Toll-like receptor 4-dependent adjuvant activity of Kakkon-to extract exists in the high molecular weight polysaccharide fraction.

Kakkon-to, a traditional herbal medicine (Kampo formula), has been used historically in China and Japan for the treatment of infectious diseases such as influenza and the common cold. However, the biological mechanism of its therapeutic action has not yet been elucidated. In this study, we investigated the immunological function of Kakkon-to and found that the high molecular weight fraction of the extract activated macrophages in vitro. This fraction was found to be composed primarily of saccharides and in vitro intensively stimulated mouse peritoneal macrophages that produce Th1 inflammatory cytokines such as tumor necrosis factor α (TNFalpha), interleukin-1beta (IL-1beta), interferon-gamma (IFN-gamma), and interleukin-6 (IL-6). The fraction did not activate macrophages from C3H/HeJ lacking Toll-like receptor 4 (TLR4) or MyD88-deficient mice, indicating that macrophage activation by the fraction was mediated by TLR4. The route of administration of the fraction into mice regulated the kinetics of TNFalpha production in immune organs. Intravenous administration induced TNFalpha production in the four target organs of spleen, liver, lung, and Peyer’s patch; however, the most abundant production occurred in the liver and peaked at 30-60 min post administration. Peritoneal administration induced similar kinetics but the most abundant production occurred in the spleen. In contrast, oral administration induced TNFalpha production in the liver, lung, and Peyer’s patch, but not in the spleen. Although liver and lung are TNFalpha-abundant organs, production peaks in these organs occurred later than in Peyer’s patch. We also found that the fraction induced antibody production as an adjuvant against a specific antigen ovalbumin (OVA) when administered simultaneously and subcutaneously in a dose-dependent manner. Interestingly, the fraction induced IgG-class antibody in response to low doses of the antigen, which induced only IgM-class antibody when administered alone, suggesting that the fraction induces a class switch of immunoglobulin as an adjuvant in vivo. The high molecular weight fraction of Kakkon-to extract could be applicable as a potent immunostimulating drug and adjuvant.

Comparison of Gd-DTPA-induced signal enhancements in rat brain C6 glioma among different pulse sequences in 3-Tesla magnetic resonance imaging.

BACKGROUND: T1-shortening contrast media are routinely used in magnetic resonance (MR) examinations for the diagnosis of brain tumors. Although some studies show a benefit of 3 Tesla (T) compared to 1.5T in delineation of brain tumors using contrast media, it is unclear which pulse sequences are optimal. PURPOSE: To compare gadopentetate dimeglumine (Gd-DTPA)-induced signal enhancements in rat brain C6 glioma in the thalamus region among different pulse sequences in 3T MR imaging. MATERIAL AND METHODS: Five rats with a surgically implanted C6 glioma in their thalamus were examined. T1-weighted brain images of the five rats were acquired before and after Gd-DTPA administration (0.1 mmol/kg) using three clinically available pulse sequences (spin echo [SE], fast SE [FSE], fast spoiled gradient echo [FSPGR]) at 3T. Signal enhancement in the glioma (E(T)) was calculated as the signal intensity after Gd-DTPA administration scaled by that before administration. Pulse sequences were compared using the Tukey-Kramer test. RESULTS: E(T) was 1.12+/-0.05 for FSE, 1.26+/-0.11 for FSPGR, and 1.20+/-0.11 for SE. FSPGR showed significantly higher signal enhancement than FSE and comparable enhancement to SE. CONCLUSION: FSPGR is superior to FSE and comparable to SE in its ability to delineate rat brain C6 glioma in the thalamus region.

Ultraviolet B induces mast cell apoptosis: a hypothetical mechanism of ultraviolet B treatment for uraemic pruritus.

