Objective: To construct the recombinant expressive vector of HSV-2-EGFP for the primary application in rapid,direct and sensitive diagnosis of herpes simplex virus infection. Methods: The gene fragment coding for HSV-2 ICP10 promoter was amplified by PCR,and the PCR product was cloned into pEGFP-1 to construct the recombinant plasmid pICP10-EGFP,which would be identified by DNA sequencing.The recombinant plasmid pICP10-EGFP were transfected into Vero cells by cation lipoid Lipofectamine 2000,then G418 was added to screen the positive cells to obtain stable cell line Vero-ICP10-EGFP.Vero-ICP10-EGFP was infected by various titers of HSV-2,and the EGFP fluorescence was detected under inverted microscope at 6,8 and 10 h post-infection.The specificity of the cell line was detected by infection with human cytomegalovirus(HCMV) and coxsackievirus. Results: The recombinant plasmid pICP10-EGFP was proved to be correct by the double restriction enzyme digestion and DNA sequencing.The Vero-ICP10-EGFP fluorescent emitting cells can be observed as early as 6 h after infection with HSV,with pronounced increase in the intensities at later hours.No induction of detectable fluorescence was detected by infection with HCMV and coxsackievirus 6,10 and 24 h post-infection. Conclusion: This novel GFP reporter system expressing the HSV-inducible EGFP reporter gene might provide a fast,easy and sensitive model for monitoring HSV in clinical specimens.

BACKGROUND:Fracture healing in diabetic patients is usual y unsatisfactory because of hormones and metabolic disorder, and an eventual multiple organ dysfunction resulting from high blood glucose. OBJECTIVE:To dynamical y observe the changes of cytokines during the fracture healing process in diabetic rats before and after insulin treatment. METHODS:A total of 120 Sprague-Dawley rats were included in this study. Of them, 90 rats intravenously injected with 5%tetraoxypyrimidine to induce rat models of diabetes were randomized into insulin treatment and diabetes groups, respectively. The remaining 30 rats were intravenously injected with equal volume of saline and selected as control group. The next day, blood glucose was determined. Healing at 1, 4, and 8 weeks after fracture were observed by the X-ray film. Biomechanical strength of the injured right tibia was measured at 4, 6, and 8 weeks after modeling. Cytokines in the osteotylus were determined by immunohistochemical staining and in situ hybridization technique. RESULTS AND CONCLUSION:The X-ray films showed that the speed of fracture healing in the diabetes group was slower than insulin treatment and control groups. Biomechanical strength of the osteotylus in the diabetes group was significantly decreased compared with the insulin treatment and control groups. However, there were no significant differences in above-mentioned parameters between the control and insulin treatment groups. Bone morphogenetic protein 2, basic fibroblast growth factor, transforming growth factor-beta, and vascular endothelial growth factor were widely expressed in the osteotylus and their expressions in diabetes group were significantly lower and slower than those in the control and insulin treatment groups. There was no statistical difference between control and insulin treatment groups. These results indicate that osteotylus formation speed, biomechanical strength, and growth factor expressions at the fracture site in diabetes rats were decreased compared with normal rats. Insulin treatment can enhance cytokine levels at the fracture site, thereby promoting the osteoblast proliferation and fracture healing.

Objective To assess the efficacy and safety of using a novel channel dilator (the Corsair microcatheter) accompanied with special occlusion guide wires for coronary chronic total occlusion (CTO) recanalization. Methods From 2011 December to 2013 August,we performed 89 cases (the study group) using channel dilator and the new special occlusion guide wires for CTO recanalization. Another 89 CTO lesions treated before using the corsair microcatheter were compared as the control group.We recorded clinical characteristic, outcome of PCI,radiation exposure time, contrast utilization and the procedure time. The MACE rate was monitored during follow up. Results The intracoronary channel dilator accompanied with special occlusion guide wires were inserted into 33 left anterior descending arteries, 17 left circumlfex arteries and 38 right coronary arteries. The success rates of procedure were signiifcantly higher in the study group than in the control group (91% vs. 67.7%, P < 0.05). Procedure and lfuoroscopy time tended to be lower in the study group than in the control group. There were no serious complications related to the catheter and no death case recorded. Conclusions The channel dilator accompanied with special occlusion guide wires may facilitate the conventional approach with a higher success rate in PCI.

The laboratory teaching of bacteriological analysis is very important for training practical analysis professionals.For better quality of laboratory teaching,in this paper,we practice a series of reform facilitating the lab teaching of bacteriological analysis,which is involved in the exploration and study of new curriculum system and teaching evaluation.The educational practices create effective teaching and learning environment and better performance.

