The RING-domain E3 ubiquitin ligase RNF146 promotes cardiac hypertrophy by suppressing the LKB1/AMPK signaling pathway.

The RING-domain E3 ubiquitin ligase RNF146 is an enzyme that plays an important role in ubiquitin-proteasomal protein degradation and participates in various pathophysiological processes. However, its role in cardiac hypertrophy is unclear. In the present work, thoracic transverse aortic constriction (TAC) was performed in transgenic mice with RNF146 knockout mice (KO) and wild-type mice, and neonatal rat cardiomyocytes (NRCMs) were subjected to angiotensin II (Ang II) stimulation to induce cardiac hypertrophy in vitro and in vivo. RNF146 expression was significantly increased in hypertrophied murine hearts and Ang II-stimulated NRCMs. RNF146-KO mice and knockdown of RNF146 NRCMs attenuated TAC- or Ang II-stimulated cardiac hypertrophy. Conversely, enforced expression of RNF146 aggravated these changes. Mechanistically, we found that RNF146 KO or knockdown increased the activation of the AMP-activated protein kinase (AMPK) pathway. Furthermore, we found that RNF146 KO or knockdown decreased ubiquitination of Liver kinase B1 (LKB1), which promoted the activation of the AMPK pathway in a dependent manner. In conclusion, RNF146 targets LKB1 protein for ubiquitin-proteasome degradation in cardiomyocytes and subsequently promotes cardiac hypertrophy by suppressing the activation of the AMPK signaling pathway.

Comparison of the biological properties between 3D-printed and decellularized tracheal grafts.

This study sought to characterize the differences between the 3D-printed and decellularized tracheal grafts, providing the basis for the synthesis of the more reasonable and effective tissue-engineered trachea. We compared the biomechanical properties and biocompatibility of the 3D-printed tracheal graft and decellularized tracheal graft in vitro and evaluated the biocompatibility, immune rejection and inflammation of the two materials through in vivo implantation experiments. Compared with the decellularized tracheal graft, the 3D-printed tracheal graft was associated with obviously higher biomechanical properties. The results demonstrated enhanced growth of BMSCs in the decellularized tracheal graft compared to the 3D-printed one when co-culture with two tracheal graft groups. Moreover, the CCK-8 assay demonstrated significant cell proliferation on the decellularized tracheal graft. Serum IgG and IgM measured in vivo by implantation testing indicated that the 3D-Printed tracheal graft exhibited the most significant inflammatory response. HE staining indicated that the inflammatory response in the 3D-printed tracheal graft consisted mainly of eosinophils, while little inflammatory cell infiltrates were observed in the decellularized tracheal graft. CD68 immunohistochemical analysis indicated that the infiltration of macrophages was not significant in both tracheal grafts. Our findings suggest that the biomechanical properties of the 3D-printed tracheal grafts are better than the decellularized tracheal grafts. Nonetheless, the decellularized tracheal graft exhibited better biocompatibility than the 3D-printed tracheal graft.

[Inhibition of GMA-induced ubiquitination process in 16HBE malignantly transformed cells on protein CD44 and MMP14].

