Objective To compare the morphology and ultrastructure changes of human gastric adenocarcinoma cell line treated by tachyplesin and n-sodium butyrate. Methods Light microscope,scanning electron microscope and transmission electron microscope were used to observe human gastric adenocarcinoma cell line BGC-823 treated by 2.0?mg/L tachyplesin,2.0?mmol/L n-sodium butyrate and 1.0?mg/L tachyplesin in combination with 1.0?mmol/L n-sodium butyrate respectively. Results Light microscope observation showed that BGC-823 cells treated by tachyplesin,n-sodium butyrate and tachyplesin+n-sodium butyrate possessed the similar cell modality as follow: the volume of cell increased,the shape of cell became flat and outspread,nucleo-cytoplasmic ratio decreased,the shape of nucleus was rounde,the number of nucleolus decreased.Scanning electron microscope and transmission electron microscope observation showed that,in the BGC-823 cells which were treated with tachyplesin,n-sodium butyrate and tachyplesin in combination with n-sodium butyrate respectively,microvilli and filopodia reduced,sheed pseudopodia increased,the shape of nucleus became regular,heterochromatin decreased while euchromatin increased,the number of mitochondria increased and its structure appeared consistent,Golgi complex turned to be typical,the amount of rough endoplasmic reticulum increased and that of polyribosome decreased.Conclusion All of these results showed that tachyplesin possessed the similar effects to n-sodium butyrate on changing morphology and ultrastructure of human gastric adenocarcinoma cells and possessed an additive role of inducing tumor cells to differentiate cooperatively with n-sodium butyrate to induce carcinoma cell differentiation.

Objective To study the value of transurethral electro-resection for renal pelvic tumor.Methods Transurethral electro-resection was performed on 11 patients after resection of the kidney.A F8 catheter was inserted into the bladder via the distal end of the ureter.The mucosal tissues around the ureter were then cut,and the ureter was fixed to the catheter and removed by pulling out the catheter.Results The operation was completed in all of the cases with a mean operation time of 115 min(65 to 170 min).None of the patients developed infection or hemorrhage after the surgery.11 patients were followed up for 6 to 18 months(mean,11 months),and no one died during the period.No tumor implantation or other intra-bladder masses were found.ConclusionsTransurethral electro-resection is feasible and safe for renal pelvic tumor.

Objective To investigate the effect of chloroform extract of Zingiber mioga Roscoe on human dermal microvascular endothelial cell (HDMEC) expression of cell adhesion molecules and adhesivity to lymphocytes.Methods HDMECs were isolated from human foreskin tissue and subjected to subculture.After several passages,some HDMECs were treated with chloroform extract from Zingiber mioga Roscoe (CFMG) of 200 mg/L for 30 minutes before or after 24-hour stimulation with tumor necrosis factor (TNF)-α or interleukin (IL)-1α.Subsequently,enzyme-linked immunosorbent assay was performed to determine the expression levels of intercellular adhesion molecule-1 (ICAM-1),vascular cell adhesion molecule-1 (VCAM-1) and E-selectin on HDMECs.Both T lymphocytes isolated from venous blood of healthy adults and Ramos Burkitt's lymphoma B cells were labelled with chromium-51 and incubated with the HDMECs for four hours followed by the determination of radiation intensity of lymphocytes adhering to HDMECs using a gamma-counter.Results Compared with 0.2％ dimethyl sulfoxide (DMSO),CFMG slightly down-regulated the expression of ICAM-1,VCAM-1 and E-selectin on resting HDMECs (all P ＞ 0.05).In comparison with culture medium,TNF-α enhanced the expressions of ICAM-1,VCAM-1 and E-selectin significantly (all P ＜ 0.01),and IL-1α elevated the expressions of ICAM-1 and E-selectin markedly(both P ＜ 0.01) as well as the expression of VCAM-1 slightly (P ＞ 0.05).The upregulating effects of TNF-α and IL-1α on the expressions of adhesion molecules were notably suppressed by the CFMG treatment prior to the stimulation with TNF-α and IL-1α(all P ＜ 0.01),but not affected by that after the stimulation (all P ＞ 0.05).The adhesivity of HDMECs to T lymphocytes and Ramos cells was slightly decreased by CFMG treatment compared with 0.2％ DMSO (both P ＞ 0.05),but was markedly increased by TNF-α and IL-1α compared with the culture medium (both P ＜ 0.01).Conclusions CFMG may play an antiinflammatory role via blocking the up-regulating effect of pre-inflammatory cytokines such as TNF-α and IL-1α on the expression of adherence molecules on HDMECs and,hence,inhibiting inflammatory cell infiltration in tissue.

