LncRNA EWSAT1 Regulates the Tumorigenesis of NSCLC as a ceRNA by Modulating miR-330-5p/ITGA5 Axis.

The aim of the study is to investigate how lncRNA EWSAT1 regulates the tumorigenesis of non-small cell lung cancer (NSCLC) as a ceRNA by modulating miR-330-5p/ITGA5 axis. qRT-PCR was conducted to evaluate the expression of EWSAT1 in NSCLC tissue. Then, A549 cells were selected and divided into Blank shScramble, shEWSAT1, miR-330-5p inhibitor, shEWSAT1 + miR-330-5p inhibitor, and siITGA5 and miR-330-5p inhibitor + siITGA5 groups. Besides, a series of in-vitro experiments were carried out to determine the changes in cell proliferation, apoptosis, invasion, and migration in each group. In addition, xenograft models were also constructed on nude mice to detect the tumor volume and weight, and the expression of Ki67 and apoptosis in xenograft tumor were evaluated. In NSCLC tissue and cell, EWSAT1 was upregulated significantly, demonstrating a correlation with tumor diameter, differentiation, lymph node metastasis, and TNM stage. Dual luciferase reporter gene assay confirmed targeting relationships among miR-330-5p, EWSAT1, and ITGA5. In comparison with the Blank group, the number of cell clones in the shEWSAT1 group and siITGA5 decreased, with declined invasion and migration but increased apoptotic rate. Meanwhile, ITGA5, MMP-2, and MMP-9 were downregulated with upregulated cleaved caspase-3. However, the changes above were totally reversed in the miR-330-5p inhibitor group, and miR-330-5p inhibitor transfection abolished the effect of shEWSAT1. In addition, subcutaneous xenotransplantation showed that the tumor growth in shEWSAT1 group retarded significantly, with downregulation of Ki67 and increase apoptotic rate. Silencing EWSAT1 could inhibit the expression of ITGA5 via upregulating miR-330-5p, thus, resulting in the inhibition of NSCLC cell growth.

[A Thermotolerant and Halotolerant Sulfate-reducing Bacterium in Produced Water from an Offshore High-temperature Oilfield in Bohai Bay, China: Isolation, Phenotypic Characterization, and Inhibition].

The growth and activity of sulfate-reducing prokaryotes (SRP) in oilfield environments could produce large amounts of H2S, leading to multifaceted problems, including oilfield souring and microbially-influenced corrosion, yet knowledge about the diversity and physiology of SRP therein was quite limited. To further understand the phenotypic characteristics of SRP residing in an offshore high-temperature oilfield at Bohai Bay, China, and to explore the potential methods for control of SRP-mediated problems, we isolated, using Hungate techniques, a thermotolerant, halotolerant SRP strain, designated BQ1, from the produced water of a high-temperature. We also presented the phenotypic features of BQ1, and investigated the efficacy of five biocides, or metabolic inhibitors, in suppressing the sulfidogenic activity of BQ1. Cells of BQ1 were motile, short rod-shaped, 1.2-2.5 µm in length and 0.5-0.8 µm in width. Although BQ1 shared 99% 16S rRNA gene sequence similarity with Desulfovibrio vulgaris Hildenborough, distinct phenotypic traits between them were observed. Isolated BQ1 could grow at 14-70℃(optimum at 30℃) and pH 6.0-9.0 (optimum pH 7.0), and in the presence of 0%-10% NaCl. Isolated BQ1 utilized a wide range of carbon substrates, including sodium formate, sodium lactate, and acetate. Sulfate, sulfite, thiosulfate, and sulfur were utilized as electron acceptors, but not nitrate or nitrite. Sodium hypochlorite (600 mg·L-1), Benzyltrimethylammonium chloride (300 mg·L-1), or nitrate (800 mg·L-1) failed to inhibit H2S production by BQ1. By contrast, glutaraldehyde (50 mg·L-1), bronopol (30 mg·L-1), chlorine dioxide (50 mg·L-1), and nitrite (70 mg·L-1) inhibited H2S production by BQ1 for at least 30 d, indicating that these compounds may be suitable for the mitigation of microbial souring in this specific, high-temperature, offshore oilfield at Bohai Bay, China.

