Determination of 44 major components and chemical profiling of saccharide in Chinese medicinal formula Lanqin Oral Liquid.

INTRODUCTION: Lanqin Oral Liquid (LQL) is a traditional Chinese medicine preparation (TCMP) containing five herbal medicines and has been commonly used for the treatment of pharyngitis and hand-foot-and-mouth disease in clinic. The material basis of LQL has been reported in our previous study, but the contents of the major components and the features of saccharide in LQL are still unclear. OBJECTIVES: This study aimed to establish accurate and rapid methods for the quantification of the major components and profiling of saccharide in LQL. The quantitative results combined with similarity evaluation were applied to improve the quality control of LQL. METHODOLOGY: An ultra-high performance liquid chromatography coupled with triple-quadrupole tandem mass spectrometry (UPLC-QQQ-MS) method was utilised to determine 44 major components. Cosine similarity was used to evaluate the similarities among 20 batches of LQL based on the quantitative results of 44 major components. The physicochemical properties, structure, composition, and contents of saccharide in LQL were detected by a combination of chemical and instrumental analysis. RESULTS: A total of 44 compounds, including flavonoids, iridoid glycosides, alkaloids, and nucleosides, were accurately determined. The 20 batches of LQL were remarkably similar (> 0.95). In addition, d-glucose, galactose, d-glucuronic acid, arabinose, and d-mannose were detected in saccharide of LQL. The contents of saccharide in LQL were 13.52-21.09 mg/ml. CONCLUSIONS: The established methods can be applied for the comprehensive quality control of LQL, including characterisation of saccharide and quantification of representative components. Our study will provide a robust chemical foundation for disclosing the quality markers of its therapeutic effect.

A Noble-Metal-Free Nickel(II) Polypyridyl Catalyst for Visible-Light-Driven Hydrogen Production from Water.

The complex [Ni(bpy)3](2+) (bpy=2,2'-bipyridine) is an active catalyst for visible-light-driven H2 production from water when employed with [Ir(dfppy)2 (Hdcbpy)] [dfppy=2-(3,4-difluorophenyl)pyridine, Hdcbpy=4-carboxy-2,2'-bipyridine-4'-carboxylate] as the photosensitizer and triethanolamine as the sacrificial electron donor. The highest turnover number of 520 with respect to the nickel(II) catalyst is obtained in a 8:2 acetonitrile/water solution at pH 9. The H2 -evolution system is more stable after the addition of an extra free bpy ligand, owing to faster catalyst regeneration. The photocatalytic results demonstrate that the nickel(II) polypyridyl catalyst can act as a more effective catalyst than the commonly utilized [Co(bpy)3 ](2+). This study may offer a new paradigm for constructing simple and noble-metal-free catalysts for photocatalytic hydrogen production.

Association between the receptor for advanced glycation end products gene polymorphisms and cancer risk: a systematic review and meta-analysis.

PURPOSE: Polymorphisms in the receptor for advanced glycation end products (RAGE) gene may influence the risk of cancer, but the results are inconsistent. Therefore, we performed a systematic review to identify statistical evidence of the association between the 3 polymorphisms rs2070600 G/S (82G>S), rs1800624 T/A ( -374 T>A) and rs1800625C/T (-429 C>T) and the risk of cancer. METHODS: We searched PubMed database (http://www.ncbi. nlm.nih.gov/pubmed/), EMBASE database (http://www.elsevier.com/online-tools/embase ) and China National Knowledge Infrastructure (CNKI) database (http://www.cnki.net/) until Aug 30, 2014 to identify eligible studies. RESULTS: The pooled analysis revealed positive association between RAGE rs2070600 polymorphism and cancer risk in all genetic models (homozygous: OR=1.831, 95%CI: 1.548-2.166, p<0.001, allele: OR=1.321, 95%CI: 1.164-1.499, p<0.001, heterozygous: OR=1.42, 95%CI:1.126-1.792, p=0.003, dominant: OR=1.499, 95%CI: 1.200-1.874 ; p<0.001, recessive: OR=1.376, 95%CI: 1.197-1.583, p<0.001). We failed to get an effective conclusion about the association between the rs1800624 and rs1800625 polymorphisms and cancer risk in overall comparison. But in subgroup analysis, the rs1800624 polymorphism significantly increased lung cancer susceptibility in the homozygous model (OR=1.486, 95%CI:1.147-1.924, p=0.003) and the allele model (OR=1.15, 95%CI:1.029-1.285, p=0.014), but most likely contributed to decreased susceptibility to breast cancer in the allele model (OR=0.791 95%CI: 0.648-0.965, p=0.021), the heterozygous model (OR=0.733, 95%CI:0.577-0.931, p=0.011) and the dominant model (OR=0.741, 95%CI:0.588-0.934, p=0.011). No significant association was found between RAGE rs1088625 polymorphism and cancer risk in Caucasians, but these results should be interpreted with caution. CONCLUSION: The polymorphism of rs2070600 in the RAGE gene may increase the susceptibility to several human cancers, especially to lung cancer and to Asians. The rs1800264 most likely contributes to decreased susceptibility to breast cancer but increased susceptibility to lung cancer. However, large-scale studies involving various cancer types and different populations are needed for a precise conclusion.

