To improve the status of management in respiratory failure in China,the project of Study on Pathogenesis and Treatment of Respiratory Failure was designed and conducted by three medical centers(Beijing Institute of Respiratory Medicine-Beijing Chaoyang Hospital,Affiliated to Capital Medical University,Zhongshang Hospital-Fudan University,Guangzhou Institute of Respiratory Medicine-First Guangzhou Medical College)for more than ten years.This project was focused on pathogenesis and treatment strategies of respiratory failure and achieved the following important innovations:(1)Pulmonary Infection Control Window(PIC Window)was firstly proposed and used to determine the time switching point of sequential invasive-noninvasive ventilation;(2)The largest sample size of early use of noninvasive positive pressure ventilation(NPPV)for acute exacerbated COPD(AECOPD)on general ward provided the evidence-based data for expanding the indication of NPPV from treating respiratory failure to alleviating respiratory muscle fatigue;(3)Three new types of masks with intellectual property for NPPV were developed;(4)Designing of intrinsic expiratory end positive pressure(PEEPi)lung model with property of expiratory flow limitation confirmed that PEEPi was the most important factor that increased inspiratory difficulty;(5)The systematic measurement was established for diaphragm strength and endurance;(6)Aquaporin 1(AQP1)was firstly proved the key channel of fluid transportation in the lung;(7)A multicenter prospective cohort study provided objective data that depression had causal effect on COPD exacerbation and hospitalization;(8)Two guidelines for NPPV and mechanical ventilation of AECOPD were initiated by this group.This project has been widely used in clinical practice and promoted the research and treatment of respiratory failure in China.

Objective To investigate the possibility of human papillomavirus late 1 (HPV-L1) protein detection in predicting the prognosis of cervical intraepithelial neoplasia (CIN). Methods (1)Through immunocytochemical method to detect expression of HPV-L1 protein in diagnosis of CIN before treatment. (2) Through hybridization and gene chip technology to detect expression of high-risk human papilloma virus (HR-HPV) in diagnosis of CIN after treatment for twelve months. Results (1) Comparing the three treatment methods in CIN patients, HR-HPV infection sustained no significant difference (P > 0.05). (2) The positive expression rate of HPV-L1 protein was 75.00% in CINⅠand 36.13%in CINⅡ~Ⅲbefore treatment, with statistically significant (P<0.05). (3) Persistent infection rate of HR-HPV was 5.97%in HPV-L1 positive expression group after treatment for twelve months and 19%in HPV-L1 negative expression group. The difference between the two was statistically significant (P<0.05). (4) The expression of HPV-L1 capsid protein before treatment had a negative relationship with HR-HPV persistent infection after 12 months of treatment. Conclusion Detection of HPV-L1 protein before treatment can predict the prognosis of CIN which had been treated.

Objective To develop a suitable extraction and purification method for detecting the residue of 20 kinds of organochlorine pesticides(OCPs) in suppositories of Chinese medicinal materials which are made of fat-soluble bases.Methods Different methods were applied for the extraction, and GPC was used for the purification of the sample Huazhi Shuan, and then the residue contents of OCPs were determined by GC- ECD method.20 kinds of OCPs were added to the samples and the feasibilities of the methods were evaluated by the obtained recovery. The established method were used in the analysis of the residue of OCPs in other Chinese herbal suppositories.Results The recovery and the reproducibility of this method met the requirements for the analysis of pesticides residues. Conclusion This developed method may provide reference for the detection of organochlorine pesticides residue in these kinds of Chinese medicinal preparations.

Objective To develop a suitable solid-phase extraction(SPE)method for the further purification of the test solu- tion in suppositories of Chinese medicinal materials which are made of lipid soluble bases,and to detect the residues of 20 kinds of organochlorine pesticides(OCPs).Methods Different kinds of SPE columns were selected and several elution solvents were used for the purification of the sample of herbal medicine of Huazhi Shuan,and then the residue contents of OCPs were determined by GC-ECD method.Twenty kinds of OCPs were added into the samples and the efficacy of the methods were eval- uated by the obtained recoveries.Results The recoveries of the 20 kinds of OCPs being detected were basically comply with the requirements for the analysis of pestiside residues.Conclusion After the tested solution is obtained from GPC column and is further purified through florisil SPE column,the impurity can be removed and the high accuracy of the quantitative analysis of the compounds can be achieved.

