Multiomics integrated analysis and experimental validation identify TLR4 and ALOX5 as oxidative stress-related biomarkers in intracranial aneurysms.

BACKGROUND: Intracranial aneurysm (IA) is a severe cerebrovascular disease, and effective gene therapy and drug interventions for its treatment are still lacking. Oxidative stress (OS) is closely associated with the IA, but the key regulatory genes involved are still unclear. Through multiomics analysis and experimental validation, we identified two diagnostic markers for IA associated with OS. METHODS: In this study, we first analyzed the IA dataset GSE75436 and conducted a joint analysis of oxidative stress-related genes (ORGs). Differential analysis, functional enrichment analysis, immune infiltration, WGCNA, PPI, LASSO, and other methods were used to identify IA diagnostic markers related to OS. Next, the functions of TLR4 and ALOX5 expression in IA and their potential targeted therapeutic drugs were analyzed. We also performed single-cell sequencing of patient IA and control (superficial temporal artery, STA) tissues. 23,342 cells were captured from 2 IA and 3 STA samples obtained from our center. Cell clustering and annotation were conducted using R software to observe the distribution of TLR4 and ALOX5 expression in IAs. Finally, the expression of TLR4 and ALOX5 were validated in IA patients and in an elastase-induced mouse IA model using experiments such as WB and immunofluorescence. RESULTS: Through bioinformatics analysis, we identified 16 key ORGs associated with IA pathogenesis. Further screening revealed that ALOX5 and TLR4 were highly expressed to activate a series of inflammatory responses and reduce the production of myocytes. Methotrexate (MTX) may be a potential targeted drug. Single-cell analysis revealed a notable increase in immune cells in the IA group, with ALOX5 and TLR4 primarily localized to monocytes/macrophages. Validation through patient samples and mouse models confirmed high expression of ALOX5 and TLR4 in IAs. CONCLUSIONS: Bioinformatics analysis indicated that ALOX5 and TLR4 are the most significant ORGs associated with the pathogenesis of IA. Single-cell sequencing and experiments revealed that the high expression of ALOX5 and TLR4 are closely related to IA. These two genes are promising new targets for IA therapy.

Endoscopic submucosal dissection for anastomotic lesions after colorectal surgery.

BACKGROUND: Endoscopic submucosal dissection (ESD) for anastomotic lesions is technically challenging. We aimed to characterize the clinicopathologic characteristics, feasibility, and effectiveness of ESD for anastomotic lesions of the lower gastrointestinal tract. METHOD: We retrospectively investigated 55 patients with anastomotic lesions of the lower gastrointestinal tract who underwent ESD from February 2008 to January 2021. The lesions involving one or both sides of anastomoses were classified into the unilaterally involving anastomosis (UIA) or straddling anastomosis (SA) group, respectively. We collected clinicopathological characteristics, procedure-related parameters and outcomes, and follow-up data and analyzed the impact of anastomotic involvement. RESULTS: The mean age was 62.5 years, and the median procedure duration was 30 min. The rates of en bloc resection and R0 resection were 90.9% and 85.5%, respectively. Four patients (7.3%) experienced major adverse events (AEs). During a median follow-up of 66 months (range 14-169), seven patients had local recurrence, and six patients had metastases. The 5-year disease-free survival and overall survival rates were 82.4% and 90.7%, respectively. The 5-year disease -specific survival (DSS) rate was 93.3%. Compared with the UIA group, the SA group had significantly longer procedure duration, larger specimen, lower rates of en bloc resection and R0 resection, and shorter disease-free survival (all P < 0.05). However, rates of AEs did not differ significantly between the two groups. CONCLUSIONS: The short-term and long-term outcomes of ESD for colorectal anastomotic lesions were favorable. Although with technically challenging, ESD could be performed safely and effectively for lesions at the anastomoses.

Endoscopic submucosal dissection for lesions involving the anastomosis after upper gastrointestinal surgery.