The defects of cholangiocyte primary cilia in patients with graft cholangiopathies.

PURPOSE: To observe the morphologic changes in intrahepatic bile ducts and the defects of cholangiocyte primary cilia in patients with graft cholangiopathies. METHODS: Four patients who were diagnosed as graft cholangiopathies and underwent retransplantation were chosen as the study group; another four patients who underwent liver transplantation during the same period and recovered normally six months after the operation were the control group. The serum levels of biochemical indicators were measured, the morphologic changes in intrahepatic bile ducts and cholangiocyte primary cilia were observed, and the ciliary marker (α-tubulin) and membrane proteins (polycystin-1, TPPV4) were detected by immunofluorescence analysis and Western blot. RESULTS: In the study group, biliary structures were vague and some bile ducts disappeared in portal areas; some epithelial cells were lost; lots of collagen was deposited and many phlogocytes infiltrated; microliths were found in some ductal lumens; partial biliary epithelial cells were necrosed; primary cilia and microvilli disappeared. In the control group, the structures of intrahepatic bile ducts and biliary epithelial cells were integrated and the primary cilia were present. CONCLUSIONS: The morphologic changes in biliary epithelial cells and the defects of cholangiocyte primary cilia have a close correlation with graft cholangiopathies in liver transplantation.

α-Mangostin suppresses human gastric adenocarcinoma cells in vitro via blockade of Stat3 signaling pathway.

AIM: To investigate the anti-tumor effects of α-mangostin, a major xanthone identified in the pericarp of mangosteen (Garcinia mangostana Linn), against human gastric adenocarcinoma cells in vitro, and the mechanisms of the effects. METHODS: Human gastric adenocarcinoma cell lines BGC-823 and SGC-7901 were treated with α-mangostin. The cell viability was measured with MTT assay, and cell apoptosis was examined using flow cytometry and TUNEL assay. The expression of the relevant proteins was detected using Western blot. RESULTS: Treatment with α-mangostin (3-10 µg/mL) inhibited the viability of both BGC-823 and SGC-7901 cells in dose- and time-manners. Furthermore, α-mangostin (7 µg/mL) time-dependently increased the apoptosis index of the cancer cells, reduced the mitochondrial membrane potential of the cancer cells, and significantly increased the release of cytochrome c and AIF into cytoplasm. Moreover, the α-mangostin treatment markedly suppressed the constitutive Stat3 protein activation, and Stat3-regulated Bcl-xL and Mcl-1 protein levels in the cancer cells. CONCLUSION: The anti-tumor effects of α-mangostin against human gastric adenocarcinoma cells in vitro can be partly attributed to blockade of Stat3 signaling pathway.

An effective strategy for assembling the sex-limited chromosome.

