P. Zhang

Diagnostic and prognostic implications of microRNAs in human hepatocellular carcinoma

MicroRNAs (miRNAs) are important gene regulators, which are often deregulated in cancers. In this study, the authors analyzed the microRNAs profiles of 78 matched cancer/noncanerous liver tissues from HCC patients and 10 normal liver tissues and found that 69 miRNAs were differentially expressed between hepatocellular carcinoma (HCC) and corresponding noncancerous liver tissues (N). Then the expressions of 8 differentially expressed miRNAs were validated by real time RT PCR. The set of differentially expressed miRNAs could distinctly classify HCC, N and normal liver tissues (NL). Moreover, some of these differentially expressed miRNAs were related to the clinical factors of HCC patients. Most importantly, Kaplan‐Meier estimates and the log‐rank test showed that high expression of hsa‐miR‐125b was correlated with good survival of HCC patients (hazard ratio, 1.787, 95% confidence interval, 1.020–3.133, p = 0.043). The transfection assay showed that overexpression of miR‐125b in HCC cell line could obviously suppress the cell growth and phosporylation of Akt. In conclusion, the authors have demonstrated the diagnostic miRNA profile for HCC, and for the first time, identified the miR‐125b with predictive significance for HCC prognosis.

Upregulation of miR-23a∼27a∼24 decreases transforming growth factor-beta-induced tumor-suppressive activities in human hepatocellular carcinoma cells

Transforming growth factor‐beta (TGF‐beta) plays a dual and complex role in human cancer. In this report, we observe a specific set of MicroRNAs (miRNAs) changed in response to TGF‐beta in human hepatocellular carcinoma (HCC) cells by miRNA microarray screening. A cluster of miRNA, miR‐23a∼27a∼24, is induced in an early stage by TGF‐beta in Huh‐7 cells. Knockdown of Smad4, Smad2 or Smad3 expression by RNA interference can attenuate the response of miR‐23a∼27a∼24 to TGF‐beta addition, indicating that this induction is dependent on Smad pathway. We also explore that miR‐23a∼27a∼24 can function as an antiapoptotic and proliferation‐promoting factor in liver cancer cells. In addition, expression of this miRNA cluster is found to be remarkably upregulated in HCC tissues versus normal liver tissues. These findings suggest a novel, alternative mechanism through which TGF‐beta could induce specific miRNA expression to escape from tumor‐suppressive response in HCC cells.

Upregulation of miR-23a approximately 27a approximately 24 decreases transforming growth factor-beta-induced tumor-suppressive activities in human hepatocellular carcinoma cells.

Transforming growth factor‐beta (TGF‐beta) plays a dual and complex role in human cancer. In this report, we observe a specific set of MicroRNAs (miRNAs) changed in response to TGF‐beta in human hepatocellular carcinoma (HCC) cells by miRNA microarray screening. A cluster of miRNA, miR‐23a∼27a∼24, is induced in an early stage by TGF‐beta in Huh‐7 cells. Knockdown of Smad4, Smad2 or Smad3 expression by RNA interference can attenuate the response of miR‐23a∼27a∼24 to TGF‐beta addition, indicating that this induction is dependent on Smad pathway. We also explore that miR‐23a∼27a∼24 can function as an antiapoptotic and proliferation‐promoting factor in liver cancer cells. In addition, expression of this miRNA cluster is found to be remarkably upregulated in HCC tissues versus normal liver tissues. These findings suggest a novel, alternative mechanism through which TGF‐beta could induce specific miRNA expression to escape from tumor‐suppressive response in HCC cells.

Analysis of acetylcholine, choline and butyrobetaine in human liver tissues by hydrophilic interaction liquid chromatography-tandem mass spectrometry.