BACKGROUND AND AIMS: Endoscopic submucosal dissection (ESD) for anastomotic lesions is technically challenging due to severe fibrosis, deformity, staples, and limited space for procedure. We aimed to characterize the clinicopathological characteristics, feasibility, and effectiveness of ESD for anastomotic lesions of the upper gastrointestinal tract. METHODS: We retrospectively investigated 43 patients with lesions involving the anastomoses of the upper GI tract who underwent ESD from April 2007 to February 2021. We collected clinicopathological characteristics, procedure­related parameters and outcomes, and follow­up data and analyzed the impact of anastomotic involvement. RESULTS: The median duration from previous upper GI surgery was 60 months and the median procedure duration was 30 min. The rate of en bloc resection and en bloc with R0 resection was 90.7% and 81.4%, respectively. Two patients (4.7%) experienced major adverse events, including delayed bleeding and febrile episode. During a median follow-up of 80 months, 3 patients had local recurrence and 4 patients had metastases. The 5-year disease-free survival (DFS) and overall survival (OS) rates were 89.6% and 95.1%, respectively. Compared with the unilaterally involving group, the straddling anastomosis group had significantly longer procedure duration, larger specimen, lower rates of en bloc resection and en bloc with R0 resection, and shorter DFS and OS (all P < 0.05). However, rates of adverse events did not differ significantly between the two groups. CONCLUSIONS: The short­ and long-term outcomes of ESD for upper GI anastomotic lesions were favorable. Although with technically challenging, ESD could be performed safely and effectively for anastomotic lesions.

Submucosal tunneling endoscopic resection treatment of multiple gastrointestinal submucosal tumors.

BACKGROUND AND AIM: Gastrointestinal submucosal tumors (SMTs) can be removed by submucosal tunneling endoscopic resection (STER). However, limited studies have evaluated STER for the removal of multiple upper gastrointestinal SMTs. The aim of this study was to evaluate the feasibility and outcomes of STER in the treatment of multiple upper gastrointestinal SMTs. METHODS: From January 2011 to April 2020, the cases of patients with multiple upper gastrointestinal SMTs undergoing STER were retrospectively analyzed. Variables of clinicopathological characteristics, major adverse events (mAEs), and follow up were collected and analyzed. RESULTS: Submucosal tunneling endoscopic resection was performed in 54 patients (48 male and 6 female patients) with 120 SMTs. Forty-four patients had two tumors, eight patients had three tumors, and two patients had four tumors. The median size of each patient was 1.8 cm (range 0.7 to 3.5 cm). Forty-five patients had tumors removed by one tunnel, and nine patients by two tunnels. The median procedure time was 50 min (range 14 to 120 min), and the mAE rate was 16.7% (9/54). No significant differences were found between patients with two tumors and those with > 2 tumors in terms of tunnel length, hospital stay, procedure time, and mAEs (all P > 0.05). In addition, patients with two tunnels had procedure time, hospital stay, and mAE rates comparable with those with one tunnel (all P > 0.05). No local recurrence or distant metastasis occurred during a median follow up of 64 months. CONCLUSIONS: Submucosal tunneling endoscopic resection is a safe and effective technique for the resection of multiple upper gastrointestinal SMTs.

IncRNA MVIH correlates with disease features, predicts treatment response and survival in pediatric acute myeloid leukemia.

