Polymer-acid-metal quasi-ohmic contact for stable perovskite solar cells beyond a 20,000-hour extrapolated lifetime.

The development of a robust quasi-ohmic contact with minimal resistance, good stability and cost-effectiveness is crucial for perovskite solar cells. We introduce a generic approach featuring a Lewis-acid layer sandwiched between dopant-free semicrystalline polymer and metal electrode in perovskite solar cells, resulting in an ideal quasi-ohmic contact even at elevated temperature up to 85 °C. The solubility of Lewis acid in alcohol facilitates nondestructive solution processing on top of polymer, which boosts hole injection from polymer into metal by two orders of magnitude. By integrating the polymer-acid-metal structure into solar cells, devices exhibit remarkable resilience, retaining 96% ± 3%, 96% ± 2% and 75% ± 7% of their initial efficiencies after continuous operation in nitrogen at 35 °C for 2212 h, 55 °C for 1650 h and 85 °C for 937 h, respectively. Leveraging the Arrhenius relation, we project an impressive T80 lifetime of 26,126 h at 30 °C.

Approach to Significantly Enhancing the Electrochromic Performance of PANi by In Situ Electrodeposition of the PANi@MXene Composite Film.

Electrochromic materials (ECMs) are capable of reversibly adjusting their transmittance or reflectance properties in response to changes in the external biasing voltages. In this study, we enhanced the electrochromic and electrochemical properties of polyaniline (PANi) effectively through the incorporation of MXene Ti2CTx using an in situ composite strategy. This improvement in the electrochromic and electrochemical properties observed can be attributed to the intermolecular forces between the aniline group of PANi and the terminal groups of MXene Ti2CTx sheets. The presence of hydrogen bonds between the PANi monomers and the MXene sheets was confirmed through theoretical calculations and photoluminescence results, which effectively improved the composite interfaces. Additionally, the PANi@MXene composite films were successfully prepared through a simple one-step in situ polymerization process, as verified by SEM and XPS characterization. The electrochemical studies revealed enhanced electronic conductivity, a high ion diffusion coefficient, and a narrow energy redox gap, all contributing to the excellent electrochemical properties observed. Overall, our results demonstrate that the MXene Ti2CTx composition effectively enhances the electrochromic performance of PANi. The PANi@MXene composite films exhibited a high optical modulation range, rapid switching response time, good thermal radiation regulation, and excellent operational stability. This composite strategy significantly improves the performance and practical applicability of ECMs.

Trace Doping: Fluorine-Containing Hydrophobic Lewis Acid Enables Stable Perovskite Solar Cells.

With the rapid development in perovskite solar cell (PSC), high efficiency has been achieved, but the long-term operational stability is still the most important challenges for the commercialization of this emerging photovoltaic technology. So far, bi-dopants lithium bis(trifluoromethylsulfonyl)-imide (Li-TFSI)/4-tert-butylpyridine (t-BP)-doped hole-transporting materials (HTM) have led to state-of-the art efficiency in PSCs. However, such dopants have several drawbacks in terms of stability, including the complex oxidation process, undesirable ion migration and ultra-hygroscopic nature. Herein, a fluorine-containing organic Lewis acid dopant bis(pentafluorophenyl)zinc (Zn-FP) with hydrophobic property and high migration barrier has been employed as a potential alternative to widely employed bi-dopants Li-TFSI/t-BP for poly[bis(4-phenyl)(2,4,6-trimethylphenyl)amine] (PTAA). The resulting Zn-FP-based PSCs achieve a maximum PCE of 20.34 % with hysteresis-free current density-voltage (J-V) scans. Specifically, the unencapsulated device exhibits a significantly advanced of operational stability under the International Summit on Organic Photovoltaic Stability protocols (ISOS-L-1), maintaining over 90 % of the original efficiency after operation for 1000 h under continuous 1-sun equivalent illumination in N2 atmosphere in both forward and reverse J-V scan.

Integrated System Built for Small-Molecule Semiconductors via High-Throughput Approaches.