Objective To investigate the pathogenesis of small cell lung cancer ( SCLC) and to ex-plore the expression of cell cycle related kinase ( CCRK) in SCLC and its clinical significance.Methods Reverse transcription polymerase chain reaction ( RT-PCR) and Western blot were performed to examine ex-pression of CCRK in SCLC and normal tissues.Results The expressions of gene [(0.51 ±0.11)IU/L] and protein [(0.61 ±0.13)IU/L] of CCRK in SCLC tissues were significantly higher than normal tissues [(0.30 ±0.08)IU/L, (0.34 ±0.09)IU/L] ( P <0.05).The expression of CCRK was closely correlated with the clinical curative effect ( P <0.05 ) rather than the clinical stages ( P >0.05 ) .Conclusions The expressions of gene and protein of CCRK in SCLC tissues were significantly higher than normal tissues. CCRK promoted the occurrence and progress of SCLC.Chem can restrain effectually the excessive expres-sion of CCRK.The expressions of gene and protein of CCRK in the different clinical curative effect group had significant difference.

ObjectiveTo study the apoptosis of CD4 + T lymphocytes and the detection of immune function in patients with pulmonary tuberculosis and explore the clinical significance.MethodsThe mononuclear cells were separated from the blood of the tuberculosis patients or the healthy.The flow cytometry was used to measure the percentage of apoptotic CD4 + T lymphocytes,and the standard of T-lymphocyte subsets were detected by using SAP technology.The red cell immune function were determined by using yeast wreath way.Results The apoptosis rate of CD4 + T lymphocytes and CD8 + T lymphocyte was ( 15.882 ± 4.65 ) ％,and (27.69 ± 0.74) ％.The Immune complex positive rate ( 19.40 ± 0.58) ％ in patients with tuberculosis was significantly higher than those in controls ( P ＜ 0.01 ).C3b receptor positive rate in red blood cells was ( 17.73 ± 0.63 ) ％,( 46.48 ± 1.34 ) ％ in CD3 + T lymphocyte,( 28.12 ±0.69 ) ％ in CD4 + T lymphocyte,and the ratio of CD4/CD8 ( 1.0223 ± 0.09362) in the patients with tuberculosis was lower than the control group( P ＜ 0.01 ).There were certain relationships between the apoptosis rate of CD4 + T lymphocytes and the percentages of CD4 + T lymphocyte,the standard of T lymphocyte subsets and the red cell immune function.ConclusionsThe apoptosis rate of CD4 + T lympho,cytes in patients with tuberculosis were significantly higher than the healthy,which led to reducing the number of CD4 + T lymphocytes.There was positive correlation between red cell immunity and T-lymphocyte immunity,and the immunity in red cell and T- lymphocyte was lower than normal controls,which may be related to the immune pathogenesis of pulmonary tuberculosis.

Refractory Crohn’s-like enterocutaneous fistula indicates the aggressive manifestation and lead to poor prognosis of patients. The development of multidisciplinary strategies for fistula administration largely subjects to the deficiency of animal model for disease remodeling and the underlying pathogenic mechanism. For the purpose, infected anal fistula model was conducted by BLV single-core electrolytic aluminum combined with dextran sodium sulfate. Notably, the inflammatory granulation tissue and inflammatory cell infiltration in the perianal tissue were arised on day 7 of the model by utilizing the Hematoxylin–eosin staining. With the aid of magnetic resonance imaging and signals of high-brightness. We intuitively observed the thickening and edema appeared in the fistula wall, which collectively suggested the formation of a fistula in the perianal area of the rat. Distinguish from the current models of anal fistula modeling including the body surface of fistula, backside of fistula and drainage wire of fistula, our model revealed multifaceted advantages such as quicker generation, higher modeling rate, preferable stability, better consistency, cost-effective, and in particular, more convenient to mimic clinical manifestations of anal fistula.

