BACKGROUND: The method of culture purified sinoatrial node cell is important in investigating its ultrastructural characteristics and autorhythmic mechanisms.However,the corresponding method has not been standardized.OBJECTIVE: To summarize the localization,cultivation and purification of sinoatrial nodes isolated from newborn rabbits,and to study the morphological characters of primary cultured pacemaker cells.METHODS: Hearts of the newborn rabbits(within 24 hours)were embedded in paraffin for hematoxylin-eosin staining,The location of sinoatrial nodes was observed under an optical microscope,the morphology of sinoatrial nodes cells were observed by light microscope and electron microscope.RESULTS AND CONCLUSION: Sinoatrial nodes localized in the anterior wall of the superior vena cava and the posterior laterial wall of right atrium.There was about 0.32 mm between its lowest point and sulcus terminalis.Three distinct types of cells were observed among the cultured cells of sinoatrial nodes: spindle,spider and polygon.The spindle cells occupied the greatest proportion of the cultured cells(59.6±7.3)%.The spontaneous contraction frequency of spindle cells was the highest among the constracting cells(145±9)times per minute.The ultrastructure observation showed that myofibrils and other organelles in spindle cells were sparse and significantly decreased in number compared with triangle cells.There was no significant difference between triangle cells isolated from sinoatrial nodes and from atrial muscle.Sinoatrial nodes could be harvested along the anterior root of the superior vena cava down to the posterolateral sulcus.Among the cultured cells from neonatal rabbit sinoatrial nodes,the spindle cells with small body and fast pulse frequency are pacemaker cells.

OBJECTIVE: To establish a method for determination of the contents of puerarin in infant Fuxieling effervescent granules with HPLC.METHODS:The puerarin was extracted by ultrasonics, using methanol as solvate.The stratographic column was Shim - Pack C1s with acetonitrile - water( 10: 90) as mobile phase. The flow rate was 2.0ml/min and the detecting wavelenghth was 250nm.The temperature of column was 25℃ .RESULTS:The resolution between the puerarin and the adjacent peaks was ＞1.5 .The number of theoretical plates was 18 550 .The standard curve was linear in the range of 0.964μg～7. 712μg .The average recovery was 100.2% (RSD = 1.76%, n = 6).CONCLUSION:The method is simple, rapid and accurate. It can be used for quality control of the preparation.

AIM: To establish the quality standard for Gujing Tablets (Olibanum, Rhizoma Cyperi, Radix Aucklandia, etc.). METHODS: Olibanuln, Rhizoma Cyperi, Radix Aucklandia were identified by TLC, and costuslactone content in Radix Aucklandia was determined by HPLC. RESULTS: The TLC identification was highly specific and spots were clear. Costuslactone in Radix Aucklandia showed a good linear relationship. The average recovery was 99.1% (RSD=1.2% and n=5). CONCLUSION: The methods are simple, reliable and have a good reproducibility and can be used for the quality control of Gujing Tablets.

Objective: To investigate the value of single and combination of virtual touch tissue imaging quantification (VTIQ) technique and mammography in diagnosis of benign and malignant breast lesions. Methods: VTIQ and mammography were performed on 99 patients (110 breast lesions).The mean value of shear wave velocity (SWVmean) of breast lesions were obtained. Mammographic classification of breast lesions was obtained with breast imaging reporting and data system (BI-RADS). Taken pathology as gold standard, the ROC curve was used to evaluate the diagnostic efficacy of SWVmean, mammography and the combination of these two methods. The differences of AUC among single and combined VTIQ technique and mammography in diagnosis of benign and malignant breast lesions were compared. Results: SWVmean of benign breast lesions was (3.03±0.78) m/s, of malignant ones was (5.61±2.11) m/s (P<0.001). The cutoff value of SWVmean in diagnosis of benign and malignant breast lesions was 3.93 m/s, and of mammography was BI-RADS 4B. AUC of VTIQ, mammography and combination was 0.870, 0.749 and 0.873, respectively. The differences of AUC between VTIQ technique and mammography, combination and mammography were statistically significant (P=0.036, 0.015), while of AUC between the combination and VTIQ technique was not statistically significant (P=0.908). Conclusion: Combination of VTIQ technique and mammography has high efficacy in diagnosis of benign and malignant breast lesions.

