Biomedical subjects

J Lu

Publications and source records attributed to J Lu.

At least 595 records · Page 33Linked to original sources

PubMed1993

Inhibition kinetics of chloramphenicol acetyltransferase by selected detergents.

Kinetic analyses indicate that the inhibitory effects of the nonionic detergents Triton X-100 and Nonidet P-40 on chloramphenicol acetyltransferase are exerted by a competitive and a non-competitive mechanism with respect to the substrates chloramphenicol and acetyl-CoA, respectively. Comparison with nonionic detergents without an aromatic moiety like that present in Triton X-100 and Nonidet P-40 suggests that the aromatic groups in these two detergents may compete with chloramphenicol for binding to the hydrophobic, active site in the chloramphenicol acetyltransferase.

Acetyl Coenzyme A↗

PubMed1993

IA-1, a new marker for neuroendocrine differentiation in human lung cancer cell lines.

IA-1 is a recently isolated novel complementary DNA which encodes a protein of 510 amino acids that contains both a zinc finger DNA-binding domain and a putative prohormone domain. mRNA expression of IA-1 has been found thus far only in tumors of neuroendocrine origin. In this report we describe the expression of IA-1 mRNA in a panel of 64 human lung cancer cell lines. IA-1 mRNA was detected by Northern blot analysis in 97% (30 of 31) of small cell lung cancer cell lines. In contrast, IA-1 mRNA was detected in only 13% (4 of 30) of non-small cell lung cancer cell lines. Nine of the 30 (30%) expressed either chromogranin A mRNA or produced L-dopa decarboxylase. Four of these 9 (44%) had detectable levels of IA-1 mRNA. In most of the lung cancer cell lines examined, IA-1 showed high concordance with the other neuroendocrine markers, L-dopa decarboxylase, and chromogranin A. The one exception was a variant small cell lung cancer cell line which expressed low or nondetectable levels of L-dopa decarboxylase. IA-1 is a candidate marker of neuroendocrine differentiation of human lung tumors.

Biomarkers, Tumor↗

PubMed1993

Structural similarity between lung surfactant protein D and conglutinin. Two distinct, C-type lectins containing collagen-like sequences.

Preparations of bovine lung surfactant D (SP-D) and conglutinin were examined by electron microscopy, gel-filtration and SDS/PAGE. SP-D is composed of non-covalently linked subunits, of 160 kDa, which each contain three, disulphide-linked, 44-kDa polypeptide chains. In the electron microscope a single 160-kDa subunit of SP-D appears as a 45.8 +/- 3-nm-long rod connected to a small globular 'head'. Particles were also seen which correspond to non-covalently linked dimers, trimers and tetramers of the 160-kDa monomer subunit of SP-D. The tetramer structure contains 12 polypeptide chains and is very similar to the electron microscopy images and model reported by Strang et al. [Strang, C. J., Slayter, US., Lachmann, P. J. and Davis, A. E. (1986) Biochem. J. 236, 3811-389] for bovine conglutinin in which four 160-kDa subunits are disulphide-linked to give a molecule of expected molecular mass of 528 kDa. This study confirmed the findings by Strang et al. in the above paper for intact conglutinin and also emphasised that the rod-like structures, of length 37.6 +/- 3.7 nm, seen in the conglutinin subunits were significantly shorter than those in SP-D despite the close similarity in amino acid sequence (79% identify) and chain length between the two proteins. In addition, a truncated form of conglutinin was found in the conglutinin preparations, due to limited proteolysis of the Arg-Ala bond at position 54 in the 44-kDa chains. These truncated conglutinin chains yield a subunit composed of three shortened, non-disulphide-linked, chains and this subunit appears as a monomer with a rod length of 34.2 +/- 2.8 nm in the electron microscope. On gel-filtration, a proportion of the SP-D preparation behaved, as expected, as a molecule with an apparent molecular mass of 600 kDa. The remainder of the SP-D preparation behaved as aggregated material with a molecular mass greater than 900 kDa which yielded no distinct structures in the electron microscope. Intact conglutinin was eluted at a position greater than 900 kDa but yet provided clear electron microscopy images of the tetramer structure described above.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗

PubMed1993

Spontaneous nucleosomal DNA fragmentation in murine leukemic L1210 cells.

