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.

Peptides that are composed of dextrorotary (d)-amino acids have gained increasing attention as a potential therapeutic class. However, our understanding of the in vivo fate of d-peptides is limited. This highlights the need for whole-body, quantitative tracking of d-peptides to better understand how they interact with the living body. Here, we used mouse models to track the movement of a programmed death-ligand 1 (PD-L1)-targeting d-dodecapeptide antagonist (DPA) using positron emission tomography (PET). More specifically, we profiled the metabolic routes of [⁶⁴Cu]DPA and investigated the tumor engagement of [⁶⁴Cu/⁶⁸Ga]DPA in mouse models. Our results revealed that intact [⁶⁴Cu/⁶⁸Ga]DPA was primarily eliminated by the kidneys and had a notable accumulation in tumors. Moreover, a single dose of [⁶⁴Cu]DPA effectively delayed tumor growth and improved the survival of mice. Collectively, these results not only deepen our knowledge of the in vivo fate of d-peptides, but also underscore the utility of d-peptides as radiopharmaceuticals.

Central nervous system (CNS) injuries, including stroke, traumatic brain injury, and spinal cord injury, are leading causes of long-term disability. It is estimated that more than half of the survivors of severe unilateral injury are unable to use the denervated limb. Previous studies have focused on neuroprotective interventions in the affected hemisphere to limit brain lesions and neurorepair measures to promote recovery. However, the ability to increase plasticity in the injured brain is restricted and difficult to improve. Therefore, over several decades, researchers have been prompted to enhance the compensation by the unaffected hemisphere. Animal experiments have revealed that regrowth of ipsilateral descending fibers from the unaffected hemisphere to denervated motor neurons plays a significant role in the restoration of motor function. In addition, several clinical treatments have been designed to restore ipsilateral motor control, including brain stimulation, nerve transfer surgery, and brain-computer interface systems. Here, we comprehensively review the neural mechanisms as well as translational applications of ipsilateral motor control upon rehabilitation after CNS injuries.
Animals ; Spinal Cord Injuries/therapy* ; Motor Neurons/physiology* ; Brain ; Stroke ; Recovery of Function/physiology*

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

Ubiquitin specific peptidase 28 (USP28) is closely associated to the occurrence and development of various malignancies, and thus has been validated as a promising therapeutic target for cancer therapy. To date, only few USP28 inhibitors with moderate inhibitory activity have been reported, highly potent and selective USP28 inhibitors with new chemotypes remain to be discovered for pathologically investigating the roles of deubiquitinase. In this current study, we reported the synthesis and biological evaluation of new [1,2,3]triazolo[4,5-]pyrimidine derivatives as potent USP28 inhibitors. Especially, compound potently inhibited USP28 (IC = 1.10 ± 0.02 μmol/L,  = 40 nmol/L), showing selectivity over USP7 and LSD1 (IC > 100 μmol/L). Compound was cellularly engaged to USP28 in gastric cancer cells. Compound reversibly bound to USP28 and directly affected its protein levels, thus inhibiting the proliferation, cell cycle at S phase, and epithelial-mesenchymal transition (EMT) progression in gastric cancer cell lines. Docking studies were performed to rationalize the potency of compound . Collectively, compound could serve as a new tool compound for the development of new USP28 inhibitors for exploring the roles of deubiquitinase in cancers.

Objective To investigate the role of Toll-like receptor 4 ( TLR4 ) and NOD-like receptor 3 (NLRP3)inflammasome in the liver injury of acute necrotizing pancreatitis (ANP) rat with obesity. Methods Twenty-four SD rats were randomly divided into normal group, ANP group, obesity group and obesity ANP group. The obesity rat model was established by continuously feeding high fat diet and the ANP model was induced by retrograde infusion of 5％ sodium taurocholate into the biliopancreatic duct. Rats were killed at 12 h after model establishment, and automatic biochemical immune analyzer were used for detecting serum AMY, LIP, ALT, AST, TG and TC. Pathological changes of pancreas and liver tissue samples were observed by miscroscopy and pathological score was recorded. The levels of MPO, CD68 , TLR4, NLRP3 and IL-1βin liver tissue were detected by immunofluorescence, and NF-κB and caspase-3 in liver tissue were detected by immunohistochemistry. Results The serum ALT and AST in obesity ANP group were significantly increased than those in ANP group (233. 00 ± 34. 44 U/L vs 102. 83 ± 8. 90 U/L,388. 00 ± 41. 60 U/L vs 282. 00 ± 21. 06 U/L);and liver pathologic score was also significantly higher than ANP group (6. 66 ± 1. 21 vs 3. 33 ± 1. 03);and CD68 + /TLR4 +, CD68 + /NLRP3 +, TLR4 + /NLRP3 +, MPO, NF-κB, IL-1β and caspase-3 level were all greatly higher in obesity ANP than those in ANP group, respectively (24. 16 ± 1. 47 vs 6. 66 ± 1. 21, 25. 00 ± 2. 60 vs 7. 00 ± 1. 41, 14. 16 ± 1. 47 vs 5. 50 ± 1. 04, 35. 33 ± 6. 88 vs 20. 83 ± 2. 48, 58. 80 ± 6. 75 vs 37. 63 ± 2. 96, 50. 00 ± 2. 36 vs 35. 00 ± 2. 82, 66. 00 ± 4. 04 vs 55. 00 ± 2. 60); and all the differences were statistically significant (all P<0. 05). Conclusions Liver injury was more severe in ANP rats with obesity, which may be related to the fact that obesity may enhance the activation of TLR4/NLRP3 signal pathway and result in the release of more inflammatory factors.

Objective: To study the efficacy and safety of sofosbuvir and daclatasvir regimens for patients who received kidney transplantation (KT) with hepatitis C virus (HCV) infection. Methods: This study enrolled a prospective cohort of consecutive KT patients with HCV infection from March 2016 to January 2018 in the hepatology Department of the Second Hospital of Shandong University. They were given sofosbuvir combined with daclatasvir, with or without ribavirin. The course of treatment was 12 weeks or 24 weeks. Clinical assessment, conventional liver and kidney biochemical parameters, hemoglobin, serum HCV RNA, as well as the types of immunosuppressive drugs and their doses were assessed routinely as follows: at the beginning of treatment; 2, 4, and 8 wk post treatment; at the end of treatment (EOT); and at 12, 24 wk after the therapy was completed. Adverse events and adjustment of anti-rejection drugs were surveilled during the treatment period. Results: A total of 13 patients were enrolled. All patients were naive to treatment. Their mean age was 46.84±7.79 years. There were 10 males and 3 females, 3 patients had cirrhosis (1 cases had decompensated cirrhosis), 10 patients had no cirrhosis. They were infected with HCV genotype 1 (6/13 GT1b), genotype 3 (2/13 GT3a) and genotype 6 (3/13 GT6a), and genotype 2 (2/13 GT2a). Twelve patients′ estimated glomerular filtration rate (eGFR) was > 30 ml/min per 1.73 m² at the beginning of treatment, 1 patient′s eGFR was <30 ml/min·1.73 m^2; 9 patients received 12 wk therapy, 4 patients received 24 wk therapy. Twelve patients had undetectable viral load by week 4 of treatment. All patients had undetectable HCV viral load at the end of treatment. Sustained virological response (SVR) 12 rate was achieved in 100% (13/13) of the recipients. The basic renal function remained stable during the course of treatment. No serious adverse events were observed during the treatment. Antiviral therapy was not discontinued due to side effects in any patient. Conclusions Sofosbuvir and daclatasvir for treatment of KT patients with HCV infection are highly effective and safe.

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.