High-throughput synthesis of solution-processable structurally variable small-molecule semiconductors is both an opportunity and a challenge. A large number of diverse molecules provide a possibility for quick material discovery and machine learning based on experimental data. However, the diversity of the molecular structure leads to the complexity of molecular properties, such as solubility, polarity, and crystallinity, which poses great challenges to solution processing and purification. Here, we first report an integrated system for the high-throughput synthesis, purification, and characterization of molecules with a large variety. Based on the principle "Like dissolves like," we combine theoretical calculations and a robotic platform to accelerate the purification of those molecules. With this platform, a material library containing 125 molecules and their optical-electronic properties was built within a timeframe of weeks. More importantly, the high repeatability of recrystallization we design is a reliable approach to further upgrading and industrial production.

A new strategy for constructing a dispiro-based dopant-free hole-transporting material: spatial configuration of spiro-bifluorene changes from a perpendicular to parallel arrangement.

Due to the low intrinsic hole mobility caused by the orthogonal conformation of two fluorene units in Spiro-OMeTAD which is a classic hole-transporting material (HTM) in perovskite solar cells (PSCs), Spiro-OMeTAD based PSCs generally can only obtain high performances through a sophisticated doping process with dopants/additives, which adds to the cost and complicacy of device fabrication, and also adversely affects the stability of PSC devices. Herein, a novel dispiro-based HTM, WH-1, is designed by cleverly replacing the central carbon atom of Spiro-OMeTAD with cyclohexane, and the spatial configuration of the HTM is changed from vertical orthogonality of the two fluorene units to a parallel arrangement, which is beneficial for the formation of a homogeneous and compact HTM film on the surface of the perovskite film, improvement of intermolecular electronic coupling and intrinsic hole mobility. WH-1 is obtained by two-step facile synthesis with a high yield from commercially available materials. WH-1 is used in PSCs as a dopant-free HTM, which is the first time that the dispiro-based molecule has been applied as a dopant-free HTM, and a power conversion efficiency (PCE) of 19.57% is obtained, rivaling Li-TFSI/t-BP doped Spiro-OMeTAD in PCE (20.29%), and showing obvious superior long-term stability.

Intelligent Biomimetic Chameleon Skin with Excellent Self-Healing and Electrochromic Properties.

Animals such as chameleons possess a natural ability to adjust their skin color as a preventive measure to deter any potential threat and to self-heal damaged skin tissues. Inspired by this, we present here a copolymer film possessing biomimetic properties that simultaneously integrates electrochromic triphenylamine and self-healing Diels-Alder groups. The flexible and stretchable copolymer film acts like natural chameleon skin, which exhibits significant color variation and also possesses excellent self-healing properties. These remarkable features make it a promising material for overcoming the crack-generation issue inherited by conventional biomimetic chameleon skin. Moreover, a flexible and wearable skin device based on the copolymer film with silver fabric as a electrode has also been fabricated. The electrochromic and self-healing properties were verified for the copolymer film, and it has been elucidated that the intelligent biomimetic "chameleon skin" was a new step toward the development of highly advanced biomimetic materials and devices.

Immunohistochemical profile of ING3 protein in normal and cancerous tissues.

The inhibitor of growth family, member 3 (ING3) protein may be capable of blocking the cell cycle via activating p53-transactivated promoters of p21 and Bcl2-associated X protein, and may induce apoptosis via a Fas/caspase-8-dependent signaling pathway. In the present study, immunohistochemistry was performed in order to characterize the expression profile of ING3 protein in tissue microarrays containing mouse and human normal tissue, human hepatocellular (n=62), renal clear cell (n=62), pancreatic (n=62), esophageal squamous cell (n=45), cervical squamous cell (n=31), breast (n=144), gastric (n=196), colorectal (n=96), ovarian (n=208), endometrial (n=96) and lung carcinoma (n=192). In mouse tissue, ING3 protein was positively detected in the cytoplasm of cardiomyocytes, kidney and skeletal muscle cells, and was additionally detected in the cytoplasm and nucleus of bronchial and alveolar epithelium, gastric and intestinal gland, and mammary gland cells. In human tissues, ING3 protein was principally distributed in the cytoplasm, but was observed in the cytoplasm and nucleus of tongue, esophagus, stomach, intestine, lung, skin, appendix, bladder, cervix and breast cells. ING3 immunoreactivity was strongly detected in the stomach, skin and cervical tissues, whereas a weak signal was detected in the cerebellum, brain stem, thymus, liver, skeletal muscle, testis and prostate. In total, ING3-positive specimens were identified in 424 of 1,194 tested cancer entities (35.5%). In a number of cases, ING3 expression was observed to be restricted to the cytoplasm and nucleus, excluding the cytoplasmic distribution identified in breast and hepatocellular carcinoma. Among these cases, ING3 was more frequently expressed in breast and gynecological types of cancer, including ovarian (59.2%), endometrial (47.9%), breast (38.9%) and cervical (35.5%) cancer. ING3-positive cases were more rare in renal clear cell (17.7%), hepatocellular (16.1%) and esophageal carcinoma (17.8%). It is suggested that ING3 may be involved in the repair and regeneration of organs or tissues, and may be closely associated with gynecological carcinogenesis.

