Search PubMed⌕ Search

Biomedical subjects

L Lin

Publications and source records attributed to L Lin.

At least 271 records · Page 15Linked to original sources
PubMed1995

Basic fibroblast growth factor regulates type I collagen and collagenase gene expression in human smooth muscle cells.

Basic fibroblast growth factor (bFGF) is a multifunctional peptide well known for angiogenic, neurotropic, and mesoderm-inducing effects. In the present study, we have investigated the effects of bFGF on collagen and collagenase gene expression in human iliac arterial smooth muscle cells. We report that bFGF inhibits type I collagen gene expression and collagen biosynthesis, with concomitant stimulation of collagenase gene expression. The smooth muscle cells incubated with human recombinant bFGF decreased the mRNA steady state levels of pro-alpha 1(I) type I collagen by as much as 72%. [3H]Hydroxyproline synthesis was also suppressed by 59% compared with untreated control cultures. Indirect immunofluorescence confirmed corresponding changes at the protein level. In contrast to the down-regulation of type I collagen gene expression, collagenase gene expression was found to be up-regulated severalfold by bFGF. The data suggest that bFGF is capable of regulating collagen and collagenase gene expression divergently in human smooth muscle cells and that the effects appear to be mediated at a pretranslational level.

Cells, Cultured↗
PubMed1995

[Effects of morphine on monosodium glutamate neurotoxicity and its mechanism].

The enhancing effects of morphine on monosodium glutamate (MSG) neurotoxicity and its blocking by naloxone were studied through morphological observation, together with detection of concentrations of intracellular free Ca2+ ([Ca2+]i) by Ca2+ indicator Fura-2/AM and lactate dehydrogenase (LDH) efflux in the bathing medium in primary cultures from 14-17 d old mouse fetal cortex. It was found that 10 min pre-incubation of young cortical neurons (7 day in vitro) with morphine 10(-7) or 10(-6) mol.L-1 substantially increased LDH release from 105.7% +/- 19.0% (treated with MSG alone) to 194.5% +/- 17.7% and 214.0% +/- 9.5% respectively after exposure to MSG 0.1 mmol.L-1, but pre-incubation with morphine (10(-7) or 10(-6) mol.L-1) plus naloxone (0.1 mmol.L-1) reversed the LDH release after treatment with the same concentration of MSG. Morphine (10(-7) or 10(-6) mol.L-1) produced little elevation of [Ca2+]i. However, when combined with MSG (0.1 mmol.L-1) morphine elevated the [Ca2+]i level much more than MSG alone. These results suggest that morphine markedly enhances excitotoxic neuron damage, which can be reversed by naloxone. Overloading of intracellular Ca2+ may be a simultaneous pathological mechanism underlying the neuronal damage and death that occur in excitatory toxicity.

Animals↗
PubMed1994

Cooperative binding of androgen receptors to two DNA sequences is required for androgen induction of the probasin gene.

The functional and structural interactions of two androgen receptor-binding sites in the 5'-flanking DNA of the rat probasin gene were determined. Deletion mapping and DNase I footprinting analysis had previously identified two androgen receptor-binding sites (ARBS) necessary for androgen induction of the probasin gene: ARBS-1, which resembled a glucocorticoid-responsive element, and ARBS-2, which had a unique sequence. In this study, maximal androgen induction in transient transfection studies only occurred when both sites were present. Neither binding site functioned independently, and deletion of the DNA sequence between the sites resulted in a 60% loss of androgen inducibility. Moreover, point mutations in either ARBS-1 or ARBS-2 led to > 90% loss in activity. Scatchard analysis indicated that ARBS-1 and ARBS-2 bound a synthetic androgen receptor, AR2, with Kd values of 20.0 and 6.7 nM, respectively. Consistent with the higher affinity, ARBS-2 bound AR2 at half the threshold concentration (200 ng) of that required in reciprocal DNase I footprinting experiments with ARBS-1. By comparison, protection occurred at a much lower threshold concentration of AR2 (60 ng) and to the same extent over each site when both sites were present, suggesting a cooperative interaction between the two sites. The cooperative effect was further substantiated when a point mutation in ARBS-1 blocked AR2 binding not only to ARBS-1, but also to ARBS-2. Similarly, a point mutation in ARBS-2 also prevented receptor binding to both sites. Androgen-specific regulation of probasin gene transcription therefore required an androgen-responsive region (positions -286 and +28) containing two androgen receptor-binding sites, where the binding of the androgen receptor to both sites occurred in a cooperative, mutually dependent manner.