Spontaneous nucleosomal DNA fragmentation (SNDF), characteristic of apoptotic cell death was observed in a murine leukemic cell line grown to high density. This effect was eliminated by permitting free diffusional exchange with a large amount of complete medium. Cells seeded at low density did not show SNDF in a conditioned medium that had been used to culture cells with severe SNDF. Cells seeded at high density in the same conditioned medium showed pronounced SNDF, but addition of 10% fetal bovine serum to such a medium significantly reduced SNDF. These observations indicate that neither physical contact among cells in high density culture per se nor secretion of a death factor by the high density culture could account for the induction of SNDF. The results support a model in which the gradual depletion of a serum factor(s) leads to the activation of a cell death program that is manifested as SNDF.

Animals↗

PubMed1993

PEBP2/PEA2 represents a family of transcription factors homologous to the products of the Drosophila runt gene and the human AML1 gene.

cDNAs representing the alpha subunit of polyomavirus enhancer binding protein 2 (PEBP2; also called PEA2) were isolated. The products of the cDNAs are highly homologous to that of Drosophila segmentation gene runt (run) for an N-proximal 128-amino acid region showing 66% identity. The run homology region encompasses the domain capable of binding to a specific nucleotide sequence motif and of dimerizing with the companion beta subunit. The human AML1 gene related to t(8;21) acute myeloid leukemia also had a run homology region. Together with the beta subunit, which increases the affinity of the alpha subunit to DNA without binding to DNA by itself, PEBP2 represents a newly discovered family of transcription factor. The major species of PEBP2 alpha mRNA was expressed in T-cell lines but not in B-cell lines tested. Evidence indicated that PEBP2 functions as a transcriptional activator and is involved in regulation of T-cell-specific gene expression.

Amino Acid Sequence↗

PubMed1993

The cDNA cloning of conglutinin and identification of liver as a primary site of synthesis of conglutinin in members of the Bovidae.

Bovine conglutinin is a collagen-like, C-type, plasma lectin which belongs to the group of proteins called 'collectins'. Two inosine-containing oligonucleotides were synthesized, based on the published protein sequence for bovine conglutinin [Lee, Leiby, Allar, Paris, Lerch and Okarma (1991) J. Biol. Chem. 266, 2715-2723], and PCR on target DNA from a bovine liver lambda gt 11 cDNA library yielded a product of the expected size of 210 bp. Screening of the library with this cDNA fragment identified a single positive clone, with an insert of 0.9 kb, coding for bovine conglutinin from residue 70 to the C-terminus. The 5' cDNA sequence, encompassing 150 bp of the 5' non-translated sequence plus the sequence encoding the leader peptide and the N-terminal residues 1-70, was completed by the use of PCR techniques. The cDNA sequence of bovine conglutinin showed 86% identity with that of bovine lung surfactant protein D (SP-D), and the derived amino acid sequence of bovine conglutinin showed 78% identity with that of bovine SP-D, which included complete identity of the leader-peptide sequences. The amino acid sequence derived from the cDNA sequence differs from the published protein sequence at four positions. Northern-blot analysis on total RNA, purified from various tissues from cattle, sheep, humans, rats and mice, showed that a strong signal of approx. 1.8 kb is present in bovine liver RNA. A weak signal of similar size was also observed in sheep liver, but not in human, rat and mouse livers. A weak signal, also of 1.8 kb, is present in the lung RNAs of all the species tested. The signals from the lung tissues are likely to be due to the cross-hybridization of the bovine conglutinin cDNA to the SP-D mRNAs of the respective species. The finding of significant signals in only the bovine and sheep liver RNA samples is indicative that serum conglutinin may be present in significant amounts only in members of the Bovidae (the family encompassing cattle, antelopes, sheep and goats) and closely related species.

Amino Acid Sequence↗

PubMed1993

Enhancement of the stability and activity of aspartase by random and site-directed mutagenesis.

