BACKGROUND: Vitrification is a comparatively new technology which applies high concentration cryoprotectant and rapid refrigeration. By the method, the cells were quickly frozen and to avoid damage by ice crystals inside and outside. OBJECTIVE: To compare the effect of four cryoprotectants on morphology and function of ovarian tissue in rats after vitrification. METHODS: The rats were randomly assigned into six groups with 6 rats for each: DMSO + EG, DMSO + EG + sucrose, DMSC +EG + sucrose + acetamide, EG + sucrose + acetamide, ovariectomized, and normal control groups. The ovarian tissues of four freezing groups were treated with the corresponding cryoprotectants, the vitrified ovarian tissues were then resected but not frozen and transplanted; otherwise, tissues were not treated with any treatment in the normal control group. Two weeks after freezing, the tissues were thawed and heterotopic-transplanted into femoribus intemus of hind limb. At 30 days after implantation, vaginal epithelial cells and estrus cycle were observed, while after three months, blood were collected to detect the level of estradiol (E2) and the ovarian tissues were reclaimed to analyze their morphological changes. RESULTS AND CONCLUSION: All ovarian tissues were damaged after cryoprersarvation in four freezing groups. The rates ot healthy primordial follicles were 67.9%, 71.6%, 80.5%, and 59.4%, respectively, while healthy primary follicles were 41.6%, 52.3%, 55.9%, and 36.7%, respectively. In all freezing groups, the rate of the healthy follicles in DMSO + EG + sucrose + acetamide group was higher than DMSO + EG group and EG + sucrose + acetamide group (P < 0.05). No significant difference was found in the proportion of follicles at different development stages among four groups. The typical secondary follicle was not found in four groups. Damaged ovotid showed oocyte pyknosis and vacuolation in cytoplasmic area. There was not typical cell type of all freezing groups. Ovarian autografting gained visible vascularity from surrounding tissue that connected ovarian tissue to form net. There was a lot of blood capillary in transplanted ovarian tissues and clumped primordial follicles in cortical substance. The rates of primary follicles and secondary follicles were lower than primordial follicles. The level of serum estradiol was obviously decreased compared with normal control group (P < 0.01). There was significant difference between DMSO + EG + sucrose + acetamide group and other three freezing groups (P < 0.05). Four kinds of freezing methods have poor effects on different stages of follicles and the structure of ovariarn tissue. DMSO + EG + sucrose + acetamide group is an optimal protocol for cryoprerserving ovarian tissue. Freezing methods still need to explore further because the rats had not appeared disciplinary estrus cycle after ovarian autoqrafting.

Objective To investigate the reversal effect of hMSH2 small interference RNA(siRNA) on chemo-resistance of ovarian carcinoma cell line OC3/TAX300,explore the clinical significance.Methods The specific hMSH2 siRNA (experimental group) and non-specific hMSH2 siRNA (negative control group) was designed,synthesized and transfected into ovarian carcinoma cell line OC3/TAX300.The expression mRNA and protein levels of hMSH2 were detected by real-time reverse transcription (RT)-PCR and western blot.The cell proliferation was detected by methyl thiazolyl tetrazolium(MTT) method after 12,24,48,72 hours of 2 μg/ml taxol,the apoptosis rate after 24,48 hours of 2 μg/ml taxol was analyzed by flow cytometry.Morphological changes and ultramicrostructure of cells after 48 hours of 2 μg/ml taxol were observed with transmission electron microscope.Results (1) The mRNA levels of hMSH2 were 0.004 ± 0.000,0.053 ±0.006 and 0.057 ± 0.012 in experimental group,negative control group and non-infected group,respectively.The protein levels of hMSH2 were 0.19 ± 0.04,1.00 ± 0.07 and 0.95 ± 0.03 in experimental group,negative control group and non-infected group,respectively.(2) Compared with the noninfected group and the negative control group.The cell proliferation was effectively inhibited after 12,24,48,72 hours of 2 μg/ml taxol(P ＜0.05).The cell cycle was arrested at G2/M phase,the apoptotic rate was significantly increased after 24,48 hours of 2 μg/ml taxol (P ＜ 0.05).The experimental group after 48 hours of 2 μg/ml taxol was found to have more visible cell shrinkage,more serious chromatin margination,nucleus condensation,fragmentation and apoptotic body formation,nucleolus disappeared,markedly swollen mitochondria,mitochondrial cristae disappeared and other signs of apoptosis.While the nucleus was located in the cells of the central and nucleolus is clear,only mild chromatin pyknosis and marginalized,mild swelling of mitochondria in the control group and blank group.Conclusion siRNA targeting hMSH2 may reverse the chemo-resistance of ovarian carcinoma cell line OC3/TAX300 and may become a treatment or a new direction in the adjuvant therapy of ovarian cancer.

