The feasibility and accuracy of the method for selecting the optimal size of double-lumen tube in thoracic surgery: a prospective, randomized controlled trial.

Double-lumen tubes (DLTs) are commonly used for one-lung ventilation (OLV) in thoracic surgery and the selection of an optimal size of DLTs is still a humongous task. The purpose of this study was to assess the feasibility and accuracy of the method for selecting an optimal size of DLTs in thoracic surgery. Sixty adult patients requiring a left side double-lumen tube (LDLT) for elective thoracoscopic surgery were included in this study. All patients were randomly allocated to the following two groups: Cuffs Collapsed group (CC group, n = 30) and Cuffs Inflated group (CI group, n = 30). In the Cuffs Collapsed group, the outer diameter of LDLT (the outer diameter of the tracheal and bronchial cuffs when they were collapsed as the outer diameter of the LDLT) matched with the inner diameter of the trachea and bronchus measured by the anesthesiologist on the chest CT slice; In the Cuffs Inflated group, the outer diameter of LDLT (the outer diameter of the tracheal and bronchial cuffs when they were inflated as the outer diameter of the LDLT) matched with the inner diameter of the trachea and bronchus measured by the anesthesiologist on the chest CT slice. The primary outcomes were the incidences of airway complications postoperative such as hoarseness and sore throat. The time of intubation and alignment, the incidences of LDLT displacement and adjustment, the peak airway pressure, the plateau airway pressure and the end-tidal carbon dioxide were also recorded. The incidences of airway complications postoperative such as sore throat and hoarseness were lower in the CI group than the CC group (P < 0.05), the intubation times was shorter in the CI group than the CC group (P < 0.05), while the peak airway pressure, the plateau airway pressure and the end-tidal carbon dioxide during two-lung ventilation and one-lung ventilation were no significant difference between two groups (P > 0.05). The method which matched the inner diameter of the trachea and bronchus measured on chest CT slice with the outer diameter of the tracheal and bronchial cuffs when they were inflated to select an appropriate size of LDLT can reduce the incidence of airway complications.Trials registration: Clinical Trials: gov. no. NCT05739318. Registered at https://classic.clinicaltrials.gov 22/02/2023.

Dual regulation effect and mechanism of human myeloid-derived suppressor cells on anticolorectal cancer cells activity of Vγ9Vδ2 T cells.

Myeloid-derived suppressor cells (MDSC) are a group of immature inhibitory cells of bone marrow origin. Human γδ T cells (mainly Vγ9Vδ2 T cells) have emerged as dominant candidates for cancer immunotherapy because of their unique recognition pattern and broad killing activity against tumor cells. Intestinal mucosal intraepithelial lymphocytes are almost exclusively γδ T cells, so it plays an important role in inhibiting the development of colorectal cancer. In this study, we investigated the effects and molecular mechanism of human MDSC on anticolorectal cancer cells activity of Vγ9Vδ2 T cells. Our results suggested that MDSC can reduce the NKG2D expression of Vγ9Vδ2 T cells through direct cell-cell contact, which is associated with membrane-type transforming growth factor-ß. In contrast, MDSC can increase Vγ9Vδ2 T cells activation and production of IFN-γ, perforin, Granzyme B through direct cell-cell contact. This may be related to the upregulation of T-bet in Vγ9Vδ2 T cells by MDSC. However, MDSC had a dominant negative regulatory effect on the anticolorectal cancer cells activity of Vγ9Vδ2 T cells. Our study provides a theoretical basis for the immune regulatory function of human MDSC on γδ T cells. This will be conducive to the clinical development of a new antitumor therapy strategy.

3-Methyladenine ameliorates surgery-induced anxiety-like behaviors in aged mice by inhibiting autophagy-induced excessive oxidative stress.