[Surgical treatment of primary tracheal tumors in 63 cases].

OBJECTIVE: To summarize the experience in diagnosis and treatment of primary tracheal tumors, and to improve the life quality of patients. METHODS: Sixty-three patients with primary tracheal tumors treated in the First Affiliated Hospital of China Medical University during the past 40 years were included in this study, among them, there were 42 cases of malignant tumors and 21 cases of benign tumors. The 61 patients underwent surgery including tracheal sleeve resection (22), carinal resection and reconstruction (6), semi-carinal resection and reconstruction (6), tracheal resection for tracheal tumors (17); tracheostomy (4), tracheal resection, partial resection of the thyroid (goiter) and esophagomyotomy (1), tracheal tumor resection and vertical hemilaryngectomy with reconstruction of laryngeal ventricle and trachea by sternocleidomastoid flap (2), cervical trachea and laryngeal resection (1), and carinal scrape (2). RESULTS: Fifty-five patients had an uneventful recovery. Eight patients suffered from postoperative complications, among them 3 patients died postoperatively. CONCLUSIONS: Primary tracheal tumors often present atypical symptoms, are easily misdiagnosed and with poor prognosis. The main aim of treatment remains to remove the airway obstruction.

[Prokaryotic expression and bioactivity of human platelet-derived growth factor B chain mature peptide].

OBJECTIVE: To express human platelet-derived growth factor (hPDGF) B chain mature peptide gene in a prokaryotic expression system and detect the bioactivity of the expressed protein. METHODS: hPDGF B chain mature peptide gene was amplified and expressed in E. coli, and the recombinant protein, rhPDGF-BB, was purified and renatured in GSSG/GSS system. The bioactivity of rhPDGF-BB in vitro was evaluated with SD rat osteoblasts. RESULTS: The full-length PDGF-B mature peptide gene was obtained and verified, and successfully expressed in E. coli. Bioactivity detection results showed that the expressed rhPDGF-BB obviously promoted the proliferation and DNA replication of SD rat osteoblasts in vitro (P<0.01). CONCLUSION: he PDGF-B chain mature peptide cDNA has been successfully cloned and the PDGF-B precursor highly expressed in E. coli, and renatured rhPDGF-BB displays high bioactivity as shown by MTT assay and flow cytometry. This success provides the basis for production of functional PDGF-BB and facilitates further studies of its role in fracture healing and trauma reconstruction.

[Effects of soluble matrix of nacre on bone morphogenetic protein-2 and Cbfa1 gene expressions in rabbit marrow mesenchymal stem cells].

OBJECTIVE: To provide insights into the mechanisms and pathways of osteogensis by observing the effects of water-soluble matrix of nacre (WSM) on bone morphogenetic protein-2 (BMP-2) and Cbfa1 gene expressions in rabbit marrow mesenchymal stem cells (BMSCs). METHODS: New Zealand rabbit BMSCs cultured in vitro were stimulated with different concentrations of WSM extracted at low temperature, and the activity of AKP in the cells was evaluated with the dose-effect curve generated. BMP-2 and Cbfa1 gene expressions in rabbit BMSCs exposed to WSM were assayed with one-step RT-PCR. RESULTS: The activity of AKP in rabbit BMSCs increased after stimulation with different concentrations of WSM, and the effects were the most obvious with the WSM concentration ranging from 150 to 200 microg/ml. BMP-2 gene expression in the BMSCs increased after WSM exposure, but which did not result in obvious changes in Cbfa1 gene expression. CONCLUSION: WSM induces differentiation of rabbit BMSCs towards osteoblasts by increasing BMP-2 gene expression, in which process Cbfa1 gene does not seem to play a significant role.

Single and joint effects of pesticides and mercury on soil urease.

