Ethanol exposure exacerbates 4-nitroquinoline-1-oxide induced esophageal carcinogenesis and induces invasive carcinoma with muscularis propria infiltration in a mouse model.

Esophageal squamous cell carcinoma (ESCC) is one of the most fatal cancers worldwide. Most ESCC patients are diagnosed at an advanced stage; however, current research on in vivo animal models accurately reflecting their clinical presentation is lacking. Alcohol consumption is a major risk factor for ESCC and has been used in several disease models for disease induction. In this study, we used 4-nitroquinoline-1-oxide in combination with ethanol to induce an in vivo ESCC mouse model. Esophageal tissues were stained with hematoxylin and eosin for histopathological examination and lesion scoring. In cellular experiments, cell adhesion and migration invasion ability were observed using phalloidin staining, cell scratch and transwell assays, respectively, and the expression of epithelial-mesenchymal transition-related markers was detected using quantitative reverse transcription polymerase chain reaction and western blotting. The results showed that ethanol-exposed mice lost more weight and had an increased number of esophageal nodules. Histological examination revealed that the lesion scores of the ethanol-exposed esophageal samples were significantly higher than those of the unexposed esophageal samples. Furthermore, ethanol-exposed esophageal cancer samples had more severe lesions with infiltration of tumor cells into the muscularis propria. In vitro cellular experiments showed that ethanol exposure induced cytoskeletal microfilament formation, promoted cell migration invasion elevated the expression of N-cadherin and Snail, and decreased the expression of E-cadherin. In conclusion, ethanol exposure exacerbates ESCC, promotes tumor cell infiltration into the muscularis propria, and could be an effective agent for establishing innovative models of invasive carcinoma.

Personalized Nanovaccines Enhance Lymph Node Accumulation and Reprogram the Tumor Microenvironment for Improved Photodynamic Immunotherapy.

Tumor immunotherapy has emerged as an efficacious therapeutic approach that mobilizes the patient's immune system to achieve durable tumor suppression. Here, we design a photodynamic therapy-motivated nanovaccine (Dex-HDL/ALA-Fe3O4) co-delivering 5-aminolevulinic acid and Fe3O4 nanozyme that demonstrate a long-term durable immunotherapy strategy. After vaccination, the nanovaccine exhibits obvious tumor site accumulation, lymph node homing, and specific and memory antitumor immunity evocation. Upon laser irradiation, Dex-HDL/ALA-Fe3O4 effectively generates reactive oxygen species at the tumor site not only to induce the immunogenic cell death-cascade but also to trigger the on-demand release of full types of tumor antigens. Intriguingly, Fe3O4 nanozyme-catalyzed hydrogen peroxide generated oxygen for alleviating tumor hypoxia and modifying the inhibitory tumor microenvironment, thereby exhibiting remarkable potential as a sensitizer. The intravenous administration of nanovaccines in diverse preclinical cancer models has demonstrated remarkable tumor regression and inhibition of postoperative tumor recurrence and metastasis, thereby enabling personalized treatment strategies against highly heterogeneous tumors.

Fabrication of MnO2 coating on aluminum honeycomb for fast catalytic decomposition of ozone at room temperature.

Herein, the coating of MnO2 nanomaterials on the surface of aluminum honeycomb was carried out to meet the requirements of high air velocity, low pressure drop and high activity in ozone removal scenarios. A commercially readily available waterborne silica sol mixed with waterborne acrylate latex was creatively utilized as the binder. A series of coating samples were prepared by spray coating method and evaluated focusing on their adhesion strength and catalytic activity towards ozone decomposition in an air duct at room temperature, by varying MnO2/binder mass ratio and number of sprayings. It was found that the adhesion strength of the catalytic coatings on the aluminum honeycomb increased with the increase of binder mass ratio, but the increased binder made the catalyst particles closely packed, resulting in reduced exposure of active sites and decrease of ozone conversion. Accordingly, catalyst slurry with 81.8 wt.% MnO2 in dry coating and spraying times of two were determined as the optimal process parameters. As-prepared aluminum honeycomb filter with MnO2 layer of 50 µm thickness achieved ozone conversion of 29.3%±1.7% under conditions of air velocity 3.0 m/sec, relative humidity ∼50%, room temperature (26°C) and initial ozone concentration of 200 ppbV. This filter can be well adaptable to indoor air purification equipment operating at high air velocity with low wind resistance.

