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Biomedical subjects

L Ma

Publications and source records attributed to L Ma.

At least 487 records · Page 27Linked to original sources
PubMed1994

Enhancement of migration activity in cholesterol-poor endothelial cells by pre-treating with HMG-CoA reductase inhibitors.

When bovine endothelial cells isolated from carotid arteries were cultured with a low density lipoprotein(LDL)-deficient serum for 2 days, their intracellular free cholesterol levels decreased. Under this condition, exposure of the endothelial cells to simvastatin and pravastatin, HMG-CoA reductase inhibitors used clinically, resulted in further decrease in the free cholesterol levels. The migration activity was enhanced in the cells cultured with the LDL-deficient serum and it was more enhanced in the cells treated with HMG-CoA reductase inhibitors under the LDL-deficient serum condition. These results suggest that cholesterol plays a negative role in wound healing in arterial walls and acts as one of the risk factors for arteriosclerosis.

Animals↗
PubMed1994

Defective assembly of membrane proteins in erythroid precursors of beta-thalassemic mice.

beta-Thalassemic mice provide a useful model for studying the pathophysiology of human beta-thalassemia in that one can perform experiments that are difficult to perform in humans. The ease of access to beta-thalassemic mouse marrow provided the opportunity to explore the cause of the ineffective erythropoiesis that characterizes severe beta-thalassemia in mouse and man. We hypothesized that the accumulation of excess alpha-globin might interfere with the normal assembly of red blood cell (RBC) membrane proteins, thus contributing to the severe intramedullary lysis. Femoral marrow was obtained from normal and beta-thalassemic mice, and RBC precursors were purified (> 90%) by panning and harvesting CD45- cells. The assembly of RBC membrane proteins was assessed by observing immunofluorescence patterns obtained on fixed permeabilized precursors using rabbit polyclonal antibodies directed against human spectrin, and band 4.1, and murine band 3. The distribution of the proteins was shown with a fluorescein-tagged goat antirabbit antibody. In contrast to normal mice, about 30% of intermediate and late stage erythroblasts in beta-thalassemic mice appear abnormal. Neither spectrin nor band 4.1 formed crisp rim fluorescence in these erythroid precursors of thalassemic mice, whereas assembly of band 3 appeared normal. Therefore, the assembly of membrane skeletal proteins is abnormal in murine beta-thalassemic erythroid precursors perhaps because of the deposition of unmatched alpha-globin chains.

Animals↗
PubMed1994

Large scale isolation and characterization of the molybdenum-iron cluster from nitrogenase.

Here we report the large scale isolation and characterization of a species, designated MoFe cluster, that exhibits an S = 3/2 EPR signal, and the comparison of this entity to isolated FeMo cofactor in N-methylformamide and to the active site of the enzyme nitrogenase. MoFe cluster is isolated from purified nitrogenase by extraction into acidic methyl ethyl ketone and it is stable in that solvent in the absence of thiols. As initially isolated, MoFe cluster solutions exhibit an S = 1/2 EPR signal that arises from an oxidized species that can be reduced by dithionite or thiols to an EPR silent state and then to a state that exhibits an S = 3/2 EPR signal. The S = 3/2 signal is as sharp as the signal exhibited by the protein and much sharper than the signal exhibited by isolated FeMo cofactor. Circular dichroism experiments indicate that unlike the last two species, MoFe cluster does not contain the endogenous ligand R-homocitrate and thus, the sharpness of the S = 3/2 signal is an intrinsic property of the metal center and does not depend upon specific interactions with this organic ligand or with the protein. Metal analyses indicate that the metal core responsible for the S = 3/2 signal contains 6 Fe atoms per molybdenum. X-ray absorption spectroscopy experiments show that although the molybdenum atom in MoFe cluster retains its pseudo-octahedral geometry, its first coordination shell has one less iron atom than that of FeMo cofactor and there has been a significant change in the long range order of the cluster.

Binding Sites↗
PubMed1994

Presence of constitutive type nitric oxide synthase in cultured astrocytes isolated from rat cerebra.

To define whether astrocytes express a constitutive nitric oxide synthase (NOS), nitric oxide (NO) producing activity in astrocytes derived fetal rat cerebra was examined. We found that the addition of an NOS inhibitor to cultures caused decrease in the basal cGMP levels in unstimulated astrocytes and the decrease was dose-dependent. Further, the expression of cNOS activity in unstimulated astrocytes was confirmed by histochemical staining for NADPH diaphorase and by measuring conversion of L-arginine to L-citrulline. We conclude that cultured astrocytes express constitutive NOS, in addition to inducible one.

Amino Acid Oxidoreductases↗
PubMed1994

Evaluation of a new photosensitizer, meso-tetra-hydroxyphenyl-chlorin, for use in photodynamic therapy: a comparison of its photobiological properties with those of two other photosensitizers.

