Search PubMed⌕ Search

Biomedical subjects

M Ling

Publications and source records attributed to M Ling.

At least 55 records · Page 3Linked to original sources
PubMed1997

The human mitochondrial elongation factor tu (EF-Tu) gene: cDNA sequence, genomic localization, genomic structure, and identification of a pseudogene.

The human mitochondrial elongation factor Tu (EF-Tu) is nuclear-encoded and functions in the translational apparatus of mitochondria. The complete human EF-Tu cDNA sequence of 1677 base pairs (bp) with a 101 bp 5'-untranslated region, a 1368 bp coding region, and a 207 bp 3'-untranslated region, has been determined and updated. The predicted protein from this cDNA sequence is approximately 49.8 kDa in size and is composed of 455 amino acids (aa) with a putative N-terminal mitochondrial leader sequence of approximately 50 aa residues. The predicted amino acid sequence shows high similarity to other EF-Tu protein sequences from ox, yeast, and bacteria, and also shows limited similarity to human cystolic elongation factor 1 alpha. The complete size of this cDNA (1677 bp) obtained by cloning and sequencing was confirmed by Northern blot analysis, which showed a single transcript (mRNA) of approximately 1.7 kb in human liver. The genomic structure of this EF-Tu gene has been determined for the first time. This gene contains nine introns with a predicted size of approximately 3.6 kilobases (kb) and has been mapped to chromosome 16p11.2. In addition, an intronless pseudogene of approximately 1.7 kb with 92.6% nucleotide sequence similarity to the EF-Tu gene has also been identified and mapped to chromosome 17q11.2.

Amino Acid Sequence↗
PubMed1997

Cloning, characterization, and chromosomal localization of human liver form cytochrome c oxidase subunit VIa related genes.

The chromosomal location of human cytochrome c oxidase (COX) subunit VIa Liver (VIa-L) isoform related sequences has been determined by a combination of in situ hybridization and analysis of human-hamster somatic cell hybrid panels. COX VIa-L related sequences were present on chromosomes 6 and 12. It has been verified that at least two COX VIa-L genes are on chromosome 6, one of which is a pseudogene. In total, four COX VIa-L related sequences have been cloned and their nucleotide sequences analyzed. At least three of the sequences represent pseudogenes; their relatedness to the COX VIa-L cDNA is discussed.

Animals↗
PubMed1997

The effects of hypoxia on the bioenergetics of liver in situ in chronic ethanol-treated rats: a noninvasive in vivo 31P magnetic resonance spectroscopy study.

OBJECTIVE: The effects of hypoxia on bioenergetic parameters of the livers of rats with and without chronic ethanol (CE) treatment has been studied in a totally noninvasive manner, using in vivo 31P magnetic resonance spectroscopy (MRS). METHOD: The effects of hypoxia (10% oxygen for 55 minutes) on the levels of ATP, inorganic phosphate (Pi), phosphomonoesters (PME) and phosphodiesters (PDE) were followed with time. The CE and control rats (four pairs) were starved for 16-20 hours to eliminate the effects of any ethanol still present within the liver. To complement the in vivo studies, perchloric acid extracts from rats (four pairs for each stage) at the three stages, initial (normoxic), hypoxic and recovery, were analyzed using in vitro high resolution 31P MRS. RESULTS: After hypoxic challenge, the hepatic Pi and PME levels increased much more significantly and more quickly for the CE rats compared to those of controls, and ATP levels decreased more dramatically for CE rats. The recoveries of Pi and PME levels after return to normoxic conditions were incomplete for the CE rats in contrast to the controls. The hepatic extract analyses showed that several different PMEs increased during hypoxia, including alpha-glycerophosphate, phosphocholine, AMP, IMP and glycolytic metabolites, and that total hepatic Pi increased during hypoxia in addition to cytoplasmic Pi. CONCLUSIONS: CE-treated rat livers in situ are bioenergetically more sensitive to a hypoxic challenge than are control rat livers, even in the absence of endogenous ethanol. This is consistent with a model of CE-induced liver damage in which CE-treated livers already have centrolobular hypoxia under normoxic conditions and cannot adequately compensate for additional hypoxic stress.

Adenosine Triphosphate↗
PubMed1996

FTIR studies of recombinant human granulocyte-macrophage colony-stimulating factor in aqueous solutions: secondary structure, disulfide reduction and thermal behavior.