BTG1 expression correlates with pathogenesis, aggressive behaviors and prognosis of gastric cancer: a potential target for gene therapy.

Here, we found that BTG1 overexpression inhibited proliferation, migration and invasion, induced G2/M arrest, differentiation, senescence and apoptosis in BGC-823 and MKN28 cells (p < 0.05). BTG1 transfectants showed a higher mRNA expression of Cyclin D1 and Bax, but a lower mRNA expression of cdc2, p21, mTOR and MMP-9 than the control and mock (p < 0.05). After treated with cisplatin, MG132, paclitaxel and SAHA, both BTG1 transfectants showed lower mRNA viability and higher apoptosis than the control in both time- and dose-dependent manners (p < 0.05) with the hypoexpression of chemoresistance-related genes (slug, CD147, GRP78, GRP94, FBXW7 TOP1, TOP2 and GST-π). BTG1 expression was restored after 5-aza-2'-deoxycytidine treatment in gastric cancer cells. BTG1 expression was statistically lower in gastric cancer than non-neoplastic mucosa and metastatic cancer in lymph node (p < 0.05). BTG1 expression was positively correlated with depth of invasion, lymphatic and venous invasion, lymph node metastasis, TNM staging and worse prognosis (p < 0.05). The diffuse-type carcinoma showed less BTG1 expression than intestinal- and mixed-type ones (p < 0.05). BTG1 overexpression suppressed tumor growth and lung metastasis of gastric cancer cells by inhibiting proliferation, enhancing autophagy and apoptosis in xenograft models. It was suggested that down-regulated BTG1 expression might promote gastric carcinogenesis partially due to its promoter methylation. BTG1 overexpression might reverse the aggressive phenotypes and be employed as a potential target for gene therapy of gastric cancer.

ING5 suppresses proliferation, apoptosis, migration and invasion, and induces autophagy and differentiation of gastric cancer cells: a good marker for carcinogenesis and subsequent progression.

Here, we found that ING5 overexpression increased autophagy, differentiation, and decreased proliferation, apoptosis, migration, invasion and lamellipodia formation in gastric cancer cells, while ING5 knockdown had the opposite effects. In SGC-7901 transfectants, ING5 overexpression caused G1 arrest, which was positively associated with 14-3-3 overexpression, Cdk4 and c-jun hypoexpression. The induction of Bax hypoexpression, Bcl-2, survivin, 14-3-3, PI3K, p-Akt and p70S6K overexpression by ING5 decreased apoptosis in SGC-7901 cells. The hypoexpression of MMP-9, MAP1B and flotillin 2 contributed to the inhibitory effects of ING5 on migration and invasion of SGC-7901 cells. ING5 overexpression might activate both ß-catenin and NF-κB pathways in SGC-7901 cells, and promote the expression of down-stream genes (c-myc, VEGF, Cyclin D1, survivin, and interleukins). Compared with the control, ING5 transfectants displayed drug resistance to triciribine, paclitaxel, cisplatin, SAHA, MG132 and parthenolide, which was positively related to their apoptotic induction and the overexpression of chemoresistance-related genes (MDR1, GRP78, GRP94, IRE, CD147, FBXW7, TOP1, TOP2, MLH1, MRP1, BRCP1 and GST-π). ING5 expression was higher in gastric cancer than matched mucosa. It was inversely associated with tumor size, dedifferentiation, lymph node metastasis and clinicopathological staging of cancer. ING5 overexpression suppressed growth, blood supply and lung metastasis of SGC-7901 cells by inhibiting proliferation, enhancing autophagy and apoptosis in xenograft models. It was suggested that ING5 expression might be employed as a good marker for gastric carcinogenesis and subsequent progression by inhibiting proliferation, growth, migration, invasion and metastasis. ING5 might induce apoptotic and chemotherapeutic resistances of gastric cancer cells by activating ß-catenin, NF-κB and Akt pathways.

