The aim of this study is to evaluate the in vitro and in vivo antibacterial potential of Ly-sZHSHW,a phage lysin against Acinetobacter baumannii infections,and to study its characteris-tics.The pET28a-Lys recombinant plasmid containing LysZHSHW coding gene was constructed by PCR,enzyme digestion and ligation using the expression plasmid pET28a as the backbone and ex-pressed in E.coli BL21(DE3).After confirming the expression of the LysZHSHW through West-ern blot analysis,its characterization and potential applications were assessed both in vitro and in vivo.The results showed that the pET28a-Lys recombinant plasmid was successfully constructed and the LysZHSHW protein was expressed correctly.The mass concentration of the purified en-zyme was 4 086 mg/L,which could be used for subsequent experiments.The enzymatic activity of LysZHSHW was determined to be 630 U/μg,with maximum activity observed at 25 ℃ and pH9.0.In vitro experiments demonstrated that 1 000 or 750 mg/L of LysZHSHW,in the presence of EDTA,resulted in a four-log reduction in bacterial counts without any cytotoxicity.In vivo,2.5 μg of LysZHSHW combined with EDTA could increase the survival rate of Galleria mellonel-la larvae infection model to 92.86％after 24 hours,and 0.15 mg of LysZHSHW reduced the bacte-rial load in the thighs of mice by 2.8 logs and alleviated the inflammatory response in muscle fi-bers.In conclusion,LysZHSHW derived from Acinetobacter baumannii bacteriophage exhibited characteristics such as stability at room temperature,alkaline pH,and safety,making it a promis-ing candidate as a novel antimicrobial agent.

Objective:A dose verification system of two-dimensional semiconductor matrix(SRS MapCHECK)was used to verify the dose of the clinical treatment plan of patients who underwent CyberKnife(CK),which realized rapid verification for individualization of radiotherapy plans of patients through analyzed the γ-passing rates of them.Methods:A total of 253 patients with tumor who received CK clinical treatment in the First Medical Center of Chinese PLA General Hospital from March 2021 to May 2023 were selected.Among of them,121 cases received CK treatment on head,and 30 cases received that on lung,and 102 cases received CK treatment on abdomen and other metastatic tumor.In the MultiPlan treatment plan system,the plan of patient was mapped to the integrated model composed of StereoPHAN model and SRS MapCHECK matrix dose verification system by the means of the plan image center overlap.The dose verification was conducted on the plan of each patient on the basis of ensuring the consistency of the number of beam,direction of beam and the monitor unit.The different γ analysis standards(1%/1 mm,2%/1 mm,3%/1 mm,1%/2 mm,2%/2 mm,3%/2 mm,1%/3 mm,2%/3 mm and 3%/3 mm)were adopted to conduct global analysis of absolute dose for each verification plan,and the threshold(TH)of low dose was set as 10%.Results:The γ passing rates of phantom verification plans of 253 patients were respectively(88.64±5.91)%,(95.43±3.40)%,(97.90±2.06)%,(96.51±2.35)%,(98.15±1.68)%,(99.06±1.12)%,(98.30±1.39)%,(99.09±0.97)%and(99.52±0.63)%under different analysis standards.The γ passing rates of other standards of patients with tumor on different parts were larger than 95%except the analysis result of 1%1 mm standard.The overall analysis result of the deviation of central point dose was(-1.30±2.17)%,among of which the tumor of head,abdominal tumors and other metastatic tumor were about approximately-2%,while that of lung tumors were approximately-3%.The deviation of abdominal and other metastatic tumor was the minimum.The correlation analysis showed that the target volume and the size of the minimum collimator were respectively correlated to the dose deviation of the center.Conclusion:SRS MapCHECK dose verification system can conveniently and quickly realize the individualized verification for the plan of patients who receive CK treatment.

