Crystal structure of human factor VIIa/tissue factor in complex with peptide mimetic inhibitor

The 3D structure of human factor VIIa/soluble tissue factor in complex with a peptide mimetic inhibitor, propylsulfonamide-D-Thr-Met-p-aminobenzamidine, is determined by X-ray crystallography. As compared with the interactions between thrombin and thrombin inhibitors, the interactions at S2 and S3 sites characteristic of factor VIIa and factor VIIa inhibitors are revealed. The S2 site has a small pocket, which is filled by the hydrophobic methionine side chain in P2. The small S3 site fits the small size residue, D-threonine in P3. The structural data and SAR data of the peptide mimetic inhibitor show that these interactions in the S2 and S3 sites play an important role for the improvement of selectivity versus thrombin. The results will provide valuable information for the structure-based drug design of specific inhibitors for FVIIa/TF.     

A dynamic neural network with temporal coding and functional connectivity

A neural network model capable of altering its pattern classifying properties by program input is proposed. Here the “program input” is another source of input besides the pattern input. Unlike most neural network models, this model runs as a deterministic point process of spikes in continuous time; connections among neurons have finite delays, which are set randomly according to a normal distribution. Furthermore, this model utilizes functional connectivity which is dynamic connectivity among neurons peculiar to temporal-coding neural networks with short neuronal decay time constants. Computer simulation of the proposed network has been performed, and the results are considered in light of experimental results shown recently for correlated firings of neurons. Received: 6 December 1996 / Accepted in revised form: 15 September 1997     

Accumulation and degradation of thiamin-binding protein and level of thiamin in wheat seeds during seed maturation and germination

Changes in the levels of thiamin-binding globulin and thiamin in wheat seeds during maturation and germination were studied. The thiamin-binding activity of the seed proteins increased with seed development after flowering. The thiamin content of the seeds also increased with development. Thiamin-binding activity decreased during seed germination. On the other hand, immunological analysis using an antibody directed against the thiamin-binding protein isolated from wheat seeds showed that the thiamin-binding globulin accumulated in the aleurone layer of the seeds during maturation, and then the protein was degraded and disappeared during seed germination. These results suggested that the thiamin-binding globulin of wheat seeds was synthesized and accumulated in the aleurone layer of the seeds with seed development, similar to the thiamin-binding albumin in sesame seeds, and that thiamin bound to the thiamin-binding globulin in the dormant wheat seeds for germ growth during germination.     

Protective roles of redox-active protein thioredoxin-1 for severe acute pancreatitis

Severe acute pancreatitis is a disease with high mortality, and infiltration of inflammatory cells and reactive oxygen species have a crucial role in the pathophysiology of this disease. Thioredoxin-1 (TRX-1) is an endogenous redox-active multifunctional protein with antioxidant and anti-inflammatory effects. TRX-1 is induced in various inflammatory conditions and shows cytoprotective effects. The aim of the present study was to clarify the protective roles of TRX-1 in the host defense mechanism against severe acute pancreatitis. Experimental acute pancreatitis was induced by intraperitoneal administration of cerulein, a CCK analog, and aggravated by lipopolysaccharide injection in transgenic mice overexpressing human TRX-1 (hTRX-1) and control C57BL/6 mice. Transgenic overexpression of hTRX-1 strikingly attenuated the severity of experimental acute pancreatitis. TRX-1 overexpression suppressed neutrophil infiltration as determined by myeloperoxidase activity, oxidative stress as determined by malondialdehyde concentration, and cytoplasmic degradation of inhibitor of kappaB-alpha, thereby suppressing proinflammatory cytokines, tumor necrosis factor-alpha, interleukin-1beta, and interleukin-6; a neutrophil chemoattractant, keratinocyte-derived chemokine; and inducible nitric oxide synthase in the pancreas. Administration of recombinant hTRX-1 also suppressed neutrophil infiltration, reduced the inflammation of the pancreas and the lung, and improved the mortality rate. The present study suggests that TRX-1 has potent antioxidant and anti-inflammatory actions in experimental acute pancreatitis and might be a new therapeutic strategy to improve the prognosis of severe acute pancreatitis.     

Immunohistochemical staining patterns of MUC1, MUC2, MUC4, and MUC5AC mucins in hyperplastic polyps, serrated adenomas, and traditional adenomas of the colorectum.

