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Biomedical subjects

H Y Kim

Publications and source records attributed to H Y Kim.

At least 19 recordsLinked to original sources

Postnatal selenium repletion protects lungs of neonatal rats from hyperoxia.

We reported previously that Se-adequate neonatal rat pups born to Se-adequate dams were resistant to lung damage by hyperoxia. To assess whether early postnatal Se repletion could also protect developing pups reared under hyperoxia, female Sprague-Dawley rats (n = 20) were bred and fed a Se-deficient (0.04 microgram/g) diet during pregnancy. On d 1 postpartum, dams were divided into two groups and fed either a Se-deficient diet or a Se-repleted (0.5 microgram/g) diet. On d 4 postpartum, litters in each group were randomly assigned to either air or high oxygen (greater than 95% O2) environments. Histologic evaluation of lungs from d-8 pups indicated that Se repletion significantly reduced the incidence of lung lesions caused by hyperoxia. Selenium-repleted pups also had significantly greater lung volumes and internal surface areas. The 7-d period of Se repletion resulted in significantly elevated maternal milk Se concentrations compared with a Se-deficient group, which was reflected in the pups by elevated plasma and hepatic Se concentrations and Se-dependent glutathione peroxidase (SeGPx) activities. Pulmonary glutathione concentration and SeGPx activity in pups were affected by oxygen exposure only, not by Se nutrition. Therefore, early postnatal Se repletion can protect the developing lung from oxygen-induced injury, a protection that is not entirely due to the effects of Se on pulmonary SeGPx activity and glutathione concentration.

Animals

Three clustered origins of replication in a promiscuous-plasmid replicon and their differential use in a PolA+ strain and a delta PolA strain of Escherichia coli K-12.

A 1,197-bp region of the broad-host-range plasmid pCU1 is adequate for its replication. Analysis of replicating molecules containing this region reveals three clustered origins of vegetative replication and replication proceeds bidirectionally from each in a theta mode. In an Escherichia coli polymerase I deletion mutant, utilization of one of these three origins was not detected. The potentiality for origin utilization may therefore be a determinant of replicon host range.

Cloning, Molecular

Interferon alpha therapy in patients with chronic type C hepatitis: changes of serum ALT, anti-HCV & HCV-RNA.

BACKGROUND: After the discovery of type C hepatitis virus, the studies on this virus are extensively progressing. The treatment of this viral infection is also widely progressing. Among many agents, recombinant interferon alpha therapy is generally accepted as an effective single agent. To evaluate the efficacy of interferon and to observe the changes of serum aminotransferase (ALT), antibody to hepatitis C virus (anti-HCV) and HCV ribonucleic acid (HCV-RNA), we treated 10 patients with chronic type C hepatitis for 6 months. METHODS: Patients were randomly divided into 2 groups: 5 patients in group A received interferon and the other 5 in group B received no therapy. Interferon was administered at a dose of 3 million units (MU) daily for the first month and thrice weekly for the following 5 months, and followed up for 2 years. RESULTS: In group A, serum ALT returned to normal in 4: 3, starting at the first month and one at the 3rd month of therapy and maintained normal throughout the follow-up period. In contrast, serum ALT level persistently fluctuated in 4 patients in group B. In one patient, serum ALT returned to normal one and a half years later. Regardless of therapy, serum anti-HCV titer remained unchanged in all patients. However, HCV-RNA, using polymerized chain reaction (PCR), became undetectable in all responded patients and in one untreated patient whose serum ALT returned to normal spontaneously. CONCLUSION: This study suggested that interferon alpha therapy in patients with chronic type C hepatitis may be clinically effective. Our study also indicated that the detection of HCV-RNA by PCR is useful to predict the prognosis of chronic type C hepatitis.

Adult

Arginine-395 is required for efficient in vivo and in vitro aminoacylation of tRNAs by Escherichia coli methionyl-tRNA synthetase.