The influence of two pesticides including chlorimuron-ethyl and furadan and mercury (Hg) on urease activity in 4 soils (meadow burozem and phaeozem) was investigated. The soils were exposed to various concentrations of the two pesticides and Hg individually and simultaneously. Results showed that there was a close relationship between urease activity and organic matter content in soil. Chlorimuron-ethyl and furadan could both activate urease in the 4 soils. The maximum increment of urease activity by chlorimuronethyl was up to 14%-18%. There was almost an equal increase (up to 13%-21%) in the urease activity by furadan. On the contrary, Hg markedly inhibited soil urease activity. A logarithmic equation was used to describe the relationship (P<0.05) between the concentration of Hg and the activity of soil urease in the 4 tested soils. Semi-effect dose (ED50) values by the stress of Hg based on the inhibition of soil urease in the 4 soils were 88, 5.5, 24 and 20 mg/kg, respectively, according to the calculation of the corresponding equations. The interactive effect of chlorimuron-ethyl or furadan with metal Hg on soil urease was mainly synergic at the highest tested concentrations.

[Expression of SKP2 protein in lung carcinoma and its implication for prognosis].

OBJECTIVE: To investigate the characteristics of SKP2 protein expression in lung carcinoma tissues and its implication for prognosis. METHODS: The expression of SKP2 protein was detected in 89 NSCLC, 13 SCLC, 5 benign lung neoplasms, 5 normal bronchus and lung tissues by tissue chip and immunohistochemical techniques. RESULTS: The positive rate of SKP2 staining was (23.52 +/-13.57)% in NSCLC tissues and (53.85 +/- 12.26)% in SCLC tissues, significantly higher than (2.91 +/- 1.27)% in benign lung neoplasms and normal bronchus and lung tissues. Its expression was highest in SCLC tissues and lowest in benign lung tissues, with a significant difference between them (P <0.01). The expressive level of SKP2 protein in lung carcinoma tissues was closely related to cell differentiation and lymph node metastasis, but not to age, sex, smoking history, tumor site and size, and TNM staging, etc. The survival analysis revealed that the 5-year survival rate of lung carcinoma patients was much lower in SKP2 protein positive expression group than that in negative expression group (P < 0.01). CONCLUSION: The positive expression of SKP2 protein is higher in lung carcinoma than in benign or normal lung tissues, in particular, much higher in SCLC tissue. Moreover, it may be an independent factor to exert negative influence on prognosis of patients with lung carcinoma.

[Sorption-desorption behavior of Cd2 + and Pb2+ in rhizosphere and bulk soil].

Sorption-desorption behavior of Cd2+ and Pb2+ in the rhizosphere and bulk soil of wheat was studied by the batch method. The results indicated that the sorption capacity of rhizosphere soil to Cd2+ and Pb2+ was higher than that of bulk soil. Isothermal curves of Cd2+ sorption by rhizosphere and bulk soil fitted Freundlich equation well. Isothermal sorption process of Pb2+ could be well described by Langmuir equation and Freundlich equation. Two-constant equation was the best model to describe the sorption kinetics of Cd2+ and Pb2+ followed by Elovich equation, and the worst model was first-order dynamics equation. There was sorption-desorption hysteresis of Cd2+ and Pb2+. The desorptive rates of Cd2+ and Pb2+ in rhizosphere soil was lower than that of bulk soil. The relationship between desorptive quantity of Cd2+ and Pb2+ in two soils and the initial sorptive quantity coincided with quadratic equation. The desorptive rates of Cd2+ and Pb2+ in two soils increased with the higher initial concentrations of Cd2+ and Pb2+ in the treatments, and decreased with the increasing desorptive time. Two-constant equation was the optimal model to describe the desorptive kinetics of Cd2+ and Pb2+ in the two soils, and followed by Elovich equation, while the first-order dynamics equation fitted not well.

[Effects of Hg on soil enzyme activity].

