Expression, purification and refolding of pro-MMP-2 from inclusion bodies of E. coli.

MMP-2 has been reported as the most validated target for cancer progression and deserves further investigation. However, due to the lack of methods for obtaining large amounts of highly purified and bioactive MMP-2, identifying specific substrates and developing specific inhibitors of MMP-2 remains extremely difficult. In this study, the DNA fragment coding for pro-MMP-2 was inserted into plasmid pET28a in an oriented manner, and the resulting recombinant protein was effectively expressed and led to accumulation as inclusion bodies in E. coli. This protein was easy to purify to near homogeneity by the combination of common inclusion bodies purification procedure and cold ethanol fractionation. Then, our results of gelatin zymography and fluorometric assay revealed that pro-MMP-2 at least partially restored its natural structure and enzymatic activity after renaturation. We obtained approximately 11 mg refolded pro-MMP-2 protein from 1 L LB broth, which was higher than other strategies previously reported. In conclusion, a simple and cost-effective procedure for obtaining high amounts of functional MMP-2 was developed, which would contribute to the progress of studies on the gamut of biological action of this important proteinase. Furthermore, our protocol should be appropriate for the expression, purification, and refolding of other bacterial toxic proteins.

Phenotypic switching of vascular smooth muscle cells in the 'normal region' of aorta from atherosclerosis patients is regulated by miR-145.

Switching of vascular smooth muscle cells (VSMCs) from a contractile phenotype to an adverse proliferative phenotype is a hallmark of atherosclerosis or vascular restenosis. However, the genetic modulators responsible for this switch have not been fully elucidated in humans nor have they been correlated with clinical abnormalities. This study investigated genetic mechanisms involved in phenotypic switching of VSMCs at non-defect areas of the aorta in patients with atherosclerosis. Aortic wall samples were obtained from patients with (N = 53) and without (N = 27) atherosclerosis undergoing cardiovascular surgery. Vascular smooth muscle cell cultures were generated, and expression of microRNA-145 (miR-145), its target gene Kruppel-Like Factor 5 (KLF5) and Myocardin (MYOCD, a smooth muscle-specific transcriptional coactivator) were analysed using RT-qPCR, along with expression of relevant proteins. Vascular smooth muscle cells were transduced with miR-145 inhibitor and mimic to determine the effect of miR-145 expression on VSMC proliferation. miR-145 expression decreased while KLF5 expression increased in atherosclerotic aortas. Atherosclerotic samples and VSMCs had decreased expression of contractile markers calponin and alpha smooth muscle actin (α-SMA) and MYOCD. miR-145 inhibitor-transduced VSMCs from non-atherosclerotic patients showed decreased expression of calponin and α-SMA and increased proliferation compared with non-transduced controls, and these levels were close to those of atherosclerotic patients. miR-145 mimic-transduced VSMCs from atherosclerotic patients showed increased expression of calponin and α-SMA and decreased proliferation compared with non-transduced controls, and these levels were close to those found in non-atherosclerotic patients. These data demonstrate that miR-145 modulates the phenotypic switch of VSMCs from a contractile to a proliferative state via KLF5 and MYOCD in atherosclerosis.

CP-31398 inhibits the growth of p53-mutated liver cancer cells in vitro and in vivo.

The tumor suppressor p53 is one of the most frequently mutated genes in hepatocellular carcinoma (HCC). Previous studies demonstrated that CP-31398 restored the native conformation of mutant p53 and trans-activated p53 downstream genes in tumor cells. However, the research on the application of CP-31398 to liver cancer has not been reported. Here, we investigated the effects of CP-31398 on the phenotype of HCC cells carrying p53 mutation. The effects of CP-31398 on the characteristic of p53-mutated HCC cells were evaluated through analyzing cell cycle, cell apoptosis, cell proliferation, and the expression of p53 downstream genes. In tumor xenografts developed by PLC/PRF/5 cells, the inhibition of tumor growth by CP-31398 was analyzed through gross morphology, growth curve, and the expression of p53-related genes. Firstly, we demonstrated that CP-31398 inhibited the growth of p53-mutated liver cancer cells in a dose-dependent and p53-dependent manner. Then, further study showed that CP-31398 re-activated wild-type p53 function in p53-mutated HCC cells, which resulted in inhibitive response of cell proliferation and an induction of cell-cycle arrest and apoptosis. Finally, in vivo data confirmed that CP-31398 blocked the growth of xenografts tumors through transactivation of p53-responsive downstream molecules. Our results demonstrated that CP-31398 induced desired phenotypic change of p53-mutated HCC cells in vitro and in vivo, which revealed that CP-31398 would be developed as a therapeutic candidate for HCC carrying p53 mutation.