Gluconobacter oxydans is known to oxidize glucose to gluconic acid (GA), and subsequently, to 2-keto-gluconic acid (2KGA) and 5-keto-gluconic acid (5KGA), while 5KGA can be converted to L-(+)-tartaric acid. In order to increase the production of 5KGA, Gluconobacter oxydans HGI-1 that converts GA to 5KGA exclusively was chosen in this study, and effects of carbon sources (lactose, maltose, sucrose, amylum and glucose) and nitrogen sources (yeast extract, fish meal, corn steep liquor, soybean meal and cotton-seed meal) on 5KGA production were investigated. Results of experiment in 500 mL shake-flask show that the highest yield of 5KGA (98.20 g/L) was obtained using 100 g/L glucose as carbon source. 5KGA reached 100.20 g/L, 109.10 g/L, 99.83 g/L with yeast extract, fish meal and corn steep liquor as nitrogen source respectively, among which the optimal nitrogen source was fish meal. The yield of 5KGA by corn steep liquor is slightly lower than that by yeast extract. For the economic reason, corn steep liquor was selected as nitrogen source and scaled up to 5 L stirred-tank fermentor, and the final concentration of 5KGA reached 93.80 g/L, with its maximum volumetric productivity of 3.48 g/(L x h) and average volumetric productivity of 1.56 g/(L x h). The result obtained in this study showed that carbon and nitrogen sourses for large-scale production of 5KGA by Gluconobacter oxydans HGI-1 were glucose and corn steep liquor, respectively, and the available glucose almost completely (85.93%) into 5KGA.
Bioreactors ; Carbon ; chemistry ; Culture Media ; chemistry ; Fermentation ; Gluconates ; metabolism ; Gluconobacter oxydans ; metabolism ; Industrial Microbiology ; Nitrogen ; chemistry

Objective To analyze the reasons of catheter fracture of implantable vascular access devices and to explore the prevention and handling measures. Methods A retrospective analysis was carried in 3 887 adult patients with implantable vascular access devices and 13 had catheter fracture. The clinical features and causes were analyzed and the successful handling measures were summarized. Results Of 3 887 cases, a total of 13 (0.33%) catheter fracture occurred. The incidence rate of catheter fracture via subclavian jugular venipuncture was 3.70%,(6/162) , via internal jugular venipuncture was 0.18% (7/3 725), via internal jugular venipuncture was significantly lower than that via the subclavian venipuncture (χ2=47.505,P=0.000). There had no statistical differences between the left and right of the two puncture ways(P=0.707,0.682). Conclusions Catheter fracture is one of the serious complications in the process of use and maintenance of implantable vascular access device. Choosing appropriate surgical method, strengthen maintenance education, specificating the operation procedure, closing observation and other measures can not only reduce the occurrence of the catheter fracture, but also can dealt with catheter fracture in time, which could ensure the safety of patients' life.

Objective To investigate the distribution and antimicrobial resistance profile of clinical gram-negative bacterial isolates in the First Afifliated Hospital of Nanjing Medical University during 2014.Methods Bacteria identiifcation was performed by API system or the VITEK-2 Compact automatic identiifcation system. Disk diffusion susceptibility testing or VITEK-2 Compact automatic identification system was used to determine the susceptibility to antimicrobial agents. All data were analyzed using WHONET 5.6 software.Results Among the total 7 931 clinical isolates in 2014, gram-negative bacteria accounted for 64.2% (5 088/7 931). The top three pathogens wereE. coli,A. baumannii andK. pneumoniae. Notably, during the year 2014, 195 strains of carbapenem-resistantEnterobacteriaceaewere isolated, about 6.9% of all theEnterobacteriaceae isolates. Meanwhile, 613 (66.5%) strains of multiple drug resistantA. baumannii and 197 (28.7%) strains of multiple drug resistantP. aeruginosa were isolated.Conclusion During the year 2014, the resistance of the gram-negative bacteria in this hospital is mainly characterized by carbapenem-resistantEnterobacteriaceae, multiple drug resistant A. baumanniiand multiple drug resistantP. aeruginosa. Surveillance of antimicrobial resistance is beneifcial for rational use of antibiotics.

Objective To observe the formation of Staphylococcus aureus biofilm and the inhibitory and dispersive effects of betaine on the biofilm.Methods The inhibitory and dispersive effects of 0.1％ betaine on the biofilm from 20 strains of Staphylococcus aureus were examined by crystal violet assay.Results All the 20 strains of Staphylococcus aureus formed biofilm.The biofilm of methicillinsensitive Staphylococcus aureus (MSSA) was formed in 24 hours with peak value of absorbance (A590 nm) (1.99 ± 0.53).The biofilm of methicillin-resistant Staphylococcus atureus(MRSA) was formed in 48 hours with peak value of absorbance(A590 nm) (1.13 ±0.47).After adding betaine,the absorbance(A590 nm) of MSSA biofilm fell down to(1.74 ± 0.61) in 24 hours,while the absorbance(A590 nm) of MRSA biofilm fell down to(0.40 ± 0.12) in 48 hours,which was significantly reduced compared with the controls (t =2.43,5.84,P ＜ 0.05 respectively).When adding betaine after the biofilm formed,the absorbancies (A590 nm) of both MSSA and MRSA showed no significant difference compared with the controls (P ＞ 0.05).Conclusion Betaine could inhibit biofilm formation of Staphylococcus aureus at concentration of 0.1％,but it could not disperse the mature biofilm of Staphylococcus aureus.