Fourier transform infrared spectroscopy (FTIR) has been used to investigate the secondary structure, disulfide reduction and thermal behavior of recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) in aqueous solutions. The contributions of amino-acid side-chain groups to the amide I bands of rhGM-CSF in H2O and in D2O solutions were carefully scrutinized, as 40% of the total 127 amino-acid residues of rhGM-CSF is side-chain absorptive (asparagine, glutamine, etc.). The FTIR results indicated that rhGM-CSF is composed of 46% alpha-helix, 7% beta-sheet, 23% turn and 24% loop/irregular structures which are in good agreement with the X-ray diffractional data. Reduction of rhGM-CSF with dithiothreitol caused apparent unfolding of the native conformation followed by the time-dependent increase of beta-aggregation bands which arose at 1622 and 1693 cm(-1) in H2O, 1613 and 1684 cm(-1) in D2O solutions. The result also showed that tertiary structure can change independently of the secondary structure. Thermal denaturation of rhGM-CSF took place at 55 to 70 degrees C and the denatured protein adopted an irregular structure as revealed by the FTIR spectra. The thermal denaturation did not show the formation of intermolecular beta-aggregates which is typical of most thermal denatured proteins. Moreover, it is partly reversible, indicating a special thermal stability of rhGM-CSF.

Deuterium Oxide↗
PubMed1996

Disorders of pyruvate carboxylase and the pyruvate dehydrogenase complex.

The most common defect associated with deficiency of the pyruvate dehydrogenase (PDH) complex occurs in the E1 component, specifically due to mutations in the X-linked E1 alpha gene. Clinical sequelae of these mutations, which range from severe neonatal lactic acidosis to carbohydrate-sensitive ataxia, can be different in males and females depending on the nature of the mutation and, in the case of females, on the X-inactivation pattern in different tissues. Males have a high representation of missense mutations among the patient cohort, while females are much more likely to have DNA rearrangements, particularly toward the 3' end of the coding sequence of the gene. Missplicing mutations involving exon 6 deletion have been reported, as has a missense mutation conferring true thiamin-responsiveness of the enzyme and the patient's clinical symptoms. Pyruvate carboxylase deficiency, on the other hand, is a true autosomal recessive disease, though it has high occurrences in particular ethnic groups, especially in Algonkian-speaking Amerindians and in Arabs. In the former group the defect is a simple type in which material cross-reactive to pyruvate carboxylase antibody is present in cultured cells (CRM+ve). In the latter group, cross-reacting material is rarely present (CRM-ve). The CRM+ve patients can survive into teenage years with careful supervision, while the CRM-ve patients have complications due to hyperammonaemia and dysfunction of the urea cycle and rarely survive beyond 3 months of life.

Amino Acid Sequence↗
PubMed1995

The development of a behavioural science course into the dental curriculum at Liverpool Dental School.

The behavioural science course for the undergraduate dental curriculum at Liverpool university Dental School is designed to be integrated into the basic science and clinical teaching. The course is taught by a clinical psychologist and social anthropologist. The course has an input to each of the 5 years of the student's training. This paper presents three examples (community dental visits, communication skills training and special symposia) of how the course is being developed, illustrating the strengths and some shortcomings requiring further development.

Behavioral Sciences↗
PubMed1995

A one-step polymerase chain reaction site-directed mutagenesis method for large gene-cassettes with high efficiency, yield, and fidelity.

A one-step three-stage efficient PCR method for site-directed mutagenesis has been developed. This method involves one PCR reaction with no need to add more reagents during this reaction or to do a second PCR reaction. This method utilizes a three-stage PCR cycling profile, a ddNTP-blocked restriction endonuclease fragment, and Pfu DNA polymerase. This method allows the amplification of at least 2 kb of final mutant product via an intermediate megaprimer as large as 1.3 kb with high fidelity, high efficiency, high yield, and high success rate. A 3.2-kb large final mutant product has also been obtained by using Taq and Taq Extender (Stratagene) instead of Pfu. This method is particularly useful when there are no unique restriction sites in the gene of interest to reduce the large sizes of the final mutant product and/or the megaprimer.

DNA Primers↗
PubMed1995

A component of the medicinal herb ephedra blocks activation in the classical and alternative pathways of complement.