Immune Exosomes Loading Self-Assembled Nanomicelles Traverse the Blood-Brain Barrier for Chemo-immunotherapy against Glioblastoma.

Effective drug delivery and prevention of postoperative recurrence are significant challenges for current glioblastoma (GBM) treatment. Poor drug delivery is mainly due to the presence of the blood-brain barrier (BBB), and postoperative recurrence is primarily due to the resistance of GBM cells to chemotherapeutic drugs and the presence of an immunosuppressive microenvironment. Herein, a biomimetic nanodrug delivery platform based on endogenous exosomes that could efficiently target the brain without targeting modifications and co-deliver pure drug nanomicelles and immune adjuvants for safe and efficient chemo-immunotherapy against GBM is prepared. Inspired by the self-assembly technology of small molecules, tanshinone IIA (TanIIA) and glycyrrhizic acid (GL), which are the inhibitors of signal transducers and activators of transcription 3 from traditional Chinese medicine (TCM), self-assembled to form TanIIA-GL nanomicelles (TGM). Endogenous serum exosomes are selected to coat the pure drug nanomicelles, and the CpG oligonucleotides, agonists of Toll-like receptor 9, are anchored on the exosome membrane to obtain immune exosomes loaded with TCM self-assembled nanomicelles (CpG-EXO/TGM). Our results demonstrate that CpG-EXO/TGM can bind free transferrin in blood, prolong blood circulation, and maintain intact structures when traversing the BBB and targeting GBM cells. In the GBM microenvironment, the strong anti-GBM effect of CpG-EXO/TGM is mainly attributed to two factors: (i) highly efficient uptake by GBM cells and sufficient intracellular release of drugs to induce apoptosis and (ii) stimulation of dendritic cell maturation and induction of tumor-associated macrophages polarization by CpG oligonucleotides to generate anti-GBM immune responses. Further research found that CpG-EXO/TGM can not only produce better efficacy in combination with temozolomide but also prevent a postoperative recurrence.

Sensitive electrochemical detection of A549 exosomes based on DNA/ferrocene-modified single-walled carbon nanotube complex.

Exosome is an emerging tumor marker, whose concentration level can reflect the occurrence and development of tumors. The development of rapid and sensitive exosome detection platform is of great significance for early warning of cancer occurrence. Here, a strategy for electrochemical detection of A549-cell-derived exosomes was established based on DNA/ferrocene-modified single-walled carbon nanotube complex (DNA/SWCNT-Fc). DNA/SWCNT-Fc complexes function as a signal amplification platform to promote electron transfer between electrochemical signal molecules and electrodes, thereby improving sensitivity. At the same time, the exosomes can be attached to DNA/SWCNT-Fc nanocomposites via the established PO43--Ti4+-PO43- method. Moreover, the application of EGFR antibody, which can specifically capture A549 exosomes, could improve the accuracy of this sensing system. Under optimal experimental conditions, the biosensor showed good linear relationship between the peak current and the logarithm of exosomes concentration from 4.66 × 106 to 9.32 × 109 exosomes/mL with a detection limit of 9.38 × 104 exosomes/mL. Furthermore, this strategy provides high selectivity for exosomes of different cancer cells, which can be applied to the detection of exosomes in serum samples. Thus, owing to its advantages of high sensitivity and good selectivity, this method provides a diversified platform for exosomes identification and has great potential in early diagnosis and biomedical applications.

Dual signal amplification based on polysaccharide-initiated ring-opening polymerization and click polymerization for exosomes detection.

