Proanthocyanidin oligomers isolated from Salacia reticulata leaves potently inhibit pancreatic lipase activity.

UNLABELLED: The pancreatic lipase inhibitory (PLI) activity of leaf extracts (aqueous, 60 and 99.8 (v/v)% EtOH) of Salacia reticulata Wight, referred to "Kothala himbutu" (KT) in Singhalese, was compared with that of KT stem extracts. Evaporated residue contents and PLI activity of each leaf extract were higher than those of each stem extract, respectively. Among the extracts, the 60% EtOH leaf extract showed the most potent PLI activity. The 60% EtOH leaf extract was separated by a Diaion HP20/water-acetone system and furthermore the most potent fraction by a Sephadex LH20/water-ethanol-acetone system. The 60% acetone fraction from the LH20/water-ethanol-acetone system had the most potent PLI activity (IC(50) value; 15 ppm). The active compounds in the active fraction of KT leaves were most likely a polyphenol, as assessed by the Folin-Ciocalteu method. Based on these spectroscopic and chemical examinations, the active fraction was shown to be proanthocyanidin oligomers composed of epigallocatechin, epicathechin, and epiafzelechin as main constituents. The degree of polymerization was estimated to be about 5 from the ratio of the peak area of the thio ethers/flavan-3-ols at 230 nm. This was consistent with the results of matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) MS, which showed the [M+Na](+) peaks corresponding to trimers-octamers. From the average molecular weight and IC(50) value of the active compounds estimated on these results, the active compounds from the KT leaf extract were one of the stronger effective lipid-lowering therapeutic agent, of which PLI activity (µM/L) was almost the same as epigallocatechin gallate. PRACTICAL APPLICATION: Proanthocyanidin oligomers isolated from Salacia reticulata, referred to "Kothala himbutu" (KT) in Singhalese, leaves was proved to potently inhibit pancreatic lipase activity. After confirming in vivo examination, healthy foods, teas, and liquors containing the extracts of KT leaves are expected to be on market.

Magnetic compression anastomosis for benign obstruction of the common bile duct.

Advances in interventional radiology have made possible magnetic compression anastomosis between the bile duct and the small intestine as a novel treatment. A 70-year-old man who had undergone subtotal gastrectomy for gastric cancer 2 years previously experienced recurring cholangitis with high fever and jaundice. Diagnostic evaluation subsequently demonstrated complete obstruction of the common bile duct which was attributed to recurrent cholangitis. A parent magnet was placed endoscopically into the afferent loop of the duodenum through the gastrojejunostomy with Billroth II reconstruction. The daughter magnet attached to a guide wire was placed in the obstructed common bile duct through a percutaneous transhepatic cholangiographic drainage tube. Two magnets were immediately attracted towards each other transmurally, and anastomosis was established on day 32 after the procedure. This novel method of magnetic compression anastomosis has the advantages of noninvasiveness and simplicity, as well as being a well-tolerated procedure for indications such as biliary obstruction.

In situ identification of polyphosphate- and polyhydroxyalkanoate-accumulating traits for microbial populations in a biological phosphorus removal process.

Polyphosphate- and polyhydroxyalkanoate (PHA)-accumulating traits of predominant microorganisms in an efficient enhanced biological phosphorus removal (EBPR) process were investigated systematically using a suite of non-culture-dependent methods. Results of 16S rDNA clone library and fluorescence in situ hybridization (FISH) with rRNA-targeted, group-specific oligonucleotide probes indicated that the microbial community consisted mostly of the alpha- (9.5% of total cells), beta- (41.3%) and gamma- (6.8%) subclasses of the class Proteobacteria, Flexibacter-Cytophaga (4.5%) and the Gram-positive high G+C (HGC) group (17.9%). With individual phylogenetic groups or subgroups, members of Candidatus Accumulibacter phosphatis in the beta-2 subclass, a novel HGC group closely related to Tetrasphaera spp., and a novel gamma-proteobacterial group were the predominant populations. Furthermore, electron microscopy with energy-dispersive X-ray analysis was used to validate the staining specificity of 4,6-diamino-2-phenylindole (DAPI) for intracellular polyphosphate and revealed the composition of polyphosphate granules accumulated in predominant bacteria as mostly P, Ca and Na. As a result, DAPI and PHA staining procedures could be combined with FISH to identify directly the polyphosphate- and PHA-accumulating traits of different phylogenetic groups. Members of Accumulibacter phosphatis and the novel gamma-proteobacterial group were observed to accumulate both polyphosphate and PHA. In addition, one novel rod-shaped group, closely related to coccus-shaped Tetrasphaera, and one filamentous group resembling Candidatus Nostocoidia limicola in the HGC group were found to accumulate polyphosphate but not PHA. No cellular inclusions were detected in most members of the alpha-Proteobacteria and the Cytophaga-Flavobacterium group. The diversified functional traits observed suggested that different substrate metabolisms were used by predominant phylogenetic groups in EBPR processes.