Androgen-Binding Protein↗
PubMed1994

Determination of the catalytic site of creatine kinase by site-directed mutagenesis.

Site-directed mutagenesis was used to alter the amino-acid residues at the presumed catalytic site Cys-283 and ATP binding site Asp-340 of human creatine kinase B cDNA. In addition, a highly conserved arginine residue, Arg-292, was also mutated. Transfection of 0.1 to 1 microgram of recombinant plasmid into COS cells produced increasing creatine kinase activity in the cell lysate. The expression of mutant Cys283-Tyr and Cys283-Ser resulted in complete abolition of homodimer BB isoform enzymatic activity without alteration of the capacity for dimerization. Expression of mutants Arg292-His, Arg292-Leu, and Arg292-Gln produced non-functional homodimers, whereas expression of mutant Arg292-Lys produced a homodimer with enzymatic activity that was 42% of the enzymatic activity of the wild type. Expression of the Asp340-Glu mutant creatine kinase did not alter enzyme activity as compared to the wild type. Following heterodimerization, there was inhibition of the normal subunit by the mutant subunit, for both the BB and the MB dimer. The results showed residues Cys-283 and Arg-292 are essential for enzyme catalysis. The best fit model for the dimer is one in which there is close apposition of the two catalytic sites. The interaction of the individual subunits during dimerization provides a molecular approach for dominant negative modulation of the creatine kinase isozyme system in future genetic manipulative experiments.

Base Sequence↗
PubMed1994

Photochemical inactivation of pathogenic bacteria in human platelet concentrates.

Platelet concentrates (PC) may be infrequently contaminated with low levels of bacteria that can cause septicemia and death in patients receiving transfusion therapy. We evaluated the efficacy of a photochemical decontamination (PCD) technique using 8-methoxypsoralen (8-MOP) and long wavelength UV light (UVA) to inactivate bacteria in standard therapeutic PC. Twelve phylogenetically distinct pathogenic bacteria, 5 gram-positive and 7 gram-negative organisms, were seeded into PC to a final challenge dose ranging from 10(5) to 10(7) colony-forming units (CFU)/mL. Contaminated PC were treated with 8-MOP (5 micrograms/mL) and 5 J/cm2 of UVA, a PCD treatment regimen found to adequately preserve in vitro platelet function. Greater than 10(5) CFU/mL of all 5 gram-positive (Staphylococcus aureus, Streptococcus epidermidis, Streptococcus pyogenes, Listeria monocytogenes, and Corynebacterium minutissimum) and 2 of the gram-negative (Escherichia coli and Yersinia enterocolitica) organisms were inactivated. The remaining 5 gram-negative organisms were more resistant, with less than 10(1) to 10(3.7) CFU/mL inactivated under these conditions. The inactivation efficiency for this resistant group of gram-negative organisms was improved when PC were resuspended in a synthetic storage medium with reduced plasma protein concentration (15%) and an increased 8-MOP concentration (23.4 micrograms/mL). Illumination with 3 J/cm2 of UVA in this system inactivated greater than 10(5) CFU/mL of 4 resistant gram-negative organisms (Salmonella choleraesuis, Enterobacter cloacae, Serratia marcescens, and Klebsiella pneumoniae) and 10(4.1) CFU/mL of the most resistant gram-negative organism (Pseudomonas aeruginosa). This level of PCD treatment did not adversely affect in vitro platelet function. These results demonstrate that PCD using 8-MOP (5 to 23.4 micrograms/mL) effectively inactivated high levels of pathogenic bacteria in PC with adequate preservation of in vitro platelet properties.

