Severe acute pancreatitis(SAP)is a rapid onset and complex acute digestive system emergency,often accompanied by severe local and systemic complications,leading to a high mortality rate.In recent years,with the continuous update of treatment concepts,especially the advancement of endoscopic and minimally invasive surgical techniques,the prognosis of SAP patients has significantly improved.However,in clinical practice,particularly in the early stages of the disease,there are still widespread issues of non-standard and imprecise diagnostic and therapeutic practices,which are important factors contributing to the occurrence of severe complications in the later stages of the disease and a major reason for the poor prognosis of SAP patients.In light of this,this article systematically reviews high-quality literature from recent years and combines the latest clinical research findings from the authors'team to summarize advancements in early precise diagnosis and treatment of SAP,providing new evidence for standardized treatment of SAP,which will undoubtedly help further improve the overall treatment outcomes for SAP.

Background and Aims:Accurate early pathogen diagnosis is a breakthrough for improving the prognosis of infectious pancreatic necrosis(IPN)patients.However,there is currently a lack of efficient methods for early identification of IPN in clinical settings.This study was performed to assess the application value of next-generation sequencing technology based on metagenomic capture(MetaCAP)in the pathogen diagnosis of IPN. Methods:A prospective study was conducted on 29 patients suspected of having acute necrotizing pancreatitis at Xiangya Hospital of Central South University between January and July 2024.Blood samples were tested using MetaCAP and conventional pathogen culture.The results of peritoneal fluid pathogen culture were used as the gold standard to compare the diagnostic efficacy of the two methods. Results:Due to three cases lacking peritoneal fluid culture results,a total of 26 cases were included in the final analysis.The overall mortality rate was 23.1％(6/26).During hospitalization,9 cases(34.6％)were diagnosed with IPN.The sensitivity and negative predictive value of MetaCAP for diagnosing IPN were significantly higher than those of conventional pathogen culture(77.8％vs.11.1％,P=0.031;86.7％vs.65.2％,P=0.032),while the differences in specificity(76.5％vs.88.2％,P=0.689)and positive predictive value(63.6％vs.33.3％,P=0.347)between the two methods were not statistically significant.The average detection time for MetaCAP was 33(20-49)h,while microbial culture took 125(45-142)h,with a significant difference(P＜0.001).The average cost for blood MetaCAP testing was 2 500 yuan per case,but it accounted for only 1.19％of the average hospitalization cost. Conclusion:MetaCAP has significant value in the early pathogen diagnosis of IPN,with a shorter detection time,good testing efficacy,and health-economic value,demonstrating a promising clinical application prospect.

Objective:To investigate the application value of metagenomic next-genera-tion sequencing (mNGS) in pathogenic diagnosis of suspected infected severe acute pancreatitis (SAP).Methods:The prospective study was conducted. The clinical data of 25 patients with suspected infected SAP who were admitted to the Xiangya Hospital of Central South University from May to September 2023 were collected. Upper limb venous blood samples of all the patients were collected for both of mNGS and routine pathogen microbial culture. Observation indicators: (1) grouping situations of the enrolled patients; (2) comparison of the diagnostic efficiency of mNGS and routine pathogen microbial culture; (3) results of peripheral blood pathogen microbial testing and peri-pancreatic effusion microbial culture; (4) testing time and cost. Measurement data with normal distribution were represented as Mean± SD, and comparison between groups was conducted using the independent sample t test. Measurement data with skewed distribution were represented as M( Q1, Q3). Count data were expressed as absolute numbers or percentages, and comparison between groups was conducted using the chi-square test. Results:(1) Grouping situations of the enrolled patients. A total of 25 patients were selected for eligibility. There were 18 males and 7 females, aged 48(40,59)years. The duration of hospital stay of 25 patients was 30(20,50)days. The etiologies of 25 patients included 14 cases of hyperlipidemic pancreatitis, 8 cases of biliary pancreatitis, 1 case of alcohol-induced acute pancreatitis, and 2 cases of pancreatitis caused by other causes. Of the 25 patients, there were 17 cases with infected pancreatic necrosis (IPN) including 7 cases of death, and 8 cases with sterile pancreatic necrosis including no death. (2) Comparison of the diagnostic efficiency of mNGS and routine pathogen microbial culture. The positive rates of mNGS and routine pathogen microbial culture in diagnosis of suspected infected SAP were 72.0%(18/25) and 32.0%(8/25), respectively, showing a significant difference between them ( χ2=8.01, P<0.05). The sensitivity and negative predic-tive value of mNGS and routine pathogen microbial culture in diagnosis of IPN were 94.1%(16/17), 35.3%(6/17) and 85.7%(6/7), 35.3%(6/17), showing significant differences between them ( χ2=12.88, 5.04, P<0.05). The specificity and positive predictive value of mNGS and routine pathogen microbial culture in diagnosis of IPN were 75.0%(6/8), 75.0%(6/8) and 88.9%(16/18), 75.0%(6/8), showing no significant difference between them ( χ2=0, 0.82, P>0.05). (3) Results of peripheral blood pathogen microbial testing and peripancreatic effusion microbial culture. Of the 17 patients with IPN, 36 strains of pathogenic bacteria were detected by mNGS, and 6 strains were detected by routine pathogen microbial culture. There were 16 of 17 patients with IPN showing positive mNGS pathogenic testing, of which 13 cases were consistent with the pathogenic testing results of peri-pancreatic effusion microbial culture, showing a consistency rate of 76.5%(13/17). There were 6 pati-ents with IPN showing positive routine pathogen microbial culture, with a consistency rate of 35.3%(6/17) to peripancreatic effusion microbial culture. (4) Testing time and cost. Testing time of mNGS and routine pathogen microbial culture were (43±17)hours and (111±36)hours, showing a signifi-cant difference between them ( t=9.31, P<0.05). Testing cost of mNGS was (2 267±0)yuan/case, accoun-ting for 1.7% of the hospitalization expenses of (133 759±120 744)yuan/case. Testing cost of routine pathogen microbial culture was (240±0)yuan/case, accounting of 0.2% of the hospitalization expenses. Conclusion:mNGS has important value for early pathogenic diagnosis of suspected infected SAP, and has a high timeliness.

