Abstract: Objective To establish a sensitive and specific nucleic acid detection method for Schistosoma japonicum based on loop-mediated isothermal amplification (LAMP) and clustered regularly interspaced short palindromic repeats (CRISPR) technology. Methods The LAMP primers, gRNA and ssDNA probe that target Schistosoma japonicum SjR2 genes were designed according to the principles of LAMP and CRISPR. The LAMP-CRISPR reaction system was established and optimized. The sensitivity and specificity of the method were evaluated against the ten-fold serial dilutions of plasmid containing SjR2 target sequences, as well as genomic DNA at different stages of Schistosoma japonicum and other parasites, including Fasciola hepatica, Schistosoma mansoni, Taenia saginata, Clonorchis sinensis, Ascaris lumbricoides, Necator americanus, Paragonimus westermani, and Echinococcus granulosus. Additionally, 15 schistosome-infected snail and 30 uninfected samples were tested by LAMP-CRISPR and LAMP methods, respectively, to evaluate the potential of this method for screening for infected snails. Results　The developed LAMP-CRISPR method was able to specifically amplify and detect the SjR2 gene of S. japonicum. The optimal reaction temperature was 37 ℃, and the optimal reaction concentrations were both 40 nmol/L for gRNA and Cas12a protein. No cross-reaction was observed with genomic DNA from other parasites such as F. hepatica. The detection limit of the method was 10 copies/μL when testing 10-fold dilutions of recombinant plasmids as a template. Furthermore, the LAMP-CRISPR method was able to accurately detect genomic DNA from S. japonicum at various stages of development, including eggs, cercariae, schistosomula, juvenile worms, and adult worms. The results of testing 45 snail samples showed no significant difference between the LAMP-CRISPR and LAMP methods for detecting infected snails (χ2=0.05, P>0.05). The sensitivity and specificity of the LAMP-CRISPR method were 100.00% (15/15) and 96.67% (29/30), respectively, compared to the gold standard, while the sensitivity and specificity of the LAMP method were 100.00% (15/15) and 93.33% (28/30), respectively. Conclusions　This established LAMP-CRISPR detection method presented good sensitivity, specificity and reliability, making it a promising tool for rapid detection and risk monitoring of S. japonicum.

Objective: To investigate the current situation of stress ulcer prevention in patients with orthopedic joint replacement in our hospital and explore the necessity of preventing bleeding of peptic ulcer in patients with hip/knee replacement. Methods: The ca-ses of hip/knee replacement discharged from our hospital in September 2017 were retrospectively analyzed. The age and gender of the patients were analyzed. The underlying medical conditions, the related laboratory indices and the uses of stress ulcer prevention drugs, analgesic and anticoagulation drugs were statistically analyzed. Results: A total of 109 cases were collected including 40 cases of hip replacement and 69 cases of knee replacement. The median of age was 63. The use rate of non-steroidal analgesic drugs was 98. 16% , that of deep venous thrombosis prevention drugs was 95. 4% , and that of stress ulcer prevention drugs was 67. 9% . No patient suffered from gastrointestinal bleeding. Conclusion: The majority of patients undergoing hip /knee replacement are the elderly, and the knee replacement patients are older with more operation duration and more bleeding during the hip replacement. Nonsteroidal anti-inflamma-tory drugs and the drugs for preventing deep venous thrombosis are widely used during perioperative period. At present, it is still uncer-tain whether the population needs regular prevention of stress ulcers. However, the combination of aspirin and NSAID is a risk factor. The other high-risk factors, as well as the necessity of prevention medication, still need to be investigated in a larger scale.