BACKGROUND: Most currently available reference genomes lack the sequence map of sex-limited (such as Y and W) chromosomes, which results in incomplete assemblies that hinder further research on sex chromosomes. Recent advancements in long-read sequencing and population sequencing have provided the opportunity to assemble sex-limited chromosomes without the traditional complicated experimental efforts. FINDINGS: We introduce the first computational method, Sorting long Reads of Y or other sex-limited chromosome (SRY), which achieves improved assembly results compared to flow sorting. Specifically, SRY outperforms in the heterochromatic region and demonstrates comparable performance in other regions. Furthermore, SRY enhances the capabilities of the hybrid assembly software, resulting in improved continuity and accuracy. CONCLUSIONS: Our method enables true complete genome assembly and facilitates downstream research of sex-limited chromosomes.

Nuclear-Targeted Nanostrategy Regulates Spatiotemporal Communication for Dual Antitumor Immunity.

Intercellular communication between tumor cells and immune cells regulates tumor progression including positive communication with immune activation and negative communication with immune escape. An increasing number of methods are employed to suppress the dominant negative communication in tumors such as PD-L1/PD-1. However, how to effectively improve positive communication is still a challenge. In this study, a nuclear-targeted photodynamic nanostrategy is developed to establish positive spatiotemporal communication, further activating dual antitumor immunity, namely innate and adaptative immunity. The mSiO2 -Ion@Ce6-NLS nanoparticles (NPs) are designed, whose surface is modified by ionic liquid silicon (Ion) and nuclear localization signal peptide (NLS: PKKKRKV), and their pores are loaded with the photosensitizer hydrogen chloride e6 (Ce6). Ion-modified NPs enhance intratumoral enrichment, and NLS-modified NPs exhibit nuclear-targeted characteristics to achieve nuclear-targeted photodynamic therapy (nPDT). mSiO2 -Ion@Ce6-NLS with nPDT facilitate the release of damaged double-stranded DNA from tumor cells to activate macrophages via stimulator of interferon gene signaling and induce the immunogenic cell death of tumor cells to activate dendritic cells via "eat me" signals, ultimately leading to the recruitment of CD8+ T-cells. This therapy effectively strengthens positive communication to reshape the dual antitumor immune microenvironment, further inducing long-term immune memory, and eventually inhibiting tumor growth and recurrence.

Single-Component Dual-Functional Autoboost Strategy by Dual Photodynamic and Cyclooxygenase-2 Inhibition for Lung Cancer and Spinal Metastasis.

Coloading adjuvant drugs or biomacromolecules with photosensitizers into nanoparticles to enhance the efficiency of photodynamic therapy (PDT) is a common strategy. However, it is difficult to load positively charged photosensitizers and negatively charged adjuvants into the same nanomaterial and further regulate drug release simultaneously. Herein, a single-component dual-functional prodrug strategy is reported for tumor treatment specifically activated by tumor microenvironment (TME)-generated HOCl. A representative prodrug (DHU-CBA2) is constructed using indomethacin grafted with methylene blue (MB). DHU-CBA2 exhibited high sensitivity toward HOCl and achieved simultaneous release of dual drugs in vitro and in vivo. DHU-CBA2 shows effective antitumor activity against lung cancer and spinal metastases via PDT and cyclooxygenase-2 (COX-2) inhibition. Mechanistically, PDT induces immunogenic cell death but stimulates the gene encoding COX-2. Downstream prostaglandins E2 and Indoleamine 2,3 dioxygenase 1 (IDO1) mediate immune escape in the TME, which is rescued by the simultaneous release of indomethacin. DHU-CBA2 promotes infiltration and function of CD8+ T cells, thus inducing a robust antitumor immune response. This work provides an autoboost strategy for a single-component dual-functional prodrug activated by TME-specific HOCl, thereby achieving favorable tumor treatment via the synergistic therapy of PDT and a COX-2 inhibitor.

GAEP: a comprehensive genome assembly evaluating pipeline.

