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Biomedical subjects

H C Tseng

Publications and source records attributed to H C Tseng.

At least 19 recordsLinked to original sources

A cloned ompR-like gene of Streptomyces lividans 66 suppresses defective melC1, a putative copper-transfer gene.

Expression of tyrosinase in Streptomyces requires functional MelC1 protein, which is postulated to transfer copper to apotyrosinase. We have previously isolated a mutant of Streptomyces lividans, HT32, that phenotypically suppressed mutations in cloned melC1 (H.-C. Tseng and C. W. Chen, in preparation). Plasmid pLUS132, containing an ATG to ATA transition at the initiation codon of melC1, was used for cloning the suppressor gene from HT32. A 1687 bp suppressor DNA was isolated that contained two characteristic Streptomyces coding sequences: a 217-amino-acid open reading frame (cutR) and a truncated open reading frame (cutS) downstream. Subcloning analysis attributed the phenotypic suppression activity to the putative cutR gene from HT32. The putative CutR exhibited similarity to the response regulator OmpR of the osmoregulatory signal-transduction system in Escherichia coli. The truncated CutS resembled, to a lesser degree, the N-terminus of EnvZ, the histidine protein kinase counterpart of OmpR. DNA hybridizing to the cloned cutR-cutS sequence was detected in 16 other Streptomyces species. We postulate that the putative cutR-cutS operon regulates copper metabolism in Streptomyces.

Amino Acid Sequence

Neural regulation of lysozyme secretion from tracheal submucosal glands of ferrets in vivo.

To investigate how central and peripheral nerves affect lysozyme secretion from tracheal submucosal glands in ferrets we injected substance P (20 nmol/kg in 200 microliters) intracisternally or intravenously into anesthetized artificially ventilated ferrets. We collected 3-ml samples from a perfused (3 ml/5 min) segment of trachea in situ during 15 min before and 45 min after injection of substance P. Content of lysozyme, a specific marker of tracheal submucosal gland serous cell secretion in ferrets, was measured spectrophotometrically in each sample. Intracisternal substance P increased peak lysozyme output threefold compared with baseline. This increase was abolished completely by cutting both superior laryngeal nerves (SLN) and was partially inhibited by atropine, phentolamine, or propranolol. Intravenous substance P increased peak lysozyme output 10-fold compared with baseline. This increase was partly abolished by cutting both SLN. We concluded that intracisternal substance P stimulated the central nervous system (CNS) and activated cholinergic, adrenergic, and nonadrenergic noncholinergic secretomotor nerves to tracheal glands and that intravenous substance P increased lysozyme secretion both by acting directly on tracheal glands and indirectly on the CNS to activate secretomotor nerves.

Animals

The melanin operon of Streptomyces antibioticus: expression and use as a marker in gram-negative bacteria.

The melC operon of Streptomyces antibioticus contains two genes, melC1 and melC2, necessary for the production of melanin pigment. We transferred the coding sequence of melC1 and melC2 to Escherichia coli plasmid pMTL23 such that its transcription was under the control of the lac promoter and melC1 was translationally fused to the lacZ alpha fragment. E. coli cultures containing this plasmid, pIF413, produced melanin after overnight incubation on 2YT agar supplemented with 0.1 mM CuCl2, 0.36 mM IPTG (or 0.2% lactose), and 2 mM tyrosine. Erwina carotovora could also be transformed by pIF413 to produce melanin. Two shuttle vectors were constructed: pLUS415 for E. coli and Streptomyces, and pLAF413 for E. coli and Xanthomonas campestris. These vectors confer melanin pigmentation in all the hosts that harbor them. The melC sequence provides the vectors with a convenient cloning marker for insertional or replacement inactivation.

Catechol Oxidase

Structure-activity relationships of chloride-sensitive fluorescent indicators for biological application.

