounding tissues, espe-cially for patients with large extent of respiratory mobility.

Objective To retrospectively compare the safety and local efficacy of multiple-electrode switching based radiofrequency ablation (RFA) and the conventional RFA in treating single early-stage hepatocellular carcinoma (HCC).Methods A total of 82 patients with single early-stage HCC received either RFA with a multiple-electrode switching system (n =43) or conventional RFA (n =39) as the first-line treatment.The rate of initial local complete response,major complications and local tumor progression (LTP) were compared between two groups.Results The total ablation time was significantly shorter in the switching-RFA group [(16.7 ± 3.4) mins] than in the conventional RFA group [(29.8 ± 10.4) mins] (P ＜ 0.05).The rate of initial local complete response was 100％ (43/43) in the switching-RFA group and 94.9％ (37/39) in the conventional RFA group (P ＞0.05).After a mean follow-up period of (26.4 ± 21.8)months (ranging 3.0-91.6 months),the rates of LTP in the switching-RFA group and conventional RFA group were 16.3％ (7/43) and 17.9％ (7/39),respectively.The LTP rates in two groups were 16.1％ versus 11.2％ atyear1,and20.5％ versus 20.6％ at year2 (P=0.666).Conclusions The multiple-electrode switching based RFA is safe and effective with shorter ablation time in treating single early-stage hepatocellular carcinoma.

Objective To identify the mutation of human ether-a-go-go-related gene (hERG) and analyze the clinical characteristics of a Chinese family with long ST syndrome (LQTS). Methods The electrocardiogram and DNA samples were obtained from a Chinese LQTS family of 26 members. Genotype was performed with polymorphic short tandem repeat (STR) markers at the known LQT1, LQT2, and LQT3 loci. SSCP analysis was used to find aberrant conformers. hERG mutation was confirmed by cloning and sequencing. Results Three gene carriers were linked to chromosome 7q35-36, where the potassium channel gene hERG was encoded. A 19-base pair deletion was identified. The mutation was located at nucleotide position 1 619-1 637 between transmembrane domains S4 and S5. Furthermore, A1692G polymorphism was found both in the normal control and patients. Conclusion A novel 19 bp deletion mutation of hERG is identified in a Chinese family. All gene carriers are demonstrated to be typical LQT2 ECG phenotype.