The properties of a new photosensitizer, meso-tetra-hydroxyphenyl-chlorin (mTHPC), were studied using V79 cells (Chinese-hamster lung fibroblasts). Comparisons were made with 2 other photosensitizers: photofrin II (PII) and meso-tetra-hydroxyphenyl-porphyrin (mTHPP). A main advantage of mTHPC is that it has a strong absorption at 652 nm. Maximal cellular uptake of the dye was observed after 24 hr incubation of the cells with the drug. Using a confocal laser-scanning fluorescence microscope, we observed a diffuse distribution of mTHPC in the cytoplasm. Furthermore, the lipophilicity of mTHPC was compared with that of the components of PII by means of high-pressure liquid chromatography (HPLC). Absorption and fluorescence spectroscopy indicated that aggregated as well as monomeric mTHPC was bound to the cells. The action spectrum for photo-inactivation of the cells showed that aggregated mTHPC did not contribute significantly to its photosensitizing effects. In the present cellular system, the efficiency of photodynamic therapy (PDT) with mTHPC (cells were irradiated at a wavelength of 652 nm) was higher than with PII (irradiation at 630 nm) or with mTHPP (648 nm). The quantum yield for photo-inactivation of cells was smaller for mTHPC than for mTHPP and PII. The addition of 1,3-diphenylisobenzofuran (DPBF) reduced cell inactivation during PDT. Thus, PDT with mTHPC seems to act at least partly via a type-II process.

Animals↗
PubMed1994

A new kind of immobilized lipase in organic solvent and its structure model.

In this paper, we used Ca-alginate gel beads coated with polyetheneimine and glutaraldehyde to adsorb Expansum penicillium lipase. The immobilized lipase catalyzed esterification of 1-dodecanol with dodecanoic acid in benzene. The results show that when the concentration of Ca-alginate, polyetheneimine (PEI) and glutaraldehyde is 1%, 6% and 1%, respectively, the activity of the immobilized lipase and the amount of adsorbed protein are the highest. The immobilized lipase is better than the SDS-immobilized lipase. The activity of the immobilized lipase connected by glutaraldehyde is higher than the activity of that without glutaraldehyde. The initial rate of the immobilized lipase and lyophilized lipase powder is 5.9 x 10(2) nmol/min.mgpr and 2.8 x 10(1) nmol/min.mgpr, respectively. After the immobilized lipase catalyzed the esterification reaction at 37 degrees C for about 12 hours, 93.3% of 1-dodecanol was converted to ester, but for lyophilized lipase powder, only 17.5% converted. Based on all above results, we have presumed and explained the structure of this kind of immobilized lipase.

Alginates↗
PubMed1994

Correction of xeroderma pigmentosum repair defect by basal transcription factor BTF2 (TFIIH).

ERCC3 was initially identified as a gene correcting the nucleotide excision repair (NER) defect of xeroderma pigmentosum complementation group B (XP-B). The recent finding that its gene product is identical to the p89 subunit of basal transcription factor BTF2(TFIIH), opened the possibility that it is not directly involved in NER but that it regulates the transcription of one or more NER genes. Using an in vivo microinjection repair assay and an in vitro NER system based on cell-free extracts we demonstrate that ERCC3 in BTF2 is directly implicated in excision repair. Antibody depletion experiments support the idea that the p62 BTF2 subunit and perhaps the entire transcription factor function in NER. Microinjection experiments suggest that exogenous ERCC3 can exchange with ERCC3 subunits in the complex. Expression of a dominant negative K436-->R ERCC3 mutant, expected to have lost all helicase activity, completely abrogates NER and transcription and concomitantly induces a dramatic chromatin collapse. These findings establish the role of ERCC3 and probably the entire BTF2 complex in transcription in vivo which was hitherto only demonstrated in vitro. The results strongly suggest that transcription itself is a critical component for maintenance of chromatin structure. The remarkable dual role of ERCC3 in NER and transcription provides a clue in understanding the complex clinical features of some inherited repair syndromes.

Animals↗
PubMed1994

Isolation and characterization of cDNA clones for chloroplast translational initiation factor-3 from Euglena gracilis.