The pathogenesis of uraemic pruritus is unclear, although there is some evidence that an increased number of skin-infiltrating mast cells may play a role. Ultraviolet B reduces itchy sensation of uraemic patients by leading to depletion of cutaneous mast cells. This study presents data that both broad-band and narrow-band ultraviolet B irradiation are able to induce apoptosis in transformed mast cells (murine mastocytoma cell line P815) in a dose-dependent manner at a time point of 24 hours. The positive apoptotic rates were as follows: sham-exposed cells (controls) -- 13.3% +/- 0.6%; with broad-band ultraviolet B irradiation -24.5% +/- 1.1% with 10mJ/cm(2), 57.9% +/- 4.6% with 20mJ/cm(2) and 70.9% +/- 4.5% with 30mJ/cm(2); with narrow-band ultraviolet B irradiation -- 29.6% +/- 2.3% with 100mJ/cm(2), 57.3% +/- 4.1% with 200mJ/cm(2) and 81.5% +/- 1.9% with 300mJ/cm(2). The difference between the number of apoptotic cells in all groups of ultraviolet B-irradiated cells and sham-exposed cells was highly significant (P<0.001). Based on these findings, it is hypothesized that ultraviolet B induced mast cell apoptosis could be an important factor in phototherapy for the diseases dependent on increased number of cutaneous mast cells, including uraemic pruritus.

Enhanced anti-Shigella activity of erythromycin supplemented with sulfadiazine.

Sulfadiazine enhanced the anti-Shigella activity of erythromycin. Erythromycin passes through the type III secretion apparatus and suppresses the growth of invasive Shigella organisms. Sulfadiazine enhanced this effect at the concentration under minimum inhibitory concentration and it came from not only the folate-inhibiting activity but also from a new function. It has proved that sulfadiazine stimulated type III secretion in Shigella as determined from the secretion of the pathogenic protein IpaB. As Congo red induced secretion of Ipa proteins and uptake of erythromycin through the type III secretion gate, sulfadiazine which is similar to Congo red in chemical structure may induce the uptake in the same way.

Oral glucocorticoid-induced fall in cortical bone volume and density in postmenopausal asthmatic patients.

Despite a deepening understanding of the influence of glucocorticoids (GC) on trabecular bone, little is known about GC-induced cortical bone loss. To elucidate the mechanism of GC-induced loss of cortical bone strength with particular reference to cortical bone loss, changes in cortical density, relative cortical volume, and the Strength Strain Index (SSI) based on biomechanical analyses of the geographic distribution of cortical bone material were measured. These parameters were compared, using peripheral quantitative computed tomography (pQCT), among the following age-matched groups: 68 postmenopausal asthmatic patients receiving high-dose oral GC in addition to inhaled GC (oral GC group), 68 postmenopausal asthmatic patients receiving only inhaled GC (inhaled GC group) and 69 postmenopausal controls without asthma or GC therapy (control group). Cortical bone mineral density (BMD) was measured, relative cortical volume was obtained by dividing the cortical area by the total bone area using pQCT (Stratec XCT960), and the Strength Strain Index (SSI) was calculated in the radius based on the density distribution around the axis. Spinal fracture was assessed on lateral radiographs. The number of vertebral fractures per patient correlated highly with cortical BMD, relative cortical volume and SSI values at the radius. The number of vertebral fractures per patient and the number of patients with fracture were similar between the control and inhaled GC group, both being significantly lower than those in the oral GC group. Total BMD, trabecular BMD, cortical BMD, relative cortical volume and SSI were similar between the first two, being significantly higher than in the last group. The slopes of cortical volume-density relationship, however, were identical among the three groups, indicating the persistence of cortical bone remodeling and a similar degree of calcification regardless of GC administration.

Surgical treatment and subsequent outcome of patients with carcinoma of the splenic flexure.

Extended resection, comprising extended right hemicolectomy, splenectomy, and distal pancreatectomy, has been advocated for carcinoma of the splenic flexure because the lymphatic drainage at this site is variable. The present study addresses the problems associated with selecting the most appropriate operative procedure to achieve cure of splenic flexure cancers. We conducted a retrospective review of 27 patients with splenic flexure cancer who underwent curative resection. Left partial colectomy was performed in 20 patients and partial resection of the transverse/descending colon was performed in 7 patients. The combined resection of adjacent organs due to tumor adherence was performed in three patients. The spleen and distal pancreas were the organs most frequently resected among a collective total of six adjacent organs. The median duration of follow-up was 60.9 months after resection for splenic flexure cancer. No patient developed local recurrence. There was no significant difference in 5-year survival between patients with splenic flexure cancers and those with colon cancers at other sites. In conclusion, splenic flexure cancer resected by left partial colectomy or partial resection of the transverse/descending colon without routine extended resection was not associated with a worse prognosis than colon cancers at other sites.