Exosomes, as a biomarker with enhancing tumor invasion and spread, play an essential role for lung cancer diagnosis, therapy, and prognosis. In this work, a novel electrochemical sensor was fabricated for detecting exosomes secreted by lung cancer cells based on polysaccharide-initiated ring-opening polymerization (ROP) and click polymerization. First, MPA formed a self-assembled monolayer on the gold electrode surface, and then anti-EGFR was immobilized on the electrode surface by amide bond. Subsequently, a lot of phosphate groups were introduced by the specific recognition between anti-EGFR and exosomes, then sodium alginate grafted Glycidyl propargyl ether (SA-g-GPE) prepared via ROP was attached to the exosomes through PO43-Zr4+-COOH coordination bond. After that, click polymerization was initiated by alkyne groups on the SA-g-GPE polymerization chain to realize highly sensitive detection of A549 exosomes. Under the optimum conditions, the fabricated sensor showed a good linear relationship between the logarithm of exosomes concentration and peak current in the range of 5 × 103 - 5 × 109 particles/mL, and the limit of detection (LOD) was as low as 1.49 × 102 particles/mL. In addition, this method had the advantages of high specificity, anti-interference, high sensitivity, simplicity, rapidity and green economy, which proposed a novel avenue for the detection of exosomes, and also had potential applications in early cancer diagnosis and biomedicine.

Perfluorosulfonic acid polymer based eATRP for ultrasensitive detection of CYFRA21-1 DNA.

The sensitive detection of biomarker cytokeratin fragment antigen 21-1 (CYFRA21-1) is crucial for early diagnosis and screening of non-small cell lung cancer (NSCLC). In this work, an electrochemical biosensor based on Nafion-initiated eATRP has been built for ultrasensitive detection of CYFRA21-1 DNA for the first time. Specifically, peptide nucleic acid (PNA) probes are immobilized onto a gold electrode surface and then hybridized with target DNA to form PNA/DNA heteroduplexes for the subsequent attachment of Nafion by the identified carboxyl-Zr4+-phosphoric acid chemistry. Finally, polymer chains are obtained by linking the monomer of ferrocenylmethyl methacrylate to the PNA/MCH/DNA/Zr4+/Nafion probes via eATRP. Under optimized steady-state conditions, the sensor offers a wide current response for CYFRA21-1 DNA from 10-11 to 10-16 M with a detection limit of 6.42 × 10-17 M. The proposed method of using Nafion as the eATRP initiator exhibits high sensitivity, reproducibility and stability and is a promising strategy for early diagnosis of NSCLC.

Electrochemical CYFRA21-1 DNA sensor with PCR-like sensitivity based on AgNPs and cascade polymerization.

In this work, a new method of CYFRA21-1 DNA (tDNA) detection based on electrochemically mediated atom transfer radical polymerization (e-ATRP) and surface-initiated reversible addition-fragmentation chain transfer polymerization (SI-RAFT) cascade polymerization and AgNP deposition is proposed. Firstly, the peptide nucleic acid (PNA) probe is captured on a gold electrode by Au-S bonds for specific recognition of tDNA. After hybridization, PNA/DNA strands provide high-density phosphate groups for the subsequent ATRP initiator by the identified carboxylate-Zr4+-phosphate chemistry. Then, a large number of monomers are successfully grafted from the DNA through the e-ATRP reaction. After that, the chain transfer agent of SI-RAFT and methacrylic acid (MAA) are connected by recognized carboxylate-Zr4+-carboxylate chemistry. Subsequently, through SI-RAFT, the resulting polymer introduces numerous aldehyde groups, which could deposit many AgNPs on tDNA through silver mirror reaction, causing significant amplification of the electrochemical signal. Under optimal conditions, this designed method exhibits a low detection limit of 0.487 aM. Moreover, the method enables us to detect DNA at the level of PCR-like and shows high selectivity and strong anti-interference ability in the presence of serum. It suggests that this new sensing signal amplification technology exhibits excellent potential of application in the early diagnosis of non-small cell lung cancer (NSCLC). Graphical abstract Electrochemical detection principle for CYFRA21-1 DNA based on e-ATRP and SI-RAFT signal amplification technology.