Selective occlusion of tumor vasculature has proven to be an effective strategy for cancer therapy. Among vascular coagulation agents, the extracellular domain of coagulation-inducing protein tissue factor, truncated tissue factor (tTF), is the most widely used. Since the truncated protein exhibits no coagulation activity and is rapidly cleared in the circulation, free tTF cannot be used for cancer treatment on its own but must be combined with other moieties. We here developed a novel, tumor-specific tTF delivery system through coupling tTF with the DNA aptamer, AS1411, which selectively binds to nucleolin receptors overexpressing on the surface of tumor vascular endothelial cells and is specifically cytotoxic to target cells. Systemic administration of the tTF-AS1411 conjugates into tumor-bearing animals induced intravascular thrombosis solely in tumors, thus reducing tumor blood supply and inducing tumor necrosis without apparent side effects. This conjugate represents a uniquely attractive candidate for the clinical translation of vessel occlusion agent for cancer therapy.

Objective Human mitochondrial transcription termination factor 3 ( MTERF3 ) is a negative regulator of mito?chondrial gene expression and energy metabolism. This study was to construct a prokaryotic expression system for MTERF3 in Esche?richia coli ( E. coli ) and prepare its mouse?anti?human polyclonal antibody. Methods The complete open reading frame ( ORF) of human MTERF3 cDNA was amplified by RT?PCR and subcloned into prokaryotic expression vector pET28b. Then the recombinant plasmid pET28b?MTERF3 was transformed into competent E.coli BL21(DE3) and IPTG induced the expression of 6×his fusion protein. The recom?binant human MTERF3 protein was purified through Ni2+?NTA agar?ose gel column affinity chromatography and the purified recombinant protein was used as immunogen to immunize the BALB/c mice to pre?pare its specific polyclonal antibody. The titer and specificity of the antibody were analyzed by ELISA, Western blot and cellular immuno?fluorescence, respectively. Results The recombinant human MTERF3 protein was successfully expressed in E. coil and the mouse?anti?human MTERF3 polyclonal antibody with high quality was successfully prepared. ELISA showed that the titer of the antibody was 1:105 . Western blot and immunofluorescence detection revealed that the mouse?anti?human MTERF3 antibody could recognize the native MTERF3 antigen specifically. Conclusion Human MTERF3 expressed in the prokaryotic system has strong immunogenicity and the polyclonal antibody obtained from immunizing mice has high titer and specificity. The prokaryotic expression of human MTERF3 and the preparation of its antibody lay the foundation for further function research of human MTERF3.

BACKGROUNDnm23-H1 gene is a well-known tumor metastasis suppression gene. Our previous study has found that transfection of wild type nm23-H1 gene can significantly downregulate the ERK1/2 activity of human high-metastatic large cell lung cancer cell line L9981. The aim of this study is to investigate the influence of nm23-H1 and exogenous ERK1/2 pathway inhibitor U0126 on the extracellular signal-regulated kinase (ERK1/2) of human high-metastatic large cell lung cancer cell line L9981 and its malignant biological behaviors.