With the rapid development of sequencing technologies, especially the maturity of third-generation sequencing technologies, there has been a significant increase in the number and quality of published genome assemblies. The emergence of these high-quality genomes has raised higher requirements for genome evaluation. Although numerous computational methods have been developed to evaluate assembly quality from various perspectives, the selective use of these evaluation methods can be arbitrary and inconvenient for fairly comparing the assembly quality. To address this issue, we have developed the Genome Assembly Evaluating Pipeline (GAEP), which provides a comprehensive assessment pipeline for evaluating genome quality from multiple perspectives, including continuity, completeness, and correctness. Additionally, GAEP includes new functions for detecting misassemblies and evaluating the assembly redundancy, which performs well in our testing. GAEP is publicly available at https://github.com/zy-optimistic/GAEP under the GPL3.0 License. With GAEP, users can quickly obtain accurate and reliable evaluation results, facilitating the comparison and selection of high-quality genome assemblies.

Observation of Strong Anisotropic Interlayer Excitons.

Anisotropic interlayer excitons had been theoretically predicted to exist in two-dimensional (2D) anisotropy/isotropy van der Waals heterojunctions. However, experimental results consolidating the theoretical prediction and exploring the related anisotropic optoelectronic response have not been reported so far. Herein, strong photoluminescence (PL) of anisotropic interlayer excitons is observed in a symmetric anisotropy/isotropy/anisotropy heterojunction exemplified by 3L-ReS2/1L-MoS2/3L-ReS2 using monolayer (1L) MoS2 and trilayer (3L) ReS2 as components. Sharp interlayer exciton PL peaks centered at ∼1.64, ∼1.61, and ∼1.57 eV are only observed at low temperatures of ≤120 K and become more pronounced as the temperature decreases. These interlayer excitons exhibit strong anisotropic PL intensity variations with periodicities of 180° as functions of the incident laser polarization angles. The polarization ratios of these interlayer excitons are calculated to be 1.33-1.45. Our study gives new insight into the manipulation of excitons in 2D materials and paves a new way for a rational design of novel anisotropic optoelectronic devices.

An Environmental-Inert and Highly Self-Healable Elastomer Obtained via Double-Terminal Aromatic Disulfide Design and Zwitterionic Crosslinked Network for Use as a Triboelectric Nanogenerator.

Due to the ongoing development of portable/mobile electronics, sources to power have received widespread attention. Compared to chemical batteries as power sources, triboelectric nanogenerators (TENGs) possess lots of advantages, including the ability to harvest energy via human motions, flexible structures, environment-friendliness, and long-life characteristics. Although many self-healable TENGs are reported, the achievement of a muscle-like elasticity and the ability to recover from inevitable damage under extreme conditions (such as a high/low temperature and/or humidity) remain a challenge. Herein, a "double-terminal aromatic disulfide" on a structure with zwitterions as branched chains is reported to engineer the high-efficient self-healable elastomer for application in a flexible TENG. The as-designed material exhibits a repeatable elastic recovery (at 250% elongation) and a self-healing efficiency with an ultimate tensile stress of 96% over 2 h, representing an improvement on previously reported disulfide-based elastomers. The elastomer can autonomously recover by 50% even at a subzero temperature of -30 °C within 24 h. The elastomer-based TENG, as a self-driven sensor for detecting human behavior, is demonstrated to exhibit stable outputs and self-healing in the temperature range of -30 to 60 °C, and so is expected to promote the development of self-powered electronics for next-generation human-machine communications.

Golgi Apparatus-Targeted Photodynamic Therapy for Enhancing Tumor Immunogenicity by Eliciting NLRP3 Protein-Dependent Pyroptosis.

Innate and adaptive immunity is important for initiating and maintaining immune function. The nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome serves as a checkpoint in innate and adaptive immunity, promoting the secretion of pro-inflammatory cytokines and gasdermin D-mediated pyroptosis. As a highly inflammatory form of cell death distinct from apoptosis, pyroptosis can trigger immunogenic cell death and promote systemic immune responses in solid tumors. Previous studies proposed that NLRP3 was activated by translocation to the mitochondria. However, a recent authoritative study has challenged this model and proved that the Golgi apparatus might be a prerequisite for the activation of NLRP3. In this study, we first developed a Golgi apparatus-targeted photodynamic strategy to induce the activation of NLRP3 by precisely locating organelles. We found that Golgi apparatus-targeted photodynamic therapy could significantly upregulate NLRP3 expression to promote the subsequent release of intracellular proinflammatory contents such as IL-1ß or IL-18, creating an inflammatory storm to enhance innate immunity. Moreover, this acute NLRP3 upregulation also activated its downstream classical caspase-1-dependent pyroptosis to enhance tumor immunogenicity, triggering adaptive immunity. Pyroptosis eventually led to immunogenic cell death, promoted the maturation of dendritic cells, and effectively activated antitumor immunity and long-lived immune memory. Overall, this Golgi apparatus-targeted strategy provided molecular insights into the occurrence of immunogenic pyroptosis and offered a platform to remodel the tumor microenvironment.