A complete cDNA clone encoding Euglena gracilis chloroplast translational initiation factor 3 (IF-3chl) has been obtained. Analysis of the sequence indicates that the IF-3chl mRNA contains the spliced leader found at the 5' end of nuclear encoded mRNAs in E. gracilis. The open reading frame for IF-3chl encodes a 537-amino acid protein. IF-3chl appears to be divided into four domains. The first 140 amino acids correspond to a transit peptide required for the import of IF-3chl into the chloroplast. The mature form of IF-3chl encompasses domains 2-4 and is about twice the size of Escherichia coli IF-3. The second domain has no homology to other known proteins. It begins with a stretch of 35 residues, of which about 30% are proline. Downstream from this region is a stretch of about 25 amino acids with a repeating (GX)n motif followed by a very acidic region. The third domain comprises a region of about 175 residues and has between 31 and 37% homology to the IF-3s found in other organisms. The IF-3 homology domain is followed by an acidic region which has no detectable homology to other sequences. Analysis of E. gracilis genomic DNA suggests that there are about four copies of the IF-3chl gene, one of which is probably a pseudogene. The activity of IF-3chl is inducible by light. However, the IF-3chl mRNA is present in approximately equal amounts in both dark- and light-grown cells, suggesting that the light-dependent induction of IF-3chl activity is post-transcriptional.

Amino Acid Sequence↗
PubMed1994

Redox mechanism as alternative to ligand binding for receptor activation delivering disregulated cellular signals.

Cross-linking with specific ligand is a general requirement for ordered activation of cell surface receptors. In this study we demonstrated a novel pathway for disregulated receptor activation through a redox mechanism. Treatment of murine thymocytes or spleen cells with thiol-reactive HgCl2, a known inducer of autoimmune proliferative lymphocyte disorders in rodents, was found to induce tyrosine phosphorylation of several cellular proteins, which was up to 100 times as extensive as that triggered by stimulation with antireceptor antibody or mitogen. Through the cross-linkage by thiol-reactive bivalent mercury, transmembrane CD4, CD3, and CD45 and glycosylphosphatidylinositol-anchored Thy-1 were aggregated together on thymocytes or T lymphocytes. Along with the aggregation of Thy-1 and CD4, nonreceptor protein tyrosine kinase p56lck was aggregated and activated. These events were linked to extensive protein tyrosine phosphorylation, which was visualized as a well localized spot beneath the membrane. Under appropriate conditions, this novel pathway of multiple receptor aggregation delivered a disregulated signal into T lymphocytes, which cross-talked to the antireceptor antibody-induced signal, for prolonged cell proliferation and IL-2 production. These results suggest a novel mechanism of disregulation of the ligand-dependent receptor function.

Animals↗
PubMed1994

Further evidence that the failure to cleave the aminopropeptide of type I procollagen is the cause of Ehlers-Danlos syndrome type VII.

Dermal fibroblasts from a Chinese Ehlers-Danlos syndrome type VII patient synthesized approximately equal amounts of normal pro-alpha 2(I) chains of type I procollagen and abnormal ones with electrophoretic mobility of pN alpha 2(I) chains, in which the amino-propeptide (N-propeptide) was retained. Reverse-transcriptase PCR analysis of the proband's RNA showed outsplicing of the 54 base exon 6 in half of the pro-alpha 2(I) mRNAs. Exon 6 encodes 18 amino acids of the N-telopeptide which contains the procollagen N-proteinase cleavage site and a cross-link precursor lysine. Loss of these sequences would result in failure to cleave the amino-propeptide of pro-alpha 2(I) and the accumulation of pN-alpha 2(I) chains. Nucleotide sequencing analyses of the proband's COL1A2 gene showed the presence of a T to C transition at position +2 of intron 6 in one allele and the proband is heterozygous for the defect. This mutation which destroyed the consensus GT dinucleotide at the 5' splice donor site of the intron is responsible for the loss of exon 6 by exon skipping. Electron microscopic analysis of the patient's dermis showed the presence of abnormal collagen I fibrils of irregular diameter and circularity. This mutation in COL1A2 in an EDS VII patient is the first reported case in the Chinese population and is identical to one reported for another EDS-VII (Libyan) patient. The occurrence of an identical mutation in two probands of different ethnic origin is direct evidence that the mutant genotype is the cause of the EDS VII phenotype.(ABSTRACT TRUNCATED AT 250 WORDS)

Base Sequence↗
PubMed1994

Involvement of nitric oxide and free radical (O2-) in neuronal injury induced by deprivation of oxygen and glucose in vitro.

Nitrix oxide (NO) is a free radical that has been recently proposed as a messenger molecule in the central nervous system. Since its involvement in glutamate neurotoxicity in vitro has been recently reported, using rat cortical cultures, we tested the hypothesis that NO also plays a role in neuronal injury induced by deprivation of oxygen and glucose. About 80-90% of neurons were killed in less than 12 h after a 4-6 h period of oxygen and glucose deprivation. N-nitro-L-arginine (L-NNA), an inhibitor of nitric oxidase synthase (NOS), significantly ameliorated this neuronal injury in a dose dependent manner. Since it has been suggested that NO is inactivated in a short time period by interaction with superoxide anions (O2-), which are generated during ischemia-reperfusion in vivo, we further evaluated the effect of superoxide dismutase (SOD) on neuronal injury in this test system. SOD failed, however, to protect against neuronal death. Furthermore, concomitant addition of SOD and L-NNA rather reduced the beneficial effects of L-NNA. Our results suggest therefore that NO, at least in part, mediates neuronal injury secondary to deprivation of oxygen and glucose in vitro and that superoxide anions may have a protective role by inactivating NO.