OBJECTIVE: To observe the effect of the ubiquitination process on the expression of CD44 antigen(CD44) and matrix metalloproteinase-14(MMP14) in human bronchial epithelial(16HBE) malignantly transformed cells induced by glycidyl methacrylate(GMA). METHODS: Successfully resuscitated 16HBE cells were cultured using a final concentration of 8 µg/mL GMA as the treatment group and 1 µg/mL dimethyl sulfoxide as the solvent control group, each time stained for 72 h, and then stained again after an interval of 24 h. After repeating the staining three times, the cells were cultured in passages respectively. The 40th generation(P40) GMA-treated group and the same-generation solvent control group were subjected to soft agar colony formation assay and concanavalin A(ConA) agglutination test to confirm that the 40th generation of GMA-induced malignant transformed 16HBE cells possessed malignant transformed cell characteristics.5, 10, 20, 40, 60 µmol/L anacardic acid were used to inhibit the ubiquitination process of GMA-induced malignant transformed 16HBE cells. The protein expression of CD44 and MMP14 were detected by western blotting, while the transcript levels of CD44, MMP14, and TFAP2A were assessed by real-time fluorescence quantitative PCR(qPCR). RESULTS: (1) In the soft agar colony formation assay, the number of clones formed by the cells in the solvent control group was 22, and the number of clones created by the malignantly transformed cells in the GMA-treated group was 208. In the ConA agglutination test, the cells in the solvent control group were uniformly dispersed in ConA solution, and no obvious agglutination occurred for 30 min, whereas the cells in the GMA-treated group were agglutinated in the 5th min, and the agglutinated cells were larger and more rapidly agglutinated. The agglomerates were more significant and faster, and the sensitivity of agglutination was increased. (2) After differential inhibition of GMA-induced ubiquitination in malignantly transformed 16HBE cells, the expression levels of CD44 and MMP14 were reduced in GMA-induced malignantly transformed 16HBE cells compared with the control group(P<0.05). The transcript levels of MMP14 and CD44 decreased with increasing inhibitor concentration(P<0.05), and the transcript levels of the upstream transcription factor TFAP2A were also simultaneously reduced(P<0.05). CONCLUSION: Inhibition of the cellular ubiquitination process mediates the down-regulation of protein expression and transcriptional expression of CD44 and MMP14 in GMA-induced malignantly transformed 16HBE cells.

[Role of LncRNA CASC11 in the malignant transformation of 16HBE cells induced by glycidyl methacrylate].

OBJECTIVE: To investigate the effect of trending up-regulation LncRNA CASC11 which is differentially expressed during the malignant transformation of human bronchial epithelial cells(16 HBE) induced by glycidyl methacrylate(GMA). METHODS: After 16 HBE cells were repeatedly exposed to low concentration GMA(8 µg/mL), the 10 th, 20 th and 30 th passage cells of the GMA group and the DMSO solvent control group were collected. High throughput LncRNA microarray were used to screen the differentially expressed LncRNAs at different stages. Short Time-series Expression Miner and bioinformatics tool RPISeq were used to screen and predict the potential targets of specific LncRNA. Real-time fluorescent quantitative PCR(qPCR) and Western blot were used to detect the relative expression of specific LncRNA and the content of its target protein respectively. RESULTS: The specific differential expression of LncRNA CASC11 in the process of GMA-induced malignant transformation of 16 HBE cells showed a trend of up-regulation. Compared with the control cells in the same period, the P10, P20 and P30 generation cells were down-regulated by 1.64 times, up-regulated by 2.01 times, and up-regulated by 2.66 times, respectively. Western blot showed that the levels of cyclin-dependent kinase 1(CDK1), cyclinB1 and cyclinB2 in P10, P20 and P30 cells after exposure were lower than those in DMSO control group during the same period, which was consistent with the microarray results. CONCLUSION: The differential expression of LncRNA CASC11 in the process of GMA-induced 16 HBE cell malignant transformation was up-regulated trendingly. It is speculated that it may block or delay cell cycle progression by inhibiting CDK1.

Downregulated CHI3L1 alleviates skeletal muscle stem cell injury in a mouse model of sepsis.

Sepsis is an acute systemic inflammatory response of the body to microbial infection and a life-threatening condition associated with multiple organ failure. Recent data suggest that sepsis survivors present with long-term myopathy due to the dysfunction of skeletal muscle stem cells and satellite cells. Accumulating studies have implicated chitinase-3-like-1 protein (CHI3L1) in a variety of infectious diseases, specifically sepsis. Therefore, the aim of the present study is to elucidate the potential mechanism by which CHI3L1 is involved in the injury of skeletal muscle stem cells in mouse models of sepsis. An in vitro cell model was developed by lipopolysaccharide (LPS) and in vivo mouse model of sepsis was induced by CRP-like protein (CLP). To elucidate the biological significance behind the silencing of CHI3L1, modeled skeletal muscle stem cells and mice were treated with siRNA against CHI3L1 or overexpressed CHI3L1. Highly expressed CHI3L1 was found in skeletal muscle tissues of mice with sepsis. Besides, siRNA-mediated silencing of CHI3L1 was revealed to increase Bcl-2 expression along with cell proliferation, while diminishing Bax expression, cell apopstosis as well as serum levels of TNF-α, IL-1ß, INF-γ, IL-10, and IL-6. Taken conjointly, this present study provided evidence suggesting that downregulation of CHI3L1 has the potential to prevent the injury of skeletal muscle stem cells in mice with sepsis. Collectively, CHI3L1 may serve as a valuable therapeutic strategy in alleviating sepsis.