Objective:To analyze the expression of sequence similarity family 3 member C (FAM3C) in pancreatic cancer and its effects on pancreatic cancer cell proliferation, migration, invasion and the underlying mechanisms.Methods:Tissue samples from four pancreatic cancer patients undergoing surgical resection at Affiliated Hospital of Jiangnan University from July 2022 to June 2023 were collected. Expression of FAM3C was assessed in both cancer and adjacent tissues using polymerase chain reaction and Western blot techniques. High FAM3C-expressing pancreatic cancer cells PANC-1 and MIA PaCa-2 were selected for further experiments involving FAM3C-siRNA and methyltransferase-like protein 3 (METTL3)-siRNA groups, alongside a negative control group. Cell counting, Transwell assays, and co-immunoprecipitation with methylation antibodies were utilized to evaluate the proliferation, migration, invasion, and methylation status of FAM3C.Results:The relative expression of FAM3C mRNA in cancer tissues of the four patients with pancreatic cancer was higher than that in adjacent tissues (1.29±0.19 vs 0.47±0.17, t=-6.48, P=0.001). Compared with the negative control group A, the proliferation ability of the FAM3C-siRNA interference group decreased, and the number of migrating and invasive cells decreased (all P<0.05). In the negative control group B of PANC-1 cells, the relative expression of FAM3C mRNA in IgG-treated cells was lower than that in the methylated antibody-treated group (1.05±0.53 vs 30.57±1.09, t=-42.04, P=0.001). After PANC-1 cells were treated with methylated antibodies, the relative expression of FAM3C mRNA in the negative control group B was higher than that in the METTL3-siRNA interference group (30.57±1.09 vs 18.17±0.50, t=17.89, P=0.001). Compared with the negative control group B, the proliferation ability, migration and invasion cell numbers of PANC-1 cells in the METTL3-siRNA interference group were reduced (all P<0.05). The results of MIA PaCa-2 and PANC-1 cells were consistent. Conclusions:FAM3C is highly expressed in pancreatic cancer and promotes the proliferation, migration and invasion of pancreatic cancer cells. METTL3 affects FAM3C expression by methylation and proliferation, migration and invasion of pancreatic cancer cells.

Background and Objectives: Adipose tissue-derived mesenchymal stem cells (ASCs) are recognized as an advantaged source for the prevention and treatment of diverse diseases including type 2 diabetes mellitus (T2DM). However, alterations in characteristics of ASCs from the aforementioned T2DM patients are still obscure, which also hinder the rigorous and systematic illumination of progression and pathogenesis. Methods: and Results: In this study, we originally isolated peripancreatic adipose tissue-derived mesenchymal stem cells from both human type 2 diabetic and non-diabetic donors (T2DM-ASCs, ND-ASCs) with the parental consent, respectively. We noticed that T2DM-ASCs exhibited indistinguishable immunophenotype, cell vitality, chondrogenic differentiation and stemness as ND-ASCs. Simultaneously, there’s merely alterations in migration and immunoregulatory capacities in T2DM-ASCs. However, differing from ND-ASCs, T2DM-ASCs exhibited deficiency in adipogenic and osteogenic differentiation, and in particular, the delayed cell cycle and different cytokine expression spectrum. Conclusions The conservative alterations of T2DM-ASCs in multifaceted characteristics indicated the possibility of autologous application of ASCs for cell-based T2DM treatment in the future.

Objective To explore the value of echocardiography for preoperative diagnosis and surgical planning of Ebstein anomaly(EA)in children.Methods Echocardiography data of 216 EA children were retrospectively analyzed.The mitral valve-tricuspid valve distance(MTD),the ratio of distance from the root of anterior mitral leaflets to cardiac apex and distance from the root of the tricuspid valve septal leaflets to cardiac apex(MTR)were measured,and Glasgow outcome scale extended(GOSE)scores were calculated.The accuracy of MTD and MTR for diagnosing EA in children were compared,and the value of GOSE for surgical planning of EA was analyzed.Results Among 216 cases,descending tricuspid valve septum was observed with echocardiography in 199 cases.The accuracy of MTD＞15 mm,MTR＞1.2 and MTR＞1.5 for diagnosing EA in children was 81.91％(163/199),95.98％(191/199)and 68.34％(136/199),respectively,being significantly different(x2=9.802-51.856,all P＜0.001).Among 178 cases who underwent surgical operations,biventricular repair was performed in 101 cases,among them 90 cases with GOSE score≤1.00 recovered well after operation,while adverse events occurred in 5 cases among 11 cases(5/11,45.45％)with GOSE score＞1.00.One-and-a-half ventricle repair was performed in 62 cases who covered well after operation,including 22 cases with GOSE score≤1.00 and 40 cases＞1.00.Single-ventricular repair was performed in 15 cases with GOSE score＞1.00,and the patients recovered well after operation.Conclusion Preoperative echocardiography measured MTR＞1.2 was relatively reliable for accurately diagnosing EA in children.GOSE score was helpful for surgical planning.