Objective To investigate the effects of betaine on the formation and dispersion of biofilm of Pseudomonas aeruginosa and its drug-resistance.Methods A total of 20 strains of Pseudomonas aeruginosa were obtained from clinical inpatients.The biofilm formation abilities of the Pseudomonas aeruginosa were evaluated by violet staining,and the effects of betaine on the formation and dispersion of biofilm were studied.The minimum inhibitory concentration (MIC) values of Pseudomonas aeruginosa on ciprofloxacin were compared with the controls when biofilm was formed and inhibited.Results Biofilm was formed in all the 20 strains of Pseudomonas aeruginosa in 24 hours with absorbance (A590 nm) (1.90 ± 0.66).Betaine significantly inhibited biofilm formation of Pseudomonasaeruginosa in 24 hours compared with control group(t =4.36,P ＜ 0.01) and the maximum inhibition reached in 48 hours with absorbance(A590 nm) (1.12 ±0.60).The maximum dispersion of betaine on mature biofilm of Pseudomonas aeruginosa reached in 24 hours.The MIC range of ciprofloxacin to the 20 strains of Pseudomonas aeruginosa was 0.03 to 4 μg/mL with 0.25 μg/mL of MIC50 and 2 μg/mL of MIC90.After the biofilm was inhibited by belaine,the MIC of ciprofloxacin to Pseudomonas aeruginosa did not changed.The MIC of ciprofloxacin to biofilm-formed Pseudomonas aeruginosa was more than 16 μg/mL.Conclusion Betaine could effectively inhibit the formation of biofilm and disperse the mature biofilm of Pseudomonas aeruginosa,which may provide more choices for the treatment of clinical infection.The germicidal efficacy of ciprofloxacin has no changed on the biofilm-formed bacteria when inhibition of betaine was involved.

Objectives To investigate the effects of different degrees of carotid artery stenosis on cognitive function and neuronal apoptosis of hippocampal CA1 region in rats and to analyze the possible mechanisms of cognitive impairment. Methods According to the random number table,50 male Wistar rats were allocated into a sham operation group,a unilateral mild stenosis group,a unilateral moderate stenosis group,a unilateral severe stenosis group,a bilateral mild stenosis group,a bilateral moderate stenosis group,
a bilateral severe stenosis group,and a sham operation group. Aneedle-controlled suture method was used to induce a carotid stenosis model with different degrees of stenosis in rats. Water maze was to localize navigation and spatial search test was used to evaluate the cognitive function with different degrees of carotid artery stenosis in rats. Immunohistochemical method was used to observe the numbers of positive cells of P75 neurotrophin receptor (p75NTR),Bax,Bcl-2,neurotrophic factor-3 (NT-3 ),nerve growth factor (NGF)in hippocampal CA1 region under the light microscope. The conditions of neuronal apoptosis were observed. Results In 50 rats,6 died,1 rat in poor health failed to complete the Morris water maze test. The degree of bilateral carotid artery stenosis in 1 rat failed to meet 30%-60% at the same time,they were all removed. The remaining 42 rats were 6 in each group. (1)Compared with the sham operation group,the mean escape latency was prolonged (39 ± 6 s,32 ± 5 s,69 ± 7 s,respectively vs. 23 ± 4 s;all P < 0. 01),and the percentage of swimming distance in the platform quadrant was decreased (35 ± 4%,44 ± 4%,22 ± 5%,respectively vs. 53 ± 7%;all P < 0. 01),and the cognitive function was decreased with the degree of stenosis in the unilateral severe stenosis group,bilateral moderate stenosis group,and bilateral severe stenosis group. Compared with the unilateral severe stenosis group,the mean escape latency was prolonged and the percentage of swimming distance in the platform quadrant was decreased in the bilateral severe stenosis group (P < 0. 01). (2)Compared with the numbers of positive cells of Bax,p75NTR and Bcl-2 (8. 8 ± 3. 1,4. 2 ± 2. 3,and 5. 8 ± 1. 8,respectively)in the sham operation group,the numbers of positive cells of Bax,p75NTR,and Bcl-2 were increased (25. 5 ± 3. 5,11. 0 ± 2. 2,12. 3 ± 2. 7;15. 8 ± 3. 7,8. 9 ± 2. 2, 10. 5 ± 2. 9;and 47. 9 ± 6. 3,24. 7 ± 3. 0,12. 8 ± 2. 5,respectively)in the unilateral severe stenosis group, bilateral moderate stenosis group,and bilateral severe stenosis group (all P < 0. 01). The numbers of Bax and p75NTR positive cells were increased with the degree of stenosis. When the stenosis was severe,the numbers of Bax and p75NTR positive cells were increased in the bilateral severe stenosis group compared with those of the unilateral severe stenosis group (P < 0. 01). The numbers of NGF and NT-3 positive cells in each stenosis group had an increased trend compared with sham operation group,but there were no significant differences (F =1. 034,and 1. 358;P = 0. 420 and 0. 259 respectively). Conclusions Carotid stenosis can cause cognitive disorder in rats,and it is correlated with the degree of carotid stenosis. Ischemia caused neuronal apoptosis in hippocampal CA1 region may be one of the mechanisms of cognitive impairment after carotid artery stenosis in rats.