NEDD4 protects vascular endothelial cells against Angiotensin II-induced cell death via enhancement of XPO1-mediated nuclear export.

NEDD4 is an E3 ubiquitin ligase containing the HECT domain, which regulates various cellular processes, but its role in vascular endothelial cells is unknown. In the present study, we found that NEDD4 bound directly to XPO1 by co-immunoprecipitation screening. In HUVECs (human umbilical vein endothelial cells), overexpression of NEDD4 reduced Ang II-induced ROS level and cell apoptosis. Ang II stimulation led to nuclear accumulation of cargoes, while overexpression of NEDD4 enhanced the XPO1-dependent nuclear export of its cargoes. KPT185, an inhibitor of XPO1, can abolished the protective effect of NEDD4 under Ang II treatment. In addition, NEDD4 could promote the interaction between XPO1 and RanBP3 via K63-linked ubiquitination of XPO1. These results suggested that NEDD4 played a protective role in vascular endothelial cell injury through regulating XPO1-mediated nuclear export.

[Role of LINC00310 in the glycidyl methacrylate-induced malignant transformation of human bronchial epithelial cell].

OBJECTIVE: To investigate the expression and biological significance of LINC00310 in the malignant transformation of human bronchial epithelial cells(16 HBE) induced by glycidyl methacrylate(GMA). METHODS: The 16 HBE cells recovered successfully used 1 µg/mL dimethyl sulfoxide(DMSO) as the solvent control group, and the final concentration was 8 µg/mL GMA as the treatment group, and were subcultured after repeated exposure 3 times for 72 hours each time. The 10 th, 20 th and 30 th generation cells of the GMA treatment group and corresponding DMSO control group were collected. The LncRNA microarrays was used to analyze the expression changes of LINC00310 in different periods, and the target gene and function prediction was performed by NCBI and cBioPortal bioinformatics database, and real-time quantification PCR(qPCR) was used to detect the relative expression levels of LINC00310 and predicted target genes. RESULTS: The result of the microarray showed that LINC00310 in the GMA-treated group was down-regulated by 2. 02-fold, up-regulated by 6. 17-fold, and up-regulated by 2. 03-fold in the pre-transformation, mid-term, and late, respectively. The result of qPCR confirmed that the expression of LINC00310 relative expression level of 10 th, 20 th and 30 th generation cells was consistent with the microarray result, which were down-regulated by 2. 76-fold, up-regulated by 2. 68-fold, and up-regulated by 3. 09-fold. Consistently, the relative expression of the target gene C-Myc was statistically significant in 20 th and 30 th generation cells. CONCLUSION: LINC00310 induced low expression in the early stage of malignant transformation of 16 HBE cells induced by GMA, and was highly expressed in the middle and late stages. It indicated that LINC00310 may play a cancer-promoting role in the process of cell malignant transformation through C-Myc.

HMGB1 promotes HLF-1 proliferation and ECM production through activating HIF1-α-regulated aerobic glycolysis.