ObjectiveTo observe the effects of different angles between tibial tunnel and tibial platform on “killer turn” in posterior cruciate ligament (PCL) reconstruction,and primarily discuss a safe and reasonable tunnel technology. Methods Eighteen fresh tendon grafts were used to reconstruct the PCL on the tibial side of fresh cadavers.The tibial tunnels of all specimens were built via anteromedial approach.Based on the different angles between tibial tunnel and tibial platform,all specimens were divided into Group A (30°),Group B (40°) and Group C (50°),with six specimens in each group.Area of tibial tunnel exit,pressure of tibia tunnel exit and circulation characteristics of tendons under the cyclic load before and after biomechanical test were recorded.ResultsThe area of tibial tunnel exit had statistical difference among three groups after the test ( F =8.80,P ＜ 0.05 ).The pressure of tibial tunnel exit had statistical difference among three groups (F =3.91,P ＜ 0.05 ).The cyclic frequency and fatigue strength of the transplanted tendons had statistical difference among three groups under the same cyclic load ( 256 N ) and same frequency ( 126 Hz ) ( F =4.25,P ＜ 0.05 ).Conclusions The angle between tibial tunnel and tibial platform has negative correlation with the area and pressure of tibial tunnel exit,and has positive correlation with the cyclic frequency and fatigue strength of the transplanted tendons under cyclic load.The ideal anatomy position of the tibial tunnel is the anteromedial tunnel with the angle of 40° between the tibial tunnel and the tibial platform.

Objective To further intensify the reform of public hospitals,promote talents team building and scientific research management innovation,enhancing the overall capacity of care delivery,as well as the development of science and technology in grass-roots hospitals.Methods A series of measures were adopted to arouse the enthusiasm of personnel to conduct research and finally increase the research outcomes.Concrete measures include talent training program and scientific research man-agement innovation,construction funding assurance,whole-process dynamic management,clarification of the quantitative evaluation index,research rewards,research funding management,as well as performance management.Results After implementation of such measures,the academic atmosphere changed a lot,medical technology improved,the quality of scientific research,project application and research outcomes increased dramatically which has statistical significance compare to previous situations.Conclusions The establishment of the incentive system plays a significant role.It helps in talent agglomeration during a relatively short period of time,exploring the potential capabilities of scientific research,enhancing the core competitiveness of hospital scientific research,which provide strong intellectual support and talent guarantee for hospitals development.