OBJECTIVE: Long non-coding RNA microvascular invasion in hepatocellular carcinoma (lnc-MVIH) is correlated with unfavorable prognosis in several malignancies, while limitedly studied in pediatric acute myeloid leukemia (AML). This study aimed to investigate the correlation of lnc-MVIH with disease features, response to induction therapy, and survival in pediatric AML patients. METHODS: A total of 129 de novo pediatric AML patients who were retrospectively analyzed and 60 children with non-malignant hematological diseases who underwent bone marrow examination were reviewed as controls. Bone marrow mononuclear cells (BMMCs) were isolated from all participants to detect lnc-MVIH expression by reverse transcription-quantitative polymerase chain reaction. The complete remission status after 1 course of induction therapy, event-free survival, and overall survival of pediatric AML patients were recorded. RESULTS: Lnc-MVIH was upregulated in pediatric AML patients compared with controls (p < 0.001). In pediatric AML patients, lnc-MVIH was correlated with increased bone marrow blasts, less inv(16) or t(16;16) abnormity, and higher Chinese Medical Association (CMA) risk stratification (all p < 0.05), whereas its correlation with National Comprehensive Cancer Network (NCCN) risk stratification was not statistically significant (p = 0.098). As for prognosis, lnc-MVIH high expression patients presented with lower complete response rate to 1 course of induction therapy (61.5% vs. 79.7%, p = 0.024), shorter event-free survival (median 12.0 months vs. 22.0 months, p = 0.006), and overall survival (median 28.0 months vs. 42.0 months, p = 0.043) compared with lnc-MVIH low expression patients. CONCLUSION: Lnc-MVIH correlates with poor treatment response and unfavorable survival in pediatric AML.

Efficacy and safety of endoscopic submucosal dissection for submucosal tumors of the colon and rectum.

BACKGROUND AND AIMS: Endoscopic submucosal dissection (ESD) of colorectal submucosal tumors (SMTs) is becoming increasingly common; however, there have been few large consecutive studies analyzing its therapeutic efficacy and safety. The aim of this study was to evaluate the efficacy, safety, and long-term outcomes of ESD for colorectal SMTs. METHODS: This retrospective study included 412 consecutive patients with colorectal SMTs who underwent ESD at the Zhongshan Hospital of Fudan University from January 2008 to July 2014. Tumor histopathology, completeness of resection, adverse events, tumor recurrence, and distant metastasis were analyzed. RESULTS: Complete resection was achieved for 358 lesions (86.9%). Thirteen patients had serious adverse events (3.2%) including bleeding and perforation, and 28 patients (6.8%) had post-ESD electrocoagulation syndrome (PEECS). Because more ESDs for colorectal SMTs were performed by endoscopists, the rate of complete resection increased (78.5% vs 88.5%), and the rate of serious adverse events decreased (9.2% vs 2.0%). SMTs in the colon increased the risk of incomplete resection (19.6% vs 11.3%), serious adverse events (8.7% vs 1.6%), and PEECS (16.3% vs 4.1%). SMTs originating from the muscularis propria and sized ≥20 mm increased the rate of PEECS (22.7% vs 5.9% and 31.3% vs 5.8%, respectively). CONCLUSION: ESD is effective for resection of colorectal SMTs and rarely causes serious adverse events. Tumor location and the experience of endoscopists influence the complete resection rate and the development of serious adverse events. ESD is feasible for large tumors and tumors in the muscularis propria, but this is associated with relatively high risks of adverse events.

Endoscopic management of duodenal perforation caused by foreign bodies in adults: A retrospective study.

BACKGROUND AND AIMS: Duodenal perforation caused by foreign bodies (FBs) is very rare but is an urgent emergency that traditionally requires surgical intervention. Several case reports have reported the successful endoscopic removal of duodenal perforating FBs. Here we aimed to evaluate the safety and efficacy of endoscopic management of duodenal perforating FBs in adults. METHODS: Between October 2004 and October 2022, 12,851 patients with endoscopically diagnosed gastrointestinal FBs from four tertiary hospitals in China were retrospectively reviewed. Patients were enrolled if they were endoscopically and/or radiographically diagnosed with duodenal perforating FBs. RESULTS: The incidence of duodenal total FBs and perforating FBs was 1.9% and 0.3%, respectively. Thirty-four patients were enrolled. Endoscopic removal was achieved in 25 patients (73.5%), and nine patients (26.5%) received surgery. For the endoscopic group, most perforating FBs were located in the duodenal bulb (36.0%) and descending part (28.0%). The adverse events included 3 mucosal injuries and 1 localized peritonitis. All patients were cured after conventional treatment. In the surgical group, most FBs were lodged in the descending part (55.6%). One patient developed localized peritonitis and one patient died of multiple organ failure. The significant features of FBs requiring surgery included FB over 10 cm, both sides perforation, multiple perforating FBs and massive pus overflow. CONCLUSION: Endoscopic removal of duodenal perforating FBs is safe and effective, and can be the first choice of treatment for experienced endoscopists. Surgical intervention may be required for patients with FBs over 10 cm, both sides perforation, multiple perforating FBs, or severe infections.