Amino Acid Oxidoreductases↗
PubMed1994

Regulation of the Msx2 homeobox gene during mouse embryogenesis: a transgene with 439 bp of 5' flanking sequence is expressed exclusively in the apical ectodermal ridge of the developing limb.

Msx2, a member of the highly conserved and widely distributed msh homeobox gene family, is expressed in a variety of sites in the vertebrate embryo, including craniofacial structures, heart, limb buds and otic and optic vesicles. In many of these sites, its expression is regulated by tissue interactions. Here we address the cis-trans regulatory interactions that direct Msx2 expression to specific regions of the embryo and enable it to respond to tissue interactions. We created a series of Msx2-lacZ fusion constructs with varying amounts of Msx2 genomic sequences. These were introduced into mouse embryos and their expression monitored by staining for beta-galactosidase activity. A construct bearing 5.2 kb of 5' flanking sequence, the intron, both exons and 3 kb of 3' flanking sequence was expressed in a pattern that closely resembled that of the endogenous Msx2 gene. In the E12.5 embryo, sites of expression included craniofacial mesenchyme, portions of the neural ectoderm, mesoderm in the distal limb bud and the overlying apical ectodermal ridge (AER). Removal of intronic and 3' UTR sequences slightly altered the pattern of Msx2 expression in the neural ectoderm of the E12 embryo. Deletion of 5' flanking sequences to -0.5 kb eliminated Msx2 expression in all sites except the AER. The proximal Msx2 promoter, including sequences required for the AER-specific expression of the -0.5 lacZ transgene, is highly conserved between mouse and human, one stretch exhibiting 100% identity over 72 bp. This conservation suggests that the AER element is under remarkably tight evolutionary constraint.

Amino Acid Sequence↗
PubMed1994

Mutational analysis of ERCC3, which is involved in DNA repair and transcription initiation: identification of domains essential for the DNA repair function.

The human ERCC3 gene, which corrects specifically the nucleotide excision repair defect in human xeroderma pigmentosum group B and cross-complements the repair deficiency in rodent UV-sensitive mutants of group 3, encodes a presumed DNA helicase that is identical to the p89 subunit of the general transcription factor TFIIH/BTF2. To examine the significance of the postulated functional domains in ERCC3, we have introduced mutations in the ERCC3 cDNA by means of site-specific mutagenesis and have determined the repair capacity of each mutant to complement the UV-sensitive phenotype of rodent group 3 cells. A conservative substitution of arginine for the invariant lysine residue in the ATPase motif (helicase domain I), six deletion mutations in the other helicase domains, and a deletion in the potential helix-turn-helix DNA-binding motif fail to complement the ERCC3 excision repair defect of rodent group 3 mutants, which implies that the helicase domains as well as the potential DNA-binding motif are required for the repair function of ERCC3. Analysis of carboxy-terminal deletions suggests that the carboxy-terminal exon may comprise a distinct determinant for the DNA repair function. In addition, we show that a functional epitope-tagged version of ERCC3 accumulates in the nucleus. Deletion of the putative nuclear location signal impairs neither the nuclear location nor the repair function, indicating that other sequences may (also) be involved in translocation of ERCC3 to the nucleus.

Amino Acid Sequence↗
PubMed1994

An experimental trial of artemether in treatment of Pneumocystis carinii in immunosuppressed rats.

An immunosuppressed rat model was established by injecting cortisone acetate 25 mg/rat twice a week for 4 weeks and 12.5 mg/rat for another 2 weeks subcutaneously. A development of Pneumocystis carinii pneumonia (PCP) was found at the end of the 6th week in all rats. These rats were injected intramuscularly with artemether at 100 mg/kg once a day for 5 consecutive days. All rats were necropsied at the end of the 8th week. The lung impression smears were stained with Gomori's stain, and Pneumocystis carinii cysts were counted. The ultrastructural changes of trophozoite, precyst and cyst were investigated by transmission electron microscopy on the 7th day after treatment.

Animals↗
PubMed1994

[Analysis on refraction status of eyes with normal vision].

Refractive status of 1,324 eyes with normal vision of the juvenile students in four grades was analysed. It was raised that the meaning of refractive status was different between the medical physiological optics and the physical or geometrical optics. "Emmetropia" did not really mean "the normal eye" in refraction as viewed from normal physiology, and the problem about the physiological refractive status of eyes in the juvenile was discussed preliminarily.

Adolescent↗