Objective To investigate the effect of glycine N-methyl transferase (GNMT)on intrauterine adhesion (IUA)fibrosis and its related mechanism.Methods In vivo experiment:A total of 36 healthy female SD rats (SPF grade,6～8 weeks old and weighing from 180～220 g)were subjected in this study.IUA model of SD rats and IUA model of GNMT overexpressed rats were established.RT-qPCR and immunofluorescence assay were applied to detect GNMT expression level in normal uterus and model group.RT-qPCR and Western blotting were used to detect the mRNA and protein levels of fibrosis-related molecules and the activation of TGF-β1/Smad3 signaling pathway in each group.The number of endometrial glands in each group was observed by HE staining.Masson staining was used to analyze the severity of endometrial fibrosis in each group.In vitro experiment:transformed human endometrial stromal cells (THESCs)fibrotic phenotype model was constructed using TGF-β1,and THESCs stably transfected with GNMT overexpression lentvirus were treated with TGF-β1.RT-qPCR and Western blotting were used to detect the mRNA and protein expression of fibrosis-related molecules.The expression of TGF-β1/Smad3 signaling pathway was detected by Western blotting.TGF-β1/Smad3 signaling pathway was activated by TGF-β1/Smad signaling pathway activator (SRI-011381),and the expression of TGF-β1/Smad3 signaling pathway and key molecular proteins of fibrosis phenotype was measured with Western blotting.Results In vivo experiment,the mRNA and protein expression levels of GNMT were significantly decreased in the IUA rats than the control rats (P<0.05).Overexpression of GNMT decreased the mRNA and protein levels of fibrosis related molecules,Collagen Ⅰ,Collagen Ⅲ and FN in the IUA rats (P<0.05),and decreased the phosphorylation levels of TGF-β1 and its downstream Smad3 protein (P<0.05).HE and Masson staining showed that overexpression of GNMT could increase the number of endometrial glands and reduce the severity of fibrosis in the IUA rats (P<0.05).In vitro experiments:overexpression of GNMT decreased the mRNA and protein levels of Collagen Ⅰ,Collagen Ⅲ and FN associated with fibrotic phenotype of THESCs (P<0.05),and reduced the phosphorylation level of Smad3 protein,downstream of TGF-β1 (P<0.05).After activation of TGF-β1/Smad3 signaling pathway,the protein levels of TGF-β1/Smad3 signaling pathway and downstream fibrosis phenotype molecules,Collagen Ⅲ and FN,were significantly decreased in the LV-GNMT+SRI-011381 group.Conclusion Overexpression of GNMT can inhibit endometrial fibrosis by regulating TGF-β1/Smad3 signaling pathway,thus achieving therapeutic effect on IUA.

Objective:To understand the prevalence of arboviruses in mosquito samples in Xichang City, Sichuan Province, and enrich the data of arbovirus activity and genetic characteristics in southwestern Sichuan Province.Methods:In June 2018, the nucleic acid was extracted from Culex tritaeniorhynchus mosquitoes collected from different pigsties in three villages and suburbs of Xichang City. The specific primers of Yunnan orbivirus, Banna virus, Tibet orbivirus (S7, S10), Flavivirus and alphavirus were used for quantitative polymerase chain reaction examination, and the positive product was cloned for sequencing analysis. Results:A total of 9 012 mosquitoes were collected, of which Cx. tritaeniorhynchus was the dominant species. A number of 88 batches of these mosquitoes were amplified, and 2 strains of Japanese encephalitis virus (JEV), 7 strains of Banna virus (BAV), 7 strains of Tibet orbivirus (TIBOV) and 1 strain of Yunnan orbivirus virus (YOUV) were detected, respectively. By the results of cluster analysis and evolutionary tree analysis, the 17 newly found virus strains were close to the Yunnan isolates, and 2 JEV strains were located in the GI-b clade. The other 7 strains of BAV were A2 evolutionary clades. Of the 7 TIBOV plants, 6 were located in the same clade. One TOUV was in the same clade as the Yunnan strain. Conclusions:Culex tritaeniorhynchus mosquitoes in Xichang city might carry JEV, BAV, YOUV and TIBOV, among them JEV was GI-b type and BAV was A2 type. The results provide data supporting the detection and analysis of arboviruses in Xichang city.

OBJECTIVE: To explore the genetic basis for a child presented with renal failure and multi-cystic dysplastic kidney without anal atresia. METHODS: Peripheral blood sample of the child and his parents were collected and subjected to whole exome sequencing. Candidate variant was verified by Sanger sequencing. RESULTS: The 40-day-old infant had presented with vomiting brown matter in a 7 days neonate and was transferred for kidney failure. Clinical examination has discovered renal failure, polycystic renal dysplasia, congenital hypothyroidism, bilateral thumb polydactyly, sensorineural hearing loss and preauricular dermatophyte. Genetic testing revealed that he has harbored a previously unreported c.824delT, p.L275Yfs*10 frameshift variant of SALL1 gene, which was confirmed by Sanger sequencing as de novo. CONCLUSION The patient was diagnosed with Townes-Brocks syndrome due to the novel de novo variant of SALL1 gene. Townes-Brocks syndrome without anal atresia is rare. Above finding has also enriched the mutational spectrum of the SALL1 gene.
Abnormalities, Multiple ; Anus, Imperforate/genetics* ; Child ; Female ; Hearing Loss, Sensorineural/genetics* ; Humans ; Infant ; Infant, Newborn ; Male ; Renal Insufficiency ; Thumb/abnormalities* ; Transcription Factors/genetics*