Aerobic glycolysis is a crucial event in fibroblast differentiation, and extracellular matrix (ECM) production in the progression of pulmonary fibrosis (PF). Abnormal high mobility group protein B1 (HMGB1) activation is involved in the pathogenesis of PF. However, whether aerobic glycolysis contributes to HMGB1-induced fibroblast proliferation and ECM production in PF has not yet been determined. In this study, we investigated the effects of HMGB1 on human embryonic lung fibroblast (HLF-1) proliferation, ECM production, and aerobic glycolysis. The lactate dehydrogenase inhibitor oxamic acid (OA), and PFKFB3 inhibitor 3PO were used to block certain crucial steps of aerobic glycolysis. As a result, we observed an increase of HMGB1 in bronchoalveolar lavage fluid (BALF) in bleomycin (BLM)-treated rats as compared to non-treated rats (control group). A concentration-dependent increase of HLF-1 proliferation and expression of α-SMA and α-collagen I were observed in the HMGB1 group, as well as increases of LDHA activation, glucose uptake levels, glycolytic rate, lactate level, and ATP production. OA and 3PO, or suppression of HIF1-α, blocked the effects of HMGB1. In summary, HMGB1 promotes fibroblast proliferation and ECM production though upregulating expression of HIF1-α to induce an increase of aerobic glycolysis.

[Anti-oncogene of opioid binding protein/cell adhesion molecule-like methylation status in malignant transformation of human bronchial epithelial cells induced by glycidyl methacrylate].

OBJECTIVE: To analyze the opioid binding protein/cell adhesion molecule-like (OPCML) methylation status at different stages of malignant transformation of human bronchial epithelial (16HBE) cells induced by Glycidyl Methacrylate (GMA) and to explore the effect of OPCML methylation in the process of malignant transformation. METHODS: Cells were harvested at different stages (the 10th generation, the 20th generation and the 30th generation). To verify the Methylation chip result of OPCML methylation status in the process of malignant transformation, we detected it by methylation-specific-PCR (MSP); Real-time fluorescence Quantitative PCR (qPCR) were applied to measure the gene expression levels of OPCML at different transformed stage, and compared with the control groups (treated with DMSO). RESULTS: Based on the result of methylation chip, the gene of OPCML methylation occurred in all stages, which was consistent to the result of MSP; qPCR showed that the levels of gene expression decreased in the 20th generation and 30th generation. CONCLUSION: Methylation status of OPCML gene promoter could be considered as a stable and specific biomarker in the transformation process.

[Determination of 2, 4-dichlorophenoxyacetic acid in air of workplace by high-performance liquid chromatography].

OBJECTIVE: To develop a method for determination of 2, 4-dichlorophenoxyacetic acid (2, 4-D) in the air of workplace by high-performance liquid chromatography. METHODS: 2, 4-D was collected by ultrafine glass filters, desorbed by methanol, separated by a C18 column, and detected by a UV detector. Identification and quantification of 2, 4-D were performed by retention time and peak areas, respectively. RESULTS: The linear range of the test was 2∼200 µg/ml; the elution efficiency was 94.6%- 95.9%; the limit of detection (S/N = 3) was 0.034 µg/ml (injection volume of 20 µl eluant); the lower limit of quantification (S/N = 10) was 0.11 µg/ml; the minimum detectable concentration was 0.011 mg/m(3); the minimum quantifiable concentration was 0.037 mg/m(3) (with sampled air volume of 45 L). CONCLUSION: This method is convenient and simple in sample collection and preparation, and satisfies all methodological requirements. Therefore, this method is useful for the determination of 2, 4-D in the air of workplace.

[Method for determining brodifacoum in workplace air by high-performance liquid chromatography].

OBJECTIVE: To establish a method for determining brodifacoum in workplace air by high-performance liquid chromatography (HPLC). METHODS: Brodifacoum in workplace air was collected with a polytetrafluoroethylene filter and desorbed by mixed solution of methanol and dichloromethane (20:80, V:V), and was then separated using an ODS column and determined by an ultraviolet detector; retention time was used for identification, and peak area was used for quantification. RESULTS: The concentration of brodifacoum showed a linear relationship with peak area within 0.2∼10.0 µg/ml; the elution efficiency was 91.6%∼95.1%; the detection limit was 0.08 µg/ml (injection volume: 20 µl eluate); the minimum detectable concentration was 0.000 67 mg/m(3) (calculated by 240 L air sample). CONCLUSION: This HPLC method is convenient and simple for air collection and sample preparation and meets the methodological requirements. Therefore, this method can be used for the determination of brodifacoum in workplace air.

