Temperature-dependent higher order structures of the (Pro-Pro-Gly)₁₀-modified dendrimer

Collagen is the most abundant protein in mammals and is widely used as a biomaterial for tissue engineering and drug delivery. We previously reported that dendrimers and linear polymers, modified with collagen model peptides (Pro‐Pro‐Gly)₅, form a collagen‐like triple‐helical structure; however, its triple helicity needs improvement. In this study, a collagen‐mimic dendrimer modified with the longer collagen model peptides, (Pro‐Pro‐Gly)₁₀, was synthesized and named PPG10‐den. Circular dichroism analysis shows that the efficiency of the triple helix formation in PPG10‐den was much improved over the original. The X‐ray diffraction analysis suggests that the higher order structure was similar to the collagen triple helix. The thermal stability of the triple helix in PPG10‐den was higher than in the PPG10 peptide itself and our previous collagen‐mimic polymers using (Pro‐Pro‐Gly)₅. Interestingly, PPG10‐den also assembled at low temperatures. Self‐assembled structures with spherical and rod‐like shapes were observed by transmission electron microscopy. Furthermore, a hydrogel of PPG10‐den was successfully prepared which exhibited the sol‐gel transition around 45°C. Therefore, the collagen‐mimic dendrimer is a potential temperature‐dependent biomaterial. © 2010 Wiley Periodicals, Inc. Biopolymers 95: 270–277, 2011.     

Interspecies Transmission of Feline Immunodeficiency Virus from the Domestic Cat to the Tsushima Cat (Felis bengalensis euptilura) in the Wild

Feline immunodeficiency virus (FIV) was isolated from a wild-caught Tsushima cat (Felis bengalensis euptilura), an endangered Japanese nondomestic subspecies of leopard cat (F. bengalensis). Phylogenetic analysis of the env gene sequences indicated that the FIV from the Tsushima cat belonged to a cluster of subtype D FIVs from domestic cats. FIVs from both the Tsushima cat and the domestic cat showed similar levels of replication and cytopathicity in lymphoid cell lines derived from these two species. The results indicated the occurrence of interspecies transmission of FIV from the domestic cat to the Tsushima cat in the wild.     

Advanced glycation end product-induced apoptosis and overexpression of vascular endothelial growth factor and monocyte chemoattractant protein-1 in human-cultured mesangial cells

Advanced glycation end products (AGE) have been implicated in the pathogenesis of glomerulosclerosis in diabetes. However, their involvement in the development of the early phase of diabetic nephropathy has not been fully elucidated. We investigated the effects of AGE on growth and on vascular endothelial growth factor (VEGF) and monocyte chemoattractant protein-1 (MCP-1) expression in human cultured mesangial cells. We prepared three immunochemically distinct AGE by incubating bovine serum albumin (BSA) with glucose, glyceraldehyde, or glycolaldehyde. When human mesangial cells were cultured with various types of AGE-BSA, viable cell numbers as well as DNA syntheses were significantly decreased. All of the AGE-BSA were found to significantly increase p53 and Bax protein accumulations and subsequently induce apoptotic cell death in mesangial cells. An antioxidant, N-acetylcysteine, significantly prevented the AGE-induced apoptotic cell death in mesangial cells. Human mesangial cells stimulated prostacyclin production by co-cultured glomerular endothelial cells. Furthermore, various types of AGE-BSA were found to up-regulate the levels of mRNAs for VEGF and stimulate the secretion of VEGF and MCP-1 proteins in mesangial cells. The results suggest that AGE disturbed glomerular homeostasis by inducing apoptotic cell death in mesangial cells and elicited hyperfiltration and microalbuminuria by stimulating the secretion of VEGF and MCP-1 proteins, thereby being involved in the pathogenesis of the early phase of diabetic nephropathy.     

Authors’ reply

Article: http://dx.doi.org/10.1002/bimj.201100221 Letter to the Editor: http://dx.doi.org/10.1002/bimj.201200254     

Baculovirus activates murine dendritic cells and induces non-specific NK cell and T cell immune responses

We previously reported that the baculovirus induced a strong host immune response against infections and malignancies. Among the immune cells, the dendritic cells were most strongly infected and activated by the baculovirus, although the exact mechanism remained unclear. Here, we evaluated the non-specific immune responses of bone marrow-derived dendritic cells (BMDCs) after infection by a wild-type baculovirus. MHC class I and II molecules and co-stimulation molecules (CD40, CD80, and CD86) on BMDCs were up-regulated by baculovirus infection. At the same time, the BMDCs produced pre-inflammatory cytokines (IL-6, IL12p70, and TNF-α) and IFN-α. NK cells showed IFN-γ production, CD69 up-regulation, and enhanced cytotoxicity when they were co-cultured with baculovirus-infected BMDCs. T cells showed IFN-γ production, CD69 up-regulation, and cell proliferation. Ex vivo analysis performed in vitro produced similar results. These findings suggested that baculovirus-infected dendritic cells induce non-specific immune responses and can be used as an immunotherapeutic agent against viral infections and malignancies, together with present therapeutic drug regimens.     

Heat stress is a potent stimulus for enhancing rescue efficiency of recombinant Borna disease virus

Recently developed vector systems based on Borna disease virus (BDV) hold promise as platforms for efficient and stable gene delivery to the central nervous system (CNS). However, because it currently takes several weeks to rescue recombinant BDV (rBDV), an improved rescue procedure would enhance the utility of this system. Heat stress reportedly enhances the rescue efficiency of other recombinant viruses. Here, heat stress was demonstrated to increase the amount of BDV genome in persistently BDV‐infected cells without obvious cytotoxicity. Further analyses suggested that the effect of heat stress on BDV infection is not caused by an increase in the activity of BDV polymerase. More cells in which BDV replication occurs were obtained in the initial phase of rBDV rescue by using heat stress than when it was not used. Thus, heat stress is a useful improvement on the published rescue procedure for rBDV. The present findings may accelerate the practical use of BDV vector systems in basic science and the clinic and thus enable broader adoption of this viral vector, which is uniquely suited for gene delivery to the CNS.     

Acid triacyglycerol lipase inhibitor in chicken plasma: purification and properties

1. Acid triacylglycerol lipase inhibitor was highly purified from chicken plasma by ammonium sulfate fractionation (0.6-1 saturation) followed by successive chromatographies on Hydroxyapatite, Blue-Cellulofine, Phenyl-Sepharose and Cellulofine GCL-2000 columns, and isoelectric focusing. 2. The lipase inhibitor showed its inhibitory action on triacylglycerol lipases in chicken erythrocytes ghosts and in chicken liver lysosome, but did not on pancreatic lipase, Rhizopus arrhizus lipase, or wheat germ lipase. 3. The inhibitor showed its molecular weight of 32,000 by Cellulofine GCL-2000 gel filtration. The inhibitor showed some heterogeneity on isoelectric focusing, and the main band had a pI of 5.10. 4. The lipase inhibitor did not show any inhibitory action on trypsin or chymotrypsin.     

Highly sensitive chemiluminescence method for determining myeloperoxidase in human polymorphonuclear leukocytes.

Myeloperoxidase activity was assayed by a chemiluminescence method, using a cypridina luciferin analog as a chemiluminescence probe, after extraction from peripheral human polymorphonuclear leukocytes. The chemiluminescence method was based on the detection of 1O2 generated by myeloperoxidase-catalyzed HOBr formation followed by the interaction of HOBr with H2O2 at pH 4.5. With this method, myeloperoxidase in less than 100 polymorphonuclear leukocytes could be detected and myeloperoxidase in 10(6) polymorphonuclear leukocytes would be calculated to be 14.4 pmol. Eosinophil extract, which contains eosinophil peroxidase, catalyzed 1O2 generation to a great extent, compared with the polymorphonuclear leukocyte extract at pH 4.5. Myeloperoxidase activity in extract of neutrophil fraction could be greatly influenced by eosinophil contamination.