[Efficacy of lidocaine cream in protecting against thermal stress during hyperthermia].

Hyperthermia is performed in combination with chemotherapy as multimodal treatment for recurrent and advanced cancer. It is generally believed that the temperature cannot be raised higher because of thermal stress. In this study, we examined the efficacy of lidocaine cream in protecting against thermal stress during hyperthermia. We devised a new local anesthetic cream containing 5% lidocaine. The subjects were eighteen patients with stomach cancer, liver cancer, or large intestine cancer. This cream was applied locally to the skin with an occlusive dressing for about one hour before hyperthermia was performed, and was wiped away just before hyperthermia. The pain scores in the treatment group were significantly lower than in the no-treatment group (p < 0.05). The scores for sensation of heat in the treatment group were lower, though not to a significant extent, than those in the no-treatment group. No adverse effects were observed. Plasma concentrations of lidocaine were lower than 0.5 microgram/ml, and percutaneous absorption of lidocaine from the lidocaine cream was minimal.

Effects of zinc deficiency/zinc supplementation on ammonia metabolism in patients with decompensated liver cirrhosis.

Hepatic encephalopathy is one of the major complications in decompensated liver cirrhosis. The current study was conducted to clarify the mechanisms of zinc deficiency in liver cirrhosis and its involvement in hepatic encephalopathy via ammonia metabolism. Ten patients each with compensated or decompensated liver cirrhosis and 11 healthy volunteers were enrolled in the study. Serum zinc levels and its daily urinary excretion were measured, an oral zinc-tolerance test was performed to examine zinc malabsorption, and the effects of diuretics on zinc excretion and of zinc supplementation on ammonia metabolism in the skeletal muscle were studied. The mean serum zinc levels in patients with decompensated liver cirrhosis were found to be significantly lower than the levels in controls and patients with compensated liver cirrhosis. The serum zinc levels were inversely correlated with blood ammonia in the fasting state. In the oral zinc-tolerance test, the percent increase in serum zinc levels 120 and 180 min after ingestion was less in cirrhotic patients than in controls. A diuretic administration resulted in a significant reduction in serum zinc levels. An increased uptake of ammonia by and an increased release of glutamine from leg skeletal muscle after oral supplementation of zinc sulfate were evident. Taken together, zinc deficiency in decompensated cirrhotic patients appears to be due to low absorption and to high urinary excretion, for which excessive diuretic administration is, in part, responsible, and zinc supplementation might play an important role in the prevention of hepatic encephalopathy by activating glutamine synthetase.

Ultraviolet A1 (340-400 nm) phototherapy for scleroderma in systemic sclerosis.

BACKGROUND: The presence of an inflammatory infiltrate consisting of helper T cells and a dysregulated matrix metabolism leading to excessive deposition of collagen are two pathogenetic factors responsible for the developments of fibrosis and sclerosis in patients with systemic sclerosis. In previous studies, ultraviolet A1 (UVA1) radiation phototherapy was shown to deplete skin-infiltrating T cells through the induction of T-cell apoptosis and to up-regulate the expression of matrixmetalloproteinase-1 (collagenase-1) in dermal fibroblasts. OBJECTIVE: Our purpose was to determine whether UVA1 phototherapy is effective for systemic sclerosis. METHODS: Lesional skin on the forearms of patients with systemic sclerosis (diffuse type, n =3; limited type, n =1) was exposed to medium-dose UVA1 radiation (60 J/cm(2)) daily. RESULTS: In all patients studied, UVA1 phototherapy-treated skin lesions were markedly softened after 9 to 29 exposures. Clinical improvement was associated with an increase in (1) joint passive range of motion values (P <.05), (2) skin temperature (thermography, P <.05), and (3) cutaneous elasticity (cutaneous elastometry, P <.05). Histologic evaluation of skin specimens obtained before and after UVA1 phototherapy revealed loosening of collagen bundles and the appearance of small collagen fibers. CONCLUSION: These studies indicate that UVA1 phototherapy is effective for patients with systemic sclerosis.