Identification of a novel thioredoxin-related transmembrane protein.

We recently identified a series of transforming growth factor-beta-responsive genes in A549 human adenocarcinoma cell line by a gene trap screening method. Here we report the molecular cloning and characterization of one of these genes, designated TMX, that encodes a novel protein of 280 amino acid residues. The TMX protein possesses an N-terminal signal peptide followed by one thioredoxin (Trx)-like domain with a unique active site sequence, Cys-Pro-Ala-Cys, and a potential transmembrane domain. There are putative TMX homologs with identical active site sequences in the Caenorhabditis elegans and Drosophila genomes. Using recombinant proteins expressed in Escherichia coli, we demonstrated the activity of the Trx domain of TMX to cleave the interchain disulfide bridges in insulin in vitro. The TMX transcript is widely expressed in normal human tissues, and subcellular fractionation and immunostaining for an epitope-tagged TMX protein suggest that TMX is predominantly localized in the endoplasmic reticulum (ER). When TMX was expressed in HEK293 cells, it significantly suppressed the apoptosis induced by brefeldin A, an inhibitor of ER-Golgi transport. This activity was abolished when two Cys residues in the active site sequence were mutated to Ser, suggesting that the Trx-like activity of TMX may help relieve ER stress caused by brefeldin A.

Experimental sensitization with Japanese cedar pollen in dogs.

Japanese cedar pollinosis is a type I allergic disease mediated by immunoglobulin E (IgE) antibodies to Japanese cedar (Cryptomeria japonica) pollen antigen (CPAg). By using 22 dogs consisting of 20 dogs aged 3 months and 2 dogs aged 3 years, immunization was performed by subcutaneous injections of CPAg with aluminum hydroxide gel. Variable levels of CPAg-specific IgE antibody response were detected in 21 of the 22 immunized dogs two weeks after the second immunization. This study provided an experimental sensitization system with CPAg in dogs, which will be useful for further immunological studies on Japanese cedar pollinosis.

Identification of a nuclear export signal in MKK6, an activator of the carp p38 mitogen-activated protein kinases.

Carp homologues of p38 mitogen-activated protein kinase (MAPK) and its activator MAPK kinase 6 (MAPKK6, referred to as MKK6) were identified. There exist at least two distinct carp p38s, cp38a and cp38b, both of which consist of 361 amino acids. The transcript of c38a was exclusively expressed in the ovary, whereas that of cp38b was ubiquitously expressed. Western blot analysis with anti-(phosphorylated MAPK) Ig specific to the active p38 or JNK has shown that p38 was activated in response to hypertonic stress (1 M sorbitol) in epithelioma papilosum cyprini carp epithelial cells (EPC) and that the activation of p38 proceeded faster to the maximal level than that of JNK. Carp homologue (cMKK6) of p38 activator MKK6 consists of 404 amino acids. It was expressed ubiquitously but was most abundant in the ovary. An in vitro kinase assay demonstrated that cMKK6 is an upstream activator of cp38 and cp38b in carp because it specifically phosphorylated and activated cp38a and cp38b. Interestingly, we found that cMKK6 has a nuclear export signal (NES) sequence in its N-terminal region although upstream activators of stress-activated MAPKs, p38 and JNK, do not in other animals. The NES sequence facilitated nuclear export of cMKK6 and ovalbumin. Leucine residues in the sequence were crucial for the NES activity, as the activity was lost on replacement of the leucines to alanines. The existence of an NES in cMKK6 implies the requisite of strict regulation of the p38 MAPK pathway in carp. The abundance of these components for the stress-activated pathway in the ovary might be related to ectogenetic early development.