Enzymatic generation of mutant libraries for random mutagenesis of aspartase gene from E. coli J2 was made. A mutant enzyme with 4-fold increase in aspartase activity was found. It is stable at pH7.5-9.0 (wild-type: pH7.0-8.0); heat stability and alpha-helicity are higher than those of the wild-type. By using site directed mutagenesis, the aspartase was activated by replacement of Lys-126 with an arginine residue. The mutation produced functional alterations without appreciable structure changes. The optimum pH for the mutant enzyme is 8.5. The stable pH range is 7.0-9.0. Heat stability is higher than that of the wild-type one. Activity of the mutant enzyme is about 5-fold as much as that of wild-type one.

Amino Acid Sequence↗

PubMed1993

Intrathecal treatment with dextrorphan or ketamine potently reduces pain-related behaviors in a rat model of peripheral mononeuropathy.

The therapeutic effects of dextrorphan and ketamine, two non-competitive N-methyl-D-aspartate (NMDA) receptor antagonists, on neuropathic pain-related behaviors were examined in rats with peripheral mononeuropathy induced by loose ligation of the common sciatic nerve (chronic constrictive injury, CCI). Four daily intrathecal treatments (beginning 1 h after nerve ligation) with dextrorphan or ketamine (12.5-100 nmol) reliably attenuated hyperalgesia to radiant heat and spontaneous pain-related behaviors in CCI rats. Thermal hyperalgesia also was reduced in CCI rats receiving a single intrathecal treatment with either dextrorphan or ketamine (50 and 100 nmol for each compound) on day 3 after nerve ligation when thermal hyperalgesia was well developed. Since both dextrorphan and ketamine are currently utilized in other clinical applications, the results suggest a new therapeutic utility of these 'old' compounds in treatment of neuropathic pain syndromes resulting from peripheral nerve injury.

Animals↗

PubMed1993

Assignment of the human pulmonary surfactant protein D gene (SFTP4) to 10q22-q23 close to the surfactant protein A gene cluster.

Pulmonary surfactant consists of a complex mixture of phospholipids and several proteins essential to normal respiratory function. Two of the surfactant proteins, SP-A and SP-D, appear to have lectin-like activity relevant to the local phagocytic defense. Using polymerase chain reaction (PCR)-based somatic cell hybrid mapping, the human SP-D gene (SFTP4) was assigned to chromosome 10. A regional mapping panel was assembled and characterized using sequence tagged sites for five loci previously mapped to 10q. SFTP4, the SP-A gene (SFTP1), and the microsatellite D10S109 were placed in the interval 10q22-q23. Low-stringency PCR using the SFTP1 primer pair suggested the presence of at least two additional SP-A-related genes in the same region. With the locus for mannose-binding lectin (MBL) at 10q21, this may be indicative of this region's central role in the evolutionary history of carbohydrate-binding proteins containing collagen-like regions.

Animals↗

PubMed1993

Myocardial perfusion scintigraphy in left bundle branch block: a perspective on the issue from image analysis in a clinical context.

Among selected study populations, myocardial perfusion scintigraphy (MPS) in patients with left bundle branch block (LBBB) has been reported to show a low specificity for the diagnosis of coronary disease. However, the stress electrocardiogram (ECG) is nondiagnostic in this setting. To place this method in its appropriate clinical context, we evaluated MPS in all 69 consecutive patients with LBBB studied with scintigraphy for clinical reasons during a 4-year period. Among 32 patients who underwent coronary angiography for clinical indications, per patient sensitivity, 96%; per vessel sensitivity, 84%, 50%, and 100% for left anterior descending (LAD), left circumflex (LCX), and right coronary artery (RCA) involvement, respectively; and per vessel specificity, 95% and 68% for LCX and RCA disease, respectively, were not significantly different from those previously published for the method in patients without LBBB. Although per patient specificity, 38%, and specificity, 39%, for LAD disease were low, the predictive value of a positive test remained relatively high (83%) owing to the small number of patients selected for angiography, in part based on scintigraphic findings, with normal coronary anatomy. In addition to a possible specific pathophysiologic cause related to LBBB, apparent perfusion abnormalities in the LAD distribution may relate to generic conditions that can make scintigraphic interpretation ambiguous, often in the anterior distribution, regardless of the clinical setting. Additionally, the apparent lack of scintigraphic specificity in the LAD distribution could relate in part to a selection bias toward catheterization of patients with induced scintigraphic abnormalities, especially in the LAD distribution.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed1993

