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Biomedical subjects

R H Chen

Publications and source records attributed to R H Chen.

At least 109 records · Page 6Linked to original sources
PubMed1991

Coordinate regulation of pp90rsk and a distinct protein-serine/threonine kinase activity that phosphorylates recombinant pp90rsk in vitro.

Protein kinase assays that use recombinant pp90rsk as a substrate were developed in an attempt to identify growth-regulated enzymes responsible for the phosphorylation and activation of pp90rsk S6 phosphotransferase activity. With this assay we have ientified a pp60v-src-, growth factor-, phorbol ester-, and vanadate-regulated serine/threonine protein kinase activity that is not related to two other cofactor-independent, growth-regulated protein kinases, pp70-S6 protein kinase and pp90rsk. The pp90rsk-protein kinase activity (referred to as rsk-kinase) is also not related to cofactor-dependent signal transducing protein kinases such as the cyclic AMP-dependent protein kinases, members of the protein kinase C family, or other Ca2(+)-dependent protein kinases. In vitro, partially purified rsk-kinase phosphorylates several of the sites (serine and threonine) that are phosphorylated in growth-stimulated cultured cells. A detailed examination of the mitogen-regulated activation kinetics of rsk-kinase and pp90rsk activities demonstrated that they are coordinately regulated. In addition, protein kinase C is not absolutely required for epidermal and fibroblast growth factor-stimulated activation of rsk-kinase, whereas, like pp90rsk, platelet-derived growth factor- and vanadate-stimulated rsk-kinase activity exhibits a greater dependence on protein kinase C-mediated signal transduction. The characterization and future purification of the rsk-kinase(s) will improve our understanding of the early signaling events regulating cell growth.

Animals↗
PubMed1991

Cell cycle-dependent strand bias for UV-induced mutations in the transcribed strand of excision repair-proficient human fibroblasts but not in repair-deficient cells.

To study the effect of nucleotide excision repair on the spectrum of mutations induced in diploid human fibroblasts by UV light (wavelength, 254 nm), we synchronized repair-proficient cells and irradiated them when the HPRT gene was about to be replicated (early S phase) so that there would be no time for repair in that gene before replication, or in G1 phase 6 h prior to S, and determined the kinds and location of mutations in that gene. As a control, we also compared the spectra of mutations induced in synchronized populations of xeroderma pigmentosum cells (XP12BE cells, which are unable to excise UV-induced DNA damage). Among the 84 mutants sequenced, base substitutions predominated. Of the XP mutants from S or G1 and the repair-proficient mutants from S, approximately 62% were G.C----A.T. In the repair-proficient mutants from G1, 47% were. In mutants from the repair-proficient cells irradiated in S, 71% (10 of 14) of the premutagenic lesions were located in the transcribed strand; with mutants from such cells irradiated in G1, only 20% (3 of 15) were. In contrast, there was no statistically significant difference in the fraction of premutagenic lesions located in the transcribed strand of the XP12BE cells; approximately 75% (24 of 32) of the premutagenic lesions were located in that strand, i.e., 15 of 19 (79%) in the S-phase cells and 9 of 13 (69%) in the G1-phase cells. The switch in strand bias supports preferential nucleotide excision repair of UV-induced damage in the transcribed strand of the HPRT gene.

Base Sequence↗
PubMed1991

Pharmacological studies of meisoindigo: absorption and mechanism of action.

Meisoindigo, an indirubin derivative, is a new type of cancer chemotherapeutic agent. It exhibited higher activity against rodent tumors than indirubin itself. Experiments have shown the improved absorption of meisoindigo, compared to indirubin to be one of the major reasons for the enhancement of antitumor activity. Studies on the mechanism of meisoindigo action indicate that it strongly inhibits DNA biosynthesis in tumor cells. Strong inhibition of the drug on assembly of microtubule protein was also obtained. By means of FCM technique the effects of meisoindigo on mouse leukemia L1210 cell cycle were examined. Experimental results showed that under the action of meisoindigo the S phase cells accumulated and the traverse of the cells in G2 + M phase to G1 phase may also be blocked to some extent.

Animals↗
PubMed1990

Effect of excision repair by diploid human fibroblasts on the kinds and locations of mutations induced by (+/-)-7 beta,8 alpha-dihydroxy-9 alpha,10 alpha-epoxy-7,8,9,10- tetrahydrobenzo[a]pyrene in the coding region of the HPRT gene.