We have previously shown that the anticodon of methionine tRNAs contains the major recognition site required for aminoacylation of tRNAs by Escherichia coli methionyl-tRNA synthetase (MetRS) and have located part of the anticodon binding domain on the enzyme at a site close to Trp461 [Schulman, L. H., & Pelka, H. (1988) Science 242, 765-768; Ghosh, G., Pelka, H., & Schulman, L.H. (1990) Biochemistry 29, 2220-2225]. In order to gain information about other possible sites of contact between MetRS and its tRNA substrates, we have examined the effects of mutations at a series of positively charged residues on the surface of the C-terminal domain of the enzyme. Conversion of Arg356, Arg366, Arg380, or Arg453 to Gln had little or no effect on enzyme activity. Similarly, conversion of Lys402 or Lys439 to Asn failed to significantly alter aminoacylation activity. Conversion of Arg380 to Ala or Arg442 to Gln produced a 5-fold reduction in kcat/Km for aminoacylation of tRNAfMet, with no effect on methionine activation, indicating a possible minor role for these residues in interaction of the enzyme with the tRNA substrate. In contrast, mutation of a phylogenetically conserved residue, Arg395, to Gln increased the Km for aminoacylation of tRNAfMet about 30-fold and reduced kcat/Km by 25,000-fold. The mutant enzyme was also shown to be highly defective by its inability to complement a strain of E. coli having an altered chromosomal MetRS gene.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence

Lipoxygenation in rat brain?

It has been previously claimed that rodent brain possesses lipoxygenase activity, based upon the structure of products which were formed from arachidonic acid and the inhibition of this activity by "lipoxygenase inhibitors." Our studies confirm that various positional isomers of hydroxyeicosatetraenoic acids (HETE) are formed (e.g., 15-, 12-, 11-, 9-, 8- and 5-HETE) by brain homogenate and that their production is inhibited by certain lipoxygenase inhibitors, such as nordihydroguaiaretic acid (NDGA) but not by cyclooxygenase or cytochrome P-450 inhibitors. However, stereochemical analysis indicated racemic distributions of these products suggesting that they were not formed by a lipoxygenase enzyme but rather by a peroxidative process. It should also be noted that the presence of 12(S)-lipoxygenase activity could be demonstrated by stereochemical analysis only when the brain was not perfused properly, indicating this activity was due to blood cell contamination. It is known that many lipoxygenase inhibitors are also capable of inhibiting peroxidative reactions apparently due to their free radical scavenging properties. For these reasons, it is essential that the stereochemical purity of purported lipoxygenase products be determined and that previous claims of lipoxygenase activity in mammalian brain be reexamined.

5,8,11,14-Eicosatetraynoic Acid

DNA cross-linking in mammalian cells by pyrrolizidine alkaloids: structure-activity relationships.

Pyrrolizidine alkaloids (PAs) are common constituents of many species of flowering plants which possess carcinogenic as well as anticarcinogenic activity in vivo. Pyrrolizidine alkaloids are genotoxic in various short-term assays. The mechanisms by which these compounds exert these effects is still unclear. In this study, we characterized the ability of eight bifunctional PAs, with differing stereochemistry and functional groups, to cross-link cellular DNA in cultured bovine kidney epithelial cells. PAs representative of three major structural classes, the macrocycles (seneciphylline, riddelline, retrorsine, senecionine, monocrotaline), the open diesters (heliosupine, latifoline), and pyrrolizidine base (retronecine) were cultured for 2 hr with cells and an external metabolizing system. Every PA induced DNA cross-links which consisted primarily of proteinase-sensitive cross-links (DPC), but also to a smaller extent, DNA interstrand cross-links (ISC). None of the PAs induced detectable amounts of DNA single-strand breaks. The PAs which produced DPC and/or ISC (ranked from most potent to least) were: seneciphylline (DPC greater than ISC); riddelline (DPC greater than ISC); retrorsine (DPC greater than ISC); senecionine (DPC greater than ISC); heliosupine (DPC greater than ISC); monocrotaline (ISC = DPC); latifoline (DPC greater than ISC); and retronecine (ISC greater than DPC). Although the PAs induced DNA cross-linking to varying degrees, cell viabilities for all treatment groups were greater than 90% as determined by trypan blue dye exclusion. Since the cross-linking ability of these PAs paralleled their ability to inhibit colony formation, cross-link formation may be involved in the biological activity of these compounds. Two structural determinants of biological activity appear to be the presence of both a macrocyclic necic acid ester and an alpha,beta-unsaturated ester function since the cross-linking ability of seneciphylline, riddelline, retrorsine, and senecionine far exceeded that of monocrotaline, heliosupine, latifoline, and retronecine. In addition, the stereochemical orientation of the ester linkage was found to have no effect on biological activity.