Adult↗
PubMed1994

C-reactive proteins, limunectin, lipopolysaccharide-binding protein, and coagulin. Molecules with lectin and agglutinin activities from Limulus polyphemus.

In 1964, Levin and Bang discovered that gram-negative bacterial endotoxin could rapidly induce gelation of Limulus amebocyte lysate. This observation has led to the development of the most sensitive and specific method for the detection of bacterial endotoxin in pharmaceuticals and drugs intended for human use. Over 10 years ago, Bang injected endotoxin into young horseshoe crabs and observed a time and dose-dependent coagulation of the whole hemolymph. Limunectin, LEBP-PI, and Limulus CRP are found together with coagulin as part of the hemolymph clot at the time of endotoxin-induced exocytosis of amebocytes. In this manner, these molecules with agglutinin/lectin activities could work in concert to assist in the recognition and eventual removal of invading microorganisms from the circulating system. Although the mechanism of endotoxin-induced clot formation is to a large extent understood, the mechanism of clot dissolution and removal in the Limulus hemolymph remains to be clarified.

Acute-Phase Proteins↗
PubMed1994

Stabilization of myoglobin by multiple alanine substitutions in helical positions.

We have carried out a series of multiple Xaa-->Ala changes at nonadjacent surface positions in the sequence of sperm whale myoglobin. Although the corresponding single substitutions do not increase the thermal stability of the protein, multiple substitutions enhance the stability of the resulting myoglobins. The effect observed is an increase in the observed Tm (midpoint unfolding temperature) relative to that predicted from assuming additivity of the free energy changes corresponding to single mutations. The stabilization occurs in the presence of urea, as measured by the dependence of the unfolding temperature on urea concentration. The sites that have been altered occur in different helices and are not close in sequence or in the native structure of myoglobin. The observed effect is consistent with a role of multiple alanines in residual interactions in the unfolded state of the mutant proteins.

Alanine↗
PubMed1994

Management of avulsion injury of the PCL with reattachment.

Twenty-eight cases of avulsion injury of the posterior cruciate ligament (PCL) treated by reattachment were followed up to evaluate the results. Evaluation was based on appropriate functional and dynamic criteria: the results are classified as good, fair or poor. The interval between injury and repair ranged from 3 days to 2 months with a mean 4 weeks. The follow-up period ranged from 3 to 10 years with a mean of 5 years. The evaluation revealed 27 (96 per cent) good, one (4 per cent) fair and no poor results. It is apparent that operative reattachment is the treatment of choice for avulsion injury of the PCL, especially in early cases.

Adolescent↗
PubMed1994

Nephrotoxicity of low doses of tobramycin in rats: effect of the time of administration.

The circadian and the circannual variations of the nephrotoxicity of tobramycin were studied in female Sprague-Dawley rats. Animals were maintained on a light-dark period of 14/10 hrs (light on: 06h00 to 20h00). They were injected once daily for 4 and 10 days with saline or tobramycin at a dose of 40 mg/kg/day i.p. at either 08h00, 14h00, 20h00 and 02h00, in April 1991, July 91, October 91, January 92. In April 91, tobramycin injected at 14h00 during 10 days induced a significant increase of [3H]-thymidine incorporation into DNA of renal cortex as compared to other groups (p < 0.01): toxicity was highest at 14h00 and lowest at 02h00. No temporal change was observed in the renal cortical accumulation of tobramycin, and in serum creatinine after the 4 or 10 days of treatment. In experiments done in April, July and October 1991 and in January 1992, no circannual variation was found in tobramycin cortical levels but peaks of toxicity were observed at 02h00 in April and October 1991 and at 14h00 in July 1991 and January 1992. There was no linear correlation between the toxicity and the tobramycin accumulation in the renal cortex (r = 0.21). The data suggest that the circadian changes in tobramycin toxicity are due to temporal changes in the susceptibility of renal cells to tobramycin.