(+/-)-7 beta,8 alpha-Dihydroxy-9 alpha,10 alpha-epoxy-7,8,9,10- tetrahydrobenzo[a]pyrene (BPDE) is a direct-acting carcinogen that forms DNA adducts only with purines, predominantly (greater than 95%) with guanine. To investigate the effect of nucleotide excision repair on the kinds and locations (spectra) of mutations induced in diploid human fibroblasts by BPDE, we synchronized cells and exposed them to BPDE either at the beginning of S phase just when the target gene hypoxanthine (guanine) phosphoribosyltransferase (HPRT) is replicated or 12 hr prior to the beginning of S phase (early G1 phase). Clones resistant to 6-thioguanine were isolated, and the mRNA in lysates of 100-500 cells from each mutant clone was used to synthesize cDNA. HPRT cDNA was amplified 10(11)-fold by the polymerase chain reaction and then sequenced directly. The mutants derived from the two populations did not differ in the kinds of mutations; 19/20 of the base substitutions in cells taken from S phase and 19/19 of those from G1 phase involved G.C base pairs, predominantly G.C----T.A. However, they differed significantly in the distribution of the mutations in the coding region of the gene. In the cells from G1 phase, 29% of the mutations were clustered within a unique run of six guanine bases; in the S-phase cells, only 4% were located there. Assuming that the premutagenic BPDE-induced lesions involved purines, in the cells treated at the beginning of S phase, 24% of these lesions were located in the transcribed strand, whereas in the G1-treated cells, none were. This suggests that in the HPRT gene of diploid human cells excision repair of BPDE adducts occurs preferentially on the transcribed strand.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗
PubMed1990

Identification of Xenopus S6 protein kinase homologs (pp90rsk) in somatic cells: phosphorylation and activation during initiation of cell proliferation.

We have identified human, mouse, and chicken homologs to Xenopus S6 protein kinase II (S6KII). In quiescent cells, the apparent molecular mass of the Xenopus homologs (referred to as pp90rsk) increased from a range of 81 to 91 to a range of 85 to 92 kilodaltons following serum addition, which is consistent with an increase in protein phosphorylation. Indeed, serum growth factors stimulated pp90rsk phosphorylation at multiple serine and threonine residues. Furthermore, pp90rsk activity was stimulated within seconds of serum addition. Distinct molecular sizes, chromatographic properties, phosphopeptide maps, and kinetics of activation, the lack of immunological cross-reactivity, and analysis of S6 kinase activities in cells that overexpressed pp90rsk suggest that pp90rsk and pp70-S6 protein kinase, a previously identified mitogen- and oncogene-regulated S6 kinase in cultured cells, are distinct and differentially regulated. The notion that both enzymes are regulated by protein phosphorylation was supported by the ability to inactivate their S6 phosphotransferase activities with potato acid phosphatase. These data demonstrate that homologs to the Xenopus S6 protein kinases are produced and regulated by protein phosphorylation in somatic cells and that, in addition to a proposed role in Xenopus oocyte maturation, these homologs may participate in the initiation of animal cell proliferation.

Acid Phosphatase↗
PubMed1990

[Ultrastructural studies of visual cells in the human fetus].

The retinas of 15 human fetuses 14 weeks to full gestation were observed by transmission electron microscopy, with the findings that rod cells and cone cells were indistinguishable in the 3rd month of fetal age, and the outer segments were not developed while the centriole was discernible in the apex of the ellipsoid. During the 4th month of gestation, cilia grew from the centriole towards the pigment epithelium. In the 5th month, primordia of the outer segment began to form. The outer segment continued to elongate during the 6th month, but the membranous discs in the outer segment were in disorder and not uniform. During the 7th month, the outer segment increased in length, the cell increased in size and the cell organelles increased in number. After the 8th month of gestation, old membranous discs began to come off for neogenesis.

Fetus↗
PubMed1989

Isolation of an acidic phospholipase A2 from the venom of Agkistrodon acutus (five pace snake) and its effect on platelet aggregation.