Dual signal amplification by polysaccharide and eATRP for ultrasensitive detection of CYFRA 21-1 DNA.

Cytokeratin fragment antigen 21-1 (CYFRA 21-1) DNA is a crucial biomarker closely associated with non-small cell lung cancer. Here, we fabricated a novel electrochemical biosensor for ultrasensitive detection of CYFRA 21-1 DNA via polysaccharide and electrochemically mediated atom transfer radical polymerization (eATRP) dual signal amplification. Specifically, thiolated peptide nucleic acid (PNA) probes at 5'-terminals are immobilized on the gold electrode surface for specific recognition of CYFRA 21-1 DNA (tDNA). After hybridization, hyaluronic acid (HA) is linked to the hybridized PNA/DNA duplexes via the recognized carboxylate-Zr4+-phosphate chemistry. Then multiple initiators of the polymerization reaction are introduced via esterification reaction. Lastly, large numbers of electro-active monomers are successfully grafted from the initiation sites of functionalized HA by eATRP reaction, significantly amplifying the electrochemical signal. Under optimal conditions, the constructed sensor can detect as low as 9.04 aM tDNA. Further, this proposed biosensor can also be applied to the direct detection of double-stranded DNA (dsDNA), obtaining 0.12 fM as the detection limit. Besides, this strategy shows high selectivity for mismatched bases and excellent applicability for CYFRA 21-1 DNA detection in the serum samples. Given its high sensitivity, selectivity, ease of operation, low cost and environmental friendliness, this biosensor has considerable potential in early diagnosis and biomedical application.

The CpG Dinucleotide Adjacent to a κB Site Affects NF-κB Function through Its Methylation.

NF-κB is an important transcription factor that plays critical roles in cell survival, proliferation, inflammation, and cancers. Although the majority of experimentally identified functional NF-κB binding sites (κB sites) match the consensus sequence, there are plenty of non-functional NF-κB consensus sequences in the genome. We analyzed the surrounding sequences of the known κB sites that perfectly match the GGGRNNYYCC consensus sequence and identified the nucleotide at the -1 position of κB sites as a key contributor to the binding of the κB sites by NF-κB. We demonstrated that a cytosine at the -1 position of a κB site (-1C) could be methylated, which thereafter impaired NF-κB binding and/or function. In addition, all -1C κB sites are located in CpG islands and are conserved during evolution only when they are within CpG islands. Interestingly, when there are multiple NF-κB binding possibilities, methylation of -1C might increase NF-κB binding. Our finding suggests that a single nucleotide at the -1 position of a κB site could be a critical factor in NF-κB functioning and could be exploited as an additional manner to regulate the expression of NF-κB target genes.

Metastatic lung adenocarcinoma to the bladder: A case report.

Urothelial cancer is the most frequently diagnosed type of malignant tumor in the bladder, of which primary adenocarcinoma accounts for a small percentage. Secondary malignancies, in particular metastatic adenocarcinoma from the lung, are exceedingly rare, with only six cases previously reported in the literature. The present study describes the case of a 71-year-old Chinese male patient with known lung cancer for >2 years, who was diagnosed with metastatic adenocarcinoma to the bladder. The histopathological characteristics and immunohistochemical features of the patient are reported. It was proposed that pathologists should consider the possibility of metastatic adenocarcinoma from the lung, rather than assume a diagnosis of primary adenocarcinoma of the bladder or direct invasion of adenocarcinoma from the surrounding organs. Furthermore, it is essential to determine the medical history of each patient and observe the immunohistochemical features of all tumors prior to diagnosis.

Room-Temperature Oxidation of Formaldehyde by Layered Manganese Oxide: Effect of Water.