Animals↗
PubMed1994

Structural requirements for the biological activity of enterostatin.

A series of enterostatin analogues were tested to investigate the minimal structure required for activity to suppress the intake of high-fat (HF) diets. The dose-response curve to intracerebroventricular (ICV) enterostatin was U-shaped (maximal inhibition at 1 nmol). Removal or modification of the N-terminal valine from enterostatin (Val-Pro-Asp-Pro-Arg) abolished activity, as did C-terminal amidation. The tripeptide (Pro-Asp-Pro) and the cyclo-diketopiperazine cyclo-Asp-Pro retained activity whereas the linear Asp-Pro dipeptide was inactive. In rats adapted to a three-choice macronutrient diet, cyclo-Asp-Pro specifically inhibited fat intake and had near maximal inhibition (50%) at the 0.03 nmol dose. The enterostatin inhibitory effect on fat intake may reside in the cyclo-diketopiperazine molecule, cyclo-Asp-Pro.

Amino Acid Sequence↗
PubMed1994

Ganglioside GM1 reduces ethanol induced phospholipase A2 activity in synaptosomal preparations from mice.

The adaptation (tolerance) to chronic EtOH exposure was explained by the development of resistance to the disordering of the membrane phospholipids (PL). This phenomenon may be associated with changes in enzymes such as phospholipase A2 (PLA2) that govern PL metabolism. The data presented here, using the mouse inhalation model, supports and confirms previously reported findings that chronic exposure to EtOH substantially increased PLA2 activity in synaptosomal preparations from rat brain. We have previously reported that pretreatment with ganglioside GM1 reduced the intoxicating effect of EtOH in mice. The present study indicates that GM1 pretreatment both in vivo and in vitro reduced the EtOH-induced activation of PLA2 in synaptosomal preparations. Thus GM1 may exert its neuroprotective effects by influencing deacylation/reacylation of membrane phospholipids.

Animals↗
PubMed1994

Sequences of four splits of HLA-A10 group. Implications for serologic cross-reactivities and their evolution.

Nucleotide sequences of alleles encoding four serologically defined splits of the HLA-A10 group, A26.1, A26.3, A26.4, and A10SA, were determined. It was confirmed that the alleles coding for A26.1 and A26.3 are identical with A*2602 and A*2601, respectively. On the other hand, alleles for A26.4 and A10SA are thus far undescribed. A26.4 (A*2603) was different from the other A26 splits at three positions: 74 histidine, 76 valine, and 77 aspartate. A10SA (A*2604) was different from A26.3 (A*2601) by a single substitution of arginine by leucine at position 163. A comparison of amino acid sequences of HLA-A10 cross-reacting antigens revealed that all of the A10 group antigens share specific amino acids: 142 isoleucine, 144 glutamine, 145 arginine, 149 threonine, and 152 glutamate. Moreover, A26.1, A26.3, A10SA, and A43 share 76 alanine and 77 asparagine, which is consistent with the reported serologic cross-reactivity. The close relationship between the alleles for the A10 cross-reacting group was supported by a phylogenetic tree analysis for the HLA-A alleles.

Alleles↗
PubMed1994

Modulation of hepatocyte growth factor gene expression by estrogen in mouse ovary.

Hepatocyte growth factor (HGF) is expressed in a variety of tissues and cell types under normal conditions and in response to various stimuli such as tissue injury. In the present study, we demonstrate that the transcription of the HGF gene is stimulated by estrogen in mouse ovary. A single injection of 17 beta-estradiol results in a dramatic and transient elevation of the levels of mouse HGF mRNA. Sequence analysis has found that two putative estrogen responsive elements (ERE) reside at -872 in the 5'-flanking region and at +511 in the first intron, respectively, of the mouse HGF gene. To test whether these ERE elements are responsible for estrogen induction of HGF gene expression, chimeric plasmids containing variable regions of the 5'-flanking sequence of HGF gene and the coding region for chloramphenicol acetyltransferase (CAT) gene were transiently transfected into both human endometrial carcinoma RL 95-2 cells and mouse fibroblast NIH 3T3 cells to assess hormone responsiveness. Transfection results indicate that the ERE elements of the mouse HGF gene can confer estrogen action to either homologous or heterologous promoters. Nuclear protein extracts either from RL95-2 cells transfected with the estrogen receptor expression vector or from mouse liver bound in vitro to ERE elements specifically, as shown by band shift assay. Therefore, our results demonstrate that the HGF gene is transcriptionally regulated by estrogen in mouse ovary; and such regulation is mediated via a direct interaction of the estrogen receptor complex with cis-acting ERE elements identified in the mouse HGF gene.