Animals

Brain microvessel 12-hydroxyeicosatetraenoic acid is the (S) enantiomer and is lipoxygenase derived.

12-Hydroxyeicosatetraenoic acid (12-HETE) production from arachidonic acid by cerebral microvessels isolated from perfused adult murine brain was reduced by the lipoxygenase inhibitors baicalein, esculetin, gossypol, nordihydroguaiaretic acid, and quercetin. Except for quercetin and gossypol, the IC50 did not exceed 10 microM. Each inhibitor, except baicalein, also decreased microvessel prostaglandin production when present in concentrations above their IC50 value for 12-HETE. In contrast, inhibitors of the cytochrome P450 monooxygenase system, clotrimazole, metyrapone, and proadifen (SKF-525A), had little effect on microvessel 12-HETE production. Chiral phase HPLC analysis revealed that only the (S) enantiomer of 12-HETE was formed. The major microvessel metabolite of eicosapentaenoic acid co-eluted with 12-hydroxyeicosapentaenoic acid (12-HEPE) on reverse-phase HPLC and the (S) enantiomer of 12-HEPE on chiral phase HPLC. Furthermore, like 12-HETE, 12-HEPE production was blocked by lipoxygenase inhibitors. These studies demonstrate that brain microvessels produce only the (S) enantiomeric 12-hydroxy derivatives of both arachidonic acid and eicosapentaenoic acid by the action of a lipoxygenase that can be selectively inhibited by baicalein. Since arachidonic acid and eicosapentaenoic acid are available to cerebral blood vessels in certain pathological settings, these 12-hydroxy acid lipoxygenase products may mediate some of the cerebrovascular dysfunction that occurs following stroke, brain trauma, or seizures.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid

The role of selenium nutrition in the development of neonatal rat lung.

Our study was designed to assess the role of selenium (Se) in development of neonatal lungs under conditions of normoxia and hyperoxia. Thirty-six female Sprague Dawley rats were bred and fed a Se-deficient (0.03 ppm Se) or a Se-adequate (0.5 ppm Se) diet during pregnancy and lactation. At d 2 postpartum, 24 litters were randomly assigned to either high oxygen (greater than 95%) or air and were cross-fostered for 4 d. Lung weight was significantly enhanced in Se-adequate pups and was not related to high oxygen or air exposure of either the pups or dams. Two types of histologic lesions were observed in the lungs of the pups: septal attenuation and interstitial inflammation. When reared in oxygen, all (17 of 17) Se-deficient pups had lesions. In contrast, only 60% (9 of 15) of Se-adequate pups were affected (p less than 0.01). Lung lesions also were more severe in Se-deficient pups. Se-deficient pups also displayed a significant degree of septal attenuation when reared in air. Se-dependent glutathione peroxidase activity in the pup lung was significantly elevated in response to hyperoxia and was unrelated to Se nutriture. No differences in activities of lung superoxide dismutase, catalase, and glutathione s-transferase were noted between Se-deficient and Se-adequate pups reared in air or high oxygen environments. These data indicate that Se has an important role in the development of neonatal lungs, a role that is even more pronounced during conditions of hyperoxia. The protective role of Se in developing lung tissue cannot be completely explained by enhanced glutathione peroxidase activity.

Animals

A case of intestinal hemorrhage due to small intestinal metastases from primary lung cancer.

Although intestinal metastases from lung cancer are not rare at postmortem studies, the development of clinically significant symptoms from the gastrointestinal metastases is very unusual. We report a case of small intestinal hemorrhage leading to intestinal perforation secondary to metastases from a large cell carcinoma of the lung in a 31-year-old man along with a review of the literature.

