Medical microbiology experiment is faced with many problems in online teaching. This study adopts the teaching mode of online live broadcast + operation video + virtual experiment, and make up the operation gap to some extent through operation video and virtual experiment. The mode of assessment is subjective thinking question (closely following the operation process) + experiment design + literature review (focusing on the key technology or new technology of clinical assessment that cannot be carried out due to the limitation of conditions in traditional experiments, such as mass spectrometry, fluorescence quantitative PCR, and G-test), and it is helpful to understand students' mastery of teaching objectives, and the ability of comprehensive application and innovative thinking. The student questionnaire shows that most students hold a positive attitude towards the online experimental teaching mode, and the quality of students' homework shows that most students have a good learning effect.

The evolution, structure and antigenic epitopes prediction of Rana dybowskii antimicrobial peptide dybowskin-1ST were carried out using bioinformatics software available online. Its antibacterial mechanism and structural properties were analyzed, and its activity was verified by applying wound healing assay in mice and bacteriostatic assay in vitro. This provides the theoretical basis for the improvement of parental peptide and the development of novel derivative peptides. The software MEGA_X were used to conduct homology alignment and to construct a phylogenetic tree. The online software ProtParam, ProtScale, PeptideCutter, signal, TMHMM Server were respectively used to predict the physicochemical parameters, hydrophilia/hydrophobicity, shear sites, signal peptides, and transmembrane domains of dybowskin-1ST. The online software SOPMA, Jpred4, DNAstar Protean were used to predict the secondary structure of dybowskin-1ST, and SWISS-MODEL, I-TASSER were used to predict the tertiary structure. ABCpred and SYFPEITHI were respectively used to predict its B-and T-cell epitopes. The effect of dybowskin-1ST on the wound healing was observed on experimental mice. Kirby-Bauer method and dilution method were used to determine the bacteriostatic activity of dybowskin-1ST. The dybowskin-1ST consists of 59 amino acid residues, of which leucine accounts for 16.9%, with a molecular formula of C₃₁₈H₅₁₀N₈₀O₉₃S₂. Its theoretical isoelectric point is 5.10 and the charge is -2. The dybowskin-1ST and dybowskin-1CDYa are closely related phylogenetically. The secondary structure of dybowskin-1ST predicted by the three methods were similar, which consisted of α-helix (44.07%), extended strand (16.95%), β-turns (3.39%), and random coil (35.39%). The prediction of tertiary structure showed that dybowskin-1ST was mainly composed of α-helix, and it was regarded as a hydrophilic protein with signal peptide sequence. Subcellular localization analysis showed that the probability of secreting the mitochondrial targeted peptides was 0.944. Dybowskin-1ST is an extracellular protein with no transmembrane structure region, but contains seven phosphorylation sites, three T-cell epitopes and eight B-cell epitopes. The dybowskin-1ST promoted wound healing and effectively inhibited the growth of Escherichia coli and Staphylococcus aureus. However, it had limited antibacterial activity against fungi and drug-resistant bacteria. Although the structure of dybowskin-1ST is rich in α-helix, the verification experiments showed that its antibacterial ability needs to be enhanced. The reason may be that it is a negatively charged and hydrophilic protein, and amino acid modification with the aim of increasing the number of positive charges and changing the hydrophobicity may be used to obtain derived peptides with enhanced activity.
Amino Acid Sequence ; Animals ; Mice ; Phylogeny ; Pore Forming Cytotoxic Proteins ; Protein Structure, Secondary ; Ranidae