TMT-based quantitative proteomic analysis reveals the underlying mechanisms of glycidyl methacrylate-induced 16HBE cell malignant transformation.

Glycidyl methacrylate (GMA) has been widely used as tackifying/crosslinking copolymer monomer in the industrial section. Occupational and environmental exposure to GMA is inevitable. GMA is classified as a Group 2 A carcinogen. However, it still lacks a sufficient understanding of its carcinogenicity at the protein level. The major pathways and players during the malignant transformation process remain unknown. In this study, we first established and characterized a malignant transformation model using human bronchial epithelial (16HBE) cells exposed to 8 µg/mL GMA. Then the proteomics approach, western-blot analysis as well as quantitative PCR (qPCR) analysis were employed to investigate its underlying mechanisms of carcinogenicity. Our results showed that the 16HBE cells exposed to GMA and passaged to the 40th generation had undergone a malignant transformation. Proteomic analysis revealed that 123 proteins were significantly up-regulated while 160 proteins were down-regulated during the process of malignant transformation. Importantly, further pathway analysis identified the extracellular matrix-receptor (ECM-receptor) interaction pathway to be one of the major players mediating the process and most of the differentially expressed proteins (DEPs) were up-regulated, including two vital proteins, CD44 and MMP14, as well as members from integrin family. These results provide direct proteomic evidence that DEPs related to the ECM-receptor interaction pathway play an active role in reinforcing the carcinogenicity of GMA. The findings of this study might deepen our understanding of the underlying mechanisms of GMA carcinogenicity and thus facilitate the risk assessment of GMA.

[Methylation status of P16 gene during malignant transformation of human bronchial epithelial cells induced by glycidyl methacrylate].

OBJECTIVE: To analyze the methylation status of P16 gene at the different stages of malignant transformation of human bronchial epithelial cells (16HBE) induced by glycidyl methacrylate (GMA) and to explore the DNA methylation mechanisms. METHODS: The cells exposed to GMA were harvested at the end of exposure (early stage), the 10th generation (protophase) and the 30th generation (anaphase), respectively. The methylation status of P16 promotor was detected by Methylation-specific PCR (MSP). The transformed 16HBE cells were compared with the normal 16HBE cells and the cells exposed to DMSO for methylation status. RESULTS: At the early stage and protophase stage, the non-methylation status in P16 gene promotor of the normal 16HBE cells and the cells exposed to DMSO appeared, the methylation status in P16 gene promotor of the 16HBE cells exposed to GMA was detected to some extension. At the anaphase stage, the methylation status in P16 gene promotor of the 16HBE cells exposed to GMA or DMSO was detected to some extension. CONCLUSION: Methylation status of P16 gene promoter was specific at the early stage and protophase stage of malignant transforming in 16HBE cells induced by GMA, which can serve as an early sensitive biological indicator for malignant transforming in 16HBE cells induced by GMA.

[Determination of brodifacoum in rat plasma by HPLC].

OBJECTIVE: A determination method of brodifacoum in rat plasma with bromadiolone as an internal standard was developed. METHODS: A volume of 10 microl internal standard (bromadiolone) was added into rat plasma, and then extracted by 0.5 ml of acetonitrile by shaking for 2 min. The residue was dissolved with 200 microl of mobile phase after centrifugation for 10 min, and evaporation to dryness by Nitrogen blowing. A C18 column and PDA detector were used for separating and detecting. The wavelength was 254 nm, the flow rate was 1.0 ml/min, and the injection volume was 20 microl. RESULTS: The liner range was 1.0-20 microg/ml, and the correlation coefficient was 0.9992. The detection limit was 0.3 microg/ml in plasma (S/N=3). The intra-assay and inter-assay coefficients of variation were 1.89%-2.45% and 2.51%-3.61% respectively. The recoveries in plasma at levels of low, middle and high concentrations were (80.8 +/- 3.1)%, (81.8 +/- 2.7)% and (87.9 +/- 3.6)% (n=6), respectively. The accuracies were 84.1%-91.5% and 86.7%-93.2%, respectively. CONCLUSION: This method is simple, fast and accurate for the determination of brodifacoum in rat plasma.