Enhanced removal of Cr(VI) by polyethyleneimine-modified bamboo hydrochar.

Hydrochar is an environmentally friendly and cheap adsorbent, but its adsorption amounts for anions is very limited. The functionalized hydrochar can overcome this shortcoming. Herein, polyethyleneimine-modified hydrochar (PEI-HC) was synthesized from hydrothermal carbonization (HTC) of methyl acrylate and bamboo after addition of initiator ammonium persulfate, and then modified by polyethyleneimine (PEI), which was used to treat Cr(VI). PEI-HC was tested by XANES, EXAFS, SEM-EDS, XPS, FTIR, N2 sorption isotherms, zeta potential, and elemental analyses. The characterizations showed that PEI was successfully grafted onto hydrochar, and the PEI-HC was rich in N and O functional groups, which presented high Cr(VI) sorption ability (528.41 mg·g-1 at pH 2). The bath experiments found the pseudo-second-order kinetic and Freundlich equations can well describe the adsorption kinetics and isotherm of the Cr(VI) adsorption onto PEI-HC, respectively. Electrostatic interaction, reduction, complexation, and H-bonding are the main removal mechanisms as supported by XANES, EXAFS, XPS, and FTIR. This study provides a strategy of combining HTC and free radical graft polymerization to convert agricultural and forestry wastes into functionalized hydrochar, showing highly efficient removal of Cr(VI).

Enhanced removal of Cr(VI) by cation functionalized bamboo hydrochar.

Chemical modification on hydrochars can significantly improve their ability of removing heavy metal ions from wastewater, but so far no research has focused on the chemical modification through free radical reaction. In this work, a cation functionalized hydrochar (CFHC) bearing - N+H2R was synthesized by grafting-polymerization of glycidyl methacrylate (GMA) onto bamboo hydrochar under initiation by benzoyl peroxide, followed by the amination with the introduced epoxy group and diethylenetriamine and a subsequent hydrochloric acid treatment. The resulted CFHC exhibited a superior removal capacity of 424.09 mg·g-1 for Cr(VI), and the highest sorption occurred at pH of 2. Combining a series of characterizations and tests, it was concluded that the sorption conformed to the pseudo-second-order and Freundlich equations, indicating a multilayer chemisorption process that mainly driven by electrostatic reaction, reduction, and surface complexation. This research proved that a free radical polymerization treatment could effectively transform hydrochars into super adsorbents for wastewater treatment.

Preparation of hydrochar with high adsorption performance for methylene blue by co-hydrothermal carbonization of polyvinyl chloride and bamboo.

Polyvinyl chloride (PVC) was blended into bamboo powder during co-hydrothermal carbonization (Co-HTC) to understand the effects on the physicochemical properties and adsorbing ability of hydrochar. The properties of hydrochar were characterized by Zeta potential, elemental analyses, BET, FTIR, XPS, Boehm titration and SEM. The addition of PVC into bamboo in Co-HTC decreased the BET area, and pore volume and radius of hydrochar, but increased the contents of surface hydroxyl and carboxyl groups. The adsorption ability of hydrochar produced by addition of PVC at 473 K over methylene blue (MB) increased significantly. The main adsorption mechanism was electrostatic attraction by -N(CH3)2+ of MB and carboxylate of hydrochar, and hydrogen-bonding interaction through N atom of phenothiazine in MB and C-OH of hydrochar. Thus, Co-HTC offers a facile, green and economical alternative for conversion of waste into high-value adsorbents.

miR-195 inhibits cell proliferation via targeting AEG-1 in hepatocellular carcinoma.