Layered manganese oxide, i.e., birnessite was prepared via the reaction of potassium permanganate with ammonium oxalate. The water content in the birnessite was adjusted by drying/calcining the samples at various temperatures (30 °C, 100 °C, 200 °C, 300 °C, and 500 °C). Thermogravimetry-mass spectroscopy showed three types of water released from birnessite, which can be ascribed to physically adsorbed H2O, interlayer H2O and hydroxyl, respectively. The activity of birnessite for formaldehyde oxidation was positively associated with its water content, i.e., the higher the water content, the better activity it has. In-situ DRIFTS and step scanning XRD analysis indicate that adsorbed formaldehyde, which is promoted by bonded water via hydrogen bonding, is transformed into formate and carbonate with the consumption of hydroxyl and bonded water. Both bonded water and water in air can compensate the consumed hydroxyl groups to sustain the mineralization of formaldehyde at room temperature. In addition, water in air stimulates the desorption of carbonate via water competitive adsorption, and accordingly the birnessite recovers its activity. This investigation elucidated the role of water in oxidizing formaldehyde by layered manganese oxides at room temperature, which may be helpful for the development of more efficient materials.

Tumor suppressor miR-34a targets PD-L1 and functions as a potential immunotherapeutic target in acute myeloid leukemia.

miRNA (miR) 34a has been shown to modulate critical gene transcripts involved in tumorigenesis, but its role in tumor-mediated immunosuppression is largely unknown. PD-L1 plays an important role in immune responses, however, presently its transcriptional regulatory mechanisms are not well understood. In the present study, we analyzed the expression of PD-L1 and miR-34a in 44 acute myeloid leukemia (AML) samples, and observed an inverse correlation between PD-L1 and miR-34a expression. Overexpression of miR-34a in HL-60 and Kasumi-1 cells blocked PD-L1 expression, and reduced PD-L1 surface expression. Using luciferase reporter assay and mutagenesis, we identified miR-34a as a putative binder of the PD-L1-3'UTR. Surface expression of PD-L1 induced by chemotherapeutic agents could also be reversed by miR-34a; furthermore, PD-L1 specific T cell apoptosis was reduced as well following miR-34a transfection. We also found that there is a positive feedback between PD-L1 expression and AKT activation. Our data suggest that miR-34a can regulate PD-L1 expression by targeting PD-L1 mRNA, and our present findings shed new light on the complex regulation of PD-L1 in human tumors, and on miR-34a in cancer immuno-based therapy.

Overexpression of PrPc, combined with MGr1-Ag/37LRP, is predictive of poor prognosis in gastric cancer.

Prion protein (PrPc) has been previously reported to be involved in gastric cancer (GC) development and progression. However, the association between expression of PrPc and GC prognosis is yet poorly characterized. In the present study, the expressions of PrPc and MGr1-Ag/37LRP, a protein interacting with PrPc, were detected using the tissue microarray technique and immunohistochemical method to compare clinicopathological parameters of 238 GC patients. We found that the expressions of PrPc and MGr1-Ag/37LRP were upregulated in GC lesions compared with their expressions in adjacent noncancerous tissues (p<0.01). High expression of PrPc was detected in 37.39% (89/238) of GC patients and positively correlated with the expression of MGr1-Ag/37LRP (r=0.532, p<0.001). PrPc expression was associated with a number of clinicopathological parameters including depth of invasion and lymph node metastasis of the tumor (p<0.001). High expression of PrPc brought a poorer prognosis than low PrPc expression. Moreover, GC patients with high level of PrPc and high level of MGr1-Ag/37LRP had the poorest prognosis. Multivariate survival analysis suggested that, along with other parameters, combined expression of PrPc and MGr1-Ag/37LRP was independent prognostic factors for GC patients. These data indicates that overexpression of PrPc, combined with MGr1-Ag/37LRP, is predictive of poor prognosis in GC and thereby could be used to guide the clinical decision.