3T3 Cells↗
PubMed1994

Molten globular characteristics of the native state of apomyoglobin.

Apomyoglobin, myoglobin lacking the haem group, is a natural intermediate in biosynthesis of myoglobin, and has some structural features in common with the haem-containing native state. Unfolding or refolding studies of apomyoglobin have identified a molten globule intermediate at acid pH. We show here that both the native state of apomyoglobin and the molten globule intermediate have highly plastic structures. Substitution of single amino acids on the surface or in the interior of helices in the native protein produce dramatic changes in the helix content and tryptophan emission of apomyoglobin at neutral and acidic pH. The signals from the intermediate and native apomyoglobin correlate closely suggesting that apomyoglobin itself has a molten globule-like character, its structure representing a population of interconverting substates rather than a fixed conformation.

Animals↗
PubMed1994

Localization of pituitary adenylate cyclase-activating polypeptide (PACAP) in the hypothalamus-pituitary system in rats: light and electron microscopic immunocytochemical studies.

The localization of pituitary adenylate cyclase-activating polypeptide (PACAP) in the hypothalamus-pituitary system in rats was examined in light and electron microscopic immunocytochemistry using a specific antiserum to synthetic PACAP 1-38 (R0831). In light microscopic study, intensely PACAP-immunostained perikarya were observed in the supraoptic and paraventricular magnocellular nucleus in the hypothalamus. In the median eminence, many immunoreactive nerve fibers were observed in the internal layer, but a few immunoreactive terminals were noticed in the external layer. In the pituitary gland, numerous immunoreactive nerve fibers were observed in the posterior lobe. In the intermediate lobe, moderately immunostained cells were observed, but in the anterior lobe no immunostained cells were noticed. In electron microscopic study, PACAP-immunoreactivity was examined by avidin-biotin peroxidase complex method. In the perikarya of the supraoptic and paraventricular magnocellular nucleus, DAB-reaction products were distributed diffusely in the cytoplasmic matrix, frequently attaching to the rough-surfaced endoplasmic reticulum. In the nerve terminals of the posterior lobe, reaction products were observed among the secretory granules, but sometimes upon them. In the cells of the intermediate lobe, reaction products were also distributed in the cytoplasmic matrix.

Animals↗
PubMed1994

The opaH locus of Neisseria gonorrhoeae MS11A is involved in epithelial cell invasion.

In order to produce a successful infection, Neisseria gonorrhoeae (GC) must attach to and invade mucosal epithelial cells. To identify GC gene products involved in this early interaction with host cells we constructed a gene bank derived from a clinical isolate of GC, and isolated a clone which had the capacity to adhere to the human endometrial adenocarcinoma tissue-culture line HEC-1-B. The cloned sequence was identified as a member of the opa gene family whose protein products have been associated with virulence. The GC chromosome contains numerous variant opa genes which, in MS11, are designated opaA-K. Previous work showed that expression of opaC confers a highly invasive phenotype upon strain MS11. When our cloned opa gene was mutated and returned to the GC MS11A chromosome by transformation and homologous recombination, we isolated one transformant that was significantly reduced in its invasive capacity. The locus mutated in this transformant was identified as opaH. Our results indicate that invasiveness of GC for human epithelial cells can be determined by more than one opa gene in strain MS11A.

Amino Acid Sequence↗