Activation of the dorsal premotor cortex and pre-supplementary motor area of humans during an auditory conditional motor task.

Using functional magnetic resonance imaging (fMRI), we measured regional blood flow to examine which motor areas of the human cerebral cortex are preferentially involved in an auditory conditional motor behavior. As a conditional motor task, randomly selected 330 or 660 Hz tones were presented to the subjects every 1. 0 s. The low and high tones indicated that the subjects should initiate three successive opposition movements by tapping together the right thumb and index finger or the right thumb and little finger, respectively. As a control task, the same subjects were asked to alternate the two opposition movements, in response to randomly selected tones that were presented at the same frequencies. Between the two tasks, MRI images were also scanned in the resting state while the tones were presented in the same way. Comparing the images during each of the two tasks with images during the resting state, it was observed that several frontal motor areas, including the primary motor cortex, dorsal premotor cortex (PMd), supplementary motor area (SMA), and pre-SMA, were activated. However, preferential activation during the conditional motor task was observed only in the PMd and pre-SMA of the subjects' left (contralateral) frontal cortex. The PMd has been thought to play an important role in transforming conditional as well as spatial visual cues into corresponding motor responses, but our results suggest that the PMd along with the pre-SMA are the sites where more general and extensive sensorimotor integration takes place.

A preliminary report of the treatment of blue nevus with dermal injection of riboflavin and exposure to near-ultraviolet/visible radiation (ribophototherapy).

Dye lasers are useful for treating pigmented skin lesions, but their equipment is expensive and bulky. A simple and cheap phototherapy would be acceptable to dermatologists for treating pigmented skin lesions such as nevus of Ota. We investigated as a pilot study whether dermal injection of riboflavin and exposure to near-ultraviolet/visible radiation (ribophototherapy) decreases the dermal pigment of blue nevi which are recalcitrant to laser therapy. The therapeutic efficacy was assessed by comparison of the amount of dermal pigment in hematoxylin-eosin specimens taken before and after treatment. Pigmentation of the nevus became faint to the depth of 1 mm with little noticeable epidermal change after 21 treatments. At the deeper dermis somewhere between 3 and 4 mm from the epidermis, ballooning degeneration of the dermal cells was observed in hematoxylin-eosin specimens. Ribophototherapy is hopeful for treating pigmented skin lesions.

Videoendoscopic laryngeal surgery.

This paper introduces videoendoscope-assisted laryngeal surgery with office-based equipment. With this technique, a patient is seated and the nose, pharynx, and larynx are topically anesthetized. A flexible videoendoscope with a light-sensitive charge-coupled device chip built into the tip is transnasally inserted by an assistant. Specially designed fine-tipped forceps and scalpels were developed for removal of laryngeal lesions. Videoendoscopic laryngeal surgery was undertaken in 114 cases of laryngeal lesions such as polyps, granuloma, and cancer. For benign vocal fold lesions, postoperative vocal function was shown to be improved on aerodynamic and perceptual analyses. For laryngeal tumors, biopsy of the lesion was easily undertaken. Videoendoscopic laryngeal surgery presents the following advantages. It is applicable to outpatients not requiring general anesthesia, it enables functional monitoring of the patient's voice and vocal fold during phonation, it allows for delicate manipulations with both hands, and it gives high-resolution images in comparison to conventional fiberscopy.

Ultraviolet A1 (340-400 nm) phototherapy for cutaneous T-cell lymphoma.

BACKGROUND: The results of a recent study suggested that ultraviolet A1 radiation (UVA1R; 340-400 nm) phototherapy for atopic dermatitis works through induction of apoptosis in skin-infiltrating helper T cells, indicating the possibility that other helper T cell-mediated skin diseases may respond to UVA1R as well. OBJECTIVE: The purpose of this open pilot study was to assess the therapeutic effectiveness of UVA1 phototherapy for cutaneous T-cell lymphoma (CTCL). METHODS: UVA1 phototherapy was used as monotherapy in patients (n = 3) with histologically proven CTCL (stages IA and IB). For daily whole body UVA1 irradiations, either a high-dose (n = 2; 130 J/cm2 UVA1 per exposure) or medium-dose (n = 1; 60 J/cm2 UVA1) regimen was used. Therapeutic effectiveness was assessed clinically and histologically. RESULTS: In each of the 3 patients, skin lesions began to resolve after only a few UVA1 radiation exposures. Complete clearance was observed between 16 and 20 exposures, regardless of whether the high- or medium-dose regimen had been employed. CONCLUSION: These studies suggest that patients with CTCL stages IA and IB can be treated effectively with UVA1 phototherapy.