We studied the distribution of the four human apomucins MUC1, MUC2, MUC4, and MUC5AC in hyperplastic polyps, serrated adenomas, and traditional adenomas of the colorectum using immunohistochemical techniques, with the aim of comparing and contrasting their patterns of expression. A series of 12 hyperplastic polyps, 27 serrated adenomas, and 20 traditional adenomas was studied. No significant change in apomucin expression was observed in traditional adenomas compared with normal colorectal epithelium, except for MUC5AC, which was present in 12 of the adenomas (60%) and only 20% of the normal samples. In both hyperplastic polyps and serrated adenomas, MUC2 and MUC5AC mucin expression was consistently and markedly increased. In 50% of the hyperplastic polyps, MUC4 was reduced but in the remaining cases was similar to normal. Loss of MUC4 expression was observed in all serrated adenomas. MUC1 was not increased in the hyperplastic polyps but increased expression was seen in 17 of the serrated adenomas (63%). Similar altered distribution patterns of MUC2, MUC4, and MUC5AC were seen in hyperplastic polyps and serrated adenomas, whereas traditional adenomas showed little change from normal patterns of expression. Although hyperplastic polyps are commonly defined as benign lesions without neoplastic potential, the similar phenotypes of hyperplastic and serrated adenomas and the existence of mixed polyps suggest that these lesions may represent a histogenetic continuum.     

The RSC chromatin remodelling enzyme has a unique role in directing the accurate positioning of nucleosomes

Nucleosomes impede access to DNA. Therefore, nucleosome positioning is fundamental to genome regulation. Nevertheless, the molecular nucleosome positioning mechanisms are poorly understood. This is partly because in vitro reconstitution of in vivo-like nucleosome positions from purified components is mostly lacking, barring biochemical studies. Using a yeast extract in vitro reconstitution system that generates in vivo-like nucleosome patterns at S. cerevisiae loci, we find that the RSC chromatin remodelling enzyme is necessary for nucleosome positioning. This was previously suggested by genome-wide in vivo studies and is confirmed here in vivo for individual loci. Beyond the limitations of conditional mutants, we show biochemically that RSC functions directly, can be sufficient, but mostly relies on other factors to properly position nucleosomes. Strikingly, RSC could not be replaced by either the closely related SWI/SNF or the Isw2 remodelling enzyme. Thus, we pinpoint that nucleosome positioning specifically depends on the unique properties of the RSC complex.     

Influence of segmental and extra-segmental conditioning stimuli on cortical potentials evoked by painful electrical stimulation

This study evaluated the effects of two different types of segmental/extra-segmental conditioning stimuli (tonic muscle pain and non-painful vibration) on the subjective experience (perceived pain intensity) and on the cortical evoked potentials to standardized test stimuli (cutaneous electrical stimuli). Twelve subjects participated in two separate sessions to investigate the effects of tonic muscle pain or cutaneous vibration on experimental test stimuli. The experimental protocol contained a baseline registration (test stimuli only), a registration with the test stimuli in combination with the conditioning stimuli, followed by a registration with the test stimuli only. In addition, the effects of the conditioning stimuli were examined at two anatomically separated locations (segmental and extra-segmental). Compared with the test stimulus alone, the perceived pain intensity and peak-to-peak amplitudes of the evoked potentials were unchanged in the presence of non-painful conditioning stimuli at either location. In contrast, a significant decrease of the perceived pain intensity and peak-to-peak amplitudes was found in the presence of painful conditioning stimuli at the extra-segmental sites. Moreover, the topographic maps of the 32-channel recordings suggested that the distribution of the scalp evoked potentials was almost symmetrical around the vertex Cz in the baseline registration. The evoked potentials were generally decreased during hypertonic saline infusion at the extra-segmental sites, but the distribution of the topographic maps did not appear to change. Vibration has previously been shown to inhibit pain, but in the present study the perceived intensity of phasic painful electrical stimuli was unchanged. The reduced perceived pain intensity and the smaller peak-to-peak amplitude of the evoked potential in the presence of extra-segmental conditioning pain are in accordance with the concept of diffuse noxious inhibitory control.     

Identification of a collagen production-promoting factor from an extract of royal jelly and its possible mechanism

We have previously shown that royal jelly (RJ) promoted collagen production by skin fibroblasts in the presence of ascorbic acid-2-O-alpha-glucoside (AA-2G). In this study, we purified the honeybee RJ-derived collagen production-promoting factor (HBRJ-CPF) from an alkali-solubilized fraction of RJ by C18 reverse-phase column chromatography. The elution profile by the C18 column chromatography and the molecular mass of the purified HBRJ-CPF material coincided with those of 10-hydroxy-2-decenoic acid (10H2DA). We then examined the collagen production-promoting activities of several commercially available fatty acids contained in RJ. We found that 10H2DA and 10-hydroxydecanoic acid increased the collagen production in a dose-dependent manner. Furthermore, 10H2DA induced the fibroblast cell line, NHDF, to produce transforming growth factor-beta 1 (TGF-beta 1) which is an important factor for collagen production. As expected, the collagen production-promoting activity of 10H2DA was neutralized by the anti-TGF-beta 1 antibody. These result suggest that HBRJ-CPF identified as 10H2DA promoted the collagen production of AA-2G-treated fibroblasts by inducing TGF-beta 1 production.