Improving bowel preparation for colonoscopy with a smartphone application driven by artificial intelligence.

Optimal bowel preparation is a prerequisite for a successful colonoscopy; however, the rate of inadequate bowel preparation remains relatively high. In this study, we establish a smartphone app that assesses patient bowel preparation using an artificial intelligence (AI)-based prediction system trained on labeled photographs of feces in the toilet and evaluate its impact on bowel preparation quality in colonoscopy outpatients. We conduct a prospective, single-masked, multicenter randomized clinical trial, enrolling outpatients who own a smartphone and are scheduled for a colonoscopy. We screen 578 eligible patients and randomize 524 in a 1:1 ratio to the control or AI-driven app group for bowel preparation. The study endpoints are the percentage of patients with adequate bowel preparation and the total BBPS score, compliance with dietary restrictions and purgative instructions, polyp detection rate, and adenoma detection rate (secondary). The prediction system has an accuracy of 95.15%, a specificity of 97.25%, and an area under the curve of 0.98 in the test dataset. In the full analysis set (n = 500), adequate preparation is significantly higher in the AI-driven app group (88.54 vs. 65.59%; P < 0.001). The mean BBPS score is 6.74 ± 1.25 in the AI-driven app group and 5.97 ± 1.81 in the control group (P < 0.001). The rates of compliance with dietary restrictions (93.68 vs. 83.81%, P = 0.001) and purgative instructions (96.05 vs. 84.62%, P < 0.001) are significantly higher in the AI-driven app group, as is the rate of additional purgative intake (26.88 vs. 17.41%, P = 0.011). Thus, our AI-driven smartphone app significantly improves the quality of bowel preparation and patient compliance.

Research on the Impact of Members' Social Capital within Agricultural Cooperatives on Their Adoption of IPM in China.

Agricultural cooperatives are effective facilitators of green production technology promotion. What is the role of social capital within agricultural cooperatives with the most competitive advantage in technology promotion? Using the survey data of 465 citrus-planting cooperative members in Sichuan Province, this study uses the IV-probit model and mediating effect model to analyze the impact role of social capital within agricultural cooperatives on its members' adoption of integrated pest management (IPM) technology. The bootstrap method is also used to test the robustness of the parameter estimates. The results show that: (1) the social capital within agricultural cooperatives has a significant positive impact on IPM adoption; (2) cooperative members' IPM cognition has a partial mediating effect on the impact of the social capital within agricultural cooperatives on its members' adoption of IPM technology (more than 51.37%). Therefore, among all the optional IPM technology promotion measures of cooperatives, multi-dimensional accumulation of the social capital within agricultural cooperatives and promotion of IPM technology awareness level of members is a viable path.

Fluorescent biosensor based on FRET and catalytic hairpin assembly for sensitive detection of polysialic acid by using a new screened DNA aptamer.