Objective:This research was designed to investigate the safety and efficiency of laparoscopic simultaneous resection in the treatment of synchronous colorectal cancer liver metastases (sCRLM).Methods:From January 2009 to December 2019, 121 patients diagnosed as sCRLM received radical resection in Peking University Third Hospital were included in the research. According to the surgery approaches, the patients were divided into laparoscopic surgery group and open surgery group. Statistical analysis of general patient data, surgical data, postoperative complications and follow-up of the two groups of patients.Results:There were 79 cases in the laparoscopic surgery group including 30 females and 49 males, and the average age was 61.5 years. There were 42 cases in the open surgery group including 15 females and 27 males, and the average age was 63.2 years. There were no significant differences in the preoperative demographic characteristics, the location of primary tumor, gene status, the size of liver metastases, the proportion of multiple liver metastases, the level of tumor markers and the proportion of neoadjuvant chemotherapy between laparoscopic surgery group and the open surgery group ( P>0.05). The complication rate was 15.2% (12/79) in the laparoscopic surgery group and 23.8% (10/42) in the open surgery group. There were no significant differences between the two groups ( P>0.05). The 3-year and 5-year survival rates in laparoscopic surgery group were 52.9% and 44.4%, which were 42.5% and 23.0% respectively in open surgery group. There were no significant differences between the two groups ( P>0.05). The 1-year and 3-year disease-free survival rate in laparoscopic surgery group were 50.6% and 41.2%, which were 44.7% and 19.4% respectively in open surgery group. There were no significant differences between the two groups ( P>0.05). Conclusions:Laparoscopic simultaneous resection was safe and feasible for patients with sCRLM. Comparing with the open surgery, the laparoscopic surgeries had similar incidence of perioperative complications and long-term oncological efficiency.

BACKGROUNDDiabetes-related pathogenic factors can cause retinal ganglion cell (RGC) apoptosis, but the specific mechanism is not very clear. The aim of this study is to investigate the correlation between glycogen synthase kinase-3 (GSK-3) activation and retinal neuron apoptosis.

METHODSIn an in vitro experiment, the number of apoptotic RGC-5 cells differentiated by staurosporine was evaluated via flow cytometry and nuclei staining using Hoechst 33258. GSK-3 phosphorylation and caspase-3 activation in RGC-5 cells after serum deprivation were determined using Western blotting. Mitochondrial membrane potential was detected using the dye 5, 5', 6, 6'tetrachloro-1, 1', 3, 3'-tetrethyl benzimidalyl carbocyanine iodide, and reactive oxygen species (ROS) levels were measured with dihydroethidium. In an in vivo experiment, the number of apoptotic retinal neurons was evaluated via terminal transferase dUTP nick-end labeling (TUNEL), and GSK-3 phosphorylation was determined using Western blotting, in the retinal nerve epithelial tissue of rats in which diabetes was induced by intravenous tail-vein injection of streptozotocin for 4 weeks.

RESULTSThe levels of phosphorylated Ser21/9 in GSK-3α/β and p-T308/S473-AKT were lower and the cleaved caspase-3 levels were higher in the serum-deprived model (P < 0.05). Lithium chloride treatment was associated with a slower rate of apoptosis, increased mitochondrial membrane potential, and decreased ROS levels in differentiated RGC-5 cells (P < 0.05). The level of blood glucose and the number of TUNEL-positive cells in the whole-mounted retinas were higher (P < 0.01), and the levels of phosphorylated Ser21/9 in GSK-3α/β and body weight were lower (P < 0.05). However, the thickness of the retinal nerve epithelial layer was not significantly less in diabetic rats compared with control group. Lithium chloride intravitreal injection increased the levels of phosphorylated Ser21/9 in GSK-3α/β and decreased TUNEL-positive cells in the whole-mounted retinas.

CONCLUSIONGSK-3 kinase is closely related to retinal neuron apoptosis, and the application of the GSK-3 inhibitor lithium chloride can reduce retinal neuron apoptosis in early diabetic retinopathy.