BACKGROUND: Postoperative anxiety is a common surgical complication in older patients. Research has recently linked excessive autophagy to several neurological disorders, including anxiety. This study aimed to determine whether 3-Methyladenine (3-MA) administration reduced anxiety-like behaviors in a mouse model following abdominal exploratory laparotomy. METHODS: An abdominal exploratory laparotomy model of postoperative anxiety was established using male C57BL/6 mice aged 20 months. 3-MA (6, 30, and 150 mg/ml) was administered via intracerebroventricular immediately following surgery. The mice were assessed 14 days after surgery using the marble burying, elevated plus maze tests, and local field potential recording in the amygdala. The levels of expression of phosphorylated-Akt, Beclin-1, LC3B, nuclear factor erythroid 2-related factor 2 (Nrf2)-occupied regions in NeuN-positive cells, superoxide dismutase (SOD) activity, malondialdehyde (MDA), and glutathione (GSH) were measured at 24 h after surgery. RESULTS: The injection of 3-MA reversed the increased number of marbles buried, decreased time spent in the open arm, and enhanced θ oscillation power after 14 days of abdominal exploratory laparotomy. In addition, administration of 3-MA reduced the ratio of phosphorylated- to total-Akt, decreased expression in Beclin-1 and LC3B, attenuated MDA levels, and increased the ratio of Nrf2-occupied areas in NeuN-positive cells, SOD activity, and GSH levels under abdominal exploratory laparotomy conditions. CONCLUSIONS: 3-MA improved anxiety-like behaviors in aged mice undergoing abdominal exploratory laparotomy by inhibiting excessive autophagy-induced oxidative stress. These results suggest that 3-MA could be an effective treatment for postoperative anxiety.

Appropriate tube temperature for fiberoptic bronchoscope-guided intubation of thermally softened double-lumen endotracheal tubes: A CONSORT-compliant article.

BACKGROUND: To compare the effects of thermal softening of double-lumen endotracheal tubes (DLT) at different temperatures during fiberoptic bronchoscopy (FOB)-guided intubation. METHODS: We randomly divided 144 patients undergoing thoracic surgery into 4 groups as follows: T1 (T = 24 ± 1°C, n = 36), T2 (T = 36 ± 1°C, n = 36), T3 (T = 40 ± 1°C, n = 36), and T4 (T = 48 ± 1°C, n = 36). All groups underwent FOB-guided double-lumen endotracheal intubation and positioning. We recorded the duration of positioning and intubation using DLT, intubation resistance (IR), the success rate of the first attempt at endotracheal intubation, and the incidence of postoperative vocal cord injury and hoarseness. RESULTS: The time to intubation was longer in the T1 group than that in the T2, T3, and T4 groups (P < .05). The time for positioning was longer in the T4 group than that in the T1, T2, and T3 groups (P < .05). IR was lower in the T3 and T4 groups than those in T1 and T2 groups (P < .05). The success rate of the first attempt at endotracheal intubation was higher in the T2, T3, and T4 groups than that in the T1 group (P < .05). Postoperative glottic injury and hoarseness were higher in the T1 and T2 groups than those in the T3 and T4 groups (P < .05). CONCLUSION: A thermally softened DLT shortened the time to intubation, reduced the IR, improved the success rate of the first attempt at endotracheal intubation, and lowered the incidence of postoperative glottic injury and hoarseness. The optimal tube temperature for FOB-guided intubation of thermally softened DLT was 40 ± 1°C.

Comparison of the accuracy of three methods measured the length of the right main stem bronchus by chest computed tomography as a guide to the use of right sided double-lumen tube.