A fast and efficient dsDNA library-immobilized magnetic bead-based SELEX technique was employed for selection of the aptamers against polysialic acid (PSA). Overall twelve rounds of screening, the pooled library was subjected to high-throughput sequencing. Five aptamer candidates with low Gibbs binding free energy and high abundance were selected for affinity evaluation. Apt3 was demonstrated to be the optimal aptamer for PSA with Kd of 114.0 nM. Furthermore, an ultrasensitive fluorescence resonance energy transfer (FRET)-based biosensor for PSA was constructed by employing the newly selected aptamer and catalytic hairpin assembly (CHA) amplification strategy. The linear detection range for PSA is from 10 pM to 1 µM and the limit of detection is 0.63 pM. The fluorescent biosensor is able to detect the target in the complex biological samples, which indicates that Apt3 has good application prospect for the biological detection and clinical diagnostics.

Endoscopic full-thickness resection without laparoscopic assistance for gastric submucosal tumors originated from the muscularis propria.

BACKGROUND: This study was designed to evaluate the clinical efficacy, safety, and feasibility of endoscopic full-thickness resection (EFR) for gastric submucosal tumors (SMTs) originated from the muscularis propria. METHODS: Twenty-six patients with gastric SMTs originated from the muscularis propria were treated by EFR between July 2007 and January 2009. EFR technique consists of five major procedures: (1) injecting normal saline into the submucosa and precutting the mucosal and submucosal layer around the lesion; (2) a circumferential incision as deep as muscularis propria around the lesion by the endoscopic submucosal dissection (ESD) technique; (3) incision into serosal layer around the lesion with Hook knife; (4) completion of full-thickness incision to the tumor including the serosal layer with Hook, IT, or snare by gastroscopy without laparoscopic assistance; (5) closure of the gastric-wall defect with metallic clips. RESULTS: EFR was successfully performed in all 26 patients without laparoscopic assistance. The complete resection rate was 100%, and the mean operation time was 105 (range, 60-145) min. The mean resected lesion size was 2.8 (range, 1.2-4.5) cm. Pathological diagnosis of these lesions included gastrointestinal stromal tumors (GISTs) (16/26), leiomyomas (6/26), glomus tumors (3/26), and Schwannoma (1/26). No gastric bleeding, peritonitis sign, or abdominal abscess occurred after EFR. No lesion residual or recurrence was found during the follow-up period (mean, 8 months; range, 6-24 months). CONCLUSIONS: EFR seems to be an efficacious, safe, and minimally invasive treatment for patients with gastric SMT, which makes it possible to resect deep gastric lesion and provide precise pathological diagnosis of it. With the development of EFR, the indication of endoscopic resection may be expanded.

Simulated microgravity, erythroid differentiation, and the expression of transcription factor GATA-1 in CD34+ cells.

INTRODUCTION: The microgravity environment of spaceflight leads to a series of changes in the human blood system. The aim of the present study was to examine the influence of simulated microgravity on the differentiation of CD34+ cells and to explore whether transcription factor GATA-1, required for the terminal differentiation of committed erythroid progenitor cells, is involved in this process. METHODS: CD34+ cells were cultured in the simulated microgravity conditions created by a rotary cell-culture system (RCCS). The effects of simulated microgravity on the differentiation and apoptosis of CD34+ cells were analyzed using flow cytometry and propidium iodide (PI) staining, respectively. Expression of GATA-1 mRNA in CD34+ cells was determined by real-time quantitative PCR. RESULTS: In the RCCS group, GlyA+ (glycophorin A) expression was lower and CD33+ expression higher than in the 1-g liquid control group (22.21% +/- 3.02% and 60.05% +/- 3.08%, vs. 52.12% +/- 1.92% and 18.87% +/- 1.41%, respectively). The proportion of differentiated cells in the 1-g methylcellulos e group (Gly+% = 54.39% +/- 2.86%, CD33+% = 21.09% +/- 3.19%) was similar to that in the 1-g liquid control group. As shown by real-time quantitative PCR, the relative expression of GATA-1 mRNA in the RCCS group was only 20% of that in the -g control group. CONCLUSIONSs: The differentiation of CD3+ cells, and especially erythroid differentiation, was inhibited by simulated microgravity by a mechanism that appears to involve the suppression of GATA-1 mRNA expression. The results of this study may be useful in understanding the critical effect of simulated microgravity on the pathogenesis of space anemia.