Animals ; Apoptosis ; genetics ; physiology ; Cell Line ; Cell Survival ; physiology ; Diabetic Retinopathy ; enzymology ; genetics ; metabolism ; Flow Cytometry ; Glycogen Synthase Kinase 3 ; genetics ; metabolism ; Male ; Neurons ; cytology ; metabolism ; Rats ; Rats, Sprague-Dawley ; Retina ; cytology ; enzymology

Objective To explore the prognosis and management of atrial fibrillation (AF) in patients with atrial septal defect(ASD) accompanied by AF after transcatheter closure of ASD. Methods During the period from July 2010 to May 2013, a total of 24 patients with ASD accompanied by AF were admitted to authors’ hospital to receive transcatheter closure of ASD. Electrocardiogram (ECG), chest X-ray film and transthoracic echocardiography (TTE) were performed before and one day after the operation. Follow-up information was obtained through telephone or at out-patient clinic interview. Results Successful occlusion of ASD was obtained in all patients, and in no patient the AF rhythm turned to sinus rhythm after the procedure. In one patient preoperative AF turned to postoperative atrial flutter, and AF recurred in one case who had received transcatheter ablation of AF before the procedure. One female patient developed gastric bleeding during the course of orally taking warfarin, and she died of cerebral infarction at three days after ceasing the use of warfarin. Of the 24 patients, no anticoagulant drug was used in 5 (20.8%), oral administration of aspirin was given in 7 (29.2%), and oral medication of warfarin was employed only in 11 (45.8%). Conclusion The spontaneous conversion rate of AF is very low in patients with ASD complicated by AF after transcatheter closure of ASD. Postoperative medication of anticoagulation should be strictly standardized and carefully managed.

Objective Prepare biomimetic muitilayered scaffold which has similar structure of natural cartilage.Method By lyophilizing the scaffolds which were prefrozen at-20℃ and in liquid nitrogen successively,we prepared double-layered spongy scaffolds.By partially thawing the prefrozen samples and refreezing them in liquid nitrogen before the final liyophilization,we prepared biomimetic multilayered scaffolds with about 2mm thickness.XRD and FT-IR were used to confirm the interaction between collagen and chitosan.SEM was used to observe the morphologies of the scaffolds.The mechanical properties of pure chitosan scaffolds,pure collagen scaffolds,composite single-layered scaffolds and biomimetic multilayered scaffolds were compared both in dry and wet conditions.Results There was chemical interaction between collagen and chitosan.Composite materials will form better pore structure.The biomimetic multilayered scaffolds have upright pores,round pores and a dense layer from bottom to top of the scaffolds.The scaffolds have quite different mechanical properties between dry and wet state.Under wet state,the different layers of the biomimetic muitilayered scaffold have different mechanical properties.Results The biomimetic structure of the multilayered scaffold is very close to that of the natural articular cartilage,and the different layers of the biomimetic muitilayered scaffold had different mechanical properties under wet state.These are hopefully beneficial to help maintain the phenotypes of chondrocytes and promote the repairing effect of cartilage defects.

Objective To evaluate the feasibility and effectiveness of endoluminal grafting for the treatment of abdominal aortic aneurysms in high-risk patients with serious co-morbidities.Methods Endoluminal stent grafting was performed in fifty-one patients(45 males and 6 females,with a mean age of 71.6±7.5 years)with abdominal aortic aneurysms.Of all the patients,21(37.7%)were high-risk surgical candidates because of associated co-morbidities.These patients were classified in grade Ⅱ and Ⅲ according to the criteria assigned by the"Society for Vascular Surgery"and"International Society for Cardiovascular Surgery".Based on the preoperative CT and DSA findings,the appropriate stent was selected for every patient.Post-operative clinical observation and CT scan were regularly carried out,the occurrence of complications and the morphological changes of the aneurysms were observed.The results were evaluated and analyzed.Results Primary technical success was achieved in all patients(100%).No death occurred during the procedure or in 30 days after the procedure.An average follow-up period of(29.1±20.5)months was made.Minor endoleak was noted on CT scans in 10 patients,and the endoleak disappeared in 5 patients during the follow-up period.One patient died from unknown cause.The total mortality rate was 2.0%(1/51).The major complications rate was 9.8%(5/51),including stent thrombosis(n=2),thrombosis at femoral artery(n=1),lymphatic fistula at femoral incision(n=1) and stent dislocation(n=1).Conclusion Endoluminal stent grafting is a safe and feasible technique for the treatment of abdominal aortic aneurysms with excellent medium-term results.This technique is especially suitable for the patients with high surgical risk.