The application of the quinoline derivative 6-methoxy-N-(3-sulfopropyl)quinolinium (SPQ) to the measurement of membrane transport of chloride in biological systems was reported recently (N.P. Illsley and A.S. Verkman (1987) Biochemistry 26, 1215-1219). To understand the structure-activity relationships of compounds with chloride-sensitive fluorescence properties, 19 structural analogs of SPQ having a single quaternized nitrogen heteroatom were synthesized and characterized. The effect of variations in ring structure, length of sulfoalkyl chain, position of ring substituent, and nature of ring substituent were examined. For each compound, the water solubility, octanol:water partition coefficient, absorbance and fluorescence spectra, fluorescence lifetime, and Stern-Volmer constants (Kq) for quenching by a series of anions were measured. All compounds were quenched by chloride, bromide, iodide, and thiocyanate, but not by cations, sulfate, phosphate, nitrate, or by pH (5-8); several compounds were quenched slightly by bicarbonate (Kq = 8-12 M-1). High chloride sensitivity (Kq greater than 50 M-1) required the presence of a quinoline backbone substituted with electron-donating groups such as methyl and methoxy, but did not depend on length of the sulfoalkyl chain or on the position of ring substituents (positions 2-7). All compounds with high chloride sensitivity had fluorescence excitation spectra in the ultraviolet (excitation maximum less than 350 nm) and fluorescence lifetimes greater than 15 ns. These results establish a set of guidelines for synthesis of chloride-sensitive fluorescent indicators tailored for specific biological applications.

Chlorides

Evaluation of parenteral nutrition in the postoperative patient.

A variety of investigators have attempted to improve nitrogen balance during the postoperative period by modifying the composition of the infused nutrient solutions. This study compared the metabolic effects of administering standard amino acid solutions with a solution enriched with branched chain amino acids (BCAA). A prospective, randomized clinical study was performed in patients who had undergone subtotal gastrectomy or hemicolectomy, and subsequently cared for in the metabolic care unit. The patients were selected from specific entry criteria so that two groups of individuals were comparable. All patients underwent operation without complications. The plasma concentrations of valine and leucine were significantly increased (p less than 0.05 and p less than 0.01, respectively) two days after administration of solutions enriched with BCAA and throughout the entire postoperative period. The plasma glutamine concentrations tended to decrease in both groups; no concentration difference occurred between groups. Nitrogen balance tended to be more positive in the group receiving BCAA but there was no significant difference between groups after operation. Urinary excretion of 3-methylhistidine tended to increase postoperatively in both groups, but no difference occurred between groups. However, the urine excretion of isoleucine increased significantly in the patients receiving infusions enriched with BCAA. Both standard balanced amino acid and amino solutions enriched with BCAA were well tolerated in all patients.

Amino Acids, Branched-Chain

Insulinoma. An immunocytochemical and morphologic analysis of 95 cases.

One hundred twenty-seven insulinomas from 95 cases (1 malignant and 94 benign) were studied pathologically. Thirty-six tumors (35 cases) were examined by electron microscopy. Typical beta-cell secretory granules of crystalloid-form cores and/or atypical secretory granules were discerned in all tumors examined. A new type of secretory granule, with high electron-dense crystalloid-form cores and moderate electron-dense granular substance filling the space between the core and the limiting membrane, were observed in two cases. Among 68 insulinomas (67 cases) subjected to immunocytochemical investigations with ten peptide hormones (insulin, glucagon, somatostatin, pancreatic polypeptide (PP), gastrin, motilin, secretin, vasoactive intestinal polypeptide (VIP), gastric inhibitory polypeptide (GIP), and neurotensin), 42 were found to be multihormonal, varying from two to four peptides secreted. The hormones contained were insulin, glucagon, PP, somatostatin, and gastrin in different combinations. One patient had hyperinsulinemia and hypergastrinemia concurrently, and two islet tumors were excised at an interval of 10 months. Both electron microscopy and immunocytochemistry confirmed the presence of beta- and alpha-cells in the first tumor, whereas the second tumor revealed only G-cells by electron microscopy, and G- and beta-cells on immunocytochemical staining. The morphologic and immunocytochemical characteristics of the insulinomas in this series are discussed.