Adrenalectomy enhances Fos-like immunoreactivity within the spinal trigeminal nucleus induced by noxious thermal stimulation of the cornea.

Immunocytochemistry was used to assess the distribution of neurons within the spinal trigeminal nucleus that expressed the protein product of the proto-oncogene c-fos after thermal stimulation of the cornea in barbiturate-anesthetized rats. The influence of adrenal steroids on Fos-like immunoreactivity induced by corneal stimulation also was examined by comparison of the results obtained in adrenal intact rats to those in adrenalectomized rats and to those in adrenalectomized rats given corticosterone replacement therapy. Stimuli (42 or 52 degrees C, 20 s per min, 15 min) were applied unilaterally to the cornea by a contact thermode. At 2 h after stimulation animals were perfused with 4% paraformaldehyde and tissue sections were incubated with primary antiserum against the Fos protein and processed with the avidin-biotin method. The pattern of Fos-like immunoreactivity after 52 degrees C stimulation revealed a dominant group of cells ipsilaterally within the superficial laminae of the caudalmost portion of trigeminal subnucleus caudalis that was greatly enhanced in adrenalectomized rats. Low-intensity stimulation did not induce Fos-like immunoreactivity among cells in this caudal region. A second significant group of cells was seen more rostrally at periobex levels within the ventrolateral pole of the nucleus. The number of cells in the periobex grouping was increased after 52 or 42 degrees C corneal stimulation when compared to unstimulated controls and was not affected by levels of corticosterone. The results indicated a discontinuous pattern of Fos-like immunoreactivity within the spinal trigeminal nucleus after thermal stimulation of the cornea and a differential effect of adrenal steroids. The appearance of Fos-like immunoreactivity within caudal portions of the nucleus was increased only by noxious intensities of stimulation and was further enhanced in animals with low levels of corticosterone. In contrast, the Fos-like immunoreactivity at periobex levels was increased after noxious and innocuous thermal stimuli and was independent of the level of corticosterone. The results were consistent with the hypothesis that glucocorticoids modify the expression of immediate early genes among a select group of central trigeminal neurons. Such steroid modulation may contribute to the mechanisms that underlie long-term adaptation to noxious sensory input.

Adrenal Glands↗

PubMed1993

Brain and liver targeted overexpression of O6-methylguanine DNA methyltransferase in transgenic mice.

O6-Methylguanine DNA methyltransferase (MGMT; EC 2.1.1.63) is an unusual DNA repair protein in that it directly and specifically repairs a premutagenic DNA lesion without involving other proteins. MGMT removes the alkyl group from O6-alkylguanine in DNA in a unique stoichiometric reaction by accepting the alkyl group on a cysteine residue. The intracellular level of MGMT varies among tissues and appears to be inversely correlated to tissue-specific tumorigenesis induced by monofunctional alkylating agents. Because MGMT acts in solo, genetic manipulation of its expression may provide valuable insight into its contribution to cellular resistance to alkylation toxicity and to tumor induction. The human MGMT full length cDNA has been fused with a portion of the human transferrin (TF) 5'-flanking region (TF/MGMT). Transgenic founder mice were produced carrying the TF/MGMT transgene and then bred to establish stable transgenic lines. Human MGMT transcripts were specifically expressed in abundance in transgenic brain and liver tissues. In vitro MGMT assays revealed approximately 150-fold and approximately 25-fold increases in MGMT activity in transgenic brain and liver extracts respectively. Western blot analysis confirmed that human MGMT protein is specifically synthesized in transgenic brain and liver tissues.

Animals↗

PubMed1993