Objective To investigate the effects and mechanisms of shRNA interfered with expression of leucine-rich repeat containing G-protein coupled receptor 5 (Lgr5) on the malignant behaviors of colorectal cancer stem cells (CSCs).Methods The experimental study was conducted.The CSCs expressing Lgr5+ were sorted by fluorescence activated cell sorting.Lgr5+ cells that were transfected with Lgr5-shRNA lentiviral vector and nontarget shRNA lentiviral vector were respectively allocated into the experimental group and control group.The percentage of Lgr5+ cells was analyzed by flow cytometery.The relative expression of Lgr5 mRNA was detected by fluorescence quantitative real-time polymerase chain reaction (qRT-PCR).The capacity of self-renewal was detected by sphere forming assay.The tumorigenesis in vitro and in vivo were respectively measured by colony formation assay and xenografting experiment.The mRNA expressions of stem cells related genes (Oct4,Sox2,Nanog,KLF4),CSCs genes (CD133,CD44,ALDH) and Wnt/β-catenin pathway key genes (Axin2,Wnt5a,Wnt3a,Fzd3,c-myc,VEGF,Ascl2,claudin-1) were detected by qRT-PCR.Measurement data with normal distribution were represented as-x±s.Comparison between groups was analyzed using the t test.Results (1)Transfection efficiency of shRNA lentiviral vector induced Lgr5 by flow cytometery was respectively 6.8％± 1.0％ in the experimental group and 92.7％±3.3％ in the control group,with a statistically significant difference (t =43.148,P＜0.05).The relative expression of Lgr5 mRNA measured by qPT-PCR was respectively 0.168±0.057 in the experimental group and 1.148±0.004 in the control group,with a statistically significant difference (t=28.778,P＜0.05).(2) The capacity of self-renewal was detected by sphere forming assay.The results of sphere forming assay:the number of spheres was 29±6 in the experimental group and 410± 10 in the control group,with a statistically significant difference (t =41.070,P＜0.05).The results of colony formation assay:the numbers of colonies in the experimental group and control group were respectively 72±4 and 412± 19,showing a statistically significant difference (t =31.433,P＜ 0.05).The results of tumorigenesis:the volumes of tumors in the experimental group and control group were respectively (81± 15)mm3 and (328±24)mm3,with a statistically significant difference (t=11.304,P＜0.05).(3) The effects of Lgr5 down-regulation on related genes,results of qRT-PCR detection:① The mRNA relative expressions of Oct4,Sox2,Nanog and KLF4 (stem cells related genes) were 0.377±0.093,0.662±0.104,3.591±0.300,0.425±0.091 in the experimental group and 1.957± 0.026,2.137±0.015,5.831±0.165,1.536±0.014 in the control group,with statistically significant differences (t=23.079,22.261,8.446,19.186,P＜0.05).② The mRNA relative expressions of CD133,CD44 and ALDH (CSCs genes) were 1.490±0.155,5.535±0.487,1.640±0.039 in the experimental group and 2.488± 0.061,9.908±0.332,5.718±0.292 in the control group,with statistically significant differences (t =8.170,9.667,27.849,P＜0.05).③The mRNA relative expressions of Axin2,Wnt5a,Wnt3a,Fzd3,c-myc,VEGF,Ascl2 and claudin-1 genes in the Wnt/β-catenin pathway were respectively 1.592±0.267,0.528±0.138,2.153±0.078,1.480±0.064,0.248±0.128,1.492±0.025,0.658±0.095,1.647±0.087 in the experimental group and 3.651±0.224,2.570±0.093,2.301±0.157,1.636±0.058,1.415±0.080,2.610±0.159,2.480±0.123,3.432±0.273 in the control group.There were statistically significant differences in the mRNA relative expressions of Axin2,Wnt5a,c-myc,VEGF,Ascl2 and claudin-1 genes between the 2 groups (t =7.316,15.332,12.649,12.320,14.831,9.063,P＜0.05),and no statistically significant difference in the mRNA relative expressions of Wnt3a and Fzd3 between the 2 groups (t =2.887,2.242,P＞0.05).Conclusion The malignant behaviors of colorectal CSCs are suppressed after shRNA lentivirus interfered with expression of Lrg5,and its mechanism is related to inhibiting activity of Wnt/β-catenin pathway.

BACKGROUND:Tissue engineering has brought new hope to the clinical challenge of liver failure,and the preparation of plant-derived decellularized fiber scaffolds holds significant importance in liver tissue engineering. OBJECTIVE:To prepare apple tissue decellularized scaffold material by using fresh apple slices and a solution of sodium dodecyl sulfate,and assess its biocompatibility. METHODS:Fresh apples were subjected to decellularization using phosphate buffer saline and sodium dodecyl sulfate solution,separately.Afterwards,the decellularized apple tissues and apple decellularized scaffold materials were decontaminated with phosphate buffer saline.Subsequently,scanning electron microscopy was used to assess the effectiveness of decellularization of the apple materials.Adipose-derived mesenchymal stem cells were extracted from the inguinal fat BALB/C of mice,and their expression of stem cell-related markers(CD45,CD34,CD73,CD90,and CD105)was identified through flow cytometry.The cells were then divided into a scaffold-free control group and a scaffold group.Equal amounts of adipose-derived mesenchymal stem cells were seeded onto both groups.The biocompatibility of the decellularized scaffold with adipose-derived mesenchymal stem cells was evaluated using CCK-8 assay,hematoxylin-eosin staining,and phalloidine staining.Cell adhesion and growth on the scaffold were observed under light microscopy and scanning electron microscopy.Furthermore,the scaffold was subdivided into the non-induced group and the hepatogenic-induced group.Adipose-derived mesenchymal stem cells were cultured on the decellularized apple scaffold,and they were cultured for 14 days in regular culture medium or hepatogenic induction medium for comparison.Immunofluorescent staining using liver cell markers,including albumin,cytokeratin 18,and CYP1A1,was performed.Enzyme-linked immunosorbent assay was used to detect the secretion of alpha fetoprotein and albumin.Additionally,scanning electron microscopy was employed to observe the morphology of the induced cells on the scaffold,verifying the expression of liver cell-related genes on the decellularized scaffold material.Finally,the cobalt-60 irradiated and sterilized decellularized apple scaffolds were transplanted onto the surface of mouse liver and the degradation of the scaffold was observed by gross observation and hematoxylin-eosin staining after 28 days. RESULTS AND CONCLUSION:(1)The scanning electron microscopy results revealed that the decellularized apple scaffold material retained a porous structure of approximately 100 μm in size,with no residual cells observed.(2)Through flow cytometry analysis,the cultured cells were identified as adipose-derived mesenchymal stem cells.(3)CCK-8 assay results demonstrated that the prepared decellularized apple tissue scaffold material exhibited no cytotoxicity.Hematoxylin-eosin staining and phalloidine staining showed that adipose-derived mesenchymal stem cells were capable of adhering and proliferating on the decellularized apple tissue scaffold.(4)The results obtained from immunofluorescence staining and enzyme-linked immunosorbent assay revealed that adipose-derived mesenchymal stem cells cultured on the decellularized apple scaffolds exhibited elevated expression of liver-specific proteins,including albumin,alpha-fetoprotein,cytokeratin 18,and CYP1A1.These results suggested that they were induced differentiation into hepatocyte-like cells possessing functional characteristics of liver cells.(5)The decellularized apple scaffold implanted at 7 days has integrated with the liver,with partial degradation of the scaffold observed.By 28 days,the decellularized apple scaffold has completely degraded and has been replaced by newly-formed tissue.(6)The results indicate that the decellularized scaffold material derived from apple tissue demonstrates favorable biocompatibility,promoting the proliferation,adhesion,and hepatic differentiation of adipose-derived mesenchymal stem cells.