Effect of coenzyme Q10 in patients with mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes (MELAS): evaluation by noninvasive tissue oximetry.

We evaluated the effect of coenzyme Q10 supplementation to two patients with mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes (MELAS) by using noninvasive tissue oximetry with near-infrared spectra of hemoglobin from the quadriceps muscle during bicycle ergometer exercise. Patients showed distinct oxygen consumption patterns reflecting the defect in oxidative phosphorylation and the impairment in oxygen utilization during exercise. Based on the oxygen consumption pattern, we considered one patient as having severe mitochondrial disorder and another patient as having mild one. After coenzyme Q10 supplementation, the oxygen consumption pattern of the patient with the severe form shifted to the mild one, while that of the patient with mild form remained unchanged. The shift of the pattern to the mild form correlated well with reduction of the sum of the serum lactate and pyruvate content during exercise. Noninvasive tissue oximetry may be useful to evaluate the effect of coenzyme Q10 supplementation to patients with mitochondrial encephalomyopathy including MELAS.

Structure and expression of carp mitogen-activated protein kinases homologous to mammalian JNK/SAPK.

Two distinct stress-activated protein kinase (JNKa and b) cDNAs were isolated from a carp ovary cDNA library. These cDNAs contained a full-length open reading frame encoding 427 amino acid residues with a predicted mass of 48.6 kDa. The deduced amino acid sequences of JNKa and b were 95.8% identical, with 18 residues replaced, and showed a high degree of sequence similarity to mammalian JNK/SAPK subgroup including the common dual phosphorylation motif of TPY. By Northern blot analysis, the carp JNKs were found to be abundant in the brain and ovary. Detailed study by RT-PCR assay revealed ubiquitous expression of JNKb, although expression of JNKa was specific to the brain and ovary. The high level expression of both JNKa and b in the ovary implies that JNKs play an important role in egg maturation or ectogenetic early development.

Extended V-Y flap: patient reports and reconsideration.

The extended V-Y flap, a modified V-Y advancement flap, is very useful in closing relatively large defects on the face. Its extension limb is hinged down as a transposition flap on the end of the V-Y advancement flap to close the most distal portion of the defect. We applied this flap in closing a defect following excision of skin tumors on the face with excellent cosmetic results in 11 patients. However, this flap tended to make a distortion at the base of the flap in the primary closure site. By drawing figures, we concluded that the distortion was due to the characteristic of this technique as a V-Y advancement-rotation flap or V-Y advancement flap with rotation.

Structurally altered Evi-1 protein generated in the 3q21q26 syndrome.

Overexpression of the Evi-1 gene appears to be a consistent feature of the 3q21q26 syndrome, an association of myeloid leukemias/myelodysplastic syndrome with a specific chromosomal aberration involving both 3q21 and 3q26, such as t(3;3)(q21;q26) or inv(3)(q21q26). The rearrangement in 3q26 has been reported to occur near the Evi-1 locus, implicating that it is the critical gene deregulated in the 3q21q26 syndrome. Here we present a structural abnormality of Evi-1 protein in a case with the 3q21q26 syndrome. In this case carrying typical inv(3)(q21q26), the 3q26 breakpoint is located within an intron of the Evi-1 gene, and resulted in overexpression of normally unexpressed, an aberrant form of Evi-1 protein, in which the C-terminal 44 amino acids of wild-type Evi-1 protein were truncated and replaced by five amino acids. The truncated Evi-1 protein is shown to increase AP1 activity when expressed in NIH3T3 cells as its wild-type counterpart. We also show that the origin of this peculiar type of rearrangement of the Evi-1 gene is not an artifact during establishment of the cell line, but is the event that occurred in the primary leukemic cells. Our results strongly support that the primary target for the 3q21q26 syndrome is the Evi-1 gene, and provide the first evidence that the structurally altered Evi-1 gene may be involved in the 3q21q26 syndrome.

The dipeptide alanyl-glutamine prevents intestinal mucosal atrophy in parenterally fed rats.