Development of a deep pathomics score for predicting hepatocellular carcinoma recurrence after liver transplantation.

BACKGROUND AND PURPOSE: Tumor recurrence after liver transplantation (LT) impedes the curative chance for hepatocellular carcinoma (HCC) patients. This study aimed to develop a deep pathomics score (DPS) for predicting tumor recurrence after liver transplantation using deep learning. PATIENTS AND METHODS: Two datasets of 380 HCC patients who underwent LT were enrolled. Residual convolutional neural networks were used to identify six histological structures of HCC. The individual risk score of each structure and DPS were derived by a modified DeepSurv network. Cox regression analysis and Concordance index were used to evaluate the prognostic significance. The cellular exploration of prognostic immune biomarkers was performed by quantitative and spatial proximity analysis according to three panels of 7-color immunofluorescence. RESULTS: The overall classification accuracy of HCC tissue was 97%. At the structural level, immune cells were the most significant tissue category for predicting post-LT recurrence (HR 1.907, 95% CI 1.490-2.440). The C-indices of DPS achieved 0.827 and 0.794 in the training and validation cohorts, respectively. Multivariate analysis for recurrence-free survival (RFS) showed that DPS (HR 4.795, 95% CI 3.017-7.619) was an independent risk factor. Patients in the high-risk subgroup had a shorter RFS, larger tumor diameter and a lower proportion of clear tumor borders. At the cellular level, a higher infiltration of intratumoral NK cells was negatively correlated with recurrence risk. CONCLUSIONS: This study established an effective DPS. Immune cells were the most significant histological structure related to HCC recurrence. DPS performed well in post-LT recurrence prediction and the identification of clinicopathological features.

Probing Nanoscale Exciton Funneling at Wrinkles of Twisted Bilayer MoS2 Using Tip-Enhanced Photoluminescence Microscopy.

In twisted bilayer (t2L) two-dimensional (2D) transition metal dichalcogenides, local strain at wrinkles strongly modulates the local exciton density and PL energy resulting in an exciton funneling effect. Probing such exciton behaviors especially at nanometer length scales is beyond the limit of conventional analytical tools due to the limited spatial resolution and low sensitivity. To address this challenge, herein we applied high-resolution tip-enhanced photoluminescence (TEPL) microscopy to investigate exciton funneling at a wrinkle in a t2L MoS2 sample with a small twist angle of 0.5°. Owing to a spatial resolution of <10 nm, excitonic behavior at nanoscale sized wrinkles could be visualized using TEPL imaging. Detailed investigation of nanoscale exciton funneling at the wrinkles revealed a deformation potential of -54 meV/%. The obtained results provide novel insights into the inhomogeneities of excitonic behaviors at nanoscale and would be helpful in facilitating the rational design of 2D material-based twistronic devices.

Filling the gap between topological insulator nanomaterials and triboelectric nanogenerators.

Reliable energy modules and higher-sensitivity, higher-density, lower-powered sensing systems are constantly required to develop wearable electronics and the Internet of Things technology. As an emerging technology, triboelectric nanogenerators have been potentially guiding the landscape of sustainable power units and energy-efficient sensors. However, the existing triboelectric series is primarily populated by polymers and rubbers, limiting triboelectric sensing plasticity to some extent owing to their stiff surface electronic structures. To enrich the current triboelectric group, we explore the triboelectric properties of the topological insulator nanofilm by Kelvin probe force microscopy and reveal its relatively positive electrification charging performance. Both the larger surface potential difference and the conductive surface states of the nanofilms synergistically improve the charge transfer behavior between the selected triboelectric media, endowing the topological insulator-based triboelectric nanogenerator with considerable output performance. Besides serving as a wearable power source, the ultra-compact device array demonstrates innovative system-level sensing capabilities, including precise monitoring of dynamic objects and real-time signal control at the human-machine interface. This work fills the blank between topological quantum matters and triboelectric nanogenerators and, more importantly, exploits the significant potential of topological insulator nanofilms for self-powered flexible/wearable electronics and scalable sensing technologies.