A phospholipase A2 from the venom of the snake Agkistrodon acutus was purified by immunoaffinity chromatography and fast protein liquid chromatography (FPLC) as a single band by PAGE and SDS-PAGE. The estimated mol.wt was 16,400 by SDS-PAGE and 16,900 by gel filtration and the isoelectric point was 4.9. The ten N-terminal amino acid residues are homologous to those of the acidic phospholipases A2 from other crotalid venoms. The purified enzyme showed a potent inhibitory effect on platelet aggregation induced by ADP, collagen and sodium arachidonate in human platelet-rich plasma. The platelet aggregation by these inducers was completely suppressed when the concentration of the venom phospholipase A2 was 10-100 micrograms/ml. However, at 20 micrograms/ml, platelet aggregation could be elicited in washed human platelet suspension. Aspirin (28 micrograms/ml), an inhibitor of cyclooxygenase, inhibited the aggregating effect of the phospholipase A2. It is proposed that the stimulatory mechanism of the phospholipase A2 might be due to the liberation of arachidonic acid from phospholipids in the membrane of platelets and the formation of thromboxane A2.

Amino Acids↗
PubMed1989

Pacidamycins, a novel series of antibiotics with anti-Pseudomonas aeruginosa activity. I. Taxonomy of the producing organism and fermentation.

The pacidamycins are a new complex of nucleosidyl-peptide antibiotics with highly specific activity against Pseudomonas aeruginosa. They are produced by Streptomyces coeruleorubidus AB 1183F-64 which was isolated from a soil sample collected at Offenburg in the FRG. The mature spore masses of the producing organism are greenish gray to blue, and the spore chains are arranged in spirals. After the structures of the pacidamycins were determined, the fermentation medium was supplemented with component amino acids. This resulted in the directed biosynthesis of several members of the complex. The overall antibiotic recovered was increased from 1 approximately 2 mg/liter to more than 100 mg/liter through a combination of strain selection, medium manipulation and amino acid feeding experiments.

Anti-Bacterial Agents↗
PubMed1989

Pacidamycins, a novel series of antibiotics with anti-Pseudomonas aeruginosa activity. II. Isolation and structural elucidation.

A novel class of antibiotics was isolated from cultures of Streptomyces coeruleorubidus strain AB 1183F-64. The antimicrobial activity of the pacidamycins is selective against Pseudomonas aeruginosa. The various congeners are nucleoside peptides which differ in the terminal amino acid residues. The structures were determined using MS-MS and 2D NMR techniques.

Anti-Bacterial Agents↗
PubMed1989

Pacidamycins, a novel series of antibiotics with anti-Pseudomonas aeruginosa activity. III. Microbiologic profile.

Pacidamycins are nucleosidyl-peptide antibiotics which have activity only against Pseudomonas aeruginosa. Their MICs for other organisms such as Enterobacteriaceae, Staphylococcus aureus, most Streptococci and other Pseudomonas species are greater than 100 micrograms/ml. These compounds had no activity against erythromycin-susceptible Streptococci. The MICs for Streptococcus pyogenes with constitutive- and inducible-type of macrolide-lincosamide-streptogramin resistance were 12.5 and 25 micrograms/ml, respectively. The MICs against P. aeruginosa ranged from 8 to 64 micrograms/ml. The activity of these compounds was 1 to 2-fold less in serum than broth. Time-kill curves were performed using 4 and 8 times the MIC of pacidamycin 1. It was bactericidal against P. aeruginosa (3 log10 decrease in 4 to 6 hours). At 24 hours, resistant mutants were found in the cultures. The MICs of piperacillin and gentamicin for these mutants were the same as for the parent strain. The frequency of resistance to these compounds was less than 3.5 x 10(-6). The resistant mutants were stable after 10 transfers in antibiotic-free medium. The pacidamycins were inactive against P. aeruginosa in mouse protection test. After a single subcutaneous injection of 25 mg/kg of pacidamycin 1, the Cmax was approximately 50 micrograms/ml and the serum half-life was 0.5 hour.

Animals↗
PubMed1989

Coumamidines, new broad spectrum antibiotics of the cinodine type. III. Microbiologic activity of coumamidine gamma 1.