With simulation test, this paper studied the effects of Hg on the activities of urease, invertase and neutral phosphotase in four soils. The results showed that Hg inhibited soil urease and invertase activities markedly, but its inhibitory effect differed with test soils. There was a significant logarithmic correlation between the concentration of HgCl2 and the activities of these two enzymes (P < 0.05). In test soils, the ED50 of urease activity was 87.99, 5.47, 24.05 and 19.88 mg x kg(-1), and that of invertase activity was 76.68, 727.49, 236.52 and 316.59 mg x kg(-1), respectively. Urease was more sensitive than invertase to Hg contamination, while organic matter had a protective effect on soil enzymes. Soil neutral phosphatase was not sensitive to Hg contamination, except that it was significantly activated by Hg in the meadow brown soil applied with plenty of organic fertilizer.

[Effect of temperature on kinetic of soil urease inhibited by Hg].

Urease kinetics inhibited by Hg in two meadow burozem, fertilized by different ways and two phaeozem soils with different organic matter content was investigated at different temperatures. The results showed that the urease activity and kinetic parameter V(max) and V(max)/K(m) were higher in soils with high organic matter content than that in soils with low organic matter among the same soil type. It indicated that organic matter had great adsorption capacity to urease. Soil urease V(max) and V(max)/K(m) in phaeozem were generally higher than that in meadow organic matter had great adsorption capacity to urease. Soil urease V(max) and V(max)/K(m) in phaeozem were generally higher than that in meadow burozem, but urease activity and K(m) were not comparable among different soil types. K(m) depended on not only the organic matter content of soils, but also fertilization ways. As incubating temperature increased, urease activity, V(max) and V(max)/K(m) value enhanced under the optical catalysis temperature. Hg behaved as uncompetitive inhibitor to soil urease in this experiment. The negative effect that Hg inhibited urease activity, K(m), V(max) and V(max)/K(m) increased with temperature increasing, indicated that the protective capacity of soil on urease decreased with the temperature increasing.

[Molecular cloning and expression of human PDGF-B chain mature peptide gene].

OBJECTIVE: To acquire sufficient PDGF-BB protein and provide the basis for the further studies of its role on the fracture healing and trauma reconstruction and its clinical applications. METHODS: Constructed the prokaryotic expression vector pQE-PDGF-B with the gene rearrangement technique, and the monomeric form of recombinant PDGF-B expressed in E. coli M15. RESULTS: PDGF-B mature peptide gene was inserted into prokaryotic expression vector pQE30, which was confirmed by PCR, enzyme digestion and sequencing identification; the expressed products of pQE-PDGF-B in E. coli showed a single protein on SDS-PAGE, and their expression level was about 15% of the total bacterial protein. The molecular weight of the purified PDGF-B protein was about 15 KDs on SDS-PAGE. CONCLUSIONS: The construction of recombinant plasmid and preparation of the monomeric protein of PDGF-B provides a solid foundation for further studying the function of PDGF-BB and producing biologically PDGF-BB protein.

[Construction and identification of yeast expression vectors containing human platelet-derived growth factor B-chain gene].

OBJECTIVE: To investigate the expression efficiency of human platelet-derived growth factor B-chain (PDGF-B) gene in yeast and assess the activity of the expressed product. METHODS: A full-length complementary DNA of human PDGF-B gene was amplified from the total RNA extracted from human vascular endothelial cells using reverse transcription (RT)-PCR and then cloned into pGEM-T vector. The PCR products with specific primers were recombined into yeast expression plasmids pMETB or pMETalphaA, followed by identification with restriction endonuclease. RESULTS: The 578 bp fragment encoding mature PDGF-BB peptide with signal peptide and the 340 bp fragment without signal peptide were identified and verified by restriction endonuclease mapping and sequencing, and two yeast expression vectors pMETB-PDGFB(1) and pMETalphaA-PDGFB(2) were reconstructed. CONCLUSION: The yeast expression vectors containing human PDGF-B gene have been successfully constructed for further studies of gene expression in yeast.