Objective To investigate the antibiotic resistance of Haemophilus influenzae isolates collected from the children with respiratory tract infection for rational use of antibiotics in clinical practice .Methods The H .influenzae strains were isolated from children and subjected to antimicrobial susceptibility testing by Kirby-Bauer method .Nitrocefin disc test was used to detect the production of beta-lactamases .WHONET 5 .6 software was used to analyze the susceptibility data .Results A totalof1256strainsof H.influenzaewereisolated.About37.8% ,65.5% and16.5% ofthe1256strainsof H.influenzae were resistant to ampicillin ,trimethoprim-sulfamethoxazole ,and ampicillin-sulbactam ,respectively .Less than 10 .0% of these strains were resistant to any other antibiotics tested .Beta-lactamase was produced in 33 .5% of the 1 256 strains of H . influenzae .Conclusions The H . influenzae strains in this study are mainly resistant ampicillin and trimethoprim-sulfamethoxazole .About 80 .0% of these H . influenzae strains were still susceptible to cefaclor ,ampicillin-sulbactam , cefixime ,ceftazidime ,azithromycin ,ciprofloxacin ,meropenem and rifampin .The primary mechanism of ampicillin resistance in Haemophilus is production of beta-lactamases .

Objective To understand the status quo of Salmonella infection in the patients with infectious diarrhea in Zhongshan city .Methods The fecal samples collected from 6 417 outpatients or inpatients with infection diarrhea in our hospital from January 2010 to December 2013 were performed the Salmonella culture ,isolation and identification and at the same time the serological clas-sification and the pulsed-field gel electrophoresis (PFGE) were performed .Results Among 6 417 samples ,460 strains of Salmonel-la were detected with the isolation rate of 7 .17% .Salmonella infection was dominated by children infection .The Salmonella infec-tion rate in children aged less than 1 years was 52 .4% (241 cases) ,followed by children aged 1- < 3 years ,which was 36 .7% (169 cases) .In the serological classification ,the most of detected Salmonella was salmonella typhimurium 46 .1% ,accounting for 46 .1% , Stanley salmonella accounting for 13 .5% and Salmonella enteritidis accounting for 8 .7% .The PFGE homology analysis showed that in the XbaI enzyme digestion spectral band ,26 PFGE band types had 100% homology .Conclusion Salmonella is one of the main pathogenic bacteria causing diarrhea and serious impact on health ,which is worthy of clinicians to pay attention to .

Objective: To characterize the expression of programmed cell death 5 (PDCD5) in normal and osteoarthritic human cartilage. Methods: Articular cartilage specimens were obtained from 20 patients with osteoarthritis and 10 with femoral neck (normal cartilage) at the time of arthroplasty. Expression of PDCD5 was detected by flow cytometry, immunofluorescence, RT-PCR and immunohistochemical analysis. Results: PDCD5 expression in osteoarthritis cartilage was significantly higher than in normal cartilage especially in the nucleus. PDCD5 positive chondrocytes were mainly observed in the superficial and deep zone of osteoarthritis tissue sections,and in contrast, in the superficial and middle regions of normal controls. Conclusion: Since apoptotic chondrocyte death occurs more frequently in osteoarthritis compared to normal cartilage and PDCD5 is an apoptosis related protein, the different expression patterns of PDCD5 in osteoarthritis and normal cartilage suggest that it might be involved in the pathogenesis of osteoarthritis.