Tumors represent one of the primary threats to human life, with the dissemination of malignant tumors being a leading cause of mortality among cancer patients. Early diagnosis of tumors can reliably predict their progression, significantly reducing mortality rates. Tumor markers, including circulating tumor cells, exosomes, proteins, circulating tumor DNA, miRNAs and so on, generated during the tumor development process, have emerged as effective approach for early tumor diagnosis. Several methods are currently employed to detect tumor markers, such as polymerase chain reaction, Northern blotting, next-generation sequencing, flow cytometry, and enzyme-linked immunosorbent assay. However, these methods often suffer from time-consuming process, high costs, low sensitivity, and the requirement for specialized personnel. Therefore, a new rapid, sensitive, and specific tumor detection method is urgently needed.The clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein (Cas) system, originating from the adaptive immune system of bacteria, has found extensive applications in gene editing and nucleic acid detection. Based on the structure and function of Cas proteins, the CRISPR/Cas system can be classified into two classes and six types. Class I systems consist of multiple Cas protein complexes, including types I, III, and IV, while Class II systems comprise single, multi-domain Cas proteins mediated by RNA, including types II (Cas9), V (Cas12), and VI (Cas13). Class II systems have been widely employed in the fields of biotechnology and nucleic acid diagnostics due to their efficient target binding and programmable RNA specificity. Currently, fluorescence method is the most common signal output technique in CRISPR/Cas-based biosensors. However, this method often requires the integration of signal amplification technologies to enhance sensitivity and involves expensive and complex fluorescence detectors. To enhance the detection performance of CRISPR/Cas-based biosensors, the integration of CRISPR/Cas with some alternative techniques can be considered. The CRISPR/Cas integrated electrochemical sensor (E-CRISPR) possesses advantages such as miniaturization, high sensitivity, high specificity, and fast response speed.E-CRISPR can convert the reactions between biomolecules and detecting components into electrical signals, rendering the detection signals more easily readable and reducing the impact of background values. Therefore,E-CRISPR enhances the accuracy of detection results. E-CRISPR has been applied in various fields, including medical and health, environmental monitoring, and food safety. Furthermore, E-CRISPR holds tremendous potential for advancing the detection levels of tumor markers.Among all types of Cas enzymes, the three most widely applied are Cas9, Cas12, and Cas13, along with their respective subtypes. In this work, we provided a brief overview of the principles and characteristics of Class II CRISPR/Cas single-effector proteins. This paper focused on the various detection technologies based on E-CRISPR technique, including electrochemical impedance spectroscopy, voltammetry, photoelectrochemistry, and electrochemiluminescence. We also emphasized the applications of E-CRISPR in the field of tumor diagnosis, which mainly encompasses the detection of three typical tumor markers (ctDNA, miRNA, and proteins). Finally, we discussed the advantages and limitations of E-CRISPR, current challenges, and future development prospects. In summary, althoughE-CRISPR platform has made significant strides in tumor detection, certain challenges still need to be overcome for their widespread clinical application. Continuous optimization of the E-CRISPR platform holds the promise of achieving more accurate tumor subtyping diagnoses in clinical settings, which would be of significant importance for early patient diagnosis and prognosis assessment.

Aim To develop a simple and rapid method to monitor insulin receptor kinase activity and provide a novel cell-based model for screening anti-diabetes drugs.Methods CHO cells were co-transfected by plasmids which respectively contained insulin receptor gene,STAT5b gene and luciferase gene driven by STAT5 response elements.The expression of exogenous gene in transfected cells was examined by RT-PCR.The transfected cells were treated by insulin,and then the concentration and time-dependent response of luciferase expression to insulin induction was examined.Moreover,the specificity was identified by AG1024 treatment and PTP1B gene transfection.Results Expressions of insulin receptor and STAT5b were detected in the transfected CHO cells.The expression of luciferase in transfected cells was induced by insulin in concentration and time-dependent way.The maximal induction fold was 6.25.Moreover,the inducible expression of luciferase by insulin could be specifically blocked by tyrphostin AG1024,an inhibitor of insulin receptor kinase,or co-transfected PTP1B gene.Conclusions The insulin receptor kinase activity can be detected by expression of reporter gene with high sensitivity and specificity in this cell model,and with potential value in high throughput screening for insulin receptor activators and sensitizers.

BACKGROUND:Epidermal stem cells as special stem cells of the skin not only plays an important role in maintaining the metabolism of skin, but also is closely related to wound repair, which are the basis for the occurrence and repair of skin and its appendages. Now, epidermal stem cells have been paid great attention on the research of gene therapy and celltherapy with its specific biological advantages. OBJECTIVE:To summarize the research status on features and clinical application of epidermal stem cells. METHODS:A computer-based online retrieval of CNKI, PubMed database and Google scholar was performed for searching papers about clinical application of epidermal stem cells using the keywords of“epidermal stem cells, stem cells division, stem cells culture, clinical application”in Chinese and English. Older theoretical perspectives and repetitive research were excluded. Finally, only 41 articles were included in result analysis. RESULTS AND CONCLUSION:Epidermal stem cells have the great potential of proliferation and multi-directional differentiation, and have important significances for large-area skin defects (burn and trauma), skin tissue engineering, and gene therapy. There are stil many problems that need to be solved, such as how to screen surface markers special for epidermal stem cells and how to induce skin differentiation of epidermal stem cells.

Animal experiment is a primary part of medical education. At present , medical students are lack of education about laboratory animal ethics and they do not usually treat animals fol-lowing rules of laboratory animal ethics. To solve this problem, we focused on the education of labora-tory animal ethics and 3R theory (reduction, replacement, refinement) in order to help the medical students treat laboratory animals nicely. We enforced the construction of the laboratory rules and regu-lations, and improved animal experiments conditions. With these efforts, we successfully standardized and scientized our experiment classes.

Phospho-histone H2AX(γH2AX) has been widely used in vitro genotoxicity evaluation of ionizing radiation, carcinogenic substances and cigarette smoke as an important biomarker of DNA double stranded breaks ( DSBs) . The study developed an enzyme-linked immunesorbent assay for detection of the content ofγH2AX in cells to evaluate the genotoxicity of cigarette smoke. The study exposed CHO cells with cigarette smoke total particulate matter ( TPM) and cigarette smoke condensate ( CSC) of different dose, dose-effect relationship between smoke exposure and DNA damage and the poisonous difference of cigarette smoke components has been investigated by detecting the content change ofγH2 AX in cells under different exposure time and dose. In addition, reactive oxygen species ( ROS) in cells were also detected to study the mechanism of cigarette smoke exposure induced DSBs. The experiment result showed that the required time for the level ofγH2AX in cells grew to the peak prolonged with increased exposure dose. Once the level of γH2AX in cells grew to the peak value, then it decreased slowly; the level of γH2AX in cells rised with increasing dose of cigarette smoke;The effect of TPM on the content change of γH2AX was more than CSC; Moreover, smoke exposure could induce concentration increase of ROS in cells, and a good correlation of content change of ROS and γH2AX in cells were found, free radicals in cigarette smoke may be one major cause of DSBs.