This study was performed to determine whether the addition of alanyl-glutamine (Ala-Gln) can prevent intestinal mucosal atrophy induced by standard solution of total parenteral nutrition (S-TPN). Forty-one male Sprague-Dawley rats weighing 250 g were randomly divided into four groups: group I was killed after overnight fasting; group II received S-TPN. The other groups received S-TPN supplemented with amino acids other than glutamine (group III) or supplemented with Ala-Gln 2 g/100 mL (group IV); both solutions were isocaloric and isonitrogenous. After 1 week of TPN the rats were killed, and the duodenum, proximal jejunum, mid-small bowel, and distal ileum were obtained for morphologic and functional analysis. Weight gain did not differ significantly among these four groups, and there was no difference in nitrogen balance between groups III and IV. Serum glutamine in group IV (102.8 +/- 13.3 mumol/dL) was significantly increased (p less than .05) compared with groups I, II, and III (66.2 +/- 3.9, 55.7 +/- 7.8, and 61.3 +/- 10.8 mumol/dL, respectively). Mucosal wet weight, protein, RNA, sucrase, and maltase of group IV were significantly increased (p less than .05) compared with groups II and III. Villus height was significantly increased (p less than .05) in the jejunum of group IV rats compared with groups II and III, but not in any other segments of the intestine. No significant changes were observed in crypt depth among all groups. Diamine oxidase in groups II, III, and IV was significantly decreased (p less than .05) compared with group I in all segments except for the ileum.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of sucralfate, lansoprazole, and cimetidine on the delayed healing by hydrocortisone sodium phosphate of chronic gastric ulcers in the rat.

We have previously shown that chronic sucralfate ingestion stimulates gastric epithelial proliferation in rats, which may explain one of the beneficial effects of sucralfate in healing of peptic ulcers. In a separate study, we have found that chronic steroid administration delays the healing of experimental gastric ulcers in rats. This study was designed to test the beneficial effects of sucralfate, cimetidine, and lansoprazole (AG-1749, a new proton pump inhibitor), on the delayed healing by steroids in rat chronic gastric ulcers. Chronic gastric ulcers were produced in male Wistar rats, weighing 180 g, by the application of 100% acetic acid. The rats were randomly divided into five groups; (1) control, (2) vehicle alone, (3) 10 mg/kg lansoprazole, (4) 500 mg/kg sucralfate, and (5) 100 mg/kg cimetidine. Except for controls, all rats received daily intraperitoneal injections of 2.5 mg/kg hydrocortisone sodium phosphate. Tested drugs were administered intragastrically (lansoprazole and sucralfate) or intraperitoneally (cimetidine) twice a day for 2 weeks. Rats were sacrificed 14 days later and ulcer size was measured. Chronic administration of hydrocortisone sodium phosphate resulted in a significant delay of ulcer healing induced by acetic acid. Treatment with either lansoprazole or sucralfate abolished the deleterious effect of steroids, whereas cimetidine had no effect. These results indicate that lansoprazole and sucralfate overcome the delayed healing by steroids of chronic gastric ulcers in the rat.

Comparative studies on the usefulness of phosphate versus glycerin enema in preparation for colon examinations.

Fifty of 100 persons who had undergone health screening received phosphate enema while the other 50 received glycerin enema prior to proctoscopy and barium enema, and their usefulness for preparation for colon examination was compared by a double-blind test. There was no significant difference in the degree of colonic cleansing achieved by proctoscopy and barium enema. In the subjects who received phosphate enema, the incidence of abdominal pain was less than that in those who received glycerin enema, while the effect of phosphate enema on defecation appeared later than that of glycerin enema, indicating prolonged stool retention in the subjects given phosphate enema. To study the safety of the two enemas, either phosphate enema, glycerin enema or physiological saline solution as a control was administered at 0.35 ml/animal in the rectum by 4-h closure of the anus in 10 male 7-week-old Wistar rats, and the rectal mucosa was observed for irritation macroscopically and histopathologically. Glycerin enema produced less irritation than phosphate enema diffusely in the entire area of the rectum, while phosphate enema produced more local irritation at the end of the rectum than glycerin enema. The differences in the extent of irritation and injury between phosphate and glycerin enemas were considered to be derived from differences in the pharmacologic actions of these drugs. If the extent of injury were included in the extent of irritation, the difference in irritation between phosphate and glycerin enemas would not be significant. As described above, no specific difference seem to exist in the usefulness of phosphate and glycerin enemas as preparation for colon examination.