Multimodal Nanoscopic Study of Atomic Diffusion and Related Localized Optoelectronic Response of WS2/MoS2 Lateral Heterojunctions.

The atomic diffusion in transition metal dichalcogenides (TMDs) van der Waals heterojunctions (HJs) strongly modifies their optoelectronic properties in the nanoscale. However, probing such localized properties challenges the spatial resolution and the sensitivity of a variety of analytic tools. Herein, a multimodal nanoscopy (based on tip enhanced Raman spectroscopy (TERS) and photoluminescence (TEPL)) combined with the Kelvin probe force microscopy (KPFM) method was used to probe such nanoscale localized optoelectronic properties induced by atomic diffusion. Chemical vapor deposition (CVD)-grown lateral bilayer (2L) WS2/MoS2 HJs were imaged with a spatial resolution better than 40 nm via TERS and TEPL mapping by using intrinsic Raman and photoluminescence (PL) peaks. The contact potential difference (CPD), capacitance, and PL variation in a nanoscale vicinity of the HJ interface can be correlated to the local stoichiometry variation determined by TERS. The diffusion coefficients of W and Mo were obtained to be ∼0.5 × 10-12 and ∼1 × 10-12 cm2/s, respectively, by using Fick's second law. The obtained results would be useful to further understand the localized optoelectronic response of the TMDs HJs.

[Application of da Vinci system for biliary operation in fifteen patients].

OBJECTIVE: To evaluate the efficacy of da Vinci surgical system in the treatment of biliary diseases. METHODS: The clinical data of 15 patients with biliary diseases who had undergone operations with da Vinci surgical system from March 2009 to November 2009 at our hospital were retrospectively analyzed. RESULTS: The operations were successfully performed on all patients. And no case was converted into open laparotomy. The total operative duration was 256 +/- 151 min and the robot operative duration 224 +/- 94 min. No blood transfusion was needed. Postoperative recovery time of bowl movement was 30 +/- 18 hours. And the average postoperative hospital stay was 6 +/- 3 days. Two patients had postoperative complications and were cured by conservative treatment. CONCLUSION: Various laparoscopic operations for biliary diseases may be performed with the aid of three-dimensional imaging system and flexible surgical tools of the Da Vinci surgical system. And its superiority is more obvious for complicated biliary diseases.

Efficacy of Biocage in treating single-segment lumbar degenerative disease in patients with high risk of non-fusion: a prospective controlled study with at least 2 years' follow-up.

OBJECTIVE: To evaluate the clinical efficacy of an allogeneic bone cage (Biocage; Beijing Datsing Bio-Tech Co., Ltd., Beijing, China) for treatment of single-segment lumbar degenerative disease in patients with a high risk of non-fusion. METHODS: From January 2013 to December 2016, 67 patients who underwent lumbar fusion were divided into the Biocage group (n = 33) and polyether ether ketone (PEEK) group (n = 34). The patients were followed up for 24 to 48 months. The mean intervertebral height of the fusion level, fusion rate, height of the intervertebral foramen, visual analog scale score, and Oswestry disability index were compared. RESULTS: The PEEK group had a lower fusion rate than the Biocage group (88.24% vs. 90.91%), although the difference was not statistically significant. During follow-up, the height of the intervertebral space was similar between the Biocage and PEEK groups (12.88 ± 0.45 and 12.84 ± 1.01 mm, respectively). The height of the intervertebral foramen was larger in the Biocage than PEEK group (20.67 ± 1.34 vs. 20.00 ± 2.05 mm). Good clinical efficacy was achieved in both groups. CONCLUSION: The Biocage is efficient and safe for treatment of single-segment lumbar degenerative disease in patients with a high risk of non-fusion.