BACKGROUND: Lung cancer is the most common malignancy world-wide. Small cell lung cancer is the deadliest subtype of lung cancer, which features such as rapid growth, early metastasis, and high vascularization. Apatinib is a vascular endothelial growth factor receptor 2 inhibitor independently developed in China, which has a significant inhibition in a variety of solid tumors. The purpose of this study is to investigate the effects of Apatinib alone or Apatinib combined with mammalian target of rapamycin (mTOR) inhibitor, CCI-779, on small cell lung cancer cell line NCI-H446 in vitro. METHODS: The small cell lung cancer cell line NCI-H446 was grew in vitro. The effects of Apatinib alone or Apatinib combined with CCI-779 on proliferation, apoptosis, cell cycle and migration of NCI-H446 small cell lung cancer cells were detected by CCK8; FACS and transwell assays were also carried out; Western blot assays were used to detect vascular endothelial growth factor and cell cycle related protein expression. RESULTS: CCK8 assays showed that high concentration of Apatinib could inhibit the proliferation of NCI-H446 cells. Apoptosis assays showed that high concentration of Apatinib could induce NCI-H446 cell apoptosis. Transwell assays showed that high concentration of Apatinib could inhibit NCI-H446 cell migration. After combined with mTOR inhibitor CCI-779, low concentration of Apatinib could inhibit the proliferation and migration of NCI-H446 small cell lung cancer cells and induce apoptosis. CONCLUSIONS Apatinib has a concentration-dependent effect on the small cell lung cancer cell line NCI-H446. High concentration of Apatinib can inhibit the proliferation and migration of NCI-H446 small cell lung cancer cells, induce apoptosis. Apatinib combined with the mTOR inhibitor CCI-779 can sensitize the NCI-H446 cells to Apatinib.

BACKGROUND: Lung cancer is a malignant with high incidence and mortality and adenocarcinoma is among the most popular subtypes. Epidermal growth factor receptor (EGFR) mutation is one of the most important driver mutations for lung adenocarcinoma and EGFR-tyrosine kinase inhibitor (TKI) will benefit those patients with sensitive EGFR mutations. Recently, immune checkpoint inhibitor (ICI) therapy, provide a new breakthrough treatment for lung cancer patients. Whereas immunotherapy as an emerging treatment does not benefit patients with EGFR mutations, for which mechanistic studies are poorly defined and focused on the link of EGFR mutations and programmed cell death-ligand 1 (PD-L1) expression, we speculate that the different immune microenvironment associated with the two classes of patients. METHODS: Lung adenocarcinoma datasets were collected from the Cancer Genome Atlas (TCGA) database, and clinical information and gene expression profiles were downloaded. The immune related lymphocyte infiltration in TCGA database were generated through timer 2.0 GSEA was used to analyze the difference of pathway expression between EGFR mutant patients and wild type patients. RESULTS: EGFR mutation was more frequently among women and never smokers. Immunoinfiltration analysis showed that patients with EGFR mutation tends to have more tumor associated fibroblasts, common myeloid progenitor cells, hematopoietic stem cells, effector CD4⁺ T cells and natural killer T cells infiltration, and less memory B cells, naïve B cells, plasma B cells, plasmacytoid dendritic cells, memory CD4⁺ T cells, CD4⁺ helper T cells 2, naive CD8⁺ T cells, CD8⁺ T cells and central memory CD8⁺ T cells infiltration. Moreover, patients with more infiltration of CD8⁺ T cells, natural killer T cells, memory B cells and hematopoietic stem cells, tends have better prognosis (Log-rank test, P=0.017, 0.0093, 0.018, 0.016). However, the patients with more CD4⁺ T th2 infiltration in the tumor tends to have worse prognosis (Log-rank test, P=0.016). Furthermore, the results of gene set enrichment analysis showed that compared with the lung adenocarcinoma patients with EGFR wild type, the three pathways positive regulation of natural killer (NK) cell-mediated immune response to tumor cells, NK cell activation involved in immune response, and NK cell-mediated immune response to tumor cells related to natural killer cells in patients with EGFR mutation were down regulated, while the pathway the positive regulation of cytokine secretion involved in immune response was up-regulated in EGFR mutation patients. CONCLUSIONS The tumour microenvironment of patients with EGFR mutations lacks potent tumour killing effector cells and appears dysfunctional with effector cells. This may be a potential reason for the poor efficacy of immunotherapy in patients with EGFR mutations.