ObjectiveTo investigate the efficacy of haploidentical blood and marrow transplantation (haplo-BMT) in the treatment of advanced chronic myeloid leukemia (CML).MethodsFrom November 2002 to October 2007,35 patients with advanced CML received haplo-BMT.Eleven patients achieved the second chronic phase (CP2) after treatment with imatinib or chemotherapy or both before pre-conditioning,but there were 13 cases in accelerated phase (AP) and 11 patients in blast phase (BP) at the time of transplantation.By the last follow-up date October 31,2011,the median follow-up time among living patients was 67 months (range,49 to 100 months).ResultsThe cases of HLA-antigen mismatched between donors and recipients as 1,2,and 3 antigens were 1,12,and 22 respectively.The number of mean mononuclear cells and CD34+ cells was (7.19+ 1.37) × 108/kg and (2.54± 1.50) × 106/kg,respectively.All but one patient achieved durable hematopoietic reconstitution. Hyperacute graft-versus-host disease (GVHD) occurred in 28.6％ (10/35) patients.The cumulative incidence of grade Ⅱ to Ⅳ acute GVHD was 48％.Among 27 patients who survived longer than 100 days after transplant,16 (60 ％) had chronic GVHD.Fiveyear overall survival (OS) rate was 46.2％ and 45.5％ in CML-AP and BP (P =0.97),respectively.Five-year probability of OS rate was 81.8％,30.8％ and 27.3％ in patients with CML-CP2,CML-AP and BP at transplant,respectively.The OS of CML-CP2 was significantly higher than CML-AP and BP at transplant (P＜0.01 ).ConclusionHaplo-BMT is a feasible therapeutic mean for patients with advanced CML who have no matched donors available.It is better to perform haplo-BMT at CML-CP2 other than CML-AP or BP.

Objective To construct a carbon dioxide(CO2)laser-induced corneal injury model in mice and observe the process of corneal wound healing.Methods Twenty eyes of ten C57BL/6J mice were divided into 4 groups.The cen-tral corneas in each group were irradiated by CO2 laser with a wavelength of 10.6 μm,spot diameter of 2 mm and power of 0.94 W.The exposure doses were 3.0 J·cm-2,4.5 J·cm-2,7.5 J·cm-2 and 10.5 J·cm-2,with corresponding expo-sure durations of 0.10 s,0.15 s,0.25 s and 0.35 s.Corneal injury severity was assessed using a slit lamp microscope,opti-cal coherence tomography and histopathological evaluation at 1 day after the exposure to determine the proper exposure dose for constructing a corneal injury model.Subsequently,the corneal injury model was constructed and the same meth-ods were used to monitor wound healing before,and 0 hours to 6 months after the exposure.Results No obvious corne-al injury was observed at an exposure dose of 3.0 J·cm-2.At an exposure dose of 4.5 J·cm-2,an off-white injury area appeared on the central cornea with loss of epithelium and endothelium.At an exposure dose of 7.5 J·cm-2 or 10.5 J·cm-2,the injury area became porcelain white,the cornea was thickened,and the iris was seen adhering to the margin of the cornea.Therefore,4.5 J·cm-2 CO2 laser was selected to construct a corneal injury model.At this exposure dose,the cornea swelled and thickened rapidly after injury,reached the maximum thickness 1 day after the exposure,and then grad-ually recovered,returning to normal by 14 days after the exposure.In the early stage(0 hours to 3 days after the expo-sure),the cornea showed shedding of injured epithelium and endothelium,migration of new epithelium and endothelium,and infiltration and regression of inflammatory cells.At the late stage(7 days to 6 months after the exposure),the cornea gradually returned to a normal physiological state,but some of the injured cornea exhibited stromal hyperplasia.Conclu-sion A CO2 laser with an exposure dose of 4.5 J·cm-2 can be used to construct a corneal injury model in mice.The a-cute phase of corneal injury primarily occurs within 3 days after the exposure.The cornea tends to restore its original physi-ological structure,but the corneal transparency cannot return to a normal state.

Background: In China, secondary cytoreductive surgery (SCR) has been widely used in ovarian cancer (OC) over the past two decades. Although Gynecologic Oncology Group-0213 trial did not show its overall survival benefit in first relapsed patients, the questions on patient selection and effect of subsequent targeting therapy are still open. The preliminary data from our pre-SOC1 phase II study showed that selected patients with second relapse who never received SCR at recurrence may still benefit from surgery. Moreover, poly(ADP-ribose) polymerase inhibitors (PARPi) maintenance now has been a standard care for platinum sensitive relapsed OC. To our knowledge, no published or ongoing trial is trying to answer the question if patient can benefit from a potentially complete resection combined with PARPi maintenance in OC patients with secondary recurrence. Methods SOC-3 is a multi-center, open, randomized, controlled, phase II trial of SCR followed by chemotherapy and niraparib maintenance vs chemotherapy and niraparib maintenance in patients with platinum-sensitive second relapsed OC who never received SCR at recurrence. To guarantee surgical quality, if the sites had no experience of participating in any OC-related surgical trials, the number of recurrent lesions evaluated by central-reviewed positron emission tomography–computed tomography image shouldn't be more than 3. Eligible patients are randomly assigned in a 1:1 ratio to receive either SCR followed by 6 cyclesof platinum-based chemotherapy and niraparib maintenance or 6 cycles of platinum-based chemotherapy and niraparib maintenance alone. Patients who undergo at least 4 cycles of chemotherapy and must be, in the opinion of the investigator, without disease progression, will be assigned niraparib maintenance. Major inclusion criteria are secondary relapsed OC with a platinum-free interval of no less than 6 months and a possibly complete resection. Major exclusion criteria are borderline tumors and non-epithelial ovarian malignancies, received debulking surgery at recurrence and impossible to complete resection. The sample size is 96 patients. Primary endpoint is 12-month non-progression rate.