Silencing NOB1 Can Affect Cell Proliferation and Apoptosis Via the C-Jun N-Terminal Kinase Pathway in Colorectal Cancer.

OBJECTIVES: To investigate the bio-functions and the molecular mechanisms of NIN1/proteasome 26S subunit non-ATPase 8 binding protein 1 homolog (NOB1) in colorectal cancer cells. METHODS: NOB1 expression was silenced using si-RNA in SW480 and LoVo cells. The transfection efficiency was measured by western blotting and RT-qPCR. Subsequently, the proliferation of SW480 and LoVo cells was determined using both MTT assay and colony-formation assay. Apoptosis and cell cycle analysis were determined using flow cytometry. RESULTS: Compared with the normal control (NC) and scramble cells, si-NOB1 could significantly attenuate the proliferation, colony-formation ability and cell percentage of S stage (p < 0.05). Additionally, at the phosphorylation level, si-NOB1 could notably increase the expression of c-Jun N-terminal kinase (JNK), extracellular signal-regulated kinase (ERK) and p38. CONCLUSIONS: Inhibition of NOB1 expression suppressed the proliferation, and promoted the apoptosis through regulation of the JNK signaling pathway.

Prognostic significance of long non coding maternally expressed gene 3 in pediatric acute myeloid leukemia.

ABSTRACT: The purpose of this study was to evaluate the correlation of long non-coding RNA maternally expressed gene 3 (Lnc-MEG3) with disease features, treatment response, and survival in pediatric acute myeloid leukemia (AML) patients.Among 92 de novo pediatric AML patients (before treatment and after 1 course of induction) and 40 controls, bone marrow mononuclear cells were obtained. Then, Lnc-MEG3 expression was determined by reverse transcription quantitative polymerase chain reaction. After 1 course of standard induction therapy of pediatric AML patients, complete remission (CR) was assessed. Furthermore, event-free survival (EFS) and overall survival (OS) were determined according to follow-up data.Lnc-MEG3 was reduced in pediatric AML patients compared with controls. In pediatric AML patients, Lnc-MEG3 was correlated with French-American-Britain subtypes and lower Chinese Medical Association risk stratification, while it was not associated with cytogenetic features, FLT3-ITD mutation, CEBPA mutation, NPM1 mutation, WT1 mutation, or National Comprehensive Cancer Network risk stratification. After 1 course of treatment, Lnc-MEG3 exhibited an up-regulation trend. Furthermore, Lnc-MEG3 was of no difference before treatment between patients with and without CR, while elevated Lnc-MEG3 and change of Lnc-MEG3 after 1 course of treatment were associated with increased CR rate. Additionally, increased Lnc-MEG3 expression before treatment was associated with longer EFS but not OS, while enhanced Lnc-MEG3 expression after 1 course of treatment was correlated with both prolonged EFS and OS.Lnc-MEG3 may have clinical significance as a biomarker for assisting with disease management, treatment optimization, and prognosis improvement in pediatric AML patients.

Decreased lncRNA, TINCR, promotes growth of colorectal carcinoma through upregulating microRNA-31.

Abnormal expression in terminal differentiation-induced noncoding RNA (TINCR), a long non-coding RNA (lncRNA), has been reported in different human cancers, including colorectal carcinoma (CRC). Moreover, the molecular mechanisms that underlie the effects of TINCR on CRC remain unclear. Here, by a set of bioinformatics studies, we found that microRNA-31 (miR-31), the oncogenic miRNA that robustly upregulates in CRC, was a sponge miRNA for TINCR. TINCR and miR-31 levels were inversely correlated in both CRC tissues and CRC cell lines. Luciferase reporter assay revealed a specific binding site on TINCR for miR-31. Suppression of TINCR promoted CRC cell growth and migration in vitro, while overexpression of TINCR inhibited CRC cell growth and migration in vitro. TINCR depletion increased tumor xenograft growth in vivo, while TINCR overexpression inhibited it. Together, our study suggests that re-expressing TINCR may suppress invasive outgrowth of CRC through miR-31.