The roles of BTG3 expression in gastric cancer: a potential marker for carcinogenesis and a target molecule for gene therapy.

BTG (B-cell translocation gene) can inhibit cell proliferation, metastasis and angiogenesis, cell cycle progression, and induce differentiation in various cells. Here, we found that BTG3 overexpression inhibited proliferation, induced S/G2 arrest, differentiation, autophagy, apoptosis, suppressed migration and invasion in MKN28 and MGC803 cells (p < 0.05). BTG3 transfectants showed a higher mRNA expression of p27, Bax, 14-3-3, Caspase-3, Caspase-9, Beclin 1, NF-κB, IL-1, -2, -4, -10 and -17, but a lower mRNA expression of p21, MMP-9 and VEGF than the control and mock (p < 0.05). At protein level, BTG3 overexpression increased the expression of CDK4, AIF, LC-3B, Beclin 1 and p38 (p < 0.05), but decreased the expression of p21 and ß-catenin in both transfectants (p < 0.05). After treated with cisplatin, MG132, paclitaxel and SAHA, both BTG3 transfectants showed lower viability and higher apoptosis than the control in both time- and dose-dependent manners (p < 0.05). BTG3 expression was restored after 5-aza-2'-deoxycytidine or MG132 treatment in gastric cancer cells. BTG3 expression was decreased in gastric cancer in comparison to the adjacent mucosa (p < 0.05), and positively correlated with venous invasion and dedifferentiation of cancer (p < 0.05). It was suggested that BTG3 expression might contribute to gastric carcinogenesis. BTG3 overexpression might reverse the aggressive phenotypes and be employed as a potential target for gene therapy of gastric cancer.

The oncogenic role of JC virus T antigen in lens tumors without cell specificity of alternative splicing of its intron.

JC virus (JCV), a ubiquitous polyoma virus that commonly infects the human, is identified as the etiologic agent for progressive multifocal leukoencephalopathy and some malignancies. To clarify the oncogenic role of JCV T antigen, we established two transgenic mice of T antigen using either α-crystallin A (αAT) or cytokeratin 19(KT) promoter. Lens tumors were found in high-copy αAT mice with the immunopositivity of T antigen, p53, ß-catenin and N-cadherin. Enlarged eyeballs were observed and tumor invaded into the brain by magnetic resonance imaging and hematoxylin-and-eosin staining. The overall survival time of homozygous mice was shorter than that of hemizygous mice (p<0.01), the latter than wild-type mice (p<0.01). The spontaneous salivary tumor and hepatocellular carcinoma were seen in αAT5 transgenic mice with no positivity of T antigen. KT7 mice suffered from lung tumor although JCV T antigen was strongly expressed in gastric epithelial cells. The alternative splicing of T antigen intron was detectable in the lens tumor of αAT mice, gastric mucosa of KT mice, and various cells transfected with pEGFP-N1-T antigen. It was suggested that JCV T antigen might induce carcinogenesis at a manner of cell specificity, which is not linked to alternative splicing of its intron. Both spontaneous lens and lung tumor models provide good tools to investigate the oncogenic role of JCV T antigen.

Ginsenoside Rh2 inhibits metastasis of glioblastoma multiforme through Akt-regulated MMP13.