Extracts of the herb Ephedra sinica have long been used in traditional Chinese medicine for the treatment of, among other conditions, acute nephritis. In preliminary studies it was shown that extracts of ephedra caused inhibition of complement in vitro. We thus set out to isolate the active component(s) of this herb, to examine the complement-inhibiting capacity in sera from different species, and to characterize the mechanism(s) by which it inhibits complement. Aqueous extraction of the herb followed by fractionation using thin layer chromatography (TLC) demonstrated that complement-inhibiting activity resided within a single band, hereafter termed the complement-inhibiting component (CIC), which represents an as yet uncharacterized polyanionic carbohydrate molecule. TLC-purified CIC inhibited the classical complement pathway in all species tested (human, pig, guinea pig, rat and rabbit). Using erythrocyte intermediates and sera specifically depleted of individual components it was apparent that CIC inhibited C2. This finding was confirmed using purified human C2, CIC causing a dose-dependent loss of C2 haemolytic activity. At much higher doses, CIC also showed some inhibiting effect in the terminal pathway, and this was shown to be due to inhibition of C9. In the alternative pathway CIC also showed inhibitory activity, although its site of action in this pathway remains unresolved. In Chinese medicine the herb is taken orally, though no studies of complement levels in patients taking the herb have been reported. Preliminary data indicate that oral administration in rats causes a partial inhibition of serum complement activity. Given the current enthusiasm for complement inhibition as a therapy for inflammatory diseases, this non-toxic, naturally occurring agent might be of therapeutic value.

Animals↗
PubMed1995

Purification of recombinant human granulocyte-macrophage colony stimulating factor expressed in Escherichia coli.

Recombinant human granulocyte-macrophage colony stimulating factor (rhGM-CSF) was expressed as inclusion bodies (IB) in E. coli. A simple and effective protocol has been worked out for the purification. IB collected after the breakage of bacteria through sonication were subjected to repeated washing followed by solubilization in TE buffer (50 mmol/L of Tris.HCl, 1 mmol/L of EDTA, pH 8.3) containing 8 mol/L urea and 10 mmol/L DL-dithiothreitol. By means of Sephacryl-200 HR, refolding, and Q Sepharose Fast Flow, rhGM-CSF was obtained with a purity of 99%. The total protein recovery was 10% and specific activity of rhGM-CSF was 1 x 10(7) u/mg. The sequence of N-terminal 16 amino acid residues of purified rhGM-CSF was determined and found to be identical to the native protein. This study provided useful parameters for mass production of rhGM-CSF.

Amino Acid Sequence↗
PubMed1994

Sequence analysis identifies the proline dehydrogenase and delta 1-pyrroline-5-carboxylate dehydrogenase domains of the multifunctional Escherichia coli PutA protein.

The PutA protein of Escherichia coli has two enzymatic activities: proline dehydrogenase (PDH) and delta 1-pyrroline-5-carboxylate dehydrogenase (P5CDH). It associates with the cytoplasmic membrane as PDH and P5CDH and with put control region DNA as put repressor. Reduction of the PutA flavin by proline, a PutA conformational change and association of PutA with membranes are coincident. The nucleotide base sequence of E. coli putA was determined, that of S. typhimurium putA was updated and the deduced PutA protein sequences were surveyed for catalytic domains and ligand binding sites. The two sequences were very similar (80.5% and 95% on the nucleic acid and protein levels, respectively). Residues 650 through 1130 of PutA were very similar to the sequences of P5C dehydrogenases and aldehyde dehydrogenases from both prokaryotes and eukaryotes. Glutamate 883 and cysteine 917 of PutA were conserved with the corresponding residues in P5C dehydrogenases and with those proposed to be active site residues in the aldehyde dehydrogenases. Those relationships suggest that gamma-glutamic semialdehyde, believed to equilibrate spontaneously with P5C, is the substrate for P5C dehydrogenases. Residues 340 through 590 of PutA were similar in sequence to proline dehydrogenases from Saccharomyces cerevisiae and Drosophila melanogaster. Limited similarities were also found between residues 315 through 357 of PutA and a consensus sequence near a putative active site and FAD-binding region shared by succinate dehydrogenase sequences from several organisms. Since residues 228 through 358 of PutA were similar in sequence to several serine-pyruvate aminotransferases, PutA is proposed to catalyze the hydrolysis of P5C (a Schiff's base intermediate) to gamma-glutamic semialdehyde. A carboxyl-terminal sequence that resembles a leucine zipper motif may be involved in association of PutA with put control region DNA.

1-Pyrroline-5-Carboxylate Dehydrogenase↗
PubMed1993

The water-soluble component of Salvia miltiorrhiza Bge can promote macrophage synthesis of C4 in vivo and in vitro.

A water-soluble component extracted from Salvia miltiorrhiza Bge has been used for many years in traditional Chinese medicine ('huo xie hua yu') for the treatment of the hyperviscosity syndrome. We demonstrate here that this extract promotes macrophage proliferation in vitro and enhances macrophage synthesis of the C4 component of complement. No increase in synthesis of other complement components by macrophages was observed. In a group of 20 patients with the hyperviscosity syndrome treated with an aqueous extract of Salvia, serum levels of C4 rose significantly over the course of treatment (mean pretreatment level 37.4 mg%, mean posttreatment level, 43.5 mg%; p < 0.01). The rise in C4 levels closely correlated with the disappearance of the patients' clinical symptoms.