BACKGROUND: The variation of right main stem bronchus leads to the orifice of the right upper lobe bronchus may be obstructed or increase the incidence of malposition intraoperatively when the right sided double-lumen tube is used. Therefore, the aim of this study was to compare the accuracy of three methods measured the length of the right main stem bronchus via chest computed tomography as a guide to the use of right sided double-lumen tube. METHODS: In this study, 168 adult patients undergoing left sided thoracic surgery were included. All these patients were allocated to carina-proximal (C-P) group, carina-distal (C-D) group and carina-carina (C-C) group. The position of endobronchial cuff observed via Fiberoptic bronchoscopy after successful initial placement and after turning the patients to the lateral decubitus position, as well as the incidence of malposition of right sided double-lumen tube intraoperative were recorded to assess the accuracy of three methods in predicting the position of right sided double-lumen tube. RESULTS: The distance between the carina to the proximal margin of the right upper lobe orifice, carina to the distal margin of the right upper lobe orifice and carina to the first right interlobar carina of the right upper lobe orifice were 17.2 ± 2.3 mm, 25.4 ± 3.7 mm and 28.5 ± 3.1 mm (P < 0.05). In the C-D group, the number of endobronchial cuffs seen to be herniating out of the carina, the number of bronchoscopies during initial placement and on the lateral position, the number of total malposition intraoperative and the number of reposition manoeuvres intraoperative were significantly less than the C-P group or the C-C group (P < 0.05). CONCLUSIONS: The length of the right main stem bronchus measured by the carina to distal margin of right upper lobe orifice method was more accurate than the other two methods in guiding the use of right sided double-lumen tube. TRIALS REGISTRATION: Clinical Trials. gov. no. NCT04127903. Registered at https://register. CLINICALTRIALS: gov on 16/10/2019.

Effects of head positions on awake fiberoptic bronchoscope oral intubation: a randomized controlled trial.

BACKGROUND: There are many factors affecting the success rate of awake orotracheal intubation via fiberoptic bronchoscope. We performed this study was to investigate the effects of head positions on awake Fiberoptic bronchoscope oral intubation. METHODS: Seventy-five adult patients, received general anaesthesia were included in this study. After written informed consent, these patients were undergoing awake orotracheal intubation via fiberoptic-bronchoscope and according to the head position, the patients were randomized allocated to neutral position group (NP group), sniffing position group (SP group) or extension position group (EP group). After sedation the patients were intubated by an experienced anesthesiologist. The time to view the vocal cords, the percentage of glottic opening scores (POGO), the time to insert the tracheal tube into trachea and the visual analog scale (VAS) scores for ease experienced of passing the tracheal tube through glottis, the hemodynamic changes and the adverse events after surgery were recorded. RESULTS: The time to view the vocal cords was significantly shorter and the POGO scores was significantly higher in the EP group compared with the other two groups (P < 0.05); The SpO2 in the EP group was higher than NP group at before intubation and higher than SP group and NP group at immediate after intubation (P < 0.05); The time to insert the tracheal tube into trachea, the VAS scores for passing the tracheal tube through glottis, the coughing scores had no significant differences among groups (P > 0.05). There were also no significant differences regard to the incidence of postoperative complications, mean arterial pressure and heart rate among the groups (P > 0.05). CONCLUSIONS: The head at extension position had a best view of glottic opening than neutral position or sniffing position during awake Fiberoptic bronchoscope oral intubation, so extension position was recommended as the starting head position for awake Fiberoptic bronchoscope oral intubation. TRIAL REGISTRATION: Clinical Trials.gov. no. NCT02792855. Registered at https://register.clinicaltrials.gov on 23 september 2017.

Fc-specific and covalent conjugation of a fluorescent protein to a native antibody through a photoconjugation strategy for fabrication of a novel photostable fluorescent antibody.

Fluorophore-antibody conjugates with high photobleaching resistance, high chemical stability, and Fc-specific attachment is a great advantage for immunofluorescence imaging. Here, an Fc-binding protein (Z-domain) carrying a photo-cross-linker (p-benzoylphenylalanine, Bpa) fused with enhanced green fluorescent protein (EGFP), namely photoactivatable ZBpa-EGFP recombinant, was directly generated using the aminoacyl-tRNA synthetase/suppressor tRNA technique without any further modification. By employing the photoactivatable ZBpa-EGFP, an optimal approach was successfully developed which enabled EGFP to site-selectively and covalently attach to native antibody (IgG) with approximately 90% conjugation efficiency. After characterizing the Fc-specific and covalent manner of the EGFP-photoconjugated antibody, its excellent photobleaching resistance for immunofluorescence imaging was demonstrated in a model study by monitoring the toll-like receptor 4 (TLR4) expression in HepG2 cells. The proposed approach here for the preparation of a novel fluorescent antibody is available and reliable, which would play an important role in fluorescence immunoassay, and is expected to be extended to the generation of other biomolecule-photoconjugated antibodies, such as other fluorescent proteins for multiplex immunofluorescence imaging or reporter enzymes for highly sensitive enzyme immunoassays.Graphical abstract.