METHODSThe expressive levels of total-ERK1/2, dually phosphorylated ERK1/2 and ERK1/2 relative activity of the human high-metastatic large cell lung cancer cell lines, L9981 (parent cell line with nm23-H1 gene hetero-deletion), L9981-nm23-H1 (transfected with nm23-H1 gene ) and L9981-PLXSN (transfected with vector) were detected by Western blot and immunoprecipitation technique after treating with U0126 (40μmol/L for 20 minutes). The in vitro proliferative and invasive abilities among the above three lung cancer cell lines were determined by MTT and improved Boyden chamber methods.

RESULTSThe phosphorylated ERK1/2 expression level and relative activity in L9981-nm23-H1 lung cancer cell line were remarkably lower than those in L9981 and L9981-PLXSN lung cancer cell lines after being treated with U0126 (P < 0.01), but there was no significant difference between L9981 and L9981-PLXSN lung cancer cell lines. No significant difference of total ERK1/2 expression level was observed among the three lung cancer cell lines (P > 0.05) after being treated with U0126. The in vitro proliferation and invasion of L9981-nm-23H1 lung cancer cell line were remarkably lower than those of L9981 and L9981-PLXSN lung cancer cell lines (P < 0.01 ), but no significant difference was found between L9981 and L9981-PLXSN lung cancer cell lines (P > 0.05 ); U0126 could significantly down-regulate the in vitro proliferation and invasion of L9981 lung cancer cell line (P < 0.01).

CONCLUSIONSBlocking the activity of ERK1/2 in L9981 lung cancer cell line and transfecting the nm23-H1 gene into the L9981 lung cancer cell line may produce similar cell biological behavior changes, namely the significant reduction of in vitro proliferation and invasion of L9981 lung cancer cell line. These results indicate that the molecular mechanism which nm23-H1 gene reverses invasion and proliferation of the human high-metastatic large cell lung cancer cell line may be related to its effects of down-regulating the activity of the key kinase ERK1/2 of Ras-to-MAPK signal transduction pathway.

BACKGROUNDIt has been proved that tumor development and metastasis are dependent on angiogenesis. Suppression of tumor angiogenesis can inhibit tumor growth and metastasis. Collagen X VIII/endostatin is one of the most effective inhibitors of angiogenesis at present. The aim of this study is to study the relationship between transcription expression of endostatin mRNA and clinical and pathophysiological characteristics in non-small cell lung cancer (NSCLC).

METHODSThe transcription expression of endostatin mRNA was detected in 46 lung cancer tissues and paracancerous lung tissues, 14 benign pulmonary lesion tissues as control by RT-PCR method.

RESULTS(1)The transcription expression of endostatin mRNA in lung cancer tissues (0.872±0.071) was significantly higher than that in paracancerous tissues (0.717±0.073) and benign pulmonary lesion tissues (0.611±0.026) (P < 0.001).(2)The transcription expression of endostatin mRNA in lung cancer tissues was closely related to P-TNM stages, distant metastasis, grade of cell differentiation and size of the primary tumors (P < 0.05), but not to location of tumor, lymph node status, histological classification, age and sex of the patients and smoking or not (P > 0.05).

CONCLUSIONSThe transcription expression of endostatin mRNA in NSCLC tissues is significantly higher than that in paracancerous tissues and benign pulmonary lesion tissues, and is closely related to P-TNM stages, distant metastasis, grade of cell differentiation and size of the primary tumors, hence it might be helpful to evaluate the biological behavior of lung cancer.

BACKGROUNDIt has been known that oncogenesis and development of lung can- cer is a complex process regulated many genes and involved in multistages. Recent studies have demonstrated that signal transduction abnormality may play a very important role in these procedures. The aim of this study is to investigate the influence of exogenous MEK1/2 pathway inhibitor U0126 on the expression and activity of extracellular signal-regulated kinase (ERK1/2) of human high-metastatic large cell lung cancer cell line L9981 and its malignant biological behaviors.

METHODSThe expressive levels of total-ERK1/2, dually phosphorylated ERK1/2 and ERK1/2 relative activity of the human high-metastatic large cell lung cancer cell line L9981 (parent cell line with nm23-H1 gene hetero-deletion ) were detected by Western blot and immuno-precipitation technique after treating with different doses of U0126. The in vitro proliferative and invasive abilities of the lung cancer cell line were determined by MTT and modified Boyden chamber methods.