Objective To investigate the distribution characteristics of pathogenic bacteria among different seasons and ages in children lower respiratory tract infection.Methods To retrospectively analyze the results of bacterial culture in pediatric outpatients and inpatients with lower respiratory tract infections in our hospital from September 2014 to August 2015.The pathogenic bacterial detection situation of lower respiratory tract infection among different age groups and different were statistically analyzed.Results A total of 2 809 pathogenic bacterial strains were isolated from 4 629 lower respiratory tract specimens,and detection rate was 60.7%.The pathogenic bacteria distribution was different among different age groups.Haemophilus influenzae (33.3%) ranked the first place in the lower respiratory infection among children aged from 19 d to 7 year old.The detection rates of Haemophilus parainfluenzae,Moraxella catarrhalis and staphylococcus aureus had obvious seasonality,while Streptococcus pneumoniae had no obvious seasonality.Conclusion Pathogenic bacteria causing lower respiratory tract infections in children are different from those in adults,which are affected by age and seasonal change.

Objective To investigate the changes of serum concentration of amylase(AMY) and pathological of pancreas in severe acute pancreatitis of SD-rats treated with Ulinastatin combined with qingyitang and to provide the basis for clinical application.Methods Ninty male SD-rats with average weight of(271.59±11.13) g were conventional feeding for a week.Twelve hours before experiment,they were forbidden to take food.They were randomly divided into five groups(with 18 in every group):sham operation group,severe acute pancreatitis group,Ulinastatin treated group,qingyitang treated group,Ulinastatin combined with qingyitang treated group.Then eighteen rats in every group were divided randomly into 6 h,12 h and 24 h groups with six rats in each group.Severe acute pancreatitis model was created by retrograde injection of 5％ sodium taurocholate (0.1 ml/100 g) through pancreatic duct,in sham operation group,only pancreas were flipped.The three treated groups were treated with vein injection of Ulinastatin(20 000 U/kg)or qingyitang perfusion(1 ml/100 g)or Ulinastatin combined with qingyitangat 0 h,5 h and 11 h after severe acute pancreatitis model successfully created.AMY and patholigical changes of pancreas were measured.Results There was significant difference on AMY level at 6 h,12 h and 24.h between severe acute pancreatitis group and sham operation group[6 h:(16 518±327)U/L vs(1671±175)U/L,F =3296.24,P ＜0.01;12 h:(16 796±338) U/L vs(1751±223)U/L,F =3802.44,P ＜0.01;24 h:(1554±373) U/L vs(1724±187) U/L,F =2382.54,P ＜0.01].Compared with severe acute pancreatitis group,the AMY level and pathologic score both decreased significantly in Ulinastatin treated group,qingyitang treated group and Ulinastatin combined with qingyitang treated group(P ＜0.05 or P ＜ 0.01).At the same time,the AMY level and pathologic score both decreased more in Ulinastatin combined with qingyitang treated group than Ulinastatin treated group and qingyitang treated group(P ＜0.05 or P ＜ 0.01).Conclusion Combination of Ulinastatin with qingyitang in treatment of severe acute pancreatitis of rats is superior to treatment with single medicine.

Objective To understand the carrying status and drug resistance of streptococcus agalactiae in the genital tract of pregnant women.Methods The vaginal discharge from pregnant women was collected for conducting the bacterial culture.The iso-lated streptococcus agalactiae was performed the identification and the drugs susceptibility test by the VITEK 2 fully automatice bacterial identification analyzer.Results Among 1 042 samples,streptococcus agalactiae was isolated in 6 cases with the separation rate of 5.4%.The drug sensitivity test showed the Streptococcus agalactiae was highly sensitive to penicillin,cefazolin,vancomycin, linezolid and nitrofurantoin.The intermediate strains or drug resistant strains were not found.The sensitive rate to levofloxacin, moxifloxacin and clindamycin,erythromycin,tetracycline were 84.0%,84.0%,71.4%,39.3% and 8.9% respectively.Conclusion The carrying rate of streptococcus agalactiae in pregnant women is not high in our hospital.Penicillin and cefazolin should be as the drug of first choice for therapy and the clincal and laboratory nould payattention to detection of streptococcus agalactiae in pregnant women.