The strong polar quaternary ammoniums, acetylcholine (ACh), choline (Ch) and butyrobetaine (BB, (3-carboxypropyl)trimethylammonium), are believed playing important roles in liver metabolism. These metabolites are at low levels and are weakly retained on reversed-phase liquid chromatographic (RP-LC) columns. Several hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS) methods have been reported to analyze these compounds from different samples. However, no application to human liver tissues has been published. In this study, HILIC-MS/MS method was developed to simultaneously determine these three metabolites in human liver tissues. They were simply extracted from tissue, separated on a HILIC column, and detected by tandem MS in the mode of multiple reaction monitoring (MRM). Further studies on the recovery and repeatability based on real samples indicated the method was accurate and reliable. This method was successfully applied to measure the levels of ACh, Ch and BB in 61 human liver tissue samples including normal, hepatocellular carcinoma (HCC) and matched non-cancerous liver tissues. By comparison of Ch and ACh contents in 29 HCC with their matched non-cancerous liver tissues, it was found that ACh content increased in 11/29 HCC cases and decreased in 13/29 cases. Furthermore, the ACh/Ch ratio increased in 16/29 HCC cases, while it decreased in 8/29 cases. These results strongly indicated that there exist different patterns of ACh content in cancer tissues among HCC patients, thus highlighting the understanding of ACh and its relevant signal pathways in hepatic carcinogenesis and HCC progression.

BNIPL-2, a novel homologue of BNIP-2, interacts with Bcl-2 and Cdc42GAP in apoptosis.

The execution phase of apoptosis is characterized by marked changes in cell morphology that include contraction and membrane blebbing. Little is known about the mechanisms underlying this process. We report here the identification of a novel member of BNIPL family, designated Bcl-2/adenovirus E1B 19kDa interacting protein 2 like-2 (BNIPL-2), which interacts with Bcl-2 and Cdc42GAP. We found that the human BNIPL-2 shares homology to human BNIP-2 and also possesses a BNIP-2 and Cdc42GAP homology (BCH) domain. Deletion experiments indicated that the BCH domain of BNIPL-2 is critical for its interactions with the Bcl-2 and Cdc42GAP and also for its cell death-inducing function. Our data showed that BNIPL-2 may be a linker protein located at the front end of Bcl-2 pathway for DNA fragmentation and Cdc42 signaling for morphological changes during apoptosis. We propose that BNIPL-2 protein may play an important role in regulation of both pathways for DNA fragmentation and for formation of membrane blebs in apoptotic cells.

3D visualization of two-phase flow in the micro-tube by a simple but effective method

The present study provides a simple but effective method for 3D visualization of the two-phase flow in the micro-tube. An isosceles right-angle prism combined with a mirror located 45° bevel to the prism is employed to synchronously obtain the front and side views of the flow patterns with a single camera, where the locations of the prism and the micro-tube for clear imaging should satisfy a fixed relationship which is specified in the present study. The optical design is proven successfully by the tough visualization work at the cryogenic temperature range. The image deformation due to the refraction and geometrical configuration of the test section is quantitatively investigated. It is calculated that the image is enlarged by about 20% in inner diameter compared to the real object, which is validated by the experimental results. Meanwhile, the image deformation by adding a rectangular optical correction box outside the circular tube is comparatively investigated. It is calculated that the image is reduced by about 20% in inner diameter with a rectangular optical correction box compared to the real object. The 3D re-construction process based on the two views is conducted through three steps, which shows that the 3D visualization method can easily be applied for two-phase flow research in micro-scale channels and improves the measurement accuracy of some important parameters of the two-phase flow such as void fraction, spatial distribution of bubbles, etc.

Identification of carboxypeptidase of glutamate like-B as a candidate suppressor in cell growth and metastasis in human hepatocellular carcinoma

Purpose: We have previously done large-scale cDNA transfection screening on human hepatocellular carcinoma (HCC) cells and have identified 3,806 cDNA genes that possess the ability of either stimulating or inhibiting cell growth. In this study, we characterized one of these growth suppressor genes, carboxypeptidase of glutamate like-B (CPGL-B), in HCC. Experimental Design: Semiquantitative reverse-transcription PCR was used to examine the expression levels of CPGL-B. The cellular localization and functions of CPGL-B were investigated by enforced expression of CPGL-B in HCC cells. Results: From our previous cDNA transfection screening, we identified a gene named CPGL and its isoform, CPGL-B. With computational analysis, CPGL was located at chromosome 18q22.3 and was a homologue of peptidase family M20. CPGL was expressed in all adult and fetal tissues, whereas its isoform, CPGL-B, lacking exons 3 and 4, was expressed in all fetal tissues but only in liver and placenta of adult tissues. In HCC, CPGL-B was frequently underexpressed (35 of 90, 38.9%) in tumorous tissues compared with the corresponding nontumorous livers. Intriguingly, the underexpression was significantly associated with the presence of venous invasion (P = 0.018) and tumor microsatellite formation (P = 0.004). Stable transfection of CPGL-B in SMMC7721 HCC cells showed significant inhibition in cell viability, colony formation, cell invasion, and tumor formation in nude mice. CPGL-B also down-regulated CXCR3, matrix metalloproteinase 11, and CD44s, which are involved in cell growth and cell migration. Conclusions: These findings suggest that the frequent underexpression of CPGL-B may be associated with cell growth and metastasis of HCC.