Adult

The effect of phytochrome action on the activity of cytosolic cholinesterase in oat cells.

Cholinesterases in the oat cell were found to be distributed in the cell wall (50%) and cytoplasm (42%). Activity of the cytosolic enzyme was inhibited about 80% by 1 mM Ca2+. The enzyme activity was also inhibited by Mn2+, but no inhibition by Mg2+ was observed. Effects of red light and calcium ion on the enzyme activity were investigated in vivo to confirm the involvement of phytochrome action in the regulation process of this enzyme via Ca2+. It was observed that inhibition by red light only occurs when external Ca2+ existed in the cell medium. Based on a previous report (8) that red light stimulates the influx of Ca2+ into the cytosol of oat cell, inhibition of the enzyme activity by irradiation of red light can be suggested to occur via the influx of Ca2+.

Calcium

Stereochemical analysis of hydroxylated docosahexaenoates produced by human platelets and rat brain homogenate.

The stereochemical configuration of hydroxylated products of docosahexaenoic acid (22:6w3) formed by human platelets and rat brain homogenate were characterized for the first time. Chiral phase HPLC was employed along with autooxidized 22:6w3 as reference material. The 14- and 11-hydroxy 22:6w3 (HDHE) products produced by human platelets were in the S configuration. Rat brain homogenate produced all of the ten possible positional isomers when incubated with 22:6w3. Their retention behavior on the reversed and chiral phase HPLC columns and GC/MS/EI analysis indicated that they were 20-, 17-, 16-, 14-, 13-, 11-, 10-, 8-, 7- and 4-HDHE. However, stereochemical analysis revealed that each positional isomer was a racemic mixture, suggesting that these were not formed by lipoxygenation but mainly by peroxidation process.

Animals

Formation of 15-lipoxygenase product from docosahexaenoic acid (22:6w3) by human platelets.

The metabolism of docosahexaenoic acid (22:6w3) by 15-lipoxygenase activity of washed human platelets was investigated. Platelets produced 17-hydroxydocosahexaenoic acid (HDHE) when incubated with 22:6w3. Similarly, 15-hydroxyeicosatetraenoic acid (HETE) and 13- and 9-hydroxyoctadecadienoic acids (HODD) were produced when incubated with 20:4w6 and 18:2w6, respectively. However, these products were observed only as minor components in the platelet incubation mixture. Control studies with carefully purified platelets and mononuclear cells indicated that these products were formed by the platelets. Chiral phase HPLC analysis indicated that these compounds were mainly in the S configuration with the exception of the 9-HODD, thus, confirming that a lipoxygenase is responsible for their production. The 9-HODD produced by platelets was a racemic mixture.

Arachidonate 15-Lipoxygenase

Separation of lipid classes by solid phase extraction.

A rapid and reliable method for the separation of lipid classes is described using aminopropyl disposable columns. This method is a modification to an existing procedure that allows the separation of both neutral and acidic phospholipid fractions and a high recovery of the latter. Acidic phospholipids were eluted with a mixture of hexane-2-propanol-ethanol-0.1 M ammonium acetate-formic acid 420:350:100:50:0.5 containing 5% phosphoric acid after neutral phospholipids had been eluted with methanol. It was verified that extremely high recoveries of cholesterol (CH), triglycerides (TG), free fatty acids (FFA), phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylinositol (PI), phosphatidylserine (PS), phosphatidic acid (PA), sphingomyelin (SM), and cerebrosides were obtained with this method. In addition, there appeared to be no preferential losses or degradation of any particular molecular species as the fatty acid distribution of bovine brain PS and the molecular species profile of plant PI were unaltered by the procedure. Depending on the tissue, this method may yield fractions containing pure lipid classes and/or simple mixtures of lipid classes of similar polarity. These fractions may then be more easily separated by thin-layer chromatography or high performance liquid chromatography for a complete lipid class analysis.

Animals

Synthesis of paf-acether from exogenous precursors by the prokaryote Escherichia coli.