Identifying optimal regional solid waste management strategies through an inexact integer programming model containing infinite objectives and constraints.

The previous inexact mixed-integer linear programming (IMILP) method can only tackle problems with coefficients of the objective function and constraints being crisp intervals, while the existing inexact mixed-integer semi-infinite programming (IMISIP) method can only deal with single-objective programming problems as it merely allows the number of constraints to be infinite. This study proposes, an inexact mixed-integer bi-infinite programming (IMIBIP) method by incorporating the concept of functional intervals into the programming framework. Different from the existing methods, the IMIBIP can tackle the inexact programming problems that contain both infinite objectives and constraints. The developed method is applied to capacity planning of waste management systems under a variety of uncertainties. Four scenarios are considered for comparing the solutions of IMIBIP with those of IMILP. The results indicate that reasonable solutions can be generated by the IMIBIP method. Compared with IMILP, the system cost from IMIBIP would be relatively high since the fluctuating market factors are considered; however, the IMILP solutions are associated with a raised system reliability level and a reduced constraint violation risk level.

[Nanoparticle-mediated endostatin gene therapy targeting hepatocellular carcinoma utilizing heat-inducible promoter].

OBJECTIVE: To investigate the inhibitory effect of nanoparticle-mediated endostatin gene therapy on hepatocellular carcinoma xenografts combined with local hyperthermia utilizing heat-inducible promoter. METHODS: Heat-inducible HSP70B promoter and fusion gene of Endo/EGFP were cloned into pcDNA3.1 (+) plasmid, thus obtaining recombinant plasmid of pcDNA3.1 (+)/HSP70-Endo/EGFP using restriction endonucleases BglII/HindIII and EcoRI/SalI. The nanoparticles polylactide-grafted dextran copolymer (DEX-g-PLA) encapsulating the recombinant plasmid DNA were prepared by the method of emulsification and evaporation of organic solvent, and the surface shape of nanoparticles was observed by transmission electron microscope. Human hepatocellular cells of the lines HepG2 and ECV304 were cultured and transfected with the recombinant plasmid utilizing the nanoparticles. Following thermal induction at 37 degrees C, 39 degrees C, 41 degrees C, 43 degrees C, and 45 degrees C for 30 min, the expression of enhanced green fluorescent protein (EGFP) was detected by fluorescence microscope and flow cytometry. The concentration of endostatin protein in the supernatant was tested by ELISA, and the growth inhibition on the HepG2 and ECV304 cells was tested by MTT method. Balb/c nude mice were inoculated with HeG2 cells and then randomly divided into 2 groups to undergo intra-tumor injection of nanoparticles (heated or not heated), Lipofectamine 2000. Mice were used as controls without intra-tumor injection. Four weeks the mice were killed to observe the tumor inhibition rate. RESULTS: The nanoparticles encapsulating recombinant plasmid were of round or elliptical shape 90 approximately 120 nm in diameter. The efficiency of gene transfection mediated by nanoparticles was about 30.65%. The expression of Endo/EGFP gene in the HepG2 cells was up-regulated along with the increase of temperature, peaked at 43 degrees C (with the EGFP expression level 3.3 times as that at 37 degrees C). The concentration of endostatin protein in the supernatant of the 43 degrees C group was (177 +/- 28) microg/L, significantly higher than that of the 37 degrees C group [(41 +/- 10) microg/L]. MTT results indicated that endostatin inhibited the growth of ECV304 cells with a inhibition rate of 96.3% at the time point of 72 h in the 43 degrees C group, however, it did not show influence on HepG2 cells no matter what was the temperature The tumor inhibition rate in the mice of endostatin with thermal induction group was 58.5%, significantly higher than that of the 37 degrees C group (34.9%, P < 0.05). CONCLUSION: Low temperature thermal induction enhances the expression and secretion of endostatin in hepatocellular cells transfected by nanoparticles, and inhibits the growth of hepatocellular carcinoma xenografts.