Adenoma, Islet Cell

Regulation of digestion. I. Effects of glucose and lysine on pancreatic secretion.

Previous cell-free studies have shown that glucose selectively elicits the release of amylase from pancreatic zymogen granules, whereas lysine promotes the selective release of trypsinogen. To investigate the expression of these effects in situ, glucose or lysine was injected into the celiac artery of anesthetized rats, either alone or together with the pancreatic secretagogue cholecystokinin, to evaluate their effects on the secretion of amylase and trypsinogen by the pancreas. When given alone neither substance significantly changed the output of either enzyme. However, when given with cholecystokinin, each altered the effect observed with injection of cholecystokinin alone. The injection of glucose resulted in a twofold increase in both peak and total amylase output without significantly increasing trypsinogen secretion, whereas lysine increased both peak and total trypsinogen output by about 50%, leaving amylase output unchanged. These findings provide in situ confirmation for the selective enzyme release produced by glucose and lysine in cell-free studies and suggest that such end products of digestion can regulate the digestive process by modifying the secretory response of the pancreas to cholecystokinin.

Animals

Regulation of digestion. II. Effects of insulin and glucagon on pancreatic secretion.

The endocrine islet-cell hormones insulin and glucagon are secreted at high concentrations into an intrapancreatic portal circulation and have been reported to affect the secretion of digestive enzyme by the exocrine pancreas. In the present experiments, insulin and glucagon were injected into the celiac artery of anesthetized rats to evaluate their effects on the secretion of amylase and trypsinogen by the pancreas. Neither hormone when given alone significantly changed the output of either enzyme. However, when given with the pancreatic secretagogue cholecystokinin, each altered the effect of injection of cholecystokinin. In a dose-dependent fashion insulin increased trypsinogen output without affecting amylase output, whereas glucagon inhibited amylase output and left trypsinogen output unchanged. Thus, both hormones produced a more trypsinogen-dominant pancreatic juice than that observed with cholecystokinin alone, although in different ways. These findings suggest that the endocrine hormones insulin and glucagon may regulate secretion of digestive enzymes by the pancreas by modulating the response to stimuli of overall protein secretion such as cholecystokinin.

Animals

Food, duodenal extracts, and enzyme secretion by the pancreas.

Previous studies have reported that injection of duodenal extracts from rats fed different meals into the celiac artery of recipient rats elicited the secretion of related pancreatic enzymes. We have been unable to reproduce the enzyme-specific increases in the average output of particular enzymes that were observed but did find changes similar in direction, although not magnitude, to those reported previously. The outputs of amylase and trypsinogen were compared by plotting individual data points and performing a regression analysis on them. The injection of duodenal extracts from lactalbumin hydrolysate-fed rats led to trypsinogen secretion being favored over that of amylase and vice versa for extracts from rats fed a glucose meal. In addition, it was found that cholecystokinin-pancreozymin produced a dramatic nonparallel transport of these two enzymes with amylase secretion being augmented to a greater degree than trypsinogen secretion. The relation between their outputs was curvilinear, i.e., the amylase dominance of secretion became more pronounced as overall enzyme output (not dose of hormone) increased. Thus, this nonparallel secretion does not seem to be the results of a discontinuous switch in the character of enzyme secretion produced by the hormone but a graded effect reflecting the magnitude of the response.

Amylases

Insulinoma. Experience in surgical treatment.

Sixty cases of insulinoma were treated surgically. The possible causes for failure to find the tumor during exploratory surgery are discussed. Simple enucleation is preferred in most cases of single or a few insulinomas. A technique of enucleation with minimal chance of injury to adjacent pancreatic ducts and larger blood vessels can be used. When no tumor is found after thorough exploratory procedures, controlled staged resection of the pancreas under blood glucose monitoring is the procedure of choice. Postoperative complications are common but the late results are gratifying provided the operation is carried out early in the course of the disease.

Adenoma, Islet Cell