Glutamate receptor ionotropic, kainate 1 serves as a novel tumor suppressor of colorectal carcinoma and predicts clinical prognosis.

Colorectal cancer (CRC) is one of the most malignant cancers worldwide. However, the mechanisms of initiation and development of CRC are still largely unclear. The present study aimed to investigate the biological function and prognosis of glutamate receptor ionotropic, kainate 1 (GRIK1) in CRC. GRIK1 expression levels were analyzed in tissue microarrays containing 80 primary CRC samples using immunohistochemistry (IHC). The association between GRIK1 expression levels, clinicopathological factors and the prognosis was also investigated using Spearman's correlation analysis and Kaplan-Meier analysis, respectively. After genetic knockdown or overexpression of GRIK1, invasion/migration assays, proliferation assay, soft agar/colony formation assays, western blotting, reverse transcription-quantitative PCR and tumor xenograft models were used to investigate the function of GRIK1 both in vitro in two CRC cell lines, HCT116 and SW620, and in vivo. The results revealed that the expression levels of GRIK1 were significantly downregulated in CRC samples. Furthermore, IHC analysis indicated that the downregulated expression levels of GRIK1 were significantly associated with lymph node status and tumor size. In addition, patients with CRC with low GRIK1 expression levels demonstrated a consistently poor overall survival. The overexpression of GRIK1 inhibited the proliferation, colony formation, migration, invasion and epithelial-mesenchymal transition of HCT116 cells in vitro. In contrast, the genetic knockdown of GRIK1 promoted the proliferative, colony forming, migratory and invasive abilities of SW620 cells in vitro. Moreover, the overexpression of GRIK1 inhibited tumor growth, and liver and lung metastasis of CRC in vivo. In conclusion, the findings of the present study suggested that GRIK1 may serve as a tumor suppressor in CRC, and upregulated expression levels of GRIK1 may predict an improved prognosis for patients with CRC.

[Inhibition of tissue factor expression in endothelial cells by lentivirus mediated shRNA].

OBJECTIVE: To construct the recombinant lentivirus RNAi vector, and to determine whether the lentivirus mediated short hairpin RNA (shRNA) can inhibit the tissue factor (TF) expression in endothelial cells. METHODS: Two short hairpin RNAs targeting to human TF were cloned into pENTRTM/U6 plasmid to obtain an entry clone, and the positive clones were verified by sequencing. A recombination reaction was performed between the pENTR/U6 entry construction and pLenti6/BLOCKiTTM-DEST vector, and then the positive clones were confirmed by sequencing. The 293FT cell line was transfected by the above recombined plasmid and lentivirus packing materials, the culture supernatant was harvested, and the virus titer was determined. RT-PCR and ELISA were used to observe the inhibition of TF gene expression after the lentivirus transduction in human umbilical vein endothelial cells. RESULTS: The shRNA sequences targeting to human TF were cloned into the vectors, and an entry clone and an expression clone were constructed successfully, which were proved by sequence determination. Viral particles were packaged in the 293FT cell line, all virus stocks were collected, and the transfection titer was 5*10(5)/transduced unit. RT-PCR and enzyme linked immunosorbent assay demonstrated that the lentivirus stocks could suppress the TF expression in endothelial cells remarkably. CONCLUSION: Lentivirus RNAi vectors containing human TF gene are successfully constructed, and lentivirus mediated shRNA can inhibit the TF expression in endothelial cells, which may provide a highly effective method for the prevention and treatment of thrombo-embolic diseases.

Downregulation of miR-196b Promotes Glioma Cell Sensitivity to Temozolomide Chemotherapy and Radiotherapy.