The efficacy and adverse effects of the Uniblocker and left-side double-lumen tube for one-lung ventilation under the guidance of chest CT.

One-lung ventilation (OLV) is essential in numerous clinical procedures, in which the left-sided double-lumen tube (LDLT) is the most commonly used device. The application of bronchial blockers, including the Uniblocker or Arndt blocker, has increased in OLV. The present study aimed to compare the efficacy and adverse effects of the Uniblocker and LDLT for OLV under the guidance of chest CT. A total of 60 adult patients undergoing elective left-side thoracic surgery requiring OLV were included in the study. The patients were randomly assigned to the Uniblocker group (U group, n=30) or the LDLT group (D group, n=30). The time for initial tube placement, the number of optimal positions of the tube upon blind insertion, the number of attempts to adjust the tube to the optimal position, incidence of airway device displacement, injury to the bronchi and carina, the duration until lung collapse and the occurrence of sore throat and hoarseness over 24 h following surgery were recorded. The time for successful placement of the LDLT was 83.9±19.4 sec and that for the Uniblocker was 84.3±17.1 sec (P>0.05). The degree of lung collapse 1 min following opening of the pleura was greater in the D group than that in the U group (P<0.01) and the time required for the lung to completely collapse was shorter in the D group (3.3±0.5 min) than that in the U group (8.4±1.2 min; P<0.01). On the contrary, the incidence of injury to the bronchi and carina was lower in the U group (2/30 cases) than in the D group (10/30 cases; P=0.02); the incidence of sore throat was also lower in the U group (2/30 cases) compared with that in the D group (9/30 cases). The mean arterial pressure of patients immediately following intubation was lower in the U group (122.0±13.4 mmHg) than that in the D group (129.2±12.1 mmHg; P<0.05). The results of the present study indicated that the extraluminal use of the Uniblocker under guidance of chest CT is an efficient method with few adverse effects in left-side thoracic surgery. The study was registered at ClinicalTrials.gov on 16th December 2017 (no. NCT03392922).

Progranulin Regulates Inflammation and Tumor.

Progranulin (PGRN) mediates cell cycle progression and cell motility as a pleiotropic growth factor and acts as a universal regulator of cell growth, migration and transformation, cell cycle, wound healing, tumorigenesis, and cytotoxic drug resistance as a secreted glycoprotein. PGRN overexpression can induce the secretion of many inflammatory cytokines, such as IL-8, -6,-10, TNF-α. At the same time, this protein can promote tumor proliferation and the occurrence and development of many related diseases such as gastric cancer, breast cancer, cervical cancer, colorectal cancer, renal injury, neurodegeneration, neuroinflammatory, human atherosclerotic plaque, hepatocarcinoma, acute kidney injury, amyotrophic lateral sclerosis, Alzheimer's disease and Parkinson's disease. In short, PGRN plays a very critical role in injury repair and tumorigenesis, it provides a new direction for succeeding research and serves as a target for clinical diagnosis and treatment, thus warranting further investigation. Here, we discuss the potential therapeutic utility and the effect of PGRN on the relationship between inflammation and cancer.

A novel method of Uniblocker placement: extraluminal technique supported by trachea length measurement: A CONSORT-compliant article.

BACKGROUND: The use of bronchial blockers has been increased for one-lung ventilation; however, the placement of bronchial blockers is time consuming. The objective of this study was to compare the novel extraluminal technique of Uniblocker placement supported by trachea length measurement on computerized tomography images with conventional intraluminal Uniblocker placement method. METHODS: Seventy adult patients undergoing left side thoracic surgery were included in the study. All the patients were randomly assigned to one of two groups: conventional intraluminal intubation group (CV-IN group, n = 35) or extraluminal CT guided group (CT-EX group, n = 35). The primary endpoints were the optimal positions of Uniblocker and the injuries of bronchi and carina. The secondary outcomes included the time of Uniblocker placement, the adequacy of lung collapse, the incidences of Uniblocker displacement, sore throat, and hoarseness postoperative. RESULTS: In the CV-IN group, 19 of 35 Uniblockers went to the left main-stem bronchus on the initial blind insertion and 15 of 35 Uniblockers were considered as in optimal depth, whereas in the CT-EX group, 32 of 35 Uniblockers went to the left main-stem bronchus on the initial blind insertion and 31 of 35 Uniblockers were considered as in optimal depth (P < .01). The incidence of bronchi and carina injuries was obviously lower in the CT-EX group (occurred in 1 of 35 cases) than that in the CV-IN group (occurred in 8 of 35 cases) (P < .05). The time of Uniblocker placement took 145.4 s in the CV-IN group and 85.4 s in the CT-EX group (P < .01). The malpositions of Uniblocker, the degree of pulmonary collapse and the adverse events postoperative such as sore throat and hoarseness were not significantly different between the two groups (P > .05). CONCLUSION: The novel extraluminal technique of Uniblocker placement supported by trachea length measurement on computerized tomography images was proved to be more rapid, more accurate and less complications than conventional intraluminal Uniblocker placement method.

Leucine aminopeptidase 3 promotes migration and invasion of breast cancer cells through upregulation of fascin and matrix metalloproteinases-2/9 expression.

Overexpression of leucine aminopeptidase 3 (LAP3) is involved in proliferation, migration, and invasion of several tumor cells and plays a crucial role in tumor metastasis. However, the related mechanism remains unknown. In this study, we used MDA-MB-231 and MCF7 breast cancer cell lines to explore the role of LAP3 in the regulation of cancer cell migration and invasion by employing the natural LAP3 inhibitor bestatin and a lentivirus vector that overexpresses or knocks down LAP3. Bestatin inhibited tumor cell migration and invasion in a dose-dependent manner. Western blot assay showed that bestatin and knockdown of LAP3 upregulated phosphorylation of Hsp27 and downregulated expression of fascin. Phosphorylation of Akt and expression of matrix metalloproteinase-2/9 can also be downregulated. LAP3 overexpression showed the opposite results. Immunohistochemistry analysis was conducted to detect expression levels of LAP3 in breast cancer tissues. High LAP3 expression was correlated with the grade of malignancy. Findings of this study uncovered the molecular mechanism of LAP3 on breast cancer metastasis and indicated that LAP3 may act as a potential antimetastasis therapeutic target.

CD13 Inhibition Enhances Cytotoxic Effect of Chemotherapy Agents.

Multidrug resistance (MDR) of hepatocellular carcinoma is a serious problem. Although CD13 is a biomarker in human liver cancer stem cells, the relationship between CD13 and MDR remains uncertain. This study uses liver cancer cell model to understand the role of CD13 in enhancing the cytotoxic effect of chemotherapy agents. Cytotoxic agents can induce CD13 expression. CD13 inhibitor, bestatin, enhances the antitumor effect of cytotoxic agents. Meanwhile, CD13-targeting siRNA and neutralizing antibody can enhance the cytotoxic effect of 5-fluorouracil (5FU). CD13 overexpression increases cell survival upon cytotoxic agents treatment, while the knockdown of CD13 causes hypersensitivity of cells to cytotoxic agents treatment. Mechanistically, the inhibition of CD13 leads to the increase of cellular reactive oxygen species (ROS). BC-02 is a novel mutual prodrug (hybrid drug) of bestatin and 5FU. Notably, BC-02 can inhibit cellular activity in both parental and drug-resistant cells, accompanied with significantly increased ROS level. Moreover, the survival time of Kunming mice bearing H22 cells under BC-02 treatment is comparable to the capecitabine treatment at maximum dosage. These data implicate a therapeutic method to reverse MDR by targeting CD13, and indicate that BC-02 is a potent antitumor compound.

Response Surface Methodology for Optimizing the Ultrasound-Assisted Extraction of Polysaccharides from Acanthopanax giraldii.

The aim of this study was to investigate and optimize the most important factors affecting the extraction of Acanthopanax giraldii HARMS polysaccharides (AHPs) by ultrasound-assisted extraction (UAE) technology in a systemic manner. The ranges of four factors, including extraction temperature, liquid/solid ratio, extraction time, and ultrasonic power, were first determined by a single-factor experiment, followed by optimization of the UAE conditions using the Box-Behnken design (BBD) for maximum AHPs production. In our study, the models developed from the experimental design predicted the experimental data well and had a high determination coefficient (R2=0.9387). The optimized conditions for AHPs extraction were as follows: extraction temperature, 58°C; liquid/solid ratio, 25 : 1; extraction time, 73 min; and ultrasonic power, 85 W. Under these optimized conditions, the polysaccharide yield was 1.532±0.037% (n=3), being very close to the predicted value of 1.546% by the model. In addition, to investigate whether there was a difference of AHPs content between UAE and traditional hot water extraction (THWE), Fourier-transform (FT) IR spectral analyses was performed. The results showed that the functional groups of the polysaccharides extracted by either UAE or THWE were fundamentally identical. Furthermore, AHPs extracted by UAE could promote macrophage activation, such as enhanced phagocytosis and increased cytokine (interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α)) secretion in RAW264.7 cells. In conclusion, optimization of the UAE conditions by response surface methodology (RSM) was a promising method to improve the extraction yield of AHPs. AHPs extracted by the optimized UAE method can maintain their polysaccharide structure and biological activity.

[Knockdown of STAT3 inhibits proliferation and migration of HepG2 hepatoma cells induced by IFN1].

Objective To prepare lentiviruses expressing shRNA sequences targeting human signal transducer and activator of transcription 3 (STAT3) and detect the effect of STAT3 knockdown on type I interferon (IFN1)-induced proliferation and migration in HepG2 cells. Methods Four STAT3-targeting shRNA sequences (shRNA1-shRNA4) and one control sequence (Ctrl shRNA) were selected and cloned respectively into pLKO.1-sp6-pgk-GFP to construct shRNA-expressing vectors. Along with backbone psPAX2 and pMD2.G vectors, they were separately transfected into HEK293T cells to prepare lentiviruses. HepG2 cells were infected with the lentiviruses. Cytoplastic STAT3 level was detected by Western blotting to screen effective shRNA sequence(s) targeting STAT3. Proliferation and migration of HepG2 cells were analyzed by CCK-8 assay and TranswellTM migration and scratching assay, respectively. To detect the effect of IFN1 on cell proliferation and migration of HepG2 cells, the cells were treated with 2000 U/mL IFNα2b for indicated time and the activation of IFN-triggered STAT1 signal transduction was assayed by Western blotting. Results Two most effective STAT3-targeting shRNA sequences shRNA1 and shRNA2 were selected, and the expression of both STAT3 shRNA significantly decreased proliferation and migration of HepG2 cells. When treated with IFNα2b, 2000 U/mL of IFN1 showed more competent in attenuating growth and migration of HepG2 cells. Our data further proved that knockdown of STAT3 increased the phosphorylation of STAT1, and IFNα2b further enhanced the activation of STAT1 signaling in HepG2 cells. Conclusion Knockdown of STAT3 inhibits cell migration and growth, and rescues IFN response through up-regulating STAT1 signal transduction in HepG2 hepatoma cells.

A comparison of extraluminal and intraluminal use of the Uniblocker in left thoracic surgery: A CONSORT-compliant article.

BACKGROUND: The aim of this study was to assess the feasibility and safety issues concerning extraluminal use of the Uniblocker for one-lung ventilation (OLV) in the left thoracic surgery. METHODS: Forty patients undergoing elective left thoracic surgery were included in this study, and all patients were randomly allocated to extraluminal use of Uniblocker group (E group, n = 20) or intraluminal use of Uniblocker group (I group, n = 20). Time for intubation, time for verification of the correct position of Uniblocker, incidence of Uniblocker displacement, index of pulmonary collapse, mean arterial pressure, heart rate, peak airway pressure, oxygen saturation in two-lung ventilation, and 30 minutes after OLV, bronchial damage after OLV, sore throat, and hoarseness postoperative were recorded. RESULTS: The time for positioning Uniblocker was 112.6 ±â€Š31.2 seconds in intraluminal use group, whereas the time for positioning Uniblocker was significantly shorter in extraluminal use group (63.4 ±â€Š15.8 seconds). The incidence of main bronchial injury, the time of intubation, the incidence of Uniblocker malposition after initial placement, the time of OLV, the degree of pulmonary collapse, mean arterial pressure, heart rate, peak airway pressure, oxygen saturation in two-lung ventilation, and 30 minutes after OLV, the incidence of sore throat and hoarseness postoperative have no statistical significance (P > .05). CONCLUSION: Extraluminal use of the Uniblocker was proved to be a more rapid and more accurate method than conventional intraluminal use of the Uniblocker for OLV in left thoracic surgery.

LASP-1 promotes tumor proliferation and metastasis and is an independent unfavorable prognostic factor in gastric cancer.

PURPOSE: The LIM and SH3 protein 1 (LASP-1) is a focal adhesion protein, and its expression has been reported to be increased in many malignant tumors. However, the role of LASP-1 in gastric cancer is still unknown. The aim of this study was to determine the relationship of LASP-1 expression with the progression and prognosis of gastric cancer. METHODS: Expression of LASP-1 was evaluated in gastric cancer tissues and cell lines by immunohistochemistry and Western blot analysis. The relationship between LASP-1 expression and clinicopathological characteristics was analyzed. Using RNA interference, the effects of LASP-1 on cell proliferation, migration and invasion were investigated in gastric cancer cell lines both in vitro and in vivo. RESULTS: The LASP-1 was overexpressed in gastric cancer tissues and cell lines. LASP-1 expression was significantly associated with tumor size, invasive depth, TNM stage, lymph node metastasis and p53 expression (all P < 0.05). Multivariate survival analysis showed that LASP-1 expression was recognized as an independent prognostic factor of patient's survival. Knockdown of LASP-1 inhibited cell proliferation, migration and invasion in vitro as well as tumorigenesis and metastasis in vivo. CONCLUSIONS: Our study showed that LASP-1, overexpressed in gastric cancer and associated with poor prognosis, plays an important role in the growth and metastasis of gastric cancer.

[The expression of human IL-1ß gene containing human erythropoietin signal peptide in HepG2 cells].

OBJECTIVE: To construct two lentiviruses secreting human IL-1ß through either classical or nonclassical pathway and analyze their expressions in HepG2 cells after packaging lentiviruses and infecting hepatoma carcinoma HepG2 cells. METHODS: Human full-length IL-1ß gene and chimeric gene containing human erythropoietin(EPO) signal peptide and mature IL-1ß protein coding sequence were respectively amplified from pIRES2-EGFP-proIL-1ß and pIRES2-EGFP-epoIL-1ß using PCR. The sequences were subsequently cloned into lentiviral expression vector pLenti6/V5 to construct pLenti6/V5-proIL-1ß, which expressed IL-1ß through nonclassical pathway, and pLenti6/V5-epoIL-1ß, which expressed IL-1ß through signal-peptide mediated classical pathway. Lentiviruses expressing human IL-1ß through either classical or nonclassical pathway were packaged in HEK293T cells using a three-plasmid packaging system, and then these viruses were used to infect HepG2 cells. The level of IL-1ß in both cytoplasm and culture supernatant were detected by sandwich ELISA and Western blotting. RESULTS: pLenti6/V5-proIL-1ß expressing human full-length IL-1ß gene and pLenti6/V5-epoIL-1ß expressing human EPO signal peptide and mature IL-1ß gene were successfully constructed and confirmed through enzymatic assay and DNA sequencing. The lentiviruses expressing IL-1ß through different pathways were prepared using a three-plasmid packaging system in HEK293T cells. Compared with the cells infected with control virus, levels of supernatant and cytoplasmic IL-1ß in the cells infected with two lentiviruses expressing IL-1ß through different pathways were markedly elevated (P<0.01). However, level of mature IL-1ß in supernatant of HepG2/epoIL-1ß cells was much higher than that of HepG2/proIL-1ß cells, while total IL-1ß level in cytoplasm of HepG2/proIL-1ß cells was significantly higher than that in HepG2/epoIL-1ß cells. CONCLUSION: Both classical and nonclassical pathway secretion vectors could express human IL-1ß in HepG2 cells, but EPO signal-peptide mediated classical pathway secreted much higher mature IL-1ß than that of nonclassical pathway.

PTTG1 promotes migration and invasion of human non-small cell lung cancer cells and is modulated by miR-186.

Deeper mechanistic understanding of non-small cell lung cancer (NSCLC), a leading cause of total cancer-related deaths, may facilitate the establishment of more effective therapeutic strategies. In this study, pituitary tumor transforming gene (PTTG1) expression was associated with lymph node and distant metastasis in patients with NSCLC and was correlated with patient survival. Reduction of PTTG1 by small interfering RNA (siRNA) inhibits the migration and invasion of NSCLC cells by mediating matrix metalloproteinases expression. To the best of our knowledge, this study is the first to report that PTTG1 promotes epidermal growth factor (EGF) induced the phosphorylation of LIN-11, Isl1 and MEC-3 protein domain kinase and cofilin, a critical step in cofilin recycling and actin polymerization. Additionally, EGF-induced Akt phosphorylation was suppressed through knockdown of PTTG1. Interestingly, miR-186 can modulate PTTG1 protein expression. As observed from the animal experiment in this study, knockdown of PTTG1 through siRNA and overexpression of miR-186 inhibited invasive activity of NSCLC cells toward the SCID mice lung. In summary, our in vitro and in vivo results indicate that PTTG1 modulated by miR-186 has an important function in NSCLC invasion/metastasis. This study identified both PTTG1 and miR-186 as potential anti-invasion targets for therapeutic intervention in NSCLC.

[The expression vector of murine secreting IL-1ß promotes proliferation and migration of Hepa1-6 hepatoma cells].

AIM: To establish a murine secreted mature peptide IL-1ß expression vector, transfect into Hepa1-6 hepatoma cells, and analyze the effect of recombinant IL-1ß on proliferation, migration, and its specific expression in Hepa1-6 hepatoma cells. METHODS: The murine AFP signal peptide encoding sequence and mature IL-1ß encoding fragment were linked together through overlapping PCR, and the chimeric DNA sequence was then inserted into a liver specific expression vector pLIVE(TM); to make a recombinant pLIVE-smIL-1ß which expressed secreted murine IL-1ß of classical pathway. pLIVE-smIL-1ß, pLIVE(TM); and pLIVE-lacZ were transfected into Hepa1-6 by jetPEI respectively. Transfection of the vectors were detected by ß-gal staining using pLIVE-lacZ transfectants. Cells treated with 5 µg/mL lipopolysaccharide were used as positive control and 3 µmol/L monesin was added into culture system to block classical pathway secretion, then sandwich ELISA was employed to detect the IL-1ß levels both in supernatant and in cytoplasm of each group of transfected cells. The proliferation of Hepa1-6 hepatoma cells was determined by MTT assay and migration of Hepa1-6 cells was assessed by scratch test in vitro. RESULTS: pLIVE-smIL-1ß vector successfully expressed murine IL-1ß in Hepa1-6 cells. Expression of the recombinant vector peaked at day 3 as indicated in a ß-gal staining method. After transfection, compared with Hepa1-6/mock cells, IL-1ß expression levels both in supernatant and in cytoplasm of Hepa1-6/smIL-1ß cell were significantly increased detected by ELISA. The proliferation of Hepa1-6/smIL-1ß group was markedly promoted in vitro detected by MTT assay. CONCLUSION: The recombinant expression vector can secret IL-1ß through classical pathway which significantly promoted proliferation and migration of hepatoma cells in vitro.