A Retrospective Study of Effectiveness of Thoracoscopic Lobectomy and Segmentectomy in Patients with Early-Stage Non-Small-Cell Lung Cancer.

Background: Surgical treatment is the first choice for non-small-cell lung cancer. To date, there are only few studies on the changes in laboratory indexes in two types of surgery, namely, thoracoscopic lobectomy and segmental pneumonectomy. Aim: To investigate the clinical impact of thoracoscopic lobectomy and segmentectomy in patients with early-stage non-small-cell lung cancer. Methods: We retrospectively reviewed the medical records of 94 patients with early-stage NSCLC in our hospital from October 2017 to October 2019. The patients were divided into two groups. The patients in control and observation groups received thoracoscopic lobectomy and thoracoscopic segmentectomy, respectively. The perioperative indicators, complications, lung function, T cell subsets, tumor markers, follow-up of tumor recurrence rate, and survival rate were compared between two groups. Results: The operation time of the observation group was longer, and the chest drainage volume was less at 24-48 h after the operation, and the chest tube indwelling time and postoperative hospital stay were shorter than those of the control group. No significant differences in complication probability were observed between two groups. The levels of FEV1, FVC, and MVV in the two groups were lower than those before the operation at 3 days after surgery, but the FEV1, FVC, and MVV levels in the observation group were higher than those in the control group. The CD3+, CD4+, and CD4+/CD8+ levels in the two groups were lower than those before the operation at 24 h and 72 h after the operation, but CD3+, CD4+, and CD4+/CD8+ levels in the observation group were higher than those of the control group. Conclusion: Thoracoscopic lobectomy and segmental resection have similar clinical effects in the treatment of early-stage NSCLC patients, but segmental resection can preserve healthy lung tissue as much as possible, with less trauma, protect lung function, and promote postoperative recovery.

[Chromosome damage of human bronchial epithelial cells induced by glycidyl methacrylate].

OBJECTIVE: To observed the chromosome damage of human bronchial epithelial cells induced by glycidyl methacrylate (GMA). METHODS: Chromosome aberration analysis of human bronchial epithelial cells treated with GMA at different dosages (4, 8, 12, 16 and 20 microg/ml), times (1, 2 and 3 times), and phases (10th, 30th genetation) was detected. RESULTS: In the dosages range from 4 to 20 microg/ml, the aberration rates (3%, 6%, 7%, 11% and 14%) were demonstrated increasingly with the increase of exposure doses, and dose-effect relationship was found. Significant differences were observed when treated with GMA three times (6%, 7% and 10%). Structure aberrations were found in the transformed 10th-generation cells, while number aberrations were mainly manifested in the transformed 30th-generation cells. CONCLUSION: The chromosome aberration can be induced by GMA in the human bronchial epithelial cells, from the structure aberration at the beginning to the lack of normal nuclear style.

[Effects of chitinase-3-like protein 1 on the expression of inflammatory damage-related molecules in mouse skeletal muscle satellite cells induced by lipopolysaccharide].