Performance and applicability of a dc refrigerator powered by the photovoltaics

Due to the gradual depletion and excessive consumption of fossil fuels and the resulting environmental problems, increasing attention has been paid to the utilization of the renewable energy sources such as solar energy, etc. The development and performance investigation of a photovoltaic-powered refrigerator is presented in the paper. The field performance test results show that the refrigerator can work well in Shanghai area and the lowest temperature in the freezer can reach near −15u2009°C and the power consumption is also reasonable. Furthermore, the energy flow of the system and applicability of it to different climatic regions have been analyzed and discussed. It can be concluded that the performance of the refrigerator is very stable and it can serve for perishable food storage, vaccine, and drug depositing at low temperatures in the remote area and it can even be used as the electrical appliance in “zero-energy” building.

Development of a Cryogenic Calorimeter for Investigating Beam-Based Heat Load of Superconducting Undulators

Superconducting undulators provide higher magmatic field to increase the brilliance and photon energy of synchrotron light sources. To quantify the amount of beam-based heat load of storage rings and optimize the design of cryogenic system, Lawrence Berkeley National Laboratory (LBNL) proposed a cryogenic calorimeter to perform the working condition of superconducting undulators. The calorimeter has been developed by Shanghai Institute of Applied Physics (SINAP) and installed on storage ring of Shanghai Synchrotron Radiation Facility (SSRF). Also, online experiments started in September of 2012. This paper describes the cryogenic system and beam-based heat load measurement system. Also, some measurement results are given in the paper.

Identification of FN1BP1 as a Novel Cell Cycle Regulator through Modulating G1 Checkpoint in Human Hepatocarcinoma Hep3B Cells

A novel human gene, FN1BP1 (fibronectin 1 binding protein 1), was identified using the human placenta cDNA library. Northern blotting showed a transcript of ∼2.8 kb in human placenta, liver, and skeletal muscle tissues. This mRNA transcript length was similar to the full FN1BP1 sequence obtained previously. We established a conditionally induced stable cell line of Hep3B-Tet-on-FN1BP1 to investigate the preliminary function and mechanism of the secretory FN1BP1 protein. Cell-proliferation and colony-conformation assays demonstrated that FN1BP1 protein suppressed Hep3B cell growth and colonization in vitro. Analysis of Atlas human cDNA expression indicated that after FN1BP1 Dox-inducing expression for 24 h, 19 genes were up-regulated and 22 genes were down-regulated more than two-fold. Most of these gene changes were related to cell-cycle-arrest proteins (p21cip1, p15, and cyclin E1), transcription factors (general transcription factors, zinc finger proteins, transcriptional enhancer factors), SWI/SNF (SWItch/Sucrose NonFermentable) complex units, early-response proteins, and nerve growth or neurotrophic factors. Down-regulated genes were subject to colony-stimulating factors (e.g., GMSFs), and many repair genes were involved in DNA damage (RAD, ERCC, DNA topoisomerase, polymerase, and ligase). Some interesting genes (p21cip1, ID2, GMSF, ERCC5, and RPA1), which changed in the cDNA microarray analysis, were confirmed by semi-qRT-PCR, and similar changes in expression were observed. FCM cell-cycle analysis indicated that FN1BP1 over-expression could result in G1 phase arrest. FN1BP1 might inhibit cell growth and/or colony conformation through G1 phase arrest of the Hep3B cell cycle. These results indicate the potential role of FN1BP1 as a treatment target for hepatocellular carcinoma.