The coumamidines are novel antibiotics with activity against a wide spectrum of aerobic Gram-positive and Gram-negative bacteria. All microbiological studies were performed on coumamidine gamma 1. The MIC90s (micrograms/ml) of coumamidine are as follows: Staphylococcus aureus 1.0, Streptococcus pyogenes 8, Enterobacteriaceae 2.0, Pseudomonas aeruginosa 8, Campylobacter jejuni and Campylobacter coli 1, Legionella pneumophila 8, Haemophilus influenzae 0.5, Neisseria gonorrhoeae 0.5. Coumamidine had MICs ranging from 8 to greater 0.5, Neisseria gonorrhoeae 0.5. Coumamidine had MICs ranging from 8 to greater than 64 for most anaerobes, except some Peptostreptococcus strains. The aminoglycoside super-sensitive strain, P. aeruginosa BMH 10, was also super-sensitive to coumamidine (MIC 0.2 micrograms/ml). Coumamidine was rapidly bactericidal for S. aureus. The viable bacterial count in logarithmic phase cultures was reduced to less than 10 cfu within 2 hours after exposure to 4 times the MIC (3.12 micrograms/ml) of coumamidine. The frequency of resistance development was less than 1 X 10(-9) for Escherichia coli and S. aureus when selected at 4 and 8 times the MIC. The Cmax in mouse serum after a single subcutaneous dose of 25 mg/kg of coumamidine was 4.5 micrograms/ml and t1/2 was 1 hour. Coumamidine is stable in serum. In mouse protection tests against S. aureus NCTC 10649 the ED50 was less than 0.6 mg/kg/day when it was administered subcutaneously at 1 and 5 hours after infection. Coumamidine was not absorbed after oral administration. The antibacterial spectrum, bactericidal activity, stability in serum and low frequency of resistance make this an interesting new class of antibiotics.

Aminoglycosides↗
PubMed1989

[Ultrastructural observations on the fetal corneal epithelium].

The maturing changes in the central corneal epithelium of 18 fetuses were observed by transmission electron microscopy, revealing that the corneal epithelium comprised 5-6 layers of cells after the 8th month of gestation, when the basal cells were short columnar in shape with oval, regular nuclei, and the cytoplasmic organelles included mitochondria. Golgi complexes, rough endoplasmic reticulum, membrane coating granules, glycogen granules etc. In the middle layers were polygonal cells with flat nuclei and few cytoplasmic organelles. The free surfaces of the superficial flat cells were covered with microvilli of different heights and in random arrangement. The study confirmed that ultrastructurally the fetal corneal epithelium after the 8th month achieved the mature form of the adult cornea.

Cornea↗
PubMed1988

Premature termination of variable gene rearrangement in B lymphocytes from X-linked agammaglobulinemia.

X-linked agammaglobulinemia (XLA) results from failure of B lymphocyte development. Immature B cells from a patient with XLA were found to produce truncated mu and delta immunoglobulin H chains encoded by D-JH-C (mu delta). The 5' terminal sequence of cDNA encoding the H chains is composed of D-JH with the characteristic GGTTTGAAG/CACTGTG consensus sequence utilized for VH gene rearrangement upstream, and a leader sequence that serves for translation of this intermediate stage of rearrangement. Failure of variable region gene rearrangement may underlie the failure of B lymphoid development in XLA.

Agammaglobulinemia↗
PubMed1988

Phenelfamycins, a novel complex of elfamycin-type antibiotics. II. Isolation and structure determination.

A novel complex of elfamycin-type antibiotics has been isolated from submerged fermentation of either Streptomyces violaceoniger AB 999F-80 or Streptomyces violaceoniger AB 1047T-33. Antibiotics were extracted from the fermentation broth with ethyl acetate and from the mycelia with acetone. Purification of individual components was achieved by a combination of solvent partitions, Sephadex LH-20 exclusion, C18 bonded-phase silica gel adsorption, diol partition and liquid-liquid countercurrent chromatographies. Seven closely related components were separated and assigned structures 4, 11, 12, 13, 14, and 16 to phenelfamycins A to F respectively and structure 17 to unphenelfamycin. These structures were elucidated employing a variety of spectroscopic techniques, including extensive use of 1D and 2D NMR spectroscopy.

Aminoglycosides↗
PubMed1987

The structural characterization of endogenous factors from Vinca rosea crown gall tumors that promote cell division of tobacco cells.

The ability of two compounds, a cytokinin and an auxin, to stimulate tobacco cell growth and differentiation has been known for greater than 30 years, but the molecular mechanism of this activation is still unknown. Previous reports of factors endogenous in crown gall tumors of Vinca rosea that could replace the cytokinin requirement in tobacco cell culture has motivated an investigation of these tissues. The optimization of a reverse-phase isolation scheme has led to the purification of sufficient material to allow for the identification of six different related compounds. The structures of two of these compounds have been assigned as a set of epimeric dehydrodiconiferyl alcohol beta-D-glycosides. The structure of these compounds suggests that they would most likely be derived from the plant cell wall.

Cell Division↗