Molecular cloning and expression of the gene encoding a phospholipase A1 from Aspergillus oryzae.

Phospholipase A1 (PLA1) is a hydrolytic enzyme that catalyzes removal of the acyl group from position 1 of lecithin to form lysolecithin. The genomic DNA and cDNA encoding PLA1 from Aspergillus oryzae were cloned with the mixed deoxyribonucleotide-primed polymerase chain reaction. The PLA1 gene is composed of 1,056 bp and has four exons and three short introns (63, 54, and 51 bp). The deduced amino acid sequence of PLA1 contained the N-terminal sequence of the mature PLA1 analyzed by Edman degradation. PLA1 cDNA has an open reading frame of 885 bp encoding the PLA1 precursor of 295 amino acid residues. The mature PLA1 is composed of 269 amino acid residues, and a prepro-sequence of 26 amino acid residues is at the N-terminal region of the PLA1 precursor. PLA1 has two possible N-glycosylation sites (Asn27 and Asn55). PLA1 has a consensus pentapeptide (-Gly-His-Ser-Xaa-Gly-), which is conserved in lipases. The amino acid sequence of PLA1 showed 47% identity with that of mono- and diacylglycerol lipase from Penicillium camembertii. The PLA1 cDNA was expressed in Saccharomyces cerevisiae KS58-2D, indicating the cloned gene to be functional.

(2E,6R)-8-hydroxy-2,6-dimethyl-2-octenoic acid, a novel anti-osteoporotic monoterpene, isolated from Cistanche salsa.

(2E,6R)-8-Hydroxy-2,6-dimethyl-2-octenoic acid [(R)-HDOA], a novel monoterpene from Cistanche salsa, a Chinese herb, was found to be an anti-osteoporotic compound. The extract of Cistanche salsa significantly suppressed the bone weight loss in ovariectomized mice, a postmenopausal osteoporosis model. The active substance was then purified by using this osteoporotic model and the chemical structure was determined. The active compound from Cistanche salsa, (R)-HDOA, suppressed the decrease of bone weight and the mechanical strength in the ovariectomized mice. Furthermore, (R)- and (S)-HDOA were synthesized and the activity of each was evaluated. (R)-HDOA suppressed the bone weight loss, although (S)-HDOA did not showed any activity.

Synthesis and antiviral activity of novel acyclic nucleosides: discovery of a cyclopropyl nucleoside with potent inhibitory activity against herpesviruses.

A series of acyclic nucleosides with two hydroxymethyl groups mimicking the 3'- and 5'-hydroxyl groups of the 2'-deoxyribose moiety were prepared and evaluated for their antiherpetic activity. Among those, 9-[[cis-1', 2'-bis(hydroxymethyl)cycloprop-1'-yl]methyl]guanine (3) showed extremely potent antiviral activity against herpes simplex virus type-1 (HSV-1) with good selectivity. Both enantiomers of 3 were synthesized starting from chiral epichlorohydrins, and only one of the enantiomers with 1'S,2'R-configuration (3a) exhibited strong antiherpetic activity (IC50 of 0.020 microg/mL against HSV-1 Tomioka vs 0.81 microg/mL for acyclovir). Enantiomer 3a was also more inhibitory than acyclovir against varicella-zoster virus (VZV) but ineffective against human immunodeficiency virus (HIV). Compound 3a is phosphorylated by HSV-1 thymidine kinase (TK) very efficiently. The relationship between conformation and antiherpetic activity in this series of compounds is discussed.

Relationship between a low toxicity of the mutant A subunit of enterotoxigenic Escherichia coli enterotoxin and its strong adjuvant action.