Objective To explore the corneal wound healing mechanism after 3.74 μm infrared laser irradiation.Methods Twenty-seven C57BL/6J mice(six to eight weeks old)were randomly divided into a normal group(3 mice)and an experimental group(24 mice).Mice in the normal group were not subjected to any treatment.The corneas of mice in the experimental group were damaged by infrared laser at a wavelength of 3.74 μm.The spot diameter was 2 mm,the exposure duration 0.8 s,and the radiant exposure 23.2 J·cm-2.Pathological sectioning of corneas was performed at 3 h,6 h,12 h,1 d,3 d,7 d,14 d,and 21 d after laser irradiation in the experimental group,with 3 mice at each time point.It was the same for mice in the normal group.Immunohistochemical staining was conducted to examine neutrophil elastase,CD68,CD163 and thrombomodulin-positive cells and identify neovascularization.Results Neutrophil elastase,CD68,CD163,and thrombomodulin-positive cells were not detected in the corneal stroma of mice in the normal group.Neutrophil elastase-positive cells were detected in the damaged corneal periphery of mice in the experimental group at 3 h,migrated to the damaged area at 12 h,peaked at 1 d,and then decreased gradually with time,but still existed slightly at 21 d after laser irradiation.CD68-positive cells were detected in the damaged corneal periphery of mice in the experimental group at 12 h and in the damaged area at 1 d through 21 d after laser irradiation.CD163-positive cells were found in the damaged corneal periphery of mice in the experimental group at 7 d and in the damaged area at 14 d and 21 d after laser irradiation.Throm-bomodulin-positive cells were found in the damaged area of the corneal stroma at 14 d and 21 d after laser irradiation.Con-clusion During the wound healing process after 3.74 μm infrared laser-induced full-thickness corneal injury in mice,a large number of inflammatory cells migrate from the damaged corneal periphery or limbus to the damaged area.Neutrophils and M1 macrophages infiltrate in the early stage,while M2 macrophages are involved in the later stage,accompanied by neovascularization.

Objective: To detect the methylation status of ribosomal S6 kinase 4 (RSK4)in papillary thyroid carcinoma (PTC)and to study its correlation with mRNA expression and clinical features. Methods: 134 cases PTC tissues and corresponding paracancerous thyroid tissues were collected. DNA methylation status of RSK4 gene in PTC tissues and corresponding paracancerous tissues were analyzed by methylation specific PCR and bisulfite genomic sequencing, and mRNA expression was detected by quantitative realtime PCR. The relationship of DNA methylation status with mRNA expression and clinical features was analyzed. Results: The methylation rate of RSK4 in PTC tissues was significantly higher than that in paracancerous tissues by methylation specific PCR (P<0.05)and bisulfite genomic sequencing (P<0.01). The RSK4 mRNA level in PTC tissues was significantly lower than that in paracancerous tissues (P<0.01). In PTC tissues, RSK4 mRNA expression in methylation group was lower than that in unmethylation group (P<0.01). The RSK4 mRNA level in PTC of methylation was lower than that in paracancerous tissues of methylation (P<0.01); the RSK4 mRNA level in PTC of unmethylation was also lower than that in paracancerous tissues of unmethylated ones (P<0.01). There was relationship of RSK4 hypermethylation with lymphatic metastasis and TNM grade (P<0.05). Serum concentrations of thyroid stimulating hormone, thyroid peroxidase antibody, and thyroglobulin antibody in methylation group were higher than those in unmethylation group (P<0.05). Conclusion The hypermethylation of RSK4 promoter′s CpG islands might be one of the mechanisms for poor expression of RSK4 in PTC, which may serve as a molecular target of early diagnosis and treatment.