Blood Viscosity↗
PubMed1992

Ethanol-induced fatty liver in the rat examined by in vivo 1H chemical shift selective magnetic resonance imaging and localized spectroscopic methods.

In vivo 1H magnetic resonance imaging (MRI), chemical shift selective imaging (CSI), and localized (VOSY) 1H magnetic resonance spectroscopy (MRS) were used to study fatty infiltration in the livers of rats chronically fed an ethanol-containing all-liquid DeCarli-Lieber diet. Conventional total proton MRI showed a somewhat hyperintense liver for ethanol-fed rats, compared with pair-fed controls. CSI showed a dramatic increase in the fat signal intensity for ethanol-treated rats that was fairly homogeneous throughout the liver. However, CSI also showed a substantial decrease in the water signal intensity for the ethanol-treated rats compared to pair-fed control rats. 1H VOSY MR spectra also showed a 5.5-fold increase in the methylene resonance (1.3 ppm) of fat and a 50-70% decrease in the water resonance (4.8 ppm). Relative in vivo proton T1 and T2 relaxation times for the water resonance separate from the fat resonance, determined from modified VOSY experiments, were found to tend to increase and decrease, respectively, for ethanol-treated rat livers compared with controls. The decrease in hepatic water signal intensity could be accounted for by the decrease in T2 and decrease in water density due to the presence of accumulated hepatic fat (approximately 25 mg/g wet weight of liver). When ethanol was withdrawn from the chronically treated rats, fatty infiltration was observed by both CSI and VOSY spectra to revert toward control values with a half-life of 2-4 days. By day 16, however, the signal intensity for hepatic fat was still significantly higher than control levels. In vitro 1H MRS studies of chloroform-methanol extracts confirmed the 5.5-fold increase in total hepatic fat induced by the chronic ethanol treatment, and showed further that triacylglycerols were increased 7.7-fold, cholesterol was increased fourfold, and phospholipids were increased 3.3-fold, compared with liver extracts from pair-fed control rats.

Animals↗
PubMed1990

[Isolation and characterization of clonal cell lines with different metastatic capacity from hepato-carcinoma in mice].

By means of soft agar colonization and limiting dilution, 5 clonal cells with different metastatic capacity, [one with high metastatic rate (78.6-89.4%), three moderate (33.3-66.6%) and one low (15-15.4%)], were separated from Hca-F25/L cell line. Their common features were: All clones metastasize to lymphatic system except a small percentage (1.88%) associated with lung metastasis; five clones as well as their parents all reacted with Keratin monoclonal antibody (HK2), which indicated the epithelial character with poor differentiation; Cells of each clone have highly identical karyotype. Five clone lines have similar marker chromosomes (M1, M2, M3, M4) with different stem line karyotypes. We also found different features of five cell lines in pathomorphological indicators and cell electrophoresis rate. Our results indicated that the Hca-F25/L strain is heterogeneous in phenotype and karyotype.

Animals↗
PubMed1987

Hyperparathyroidism: comparison of MR imaging with radionuclide scanning.

Twenty-three patients with hyperparathyroidism were evaluated preoperatively with magnetic resonance (MR) imaging. Twenty patients also underwent thallium-201/technetium-99m scintigraphy. Of 22 patients with primary hyperparathyroidism, 12 had persistent or recurrent disease. One had secondary hyperparathyroidism due to end-stage renal disease. MR imaging allowed accurate localization of abnormal parathyroid glands in 64% evaluated prospectively and 82% evaluated retrospectively. Scintigraphy allowed localization of 60% evaluated prospectively and 70% retrospectively. The two imaging modalities together allowed detection of 68% evaluated prospectively and 91% retrospectively. MR imaging allowed detection of two of five mediastinal adenomas evaluated prospectively and four of five retrospectively. In patients who underwent both imaging studies, MR was more successful in those with previous neck surgery (73% evaluated prospectively and 91% retrospectively) than in those with no prior surgery (57% prospectively and 71% retrospectively). Scintigraphy allowed accurate localization in 64% evaluated prospectively and 64% retrospectively in patients with previous surgery versus 57% prospectively and 86% retrospectively in patients with no prior neck surgery. Four false-positive results were obtained with MR imaging and three with scintigraphy. MR imaging was useful for parathyroid localization in patients with hyperparathyroidism, particularly in patients requiring additional surgery.

Adult↗