Paf-acether (paf) is a potent mediator of inflammatory diseases and septic shock. Using normal-phase HPLC, a paf-like activity was found in culture supernatants from E. coli. Prokaryotic paf exhibited the same biological and physico-chemical properties as eukaryotic cells and synthetic paf. Further, reverse-phase HPLC indicates that paf generated by bacteria is predominantly of the hexadecyl and octadecyl species. When cultures were supplemented with lyso-paf, a dramatic increase in paf production was observed. The purity and molecular structure of bacterial paf were further characterized by mass spectral analysis. These results could be of importance considering the pathogenetic role of enterobacteria. Further, it appears that the competence to form and release paf is an early phylogenetic development.

Animals

Cyclopentenylcytosine triphosphate. Formation and inhibition of CTP synthetase.

Cyclopentenylcytosine (CPEC) is phosphorylated in L1210 cells with CPEC triphosphate as the major metabolite. Partially purified uridine-cytidine kinase catalyzes the initial phosphorylation of cyclopentenylcytosine with an apparent Km of 196 +/- 9 microM, and cyclopentenylcytosine is a competitive inhibitor of cytidine phosphorylation by this enzyme with a Ki value of 144 +/- 14 microM. Examination of the CTP synthetase activity in extracts of L1210 cells revealed a dose-dependent decrease on exposure of cells to CPEC. Synthesis of CPEC triphosphate by an enzymatic method permitted direct examination of the inhibition of partially purified CTP synthetase. CPEC triphosphate inhibited bovine CTP synthetase with a median inhibitory concentration of 6 microM, whereas CPEC mono- and diphosphates were ineffective. CTP synthetase showed a classical Michaelis-Menten hyperbolic plot of velocity and UTP concentration in the presence of saturating concentrations of ATP and glutamine, but CPEC triphosphate induced sigmoidal kinetic plots. The Hill coefficient was calculated to be 3.2.

Animals

Preparation and the structural determination of hydroperoxy derivatives of docosahexaenoic acid and other polyunsaturates by thermospray LC/MS.

A new method to determine the structure of lipoxygenase reaction products is presented. Thermospray mass spectra of hydroperoxy derivatives of polyunsaturates contain both molecular ion species and fragments reflecting the position of oxygenation. Data are presented for hydroperoxy-docosahexaenoic, eicosapentaenoic, arachidonic and linoleic acids in this regard. Ten positional isomers of hydroperoxy docosahexaenoic acid were prepared by autooxidation and their structures were determined by thermospray LC/MS and confirmed by electron impact GC/MS after suitable derivatives were made. This technique was particularly useful in determining the structure of unknown metabolites by direct monitoring of the reaction mixture without derivatization. In this paper, the value of this approach is demonstrated using a soybean lipoxygenase reaction mixture as a simple example.

Arachidonic Acids

Effect of short-term prednisolone therapy in patients with severe chronic type B hepatitis.

Ten patients with severe chronic type B hepatitis confirmed by liver biopsy were treated with prednisolone for eight weeks and followed up for more than one year. The patients were comprised of 6 males and 4 females, ages 17 to 45 (mean 32) yrs. Serum alanine aminotransferase (ALT) was elevated more than one month before the treatment in all (mean: 379 U/L, range: 87 to 772 U/L). Initial serological tests showed hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) in all and hepatitis B virus DNA (HBV-DNA) in 7/10 (70%). Liver biopsy showed severe chronic active hepatitis with confluent necrosis or acinar hepatitis in all. Prednisolone, 60 mg/day, was administered initially and the dose was tapered every 2 weeks over the 8 weeks period. Two to six months after cessation of treatment, 5 of 10 patients showed a disappearance of HBeAg and serum HBV-DNA and return of serum ALT level to normal (responders). The initial serum ALT level in responders was slightly higher than that of non-responders (mean: 404 vs. 355 U/L), but there was no statistical significance. Among 5 responders, serum HBV-DNA was detected in three patients initially and was transiently detected in one patient during treatment. In non-responders, HBeAg persisted during and after the treatment and serum HBV-DNA persisted in three, but serum ALT was decreased in all. One patient who did not show any clinical or serological improvement, died of jaundice, ascites and hepatic encephalopathy 4 months later.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent