Sialylated endogenous glycoconjugates in plant cells

Bioengineered plants are emerging as promising systems for the production of therapeutically valuable proteins. It has been commonly accepted that plants do not perform mammalian-like post-translational modifications, particularly sialylation of glycoconjugates, and no evidence has previously been reported to suggest that they have such capabilities. Here we report the presence of sialylated glycoconjugates in suspension-cultured cells of Arabidopsis thaliana and suggest that a genetic and enzymatic basis for sialylation exists in plants.     

Structure-based design of specific cathepsin inhibitors and their application to protection of bone metastases of cancer cells.

We report the antihypercalcemic and antimetastatic effects of CLIK-148 in vivo, which is a specific inhibitor of cathepsin L. The decalcification during bone absorption is followed by the degradation of type-1 collagen by osteoclastic cathepsins. Tumor-bearing osteoclasts or TNF-alpha-activated osteoclasts secrete large amounts of cysteine proteases, especially procathepsin L, which powerfully degrade type-1 collagen leading to tumor-associated bone absorption and release of bone calcium. The bone pit formations in vitro, which are caused by osteoclasts derived from human bone marrow cells activated by RANKL and M-CSF and also by mice osteoclasts activated by TNF-alpha, are significantly prevented by CLIK-148 treatment. We evaluated the in vivo inhibitory effect of malignant hypercalcemia induced by LJC-1 human mandibular cancer inoculation by CLIK-148 treatment, and the CLIK-148 treatment significantly protected against the tumor-induced hypercalcemia. On the protection of bone metastasis of colon 26 PMF-15 implanted to mouse calvaria, CLIK-148 treatment significantly inhibited calvaria bone absorption (direct metastasis). The CLIK-148 treatment also reduced distant bone metastasis to the femur and tibia of melanoma A375 tumors implanted into the left ventricle of the heart.     

Conspecific thistle plant selection by a herbivorous ladybird beetle, Epilachna pustulosa

Host selection by Epilachna pustulosa Kôno (Coleoptera: Coccinellidae) was surveyed in an area of about 130 ares, focusing on the role of the spatial distribution pattern of the host plant, thistle Cirsium kamtschaticum Ledeb. (Asteraceae) and the environmental conditions of habitats where thistle plants were growing. A total of 198 thistle clones were found in the area studied, and approximately 40% showed some degree of E. pustulosa infestation by July. Eggs were only oviposited on thistle clones that were fed on by adults. Adult beetles and egg masses of E. pustulosa showed an aggregated distribution in the earlier season (June) among thistle clones. The distribution of adults became more random (but still aggregated) by the later season (July), along with an increase in the number of infested clones. Multiple logistic regression analyses revealed that clone size and soil moisture were consistently important for the beetle's choice of clones to feed on. The other logistic regression analyses indicated that thistle‐clone size and sunlight condition influenced egg distribution. Thistle clone selection by E. pustulosa changed with season from a rather strict selection in June to a more obscure one in July, expanding the range of thistle clones used as feeding and oviposition substrate.     

Novel Genetic Variants of Anaplasma phagocytophilum, Anaplasma bovis, Anaplasma centrale, and a Novel Ehrlichia sp. in Wild Deer and Ticks on Two Major Islands in Japan

Wild deer are one of the important natural reservoir hosts of several species of Ehrlichia and Anaplasma that cause human ehrlichiosis or anaplasmosis in the United States and Europe. The primary aim of the present study was to determine whether and what species of Ehrlichia and Anaplasma naturally infect deer in Japan. Blood samples obtained from wild deer on two major Japanese islands, Hokkaido and Honshu, were tested for the presence of Ehrlichia and Anaplasma by PCR assays and sequencing of the 16S rRNA genes, major outer membrane protein p44 genes, and groESL. DNA representing four species and two genera of Ehrlichia and Anaplasma was identified in 33 of 126 wild deer (26%). DNA sequence analysis revealed novel strains of Anaplasma phagocytophilum, a novel Ehrlichia sp., Anaplasma centrale, and Anaplasma bovis in the blood samples from deer. None of these have been found previously in deer. The new Ehrlichia sp., A. bovis, and A. centrale were also detected in Hemaphysalis longicornis ticks from Honshu Island. These results suggest that enzootic cycles of Ehrlichia and Anaplasma species distinct from those found in the United States or Europe have been established in wild deer and ticks in Japan.     