Glioblastoma multiforme (GBM) is the most malignant type of primary brain tumor. Although the growth of the tumor cells in a relatively closed space may partially account for its malignancy, highly invasive nature of glioblastoma cells has been suggested to be the main reason for the failure of current therapeutic approaches. Ginsenoside Rh2 (GRh2) has recently been shown to significantly suppress the growth and survival of GBM through inhibiting epidermal growth factor receptor signaling, whereas its effects on the invasion and metastasis have not been examined. Here, we showed that GRh2 dose-dependently decreased GBM cell invasiveness in both scratch wound healing assay and Transwell cell migration assay. Moreover, the inhibitory effects of GRh2 on cell migration seemed to be conducted through decreased expression of matrix metalloproteinase (MMP)-13. Furthermore, using specific inhibitors, we found that GRh2 inhibited MMP13 through PI3k/Akt signaling pathway. Finally, high MMP13 levels were detected in GBM specimen from the patients. Together, these data suggest that GRh2 may suppress GBM migration through inhibiting Akt-mediated MMP13 activation. Thus, our data highlight a previous unappreciated role for GRh2 in suppressing GBM cell metastasis.

S100A4 expression is closely linked to genesis and progression of glioma by regulating proliferation, apoptosis, migration and invasion.

BACKGROUND: The calcium-binding S100A4 protein is involved in epithelial to mesenchymal transition, oncogenic transformation, angiogenesis, cytoskeletal integrity, mobility and metastasis of cancer cells. This study aimed to clarify the roles of S100A4 in genesis and progression of glioma. MATERIALS AND METHODS: S100A4 expression was examined by real-time RT-CPR and Western blot in glioma and paired normal brain tissue (n=69), and compared with clinicopathological parameters of tumors. In addition, glioma U251 cells transfected with an S100A4-expressing plasmid were examined for proliferation by MTT, apoptosis by Annexin V-FITC, and migration and invasion with Transwell chambers. RESULTS: Increased S100A4 mRNA expression was found in gliomas, compared with paired non-tumor tissue (p<0.001). Gradual elevation of overexpression of S100A4 was observed with increasing glioma grade (p<0.001). Astrocytoma showed lower S100A4 mRNA expression than oligodendrogliomas, with glioblastomas having highest values (p<0.001). Similar results were obtained for S100A4 protein, a positive link being found between mRNA and protein expression in gliomas (p<0.001). There was higher growth, lower apoptosis, stronger migration and invasion of S100A4 transfectants than control and mock transfected cells (p<0.001). CONCLUSIONS: These findings indicate that up-regulated S100A4 expression is positively linked to pathogenesis, progression and histogenesis of glioma by modulating proliferation, apoptosis, migration and invasion.

Downregulation of Heparanase Expression Results in Suppression of Invasion, Migration, and Adhesion Abilities of Hepatocellular Carcinoma Cells.

OBJECTIVE: Heparanase (HPSE) is high-expressed in most malignant tumors including hepatocellular carcinoma (HCC) and promotes cancer cell invasion and migration. The aim of the study is to explore whether HPSE enhances adhesion in metastasis of HCC cells. METHODS: HPSE expressions in human HCC cells were measured with real-time RT-PCR and Western blot analysis. Four recombinant miRNA vectors pcDNATM6.2-GW/EmGFP-miR-HPSE (pmiR-HPSE) were transfected into HCCLM3 cell. HPSE expression in transfected cell was measured. The cell invasion, migration, and adhesion abilities were detected, respectively. RESULTS: Both HPSE mRNA and protein relative expression levels were higher in HepG2, BEL-7402, and HCCLM3 cells than those in normal hepatocyte (P < 0.05). HPSE showed highest expression level in HCCLM3 cell (P < 0.05). Transfection efficiencies of four miRNA vectors were 75%-85%. The recombinant vectors significantly decreased HPSE expression in transfected HCCLM3 cells (P < 0.01), and pmiR-HPSE-1 showed best interference effect (P < 0.05). pmiR-HPSE-1 significantly decreased the penetrated and migrating cells numbers and adherence rate of HCCLM3 cells (P < 0.05). CONCLUSION: HPSE is a potentiator of cell adhesion in metastasis of HCC.

Association between -1562C>T polymorphism in the promoter region of matrix metalloproteinase-9 and coronary artery disease: a meta-analysis.