In the work described here it was determined if and how unnicking in the A subunit of Escherichia coli enterotoxin at Arg192 or nearby residues affected biological activities of the toxin. The mutant toxin was constructed to lack the nick site in the A subunit by deleting the tripeptide Arg192-Thr193-Ile194, which is essential for toxicity. The mutant toxin did not exhibit agmatine ADP-ribosyltransferase activity in the presence or absence of the ADP-ribosylation factor and had less diarrhoeal activity and lower induction of cyclic AMP than did LT. The mutant toxin exhibited a much stronger adjuvant action on antibody responses to measles virus, keyhole limpt haemocyanin, bovine immunoglobulin and ovalbumin compared with LT. The altered toxicity of the mutant toxin might be closely related to the potent adjuvant action on antibody responses to antigens. The relationship between two activities is discussed.

2B1 antigen characteristically expressed on extracellular matrices of human malignant tumors is a large chondroitin sulfate proteoglycan, PG-M/versican.

2B1 is a monoclonal antibody against a large proteoglycan isolated from human yolk sac tumor (M. Sobue et al., Histochem. J., 21: 455-460, 1989). The antigen is expressed in a variety of embryonal tissues as well as most if not all malignant tumor tissues. However, the expression in normal adult tissues is limited to some tissues, such as the smooth muscle layers of the aorta. We characterized the 2B1 antigen isolated from the conditioned medium of human malignant fibrous histiocytoma and found that immunological and biochemical properties are identical to those of a large chondroitin sulfate proteoglycan, PG-M/versican. Partial amino acid sequences of peptides obtained from the core protein by V8 protease digestion and subsequent SDS-PAGE were detected in the reported amino acid sequence of human PG-M/versican with a complete identity. Furthermore, 2B1 was distinctly reactive to the expressed protein by transfection of the cDNA for the shortest form into mouse cells. The results indicate that the antigen is the PG-M core protein, and the epitope may be in one of the globular domains. It is thus likely that PG-M/versican is one of the extracellular matrix components characteristic of human malignant tumors.

Identification of two isoforms of mouse neuropeptide Y-Y1 receptor generated by alternative splicing. Isolation, genomic structure, and functional expression of the receptors.

Two cDNA clones homologous with human neuropeptide (NP) Y-Y1 receptor have been isolated from a mouse bone marrow cDNA library. One was thought to be the cognate of the human NPY-Y1 receptor, termed Y1 alpha receptor, and the other form, termed Y1 beta receptor, differed from the Y1 alpha receptor in the seventh transmembrane domain and C-terminal tail. Analysis of the mouse genomic DNA showed that both receptors originated from a single gene. The different peptide sequences of the Y1 beta receptor were encoded by separate exons, hence, these receptors were generated by differential RNA splicing. High affinity binding of [125I]NPY to each receptor expressed in Chinese hamster ovary (CHO) cells and sequestration of [125I]NPY after binding to each receptor were observed. In the CHO cells expressing the Y1 alpha receptor, intracellular Ca2+ increase, inhibition of forskolin-induced cAMP accumulation, and mitogen-activated protein kinase (MAPK) activation were observed by stimulation of NPY, and these responses were abolished by pretreatment with pertussis toxin. Since wortmannin completely inhibited NPY-elicited MAPK activation, we speculate that wortmannin-sensitive signaling molecule(s) such as phosphoinositide 3-kinase may lie between pertussis toxin-sensitive G-protein and MAPK. In contrast, these intracellular signals were not detected in CHO cells expressing the Y1 beta receptor. Northern blots and reverse transcriptase-polymerase chain reaction analyses indicated that the Y1 alpha receptor was highly expressed in the brain, heart, kidney, spleen, skeletal muscle, and lung, whereas the Y1 beta receptor mRNA was not detected in these tissues. However, the Y1 beta receptor was expressed in mouse embryonic developmental stage (7 and 11 days), bone marrow cells and several hematopoietic cell lines. These results suggest that the Y1 beta receptor is an embryonic and a bone marrow form of the NPY-Y1 receptor, which decreases in the expression during development and differentiation.

Successful treatment of systemic sclerosis with topical PUVA.

Treatment of a 43-year-old woman with systemic sclerosis with 8-methoxypsoralen and ultraviolet. A radiation (PUVA) resulted in the significant clinical and histological improvement of skin lesions without side effects.