In vivo tracking and comparison of the therapeutic effects of MSCs and HSCs for liver injury.

BACKGROUND: Mesenchymal stem cells (MSCs) and hematopoietic stem cells (HSCs) have been studied for damaged liver repair; however, the conclusions drawn regarding their homing capacity to the injured liver are conflicting. Besides, the relative utility and synergistic effects of these two cell types on the injured liver remain unclear. METHODOLOGY/PRINCIPAL FINDINGS: MSCs, HSCs and the combination of both cells were obtained from the bone marrow of male mice expressing enhanced green fluorescent protein(EGFP)and injected into the female mice with or without liver fibrosis. The distribution of the stem cells, survival rates, liver function, hepatocyte regeneration, growth factors and cytokines of the recipient mice were analyzed. We found that the liver content of the EGFP-donor cells was significantly higher in the MSCs group than in the HSCs or MSCs+HSCs group. The survival rate for the MSCs group was significantly higher than that of the HSCs or MSCs+HSCs group; all surpassed the control group. After MSC-transplantation, the injured livers were maximally restored, with less collagen than the controls. The fibrotic areas had decreased to a lesser extent in the mice transplanted with HSCs or MSCs+HSCs. Compared with mice in the HSCs group, the mice that received MSCs had better improved liver function. MSCs exhibited more remarkable paracrine effects and immunomodulatory properties on hepatic stellate cells and native hepatocytes in the treatment of the liver pathology. Synergistic actions of MSCs and HSCs were most likely not observed because the stem cells in liver were detected mostly as single cells, and single MSCs are insufficient to provide a beneficial niche for HSCs. CONCLUSIONS/SIGNIFICANCE: MSCs exhibited a greater homing capability for the injured liver and modulated fibrosis and inflammation more effectively than did HSCs. Synergistic effects of MSCs and HSCs were not observed in liver injury.

HIF-1α up-regulates NDRG1 expression through binding to NDRG1 promoter, leading to proliferation of lung cancer A549 cells.

Hypoxia-inducible signaling pathway is involved in many pathological processes, such as adaptiveness regulation of plateau environment, myocardial ischemia and tumorigenesis. NDRG1 is a member of the N-myc downregulated gene (NDRG) family, and it has strong hypoxia stress reaction functions. Although the cellular responses to hypoxia are well known, little is known about the interaction between hypoxia-inducible transcription factor (HIF)-1α and NDRG1. In this study, we cloned HIF-1α CDS, NDRG1 promoter and its truncatures, constructed pCDNA3.0-Hif-1α and pGL3-basic-NDRG1. Reporter assay results showed that HIF-1α could bind to NDRG1 promoter to activate NDRG1 expression. Further results revealed that -1202 to -450 of NDRG1 promoter is the most important region for HIF-1α binding. Then, we constructed NDRG1 stable transfection cell line. Results from MTT, colony-forming assay and flow cytometry showed that NDRG1 overexpression results in more proliferation and less apoptosis of A549 lung cancer cells. Our study elucidates the mechanism of NGRG1 in hypoxia stress reactions and may provide new strategy for hypoxia injuries.

Dynamic microRNA profiles of hepatic differentiated human umbilical cord lining-derived mesenchymal stem cells.