Objective Using of cumulative live birth rate(CLBR)per oocytes retrieved cycle,to assess the clinical outcomes of in vitro fertilization or intracytoplasmic sperm injection(IVF/ICSI),and to explore impact factors on CLBR following utilization of all fresh and frozen embryos in one complete IVF/ICSI cycle using gonadotropin-releasing hormone(GnRH)agonist, GnRH-antagonist and clomiphene mild stimulation protocols. Methods Of the patients who underwent IVF/ICSI from January 1st, 2014 to December 31st, 2015 in the First Affiliated Hospital, Nanjing Medical University, a total of 6 142 oocytes retrieved cycles were included. The clinical and laboratory parameters of different ovarian stimulation protocols, and the effects of the age, number of oocytes retrieved and number of embryos available on the CLBR of each oocytes retrieved cycle were analyzed.Results The CLBR was 69.0%(2 004/2 906)in the GnRH-agonist protocol versus 67.4%(644/955)in the GnRH-antagonist protocol (P>0.05); the CLBR of clomiphene mild stimulation protocol was 53.2%(1 215/2 281),significantly lower than those of the other two protocols (all P<0.05). The CLBR significantly decreased with age increased. When divided into four groups according to the patients′ age, we found that CLBR were not statistically significant using three different protocols in the 20-25 years old group(all P>0.05).There was a strong association between the number of oocytes retrieved and embryos available on CLBR. CLBR rose significantly with an increasing number of oocytes up to 6, then the rising trend slowed down. Patients were categorized into four groups according to the number of oocytes retrieved,CLBR was significantly higher using GnRH-antagonist protocol (50.0%)than mild stimulation protocol(37.0%)in low ovarian responder(0-4 oocytes)group(P<0.05). The CLBR were no significant difference among three protocols in normal(10-15 oocytes)and high responders(≥15 oocytes)group(all P>0.05).The incidence rate of ovarian hyperstimulation syndrome in GnRH-agonist protocols(5.2%,152/2 906)were significantly higher than those of GnRH-antagonist(4.4%, 42/955)and clomiphene mild stimulation protocols(1.5%,34/2 281;all P<0.05).Conclusions CLBR is an important index to assess the clinical outcomes of IVF/ICSI. Age, number of oocytes retrieved and embryos available could affect CLBR obviously. According to the different age and ovarian response of patients, we should design ovarian stimulation protocols based on target oocytes number in order to get higher CLBR and reduce complications.

Objective To investigate the impact of dyslipidemia on in-vitro fertilization or intracytoplasmic sperm injection (IVF/ICSI) pregnancy outcome in patients with polycystic ovary syndrome (PCOS).Methods From July 2013 to March 2016,468 PCOS patients with antagonist protocol in IVF/ICSI of First Affiliated Hospital of Nanjing Medical University,cycles were divided into dyslipidemia group (108 cases) and normol blood lipids group (360 cases) according to the serum cholesterol,triglyceride (TG),high-density lipoprotein,low density lipoprotein levels.The general condition and clinical outcomes of the two groups were analyzed retrospectively,including the implantation rate,clinical pregnancy rate,live birth rate and the incidence of moderate to severe ovarian hyperstimulation syndrome (OHSS),etc.Besides,stratified analysis and multivariate logistic regression analysis were used to correct the impact of body mass index (BMI).Results (1) Comparing the based data of dyslipidemia group and normal blood lipids group:age,years of infertility,basic FSH,basic LH,basic estradiol and other indexes had no significant differences (all P＞0.05),but BMI of dyslipidemia group was significantly higher than normal blood lipids group [(25.0±3.0) versus (23.1±3.0) kg/m2],difference had statistical significance (P＜0.01).(2) The high score embryo rate,endometrial thickness on the day of hCG injection,progesterone and LH levels on the day of hCG injection,moderate to severe OHSS rate and miscarriage rate in the two groups did not exhibit remarkable differences (all P＞0.05).However,the number of dominant follicle,retrieved oocyte number,estrogen level on the day of hCG injection,implantation rate,biochemical pregnancy rate,clinical pregnancy rate and the live birth rate in dyslipidemia group were significantly less than those of normal blood lipids group (all P＜0.05),the dose of gonadotropin (Gn) and days of stimulation were significantly higher compared with the normal blood lipids group,there were significant differences statistically (all P＜0.05).(3) Stratified analysis showed that no matter in BMI＜24 or BMI≥24 kg/m2 group,the dose of Gn and days of stimulation were significantly higher in the dyslipidemia group than those of the normal blood lipids group,the difference was statistically significant (P＜0.05).However,the number of oocytes retrieved,estrogen level on the day of hCG injection had obvious downtrend,and the difference was statistically significant (P＜0.05) in BMI≥24 kg/m2 group.Multivariate logistic regression analysis found that,even after the correction of BMI,dyslipidemia still had negative impact on implantation rate,biochemical pregnancy rate,clinical pregnancy rate and the live birth rate (P＜0.05).(4) Further analysis of the different components of blood lipids in the clinical pregnancy group and unobtained pregnancy group revealed that the level of triglyceride (TG) in the unobtained pregnancy group was significantly higher than that in the pregnancy group,and the difference was statistically significant (P＜0.05);logistic regression analysis also showed that the increase of TG levels was negatively correlated with the clinical pregnancy rate of PCOS patients (P＜0.05).Conclusions PCOS patients combined with dyslipidemia have a higher BMI,and dyslipidemia increases the dosage of Gn,reduces the implantation rate,clinical pregnancy rate and live birth rate,especially the increase of TG level,which has adverse effects on IVF/ICSI outcome in patients with PCOS.