OBJECTIVE: To explore the potential mechanism of chitinase-3-like protein 1 (CHI3L1) involved in skeletal muscle stem cell injury induced by sepsis. METHODS: Six different concentrations of lipopolysaccharide (LPS) were used to stimulate mouse skeletal muscle satellite cells cultured in vitro. Enzyme linked immunosorbent assay (ELISA) and cell counting kit-8 (CCK-8) were used to determine the optimal concentration. The overexpression and interference vectors of CHI3L1 were constructed to transfect skeletal muscle satellite cells, and the transfection efficiency was verified by polymerase chain reaction (PCR) and Western blotting. The cells were randomly divided into blank control group (cells without any intervention), model group (LPS-stimulated untransfected cells), overexpressing CHI3L1 group (LPS-stimulated cells transfected with CHI3L1 plasmid), overexpressing CHI3L1 control group [LPS-stimulated cells transfected with negative control (NC) plasmid], CHI3L1 interference group [LPS-stimulated cells transfected with CHI3L1 small interfering RNA (siRNA)], CHI3L1 interference control group (LPS-stimulated cells transfected with CHI3L1-siRNA NC). The levels of extracellular caspase-1 and interleukin-1ß (IL-1ß) were detected by ELISA. The protein expressions of intracellular IL-1ß, signal transducters and activator of transcription 3 (STAT3), protein kinase B (Akt) and phosphorylated Akt (p-Akt) were detected by Western blotting. RESULTS: According to the results of CCK-8 and ELISA, the best concentration of 5 mg/L LPS was selected for the subsequent experiment. The transfection was validated by PCR and Western blotting. Compared with the blank control group, the levels of extracellular IL-1ß, caspase-1 and the protein expressions of intracellular Akt, p-Akt, and IL-1ß were significantly increased in the model group [IL-1ß (ng/L): 11.22±0.55 vs. 8.63±0.63, caspase-1 (pmol/L): 9.47±0.22 vs. 8.65±0.15, Akt/GAPDH: 1.36±0.12 vs. 1.06±0.15, p-Akt/GAPDH: 0.78±0.07 vs. 0.09±0.01, IL-1ß/GAPDH: 1.38±0.12 vs. 0.18±0.03, all P < 0.05]. Compared with the model group and the overexpressing CHI3L1 control group, the levels of extracellular IL-1ß, caspase-1 and the protein expressions of intracellular p-Akt and IL-1ß were significantly increased in the overexpressing CHI3L1 group [IL-1ß (ng/L): 14.93±0.97 vs. 11.22±0.55, 9.38±0.40, caspase-1 (pmol/L): 10.35±0.03 vs. 9.47±0.22, 8.46±0.24, p-Akt/GAPDH: 1.21±0.04 vs. 0.78±0.07, 0.63±0.04, IL-1ß/GAPDH: 1.87±0.08 vs. 1.38±0.12, 1.51±0.17, all P < 0.05]. Compared with the model group and the CHI3L1 interference control group, the levels of extracellular IL-1ß, caspase-1 and the protein expressions of intracellular p-Akt and IL-1ß were significantly decreased in the CHI3L1 interference group [IL-1ß (ng/L): 8.98±0.73 vs. 11.22±0.55, 10.44±0.65, caspase-1 (pmol/L): 7.61±0.63 vs. 9.47±0.22, 8.37±0.38, p-Akt/GAPDH: 0.50±0.04 vs. 0.78±0.07, 0.94±0.06, IL-1ß/GAPDH: 0.77±0.02 vs. 1.38±0.12, 1.13±0.07, all P < 0.05]. CONCLUSIONS: CHI3L1 may mediate the damage of skeletal muscle stem cells in sepsis by increasing the expression of caspase-1 and IL-1ß. CHI3L1 may be involved in the regulation of Akt signaling pathway in skeletal muscle stem cells, but has no significant effect on STAT3 signaling pathway.

[Study on malignant transformation of human bronchial epithelial cells induced by glycidyl methacrylate].