Despite the extensive hepatic differentiation potential of human umbilical cord lining-derived mesenchymal stem cells (hUC-MSC), little is known about the molecular mechanisms of hUC-MSC differentiation. At the post-transcriptional level, microRNAs are key players in the control of cell fate determination during differentiation. In this study, we aimed to identify microRNAs involved in the hepatic differentiation of hUC-MSCs. After successfully isolating hUC- MSCs, we induced hepatocyte formation in vitro with growth factors. After 26 days of induction, hUC-MSCs could express hepatocyte-specific genes, synthesize urea and glycogen and uptake low-density lipoprotein. Cellular total RNA from hUC-MSCs and hepatic differentiated hUC-MSCs was collected at 7 time points, including 2 days, 6 days, 10 days, 14 days, 22 days and 26 days, for microRNA microarray analysis. Dynamic microRNA profiles were identified that did not overlap or only partially overlapped with microRNAs reported to be involved in human liver development, hepatocyte regeneration or hepatic differentiation of liver-derived progenitor cells. A total of 61 microRNAs among 1205 human and 144 human viral microRNAs displayed consistent changes and were altered at least 2-fold between hUC-MSCs and hepatic differentiated hUC-MSCs. Among these microRNAs, 25 were over-expressed; this over-expression occurred either gradually or increased sharply and was maintained at a high level. A total of 36 microRNAs were under-expressed, with an expression pattern similar to that of the over-expressed microRNAs. The expression of the altered expressed microRNAs was also confirmed by quantitative reverse-transcription polymerase chain reaction. We also found that microRNAs involved in hepatic differentiation were not enriched in hepatocyte or hepatocellular carcinoma cells and can potentially target liver-enriched transcription factors and genes. The elucidation of the microRNA profile during the hepatic differentiation of hUC-MSCs provides the basis for clarifying the role of microRNAs in hUC-MSC hepatic differentiation and specific microRNA selection for the conversion of hUC-MSCs to hepatocytes.

[The construction and primary screening of a phage display library of HCV C and E1 genes evolved with an artificial pattern].

OBJECTIVES: To construct and screen a primarily phage display library of HCV C and E1 genes evolved with an artificial pattern. METHODS: Two genes of about 1 kb with different genotypes were evolved by DNA shuffling. The re-assembled HCV C and E1 genes were cloned into a phage vector. After being rescued with helper phage M13KO7, a phage display library was constructed. Then the library was screened with anti-C and E1 McAb. Double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) was carried out on twenty individual phage clones selected randomly to detect their binding and reactive activity with high-titer HCV-positive sera. Normal sera were used as controls. RESULTS: The phage display library of HCV C and E1 genes which evolved with an artificial pattern was constructed. Their capacity amounted to 1.64 x 10(6), and 86 percent of the clones contained C and E1 genes. After four rounds of panning, the phage library was specifically enriched. Twelve positive clones were successfully screened. CONCLUSION: The capacity and diversity of the constructed library are enough for screening. The results demonstrate the superiority of the specific binding and reactive activity and affinity of the 12 phage clones from the HCV positive sera.

[Experimental study on the effect of VEGF165 antisense RNA on human squamous cell carcinoma of esophagus].

AIM: To investigate the effect of antisense RNA against vascular endothelial growth factor 165 (VEGF165) on human esophagus squamous cell carcinoma cell line EC109. METHODS: Eukaryotic expression vector for VEGF165 antisense RNA was constructed and identified. Recombinant plasmid was transfected into EC109 cells and the transfected EC109 cells were inoculated subcutaneously to nude mice. The biological characteristics and tumorigenicity of transfected EC109 cells were observed by in situ hybridization, laser confocal microscope, transmission electron microscopy and flow cytometry. RESULTS: The eukaryotic expression vector pCEP-AVEGF165 was successfully constructed and expressed in transfected EC109 cells. The rate of VEGF165 expression dropped by 75% in transfected cells. The morphology and cell cycle of transfected EC109 cells were not affected by the antisense RNA, but the tumorigenicity and angiogenesis of transfected EC109 cells were greatly reduced in nude mice. The volume of tumors in pCEP-AVEGF165 transfected group, empty vector transfected group and control group were (820+/-112.5) mm3, (7 930+/-1 035) mm3 and (7 850+/-950) mm3, respectively. The microvessel density of the three groups were (8.5+/-1.2)/mm2, (44.3+/-9.4)/mm2 and (46.4+/-12.6)/mm2, respectively. CONCLUSION: The angiogenesis and tumorigenicity of human esophageal squamous cell carcinoma were effectively inhibited by VEGF165 antisense RNA, which may be applied to treat solid tumor in the future.