Emerging evidence has indicated that microRNAs (miRNAs) are frequently dysregulated and are fundamental in the pathogenesis of hepatocellular carcinoma (HCC). However, the roles of miR-195 in HCC have not been well elucidated. In the present study, the expression of miR-195 was determined to be markedly downregulated in HCC tissues and cell lines, as compared with normal liver cells. Restoration of miR-195 expression resulted in significant inhibition of the proliferation and tumorigenicity of HCC cells in vitro and in vivo. Gene expression data and luciferase reporter assays revealed that miR-195 is able to directly inhibit the expression of astrocyte elevated gene 1 (AEG-1) through interaction with its 3' untranslated region. Consistently, an inverse correlation between miR-195 and AEG-1 expression was observed in HCC tissues. Furthermore, the overexpression of AEG-1 was able to partially attenuate the miR-195-induced inhibition of cell growth and promotion of apoptosis. Taken together, these findings indicate that miR-195 functions as a tumor suppressor by inhibiting AEG-1. This pathway may provide new insights into the potential molecular mechanisms of HCC.

MiR-497 suppresses angiogenesis and metastasis of hepatocellular carcinoma by inhibiting VEGFA and AEG-1.

Hepatocellular carcinoma (HCC) is a worldwide malignance and displays marked vascular abnormalities and active metastasis. MicroRNAs (miRNAs) have been shown to play important roles in regulating tumor properties in cancer, however, whether miR-497 contributes to HCC angiogenesis or metastasis remains unclear. In this study, we found that miR-497 was significantly down-regulated in HCC tissue samples and cell lines. Gain-of-function and loss-of-function studies revealed that miR-497 could repress both the pro-angiogenic and metastatic ability of HCC cells. Subsequent investigations disclosed that miR-497 directly inhibited the 3'-untranslated regions (UTRs) of vascular endothelial growth factor A (VEGFA) and astrocyte elevated gene-1 (AEG-1). Furthermore, overexpression of these targets antagonized the function of miR-497. Based on nude mouse models, we demonstrated that overexpression of miR-497 significantly repressed microvessel densities in xenograft tumors and reduced pulmonary metastasis. In conclusion, our findings indicate that miR-497 downregulation contributes to angiogenesis and metastasis in HCC.

An unprecedented 3D POM-Ag architecture with intertwined and homological helical structures.

A new hybrid compound, Na[Ag6(pyttz)2(H2O)][PMo12O40] (pyttz = 3-(pyrid-3-yl)-5-(1H-1,2,4-triazol-3-yl)-1,2,4-triazolyl), has been hydrothermally synthesized and structurally characterized by routine techniques. X-ray diffraction analysis reveals that the title compound is constructed by the 2D Ag-pyttz coordination polymer and 3D Ag-POM architecture with helix. A fascinating structural feature is the assembling fashion of the right- and left-helical chain, namely, the helical chains with different orientations are intertwined with each other forming intertwined double helical layers along the c-axis, and the identical left- or right-handed helical chains are fused together in a hand-by-hand mode generating another homological helical layer along the a-axis. As a result, these helical layers intersect each other obtaining an unprecedented 3D POM-Ag inorganic architecture. Note that the 3D framework with a helix constructed by POMs and metal ions has never been observed up to date. Additionally, its photocatalytic degradation of RhB was also investigated.

A new molybdenum(V) nickel phosphate based on divacant [H30(Mo(V)16O32)Ni14(PO4)26O2(OH)4(H2O)8]12- wheel.

A new molybdenum nickel phosphate, [H(2)en](3)Na(4)[Ni(H(2)O)(3)][H(30)(Mo(V)(16)O(32))Ni(14)(PO(4))(26)O(2) (OH)(4)(H(2)O)(8)] x 8 H(2)O 1, has been hydrothermally synthesized and structurally characterized. Compound 1 crystallizes in the monoclinic space group P2(1)/c with a = 18.6118(6) A, b = 20.9879(6) A, c = 22.9360(5) A, beta = 116.678(2) degrees, V = 8005.5(4) A(3) and Z = 2. The polyoxoanion of 1 exhibits an unusual divacant wheel-type cluster in which two {NiO(6)} octahedra are lost from the well-known "saturated" {Mo(16)TM(16)P(26)} wheel. The two vacant sites are occupied by two protonated ethylenediamines (H(2)en) via the strong hydrogen-bonding interactions between the surface O atoms of the polyoxoanions and the amine group derived from the H(2)en ligands. DC susceptibility measurements show that compound 1 exhibits strong antiferromagnetic interactions inside the wheel-type cluster.