ABSTRACT:Objective To assess the effect of prolonged laparoscopic surgery on peritoneal mesothelial cells and fibrinolysis in humans.Methods We examined prospectively 1 6 consecutive patients who underwent laparoscopic surgery (LAP)and 2 1 patients who underwent conventional open surgery (OP)for high-medium rectal cancer with curative intent.During the procedure,biopsy of the parietal peritoneum was made before operation and at 45 min,90 min,and 120 min after operation.The tissue-type plasminogen activator (tPA)and plasminogen activator inhibitor-1 (PAI-1 )were determined by enzyme-linked immunosorbent assay in peritoneal tissues.The cellular injury was detected by LDH assay.The proliferation was quantified by MTT assay.Results PAI-1 activity in the peritoneal tissue was significantly lower in LAP group than in the OP group.tPA activity decreased after 45min of open surgery,but there was no significant change in the LAP group.With time extension,the LDH activity increased and the proliferation of the mesothelial cells decreased.Conclusion Preservation of a prolonged hypofibrinolytic state by inhibition of PAI-1 up-regulation during LAP may predispose patients to less postoperative peritoneal adhesion. The cellular injury becomes apparent and the proliferation is inhibited during prolonged laparoscopic surgery.

Calculation of cardiac hemodynamic parameters is based on accurate detection of feature points in impedance cardiogram. According to these parameters, doctors can determine heart conditions, so it is very important to accurately detect the feature point of impedance differential signals. This article presents a process in which we used wavelet threshold method to de-noise signals, and then detected the feature points after six layers wavelet decomposition by using bior3. 7. The experimental data were collected from healthy persons in our laboratory and twenty two clinical patients in Chongqing Daping Hospital by using KF_ICG instrument. The results indicated that this method could precisely detect feature points whether it was from healthy people or clinical patients. This helps to achieve the application of noninvasive detection cardiac hemodynamic parameters in clinical treatments by using impedance method.
Cardiography, Impedance ; Electric Impedance ; Heart ; physiology ; Hemodynamics ; Humans ; Wavelet Analysis

Objective To explore the feasibility and efficiency of the treatment of lumbar degenerative diseases after transforaminal lumbar interbody fusion(TLIF)and posterolateral fusion(PLF)procedures in which unilateral pedicle screw fixation was used.Methods From December 2006 to August 2008,78 cases with the lumbar degenerative diseases who received lumbar posterolateral fusion were analyzed.There were 48 cases of which underwent TLIF and PLF procedures with unilateral pedicle screw fixation(unilateral group),including 25 males and 23 females with an average of 47.6 years;and 30 cases of which underwent TLIF and PLF procedures with bilateral pedicle screw fixation(bilateral group),including 21 males and 9 females with an average of 50.5 years.The clinical effects between the two groups were evaluated with Oswestry disability index and visual analogue score(VAS)index.The operation time,blood loss,fusion rates and intervertebral collapse rates were also compared.Results Oswestry disability index,low back pain VAS index and skelalgia VAS index in both groups showed statistical significance between preoperation and 3 months,or 3 months and 1 year postoperatively.There was no difference in score improvement between the two groups.There were difference in operation time,blood loss and cost of hospitalization between unilateral and bilateral group.The former was lower.There was no difference in postoperative length of stay between the two groups.The fusion rate of unilateral group and bilateral group were 91.7%(44/48)and 93.3%(28/30),respectively.Conclusion Auto graft combined with unilateral pedicle screw fixation provids better spinal instant stability.TLIF and PLF with unilateral pedicle screw fixation was a satisfactory method in treating degenerative disease of lumbar vertebrae.

Objective To investigate the relationship between serum progesterone level at the day with human chorionic gonadotrophin (hCG) administration and pregnant outcome from in in-vitro fertilization-embryo transfer(IVF-ET). Methods From Mar. 2002 to Apr. 2007, 786 cycles with serum progesterone measurement on the day of hCG administration for final oocyte maturation in IVF were analyzed retrospectively in Reproductive Medicine Center in First Affiliated Hospital of Nanjing Medical University.All stimulations were down-regulated with gronadotrophin release hormone agonist (GnRH-a) in both long protocols and short protocols before gonadotrophin stimulation. When the thresholds of serum progesterone were set at 5.5, 6.0,6.5,7.0,7.5,8.0,8.5 and 9.0 nmol/L, respectively. If the level of progesterone was less than the thresholds, those patients were in lower progesterone group, on the contrary, more than the threshold value, those patients were in higher progesterone group. The laboratory results and the clinical outcomes between all patients at lower and higher progesterone group at different thresholds value were analyzed. Results The rate of normal fertilization, quality embryos, successful implantation, chemical pregnancy, clinical pregnancy and live birth did not exhibit remarkable difference between patients with higher and lower serum progesterone level at multiple thresholds on the day of hCG administration in the 786 cycles (P >0.05). However, when the thresholds of serum progesterone were at 8.5 and 9.0 nmol/L, early abortion rates of 27.3% (3/11) and 3/7 in higher progesterone group were significantly higher than 8.8% (26/297) and 8.6% (26/301) in lower progesterone group (P<0.05). And the total abortion rates of 3/7 in higher progesterone group were significantly higher than 11.0% (34/301) in lower progesterone group when the thresholds of serum progesterone were 9.0 nmol/L (P<0.05). Conclusions This study did not prove the correlationship between progesterone level at the clay with hCG administration and the probability of clinical pregnancy or live birth. However, early abortion rates or the total abortion rates were associated with higher progesterone level when the thresholds of serum progesterone were at 8.5 nmol/L or 9.0 nmoL/L.