OBJECTIVE: To study the malignant transformation of human bronchial epithelial cells induced by glycidyl methacrylate (GMA). METHODS: 16HBE cells were treated multiple times with GMA at concentrations of 1, 2, 4 and 8 microg/ml. Cellular biological characteristics of malignant transformation were identified by the tests of conA, colony forming frequency on soft agar, scanning electron microscope and tumorigenesis in nude mice. Test of immunocytochemical detection was also applied to confirm the derivation of cell and tumor. Groups of solvent control (DMSO) and positive control (MCA) were also performed at the same time. RESULTS: Transformed foci could be observed after the cells were treated by GMA at concentrations from 1 to 8 microg/ml. The number of transformation foci increased with the concentration of GMA. Transforming rate in 8 microg/ml group (8.48 x 10(-6)) was significantly higher (P < 0.01) than that of solvent control group (4.5 x 10(-7)). The transformed cells lost contact inhibition and exhibited a crossover growth in culture dish. They also could grow in semi-solid agar and showed dose-reaction relations with the concentration of GMA. The colony forming frequency in 2, 4 and 8 microg/ml group was 1.20 per thousand, 2.35 per thousand and 5.70 per thousand respectively, which were higher than that of solvent control group (P < 0.01). The transformed cells could be agglutinated by low concentration of conA. Microvilli on the surface of transformed cells increased and became strong and long under scanning electron microscope. The transformed cells could form subcutaneous tumor in nude mice which was diagnosed as squamous cell carcinoma in morphology. Expression of cytokeratin (CK) was detected in both 16HBE cells and tumor formed in nude mice. CONCLUSION: GMA could induce the malignant transformation of 16HBE cells. This research system might provide a potential tool and lay a foundation for the study of the molecular mechanism of carcinogenesis induced by GMA.

PTEN Expression, Not Mutation Status in TSC1, TSC2, or mTOR, Correlates with the Outcome on Everolimus in Patients with Renal Cell Carcinoma Treated on the Randomized RECORD-3 Trial.

PURPOSE: Genomic alterations in key components of PI3K/mTOR pathway have been proposed as candidate predictive markers for rapalog therapy in renal cell carcinoma (RCC). We tested this hypothesis in patients from a randomized phase II trial of everolimus versus sunitinib. PATIENTS AND METHODS: Archival specimens collected at baseline were analyzed with targeted next-generation sequencing (NGS). Focus of interest were alterations in key PI3K pathway components. PTEN expression was assessed by IHC. Association between molecular findings and treatment outcomes was investigated; same associations were tested for 2 everolimus-treated trial cohorts in gastric and hepatocellular carcinoma (HCC). RESULTS: Among 184 everolimus-treated patients with RCC with NGS data, mutation rates in genes of interest were 6% (TSC1), 4.4% (TSC2), and 8.2% (mTOR); 44% harbored alterations in ≥1 PI3K pathway component. For subjects with presence versus absence of mutations in TSC1, TSC2, or mTOR progression-free survival (PFS) neither differed on univariate analysis (HR, 1.0; P = 0.895) nor on multivariate testing stratified by MSKCC risk group and other established prognostic factors (HR, 1.1; P = 0.806). Everolimus-treated patients with retained (n = 50) versus lost (n = 50) PTEN IHC expression had median PFS of 5.3 months versus 10.5 months (HR, 2.5; P < 0.001). Such differences were not seen with sunitinib (10.9 months vs. 10.3 months; HR, 0.8; P = 0.475). Molecular findings did not correlate with outcomes in gastric and HCC cohorts. CONCLUSIONS: Association between mutation status for TSC1/TSC2/mTOR and therapeutic outcome on everolimus was not confirmed. Clinically meaningful differences in PFS were seen based on PTEN expression by IHC, lost in >50% of patients.

Hybrid Metal-Dielectric Nano-Aperture Antenna for Surface Enhanced Fluorescence.

A hybrid metal-dielectric nano-aperture antenna is proposed for surface-enhanced fluorescence applications. The nano-apertures that formed in the composite thin film consist of silicon and gold layers. These were numerically investigated in detail. The hybrid nano-aperture shows a more uniform field distribution within the apertures and a higher antenna quantum yield than pure gold nano-apertures. The spectral features of the hybrid nano-apertures are independent of the aperture size. This shows a high enhancement effect in the near-infrared region. The nano-apertures with a dielectric gap were then demonstrated theoretically for larger enhancement effects. The hybrid nano-aperture is fully adaptable to large-scale availability and reproducible fabrication. The hybrid antenna will improve the effectiveness of surface-enhanced fluorescence for applications, including sensitive biosensing and fluorescence analysis.