Human glioma is the most common type of primary brain tumor. The survival rate of people with a malignant glioma is extremely low, primarily due to a lack of effective treatments. We previously reported that miR-196b expression is upregulated in glioblastoma tissues and overexpression of miR-196b is associated with poor prognosis. miR-196b acts as an oncogene by enhancing cellular proliferation and increasing the expression of E2F1, which plays an important role in the PI3K/AKT signaling pathway. In the present study, we explored the effects of miR-196b expression on glioma cells and characterized the relationship between miR-196b expression and the PI3K/AKT signaling pathway. We found that downregulation of miR-196b decreased the proliferation of U87 and U251 glioma cells. When anti-miR-196b and radiotherapy were used together, cellular proliferation decreased, whereas apoptosis and caspase 3/7activity, an indicator of apoptosis, increased. Meanwhile, downregulation of miR-196b remarkably inhibited glioma cell growth and colony formation when concurrent with temozolomide administration. Further studies demonstrated that neither upregulation nor downregulation of miR-196b markedly changed the protein expression levels of downstream molecules in the PI3K/AKT signaling pathway in cellular experiments. Therefore, whether miR-196b plays a role by activating the PI3K/AKT signaling pathway has not yet been determined. Together, our findings indicate that downregulation of miR-196b increased glioma cell sensitivity to temozolomide chemotherapy and radiotherapy and may be a valuable target when treating malignant gliomas. However, further studies are required to accurately characterize the mechanism by which miR-196b elicits its pivotal role.

[Impact of additional gastrectomy after endoscopic submucosal dissection on the prognosis of early gastric cancer].

OBJECTIVE: To investigate the impact of additional gastrectomy after endoscopic submucosal dissection(ESD) on the prognosis of early gastric cancer. METHODS: Clinical data of 107 early gastric cancer patients undergoing additional gastrectomy after ESD (research group, n=44) or radical surgery (control group, n=63) from January 2008 to December 2014 in Zhongshan Hospital were retrospectively analyzed. The reasons for additional gastrectomy after ESD included positive resection margin (n=10), lymphovascular invasion (n=5), well-differentiated mucosal tumor with a diameter >3 cm (n=10), poor-differentiated mucosal tumor with a diameter >2 cm (n=4), submucosal tumor(sm1) with a diameter >3 cm (n=10), and submucosal tumor(sm2) (n=9). Operation time, length of stay, lymph node metastasis, tumor recurrence and disease-free survival rate were compared between two groups. RESULTS: Baseline data of two groups were not significantly different (all P>0.05). After evaluation, absolute and relative indications were identified in 19 cases (43.2%) and 25 cases (56.8%) of research group, and in 28 cases (44.4%) and 35 cases(55.6%) of control group without significant difference (P=0.897). Lymph node metastasis occurred in 6 patients (4.5%) after surgery in research group and 6.3% in control group (P=0.690). Operation time was (218.5±74.3) minutes in research group and (219.8±81.8) minutes in control group (P=0.932). Length of stay was (10.0±12.3) days in research group and (10.8±9.9) days in control group (P=0.687). Follow-up time was (35.5±15.0) months in research group and (29.5±18.1) months in control group (P=0.072). Tumor recurrence rate was 4.5% in research group and 9.5% in control group (χ(2)=0.928, P=0.229). Mortality was 4.5% in research group and 7.9% in control group (χ(2)=0.487, P=0.485). Besides, no significant differences of operation mode (P=0.164), lymphatic clearance mode (P=0.330), number of harvested lymph node (P=0.467), morbidity of postoperative infection or fever (P=0.923) were found. Three-year tumor-free survival rate was 95.5% and 89.2% in research and control group respectively without significant differences (P=0.571). CONCLUSION: Additional gastrectomy after endoscopic submucosal dissection has no negative influence on the prognosis of patients with early gastric cancer, whose efficacy is similar to simple radical gastrectomy.