Objective:To evaluate the correlation between the timed up and go(TUG)test and fall risks in elderly frail patients.Methods:From July to September 2019, elderly frail patients who were treated at the cardiovascular department of our hospital were enrolled.Basic clinical data and fall-related information of patients were collected.Patients were divided into the fall group and the non-fall group.Results on the body mass index(BMI), TUG, 4-meter maximum walking speed(4 m MWS)and Barthel index were compared between the two groups.The correlation between TUG and each indicator was examined.Multivariate Logistic regression analysis was used to analyze the correlation between the TUG and falls in elderly patients.Results:A total of 96 eligible patients were enrolled, including 35 in the fall group and 61 in the non-fall group.The average TUG time was longer in the fall group than in the non-fall group(16.45±6.44 s vs.10.17±2.91 s, t=-6.556, P<0.001). The correlation analysis results showed that the TUG was correlated with falls and 4 m MWS( r=0.582 and 0.875, both P<0.001). Multivariate Logistic regression analysis showed that the TUG( OR=1.201, 95% CI: 1.111-1.470, P=0.004)and 4 m MWS( OR=1.146, 95% CI: 1.063-1.244, P=0.015)were risk factors for falls. Conclusions:The TUG is correlated with fall risks in elderly frail patients and should be recommended as a routine test in clinical practice.

Objective: To investigate the clinical features and prognosis factors of acquired immunodeficiency syndrome (AIDS) patients complicated with cryptococcal meningitis (CM). Methods: Retrospective analyses were performed on clinical features, laboratory data, treatment status and related prognosis factors in 81 AIDS patients with CM admitted to the Department of Infectious Diseases, Beijing You′an Hospital, Capital Medical University from January 2010 to December 2017. The t test, rank sum test and χ² test were employed to analyze the data. Results: Of the 81 AIDS patients with CM, 71 cases were infected with human immunodeficiency virus (HIV) by sexual transmission (87.7%). The most common clinical symptoms were fever in 60 (74.1%), headache in 72 (88.9%), and nausea and vomiting in 56 (69.1%). Cerebrospinal fluid (CSF) examination results show that 60 cases (74.1%) had elevated opening pressure, the white blood cell count was 17.0 (6.0, 44.5)×10⁶/L, monocyte count was 9.0 (3.0, 29.5)×10⁶/L, the level of chloride was (117.26±5.61) mmol/L, of glucose was 2.89 (2.05, 3.41) mmol/L, of protein was 0.32 (0.21, 0.65) g/L, ink staining positive rate was 84.0% (68/81), fungal culture positive rate was 59.3% (48/81). The positive rate of serum cryptococcal antigen was 96.3% (78/81), and CSF cryptococcal antigen positive rate was 93.8% (76/81). The clinical efficacies were not significant different among different treatment regimens (χ²=1.479, P=0.533). After treatment, 60 patients survived and 21 died, with an overall mortality rate of 25.9%. Univariate analysis showed that consciousness disorder and CSF opening pressure were significantly higher in the death group than those in the survival group (χ²=22.365, t=0.317, respectively, both P<0.05), while serum albumin (Alb) and CD4⁺ T lymphocyte counts were significantly lower in the death group than those in the survival group (t=7.975, Z=-3.073, respectively, both P<0.05). Multivariate logistic regression analysis showed that consciousness disorder and Alb were independent factors influencing the clinical outcome of AIDS patients with CM. Consciousness disorder was related with poor outcome (odd ratio (OR)=26.704, P=0.011, 95% confidence interval (CI) 2.115-337.247), and higher Alb was related with good outcome (OR=0.671, P=0.005, 95%CI 0.507-0.888). The area under the receiver operating characteristic curve of serum Alb for predicting poor outcomes of AIDS patients with CM was 0.932 (95% CI 0.859-0.998, P<0.01). By using 31.7 g/L as cut-off value, the sensitivity was 95% and the specificity was 81% for predicting poor outcome. Conclusions AIDS complicated with CM has a high mortality rate, and its clinical features are lack of specificity. Consciousness disorder and Alb are independent prognosis factors.