Objective: To acknowledge the availability and rates of annual transition of outcomes during the progression and regression stages of colorectal cancer (CRC) and related diseases, by pooling global follow-up studies on the natural history of CRC. Methods: Till March, 2017, data was collected through systematic literature review over multiple databases, including PubMed, Embase, Cochrane and Chinese Biology Medicine (CBM) disc. Information regarding the characteristics, classification system of health states, related outcomes and incidence rates on CRC or high-risk adenoma for the surveillance cohorts of the studies, were extracted and summarized. Both Meta and sensitivity analyses were performed on those outcomes if they appeared in more than 3 studies, using the random effects model. Annual transition rate with 95%CI was used to estimate each of the outcomes, Quality of the studies was assessed, using the Newcastle-Ottawa Scale. Results: A total of 29 cohort studies were included, with the mean follow-up period as 5.7 years. All studies except one, focused on adenoma-carcinoma pathway and reported the outcome parameters of adenomas by different risk, and some reported the findings on different sizes (n=6) of adenomas. These cohorts were divided into three groups (normal status, with low-risk or high-risk adenoma) according to the status of baseline endoscopic pathologic findings. Their available outcome parameters, corresponding number of involved articles, aggregated sample size and pooled annual transition rates were presented. Six parameters were obtained in the normal cohorts, including those from normal to low-risk adenoma (16 articles, 58 235, 0.030: 0.024-0.037), to high-risk adenoma (17 articles, 62 089, 0.003: 0.002-0.004), to diminutive adenoma (<5 mm, 4 articles, 1 277, 0.021: 0.013-0.029), to small adenoma (6-9 mm, 4 articles, 1 277, 0.006: 0.001-0.010), to large adenoma (≥10 mm, 7 articles, 3 531, 0.002: 0.000-0.003) and to CRC (19 articles, 104 836, 0.000 3: 0.000 2-0.000 5). Three parameters were obtained in low-risk adenoma in cohorts with polypectomy findings, including recurrence (9 articles, 4 788, 0.109: 0.062-0.157) from low-risk adenoma after polypectomy to high-risk adenoma (10 articles, 5 736, 0.009: 0.004-0.013) and to CRC (12 articles, 11 347, 0.000 6: 0.000 4-0.000 8). Three parameters were obtained on high-risk adenoma from cohorts with polypectomy findings, including recurrence (12 articles, 7 030, 0.038: 0.028-0.048) from high-risk adenoma after polypectomy to low-risk adenoma (8 articles, 2 489, 0.133: 0.081-0.185) and CRC (14 articles, 14 899, 0.002: 0.001-0.003). Except for normal to low-risk adenomas, results from the sensitivity analysis for the other parameters showed stable. Of the included studies, two presented incidence rates of CRC in different clinical stages and the another two were focusing on the parameters related to serrated pathway. Conclusions Globally, follow-up studies reported data on natural history of colorectal cancer is of paucity. Compared to the "adenoma-carcinoma" pathway, transition parameters of the serrated lesion pathway are more limited. This Meta-analysis provided convincing evidence for optimizing the strategies regarding follow-up program on the disease, using the baseline endoscopic findings from global CRC Screening Program. These results also offered strong data-related support for Chinese population- specific interventional model on colorectal cancer.