Radiosensitization of hypoxic HeLa S3 cells in vitro by a new type of radiosensitizer: spermine and spermidine amides with nitro groups

A new type of hypoxic cell sensitizer (FNT-series compounds) has been developed and tested on HeLa S3 cells. They have two nitrobenzoyl groups at both ends of the spermine or spermidine in their chemical structures. Their ability to sensitize hypoxic cells is greater than that of misonidazole. Among them, N1, N10-bis(4-nitrobenzoyl)-spermidine (FNT-1) was most effective. 1 mM FNT-1 gave a corrected enhancement ratio of 1.71 compared to 1.32 for the same concentration of misonidazole. Its electron affinity, in terms of the half-wave reduction potential, was - 350 mV and higher than that of misonidazole (-395 mV). These FNT-series compounds are thought to interact with DNA in two ways; noncovalent linkage between the basic groups of the polyamine and highly acidic phosphate moieties of the nucleic acid and, secondly, the insertion of the nitrobenzoyl groups between base pairs of the DNA double helix. Since spermine and spermidine themselves did not show any sensitizing activities, it is suggested that the nitrobenzoyl groups which are introduced in spermine and spermidine, play an important role in sensitization.     

Solubility of phosphatidylcholine in chloroform. Formation of hydrogen bonding between phosphatidylcholine and chloroform

The solubility of phosphatidylcholine (PC) was studied by the spectroscopic analysis and the measurement of the solubility. The qualitative analysis of infrared absorptionspectra confirmed the existence of two types of hydrogen bondings between chloroform and PC, one between chloroform and the CO group of PC and the other between chloroform and the phosphorylcholine group of PC. The quantitative analysiss of the C6 stretching vibration bands of the chloroform-d solution of PC showed that the latter hydrogen bonding mainly contributes to the solubility and that PC dissolves in chloroform to form a complex consisting of a few or more molecules of chloroform and one molecule of PC. We discussed in this report about the molecular organization of PC in chloroform solution.     

Synthesis and evaluation of 6-methylene-bridged uracil derivatives. Part 1: discovery of novel orally active inhibitors of human thymidine phosphorylase

A series of novel 6-methylene-bridged uracil derivatives have been prepared as inhibitors of human thymidine phosphorylase (TP). To enhance the in vivo antitumor activity of fluorinated pyrimidine 2'-deoxyribonucleosides such as 2'-deoxy-5-(trifluoromethyl)uridine (F(3)dThd), a potent TP inhibitor preventing their degradation to an inactive compound, has become a target of medicinal chemistry. We present here the synthesis and evaluation of novel human TP inhibitors. Introduction of an N-substituted aminomethyl side chain at the 6-position of 5-chlorouracil has improved water solubility and enhanced inhibitory activity compared with the known TP inhibitor, 6-amino-5-chlorouracil. Compound 42 was reasonably well absorbed in mice after oral administration. When combined with F(3)dThd, compound 42 exerted its TP inhibitory potency by increasing the maximum plasma concentrations of the former as evidenced in experiments with monkeys. Positive changes in pharmacokinetic profile were accompanied by the enhanced in vivo antitumor activity of this combination when compared to F(3)dThd alone, in mice bearing human tumor xenografts. Both biochemical and pharmacological effects appeared to fit the concept as anticipated.     

Demonstration of functional role of TECK/CCL25 in T lymphocyte-endothelium interaction in inflamed and uninflamed intestinal mucosa

It has recently been suggested that C-C chemokines may play a role in the organ-specific homing of lymphocytes, but there is not enough in vivo evidence in intestinal mucosa. The aim of this study was to examine whether thymus-expressed chemokine (TECK)/CCL25 and its ligand CCR9 are involved in T-lymphocyte interaction with microvessels of murine intestinal mucosa. T lymphocytes from the small intestine were fluorescence labeled, and their adhesion to mucosal microvessels was observed by intravital microscopy. Lamina proprial lymphocytes (LPL) and intraepithelial lymphocytes (IEL) adhered to both the small intestine and colon, and desensitization of CCR9 with TECK/CCL25 or anti-TECK/CCL25 antibody significantly inhibited these adhesions only in small intestine. At both sites, TNF-alpha significantly increased LPL adhesion but not IEL adhesion. Desensitization of CCR9 or anti-TECK/CCL25 antibody also attenuated the TNF-alpha-induced LPL adhesion in the small intestine. Increased expression of TECK/CCL25 by TNF-alpha was observed in the lamina propria of small intestine. TECK/CCL25 may thus play an important role in the adherence of mucosal lymphocytes to the microvessels of the small intestine but not the colon under uninflamed as well as inflamed conditions.