Objective To analyze the clinical and CT manifestations of lung involvement of microscopic polyangiitis (MPA). Methods The clinical manifestations,laboratory ANCA examinations and CT features of 16 patients with lung involvement of MPA were retrospectively reviewed. Results (1) Clinical manifestations: 11 cases had hemoptysis or bloody sputum. Eight cases, who first presented with lung symptoms, were misdiagnosed with other lung diseases. All cases had mulfiorgans injuries involved kidney, cardiovascular and endocrine system, etc. (2) Laboratory examinations: all cases were pANCA positive and 14 cases were MPO-ANCA positive. (3) CT examinations: all cases had interstitial changes, 15 cases were interstitial predominately and 1 case was parenchymal predominately. Eight cases had pulmonary interstitial fibrosis and 11 cases had consolidation and 6 of them had both interstitial and consolidation. Two case had accompanied multiple nodulesand one of them had multiple cavitates. Six cases had mediastinal lymphoadenopathy. Conclusions Most of MPA patients have clinical manifestations of hemoptysis and bloody sputum, the CT examination show interstitial lung disease. Middle or advanced age population presented with above-mentioned manifestations should be alert to MPA, whether or not they have kidney and other organs injury.

Objective To study the inhibitory effect on the expression of regulated upon activation,normal T cell expressed and secreted (RANTES) and monocyte ehemotactic activity of ectopic endometrial stromal cells by nuclear factor(NF)-kB decoy oligonucleotides (ODN). Methods The stromal cells of ectopic endometrium were divided into 3 groups. Two groups were cultured with or without 10 μg/L of interleukin (IL)-1β. Another group was transfected with NF-kB decoy ODN with the aid of a lipofectamine reagent. After 4 h of transfection, 10 μg/L of IL-1β was added to induce the stromal cells to secrete RANTES. Concentration of RANTES in the supernatant at 4, 8, 12, 24 and 36 h was measured with the sandwich enzyme linked immunosorbent assay (ELISA). U937 monocyte chemotactic activity was assayed in Boyden chambers. The specific RANTES-neutralizing monoclonal antibodies at serial doses (0. 5, 1, 2, 4and 8 mg/L) were added into IL-1β induced medium of 24 h to detect the monocyte chemotactic activity of RANTES in supernatant. Results The concentration of RANTES secreted by stromal cells was respectively (58 ± 10), ( 150 ± 35 ), ( 360 ± 46 ) and ( 586 ± 42 ) ng/L after IL-1β stimulation for 8,12,24 and 36 h,significantly higher than that of stromal cells cultured without IL-1β. The concentrations of RANTES were respectively (86±16), ( 128±28 ) and ( 183±32) ng/L after IL-1β stimulation for 12, 24 and 36 h in stromal cells transfected with NF-kB decoy ODN, evidently lower than that of stromal cells stimulated with IL-1β alone. The monocyte ehemotactic index of 12, 24, 36 h in conditioned medium of stromal cells transfected with NF-kB decoy ODN was respectively 10. 3 ± 0. 9, 13.7 ± 1.1, 18.6 ± 1.2, which was evidently lower than that of stromal cells stimulated with IL-1β alone. The anti-RANTES antibody at 0. 5, 1,2, 4 and 8 mg/L inhibited respectively 5%, 23%, 40%, 62% and 61% of the chemotactic activity in 12 h medium treated with IL-1β. Conclusions RANTES accounts for the majority of the monocyte chemotactic activity in IL-1β induced medium of 24 h. NF-kB decoy ODN may influence the feed-forward inflammatory loop whereby IL-1β from activated macrophages can lead to RANTES production by ectopic implants and further monocyte chemotaxis.