AIMS: The present study was undertaken to determine the association between -1562C>T polymorphism in the promoter region of matrix metalloproteinase-9 (MMP-9) and coronary artery disease (CAD) risk. METHODS: This meta-analysis was on the basis of 26 studies that included 12,776 cases and 6371 controls, heterogeneity of which was assessed by the Q-statistic test and the I(2)-statistic test. Sensitivity analysis was conducted by sequentially omitting any single study and recalculating the odds ratios (ORs) and 95% confidence intervals (CIs). Funnel plots and Egger's test were performed to test the potential publication bias. All data were analyzed by using STATA version 12.0. RESULTS: We found that -1562C>T polymorphism did not contribute to the risk of CAD in the overall results. But the stratified analysis by ethnicity indicated that -1562C>T polymorphism might decrease susceptibility to CAD in Asians (OR, 0.94; 95% CI, 0.88-1.00; ph=0.956 for CC vs. CT+TT). CONCLUSIONS: Our meta-analysis supports the fact that -1562C>T polymorphism may have association with CAD risk in Asian populations. But further larger studies are required to confirm our findings.

Dyna-CT-assisted percutaneous microballoon compression for trigeminal neuralgia.

BACKGROUND: Percutaneous microballoon compression (PMC) for trigeminal neuralgia (TN) is a well-established technique. However, complications from cannulating the foramen ovale (FO) have been reported because direct puncture of the FO is sometimes difficult. Here we report our experience with Dyna-CT-assisted PMC for TN in cannulating the FO and determining the position and volume of the balloon. METHODS: Dyna-CT-assisted PMC was performed for image reconstruction in 16 cases. The optimal working projection was generated and further fluoroscopic data were used to determine the relationship of the needle with the FO during puncture. The balloon position and three-dimensional shape were verified with Dyna-CT during balloon compression and the balloon volume and puncture angle were also calculated. Patients' prognosis is discussed. RESULTS: Dyna-CT allows quick, safe and easy cannulation of the FO. It provided three-dimensional images which were more elaborate than the classic 'pear shape' images for determining correct positioning in 16 cases. The volume of the inflated balloon ranged from 568.2 mm(3) to 891.4 mm(3) (average 769.5 mm(3)). The angle of introducing the cannula ranged from 15.32° to 35.48° rotation to the midline (average 25.18°) and 38.47°-51.89° angulation to the Reid line (average 46.17°). All the patients were pain-free after PMC. Four patients had resolvable masseter weakness and fine touch loss. No recurrence of TN was reported on follow-up. CONCLUSIONS: Dyna-CT performed by digital subtraction angiography assists PMC in three ways: (1) the FO can be better visualized independent of the patient's position; (2) needle correction or insertion can be performed much more easily because of the direct fluoroscopic control; and (3) the needle position, balloon position, balloon configuration and the volume of the inflated balloon is more reliably determined. The use of Dyna-CT-assisted PMC has a low incidence of complications and a good prognosis.

Cryptotanshinone and dihydrotanshinone I exhibit strong inhibition towards human liver microsome (HLM)-catalyzed propofol glucuronidation.

Danshen is one of the most famous herbs in the world, and more and more danshen-prescribed drugs interactions have been reported in recent years. Evaluation of inhibition potential of danshen's major ingredients towards UDP-glucuronosyltransferases (UGTs) will be helpful for understanding detailed mechanisms for danshen-drugs interaction. Therefore, the aim of the present study is to investigate the inhibitory situation of cryptotanshinone and dihydrotanshinone I towards UGT enzyme-catalyzed propofol glucuronidation. In vitro the human liver microsome (HLM) incubation system was used, and the results showed that cryptotanshinone and dihydrotanshinone I exhibited dose-dependent inhibition towards HLM-catalyzed propofol glucuronidation. Dixon plot and Lineweaver-Burk plot showed that the inhibition type was best fit to competitive inhibition type for both cryptotanshinone and dihydrotanshinone I. The second plot using the slopes from the Lineweaver-Burk plot versus the concentrations of cryptotanshinone or dihydrotanshinone I was employed to calculate the inhibition parameters (Ki) to be 0.4 and 1.7µM, respectively. Using the reported maximum plasma concentration (Cmax), the altered in vivo exposure of propofol increased by 10% and 8.2% for the co-administration of dihydrotanshinone I and cryptotanshinone, respectively. All these results indicated the possible danshen-propofol interaction due to the inhibition of dihydrotanshinone I and cryptotanshinone towards the glucuronidation reaction of propofol.