RESULTSThe ERK1/2 relative activity of L9981 gradually decreased accompanied with increase of U0126 doses, and a highly significant difference of phosphorylated ERK1/2 expression level existed among the different concentration groups of U0126 (P < 0.01), but no significant difference of total ERK1/2 expression was found among the different concentration groups of U0126 (P=0.387). After treatment with same concentration of U0126 for different time, the ERK1/2 relative activity of L9981 gradually decreased as the treatment time of U0126 prolonging, and a highly significant difference of phosphorylated ERK1/2 expression level was observed among different treatment time groups (P < 0.01). But no significant difference of total ERK1/2 expression level was found among different time groups (P=0.689). The inhibition of ERK1/2 pathway by MEK1/2 specific inhibitor U0126 targeting the ERK1/2 pathway of L9981 was dose- and time-dependent. After treatment with different concentration of U0126, the proliferation of L9981 gradually decreased accompanied with increase of U0126 concentration and a highly significant difference existed among different groups of U0126 concentration (P < 0.001). The in vitro invasion of L9981 gradually decreased accompanied with increase of U0126 concentration. No significant difference of in vitro invasion of L9981 was found among 0, 10 and 20μmol/l of U0126 (P > 0.05). A highly significant difference was observed when U0126 concentration increased to 40 and 60μmol/l compared with 0, 10 and 20μmol/l of U0126 (P < 0.001).

CONCLUSIONSThe inhibition of Ras-to-MAPK pathway by Ras-to-MAPK specific inhibitor U0126 targeting the Ras-to-MAPK pathway of the human high-metastatic large cell lung cancer cell line L9981 is dose-dependent and time-dependent. Suppressing or blocking of Ras-to-MAPK signal transduction pathway can reverse the invasive and metastatic phenotype of the human high-metastatic large cell lung cancer cell line L9981. These results suggest that the key kinase MEK1/2 of the ERK1/2 pathway may be a potent therapeutic target for human lung cancer.

nm23-H1 is a proven tumor metastasis suppressive gene, tumor metastasis phenotype could be reversed by transfected nm23-H1 cDNA. This study was conducted to transfect nm23-H1 cDNA into L9981 cells and to explore the function of nm23-H1 in reversing the malignant phenotype of L9981 cells. The plasmid of pLXSN-nm23-H1-EGFP was constructed by gene clone technique, and the transfected nm23-H1 cDNA cell lines of L9981-nm23-H1 was established. The protein expression of nm23-H1 was detected by Western blot. The biologic features of L9981-nm23-H1 cells were studied in vitro and in vivo. The results showed that the fusion protein of nm23-H1-EGFP was stable, continuous and expressed with high efficiency in L9981-nm23-H1 cells. The cell proliferation, colon formation and invasive ability are significantly lowered in L9981 cells transfected nm23-H1 cDNA (P < 0.01); the tumorgenesis and the lung metastasis incidence was lower in tranfected nm23-H1 cells than in L9981 and L9981-Plxsn in nude mice (P < 0.01); the rate for inhibiting tumorgenesis of nm23 -H1 was 82.56%. These data suggest that the malignant phenotype could be reversed by wild nm23-H1 gene in L9981 cells.
Animals ; Cell Line, Tumor ; Cell Proliferation ; Humans ; Lung Neoplasms ; pathology ; Mice ; Mice, Nude ; NM23 Nucleoside Diphosphate Kinases ; genetics ; Neoplasm Metastasis ; Neoplasm Transplantation ; Recombinant Fusion Proteins ; genetics ; Transfection ; Tumor Suppressor Proteins ; genetics

BACKGROUNDTo study the regulation of nm23-H1 gene on the expression of integrins in human high-metastasis large cell lung cancer cell line L9981 by transfecting nm23-H1 into cell.

METHODSLipofect was used to transfect nm23-H1 into the L9981 cell line; semi-RT-PCR was used to detect the difference of integrin β1 and integrin β3 mRNA expressions between tranfected and non-transfected cell lines; flow cytometry was used to detect the difference of integrin β1 and integrin β3 protein expressions between transfected and non-transfected cell lines.

RESULTS(1) A transgenic cell line L9981-nm23-H1 was obtained, which could stably and effectively express nm23-H1. The mRNA expressions of integrin β1 and β3 in L9981-nm23-H1 cell line were remarkably lower than those in L9981 cell line. (2) The protein expressions of integrin β1 and β3 in L9981-nm23-H1 cell line were significantly lower than those in L9981 cell line too (P < 0.01).

CONCLUSIONSTransfection of nm23-H1 gene into the L9981 cell line can significantly down-regulate the mRNA and protein expressions of integrin β1 and β3 in this cell line, which indicates that nm23-H1 may reverse the metastasis potential of L9981 cell line through modulation of integrin expression.