The Role of Liver Resection for Patients with Liver and Unresectable Lung Colorectal Metastases (LUNA)

Annals of Surgical Oncology -     

Scavenging of hydroxyl radicals but not of peroxynitrite by inhibitors and substrates of nitric oxide synthases

1. The nitric oxide synthase inhibitor N^G-nitro-L-arginine methyl ester (L-NAME) is widely used to study the role of NO^• in physiological and pathological processes, including its role in the generation of the cytotoxic species peroxynitrite (ONOO⁻) and of reactive oxygen radicals such as hydroxyl (OH^•). Often L-NAME is applied to tissues at mM concentrations. At such high concentrations, it might act as a free radical scavenger. A similar possibility might apply to the use of high levels of arginine to study the role of NO^. in atherogenesis.
2. We therefore examined the rate of scavenging of OH^• by L-NAME and found that L-NAME reacts more quickly with OH^. than the established ‘OH^. scavenger'' mannitol and the widely used `OH^• trap'' salicylate. However, D-NAME can scavenge OH^• at rates equal to L-NAME. Both L- and D-arginine were also good OH^• scavengers, comparable in effectiveness to mannitol.
3. Neither L-NAME, D-NAME, L-arginine nor D-arginine was able to inhibit ONOO⁻-dependent nitration of tyrosine, suggesting that they are unlikely to be scavengers of ONOO⁻-derived nitrating species.
4. Neither L-NAME, D-NAME, L-arginine nor D-arginine was able to inhibit the inactivation of α₁-antiproteinase by ONOO⁻, suggesting that they cannot prevent direct oxidations by peroxynitrite.
5. We conclude that L-NAME has sufficient activity as an OH^• scavenger to confound certain pharmacological experiments. However, this explanation of its biological effects can be ruled out if control experiments show that D-NAME has no effect and that L-arginine (also a free radical scavenger) antagonizes the action of L-NAME.

     

Major histocompatibility complex class I-binding peptides are recycled to the cell surface after internalization

Cytotoxic T lymphocytes (CTL) recognize target antigens as short, processed peptides bound to major histocompatibility complex class I (MHC-I) heavy and light chains (β₂-microglobuhn; β₂ m).The heavy chain, which comprise the actual peptide binding α-1 and α-2 domains, can exist at the cell surface in different forms, either free, bound to β₂m or as a ternary complex with β₂m and peptides. MHC-I chains are also known to internalize, and recycle to the cell surface, and this has been suggested to be important in peptide presentation. Whether MHC-I-bound peptides also can recycle is not known. We have investigated this by using both peptide transporter mutant RMA-S cells and EL4 cells loaded with D^b-binding peptides, by two different approaches. First, peptides were covalently linked with galabiose (Galα4Gal) at a position which did not interfere with D^b binding or immunogenicity, and peptide recycling tested with Gal₂-specific monoclonal antibodies. By flow cytometry, a return of Gal₂ epitopes to the cell surface was found, after cellular internalization and cell surface clearance by pronase treatment. This peptide recycling could be discriminated from free fluid-phase uptake and was inhibited by methylamine, chloroquine and low temperature (18°C) but not by leupeptin. Second, specific CTL were reacted with peptide-loaded target cells after complete removal of surface D^b molecules by pronase, and after different times of incubation at 37C to allow reexpression. By this procedure, reappearance of target cell susceptibility was confirmed. The results are in agreement with a model for optimizing peptide presentation by recycling through an intracellular compartment similar to early endosomes in certain antigen-presenting cells.     

Preparation of porous hydroxyapatite scaffolds by combination of the gel-casting and polymer sponge methods

A new technique of combining the gel-casting and polymer sponge methods is introduced in this study to prepare macroporous hydroxyapatite scaffolds, which provides a better control over the microstructures of scaffolds and enhances their mechanical properties. With this technique, we were able to produce scaffolds with mechanical and structural properties that cannot be attained by either the polymer sponge or gel-casting method. The scaffolds prepared have an open, uniform and interconnected porous structure with a pore size of 200-400 microm. A compressive yield strength of approximately 5 MPa equivalent to that of cancellous bone and a compressive modulus of approximately 8 GPa similar to that of cortical bone were achieved. The pore morphology, size, and distribution of the scaffolds were characterized using a scanning electron microscope. X-ray diffraction and Fourier transform infrared spectroscopy were used to determine the crystal structure and chemical composition of scaffolds, respectively. Scaffolds with desired porosity, pore size, and geometry can be prepared by using polymer sponges of appropriate structures.     

The anti‐hepatitis B virus therapeutic potential of anthraquinones derived from Aloe vera

Although the approved hepatitis B virus (HBV)‐polymerase inhibitors (e.g., lamivudine) often lead to drug‐resistance, several natural products have shown promising efficacies. Though Aloe vera (AV) gel and its constituents are shown inhibitors of many viruses, their anti‐HBV activity still remains elusive. We therefore, tested the anti‐HBV potential of AV extract and its anthraquinones in hepatoma cells, including molecular docking, high‐performance thin layer chromatography (HPTLC), and cytochrome P450 (CYP3A4) activation analyses. Our anti‐HBV assays (HBsAg/HBeAg Elisa) showed maximal inhibition of viral antigens production by aloe‐emodin (~83%) > chrysophanol (~62%) > aloin B (~61%) > AV extract (~37%) in HepG2.2.15 cells. Interestingly, the effect of aloe‐emodin was comparable with lamivudine (~86%). Moreover, sequential treatment with lamivudine (pulse) followed by aloe‐emodin (chase) enhanced the efficacy of monotherapy by ~12%. Docking (AutoDock Vina) of the anthraquinones indicated strong interactions with HBV‐polymerase residues that formed stable complexes with high Gibbs's free energy. Further, identification of aloe‐emodin and aloin B by validated HPTLC in AV extract strongly endorsed its anti‐HBV potential. In addition, our luciferase‐reporter gene assay of transfected HepG2 cells showed moderate induction of CYP3A4 by aloe‐emodin. In conclusion, this is the first report on anti‐HBV potential of AV–derived anthraquinones, possibly via HBV‐polymerase inhibition. Of these, although aloin B exhibits novel antiviral effect, aloe‐emodin appears as the most promising anti‐HBV natural drug with CYP3A4 activating property towards its enhanced therapeutic efficacy.