Activity and tumor specificity of human heparanase gene core promoter.

Heparanase (HPSE) plays a critical role in tumor metastasis and vascularization. In addition, the human HPSE promoter has been cloned and characterized. However, the activity and specificity of the HPSE promoter in tumor cells remains unclear. The core fragment of the HPSE promoter was amplified and cloned into the multiple cloning site of the pEGFP-1 vector. The recombinant plasmid pEGFP-Hp was transfected into human umbilical vein endothelial cells (ECV304) and human hepatoma carcinoma (HepG2), laryngocarcinoma (Hep2) and chronic myelogenous leukemia (K562) cell lines. The vectors pEGFP-1 and pEGFP­N1 were used as negative and positive controls, respectively. The activity and expression of green fluorescent protein (GFP) were analyzed. Results showed that the sequence of the amplified HPSE promoter was in agreement with the GenBank data. The recombinant plasmid pEGFP-Hp was consistent with the expected result. No GFP expression was observed in the transfected cells in the pEGFP-1 group, but a high expression was observed in the pEGFP-N1 group. As regards the pEGFP-Hp group, less fluorescence was revealed in ECV cells with a relatively high fluorescence in tumor cells. The average transfection efficiencies of pEGFP-Hp in the ECV304, HepG2, Hep2 and K562 cell lines were 3.9, 21.3, 10.8 and 6.5%, respectively, while those of pEGFP-Nl were 17.1, 24.0, 14.0 and 11.0%, respectively. The HPSE gene promoter drives the expression of downstream genes in a eukaryotic vector, specifically in tumor cell lines, but its activity is relatively weak.

Knockdown of glucose-regulated protein 78 decreases the invasion, metalloproteinase expression and ECM degradation in hepatocellular carcinoma cells.

BACKGROUND: We have reported previously that overexpression of glucose-regulated protein 78 (GRP78) promotes the invasion of hepatocellular carcinoma. However, whether GRP78 knockdown affects the extracellular matrix degradation has not been elucidated. Here we are going to determine whether GRP78 knockdown affect the ECM degradation and the role of MMP-2 and MMP-9 in these process in hepatocellular carcinoma cells. METHODS: Human hepatocellular carcinoma cell line SMMC7721 and HepG2 were cultured in DMEM supplemented with 10% FBS, RT-PCR and western blot were used to detect the endogenous expression of GRP78, MMP-2, MMP-9 and TIMP-2 in SMMC7721 and HepG2. GRP78 shRNAs were transfected using lipofection2000. Transwell assay and wound healing assay were used to analyze the invasion of each transfectant. Gelatin zymography and FITC-gelatin degradation assay were employed to investigate the capabilities of ECM degradation of each transfectant. MTT assay was used to determine the proliferation status. Western blot was employed to detect the expression of matrix metalloproteinase 2(MMP-2), MMP-9, MMP-14, and tissue inhibitor of metalloproteinases 2(TIMP-2), focal adhesion kinase (FAK), ERK1/2, JNK and Src. RESULTS: According to the expression levels of GRP78, MMP-2, MMP-9, MMP-14 and TIMP-1 in hepatocellular carcinoma cell lines SMMC7721 and hepG2, we used SMMC7721 as the in vitro invasion model for further functional analysis. Using this model, we found that GRP78 knockdown decreased the invasion of tumor cells, and this inhibitory effect was independent of cell proliferation. In hepatocellular carcinoma cells, Grp78 knockdown inhibited ECM degradation and the decreased activity and expression of MMP-2, but not MMP-9 contributed largely to this impact. Further analysis revealed that the decreased activity and expression of MMP-2 is mediated by JNK. CONCLUSION: Knockdown of GRP78 decreases ECM degradation, and downregulates the expression and activity of MMP-2 and TIMP-2. These results further demonstrate that GRP78 is a potential target for inhibiting the invasion of hepatocellular carcinoma cells.